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The evaluation of a short-term holding system for the North American lobster, Homarus americanus 1983

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THE EVALUATION OF A SHORT-TERM HOLDING SYSTEM FOR THE NORTH AMERICAN LOBSTER, HOMARUS AMERICANUS by BLAIR ALAN STOCKWELL B . S c , T r e n t U n i v e r s i t y , 1977 A THESIS SUBMITTED IN PARTIAL FULFILMENT OF THE REQUIREMENTS FOR THE DEGREE OF MASTER OF SCIENCE i n THE FACULTY OF GRADUATE STUDIES (Department of A g r i c u l t u r a l Mechanics) We a c c e p t t h i s t h e s i s as c o n f o r m i n g t o the r e q u i r e d s t a n d a r d THE UNIVERSITY OF BRITISH COLUMBIA J a n u a r y , 1983 (c) B l a i r A l a n S t o c k w e l l , 1983 In presenting t h i s thesis i n p a r t i a l f u l f i l m e n t of the requirements for an advanced degree at the University of B r i t i s h Columbia, I agree that the Library s h a l l make i t f r e e l y available for reference and study. I further agree that permission for extensive copying of t h i s thesis for scholarly purposes may be granted by the head of my department or by his or her representatives. I t i s understood that copying or publication of t h i s thesis for f i n a n c i a l gain s h a l l not be allowed without my written permission. The University of B r i t i s h Columbia 1956 Main Mall Vancouver, Canada V6T 1Y3 DE-6 (3/81) i i ABSTRACT The water q u a l i t y parameters for use i n the design of lobster (Homarus americanus) holding f a c i l i t i e s were reviewed from published l i t e r a t u r e . The review indicated that several parameters (temperature, pH, s a l i n i t y , dissolved oxygen, oxygen demand, suspended s o l i d s , ammonia and n i t r i t e nitrogen) could be of major importance i n the design of commercial short-term holding f a c i l i t i e s where p a r t i a l or complete water recycle i s required. Of major inter e s t were the changes i n water q u a l i t y immediately following the introduction of lobsters into the system. Monitoring of the changes i n water q u a l i t y for d i f f e r e n t biomass loading rates and temperatures was car r i e d out at a commercial holding f a c i l i t y . The f a c i l i t y t y p i c a l l y holds approximately 1000 kg of lobster i n 13 tanks with a system water volume of 17500 1. Normally the water i s completely recycled with treatment consisting of drop aeration, sand f i l t r a t i o n and UV s t e r i l i z a t i o n . Results indicated that water temperature i s the most important factor in the maintenance of water q u a l i t y and design of a lobster holding f a c i l i t y . It has an impact on lobster i i i m e t a b o l i s m , t h e d i s s o l v e d o x y g e n c o n c e n t r a t i o n a n d b i o c h e m i c a l o x y g e n demand o f t h e h o l d i n g w a t e r , a s w e l l a s a f f e c t i n g t h e r a t e o f n i t r i f i c a t i o n o f ammonia. I t was d e m o n s t r a t e d t h a t d u r i n g t h e f i r s t 0.5 h a f t e r l o b s t e r i n t r o d u c t i o n a t t h e h i g h e x p e r i m e n t a l t e m p e r a t u r e s , w a t e r q u a l i t y o f t e n d e t e r i o r a t e d t o a d e l e t e r i o u s l e v e l , p a r t i c u l a r l y w i t h r e s p e c t t o d i s s o l v e d o x y g e n c o n c e n t r a t i o n . A c c e p t a b l e l e v e l s w e r e r e g a i n e d g r a d u a l l y a f t e r t h e c r i t i c a l p e r i o d . A t a n o r m a l o p e r a t i n g t e m p e r a t u r e o f 7°C and a l o b s t e r l o a d o f 1100 k g , o x y g e n demand was r e d u c e d , a s d i s s o l v e d o x y g e n d r o p p e d f r o m 10.5 t o 8.5 mg- 1 _ 1 c o m p a r e d t o a d r o p f r o m 9.0 t o 5.5 a t 13°C a n d f r o m 8.2 t o 3.3 m g - a t 19°C. An a t t e m p t t o d e t e c t n i t r i f i c a t i o n i n t h e s a n d f i l t e r s i n d i c a t e d t h a t v e r y l i m i t e d a c t i v i t y was p r e s e n t a t 12 a n d 17°C. The most i m p o r t a n t r e g u l a t i n g f a c t o r s w e r e p o s s i b l y t h e r e s i d e n c e t i m e o f t h e h o l d i n g w a t e r i n t h e f i l t e r u n i t s and c o m p e t i t i o n f r o m h e t e r o t r o p h i c b a c t e r i a . R e s u l t s f r o m e x p e r i m e n t s d e s i g n e d t o e s t a b l i s h t h e e f f e c t i v e n e s s o f t h e UV s t e r i l i z e r s a t c o n t r o l l i n g b a c t e r i a l l e v e l s i n t h e h o l d i n g w a t e r showed t h a t t h e u n i t s w e r e e f f e c t i v e i n c o n t r o l l i n g b a c t e r i a a t t h e t h r e e t e s t t e m p e r a t u r e s . G e n e r a l r e c o m m e n d a t i o n s r e g a r d i n g t h e d e s i g n and m a i n t e n a n c e o f a s h o r t - t e r m l o b s t e r h o l d i n g s y s t e m i n c l u d e : t e m p e r a t u r e c o n t r o l a s a n e s s e n t i a l d e s i g n f e a t u r e ; TOC c o n t r o l V TABLE OF CONTENTS Page ABSTRACT i i LIST OF TABLES V i i LIST OF FIGURES ix ACKNOWLEDGEMENTS X i i 1.0 INTRODUCTION 1 2.0 SPECIFIC RESEARCH OBJECTIVES 5 3.0 LITERATURE REVIEW 6 3.1 Introduction 6 3.2 Microbiology/Pathology 8 3.3 Temperature 10 3.4 pH 11 3.5 S a l i n i t y 13 3.6 Dissolved Oxygen 14 3.7 Solids and Organic Carbon 15 3.8 Nitrogenous Compounds 18 4.0 SYSTEM DESCRIPTION 29 5.0 MATERIALS AND METHODS 38 5.1 Introduction 38 5.2 Sampling Requirements 40 5.3 Total Organic Carbon versus Chemical Oxygen Demand 42 5.4 Time Series Experiments 43 5.5 S i l i c a Sand F i l t e r Experiments 44 5.6 U l t r a v i o l e t S t e r i l i z e r Experiments 46 6.0 EXPERIMENTAL RESULTS 49 6.1 Sampling Requirements 49 6.2 Total Organic Carbon versus Chemical Oxygen Demand 51 6.3 Time Series Experiments 53 6.4 S i l i c a Sand F i l t e r Experiments 66 6.5 U l t r a v i o l e t S t e r i l i z e r Experiments 93 Table of Contents (cont'd) 7.0 DISCUSSION 7.1 Sampling Requirements 7.2 Total Organic Carbon versus Chemical Oxygen Demand 7.3 Time Series Experiments 7.4 S i l i c a Sand F i l t e r Experiments 7.5 U l t r a v i o l e t S t e r i l i z e r Experiments 8.0 SUMMARY AND CONCLUSIONS 9.0 RECOMMENDATIONS 10.0 LITERATURE CITED 11.0 APPENDICES v i i LIST OF TABLES Table T i t l e Page 1 Important water q u a l i t y p a rameters t o be 28 m o n i t o r e d i n a l i v e l o b s t e r s t o r a g e system 2 L o b s t e r h o l d i n g system equipment and 35 s p e c i f i c a t i o n s 3 ANOVA t a b l e showing r e s u l t s o f t h r e e - l e v e l 50 n e s t e d a n a l y s i s o f v a r i a n c e on ammonia-N samples t o d e t e r m i n e sample r e q u i r e m e n t s 4 C o n d i t i o n s f o r t i m e - s e r i e s e x p e r i m e n t s 54 5 Mean (Y) w i t h 95% c o n f i d e n c e i n t e r v a l s , 68 s t a n d a r d d e v i a t i o n (S.D.) and c o e f f i c i e n t of v a r i a t i o n (V) c a l c u l a t e d f o r ammonia-N l e v e l s measured i n sample s e r i e s c o l l e c t e d p r i o r t o s i l i c a sand f i l t e r e x p e r i m e n t s 6 Mean (Y) w i t h 9 5 % c o n f i d e n c e i n t e r v a l s , 69 s t a n d a r d d e v i a t i o n (S.D.) and c o e f f i c i e n t o f v a r i a t i o n (V) c a l c u l a t e d f o r NO3-N l e v e l s measured i n sample s e r i e s c o l l e c t e d p r i o r t o s i l i c a sand f i l t e r e x p e r i m e n t s 7 Summary of d i s s o l v e d oxygen (DO), pH and 70 s a l i n i t y l e v e l s measured d u r i n g t h e s i l i c a sand f i l t e r e x p e r i m e n t s 8 I n i t i a l and f i n a l c o n c e n t r a t i o n s , r a n g e , mean 73 ( Y ) w i t h 9 5 % c o n f i d e n c e i n t e r v a l s , s t a n d a r d d e v i a t i o n (S.D.) and c o e f f i c i e n t o f v a r i a t i o n (V) of ammonia-N i n t i m e - s e r i e s samples t a k e n p r i o r t o passage t h r o u g h t h e s i l i c a sand f i l t e r s 9 I n i t i a l and f i n a l c o n c e n t r a t i o n s , range, mean 74 (Y) w i t h 9 5 % c o n f i d e n c e i n t e r v a l s , s t a n d a r d d e v i a t i o n (S.D.) and c o e f f i c i e n t of v a r i a t i o n (V) of n i t r a t e - N i n t i m e - s e r i e s samples t a k e n p r i o r t o passage t h r o u g h t h e s i l i c a sand f i l t e r s 10 I n i t i a l and f i n a l c o n c e n t r a t i o n s , range, mean 75 (Y) w i t h 9 5 % c o n f i d e n c e i n t e r v a l s , s t a n d a r d d e v i a t i o n (S.D.) and c o e f f i c i e n t of v a r i a t i o n (V) of n i t r i t e - N i n t i m e - s e r i e s samples t a k e n p r i o r t o passage t h r o u g h t h e s i l i c a sand f i l t e r s T i t l e H i g h e s t NH3-N c o n c e n t r a t i o n s t h a t o c c u r r e d d u r i n g t h e t i m e - s e r i e s e x p e r i m e n t s c a l c u l a t e d from measured t o t a l ammonia-N v a l u e s and pH. i x L I S T OF FIGURES F i g u r e T i t l e Page 1 T h r e e d i m e n s i o n a l d i a g r a m o f t h e b o u n d a r y o f 16 l e t h a l c o n d i t i o n s f o r l o b s t e r s u n d e r v a r i o u s c o m b i n a t i o n s o f t e m p e r a t u r e , s a l i n i t y a n d o x y g e n . S y s t e m c o m p o n e n t s and w a t e r f l o w p a t t e r n o f l o b s t e r h o l d i n g u n i t . 30 3 One o f t h e t h r e e s e t s o f s t a c k e d f i b e r g l a s s 31 h o l d i n g t a n k s u s e d f o r l o b s t e r s t o r a g e . 4 One o f t h r e e s i l i c a s a n d f i l t e r s u s e d f o r 32 t r e a t m e n t o f t h e l o b s t e r h o l d i n g w a t e r . 5 One o f t h e t h r e e UV s t e r i l i z e r u n i t s s h o w i n g 33 c i r c u l a r a r r a n g e m e n t o f UV l a m p s . 6 R e f r i g e r a t i o n u n i t u s e d t o m a i n t a i n l o b s t e r 34 h o l d i n g w a t e r t e m p e r a t u r e . 7 R e l a t i o n s h i p b e t w e e n TOC a n d COD o f l o b s t e r 52 h o l d i n g w a t e r . 8 C h a n g e s i n TOC c o n c e n t r a t i o n s r e c o r d e d d u r i n g 56 r e p r e s e n t a t i v e t i m e - s e r i e s e x p e r i m e n t s . 9 C h a n g e s i n d i s s o l v e d o x y g e n c o n c e n t r a t i o n s 58 r e c o r d e d d u r i n g r e p r e s e n t a t i v e t i m e - s e r i e s e x p e r i m e n t s . 10 pH c h a n g e s r e c o r d e d d u r i n g r e p r e s e n t a t i v e t i m e - 60 s e r i e s e x p e r i m e n t s . 11 C h a n g e s i n ammonia-N c o n c e n t r a t i o n s r e c o r d e d 62 d u r i n g r e p r e s e n t a t i v e t i m e - s e r i e s e x p e r i m e n t s . 12 C h a n g e s i n n i t r i t e - N c o n c e n t r a t i o n s r e c o r d e d 63 d u r i n g r e p r e s e n t a t i v e t i m e - s e r i e s e x p e r i m e n t s . 13 C h a n g e s i n n i t r a t e - N r e c o r d e d d u r i n g 64 r e p r e s e n t a t i v e t i m e - s e r i e s e x p e r i m e n t s . 14 D i f f e r e n c e s b e t w e e n f i l t e r 1 i n f l o w a n d o u t f l o w 76 ammonia-N a n d n i t r a t e - N c o n c e n t r a t i o n s d u r i n g e x p e r i m e n t 1 a t 7°C. T i t l e D i f f e r e n c e s between f i l t e r 2 i n f l o w and o u t f l o w ammonia-N and n i t r a t e - N c o n c e n t r a t i o n s d u r i n g experiment 1 a t 7°C. D i f f e r e n c e s between f i l t e r 3 i n f l o w and o u t f l o w ammonia-N and n i t r a t e - N c o n c e n t r a t i o n s d u r i n g e x periment 1 a t 7°C. D i f f e r e n c e s between f i l t e r 1 i n f l o w and o u t f l o w ammonia-N and n i t r a t e - N c o n c e n t r a t i o n s d u r i n g e x periment 2 a t 7°C. D i f f e r e n c e s between f i l t e r 2 i n f l o w and o u t f l o w ammonia-N and n i t r a t e - N c o n c e n t r a t i o n s d u r i n g e x p e r i m e n t 2 a t 7°C. D i f f e r e n c e s between f i l t e r 3 i n f l o w and o u t f l o w ammonia-N and n i t r a t e - N c o n c e n t r a t i o n s d u r i n g e x periment 2 a t 7°C. D i f f e r e n c e s between f i l t e r 1 i n f l o w and o u t f l o w ammonia-N and n i t r a t e - N c o n c e n t r a t i o n s d u r i n g e x p e r i m e n t 1 a t 12°C. D i f f e r e n c e s between f i l t e r 2 i n f l o w and o u t f l o w ammonia-N and n i t r a t e - N c o n c e n t r a t i o n s d u r i n g e x periment 1 a t 12°C. D i f f e r e n c e s between f i l t e r 3 i n f l o w and o u t f l o w ammonia-N and n i t r a t e - N c o n c e n t r a t i o n s d u r i n g e x p e r i m e n t 1 a t 12°C. D i f f e r e n c e s between f i l t e r 1 i n f l o w and o u t f l o w ammonia-N and n i t r a t e - N c o n c e n t r a t i o n s d u r i n g e x periment 2 a t 12°C. D i f f e r e n c e s between f i l t e r 2 i n f l o w and o u t f l o w ammonia-N and n i t r a t e - N c o n c e n t r a t i o n s d u r i n g e x periment 2 a t 12°C. D i f f e r e n c e s between f i l t e r 3 i n f l o w and o u t f l o w ammonia-N and n i t r a t e - N c o n c e n t r a t i o n s d u r i n g e x periment 2 a t 12°C. D i f f e r e n c e s between f i l t e r 1 i n f l o w and o u t f l o w ammonia-N and n i t r a t e - N c o n c e n t r a t i o n s d u r i n g e x p e r i m e n t 1 a t 17°C. XI F i g u r e T i t l e Page 27 D i f f e r e n c e s b e t w e e n f i l t e r 2 i n f l o w a n d o u t f l o w 91 ammonia-N and n i t r a t e - N c o n c e n t r a t i o n s d u r i n g e x p e r i m e n t 1 a t 17°C. 28 D i f f e r e n c e s b e t w e e n f i l t e r 3 i n f l o w a n d o u t f l o w 92 ammonia-N a n d n i t r a t e - N c o n c e n t r a t i o n s d u r i n g e x p e r i m e n t 1 a t 17°C. 29 D i f f e r e n c e s b e t w e e n f i l t e r 1 i n f l o w a n d o u t f l o w 94 ammonia-N and n i t r a t e - N c o n c e n t r a t i o n s d u r i n g e x p e r i m e n t 2 a t 17°C. 30 D i f f e r e n c e s b e t w e e n f i l t e r 2 i n f l o w a n d o u t f l o w 95 ammonia-N and n i t r a t e - N c o n c e n t r a t i o n s d u r i n g e x p e r i m e n t 2 a t 17°C. 31 D i f f e r e n c e s b e t w e e n f i l t e r 3 i n f l o w a n d o u t f l o w 96 ammonia-N and n i t r a t e - N c o n c e n t r a t i o n s d u r i n g e x p e r i m e n t 2 a t 17°C. 32 C h a n g e s i n b a c t e r i a l c o n c e n t r a t i o n r e c o r d e d o v e r 97 t i m e i n t h e a q u a r i u m a nd t a n k d u r i n g UV e x p e r i m e n t 1 a t 7°C. 33 C h a n g e s i n b a c t e r i a l c o n c e n t r a t i o n r e c o r d e d o v e r 98 t i m e i n t h e a q u a r i u m a nd t a n k d u r i n g UV e x p e r i m e n t 2 a t 7°C. 34 C h a n g e s i n b a c t e r i a l c o n c e n t r a t i o n r e c o r d e d o v e r 99 t i m e i n t h e a q u a r i u m a n d t a n k d u r i n g UV e x p e r i m e n t a t 12°C. 35 Changes i n b a c t e r i a l c o n c e n t r a t i o n r e c o r d e d o v e r 100 t i m e i n t h e a q u a r i u m a nd t a n k d u r i n g UV e x p e r i m e n t a t 17°C. 36 E f f e c t o f t e m p e r a t u r e on o x y g e n c o n s u m p t i o n 112 d u r i n g t h e f i r s t 0.25 h r o f r e p r e s e n t a t i v e t i m e - s e r i e s e x p e r i m e n t s , e x p r e s s e d a s mg o f o x y g e n consumed p e r g o f l o b s t e r p e r h r , c o m p a r e d t o p u b l i s h e d v a l u e s f r o m t w o l i t e r a t u r e s o u r c e s . 37 E f f e c t o f t e m p e r a t u r e on b a c t e r i a l g r o w t h 128 r a t e i n t h e a q u a r i u m . x i i ACKNOWLEDGEMENTS My s i n c e r e s t a p p r e c i a t i o n must go t o t h e f o l l o w i n g : Dr. N.R. B u l l e y , who d i r e c t e d t h i s r e s e a r c h and p r o v i d e d encouragement and a d v i c e . P r o f e s s o r L.M. S t a l e y , Dr. R. Bose and Dr. T. Howard f o r s e r v i n g on the t h e s i s committee and r e v i e w i n g t h i s t h e s i s . The s t a f f of P a c i f i c Rim S h e l l f i s h f o r t h e i r s u p p o r t and c o o p e r a t i o n t h r o u g h o u t t h e d u r a t i o n of t h i s r e s e a r c h . B e r n i e S o u t h c o t t , f o r her generous s u p p o r t and t e c h n i c a l a d v i c e d u r i n g the m i c r o b i o l i c a l work a s s o c i a t e d w i t h t h i s t h e s i s . F r a n c e s Buchan, f o r her word p r o c e s s i n g w i z a r d r y and her generous commitment o f t i m e and e f f o r t i n p r e p a r i n g t h i s document. Dr. P. L i a o , N. J a c k s o n and J . P e h l k e , f o r t h e i r t e c h n i c a l a s s i s t a n c e d u r i n g t h i s r e s e a r c h . L e s l i e Matthews, f o r her h e l p w i t h one of t h e f i g u r e s . M i c h a e l F a t t o r i , f o r h i s v a l u a b l e g u i d a n c e i n t e c h n i c a l and n o n - t e c h n i c a l m a t t e r s t h r o u g h o u t t h e t h e s i s work. B.C. S c i e n c e C o u n c i l , f o r t h e i r f i n a n c i a l s u p p o r t t h r o u g h t h e Graduate Research E n g i n e e r i n g and Technology Award programme. C h r i s H a t f i e l d , f o r a l l o w i n g me a c c e s s t o HCL o f f i c e equipment and s u p p l i e s f o r t h e p r e p a r a t i o n of t h i s document. 1 1.0 INTRODUCTION The American Lobster (Homarus americanus) has lony been and continues to be a very popular food item i n North America and around the globe. This popularity ensures that the lobster fi s h e r y w i l l continue to be one of the most economically important on the east coast of North America. A summary of s t a t i s t i c s from 1954-1974 by Cobb (1976) indicates that the continuously increasing demand for lobster i s r e f l e c t e d i n i t s increasing value but not in the quantities landed in the U.S. A similar s i t u a t i o n exists on the east coast of Canada (DeWolf 1974; Campbell and Duggan 1980). Numerous investigators have reported apparently declining lobster populations (DeWolf 1974; Thomas 1973) and i t i s obvious from their data that e f f e c t i v e management regulations are required to a l l e v i a t e the c r i t i c a l s i t u a t i o n i n t h i s f i s h e r y . It i s clear that in recent years f i s h i n g e f f o r t has increased dramatically, whereas lobster landings have slowly declined. The apparent decline i n stocks, coupled with increasing consumer demand, has generated a great deal of interest i n commercial lobster rearing. A considerable amount of federal money has recently been made available for research into the various biochemical, technological and economic aspects of lobster aquaculture i n Canada, through the Department of Fisheries and Oceans. Similar research i s being funded in the United States, through the Sea Grant Program. U n t i l t h i s 2 r e s e a r c h l e a d s t o c o m m e r c i a l l y v i a b l e l o b s t e r o p e r a t i o n s , the consumer remains dependent upon t h e n a t u r a l l o b s t e r s t o c k s . C o m m e r c i a l l y v i a b l e l o b s t e r c u l t u r i n g o p e r a - t i o n s w i l l most p r o b a b l y emerge i n t h e near f u t u r e p r o v i d e d t h a t s o l u t i o n s are found f o r a number of i m p o r t a n t problems. The a r e a s r e q u i r i n g t h e most i n t e n s i v e r e s e a r c h e f f o r t s i n c l u d e l o b s t e r n u t r i t i o n , energy a c q u i s i t i o n and u t i l i z a t i o n , and water t r e a t m e n t t e c h n o l o g y ( J o h n s t o n and B o t s f o r d 1980). Host of the c ommercial c a t c h of Homarus i s s o l d l i v e , r e q u i r i n g s t o r a g e f o r p e r i o d s o f up t o s i x months i n v a r i o u s t y p e s of h o l d i n g f a c i l i t i e s . T r a d i t i o n a l l y , t h e s e h o l d i n g u n i t s u t i l i z e ambient t e m p e r a t u r e sea w a t e r , a r e r e l a t i v e l y l a r g e , and a r e r e s t r i c t e d t o s e l e c t e d c o a s t a l i n t e r t i d a l a r e a s . W i t h th e e x p a n s i o n of t h e l i v e l o b s t e r market t o i n c l u d e numerous n o n - l o b s t e r p r o d u c i n g c o a s t a l a r e a s , as w e l l as i n l a n d s i t e s , has come a more s o p h i s t i c a t e d approach t o l i v e l o b s t e r s t o r a g e i n g e n e r a l , and s p e c i f i c a l l y i n water q u a l i t y manage- ment. I n an attempt t o reduce m o r t a l i t y and m a i n t a i n a q u a l i t y p r o d u c t , water t r e a t m e n t t e c h n o l o g y u t i l i z e d i n o t h e r f i e l d s has been borrowed and m o d i f i e d , when n e c e s s a r y , t o s u i t t h e r e q u i r e m e n t s o f l o b s t e r h o l d i n g systems. The most complex h o l d i n g f a c i l i t i e s , w i t h r e s p e c t t o water q u a l i t y c o n t r o l , a r e t h o s e t h a t a r e s i t u a t e d i n l a n d , as t h e s e o p e r a t i o n s r e q u i r e , out o f n e c e s s i t y , r e c i r c u l a t i o n of t h e s a l t water medium. P a r t i a l o r complete r e c y c l e may a l s o be u s e f u l i n c o a s t a l o p e r a t i o n s f o r t e m p e r a t u r e c o n t r o l and energy c o n s e r v a t i o n . 3 With r e c i r c u l a t i o n though comes deteriorating water q u a l i t y over time due to eventual build-up of metabolities (Hughes et a l . 1972; Shlesser and Tchobanoglous 1974; Gravitz et a l . 1975) and increased pathogen concentration (Fisher et a l . 1978). A number of units and operating procedures have been developed to cope with d i f f e r e n t aspects of these problems (Getchell 1953; Wilder 1953; Wilder and McLeese 1957; Thomas 1962, MeLeese and Wilder 1964; Ayres and Wood 1977; and Cormick and Stewart 1977). Although the objectives in lobster aqua- culture are somewhat divergent from those set out in lobster holding, many of the problems are common to both operations. As a r e s u l t , the increased interest in lobster culture that has taken place during the l a s t few years has s i g n i f i c a n t l y aided in the development of systems designed s t r i c t l y for the maintenance of market size lobsters (Sastry 1975; Hand 1977). From a d i s t r i b u t o r ' s or r e t a i l e r ' s point of view, there are a number of economic advantages to successful l i v e storage. These include (1) increased f l e x i b i l i t y of marketing and dispatch which can be carr i e d out at convenient times or on demand, (2) reduced mortality by maintaining animals i n water, and (3) insurance that the produce arrives at i t s destination in good condition. Long-term storage brings further benefits, p a r t i c u l a r l y to those producers who are geographically isolated from major markets. Large consignments can be b u i l t up, customers can be supplied with equally graded shipments, advantage can be taken of bulk packing and transportation and, 4 most i m p o r t a n t o f a l l , l o b s t e r s may be pu r c h a s e d when cheaper and more abundant, and s t o r e d and s o l d when demand and p r i c e s a r e a t a peak. I t i s a l o b s t e r h o l d i n g system c h a r a c t e r i z e d by many of the f e a t u r e s l i s t e d above t h a t i s t h e s u b j e c t o f t h i s t h e s i s . The f a c i l i t y * e x i s t s s t r i c t l y as a h o l d i n g o p e r a t i o n f o r whole- s a l e and r e t a i l t r a d e , and i s r e l a t i v e l y s i m p l e when compared t o p r o t o t y p e l o b s t e r c u l t u r i n g f a c i l i t i e s . I t i s t h i s a p p arent s i m p l i c i t y of d e s i g n and t e c h n o l o g y t h a t makes t h i s o p e r a t i o n worthy of s t u d y . A h o l d i n g system t h a t u t i l i z e s t e s t e d and e s t a b l i s h e d t e c h n o l o g y , and i s r e l a t i v e l y s i m p l e t o f a b r i c a t e and manage may have g r e a t p o t e n t i a l i n a v a r i e t y of o t h e r s i t u a t i o n s and l o c a t i o n s . T h e r e f o r e t h e o v e r l y i n g o b j e c t i v e s of t h e r e s e a r c h r e p o r t e d here were i n i t i a l l y , t o e s t a b l i s h t h e demands on t h e e x i s t i n g system m a i n l y from a water q u a l i t y v i e w p o i n t , and s e c o n d l y t o e v a l u a t e system response t o t h e s e demands so t h a t recommendations t o upgrade the system c o u l d be f o r m u l a t e d and p r e s e n t e d t o t h e managers of t h e f a c i l i t y . I t was f e l t t h a t t h e knowledge a c q u i r e d from t h e s t u d y o f t h i s p a r t i c u l a r l i v e l o b s t e r h o l d i n g o p e r a t i o n c o u l d be a p p l i e d t o the d e s i g n and maintenance o f f u t u r e l i v e s e a f o o d s t o r a g e u n i t s and even i n t e n s i v e and e x t e n s i v e a q u a c u l t u r e p r o j e c t s . • P a c i f i c Rim S h e l l f i s h L t d . - The L o b s t e r Man 1604 D u r a n l e a u , Vancouver, B.C. 5 2.0 SPECIFIC RESEARCH OBJECTIVES In r e s p onse t o t h e o v e r l y i n g o b j e c t i v e s o u t l i n e d i n t h e p r e c e e d i n g s e c t i o n , f o u r s p e c i f i c r e s e a r c h o b j e c t i v e s were f o r m u l a t e d and a r e as f o l l o w s : 1. To e s t a b l i s h , from a l i t e r a t u r e r e v i e w , t h e most i m p o r t a n t water q u a l i t y p a rameters t o be c o n s i d e r e d i n t h e d e s i g n of a s h o r t - t e r m l o b s t e r h o l d i n g f a c i l i t y . 2. To d e t e r m i n e changes i n t h e l e v e l s o f e s t a b l i s h e d water q u a l i t y parameters i m m e d i a t e l y a f t e r i n t r o d u c t i o n of l o b s t e r s i n t o an e x i s t i n g s h o r t - t e r m h o l d i n g system. 3. To d e t e r m i n e changes i n t h e l e v e l s o f e s t a b l i s h e d water q u a l i t y p a rameters d u r i n g a p e r i o d o f s h o r t - t e r m l o b s t e r h o l d i n g . 4. To propose b a s i c d e s i g n c r i t e r i a f o r the maintenance of a c c e p t a b l e water q u a l i t y i n a s h o r t - t e r m l o b s t e r h o l d i n g f a c i l i t y based on t h e r e s u l t s o b t a i n e d d u r i n g t h e f u l f i l l m e n t of O b j e c t i v e s 2 and 3. 6 3.U LITERATURE REVIEW 3.1 I n t r o d u c t i o n Due i n p a r t t o r e c e n t e f f o r t s a t commercial l o b s t e r r e a r i n g and n a t u r a l s t o c k management and enhancement, a s u b s t a n t i a l amount of l i t e r a t u r e r e g a r d i n g e n v i r o n m e n t a l r e q u i r e m e n t s of Homarus (amer i c a n u s and gammarus) i s a v a i l a b l e . A l t h o u g h a g r e a t d e a l of a d d i t i o n a l i n v e s t i g a t i o n i s needed b e f o r e i n t e n s i v e l o b s t e r c u l t u r e becomes a r e a l i t y , a g e n e r a l p i c t u r e d e s c r i b i n g t h e b a s i c e n v i r o n m e n t a l needs of l o b s t e r s has d e v e l o p e d , and i t i s t h i s k i n d o f i n f o r m a t i o n t h a t i s p r o v i n g t o be i n v a l u a b l e t o d e s i g n e r s and o p e r a t o r s of l o b s t e r h o l d i n g f a c i l i t i e s . The p r i m a r y o b j e c t i v e of t h i s l i t e r a t u r e r e v i e w i s t o e s t a b l i s h w hich water q u a l i t y p a rameters a r e of g r e a t e s t i m p o r t a n c e i n l o b s t e r h o l d i n g systems, and over what range t h e s e e s t a b l i s h e d p a r a m e t e r s must be m a i n t a i n e d . I n f o r m a t i o n concerned w i t h parameter m o n i t o r i n g methods, and s a m p l i n g t e c h n i q u e s and i n t e r v a l s was a l s o o b t a i n e d from the l i t e r a t u r e and w i l l be o u t l i n e d i n more d e t a i l i n a l a t e r s e c t i o n . P h y l o g e n e t i c a l l y , t h e l o b s t e r i s a c r u s t a c e a n b e l o n g i n g t o the s u b - c l a s s M a l a c o s t r a c a , which c o n t a i n s most of t h e commer- c i a l l y i m p o r t a n t s p e c i e s . There a r e two s p e c i e s of l o b s t e r : Homarus gammarus, t h e European l o b s t e r , o c c u r i n g i n the e a s t e r n A t l a n t i c w a t e r s from t h e A r c t i c C i r c l e t o Morocco and i n t o t h e M e d i t e r r a n e a n , w i t h i t s c e n t r e of d i s t r i b u t i o n the B r i t i s h I s l e s ; and Homarus americanus found on t h e A t l a n t i c c o a s t of 7 Northern United States and Canada (Headstrom 1979). There exists a variety of good general publications on Hoiaarus that provide useful background information on lobster biology, ecology and behavior, as well as numerous references concerned with s p e c i f i c problems i n lobster research (Herrick 1909; Doliber 1973; Taylor 1975; Cobb 1976; Richards and Wickins 1979). These publications act as an excellent resource for a more in-depth look at the water q u a l i t y problems associated with l i v e lobster storage. Unsatisfactory conditions in a lobster holding system aris e as a result of either animal related a l t e r a t i o n s to the environment or a l t e r a t i o n s caused by external influences. In a land-based holding operation, employing moderately sophisticated technology, external influence, such as ambient a i r temperature, can be kept to a minimum, providing a suitable water supply i s a v a i l a b l e . Therefore, i t i s the biochemical and behavioral c h a r a c t e r i s t i c s of the animal that complicate l i v e lobster storage procedures. Research e f f o r t s have reacted i n two ways to t h i s s i t u a t i o n . I n i t i a l l y investigators established which biochemical and behavioral c h a r a c t e r i s t i c s were causing problems in lobster holding, and secondly what l e v e l of perturbation was acceptable in each case. One very important consideration i s that environmental q u a l i t y i s very much dependent upon animal density, the more lobsters there are per unit area or volume the more concentrated or intense the associated problems are l i k e l y to be (Sastry and Z e i t l i n - H a l e 1975; Shlesser 1974). 8 The aggressive nature and c a n n i b a l i s t i c tendencies of both juvenile and adult lobsters ex i s t as the most s i g n i f i c a n t behavioral problems that occur in a holding or culturing system. A popular approach in the design of lobster culturing systems i s to provide i n d i v i d u a l growth chambers for each animal so that behavioral a f f e c t s are minimized (Chanley and Terry 1974). The economics of lobster holding w i l l not allow t h i s approach, so alter n a t i v e methods must be employed to ensure that these factors are controlled. 3. 2 Microbiology/Pathology Most l i t e r a t u r e related to lobster culture and storage states that b a c t e r i a l populations must be cl o s e l y controlled to reduce or eliminate disease. When lobsters are held at high d e n s i t i e s , r e s u l t i n g in s t r e s s f u l conditions, diseases that are v i r t u a l l y unknown i n natural environments may appear. A recent review of lobster microbial diseases (Fisher et a l . 1978) states that there are b a s i c a l l y six diseases that could create problems in holding systems. Included in the review are s h e l l disease, Gaffkemia, microbial epibiont disease, Lagenidium disease, Haliphthoros disease, and Fusarium disease. Gaffkemia and s h e l l disease appear to be the most common diseases af f e c t i n g lobsters held in a high density s i t u a t i o n . Gaffkemia i s a systemic disease and i s caused by a Gram- p o s i t i v e , tetrad forming bacterium known as Aerococcus viridans variety homari (Steward and Zwicker 1974). A. viridans i s a 9 n o r m a l component o f t h e f l o r a o f t h e e x o s k e l e t o n and p o s s e s s e s no m e c h a n i s m f o r i n v a s i o n o f t h e h o s t . However i t c a n be i n t r o d u c e d i n t o t h e h e m o c e l by c r a c k s o r p u n c t u r e s c a u s e d by t r a d i t i o n a l ' p e g g i n g ' o f t h e c r u s h e r c l a w o r b y d r y i n g o f t h e e x o s k e l e t o n i n s h i p m e n t ( S t e e n b e r g e n a n d S c h a p i r o 1 9 7 4 ) . S t e w a r t and R a b i n (1970) r e p o r t t h a t Homarus shows an a l m o s t c o m p l e t e l a c k o f d e f e n s e a g a i n s t g a f f k e m i a . S h e l l d i s e a s e on l o b s t e r s was f i r s t r e p o r t e d by H e s s ( 1 9 3 7 ) and was b e l i e v e d t o be c a u s e d by c h i t i n o l y t i c b a c t e r i a . R o s e n (1970) h a s shown t h a t b o t h b a c t e r i a and f u n g i a r e r e s p o n s i b l e f o r s h e l l d i s e a s e o n a v a r i e t y o f c r u s t a c e a n s . The g r o s s s i g n s o f t h e d i s e a s e a r e s i m i l a r i n a l l s p e c i e s ; t h e e x o s k e l e t o n i s p i t t e d a nd m a r r e d w i t h n e c r o t i c l e s i o n s a n d , a l t h o u g h t h e d i s e a s e i s n o t i m m e d i a t e l y f a t a l , d e a t h may o c c u r . The c h i t i n o l y t i c m i c r o o r g a n i s m s do n o t p e n e t r a t e i n t o t h e s o f t t i s s u e s u n d e r l y i n g t h e c h i t i n o u s e x o s k e l e t o n ( R o s e n 1 9 6 7 ; R o s e n 1 9 7 0 ) , b u t may p r o v i d e a p o r t a l o f e n t r y t o e p i d e r m a l t i s s u e s f o r s e c o n d a r y i n v a d e r s . S a w yer a n d T a y l o r ( 1 9 4 9 ) r e p o r t e d t h a t s h e l l d i s e a s e may a l s o c a u s e d i s c o l o r a t i o n , i r r i t a t i o n , a n d e r o s i o n o f l o b s t e r g i l l s r e s u l t i n g i n i m p a i r e d g a s e x c h a n g e . The d i s e a s e i s c o n t a g i o u s e s p e c i a l l y when t h e a n i m a l s a r e h e l d i n mass c o n f i n e m e n t , b u t l o b s t e r s may o v e r c o m e m i n o r s h e l l d i s e a s e by s u c c e s s f u l l y m o l t i n g ( M c L e e s e 1 9 6 5 ) . I t i s c l e a r t h e n t h a t c o n t r o l o f m i c r o o r g a n i s m s i s e s s e n t i a l i n a l o b s t e r h o l d i n g o p e r a t i o n t o r e d u c e m o r t a l i t y 10 and preserve lobster health and physical appearance. This control can be achieved through a number of techniques includ- ing the exposure of holding water to u l t r a v i o l e t l i g h t at the appropriate wavelength and in t e n s i t y (Ayres 1978) or through ozone s t e r i l i z a t i o n (Wheaton 1977). Therefore the evaluation of any lobster holding operation must include some sort of analysis of the system's a b i l i t y to control p o t e n t i a l l y l e t h a l microorganisms. Along with the aforementioned b i o l o g i c a l and behavioral demands of stored lobsters the l i t e r a t u r e l i s t s a number of physical and chemical parameters or c h a r a c t e r i s t i c s that must be considered i n a evaluation of t h i s s o r t . The most appropriate parameters are temperature, pH, s a l i n i t y , dissolved oxygen, ammonia, n i t r i t e and n i t r a t e (Hand 1977; Ayres and Wood 1977). Suspended and dissolved s o l i d s , and organic carbon are also considered important parameters i n various holding and culture systems. Therefore these water q u a l i t y c h a r a c t e r i s t i c s w i l l be examined with respect to lobster holding. 3.3 Temperature As with most other poikilotherms, temperatures that lobsters can tolerate depend to a considerable extent on the temperature to which they were acclimated. In general, market size lobsters can withstand a wide range of water temperatures, providng that fluctuations are gradual. They can survive i n water cooled to the freezing point or heated to 32°C (Cobb 11 1 9 7 6 ) . L e t h a l w a t e r t e m p e r a t u r e l e v e l s a r e n o t d e t e r m i n e d by t h e s i z e o f t h e l o b s t e r , a n d a r e n o t a f f e c t e d by a two-month p e r i o d o f s t a r v a t i o n , a n i m p o r t a n t f a c t o r i n l i v e l o b s t e r s t o r a g e ( M c L e e s e 1 9 5 6 ) . When o t h e r w a t e r q u a l i t y p a r a m e t e r s s u c h a s d i s s o l v e d o x y g e n o r s a l i n i t y a r e l e s s t h a n o p t i m a l , t o l e r a n c e t o t e m p e r a t u r e i s r e d u c e d . D u r i n g s t o r a g e t h e w a t e r t e m p e r a t u r e s h o u l d be m a i n t a i n e d b e t w e e n 4.5° and 10°C ( A y r e s a n d Wood 1 9 7 7 ; C o r n i c k a n d S t e w a r t 1 9 7 7 ) . S i n c e t h e l o b s t e r i s a c o l d - b l o o d e d a n i m a l i t r e s p o n d s t o r e d u c e d e n v i r o n m e n t a l t e m p e r a t u r e by l o w e r i n g i t s m e t a b o l i s m a n d t h u s i t s a c t i v i t y , b o t h r e s p o n s e s a r e d e s i r a b l e i n a h o l d i n g t a n k ( M c L e e s e a n d W i l d e r 1 9 5 8 ) . A f u r t h e r a d v a n t a g e i n m a i n t a i n i n g l o w w a t e r t e m p e r a t u r e s i n a l o b s t e r h o l d i n g s y s t e m i s t h a t t h e p r e v i o u s l y m e n t i o n e d s y s t e m i c d i s e a s e g a f f k e m i a i s t e m p e r a t u r e d e p e n d e n t , t h e l o w e r t h e t e m p e r a t u r e t h e s l o w e r t h e d i s e a s e p r o g r e s s e s ( S t e w a r t e t a l . 1 9 6 9 ) . 3.4 pH The pH, o r h y d r o g e n i o n c o n c e n t r a t i o n o f a s o l u t i o n , i s d e f i n e d by t h e e q u a t i o n : pH = - l o g 1 Q [ H + ] (1) wh e r e [ H + ] = t h e h y d r o g e n i o n c o n c e n t r a t i o n (Wheaton 1 9 7 7 ) . S i n c e many d i s s o c i a t i o n c o n s t a n t s f o r t h e c h e m i c a l r e a c t i o n s o c c u r i n g i n a q u e o u s s o l u t i o n s a r e pH d e p e n d e n t , t h e c h e m i c a l e n v i r o n m e n t f o r a q u a t i c o r g a n i s m s i s s t r o n g l y i n f l u e n c e d by t h i s p a r a m e t e r . The e q u i l i b r i u m b e t w e e n ammonia (^3) a n ( ^ 12 ammonium (NH^) i n water i s a notable example of t h i s pH e f f e c t . There are a number of b i o l o g i c a l l y mediated reactions a f f e c t i n g pH i n natural water systems, two are s i g n i f i c a n t in sea water holding systems. The f i r s t process i s plant and animal r e s p i r a t i o n which tends to decrease pH by the production of free carbon dioxide (C0 2). The basic reaction i s as follows: C6 H12°6 + 6 0 2 — ^ 6 C 0 2 + 6H20 (2) The carbon dioxide then combines with water to produce carbonic acid (H 2C0 3) which results i n a pH reduction. The second process i s n i t r i f i c a t i o n , which again tends to reduce pH t h i s time through the direc t production of hydrogen ions. Equation 3 shows the o v e r a l l reaction. NH* + 2 0 2 — • N O " + H20 + 2H + (3) According to Spotte (1970) the acceptable pH range for marine culture and holding water i s 7.5-8.3. Although there seems to be general acceptance of t h i s range i n the l i t e r a t u r e , the ef f e c t of low pH on animals i s poorly understood and methods employed to deal with incorrect pH le v e l s seem to vary. Hirayama (1970) reports that when calcareous gravels are r e l i e d on exclusively to buffer water i n a closed sea water system, the water eventually equilibrates at about pH 7.5 with an a l k a l i n i t y of 1.0 m e q . l - 1 . Both values are s l i g h t l y lower than the reported optimum. P a r t i a l water changes at a rate of 10% every two weeks, and regular addition of sodium carbonate or sodium bicarbonate are usually necessary to keep the pH and 13 a l k a l i n i t y v a l u e s w i t h i n t h e a c c e p t a b l e r a n g e ( S p o t t e 1 9 7 0 ) . 3.5 S a l i n i t y S a l i n i t y c a n be d e f i n e d a s t h e t o t a l amount o f s o l i d m a t e r i a l , i n g r a m s , c o n t a i n e d i n one k i l o g r a m o f s e a w a t e r when a l l t h e c a r b o n a t e h a s b e e n c o n v e r t e d t o o x i d e , t h e b r o m i n e and i o d i n e r e p l a c e d by c h o l o r i n e , and a l l o r g a n i c m a t t e r c o m p l e t e l y o x i d i z e d ( T s u r i k o v a a n d T s u r i k o v 1 9 7 1 ) . The s a l i n i t y o f s e a - w a t e r g e n e r a l l y v a r i e s f r o m 33 t o 37 p a r t s p e r t h o u s a n d ( p p t ) . H owever, i n t i d a l e s t u a r i e s , s a l i n i t i e s a r e g e n e r a l l y l o w e r and s u b j e c t t o c o n s i d e r a b l e v a r i a t i o n . The most c r i t i c a l a s p e c t o f s a l i n i t y i n a l i v e l o b s t e r h o l d i n g s y s t e m i s i t s i n f l u e n c e on o s m o r e g u l a t i o n . L o b s t e r s e x h i b i t l i m i t e d o s m o r e g u l a t o r y a b i l i t y when t h e o s m o t i c c o n c e n t r a t i o n o f t h e s u r r o u n d i n g w a t e r i s e i t h e r a b o v e o r b e l o w t h a t o f t h e i r b l o o d . E x c e s s s a l t i s e x c r e t e d by t h e g u t , w h i l e e x c e s s w a t e r i s e x c r e t e d by t h e a n t e n n a l g l a n d ( D a l l 1 9 7 0 ) . L o b s t e r s a r e t y p i c a l l y c o a s t a l a n i m a l s f o u n d i n w a t e r s h a v i n g a s a l i n i t y o f 33 p p t o r more. A l t h o u g h t h e y c a n be a c c l i m a t e d t o t o l e r a t e l o w s a l i n i t i e s t h e y a r e u s u a l l y n o t f o u n d n a t u r a l l y i n b r a c k i s h w a t e r . I t i s p o s s i b l e t o s t o r e l o b s t e r s i n w a t e r h a v i n g a s a l i n i t y down t o 20 p p t and l e s s when w a t e r t e m p e r a t u r e s a r e b e l o w 10°C, b u t t h e minimum v a l u e u s u a l l y c o n s i d e r e d a c c e p t a b l e i n c o m m e r c i a l s t o r a g e u n i t s i s 27 p p t (Wood and A y r e s 1 9 7 7 ) . S a l i n i t y o f a b o u t 8 p p t i s t h e l o w e r l i m i t o f t o l e r a n c e f o r j u v e n i l e and a d u l t Homarus 14 ( P h i l l i p s e t a l . 1980). 3. 6 D i s s o l v e d Oxygen The e x t r a c t i o n o f oxygen from water and t h e a d d i t i o n of t h i s gas t o water a r e o p e r a t i o n s of c r i t i c a l i m p o r t a n c e t o a q u a t i c a n i m a l s . In a c o l d - b l o o d e d a n i m a l such as t h e l o b s t e r , oxygen demand r i s e s w i t h i n c r e a s i n g e n v i r o n m e n t a l t e m p e r a t u r e , however, t h e s o l u b i l i t y of oxygen i n sea water i s reduced as the t e m p e r a t u r e r i s e s . The s y n e r g i s t i c e f f e c t produced by the c o m b i n a t i o n of t h e s e two p r i n c i p l e s i s t h a t t h e amount of s e a - water l o b s t e r s r e q u i r e t o s a t i s f y t h e i r oxygen r e q u i r e m e n t s d u r i n g s t o r a g e i n c r e a s e s d i s p r o p o r t i o n a t e l y as the t e m p e r a t u r e r i s e s ( A y r e s and Wood 1977). I n c r e a s e d s a l i n i t y a l s o has the a f f e c t o f d e c r e a s i n g t h e oxygen c a r r y i n g c a p a c i t y o f w a t e r . In s t u d i e s o f oxygen consumption and t o l e r a n c e i t has been shown t h a t when o t h e r p a r a m e t e r s a r e o p t i m a l , l o b s t e r s can s u r v i v e oxygen l e v e l s as low as about 1 mg oxygen per l i t e r (McLeese 1956). The amount of oxygen consumed per gram of body weight d e c r e a s e s w i t h i n c r e a s i n g body w e i g h t , and i n c r e a s e s w i t h i n c r e a s i n g water t e m p e r a t u r e (McLeese 1964). Under normal c o n d i t i o n s , c o a s t a l w a t e r s c o n t a i n from 7-13 ppm of d i s s o l v e d oxygen depending on a number of f a c t o r s i n c l u d i n g t e m p e r a t u r e , s a l i n i t y and b i o c h e m i c a l oxygen demand. I n l o b s t e r s t o r a g e systems oxygen r e q u i r e m e n t s depend on s t o r a g e d e n s i t y , a n i m a l a c t i v i t y and s t r e s s f a c t o r s . The g e n e r a l l y a c c e p t e d d i s s o l v e d oxygen l e v e l i s 7 or 9 ppm but t h e c l o s e r t o s a t u r a t i o n l e v e l s 15 t h e b e t t e r . The t o l e r a n c e o f l o b s t e r s t o o x y g e n , s a l i n i t y , a n d t e m p e r a t u r e i s shown i n t h e t h r e e - d i m e n s i o n a l g r a p h i n F i g u r e 1 ( a n y p o i n t i n t h e g r a p h r e p r e s e n t s a s p e c i f i c c o m b i n a t i o n o f o x y g e n t e m p e r a t u r e and s a l i n i t y ) . 3.7 S o l i d s a nd O r g a n i c C a r b o n The g e n e r a l t e r m " s o l i d s " u s u a l l y i s t a k e n t o mean t h e t o t a l s o l i d s o r r e s i d u e c o n t e n t o f t h e h o l d i n g w a t e r . I n most i n s t a n c e s , h o w e v e r , t h e s p e c i f i c f o r m i n w h i c h t h e s o l i d s a r e p r e s e n t i n t h e w a t e r must be d e t e r m i n e d . T h e r e f o r e , m e t h o d s h a v e b e e n d e v e l o p e d t o d i f f e r e n t i a t e b e t w e e n s e t t l e a b l e s o l i d s , s u s p e n d e d s o l i d s , d i s s o l v e d s o l i d s a nd v o l a t i l e s o l i d s ( A m e r i c a n P u b l i c H e a l t h A s s o c i a t i o n e t a l . 1 9 7 6 ) . I n a l o b s t e r h o l d i n g s y s t e m most o f t h e l a r g e p a r t i c u l a t e m a t t e r o r s e t t l e a b l e f r a c t i o n s h o u l d be r e m o v e d by m e c h a n i c a l f i l t r a t i o n , a n d a s s u m i n g t h a t t h e a v a i l a b l e h o l d i n g w a t e r s u p p l y i s o f g o o d q u a l i t y , t o t a l d i s s o l v e d s o l i d s s h o u l d n o t be a s e r i o u s p r o b l e m . The m a j o r c o n c e r n l i e s i n t h e s m a l l s u s p e n d e d and v o l a t i l e o r g a n i c f r a c t i o n s . T h e s e f r a c t i o n s may h a v e t h e f o l l o w i n g e f f e c t s i n a l o b s t e r h o l d i n g s y s t e m : 1. i r r i t a t i o n a n d / o r damage t o l o b s t e r g i l l s ( b e c o m i n g more s e v e r e i n t h e p r e s e n c e o f t o x i c s u b s t a n c e s ) , 2. e x e r t i o n o f b i o c h e m i c a l o x y g e n demand a n d c r e a t i o n o f n o x i o u s c o n d i t i o n s , 3. t r a n s p o r t o f a d s o r b e d p o l l u t a n t s , 16 Figure 1. Three dimensional diagram of the boundary of lethal conditions for lobsters under various combinations of temperature, salinity and oxygen. T - region in which temperature alone acts as a lethal factor. S - region in which salinity alone acts as a lethal factor. 0 - region in which oxygen alone acts as a lethal factor. [redrawn from McLeese(1956)] 17 4. i n c r e a s e d s y s t e m m a i n t e n a n c e . The f i r s t two e f f e c t s a r e t h e most c r i t i c a l . The m e asurement o f b i o c h e m i c a l o x y g e n demand (BOD), w h i c h d e f i n e s t h e b i o d e g r a d a b l e o r g a n i c c o n t e n t o f a w a s t e w a t e r , and c h e m i c a l o x y g e n demand, w h i c h m e a s u r e s t h e t o t a l c h e m i c a l l y o x i d i z a b l e o r g a n i c c o n t e n t , b o t h d e g r a d a b l e a n d r e f r a c t o r y , i s d i f f i c u l t when u s i n g l o w c o n c e n t r a t i o n w a s t e w a t e r s u c h a s w a t e r f r o m a l o b s t e r h o l d i n g s y t e m . BOD d e t e r m i n a t i o n s a r e p a r t i c u - l a r l y d i f f i c u l t t o o b t a i n when u s i n g a s a l t w a t e r medium ( I v e r s o n 1973) . S u s p e n d e d s o l i d s m e a s u r e m e n t s a r e c o m p l i c a t e d by f l u c - t u a t i n g s a l i n i t y l e v e l s i n t h e h o l d i n g w a t e r . S i n c e m e t a b o l i s m i s l o w and f e e d i n g i s n o t t a k i n g p l a c e , a v e r y s m a l l amount o f s o l i d m a t t e r w o u l d be e x p e c t e d t o be g e n e r a t e d i n a l o b s t e r h o l d i n g u n i t . As a r e s u l t , v e r y l i t t l e e m p h a s i s h a s b e e n p l a c e d on m e a s u r i n g t h i s p a r a m e t e r i n e s t a b - l i s h e d s y s t e m s . D a t a on l o b s t e r t o l e r a n c e l e v e l s t o s u s p e n d e d s o l i d s i n r e c i r c u l a t i n g h o l d i n g o p e r a t i o n s a r e u n a v a i l a b l e a t p r e s e n t . The N a t i o n a l Academy o f S c i e n c e s a n d The N a t i o n a l Academy o f E n g i n e e r i n g ( 1 9 7 4) s u g g e s t t h a t a q u a t i c c o m m u n i t i e s s o u l d be p r o v i d e d w i t h a h i g h l e v e l o f p r o t e c t i o n a t s u s p e n d e d s o l i d s c o n c e n t r a t i o n s o f g r e a t e r t h a n 25 mg«l ^. I n a summary o f w a t e r q u a l i t y c r i t e r i a f o r s a l m o n i d h a t c h e r i e s t h e C a n a d i a n D e p a r t m e n t o f F i s h e r i e s a n d O c e a n s ( 1 9 7 9 ) s t a t e s t h a t t h e maximum c o n c e n t r a t i o n o f s u s p e n d e d s o l i d s i n a s a l m o n i d h o l d i n g p o n d s h o u l d n o t e x c e e d 25 m g - l - 1 . T h i s f i g u r e i s l i k e l y more t h a n a d e q u a t e f o r l o b s t e r s t o r a g e a s t h e l o b s t e r i s 18 g e n e r a l l y more t o l e r a n t o f poor water q u a l i t y t h a n most s a l m o n i d s p e c i e s . S i n c e BOD, COD and suspended s o l i d s d e t e r m i n a t i o n s a r e c o m p l i c a t e d by v a r i o u s c o n d i t i o n s i n h e r e n t i n a l o b s t e r h o l d i n g system, t o t a l o r g a n i c c a r b o n (TOC) l e v e l s can be c o n s i d e r e d as a s u b s t i t u t e f o r t h e d e t e r m i n a t i o n o f suspended m a t t e r and b i o c h e m i c a l oxygen demand. R e l a t i o n s h i p s between TOC and COD i n v a r i o u s t y p e s of wastewaters have been deve l o p e d ( E c k e n f e l d e r 1970) and t h e s e r e l a t i o n s h i p s are d i s c u s s e d i n more d e t a i l i n a l a t e r s e c t i o n . 3.8 N i t r o g e n o u s Compounds The f i n a l a r e a of e v a l u a t i o n i n t h i s s t u d y i s t h a t of n i t r o g e n o u s compounds i n l o b s t e r h o l d i n g w a t e r . The t h r e e major e n d - p r o d u c t s of n i t r o g e n m e t a b o l i s m i n a n i m a l s a r e ammonia, u s u a l l y as t h e ammonium i o n (NH^), u r e a , and u r i c a c i d (Campbell 1973). The major n i t r o g e n o u s end-product of marine i n v e r t e b r a t e s , i n c l u d i n g Homarus, i s ammonia, making t h e s e a n i m a l s a m m o n i o t e l i c . The term ammonia u s u a l l y r e f e r s t o t h e sum of ammonium i o n and f r e e ammonia (NH-j), d e t e r m i n e d a n a l y t i c a l l y as t o t a l NH^-N. As w e l l as d i r e c t e x c r e t i o n of ammonia, t h e l o b s t e r does produce minor amounts of u r e a , u r i c a c i d , and amino a c i d s which a l l e v e n t u a l l y undergo m i n e r a l i z a t i o n or d e a m i n a t i o n t o form ammonia ( S p o t t e 1979). N i t r o g e n o u s wastes a r e e x c r e t e d by t h e g i l l s , t h e g u t , and, t o a l e s s e r e x t e n t , by t h e a n t e n n a l g l a n d s i n t h e a n t e r i o r v e n t r a l 19 p a r t of the body t h r o u g h a pore on t h e lower s i d e of the b a s a l segment of t h e f i r s t antennae. When u r i n e a l o n e was examined from t h e s p i n y l o b s t e r ( J a s u s e d w a r d s i i ) , u r e a , ammonia and amino compounds t o g e t h e r made up o n l y 21% o f t h e t o t a l u r i n e n i t r o g e n , t h e remainder b e i n g u n i d e n t i f i e d ( B i n n s and P e t e r s o n 1969). The r a t e of u r i n e o u t p u t has been e s t i m a t e d a t a p p r o x i m a t e l y 0.5% of t h e body weight per day f o r Homarus (Burger 1957). In a c u l t u r e or n a t u r a l s i t u a t i o n , where t h e l o b s t e r s a r e consuming f o o d , ammonia p r o d u c t i o n i s c l o s e l y r e l a t e d t o the f e e d i n g r a t e , d i e t a r y p r o t e i n - n i t r o g e n l e v e l , p r o t e i n u t i l i z - a t i o n o r c o n v e r s i o n , and n i t r o g e n e x c r e t e d as ammonia or compounds r e a d i l y c o n v e r t e d t o ammonia. A t h e o r e t i c a l t o t a l ammonia-ammonium p r o d u c t i o n r a t e can be d e t e r m i n e d from t h e q u a n t i t y o f f o o d s u p p l i e d and t h e p r o t e i n c o n t e n t of t h e f e e d : N = (P/6.25) Q f (4) where N i s t h e r a t e of i n t a k e of n i t r o g e n , i n g-day-"*", P i s th e p r o p o r t i o n o f p r o t e i n i n t h e r a t i o n , and i s t h e q u a n t i t y o f f e e d s u p p l i e d i n g*day~"*" ( A l l e n and J o h n s t o n 1976). P r o d u c t i o n o f ammonia gas, t h e more t o x i c m e t a b o l i t e , i s c a l c u l a t e d w i t h t h e t h e o r e t i c a l l y a v a i l a b l e n i t r o g e n e s t i m a t e from E q u a t i o n 4. The p r o p o r t i o n o f t o t a l ammonium d i s s o c i a t e d as gas c a n be d e r i v e d from t h e Henderson-Hasselbach e q u a t i o n ( A l l e n and J o h n s t o n 1976): PH - P K a + . - 5 * r - FKa + l o g -ffffff ; (5) 20 where C i s the molar concentrat ion of ac id or base and X i s the amount of strong base that must be added to make the so lu t ion e l e c t r i c a l l y n e u t r a l . No e m p i r i c a l l y determined nitrogen conversion factor i s presented by A l l e n and Johnston, therefore , i t i s impossible to ca lcu la te actual ammonia- ammonium product ion. Colt and Armstrong (1981) state that the prote in conversion factor for aquatic animals ranges from 0.65 to 0.80. The est imation of ammonia production by f i s h has received far more a t tent ion due to the importance of th i s para- meter in hatchery and growout f a c i l i t i e s . As a r e s u l t , the var iab les involved in f i s h cu l ture have been defined in a much c learer fashion (Pettigrew et a l . 1978). When lobsters are being held and not provided with food, i t becomes very d i f f i c u l t to estimate ammonia product ion. Compared to a lobster growout operat ion , where feeding i s tak- ing place and metabolism i s h igh, the concentration of ammonia i n a system designed s t r i c t l y for lobster storage should be qui te low, assuming both systems u t i l i z e s i m i l a r water recyc le pat terns . Even assuming a low rate of ammonia production i n a lobster holding u n i t , tox ic l eve l s of t h i s compound can develop when the holding water i s continuously recycled without t r e a t - ment or d i l u t i o n . The hydro lys i s of ammonia i n natura l waters, as shown by the react ion below, has a pK value of about 9.0, so that the percentage of ammonium i s always greater than that of free ammonia (Spotte 1970). NH* + H 20 ^ w NH 3 + H 3 0 + (6) 21 F a c t o r s a f f e c t i n g ammonia h y d r o l y s i s a r e o f g r e a t i m p o r t a n c e i n a q u a t i c c u l t u r e a n d h o l d i n g s y s t e m s , s i n c e i t h a s b e e n shown t h a t i n most i n s t a n c e s N H 3 i s s i g n i f i c a n t l y more t o x i c t h a n NH^. E q u a t i o n 6 i s c o n t r o l l e d p r e d o m i n a t e l y by pH and t o a l e s s e r e x t e n t by t e m p e r a t u r e a n d s a l i n i t y . A pH i n c r e a s e o f one u n i t c a u s e s t h e p e r c e n t a g e o f f r e e ammonia t o i n c r e a s e a b o u t t e n f o l d ( T r u s s e l 1 9 7 2 ; Bower a n d B i d w e l l 1 9 7 8 ) . The t e m p e r a t u r e e f f e c t i s t h e r e s u l t o f i n c r e a s e d h y d r o l y s i s o f ammonium i o n s a t h i g h e r t e m p e r a t u r e l e v e l s ; t h e s a l i n i t y e f f e c t i s t h e r e s u l t o f t h e d e c r e a s i n g a c t i v i t y o f f r e e ammonia i n s o l u t i o n s o f i n c r e a s i n g i o n i c s t r e n g t h (Hampson 1 9 7 6 ) . T h r o u g h t h e b i o l o g i c a l o x i d a t i o n p r o c e s s known a s n i t r i f i - c a t i o n , ammonia i s c o n v e r t e d i n t o two o t h e r t o x i c n i t r o g e n b a s e d compounds, n i t r i t e a n d n i t r a t e . N i t r i f i c a t i o n i s c a r r i e d o u t m a i n l y b y two a u t o t r o p h i c b a c t e r i a , N i t r o s o m o n a s a n d N i t r o b a c t e r . The s t o i c h i o m e t r i c r e a c t i o n f o r o x i d a t i o n o f ammonium t o n i t r i t e by N i t r o s o m o n a s i s : N H + + 1.5 0 2 — • 2 H + + H 20 + N 0 " 2 (7) The r e a c t i o n f o r o x i d a t i o n o f n i t r i t e t o n i t r a t e by N i t r o b a c t e r i s : NO" + 0.5 0 2 — • N 0 3 (8) N i t r i t e i s t h e i o n i z e d f o r m o f n i t r o u s a c i d , a weak a c i d . T h i s r e a c t i o n c a n be w r i t t e n : N0~ + H + — • HN0 2 (9) At h o l d i n g w a t e r pH and t e m p e r a t u r e s t h e p e r c e n t a g e HN0 ?-N 22 r a n g e s f r o m .0005 t o .05 p e r c e n t ( R u s s o e t a l . 1 9 8 1 ) . Low pH and t e m p e r a t u r e s f a v o r t h e n i t r o u s a c i d f o r m . N i t r a t e s a r e i n g e n e r a l r e a d i l y s o l u b l e i n w a t e r , s h o w i n g s l i g h t t e n d a n c i e s t o f o r m c o o r d i n a t i o n compounds, and f o r most p u r p o s e s c a n be c o n s i d e r e d t o be d i s s o c i a t e d c o m p l e t e l y ( L a t i m e r and H i l d e b r a n d 1951) . The m e c h a n i s m o f ammonia t o x i c i t y i n l o b s t e r s i s a s y e t n o t f u l l y u n d e r s t o o d b u t a s i n f i s h i t may be by 'mass l a w ' p r e v e n t i o n o r r e v e r s a l o f n o r m a l n i t r o g e n m e t a b o l i s m . NH^ i s r e g a r d e d a s t h e f o r m o f ammonia t h a t i s t o x i c t o f r e s h w a t e r f i s h e s b e c a u s e i t i s l i p i d s o l u b l e , w h e r e a s t h e p e r m e a b i l i t y o f p l a s m a membranes t o NH^, h y d r a t e d ammonia i o n s , i s r e l a t i v e l y l o w ( M i l n e e t a l . 1 9 5 8 ) . C o l t a n d A r m s t r o n g ( 1 9 8 1 ) c a t e g o r i z e t h e t o x i c e f f e c t s o f ammonia t o a q u a t i c o r g a n i s m s u n d e r e i g h t g e n e r a l h e a d i n g s - e f f e c t s on a c e l l u l a r l e v e l , e f f e c t s on n i t r o g e n e x c r e t i o n , e f f e c t s on o s m o r e g u l a t i o n , e f f e c t s on o x y g e n t r a n s p o r t , e f f e c t s on t i s s u e , e f f e c t s on d i s e a s e , l e t h a l e f f e c t s and e f f e c t s on g r o w t h . On t h e c e l l u l a r l e v e l , t h e r e l e a s e o f NH^ i n t o t h e b l o o d f r o m a m b i e n t w a t e r o r m e t a b o l i c p r o d u c t i o n i s c o n v e r t e d t o NH^ w i t h t h e r e l e a s e o f an OH~. The s u b s e q u e n t e l e v a t i o n o f b l o o d a n d p e r h a p s i n t e r c e l l u l a r pH c a n h a v e a p r o n o u n c e d e f f e c t on e n z y m e - c a t a l y z e d r e a c t i o n s a nd membrane s t a b i l i t y ( C a m p b e l l 1 9 7 3 ) . C a m p b e l l a l s o s t a t e s t h a t h i g h l e v e l s o f ammonia may c a u s e a r e v e r s a l o f t h e g l u t a m a t e 23 dehydrogenase r e a c t i o n , w i t h d r a w i n g a l p h a - k e t o g l u t a r a t e from t h e t r i c a r b o x y l i c a c i d c y c l e as w e l l as d e c r e a s i n g t h e amount of NADH a v a i l a b l e f o r o x i d a t i o n . Three p r i n c i p a l r o u t e s o f m e t a b o l i c ammonia e x c r e t i o n a re a v a i l a b l e t o a q u a t i c a n i m a l s : 1) d i f f u s i o n of NH^ from the b l o o d t o the water t h r o u g h t h e g i l l s , 2) exchange t r a n s p o r t of NH* w i t h N a + , and 3) c o n v e r s i o n t o n o n - t o x i c compound l i k e u r e a . As p r e v i o u s l y s t a t e d , t h e most i m p o r t a n t r o u t e u t i l i z e d by l o b s t e r s i s d i f f u s i o n a c r o s s t h e g i l l s . Hampson (1976) s u g g e s t s t h a t h i g h ammonia l e v e l s i n t h e b l o o d i n h i b i t ammonia e x c r e t i o n . I t has been demonstrated f o r a number of a n i m a l s i n c l u d i n g rainbow t r o u t , (Salmo g a i r d n e r i ( O l s o n and Fromm 1 9 7 1 ) ; g o l d f i s h , C a r a s s i u s a u r a t u s ( O l s o n and Fromm 1 9 7 1 ) ; c r a b , C a l l i n e c t e s s a p i d u s (Mangum e t a l . 1 9 7 6 ) ; and t h e f r e s h w a t e r s h r i m p , Macrobrachium r o s e n b e r g i i (Armstrong 1 9 7 8 ) , t h a t t h e a d d i t i o n o f ammonia t o t h e e x t e r n a l medium w i l l reduce ammonia e x c r e t i o n . T h i s e f f e c t i s p a r t i c u a r l y i m p o r t a n t i n an a q u a c u l t u r e system because t h e i n i t i a l r e a c t i o n of a q u a t i c a n i m a l s t o t h i s i n h i b i t i o n of ammonia e x c r e t i o n may be the c e s s a t i o n of f e e d i n g which i n t u r n may reduce growth r a t e s . I t i s a l s o q u i t e l i k e l y t h a t s u b l e t h a l c o n c e n t r a t i o n s of ammonia may produce s e v e r e h y p e r p l a s i a o f t h e g i l l e p i t h e l i u m and an i n c r e a s e d p e r m a b i l i t y o f t h e a n i m a l t o water. T h i s e f f e c t on o s m o r e g u l a t i o n was o b s e r v e d i n f i s h by L l o y d and O r r ( 1 9 6 9 ) . Ammonia can have a s e r i o u s , e f f e c t on t h e a b i l i t y of 24 a q u a t i c s p e c i e s t o t r a n s p o r t oxygen to the t i s s u e s . These e f f e c t s i n c l u d e damage to the g i l l s , r e d u c t i o n of the b l o o d ' s c a p a c i t y to c a r r y oxygen due to a lowered pH, an i n c r e a s e d oxygen demand, and h i s t o l o g i c a l damage to the b l o o d c e l l s and the c e l l s p r o d u c i n g t i s s u e s (Smart 1978). S u b l e t h a l and l e t h a l ammonia l e v e l s can cause h i s t o l o g i c a l changes i n the k i d n e y s , l i v e r , s p l e e n , t h y r o i d t i s s u e , and b l o o d parameters of many f i s h s p e c i e s (Smith and P i p e r 1975) . E f f e c t s on d i s e a s e a r e d i f f i c u l t t o a s se s s but i t i s r e a s o n a b l e to expect t h a t an a q u a t i c an imal w i l l be more s u s c e p t i b l e to d i s e a s e i f s u f f e r i n g the a d d i t i o n a l s t r e s s a s s o c i a t e d w i t h h i g h ambient ammonia c o n c e n t r a t i o n s . The 96-h LC50 v a l u e o f u n - i o n i z e d ammonia ranges from 0.4 to 3.1 m g ' l - 1 f o r f i s h ( C o l t and Tchobanoglous 1976; B a l l 1967) , 0.40 to 2 .31 m g « l ^ f o r c r u s t a c e a n s (Armstrong et a l . 1978 ; D e l i s t r a t y e t a l . 1977 ; W i c k i n s 1976), and 3 .3 to 6.0 m g - l - ^ f o r marine m o l l u s c s ( E p i f a n i o and Srna 1975) . E f f e c t s of ammonia on growth of a q u a t i c an imal s i s d i f f i c u l t t o a s se s s q u a n t i t a t i v e l y and i s o f minor c o n c e r n i n a l o b s t e r h o l d i n g f a c i l i t y . N i t r i t e t o x i c i t y i n f i s h i s due i n p a r t to the o x i d a t i o n o f hemoglobin to f e r r i h e m o g l o b i n or methemoglobin (MHb) by n i t r i t e (Krous e t a l . 1982). Methemoglobin does not have the c a p a c i t y to c a r r y oxygen; i f s u f f i e n t methemoglobin i s formed h y p o x i a and c y a n o s i s may r e s u l t ( K i e s e 1974) . The dominant r e s p i r a t o r y t r a n s p o r t pigment i n m a l a c o s t r a c a n c r u s t a c e a n s , 25 s u c h a s Homarus, i s h e m o c y a n i n ( P i c k e t e t a l . 1 9 6 6 ) . I t i s l i k e l y t h a t t h e same o x i d a t i o n r e a c t i o n c a n o c c u r w i t h t h e c o p p e r o f c r u s t a c e a n h e m o c y a n i n b u t s u c h c o n v e r s i o n h a s n o t a s y e t b e e n r e p o r t e d f o r t h i s a n i m a l c l a s s . A l t h o u g h o n l y s c a n t y i n f o r m a t i o n e x i s t s r e l a t i n g t o t h e t o x i c i t y o f n i t r i t e and n i t r a t e t o l o b s t e r s , Hand ( 1 9 7 7 ) r e p o r t s t h a t Homarus e x h i b i t s a much g r e a t e r t o l e r a n c e t o b o t h i n o r g a n i c n i t r o g e n s p e c i e s t h a n most o t h e r a q u a t i c o r g a n i s m s , most n o t a b l y f r e s h w a t e r f i s h . T h i s s i t u a t i o n i s n o t s u r p r i s i n g a s i t h a s b e e n o b s e r v e d t h a t n i t r i t e i s s i g n i f i c a n t l y l e s s t o x i c t o f i s h i n a s e a w a t e r e n v i r o n m e n t ( C r a w f o r d and A l l e n 1 9 7 7 ) . V a r i o u s c h e m i c a l c o n s t i t u e n t s a n d p r o c e s s e s a s s o c i a t e d w i t h s e a w a t e r may be i n v o l v e d i n t h i s a p p a r e n t i n c r e a s e d t o l e r a n c e . P e r r o n e and Meade ( 1 9 7 7 ) s u g g e s t e d t h a t t h e p r o t e c t i v e e f f e c t o f e n v i r o n - m e n t a l i o n s may be t h e r e s u l t o f a c o m p e t i t i v e i n h i b i t i o n o f n i t r i t e u p t a k e t h r o u g h t h e g i l l s a n d o t h e r i n t e g u m e n t o r y t i s s u e s , a n d / o r an i n c r e a s e i n p l a s m a and b ody f l u i d i o n l e v e l s . A number o f i o n s ( C a + + , C l ~ , N a + , and SO^) h a v e b e e n t e s t e d f o r t h e i r e f f e c t i v e n e s s i n r e d u c i n g n i t r i t e - i n d u c e d m o r t a l i t y a n d t h e f o r m a t i o n o f MHb, w i t h most f i n d i n g s s u p p o r t i n g t h e s u g g e s t i o n o f a c o m p e t i t i v e i n h i b i t i o n o f n i t r a t e u p t a k e a t t h e g i l l s ( C r a w f o r d a n d A l l e n 1 9 7 7 ; Wedermeyer and Y a s u t a k e 1978; Tomasso e t a l . 1 9 7 9 ) . I t i s l i k e l y t h a t a number o f o t h e r c o m p l e x f a c t o r s s u c h a s b e h a v i o u r and b i o - c h e m i c a l p r o c e s s e s a r e i n v o l v e d i n t h e c o m p a r a t i v e l y h i g h t o l e r a n c e t o n i t r i t e a n d n i t r a t e e x h i b i t e d by l o b s t e r s b u t no 26 l i t e r a t u r e i s a v a i l a b l e a t p r e s e n t r e l a t i n g t o t h e s e f a c t o r s . A l t h o u g h h i g h l e v e l s o f w a t e r q u a l i t y must be m a i n t a i n e d , due t o t h e p o t e n t i a l b u i l d - u p o f t o x i c m e t a b o l i c w a s t e s and b e c a u s e w a t e r d i s c h a r g e d f r o m a n y c o m m e r c i a l o p e r a t i o n must meet s t a n d a r d s s e t by e n v i r o n m e n t a l l a w s , l o b s t e r s c a n s u r v i v e i n w a t e r w i t h c o m p a r a t i v e l y h i g h l e v e l s o f n i t r o g e n o u s compounds (Cobb 1 9 7 6 ) . I n f a c t , t h e y c a n t o l e r a t e c o n c e n t r a - t i o n s o f ammonia, n i t r a t e a n d n i t r i t e , c o n s i d e r a b l y h i g h e r t h a n t h o s e a c c e p t a b l e t o many o t h e r a n i m a l s i n r e a r i n g o r h o l d i n g e n v i r o m e n t s . G r a v i t z e t a l . ( 1 9 7 5 ) r e p o r t s t h a t f o r 24 h o u r s , m ost j u v e n i l e l o b s t e r s o f one t o t h r e e grams w i l l s u r v i v e c o n c e n t r a t i o n s o f t o t a l ammonia-N up t o 25 mg«l \ i f o t h e r c o n d i t i o n s a r e o p t i m a l . However, l o n g e r e x p o s u r e t o ammonia a t l o w e r c o n c e n t r a t i o n s c a n r e s u l t i n d e a t h . V a l u e s o f 96h LC^g h a v e b e e n e s t i m a t e d a t 1.2 + 0.1 mg NH^-N • l - 1 f o r l g l o b s t e r s a n d 1.4 + 0.1 mg NH 3-N • l - 1 f o r 3g l o b s t e r s b u t no r e l i a b l e i n f o r m a t i o n on c o n c e n t r a t i o n s c a u s i n g c h r o n i c t o x i c i t y i s known t o be a v a i l a b l e ( A l l e n and J o h n s t o n 1 9 7 6 ) . S i m i l a r a s s a y s w i t h n i t r i t e a n d n i t r a t e showed t h e s e s p e c i e s o f n i t r o g e n t o be n o n t o x i c a t c o n c e n t r a t i o n s o f 100 and 500 m g « l _ 1 r e s p e c t i v e l y (Hand 1 9 7 7 ) . I n e a c h c a s e t h o u g h a n i m a l s t h a t s u r v i v e d t h e t o x i c i t y t e s t s a n d w e r e a l l o w e d t o r e c o v e r i n r u n n i n g a m b i e n t s e a w a t e r w e r e f o u n d t o be f a r more s e n s i t i v e u pon r e - e x p o s u r e t o t h e s e compounds. D e s p i t e t h e r e l a t i v e l y i n o c c u o u s n a t u r e o f NC^ and NO,, m o n i t o r i n g o f t h e s e compounds i s i m p o r t a n t i n t h e 27 e v a l u a t i o n of l o b s t e r h o l d i n g systems, s i n c e t h e y a c t as i n d i c a t o r s p e c i e s o f v a r i o u s c h e m i c a l p r o c e s s e s (such as n i t r i f i c a t i o n ) t h a t t a k e p l a c e i n a r e c i r c u l a t i n g seawater system. T a b l e 1 summarizes t h e r e s u l t s o f t h i s l i t e r a t u r e r e v i e w by l i s t i n g t h e a p p r o p r i a t e water q u a l i t y p a r a m e t e r s t o be m o n i t o r e d d u r i n g e v a l u a t i o n s o f l o b s t e r h o l d i n g systems and i n d i c a t e s an a c c e p t a b l e l e v e l f o r each parameter. A l s o l i s t e d a r e p o t e n t i a l l y l e t h a l v a l u e s and l e v e l s t o be e x p e c t e d under n a t u r a l c o n d i t i o n s . Table 1. Important water q u a l i t y parameters to be monitored in a l i v e lobster storage system. Values are l i s t e d for holding, natural, and p o t e n t i a l l y l e t h a l conditions.* P a r a m e t e r H o l d i n g C o n d i t i o n s N a t u r a l C o n d i t i o n s P o t e n t i a l l y L e t h a l T e m p e r a t u r e (°C) P H S a l i n i t y ( p p t ) D i s s o l v e d O x y g en ( m g » l - 1 ) S u s p e n d e d S o l i d s (mg«l~l) N i t r o g e n o u s Compounds ( m g - l ~ l ) 6-7 8.0-8.3 27 7-8 o r j u s t b e l o w s a t u r a t i o n 0-25 7.5-8.4 29-34 4-10 -2 and 32 5 and 9 8 and 45 2 and s u p e r s a t u r a t i o n 25 P a r a m e t e r N H 3-N** 1 s t l a r v a l s t a g e 4 t h l a r v a l s t a g e 1 s t j u v e n i l e 1 2 t h j u v e n i l e H o l d i n g C o n d i t i o n s N a t u r a l C o n d i t i o n s P o t e n t i a l l y L e t h a l t o 0.3 N 0 2 - N NO3-N j u v e n i l e j u v e n i l e 1.3 100, n o t l e t h a l a t 96 h r 50 0 , n o t l e t h a l a t 96 h r 0.014 0.07-21.0 1.3 3.8 9.4 94.0 p o s s i b l y 100 p o s s i b l y 500 • F i g u r e s q u o t e d a r e r e f e r e n c e d i n t h e t e x t . * * B a s e d on L C 5 0 / 24-48 h r 29 4.0 SYSTEM DESCRIPTION The g e n e r a l s y s t e m l a y o u t and w a t e r f l o w p a t t e r n a t P a c i f i c R i m S h e l l f i s h i s o u t l i n e d i n F i g u r e 2. The b a s i c c o m p o n e n t s o f t h e s y s t e m i n c l u d e 13 f i b e r g l a s s h o l d i n g t a n k s ( F i g u r e 3 ) , a w a t e r s t o r a g e r e s e r v o i r , a c e n t r i f u g a l i n t a k e pump and a c e n t r i f u g a l c i r c u l a t i o n pump, t h r e e s i l i c a - s a n d f i l t e r s ( F i g u r e 4 ) , t h r e e u l t r a v i o l e t (UV) s t e r i l i z e r u n i t s ( F i g u r e 5 ) , a n d a r e f r i g e r a t i o n u n i t ( F i g u r e 6 ) . B o t h t h e s i l i c a - s a n d f i l t e r s a n d t h e u l t r a v i o l e t s t e r i l i z e r s a r e a r r a n g e d i n a p a r a l l e l f a s h i o n s o t h a t e a c h u n i t r e c e i v e s o n e - t h i r d o f t h e w a t e r f l o w . S y s t e m c o m p o n e n t s and t h e i r s p e c i f i c a t i o n s a r e l i s t e d i n T a b l e 2. W a t e r f l o w t h r o u g h t h e v a r i o u s c o m p o n e n t s o r i g i n a t e s a t t h e r e s e r v o i r , w h i c h i s f e d o n a demand b a s i s by t h e i n t a k e pump. From t h e r e s e r v o i r , w a t e r f l o w s t h r o u g h t h e c i r c u l a t i o n pump a n d i n t o t h e s i l i c a - s a n d f i l t e r s w h e re much o f t h e p a r t i c u l a t e m a t t e r i s r e m o v e d . A f t e r f i l t r a t i o n t h e w a t e r i s e x p o s e d t o h i g h i n t e n s i t y u l t r a v i o l e t l i g h t f r o m t h e UV s t e r i l i z e r s t o d e s t r o y a n y m i c r o o r g a n i s m s p r e s e n t . W a t e r i s t h e n pumped i n t o t h e f i b e r g l a s s h o l d i n g t a n k s w h e r e i t e v e n t u a l l y f l o w s b y g r a v i t y b a c k i n t o t h e r e s e r v o i r . The r e f r i g e r a t i o n u n i t e x i s t s a s a s e l f - c o n t a i n e d c i r c u l a t i o n l o o p . W a t e r i s e x t r a c t e d f r o m t h e r e s e r v o i r , c h i l l e d a nd r e t u r n e d t o t h e r e s e r v o i r i n a c y c l i c f a s h i o n . A v a r i e t y o f v a l v e s and b y p a s s p o i n t s e x i s t i n t h e w a t e r t r e a t m e n t s y s t e m t o A |ptofc# PIMP t Samp C Clfwifltiofi PtMip D SondFlttat* t UVSMriftian »- » * - •wraw ram Figure 2. System components and water flow pattern of lobster holding unit. 31 Figure 3. One of the three sets of stacked fiberglass holding tanks used for lobster storage. Figure 4. One of three s i l i c a sand f i l t e r s used for treatment of the lobster holding water. 33 Figure 5. One of the three UV s t e r i l i z e r units showing c i r c u l a r arrangement of UV lamps. Figure 6 . Refrigeraton unit used to maintain lobster holding water temperature. 35 Table 2: Lobster holding system equipment and sp e c i f i c a t i o n s , Component i n t a k e pump c e n t r i f u g a l c i r c u l a t i o n pump s i l i c a s a n d f i l t e r s U l t r a v i o l e t s t e r i l i z e r s r e f r i g e r a t i o n u n i t f i b e r g l a s s h o l d i n g t a n k s r e s e r v o i r S p e c i f i c a t i o n s 0.47 I s - 1 5.6 kW m o t o r r a t e d a t 9.5 l - s - i - a c t u a l a p p r o x i m a t e o u t p u t 6.5 I s - 1 o p e r a t i n g p r e s s u r e , 345 kPa i n d i v i d u a l f i l t e r a r e a , 0.64 m 2 60,000 yuWs.cm - 1 a t 2537 A 560 W m o t o r , F r e o n - 1 2 , r a t e d a t 30160 W i n s i d e d i m e n s i o n s : 1.37x3.05x0.38 m a v e r a g e w a t e r d e p t h : 0.24 m a v e r a g e t a n k v o l u m e : 1003 1 i n s i d e d i m e n s i o n s : 0.97x3.35x2.64m a v e r a g e v o l u m e : 4500 1 a v e r a g e s y s t e m v o l u m e 17,539 1 36 f a c i l i t a t e f i l t e r b a c k w a s h a n d component m a i n t e n a n c e . The s y s t e m f u n c t i o n s m a i n l y o n a c o m p l e t e r e c y c l e b a s i s (one c y c l e a p p r o x i m a t e l y e v e r y 45 m i n u t e s ) w i t h t h e a d d i t i o n o f ' f r e s h ' s e a w a t e r c o m i n g on a demand b a s i s a nd d u r i n g f i l t e r b a c k w a s h o p e r a t i o n s . The f i l t e r s a r e b a c k w a s h e d u s u a l l y o n c e d a i l y r e s u l t i n g i n t h e a d d i t i o n o f a p p r o x i m a t e l y 10% o f t h e s y s t e m v o l u m e i n f r e s h s e a w a t e r . T h e r e f o r e a c o m p l e t e w a t e r c h a n g e o c c u r s e v e r y 10 d a y s . S i n c e t h e ' f r e s h ' s e a w a t e r o b t a i n e d f r o m t h e m a r i n e i n l e t a d j a c e n t t o t h e h o l d i n g o p e r a - t i o n i s o f p o o r q u a l i t y ( i . e . l o w s a l i n i t y a n d u s u a l l y t o o warm) e n e r g y a n d m a t e r i a l s must be e x p e n d e d t o t r e a t i n t a k e w a t e r . T h e r e f o r e , i n a n a t t e m p t t o c o n s e r v e e n e r g y , w a t e r i s r e c y c l e d i n t h e s y s t e m f o r a s l o n g a p e r i o d a s p o s s i b l e . The w a t e r q u a l i t y p r o b l e m s a s s o c i a t e d w i t h t h e i n t a k e w a t e r e x h i b i t s e a s o n a l v a r i a b i l i t y , b e i n g o f m i n o r c o n c e r n d u r i n g t h e w i n t e r m o nths b u t b e c o m i n g p r o g r e s s i v e l y more a c u t e d u r i n g s p r i n g a n d summer. L o b s t e r i n t r o d u c t i o n i n t o t h e h o l d i n g t a n k s a t P a c i f i c R i m S h e l l f i s h i s a r o u t i n e p r o c e d u r e . Upon a r r i v a l o f a s h i p m e n t f r o m t h e a i r p o r t ( a p p r o x i m a t e l y 1000 kg o f l o b s t e r ) t h e i n s u l a t e d c o n t a i n e r s h o l d i n g t h e a n i m a l s a r e s o r t e d a c c o r d i n g t o w e i g h t c l a s s and moved t o a s i t e a d j a c e n t t o t a n k s a l l o c a t e d f o r t h e p a r t i c u l a r c l a s s . F o u r w e i g h t c l a s s e s a r e p r e s e n t l y b e i n g s t o c k e d : 1/ " c h i x " - one p o u n d ; 2/ " q u a r t e r s " - one and a q u a r t e r p o u n d s ; 3/ " h a l v e s " - one a n d a h a l f p o u n d s ; and 4/ " s e l e c t s " - two p o u n d s a n d o v e r . The l o b s t e r s a r e t h e n 37 i n d i v i d u a l l y p l a c e d i n t o the h o l d i n g t a n k s as q u i c k l y as p o s s i b l e . The e n t i r e p r o c e d u r e r e q u i r e s a p p r o x i m a t e l y 20 minutes depending on t h e s i z e o f t h e shipment and manpower a v a i l a b l e . 38 5.0 MATERIALS AND METHODS 5.1 I n t r o d u c t i o n P r i o r t o a d e t a i l e d d e s c r i p t i o n of m a t e r i a l s and methods, i t i s i m p o r t a n t t o p o i n t out how s e a s o n a l v a r i a t i o n i n make-up water ( F a l s e Creek) q u a l i t y , and o p e r a t i o n a l p r o c e d u r e s a t t h e h o l d i n g f a c i l i t y i n f l u e n c e d t h e e m p i r i c a l p o r t i o n o f t h i s p r o j e c t . I n g e n e r a l t h e s i t u a t i o n r e s u l t e d i n u n c o n t r o l l a b l e background c o n d i t i o n s as w e l l as d i f f i c u l t i e s i n i s o l a t i n g i n d i v i d u a l system components f o r independent s t u d y . A b r i e f o u t l i n e of some o f the o p e r a t i o n a l p r o c e d u r e s employed at the f a c i l i t y w i l l a i d i n i l l u s t r a t i n g t h e r e s e a r c h problems. Two g e o g r a p h i c a l l y d i s t a n t markets f o r l i v e l o b s t e r have been e s t a b l i s h e d by t h e company, one l o c a l l y and t h e o t h e r i n Japan. As a r e s u l t l o b s t e r s t o r a g e d u r a t i o n v a r i e s w i t h f i n a l market d e s t i n a t i o n , t h e a n i m a l s d e s t i n e d f o r Japan may o n l y r e q u i r e h o l d i n g f o r one o r two d a y s , whereas l o b s t e r s h e l d f o r l o c a l d i s t r i b u t i o n may remain i n t h e system f o r a p e r i o d of up t o s e v e r a l weeks depending on demand. T h i s dynamic s u p p l y and demand s i t u a t i o n had a d e f i n i t e i n f l u e n c e on e x p e r i m e n t a l d e s i g n . N ecessary day-to-day system maintenance such as f i l t e r backwashing a l s o a f f e c t e d t h e t i m i n g and d e s i g n of e x p e r i m e n t s . In a d d i t i o n water t r e a t m e n t equipment u p g r a d i n g and replacement made c o n t r o l l e d e x p e r i m e n t a t i o n v e r y d i f f i c u l t . A prime example of t h i s t y p e of s i t u a t i o n r e s u l t e d from t h e u p g r a d i n g of the r e f r i g e r a t i o n system a f t e r one season o f o p e r a t i o n when i t 39 became o b v i o u s t h a t t h e o r i g i n a l u n i t was i n a d e q u a t e d u r i n g the warmer months. F l u c t u a t i n g i n t a k e water q u a l i t y , r e s u l t i n g from s e a s o n a l changes i n the n a t u r a l sea water s u p p l y , a l s o had a s i g n i f i c a n t impact on t h e i n v e s t i g a t i o n s c a r r i e d out d u r i n g t h i s s t u d y . The most c o n s e q u e n t i a l and c h r o n i c problem was t h a t of f l u c - t u a t i n g s a l i n i t y l e v e l s . Temperature c o n t r o l of t h e i n t a k e water proved t o be v e r y d i f f i c u l t d u r i n g the f i r s t y e a r of o p e r a t i o n a t t h e h o l d i n g f a c i l i t y , c o n t r i b u t i n g d i r e c t l y t o h i g h l o b s t e r m o r t a l i t y a t t i m e s and c o m p l i c a t i n g e x p e r i m e n t a l d e s i g n and p r o c e d u r e s . The u p g r a d i n g of t h e r e f r i g e r a t i o n system h e l p e d t o a l l e v i a t e t h e i n t a k e water t e m p e r a t u r e problem, but an e f f i c i e n t mechanized method o f d e a l i n g w i t h t h e c o n t i n - u o u s l y c h a n g i n g s a l i n i t y l e v e l s i n t h e system has not been d e v e l o p e d . These problems a s s o c i a t e d w i t h water q u a l i t y i n v e s t i g a t i o n s a t t h e p r o j e c t s i t e s e r v e d i n p a r t as a b a s i s f o r t h e f o r m u l a t i o n o f t h e m a t e r i a l s and methods summarized below. The e x p e r i m e n t s c a r r i e d out on t h e system i n c l u d e d an i n i t i a l t e s t t o e s t a b l i s h s a m p l i n g r e q u i r e m e n t s d u r i n g t i m e - s e r i e s m o n i t o r i n g , f o l l o w e d by t h e t i m e - s e r i e s m o n i t o r i n g , and f i n a l l y a l o o k a t t h e f u n c t i o n a l c a p a b i l i t i e s o f two system components, the s i l i c a sand f i l t e r s and t h e u l t r a v i o l e t s t e r i l i z e r u n i t . The e f f i c i e n c y and f u n c t i o n a l c a p a b i l i t y of t h e r e f r i g e r a t i o n u n i t was not examined d u r i n g t h i s i n v e s t i - g a t i o n but i s i n c l u d e d i n a d e t a i l e d independent s t u d y of heat 40 b a l a n c e s on s i t e a t t h e l o b s t e r h o l d i n g f a c i l i t y (Monk 1 9 8 0 ) . A t e s t was a l s o r u n t o e s t a b l i s h a r e l a t i o n s h i p b e t w e e n t o t a l o r g a n i c c a r b o n c o n t e n t a n d c h e m i c a l o x y g e n demand o f t h e l o b s t e r h o l d i n g w a t e r . 5.2 S a m p l i n g R e q u i r e m e n t s T h i r t e e n h o l d i n g t a n k s ' w e r e i n u s e a t t h e s t u d y s i t e d u r i n g t h e r e s e a r c h o u t l i n e d i n t h i s t h e s i s . i n o r d e r t o e s t a b l i s h t h e number o f w a t e r s a m p l e s r e q u i r e d f r o m t h e s y s t e m d u r i n g t h e t i m e - s e r i e s m o n i t o r i n g e x p e r i m e n t s i t was e s s e n t i a l t o d e t e r m i n e how t h e c h e m i c a l p a r a m e t e r s , ammonia, n i t r a t e , a n d n i t r i t e v a r i e d f r o m t a n k t o t a n k a n d w i t h i n i n d i v i d u a l t a n k s . T h i s was a c c o m p l i s h e d b y c o m p a r a t i v e c h e m i c a l a n a l y s i s o f a number o f w a t e r s a m p l e s t a k e n a t a p p r o x i m a t e l y t h e same t i m e f r o m f i v e common l o c a t i o n s i n e a c h t a n k . S a m p l e s w e r e a n a l y s e d f o r ammonia-N ( t o t a l ammonia) o n l y . Ten s a m p l e s w e r e t a k e n , two f r o m e a c h o f f i v e l o c a t i o n s i n 12 o f t h e 13 t a n k s f o r a t o t a l o f 120 s a m p l e s . The 12 t a n k s i n c l u d e d t h e 9 s t a c k e d t a n k s a nd 3 o f t h e 4 u n s t a c k e d t a n k s . One o f t h e u n s t a c k e d t a n k s was r a n d o m l y o m i t t e d t o c r e a t e a b a l a n c e d e x p e r i m e n t a l d e s i g n . The f i v e s a m p l e l o c a t i o n s i n c l u d e d t h e f o u r c o r n e r a r e a s a nd t h e c e n t r e , w i t h s a m p l e s a t a l l l o c a t i o n s c o m i n g f r o m m i d - w a t e r . A l l s a m p l e s w e r e c o l l e c t e d when t h e s y s t e m was o p e r a t i n g a t f u l l h o l d i n g c a p a c i t y . The r e s u l t s w e r e s t a t i s t i - c a l l y a n a l y s e d a nd a d e c i s i o n on s a m p l e r e q u i r e m e n t s was made. The o b j e c t i v e i n t h e s t a t i s t i c a l a n a l y s i s was t o f i n d t h e most 41 s u i t a b l e t a n k ( s ) a n d l o c a t i o n w i t h t h e t a n k s t o s a m p l e . T h i s was a c c o m p l i s h e d by c a r r y i n g - o u t a t h r e e - l e v e l n e s t e d a n a l y s i s o f v a r i a n c e a s o u t l i n e d i n S o k a l a n d R o h l f ( 1 9 6 9 ) . The t h r e e f a c t o r s t h a t w e r e t e s t e d i n c l u d e d : t h e s t a c k e d v e r s u s t h e u n s t a c k e d t a n k f o r m a t ; t a n k p o s i t i o n - t o p , m i d d l e , b o t t o m o r a l o n e ; a n d t h e l o c a t i o n w i t h i n e a c h t a n k . A l l n i t r o g e n a n a l y s e s r e q u i r e d f o r t h i s e x p e r i m e n t a n d t h e o t h e r e x p e r i m e n t s o u t l i n e d i n t h i s s e c t i o n w e r e p e r f o r m e d i n (R) d u p l i c a t e on a T e c h n i c o n A u t o A n a l y s e r I I . The m e t h o d s u s e d a r e o u t l i n e d i n T e c h n i c o n A u t o A n a l y z e r I I . I n d u s t r i a l M e t h o d No. 33-69W ( 1 9 6 9 ) a n d T e c h n i c o n A u t o A n a l y z e r I I . I n d u s t r i a l M e t h o d No. 98-70W ( 1 9 7 1 ) . D e t e c t i o n l i m i t s f o r ammonia-N, n i t r i t e a n d n i t r a t e d e t e r m i n a t i o n s on t h e a u t o a n a l y s e r a r e 0.2 mg-1" 1, 0.04 ^ u g » l - 1 and 0.04 m g « l - 1 r e s p e c t i v e l y . R e s p e c t i v e c o e f f i c i e n t s o f v a r i a t i o n a r e 0.25% f o r ammonia-N, 0.95% f o r n i t r i t e a n d 1.8% f o r n i t r a t e . The f o l l o w i n g c o n v e n t i o n w i l l be u s e d i n r e p o r t i n g t h e r e s u l t s o b t a i n e d f r o m n i t r o g e n a n a l y s e s : NH^-N = u n - i o n i z e d ammonia n i t r o g e n NH*-N = i o n i z e d ammonia (ammonium) n i t r o g e n ammonia-N = u n - i o n i z e d + i o n i z e d ammonia n i t r o g e n NO~-N = n i t r i t e a s n i t r o g e n NO~-N = n i t r a t e a s n i t r o g e n I n most i n s t a n c e s t h r o u g h o u t t h i s d ocument ammonia v a l u e s a r e r e p o r t e d a s ammonia-N due t o m i n i m a l pH i n f l u e n c e on t h e f o r m a t i o n o f NH..-N. F o r e x a m p l e when pH, t e m p e r a t u r e and 42 s a l i n i t y v a l u e s o f 7.7, 17°C a n d 15°/oo r e s p e c t i v e l y a r e u s e d i n t h e c a l c u l a t i o n o f p e r c e n t NH^-N i o n i z e d t o NH*-N (Bower a n d B i d w e l l 1 9 7 8 ) , a maximum v a l u e o f 1.40% r e s u l t s , w i t h an o v e r a l l a c i d h y d r o l y s i s c o n s t a n t o f ammonium i o n s i n t h e s e a w a t e r ( p K a s ) o f 9.55. 5.3 T o t a l O r g a n i c C a r b o n v s C h e m i c a l O x y g e n Demand A l i t e r a t u r e r e v i e w r e v e a l e d t h a t v i r t u a l l y no i n f o r m a t i o n e x i s t s r e g a r d i n g t o t a l o r g a n i c c a r b o n (TOC) l e v e l s i n l o b s t e r h o l d i n g w a t e r . Some c o m p a r a t i v e d a t a h a v e b e e n r e p o r t e d on s o l i d s c h a r a c t e r i s t i c s a n d c h e m i c a l o x y g e n demand (COD) o f l o b s t e r w a s t e ( I v e r s o n 1973) b u t n o t on t h e h o l d i n g w a t e r . T h e r e f o r e , a s e r i e s o f t e s t s w e r e r u n t o e s t a b l i s h b a c k g r o u n d l e v e l s o f TOC a n d COD i n t h e l o b s t e r h o l d i n g w a t e r p r i o r t o u s e o f t h e TOC a n a l y s i s i n t h e t i m e - s e r i e s e x p e r i m e n t s . R e s u l t s o b t a i n e d w e r e u s e d t o e x a m i n e t h e r e l a t i o n s h i p b e t w e e n t h e s e two p a r a m e t e r s f o r c o m p a r t i v e p u r p o s e s . W a t e r s a m p l e s u s e d f o r t h e t e s t w e r e c o l l e c t e d f r o m t h e h o l d i n g s y s t e m d u r i n g a v a r i e t y o f b i o m a s s l o a d i n g c o n d i t i o n s i n a n a t t e m p t t o s e c u r e a r a n g e o f TOC a n d COD l e v e l s . T h r e e s a m p l e s w ere c o l l e c t e d a t e a c h l o a d i n g l e v e l a n d a n a l y s e d i n d u p l i c a t e f o r TOC u s i n g a Beckman M o d e l 915 t o t a l o r g a n i c c a r b o n a n a l y z e r , a n d f o r COD u s i n g t h e method d e s c r i b e d by B u r n s a nd M a r s h a l l ( 1 9 6 5) w h i c h i n v o l v e s t h e u s e o f a d d i t i o n a l m e r c u r i c s u l f a t e f o r t h e s u p p r e s s i o n o f c h l o r i d e o x i d a t i o n . The t o t a l o r g a n i c c a r b o n a n a l y z e r was o p e r a t e d u s i n g t h e 4 3 p r o c e d u r e s o u t l i n e d i n American P u b l i c H e a l t h A s s o c i a t i o n e t a l . ( 1 9 7 6 ) . P r i o r t o i n j e c t i o n i n t o t h e a n a l y z e r w i t h a H a m i l t o n a u t o m a t i c hypodermic s y r i n g e , t h e samples were t h o r o u g h l y homogenized. A l l samples were a n a l y s e d w i t h i n one hour of c o l l e c t i o n . 5 . 4 T i m e - S e r i e s E x p e r i m e n t s The purpose o f t h i s s e t of e x p e r i m e n t s was t o m o n i t o r , over t i m e , system response and e n v i r o n m e n t a l change r e s u l t i n g from d r a m a t i c i n c r e a s e s i n l o b s t e r biomass. These r a p i d i n c r e a s e s i n l o b s t e r biomass o c c u r d u r i n g the i n t r o d u c t i o n o f l a r g e shipments ( u s u a l l y about 1 0 0 0 kg) from e a s t c o a s t s u p p l i e r s . The incoming a n i m a l s a r e t r a n s f e r r e d r a p i d l y ( w i t h i n 1 5 m inutes of a r r i v a l a t t h e h o l d i n g f a c i l i t y ) i n t o the t a n k s , t y p i c a l l y i n c r e a s i n g a n i m a l d e n s i t y from 5 or 1 0 p e r c e n t t o 9 5 or 1 0 0 p e r c e n t of system c a p a c i t y . A t o t a l o f t e n t i m e - s e r i e s e x p e r i m e n t s were c a r r i e d out under a v a r i e t y of i n i t i a l background water q u a l i t y c o n d i t i o n s (e.g . t e m p e r a t u r e s r a n g i n g from 7 t o 1 9 ° C ) . Except f o r t h e f i r s t e x p e r i m e n t , which r a n f o r 2 4 h o u r s , d u r a t i o n was s e t a t 1 2 h ours and i n v o l v e d a t o t a l of 2 0 s a m p l i n g t i m e s . E i g h t samples were c o l l e c t e d d u r i n g t h e f i r s t two hours a t 1 5 minute i n t e r v a l s , f o l l o w e d by a s e t e v e r y 3 0 minutes f o r the next two h o u r s , and f i n a l l y a s e t e v e r y hour f o r t h e r e m a i n i n g e i g h t h o u r s . An i n i t i a l s e t o f samples was o b t a i n e d p r i o r t o each run t o e s t a b l i s h background v a l u e s . Sample r e q u i r e m e n t s were 44 e s t a b l i s h e d e x p e r i m e n t a l l y a s p r e v i o u s l y o u t l i n e d w i t h e a c h s a m p l e s e t c o n s i s t i n g o f f o u r 0.5 1 s a m p l e s o b t a i n e d f r o m t h e c e n t e r o f t a n k s 2, 5, 8 and 12 ( F i g u r e 2 ) . C o n d i t i o n s o f 100 p e r c e n t r e c y c l e w e r e i n e f f e c t t h r o u g h o u t t h e s y s t e m d u r i n g a l l t h e e x p e r i m e n t s . The w a t e r q u a l i t y p a r a m e t e r s t h a t w e r e m o n i t o r e d d u r i n g t h i s p r i n c i p l e s e t o f e x p e r i m e n t s w e r e pH, s a l i n i t y , d i s s o l v e d o x y g e n , t e m p e r a t u r e , ammonia-N, n i t r a t e - N , n i t r i t e - N , a n d t o t a l o r g a n i c c a r b o n . I n s t r u m e n t s e m p l o y e d t o d e t e r m i n e p a r a m e t e r v a l u e s i n c l u d e d a Beckman Chem-Mate p o r t a b l e pH m e t e r , a Y S I M o d e l 33 s a l i n i t y m e t e r , a Y S I M o d e l 57 d i s s o l e d o x y g e n m e t e r , a s t a n d a r d a l c o h o l t h e r m o m e t e r , a T e c h n i c o n A u t o A n a l y s e r I I ^ R ^ f o r n i t r o g e n a n a l y s i s , and a Beckman M o d e l 915 t o t a l o r g a n i c c a r b o n a n a l y s e r f o r t o t a l o r g a n i c c a r b o n d e t e r m i n a t i o n s . P r i o r t o c h e m i c a l a n a l y s i s , w h i c h t o o k p l a c e w i t h i n 18 h o u r s o f c o l l e c t i o n , a l l s a m p l e s w e r e s t o r e d i n d a r k n e s s a t 4°C. 5.5 S i l i c a S a nd F i l t e r E x p e r i m e n t s M e c h a n i c a l f i l t e r i n g e f f i c e n c y o f s i l i c a s a n d f i l t e r s h a s b e e n w e l l d o c u m e n t e d by m a n u f a c t u r e r s and i n d e p e n d e n t i n v e s t i g a t o r s . I t was t h e r e f o r e c o n s i d e r e d r e d u n d a n t t o i n c l u d e e x p e r i m e n t s d e s i g n e d t o t e s t t h i s m e c h a n i c a l f u n c t i o n d u r i n g t h e p r e s e n t s t u d y . I n s t e a d t h e f i l t e r s w e r e e v a l u a t e d w i t h r e s p e c t t o t h e i r p e r f o r m a n c e a s b i o f i l t e r s . A l o n g w i t h i t s p r i m a r y f u n c t i o n o f m e c h a n i c a l f i l t r a t i o n t h e f i l t e r m e d i a a c t s a s a s u i t a b l e s u b s t r a t e f o r t h e g r o w t h o f n i t r i f y i n g 45 b a c t e r i a w h i c h o x i d i z e ammonia t o n i t r i t e a n d n i t r a t e . I n o r d e r t o e v a l u a t e t h e p o s s i b l e s i g n i f i c a n c e o f t h i s n i t r i f i c a - t i o n p r o c e s s , ammonia, n i t r i t e a n d n i t r a t e l e v e l s i n t h e l o b s t e r h o l d i n g w a t e r i m m e d i a t e l y b e f o r e and a f t e r p a s s a g e t h r o u g h t h e s i l i c a s a n d f i l t e r s w e r e m o n i t o r e d . O n - s i t e t e s t s t o d e t e r m i n e t h e a p p r o p r i a t e f i l t e r medium g r a i n s i z e w e r e c o n d u c t e d p r i o r t o t h e n i t r i f i c a t i o n e x p e r i - m e n t s . A d e c i s i o n t o u s e a #16 s i l i c a s a n d was r e a c h e d b a s e d on a number o f p a r a m e t e r s i n c l u d i n g p a r t i c u l a t e r e m o v a l e f f i c i e n c y , i n t e r s t i t i a l b l o c k a g e , a n d b a c k w a s h f r e q u e n c y . Once t h e medium was i n s t a l l e d a n d t h e w a t e r t r e a t m e n t s y s t e m was o p e r a t i n g a t maximum c a p a c i t y a p e r i o d o f a p p r o x i m a t e l y 75 d a y s was a l l o w e d f o r b a c t e r i a l s e e d i n g a n d p o p u l a t i o n g r o w t h p r i o r t o e x p e r i m e n t a t i o n . I n c l u d e d i n t h i s c o n d i t i o n i n g p e r i o d w e r e f i v e c o n s e c u t i v e d a y s i n w h i c h t h e u l t r a v i o l e t s t e r i l i z e r u n i t was n o t o p e r a t i n g . S p o t t e ( 1 9 7 0 ) d e f i n e d a ' c o n d i t i o n e d s y s t e m 1 a s one i n w h i c h t h e n i t r i f y i n g b a c e r i a a r e i n d y n a m i c e q u i l i b r i u m w i t h t h e r o u t i n e f o r m a t i o n o f t h e i r e n e r g y s o u r c e . D u p l i c a t e e x p e r i m e n t s w e r e c o n d u c t e d a t t h r e e t e m p e r a t u r e s , 7°, 12° and 17°C, a n d w e re s c h e d u l e d f o r a p e r i o d s h o r t l y a f t e r t i m e s o f p e a k ammonia l o a d i n g i n t h e h o l d i n g s y s t e m . The peak p e r i o d u s u a l l y o c c u r r e d w i t h i n f o u r h o u r s a f t e r t h e i n t r o d u c t i o n o f a l a r g e l o b s t e r s h i p m e n t , a n d was o f r e l a t i v e l y s h o r t d u r a t i o n . T h i s p e r i o d was c h o o s e n s o t h a t a r e l a t i v e l y c o n s i s - t e n t i n f l o w l e v e l o f n i t r o g e n compounds c o u l d be e n s u r e d . S i n c e t h e t h r e e s a n d f i l t e r s w e r e a r r a n g e d i n a p a r a l l e l c o n f i g u r a - 46 t i o n , e a c h was s t u d i e d i n d i v i d u a l l y d u r i n g t h e t e s t p e r i o d s . I t was c a l c u l a t e d f r o m t h e s p e c i f i c a t i o n s o u t l i n e d i n T a b l e 2 t h a t s y s t e m t u r n o v e r t i m e ( i . e . t h e amount o f t i m e r e q u i r e d f o r a v o l u m e o f w a t e r e q u i v a l e n t t o t h e a p p r o x i m a t e v o l u m e o f t h e s y s t e m t o p a s s t h r o u g h t h e t r e a t m e n t s y s t e m ) was a p p r o x i m a t e l y one h o u r . T h e r e f o r e , t o a v o i d a s much a s p o s s i b l e t e s t i n g p r e v i o u s l y t r e a t e d w a t e r , e x p e r i m e n t d u r a t i o n was s e t a t one h o u r . E v e r y f i v e m i n u t e s d u r i n g t h e one h o u r l o n g e x p e r i m e n t s d u p l i c a t e w a t e r s a m p l e s w e r e d r a w n f r o m t h e h o l d i n g w a t e r b e f o r e a n d a f t e r p a s s a g e t h r o u g h t h e s i l i c a s a n d f i l t e r s . S a m p l i n g p o i n t s w e r e c o n v e n i e n t l y e s t a b l i s h e d i n t h e m a i n w a t e r l i n e s l e s s t h a n one m e t e r f r o m t h e f i l t e r o u t f l o w a n d i n f l o w s . S a m p l e s were a n a l y s e d f o r NO^-N, NO^-N, a n d ammonia-N. D i s s o l v e d o x y g e n a n d pH w e r e m o n i t o r e d d u r i n g e a c h e x p e r i m e n t i n t h e m a i n h o l d i n g t a n k s . P r i o r t o b o t h e x p e r i m e n t s a t e a c h o f t h e t h r e e t e m p e r a t u r e s a s e r i e s o f 10 s a m p l e s was c o l l e c t e d i n r a p i d s u c c e s s i o n f r o m one o f t h e i n p u t s a m p l i n g p o r t s a n d a n a l y s e d f o r ammonia-N and NO~N. S t a n d a r d d e v i a t i o n s and c o e f f i c i e n t s o f v a r i a t i o n w e r e c a l c u l a t e d f o r e a c h s e t o f r e s u l t s . The o b j e c t i v e o f t h i s t e s t i n g was t o e n s u r e t h a t c h a n g e s o b s e r v e d i n ammonia-N and NO~N c o n c e n t r a t i o n s were n o t t o t a l l y a r e s u l t o f i n d i v i d u a l s a m p l e v a r i a t i o n . 5.6 U l t r a v i o l e t S t e r i l i z e r E x p e r i m e n t s T h i s s e t o f e x p e r i m e n t s was d e s i g n e d t o e s t a b l i s h t h e 47 e f f e c t i v e n e s s o f t h e u l t r a v i o l e t s t e r i l i z e r u n i t s a t c o n t r o l l i n g b a c t e r i a l l e v e l s i n t h e l o b s t e r h o l d i n g w a t e r . A t i m e - s e r i e s a p p r o a c h was a d o p t e d w i t h t e s t d u r a t i o n s v a r y i n g f r o m f o u r t o s e v e n d a y s d e p e n d i n g on w a t e r t e m p e r a t u r e . F o u r e x p e r i m e n t s w e re p e r f o r m e d , t w o a t 7°C, and one e a c h a t 12° and 1 7 ° C , An a t t e m p t was made t o s c h e d u l e a l l e x p e r i m e n t s d u r i n g p e r i o d s o f m i n i m a l b i o m a s s f l u c t u a t i o n i n t h e h o l d i n g t a n k s s o t h a t v a r i a t i o n s i n b a c t e r i a l p r o d u c t i o n w o u l d r e s u l t o n l y f r o m c h a n g e s i n t e m p e r a t u r e a n d t h e i n f l u e n c e o f t h e u l t r a v i o l e t s t e r i l i z e r s . To e s t a b l i s h t h e p e r f o r m a n c e c h a r a c t e r i s t i c s o f t h e u l t r a v i o l e t u n i t i t was i m p e r a t i v e t o m o n i t o r , o v e r t i m e , b a c t e r i a l l e v e l s i n t h e r e c y c l i n g h o l d i n g w a t e r w i t h a n d w i t h o u t t h e i n f l u e n c e o f t h e s t e r i l i z e r s . Due t o p r a t i c a l c o n s t r a i n t s on t h e u s e o f t h e e n t i r e h o l d i n g s y s t e m a s a n e x p e r i m e n t a l u n i t a s u b s y s t e m was c o n s t r u c t e d t o o p e r a t e w i t h o u t u l t r a v i o l e t s t e r i l i z a t i o n b u t a t t h e same t i m e p r o v i d e a l l t h e o t h e r p r i m a r y w a t e r t r e a t m e n t f u n c t i o n s . A 341 1 g l a s s a q u a r i u m t a n k was f i l l e d w i t h l o b s t e r h o l d i n g w a t e r a n d p l a c e d i n one o f t h e m a i n h o l d i n g t a n k s , e f f e c t i v e l y i s o l a t i n g a p o r t i o n o f t h e h o l d i n g w a t e r w h i l e i n s u r i n g a c o n s i s t e n t a m b i e n t t e m p e r a t u r e . The a q u a r i u m t a n k was f i t t e d w i t h a s t a n d a r d a i r pump and c o a r s e f i l t r a t i o n u n i t t o i n s u r e a d e q u a t e d i s s o l v e d o x y g e n a nd p a r t i c u l a t e r e m o v a l r e s p e c t i v e l y . A f t e r t h e s y s t e m h a d s t a b i l i z e d ( i . e . w a t e r q u a l i t y c o n s i s t e n t w i t h m a i n s y s t e m ) t h e a q u a r i u m t a n k was s t o c k e d w i t h l o b s t e r s a t a 48 d e n s i t y e q u i v a l e n t t o t h a t o f t h e m a i n h o l d i n g s y s t e m a nd s a m p l i n g was i n i t i a t e d . D u p l i c a t e w a t e r s a m p l e s w e r e c o l l e c t e d f r o m b o t h t h e a q u a r i u m a nd t h e h o l d i n g t a n k i m m e d i a t e l y a f t e r l o b s t e r i n t r o d u c t i o n a n d t h e n on a d a i l y b a s i s u n t i l b a c t e r i a l l e v e l s r e a c h e d a l e v e l c o n s i d e r e d p o t e n t i a l l y h a r m f u l t o 'the t e s t 4 -1 a n i m a l s ( a p p r o x i m a t e l y 10 c o l o n i e s - m l ) . B a c t e r i a l l e v e l s w e r e d e t e r m i n e d by s t a n d a r d a e r o b i c p l a t e c o u n t s a c c o r d i n g t o m e t h o d s o u t l i n e d i n I n t e r n a t i o n a l A s s o c i a t i o n o f M i c r o b i o l o g i c a l S o c i e t i e s ( 1 9 7 8) and A m e r i c a n P u b l i c H e a l t h A s s o c i a t i o n ( 1 9 7 6 ) . A number o f o t h e r e x p e r i m e n t s a nd t e s t s o f d i r e c t p r a c t i c a l s i g n i f i c a n c e t o t h e s u c c e s s f u l m a n a g i n g o f t h e l i v e l o b s t e r s t o r a g e f a c i l i t y w e r e c o n d u c t e d t h r o u g h o u t t h e s t u d y p e r i o d . The r e s u l t s o b t a i n e d f r o m t h i s p e r i p h e r a l work a r e n o t r e p o r t e d h e r e b u t a r e o u t l i n e d i n p a r t i n a s e p a r a t e o p e r a t i o n s m a n u a l s u b m i t t e d t o P a c i f i c Rim S h e l l f i s h L t d . 49 6.0 EXPERIMENTAL RESULTS 6.1 Sampling Requirements Sample quantity and sampling l o c a t i o n for the time-series monitoring experiments was based on a s t a t i s t i c a l analysis of comparative water samples c o l l e c t e d throughout the holding system as outlined i n the preceeding section. Generally the vari a t i o n among samples was very low. The range of ammonia-N values for the 120 samples analysed was from 7.2 and 9.0 mg.l 1 . Despite t h i s very low range i t was f e l t that an analysis of variance was a worthwhile exercise as i t might reveal s u b t l e t i e s about the data that could be useful in determining sample requirements. To determine whether there was s i g n i f i c a n t variance within the holding system the combined results of ammonia-N determinations from the sampling runs were subjected to a three-level nested analysis of variance with unequal sample s i z e s . Calculations revealed that choice of location within each tank was s i g n i f i c a n t at the 0.01 l e v e l (Table 3). Further analysis of the data employing the multiple comparison Student-Newman-Keuls test to compare sample a c q u i s i t i o n from the f i v e sample locations was carr i e d out. Results showed that the central sampling p o s i t i o n in each tank was the most representative of the f i v e locations tested. A ser i e s of ten water samples c o l l e c t e d simultaneously at location 5 (middle of each tank) had a mean ammonia-N value of 8.14 rng'l"1 with a standard deviation of 0.07 mg^l - 1 and a 50 T a b l e 3. ANOVA t a b l e s h owing r e s u l t s o f t h r e e - l e v e l n e s t e d a n a l y s i s o f v a r i a n c e on ammonia-N s a m p l e s t o d e t e r m i n e sample r e q u i r e m e n t s . S o u r c e o f v a r i a t i o n d f SS MS Fs p o s i t i o n 3 0 .66 0 .22 0 .73 ns Tank ( w i t h i n p o s i t i o n ) 8 2 .37 0 .30 1 .07 ns L o c a t i o n ( w i t h i n t a n k ) 48 13 .52 0 .28 28 .0** e r r o r 60 0 .58 0 .01 T o t a l 119 17 .13 F.05 [ 3 . 8 ] = 4.07 F.05 [ 8 , 4 8 ] = 2.14 F.01 [ 4 8 , 6 0 ] = 1.89 51 c o e f f i c i e n t o f v a r i a t i o n (V) o f 0.86%, i n d i c a t i n g t h a t d u p l i c a t e s a m p l i n g d u r i n g t h e t i m e s e r i e s e x p e r i m e n t s h o u l d n o t be n e c e s s a r y . I n summary, s t a t i s t i c a l a n a l y s i s o f t h e m e a s u r e d ammonia-N l e v e l s t h r o u g h o u t t h e h o l d i n g t a n k s i n d i c a t e d t h a t t o o b t a i n a r e p r e s e n t a t i v e p i c t u r e o f w a t e r q u a l i t y c o n d i t i o n s i n t h e s y s t e m , w a t e r s a m p l e s s h o u l d be c o l l e c t e d f r o m t h e c e n t r a l l o c a t i o n i n any t a n k s a m p l e d . S i n c e no s i g n i f i c a n t d i f f e r e n c e b e t w e e n t h e u n s t a c k e d and s t a c k e d f o r m a t o r p o s i t i o n o f t a n k was f o u n d , an a r b i t r a r y a s s i g n m e n t o f s a m p l e t a n k s was made. E f f i c i e n t a n d a d e q u a t e c o v e r a g e o f t h e s y s t e m s h o u l d i n c l u d e s a m p l e s f r o m t h e t h r e e m i d d l e t a n k s i n t h e s t a c k e d f o r m a t and one o f t h e u n s t a c k e d t a n k s . 6.2 T o t a l O r g a n i c C a r b o n v e r s u s C h e m i c a l Oxygen Demand The r e l a t i o n s h i p b e t w e e n t o t a l o r g a n i c c a r b o n (TOC) a n d c h e m i c a l o x y g e n demand (COD) was b a s e d on a n a l y s i s o f w a t e r s a m p l e s c o l l e c t e d d u r i n g f o u r d i f f e r e n t l o b s t e r l o a d i n g r a t e s i n t h e h o l d i n g s y s t e m ( F i g u r e 7 ) . TOC l e v e l s r a n g e d f r o m a mean l o w o f 51 ppm t o a h i g h o f 192 ppm, w i t h a COD maximum and minimum o f 283 and 42 ppm r e s p e c t i v e l y . W i t h t h e e x c e p t i o n o f t h e l o w e s t c o n c e n t r a t i o n , w here mean TOC and COD l e v e l s were v i r t u a l l y i d e n t i c a l , COD l e v e l s w e r e c o n s i s t e n t l y g r e a t e r t h a n TOC v a l u e s by a p p r o x i m a t e l y 30 p e r c e n t . T r i p l i c a t e COD d e t e r - m i n a t i o n s showed s i g n i f i c a n t l y h i g h e r v a r i a t i o n t h a n TOC t r i p l i c a t e s , a s i n d i c a t e d by t h e r a n g e b a r s a s s o c i a t e d w i t h Figure 7. Relationship between TOC and COD of lobster holding water. Range bars are provided for COD concentrations. 53 e a c h p o i n t . Maximum v a r i a t i o n was r e c o r d e d f o r t h e t h r e e h i g h e s t COD v a l u e s , w here a s t h e r a n g e f o r t h e l o w e s t v a l u e was s m a l l . S i n c e v a r i a t i o n f o r TOC was m i n i m a l , r a n g e b a r s a r e n o t shown. The COD/TOC r a t i o , d e t e r m i n e d f r o m p o i n t s on t h e r e g r e s s i o n l i n e , i s 1.67. 6.3 T i m e - S e r i e s E x p e r i m e n t s A summary o f c o n d i t i o n s f o r t h e t e n t i m e - s e r i e s e x p e r i m e n t s ( T a b l e 4) shows t h a t w a t e r t e m p e r a t u r e s v a r i e d f r o m a l o w o f 7.0°C d u r i n g t h e J a n u a r y , 1980 e x p e r i m e n t t o a h i g h o f 19°C r e c o r d e d i n J u l y o f t h e p r e v i o u s y e a r . E x p e r i m e n t a l b i o m a s s l o a d a l s o v a r i e d h a v i n g a minimum o f 675 kg d u r i n g t h e J a n u a r y , 1980 e x p e r i m e n t , a maximum o f 1200 kg i n F e b r u a r y , 1980 and a mean w e i g h t o f 962.5 kg + 174.4. A c l o s e r l o o k a t t h e t e m p e r a t u r e d a t a shows t h a t e x p e r i m e n t s w e r e r u n a t b a s i c a l l y t h r e e w a t e r t e m p e r a t u r e s , a l o w o f b e t w e e n 7.0-8.0°C, a h i g h o f 18.0-19.0°C a n d a g r o u p a t m i d - r a n g e t e m p e r a t u r e s . S a l i n i t i e s r e m a i n e d c o n s t a n t d u r i n g e a c h e x p e r i m e n t r a n g i n g f r o m a h i g h o f 27 p a r t s p e r t h o u s a n d ( 0 / 0 0 ) d u r i n g e x p e r i m e n t 6 t o a l o w o f 17 0/00 d u r i n g e x p e r i m e n t 1. To a v o i d r e p e t i t i o n a n d s i m p l i f y p r e s e n t a t i o n , d a t a f r o m t h r e e e x p e r i m e n t s ( 1 , 3 a n d 10) r e p r e s e n t a t i v e o f t h e t h r e e t e m p e r a t u r e g r o u p s i s p r e s e n t e d b e l o w , t h e r e m a i n i n g d a t a i s l i s t e d e l s e w h e r e ( A p p e n d i c e s 1 , 2 , 3 , 4 , 5 , 6 ) . S e l e c t i o n o f t h e t h r e e r e p r e s e n t a t i v e d a t a s e t s was a l s o b a s e d on t h e s i z e o f t h e e x p e r i m e n t a l b i o m a s s l o a d . The e s t i m a t e d l o a d was v i r t u a l l y Table 4. Conditions for time-series experiments. E x p e r i m e n t # D a t e 1 May 29/79 2 J u n e 27/79 3 J u l y 21/79 4 Aug 24/79 5 S e p t 14/79 6 Nov 9/79 7 J a n 14/80 8 Feb 26/80 9 May 23/80 10 Aug 19/80 T e m p e r a t u r e (°C) T i m e ( h r ) W a t e r A i r 13:40 13.0 14.0 14:45 18.5 22.0 13:45 19.0 26.0 16:45 18.7 24.0 15 :50 16.0 21.0 16:05 12.0 13.0 14:35 7.0 9.0 15:20 8.0 13.0 13:30 7.5 17.0 16:15 8.0 23.0 B i o m a s s L o a d ( k g ) S a l i n i t y ( o/oo) R e s i d u a l E x p e r i m e n t a l 17 75 1050 21 85 820 23 105 1100 21 70 770 19 55 840 27 150 1080 24 50 675 26 200 1200 25 80 980 28 110 1110 55 i d e n t i c a l f o r t h e t h r e e e x p e r i m e n t s c h o o s e n , w i t h a mean o f 1087 kg and a s t a n d a r d d e v i a t i o n o f 32 k g . I n some i n s t a n c e s a d d i t i o n a l d a t a s e t s w i l l be d i s p l a y e d t o h i g h l i g h t a s p e c i f i c f e a t u r e o f t h e r e s u l t s . Due t o t e c h n i c a l p r o b l e m s w i t h t h e i n o r g a n i c c o m b u s t i o n c o l u m n i n t h e c a r b o n a n a l y s e r d u r i n g most o f t h e t i m e - s e r i e s e x p e r i m e n t s , o n l y t o t a l c a r b o n (TC) v a l u e s w e r e o b t a i n e d . T h e r e f o r e 50 s a m p l e s o f l o b s t e r h o l d i n g w a t e r w ere a n a l y s e d f o r TC and t o t a l i n o r g a n i c c a r b o n when t h e a n a l y s e r was f u l l y o p e r a t i o n a l a n d a c o r r e c t i o n f a c t o r d e v e l o p e d w h i c h a l l o w e d f o r t h e e s t i m a t i o n o f TOC f r o m t h e t i m e - s e r i e s TC v a l u e s . TC v a l u e s r a n g e d f r o m a l o w o f 19.1 t o a h i g h o f 305.8 m g - l " 1 . The r a n g e o f i n o r g a n i c c a r b o n v a l u e s was 0.1 t o 7.8 mg^l -" 1". The mean TOC c o n t e n t o f t h e s e s a m p l e s was c a l c u l a t e d t o be 97 . 0 2 % w i t h a s t a n d a r d d e v i a t i o n o f 1.39 and a c o e f f i c i e n t o f v a r i a t i o n (V) o f 1.43%. T h e s e r e s u l t s w e r e c o n s i d e r e d s i g n i f i c a n t b a s e d o n t h e l o w V v a l u e a n d t h e r e f o r e t h e TOC c o n v e r s i o n f a c t o r a p p l i e d t o a l l t h e TC r e s u l t s f r o m t h e t i m e - s e r i e s e x p e r i m e n t s was s e t a t 0.97. R e p r e s e n t a t i v e r e s u l t s f r o m t h e a n a l y s i s o f TOC d u r i n g t h e t i m e - s e r i e s e x p e r i m e n t s a r e p r e s e n t e d i n F i g u r e 8. S e v e r a l c h a r a c t e r i s t i c s a r e common t o t h e d a t a a t a l l t h r e e t e m p e r a t u r e s . An i n i t i a l d r a m a t i c i n c r e a s e i n TOC c o n c e n t r a - t i o n w i t h i n t h e f i r s t h o u r a f t e r t h e l o b s t e r i n t r o d u c t i o n was o b s e r v e d i n a l l e x p e r i m e n t s , f o l l o w e d by a g r a d u a l d e c l i n e i n c o n c e n t r a t i o n . I n most i n s t a n c e s TOC v a l u e s r e a c h e d a s t e a d y 56 TIME-SERIES TOC EXPERIMENTS I I I 1 I 1 1 I I 1 1 1 1 1 L 5_ o e EXPT 1 (13 C) h + EXPT 3 (19 C) O" 9 * EXPT 7 (7 C) §_ x x EXPT 10 (8 C) *- A o i' i Figure 8. Changes i n TOC concentxations recorded during representative time-series experiments. A discontinuous time-scale was used to emphasize changes i n TOC concentrations during the f i r s t three hours of each experiment. 57 s t a t e a t a p p r o x i m a t e l y 3 t o 4 h o u r s and f i n a l TOC l e v e l s w e r e a l l h i g h e r t h a n i n i t i a l ( T i me 0) v a l u e s . I n e x p e r i m e n t number 10 a t 8°C t h e i n i t i a l TOC c o n c e n t r a t i o n was 48.4 m g - 1 - 1 w i t h a maximum v a l u e d u r i n g t h e e x p e r i m e n t o f 405.1 m g - 1 - 1 a t 0.50 h o u r s , a n d a 12 h o u r l e v e l o f 89.4 m g - 1 - 1 . A t 13°C i n e x p e r i m e n t 1 t h e i n i t i a l c o n c e n t r a t i o n was 18.1 mg-1 ^, t h e pe a k was 316.1 m g - l - 1 a t 0.75 h o u r s a n d t h e f i n a l , 12 h o u r , v a l u e was 36.1 m g - 1 - 1 . E x p e r i m e n t 3 was r u n a t 19°C a n d h a d a n i n i t i a l c o n c e n t r a t i o n o f 25.4 m g - 1 - 1 , a p e a k o f 305.7 a t 0.50 h o u r s , and a f i n a l v a l u e o f 78.9 mg-1 1 . F o r c o m p a r a t i v e p u r p o s e s d a t a f r o m e x p e r i m e n t 7 w h i c h was r u n u n d e r much l o w e r b i o m a s s l o a d i n g ( a p p r o x i m a t e l y h a l f o f t h e p r e v i o u s l y d i s c u s s e d e x p e r i m e n t s ) c o n d i t i o n s i s a l s o p r e s e n t e d . The i n i t i a l TOC c o n c e n t r a t i o n was r e c o r d e d a t 25.1 m g - 1 - 1 , t h e p e a k v a l u e was 164.0 m g - 1 - 1 a t 0.25 h o u r s , w i t h a f i n a l 12 h o u r v a l u e o f 39.1 m g - 1 - 1 . I n a l l f o u r e x p e r i m e n t s , DO c o n c e n t r a t i o n s i n t h e h o l d i n g w a t e r d r o p p e d , i n some c a s e s d r a m a t i c a l l y , w i t h i n t h e f i r s t 15 t o 30 m i n u t e s o f t h e r u n s a n d t h e n s l o w l y i n c r e a s e d t o a s t e a d y s t a t e l e v e l w h i c h was l o w e r t h a n t h e i n i t i a l ( Time 0) v a l u e ( F i g u r e 9 ) . A t 19°C ( E x p t . 3 ) , t h e h i g h e s t e x p e r i m e n t a l t e m p e r a t u r e , a n i n i t i a l DO c o n c e n t r a t i o n o f 8.1 m g - 1 - 1 was r e c o r d e d . W i t h i n 30 m i n u t e s a f t e r t h e i n t r o d u c t i o n o f t h e l o b s t e r s h i p m e n t , t h e DO h a d d r o p p e d t o a l e v e l o f 3.4 m g - 1 - 1 . W i t h i n 2.5 h DO c o n c e n t r a t i o n s h a d r e a c h e d a s t e a d y s t a t e a t a p p r o x i m a t e l y 5.5 mg-1-"1". T h i s v a l u e r e p r e s e n t s a r e d u c t i o n 58 HUE-SERIES DO EXPERIMENTS ° I i i i i i i — i — i — i — i — i — i — i — i — i — ' — " — i — L o- e> EXPT 1 (13 C) •i + EXPT 3 (13 C) « e> EXPT 7 (7 0 x x EXPT 10 (8 C) 0 ' 0! S ' l!o ' V.6 ' 2!o ' a!6 ' 3'.0 4'i, & T!O * > «[o l i o l i O TIME (HR) Figure 9. Changes i n dissolved oxygen concentrations recorded during representative time-series experiments. A discontinuous time-scale was to emphasize changes i n dissolved oxygen during the f i r s t three hours of each experiment. 59 i n a m b i e n t DO c o n c e n t r a t i o n o f n e a r l y 3 mg-1 A t 13°C i n e x p e r i m e n t 1, t h e i n i t i a l DO v a l u e was m e a s u r e d a t 8.9 m g - 1 - 1 . A minimum DO c o n c e n t r a t i o n o f 5.3 mg-1 1 o c c u r r e d w i t h i n t h e f i r s t 1.5 h o u r s o f t h e e x p e r i m e n t . T h i s l e v e l was m a i n t a i n e d f o r an a d d i t i o n a l 0.5 h o u r s a t w h i c h t i m e t h e DO c o n c e n t r a t i o n b e g a n t o g r a d u a l l y i n c r e a s e t o a f i n a l s t e a d y s t a t e v a l u e o f 6.6 m g - 1 - 1 . E x p e r i m e n t 10 was r u n a t 8°C w i t h an i n i t i a l DO c o n c e n t r a t i o n o f 10.0 mg-1 1 . A d e c r e a s e i n DO was o b s e r v e d d u r i n g t h i s r u n r e a c h i n g a minimum v a l u e o f 7.6 i n 0.75 h. A s t e a d y s t a t e c o n c e n t r a t i o n o f a p p r o x i m a t e l y 8.5 mg-1 1 was r e a c h e d a f t e r 3 h. The t h r e e r u n s w e re a l l c a r r i e d - o u t u n d e r a p p r o x i m a t e l y t h e same b i o m a s s l o a d i n g c o n d i t i o n s . F o r c o m p a r a t i v e p u r p o s e s r e s u l t s f r o m e x p e r i m e n t 7, i n w h i c h a much l o w e r b i o m a s s l o a d o c c u r r e d , a r e p r e s e n t e d . O t h e r t h a n b i o m a s s , e x p e r i m e n t a l c o n d i t i o n s w e r e s i m i l a r t o t h o s e r e c o r d e d f o r e x p e r i m e n t 10. The i n i t i a l DO c o n c e n t r a t i o n was 10.5 m g - l " 1 w h i c h d r o p p e d t o a minimum l e v e l o f 8.5 m g - l - 1 a t t h e 0.5 h mark i n t h e e x p e r i m e n t . From t h i s p o i n t DO c o n c e n t r a t i o n g r a d u a l l y i n c r e a s e d t o a s t e a d y - s t a t e o f 9.5 m g - l " 1 . pH l e v e l s m e a s u r e d i n t h e h o l d i n g w a t e r d u r i n g e x p e r i m e n t s 1, 3 a n d 10 a r e p r e s e n t e d i n F i g u r e 10. Changes i n pH were g e n e r a l l y s m a l l b u t d e t e c t a b l e w i t h r e s u l t s f r o m e x p e r i m e n t s a t h i g h e r t e m p e r a t u r e s s h o w i n g c h a n g e s o f g r e a t e r m a g n i t u d e . A t 8°C i n e x p e r i m e n t 10 a n i n i t i a l pH o f 7.1 was r e c o r d e d w i t h 60 Figure 10. pH changes recorded during representative time-series experiments. A discontinuous time-scale was used to emphasize changes i n pH during the f i r s t three hours of each experiment. 61 subsequent measurements showing o n l y minor v a r i a t i o n . A maximum and minimum pH of 6.9, a t 1 hour, and 7.3, d u r i n g t h e f i n a l 6 hou r s of t h e e x p e r i m e n t , r e s p e c t i v e l y , were r e c o r d e d . R e s u l t s from experiment 1 (13°C) show pH b e g i n n i n g t o i n c r e a s e i m m e d i a t e l y a f t e r the l o b s t e r i n t r o d u c t i o n a t Time 0 from a l e v e l of 6.9 t o a maximum v a l u e o f 8.2 a t 2.5-3.0 h. At t h i s p o i n t , pH was ob s e r v e d t o d e c l i n e g r a d u a l l y t o a minimum l e v e l of 7.3 a t t h e t e r m i n a t i o n of t h e r u n . I n i t i a l pH i n exp e r i m e n t 3 (19°C) was 6.6. A s i m i l a r t i m e - s e r i e s p a t t e r n t o t h a t o b s e r v e d f o r experiment 1 was r e c o r d e d d u r i n g t h i s r u n . A s u b s t a n t i a l i n c r e a s e i n pH was observ e d t o t a k e p l a c e s h o r t l y a f t e r the l o b s t e r s were i n t r o d u c e d i n t o t h e system. A maximum pH l e v e l was reached a p p r o x i m a t e l y 1-2 hours a f t e r the exp e r i m e n t was i n i t i a t e d , a t which p o i n t a g r a d u a l d e c l i n e was r e c o r d e d u n t i l the t e r m i n a t i o n of t h e e x p e r i m e n t , when t h e pH was measured a t 7.6. As w i t h t h e o t h e r e x p e r i m e n t s r e p o r t e d h e r e , t h e f i n a l pH l e v e l of t h e h o l d i n g water was h i g h e r t h a n t h e l e v e l r e c o r d e d at Time 0 of t h e e x p e r i m e n t . Ammonia-N, NO~ and NO^-N c o n c e n t r a t i o n s measured d u r i n g the r e p r e s e n t a t i v e t i m e - s e r i e s e x p e r i m e n t s a r e p r e s e n t e d i n F i g u r e s 1 1 , 12, and 13 r e s p e c t i v e l y . At t h e l o w e s t tempera- t u r e o f 8°C i n exp e r i m e n t 10 t h e i n i t i a l ammonia-N c o n c e n t r a - t i o n was 1.3 mg-1 A s t e a d y i n c r e a s e i n ammonia-N was obs e r v e d f o r the f i r s t 0.75 h. of the e x p e r i m e n t , f o l l o w e d by a much more g r a d u a l i n c r e a s e t o t h e maximum l e v e l of 2.3 62 Figure 11. Changes i n ammonia-N concentrations recorded during representative time-series experiments. A discontinuous time-scale was used to emphasize changes i n ammohia-N concentrations during the f i r s t three hours of each experiment. 63 TIME-SERIES NITRITE-N EXPERIMENTS • i i i i i i i ' i i i i i i i — i — i — 1 _ e © EXPT 1 (13 C) •i + EXPT 3 (19 C) « * EXPT 10 (8 C) Z - 4—e—e—© —i 1 1 r—i 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1— . 0 as 10 13 2X) 2JS U U U U 7.0 8.0 ftO 10.0 110 iio TIME (HR) Figure 12. Changes i n nitrite-N concentrations recorded during representative time-series experiments. A discontinuous time-scale was to emphasize changes i n nitrite-N during the f i r s t three hours of each experiment. 64 TIME-SERIES NITRRTE-N EXPERIMENTS < I I I I I I I I I I I 1 1 1 1 1 L 0 © EXPT 1 (13 C) 1 + EXPT 3 (19 C) « » EXPT 10 (8 C) \ > . , in —i C ' Z q I V>-LU t— CE . or in i - " n as ' to 1 is 1 TO ' 5 1 iSo 4i u u ig ID ilo no ito iio TINE (HR) Figure 13. Changes i n nitrate-N concentrations recorded during representative time-series experiments. A discontinuous time-scale was used to emphasize changes i n nitrate-N during the f i r s t three hours of each experiment. 65 m g - l - 1 , r e c o r d e d a t 1.5 h. From t h i s p o i n t ammonia-N c o n c e n t r a t i o n s g r a d u a l l y d e c l i n e d t o 1.7 m g - l - 1 a t 12 h. A t 13°C i n e x p e r i m e n t 1 t h e ammonia-N l e v e l was r e c o r d e d a t 3.3 mg-l -"'" a t t i m e 0. I m m e d i a t e l y a f t e r t h e i n t r o d u c t i o n o f t h e l o b s t e r s ammonia-N b e g a n t o i n c r e a s e i n c o n c e n t r a t i o n r a p i d l y t o a maximum l e v e l o f 6.4 m g - l - 1 a t 1.5 h. T h i s c o n c e n t r a t i o n was m a i n t a i n e d i n s y s t e m f o r a n a d d i t i o n a l 1.5 h. a t w h i c h p o i n t a d e c l i n e was o b s e r v e d . The most d r a m a t i c d e c l i n e was r e c o r d e d o v e r t h e one h o u r p e r i o d f r o m h o u r 3 t o h o u r 4 when t h e c o n c e n t r a t i o n d r o p p e d f r o m 6.3 t o 4.9 mg-l -"''. A g e n e r a l d e c l i n e i n ammonia-N c o n c e n t r a t i o n was o b s e r v e d f o r t h e r e m a i n d e r o f t h e e x p e r i m e n t t o a f i n a l l e v e l o f 4.3 m g - l - 1 . An i n i t i a l ammonia-N c o n c e n t r a t i o n o f 1.9 mg*1~^ was r e c o r d e d f o r e x p e r i m e n t 3 a t 19°C. R e s u l t s w e r e s i m i l a r t o t h o s e p r o d u c e d d u r i n g e x p e r i m e n t 1 a s ammonia-N i n c r e a s e d r a p i d l y i m m e d i a t e l y a f t e r t h e l o b s e r i n t r o d u c t i o n t o a maximum l e v e l o f 5.0 m g ^ l " ^ a t 2.5 h, a t w h i c h p o i n t a g r a d u a l d e c l i n e was o b s e r v e d t o a minimum l e v e l o f 3.7 mg-l"''' a t 12 h. NO~-N c o n c e n t r a t i o n s m e a s u r e d d u r i n g t h e t i m e - s e r i e s e x p e r i m e n t s a r e p r e s e n t e d i n F i g u r e 12. I n e x p e r i m e n t s 10 a n d 1 a t 8°C and 13°C r e s p e c t i v e l y , NO^-N c o n c e n t r a t i o n s r e m a i n e d r e l a t i v e l y c o n s t a n t t h r o u g h o u t t h e e x p e r i m e n t s . I n o t h e r c a s e s f l u c t u a t i o n s w e r e l e s s t h a n 0.5 mg-l -'*'. A t 19°C f l u c t u a t i o n s i n NOj-N c o n c e n t r a t i o n were much more p r o n o u n c e d . A maximum NO~-N v a l u e o f 2.35 m g - l - 1 was 66 r e c o r d e d a t 3.0 h, a t w h i c h p o i n t a s l o w g r a d u a l d e c l i n e was o b s e r v e d w h i c h c o n t i n u e d u n t i l t h e t e r m i n a t i o n o f t h e r u n . NO^-N r e s u l t s f o r t h e t h r e e e x p e r i m e n t s a r e a l l s i m i l a r i n t h a t NO~-N c o n c e n t r a t i o n s r e m a i n e d b a s i c a l l y c o n s t a n t f o r t h e f i r s t 5 o r 6 h o u r s o f e a c h e x p e r i m e n t a t w h i c h p o i n t l e v e l s b e g a n t o i n c r e a s e g r a d u a l l y . A t 8°C NO^-N r e m a i n e d a t a p p r o x i m a t e l y 5 n i g - l - " * " f o r 6 h o u r s when i t b e g a n t o i n c r e a s e t o a maximum l e v e l o f 6.4 mg'! - 1 a t h o u r 12. The i n t i a l NO^-N c o n c e n t r a t i o n a t 13°C was 10.5 m g - l - 1 . The g r a d u a l i n c r e a s e i n NO^-N t h r o u g h o u t t h e e x p e r i m e n t r e s u l t e d i n a f i n a l v a l u e o f 13.3 m g » l _ 1 a t h o u r 12. A t 19°C t h e i n i t i a l a n d f i n a l NO^-N c o n c e n t r a t i o n s w e re 23.9 a n d 26.6 m g ' l - 1 r e s p e c t i v e l y . 6 . 4 S i l i c a Sand F i l t e r E x p e r i m e n t s R e s u l t s r e p o r t e d f r o m t h i s s e r i e s o f e x p e r i m e n t s a r e mean v a l u e s c a l c u l a t e d f r o m d u p l i c a t e s a m p l e s t a k e n a t e a c h s a m p l i n g p e r i o d . To e s t a b l i s h l e v e l s o f s a m p l e v a r i a t i o n a s e r i e s o f 10 w a t e r s a m p l e s w e r e t a k e n a n d a n a l y s e d f o r ammonia-N and NO^-N p r i o r t o e a c h e x p e r i m e n t as o u t l i n e d i n S e c t i o n 5.5. The mean, s t a n d a r d d e v i a t i o n a n d c o e f f i c i e n t o f v a r i a t i o n w e r e c a l c u l a t e d f o r e a c h s e t o f r e s u l t s a n d a r e p r e s e n t e d i n T a b l e 5 f o r ammonia-N a nd T a b l e 6 f o r NO^-N. I n a l l c a s e s t h e c o e f f i c i e n t o f v a r i a t i o n ( t h e s t a n d a r d d e v i a t i o n e x p r e s s e d a s a p e r c e n t a g e o f t h e mean v a l u e ) was l e s s 67 Table 5 . Mean (Y) with 95% confidence i n t e r v a l s , standard deviation (S.D.) and c o e f f i c i e n t of v a r i a t i o n (V) calculated for ammonia-N l e v e l s measured i n sample series c o l l e c t e d p r i o r to s i l i c a sand f i l t e r experiments. Temp.(°C) Expt. # Y (mg-l" 1) S.D.(mg-l"1) V (%) 1 0.4+0.02 0.01 2.5 2 1.3 + 0.22 0.11 8.5 1 1.5 + 0.18 0.09 6.0 2 7.6 + 1.35 0.69 9.1 1 13.6 + 2.63 1.34 9.9 2 6.3 + 0.71 0.36 5.7 7 12 17 68 Table 6 . Mean (Y) with 95% confidence i n t e r v a l s , standard deviation (S.D.) and c o e f f i c i e n t of v a r i a t i o n (V) calculated for NO3-N l e v e l s measured i n sample series c o l l e c t e d p r i o r to s i l i c a sand f i l t e r experiments. Temp.(°C) Expt. # _Y_(mg-l _ 1) S.D.(mg-l"1) V (%) 1 16 .4 + 2 .57 1.31 8 .0 2 24 .8 + 3 .41 1.74 7 .0 1 19 .7 + 1 .55 0.79 4 . 1 2 23 .9 + 2 .00 1.02 4 .3 1 4 .3 + 0 .22 0 .11 2 .6 2 2 9 . 5 + 3 .06 1.56 5 . 3 7 12 17 69 t h a n 10%. The s i l i c a s a n d f i l t e r e x p e r i m e n t s w e re d e s i g n e d t o d e t e c t , a n d i f p o s s i b l e q u a n t i f y , a n y n i t r i f i c a t i o n a c t i v i t y t a k i n g p l a c e i n t h e f i l t e r b e d s d u r i n g p e r i o d s o f h e a v y ammonia l o a d i n g . To a c h i e v e t h i s o b j e c t i v e ammonia-N, NO^-N and NC^-N l e v e l s i n t h e h o l d i n g w a t e r w e r e m o n i t o r e d b e f o r e and a f t e r p a s s a g e t h r o u g h t h e s a n d f i l t e r s a t t h r e e t e m p e r a t u r e s ( 7 , 12 and 1 7 ° C ) . D i s s o l v e d o x y g e n (DO), pH and s a l i n i t y were m o n i t o r e d i n t h e m a i n h o l d i n g t a n k s and a summary o f t h e r e s u l t s i s g i v e n i n T a b l e 7. S a l i n i t y l e v e l s r e m a i n e d c o n s t a n t d u r i n g e a c h e x p e r i m e n t b u t f l u c t u a t e d b e t w e e n d u p l i c a t e e x p e r i m e n t s , f o r e x a m p l e s a l i n i t i e s w e r e 25 a n d 28 p p t f o r e x p e r i m e n t s 1 and 2 r e s p e c t i v e l y a t 7°C. Maximum and minimum v a l u e s a r e p r e s e n t e d f o r d i s s o l v e d o x y g e n and pH w i t h b o t h pH a n d DO maxima o c c u r i n g u s u a l l y d u r i n g t h e i n i t i a l p h a s e o f e a c h e x p e r i m e n t a n d t h e minimum a t some p o i n t c l o s e t o t h e e n d o f t h e r u n . A DO d r o p o v e r t i m e was r e c o r d e d f o r e x p e r i m e n t s a t e a c h o f t h e t h r e e t e m p e r t u r e s . F o r b o t h r u n s a t 7°C t h e DO d r o p was m i n o r g o i n g f r o m 9.5 t o 9.1 m g - l - 1 d u r i n g t h e f i r s t e x p e r i m e n t a n d f r o m 9.8 t o 8.8 m g - l ^ i n t h e s e c o n d . A somewhat g r e a t e r d r o p was r e c o r d e d f o r t h e d u p l i c a t e r u n s a t 12°C w i t h DO g o i n g f r o m 7.3 t o 5.9 d u r i n g t h e f i r s t e x p e r i m e n t a n d f r o m 7.5 t o 5.0 m g ^ l - ^ i n t h e s e c o n d . A t 17°C maximum DO v a l u e s w e r e g e n e r a l l y l o w due t o t h e l o w c a r r y i n g c a p a c i t y o f t h e w a t e r a t t h a t t e m p e r a t u r e . A minimum DO d r o p f r o m 4.8 t o 4.1 mg«l~^ was r e c o r d e d f o r t h e f i r s t e x p e r i m e n t a t 17°C, w i t h a much more s u b s t a n t i a l l o s s r e c o r d e d f o r t h e s e c o n d r u n a s DO went 70 T a b l e 7. S u m m a r y o f d i s s o l v e d o x y g e n ( D O ) , p H a n d s a l i n i t y l e v e l s m e a s u r e d d u r i n g t h e s i l i c a s a n d f i l t e r e x p e r i m e n t s . D.O.(mg.l ) pH Temp.(°C) Expt.# max min max min S a l i n i t y (o/oo) 1 9.5 9.1 8.1 7.2 25 2 9.8 8.8 7.7 6.9 28 1 7.3 5.9 6.9 6.4 19 2 7.5 5.0 7.5 6.6 23 1 4.8 4.1 7.8 6.7 21 2 6.1 3.3 7.9 6.8 25 71 f r o m 6.1 t o 3.3 m g ' l _ J ' . E x c e p t f r o m t h e f i r s t e x p e r i m e n t a t 12°C/ pH d r o p s w e r e a l l i n t h e o r d e r o f one pH u n i t o v e r t h e one h o u r t e s t p e r i o d . A l t h o u g h ammonia-N, NO~-N and NO~-N were a l l m e a s u r e d d u r i n g t h e s e e x p e r i m e n t s o n l y ammonia-N and NO^-N v a l u e s a r e p r e s e n t e d i n t h i s s e c t i o n . A l t h o u g h some m i n o r f l u c t u a t i o n i n n i t r i t e l e v e l was m e a s u r e d d u r i n g s e v e r a l e x p e r i m e n t s , c o n c e n t r a t i o n s o f t h i s p a r a m e t e r t e n d e d t o r e m a i n f a i r l y s t a b l e t h r o u g h o u t t h e d u r a t i o n o f e a c h r u n . As a r e s u l t , t h e s e d a t a a r e e x c l u d e d f r o m t h e g r a p h i c a l p r e s e n t a t i o n . I n i t i a l d a t a a n a l y s i s h a s i n v o l v e d d e t e r m i n i n g w h e t h e r ammonia-N a n d NO^-N c o n c e n t r a t o n s i n t h e h o l d i n g w a t e r h a v e i n c r e a s e d o r d e c r e a s e d a f t e r p a s s a g e t h r o u g h t h e t h r e e f i l t e r s a t e a c h t e m p e r a t u r e . D u r i n g t h e p r o c e s s o f n i t r i f i c a t i o n ammonia-N i s o x i d i z e d t o f o r m n i t r a t e - N t h u s d e c r e a s i n g t h e c o n c e n t r a t i o n o f ammonia a n d i n c r e a s i n g t h e l e v e l o f n i t r a t e . T h i s c o n v e r s i o n i s b a s e d on t h e a s s u m p t i o n s t h a t t h e o x i d a t i o n f r o m n i t r i t e t o n i t r a t e i s r a p i d a n d t h a t no o t h e r p r o c e s s , s u c h a s v o l a t i l i z a t i o n , i s a f f e c t i n g t h e c o n c e n t r a t i o n s o f t h e s e n i t r o g e n o u s compounds. I n c r e a s i n g o r d e c r e a s i n g c o n c e n t r a t i o n s o f ammonia-N and NO^-N w e r e t h e n p l o t t e d o v e r t i m e f o r t h e t h r e e f i l t e r s a t e a c h o f t h e t e s t t e m p e r a t u r e s . P r e s e n t a t i o n o f t h e d a t a i n t h i s f a s h i o n p r e s u p p o s e s a r e l a t i v e l y c o n s t a n t i n f l o w c o n c e n t r a t i o n o f n i t r o g e n o u s compounds t h r o u g h o u t t h e d u r a t i o n o f e a c h e x p e r i m e n t . As 72 o u t l i n e d i n a p r e v i o u s s e c t i o n , f e a t u r e s w e r e i n c o r p o r a t e d i n t o t h e e x p e r i m e n t a l d e s i g n t o e n s u r e t h a t f l u c t u a t i o n was r e d u c e d t o a minimum. To i l l u s t r a t e t h e e f f e c t o f t h e s e m e a s u r e s i n f l o w c o n c e n t r a t i o n s o f ammonia-N, NO^-N and NO^-N w e r e s t a t i s t i c a l l y a n a l y s e d a n d t h e r e s u l t s a r e p r e s e n t e d i n T a b l e s 8, 9, and 10 r e s p e c t i v e l y . I t i s e v i d e n t f r o m t h e s e f i g u r e s t h a t i n f l o w c o n c e n t r a t i o n s r e m a i n e d r e l a t i v e l y c o n s t a n t d u r i n g most e x p e r i m e n t s . The c o e f f i c i e n t s o f v a r i a t i o n (V) f o r ammonia-N r a n g e d f r o m a l o w o f 1.3% i n e x p e r i m e n t 1, f i l t e r 1, a t 7°C t o a maximum o f 1 0 . 5 % i n e x p e r i m e n t 1, f i l t e r 1, a t 17°C. V v a l u e s f o r NO^-N i n f l o w s a m p l e s v a r i e d f r o m a l o w o f 1.1% i n e x p e r i m e n t 2, f i l t e r 2, a t 17°C t o a h i g h o f 2.6% i n e x p e r i m e n t 1, f i l t e r 2, a t 7°C. NO^-N V v a l u e s r a n g e d f r o m a maximum o f 7.0% a t 17°C e x p e r i m e n t 1, f i l t e r 3, t o a minimum v a l u e o f 1.9% i n e x p e r i m e n t 2, f i l t e r 2, a t t h e same t e m p e r a t u r e . F i g u r e s 14, 15 a n d 16 show t h e r e s u l t s f r o m t h e t h r e e f i l t e r s f o r e x p e r i m e n t number 1 a t 7°C. A l t h o u g h m i n o r f l u c t u a t i o n s i n ammonia-N a n d NOy-N l e v e l s w e r e d e t e c t e d i n t h e h o l d i n g w a t e r a f t e r p a s s a g e t h r o u g h f i l t e r #1 a t t h i s t e m p e r a t u r e , no g e n e r a l p a t t e r n i s e v i d e n t f r o m t h e r e s u l t s . I n i t i a l l y b o t h ammonia-N a n d NO^-N c o n c e n t r a t i o n s b e g i n t o r i s e f o l l o w e d b y a d e c l i n e i n b o t h p a r a m e t e r s a n d f i n a l l y a m i n o r i n c r e a s e i n NOj-N a c c o m p a n i e d by a e q u a l l y s m a l l d e c r e a s e i n ammonia-N. A l t h o u g h t h e m a g n i t u d e o f t h e c h a n g e s was s l i g h t l y l a r g e r , r e s u l t s o b t a i n e d f r o m f i l t e r s 2 a n d 3 Table 8. I n i t i a l and f i n a l concentrations, range, mean (Y) with 95% confidence i n t e r v a l s , standard deviation (S.D.) and c o e f f i c i e n t of variation (V) of ammonia-N in time series samples taken prior to passage through the s i l i c a sand f i l t e r s . Cone, ( m g . l - 1 ) Range (mg-l -- 1-) Temp.(°C) Ex p t . # F i l t e r # I n i t i a l F i n a l Max M i n Y S.D. V (%) 7 1 1 0.7 0.6 1.3 0.6 0.9 + 0.06 0.03 3.3 2 0.5 0.8 0.9 0.3 0.6 +0.02 0.01 1.7 3 0.9 0.9 1.1 0.5 0.8 +0.02 0.01 1.3 2 1 1.7 1.8 2.1 1.4 1.7 +0.41 0.21 12.4 2 1.8 1.5 2.0 1.2 1.7 + 0.53 0.27 15.9 3 1.2 1.2 1.5 1.0 1.2 + 0.38 0.19 15.8 12 1 1 3.4 3.5 3.8 3.0 3.4 +0.41 0.21 6.2 2 2.9 3.1 3.5 2.8 3.1 +0.43 0.22 7.1 3 3.0 2.8 3.4 2.7 3.0 +0.37 0.19 6.3 2 1 7.4 7.3 8.1 7.0 7.6 +0.59 0.30 3.9 2 7.0 7.3 7.8 7.0 7.4 + 0.57 0.29 3.9 3 7.3 7.7 7.9 7.0 7.6 + 0.53 0.27 3.6 17 1 1 2.1 2.5 2.6 1.9 2.2 +0.41 0.21 9.5 2 2.9 2.7 2.9 2.1 2.6 +0.47 0.24 9.2 3 1.8 2.3 2.3 1.7 2.0 +0.41 0.21 10.5 2 1 3.8 3.3 3.8 3.0 3.4 +0.59 0.30 8.8 2 3.4 3.1 3.9 2.9 3.4 +0.65 0.33 9.7 3 3.9 3.8 4.1 3.0 3.6 +0.65 0.33 9.2 T a b l e 9 I n i t i a l and f i n a l c o n c e n t r a t i o n s , r a n g e , mean (Y) w i t h 95% c o n f i d e n c e i n t e r v a l s , s t a n d a r d d e v i a t i o n (S.D.) and c o e f f i c i e n t o f v a r i a t i o n (V) o f n i t r a t e - N i n t i m e s e r i e s s a m p l e s t a k e n p r i o r t o p a s s a g e t h r o u g h t h e s i l i c a sand f i l t e r s . Temp.(°C) 7 Expt.# 1 F i l t e r # 1 2 3 1 2 3 Cone, (mg • I " 1 ) Range (mg •1-1) S.D. V (%) I n i t i a l F i n a l Max. M i n . Y 16.6 17.0 17.2 16.1 16.7 + 0.71 0.36 2.2 16.1 17.0 17.0 15.8 16.4 + 0.82 0.42 2.6 15.8 16.6 16.7 15.8 16.3 + 0.55 0.28 1.7 25.4 26.4 26.2 24.3 25.3 + 1.06 0.54 2.1 26.1 25.9 26.9 25.3 26.1 +0.84 0.43 1.6 24.8 26.0 26.4 24.8 25.7 + 1.04 0.53 2.1 12 1 1 2 3 19.1 18.9 19.4 19.7 19.5 20.2 19.8 19.5 20.2 18.8 18.7 18.9 19.3 19.1 19.7 +0.62 +0.51 +0.67 0.32 0.26 0.34 1.7 1.4 1.7 2 1 2 3 31.1 33.2 31.9 31.7 32.9 32.1 32.1 33.4 32.4 30.4 31.4 30.7 31.4 32.5 31. 5 + 0.95 + 1.02 +1.04 0.50 0.52 0.53 1.6 1.6 1.7 17 1 1 2 3 18.9 19.6 19.2 19.7 19.3 20.4 19.7 19 .9 20.4 18.8 18.8 18.7 19.2 19.3 19.5 + 0.55 +0.69 +0.96 0.28 0.35 0.49 1.5 1.8 2.5 2 1 2 3 39.4 38.6 39.9 39.6 39.2 39.9 41.0 39.2 40.8 38.6 37.7 38.8 39.8 38.5 39.5 + 1.14 +0.86 + 1.16 0.58 0.44 0.59 1.5 1.1 1.5 Table 10. I n i t i a l and f i n a l concentration, range, mean (Y) with 95% confidence i n t e r v a l s , standard deviation (S.D.) and c o e f f i c i e n t of variation (V) of n i t r i t e - N in time-series samples taken prior to passage through the s i l i c a sand f i l t e r s . Temp.("O Expt.# F i l t e r # 1 2 3 1 2 3 Cone, (mg-1 - 1) Range (mg-1 - 1) I n i t i a l Final Max. Min. 54 59 50 74 66 64 56 53 56 75 68 71 60 61 58 80 74 72 54 50 50 69 66 64 56 55 55 75 69 69 + 3.3 +6.5 + 4.7 +7.3 +4.7 +4.5 S.D. 1.7 3.3 2.4 3.7 2.4 2.3 V (%) 3.0 6.0 4.4 4.9 3.5 3.3 12 1 1 105 103 112 99 106 +8.2 4.2 4.0 2 100 109 110 100 106 + 5.9 3.0 2.8 3 111 108 115 108 111 + 4.5 2.3 2.1 2 1 85 94 99 85 92 +7.4 3.8 4.1 2 91 93 99 86 93 + 6.3 3.2 3.4 3 89 96 98 87 92 + 6.9 3.5 3.8 17 1 1 69 74 77 65 61 +6.5 3.3 4.6 2 65 72 75 64 69 + 6.9 3.5 5.1 3 61 75 75 61 69 +9.4 4.8 7.0 2 1 164 163 174 160 168 +8.8 4.5 2.7 2 171 174 177 165 171 + 6.3 3.2 1.9 3 158 169 174 158 168 + 9.6 4.9 2.9 76 Figure 14. Differences between f i l t e r 1 inflow and outflow ammonia-N and nitrate-N concentrations during experiment 1 at 7°C. Increasing and decreasing concentrations are represented by p o s i t i v e and negative values respectively. 77 Figure 15. Differences between f i l t e r 2 inflow and outflow ammonia-N and nitrate-N concentrations during experiment 1 at 7°C. Increasing and decreasing concentrations are represented by p o s i t i v e and negative values respectively. 78 EXPERIMENT 1 AT 1 C FILTER 3 i i — i — — i — i — i — ' — i — 1 — 1 — 1 — 1 0 © finnoNio-N 1 + NITRflTE-N Figure 16. Differences between f i l t e r 3 inflow and outflow ammonia-N and nitrate-N concentrations during experiment 1 at 7°C. Increasing and decreasing concentrations are represented by p o s i t i v e and negative values respectively. 79 a p p e a r s i m i l a r t o t h o s e m e a s u r e d f o r f i l t e r 1. R e s u l t s f r o m t h e s e c o n d e x p e r i m e n t a t 7°C a r e p r e s e n t e d i n F i g u r e s 17/ 18 and 19. A g a i n , a s w i t h t h e r e s u l t s o f t h e f i r s t r u n a t t h i s t e m p e r a t u r e , t h e r e d o e s n o t a p p e a r t o be a n y r e g u l a r p a t t e r n o f ammonia-N d e c r e a s e o r NO^-N i n c r e a s e o v e r t i m e . T h i s i s p a r t i c u l a r l y t r u e f o r f i l t e r s 2 a n d 3 w h e r e b o t h ammonia-N a n d NOj-N l e v e l s showed a l m o s t no c h a n g e a f t e r p a s s a g e t h r o u g h t h e f i l t e r s a t t h e t e r m i n a t i o n o f t h e e x p e r i m e n t . I t i s l i k e l y t h a t a n y c h a n g e s i n t h e p a r a m e t e r l e v e l s m e a s u r e d d u r i n g t h e e x p e r i m e n t a r e a r e s u l t o f s a m p l e v a r i a t i o n . A t 12°C a s i m i l a r p a t t e r n t o t h a t a t 7°C e m e r g e d . R e s u l t s f r o m t h e f i r s t r u n a t 12°C ( F i g u r e s 2 0 , 21 a n d 22) show t h a t ammonia-N a n d NOj-N l e v e l s c h a n g e d v e r y l i t t l e a f t e r p a s s a g e t h r o u g h f i l t e r 1 f o r t h e d u r a t i o n o f t h e e x p e r i m e n t . R e s u l t s f r o m f i l t e r 2 r e v e a l t h a t ammonia-N and NO^-N l e v e l s c h a n g e d v e r y l i t t l e a f t e r p a s s a g e t h r o u g h t h e s a n d f o r t h e f i r s t 20-25 m i n u t e s o f t h e e x p e r i m e n t . A t t h i s p o i n t , NO^-N o u t f l o w c o n c e n t r a t i o n s b e g a n t o i n c r e a s e a t a p p r o x i m a t e l y t h e same r a t e a s ammonia-N o u t f l o w l e v e l s w e r e d e c r e a s i n g . By t h e e n d o f t h e e x p e r i m e n t ammonia-N d i s c h a r g e c o n c e n t r a t i o n s w e r e a p p r o x i m a t e l y 1.5 m g ^ l " 1 l o w e r t h a n i n f l o w l e v e l s . The r e v e r s e was t r u e f o r NO^-N. R e s u l t s f r o m f i l t e r 3 showed t h a t ammonia-N and NO^-N o u t f l o w c o n c e n t r a t i o n s r e m a i n e d g e n e r a l l y t h e same a s c o r r e s p o n d i n g i n f l o w l e v e l s f o r a p p r o x i m a t e l y 35 m i n u t e s i n t o t h e 80 Figure 17. Differences between f i l t e r 1 inflow and outflow ammonia-N and nitrate-N concentrations during experiment 2 at 7°C. Increasing and decreasing concentrations are represented by p o s i t i v e and negative values respectively. 81 EXPERIMENT 2 AT 1 C FILTER 2 i — i — i — i — i — i — 1 — 1 — 1 — 1 — 1 e © finnONIR-N H + NITRATE-N Figure 18. Differences between f i l t e r 2 inflow and outflow ammonia-N and nitrate-N concentrations during experiment 2 at 7°C. Increasing and decreasing concentrations are represented by p o s i t i v e and negative values respectively. 82 j i i 1 1 1 EXPERIMENT 2 AT 1 C FILTER 3 i i I i i • • ' i I 1 1 1 1 1 1 L o o onnoNifl-N H + NITRRTE-N \ / — i 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 I 1 ' (LB S J B.B K J 210 2SJ) 30.0 3S.0 4 0 J 4SJ) 510 5 S J 6 0 J TinEtniN) Figure 19. Differences between f i l t e r 3 inflow and outflow ammonia-N and nitrate-N concentrations during experiment 2 at 7°C. Increasing and decreasing concentrations are represented by p o s i t i v e and negative values respectively. 83 Figure 20. Differences between f i l t e r 1 inflow and outflow ammonia-N and nitrate-N concentrations during experiment 1 at 12°C. Increasing and decreasing concentrations are represented by p o s i t i v e and negative values respectively. 84 EXPERIMENT 1 AT 12 C FILTER 2 o i I I I I i i i I I I I I L _ I I 1 1 1 1 1 1 1 L r> o o pnnoNifi-N * + NITRRTE-N r J - o "M 1 r—i 1 1 1 1 1 1 1 — T 1 1 1 1 1 1 1 1 1 1 1 1 LO 5 J V.D CJ) 20.0 2SJ) 3&D 3 S J 40.0 45J) S0.D S5.0 6S.0 TIHE(fllN) Figure 21. Differences between f i l t e r 2 inflow and outflow ammonia-N and nitrate-N concentrations during experiment 1 at 12°C. Increasing and decreasing concentrations are represented by p o s i t i v e and negative values respectively. 85 Figure 22. Differences between f i l t e r 3 inflow and outflow ammonia-N and nitrate-N concentrations during experiment 1 at 12°C. Increasing and decreasing concentrations are represented by p o s i t i v e and negative values respectively. 86 e x p e r i m e n t . A t t h a t p o i n t NO^-N r e m a i n e d r e l a t i v e l y c o n s t a n t w h e r e a s ammonia-N d r o p p e d i n c o n c e n t r a t i o n . A t 50 m i n u t e s ammonia-N i n f l o w a n d o u t f l o w c o n c e n t r a t i o n s b e g a n t o move s i g n i f i c a n t l y c l o s e r t o g e t h e r w h i c h i s c u r i o u s s i n c e NO~-N o u t f l o w l e v e l s c o n t i n u e d t o r e m a i n s i m i l a r t o i n f l o w c o n c e n t r a t i o n u n t i l t h e t e r m i n a t i o n o f t h e e x p e r i m e n t a t 60 m i n u t e s . F i l t e r s 1, 2, and 3 p r o d u c e d v e r y s i m i l a r r e s u l t s i n t h e s e c o n d e x p e r i m e n t a t 12°C ( F i g u r e s 2 3 , 24 and 2 5 ) . I n f l o w and o u t f l o w c o n c e n t r a t i o n s o f b o t h n i t r o g e n o u s compounds r e m a i n e d r e l a t i v e l y c o n s t a n t f o r t h e f i r s t 15-20 m i n u t e s o f t h e e x p e r i m e n t . A t t h a t p o i n t ammonia-N o u t f l o w c o n c e n t r a t i o n s b e g a n t o d e c r e a s e when c o m p a r e d t o i n f l o w v a l u e s . The f i n a l d i f f e r e n c e b e t w e e n i n f l o w a n d o u t f l o w c o n c e n t r a t i o n s was b e t w e e n 1.5 a n d 2.0 mg l - " ^ a t t h e e n d o f e a c h e x p e r i m e n t . R e s u l t s f r o m e x p e r i m e n t 1 a t 17°C a r e p r e s e n t e d i n F i g u r e s 26, 27 a n d 28. No s i g n i f i c a n t i n c r e a s e o r d e c r e a s e i n e i t h e r ammonia-N o r NO^-N was r e c o r d e d f o r o u t f l o w w a t e r when c o m p a r e d t o i n f l o w w a t e r f r o m f i l t e r s 2 o r 3. I n f i l t e r 1 -3 -1 NO.J -N o u t f l o w c o n c e n t r a t i o n s r o s e t o a b o u t 1 mg 1 a b o v e i n f l o w l e v e l s a t a p p r o x i m a t e l y 30-35 m i n u t e s a n d r e m a i n e d c l o s e t o t h a t l e v e l f o r t h e d u r a t i o n o f t h e e x p e r i m e n t . The r e v e r s e i s t r u e f o r ammonia-N i n f l o w a n d o u t f l o w c o n c e n t r a t i o n s a s t h e o u t f l o w l e v e l d r o p p e d t o a p p r o x i m a t e l y 1 mg l - 1 b e l o w t h e i n f l o w c o n c e n t r a t i o n . A much d i f f e r e n t p i c t u r e emerged f r o m e x p e r i m e n t 2 a t 17°C ( F i g u r e s 29, 30 a n d 3 1 ) . R e s u l t s 87 EXPERIMENT 2 AT 12 C FILTER 1 — i — i — i — i — i — i — i — i i i i i i i i i i i i i i • 0 © flnnoNifl-N 1 + NITRPJE-N / \ / \ 1 i i i — i — i — i — i — i — i — i — i — i — i — i — i — i — i — i — i — i — i — i — f - 0-0 S.0 10.0 ISO 20.0 25.0 3O.0 3S.0 40.0 45.0 50.0 55.0 60.0 TIME(tllN) Figure 23. Differences between f i l t e r 1 inflow and outflow ammonia-N and nitrate-N concentrations during experiment 2 at 12°C. Increasing and decreasing concentrations are represented by positive and negative values respectively. 88 EXPERIMENT 2 AT 12 C FILTER 2 _ i i i i i i i i i i i i i i i i i i i i i i i o c onnoNio-N H + NITRATE-N Figure 24. Differences between f i l t e r 2 inflow and outflow ammonia-N and nitrate-N concentrations during experiment 2 at 12°C. Increasing and decreasing concentrations are represented by po s i t i v e and negative values respectively. 89 EXPERIMENT 2 FIT 12 C FITER 3 1 1 I 1 1 1 1 1 1—1—1—1 1—1—1 1 — 1 — 1 — 1 — 1 — 1 — 1 — 1 — o o RnnoNiA-N +- - - * NITRRTE-N - — i — i — i — i — i — i — i — i — i — i — i — i — i — i — i — i — i — i — i — i — i — i i 0.0 5.0 10.0 6.0 20.0 25.0 3Q.D 35.0 40.0 45.0 50.0 S5.0 60.0 TlflE(MIN) F i g u r e 25. D i f f e r e n c e s between f i l t e r 3 i n f l o w and o u t f l o w ammonia-N and n i t r a t e - N c o n c e n t r a t i o n s during experiment 2 a t 12°C. Inc r e a s i n g and decreasing c o n c e n t r a t i o n s are represented by p o s i t i v e and negative v a l u e s r e s p e c t i v e l y . 90 F i g u r e 26. D i f f e r e n c e s between f i l t e r 1 i n f l o w and o u t f l o w ammonia-N and n i t r a t e - N c o n c e n t r a t i o n s d u r i n g experiment 1 a t 17°C. I n c r e a s i n g and decreasing c o n c e n t r a t i o n s are represented by p o s i t i v e and negative v a l u e s r e s p e c t i v e l y . 91 Figure 27. Differences between f i l t e r 2 inflow and outflow airmonia-N and nitrate-N concentrations during experiment 1 at 17°C. Increasing and decreasing concentrations are represented by p o s i t i v e and negative values respectively. 92 Figure 28. Differences between f i l t e r 3 inflow and outflow ammonia-N and nitrate-N concentrations during experiment 1 at 17°C. Increasing and decreasing concentrations are represented by p o s i t i v e and negative values respectively. 93 f r o m f i l t e r s 1 and 2 show a d r a m a t i c i n c r e a s e o f NO^-N o u t f l o w c o n c e n t r a t i o n w i t h r e s p e c t t o i n f l o w l e v e l s and a s u b s t a n t i a l d e c r e a s e i n ammonia-N i n t h e o u t f l o w w a t e r . B o t h e v e n t s o c c u r r e d a t a p p r o x i m a t e l y 15 m i n u t e s a f t e r t h e i n i t i a t i o n o f t h e e x p e r i m e n t . F o r b o t h f i l t e r s t h e m a g n i t u d e o f c h a n g e i s s l i g h t l y g r e a t e r f o r NO^-N t h a n t h a t r e c o r d e d f o r ammonia-N. No s i g n i f i c a n t i n c r e a s e o r d e c r e a s e was m e a s u r e d i n w a t e r p a s s i n g t h r o u g h f i l t e r 3. 6.5 U l t r a v i o l e t S t e r i l i z e r E x p e r i m e n t s U l t r a v i o l e t s t e r i l i z e r e v a l u a t i o n was c a r r i e d o u t a t t h r e e t e m p e r a t u r e s , one e x p e r i m e n t e a c h a t 12° and 17°C, and d u p l i c a t e r u n s a t 7°C. E x p e r i m e n t l e n g t h s v a r i e d f r o m a minimum o f f o u r d a y s a t 17°C t o a maximum o f s e v e n d a y s f o r e a c h o f t h e r u n s a t 7°C. M e a s u r e d p l a t e c o u n t s w e re t r a n s - f o r m e d t o Log 10 c o u n t s p e r m l f o r g r a p h i c a l p r e s e n t a t i o n ( F i g u r e s 32, 3 3 , 34, 3 5 ) . B a c t e r i a l c o n c e n t r a t i o n s i n t h e m a i n c o n t r o l t a n k r e m a i n e d a t c o n s i s t e n t l y l o w l e v e l s t h r o u g h o u t t h e e n t i r e s e r i e s o f e x p e r i m e n t s . B a c k g r o u n d c o n c e n t r a t i o n s i n c r e a s e d s l i g h t l y w i t h i n c r e a s i n g t e m p e r a t u r e , a r e s u l t w h i c h i s n o t s u r p r i s i n g s i n c e t e m p e r a t u r e i s an i m p o r t a n t f a c t o r i n c o n t r o l l i n g b a c t e r i a l p o p u l a t i o n d y n a m i c s . A c e r t a i n amount o f d a i l y f l u c t u a t i o n i n t h e c o n t r o l t a n k was r e c o r d e d , b u t v a r i a t i o n s w e r e c o n s i d e r e d i n s i g n i f i c a n t when c o m p a r e d t o t h e s t e a d i l y i n c r e a s i n g c o n c e n t r a t i o n o b s e r v e d i n t h e a q u a r i u m s y s t e m a s e x p e r i m e n t s 94 EXPERIMENT 2 RT 17 C FILTER 1 i i i > i i i i ' i i i i i i i i i i i i i i o o nnnoNifl-N i i — i — i — i — i — i — i — i — i — i — i — i — i — i — i — i — i — i — i — i — i — i — i — i — r 0.0 S.0 10.0 6.9 20.0 25.0 30.9 35.0 40.0 45.0 50.0 55.0 60.0 TinF.miN) Figure 29. Differences between f i l t e r 1 inflow and outflow ammonia-N and nitrate-N concentrations during experiment 2 at 17°C. Increasing and decreasing concentrations are represented by p o s i t i v e and negative values respectively. — © finnoNio-N - -+ NITRATE-N i -K Figure 30. Differences between f i l t e r 2 inflow and outflow^aimonia-1 and nitrate-N concentrations during experiment 2 at 17°C Increasing and decreasing concentrations are represented by p o s i t i v e and negative values respectively. 96 Figure 31. Differences between f i l t e r 3 inflow and outflow ammonia-N and nitrate-N concentrations during experiment 2 at 17°C. Increasing and decreasing concentrations are represented by p o s i t i v e and negative values respectively. 97 Figure 32. Changes i n b a c t e r i a l concentration recorded over time i n the aquarium and tank during UV experiment 1 at 7°C. Figure 33. Changes i n b a c t e r i a l concentration recorded over time i n the aquarium and tank during UV experiment 2 at 7°C. 9 Figure 34. Changes i n b a c t e r i a l concentration recorded over time i n the aquarium and tank during the UV experiment at 12°C. 100 Figure 35. Changes i n b a c t e r i a l concentration recorded over time i n the aquarium and tank during the UV experiment at 17 °C. 101 p r o g r e s s e d . A l l e x p e r i m e n t s were t e r m i n a t e d when b a c t e r i a l c o n c e n t r a - 4 t i o n s i n t h e a q u a r i u m t a n k r e a c h e d a p p r o x i m a t e l y 10 c o u n t s ml -"*". I n t h e a q u a r i u m , b a c t e r i a l c o n c e n t r a t i o n s a p p e a r e d t o be r e l a t e d t o w a t e r t e m p e r a t u r e . AT 7°C t h e 1 0 4 c o u n t s m-"1" l e v e l was r e a c h e d i n a p p r o x i m a t e l y 7 d a y s d u r i n g b o t h e x p e r i m e n t s a t t h i s t e m p e r a t u r e ( F i g u r e 32 a n d 33) w h e r e a s o n l y 4-5 d a y s were r e q u i r e d a t 12°C ( F i g u r e 34) and a p p r o x i m a t e l y 3 d a y s a t 17°C ( F i g u r e 35) t o r e a c h t h e same c o n c e n t r a t i o n . 102 7.0 DISCUSSION 7.1 S a m p l i n g R e q u i r e m e n t s A l t h o u g h t h e o v e r a l l v a r i a t i o n i n ammonia-N c o n c e n t r a t i o n s was l o w f o r t h e s a m p l e s a n a l y s e d i n t h i s segment o f t h e s t u d y , s t a t i s t i c a l a n a l y s i s o f t h e r e s u l t s d i d r e v e a l some n o t e w o r t h y f e a t u r e s o f t h e d a t a . I n i t i a l l y , i t was e x p e c t e d t h a t no s i g n i f i c a n t d i f f e r e n c e w o u l d be f o u n d b e t w e e n t h e s t a c k e d and u n s t a c k e d f o r m a t s i n c e t h e t h r e e s t a c k e d t a n k s w e re a v e r a g e and c o m p a r e d t o t h e u n s t a c k e d . I n t h e s t a c k e d f o r m a t t h e c e n t r e t a n k was n o t f o u n d t o be most r e p r e s e n t a t i v e o f t h e t h r e e , a s i t m i g h t be e x p e c t e d t o h a v e a m e d i a n l e v e l o f ammonia-N i n a c a s c a d i n g s y s t e m . A l t h o u g h t h e s e r e s u l t s may h o l d t r u e u n d e r most c a s e s , a c e r t a i n amount o f v a r i a t i o n s h o u l d o c c u r d e p e n d i n g on a number o f f a c t o r s s u c h a s t e m p e r a t u r e , a n i m a l l o a d , pH and a g g i t a t i o n o f t h e h o l d i n g w a t e r . The c e n t r a l p o s i t i o n i n e a c h t a n k was f o u n d t o be t h e most r e p r e s e n t a t i v e o f ammonia-N c o n c e n t r a t i o n s t h r o u g h o u t t h e t a n k a t t h e 0.01 l e v e l o f s i g n i f i c a n c e . An a n a l y s i s o f h y d r u a l i c movement i n t h e t a n k s w o u l d be r e q u i r e d t o f u l l y e x p l a i n t h i s r e s u l t . By r e d u c i n g t h e number o f s a m p l e s t a k e n d u r i n g t h e e x p e r i m e n t s i n t h e h o l d i n g t a n k s , s a m p l e a n a l y s i s was n o t d e l a y e d and s t o r a g e t i m e was k e p t t o a minimum. By m i n i m i z i n g t h e s e two f a c t o r s , g r e a t e r a c c u r a c y a n d p r e c i s i o n was a c h i e v e d i n t h e s a m p l e a n a l y s i s . I t i s b e l i e v e d t h a t t h e a n a l y s i s c o n d u c t e d i n t h i s segment o f t h e work p r o v i d e d a d e q u a t e 103 j u s t i f i c a t i o n f o r t h e s a m p l i n g r e g i m e a d o p t e d . 7.2 T o t a l O r g a n i c C a r b o n v s . C h e m i c a l Oxygen Demand B i o c h e m i c a l O x y g en Demand (BOD) and C h e m i c a l O x y gen Demand (COD) a r e methods t h a t h a v e b e e n t r a d i t i o n a l l y u s e d i n c u l t u r e and h o l d i n g s y s t e m s t o d e t e r m i n e o x y g e n demand o f t h e h o l d i n g w a t e r s . T h e s e m e t h o d s h a v e a s s o c i a t e d w i t h them i n h e r e n t p r o b l e m s and t i m e c o n s t r a i n t s a s o u t l i n e d i n S e c t i o n 5, t h e r e f o r e , i t was d e c i d e d t o e v a l u a t e t h e p o s s i b i l i t y o f u s i n g t h e T o t a l O r g a n i c C a r b o n (TOC) a n a l y s i s a s a r e p l a c e m e n t f o r BOD a n d COD, t h u s m i n i m i z i n g t h e t i m e c o n s t r a i n t p r o b l e m s . The TOC v a l u e s i n t u r n c o u l d be c o m p a r e d t o a s e r i e s o f COD m e a s u r e - m e n t s c a r r i e d - o u t on t h e l o b s t e r h o l d i n g w a t e r s o a r a t i o c o u l d be d e r i v e d . S i n c e no r e s u l t s f r o m TOC m e a s u r e m e n t s on l o b s t e r w a s t e s o r h o l d i n g w a t e r h a v e b e e n p u b l i s h e d t h i s r a t i o i s i m p o r t a n t i f a n y c o m p a r i s o n s a r e t o be made w i t h p u b l i s h e d COD v a l u e s . A l t h o u g h v a r i a t i o n was r e l a t i v e l y l a r g e f o r t h e COD d e t e r m i n a t i o n s , a s t r o n g c o r r e l a t i o n b e t w e e n COD and TOC was d e m o n s t r a t e d ( F i g u r e 7 ) . An e q u a l l y s t r o n g c o r r e l a t i o n was e s t a b l i s h e d f o r s w i n e w a s t e s by B u l l e y a n d Husdon ( 1 9 7 4 ) . T h i s s t r o n g c o r r e l a t i o n i s a g o o d i n d i c a t i o n t h a t TOC c a n be u s e d a s a r e l i a b l e i n d i c a t o r o f o x y g e n demand f o r t h e l o b s t e r h o l d i n g w a t e r . The r a t i o f o u n d f o r COD/TOC o f 1.67 f a l l s w i t h i n t h e r a n g e o f v a l u e s p u b l i s h e d f o r o t h e r w a s t e t y p e s . A v a l u e o f 2.20 f o r t h e COD/TOC r a t i o i s g i v e n by E c k e n f e l d e r ( 1 9 7 0 ) f o r 104 t h e e f f l u e n t f r o m b i o l o g i c a l l y t r e a t e d d o m e s t i c w a s t e . A much h i g h e r COD/TOC r a t i o o f 3.60 was r e p o r t e d by V a n H a l l a n d S t e n g e r ( 1 9 6 3) f o r m u n i c i p a l w a s t e w a t e r . A COD/TC r a t i o o f 1.50 was a l s o g i v e n , i n d i c a t i n g t h a t t h e m u n i c i p a l w a s t e w a t e r c o n t a i n e d a c o m p a r a t i v e l y h i g h c o n c e n t r a t i o n o f i n o r g a n i c c a r b o n . A l t h o u g h c o m p o s i t i o n w o u l d be e x p e c t e d t o be somewhat d i f f e r e n t t h a n l o b s t e r h o l d i n g w a t e r , u n t r e a t e d d o m e s t i c w a s t e w a t e r e x h i b i t i n g c o m p a r a b l e COD and TOC l e v e l s i s c o n s i d e r e d weak on a s c a l e o f weak, medium a n d s t r o n g ( M e t c a l f a n d Eddy 1 9 7 9 ) . I t may be e x p e c t e d t h a t t h e s t o i c h i o m e t r i c COD/TOC r a t i o o f a w a s t e w a t e r w o u l d be a p p r o x i m a t e l y e q u i v a l e n t t o t h e 3 2 m o l e c u l a r r a t i o o f o x y g e n t o c a r b o n ( 2 . 6 6 ) . E c k e n f e l d e r ( 1 9 7 0 ) s t a t e s t h a t t h e o r e c t i c a l l y t h e r a t i o l i m i t s w o u l d r a n g e f r o m z e r o , when t h e o r g a n i c m a t e r i a l i s r e s i s t a n t t o d i c h r o m a t e o x i d a t i o n , t o 5.33 f o r methane o r s l i g h t l y h i g h e r when i n o r g a n i c r e d u c i n g a g e n t s a r e p r e s e n t . The c o m p a r a t i v e l y l o w COD/TOC r a t i o d e t e r m i n e d f o r t h e l o b s t e r h o l d i n g w a t e r may be a r e s u l t o f r e s i s t e n t o r g a n i c compounds. BOD a n d COD r e s u l t s may o f t e n n o t i n c l u d e many o r g a n i c compounds t h a t a r e p a r t i a l l y o r t o t a l l y r e s i s t e n t t o b i o c h e m i c a l o r d i c h r o m a t e o x i d a t i o n . I n most i n s t a n c e s , a l l t h e o r g a n i c c a r b o n i n t h e s e compounds i s r e c o v e r e d i n t h e TOC a n a l y s i s . 7. 3 T i m e - S e r i e s E x p e r i m e n t s The w a t e r q u a l i t y p a r a m e t e r s c h o o s e n f o r s t u d y d u r i n g t h i s 105 s e r i e s o f e x p e r i m e n t s i n c l u d e t e m p e r a t u r e , s a l i n i t y , o x y g e n , pH, TOC, ammonia-N, n i t r a t e - N a n d n i t r i t e - N . A l o n g w i t h t h e b i o m a s s l o a d , b o t h t e m p e r a t u r e a n d s a l i n i t y r e m a i n e d b a s i c a l l y c o n s t a n t t h r o u g h o u t e a c h e x p e r i m e n t . T h i s c o n d i t i o n was t h e r e s u l t o f t h e r e l a t i v e l y s h o r t d u r a t i o n o f e a c h e x p e r i m e n t (12 h o u r s ) . Due t o t h e s y s t e m d e s i g n b o t h o f t h e s e p a r a m e t e r s t e n d e d t o v a r y i f m o n i t o r e d o v e r a l o n g e r t i m e p e r i o d ( d a y s a s o p p o s e d t o h o u r s ) . The t e m p e r a t u r e p r o b l e m was e v e n t u a l l y a d d r e s s e d a n d r e c t i f i e d b a s e d i n p a r t on r e c o m m e n d a t i o n s d e v e l o p e d d u r i n g t h e work done f o r t h i s t h e s i s . The r e f r i g e r a - t i o n s y s t e m w h i c h was e v e n t u a l l y i n s t a l l e d t o r e p l a c e t h e g r o s s l y u n d e r s i z e d u n i t o r i g i n a l l y i n p l a c e , now m a i n t a i n s t h e h o l d i n g w a t e r a t a t e m p e r a t u r e o f b e t w e e n 6°C a n d 8°C, a t e m p e r a t u r e w h i c h m e e t s t h e e s t a b l i s h e d l e v e l f o r t h i s p a r a m e t e r . As f a r a s t h e t i m e - s e r i e s e x p e r i m e n t s w e r e c o n c e r n e d s a l i n i t y c a n be c o n s i d e r e d a c o n s t a n t , w h i c h s i m p l i f i e s t h e a n a l y s i s o f t h e r e s u l t s . G e n e r a l l y t h o u g h , s a l i n i t y l e v e l s i n t h e h o l d i n g w a t e r w e r e l o w when c o m p a r e d t o o p e n o c e a n c o n c e n t r a t i o n s w h i c h v a r y f r o m a b o u t 30 t o 35 o/oo. As o u t l i n e d i n t h e l i t e r a t u r e r e v i e w s e c t i o n , t h e s e l o w s a l i n i t y l e v e l s c a n h a v e a d v e r s e e f f e c t s o n l o b s t e r h e a l t h , a n d a s s u c h w e r e a m a t t e r o f c o n c e r n d u r i n g t h i s w o r k . The p r o b l e m o f f l u c t u a t i n g s a l i n i t y l e v e l s i s n o t a d d r e s s e d h e r e s i n c e i t was c o n s i d e r e d a p r a c t i c a l management p r o b l e m . I s s u e s o f t h i s s o r t a r e i n c l u d e d i n a w a t e r q u a l i t y m a n u a l d e v e l o p e d d u r i n g t h e 106 c o u r s e o f t h i s s t u d y and p r e s e n t e d t o t h e m a n a g e r s o f t h e l o b s t e r h o l d i n g o p e r a t i o n . S a l i n i t y l e v e l s w e r e p a r t i c u l a r l y l o w d u r i n g t h e f i r s t f i v e e x p e r i m e n t s a f t e r w h i c h a method i n v o l v i n g t h e a d d i t i o n o f b u l k s a l t t o t h e h o l d i n g w a t e r was d e v e l o p e d and i m p l e m e n t e d d u r i n g t h e p e r i o d c o v e r i n g t h e f i v e r e m a i n i n g e x p e r i m e n t s . S a l i n i t y c a n be m a i n t a i n e d a t an a c c e p t a b l e l e v e l u s i n g t h i s t e c h n i q u e b u t t h e s y s t e m must be m o n i t o r e d c o n t i n u o u s l y due t o s a l i n i t y c h a n g e r e s u l t i n g f r o m e v a p o r a t i o n and d i l u t i o n . TOC c o n c e n t r a t i o n s w e r e c a l c u l a t e d f r o m t o t a l c a r b o n m e a s u r e m e n t s e s t a b l i s h e d d u r i n g t h e t i m e - s e r i e s e x p e r i m e n t s u s i n g a n e m p i r i c a l l y d e t e r m i n e d c o n v e r s i o n f a c t o r o f 0.97. A l t h o u g h t h e v a l u e o f 97% o r g a n i c c a r b o n i s a r e l a t i v e l y l a r g e f r a c t i o n o f t h e c a r b o n p r e s e n t i t d o e s n o t a p p e a r t o be o u t o f l i n e when c o m p a r e d t o TOC l e v e l s i n o t h e r t y p e s o f w a s t e s . F o r i n s t a n c e , B u l l e y and Husdon ( 1 9 7 4 ) d e t e r m i n e d t h a t TOC c o m p r i s e d a p p r o x i m a t e l y 90% o f t h e d e t e c t a b l e c a r b o n i n s w i n e w a s t e . A s i m i l a r v a l u e h a s b e e n f o u n d f o r d a i r y manure ( p e r s o n a l c o m m u n i c a t i o n D r . V. L o ) . I n a c t u a l i t y t h e f r a c t i o n o f TOC w i l l v a r y d e p e n d i n g on t h e m a t e r i a l e n t e r i n g t h e l o b s t e r h o l d i n g w a t e r , b u t i f t h e m a j o r i t y o f t h e m a t e r i a l i s l o b s t e r w a s t e i t i s q u i t e r e a s o n a b l e t o assume t h a t t h e r a t i o o f TOC/TC w i l l r e m a i n h i g h . The r e s u l t s f r o m t h e TOC m o n i t o r i n g show a n i m m e d i a t e a n d d r a m a t i c i n c r e a s e i n TOC c o n c e n t r a t i o n s i n t h e l o b s t e r h o l d i n g w a t e r a f t e r t h e i n t r o d u c t i o n o f t h e a n i m a l s . Two f a c t o r s may 107 be c o n t r i b u t i n g t o t h i s i n c r e a s e . The f i r s t i s t h a t the l o b s t e r s may be s u p e r f i c i a l l y c a r r y i n g m a t e r i a l , or e x c r e t i n g wastes w i t h a h i g h TOC l e v e l . I t i s i n e v i t a b l e t h a t some m a t e r i a l w i l l be t r a n s p o r t e d on t h e e x t e r i o r of t h e l o b s t e r s d u r i n g shipment and e v e n t u a l l y e n t e r the h o l d i n g w a t e r . A l t h o u g h no s u b s t a n t i a l amount of e x t e r i o r f o u l i n g was ever o b s e r v e d on t h e c a r a p a c e s of newly a r r i v e d l o b s t e r s , i t cannot be s t a t e d c o n c l u s i v e l y t h a t e l e v a t e d TOC c o n c e n t r a t i o n s r e c o r d e d i m m e d i a t e l y a f t e r l o b s t e r i n t r o d u c t i o n a r e not a t t r i b u t a b l e t o t h i s f a c t o r . I t i s a l s o d i f f i c u l t t o s t a t e c o n c l u s i v e l y t h a t newly i n t r o d u c e d l o b s t e r s a r e or a r e not e x c r e t i n g q u a n t i t i e s of f e c a l m a t t e r s u b s t a n t i a l enough t o cause t h i s TOC change. T h i s w i l l depend on how r e c e n t l y f e e d i n g has o c c u r r e d and t h e n a t u r e of t h e f o o d consumed. No f o o d i s a v a i l a b l e t o the l o b s t e r s d u r i n g the a p p r o x i m a t e l y 24 hour s h i p p i n g p e r i o d from th e e a s t c o a s t , and i t i s q u i t e l i k e l y t h a t many of t h e a n i m a l s have been h e l d p r i o r t o shipment w i t h o u t f o o d f o r a number of days. T h e r e f o r e , i t i s u n l i k e l y t h a t l a r g e q u a n t i t i e s of f e c a l m a t t e r w i l l be g e n e r a t e d i n t h e h o l d i n g system d u r i n g t h e r e l a t i v e l y s h o r t p e r i o d a l o t t e d t o t h e t i m e - s e r i e s e x p e r i m e n t s . The second, and most p r o b a b l e f a c t o r c o n t r i b u t i n g t o the r a p i d i n c r e a s e i n TOC i m m e d i a t e l y a f t e r the l o b s t e r i n t r o d u c t i o n i s t h e r e s u s p e n s i o n of p r e v i o u s l y s e t t l e d m a t e r i a l d u r i n g the l o b s t e r l o a d i n g a c t i v i t i e s . A s u b s t a n t i a l amount of m i x i n g r e s u l t s from p l a c i n g t h e a n i m a l s i n t h e t a n k s as w e l l as from 108 l o b s t e r movement d u r i n g t h i s p e r i o d . L o b s t e r a c t i v i t y was o b s e r v e d t o be more p r o n o u n c e d a t h i g h e r t e m p e r a t u r e s w h i c h s u g g e s t s t h a t TOC i n c r e a s e s s h o u l d be g r e a t e r a t t h e h i g h e r e x p e r i m e n t a l t e m p e r a t u r e s , a s s u m i n g s i m i l a r l e v e l s o f r e s i d u a l TOC. T h i s however was n o t t h e c a s e , a s t h e i n c r e a s e a t 8°C was s l i g h t l y l a r g e r t h a n t h o s e r e c o r d e d f o r e x p e r i m e n t s a t e i t h e r 13 o r 19°C. A p r o p o r t i o n a t e l y much s m a l l e r i n c r e a s e i n TOC was r e c o r d e d a t 7°C ( e x p e r i m e n t 7) when t h e b i o m a s s l o a d was r e d u c e d . T h i s r e s u l t i l l u s t r a t e s t h e s i g n i f i c a n c e o f t h e a c t u a l s i z e o f t h e b i o m a s s l o a d a n d l o a d i n g p r o c e d u r e , r a t h e r t h a n t h e a c t i v i t y o f t h e a n i m a l s , i n r e s u s p e n d i n g t h e o r g a n i c m a t e r i a l i n t h e t a n k s . The i n c r e a s e d a c t i v i t y , r e s u l t i n g f r o m t h e h i g h e r t e m p e r a t u r e s , may h a v e a n a f f e c t on t h e s e t t l i n g r a t e o f t h e m a t e r i a l . A t h i g h e r t e m p e r a t u r e s s e t t l i n g may t a k e l o n g e r w h i c h e x t e n d s t h e p e r i o d o f e l e v a t e d TOC l e v e l s . T h e r e i s some e v i d e n c e o f t h i s o c c u r i n g d u r i n g t h e h i g h e r t e m p e r a t u r e e x p e r i m e n t s . V i s u a l l y , t h e r e was no i n c r e a s e i n w a t e r t u r b i d i t y s h o r t l y a f t e r t h e l o b s t e r i n t r o d u c t i o n o v e r t h a t o b s e r v e d d u r i n g p e r i o d s o f n o r m a l o p e r a t i o n . The most s i g n i f i c a n t p o t e n t i a l e f f e c t o f t h i s TOC i n c r e a s e o n l o b s t e r h e a l t h i s t h e a s s o c i a t e d i n c r e a s e i n o x y g e n demand o f t h e h o l d i n g w a t e r . A s s u m i n g t h i s TOC i n c r e a s e i s i n p a r t a r e s u l t o f w a s t e e x c r e t i o n by t h e l o b s t e r s , t h e n i t i s p o s s i b l e t h a t ammonia-N c o n c e n t r a t i o n s w i l l r i s e due t o m i n e r a l i z a t i o n , i n t h e f o r m o f d e a m i n a t i o n , o f p r o t e i n s i n t h e w a s t e m a t e r i a l . The COD e q u i v a l e n t a s d e t e r m i n e d by t h e COD/TOC r a t i o o f 1.67 109 w o u l d be a p p r o x i m a t e l y two t h i r d s a g a i n a s h i g h a s t h e TOC v a l u e s . F o r e x a m p l e , t h e p e a k c o n c e n t r a t i o n o f TOC a t 13°C d u r i n g e x p e r i m e n t 1 was a p p r o x i m a t e l y 325 mg-l""'', w h i c h i s e q u i v a l e n t t o a COD c o n c e n t r a t i o n o f 543 mg«l~'1'. Oxygen demand i n a q u a c u l t u r e o p e r a t i o n s i s u s u a l l y e x p r e s s e d a s BOD 5. T h e r e f o r e t o c o m p a r e t h e r e s u l t s o b t a i n e d h e r e w i t h p u b l i s h e d d a t a a BOD^/COD r a t i o must be e s t i m a t e d . I t i s g e n e r a l l y known t h a t t h e BOD^/COD r a t i o v a r i e s f r o m 0.4 t o 0.8 f o r a v a r i e t y o f w a s t e s ( M e t c a l f a n d Eddy 1 9 7 2 ) . I f a m e d i a n v a l u e o f 0.6 i s assumed t h e n t h e e q u i v a l e n t BOD c o n c e n t r a t i o n o f t h e h o l d i n g w a t e r w o u l d be 326 m g - l - 1 , a l m o s t i d e n t i c a l t o t h a t d e t e r m i n e d f o r TOC. Oxygen demand m e a s u r e d a s BOD 5 f o r f i s h h a t c h e r y h o l d i n g w a t e r v a r i e s f r o m a p p r o x i m a t e l y 50 t o 100 m g - l - ^ (Brown a n d Nash 1 9 8 1 ; S l o n e e t a l 1 9 8 1 ) , v a l u e s w h i c h a r e s i m i l a r t o t h e b a c k g r o u n d o r s t e a d y s t a t e v a l u e s m e a s u r e d i n t h e l o b s t e r t a n k s . Maximum c o n c e n t r a t i o n s w h i c h w e r e o b s e r v e d d u r i n g t h e f i r s t h o u r o f e a c h t i m e - s e r i e s e x p e r i m e n t a r e s u b s t a n t i a l l y h i g h e r a s o u t l i n e d a b o v e . I t a p p e a r s t h e n t h a t t h e f i r s t h o u r a f t e r t h e l o b s e r i n t r o d u c t i o n i s t h e c r i t i c a l p e r i o d w i t h r e s p e c t t o o x y g e n demand. One a d d i t i o n a l a d v e r s e e f f e c t o f h i g h TOC c o n c e n t r a t i o n s i n s e a w a t e r s y s t e m s i s t h a t t h e d i s s o l v e d component o f t h e o r g a n i c c a r b o n t e n d s t o r e d u c e t h e b u f f e r i n g c a p a c i t y o f t h e w a t e r by a f f e c t i n g t h e s o l u b i l i t y o f t h e c a l c i u m a n d magnesium c a r b o n a t e s p r e s e n t . T h i s i n f l u e n c e on b u f f e r i n g c a p a c i t y i s 1 1 0 a d d r e s s e d i n more d e t a i l i n t h e d i s c u s s i o n on pH. The r e s u l t s from t h e d i s s o l v e d oxygen m o n i t o r i n g show v e r y c l e a r l y t h a t t h e g r e a t e s t oxygen demand o c c u r s w i t h i n the f i r s t hour a f t e r l o b s t e r i n t r o d u c t i o n . D.O. drops of a p p r o x i m a t e l y 1 . 5 , 2.2 and 3.4 m g - l " 1 were r e c o r d e d 0.25 h a f t e r l o b s t e r i n t r o d u c t i o n a t 8 ° , 1 3 ° and 1 9 ° C r e s p e c t i v e l y . S i n c e oxygen exchange a t t h e a i r - w a t e r i n t e r f a c e i s a c o n t i n u o u s p r o c e s s , D.O. drops r e c o r d e d d u r i n g the t h r e e e x p e r i m e n t s a r e i n a sense b u f f e r e d . But s i n c e t h e s e drops a r e c o m p a r a t i v e l y l a r g e and o c c u r over such a s h o r t time p e r i o d , e r r o r s i n r a t e c a l c u l a t i o n s s h o u l d be m i n i m a l . These changes i n d i s s o l v e d oxygen c o n c e n t r a t i o n can be e x p r e s s e d as a l o b s t e r r e s p i r a t i o n r a t e i n mg-g - 1 o f l o b s t e r . h - 1 . Assuming an average system water volume of 1 7 , 5 0 0 1, an average l o b s t e r w e i g h t of 7 0 0 g, and an average of 1 6 0 0 l o b s t e r s i n t h e t a n k s d u r i n g each e x p e r i m e n t , c a l c u l a t i o n s r e v e a l t h a t d i s s o l v e d oxygen c o n c e n t r a t i o n d e c r e a s e d a t a r a t e of 0 . 0 9 4 m g . g - 1 . h - 1 , 0 . 1 3 8 m g - g - 1 . h - 1 and 0 . 2 1 3 mg.g - 1 a t 8 ° , 1 3 ° , and 1 9 ° C r e s p e c i v e l y d u r i n g t h e f i r s t 15 min. of each r u n . These r a t e s were c a l c u l a t e d from a c t u a l d i s s o l v e d oxygen drops which were from 1 0 . 0 t o 8.5 a t 8 ° C , 8.9 t o 6.7 a t 1 3 ° C and 8.1 t o 4.7 a t 1 8 ° C . These v a l u e s can be compared w i t h p u b l i s h e d e s t i m a t e s of l o b s t e r oxygen consumption. Two e m p i r i c a l l y d e r i v e d e s t i m a t e s e x i s t i n t h e l i t e r a t u r e . The f i r s t i s an e q u a t i o n p r e s e n t e d i n A l l e n and J o h n s t o n ( 1 9 7 6 ) based on the work of McLeese ( 1 9 6 4 ) : I l l M = K WB (10) o o w h e r e M Q = q u a n t i t y o f o x y g e n consumed (mg«hr W = w e i g h t o f t h e l o b s t e r (g) B = an u n e x p l a i n e d p a r a m e t e r d e r i v e d f r o m u n p u b l i s h e d d a t a K Q = (0. 0 1 6 9 U - 0.0974) U = w a t e r t e m p e r a t u r e (°C) U s i n g t h e p r e v i o u s l y e s t a b l i s h e d p a r a m e t e r v a l u e s t h e f o l l o w i n g o x y g e n c o n s u m p t i o n r a t e s c a n be g e n e r a t e d f r o m E q u a t i o n 1 0 : 0.017, 0.056 a n d 0.102 mg• g " 1 o f l o b s t e r - h " 1 a t 8 °, 13° and 19°C r e s p e c t i v e l y . The s e c o n d s o u r c e o f c o m p a r a t i v e d a t a i s a n o x y g e n u p t a k e v e r s e s t e m p e r a t u r e c u r v e d e v e l o p e d by A r y e s and Wood (1 9 7 7 ) f o r m a r k e t s i z e l o b s t e r s h e l d u n d e r s t e a d y s t a t e e n v i r o n m e n t a l c o n d i t i o n s . From t h i s c u r v e o f o x y g e n c o n s u m p t i o n r a t e s v s t e m p e r a t u r e , t h e o x y g e n c o n s u m p t i o n r a t e s c o r r e s p o n d i n g t o t h e t h r e e e x p e r i m e n t a l t e m p e r a t u r e s a r e 0.048, 0.068, a n d 0.99 mq- g " 1 o f l o b s t e r - h - 1 f o r 8 ° , 13° a n d 19°C r e s p e c t i v e l y . The t h r e e s e t s o f 0^ c o n s u m p t i o n d a t a a r e p r e s e n t e d i n F i g u r e 36. The o x y g e n c o n s u m p t i o n r a t e s p r e s e n t e d i n t h e l i t e r a t u r e c o m p a r e f a v o u r a b l y o v e r a w i d e r a n g e o f t e m p e r a t u r e s . A l t h o u g h a s i m i l a r r e l a t i o n s h i p e x i s t s b e t w e e n o x y g e n c o n s u m p t i o n and t e m p e r a t u r e f o r t h e t i m e - s e r i e s e x p e r i m e n t s ( i . e . 0 2 c o n s u m p t i o n i n c r e a s e s w i t h i n c r e a s i n g t e m p e r a t u r e ) t h e a c t u a l r a t e s a t e a c h t e m p e r a t u r e a r e a p p r o x i m a t e l y d o u b l e t h e l i t e r a t u r e v a l u e s . A number o f e x p l a n a t i o n s c a n be p u t f o r t h Figure 36. E f f e c t of temperature on oxygen consumption during the f i r s t 0.25 hr of representative time-series experiments, expressed as mg of oxygen consumed per g of lobster per hr, compared to published values from two l i t e r a t u r e sources. 113 i n t h i s r e g a r d . F i r s t , t h e l i t e r a t u r e v a l u e s were d e r i v e d t h r o u g h t e s t s w i t h u n s t r e s s e d ( o t h e r t h a n t e m p e r a t u r e ) a n i m a l s , where as the l o b s t e r s used i n the t i m e - s e r i e s e x p e r i m e n t s were p o t e n t i a l l y under c o n s i d e r a b l e s t r e s s and e x h i b i t e d a b n o r m a l l y h i g h a c t i v i t y l e v e l s d u r i n g t h e e a r l y p a r t o f each t e s t . Both f a c t o r s c o n t r i b u t e t o i n c r e a s e d oxygen demand. As o u t l i n e d i n the d i s c u s s i o n on TOC, an a d d i t i o n a l i n c r e a s e i n oxygen consumption c o u l d r e s u l t from t h e suspended and d i s s o l v e d m a t e r i a l p r e s e n t i n the h o l d i n g water d u r i n g the p e r i o d i m m e d i a t e l y a f t e r l o b s t e r i n t r o d u c t i o n . These f a c t o r s c o u l d be r e s p o n s i b l e f o r the h i g h e r oxygen demand o b s e r v e d d u r i n g t h e s e e x p e r i m e n t s . A l t h o u g h d i s s o l v e d oxygen i n t h e h o l d i n g t a n k s never r e a c h e d t h e p o t e n t i a l l y l e t h a l l e v e l of 2 mg d u r i n g any of t h e e x p e r i m e n t s t h e r a p i d r a t e s a t which t h e oxygen was consumed d u r i n g the f i r s t hour o f t h e runs s h o u l d be a major c o n s i d e r a t i o n w i t h r e s p e c t t o l o b s t e r h e a l t h and s t o r a g e s u c c e s s . I t i s l i k e l y t h a t due t o the added s t r e s s a s s o c i a t e d w i t h t h e i n t r o d u c t i o n and s h o r t - t e r m s t o r a g e p r o c e s s , t h e p o t e n t i a l l y l e t h a l l e v e l may be c o n s i d e r a b l y h i g h e r . F i n a l l y t h e r e s u l t s p r e s e n t e d here i l l u s t r a t e t h e importance of m a i n t a i n i n g a low h o l d i n g water t e m p e r a t u r e so t h a t oxygen c a r r y i n g c a p a c i t y i s maximized and l o b s t e r m e t a b o l i c r a t e i s m i n i m i z e d . A l t h o u g h p o t e n t i a l l y l e t h a l pH l e v e l s d i d not occur d u r i n g any of the t i m e - s e r i e s e x p e r i m e n t s , t h e changes t h a t were 114 m e a s u r e d a r e c o n s i d e r e d s i g n i f i c a n t f o r a number o f r e a s o n s . G e n e r a l l y , t h e pH v a l u e s r e c o r d e d d u r i n g t h e e x p e r i m e n t s w e r e l o w e r t h a n t h e o p t i m u m h o l d i n g r a n g e o f 8.0 t o 8.3. S i n c e most e x p e r i m e n t s d e s i g n e d t o t e s t t h e e f f e c t s o f pH on a q u a t i c o r g a n i s m s h a v e d e a l t w i t h e x t r e m e o r l e t h a l l e v e l s , v e r y l i t t l e i s known a b o u t t h e l o n g - t e r m s u b l e t h a l e f f e c t s r e s u l t i n g f r o m m i n o r d e v i a t i o n f r o m t h e o p t i m a l pH r a n g e . The p o t e n t i a l s y n e r g i s t i c e f f e c t r e s u l t i n g f r o m v a r i a t i o n i n o t h e r w a t e r q u a l i t y p a r a - m e t e r s a l s o makes i t d i f f i c u l t t o c h a r a c t e r i z e and q u a n t i f y c h r o n i c e f f e c t s . B e t t e r known a r e t h e c a u s e and e f f e c t r e l a t i o n s h i p s a s s o c i a t e d w i t h pH and a number o f b i o l o g i c a l a n d c h e m i c a l p r o c e s s e s t h a t may t a k e p l a c e i n a l o b s t e r h o l d i n g s y s t e m . T h e s e r e a l t i o n s h i p s f o r m t h e b a s i s f o r s t u d y i n g t h e pH c h a n g e s o b s e r v e d d u r i n g t h e t i m e - s e r i e s e x p e i m e n t s . No s i g n i f i c a n t v a r i a t i o n i n pH was o b s e r v e d d u r i n g e x p e r i m e n t 10 a t 8°C. T h i s o b s e r v a t i o n i s c o n s i s t a n t w i t h t h o s e made f o r t h e o t h e r p a r a m e t e r s w h i c h a l s o e x h i b i t o n l y m i n o r v a r i a t i o n o v e r - t i m e a t l o w t e m p e r a t u r e . B i o c h e m i c a l p r o c e s s e s , s u c h a s n i t r i f i c a t i o n a n d r e s p i r a t i o n , w h i c h a r e i n f l u e n c e d by pH a r e a l s o e f f e c t e d by t e m p e r a t u r e . Low e n v i r o n m e n t a l t e m p e r a t u r e t e n d s t o e i t h e r s l o w down o r a r r e s t t h e s e p r o c e s s e s . A t t h e h i g h e r t e m p e r a t u r e s u s e d i n e x p e r i m e n t s 1 a n d 2 s i g n i f i c a n t pH c h a n g e s d i d o c c u r . C o n s i s t e n t p a t t e r n s o f pH c h a n g e e m e r g e d d u r i n g e x p e r i m e n t s a t b o t h 13° and 19°C. F o r t h e f i r s t t h r e e h o u r s o f e a c h r u n , 115 pH l e v e l s i n c r e a s e d t o a p o i n t a p p r o x i m a t e l y 1.5 u n i t s h i g h e r t h a n t h e t i m e 0 v a l u e . A s t e a d y d e c l i n e was t h e n o b s e r v e d w h i c h brought t h e pH down a g a i n t o w i t h i n 1 or 0.5 u n i t s of the time 0 l e v e l by the end of each run. I t i s q u i t e p r o b a b l e t h a t the i n i t i a l i n c r e a s e i n pH o b s e r v e d d u r i n g b o t h h i g h tempera- t u r e e x p e r i m e n t s was a r e s u l t o f e l e v a t e d ammonia-N c o n c e n t r a - t i o n which appeared i n the h o l d i n g water s h o r t l y a f t e r the l o b s t e r i n t r o d u c t i o n s ( t h e ammonia-N m o n i t o r i n g i s d i s c u s s e d b e l o w ) . The pH t h e n d e c l i n e d i n response t o a c i d i f i c a t i o n r e s u l t i n g p o s s i b l y from m i n e r a l i z a t i o n of o r g a n i c c a r b o n compounds, n i t r i f i c a t i o n and a n i m a l r e s p i r a t i o n . In g e n e r a l , b i o l o g i c a l o x i d a t i o n i n h o l d i n g systems exceeds r e d u c t i o n o v e r a l l , r e s u l t i n g i n a g r a d u a l d e c l i n e i n a l k a l i n i t y and pH. The r e c y c l e d seawater i n t h e l o b s t e r h o l d i n g system u s u a l l y had a r e l a t i v e l y low pH a t the b e g i n n i n g of each e x p e r i m e n t . The g e n e r a l l y low pH v a l u e s may be a r e s u l t of t h e s e o x i d a t i o n p r o c e s s e s or t h e y may be r e l a t e d t o low b u f f e r i n g c a p a c i t y of the h o l d i n g w a t e r . S i n c e s a l i n i t y l e v e l s were low d u r i n g t h e two h i g h t e m p e r a t u r e e x p e r i m e n t s , b u f f e r i n g c a p a c i t y may have been s i g n i f i c a n t l y reduced. In a d d i t i o n , o r g a n i c carbon c o n c e n t r a t i o n s were h i g h d u r i n g much o f t h e e x p e r i m e n t s and i t has been shown t h a t d i s s o l v e d o r g a n i c s can c o a t t h e s u r f a c e s o f c a r b o n a t e p a r t i c l e s t h u s r e d u c i n g t h e number of i o n i c exchange s i t e s (Meyers and Quin 1 9 7 1 ; Suess 1 9 7 0 ) . A v e r y i m p o r t a n t f a c t o r i s t h e m e d i a t i n g e f f e c t of pH on aqueous c h e m i c a l r e a c t i o n s , p a r t i c u l a r l y t h e h y d r o l y s i s of ammonia. 1 1 6 A l t h o u g h t o t a l ammonia c o n c e n t r a t i o n s i n c r e a s e d s i g n i f i - c a n t l y d u r i n g t h e f i r s t two hours of e x p e r i m e n t s ru n a t 13 and 1 9 ° C , c a l c u l a t i o n s showed t h a t NH^-N never reached t o x i c l e v e l s (see T a b l e 1 ) . NH-̂ -N c o n c e n t r a t i o n s were c a l c u l a t e d by u s i n g t h e f o l l o w i n g e q u a t i o n d e r i v e d by W h i t f i e l d ( 1 9 7 4 ) : % NH 3 = 1 0 0 / [ 1 + a n t i l o g ( p K a s ( T ) - p H ) ] ( 1 1 ) where p K a s = a c i d h y d r o l y s i s c o n s t a n t of ammonium i o n s i n seawater and p K a S ( T ) = p K a S ( T = 298°K) + 0 . 0 3 2 4 ( 2 9 8 - T ° K ) NH^-N v a l u e s w i t h i n the pH range 7.5 t o 8.5 were o b t a i n e d from t a b l e s p r e s e n t e d i n Bower and B i d w e l l ( 1 9 7 8 ) . The h i g h e s t c o n c e n t r a t i o n s of NH^-N c a l c u l a t e d from th e t o t a l ammonia l e v e l s p r e s e n t e d i n t h e r e s u l t s a r e o u t l i n e d i n T a b l e 1 1 . These v a l u e s a r e s u b s t a n t i a l l y l e s s t h a n t h e a c c e p t a b l e l o b s t e r h o l d i n g l e v e l of 1.3 mg N H j - N ' l " 1 . Ammonia-N c o n c e n t r a t i o n s i n c r e a s e d i n a g e n e r a l l y l i n e a r f a s h i o n d u r i n g t h e f i r s t 0.5 h of each e x p e r i m e n t . T h i s was a l s o t h e p e r i o d d u r i n g which th e g r e a t e s t r a t e i n c r e a s e was r e c o r d e d . Ammonia-N l e v e l s i n c r e a s e d by a p p r o x i m a t e l y 1 . 4 , 4.2 and 4.1 m g - l " 1 ^ - 1 d u r i n g t h e f i r s t 0.5 h of each run a t 8, 13 and 1 9 ° C r e s p e c t i v e l y . By i n c o r p o r a t i n g the p r e v i o u s l y d e f i n e d l o b s t e r biomass parameters i n t o c o n v e r s i o n c a l c u l a t i o n s t h e s e r a t e s can be e x p r e s s e d i n mg o f ammonia-N.g - 1 o f l o b s t e r . h - 1 . The r a t e s c o r r e s p o n d i n g t o t h e t h r e e tempera- t u r e s a r e 0 . 0 2 1 9 mg g " 1 h " 1 a t 8 ° C , 0 . 0 6 5 6 mg.g" 1 . h " 1 a t 1 3 ° C and 0 . 0 6 2 5 mg g - 1 h " 1 a t 1 9 ° C . The o n l y compara- 117 T a b l e 1 1 . H i g h e s t NH3-N c o n c e n t r a t i o n s t h a t o c c u r e d d u r i n g t h e t i m e s e r i e s e x p e r i m e n t s c a l c u l a t e d f r o m m easured t o t a l ammonia-N v a l u e s and pH. 8°C T e m p e r a t u r e 13°C 19°C N H 3 _ N ( m g - 1 - 1 ) 0.005 0.15 0.17 T o t a l Ammonia-N ( m g - 1 - 1 ) 2.2 6.0 4-7 pH 7.1 8.1 8.1 p K a s 9.51 9.68 9.51 Time ( h r ) 0.75 2.0 1.5 118 t i v e v a l u e a v a i l a b l e i n t h e l i t e r a t u r e i s t h a t p r e s e n t e d by L o g a n a n d E p i f a n i o ( 1 9 7 8 ) . I n t h i s w ork ammonia e x c r e t i o n r a t e s w e r e e s t a b l i s h e d f o r l a r v a l a n d j u v e n i l e l o b s t e r s f e e d i n g on A r t e m i a s a l i n a a t 22°C. The r e l a t i o n s h i p f o r b o t h d e v e l o p m e n t a l s t a g e s was e x p r e s e d i n t h e e q u a t i o n : U = -0.33 l o g W- 0.37 where U = /ag NH-j.mg d r y weight -'*", h - 1 a n d V/ = l o b s t e r w e i g h t i n mg I n d e r i v i n g t h i s r e l a t i o n s h i p L o g a n a n d E p i f a n i o f o u n d t h a t s t a g e 7 p o s t - l a r v a l l o b s t e r s e x c r e t e d a p p r o x i m a t e l y 12.46 / i g NH.j • mg d r y wt-'''* h - 1 . A g r o s s c o n v e r s i o n o f t h i s v a l u e t o mg ammonia-N. g wet w e i g h t " 1 , h - 1 c a n b e made f o r c o m p a r i s o n w i t h t h e t i m e - s e r i e s c a l c u l a t i o n s by a s s u m i n g a pH o f 8.0 and a wet w t . / d r y w t . r a t i o o f 0.1. The v a l u e r e s u l t i n g f r o m t h i s c a l c u l a t i o n i n 0.0167 m g » g - 1 . h - 1 . A l l t h r e e o f t h e ammonia-N r a t e s c a l c u l a t e d f r o m t h e t i m e - s e r i e s d a t a p a r t i c u l a r l y t h o s e a t 13 a n d 19°C, a r e s u b s t a n t i a l l y h i g h e r t h a n t h e r a t e s g e n e r a t e d f r o m t h e c o n v e r s i o n o f t h e L o g a n a n d E p i f a n i o e x c r e t i o n d a t a . As p r e v i o u s l y s t a t e d , t h e r e a r e g e n e r a l l y two i m p o r t a n t s o u r c e s o f ammonia i n a l o b s t e r h o l d i n g o f c u l t u r i n g s y s t e m , m i n e r a l i z a t i o n o f o r g a n i c compounds by b a c t e r i a a n d e x c r e t i o n by t h e l o b s t e r s . W a s t e m a t e r i a l a d h e r i n g t o t h e e x t e r n a l s u r f a c e o f t h e l o b s t e r i s an a d d i t i o n a l s o u r c e t h a t may be s i g n i f i c a n t . A l t h o u g h t h e ammonia p r o d u c t i o n r a t e s c a l c u l a t e d f r o m t h e t i m e - s e r i e s e x p e r i m e n t s a p p e a r t o be h i g h c o m p a r e d t o t h e r e s u l t s o f L o g a n 119 a n d E p i f a n i o ( 1 9 7 8 ) , t h e t w o s e t s o f d a t a a r e s u r p r i s i n g l y c l o s e c o n s i d e r i n g t h e two s e t s o f e x p e r i m e n t a l c o n d i t i o n s . The ammonia e x c r e t i o n r a t e s p r e s e n t e d by L o g a n a n d E p i f a n i o w e r e d e t e r m i n e d f r o m e x p e r i m e n t s o n j u v e n i l e l o b s t e r s i n a s t a b l e c u l t u r e e n v i r o n m e n t w i t h a r e g u l a r f o o d s u p p l y . A l t h o u g h i t i s g e n e r a l l y known t h a t j u v e n i l e a q u a t i c o r g a n i s m s h a v e a p r o p o r - t i o n a t e l y h i g h e r m e t a b o l i c r a t e a n d u t i l i z e a v a i l a b l e p r o t e i n more e f f i c i e n t l y t h a n a d u l t s , i t w o u l d be a n t i c i p a t e d t h a t ammonia e x c r e t i o n by a d u l t l o b s t e r s i n a h o l d i n g s y s t e m w i t h a s l i g h t l y l o w e r w a t e r t e m p e r a t u r e a n d l i t t l e o r no a v a i l a b l e f o o d s h o u l d be l o w e r . S i n c e t h i s d i d n o t seem t o be t h e c a s e d u r i n g t h e t i m e - s e r i e s e x p e r i m e n t s one p o s s i b l e e x p l a n a t i o n i s t h a t a s u b s t a n t i a l p o r t i o n o f t h e ammonia g e n e r a t e d i n t h e h o l d i n g w a t e r s h o r t l y a f t e r t h e l o b s t e r i n t r o d u c t i o n may be a r e s u l t o f m i n e r a l i z a t i o n o f o r g a n i c m a t t e r . A l t h o u g h i t i s p o s s i b l e t h a t some ammonia may be g e n e r a t e d i n t h i s f a s h i o n , i t i s u n l i k e l y t h a t t h e b a c t e r i a i n t h e s y s t e m c a n g e n e r a t e s u f f i - c i e n t a mounts o f ammonia t h r o u g h p r o c e s s e s s u c h a s d e a m i n a t i o n t o a c c o u n t f o r t h e q u a n t i t i e s t h a t a r e o u t s t a n d i n g b a s e d o n t h e c a l c u l a t i o n s made a b o v e , p a r t i c u l a r l y d u r i n g t h e s h o r t p e r i o d u n d e r c o n s i d e r a t i o n . I t i s c o n c l u d e d t h e r e f o r e , t h a t e x c r e t i o n by t h e l o b s t e r s i s t h e m a j o r s o u r c e o f ammonia b u t t h a t c o n - t r o l l i n g TOC may e f f e c t i v e l y r e d u c e t h e c o n c e n t r a t i o n o f t h i s compound p a r t i c u l a r l y d u r i n g t h e c r i t i c a l p e r i o d i m m e d i a t e l y a f t e r l o b s t e r i n t r o d u c t i o n . S i n c e a number o f f a c t o r s may i n f l u e n c e ammonia-N c o n c e n - 120 t r a t i o n i n t h e h o l d i n g w a t e r a f t e r t h e l o b s t e r i n t r o d u c t i o n s , i t i s d i f f i c u l t t o d e t e r m i n e what p r o p o r t i o n o f t h e r e d u c t i o n i n ammonia-N c o n c e n t r a t i o n o b s e r v e d o v e r t i m e i n t h e t i m e - s e r i e s e x p e r i m e n t s may be a t t r i b u t a b l e t o r e d u c e d e x c r e t i o n by t h e l o b s t e r s . D e s p i t e t h i s p r o b l e m t h e f a c t r e m a i n s t h a t a s u b s t a n t i a l l o a d o f ammonia-N a p p e a r s i n t h e h o l d i n g w a t e r s h o r t l y a f t e r t h e i n t r o d u c t i o n , a n d i t i s t h i s d r a m a t i c i n c r e a s e t h a t s h o u l d be o f c o n c e r n i n t h e d e s i g n a n d m a i n t e n a n c e o f a l o b s t e r h o l d i n g s y s t e m . The s t r e s s f u l c o n d i t i o n s a n d i n c r e a s e d a c t i v i t y e x p e r i e n c e d by t h e l o b s t e r s d u r i n g t h e f i r s t 15 o r 30 m i n u t e s a f t e r t h e i n t r o d u c t i o n may be i m p o r t a n t f a c t o r s i n p r o d u c i n g t h i s s h o r t - t e r m i n c r e a s e i n ammonia-N e x c r e t i o n . I t h a s b e e n shown f o r f i s h t h a t ammonia-N e x c r e t i o n w i l l o f t e n i n c r e a s e u n d e r s t r e s s ( H o a r and R a n d a l l 1 9 6 9 ) . A l t h o u g h ammonia p r o d u c t i o n r a t e s w e r e s i m i l a r a t 13 a n d 19°C, r a t e s w e r e s u b s t a n t i a l l y l e s s a t 8°C i n d i c a t i o n t h a t p r o d u c t i o n was l i m i t e d by t e m p e r a t u r e . T h i s i s n o t s u r p r i s i n g s i n c e m e t a b o l i c a c t i v i t i e s o f a l l t h e o r g a n i s m s i n t h e h o l d i n g s y s t e m w e r e r e d u c e d a t l o w e r t e m p e r a t u r e s . T h i s o b s e r v a t i o n i s p a r t i c u l a r l y i m p o r t a n t t o n o t e w i t h r e s p e c t t o l o b s t e r h o l d i n g s y s t e m d e s i g n . F i n a l l y , i n a l l t i m e - s e r i e s e x p e r i m e n t s , ammonia l e v e l s r e a c h e d a maximum and t h e n b e g a n t o g r a d u a l l y d e c l i n e . T h i s g r a d u a l r e d u c t i o n i n ammonia c o n c e n t r a t i o n c a n be a t t r i b u t e d t o one o r b o t h o f two i n c i d e n t a l p r o c e s s e s , v o l a t i l i z a t i o n o r n i t r i f i c a t i o n . V o l a t i l i z a t i o n i s a n a l o g o u s t o t h e i n d u s t r i a l 121 a n d m u n i c i p a l w a s t e t r e a t m e n t p r o c e s s o f a i r s t r i p p i n g . T h i s p r o c e s s i s pH d e p e n d e n t . As t h e pH o f t h e w a s t e w a t e r i n c r e a s e s a b o v e 7, t h e ammonia-ammonium e q u i l i b r i u m i s s h i f t e d t o t h e ammonia component w h i c h may be removed a s a g a s by a g i t a t i n g t h e w a s t e w a t e r i n t h e p r e s e n c e o f a i r . The w a s t e w a t e r pH i s commonly a d j u s t e d t o 11 p r i o r t o a e r a t i o n . A t 25°C a n d pH 11 a p r o x i m a t e l y 9 8 % o f t h e ammonia i n t h e w a s t e w a t e r i s i n t h e u n - i o n i z e d f o r m . A l t h o u g h a c o n s i d e r a b l e amount o f a e r a t i o n d o e s t a k e p l a c e a t c e r t a i n l o c a t i o n s i n t h e l o b s t e r h o l d i n g s y s t e m , pH l e v e l s t e n d t o r e m a i n c o m p a r a t i v e l y l o w , w h i c h i n t u r n means t h a t t h e r a t e o f NH^-N v o l a t i l i z e d i s a l s o l o w . N i t r i f i c a t i o n c o u l d a l s o a c c o u n t f o r t h e g r a d u a l r e d u c t i o n i n ammonia-N c o n c e n t r a t i o n s o v e r t i m e i n t h e h o l d i n g s y s t e m N i t r i t e a nd n i t r a t e c o n c e n t r a t i o n s w e r e m o n i t o r e d d u r i n g t h e t i m e - s e r i e s e x p e r i m e n t s t o e s t a b l i s h f i r s t w h e t h e r c o n c e n t r a t i o n s o f t h e s e compounds c h a n g e d o v e r t i m e i n t h e s y s t e m , w h i c h w o u l d i n d i c a t e some l e v e l o f n i t r i f i c a t i o n , a n d s e c o n d , w h e t h e r t o x i c l e v e l s w e r e e v e r r e a c h e d . M o n i t o r i n g r e v e a l e d t h a t l e v e l s o f n i t r i t e a n d n i t r a t e d i d n o t a p p r o a c h t o x i c c o n c e n t r a t i o n s d u r i n g a n y o f t h e t i m e - s e r i e s e x p e r i m e n t s b u t i t i s p o s s i b l e t h a t s u b - l e t h a l e f f e c t s may r e s u l t f r o m t h e l e v e l s d e t e c t e d i n t h e s y s t e m . S i n c e v i r t u a l l y n o t h i n g h a s b e e n r e p o r t e d r e g a r d i n g s u b - l e t h a l e f f e c t s on c o n c e n t r a t i o n s i t i s d i f f i c u l t t o e v a l u a t e t h i s p o t e n t i a l p r o b l e m . Some i n d i c a t i o n o f n i t r i f i c a t o n i s e v i d e n t f r o m t h e n i t r i t e a n d n i t r a t e a n a l y s i s . A l t h o u g h m i n o r v a r i a t i o n s i n 122 n i t r i t e c o n c e n t r a t i o n o v e r t i m e w e r e r e c o r d e d , p a r t i c u l a r l y a t 19°C, g e n e r a l l y c o n c e n t r a t i o n s r e m a i n e d r e l a t i v e l y s t a b l e d u r i n g a l l t h e t i m e - s e r i e s e x p e r i m e n t s . N i t r a t e , on t h e o t h e r h a n d , g r a d u a l l y i n c r e a s e d i n c o n c e n t r a t i o n d u r i n g t h e t i m e - s e r i e s e x p e r i m e n t s , p a r t i c u l a r l y a t t h e h i g h e r t e m p e r a t u r e s . The g r a d u a l i n c r e a s e i n n i t r a t e i s a n i n d i c a t i o n t h a t n i t r i f y i n g b a c t e r i a a r e p r e s e n t i n t h e s y s t e m o x i d i z i n g ammonia t o n i t r a t e . T h i s o b s e r v e d e f f e c t o f t e m p e r a t u r e on b a c t e r i a l p o p u l a t i o n d y n a m i c s and r e a c t i o n k i n e t i c s i s w e l l d o c u m e n t e d f o r b i o l o g i c a l n i t r i f i c a t i o n (Meade 197 4 ; S p e e c e 1 9 7 3 ) . I t i s somewhat s u r - p r i s i n g t h a t n i t r i t e , w h i c h i s t h e i n t e r m e d i a t e p r o d u c t o f n i t r i f i c a t i o n , d i d n o t c h a n g e s i g n i f i a n t l y i n c o n c e n t r a t i o n p r i o r t o t h i s g r a d u a l n i t r a t e i n c r e a s e . One e x p l a n a t i o n o f t h i s o b s e r v a t i o n may be t h a t i n a w e l l a g e d r e c y c l e d s y s t e m , t h e r a t e o f ammonia o x i d a t i o n b y N i t r o s o m o n a s i s e q u a l t o t h e r a t e o f n i t r i t e o x i d a t i o n by N i t r o b a c t e r , r e s u l t i n g i n t y p i a l l y l o w n i t r i t e c o n c e n t r a t i o n s a n d m i n o r n i t r i t e v a r i a t i o n o v e r t i m e ( C o l t and A r m s t r o n g 1 9 8 1 ) . I t a p p e a r s a s t h o u g h t e m p e r a t u r e i s t h e c r i t i c a l f a c t o r i n c o n t r o l l i n g t h e c o n c e n t r a t i o n o f a l l t h r e e n i t r o g e n o u s compunds i n t h e h o l d i n g s y s t e m due t o i t s i n f l u e n c e on t h e m e t a b o l i c a c t i v i t i e s o f b o t h t h e l o b s t e r s a n d t h e h e t e r o t r o p h i c and a u t o t r o p h i c b a c t e r i a p r e s e n t . 7.4 S i l i c a Sand F i l t e r E x p e r i m e n t s T h i s s e r i e s o f e x p e r i m e n t s was c a r r i e d - o u t t o d e t e r m i n e 123 whether ammonia-N i n t h e l o b s t e r h o l d i n g water was b e i n g o x i d i z e d t o n i t r a t e - N t h r o u g h t h e p r o c e s s of n i t r i f i c a t i o n d u r i n g passage t h r o u g h t h e s i l i c a sand f i l t e r s . R e s u l t s from t h e t i m e - s e r i e s e x p e r i m e n t s i n d i c a t e t h a t n i t r i f i c a t i o n r a t e s i n t h e s i l i c a sand f i l t e r s a r e g e n e r a l l y v e r y low. At 7°C some v a r i a t i o n i n ammonia-N and n i t r a t e - N d i s c h a r g e c o n c e n t r a t i o n s was d e t e c t e d but no r e a l p a t t e r n of i n c r e a s i n g n i t r a t e - N and d e c r e a s i n g ammonia-N was o b s e r v e d . At the two h i g h e r t e m p e r a t u r e s l i m i t e d e v i d e n c e o f n i t r i f a t i o n e x i s t s . At t h e 30 min mark i n exp e r i m e n t 1 a t 12°C, l e v e l s o f n i t r a t e - N began t o i n c r e a s e i n the h o l d i n g water a f t e r passage t h r o u g h f i l t e r 2 and ammonia-N l e v e l s began t o drop a t a p p r o x i m a t e l y the same r a t e . A s i m i l a r p a t t e r n was o b s e r v e d i n f i l t e r 1 d u r i n g e x p e r i m e n t 2 a t 17°C. An e x p l a n a t i o n o f the v e r y low n i t r i f i c a t i o n o b s e r v e d may be found i n t h e c o n d i t i o n s under which t h e s i l i c a sand f i l t e r s o p e r a t e as compared t o t h o s e o u t l i n e d i n t h e l i t e r a t u r e f o r optimum b i o f i l t e r p e rformance. A number of f a c t o r s i n f l u e n c e t h e e f f i c i e n c y of a b i o l o g i c a l f i l t e r , i n c l u d i n g water t e m p e r a t u r e , p r e s e n c e o f t o x i c compounds i n the w a t e r , pH, the c o n c e n t r a t i o n of d i s s o l v e d oxygen, s a l i n i t y , s u r f a c e a r e a s o f the f i l t r a n t m a t e r i a l , h y d r a u l i c l o a d i n g ( f l o w per u n i t volume o f f i l t e r s s u r f a c e a r e a ) , t h e r e s i d e n c e t i m e of the h o l d i n g water i n the f i l t e r , ammonia l o a d i n g , and o r g a n i c l e v e l i n t h e r e c i r c u l a t i n g water ( P e t t i g r e w e t a l . 1978; S p o t t e 1 9 7 9 ) . The s t u d i e s of 124 S r n a a n d B a g g a l e y ( 1 9 7 5 ) showed t h a t i n s e a w a t e r a q u a r i a an i n c r e a s e o f 4°C i n c r e a s e d ammonia a n d n i t r i t e o x i d a t i o n b y 50% a n d 12% r e s p e c t i v e l y . R e d u c i n g t h e t e m p e r a t u r e 1°C s l o w e d t h e o x i d a t i o n r a t e o f ammonia by 3 0 % , and a 1.5°C d e c r e a s e s l o w e d t h e r a t e o f n i t r i t e o x i d a t i o n by 8%. I t i s g e n e r a l l y a c c e p t e d t h a t i n c r e a s e s i n t e m p e r a t u r e s p e e d up b i o c h e m i c a l a c t i v i t y a n d a l a g p e r i o d i s o f t e n e v i d e n t i n a b i o f i l t e r i f t e m p r a t u r e i s a l t e r e d a b r u p t l y . S i n c e no t o x i c compounds, s u c h a s b i o c i d e s , a r e p a s s e d t h r o u g h t h e h o l d i n g s y s t e m t h i s f a c t o r s h o u l d n o t h a v e i n f l u e n c e d n i t r i f i c a t i o n d u r i n g t h e s i l i c a s a n d f i l t e r e x p e r i m e n t s . S r n a a n d B a g g a l e y ( 1 9 7 5 ) d e t e r m i n e d t h a t m a r i n e n i t r i f i e r s u s e d i n t h e i r s t u d y p e r f o r m e d most e f f i c i e n t l y a t pH 7.45, w i t h a n e f f e c t i v e r a n g e o f 7.0-8.2. A c o n s i d e r a b l e amount o f o x y g e n i s r e q u i r e d f o r n i t r i f i c a t i o n . S t a n k e w i c h ( 1 9 7 2 ) c a l c u l a t e d t h e s t o i c h i o m e t r i c o x y e n r e q u i r e m e n t f o r t h e c o n v e r s i o n o f NH^-N t o NO^-N a s 3.43 kg o f o x y g e n « k g - 1 o f NH^-N o x i d i z e d . The NO~-N t o NO~-N c o n v e r s i o n r e q u i r e s 1.14 kg o f o x y g e n - k g " 1 o f NO^-N o x i d i z e d . A l t h o u g h many s p e c i e s o f n i t r i f y i n g b a c t e r i a c a n t o l e r a t e s a l t w a t e r a nd s a l i n i t y c h a n g e s , t h e r e i s some i n d i c a t i o n t h a t n i t r i f i c a t i o n p r o c e e d s more r a p i d l y i n f r e s h w a t e r t h a n i n s e a w a t e r s y s t e m s ( K a u a i e t a l . 1 9 6 4 ; K u h l and Mann 1 9 6 2 ) . A v a i l a b l e s u r f a c e a r e a i n a b i o f i l t e r i s a n i m p o r a n t f a c t o r i n e s t a b l i s h i n g an e f f e c t i v e c o m m u n i t y o f n i t r i f y i n g b a c t e r i a . A s s u m i n g t h a t o t h e r c o n d i t i o n s a r e o p t i m u m , i t i s g e n e r a l l y a c c e p t e d t h a t t h e g r e a t e r t h e s u r f a c e a r e a t h e g r e a t e r t h e 125 c o n c e n t r a t i o n o f b a c t e r i a . I n a b i o f i l t e r h o w e v e r , i n t e r s t i c i a l g a p s must be l a r g e e nough t o a l l o w f r e e p a s s a g e o f t h e f l u i d , i n c l u d i n g a n y o r g a n i c o r i n o r g a n i c p a r t i c u l a t e s . Ammonia r e m o v a l e f f i c i e n c y i n c r e a s e s l i n e a r l y a s t h e r e t e n t i o n t i m e i n c r e a s e s a n d a p p e a r s t o be i n d e p e n d e n t o f h y d r a u l i c l o a d when t h e l o a d i n g i s b e t w e e n 1.5 a n d 2.5 gpm ( 5 . 6 8 - 7.46 -1 -2 I ) , m i n . f t o f f i l t e r s u r f a c e a r e a ( L i a o a n d Mayo 1 9 7 4 ) . L i a o a n d Mayo (1974) a l s o s u g g e s t e d t h a t an ammonia l o a d i n g o f 2 x l 0 ~ 4 l b ( 9 x l 0 ~ 5 kg) NH^-N«ft" 2 o f s p e c i f i c medium s u r f a c e a r e a ' d a y - 1 be c o n s i d e r e d t h e u p p e r l i m i t f o r f i l t e r d e s i g n . F i n a l l y , a n y o r g a n i c m a t t e r p r e s e n t i n t h e r e c i r c u l a t i n g w a t e r e n c o u r a g e s t h e g r o w t h o f h e t e r o t r o p h i c b a c t e r i a w h i c h i n t u r n i n c r e a s e s o x y g e n demand and r e d u c e s ammonia o x i d a t i o n . B a s e d on t h e e v i d e n c e p r e s e n t e d a b o v e a number o f f a c t o r s a s s o c i a t e d w i t h s i l i c a s a n d f i l t e r s a c t t o r e d u c e t h e i r s u i t a b i l i t y as b i o f i l t e r s . F i r s t , a l t h o u g h c o u r s e s a n d i s u s e d a s t h e f i l t e r medium, t h e b a s i c r e a s o n f o r h a v i n g t h e f i l t e r s i n l i n e i s t o remove p a r t i c u l a t e m a t t e r . A l a r g e p o r t i o n o f t h i s m a t e r i a l i s o r g a n i c , a s p r e v i o u s l y d e t e r m i n e d , a n d a s s u c h h a s a n e g a t i v e i n f l u e n c e on n i t r i f i c a t i o n . S e c o n d , t h e r e n t e n - t i o n t i m e i n t h e s a n d f i l t e r s ( a p p r o x i m a t e l y 1 m i n ) i s s h o r t and may l i m i t t h e e f f e c t i v e n e s s o f t h e f i l t e r s i n r e m o v i n g ammonia. S a l i n i t y , d i s s o l v e d o x y g e n and pH w e r e a l l m a i n t a i n e d a t s u i t a b l e l e v e l s d u r i n g t h e e x p e r i m e n t s . A l t h o u g h i t i s a l m o s t c e r t a i n t h a t a l i m i t e d amount o f 126 n i t r i f i c a t i o n i s t a k i n g p l a c e i n t h e s a n d f i l t e r s , i t a p p e a r s t h a t t h e r e d u c t i o n i n t h e ammonia-N c o n c e n t r a t i o n s o b s e r v e d d u r i n g t h e t i m e - s e r i e s e x p e r i m e n t s i s t o a l a r g e e x t e n t a r e s u l t o f o x i d a t i o n by n i t r i f y i n g b a c t e r i a i n s u s p e n s i o n o r a n o t h e r ammonia r e m o v i n g p r o c e s s s u c h a s a i r s t r i p p i n g . I n t h e h o l d i n g s y s t e m ammonia r e m o v a l by e i t h e r p r o c e s s n e e d n o t be a c o n c e r n p r o v i d e d w a t e r t e m p e r a t u r e s a r e k e p t b e l o w 10°C. 7. 5 U l t r a v i o l e t S t e r i l i z e r E x p e r i m e n t s The u s e o f u l t r a v i o l e t (UV) i r r a d i a t i o n , a t o r n e a r 2537 A ° , f o r t h e d e s t r u c t i o n o f a q u a t i c p a t h o g e n i c o r g a n i s m s 30 jdra o r s m a l l e r h a s r e c e i v e d a c o n s i d e r a b l e amount o f a t t e n t i o n i n t h e p a s t . R e s u l t s p r e s e n t e d h e r e show t h a t t h e c o m b i n a t i o n o f p r e s s u r e f i l t r a t i o n a n d UV i r r a d i a t i o n i s e f f e c t i v e i n c o n t r o l l i n g b a c t e r i a l l e v e l s i n a l o b s t e r h o l d i n g s y s t e m . A l t h o u g h b a c k g r o u n d b a c t e r i a l c o n c e n t r a t i o n s d i d i n c r e a s e s l i g h t l y w i t h i n c r e a s i n g t e m p e r a t u r e t h e g e n e r a l l y c o n s i s t e n t l e v e l s i n t h e h o l d i n g t a n k i s a s t r o n g i n d i c a t i o n t h a t t h e s t e r i l i z a t i o n u n i t s a r e p r e f o r m i n g e f f e c t i v e l y . T h e s e r e s u l t s a r e i n a g r e e m e n t w i t h t h e work r e p o r t e d by B u r r o w s and Combs (1 9 6 8 ) who s t a t e d t h a t s t e r i l i z a t i o n o f f i s h h a t c h e r y w a t e r i s b e s t a c c o m p l i s h e d by a c o m b i n a t i o n o f p r e s s u r e f i l t r a t i o n a n d u l t r a v i o l e t r a d i a t i o n . H e r a l d e t a l . ( 1 9 7 1 ) u s e d UV i r r a d i a t i o n t o e f f e c t a 98% r e d u c t i o n o f b a c t e r i a i n a c l o s e d s e a w a t e r a q u a r i u m s y s t e m . S i m i l a r r e s u l t s w e r e o b t a i n e d by B u l l o c k a nd S t u c k e y ( 1 9 7 7 ) w o r k i n g o n f i v e g r a m - n e g a t i v e 127 b a c t e r i a p a t h o g e n i c t o f i s h e s . The i n f l u e n c e o f t e m p e r a t u r e was p a r t i c u a r l y e v i d e n t i n t h e a q u a r i u m s y s t e m . The e f f e c t o f t e m p e r a t u r e on b a c t e r i a p o p u l a t i o n d y n a m i c s i s w e l l d o c u m e n t e d . I n t h i s e x p e r i m e n t a d i r e c t r e l a t i o n s h i p b e t w e e n b a c t e r i a l g r o w t h r a t e a nd t e m p e r a - t u r e i n t h e n o n - i r r a d i a t e d a q u a r i u m w a t e r was e s t a b l i s h e d ( F i g u r e 3 7 ) . T h i s r e l a t i o n s h i p was d e r i v e d by e x a m i n i n g e a c h o f t h e b a c t e r i a l t i m e - s e r i e s c u r v e s ( F i g u r e s 32, 33, 34, 35) t o l o c a t e a l i n e a r s e g m e n t t h a t e x t e n d e d o v e r a two d a y p e r i o d w i t h i n t h e f i r s t two o r t h r e e d a y s o f e a c h e x p e r i m e n t . From t h e s e l i n e a r s e g m e n t s p o p u l a t i o n g r o w t h r a t e s w e r e c a l c u l a t e d and p l o t t e d a g a i n s t t h e i r r e s p e c t i v e t e m p e r a t u r e s . A l t h o u g h i t was b a s e d o n o n l y f o u r p o i n t s , a l i n e a r r e l a t i o n s h i p i s o b t a i n e d w i t h a s t r o n g c o r r e l a t i o n c o e f f i c i e n t o f 0.98 a n d a s l o p e o f 0.1155. T h i s r e l a t i o n s h i p was e v i d e n t i n i t i a l l y f r o m t h e r e s u l t s o f t h e b a c t e r i a l t i m e - s e r i e s e x p e r i m e n t s , a s i t t o o k a p p r o x i m a t e l y t w i c e a s l o n g f o r t h e b a c t e r i a l c o n c e n - t r a t i o n i n t h e a q u a r i u m t o r e a c h 1 0 ^ c o u n t s ^ m l - " 1 ' a t 7°C t h a n a t 17°C. An i n t e r m e d i a t e v a l u e o f 4-5 d a y s was o b t a i n e d a t 12°C. B a c t e r i a l c o n c e n t r a t i o n s i n t h e h o l d i n g t a n k s r e m a i n e d a t 2 -1 a p p r o x i m a t e l y 10 c o u n t s - m l t h r o u g h o u t t h e e x p e r i m e n t s . Hand (1977) r e p o r t e d t h a t t h e m a r i n e i n t a k e w a t e r u s e d i n a l o b s t e r c u l t u r i n g s y s t e m h a d a b a c t e r i a l c o u n t r a n g i n g f r o m 500-2000 c o u n t s p e r m l b u t t h e n o r m a l o p e r a t i n g l e v e l o f t h e s y s t e m , w h i c h i n c l u d e d a UV s t e r i l i z e r was a p p r o x i m a t e l y 20 128 Figure 37. E f f e c t of temperature on b a c t e r i a l growth rate i n the aquarium. Fates were calculated over a two day period from the l i n e a r segment of each b a c t e r i a l time-series graph. 129 c o u n t s p e r m l . The s l i g h t l y h i g h e r c o u n t s r e c o r d e d i n t h e s y s t e m p r e s e n t e d h e r e may be a r e s u l t o f a number o f f a c t o r s . A c c o r d i n g t o t h e s p e c i f i c a t i o n s e s t a b l i s h e d f o r t h e s y s t e m s u f f i c i e n t u l t r a v i o l e t e n e r g y i s t h e o r e t i c a l l y p r o d u c e d t o e f f e c t a 100% k i l l o f a l l b a c t e r i a l s p e c i e s t h a t may be p r e s e n t . The r a t e d o u t p u t o f t h e UV s y s t e m u n d e r i n v e s t i g a t i o n i s 60,000 /iWs.cm , c o m p a r e d t o a r e q u i r e d e n e r g y l e v e l o f b e t w e e n 3,000 and 30,000 ^Ws-cm f o r t h e e l i m i n a t i o n o f most b a c t e r i a l s p e c i e s ( P h i l l i p s a n d H a n e l 1 9 6 0 ; K e l l y 1 9 6 5 ; K o l l e r 1 9 6 5 ) . Thus i t w o u l d seem t h a t o t h e r f a c t o r s a r e i n f l u e n c i n g t h e e f f i c i e n c y o f t h e UV s y s t e m . The t h r e e most p r o b a b l e f a c t o r s a r e t h e c o n d i t i o n o f t h e UV l a m p s a n d q u a r t z s l e e v e s t h e f l o w r a t e o f t h e h o l d i n g w a t e r p a s t t h e l a m p s , and t h e t u r b i d i t y o f t h e w a t e r . S i n c e t h e UV l a m p s and q u a r t z s l e e v e s a r e s e r v i c e d r e g u l a r l y ( o n c e a y e a r a s p e r s p e c i f i c i a t i o n s ) by t h e h o l d i n g o p e r a t i o n m a n a g e r s , a n d t h e f l o w r a t e p a s t t h e l a m p s i s r e g u l a t e d by an a c c u r a t e v a l v e s y s t e m , i t i s most l i k e l y t h a t i n h i b i t i o n o f UV p e n e t r a t i o n i s a t t r i b u t a b l e t o s u s p e n d e d a n d d i s s o l v e d m a t e r i a l i n t h e h o l d i n g w a t e r . The t r a n s m i s s i o n o f l i g h t t h r o u g h w a t e r f o l l o w s t h e e x p o n e n t i a l r e l a t i o n s h i p : 130 w h e r e TR = t r a n s m i s s i o n (%) e = b a s e o f n a t u r a l l o g a r i t h m a = c o e f f i c i e n t o f a b s o r p t i o n ( c m - 1 ) d = d e p t h o f w a t e r (cm) RF = s u r f a c e r e f l e c t i o n s (0.02 f o r w a t e r ) S i n c e t h e a v e r a g e d i s t a n c e t h e UV r a d i a t i o n must t r a v e l i n a UV s t e r i l i z e r i s s m a l l ( a p p r o x i m a t e l y 10 cm) t h e c o e f f i c i e n t o f a b s o r p t i o n i s t h e s i g n i f i c a n t f a c t o r i n d e t e r m i n i n g t h e p e r c e n t b a c t e r i a l k i l l i n t h e s y s t e m . The c o e f f i c i e n t o f a b s o r p t i o n w i l l most l i k e l y v a r y s u b s t a n t i a l l y w i t h t h e f i l t e r i n g e f f i c i e n c y o f t h e s i l i c a s a n d f i l t e r s , w h i c h i n t u r n v a r i e s w i t h p a r t i c u l a t e l o a d i n t h e s y s t e m . D a t a p r e s e n t e d by R o l l e r ( 1 965) i n d i c a t e t h a t e v e n s m a l l i n c r e a s e s i n t h e c o e f f i c i e n t o f a b s o r p t i o n may h a v e a s i g n i f i c a n t e f f e c t on t h e amount o f UV l i g h t r e a c h i n g t h e m i c r o o r g a n i s m s i n t h e h o l d i n g w a t e r . An i n c r e a s e o f 0.01 i n t h e c o e f f i c i e n t o f a b s o r p t i o n f r o m 0.05 t o 0.06 t h r o u g h 10 cm o f w a t e r d r o p s t h e p e r c e n t UV l i g h t t r a n s m i s s i o n f r o m a p p r o x i m a t e l y 60 t o 5 5 % . I t i s d i f f i c u l t t o e s t i m a t e t h e c o e f f i c i e n t o f a b s o r p t i o n f o r t h e l o b s t e r h o l d i n g w a t e r s i n c e i t w i l l v a r y s h a r p l y w i t h l o b s t e r l o a d and f i l t e r i n g e f f i c i e n c y . K o l l e r ( 1 9 6 5 ) a s s i g n s an a b s o r p t i o n c o e f f i c i e n t o f 0.008 t o d i s t i l l e d w a t e r , a v a l u e much l o w e r t h a n one w o u l d e x p e c t f o r a mean c o e f f i c i e n t f o r l o b s t e r h o l d i n g w a t e r . E x c l u d i n g d i s t i l l e d w a t e r t h e l o w e s t c o e f f i c i e n t p r e s e n t e d by K o l l e r i s 0.02 w h i c h i s u n d o u b t e d l y l o w e r t h a n l o b s t e r h o l d i n g w a t e r . E v e n a t t h i s l o w c o e f f i c i e n t 131 o f a b s o r p t i o n UV l i g h t t r a n s m i s s i o n t h r o u g h 10 cm o f w a t e r i s r e d u c e d by a p p r o x i m a t e l y 15% t o 2 5 % . T h e s e f i g u r e s d e m o n s t r a t e t h e i m p o r t a n c e o f p a r t i c u l a t e r e m o v a l p r i o r t o UV f i l t r a t i o n a n d p r o v i d e a p o s s i b l e e x p l a n a t i o n f o r t h e s l i g h t l y h i g h e r t h a n e x p e c t e d b a c k g r o u n d l e v e l s o f b a c t e r i a i n t h e h o l d i n g s y s t e m . A l t h o u g h t h e two most s i g n i f i c a n t l o b s t e r d i s e a s e s f r o m a l i v e s t o r a g e p o i n t o f v i e w , G a f f k e m i a , a n d s h e l l d i s e a s e , a r e c a u s e d by b a c t e r i a (some f o r m s o f s h e l l d i s e a s e may be a r e s u l t o f f u n g a l i n f e c t i o n ) s e v e r a l o t h e r l e s s common d i s e a s e s s u c h a s L a g e n i d u m d i s e a s e a n d F u s a r i u m d i s e a s e a r e f u n g a l i n f e c t i o n s . R e s e a r c h i n d i c a t e s t h a t f u n g a l o r g a n i s m s a r e g e n e r a l l y more r e s i s t a n t t o UV i r r a d i a t i o n t h a n b a c t e r i a ( P h i l l i p s and H a n e l 1 9 6 0 ; K e l l y 1 9 6 5 ; R o l l e r 1965) and a s s u c h w o u l d be more l i k e l y t o s u r v i v e i n t h e h o l d i n g w a t e r . A l t h o u g h s p e c i f i c s t u d i e s w e r e n o t c o n d u c t e d on t h e p r e s e n c e o f f u n g u s a s s o c i a t e d w i t h t h e h o l d i n g w a t e r o r t h e l o b s t e r s t h e r e w e r e no c a s e s o f f u n g a l d i s e a s e s o b s e r v e d d u r i n g t h e p e r i o d o f g e n e r a l i n v e s t i g a t i o n o f t h e h o l d i n g f a c i l i t y . A t e s t f o r G a f f k e m i a i n a s h i p m e n t o f l o b s t e r s p r o d u c e d n e g a t i v e r e s u l t s . A t v a r i o u s t i m e s s h e l l d i s e a s e i s a m i n o r p r o b l e m i n t h e h o l d i n g s y s t e m b u t u s u a l l y v e r y f e w a n i m a l s a r e i n f e c t e d a n d due i n p a r t t o t h i s s t u d y , a p p r o p r i a t e m e a s u r e s , s u c h a s r e m o v i n g i n f e c t e d a n i m a l s and m a i n t a i n i n g p r o p e r l e v e l s o f f i l t r a t i o n a n d UV i r r a d i a t i o n , a r e t a k e n t o a b a t e t h e p r o b l e m . 132 8.0 SUMMARY AND CONCLUSIONS An o v e r v i e w o f t h e work r e p o r t e d h e r e r e v e a l s a number o f i m p o r t a n t f e a t u r e s a s s o c i a t e d w i t h w a t e r q u a l i t y i n a s h o r t - t e r m l o b s t e r h o l d i n g s y s t e m . F i r s t , a g o o d c o r r e l a t i o n was f o u n d b e t w e e n t h e COD a n d TOC c o n c e n t r a t i o n i n t h e l o b s t e r h o l d i n g w a t e r . S i n c e t h i s r e l a t i o n s h i p i s s t r o n g i t i s l i k e l y t h a t TOC i s a go o d m e a s u r e o f o x y g e n demand i n t h e s y s t e m . The e m p i r i c a l l y d e t e r m i n e d COD/TOC r a t i o o f 1.67 was shown t o be l o w e r t h a n p u b l i s h e d v a l u e s f o r a v a r i e t y o f w a s t e w a t e r s . T h i s may be due i n p a r t t o o r g a n i c compounds i n t h e h o l d i n g w a t e r t h a t a r e r e s i s t a n t t o t h e COD d i g e s t i o n . TOC m o n i t o r i n g d u r i n g t h e t i m e - s e r i e s e x p e r i m e n t s g e n e r a l l y showed a d r a m a t i c i n c r e a s e i n TOC c o n c e n t r a t i o n s i n t h e h o l d i n g w a t e r i m m e d i a t e l y a f t e r t h e l o b s t e r s w e re i n t r o d u c e d i n t o t h e t a n k s , f o l l o w e d by a r a p i d d e c l i n e 15 o r 30 m i n u t e s a f t e r t h e i n t r o d u c t i o n . I t was c o n c l u d e d t h a t t h e i n c r e a s e i n TOC was a c o m b i n e d r e s u l t o f l o b s t e r e x c r e t i o n , m a t e r i a l f r o m t h e e x t e r n a l s u r f a c e o f t h e a n i m a l s a n d r e s u s p e n s i o n o f e x i s t i n g o r g a n i c s . No r e l a t i o n s h i p was shown b e t w e e n TOC c o n c e n t r a t i o n a n d t e m p e r a t u r e b u t t h e r e d i d a p p e a r t o be a b i o m a s s e f f e c t . TOC i n c r e a s e s w e re n o t n e a r l y a s e x t r e m e when a s m a l l e r b i o m a s s was i n t r o d u c e d i n t o t h e s y s t e m . C a l c u l a t i o n s w i t h t h e TOC d a t a t o a p p r o x i m a t e a BOD e q u i v a l e n t showed t h a t o x y g e n demand o f t h e l o b s t e r h o l d i n g w a t e r was 4 t o 5 t i m e s h i g h e r t h a n l e v e l s i n f i s h h a t c h e r y w a t e r . T h e s e h i g h o x y g e n demand c o n d i t i o n s 133 a r e u n d u l y s t r e s s f u l t o l o b s t e r s f o r t w o r e a s o n s . F i r s t , r e d u c e d o x y g e n a v a i l a b i l i t y , a n d s e c o n d , a c o n s i d e r a b l e amount o f s m a l l s u s p e n d e d p a r t i c u l a t e m a t e r i a l i s a s s o c i a t e d w i t h t h e h i g h TOC l e v e l s w h i c h i n t e r f e r s w i t h t h e e x c h a n g e o f o x y g e n a c r o s s t h e s u r f a c e o f t h e g i l l s . D i s s o l v e d o x y g e n c o n c e n t r a t i o n s r e c o r d e d d u r i n g t h e t i m e - s e r i e s e x p e r i m e n t s n e v e r r e a c h e d p o t e n t i a l l y l e t h a l l e v e l s , n o r d i d any o f t h e o t h e r w a t e r q u a l i t y p a r a m e t e r s . A t t h e h i g h e r e x p e r i m e n t a l t e m p e r a t u r e s , o x y g e n l e v e l s i n t h e h o l d i n g w a t e r d i d d r o p t o a p o i n t t h a t c o u l d be c o n s i d e r e d c r i t i c a l when o t h e r p a r a m e t e r l e v e l s a r e l e s s t h a n o p t i m a l . T h i s s y n e r g i s t i c e f f e c t , a l t h o u g h i t may be s i g n i f i c a n t u n d e r c e r t a i n c o n d i t i o n s , i s v e r y d i f f i c u l t t o q u a n t i f y . A d r o p i n o x y g e n c o n c e n t r a t i o n was o b s e r v e d i m m e d i a t e l y a f t e r l o b s t e r i n t r o d u c t i o n , f o l l o w e d by a g r a d u a l i n c r e a s e t o a s t e a d y s t a t e c o n d i t i o n w h i c h was u s u a l l y l o w e r t h a n t h e i n i t i a l ( t i m e 0) c o n c e n t r a t i o n . A number o f f a c t o r s may a c t t o r e d u c e d i s s o l v e d o x y g e n i n t h e s y s t e m a t t h a t p o i n t , t h e most s i g n i f i c a n t b e i n g g e n e r a l o x y g e n demand f r o m t h e w a t e r a n d c o n s u m p t i o n by t h e l o b s t e r s . O xygen l e v e l s b e g a n t o i n c r e a s e s i g n i f i c a n t l y " d u r i n g t h e t i m e - s e r i e s e x p e r i m e n t s a f t e r l o b s t e r a c t i v i t y i n t h e h o l d i n g t a n k s s u b s i d e d . R a t e s o f o x y g e n c o n s u m p t i o n c a l c u l a t e d o v e r t h e f i r s t 15 m i n u t e s o f t h e e x p e r i m e n t s when c o m p a r e d t o p u b l i s h e d v a l u e s f o r l o b s t e r o x y g e n c o n s u m p t i o n showed s i m i l a r t r e n d s w i t h r e s p e c t t o t e m p e r a t u r e ( h i g h e r c o n s u m p t i o n a t h i g h e r t e m p e r a t u r e ) b u t t h e t i m e - s e r i e s r a t e s o f o x y g e n 134 consumption were a l l h i g h e r t h a n p u b l i s h e d v a l u e s . I t i s c o n c l u d e d t h a t the h i g h e r t i m e - s e r i e s l e v e l s a r e t h e r e s u l t of i n c r e a s e d oxygen consumption by the l o b s t e r s due t o s t r e s s and a p o s s i b l e h i g h oxygen demand of t h e water which was not a f a c t o r i n the e x p e r i m e n t s r e p o r t e d i n the l i t e r a t u r e . A l t h o u g h pH was never a problem d u r i n g t h e t i m e - s e r i e s e x p e r i m e n t s , changes i n t h i s parameter were i n d i c a t i v e of v a r i o u s b i o c h e m i c a l p r o c e s s e s . pH g e n e r a l l y i n c r e a s e d d u r i n g t h e e a r l y s t a g e o f t h e e x p e r i m e n t s due t o t h e i n f l u x o f ammonia. As ammonia l e v e l dropped so d i d pH due t o p r o c e s s e s such as n i t r i f i c a t i o n and a n i m a l r e s p i r a t i o n . Due t o the r e l a t i v e l y h i g h TOC l e v e l s t h a t appear i n t h e h o l d i n g water s h o r t l y a f t e r l o b s t e r i n t r o d u c t i o n , the b u f f e r i n g c a p a c i t y of the h o l d i n g water may have been a d v e r s e l y e f f e c t e d . O r g a n i c c a r b o n has t h e c a p a c i t y f o r r e d u c i n g the s o l u b i l i t y of sodium and c a l c i u m c a r b o n a t e s . Of t h e t h r e e n i t r o g e n o u s compounds m o n i t o r e d d u r i n g t h e t i m e - s e r i e s e x p e r i m e n t s o n l y ammonia and n i t r a t e showed s i g n i f i c a n t changes over t i m e . G e n e r a l l y , ammonia c o n c e n t r a t i o n s i n c r e a s e d r a p i d l y e a r l y i n t h e e x p e r i m e n t s a t t h e h i g h e r t e m p e r a t u r e s due t o l o b s t e r e x c r e t i o n and p o s s i b l y m i n e r a l i z a t i o n of o r g a n i c m a t e r i a l . A f t e r t h e i n i t i a l p e r i o d o f h i g h s t r e s s , t h e l o b s t e r s most l i k e l y reduced t h e i r ammonia e x c r e t i o n r a t e , and n i t r i f i c a t i o n and v o l a t i l i z a t i o n combined t o reduce l e v e l s o f t h i s compound even f u r t h e r . An ammonia p r o d u c t i o n r a t e c a l c u l a t e d from the change i n ammonia 135 c o n c e n t r a t i o n d u r i n g t h e f i r s t 30 m i n u t e s o f t h e t i m e - s e r i e s e x p e r i m e n t a t 19°C was h i g h e r t h a n a r a t e c a l c u l a t e d f o r j u v e n i l e c u l t u r e d l o b s t e r s a t 22°C. A number o f f a c t o r s c a n be s u g g e s t e d t o e x p l a i n t h i s d i f f e r e n c e b u t t h e h i g h s t r e s s c o n d i t i o n s u n d e r w h i c h t h e t i m e - s e r i e s l o b s t e r w e r e h e l d p r o b a b l y c o n t r i b u t e d most s i g n i f i c a n t l y t o t h e h i g h r a t e o f ammonia p r o d u c t i o n i n t h e s y s t e m . No s i g n i f i c a n t c h a n g e s i n n i t r i t e c o n c e n t r a t i o n s were o b s e r v e d d u r i n g any o f t h e t i m e - s e r i e s e x p e r i m e n t s . Any n i t r i t e p r o d u c e d d u r i n g t h e n i t r i f i c a t i o n p r o c e s s w i l l be o x i d i z e d t o n i t r a t e r a p i d l y t h u s m a i n t a i n i n g t h e c o n c e n t r a t i o n o f t h i s compound a t a r e l a t i v e l y c o n s i s t e n t l e v e l . The c o n c e n t r a t o n o f n i t r a t e was o b s e r v e d t o i n c r e a s e g r a d u a l l y d u r i n g t h e t i m e - s e r i e s e x p e r i m e n t s c o n d u c t e d a t t h e two h i g h e r t e m p e r a t u r e . I t i s c o n c l u d e d t h a t n i t r i f i c a t i o n was o c u r i n g a t a l o w b u t d e t e c t a b l e r a t e . No e v i d e n c e o f n i t r i f i c a t i o n r e s u l t e d f r o m e x p e r i m e n t s on t h e s i l i c a s a n d f i l t e r a t 7°C. A number o f f a c t o r s a r e r e s p o n s i b l e f o r t h i s , t h e most s i g n i f i c a n t b e i n g t h e l o w t e m p e r a t u r e a nd t h e h i g h f l o w r a t e t h r o u g h t h e f i l t e r s . A t 12 and 17°C n i t r i f i c a t i o n was e v i d e n t t o a l i m i t e d e x t e n t . The most i m p o r t a n t r e g u l a t i n g f a c t o r s a r e l i k e l y t h e h i g h f l o w r a t e a n d c o m p e t i t i o n f r o m h e t e r o t r o p h i c b a c t e r i a . A t t h e h i g h e r t e m p e r a t u r e s a s i g n i f i c a n t amount o f n i t r i f i c i a t i o n may be t a k i n g p l a c e i n s u s p e n s i o n . I t i s c o n c l u d e d t h a t t h e UV s t e r i l i z e r s y s t e m i s e f f e c t i v e a t c o n t r o l l i n g b a c t e r i a l p o p u l a t i o n s i n t h e h o l d i n g w a t e r , 136 a l t h o u g h l e v e l s d i d i n c r e a s e s l i g h t l y w i t h i n c r e a s i n g t e m p e r a - 2 -1 t u r e ( f r o m a p p r o x i m a t e l y 10 c o u n t s ^ m l a t 7°C t o a b o u t 2 5 -1 10 * c o u n t s . m l a t 1 7 ° C ) . I n t h e n o n - i r r a d i a t e d a q u a r i u m w a t e r b a c t e r i a l p o p u l a t i o n s w e re n o t c o n t r o l l e d a t any t e m p e r a - t u r e b u t g r o w t h r a t e s w e r e s i g n i f i c a n t l y s l o w e r a t l o w e r t e m p e r a t u r e s . S e v e r a l c o n c l u s i o n s c a n be d r a w n f r o m t h e work p r e s e n t e d h e r e . F i r s t , i t a p p e a r s t h a t s h o r t t e r m e f f e c t s o f l o b s t e r i n t r o d u c t i o n a r e t h e most d r a m a t i c a n d p r o b a b l y t h e most s i g n i f i c a n t . I n v i r t u a l l y a l l i n s t a n c e s , p a r t i c u l a r l y a t t h e h i g h e r t e m p e r a t u r e s , s u b s t a n t i a l d e t e r i o r a t i o n i n w a t e r q u a l i t y , a s m e a s u r e d b y t h e e s t a b l i s h e d w a t e r q u a l i t y p a r a m e t e r s , was o b s e r v e d d u r i n g t h e f i r s t 15-30 m i n u t e s a f t e r l o b s t e r i n t r o - d u c t i o n . C o n t i n u e d m o n i t o r i n g f o r a 12 h p e r i o d showed t h a t w a t e r q u a l i t y o f t e n i m p r o v e d d r a m a t i c a l l y a f t e r t h e 15 t o 30 m i n u t e s c r i t i c a l p e r i o d , b u t i n most c a s e s n e v e r r e a c h e d t h e l e v e l r e c o r d e d a t t i m e 0. T h e r e f o r e i t i s t h i s s h o r t c r i t i c a l p e r i o d t h a t i s l i k e l y t h e most s t r e s s f u l f o r t h e l o b s t e r s d u r i n g t h e h o l d i n g p e r i o d . Over t h e l o n g e r - t e r m , w a t e r q u a l i t y g e n e r a l l y i m p r o v e d t o a l e v e l a t w h i c h a n y e f f e c t s t h a t may h a v e r e s u l t e d w e r e i n s i g n i f i c a n t c o m p a r e d t o t h e s h o r t t e r m i m p a c t s . T h e s e l o n g t e r m e f f e c t s may be i m p o r t a n t i n c u l t u r i n g o p e r a t i o n b u t n o t i n a s h o r t t e r m h o l d i n g s y s t e m . A s e c o n d g e n e r a l c o n c l u s i o n i s t h a t t e m p e r a t u r e i s t h e most i m p o r t a n t f a c t o r c o n t r o l l i n g w a t e r q u a l i t y i n t h e l o b s t e r h o l d i n g s y s t e m . T e m p e r a t u r e d i r e c t l y e f f e c t s t h e o x y g e n c a r r y - 137 ing capacity of the water, and more importantly, the metabolic a c t i v i t y of the lobsters. Therefore, i f temperature can be kept low (6-8°C) most of the other important water q u a l i t y parameters can be maintained at suitable l e v e l s . Although temperature control appears to be the c r i t i c a l factor in the design of a successful short-term holding system for lobsters, i t i s important to monitor a wide range of water qu a l i t y parameters during operation of the system since low temperatures are not absolute insurance against deteriorating holding water q u a l i t y . F i n a l l y , although p o t e n t i a l l y l e t h a l l e v e l s were never reached for any parameter during the c r i t i c a l period estab- lished during the time-series experiments, combined e f f e c t s from several parameters at l e s s than acceptable l e v e l s may prove to be f a t a l to stress weakened lobsters. In fact t h i s r e s u l t was observed on a number of occasions during the experiments at the higher temperatures. A number of mortal- i t i e s occured which were apparently not a t t r i b u t a b l e to unacceptable l e v e l s of any one water q u a l i t y parameter. 138 9.0 RECOMMENDATIONS A number o f g e n e r a l r e c o m m e n d a t i o n s r e g a r d i n g t h e d e s i g n and m a i n t e n a n c e o f a s h o r t - t e r m l o b s t e r h o l d i n g s y s t e m c a n be p u t f o r t h b a s e d o n t h e r e s u l t s o f t h i s s t u d y . 1. T e m p e r a t u r e c o n t r o l i s a n e s s e n t i a l f e a t u r e i n h o l d i n g s y s t e m d e s i g n . The r e f r i g e r a t i o n u n i t i n c o r p o r a t e d i n t o t h e d e s i g n o f a n y h o l d i n g s y s t e m s h o u l d be c a p a b l e o f m a i n t a i n i n g t h e h o l d i n g w a t e r t e m p e r a t u r e a t b e t w e e n 6 and 8°C. 2. To h e l p c o n t r o l h i g h TOC a n d p a r t i c u l a t e l e v e l s i n t h e h o l d i n g w a t e r i m e d i a t e l y a f t e r l o b s t e r i n t r o d u c t i o n / t a n k s s h o u l d be vacuum c l e a n e d p r i o r t o t h e i n t r o d u c t i o n o f e a c h new s h i p m e n t . 3. D u r i n g t h e c r i t i c a l p e r i o d i m m e d i a t e l y a f t e r i n t r o d u c t i o n o f t h e l o b s t e r s , a d d i t i o n a l o x y g e n s h o u l d be a d d e d t o t h e h o l d i n g w a t e r t o s a t i s f y t h e h i g h demand t h a t o c c u r s a t t h i s t i m e . 4. To m i n i m i z e d e t e r i o r a t i o n o f w a t e r q u a l i t y upon l o b s t e r i n t r o d u c t i o n i t i s s u g g e s t e d t h a t t h e i n t r o d u c t i o n p r o c e s s t a k e p l a c e o v e r a p e r i o d o f a p p r o x i m a t e l y one h o u r . T h i s r e d u c t i o n i n i n t r o d u c t i o n r a t e w i l l h a v e a b u f f e r i n g e f f e c t 139 on w a t e r q u a l i t y i m p a c t s t h a t r e s u l t f r o m r a p i d i n t r o d u c t i o n o f l o b s t e r s i n t o a h o l d i n g s y s t e m . An a c c l i m a t i o n p e r i o d may be r e q u i r e d p r i o r t o i n t r o d u c t i o n o f l o b s t e r s i n t o a h o l d i n g s y s t e m i f t h e d i f f e r e n c e b e t w e e n t h e s h i p p i n g t e m p e r a t u r e and t h e h o l d i n g t e m p e r a t u r e i s g r e a t e r t h a n 5°C. S i n c e w a t e r q u a l i t y t e n d e d t o r e a c h a m o r e - o r - l e s s a c c e p t a b l e l o n g - t e r m s t e a d y - s t a t e l e v e l a f t e r t h e 30 m i n u t e c r i t i c a l p e r i o d , i t i s assumed t h a t t h e a d d i t i o n o f f r e s h s e a w a t e r a t t h e r a t e o f 10% o f t h e s y s t e m v o l u m e p e r d a y , was s u f f i c i e n t t o e n s u r e a c c e p t a b l e w a t e r q u a l i t y i n a l o b s t e r h o l d i n g s y s t e m o v e r a p e r i o d o f s e v e r a l w e e k s . D e p e n d i n g on a v a r i e t y o f f a c t o r s , s u c h a s w a t e r t e m p e r a t u r e a n d b u f f e r i n g c a p a c i t y , w a t e r q u a l i t y may d e t e r i o r a t e o v e r a l o n g e r p e r i o d , t h e r e f o r e i t i s recommended t h a t a c o m p l e t e w a t e r c h a n g e be c a r r i e d - o u t on t h e s y s t e m a p p r o x i m a t e l y o n c e e v e r y m onth. A l t h o u g h e a c h i n d i v i d u a l l o b s t e r h o l d i n g s y s t e m w i l l r e q u i r e u n i q u e d e s i g n f e a t u r e s t o d e a l w i t h s i t e s p e c i f i c w a t e r q u a l i t y p r o b l e m s , i t i s e s s e n t i a l t h a t a b a s i c w a t e r q u a l i t y m o n i t o r i n g programme be a p a r t o f t h e d a y - t o - d a y o p e r a t i o n o f a l l s y s t e m s . 140 10.0 LITERATURE CITED A l l e n , P. G. a n d W. E. J o h n s t o n . 1976. R e s e a r c h d i r e c t i o n a n d e c o n o m i c f e a s i l i b i t y : a n e x a m p l e o f s y s t e m s a n a l y s i s f o r l o b s t e r a q u a c u l t u r e . A q u a c u l t u r e 9:155-180. A m e r i c a n P u b l i c H e a l t h A s s o c i a t i o n . 1976. Compendium o f M e t h o d s f o r t h e M i c r o b i o l g i c a l E x a m i n a t i o n o f F o o d s . M.L. S p e c k , E d i t o r . A m e r i c a n P u b l i c H e a l t h A s s o c i a t i o n , W a s h i n g t o n , D.C. A m e r i c a n P u b l i c H e a l t h A s s o c i a t i o n , A m e r i c a n W a t e r Works A s s o c i a t i o n , a n d t h e W a t e r P o l l u t i o n C o n t r o l F e d e r a t i o n . 1976. S t a n d a r d m e t h o d s f o r t h e e x a m i n a t i o n o f w a t e r and w a s t e w a t e r , 1 4 t h e d . Am. Pub. H e a l t h A s s o c . , W a s h i n g t o n , D.C., 1193 p p . A r m s t r o n g , D. A. 1978. T o x i c i t y a n d m e t a b o l i s m o f n i t r o g e n compounds: E f f e c t s on s u r v i v a l , g r o w t h a n d o s m o r e g u l a t i o n o f t h e p r a w n M a c r o b r a c h i u m r o s e n b e r g i i . PhD. d i s s e r t a t i o n , E c o l o g y G r o u p , U n i v . C a l i f , D a v i s . 94 p p . A r m s t r o n g , D. A., D. C h i p p e n d a l e , A. W. K n i g h t and J . E. C o l t . 1 9 7 8 . I n t e r a c t i o n o f i o n i z e d a nd u n i o n i z e d ammonia on s h o r t - t e r m s u r v i v a l and g r o w t h o f p r a w n l a r v a l , M a c r o b r a c h i u m r o s e n b e r g i i . B i o l . B u l l . 1 5 4 : 1 5 - 3 1 . A y r e s , p.A. 1978. S h e l l f i s h P u r i f i c a t i o n . Lab L e a f l e t #38, M.A.F.F. D i r e c t F i s h . R e s . , L o w e s t o f t . 20 p p . A y r e s , p.A. a n d P.C. Wood. 1977. The L i v e S t o r a g e o f L o b s t e r . L a b L e a f l e t #37, M.A.F.F. D i r e c t . F i s h . R e s . , L o w e s t o f t . 9 p p . B a l l , I . R. 1 9 6 7 . The r e l a t i v e s u s c e p t i b i l i t y o f some s p e c i e s o f f r e s h w a t e r f i s h t o p o i s o n s - I . Ammonia .Water R e s . 1 ( 1 1 / 1 2 ) : 7 6 7 - 7 7 5 . B i n n s , R. a n d A. J . P e t e r s o n . 1969. N i t r o g e n e x c r e t i o n by t h e s p i n y l o b s t e r J a s u s e d w a r d s i i ( H u t t o n ) : The r o l e o f t h e a n t e r n a l g l a d . B i o l . B u l l . (Woods H o l e , M a s s . ) 1 3 6 : 1 4 7 - 1 5 3 . Bower, C. E. and J . P. B i d w e l l . 1978. I o n i z a t i o n o f ammonia i n s e a w a t e r : e f f e c t s o f t e m p e r a t u r e , pH, and s a l i n i t y . J . F i s h . R e s . B o a r d Can. 3 5 : 1 0 1 2 - 1 0 1 6 . B r o w n , C. M. a n d C. E. N a s h . 1 9 8 1 . A q u a c u l t u r e a s a method f o r m e e t i n g h a t c h e r y d i s c h a r g e s t a n d a r d s . P a g e s 183-189 i n L . J . A l l e n a n d E.C. K i n n e y e d s . P r o c e e d i n g s o f t h e B i o - E n g i n e e r i n g Symposium f o r F i s h C u l t u r e . F i s h C u l t u r e S e c t i o n , A m e r i c a n F i s h e r i e s S o c i e t y . T r a v e r s e C i t y , M i c h i g a n . 141 B u l l e y , N. R. a n d J . T. R. H u sdon. 1964. Use o f T o t a l o r g a n i c C a r b o n a n a l y s i s a s a m e a s u r e o f c a r b o n r e d u c t i o n d u r i n g a e r o b i c d i g e s t i o n o f s w i n e w a s t e . C a n a d i a n A g r i c u l t u r a l E n g i n e r i n g 1 6 : 2 - 5 . B u l l o c k , G.L., a n d H.M. S t u c k e y . 1977. U l t r a v i o l e t t r e a t m e n t o f w a t e r f o r d e s t r u c t i o n o f f i v e g r a m - n e g a t i v e b a c t e r i a p a t h o g e n i c t o f i s h e s . J . F i s h . R e s . B o a r d Can. 34 :1244-1249. B u r g e r , J . W. 1957. The g e n e r a l f o r m o f e x c r e t i o n i n t h e l o b s t e r , Homarus. B i o l . B u l l . (Woods H o l e , Mass.) 1 1 3 : 2 0 7 - 2 2 3 . B u r n s , E. R. a n d C. M a r s h a l l . 1965. C o r r e c t i o n f o r c h l o r i d e i n t e r f e r e n c e i n t h e C h e m i c a l O x y gen Demand T e s t . J . W a t e r P o l l . C o n t r o l F e d . 37:1716-17 21. B u r r o w s , R.E. and B.D. Combs. 1969. 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U n i v e r s i t y o f Rhode i s l a n d S ea G r a n t P u b l i c a t i o n 4 9. 32 pp. C o l t , T. a n d G. T c h o b a n o g l o u s . 1976. E v a l u a t i o n o f t h e s h o r t - t e r m t o x i c i t y o f n i t r o g e n o u s compounds t o c h a n n e l c a t f i s h , i c t a l u r u s p u n c t a t u s . A q u a c u l t u r e 8:209-224. C o l t , J . E . a n d D. A. A r m s t r o n g . 1 9 8 1 . N i t r o g e n t o x i c i t y t o c r u s t a c e a n s , f i s h a n d m o l l u s c s . P a g e s 34-47 i n L . J . A l l e n a n d E.C. K i n n e y e d s . P r o c e e d i n g s o f t h e B i o - E n g i n e e r i n g Symposium f o r F i s h C u l t u r e . F i s h C u l t u r e S e c t i o n , A m e r i c a n F i s h e r i e s S o c i e t y . T r a v e r s e C i t y , M i c h i g a n . C o r n i c k , J.W. and J . E. S t e w a r t . 1977. 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Aquacultural Engineering. Wiley Inter- science, Inc., New York, N.Y. Whit f i e l d , M. 1974. The hydrolysis of ammonia ions in sea water. A theopretical study. J. Mar. B i o l . Assoc. U.K. 54:565-580. Wickins, J. F. 1976. The tolerance of warm-water prawns to rec i r c u l a t e d water. Aquaculture 9:19-37. Wilder, D.G. 1953. Holding l i v e lobsters i n aerated, a r t i f i c i a l seawater. Fish. Res. Board Can., A t l . B i o l . Stn. Gen. Ser. C i r c . 21:4 p. Wilder, D.G. and D.W. McLeese. 1957. How temperature and crowd- ing e f f e c t the holding of lobsters i n a r t i f i c i a l seawater. Fish Res. Board Can., A t l . Prog. Rep. 66:24-25. Wood, P. C. and P. A. Ayres. 1977. A r t i f i c i a l Sea Water for S h e l l f i s h Tanks. Lab Leaflet #39, M.A.F.F. Dire c t . Fish. Res., Lowestorft. 11 pp. 150 11.0 APPENDICES A p p e n d i x 1. R e s u l t s o f TOC m o n i t o r i n g i n mg l - 1 , d u r i n g t h e t i m e - s e r i e s e x p e r i m e n t s • Time ( h r ) 1 2 3 4 E x p e r i m e n t # 5 6 7 8 9 10 0 18.1 36.1 25.4 101.1 26.5 57.6 25.1 34.5 19.5 48.4 0.25 204. 4 261.9 310. 3 241.9 195.6 224.3 164.0 461.7 241.4 385.4 0.50 222.3 244.3 305.7 256.8 220.4 386.5 89.3 489.8 304.5 405.1 0.75 316.1 201.7 272.8 268.6 231.5 291.6 43.7 451.9 276.7 261.7 1.0 184.9 179.8 224.7 287.5 222.6 205.7 45.9 369.4 201.4 144.1 1.5 170.2 185.6 206.4 256.1 191.3 211.8 38.5 195.7 169.9 139.7 2.0 166.6 164.5 139.3 224. 3 176.1 225.1 33.2 181.2 144.6 117.5 2.5 161.1 143.8 9 1 . 3 185.2 155.9 198.7 27.9 177.3 101.4 118.8 3.0 84. 4 150.2 82.9 168.7 138.4 169.4 36.5 186.4 93.3 105.9 4.0 86.4 138.7 84.1 177.8 115.6 181.7 39.4 169.5 56.5 102.6 5.0 42.8 121.4 78.5 191.2 96.4 176.4 32.9 171.7 50.1 112.1 6.0 46.3 126.3 75.4 176.4 105.1 144.9 40.1 168.1 52.4 101.9 7.0 36.7 109.9 74.3 165.1 86.7 150.1 35.7 156.9 46.5 98.5 8.0 38.9 84.1 83.7 154.6 101.9 139.3 29.9 159.4 56.7 87.6 9.0 50.1 77.6 80.2 149.8 68.9 127.4 34.8 145.6 60.9 94.5 10.0 40.5 89.8 75.1 159.6 76.4 115.5 39.4 151.3 55.4 96.8 11.0 38.0 81.3 81.5 150.1 56.8 111.2 36.8 149.6 52.3 101.6 12.0 36.1 75.7 78.9 145.9 6.1.5 104.1 39.1 141.8 48.5 89.4 A p p e n d i x 2 . R e s u l t s o f d i s s o l v e d o x y g e n m o n i t o r i n g , i n mg l " 1 , d u r i n g t h e t i m e - s e r i e s e x p e r i m e n t s . E x p e r i m e n t # Time ( h r ) 1 2 3 4 5 6 7 8 9 10 0 8.9 8.2 8.1 7.8 8.8 9.1 10.5 10.2 10.8 10.0 0.25 6.7 4.6 4.7 3.4 4.1 7.5 8.9 9.1 8.6 8.5 0.50 5.6 2.9 3.4 2.1 5.0 7.6 8.5 9.0 8.4 7.9 0.75 5.3 3.8 3.5 2.6 5.3 7.8 8.6 9.0 8.5 7.6 1.0 5.2 4.2 3.9 3.9 5.4 8.3 8.7 9.3 8.6 8.1 1.5 5.0 4.9 4.7 4.8 5.8 8.4 8.9 9.5 8.9 7.8 2.0 5.4 5.1 4.9 4.9 6.2 8.5 9.3 9.2 8.7 7.9 2.5 6.0 5.1 5.4 5.4 6.6 8.5 9.4 9.4 8.7 8.0 3.0 6.6 5.8 5.6 5.5 6.8 8.4 9.4 9.5 8.5 8.5 4.0 6.7 5.9 5.9 5.7 6.8 8.4 9.5 9.6 8.9 8.2 5.0 6.6 6.2 5.8 6.3 7.0 8.5 9.7 9.5 8.7 8.6 6.0 6.6 6.3 5.5 6.2 7.1 8.5 9.5 9.7 8.9 8.5 7.0 6.5 6.5 5.4 6.4 7.2 8.6 9.7 9.3 8.9 8.7 8.0 6.6 6.5 5.7 6.3 7.2 8.4 9.6 9.4 9.3 8.4 9.0 6.6 6.7 5.5 6.3 7.2 8.5 9.6 9.3 9.5 8.6 10.0 6.5 6.8 5.9 6.3 7.3 8.5 9.5 9.5 9.1 8.8 11.0 6.5 6.8 5.8 6.4 7.4 8.7 9.7 9.7 9.3 8.5 12.0 6.6 7.1 5.3 6.8 7.5 8.4 9.4 9.6 9.6 8.7 A p p e n d i x 3. R e s u l t s o f pH m o n i t o r i n g d u r i n g t h e t i m e - s e r i e s e x p e r i m e n t s . E x p e r i m e n t # Time ( h r ) 1 2 3 4 5 6 7 8 9 10 0 6.9 7.6 6.6 7.3 7.2 8.0 7.7 7.6 7.8 7.1 0.25 7.4 7.7 7.2 7.3 7.4 8.1 7.8 7.6 7.7 7.2 0.50 7.5 7.9 7.3 7.5 7.2 8.0 7.6 7.5 7.9 7.0 0.75 7.8 8.0 7.5 7.6 7.4 8.3 7.7 7.7 7.9 7.1 1.0 7.7 8.0 8.0 7.6 7.3 8.4 7.7 7.8 8.0 6.9 1.5 7.9 7.8 8.1 7.4 7.5 8.6 7.6 7.9 7.9 7.0 2.0 8.1 8.0 8.0 7.6 7.7 8.6 7.6 7.9 8.1 7.1 2.5 8.2 7.9 8.0 7.7 7.8 8.7 7.6 8.2 8.0 7.2 3.0 8.2 7.7 8.1 7.6 7.7' 8.5 7.8 8.3 8.1 7.1 4.0 7.7 7.9 7.9 7.6 7.7 8.4 7.7 8.5 8.1 7.1 5.0 7.6 7.8 7.7 7.5 7.8 8.3 7.8 8.4 8.2 7.2 6.0 7.8 7.9 7.9 7.6 7.6 8.4 7.9 8.5 8.0 7.3 7.0 7.5 7.7 7.8 7.7 7.7 8.5 7.6 8.5 8.0 7.3 8.0 7.7 7.7 7.7 7.5 7.6 8.4 7.8 8.4 7.9 7.1 9.0 7.5 7.6 7.7 7.5 7.6 8.4 7.7 8.1 8.0 7.0 10.0 7.4 7.6 7.6 7.4 7.7 8.3 7.7 8.0 7.8 7.3 11.0 7.4 7.4 7.7 7.4 7.5 8.4 7.6 8.1 7.8 7.2 12.0 7.3 7.5 7.6 7.4 7.5 8.4 7.8 7.8 7.7 7.3 to A p p e n d i x 4. R e s u l t s o f ammonia-N m o n i t o r i n g , i n mg l " 1 , d u r i n g t h e t i m e - s e r i e s e x p e r i m e n t s . E x p e r i m e n t # Time ( h r ) 1 2 3 4 5 6 7 8 9 10 0 3.3 4.8 1.9 1.7 3.6 2.3 4.8 6.8 4.7 1.3 0.25 4.5 4.7 3.3 1.8 3.4 2.7 4.9 6.5 5.0 1.6 0.50 5.4 4.9 3.9 2.3 3.5 2.9 5.0 6.7 4.9 2.0 0.75 5.3 5.6 4.2 2.4 3.7 3.0 4.7 6.9 5.3 2.2 1.0 5.9 5.8 4.4 2.8 3.9 2.9 4.9 7.4 5.2 2.0 1.5 6.4 6.0 4.7 3.2 4.0 3.4 5.1 7.5 5.6 2.3 2.0 6.0 6.8 4.8 3.1 4.1 3.7 5.0 7.9 5.8 2.1 2.5 5.9 6.7 5.0 3.3 3.8 3.9 4.9 8.1 5.6 1.9 3.0 6.3 7.1 4.7 3.0 3.8 4.1 5.6 8.3 5.9 2.0 4.0 4.9 6.9 4.8 3.2 4.0 4.2 5.0 8.0 5.7 1.8 5.0 4.6 6.8 5.0 3.1 3.5 4.0 5.1 8.5 6.0 2.1 6.0 4.8 6.4 4.6 3.0 3.7 4.2 4.8 8.6 5.7 2.0 7.0 4.3 6.3 4.5 3.1 3.8 3.9 4.7 8.4 5.5 1.9 8.0 4.4 6.2 4.3 2.8 3.4 4.2 4.9 8.0 5.7 1.9 9.0 4.0 6.2 4.2 2.7 3.6 3.8 4.5 8.3 5.3 1.8 10.0 3.9 6.3 4.0 3.0 3.7 3.9 4.7 8.1 5.2 1.7 11.0 4.1 6.1 3.9 2.5 3.3 3.6 4.9 7.9 5.1 1.8 12.0 4.3 5.9 3.7 2.3 3.5 3.7 4.8 7.6 4.8 1.7 Appendix 5. Results of n i t r i t e - N monitoring, i n mg l " 1 , during the time-series experiments. Experiment # Time (hr) 1 2 3 4 5 6 7 8 9 10 0 1 .33 0 . 1 9 1 .89 0 . 4 2 0 .16 1.66 0 .11 0 .24 0 . 3 6 0 . 4 5 0 . 2 5 1 .34 0 .24 2.11 0 .51 0 . 2 9 1.50 0 .21 0 .30 0 .61 0 .41 0 . 5 0 1 .34 0 .20 2 .34 0 .64 0 .21 1.61 0 .15 0 .41 0 . 4 8 0 . 4 8 0 . 7 5 1 .35 0. 21 1 .86 0 .48 0 .33 1.55 0 .22 0 .29 0 . 2 9 0 . 4 7 1.0 1 .38 0 . 4 3 2 .28 0 . 7 7 0 .25 1. 49 0 .09 0 .38 0 .44 0 . 5 4 1.5 1. 41 0 . 8 5 1 .77 0 .86 0 .47 1.50 0 .14 0 .44 0 .71 0 . 3 9 2 .0 1 .44 0 . 7 8 1 .59 0 .91 0 . 4 0 1.39 0 . 2 0 0 .52 0 . 7 8 0 .41 2.5 1 .49 0 .94 1 .56 1.04 0 .56 1.47 0 .18 0 .43 0 .68 0 .33 3 .0 1. 53 0 . 7 5 2 .35 1 .09 0 .61 1.36 0 .26 0 . 4 0 0 . 5 9 0 . 3 7 4 . 0 1 .56 0 .85 2 .14 1.07 0 . 4 7 1.45 0 .10 0. 51 0 . 5 0 0 .43 5 .0 1 .59 0 . 5 7 2. 22 0 . 7 9 0 .41 1.33 0 .16 0 .64 0 . 6 9 0 .39 6 .0 1 .57 0 .67 1.96 0 .84 0 .30 1.29 0 .24 0 .60 0 .82 0 .42 7.0 1 .55 0 .71 2 .05 0 .51 0 . 2 9 1. 20 0.31 0 . 5 9 0 . 7 5 0 . 4 7 8 .0 1 .52 0 . 6 9 1 .79 0 . 6 9 0 .30 1.31 0 .23 0 .70 0 . 7 9 0 . 5 0 9 .0 1 .48 0 . 8 0 1.91 0 . 7 5 0 .26 1.37 0 .15 0 .65 0 . 5 8 0 . 5 6 1 0 . 0 1 .45 0 .64 1 .83 0 .86 0 .39 1.25 0 . 1 9 0 .49 0 .51 0 . 5 8 1 1 . 0 1 .43 0 . 5 9 1 .64 0 .70 0 . 2 4 1.17 0 .22 0 .38 0 . 4 3 0 . 5 8 1 2 . 0 1 .42 0 .72 1 .59 0 .62 0 .32 1 .28 0 .17 0 .45 0 . 4 9 0 . 5 6 cn Appendix 6. Results of nitrate-N monitoring, in mg l " 1 , during the time- experiments. Experiment # Time (hr) 1 2 3 4 5 6 7 8 9 1 0 0 1 0 . 5 1 6 . 5 2 3 . 9 8 3 . 4 2 1 . 8 1 8 . 3 1 1 . 6 1 5 . 9 5 . 9 4 . 8 0 . 2 5 1 1 . 0 1 8 . 4 2 4 . 6 8 0 . 6 2 2 . 4 1 7 . 4 9 . 6 1 6 . 4 4 . 9 4 . 7 0 . 5 0 1 1 . 1 1 6 . 8 2 2 . 3 7 9 . 4 2 3 . 8 1 6 . 9 9 . 9 1 5 . 1 5 . 1 5 . 1 0 . 7 5 1 1 . 2 1 5 . 6 2 3 . 1 8 0 . 9 2 2 . 6 1 8 . 1 1 0 . 4 1 4 . 8 6 . 4 4 . 5 1 . 0 1 1 . 1 1 7 . 5 2 3 . 8 7 6 . 6 2 4 . 1 1 9 . 5 1 1 . 6 1 5 . 5 5 . 8 4 . 9 1 . 5 1 1 . 3 1 8 . 1 2 3 . 6 8 1 . 4 2 4 . 8 1 9 . 0 1 0 . 8 1 6 . 6 5 . 5 5 . 3 2 . 0 1 1 . 6 1 7 . 4 2 4 . 0 8 1 . 5 2 5 . 3 2 1 . 4 1 1 . 5 1 6 . 0 5 . 9 5 . 0 2 . 5 1 1 . 2 1 6 . 9 2 3 . 9 7 5 . 4 2 4 . 3 1 8 . 6 1 0 . 8 1 6 . 8 6 . 1 4 . 5 3 . 0 1 1 . 7 1 8 . 9 2 4 . 4 7 6 . 1 2 5 . 6 2 2 . 7 9 . 4 1 5 . 9 6 . 6 4 . 6 4 . 0 1 1 . 4 1 7 . 8 2 4 . 6 7 0 . 9 2 7 . 1 2 3 . 8 1 0 . 7 1 6 . 7 5 . 7 4 . 9 5 . 0 1 1 . 9 1 8 . 5 2 4 . 1 7 3 . 8 2 7 . 9 2 3 . 0 1 1 . 0 1 7 . 1 6 . 8 5 . 4 6 . 0 1 1 . 8 1 9 . 0 2 4 . 3 6 9 . 5 2 8 . 0 2 3 . 5 1 0 . 4 1 7 . 4 7 . 0 4 . 8 7 . 0 1 1 . 6 1 9 . 1 2 4 . 9 7 3 . 4 2 7 . 3 2 4 . 1 1 0 . 8 1 7 . 9 6 . 4 5 . 6 8 . 0 1 1 . 8 1 9 . 6 2 5 . 2 6 9 . 6 2 8 . 4 2 3 . 2 1 1 . 6 1 8 . 0 6 . 8 5 . 8 9 . 0 1 2 . 4 1 9 . 5 2 5 . 7 7 0 . 8 2 9 . 9 2 3 . 6 1 1 . 9 1 8 . 9 6 . 3 6 . 2 1 0 . 0 1 2 . 8 2 0 . 1 2 6 . 0 6 8 . 4 2 8 . 6 2 2 . 9 1 2 . 1 1 7 . 8 7 . 2 6 . 5 1 1 . 0 1 2 . 9 1 9 . 7 2 6 . 6 6 7 . 7 2 8 . 8 2 3 . 8 1 1 . 2 1 8 . 4 6 . 9 6 . 3 1 2 . 0 1 3 . 3 1 9 . 8 2 6 . 4 6 9 . 1 2 9 . 0 2 2 . 5 1 1 . 6 1 8 . 1 6 . 7 6 . 4 r-1 U l cn

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