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The effect of heat damage on the digestibility and nitrogen utilization of grass silages by sheep Pennells, Gerald Charles Louis 1984

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THE EFFECT OF HEAT DAMAGE ON THE DIGESTIBILITY AND NITROGEN UTILIZATION OF GRASS SILAGES BY SHEEP by - V'* , GERALD CHARLES LOUIS PENNELLS B.Sc, Edinburgh University, 1973 - 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 Animal Science We accept this thesis as conforming to the required standard THE UNIVERSITY OF BRITISH COLUMBIA April 1984 Q Gerald Charles Louis Pennells, 1984 In p r e s e n t i n g t h i s t h e s i s i n p a r t i a l f u l f i l m e n t of the requirements f o r an advanced degree at the U n i v e r s i t y o f B r i t i s h Columbia, I agree t h a t the L i b r a r y s h a l l make i t f r e e l y a v a i l a b l e f o r r e f e r e n c e and study. I f u r t h e r agree t h a t p e r m i s s i o n f o r e x t e n s i v e copying of t h i s t h e s i s f o r s c h o l a r l y purposes may be granted by the head o f my department o r by h i s or her r e p r e s e n t a t i v e s . I t i s understood t h a t copying o r p u b l i c a t i o n o f t h i s t h e s i s f o r f i n a n c i a l g a i n s h a l l not be allowed w i t h o u t my w r i t t e n p e r m i s s i o n . Department o f Animal Science The U n i v e r s i t y of B r i t i s h Columbia 1956 Main Mall Vancouver, Canada V6T 1Y3 Date May 3. 1984 - i i -ABSTRACT The e f f e c t s o f h e a t damage , e x p r e s s e d as t h e p e r c e n t a g e o f t o t a l n i t r o g e n l o c a t e d i n t h e a c i d - d e t e r g e n t f i b r e f r a c t i o n ( AD IN%N ) , on t h e d i g e s t i b i l i t y and n i t r o g e n u t i l i z a t i o n o f g r a s s s i l a g e s by s h e e p was i n v e s t i g a t e d u s i n g b o t h n a t u r a l l y h e a t e d ( E x p e r i m e n t 1) and a r t i f i c i a l l y h e a t e d ( E x p e r i m e n t 2) s a m p l e s . F o r E x p e r i m e n t 1, 6 l o c a l l y p r o d u c e d g r a s s s i l a g e s we re s e l e c t e d f o r t h e i r v a r y i n g l e v e l s o f h e a t damage e x p r e s s e d as ADIN%N: l e v e l s r a n g e d f r o m a l ow o f 4 . 8 % t o a h i g h o f 2 2 . 9 % . The a p p a r e n t d i g e s t i b i l i t y o f e a c h s i l a g e was m e a s u r e d i n d i g e s t i o n t r i a l s u s i n g 4 t o 6 s h e e p p e r s i l a g e and a 14 day a d a p t a t i o n p e r i o d f o l l o w e d by a 7 day t o t a l c o l l e c t i o n t r i a l . M e a s u r e s o f t o t a l n i t r o g e n ( N ) , a c i d - d e t e r g e n t f i b r e (ADF) and n e u t r a l -d e t e r g e n t f i b r e (NDF) as a p e r c e n t a g e o f s i l a g e d r y m a t t e r (DM) we re p o o r i n d i c a t o r s o f a p p a r e n t d i g e s t i b i l i t y . F o r e x a m p l e , s i l a g e s 3 and 6 had N, ADF, and NDF c o n t e n t s o f 1 . 9 , 4 4 . 4 , 6 6 . 3 and 2 . 5 , 4 3 . 4 , 5 9 . 6 % r e s p e c t i v e l y , w h e r e a s t h e a p p a r e n t d i g e s t i b i l i t y c o e f f i c i e n t s f o r d r y m a t t e r (DM) , o r g a n i c m a t t e r (OM) , N, ADF and NDF were 6 5 . 5 , 6 8 . 7 , 5 9 . 8 , 6 8 . 6 , 7 0 . 0 and 5 6 . 1 , 5 8 . 6 , 4 1 . 1 , 5 4 . 6 , 5 9 . 0 % r e s p e c -t i v e l y . V a r i a t i o n i n t h e l e v e l o f ADIN%N was a b l e t o a c c o u n t f o r 9 1 , 9 6 , 9 1 , 61 and 66% o f t h e v a r i a t i o n i n t h e a p p a r e n t d i g e s t i b i l i t y o f DM, OM, N, ADF and NDF r e s p e c t i v e l y . ADIN%N was s i g n i f i c a n t l y ( P < . 01 ) n e g a t i v e l y - i i i -c o r r e l a t e d w i t h t h e a p p a r e n t d i g e s t i b i l i t y o f DM ( - . 9 6 ) , OM ( - . 9 8 ) , N ( - . 9 5 ) , ADF ( - . 7 8 ) and NDF ( - . 8 1 ) . To r e d u c e t h e number o f v a r i a b l e s a s s o c i a t e d w i t h E x p e r i m e n t 1 and t o a s s e s s t h e f e a s i b i l i t y o f a r t i f i c i a l l y i n d u c i n g h e a t damage i n p r e v i o u s l y f e r m e n t e d s i l a g e , E x p e r i m e n t 2 was c o n d u c t e d u s i n g one l o c a l l y p r o d u c e d o r c h a r d g r a s s s i l a g e w h i c h was e i t h e r n o t h e a t e d ( t r e a t m e n t C ) , h e a t e d a t 49°C f o r 30 day s ( t r e a t m e n t H I ) , o r h e a t e d a t 50°C f o r 33 days f o l l o w e d by 30 day s a t 63°C ( t r e a t m e n t H 2 ) . The s i l a g e s we re s t o r e d i n 2 .2 by lm p a p e r f i b r e f o r m c y l i n d e r s w h i c h c o n t a i n e d c o i l e d h e a t i n g c a b l e s a t 5 t o 10cm s p a c i n g s a l o n g s i d e t h e p o l y e t h y l e n e bag w i t h i n s i l o s HI and H2. The o u t s i d e o f e a c h h e a t e d s i l o was i n s u l a t e d w i t h a 15cm t h i c k n e s s o f r o c k w o o l . T h i s me thod o f h e a t i n g w o r k e d w e l l w i t h t h e e x c e p t i o n o f l o c a l p e r f o r a t i o n o f t h e p o l y e t h y l e n e s e a l c a u s e d by d i r e c t c o n t a c t w i t h t h e h e a t i n g c a b l e s . T h i s p r o b l e m c o u l d be e l i m i n a t e d by c o i l i n g t h e c a b l e s a r o u n d t h e o u t s i d e o f t h e s i l o s . A r t i f i c i a l h e a t i n g had no e f f e c t on t o t a l N c o n t e n t , b u t c a u s e d a s i g n i f i c a n t ( P< . 01 ) i n c r e a s e i n ADIN%N l e v e l s f r o m 8 . 7 , t o 14 .1 and 2 4 . 3 % f o r t r e a t m e n t s C, HI and H2 r e s p e c t i v e l y . L i n k e d t o t h e r i s e i n ADIN%N l e v e l s was a s i g n i f i c a n t ( P < . 01 ) i n c r e a s e i n ADF c o n t e n t and a s i g n i f i c a n t ( P< . 01 ) d e c r e a s e i n h e m i c e l l u l o s e , a known s u b s t r a t e f o r t h e b r o w n i n g r e a c t i o n , w i t h i n c r e a s e d h e a t i n g . H e a t i n g e x p r e s s e d as ADIN%N s i g n i f i c a n t l y ( P< .01 ) d e c r e a s e d a p p a r e n t N d i g e s t i b i l i t y f r o m 6 2 . 9 , t o 5 7 . 2 and 3 9 . 1 % f o r t r e a t m e n t s C, HI and H2 r e s p e c t i v e l y . ADIN%N was s i g n i f i c a n t l y ( P < . 01 ) n e g a t i v e l y c o r r e l a t e d w i t h t h e a p p a r e n t d i g e s t i b i l i t y o f N ( r = - . 9 8 ) . V a r i a t i o n i n t h e l e v e l o f ADIN%N was a b l e t o a c c o u n t f o r 96% o f t h e v a r i a t i o n i n N d i g e s t i b i l i t y , b u t , i n c o n t r a s t t o E x p e r i m e n t 1, was a p o o r p r e d i c t o r f o r t h e o t h e r d i g e s t i b i l i t y c o e f f i c i e n t s . W i t h s i m i l a r N i n t a k e s , s h e e p w h i c h we re f e d t r e a t m e n t s HI and e x c r e t e d 17 t o 63% more N i n t h e i r f e c e s , b u t 8 t o 31% l e s s N i n t h e i r u r i n e , a s c o m p a r e d t o t r e a t m e n t C. ADIN%N c o n t e n t o f t h e s i l a g e s was s i g n i f i c a n t l y ( P< .01 ) c o r r e l a t e d w i t h b o t h t h e s e m e a s u r e s and e x p l a i n e d 96 and 66% o f t h e v a r i a t i o n i n f e c a l - N and u r i n a r y - N r e s p e c t i v e l y . R e d u c e d N d i g e s t i b i l i t y was c o m p e n s a t e d f o r by an i n c r e a s e i n t h e e f f i c i e n c y o f N r e t e n t i o n i n t r e a t m e n t HI c o m p a r e d t o C, so t h a t t h e r e was no d i f f e r e n c e i n o v e r a l l N r e t e n t i o n b e t w e e n t h e s e 2 t r e a t m e n t s ( . 9 4 g / d a y o f N f o r b o t h t r e a t -m e n t s ) . More s e v e r e h e a t i n g (H2) l e d t o a s i g n i f i c a n t ( P< . 05 ) r e d u c t i o n i n N r e t e n t i o n ( . 1 5 g / d a y o f N ) . The r e g r e s s i o n e q u a t i o n s f r o m E x p e r i m e n t s 1 and 2 f o r p r e d i c t i n g N d i g e s t i b i l i t y f r o m ADIN%N w e r e i n c l o s e a g r e e m e n t and t h e r e s u l t s we re p o o l e d t o o b t a i n t h e f o l 1 o w i ng : -D i g e s t i b i l i t y o f N (%) = 8 1 . 7 6 - 1 .83 (AD IN%N) ( r 2 = . 92 ) Taken t o g e t h e r , t h e s e r e s u l t s c o n f i r m t h a t ADIN%N m e a s u r e m e n t s s h o u l d be a p a r t o f any r o u t i n e a n a l y s i s o f g r a s s s i l a g e , p a r t i c u l a r l y when h e a t damage i s s u s p e c t e d . - V -TABLE OF CONTENTS Page A b s t r a c t i i T a b l e o f C o n t e n t s v L i s t o f T a b l e s v i i i L i s t o f F i g u re s x A c k n o w l e d g e m e n t s x i INTRODUCTION 1 REVIEW OF L ITERATURE 5 1. M e t h o d s o f d e t e c t i n g h e a t damage i n f o r a g e s a m p l e s 7 2 . F a c t o r s a f f e c t i n g t h e s u s c e p t i b i l i t y o f c r o p s t o h e a t damage 10 3. The e f f e c t s o f f e e d i n g h e a t damaged f o r a g e s on a n i m a l p e r f o r m a n c e 20 4 . S u r v e y s o f t h e o c c u r r e n c e o f h e a t damage among f o r a g e s 26 MATERIALS AND METHODS 31 1. I n t r o d u c t i o n 31 2 . E x p e r i m e n t 1 32 2.1 S i l a g e s 32 2 .2 A n i m a l s 33 2.3 D i g e s t i b i l i t y t r i a l s 34 2 . 4 S t a t i s t i c a l a n a l y s i s 35 3. E x p e r i m e n t 2 36 3.1 A c q u i s i t i o n o f e x p e r i m e n t a l s i l a g e . . . . 36 3.2 E x p e r i m e n t a l s i l o s 37 3.3 S i l a g e t e m p e r a t u r e 41 3.4 T r e a t e d s i l a g e 43 - vi -Page 3.5 Animals 43 3.6 D i g e s t i b i l i t y t r i a l s 45 3.7 Nitrogen balance 45 3.8 S t a t i s t i c a l analysis 46 RESULTS 47 1. Experiment 1 47 1.1 Silage composition 47 1.2 P a l a t a b i l i t y and intake 49 1.3 Apparent d i g e s t i b i l i t y 50 1.4 Correlations and regressions between apparent d i g e s t i b i l i t y measures and ADI N%N 51 2. Experiment 2 53 2.1 Silage temperature 53 2.2 Effects of heating on oven dry matter and ADIN%N levels 55 2.3 Silage composition 60 2.4 D i g e s t i b i l i t y t r i a l s 61 2.4.1 P a l a t a b i l i t y and intake 61 2.4.2 Ewe body weight change 62 2.4.3 Apparent d i g e s t i b i l i t y 64 2.4.4 Correlations between apparent d i g e s t i b i l i t y measures and ADIN%N 65 2.4.5 Nitrogen u t i l i z a t i o n 66 2.4.6 Regression equations 67 3. Regression equations based on pooled data . . . . 68 DISCUSSION 70 1. Experiment 1 70 - v i i -Page 2. Experiment 2 75 2.1 Experimental silage 75 2.2 Experimental s i l o s 75 2.3 Effects of heating on oven DM of sila g e . . 77 2.4 Effects of heating on silage composition . 78 2.5 Effects of heating on p a l a t a b i l i t y and intake 8 0 2.6 The effects of heating on apparent d i g e s t i b i l i t y c o e f f i c i e n t s 82 2.7 The effects of heating on nitrogen u t i l i z a t i o n 87 SUMMARY 8 9 CONCLUSIONS 9 4 BIBLIOGRAPHY 9 6 - v i i i L I S T OF TABLES T a b l e Page 1 S q u a r e d c o r r e l a t i o n c o e f f i c i e n t s ( r ) b e t w e e n ADIN%N (Y ) and v a r i o u s d i g e s t i b i l i t y c o e f f i c i e n t s (X ) 22 2 S q u a r e d c o r r e l a t i o n c o e f f i c i e n t s ( r ) b e t w e e n ADIN%DM (Y ) and v a r i o u s d i g e s t i b i l i t y c o e f f i c i e n t s (X ) 22 3 R e g r e s s i o n s o f t o t a l n i t r o g e n d i g e s t i o n c o e f f i c i e n t s ( Y ) on ADIN%N"(X) 23 4 P e r c e n t a g e o f h e a t damaged s a m p l e s i n e a c h s i l o t y p e f o r 146 h a y - c r o p s i l a g e s i n O n t a r i o 28 5 Mean ADIN%DM and DM l e v e l s f o r B . C . f o r a g e s c o n s e r v e d by d i f f e r e n t me thod s 29 6 H e a t damage e x p r e s s e d as ADIN%N f o r t h e s i x s i l a g e s u s e d i n E x p e r i m e n t 1 32 7 A v e r a g e t e m p e r a t u r e s and l e n g t h o f h e a t i n g p e r i o d f o r t h e 3 t r e a t m e n t s i n E x p e r i m e n t 2 42 8 C o m p o s i t i o n o f t h e s i x s i l a g e s u s e d i n E x p e r i m e n t 1 48 9 I n t a k e s and p a l a t a b i l i t y o f E x p e r i m e n t 1 s i l a g e s . . 49 10 A p p a r e n t d i g e s t i b i l i t y o f E x p e r i m e n t 1 s i l a g e s . . . 50 11 C o r r e l a t i o n s b e t w e e n ADIN%N and v a r i o u s d i g e s t i -b i l i t y c o e f f i c i e n t s f o r E x p e r i m e n t 1 s i l a g e s . . . . 51 12 R e g r e s s i o n s o f v a r i o u s d i g e s t i o n c o e f f i c i e n t s (Y ) on ADI N%N (X ) " 52 13 A v e r a g e t e m p e r a t u r e s and l e n g t h o f h e a t i n g p e r i o d f o r t r e a t m e n t s HI and H2 53 14 Range o f o v e n DM and ADIN%N i n s i l o s HI and H2. . . 55 15 Oven DM and ADI N%N l e v e l s i n s i l o C 55 16 C o m p o s i t i o n o f t h e 3 s i l a g e s u s e d i n E x p e r i m e n t 2 . 60 - i x -T a b l e Page 17 I n t a k e and p a l a t a b i l i t y o f E x p e r i m e n t 2 s i l a g e s . . 62 18 A p p a r e n t d i g e s t i b i l i t y c o e f f i c i e n t s f o r t h e t h r e e t r e a t m e n t s i n E x p e r i m e n t 2 64 19 C o r r e l a t i o n s b e t w e e n ADINXN and v a r i o u s d i g e s t i -b i l i t y c o e f f i c i e n t s f o r E x p e r i m e n t 2 s i l a g e s . . . . 65 20 E f f e c t s o f E x p e r i m e n t 2 t r e a t m e n t s on n i t r o g e n u t i l i z a t i o n 66 21 R e g r e s s i o n s o f v a r i o u s d i g e s t i o n c o e f f i c i e n t s and m e a s u r e s o f n i t r o g e n u t i l i z a t i o n (Y) on ADIN%N (X) 67 22 R e g r e s s i o n s o f v a r i o u s d i g e s t i o n c o e f f i c i e n t s (Y) f r o m E x p e r i m e n t s 1 and 2 on ADIN%N (X) 68 23 R e g r e s s i o n s o f DM d i g e s t i b i l i t y c o e f f i c i e n t s (Y) o n ADI N%N (X) 71 \ L I S T OF FIGURES F i g u r e Pag 1 E f f e c t o f t e m p e r a t u r e on t h e e x t e n t o f b r o w n i n g a s s a y e d as ADIN%N. lOg s a m p l e s o f o r c h a r d g r a s s were h e a t e d i n a 10ml f l a s k f o r 24 h o u r s a t 53% m o i s t u r e c o n t e n t 11 2 The e f f e c t o f l e n g t h o f h e a t e x p o s u r e a t 80°C on 2g s a m p l e s o f o r c h a r d g r a s s and a l f a l f a a t 53% m o i s t u r e c o n t e n t i n a 10ml f l a s k 12 3 Changes i n ADIN when 47% DM a l f a l f a was e n s i l e d a t one o f 3 t e m p e r a t u r e l e v e l s e i t h e r w i t h o u t a i r e x p o s u r e o r a t one o f 3 r a t e s o f a e r a t i o n . . . 18 4 C r o s s - s e c t i o n t h r o u g h one o f t h e h e a t e d s i l o s . . . 38 5 F i b r e f o r m c y l i n d e r , p o l y p r o p y l e n e mesh and e l e c t r i c h e a t i n g c a b l e u s e d i n t r e a t m e n t s HI and H2 39 6 T r e a t m e n t s HI and H2 s h o w i n g o u t e r i n s u l a t i n g l a y e r o f r o c k w o o l and t h e a u t o - t e m p e r a t u r e r e c o r d e r c o n n e c t e d t o t h e t h e r m o c o u p l e s 40 7 T e m p e r a t u r e s r e c o r d e d i n t h e o u t e r l a y e r and c o r e o f s i l o s HI and H2 54 8 S i l o H I : o v e n d r y m a t t e r a t v a r i o u s l o c a t i o n s . . . 56 9 S i l o H2: o ven d r y m a t t e r a t v a r i o u s l o c a t i o n s . . . 57 10 S i l o H I : ADIN%N a t v a r i o u s l o c a t i o n s 58 11 S i l o H2: ADI N%N a t v a r i o u s l o c a t i o n s . 59 12 Ewe body w e i g h t c h a n g e d u r i n g E x p e r i m e n t 2 63 13 R e g r e s s i o n s o f t o t a l n i t r o g e n d i g e s t i o n c o e f f -i c i e n t s ( Y ) on ADIN%N (X ) f r o m v a r i o u s s o u r c e s . . . 72 14 R e g r e s s i o n s o f DM d i g e s t i b i l i t y c o e f f i c i e n t s ( Y ) on ADI N%N (X ) f r o m two s o u r c e s 73 15 R e g r e s s i o n s o f t o t a l n i t r o g e n d i g e s t i o n c o e f f -i c i e n t s ( Y ) on ADIN%N (X ) f r o m v a r i o u s s o u r c e s . . . 86 - x i -ACKNOWLEDGEMENTS The a u t h o r w o u l d l i k e t o t a k e t h i s o p p o r t u n i t y t o a c k n o w l e d g e t h e e f f o r t s o f t h o s e p e o p l e who were o f p r i m e i m p o r t a n c e t o t h e r e s e a r c h and p r e p a r a t i o n o f t h i s t h e s i s . In p a r t i c u l a r , I s h o u l d l i k e t o e x p r e s s my s i n c e r e t h a n k s t o D r . J . A . S h e l f o r d , D e p a r t m e n t o f A n i m a l S c i e n c e U . B . C , u n d e r whose s u p e r v i s i o n t h i s work was done and whose u n f a i l i n g e n c o u r a g e m e n t and mos t p a t i e n t a s s i s t a n c e was g r e a t l y a p p r e c i a t e d . Thank s a r e a l s o due t o t h e members o f my r e s e a r c h c o m m i t t e e , e s p e c i a l l y D r . L . J . F i s h e r and D r . R.M. T a i t f o r t h e i r h e l p f u l a d v i c e and s u g g e s t i o n s . F i n a l l y , I s h o u l d l i k e t o t h a n k M r s . J e a n Thomas , D e p a r t m e n t o f A n i m a l S c i e n c e U . B . C , f o r h e r h e l p w i t h t h e p r e l i m i n a r y s e l e c t i o n o f t h e s i l a g e s and s u b s e q u e n t a n a l y t i c a l w o r k . - 1 -INTRODUCTION E n s i l a g e i s t h e a n a e r o b i c p r e s e r v a t i o n o f m o i s t f o r a g e by o r g a n i c a c i d s w h i c h a r e p r o d u c e d by t h e f e r m e n t a t i o n o f w a t e r - s o l u b l e c a r b o h y d r a t e s (ADAS , 1 9 7 4 ) . W h i l e t h e e n s i l a g e o f c r o p s has been p r a c t i s e d f o r a t l e a s t 3000 y e a r s ( S c h u k k i n g , 1 9 7 6 ) , i t i s o n l y d u r i n g t h e l a s t 20 y e a r s t h a t t h e d e t r i m e n t a l e f f e c t s o f o v e r h e a t i n g on t h e d i g e s t i b i l i t y o f t h e e n s i l e d mass have been b o t h i d e n t i f i e d and q u a n t i f i e d . T h i s i s p r o b a b l y as a r e s u l t o f t h e i n c r e a s e i n p o p u l a r i t y o f l o w m o i s t u r e s i l a g e s w i t h t h e i r g r e a t e r s u s c e p t i b i l i t y t o o v e r h e a t i n g . The w i n t e r f e e d i n g o f r u m i n a n t s i n B r i t i s h C o l u m b i a i s l a r g e l y b a s e d on c o n s e r v e d f o r a g e s . T r a d i t i o n a l l y t h i s has been m o s t l y i n t h e f o r m o f h a y . I t i s d i f f i c u l t , h o w e v e r , t o e n s u r e a l a r g e q u a n t i t y o f g o o d , w e l l made hay s i n c e t h e p r o c e s s i s d e p e n d e n t on s e v e r a l c o n s e c u t i v e days o f f i n e w e a t h e r t o d r y t h e c r o p p r i o r t o b a l i n g . A c o n s e r v a t i o n s y s t e m b a s e d on s i l a g e i s l e s s w e a t h e r -d e p e n d e n t and a l l o w s h i g h y i e l d i n g c r o p s t o be more e a s i l y c o n s e r v e d a t a s t a g e o f g r o w t h when f e e d i n g v a l u e i s h i g h (ADAS , 1 9 7 4 ) . In c o n s e q u e n c e , c r o p a c r e a g e s h a r v e s t e d as s i l a g e have i n c r e a s e d d r a m a t i c a l l y o v e r t h e p a s t 20 t o 25 y e a r s . A m a j o r v a r i a b l e i n t h e m a k i n g o f good s i l a g e i s m o i s t u r e c o n t e n t . H i g h m o i s t u r e s i 1 a g e s . a re p r o n e t o an u n d e s i r a b l e c l o s t r i d i a l t y p e o f f e r m e n t a t i o n , r e s u l t i n g i n l o w - 2 -d r y m a t t e r i n t a k e s , and t h e p r o d u c t i o n o f l a r g e q u a n t i t i e s o f p o l l u t i n g e f f l u e n t l e a d i n g t o e x c e s s i v e n u t r i e n t l o s s ( W a t s o n & N a s h , 1 9 6 0 ) . A c c o r d i n g t o P e l l e t i e r ( 1 9 7 3 ) , t o t a l d r y m a t t e r l o s s e s o f a t l e a s t 20% s h o u l d be e x p e c t e d i n s i l a g e s above 80% m o i s t u r e c o n t e n t . He goes on t o s t a t e t h a t : "As m o i s t u r e c o n t e n t i s d e c r e a s e d down t o t h e 50% r a n g e , t o t a l l o s s e s w i l l d e c r e a s e t o a min imum o f 2 t o 5% w h i c h r e s u l t s f r o m no s e e p a g e , no s p o i l a g e , and a v e r y l i m i t e d amount o f f e r m e n t a t i o n and r e s p i r a t i o n . " U n f o r t u n a t e l y , a h i g h d r y m a t t e r c r o p has a l ow d e n s i t y w h i c h makes i t d i f f i c u l t t o c o n s o l i d a t e . T h e r e f o r e , f a r m e r s t r y i n g t o p r o d u c e h i g h d r y m a t t e r s i l a g e u s i n g l e s s t h a n o p t i m a l s t o r a g e s i l o s a n d / o r p o o r f i l l i n g t e c h n i q u e s have e n c o u n t e r e d a new p r o b l e m o f e x c e s s i v e a i r i n f i l t r a t i o n c a u s i n g a e r o b i c d e t e r i o r a t i o n . T h i s d e t e r i o r a t i o n i n v o l v e s t h e o x i d a t i o n o f o r g a n i c a c i d s and w a t e r - s o l u b l e c a r b o h y d r a t e s by m i c r o - o r g a n i s m s w i t h t h e p r o d u c t i o n o f c a r b o n d i o x i d e , w a t e r a n d , most i m p o r t a n t l y , l a r g e q u a n t i t i e s o f h e a t ( M c D o n a l d , 1 9 8 1 ) . W i t h t h e c o m p l e t e o x i d a t i o n o f g l u c o s e , t h e t e m p e r a t u r e r i s e (T ) i n t h e s i l a g e m a s s , a s s u m i n g a s p e c i f i c h e a t c a p a c i t y o f 1 . 8 9 k J k g " l o C - 1 f o r t h e s i l a g e d r y m a t t e r and no l o s s o f h e a t t o t h e a t m o s p h e r e , i s g i v e n by t h e e x p r e s s i o n : -2 6 7 . 5 - 0 . 147D - 3 -w h e r e D i s d r y m a t t e r c o n t e n t ( g / k g ) and G i s t h e amount o f g l u c o s e ( g / k g _ 1 D M ) o x i d i s e d ( M c D o n a l d & W h i t t e n b u r y , 1 9 7 3 ) . F o r e x a m p l e , t h e p o t e n t i a l t e m p e r a t u r e r i s e t h r o u g h t h e c o m p l e t e o x i d a t i o n o f s u g a r s i n an i n s u l a t e d s i l a g e o f DM c o n t e n t 4 0 0 g / k g - 1 and s u g a r c o n t e n t ( a s g l u c o s e ) o f 1 0 0 g / k g _ 1 D M , w o u l d be 1 9 2 ° C . O b v i o u s l y t e m p e r a t u r e r i s e s o f t h i s •>. m a g n i t u d e n e v e r o c c u r i n p r a c t i c e b e c a u s e t h e r e s p i r i n g m i c r o - o r g a n i s m s w o u l d be k i l l e d w e l l b e l o w t h i s t e m p e r a t u r e and t h e h e a t p r o d u c e d t e n d s t o be d i s s i p a t e d i n t o t h e a t m o s p h e r e . In p r a c t i c e , t e m p e r a t u r e s b e t w e e n 50 and 6 0 ° C a r e r e a d i l y a t t a i n a b l e ( W a t s o n & N a s h , 1 9 6 0 ; W i e r i n g a e t a l . , 1 9 6 1 ) . F o r e x a m p l e , W i e r i n g a e t a l . ( 1 9 6 1 ) i n t h e i r s u r v e y o f 112 s i l a g e s on f a r m s i n t h e N e t h e r l a n d s f o u n d t h a t 27% o f a l l s a m p l e s r e c o r d e d a maximum t e m p e r a t u r e o f 5 1 - 6 0 ° C , w h i l e a f u r t h e r 17% r e a c h e d a maximum t e m p e r a t u r e g r e a t e r t h a n 6 0 ° C . They went on t o show t h a t h e a t i n g a t t e m p e r a t u r e s above 50°C s e r i o u s l y r e d u c e d t h e d i g e s t i b i l i t y o f c r u d e p r o t e i n . More r e c e n t l y , Yu and Thomas ( 1 9 7 5 ) w o r k i n g w i t h a l f a l f a h a y l a g e w i l t e d t o a b o u t 50% m o i s t u r e c o n t e n t and s t o r e d i n 3 by 6.1m c o n c r e t e s t a v e t o w e r s i l o s , f o u n d t h a t h e a t i n g , e x p r e s s e d as d e g r e e day s above 3 5 ° C , was s i g n i f i c a n t l y ( P < . 0 5 ) n e g a t i v e l y c o r r e l a t e d w i t h n i t r o g e n d i g e s t i b i l i t y ( r = - . 8 1 ) and n i t r o g e n b a l a n c e ( r = - . 4 9 ) . - 4 -The p u r p o s e o f t h e e x p e r i m e n t s d e s c r i b e d h e r e i n was t w o - f o l d . F i r s t , t o o b t a i n l o c a l l y - p r o d u c e d g r a s s s i l a g e s w i t h known amount s o f h e a t damage ( e x p r e s s e d as t h e amount o f n i t r o g e n i n t h e a c i d - d e t e r g e n t f i b r e f r a c t i o n as a p e r c e n t a g e o f t h e t o t a l n i t r o g e n i n t h e d r y m a t t e r o r ADIN%N) and r e l a t e t h i s h e a t damage t o t h e a p p a r e n t d i g e s t i b i l i t y by s h e e p o f v a r i o u s f r a c t i o n s i n t h e s i l a g e s . S e c o n d l y , t o i n v e s t i g a t e t h e f e a s i b i l i t y o f a r t i f i c i a l l y i n d u c i n g h e a t damage i n a p r e v i o u s l y f e r m e n t e d g r a s s s i l a g e and t h e n r e l a t i n g amount o f h e a t i n g t o t h e c h e m i c a l c o m p o s i t i o n , a p p a r e n t d i g e s t i b i l i t y and n i t r o g e n u t i l i z a t i o n o f t h i s s i l a g e . 0 - 5 -REVIEW OF L ITERATURE H e a t damage i n f o r a g e s can o f t e n be v i s u a l l y i d e n t i -f i e d by a d e f i n i t e d a r k e n i n g o r b r o w n i n g o f t h e s a m p l e ; h e n c e t h e f r e q u e n t s u b s t i t u t i o n o f t h e wo rd s " b r o w n i n g " o r " n o n - e n z y m i c b r o w n i n g r e a c t i o n " f o r h e a t damage. Hodge ( 1 9 5 3 ) i n h i s r e v i e w showed t h a t n o n - e n z y m i c b r o w n i n g i s an i n t r i c a t e s u b j e c t i n v o l v i n g a s e r i e s o f c o m p l e x d e g r a d a t i o n and r e c o m b i n a t i o n r e a c t i o n s s t a r t i n g w i t h a l d o s e s u g a r s and amino a c i d s i n an e q u i m o l a r r a t i o , and t e r m i n a t i n g w i t h t h e f o r m a t i o n o f u n s a t u r a t e d , c o l o u r e d p o l y m e r s o f v a r y i n g c o m p o s i t i o n . By u s i n g a g e n t s known t o e n h a n c e ( f u r f u r a l ) and i n h i b i t ( b i s u l p h i t e ) t h e r e a c t i o n , Van S o e s t ( 1 9 6 5 ) was a b l e t o p r o v e t h a t h e a t damage a r t i f i c i a l l y i n d u c e d i n f o r a g e s was c a u s e d by t h e n o n - e n z y m i c b r o w n i n g •. r e a c t i o n . M o i s t e n e d a i r - d r i e d f o r a g e was h e a t e d a l o n e , w i t h f u r f u r a l , w i t h s o d i u m m e t a b i s u l p h i t e , and w i t h s o d i u m m e t a b i s u l p h i t e and f u r f u r a l t o g e t h e r . The e x t e n t o f h e a t damage, e x p r e s s e d as p e r c e n t a g e o f a c i d - d e t e r g e n t i n s o l u b l e p r o t e i n was 4 . 8 , 1 0 . 9 , 1.8 and 2 . 6% f o r e a c h t r e a t m e n t r e s p e c t i v e l y . The i n i t i a l p h a s e o f h e a t damage c o n s i s t s o f M a i l l a r d r e a c t i o n s i n v o l v i n g , i n p a r t i c u l a r , am ino g r o u p s ( f o r e x a m p l e , o f l y s i n e ) and t h e a l d e h y d e g r o u p s o f r e d u c i n g s u g a r s ( K n i p f e l , 1 9 8 1 ) . T h i s r e a c t i o n i s r e v e r s i b l e by a c i d i n i t s e a r l y s t a g e s , b u t n o t by - 6 -e n z y m i c h y d r o l y s i s (Donoso e t a l . , 1 9 6 2 ) . W i t h i n c r e a s i n g s e v e r i t y o f h e a t i n g , t h e N - s u b s t i t u t e d g l y c o s y l a m i n e s p r o d u c e d by t h e M a i l l a r d r e a c t i o n s a r e c o n v e r t e d t h r o u g h a number o f c o m p l e x p a t h w a y s t o d a r k -c o l o u r e d m e l a n o i d i n s , w h i c h a r e brown n i t r o g e n o u s p o l y m e r s and c o - p o l y m e r s ( H o d g e , 1953 ; L i v i n g s t o n e t a l . , 1 9 7 1 ) . These m e l a n o i d i n s have been r e f e r r e d t o as " a r t i f a c t l i g n i n " (Van S o e s t , 1965) s i n c e t h e y a p p e a r i n t h e s u l p h u r i c a c i d - i n s o l u b l e o r a c i d - d e t e r g e n t f i b r e (ADF) p o r t i o n d u r i n g r o u t i n e f e e d a n a l y s i s . A r t i f a c t l i g n i n p r o d u c e d by h e a t damage can be d i s t i n g u i s h e d f r o m t r u e l i g n i n by i t s c o n s i d e r a b l y h i g h e r n i t r o g e n c o n t e n t ( 1 1 % v e r s u s 2%) (Van S o e s t , 1 9 8 2 ) . I t s o t h e r i m p o r t a n t c h a r a c t e r i s t i c i s i t s r e l a t i v e i n d i g e s t i b i 1 i t y w h i c h t h e r e f o r e l e a d s t o a r e d u c t i o n i n n i t r o g e n and e n e r g y d i g e s t i b i l i t y o f h e a t damaged f e e d s t u f f s ( G o e r i n g e t a l . , 1 9 7 2 ; Yu & Thomas , 1 9 7 5 ) . H i s t o r i c a l l y , h e a t i n g o r b r o w n i n g o f f o r a g e s was c o n s i d e r e d b e n e f i c i a l ( F r y , 1 8 8 5 ; Wa t son & N a s h , 1 9 6 0 ) , b o t h p r a c t i c a l l y b e c a u s e h i g h t e m p e r a t u r e s p r e v e n t an u n d e s i r a b l e f e r m e n t a t i o n , and a e s t h e t i c a l l y b e c a u s e o f t h e p l e a s a n t c a r a m e l i z e d o r " t o a s t e d " o d o u r t h a t a c c o m p a n i e s t h e b r o w n i n g r e a c t i o n . I n d e e d , as r e c e n t l y as 1 9 4 5 , B e c h t e l e t a l . ( 1 9 4 5 ) r e p o r t e d t h a t many Kan sa s f a r m e r s r e g a r d e d b rown h e a t e d hay " a s e q u a l , i f n o t s u p e r i o r , t o g r e e n hay f o r c a t t l e - f e e d i n g p u r p o s e s . " - 7 -U n e x p e c t e d l y p o o r r e s u l t s f r o m f e e d i n g b r o w n e d f o r a g e l e d r e s e a r c h w o r k e r s t o q u a n t i t a t i v e l y i n v e s t i g a t e t h e d i g e s t i b i l i t y o f h e a t e d and u n h e a t e d f e e d s t u f f s . F o r e x a m p l e , Wa t s on ( 1 9 3 9 ) , r e p o r t i n g on work c a r r i e d o u t a t J e a l l o t ' s H i l l i n 1 9 3 1 - 3 2 , f o u n d t h a t h e a t i n g d r a s t i c a l l y r e d u c e d t h e a p p a r e n t p r o t e i n d i g e s t i b i l i t y o f g r a s s s i l a g e . The t e m p e r a t u r e i n t h e s i l o r o s e t o 6 6 - 7 1 ° C , and i n one s p o t r e a c h e d 7 3 ° C . The a p p a r e n t p r o t e i n d i g e s t i b i l i t y o f t h e u p p e r l a y e r s , w h i c h had h e a t e d t h e m o s t , was 2 4 . 8 % , c o m p a r e d w i t h 3 8 . 7 % f o r t h e l o w e r l a y e r s and 70% f o r t h e f r e s h h e r b a g e . As t h e e v i d e n c e o f t h e d e t r i m e n t a l e f f e c t s o f h e a t i n g on f o r a g e d i g e s t i b i l i t y a c c u m u l a t e d , r e s e a r c h on t h e t o p i c c o n c e n t r a t e d on f o u r m a i n a r e a s : -1. Me thod s o f d e t e c t i n g h e a t damage i n f o r a g e s a m p l e s . 2 . F a c t o r s a f f e c t i n g t h e s u s c e p t i b i l i t y o f c r o p s t o h e a t damage. 3. The e f f e c t s o f f e e d i n g h e a t damaged f o r a g e s on a n i m a l p e r f o r m a n c e . 4 . S u r v e y s o f t h e o c c u r r e n c e o f h e a t damage among f o r a g e s . 1. M e t h o d s o f D e t e c t i n g H e a t Damage i n F o r a g e S a m p l e s A s i d e f r o m s l i g h t l y e l e v a t e d f i b r e l e v e l s , h e a t damage c a n n o t be d e t e c t e d by n o r m a l p r o x i m a t e a n a l y s i s ( B e c h t e l e t a l . , 1 9 4 3 , 1 9 4 5 ; G o e r i n g e t a l . , 1 9 7 4 ; H a r t m a n n , 1 9 7 3 ; Ma r sh & M u r d o c h , 1 9 7 5 ; M i l l e r e t a l . , 1967 ; Yu & V e i r a , 1977) . - 8 -S e v e r e h e a t damage can be q u a l i t a t i v e l y d e t e c t e d by t h e brown a p p e a r a n c e o f t h e f o r a g e s a m p l e . E a r l y r e s e a r c h t r i e d t o l i n k t h e i n t e n s i t y o f t h e brown c o l o u r a t i o n t o f e e d i n g v a l u e ( W a t s o n , 1 9 3 9 ) . In h i s r e v i e w Wa t s on showed t h a t b rown hay may r e s e m b l e n o r m a l hay i n c h e m i c a l c o m p o s i t i o n , b u t t h a t t h e b r o w n i n g p r o c e s s i s a s s o c i a t e d w i t h l o s s e s i n o r g a n i c m a t t e r and i n f e e d i n g v a l u e " p r o p o r t i o n a l t o t h e i n t e n s i t y o f t h e brown c o l o u r d e v e l o p e d by t h e h a y " ( W a t s o n , 1 9 3 9 ) . A more q u a n t i t a t i v e m e t h o d b a s e d on d e g r e e o f b rown c o l o u r a t i o n was d e s c r i b e d by Yu and Thomas ( 1 9 7 6 ) i n w h i c h t h e o p t i c a l d e n s i t y o f a p r e p a r e d f o r a g e s a m p l e was d e t e r m i n e d a t 440 m i l l i m i c r o n s . U s i n g t h i s t e c h n i q u e t h e y o b t a i n e d a s q u a r e d c o r r e l a t i o n c o e f f i c i e n t ( r ) b e t w e e n i n v i v o n i t r o g e n d i g e s t i b i l i t y and d e g r e e o f f o r m a t i o n o f brown c o l o u r o f 0 . 5 3 s u g g e s t i n g t h a t t h i s me thod had a low p r e d i c t i v e v a l u e . Mos t work on d e t e c t i n g h e a t damage and e s t i m a t i n g i t s e f f e c t s on f o r a g e d i g e s t i b i l i t y has c o n c e n t r a t e d on m e a s u r e m e n t s o f n i t r o g e n c o n t a i n e d i n t h e ADF p o r t i o n o f f e e d . Van S o e s t ( 1 9 6 5 ) showed t h a t e x p o s i n g f e e d s a m p l e s t o h i g h t e m p e r a t u r e s c a u s e d an i n c r e a s e i n ADF and t h a t t h i s ADF f r a c t i o n c o n t a i n e d an u n e x p e c t e d l y h i g h l e v e l o f n i t r o g e n . He s u g g e s t e d t h a t t h e n i t r o g e n c o n t e n t o f t h e ADF f r a c t i o n was " a s e n s i t i v e a s s a y f o r n o n - e n z y m i c b r o w n i n g due t o o v e r h e a t i n g o f f e e d s . " G o e r i n g e t a l . ( 1 9 7 2 ) i n v e s t i g a t e d t h e r e l a t i o n s h i p s o f b o t h p e p s i n - i n s o l u b l e n i t r o g e n ( P I N ) and a c i d - d e t e r g e n t - 9 -i n s o l u b l e n i t r o g e n (AD IN) w i t h n i t r o g e n and e n e r g y d i g e s t i b i l i t i e s o f 44 h e a t damaged f o r a g e s . Whe rea s t o t a l n i t r o g e n as a p e r c e n t a g e o f DM e x p l a i n e d o n l y 32% o f t h e v a r i a t i o n i n n i t r o g e n d i g e s t i b i l i t y , ADIN as a p e r c e n t a g e o f t o t a l n i t r o g e n (ADIN%N) and P IN as a p e r c e n t a g e o f t o t a l n i t r o g e n (P IN%N) e x p l a i n e d 86 and 83% o f t h e v a r i a t i o n r e s p e c t i v e l y . ADIN as a p e r c e n t a g e o f t h e DM (ADIN%DM) and P IN as a p e r c e n t a g e o f t h e DM (PIN%DM) e x p l a i n e d 81 and 63% o f t h e v a r i a t i o n i n e n e r g y d i g e s t i b i l i t y r e s p e c t i v e l y . Yu and Thomas ( 1 9 7 6 ) i n v e s t i g a t e d t h e p o t e n t i a l o f 30 l a b o r a t o r y p r o c e d u r e s t o p r e d i c t t h e f o r a g e q u a l i t y o f 21 hay o r h a y l a g e s a m p l e s o f known i n v i v o n i t r o g e n and DM d i g e s t i b i l i t i e s , and w i t h v a r y i n g d e g r e e s o f h e a t damage. T o t a l n i t r o g e n as a p e r c e n t a g e o f DM e x p l a i n e d o n l y 4% o f t h e v a r i a t i o n i n n i t r o g e n d i g e s t i b i l i t y , w h e r e a s ADIN%N and PIN%N e x p l a i n e d 86 and 79% o f t h e v a r i a t i o n r e s p e c t i v e l y . S h e l f o r d ( 1 9 8 0 ) i n r e v i e w i n g b o t h t h e s e s t u d i e s c o n c l u d e d t h a t "AD IN e x p r e s s e d as a p e r c e n t a g e o f t h e t o t a l n i t r o g e n i s most l i k e l y t h e b e s t me thod o f p r e d i c t i n g n i t r o g e n d i g e s t i o n c o e f f i c i e n t s f o r C o n s e r v e d f o r a g e s . " Shenk e t a l . ( 1 9 7 9 ) have had some s u c c e s s i n a s s e s s i n g h e a t damage by t h e use o f i n f r a r e d r e f l e c t a n c e t e c h n i q u e s . One h u n d r e d and e l e v e n s a m p l e s o f c o m m e r c i a l l y d e h y d r a t e d a l f a l f a showed m a r k e d d i f f e r e n c e s i n i n f r a r e d s p e c t r a b e t w e e n h e a t damaged and undamaged - 10 -s a m p l e s . The c a l i b r a t i o n e q u a t i o n s t h e y f o r m u l a t e d e x p l a i n e d 74 and 71% o f t h e v a r i a t i o n i n ADIN and PIN l e v e l s . They c o n c l u d e d t h a t i n f r a r e d r e f l e c t a n c e w o u l d be h e l p f u l i n d e t e c t i n g s a m p l e s w i t h h i g h o r l o w h e a t damage, b u t , a t p r e s e n t , i n t e r m e d i a t e v a l u e s c a u s e p rob1 ems. I n summary , t h e m a j o r i t y o f r e s e a r c h e r s who have i n v e s t i g a t e d h e a t damage i n f o r a g e s f a v o u r ADIN e x p r e s s e d as a p e r c e n t a g e o f e i t h e r DM (ADIN%DM) o r t o t a l n i t r o g e n (ADIN%N) as a m e a s u r e o f t h e d e g r e e o f h e a t damage o f a s a m p l e . G o e r i n g and Adams ( 1 973 ) s u g g e s t t h a t f o r a g e s w i t h a l e v e l o f ADIN%DM g r e a t e r t h a n 0 . 2 9 % a r e h e a t damaged . O t h e r w o r k e r s ( Van S o e s t , 1 9 6 5 ; Yu & Thomas , 1976) s u g g e s t t h a t n o r m a l u n h e a t e d f o r a g e s c o n t a i n up t o 7 -9% o f t h e i r t o t a i l n i t r o g e n i n t h e ADF f r a c t i o n , so t h a t f o r a g e s w i t h ADIN%N l e v e l s g r e a t e r t h a n 9% can be c o n s i d e r e d h e a t damaged (Yu & Thomas , 1 9 7 6 ) . 2 . F a c t o r s A f f e c t i n g t h e S u s c e p t i b i l i t y o f C r o p s t o H e a t Damage O b v i o u s l y t h e mo s t i m p o r t a n t f a c t o r a f f e c t i n g d e g r e e o f h e a t damage w i l l be t h e e x t e n t o f t h e h e a t i n g p r o c e s s b o t h i n t e r m s o f maximum t e m p e r a t u r e a t t a i n e d and l e n g t h o f e x p o s u r e . R e s u l t s can be c o n f o u n d e d , h o w e v e r , by m o i s t u r e c o n t e n t , pH , a e r a t i o n , and p l a n t s p e c i e s ( G o e r i n g e t a l . , 1 9 7 3 ; G o r d o n , 1 9 6 7 ; M i d d l e t o n & Thomas , 1 9 8 3 ; Y u , 1 9 7 6 ) . - 11 -G o e r i n g e t a l . ( 1 973 ) m e a s u r e d t h e f o r m a t i o n o f ADIN%N i n l O g s a m p l e s o f o r c h a r d g r a s s h e a t e d f o r 24 h o u r s a t 4 0 , 6 0 , 80 and 1 0 0 ° C . T h e i r r e s u l t s ( r e p r o d u c e d i n F i g u r e 1) showed t h a t ADIN%N i n c r e a s e d r a p i d l y as t e m p e r a t u r e s were i n c r e a s e d above 6 0 ° C f o r 24 h o u r s . They s u g g e s t e d t h a t l o w t e m p e r a t u r e s ( 4 0 - 6 0 ° C ) can c a u s e h e a t damage i f m a i n t a i n e d f o r l o n g p e r i o d s and t h e y r e c o g n i z e d t h a t 24 h o u r s i s a s m a l l f r a c t i o n o f t h e u s u a l s t o r a g e t i m e f o r s i l a g e . F i g u r e 1: E f f e c t o f t e m p e r a t u r e on t h e e x t e n t o f b r o w n i n g a s s a y e d as ADIN%N. l O g s a m p l e s o f o r c h a r d g r a s s were h e a t e d i n a 10ml f l a s k f o r 24 h o u r s a t 53% m o i s t u r e c o n t e n t . ( G o e r i n g e t a l . , 1 9 7 3 ) . 60h 40 3 soh e I" 20 10-INSOLUBLE NITROGEN _l_ 40 60 80 100 TEMPERATURE. °C In a f u r t h e r e x p e r i m e n t t o r e l a t e l e n g t h o f h e a t i n g p e r i o d t o ADIN f o r m a t i o n , G o e r i n g e t a l . ( 1 9 7 3 ) h e a t e d 2g s a m p l e s o f o r c h a r d g r a s s o r a l f a l f a a t 8 0°C f o r 0 , 2 , - 12 -4 , 8 , 1 6 , 32 and 70 h o u r s . T h e i r r e s u l t s a r e r e p r o d u c e d i n F i g u r e 2 . ADIN%N i n c r e a s e d w i t h t i m e f o r b o t h f o r a g e s , b u t t h e r a t e o f i n c r e a s e and t h e r e f o r e t o t a l amount f o r m e d was c o n s i d e r a b l y g r e a t e r f o r t h e o r c h a r d g r a s s s a m p l e , w h i c h i n d i c a t e d t h e i m p o r t a n c e o f f o r a g e s p e c i e s . F i g u r e 2 : The e f f e c t o f l e n g t h o f h e a t e x p o s u r e a t 80°C on 2g s a m p l e s o f o r c h a r d g r a s s and a l f a l f a a t 53% m o i s t u r e c o n t e n t i n a 10ml f l a s k . ( G o e r i ng e t a l . , 1973) . M i d d l e t o n and Thomas ( 1 9 8 3 ) e x p o s e d a l f a l f a - g r a s s h a y l a g e t o 3 0 , 3 8 , 5 0 , 55 o r 70°C f o r 7 o r 14 day p e r i o d s and t h e n m e a s u r e d ADIN%DM. ADIN%DM l e v e l s we re s i m i l a r i n s a m p l e s s t o r e d a t 30 t o 5 5 ° C . S a m p l e s s t o r e d a t 70°C f o r 7 day s had ADIN%DM l e v e l s a b o u t 3 t i m e s h i g h e r t h a n o f a l l s a m p l e s a t t h e l o w e r t e m p e r a t u r e s . T h i s r e s u l t - 13 -seemed t o c o n f i r m t h e e a r l i e r work o f G o e r i n g e t a l . ( 1 9 7 3 ) t h a t t h e r e was l i t t l e i n c r e a s e i n ADIN f o r m a t i o n a t t e m p e r a t u r e s b e l o w 6 0 ° C , a l t h o u g h a g a i n t h e r e s e a r c h e r s c o n c e d e d t h a t s h o r t d u r a t i o n h e a t i n g t r i a l s "may be o f l i m i t e d v a l u e i n d e t e r m i n i n g r e l a t i o n s h i p s b e t w e e n t e m p e r a t u r e and h e a t damage i n a s i l o , w h e r e i n c r e a s e d t e m p e r a t u r e s a r e m a i n t a i n e d f o r l o n g p e r i o d s " ( M i d d l e t o n & Thomas , 1 9 8 3 ) . ^ Yu ( 1 9 7 6 ) s t u d i e d t h e r e l a t i o n s h i p b e t w e e n v a r i o u s m e a s u r e m e n t s o f h e a t i n g and ADIN f o r m a t i o n i n f r e s h a l f a l f a , h a y l a g e and h a y . The r a n g e o f h e a t i n g t i m e was 6 - 4 8 h o u r s (6 h o u r i n c r e m e n t s ) and f o r t e m p e r a t u r e was 2 0 - 8 8 ° C . Maximum t e m p e r a t u r e e x p l a i n e d o n l y 36 t o 56% o f t h e v a r i a t i o n i n b o t h ADIN%N and ADIN%DM, a l t h o u g h h i s r e s u l t s s u g g e s t e d t h a t h e a t damage becomes d e f i n i t i v e a t 50°C o r a b o v e , and i s a l s o m e a s u r a b l e a t 3 5 ° C . M e a s u r e s o f l e n g t h o f h e a t i n g p e r i o d , e s p e c i a l l y h o u r s above 60 o r 7 0 ° C , were t h e b e s t p r e d i c t o r s o f b o t h ADIN%N and ADIN%DM ( r 2 f o r a l f a l f a h a y l a g e = 0 . 8 7 t o 0 . 9 2 ) . Work by Yu and Thomas ( 1 9 7 5 ) , who s t o r e d a l f a l f a h a y l a g e i n 3 by 12m c o n c r e t e s t a v e t o w e r s i l o s f o r 6 week p e r i o d s and m o n i t o r e d t e m p e r a t u r e a t d i f f e r e n t v e r t i c a l a r e a s w i t h i n t h e same s i l o , showed t h e i m p o r t a n c e o f l e n g t h o f h e a t i n g p e r i o d a t l o w e r t e m p e r a t u r e s . They r e p o r t e d t h a t ADIN i n s a m p l e s e x p o s e d t o 48°C f o r 6 weeks was a p p r o x i m a t e l y t w i c e t h a t o f h a y l a g e f r o m t h e same s i l o a t 33°C (ADIN%N l e v e l o f 1 5 . 4% a t 4 8 ° C and 8 . 7 % a t 3 3 ° C . ) - 14 -The n o n - e n z y m i c b r o w n i n g r e a c t i o n s i n v o l v e d i n h e a t damage a r e known t o r e q u i r e w a t e r s i n c e many o f them o c c u r o n l y i n a q u e o u s s o l u t i o n ( H o d g e , 1 9 5 3 ) . Van S o e s t ( 1 9 6 5 ) n o t e d t h a t t h e r a t e o f ADIN f o r m a t i o n i n d r y m a t e r i a l s was v e r y s l o w . E x p o s i n g o r c h a r d g r a s s -c l o v e r s a m p l e s o f 9% m o i s t u r e c o n t e n t t o 100°C f o r 20 h o u r s i n e i t h e r an open d i s h o r a c o v e r e d d i s h w i t h a d d e d w a t e r r e s u l t e d i n t h e p r o d u c t i o n o f a l m o s t f o u r t i m e s as much ADIN%DM i n t h e p r e s e n c e o f a d d e d w a t e r ( 1 . 4 9 v s . 0 . 3 5% ) . G o e r i n g e t a l . ( 1973) s t u d i e d t h e e f f e c t o f m o i s t u r e c o n t e n t on ADIN f o r m a t i o n w i t h o r c h a r d g r a s s ( a i r d r i e d ) h e a t e d a t 80°C f o r 24 o r 48 h o u r s . They c o n c l u d e d t h a t t h e g r e a t e s t s u s c e p t i b i l i t y a p p e a r e d t o be o v e r t h e r a n g e o f 20 t o 70% m o i s t u r e c o n t e n t . T h e s e f i n d i n g s we re i n a g r e e m e n t w i t h t h o s e o f M i d d l e t o n and Thomas ( 1 9 8 3 ) who r e p o r t e d t h a t s i l a g e m o i s t u r e c o n t e n t had r e l a t i v e l y l i t t l e e f f e c t on ADIN p r o d u c t i o n o v e r t h e r a n g e o f 4 1 - 7 5 % m o i s t u r e c o n t e n t . The l i m i t i n g f a c t o r s l o w i n g ADIN f o r m a t i o n a t m o i s t u r e l e v e l s a b o v e 75% i s u n l i k e l y t o be m o i s t u r e p e r se s i n c e b r o w n i n g c a n p r o c e e d r a p i d l y i n b o i l i n g w a t e r (Van S o e s t , 1 9 6 5 ) . S i n c e t h e g r e a t m a j o r i t y o f s i l a g e s f a l l w i t h i n t h e r a n g e o f 2 0 - 7 5 % f o r m o i s t u r e c o n t e n t , i t w o u l d a p p e a r t h a t a l l s h o u l d be e q u a l l y s u s c e p t i b l e t o h e a t damage. Many r e s e a r c h e r s , h o w e v e r , have l i n k e d i n c r e a s i n g l e v e l s - 15 -o f " o n - f a r m " h e a t damage w i t h t h e t r e n d t o w a r d s h i g h DM s i l a g e o r h a y l a g e ( G o e r i n g & Adams , 1 9 7 3 ; P i e r s o n e t a l . , 1 9 7 1 ; Thomas e t a l . , 1 9 7 2 ) . T h i s i s due t o t h e i n d i r e c t e f f e c t s o f h i g h DM l e v e l s on t h e h e a t i n g p r o c e s s . The se e f f e c t s may be s u m m a r i z e d as f o l l o w s : -a) H i g h DM s i l a g e s t e n d t o have u n d e r g o n e a l i m i t e d f e r m e n t a t i o n as m e a s u r e d by pH ( G o r d o n e t a l . , 1 9 6 5 ; M c D o n a l d e t a l . , 1 9 6 8 ) . The f e r m e n t a t i o n p r o c e s s i n v o l v e s t h e c o n v e r s i o n o f w a t e r - s o l u b l e c a r b o h y d r a t e s (WSC) t o o r g a n i c a c i d s . S i n c e t h e s e WSC a r e a m a j o r p a r t i c i p a n t i n t h e b r o w n i n g r e a c t i o n , a n y t h i n g w h i c h f a v o u r s t h e i r p r e s e r v a t i o n must a l s o i n d i r e c t l y f a v o u r t h e b r o w n i n g p r o c e s s . T h e r e f o r e , h i g h DM s i l a g e s w i t h t h e i r l i m i t e d f e r m e n t a t i o n w i l l t e n d t o have h i g h e r t h a n a v e r a g e l e v e l s o f WSC and so be more s u s c e p t i b l e t o h e a t damage. b) The s p e c i f i c h e a t o f s i l a g e d e c r e a s e s w i t h d e c r e a s i n g m o i s t u r e c o n t e n t . F o r e x a m p l e , M c D o n a l d and W h i t t e n b u r y ( 1 9 7 3 ) e s t i m a t e d t h e s p e c i f i c h e a t c a p a c i t y o f r y e g r a s s s i l a g e a t 80 and 50% m o i s t u r e c o n t e n t t o be 3 .73 and 3 . 0 4 k J / k g ~ l o C _ 1 r e s p e c t i v e l y . C o n s e q u e n t l y , a g i v e n amount o f h e a t p r o d u c e d w i l l i n c r e a s e t h e t e m p e r a t u r e o f a d r i e r s i l a g e more t h a n a w e t t e r o n e , and more damage may t h e r e f o r e o c c u r . c ) D r i e r s i l a g e s a r e d i f f i c u l t t o c o n s o l i d a t e . T h e r e f o r e h i g h DM s i l a g e s a r e p r o n e t o a i r i n f i l t r a t i o n and - 16 -a e r o b i c d e t e r i o r a t i o n w i t h i t s a s s o c i a t e d r e l e a s e o f l a r g e amount s o f h e a t e n e r g y ( M c D o n a l d & W h i t t e n b u r y , 1 9 7 3 ) . I t i s s u g g e s t e d t h a t t h e e x t e n t o f t h e b r o w n i n g r e a c t i o n i n c r e a s e s w i t h pH, b u t t h e r a t e i s d e c r e a s e d ( H o d g e , 1 9 5 3 ) . Work by G o e r i n g e t a l . ( 1 9 7 3 ) seemed t o c o n f i r m t h i s : when s a m p l e s we re e x p o s e d t o 80°C f o r 24 h o u r s , ADIN%N i n c r e a s e d ( f r o m an i n i t i a l 4 . 8 % t o 1 1 . 9 , 2 3 . 4 and 3 2 . 3 % ) w i t h pH ( 4 . 4 , 5.4 and 6.5 r e s p e c t i v e l y ) f o r s m a l l 2g s a m p l e s o f o r c h a r d g r a s s , b u t d e c r e a s e d ( f r o m 4 . 8 % t o 2 1 . 6 , 15 .6 and 13 . 8% ) w i t h t h e same r a n g e o f pH f o r l a r g e r 300g s a m p l e s . They s u g g e s t e d t h a t t h e s m a l l s a m p l e s q u i c k l y r e a c h e d h e a t t r a n s f e r e q u i l i b r i u m so t h a t t h e e x t e n t o f ADIN%N f o r m a t i o n was l i m i t e d by l o w p H , w h e r e a s t h e s l o w e r r a t e o f h e a t t r a n s f e r and t h e s l o w i n g o f t h e r e a c t i o n w i t h i n c r e a s e i n pH i n t h e l a r g e r s a m p l e s meant t h a t t h e l e n g t h o f h e a t i n g p e r i o d became t h e l i m i t i n g f a c t o r , so t h a t i f t h e s e s a m p l e s had been e x p o s e d t o h e a t i n g f o r l o n g e r p e r i o d s , ADIN%N p r e s u m a b l y w o u l d h a v e i n c r e a s e d w i t h pH. In p r a c t i c a l s i t u a t i o n s t h e e x t e n t o f t h e b r o w n i n g r e a c t i o n i s more i m p o r t a n t t h a n t h e r a t e b e c a u s e o f t h e l o n g s t o r a g e p e r i o d s a s s o c i a t e d w i t h s i l a g e . Go rdon ( 1 9 6 7 ) a n a e r o b i c a l l y s t o r e d f r e s h a l f a l f a and w e l 1 - f e r m e n t e d a l f a l f a s i l a g e a t 6 0 ° C f o r 4 w e e k s . Whereas h e a t damage was n o t e d i n t h e f r e s h a l f a l f a s a m p l e s (ADIN%N = 1 3 . 4 % ) , r e l a t i v e l y l i t t l e damage o c c u r r e d i n t h e w e l 1 - f e r m e n t e d , l ow pH a l f a l f a s i l a g e (ADIN%N = 8 . 6 % ) . - 17 -As w i t h m o i s t u r e c o n t e n t , pH need s t o be c o n s i d e r e d t o g e t h e r w i t h l e v e l o f WSC. Low pH i s a s s o c i a t e d w i t h a w e l 1 - f e r m e n t e d s i l a g e whe re most o f t h e WSC w i l l have been c o n v e r t e d t o o r g a n i c a c i d s . WSC, i n p a r t i c u l a r r e d u c i n g s u g a r s , a r e r e q u i r e d f o r t h e i n i t i a l p h a s e o f t h e b r o w n i n g r e a c t i o n ( K n i p f e l , 1 9 8 1 ) . H i g h pH i s i n d i c a t i v e o f l i t t l e f e r m e n t a t i o n and t h e r e f o r e h i g h l e v e l s o f WSC w h i c h i n c o n s e q u e n c e i n c r e a s e t h e p o t e n t i a l e x t e n t o f h e a t damage. A e r a t i o n has a v e r y i m p o r t a n t i n f l u e n c e on t h e e x t e n t o f h e a t damage b e c a u s e o f i t s e f f e c t s on h e a t p r o d u c t i o n w i t h i n t h e s i l o . The e n e r g y r e l e a s e d by t h e a e r o b i c r e s p i r a t i o n o f c a r b o h y d r a t e s i s r e s p o n s i b l e f o r much o f t h e r i s e i n t e m p e r a t u r e o f a s i l a g e . The maximum t e m p e r a t u r e r e a c h e d w i t h i n a s i l o w i l l d epend on t h e q u a n t i t y o f a i r p r e s e n t and t h e i n s u l a t i n g p r o p e r t i e s and s p e c i f i c h e a t o f t h e h e r b a g e mass ( M c D o n a l d , 1 9 8 1 ) . In a s e a l e d s i l o , a s s u m i n g no h e a t l o s s t o t h e e n v i r o n m e n t and a s p e c i f i c h e a t c a p a c i t y o f 1 . 8 9 k J / k g " * ° C " ' ' ' f o r f o r a g e DM, t h e r i s e i n t e m p e r a t u r e (T ) i n °C o f t h e s i l a g e i s g i v e n by : -T = 1.002Y 1 - 0 . 0 0 0 5 5 D w h e r e Y i s t h e a i r / h e r b a g e ( v / v ) r a t i o i n t h e s i l o and D i s t h e DM c o n t e n t ( g / k g - 1 ) ( M c D o n a l d & W h i t t e n b u r y , 1 9 7 3 ) . From t h e above e q u a t i o n i t c an be c a l c u l a t e d - 18 -t h a t when t h e a i r / h e r b a g e r a t i o i s 4 ( p o o r l y c o m p a c t e d ) , t h e t e m p e r a t u r e i n c r e a s e i n a 40% DM c r o p w o u l d be o n l y 5 . 1 ° C . Of f a r g r e a t e r c o n s e q u e n c e i n t h e g e n e r a t i o n o f e x t r e m e h i g h t e m p e r a t u r e s i n s i l o s i s a i r i n f i l t r a t i o n a f t e r e n s i l i n g b e c a u s e o f p o o r s e a l i n g . T h i s a i r i n f i l t r a t i o n i s p a r t i c u l a r l y s e r i o u s when t h e c r o p i s p o o r l y c o n s o l i d a t e d , w h i c h i s o f t e n t h e c a s e w i t h h i g h DM s i 1 a g e s . W i t h r e g a r d t o t h e e f f e c t s o f a e r a t i o n on t h e b r o w n i n g p r o c e s s i t s e l f , s e v e r a l w o r k e r s have r e p o r t e d t h a t a e r a t i o n i n c r e a s e s t h e r a t e o f ADIN f o r m a t i o n ( G o r d o n , 1 9 6 7 ; M i d d l e t o n & Thomas , 1 9 8 3 ) . In t h e l a t t e r s t u d y , 47% DM c h o p p e d a l f a l f a was p l a c e d i n g l a s s j a r s and i n c u b a t e d a t 3 8 , 55 o r 7 0 ° C . A i r was f o r c e d t h r o u g h t h e j a r s a t 0 , 1 0 , 15 o r 3 0 m l / m i n . The c o n t e n t s o f t h e j a r s w e r e r emoved a f t e r 14 day s o f t r e a t m e n t , a i r d r i e d and a n a l y z e d f o r ADIN%DM. T h e i r r e s u l t s ( F i g u r e 3) s u g g e s t e d t h a t a e r a t i o n had a much g r e a t e r i n f l u e n c e on ADIN f o r m a t i o n a t l o w e r t e m p e r a t u r e s . F i g u r e 3: Changes i n ADIN when 47% DM a l f a l f a was e n s i l e d a t one o f 3 t e m p e r a t u r e l e v e l s e i t h e r w i t h o u t a i r e x p o s u r e o r a t one o f 3 r a t e s o f a e r a t i o n . ( M i d d l e t o n & Thomas , 1 9 8 3 ) . o"* ^ ^ AIR FLOW, ml/min - 19 -G o e r i n g e t a l . ( 1 9 7 3 ) i n v e s t i g a t e d v a r i a t i o n s i n t h e s u s c e p t i b i l i t y t o b r o w n i n g o f 11 d i f f e r e n t s a m p l e s o f l a d i n o c l o v e r , a l f a l f a , b r o m e g r a s s and o r c h a r d g r a s s . F o r a g e s v a r i e d w i d e l y w i t h r e s p e c t t o amount o f i n i t i a l A D I N , p l a n t s p e c i e s , and t h e c o n t e n t o f t o t a l N. They w e r e , h o w e v e r , u n a b l e t o c o n s i s t e n t l y r e l a t e s u s c e p t i b i l i t y t o b r o w n i n g t o any o f t h e s e v a r i a b l e s . Two a l f a l f a s a m p l e s w e r e r e s i s t a n t t o b r o w n i n g and one was s u s c e p t i b l e . The a l f a l f a w i t h t h e g r e a t e s t s u s c e p t i b i l i t y had t h e l o w e s t t o t a l N c o n t e n t , w h i l e t h e b r o m e g r a s s mos t s u s c e p t i b l e had t h e h i g h e s t t o t a l N c o n t e n t . L a d i n o c l o v e r was t h e mos t s u s c e p t i b l e f o r a g e . They c o n c l u d e d t h a t t h e c o n s i d e r a b l e v a r i a t i o n i n s u s c e p t i b i l i t y t o b r o w n i n g shown by s i m i l a r f o r a g e s s u g g e s t e d " t h a t t h e r e a c t i o n i n f o r a g e s i s v e r y c o m p l e x and r e l a t e d , i n p a r t , t o d i f f e r e n c e s i n c o m p o s i t i o n n o t y e t m e a s u r e d o r r e c o g n i z e d as- i m p o r t a n t " ( G o e r i n g e t a l . , 1 9 7 3 ) . G o e r i n g and Adams ( 1 9 7 3 ) i n v e s t i g a t e d t h e f r e q u e n c y o f h e a t damage i n h a y - c r o p s i l a g e and c o r n s i l a g e i n P e n n s y l v a n i a . Wherea s 40% o f t h e 107 h a y - c r o p s i l a g e s we re h e a t damaged , none o f t h e 111 c o r n s i l a g e s e x h i b i t e d s i g n i f i c a n t h e a t damage. T h i s w o u l d s u g g e s t t h a t c o r n s i l a g e may be r e l a t i v e l y r e s i s t a n t t o h e a t damage , a l t h o u g h t h e s e r e s u l t s a r e q u i t e p r o b a b l y due t o t h e r a p i d s i l o f i l l i n g , more e ven chop l e a d i n g t o b e t t e r c o n s o l i d a t i o n , and l o w p r o t e i n c o n t e n t a s s o c i a t e d w i t h t h i s c r o p . - 20 -The f i n a l f a c t o r w h i c h has some i n f l u e n c e on t h e s u s c e p t i b i l i t y o f a f o r a g e t o b r o w n i n g i s t h e p r e s e r v a t i o n m e t h o d . R e s e a r c h s u g g e s t s ( Y u , 1976) t h a t s i l a g e i s more s u s c e p t i b l e t h a n h a y , p r o b a b l y b e c a u s e o f i t s h i g h e r m o i s t u r e c o n t e n t (Van S o e s t , 1 9 6 5 ; G o e r i n g e t a l . , 1 9 7 3 ) . F o r m a t i o n o f ADIN%N i n a l f a l f a h a y l a g e ( 4 2 % DM) was n i n e t i m e s f a s t e r t h a n i n a l f a l f a hay ( 89% DM) h e a t e d a t 88°C f o r 6 h o u r s ( Y u , 1 9 7 6 ) . A f t e r 48 h o u r s a t 8 8 ° C , ADIN%N o f t h e h a y l a g e had i n c r e a s e d f r o m 8 t o 3 8 % , w h i l e t h a t o f t h e hay had i n c r e a s e d f r o m 5 t o 27% . In summary , t h e t y p e o f f o r a g e mos t s u s c e p t i b l e t o h e a t damage w o u l d a p p e a r t o be a h i g h DM g r a s s , l e gume o r g r a s s / l e g u m e s i l a g e ( h a y l a g e ) , t h a t had u n d e r g o n e a l i m i t e d f e r m e n t a t i o n and t h e r e f o r e had a r e l a t i v e l y h i g h pH and WSC l e v e l , and t h a t was p o o r l y c o n s o l i d a t e d a n d / o r s t o r e d u n d e r c o n d i t i o n s w h i c h a l l o w e d a i r i n f i l t r a t i o n . 3. E f f e c t s o f F e e d i n g Hea t Damaged F o r a g e s on A n i m a l P e r f o r m a n c e S i n c e t h e b r o w n i n g r e a c t i o n i n v o l v e s t h e f o r m a t i o n o f i n d i g e s t i b l e p o l y m e r s w i t h a r e l a t i v e l y h i g h n i t r o g e n c o n t e n t , i t i s t o be e x p e c t e d t h a t n i t r o g e n u t i l i z a t i o n w i l l be t h e f a c t o r most s e v e r e l y a f f e c t e d by h e a t damage. Many r e s e a r c h e r s have shown t h a t b o t h a p p a r e n t n i t r o g e n d i g e s t i b i l i t y and a n i m a l p e r f o r m a n c e we re d e p r e s s e d when a n i m a l s were f e d h e a t damaged f o r a g e s ( B e c h t e l e t a l . , - 21 -1 9 4 3 , 1 9 4 5 ; Donoso e t a l . , 1 9 6 2 ; D u c k w o r t h & Woodham, 1 9 6 1 ; G o r d o n , 1 9 6 8 ; G o e r i n g e t a l . , 1 9 7 2 ; M a r s h , 1 9 7 6 ; Yu & Thomas , 1 9 7 5 ; Yu & V e i r a , 1 9 7 7 ) . B e c h t e l e t a l . ( 1945) c o m p a r e d t h e p e r f o r m a n c e o f d a i r y cows whose s o l e d i e t c o n s i s t e d o f " n o r m a l " , " b r o w n " o r " b l a c k " a l f a l f a h a y s . Deg ree o f b r o w n i n g was r e l a t e d t o amount o f h e a t i n g i n t h e s t a c k . E x c e s s i v e h e a t i n g d u r i n g s t o r a g e c o n s i s t e n t l y l o w e r e d t h e a p p a r e n t d i g e s t i b i l i t y o f DM, c r u d e p r o t e i n , c r u d e f i b r e and n i t r o g e n - f r e e e x t r a c t . P r o t e i n was t h e mos t s e v e r e l y a f f e c t e d c o m p o n e n t , w i t h a v e r a g e d i g e s t i b i l i t y c o e f f i c i e n t s f o r n o r m a l , brown o r b l a c k hay s o f 6 7 , 16 and 3% r e s p e c t i v e l y . V o l u n t a r y f e e d i n t a k e d e c r e a s e d w i t h i n c r e a s e i n d e g r e e o f b r o w n i n g w h i c h was i n a g r e e -ment w i t h an e a r l i e r s t u d y u s i n g s o r ghum s i l a g e ( B e c h t e l e t a l . , 1 9 4 3 ) . More r e c e n t work has c o n f i r m e d t h e s e f i n d i n g s and good c o r r e l a t i o n s have been o b t a i n e d b e t w e e n ADIN m e a s u r e s and t h e d i g e s t i b i l i t y o f n i t r o g e n ( N ) , DM, o r g a n i c m a t t e r (OM) , e n e r g y ( E ) , and a c i d - d e t e r g e n t f i b r e ( A D F ) . Some o f t h e s e c o r r e l a t i o n s a r e s u m m a r i z e d i n T a b l e s 1 and 2. - 22 -T a b l e 1: S q u a r e d c o r r e l a t i o n c o e f f i c i e n t s ( r ) b e t w e e n ADIN%N (Y ) and v a r i o u s d i g e s t i b i l i t y c o e f f i c i e n t s ( X ) . 2 P r e d i c t i n g V a r i a b l e (X ) r_ n_ S o u r c e A p p a r e n t D i g e s t i b i l i t y % DM .67 21 Yu and Thomas , 1976 DM .79 - Yu and V e i r a . 1977 DM . 19 36 Yu and Thomas , 1975 N .50 36 Yu and Thomas , 1975 N .86 44 G o e r i ng e t a l . , 1972 N .94 - Yu and V e i r a , 1977 E .51 20 G o e r i ng e t a l . , 1972 ADF .38 - Yu and V e i r a , 1977 ADIN .01 - Yu and V e i r a , 1977 T a b l e 2 : S q u a r e d c o r r e l a t i o n c o e f f i c i e n t s ( r ) b e t w e e n ADIN%DM (Y ) and v a r i o u s d i g e s t i b i l i t y c o e f f i c i e n t s ( X ) . P r e d i c t i n g V a r i a b l e ( X ) r_ n_ S o u r c e A p p a r e n t D i g e s t i b i 1 i t y (%) DM .64 10 Thomas e t a l . , 1972 DM .81 36 Yu and Thomas , 1975 DM .59 21 Yu and Thomas , 1976 N .74 47 G o e r i n g e t a l . , 1972 N .84 10 Thomas e t a l . , 1972 N .67 36 Yu and Thomas , 1975 OM .53 10 Thomas e t a l . , 1972 - 23 -The best correlations obtained have generally been between ADIN%N and nitrogen d i g e s t i b i l i t y . Some regression equations based on these measures are shown in Table 3. Table 3: Regressions of total nitrogen digestion c o e f f i c i e n t s (Y) on ADIN%N (X). Equation r_ n 1. Y = 72.96 - 1.02X .86 44 2. Y = 94.0 - 2.3X .85 3. Y = 91.23 - 2 .19X .86 21 Range of ADIN%N Source 1.78 - 74.54 Goeri ng et al Thomas et a l . 7.1 - 21.9 Yu and Thomas The large difference between equation 1 and the other two is probably related to differences in the forages studied: the range of heat damage in the forages used for equation 1 was much wider than for the other two, and a much wider variety of forages was used. Whereas nitrogen d i g e s t i b i l i t y shows how much nitrogen is absorbed by the animal, i t does not show how much is retained. Several workers have studied the effects of feeding heat damaged forages on nitrogen balance (Goering et al . , 1974 ; Thomas et a l . , 1972; Yu & Thomas, 1975; Yu & Veira, 1977). In a l l cases, nitrogen retention and the e f f i c i e n c y of u t i l i z a t i o n of digested nitrogen were s i g n i f i c a n t l y reduced by heating and were negatively correlated with ADIN and extent of heating. In their study, Yu and Veira (1977) heated 47% DM - 24 -a l f a l f a h a y l a g e i n s e a l e d c o n t a i n e r s a t 88°C f o r 0 , 24 o r 48 h o u r s , and s u b s e q u e n t l y m e a s u r e d n i t r o g e n u t i l i z a t i o n u s i n g s h e e p . T h e r e were no s i g n i f i c a n t d i f f e r e n c e s b e t w e e n n i t r o g e n i n t a k e s , b u t t h e s h e e p f e d h e a t e d h a y l a g e s e x c r e t e d 33 t o 74% more n i t r o g e n i n t h e i r f e c e s ( P < . 0 5 ) , and 26 t o 29% l e s s n i t r o g e n i n t h e i r u r i n e ( P< . 05 ) as c o m p a r e d t o s h e e p f e d t h e u n h e a t e d h a y l a g e . R e t e n t i o n o f n i t r o g e n was r e d u c e d by h e a t i n g and was n e g a t i v e l y c o r r e l a t e d w i t h ADIN%N o r d e g r e e h o u r s above 60°C ( r = - 0 . 7 4 and - 0 . 6 5 r e s p e c t i v e l y ) . They p o s t u l a t e d t h a t t h e d e c r e a s e i n n i t r o g e n r e t e n t i o n was p r o b a b l y t h e r e s u l t o f r e d u c e d i n t a k e s o f t o t a l d i g e s t i b l e n u t r i e n t s and d i g e s t i b l e c r u d e p r o t e i n , a n d / o r t h e u n a v a i l a b i l i t y ( a n d so d e f i c i e n c y ) o f one o r more o f t h e e s s e n t i a l amino a c i d s due t o t h e i r b e i n g t i e d up by t h e b r o w n i n g p r o c e s s . The u n a v a i l a b i l i t y o f e s s e n t i a l amino a c i d s c o u l d a l s o have e x p l a i n e d why t h e y o b s e r v e d a s i g n i f i c a n t d e c r e a s e ( P < . 05 ) i n t h e e f f i c i e n c y o f u t i l i z a t i o n o f d i g e s t e d n i t r o g e n w i t h h e a t i n g ( r w i t h ADIN%N = - 0 . 5 2 ) . L y s i n e i s t h e p r i n c i p a l am ino a c i d i n v o l v e d i n t h e M a i l l a r d r e a c t i o n (Van S o e s t , 1982) so i t i s n o t s u r p r i s i n g t h a t r e s u l t s f r o m s e v e r a l t r i a l s have i n d i c a t e d a s i g n i f i c a n t r e d u c t i o n i n a v a i l a b l e l y s i n e f o l l o w i n g h e a t i n g o f a l f a l f a o r g r a s s ( M a r s h , 1976 ; S a u n d e r s e t a l . , 1 9 7 3 ) . Donoso e t a l . ( 1 9 6 2 ) s t u d i e d t h e e f f e c t o f h e a t i n g - 25 -po r k p r o t e i n on t h e u t i l i z a t i o n o f l y s i n e and t h e s u l p h u r amino a c i d s by r a t s . H e a t i n g r e d u c e d t h e d i g e s t i b i l i t y o f l y s i n e by 2 7 % , w h i l e t h e a p p a r e n t d i g e s t i b i l i t y o f t h e s u l p h u r am ino a c i d s f e l l by o n l y 17%. The h i g h l e v e l s o f s u l p h u r p r e s e n t i n t h e u r i n e l e d t h e r e s e a r c h e r s to c o n c l u d e t h a t w h i l e h e a t i n g d i d n o t s i g n i f i c a n t l y r e d u c e t h e d i g e s t i b i 1 i t y o f t h e s u l p h u r am ino a c i d s , i t d i d d r a m a t i c a l l y r e d u c e t h e i r b i o l o g i c a l v a l u e . T h o r b e k ( 1 977 ) showed t h a t a d e f i c i e n c y o f one o r more o f t h e s e e s s e n t i a l am ino a c i d s c o u l d have d r a m a t i c n e g a t i v e e f f e c t s on n i t r o g e n r e t e n t i o - n . S e v e r a l w o r k e r s have r e p o r t e d t h a t some h e a t i n g o f f o r a g e c a n . b e a d v a n t a g e o u s s i n c e i t r e d u c e s n i t r o g e n l o s s as ammonia f r o m t h e rumen by c o n v e r t i n g t h e h i g h l y s o l u b l e p r o t e i n s i n t h e f o r a g e t o a f o r m w h i c h i s l e s s e a s i l y d e g r a d a b l e by t h e rumen m i c r o f l o r a , t h u s f o r c i n g them t o use more n o n - p r o t e i n n i t r o g e n (NPN) and t h e r e f o r e i n c r e a s i n g t h e amount o f u n d e g r a d e d t r u e p r o t e i n r e a c h i n g t h e s m a l l i n t e s t i n e ( B e e v e r e t a l . , 1 9 7 6 ; G o e r i n g & W a l d o , 1974 , 1 9 7 8 ; H a r t m a n n , 1 9 7 3 ) . T h i s s h o u l d be o f p a r t i c u l a r i m p o r t a n c e f o r s i l a g e s i n c e t h e p r o t e i n i n s i l a g e i s h i g h l y s o l u b l e ( W i l k i n s , 1 9 7 8 ) . G o e r i n g and Wa ldo ( 1 9 7 8 ) r e p o r t e d t h a t when d i e t s c o n t a i n i n g h i g h l y s o l u b l e p r o t e i n s o r NPN, s u c h as g r a s s s i l a g e , a r e f e d , " a s u r p l u s o f ammonia w i l l be a v a i l a b l e , and t h e m i c r o b i a l p r o t e i n s y n t h e s i s w i l l be a t a maximum f o r t h e d i g e s t i b l e 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 d i e t . E x c e s s ammonia w i l l be a b s o r b e d t h r o u g h t h e w a l l o f t h e r u m e n , c o n v e r t e d t o - 26 -u r e a i n t h e l i v e r , and e x c r e t e d i n t h e u r i n e ; and a n e t l o s s o f n i t r o g e n w i l l r e s u l t . " B e e v e r ( 1 9 8 0 ) r e p o r t e d t h a t a n i m a l s f e d g r a s s s i l a g e s c o n t a i n i n g h i g h l e v e l s o f rumen d e g r a d a b l e p r o t e i n we re p a s s i n g as l i t t l e as 15% o f u n d e g r a d e d d i e t a r y n i t r o g e n i n t o t h e duodenum, w i t h t h e c o n s e q u e n c e t h a t t h e y we re p a r t i c u l a r l y d e p e n d e n t on m i c r o b i a l p r o t e i n f o r t h e i r amino a c i d s u p p l y , t h u s g i v i n g p r o b l e m s i n s a t i s f y i n g t h e n i t r o g e n r e q u i r e m e n t s o f h i g h p r o d u c i n g l i v e s t o c k s u c h as cows i n e a r l y l a c t a t i o n and r a p i d l y g r o w i n g b e e f c a t t l e . T h i s work has been mos t u s e f u l l y a p p l i e d t o t h e a r t i f i c i a l d e h y d r a t i o n o f f o r a g e s ( B e e v e r e t a l , 1976 ; G o e r i n g & W a l d o , 1 9 7 8 ; H a r t m a n n , 1 9 7 3 ) . U n f o r t u n a t e l y , i t i s o f v e r y l i m i t e d use i n h e a t i n g s t u d i e s w i t h s i l a g e s b e c a u s e o f t h e i m p o s s i b i l i t y o f e i t h e r p r e d i c t i n g o r c o n t r o l l i n g t h e amount o f h e a t p r o d u c t i o n w i t h i n a s i l o , as w e l l as t h e i n a b i l i t y o f m o i s t u r e t o e s c a p e f r o m a s e a l e d s i l o and t h e l o n g e r p e r i o d o f e x p o s u r e t o h e a t i n g a s s o c i a t e d w i t h e n s i l a g e . H e a t damage r e s u l t i n g i n d e p r e s s i o n i n d i g e s t i b i l i t y i s v i r t u a l l y i n e v i t a b l e when f o r a g e c o n t a i n i n g more t h a n 20% m o i s t u r e i s h e a t e d i n a s e a l e d c o n t a i n e r f o r e ven a few h o u r s ( G o e r i n g e t a l . , 1973 ; Van S o e s t , 1965 ) . 4 . S u r v e y s o f t h e O c c u r r e n c e o f H e a t Damage Among F o r a g e s A l t h o u g h r e s u l t s f r o m f a r m s u r v e y s have f a i l e d t o c o n s i s t e n t l y l i n k d e g r e e o f h e a t damage ( e x p r e s s e d as - 27 -e i t h e r ADIN%DM o r ADIN%N) t o p r o t e i n c o n t e n t o r s i l o t y p e , t h e y have shown t h a t h e a t damage common ly o c c u r s i n g r a s s and l e g u m e s i l a g e s , i s l e s s common i n h a y s , and i s o f t e n p o s i t i v e l y c o r r e l a t e d w i t h s i l a g e DM ( G o e r i n g & Adams, 1 9 7 3 ; Kung e t a l . , 1 9 8 2 ; O n t a r i o M . A . F . , 1 9 7 6 ; S h e l f o r d , 1982 ; Thomas e t a l . , 1972) . Thomas e t a l . ( 1 9 7 2 ) s a m p l e d 34 a l f a l f a h a y l a g e s f r o m M i c h i g a n f a r m s and f o u n d t h a t 33% o f them had ADIN l e v e l s i n e x c e s s o f .36% o f DM. V a l u e s e x c e e d i n g . 29% a r e i n d i c a t i v e o f h e a t damage ( G o e r i n g & Adams , 1 9 7 3 ) . Mean ADIN l e v e l s were s i m i l a r f o r h a y l a g e s f r o m a l l t y p e s o f s i l o s and ADIN was s i g n i f i c a n t l y ( P< . 01 ) c o r r e l a t e d w i t h h a y l a g e DM ( r = 0 . 4 4 ) . G o e r i n g and Adams ( 1 9 7 3 ) i n t h e i r s u r v e y o f 107 h a y - c r o p s i l a g e s f r o m P e n n s y l v a n i a f a r m s , f o u n d t h a t 40% o f t h e s i l a g e s had h e a t damage (AD IN>.29% o f DM) w i t h 3 s a m p l e s s e v e r e l y damaged (ADIN>1% o f DM). In c o n t r a s t t o t h e r e s u l t s o b t a i n e d by Thomas e t a l . ( 1 9 7 2 ) , t h e y we re u n a b l e t o r e l a t e c r o p DM t o e i t h e r ADIN l e v e l s o r h e a t damage (AD IN>.29% o f DM) o v e r t h e r a n g e 2 0 - 7 9 % . A C a n a d i a n s u r v e y ( O n t a r i o M . A . F . , 1976 ) o f 146 h a y - c r o p s i l a g e s f r o m O n t a r i o f a r m s , f o u n d 1 7 . 1 % o f a l l s a m p l e s t o be h e a t damaged (AD IN>.29% DM). No r e l a t i o n -s h i p was f o u n d b e t w e e n h e a t damage and c r o p DM, b u t s i l a g e s s t o r e d i n h o r i z o n t a l s i l o s we re much more s u s c e p t i b l e t o b r o w n i n g t h a n t h o s e s t o r e d i n t o w e r s ( T a b l e 4 ) . The r e s u l t s o f t h e s u r v e y i n d i c a t e d t h a t h i g h q u a l i t y s i l a g e c o u l d be made i n a l l t y p e s o f s i l o s - 28 -p r o v i d i n g p r o p e r a t t e n t i o n i s p a i d t o management f a c t o r s s u c h as s h o r t chop l e n g t h , good c o n s o l i d a t i o n , r a p i d f i l l i n g and e f f i c i e n t s e a l i n g , b u t t h a t u n d e r l e s s t h a n i d e a l management , t h e p r o b a b i l i t y o f h e a t damage i s s l i g h t l y g r e a t e r i n c o n v e n t i o n a l t o w e r s i l o s and much g r e a t e r i n h o r i z o n t a l b u n k e r s and p i t s c o m p a r e d t o o x y g e n - l i m i t i n g t o w e r s . T a b l e 4 : P e r c e n t a g e o f h e a t damaged s a m p l e s i n e a c h s i l o t y p e f o r 146 h a y - c r o p s i l a g e s i n O n t a r i o . ( O n t a r i o M . A . F . , 1976) . S i l o Type n % o f S a m p l e s w i t h AD IN%DM> .29% O x y g e n - 1 i m i t i n g t o w e r 5 3 9 .4 C o n v e n t i o n a l t o w e r 72 11 .1 H o r i z o n t a l b u n k e r s & 22 3 6 . 4 p i t s S h e l f o r d ( 1 9 8 2 ) i n v e s t i g a t e d s a m p l e s o f c o r n , g r a s s and l e gume f o r a g e s c o n s e r v e d as hay o r s i l a g e f r o m 153 f a r m s i n B r i t i s h C o l u m b i a . Some o f h i s r e s u l t s a r e s u m m a r i z e d i n T a b l e 5. Mean v a l u e s o f ADINXDM f o r a l l c o n s e r v a t i o n method s e x c e p t l o a f e x c e e d e d . 2 9 % , i n d i c a t i n g t h a t h e a t damage was w i d e s p r e a d t h r o u g h o u t B . C . i n t h e 1981 c r o p p i n g s e a s o n . Among t h e s i l a g e s , h e a t damage was r e s t r i c t e d p r i m a r i l y t o g r a s s and l e gume f o r a g e s p e c i e s . - 29 -T a b l e 5: Mean ADIN%DM and DM l e v e l s f o r B .C . f o r a g e s c o n s e r v e d by d i f f e r e n t m e t h o d s . ( S h e l f o r d , 1 9 8 2 ) . C o n s e r v a t i o n Me thod n_ Mean ADIN%DM ( S . D . ) Mean DM ( S . D . ) I ' S q u a r e B a l e 106 % .30 ( . 1 1 ) % 86 .2 ( 3 . 3 ) HAYS ^ >Round B a l e 67 .42 ( . 46 ) 81 .2 ( 1 0 . 8 ) I k L o a f 13 .28 ( . 1 1 ) 85 .4 ( 3 . 1 ) r T o w e r 21 .31 ( . 1 2 ) 27 .8 ( 7 . 8 ) S I LAGES < 1 O x y g e n - 1 i m i t i ng B u n k e r 20 70 .57 .43 ( . 20 ) ( . 30 ) 45 27 .9 .6 ( 1 3 . 1 ) ( 7 . 3 ) <Pi t 40 .54 ( .30 ) 34 .6 ( 8 . 5 ) The DM c o n t e n t o f t h e s i l a g e s was s i g n i f i c a n t l y c o r r e l a t e d ( P< . 01 ) w i t h h e a t damage (ADIN%DM) w i t h i n t h e o x y g e n -l i m i t i n g s i l o s ( r = .62 ) and t h e b u n k e r s i l o s ( r = . 4 3 ) , b u t c o r r e l a t i o n s f o r t h e o t h e r c o n s e r v a t i o n me thod s were n o t s i g n i f i c a n t . The h i g h e r ADIN l e v e l s i n b u n k e r s i l o s c o m p a r e d w i t h t o w e r s i l o s a t s i m i l a r a v e r a g e m o i s t u r e c o n t e n t s was t h o u g h t t o be due t o t h e g r e a t e r s u r f a c e a r e a e x p o s e d t o t h e a i r i n b u n k e r s i l o s . In s ummary , t h e s e s u r v e y s s u g g e s t t h a t h e a t damage among h a y - c r o p s i l a g e s i s w i d e s p r e a d a f f e c t i n g 1 7 - 4 0 % o f a l l s a m p l e s and i s more l i k e l y i n h i g h DM g r a s s - l e g u m e s i l a g e s , u n l e s s t h e y a r e w e l 1 - c o n s o l i d a t e d and s t o r e d i n s t r u c t u r e s w h i c h p e r m i t no a i r i n f i l t r a t i o n ( s u c h as o x y g e n - l i m i t i n g t o w e r s ) . - 30 -To c o n c l u d e , a f t e r e a r l y s t u d i e s i n d i c a t e d t h a t e x c e s s i v e h e a t i n g o f f o r a g e s c o u l d have d e t r i m e n t a l e f f e c t s on a p p a r e n t d i g e s t i b i l i t y and a n i m a l p e r f o r m a n c e , s u b s e q u e n t r e s e a r c h c o n c e n t r a t e d on t h e b i o c h e m i s t r y o f t h e b r o w n i n g r e a c t i o n , me thod s o f d e t e c t i n g h e a t damage i n f o r a g e s , f a c t o r s a f f e c t i n g t h e s u s c e p t i b i l i t y o f c r o p s t o h e a t damage, and t h e e f f e c t s o f f e e d i n g h e a t damaged f o r a g e s on a n i m a l p e r f o r m a n c e . W h i l e t h i s r e s e a r c h on h e a t damage has been q u i t e e x t e n s i v e , s t u d i e s have t e n d e d t o f o c u s on a l f a l f a h a y l a g e s . In a d d i t i o n , mos t i n v e s t i g a t i o n s u t i l i z i n g a r t i f i c i a l h e a t i n g have been o f s h o r t d u r a t i o n ( t y p i c a l l y 48 h o u r s o f h e a t i n g ) w i t h s m a l l amount s o f m a t e r i a l ( t y p i c a l l y 2 - 3 0 0 g o f DM). W i t h t h e p r e s e n t s t u d y i t was hoped t o g a i n more i n f o r m a t i o n on h e a t damage i n g r a s s s i l a g e u s i n g b o t h n a t u r a l l y h e a t e d and l o n g t e r m a r t i f i c i a l l y h e a t e d ( 4 - 8 weeks a t 5 0 - 6 0 ° C ) s a m p l e s i n s u f f i c i e n t q u a n t i t i e s t o c a r r y o u t d i g e s t i b i l i t y t r i a l s w i t h s h e e p . F u r t h e r -more i t was p l a n n e d t o c ompa re c o r r e l a t i o n s b e t w e e n s i l a g e ADIN%N l e v e l s and a p p a r e n t d i g e s t i b i l i t y o f v a r i o u s f r a c t i o n s w i t h i n b o t h t h e n a t u r a l l y and t h e a r t i f i c i a l l y h e a t e d s i l a g e s i n o r d e r t o g a i n some i n s i g h t i n t o t h e v a l i d i t y o f s t u d i e s t h a t s eek t o a r t i f i c i a l l y i n d u c e h e a t damage i n p r e v i o u s l y f e r m e n t e d s i l a g e s . - 31 -MATERIALS AND METHODS 1. INTRODUCTION To s t u d y t h e e f f e c t s o f h e a t damage on g r a s s s i l a g e s , two s e p a r a t e e x p e r i m e n t s we re c a r r i e d o u t i n s u c c e s s i v e y e a r s . The a im o f t h e f i r s t y e a r ' s i n v e s t i g a t i o n ( 1 9 8 1 ) was a t t e m p t t o e s t a b l i s h a l i n k b e t w e e n h e a t damage and t h e a p p a r e n t d i g e s t i b i l i t y o f v a r i o u s f r a c t i o n s w i t h i n g r a s s s i l a g e . To t h i s e n d , s i x d i f f e r e n t s i l a g e s f r o m t h e F r a s e r V a l l e y a r e a o f B r i t i s h C o l u m b i a were s e l e c t e d b a s e d on t h e i r v a r y i n g d e g r e e s o f h e a t damage as d e t e r m i n e d by t h e i r l e v e l s o f AD IN%N, and i n v i v o a p p a r e n t d i g e s t i b i l i t y c o e f f i c i e n t s were m e a s u r e d i n t r i a l s u s i n g s h e e p . To r e d u c e t h e number o f v a r i a b l e s a s s o c i a t e d w i t h t h e Y e a r 1 e x p e r i m e n t and t o a s s e s s t h e f e a s i b i l i t y o f a r t i f i c i a l l y i n d u c i n g h e a t damage , a f u r t h e r e x p e r i m e n t was c a r r i e d o u t i n 1 982 . A l o c a l l y p r o d u c e d g r a s s s i l a g e was s e l e c t e d f o r i t s a p p a r e n t a b s e n c e o f h e a t damage (ADIN%N = 7 .7%) and was a r t i f i c i a l l y h e a t damaged to v a r y i n g l e v e l s . D i g e s t i b i l i t y t r i a l s w i t h s h e e p we re a g a i n u s e d t o i n v e s t i g a t e l i n k s b e t w e e n h e a t damage and i n v i v o d i g e s t i b i l i t y c o e f f i c i e n t s . U r i n e was a l s o c o l l e c t e d so t h a t m e a s u r e s o f n i t r o g e n u t i l i z a t i o n c o u l d be c o m p u t e d . F i n a l l y , t h e r e s u l t s f r o m b o t h e x p e r i m e n t s we re c o m p a r e d i n o r d e r t o t e s t t h e v a l i d i t y o f t h e a r t i f i c i a l h e a t t r e a t m e n t . - 32 -2 . EXPERIMENT 1 2 .1 S i 1 age s S i x s i l a g e s f r o m t h e F r a s e r V a l l e y a r e a o f B r i t i s h C o l u m b i a we re s e l e c t e d f o r t h e i r v a r y i n g d e g r e e s o f h e a t damage as m e a s u r e d by t h e p e r c e n t a g e o f a c i d - d e t e r g e n t i n s o l u b l e n i t o g e n i n t h e t o t a l n i t r o g e n (ADIN%N) ( T a b l e 6 ) . T a b ! e 6 : H e a t damage e x p r e s s e d as ADIN%N f o r t h e s i x s i l a g e s u s e d i n E x p e r i m e n t 1. S i 1 age ADIN%N 1 4 . 8 2 5.4 3 8 .9 4 9 . 0 5 17 . 3 6 2 2 . 9 The s i l a g e s we re p r e d o m i n a n t l y o r c h a r d g r a s s - c l o v e r mi x t u r e s . Once s e l e c t e d as s u i t a b l e , a s i l a g e was p l a c e d i n p o l y e t h y l e n e b a g s , e a c h h o l d i n g enough f o r one day s f e e d i n g o f 5 o r 6 s h e e p . B e t w e e n 21 and 24 bag s o f e a c h s i l a g e we re w e i g h e d o u t and s t o r e d i n a f r e e z e r u n t i l r e q u i r e d . D u r i n g t h e d i g e s t i b i l i t y t r i a l s , f e e d s a m p l e s were t a k e n a t random on a d a i l y b a s i s and we re o v e n d r i e d a t 50°C f o r 72 h o u r s i n a f o r c e d a i r o v e n . The d r i e d - 33 -s a m p l e s we re g r o u n d i n a W i l e y m i l l , c o m p o s i t e d and a n a l y z e d i n d u p l i c a t e f o r t o t a l n i t r o g e n (N) by b l o c k d i g e s t i o n and t h e T e c h n i c o n a u t o - a n a l y z e r p r o c e d u r e s ( m e t h o d 2 9 - 6 9 A ) , f o r a c i d - d e t e r g e n t f i b r e (ADF) and n e u t r a l - d e t e r g e n t f i b r e (NDF) by t h e m i c r o - d i g e s t i o n t e c h n i q u e s o f W a l d e r n ( 1 9 7 1 ) , f o r a c i d - d e t e r g e n t i n s o l u b l e n i t r o g e n (AD IN) by m e a s u r i n g t h e t o t a l N i n t h e ADF p o r t i o n ( G o e r i n g & Van S o e s t , 1 9 7 0 ) , and f o r ash by a s h i n g a 2g s a m p l e a t 6 0 0 ° C f o r 2 h o u r s i n a m u f f l e f u r n a c e . F u r t h e r random s a m p l e s t a k e n d u r i n g e a c h d i g e s t i b i l i t y t r i a l were f r o z e n i m m e d i a t e l y and u sed f o r d e t e r m i n i n g d r y m a t t e r (DM) and v o l a t i l e n i t r o g e n ( V o l . N ) . F r o z e n s a m p l e s we re m i x e d w i t h c h o p p e d d r y i c e and f i n e l y g r o u n d w i t h a h i g h s p e e d k i t c h e n b l e n d e r . A t l e a s t 2 day s we re a l l o w e d f o r t h e d r y i c e t o d i s s i p a t e b e f o r e s u b j e c t i n g t h e s a m p l e s t o f u r t h e r a n a l y s i s . DM was e s t i m a t e d u s i n g t h e t o l u e n e d i s t i l l a t i o n p r o c e d u r e o f Dewar & M c D o n a l d ( 1 9 6 1 ) , w h i l e t h e K j e h l d a l m e t h o d , m o d i f i e d by o m i t t i n g t h e i n i t i a l d i g e s t i o n , was u s e d t o d e t e r m i n e V o l . N. 2.2 A n i ma!s S i x S u f f o l k X D o r s e t ewe l ambs a v e r a g i n g 50kg body w e i g h t we re s e l e c t e d on t h e b a s i s o f s i m i l a r i t i e s i n age and s i z e . P r i o r t o t h e e x p e r i m e n t , t h e s h e e p had been r e c e i v i n g a r a t i o n o f a l f a l f a hay and a l f a l f a c u b e s . - 34 -They we re g r a d u a l l y s w i t c h e d o v e r t o g r a s s s i l a g e u s i n g a l f a l f a c u b e s and i n c r e a s i n g p r o p o r t i o n s o f s i l a g e . A r a n d o m i z e d b l o c k d e s i g n was u s ed t o c o m p a r e t h e 6 s i l a g e s i n 6 b l o c k s . Each ewe s h o u l d h a v e r e c e i v e d a l l 6 s i l a g e s . Due t o s h o r t a g e o f e x p e r i m e n t a l f e e d s , h o w e v e r , s i l a g e s 1 and 2 we re f e d t o a l l s i x , w h i l e s i l a g e s 3, 4 and 6 we re f e d t o f i v e , and s i l a g e 5 was f e d t o o n l y f o u r o f t h e s h e e p . R e s i d u a l e f f e c t s c o u l d n o t be e s t i m a t e d b u t we re m i n i m i z e d by a l l o w i n g a l o n g 14 day a d a p t a t i o n p e r i o d b e t w e e n t r i a l s . 2 . 3 D i g e s t i b i l i t y T r i a l s The ewes w e r e i n d i v i d u a l l y p e n n e d d u r i n g e a c h 14 day a d a p t a t i o n p e r i o d and we re s u b s e q u e n t l y p l a c e d i n d i g e s t i b i l i t y c r a t e s f o r a 7 day t o t a l f e c a l c o l l e c t i o n p e r i o d i n o r d e r t o e s t i m a t e t h e a p p a r e n t d i g e s t i b i l i t y o f e a c h s i l a g e . D u r i n g t h e a d a p t a t i o n p e r i o d s i n t a k e s were a d j u s t e d t o t r y and a c h i e v e a u n i f o r m d a i l y i n t a k e o f 1200g o f DM f o r e a c h e w e . The d e f r o s t e d s i l a g e f r o m one bag was w e i g h e d o u t e a c h m o r n i n g and f e d i n two e q u a l a m o u n t s : one a t 8am and one a t 3pm. The s h e e p had a c c e s s t o w a t e r and i o d i z e d s a l t a t a l l t i m e s . The f i r s t d i g e s t i b i l i t y t r i a l s t a r t e d on A p r i l 6 , 1 9 8 1 , w h i l e t h e l a s t was c o m p l e t e d on A u g u s t 1 0 , 1 9 8 1 . The s h e e p we re t r a n s f e r r e d f r o m i n d i v i d u a l pens t o d i g e s t i b i l i t y c r a t e s one day b e f o r e t h e c o l l e c t i o n - 35 -s t a r t e d . F e c e s we re c o l l e c t e d i n a p l a s t i c bowl b e n e a t h e a c h c r a t e and were w e i g h e d and r e c o r d e d a t 9am e v e r y m o r n i n g . In o r d e r t o c a r r y o u t f u r t h e r a n a l y s i s , t h e f e c e s we re p l a c e d i n a l u m i n u m t r a y s and d r i e d i n a f o r c e d a i r o v e n a t 90°C f o r 48 h o u r s . F o l l o w i n g d r y i n g , t h e 7 d a i l y s a m p l e s : f r o m e a c h ewe w e r e g r o u n d i n a W i l e y m i l l , c o m p o s i t e d and a n a l y z e d i n d u p l i c a t e f o r t o t a l N, ADF, NDF, ADIN and a sh c o n t e n t u s i n g t h e same p r o c e d u r e s as d e t a i l e d f o r t h e s i l a g e s . P r i o r t o t h e m o r n i n g f e e d i n g , f e e d r e f u s a l s we re c o l l e c t e d and o v e n d r i e d t o g a i n an a c c u r a t e e s t i m a t e o f DM i n t a k e . B e c a u s e some o f t h e ewes we re s e l e c t i v e i n wha t p a r t s o f a s i l a g e t h e y w o u l d e a t ( f o r e x a m p l e , some l e f t s t a l k y m a t e r i a l ) , d r i e d f e e d r e f u s a l s a m p l e s were g r o u n d and a n a l y z e d f o r t o t a l N, ADF, NDF, ADIN and a sh c o n t e n t . A f t e r a l l s a m p l e s had been a n a l y z e d , t h e a p p a r e n t d i g e s t i b i l i t i e s o f DM, OM, N, ADF, NDF and ADIN were c a l c u l a t e d f o r e a c h s i l a g e u s i n g DM i n t a k e s c a l c u l a t e d by t h e t o l u e n e d i s t i l l a t i o n p r o c e d u r e . 2 .4 S t a t i s t i c a l A n a l y s i s A n a l y s i s o f v a r i a n c e u s i n g t h e UBC p a c k a g e p r o g r a m BMD-10V ( G e n e r a l L i n e a r H y p o t h e s i s ) was u s e d t o d e t e c t d i f f e r e n c e s i n t h e two m a i n e f f e c t s - s i l a g e s and s h e e p ; s i g n i f i c a n c e o f d i f f e r e n c e s b e t w e e n means w e r e d e t e r m i n e d u s i n g a N e w m a n - K e u l s 1 T e s t . - 36 -R e l a t i o n s h i p s o f ADIN%N and o t h e r s i l a g e f r a c t i o n s w i t h a p p a r e n t d i g e s t i b i l i t i e s we re d e t e r m i n e d u s i n g c o r r e l a t i o n and s i m p l e r e g r e s s i o n a n a l y s i s w i t h t h e UBC p a c k a g e p r o g r a m *TRP ( T r i a n g u l a r R e g r e s s i o n P a c k a g e , 1 9 7 7 ) . 3 . EXPERIMENT 2 3.1 A c q u i s i t i o n o f E x p e r i m e n t a l S i l a g e B a s e d on i t s a p p a r e n t l a c k o f h e a t damage (ADIN%N = 7 . 7 % ) , o r c h a r d g r a s s s i l a g e was p u r c h a s e d f r o m t h e B r i t i s h C o l u m b i a M i n i s t r y o f A g r i c u l t u r e f a r m a t C o l o n y F a r m , B . C . f o r t h e s e c o n d y e a r ' s s t u d y . A f u r t h e r s e l e c t i o n p a r a m e t e r was i t s r e a s o n a b l y h i g h c r u d e p r o t e i n c o n t e n t ( 14% o f DM). S u b s e q u e n t l y i t was f o u n d t h a t t h e c r u d e p r o t e i n c o n t e n t had been c a l c u l a t e d i n c o r r e c t l y and was i n f a c t o n l y 9%. S i n c e t h e i n v e s t i g a t i o n had a l r e a d y commenced and s i n c e s i l a g e s o f t h i s c r u d e p r o t e i n l e v e l a r e s o m e t i m e s p r o d u c e d i n B . C . , i t was d e c i d e d t o p r o c e e d . The e x p e r i m e n t c o n s i s t e d o f 3 t r e a t m e n t s e a c h o f w h i c h r e q u i r e d t h e s e l e c t e d s i l a g e t o be t r a n s f e r r e d t o i a s m a l l - s c a l e s i l o ( s e e S e c t i o n 3 . 2 ) . The 3 e x p e r i m e n t a l s i l o s we re t r a n s p o r t e d on a f l a t - b e d t r u c k f r o m t h e r e s e a r c h f a r m a t UBC t o C o l o n y F a r m . A t t h e f a r m t h e s i l a g e was u n l o a d e d f r o m t h e c o n c r e t e s t a v e t o w e r s i l o i n w h i c h i t was s t o r e d and p a c k e d t i g h t l y i n t o e a c h e x p e r i m e n t a l s i l o . Two p e o p l e f i l l e d w h i l e two o t h e r s - 37 -c o n t i n u a l l y t r a m p e d t h e s i l a g e down t o m i n i m i z e a e r o b i c d e t e r i o r a t i o n . In o r d e r t o e n s u r e u n i f o r m i t y , a l l 3 s i l o s were f i l l e d s i m u l t a n e o u s l y . Once f u l l , t h e s i l o s we re s e a l e d , t h e t o p s we re w e i g h t e d down, and t h e y we re t r a n s p o r t e d back t o t h e r e s e a r c h f a r m a t UBC. 3.2 E x p e r i m e n t a l S i l o s The 3 e x p e r i m e n t a l s i l o s we re p a p e r f i b r e f o r m ( " P e r m a f o r m " ) c y l i n d e r s , lm i n d i a m e t e r and 2.2m t a l l . They w e r e o f s u f f i c i e n t s t r e n g t h t o p r o v i d e s t r u c t u r a l s u p p o r t and t h e y were moun ted on p a l l e t s t o f a c i l i t a t e t r a n s p o r t . The s i l a g e was c o m p l e t e l y s e a l e d by a p o l y e t h y l e n e bag w h i c h was i n s i d e t h e f i b r e f o r m c y l i n d e r . The t r e a t m e n t s r e q u i r e d one s i l o as a c o n t r o l (C ) and t h e o t h e r two t o be h e a t e d t o two l e v e l s (H I and H 2 ) . V a r i o u s me thod s o f h e a t i n g s u c h as ho t w a t e r p i p e s and e l e c t r i c b l a n k e t s were i n v e s t i g a t e d b e f o r e i t was d e c i d e d t o use t h e r m o s t a t i c a l l y c o n t r o l l e d e l e c t r i c h e a t i n g c a b l e s . S i l o s HI and H2 we re e a c h f i t t e d w i t h one t y p e GP 250 h e a t i n g c a b l e ( 2235 w a t t s a t 240 v o l t s ; 76m l o n g ) c o n t r o l l e d by a t y p e T 3246 t h e r m o s t a t , a l l s u p p l i e d by C h r o m a l o x I n c . o f R e x d a l e , O n t a r i o . The h e a t i n g c a b l e s we re t h r e a d e d t h r o u g h a p o l y p r o p y l e n e 5cm s q u a r e mesh f o r m e d i n t o a c y l i n d e r w h i c h f i t t e d s n u g l y i n t o t h e i n s i d e o f t h e f i b r e f o r m s h e l l p r i o r t o f i l l i n g ( F i g u r e s ) . Thus t h e h e a t i n g c a b l e was a r r a n g e d i n c o i l s a t 5 t o 10cm s p a c i n g s a r o u n d t h e s i l a g e m a s s . To r e d u c e - 38 -Figure 4: Cross-section through one of the heated s i l o s . (Scale 1:10) - 40 -Fj gure 6: Treatments HI and H2 showing outer insulating layer of rock wool and the auto-temperature recorder connected to the thermocouples. - 41 -h e a t l o s s , t h e o u t s i d e o f e a c h o f t h e two h e a t e d s i l o s was c o m p l e t e l y c o v e r e d w i t h a 15cm t h i c k n e s s o f r o c k woo l i n s u l a t i o n ( s e e F i g u r e 6 ) . In o r d e r t o m o n i t o r s i l a g e t e m p e r a t u r e a c c u r a t e l y w i t h i n t h e h e a t e d s i l o s , t h r e e t h e r m o c o u p l e s we re i n s e r t e d h a l f way down e a c h s i l o : one i n t h e c e n t r e , one 5cm f r o m t h e o u t s i d e e d g e , and one a t t h e m i d way p o i n t b e t w e e n t h e s e t w o . A c r o s s -s e c t i o n a l d i a g r a m o f one o f t h e h e a t e d s i l o s i s shown i n F i g u r e 4 . 3.3 S i l a g e T e m p e r a t u r e I n i t i a l l y , s i 1 age t e m p e r a t u r e s we re c h e c k e d t w i c e d a i l y i n o r d e r t o a d j u s t t h e t h e r m o s t a t s t o a c h i e v e t h e d e s i r e d t e m p e r a t u r e . F o l l o w i n g t e m p e r a t u r e s t a b i l i z a t i o n , t h e s i l o s we re c h e c k e d o n c e d a i l y . W h i l e t h e o u t e r l a y e r o f s i l a g e n e e d e d o n l y 12 h o u r s t o warm u p , t h e c o r e t o o k a b o u t 3 day s t o r e a c h t h e d e s i r e d t e m p e r a t u r e . A t t h e end o f t h e i r h e a t i n g p e r i o d b o t h s i l a g e s HI and H2 t o o k a b o u t 4 day s t o c o o l down. In b e t w e e n s t a r t up and s h u t down i t was p o s s i b l e t o m a i n t a i n r e m a r k a b l y c o n s t a n t t e m p e r a t u r e s u s i n g t h i s a p p a r a t u s . No t e m p e r a t u r e r e a d i n g s we re t a k e n f o r t h e c o n t r o l s i l a g e ( t r e a t m e n t C ) . A v e r a g e t e m p e r a t u r e s and l e n g t h o f h e a t i n g f o r e a c h t r e a t m e n t a r e shown i n T a b l e 7. - 42 -T a b l e 7 : A v e r a g e t e m p e r a t u r e and l e n g t h o f h e a t i n g p e r i o d f o r t h e 3 t r e a t m e n t s i n E x p e r i m e n t 2. T r e a t m e n t A v e r a g e T e m p e r a t u r e and L e n g t h o f P e r i o d C o n t r o l (C) No h e a t i n g Hea t 1 ( H I ) 30 day s a t 49°C Hea t 2 (H2) 33 day s a t 50°C f o l l o w e d by 30 da y s a t 6 3 ° C The r a t i o n a l e f o r s e l e c t i n g t h e s e t e m p e r a t u r e s and t i m e s was b a s e d on t h e f o l l o w i n g : -a) The r e s u l t s f r o m f a r m s u r v e y s ( W a t s o n & N a s h , 1 9 6 0 ; W i e r i n g a e t a l . , 1961) t h a t f o u n d t h a t t e m p e r a t u r e s o f 5 0 - 6 0 ° C were r e a d i l y a t t a i n a b l e w i t h i n s i l o s . b) The e x p e r i m e n t a l e v i d e n c e ( W i e r i n g a e t a l . , 1 9 6 1 ; Yu & Thomas , 1 9 7 5 ; Y u , 1976) t h a t i n d i c a t e s t h a t t h e e x t e n t o f h e a t damage as m e a s u r e d by ADIN f o r m a t i o n i s n o t s i g n i f i c a n t a t t e m p e r a t u r e s b e l o w 3 5 - 4 0 ° C . c ) The e x p e r i m e n t a l e v i d e n c e ( G o e r i n g e t a l . , 1 9 7 3 ; M i d d l e t o n & Thomas , 1983) t h a t s u g g e s t s t h a t t h e e x t e n t o f h e a t damage as m e a s u r e d by ADIN f o r m a t i o n i n c r e a s e s d r a m a t i c a l l y a t t e m p e r a t u r e s above 6 0 ° C ( t h i s was t h e r a t i o n a l e f o r t h e h i g h e r t e m p e r a t u r e s e l e c t e d f o r t h e l a t t e r p a r t o f t r e a t m e n t H 2 ) . d) The e x p e r i m e n t a l e v i d e n c e ( W i e r i n g a e t a l . , 1 9 6 1 ; Y u , 1976) t h a t s u g g e s t s t h a t t h e amount o f ADIN f o r m e d i n c r e a s e s w i t h t h e l e n g t h o f t h e h e a t i n g p e r i o d . e) The e v i d e n c e f r o m b o t h e x p e r i m e n t s (Yu & Thomas , 1975) - 43 -and f a r m s u r v e y s ( W i e r i n g a e t a l . , 1961) t h a t f o u n d t h a t t e m p e r a t u r e s i n t h e 3 5 - 5 0 ° C r a n g e c o u l d be m a i n t a i n e d f o r e x t e n d e d p e r i o d s ( 3 0 - 7 0 d a y s ) u n d e r n a t u r a l s i l o c o n d i t i o n s . F o r t h e s e r e a s o n s i t was d e c i d e d t o h e a t t h e 2 s i l o s a t a p p r o x i m a t e l y 50°C f o r 30 day s and t h e n h e a t s i l o H2 a t t e m p e r a t u r e s s l i g h t l y i n e x c e s s o f 6 0°C f o r a f u r t h e r 30 d a y s . 3.4 T r e a t e d S i l a g e When r e a d y f o r f e e d i n g , e a c h s i l o was u n l o a d e d and t h e s i l a g e p l a c e d i n p o l y e t h y l e n e bags e a c h h o l d i n g s u f f i c i e n t f o r one day s f e e d i n g o f 6 s h e e p . T w e n t y - o n e bags o f e a c h s i l a g e were w e i g h e d o u t and p l a c e d i n a f r e e z e r u n t i l r e q u i r e d . D u r i n g s i l o u n l o a d i n g , s a m p l e s we re t a k e n a t r e c o r d e d d e p t h s and p o s i t i o n s , and a n a l y z e d f o r DM and ADIN%N i n o r d e r t o g a i n some i n s i g h t i n t o t h e e f f e c t s o f t h e h e a t i n g t r e a t m e n t on t h e s e two v a r i a b l e s . The c o l l e c t i o n and a n a l y s i s o f f e e d s a m p l e s d u r i n g t h e d i g e s t i b i l i t y t r i a l s was e x a c t l y t h e same as d e s c r i b e d f o r E x p e r i m e n t 1. 3.5 A n i ma 1s S i x o n e - y e a r o l d u n b r e d S u f f o l k X D o r s e t ewes a v e r a g i n g 60kg body w e i g h t we re s e l e c t e d on t h e b a s i s o f s i m i l a r i t i e s i n s i z e . - 44 -A r a n d o m i z e d b l o c k d e s i g n was u sed t o compa re t h e 3 s i l a g e s i n 3 b l o c k s , so t h a t e a c h ewe r e c e i v e d a l l 3 s i l a g e s . The s i l a g e s we re f e d i n t h e o r d e r H I , C and f i n a l l y H2. A 3X3 L a t i n - s q u a r e d e s i g n w i t h 2 g r o u p s o f 3 s h e e p and w i t h a l l 3 t r e a t m e n t s b e i n g f e d s i m u l t a n e o u s l y was i m p o s s i b l e b e c a u s e o f t h e n e c e s s i t y o f c o m p l e t e l y u n l o a d i n g e a c h s i l o o n c e t h e p o l y e t h y l e n e s e a l had been b r o k e n i n o r d e r t o p r e v e n t s p o i l a g e , c o m b i n e d w i t h t h e l a c k o f f r e e z e r s p a c e t o h o l d a l l 3 t r e a t m e n t s a t o n c e . The ewes w e r e a d a p t e d t o e a c h s i l a g e f o r 14 day s p r i o r t o b e i n g p l a c e d i n d i g e s t i b i l i t y c r a t e s f o r a 7 day t o t a l c o l l e c t i o n t r i a l . D u r i n g t h e a d a p t a t i o n p e r i o d , i n t a k e s , w e r e a d j u s t e d t o t r y and a c h i e v e a u n i f o r m d a i l y i n t a k e o f 750g o f DM f o r e a c h ewe. T h i s was c o n s i d e r a b l y l o w e r t h a n t h e 1200g a v e r a g e i n t a k e o f DM a c h i e v e d d u r i n g E x p e r i m e n t 1 and was n e c e s s a r i l y c h o s e n b e c a u s e o f t h e p o o r v o l u n t a r y f e e d i n t a k e a s s o c i a t e d w i t h a l l 3 t r e a t m e n t s . I n d e e d two o f t h e ewes c o n s i s t e n t l y a t e c o n s i d e r a b l y l e s s t h a n t h i s (ewe 3, 690g and ewe 4 , 610g a p p r o x i m a t e a v e r a g e d a i l y DM i n t a k e o v e r t h e 3 t r i a l s ) . P r o b a b l y as a r e s u l t o f h e r l o w i n t a k e , ewe 4 was s i c k ( o f f -f e e d and s l i g h t l y l o o s e ) o v e r t h e l a t t e r h a l f o f f i n a l t r i a l . Ewe body w e i g h t s we re r e c o r d e d a t t h e b e g i n n i n g and end o f t h e e x p e r i m e n t , and i n m i d - J u n e a f t e r s i l a g e s HI and C had been f e d . The f e e d i n g m e t h o d was i d e n t i c a l t o t h a t u s e d i n E x p e r i m e n t 1. - 45 -3 .6 D i g e s t i b i l i t y T r i a l s A p p a r e n t d i g e s t i b i l i t y c o e f f i c i e n t s f o r t h e t h r e e t r e a t m e n t s w e r e c a l c u l a t e d u s i n g 7 day t o t a l c o l l e c t i o n t r i a l s as i n E x p e r i m e n t 1. The d i g e s t i b i l i t y t r i a l s r a n t h r o u g h o u t May and J u n e , 1 9 8 2 . C o l l e c t i o n , p r e p a r a t i o n and a n a l y s i s o f f e e d , f e c e s and f e e d r e f u s a l s a m p l e s we re i d e n t i c a l t o t h o s e m e t h o d s u s e d i n E x p e r i m e n t 1. 3.7 N i t r o g e n B a l a n c e In c o n t r a s t t o E x p e r i m e n t 1, some m e a s u r e s o f n i t r o g e n u t i l i z a t i o n we re m e a s u r e d by a r r a n g i n g f o r t o t a l u r i n e c o l l e c t i o n s i n a d d i t i o n t o t h e f e c e s f r o m e a c h ewe . The u r i n e was f u n n e l l e d and f i l t e r e d t h r o u g h c l e a n g a u z e t o p r e v e n t c o n t a m i n a t i o n by t h e few s m a l l f e c a l p e l l e t s t h a t o c c a s i o n a l l y p a s s e d t h r o u g h t h e s e p a r a t o r g r i d i n t o t h e u r i n e , i n t o a p o l y e t h y l e n e v e s s e l c o n t a i n i n g 50ml o f s u l p h u r i c a c i d ( 5 0% v / v ) . A t t h e end o f e a c h c o l l e c t i o n d a y , t h e u r i n e was w e i g h e d and m i x e d , and a 25ml s u b s a m p l e was t a k e n f r o m e a c h ewe and s t o r e d a t 2 ° C . F o l l o w i n g t h e c o m p l e t i o n o f e a c h t r i a l , t h e u r i n e s a m p l e s f o r e a c h ewe we re p o o l e d and a n a l y z e d f o r t o t a l n i t r o g e n u s i n g t h e K j e h l d a l m e t h o d . In t h i s w a y , n i t r o g e n r e t e n t i o n and t h e e f f i c i e n c y o f n i t r o g e n r e t e n t i o n ( m e a s u r e d as a p e r c e n t a g e o f n i t r o g e n d i g e s t e d ) we re b o t h e s t i m a t e d . - 46 -3.8 S t a t i s t i c a l A n a l y s i s S t a t i s t i c a l t r e a t m e n t o f t h e r e s u l t s was i d e n t i c a l t o t h o s e method s u s e d i n E x p e r i m e n t 1 w i t h t h e a d d i t i o n o f a n a l y s i s o f v a r i a n c e on s i l a g e c o m p o s i t i o n t o d e t e c t d i f f e r e n c e s due t o t h e h e a t i n g t r e a t m e n t s . - 47 -RESULTS 1. EXPERIMENT 1 1.1 S i l a g e Compos i t i o n C o m p o s i t i o n o f t h e s i x s i l a g e s s e l e c t e d f o r t h e d i g e s t i b i l i t y t r i a l s a r e shown i n T a b l e 8. T h e r e were c o n s i d e r a b l e d i f f e r e n c e s b e t w e e n t h e s i l a g e s i n l e v e l s o f t o t a l n i t r o g e n and ADIN as a p e r c e n t a g e o f t o t a l N. L e v e l s o f ADIN%N were u n r e l a t e d t o b o t h t o l u e n e DM and t o t a l N c o n t e n t . The s i l a g e s d i f f e r e d m a r k e d l y i n a p p e a r a n c e and s m e l l . S i l a g e s 3 and 5 we re p a r t i c u l a r l y s t r o n g s m e l l i n g and mos t l i k e l y had u n d e r g o n e a c l o s t r i d i a l f e r m e n t a t i o n a l t h o u g h u n f o r t u n a t e l y no m e a s u r e s o f pH o r o r g a n i c a c i d s were t a k e n . S i m i l a r l y , s i l a g e 4 had odd p a t c h e s o f f o u l -s m e l l i n g , b a d l y f e r m e n t e d m a t e r i a l i n i t . The h i g h d r y m a t t e r s i l a g e 6 had a v e r y d i s t i n c t b rown c o l o u r and a r a t h e r p l e a s a n t b u r n t o r t o b a c c o - l i k e a r o m a . T a b l e 8: C o m p o s i t i o n o f t h e s i x s i l a g e s u s e d i n E x p e r i m e n t 1 ( n = 3). S ILAGE NUMBER 1 (S.D.*) 2 (S.D.) 3 (S.D.) 4 (S.D.) 5 (S.D.) 6 (S.D. Toluene DM {%) 34.94 (.10) 22.15 (.01) 25.80 (.15) 22.83 (.38) 29.95 (.42) 58.11 (.86) Composition of DM (%) Ash 11.35 (.03) 12.52 (.16) 11.82 (.10) 12.43 (.03) 10.00 (.16) 14.32 (.22) NDF 45; 83 (.26) 51.25 (.48) 66.27 (.04) 52.56 (.06) 72.36 (.56) 59.64 (.22) ADF 32.68 (.12) 39.61 (.27) 44.45 (.40) 39.80 (.39) 47.01 (.12) 43.36 (.46) Total N 3.35 (.02) 3.12 (.04) 1.88 (.02) 2.66 (.03) 1.67 (.01) 2.52 (.01) V o l a t i l e N .36 (.01) .49 (.01) .89 (.02) .92 (.02) .86 (.01) .22 (.01) ADIN .50 (.01) .42 (.01) .38 (.01) .60 (.01) .61 (.02) 1.33 (.02) ADINXTotal N 4.81 5.37 8.96 9.02 17.31 22.91 CO * S . D . = S t a n d a r d D e v i a t i o n - 4 9 -1. 2 P a l a t a b i l i t y and I n t a k e As i t was d e c i d e d t o f e e d t h e ewes a t j u s t above t h e m a i n t e n a n c e l e v e l and t o e s t a b l i s h t h i s a t a r o u n d 1200g o f DM f o r a l l 6 s i l a g e s , m e a s u r e s o f v o l u n t a r y f e e d i n t a k e were n o t made. P a l a t a b i l i t y was a s s e s s e d q u a l i t a t i v e l y by c o m p a r i n g s p e e d o f e a t i n g , p r e s e n c e o r a b s e n c e o f f e e d r e f u s a l s and t h e r e f o r e t h e number o f ewes who r e g u l a r l y a t e t h e i r t o t a l d a i l y a l l o w a n c e . Q u a l i t a t i v e and q u a n t i t a t i v e m e a s u r e s o f s i l a g e i n t a k e and r e l a t i v e p a l a t a b i l i t y a r e shown i n T a b l e 9 . T a b l e 9 : I n t a k e and r e l a t i v e p a l a t a b i l i t y o f E x p e r i m e n t 1 s i l a g e s . DM Of fered DM Consumed F r a c t i on of Ewes R e l a t i v e Average Range That Normally Ate P a l a t a b i l i t y Da i l y Al lowance g g g S i l a ge No. 1 1223 1218 1195-1223 5/6 Excel l e n t 2 1247 1247 1247 6/6 Excel l e n t 3 1170 1147 1080-1170 2/5 Good 4 1299 1246 1233-1258 1/5 * E x c e l l e n t 5 1201 1174 1161-1188 1/4 Good 6 1136 1136 1136 5/5 Excel l e n t * T h e p a t c h e s o f b a d l y f e r m e n t e d m a t e r i a l m e n t i o n e d i n t h e p r e v i o u s s e c t i o n 1.1 we re i n v a r i a b l y r e j e c t e d by a l 1 e w e s . A l t h o u g h p a l a t a b i l i t y was n o t m e a s u r e d q u a n t i t a t i v e l y , t h e r e was no a p p a r e n t r e l a t i o n s h i p b e t w e e n r e l a t i v e p a l a t a b i l i t y and ADIN%DM o r ADIN%N. - 50 -1.3 Apparent D i g e s t i b i l i t y The apparent d i g e s t i b i l i t y results for the 6 silages are shown in Table 10. Table 10: Apparent d i g e s t i b i l i t y of Experiment 1 silages. Digestibility Silage Number % 1 2 3 4 5 6 S.E.! of Mean DM 72.83a 69.61b 65.47d 67.38C 62.29e 56.ll f .33** OM 74.89a 73.24b 68.73d 69.86C 63.33e 58.55f .30** N 77.93a 75.98b 59,84C 60.83C 47.78d 41.146 .40** NDF 71.689 68.11C 69.99b 69.03bc 67.97C 59.03d .45** ADF 68.89a 67.14b 68.63ab 65.02c 67.18ab 54.62d .52** ADIN -23.88C -8.17C 11.20b • -37.32d 22.30a 10.90b 2.28** ADIN%N of Silage 4.81 5.37 8.96 9.02 17.31 22.91 Means -within rows followed by diff e r e n t superscripts are d i f f e r e n t (P<.05) ** = Si g n i f i c a n t at P<.01 Since silages 1 to 6 are ranked in order of ADIN%N le v e l s , increasing ADIN%N levels appeared^to s i g n i f i c a n t l y depress the apparent d i g e s t i b i l i t y of N, DM and 0M. - 51 -1.4 C o r r e l a t i o n s and R e g r e s s i o n s B e t w e e n A p p a r e n t D i g e s t i b i l i t y M e a s u r e s and ADIN%N. T a b l e 11 i l l u s t r a t e s t h a t l e v e l o f h e a t damage e x p r e s s e d as ADIN%N was h i g h l y n e g a t i v e l y c o r r e l a t e d w i t h mos t o f t h e a p p a r e n t d i g e s t i b i l i t y f i g u r e s , p a r t i c u l a r l y w i t h t h e d i g e s t i b i l i t y o f DM ( r = - . 9 6 ) , OM ( r = - . 9 8 ) and N ( r '= - . 9 5 ) . T a b l e 1 1 : C o r r e l a t i o n s b e t w e e n ADIN%N and v a r i o u s d i g e s t i b i l i t y c o e f f i c i e n t s . (31 p a i r e d o b s e r v a t i o n s ) . D i g e s t i b i l i t y C o e f f i c i e n t C o r r e l a t i o n w i t h ADIN%N DM - . 9 6 * * - . 9 8 * * N - . 9 5 * * ADF - . 7 8 * * NDF - . 8 1 * * ADIN . 5 9 * * * * = S i g n i f i c a n t a t P<.01 R e g r e s s i o n e q u a t i o n s b a s e d on t h e s e m e a s u r e s a r e shown i n T a b l e 1 2 . - 52 -T a b l e 12 : R e g r e s s i o n s o f v a r i o u s d i g e s t i o n c o e f f i c i e n t s ( Y ) on ADIN%N ( X ) . (n = 31) Dependent Va r i ab l e Equation C o e f f i c i e n t o f Standard E r r o r o f Determinat ion the Est imate (Y) ( r 2 ) DM D i g e s t i b i l i t y Y = 74.74 - .8012(X) .91 1.68 OM D i g e s t i b i l i t y Y = 77.85 - .8559(X) .96 1.14 N D i g e s t i b i l i t y Y = 83.42 - 1.987(X) .91 4.32 ADF D i g e s t i b i l i t y Y = 71.94 - .6140(X) .61 3.29 NDF D i g e s t i b i l i t y Y = 73.50 - .5222(X) .66 2.52 ADIN D i g e s t i b i l i t y Y = 73.55 + 1.917(X) .35 17.70 ADIN%\N was a good p r e d i c t o r o f t h e a p p a r e n t d i g e s t i b i l i t y c o e f f i c i e n t s f o r a l l f r a c t i o n s w i t h t h e e x c e p t i o n o f ADIN i t s e l f . The l e v e l o f ADIN%N e x p l a i n e d 96% o f t h e v a r i a t i o n i n OM d i g e s t i b i l i t y and 91% o f t h e v a r i a t i o n i n b o t h DM and N d i g e s t i b i l i t y . - 53 -2 . EXPERIMENT 2 2.1 S i l a g e T e m p e r a t u r e The r a n g e o f t e m p e r a t u r e s r e c o r d e d o v e r t h e h e a t i n g p e r i o d a r e shown i n F i g u r e 7 . From t h e s e r e s u l t s t h e a v e r a g e amoun t s o f h e a t i n g f o r t h e two t r e a t m e n t s HI and H2 we re c a l c u l a t e d ( T a b l e 1 3 ) . T a b ! e 1 3 : A v e r a g e t e m p e r a t u r e s and l e n g t h o f h e a t i n g p e r i o d f o r t r e a t m e n t s HI and H2. T r e a t m e n t A v e r a g e T e m p e r a t u r e and L e n g t h o f P e r i o d HI H2 30 day s a t 4 9°C 33 day s a t 50°C f o l l o w e d by 30 day s a t 6 3 ° C - 54 -F i g u r e 7: T e m p e r a t u r e s r e c o r d e d i n o u t e r l a y e r and c o r e o f s i l o s HI and H2 . T 1 1 1 1 1 r 10 20 30 40 50 60 70 DAYS - 55 -2.2 E f f e c t s o f H e a t i n g on Oven DM and ADIN%N L e v e l s By t a k i n g s a m p l e s a t d i f f e r e n t d e p t h s and p o s i t i o n s w i t h i n s i l o s HI and H2, i t was p o s s i b l e t o e s t i m a t e t h e e f f e c t s o f h e a t i n g on oven DM and on ADIN%N f o r m a t i o n . The se r e s u l t s a r e s u m m a r i z e d i n F i g u r e s 8 t o H a n d T a b l e 14. T a b l e 14 : Range o f o v e n DM and ADIN%N i n s i l o s HI and H2. T r e a t m e n t Oven DM ADIN%N Number o f S a m p l e s % HI 26.6-46.1 12.5-17.2 19 H2 26.4-75.9 19.8-26.3 18 S i m i l a r v a r i a t i o n s i n o v e n DM and ADIN%N l e v e l s we re no t n o t e d w i t h i n t h e c o n t r o l s i l o C ( T a b l e 1 5 ) . T a b l e 1 5 : Oven DM and ADIN%N l e v e l s i n s i l o C. Mean V a l u e S t a n d a r d E r r o r Number o f S a m p l e s Oven DM {%) 3 0 . 7 .08 9 ADIN%N 8 .71 .15 9 H e a t i n g t e n d e d t o d r y o u t t h e o u t e r l a y e r s and f o r c e t h e m o i s t u r e t o t h e t o p o f t h e s i l o . P e r f o r a t i o n o f t h e p o l y e t h y l e n e s e a l c a u s e d by c o n t a c t w i t h t h e h e a t i n g c a b l e was a p r o b l e m i n s i l o H2. C o n s e q u e n t l y - 56 -Figure 8: Silo HI: oven dry matter at various locations (Scale 1 :10) 22.6% 29.4 [27.5[ l2"9~4" [2676 32 .5 42.5 39.2 38.7 ftTHl /32~4l ft1.5 38.8 36 .0 36.5 37.8 37 .7 37 .8 J f26T6 /46~T] •NO HEATING ' CABLES Sample Area <—37 .8—> = Area within the arrows (3 7.8] = Area within the box - 57 -Figure 9; Silo H2: oven dry matter at various locations (Scale 1:10) < 22.8% =H 26 .4 /75 .91 r3TH 128.31 30 .4 42 .6 40.3 46.7 53.1 47.0 48.2 L34 7v 45 .8 t [61 .5j IO/ /30| 1207 /61.5/ NO HEATING CABLES Sample Area 45.8'—<> = Area within the arrows = Area within the box - 58 -V Figure 10: Silo HI: ADIN%N at various locations. (Scale 1:10) 15.9% 16.7 Al7 .21 /1478 14.9 i 14.3 ^ Sample Area -12 .5—^ = Area within the arrows I n 2 .51 = Area within the box - 59 -Figure 11: Silo H2 ADIN%N at various locations (Scale .1:10) /26~3) [2470 go 27.1% 21.5 22.6 24.5 26 .2 25 .5 26.2 24. 5 /22~9l 24. 3 [22.2 /2470 frrTp] fro 3| 19 .8 rNO HEATING CABLES M Sample Area T ^—24.3—^ = Area within the arrows I (2 4. 3( = Area within the box - 60 -much m o i s t u r e was l o s t f r o m t h i s s i l o w h i c h h e l p e d t o a c c o u n t f o r t h e w i d e v a r i a t i o n s i n DM w i t h i n t h i s s i l a g e m a s s . 2 .3 S i l a g e C o m p o s i t i o n The r e s u l t s o f t h e a n a l y s i s o f t h e t h r e e s i l a g e s a r e shown in T a b l e 16 . T a b l e 1 6 : C o m p o s i t i o n o f t h e 3 s i l a g e s u s e d i n E x p e r i m e n t 2 . SILAGE S.E. of Mean C MI H2 Toluene DM (%) 34 .22 c 40.85 b 48.46 a .82 * * Composition of DM (%) Ash 6 . 82 b 6 . 73 c 6 .90 a . 02 * * NDF 72.45 a 72.12 3 71.82 a .47 n.s. ADF 44.79 c 46.63 b 49 .04 a .42 * * Hemice l l u l o se 27.66 a 25 .49 b 22.78 c .36* * Tota l N 1.48 a 1.48 a 1.40 a .28 n.s. V o l a t i l e N .16 c . 20 b .24 a . 0 1 * * ADIN .29° . 45 b . 69 a . 03 * * ADIN% o f Tota l N 8 . 71 C 14.09 b 24.32 a 1.15** a , b ' c m e a n s w i t h i n rows d i f f e r e n t ( P< . 05 ) f o l 1 owed by d i f f e r e n t s u p e r s c r i p t s a r e * * = S i g n i f i c a n t a t P<. 0 1 ; n . s . = i o t s i g n i f i c a n t - 61 -H e a t i n g s i g n i f i c a n t l y (P<.01) i n c r e a s e d DM, ADF, v o l a t i l e N, ADIN and ADIN%N, w h i l e i t s i g n i f i c a n t l y (P<.01) r e d u c e d h e m i c e l l u l o s e l e v e l s . D i f f e r e n c e s b e t w e e n t o t a l N were n o t s i g n i f i c a n t . T h e r e we re m a r k e d d i f f e r e n c e s i n v i s u a l a p p e a r a n c e b o t h w i t h i n and b e t w e e n s i l a g e s . Whereas t r e a t m e n t C was a u n i f o r m y e l l o w / l i g h t b rown c o l o u r , t h e a v e r a g e d a r k n e s s o f t h e b rown c o l o u r a t i o n i n c r e a s e d w i t h h e a t i n g . Thus H2 on a v e r a g e was d a r k e r b rown t h a n H I , w h i c h was s u b s t a n t i a l l y d a r k e r t h a n C. The b r owned s i l a g e , p a r t i c u l a r l y t r e a t m e n t H2 , had t h e c h a r a c t e r i s t i c b u r n t o r t o b a c c o - l i k e a r o m a . 2.4 D i g e s t i b i l i t y T r i a l 2 . 4 . 1 P a l a t a b i l i t y and I n t a k e A l l t h r e e s i l a g e s were con sumed i n d i s a p p o i n t i n g l y s m a l l a m o u n t s . D u r i n g t h e p r e - t r i a l a d a p t a t i o n p e r i o d , t h e amount o f s i l a g e o f f e r e d had t o be c o n t i n u a l l y a d j u s t e d downwards f r o m t h e i n i t i a l t a r g e t i n t a k e o f 1200g DM. D e s p i t e t h e i r l o w DM i n t a k e s , t h e ewes n e v e r showed s i g n s o f h u n g e r and seemed c o n t e n t e d . Q u a l i t a t i v e and q u a n t i t a t i v e m e a s u r e s o f p a l a t a b i l i t y and i n t a k e a r e shown i n T a b l e 1 7 . H e a t i n g a p p e a r e d t o i n c r e a s e t h e r e l a t i v e p a l a t a b i l i t y o f t h e s i l a g e , j u d g e d by f e e d r e f u s a l s and t h e s p e e d w i t h w h i c h t h e ewes a t e t h e i r a l l o w a n c e . The ewe whose a v e r a g e d a i l y DM i n t a k e o f s i l a g e H2 was o n l y 5 5 0 g , was u n w e l l - 62 -T a b l e 17 : I n t a k e and p a l a t a b i l i t y o f E x p e r i m e n t 2 s i l a g e s . DM O f fe red DM Consumed No. o f Ewes who Re l a t i v e Mpan Ranap Ate Tota l Al lowance P a l a t a b i l i t y g g g Treatment C 821 754 596-821 2 Poor HI 820 775 682-820 3 Poor H2 872 797 550-872 4 Fai r d u r i n g mo s t o f t h i s t r i a l and s u f f e r e d a d r a s t i c l o s s o f body w e i g h t ( s e e s e c t i o n 2 . 4 . 2 - ewe number 4 ) . 2 . 4 . 2 Ewe Body H e i g h t Change The t r i a l s w e r e r u n i n t h e o r d e r H I , C and H2. Body w e i g h t s w e r e r e c o r d e d a t t h e v e r y b e g i n n i n g o f t h e e x p e r i m e n t , a f t e r t r i a l C, and f i n a l l y a f t e r t r i a l H2 ( s e e F i g u r e 12 ) . The a v e r a g e l o s s i n body w e i g h t o v e r t h e 9 week e x p e r i m e n t a l p e r i o d was 8kg w h i c h i n d i c a t e s t h a t t h e d r y m a t t e r o f f e r e d was c o n s i d e r a b l y b e l o w m a i n t e n a n c e r e q u i r e m e n t s . The p o o r d r y m a t t e r i n t a k e s o f ewes 3 and 4 a r e r e f l e c t e d i n t h e i r d r a m a t i c l o s s o f body w e i g h t . On a v e r a g e , t h e o t h e r ewes con sumed a l l f e e d o f f e r e d , b u t were u n w i l l i n g t o e a t s i g n i f i c a n t f u r t h e r a m o u n t s . Ewe 4 was u n w e l l d u r i n g mos t o f t r i a l H2. He r v e r y l o w d r y m a t t e r i n t a k e r e s u l t e d i n a d r a s t i c l o s s o f body w e i g h t o v e r t h e f i n a l 3 w e e k s . - 63 -F i g u r e 12 : Ewe body w e i g h t c h a n g e d u r i n g E x p e r i m e n t 2. TREATMENT -t 1 r 3 ' 6 9 Weeks On a v e r a g e , t h e g r e a t e s t w e i g h t l o s s o c c u r r e d w i t h t h e s e v e r e l y h e a t e d s i l a g e H2. T h i s was t h e a l s o t h e s i l a g e w i t h t h e g r e a t e s t a v e r a g e i n t a k e . - 64 -2.4.3 Apparent D i g e s t i b i l i t y Apparent d i g e s t i b i l i t y c o e f f i c i e n t s for the three treatments are shown in Table 18. Table 18: Apparent d i g e s t i b i l i t y c o e f f i c i e n t s for the three treatments in Experiment 2. Di ges t i bi 1 i ty Treatment % C HI H2 S.E. of Mean DM 53 .00b 55 .16a 51.26° .22** OM 53.31 b 55.6l a 51.65C _ 24** N 62.93 a ' 57.24 b 39.07 C .43** ADF 47 .65° 53 .39a 49.66b .33** NDF 50.73 C 55.08a 52.18 b .34** ADIN 7.47C 15.39b 23.19 a 1.80** ADIN%N of Silage 8.71° 14.09b 24.32a 1. 15** 3 b c ' ' 'means within rows followed by different superscripts are diffe r e n t (P<.05) ** = Sig n i f i c a n t at P<. 01 A l l apparent d i g e s t i b i l i t y values varied s i g n i f i c a n t l y (P<.05) with the heat treatments. Heating depressed nitrogen d i g e s t i b i l i t y and the depression increased with the severity of heating. As previously shown, heating increased ADIN levels and the apparent d i g e s t i b i l i t y of the ADIN fra c t i o n increased with heating. The d i g e s t i b i l i t y of the f i b r e fraction (NDF and ADF) increased s i g n i f i c a n t l y (P<.05) with moderate heating (HI compared to C). More - 65 -s e v e r e h e a t i n g (H2) c a u s e d t h e d i g e s t i b i l i t y o f t h e s e f r a c t i o n s t o f a l l , a l t h o u g h t o l e v e l s t h a t we re s t i l l s i g n i f i c a n t l y ( P< . 05 ) h i g h e r t h a n t h o s e o f t h e u n h e a t e d s i l a g e (C).. 2 . 4 . 4 C o r r e l a t i o n s B e t w e e n A p p a r e n t D i g e s t i b i l i t y  M e a s u r e s and ADIN%N. T a b l e 19 i l l u s t r a t e s t h a t l e v e l o f h e a t damage e x p r e s s e d as ADIN%N was s i g n i f i c a n t l y ( P < . 01 ) and n e g a t i v e l y c o r r e l a t e d w i t h a p p a r e n t n i t r o g e n d i g e s t i b i l i t y ( r = - . 9 8 ) and p o s i t i v e l y c o r r e l a t e d ( P< . 01 ) w i t h t h e a p p a r e n t d i g e s t i b i l i t y o f ADIN ( r = . 8 4 ) . C o r r e l a t i o n s w i t h o t h e r p a r a m e t e r s were n o t s i g n i f i c a n t w i t h t h e e x c e p t i o n o f DM d i g e s t i b i l i t y ( r = - . 4 7 ) ( P < . 0 5 ) . T a b l e 19 : C o r r e l a t i o n s b e t w e e n ADIN%N and v a r i o u s d i g e s t i b i l i t y c o e f f i c i e n t s (18 p a i r e d o b s e r v a t i o n s ) D i g e s t i b i l i t y C o e f f i c i e n t C o r r e l a t i o n w i t h ADIN%N DM OM N ADF NDF ADIN - . 4 7 * - . 4 4 - . 9 8 * * . 15 .11 . 8 4 * * * = s i g n i f i c a n t a t P < . 05 ; * * = s i g n i f i c a n t a t P<.01 - 66 -2.4.5 Nitrogen U t i l i z a t i o n The nitrogen u t i l i z a t i o n results are summarized in Table 20. Table 20: Effects of Experiment 2 treatments on nitrogen u t i l i z a t i o n . - I tern Nitrogen (g/day) C Treatment H_l H2 Correlati on with ADIN%N Intake 11.343 11.56a 11.28a -.04 Feces 4.22c 4.95b 6.88a . 98** Urine 6.18a 5.67b 4.25C -.81** Retenti on .94a .94a .15b - .49* N Retention (% of N Digested) 13.209 14.2 2a 6 .82b -.42 a , b, c • • i • ' 'means within rows fol1 owed by di f fe rent superscripts are different (P< .05) * = Si gni f i can t at P<.05 ; ** = si gni f i cant at P<.01 With similar nitrogen intakes, sheep fed the heated silages excreted 17 to 63% more nitrogen in their feces (P<*05), but 8 to 31% less nitrogen in their urine (P<.05) as compared to when they received the control silage (C). ADIN%N. was s i g n i f i c a n t l y ( P<. 01) p o s i t i v e l y correlated with fecal nitrogen (r = .98) and negatively correlated with urinary nitrogen (r = -.81). Overall nitrogen retention was unaffected by moderate heating (treatment HI), but was s i g n i f i c a n t l y (P<.05) depressed with more - 67 -severe heating (treatment H2). Moderate heating (HI) s l i g h t l y increased the e f f i c i e n c y of u t i l i z a t i o n of digested nitrogen although the difference was not s i g n i f i c a n t , whereas more severe heating (H2) s i g n i f i c a n t l y (P<.05) reduced this e f f i c i e n c y . 2.4.6 Regression Equations Regression equations based on correlations between ADIN%N and various measures of apparent d i g e s t i b i l i t y and nitrogen u t i l i z a t i o n are shown in Table 21. Table 21: Regressions of various digestion c o e f f i c i e n t s and measures of nitrogen u t i l i z a t i o n (Y) on ADIN%N (X). (n = 18). Dependent Variable Equation Coefficient of Standard Error of Determination Retained N as % of N Digested Y = 41.38 - .7442(X) ,17 the Estimate (Y) (r 2) DM Digestibility Y = 55.45 - .1469(X) .23 1.87 OM Digestibility Y = 55.78 - .1433(X) .19 2.03 N Digestibility Y = 77.60 - 1.561(X) .96 2.29 ADF Digestibility Y = 49.23 + .0637(X) .02 2.92 NDF Digestibility Y = 51.98 + .0434(X) .01 2.75 ADIN Digestibility Y = .3764 + .9749(X) .71 4.31 Fecal N Y = 8.878 + .0374(X) .96 .13 Urinary N Y = 7.341 - .1255(X) .66 .62 Retained N Y = 2.523 - .0542(X) .24 .66 11.11 - 68 -In contrast to Experiment 1, ADIN%N was a poor predictor of apparent d i g e s t i b i l i t y for a l l components with the notable exception of nitrogen for which ADIN%N explained 96% of the variation in nitrogen d i g e s t i b i l i t y between the treatments. 3. REGRESSION EQUATIONS BASED ON POOLED DATA Since both Experiments 1 and 2 used orchardgrass silages in similar d i g e s t i b i l i t y t r i a l s with sheep, i t was decided to combine the observations in order to calculate regressions of apparent d i g e s t i b i l i t y c o e f f i c i e n t s on ADIN%N for the pooled data (Table 22). Table 22: Regressions of various digestibility coefficients (Y) from! Experiments 1 and 2 on ADIN%N (X). (n = 49). Dependent Variable Equation Coefficient of Standard Error Determination the Estimate (Y) (r 2) DM Digestibility Y = 71.48 - .8053(X) .52 5.43 OM Digestibility Y = 74.23 - .8859(X) .50 6.25 N Digestibility Y = 81.76 - 1.830(X) .92 3.89 ADF Digestibility Y = 68.38 - .6832(X) .31 7.21 NDF Digestibility Y = 70.35 - .6407(X) .29 7.04 ADIN Digestibility Y = 78.16 + 1.892(X) .42 15.72 Using the pooled data, ADIN%N explained 92% of the variation in nitrogen di ges t i bi1i ty However, i t expl ai ned only 50 and 52% of the variation in the d i g e s t i b i l i t y of - 69 -OM and DM respectively, and i t was a r e l a t i v e l y poor predictor for the d i g e s t i b i l i t y of ADF, NDF and ADIN. - 70 -DISCUSSION Heating of forages encourages the Maillard or non-enzymic browning reaction. This reaction involves the condensation of sugar residues with amino acids followed by polymerization to form a brown a r t i f a c t l i g n i n containing about 11% nitrogen (Van Soest, 1982). This fibre a r t i f a c t is i d e n t i f i e d in the ADIN component during feed analysis and is largely i n d i g e s t i b l e . Thus ^ heat damage leads to a decrease in the apparent d i g e s t i b i l i t y of feedstuffs. Protein d i g e s t i b i l i t y i s normally the 1 factor most severely depressed. The purpose of these experiments was two-fold. F i r s t relate heat damage expressed as ADIN%N to the apparent d i g e s t i b i l i t y of various fractions in grass silages, and second, to investigate the f e a s i b i 1 i t y of a r t i f i c i a l l y inducing heat damage in an already fermented grass silage and then rel a t i n g this heat damage to i t s apparent d i g e s t i b i l i t y and nitrogen u t i l i z a t i o n . The results of the two experiments are discussed below. EXPERIMENT 1 Despite marked differences in composition, degree of preservation and type of fermentation between the six experimental silages, highly s i g n i f i c a n t (P<.01) negative correlations existed between ADIN%N and the apparent d i g e s t i b i l i t y c o e f f i c i e n t s of DM, OM, N, ADF and NDF (Table 11). Similar findings were reported by Yu and - 71 -Veira (1977), and many workers have shown a close link between measures of ADIN and reduced protein d i g e s t i b i l i t y (Goering et a l . , 1972; Gordon, 1968; Thomas et a l . , 1972; Yu & Thomas, 1975) . ADIN%N explained 96% of the variation in OM d i g e s t i -b i l i t y and 91% of the variation in both DM and N d i g e s t i -b i l i t y (Table 12). The regression equation for total N d i g e s t i b i l i t y based on ADIN%N was in f a i r l y close agreement with those of Thomas et al . ( 1972) and Yu and Thomas ( 1976 ), who studied a variety of a l f a l f a hay and haylage samples (Figure 13). The prediction equation formulated by Goering et al . ( 1972 ) was in less close agreement particu-l a r l y at ADIN%N levels in excess of 20%, but this was probably due to the much wider range of heat damaged material used in th e i r study (ADIN%N ranged between 1.78% and 74.54% compared with 4.8-22.9% in the present study). When the regression equation of DM d i g e s t i b i l i t y on ADIN%N from Experiment 1 was compared to that reported by Yu and Veira (1977), differences were more apparent (Table 23), although the slopes of the two lines were simi l a r (Figure 14). The difference was possibly Table 23: Regressions of DM d i g e s t i b i l i t y c o e f f i c i e n t s (Y) on ADIN%N (X). Source Equation 2 Range of ADIN%N n r Experiment 1 31 Y = 74.74 - .8(X) .91 4.8 - 22.9 Yu & Veira (1977) 9 Y = 67.24 - .73(X) .80 7.7 - 24.1 - 72 -F i g u r e 1 3 : R e g r e s s i o n s o f t o t a l n i t r o g e n d i g e s t i b i l i t y c o e f f i c i e n t s ( Y ) on ADIN%N (X ) f r o m v a r i o u s s o u r c e s . 80 " Qj , , , — , 1 X 5 10 15 20 25 30 ADI MSN - 73 -F i q u r e 1 4 : R e g r e s s i o n s o f DM d i g e s t i b i l i t y c o e f f i c i e n t s (Y ) — on ADI N%N (X ) f r o m two s o u r c e s . 751 70 651 DM DIGESTIBILITY 6 0 % 55 50 S l o p e # 1 Sou r ce E x p e r i m e n t 1 1977 -1— 10 i— 15 20 —i 25 ADIN5&N - 74 -due to experimental technique and/or type of forage used in that Yu and Veira used one a l f a l f a haylage (47% DM) which they a r t i f i c i a l l y heat damaged to varying l e v e l s , while Experiment 1 used naturally heat damaged orchard-grass/clover silages from varying sources. The negative d i g e s t i b i l i t y figures for ADIN in silages 1, 2 and 4 (Table 10) were probably a result of oven drying the feces at 90°C for 48 hours. These high quality silages with low ADIN l e v e l s , but high N contents, most l i k e l y produced feces that were capable of being s i g n i f i c a n t l y heat damaged by oven drying. Van Soest (1965) found that drying sheep and cow feces in a forced ai r oven at temperatures in excess of 50°C led to s i g n i f i c a n t increases in ADIN levels within his samples. Although measures of p a l a t a b i l i t y were largely subjective, heat damage appeared to have no effect on p a l a t a b i l i t y for the silages studied (Table 9). Of far greater consequence was the type of fermentation and the levels of NDF within each si l a g e . Silages 3 and 5 had the highest NDF contents and appeared to have the lowest voluntary intake. Van Soest (1966) reported that increasing NDF levels in forages led to corresponding decreases in intake and that NDF gave the highest squared correlation c o e f f i c i e n t (.58) for voluntary intake of fo rages. - 75 -. EXPERIMENT 2 In o r d e r t o g a i n a c l e a r e r u n d e r s t a n d i n g o f t h e e f f e c t s o f h e a t damage on t h e a p p a r e n t d i g e s t i b i l i t y c o e f f i c i e n t s and n i t r o g e n u t i l i z a t i o n o f g r a s s s i l a g e , i t was d e c i d e d t o c a r r y o u t a f u r t h e r e x p e r i m e n t i n w h i c h one s i l a g e w o u l d be a r t i f i c i a l l y h e a t e d t o i n c r e a s e t h e ADIN%N l e v e l s . In t h i s way i t was hoped t o a v o i d t h e p r o b l e m o f t h e v a r i a b l e b a c k g r o u n d o f s i l a g e s a s s o c i a t e d w i t h E x p e r i m e n t 1. 1 E x p e r i m e n t a l S i l a g e A c o n c e r n t h r o u g h o u t t h e e x p e r i m e n t was t h e v e r y l o w p r o t e i n c o n t e n t and h i g h f i b r e l e v e l s o f t h e s e l e c t e d s i l a g e . O b v i o u s l y t h i s meant t h a t t h e amount o f p r o t e i n a v a i l a b l e t o be h e a t damaged was l i m i t e d . The r e s u l t s o f t h e s t u d y we re s t i l l v a l u a b l e , h o w e v e r , s i n c e s i l a g e s w i t h l o w p r o t e i n and h i g h f i b r e l e v e l s a r e n o t uncommon i n B .C . 2 E x p e r i m e n t a l S i l o s The use o f t h e p a p e r f i b r e f o r m s h e l l s was c o n s i d e r e d t o be a g r e a t s u c c e s s . Each c y l i n d e r h e l d a :more t h a n a d e q u a t e amount o f s i l a g e f o r a 3 t o 4 week f e e d i n g t r i a l f o r 6 s h e e p . The c y l i n d e r s w e r e e a s i l y t r a n s p o r t e d on p a l l e t s f r o m one l o c a t i o n t o a n o t h e r . The f i l l i n g o f them was v e r y s t r a i g h t f o r w a r d and i t was a s i m p l e m a t t e r t o c o m p a c t t h e f o r a g e m a n u a l l y t h r o u g h o u t t h e f i l l i n g - 76 process. In this way maximum exclusion of a i r was achieved and was maintained by the use of the polyethylene l i n i n g which was custom made. One small problem encountered was that excessive rocking of the f i l l e d s i l o s during the 32km journey back to U.B.C. encouraged the silage within the polyethylene l i n i n g to work i t s way out at the base of the cylinder. This could be prevented in future t r i a l s by fastening some sort of base to the cylinder. The heating method e a s i l y developed s u f f i c i e n t heat to achieve the desired temperatures within s i l o s HI and H2. Indeed a complication was local perforation of the polyethylene seal caused by direct contact with the heating cable, p a r t i c u l a r l y for treatment H2. In view of this i t was f e l t that any future investigations should proceed with the heating cables coiled around the outside of the fi b r e form cylinder. By careful coordination between the thermostat and the temperatures.as recorded by the thermistor probes, i t was possible to maintain reasonably constant temperatures within the heated silages. After the i n i t i a l warm-up period, there were no differences in temperature between the core and outer layer of s i l o HI (Figure 7). The same was not true for s i l o H2 where a 9 to 12°C difference existed throughout the major part of the treatment. The reasons for the difference are not known although i t may have been due to differences in the location of the probes (the outer probe for H2 may have been closer to a heating - 77 -c a b l e t h a n t h a t f o r H I ) . A g a i n i t may have been c a u s e d by t h e p e r f o r a t i o n o f t h e p o l y e t h y l e n e s e a l i n s i l o H2 r e s u l t i n g i n a v e r y d r y o u t e r l a y e r w i t h a l o w e r s p e c i f i c h e a t c a p a c i t y t h a n t h e damper c o r e , a l t h o u g h t h i s i s u n l i k e l y s i n c e t h e t e m p e r a t u r e d i f f e r e n c e e x i s t e d f r o m t h e v e r y s t a r t o f t h e t r i a l o n w a r d s . 2 . 3 E f f e c t s o f H e a t i n g on Oven DM o f S i l a g e M i g r a t i o n o f m o i s t u r e w i t h i n t h e h e a t e d s i l o s was a p r o b l e m p a r t i c u l a r l y f o r t r e a t m e n t H2 whose p e r f o r a t e d s e a l a l l o w e d a c o n s i d e r a b l e amount o f m o i s t u r e t o e s c a p e ( F i g u r e 9 ) . I n d e e d i t was p o s s i b l e on c o o l m o r n i n g s t o see c o n d e n s e d w a t e r v a p o u r e s c a p i n g f r o m t h e t o p o f t h i s s i l o . F o l l o w i n g u n l o a d i n g , v e r y c a r e f u l and t h o r o u g h m i x i n g was e s s e n t i a l i n o r d e r t o e n s u r e u n i f o r m d a i l y DM i n t a k e s f o r t h e d i g e s t i b i l i t y t r i a l s . V a r i a t i o n s i n ADIN%N showed no c o n s i s t e n t r e l a t i o n -s h i p w i t h m o i s t u r e c o n t e n t ( F i g u r e s 8 , 9 , 10 and 1 1 ) . O t h e r w o r k e r s have r e p o r t e d t h a t s i l a g e DM c o n t e n t had r e l a t i v e l y l i t t l e e f f e c t on ADIN p r o d u c t i o n o v e r t h e r a n g e 2 5 - 5 9 % ( M i d d l e t o n & Thomas , 1983) and 3 0 - 7 0 % ( G o e r i n g e t a l . , 1 9 7 3 ) . They c o n c l u d e d t h a t above and b e l o w t h e s e l e v e l s , h e a t damage t e n d s t o be r e d u c e d . T h e r e f o r e i t i s o f i n t e r e s t t h a t t h e s a m p l e i n t r e a t m e n t H2 w i t h a DM c o n t e n t o f 7 5 . 9 % had a c o n s i d e r a b l y l o w e r l e v e l o f ADIN%N t h a n t h e a v e r a g e ( 1 9 . 8 % v s . 2 4 . 3 % ) and i n d e e d had t h e l o w e s t l e v e l o f h e a t damage o f a l l s a m p l e s f r o m t h i s t r e a t m e n t . - 78 -2 .4 E f f e c t s o f H e a t i n g on S i l a g e C o m p o s i t i o n ( T a b l e 16) The i n c r e a s e i n ADIN w i t h i n c r e a s e s i n amount o f h e a t i n g was i n a g r e e m e n t w i t h numerous o t h e r s t u d i e s ( G o e r i n g e t a l . , 1 9 7 3 ; M i d d l e t o n & Thomas , 1 9 8 3 ; Van S o e s t , 1 9 6 5 ; Yu & Thomas , 1 9 7 5 ; Yu & V e i r a , 1 9 7 7 ) . I n c r e a s e d ADIN r e s u l t s f r o m n o n - e n z y m i c b r o w n i n g r e a c t i o n s r e s u l t i n g i n t h e f o r m a t i o n o f an a r t i f a c t l i g n i n f r o m p r o t e i n s and s u g a r r e s i d u e s ( i n c l u d i n g h e m i e e l 1 u l o s e s ) ( M i d d l e t o n & Thomas , 1 9 8 3 ) . T h i s w o u l d h e l p t o a c c o u n t f o r t h e i n c r e a s e i n ADF and d e c r e a s e i n h e m i e e l 1 u l o s e a s s o c i a t e d w i t h i n c r e a s e d h e a t e x p o s u r e . Van S o e s t ( 1 9 6 5 ) d e m o n s t r a t e d a c l o s e r e l a t i o n s h i p b e t w e e n t h e e x t e n t o f h e a t i n g u n d e r l a b o r a t o r y c o n d i t i o n s and v a l u e s f o r ADF: f o r e x a m p l e , when 2g s a m p l e s o f a f r e s h o r c h a r d - ! a d i n o m i x t u r e ( 20% DM) we re d r i e d i n an open d i s h a t 100°C f o r 20 h o u r s , ADF l e v e l s i n c r e a s e d f r o m 3 1 . 1 % ( u n h e a t e d s a m p l e s ) t o 3 6 . 8 % . The f a l l i n h e m i e e l 1 u l o s e w i t h r i s e i n ADIN was i n c l o s e a g r e e m e n t w i t h e a r l i e r work by G o e r i n g e t a l . ( 1 9 7 3 ) and Yu and V e i r a ( 1 9 7 7 ) . The i n c r e a s e i n v o l a t i l e o r s o l u b l e n i t r o g e n w i t h h e a t i n g was e x p e c t e d . A l t h o u g h work w i t h a r t i f i c i a l l y d e h y d r a t e d a l f a l f a and g r a s s i n d i c a t e s t h a t v o l a t i l e n i t r o g e n d e c r e a s e s w i t h i n c r e a s e s i n maximum t e m p e r a t u r e and l e n g t h o f h e a t i n g p e r i o d ( G o e r i n g & W a l d o , 1 9 7 8 ; H a r t m a n n ( 1 9 7 3 ) , Yu and Thomas ( 1 9 7 5 ) , w o r k i n g w i t h i s i l a g e , f o u n d t h a t v o l a t i l e n i t r o g e n i n c r e a s e d f r o m 1.27% t o 1.35% w i t h an i n c r e a s e i n ADIN%N f r o m 9 .6% t o - 79 -1 5 . 4 % w i t h i n t h e s a m e s i l o . T h e a m o u n t o f v o l a t i l e n i t r o g e n u s u a l l y r e f l e c t s t h e e x t e n t o f f e r m e n t a t i o n o r p r o t e i n d e g r a d a t i o n ( W a l d o , 1 9 7 3 ) . I t i s l i k e l y t h a t h e a t i n g c a u s e d s o m e p r o t e i n d e g r a d a t i o n t o v o l a t i l e n i t r o g e n w h i c h w a s u n a b l e t o e s c a p e i n t h e s e a l e d c o n d i t i o n s w i t h i n a s i l o . T o t a l n i t r o g e n c o n t e n t w a s n o t a f f e c t e d ( P < . 0 5 ) b y e i t h e r o f t h e h e a t t r e a t m e n t s . T h i s h a s b e e n o b s e r v e d i n o t h e r s t u d i e s w i t h h e a t d a m a g e d f o r a g e s ( B e c h t e l e t a l . , 1 9 4 3 , 1 9 4 5 ; G o r d o n , 1 9 6 7 ; H a r t m a n n , 1 9 7 3 ; M i l l e r e t a l . , 1 9 6 7 ; N e l s o n , 1 9 6 6 ; Y u & V e i r a , 1 9 7 7 ) . I t i s i m p o s s i b l e t o s a y w h e t h e r t h e i n c r e a s e i n A D I N l e v e l s o f t r e a t m e n t H 2 c o m p a r e d w i t h t r e a t m e n t H I w a s d u e t o t h e l o n g e r h e a t i n g p e r i o d o r t h e h i g h e r m a x i m u m t e m p e r a t u r e a s t h e s e e f f e c t s w e r e n o t i s o l a t e d . T h e e f f e c t o f t h e d e g r e e o f h e a t i n g o n t h e e x t e n t o f A D I N f o r m a t i o n i s d i f f i c u l t t o c o m p a r e w i t h o t h e r p u b l i s h e d r e s u l t s s i n c e m o s t o t h e r w o r k h a s c o n c e n t r a t e d o n s h o r t e r d u r a t i o n , b u t h i g h e r t e m p e r a t u r e t r e a t m e n t s . I n d e e d m o s t o f t h e s e s t u d i e s h a v e c o n c l u d e d t h a t w i t h s h o r t d u r a t i o n h e a t i n g t h e r e i s l i t t l e i f a n y i n c r e a s e i n A D I N f o r m a t i o n a t t e m p e r a t u r e s b e l o w 6 0 ° C ( G o e r i n g e t a l . , 1 9 7 3 ; M i d d l e t o n & T h o m a s , 1 9 8 3 ; Y u , 1 9 7 6 ) . U n d e r t h e s e c i r c u m s t a n c e s o n e w o u l d h a v e e x p e c t e d v i r t u a l l y n o r i s e i n A D I N l e v e l s f o r t r e a t m e n t H I w h e r e a v e r a g e t e m p e r a t u r e n e v e r r o s e a b o v e 5 0 ° C a n d t h e m a x i m u m t e m p e r a t u r e r e c o r d e d w a s 5 4 ° C ( F i g u r e 7 ) . I n f a c t A D I N % N l e v e l s i n t r e a t m e n t H I r o s e b y o v e r 6 0 % w h e n c o m p a r e d t o t h e u n h e a t e d c o n t r o l - 80 -treatment C. This result was in agreement with e a r l i e r work by Yu and Thomas (1975) who found that ADIN in samples exposed to 48°C for 6 weeks was approximately twice that of haylage from the same s i l o at 33°C. 2.5 Effects of Heating on P a l a t a b i l i t y and Intake (Table 17) The very low intakes had l i t t l e to do with heating since the problem was common to a l l three treatments. Milford and Minson (1965) demonstrated a precipitous decline in intake when forage crude protein levels f e l l below 7% of DM. In the present study, i f one subtracts the e s s e n t i a l l y i n d i g e s t i b l e ADIN from total N to calculate crude protein, the adjusted crude protein levels for treatments C, HI and H2 were only 7.4%, 6.4% and 4.4% respectively. The minimum crude protein require-ment for rumen bacteria is 8% (Van Soest, 1982). Nitrogen deficiency coupled with the very high f i b r e levels associated with the silages would most l i k e l y have caused a very slow rate of fermentation in the rumen. Certainly i t was true that, despite their low intakes, the. sheep never appeared to be hungry although they a l l lost a considerable amount of weight (Figure 12). Mertens cited by Van Soest (1982), investigated 187 dif f e r e n t forages and found a close negative correlation (r = -.76) between NDF levels and intake. The very high levels of NDF in the experimental silages would thus have been expected to cause low intakes. - 81 -The apparently higher voluntary intake associated with treatment H2, which was the f i n a l silage to be fed in Experiment 2, may have been due to stretching of the rumen wall into the abdominal cavity, thus allowing a greater intake into the expanded rumen. This e f f e c t has been demonstrated by Welch (1967) who introduced poly-propylene fibres into the rumen of sheep to decrease the e f f e c t i v e size of the rumen and therefore decrease intake. He found that after about 20 days the rumen had stretched to a size s u f f i c i e n t to allow near normal intakes. Silage H2 started to be fed 40 days after the commencement of Experiment 2. Since the sheep were not given any choice of forage, i t is d i f f i c u l t to objectively assess p a l a t a b i l i t y . Subjective observations tended to suggest that heating increased p a l a t a b i l i t y although this may have been due to higher intakes as a result of the rumen stretching outlined above. This observation runs counter to results obtained by Bechtel et a l . (1943, 1945) and Yu and Veira (1977). The l a t t e r study quantitatively compared palata-b i l i t y of an a l f a l f a haylage a r t i f i c i a l l y heat damaged to ADIN%N levels of 7.7, 15.2 and 24.1%, in cafeteria t r i a l s with sheep. They found that sheep consumed about ten times more of the unheated forage (7.7% ADIN%N) compared to the same forage heated to 88°C for 48 hours (24.1% ADIN%N) and that intake was closely and negatively correlated to ADIN%N (r = -.78 to -.85). They suggested - 82 -that b i t t e r compounds produced by the Maillard reaction decreased p a l a t a b i l i t y . With no choice, however, the voluntary feed intake of their sheep was the same for a l l three treatments. In the present t r i a l , the sheep were never given a c a f e t e r i a - s t y l e choice. 2.6 The Effects of Heating on Apparent D i g e s t i b i l i t y Coefficients (see Tables 18 and 19) The effects of the heat treatment on apparent d i g e s t i -b i l i t y values were far from clear since moderate heating (treatment HI) increased the apparent d i g e s t i b i l i t y of DM, OM, ADF and NDF when compared to the unheated treatment C and the more severely heated treatment H2. Indeed, treatment C had the lowest apparent d i g e s t i b i l i t y figures for both ADF and NDF. As expected, the apparent d i g e s t i b i l i t y of nitrogen was markedly reduced with increasing severity of heat treatment and a close negative correlation with ADIN%N existed (r = -.98). This was in agreement with several other studies (Goering et a l . , 1972; Yu & Thomas, 1975, 1976 ; Yu & Vei ra, 1977) . The fact that the apparent d i g e s t i b i l i t y of ADIN increased with heating indicates that i t s fate within the digestive tract is far from clear. Yu and Veira (1977) in a si m i l a r study found that the d i g e s t i b i l i t y of ADIN varied from 11.3% to 22.4% and was unrelated to either extent of heating or, therefore, to ADIN levels in the dry matter. In the present study, a close positive - 83 -correlation (r = .84) existed between ADIN%N and the d i g e s t i b i l i t y of ADIN. The variation in this measure between sheep for each t r i a l was wide, but was i n s u f f i c i e n t to affect the significance of this result. The rise in the apparent d i g e s t i b i l i t y of OM and DM with moderate heating (treatment HI) was probably a result of the unexpected increase in fibre d i g e s t i b i l i t y for this treatment. It would appear that any increase in fibr e d i g e s t i b i l i t y was i n s u f f i c i e n t to overcome the dramatic f a l l in nitrogen d i g e s t i b i l i t y for treatment H2. Consequently, the apparent d i g e s t i b i l i t y of DM and OM was lower for treatment H2 than for treatment C. The anomalous rise in OM and DM d i g e s t i b i l i t y associated with moderate heating (HI) meant that these measures were r e l a t i v e l y poorly correlated with ADIN%N when compared to the results obtained in Experiment 1 (see Tables 11 and 19) . The increase in the apparent d i g e s t i b i l i t y of fibre with both moderate heating (HI) and more severe heating (H2) is d i f f i c u l t to explain. Yu and Veira (1977) working with 47% DM a l f a l f a haylage sealed in 37.81 containers and heated in a forced a i r oven at 88°C for 0, 24 or 48 hours, found that, as in the present study, heating increased ADF levels (from 38.9 to 44.9 and 46.3% of the DM) but decreased ADF d i g e s t i b i l i t y from 54.4 to 51.3 and 46.2% respectively (P<.05). - 84 Heating under the moisture-confining conditions within a sealed s i l o may lead to some sort of "steam cooking" process which increases the d i g e s t i b i l i t y of components within the fibr e f r a c t i o n . Bender et a l . ( 1970) investigated the potential of a steam heat treatment for increasing the dry matter d i g e s t i b i l i t y of wood, which i s of course largely composed of f i b r e . Exposing samples of aspen to steam at 140°C for 1 hour increased the DM d i g e s t i b i l i t y from 9.2 to 43.3%. This temperature was twice as high as any used in the present study, but lower temperatures over much longer periods of time may cause a s i g n i f i c a n t , a l b e i t small, increase in f i b r e d i g e s t i b i l i t y . The f a l l in fibre d i g e s t i b i l i t y with more severe heating (H2) when compared to treatment HI may have been due to the increased formation of a r t i f a c t l i g n i n (ADF content of the DM for HI and H2 were 46.6 and 49.0% respectively at a significance of P<.01). Thus the difference in the d i g e s t i b i l i t y of the f i b r e fraction less the a r t i f a c t l i g n i n formed by heating may have been negligible between these two treatments. It is more l i k e l y , however, that the f a l l in f i b r e d i g e s t i b i l i t y was due to the lower levels of available nitrogen associated with treatment H2, since the minimum crude protein requirement for rumen bacteria is 8% (Van Soest, 1982) and the available protein in treatment H2 was about 4.4% (Available Protein = (Total N - ADIN) X 6.25). - 85 -F i n a l l y , the s l i g h t increase in soluble or v o l a t i l e nitrogen associated with increased heating (Table 16) may have had a small effect on f i b r e digestion by increasing the level of rumen-degradabl.e nitrogen for the rumen bacteria. In contrast to Experiment 1, ADIN%N was a poor predictor of apparent d i g e s t i b i l i t y for a l l components except total nitrogen and ADIN (Table 21). The level of ADIN%N explained 96% of the variation in nitrogen d i g e s t i b i l i t y between the treatments, whereas i t explained only 23% and 19% of the variation in the d i g e s t i b i l i t y of DM and OM respectively. This was presumably because of the unexpected increase.in the d i g e s t i b i l i t y of ADF and NDF associated with heating. The prediction equation for nitrogen d i g e s t i b i l i t y based on ADIN%N was in good agreement with the results from Experiment 1. Figure 15 shows the regression equations for nitrogen d i g e s t i b i l i t y predicted by ADIN%N from the present study and from two other sources. The data for slope #1 (Thomas et a l . , 1972) was based on samples from 5 small stacks of a l f a l f a haylage at Michigan State University Farm, while slope #2 (Yu & Thomas, 1976) was based on data from 21 forage samples, predominantly a l f a l f a haylages, from the same farm. From Figure 15 i t is apparent that the prediction equations are in f a i r l y close agreement at ADIN%N levels between about 17 to 25%, but that there is increasing - 86 -Figure 15: Regressions of total nitrogen d i g e s t i b i l i t y c o e f f i c i e n t s (Y) on ADIN%N (X) from various sources. 80 l ~i 1 1 r 1 1 5 10 15 20 25 30 ADIN%N 87 -disagreement with the present study at values outside these l i m i t s . This disagreement is most l i k e l y due to differences in the type of forage used in that slopes #1 and #2 relate to a l f a l f a haylages while the slopes •; from Experiments 1 and 2 are based on data from grass si 1 ages . 2.7 The Effects of Heating on Nitrogen U t i l i z a t i o n (Tab!e 20). , A near-perfect positive correlation existed between ADIN%N and fecal-N (r = .98). Results from many other studies have shown that heating increases the amount of nitrogen excreted in the feces (Bechtel et a l . , 1943, 1945; Hartmann, 1973; Sutton & Vetter, 1971; Yu & Thomas, 1975; Yu & Veira, 1977). S i m i l a r l y , the severe depression in nitrogen balance associated with the greatest heating (treatment H2) was in good agreement with other t r i a l s (Goering et a l . , 1974 ; M i l l e r et a l . , 1967 ; Thomas et a l . , 1972; Yu & Thomas, 1975; Yu & Veira, 1977). The decrease in nitrogen loss in the urine with increase in heating meant that the moderate heat treated silage (HI) had the same overall nitrogen retention as the unheated control (C). Similar depressions in urinary-N output with heating have been noted in work using dehydrated forages (Hartmann, 1973 ; Jasiorowski , 1960) and a r t i f i c i a l l y heated a l f a l f a haylage ( Yu & Veira, 1977). Yu and Veira (1977), working with a l f a l f a haylage heated to 88°C for 0, 24 or 48 hours, found that - 88 -urinary-N was 23.6, 17.3 and 16.6g/day respectively, while nitrogen retained by their sheep was 10.9, 7.2 and 6.3g/day respectively. Their negative correlation of -.74 between ADIN%N and nitrogen retention was considerably higher than the value of -.49 obtained in the present study. Heating may decrease urinary-N loss by decreasing ammonia-N loss in the rumen. Goering and Waldo (1978) in t h e i r review paper, suggested that some heating of forage protein may improve nitrogen u t i l i z a t i o n by making the protein less degradable in the rumen. In the present study, moderate heating (treatment HI) reduced nitrogen d i g e s t i b i l i t y , but a greater proportion of the nitrogen digested was retained, so that overall nitrogen retention remained the same when compared to the unheated control (treatment C). Greater heat damage associated with more severe heating (treatment H2) meant that less nitrogen was absorbed so that the decrease in urinary-N output was i n s u f f i c i e n t to compensate leading to a net reduction in overall nitrogen retention. In f a c t , two sheep on t r i a l H2 had an overall negative nitrogen balance of -.5 and -.7g/day. - 89 -SUMMARY The effects of heat damage on the feeding value of grass silages, in p a r t i c u l a r nitrogen d i g e s t i b i l i t y and u t i l i z a t i o n , were studied in two successive experiments. Heat damage was measured as the nitrogen content of the ADF f r a c t i o n expressed as a percentage of total nitrogen (ADIN%N). Forages with levels of ADIN%N below 9% were considered to be undamaged. EXPERIMENT 1 Six grass silages with varying degrees of heat damage (ADIN%N ranged between 4.8 and 22.9%) were studied in total c o l l e c t i o n sheep digestion t r i a l s . The major findings are summarized below:-1. Voluntary feed intake and p a l a t a b i l i t y appeared to be unrelated to ADIN%N le v e l s , although p a l a t a b i l i t y was not objectively measured. 2. ADIN%N was highly s i g n i f i c a n t l y (P<.01) negatively correlated with most of the apparent d i g e s t i b i l i t y c o e f f i c i e n t s , in p a r t i c u l a r with the c o e f f i c i e n t s for dry matter (r = -.96), organic matter (r = -.98) and nitrogen (r = -.95). 3. The amount of ADIN%N present in the silages explained 96%, 91% and 91% of the variation in the d i g e s t i b i l i t y of organic matter, dry matter and nitrogen respectively. 4. An increase in ADIN%N of 10 percentage units resulted - 90 -in a depression in nitrogen d i g e s t i b i l i t y of 20 percentage units and a depression in dry matter d i g e s t i b i l i t y of 8 percentage units. 5. ADIN had a negative apparent d i g e s t i b i l i t y for three of the six silages. It was thought that this was probably due to oven drying at 90°C heat damaging the feces for these three high quality sil a g e s . EXPERIMENT 2 The major problem associated with Experiment 1 was the variable background of the experimental sil a g e s . To avoid this problem one silage of low ADIN%N content was obtained and a r t i f i c i a l l y heated which resulted in an increase in ADIN%N levels from 8.7% to 14.1% and 24.3%. These materials were then fed to sheep in a digestion t r i a l as in Experiment 1, with the addition of urine collections so that nitrogen balance could be measured. The major points associated with this experiment are summarized below:-1. The method of heating employed generated a more than adequate amount of heat and was r e l a t i v e l y easy to control. 2. Future t r i a l s should proceed with the heating cables outside the paper fibr e form cylinder to avoid perforation of the polyethylene l i n i n g by direct contact with the cable. - 91 Perforation of the seal meant that the heated silages l o s t a considerable amount of moisture resulting in an unwanted divergence of moisture contents for the three treatments(34.2%, 40.8% and 48.5% for treatments C, HI and H2 respectively). Heating s i g n i f i c a n t l y increased the amount of ADIN%N from 8.7% for the unheated treatment C, to 14.1% for the moderately heated treatment HI (30 days at 49°C), and 24.3% for the severely heated treatment H2 (33 days at 50°C followed by 30 days at 63°C). ADIN%N increased with both maximum temperature and length of exposure to heat, but i t was not possible to iso l a t e these two e f f e c t s . The level of ADF increased while hemicellulose, a known substrate for the browning reaction, decreased with increase in heating. Increased heating led to increased darkening or browning of the silages. Due to their high f i b r e and low nitrogen contents, a l l three silages were consumed in disappointingly 1ow amounts. As a result of low intakes, each ewe l o s t an average of 8kg of bodyweight over the 9 week experimental pe ri od. The results for apparent nitrogen d i g e s t i b i l i t y were very similar to those obtained in Experiment 1, and - 92 a negative correlation of -.98 existed between nitrogen d i g e s t i b i l i t y and level of ADIN%N. 10. Other d i g e s t i b i l i t y c o e f f i c i e n t s were in marked disagreement with the results of Experiment 1. Moderate heating (HI) increased the d i g e s t i b i l i t y of ADF and NDF s u f f i c i e n t l y to cause the dry matter and organic matter of silage HI to be more digestible than that of the unheated control (C). More severe heating (H2) caused a depression in fi b r e d i g e s t i -b i l i t y compared to HI, although both NDF and ADF remained more digestible than in the unheated control (C). The rise in fibre d i g e s t i b i l i t y was d i f f i c u l t to explain as i t was in disagreement with other published results. It may have been due to some sort of "cooking" process associated with the heat treatment making some components of the fibre more diges t i b l e . Whatever the reasons, these results raise some questions about the v a l i d i t y of a r t i f i c i a l l y heating already fermented silage to produce heat damage. 11. With similar nitrogen intakes, sheep fed the heated treatments excreted 17 to 63% more nitrogen in their feces, but 8 to 31% less nitrogen in their urine as compared to when they received the unheated control treatment C. ADIN%N was s i g n i f i c a n t l y correlated with both these measures (P<.01). - 93 12. The amount of ADIN%N in the silages explained 66% and 96% of the variation in urinary-N and fecal-N respectively. 13. Reduced nitrogen d i g e s t i b i l i t y was compensated for by an increase in the e f f i c i e n c y of nitrogen retention in treatment HI compared to treatment C, so that there was no difference in overall nitrogen retention between these two treatments. More severe heating (treatment H2) led to a s i g n i f i c a n t (P<.05) reduction in nitrogen retention. 14. The regression equation for predicting nitrogen d i g e s t i b i l i t y from level of ADIN%N was in close agreement with that obtained in Experiment 1 and the following equation was obtained by pooling the results from both t r i a l s , : -% Nitrogen D i g e s t i b i l i t y = 81.76 ( r 2 = .92) - 1.83(ADIN%N) - 94 CONCLUSIONS In the f i n a l analysis, any brief conclusions about such a complex subject as heat damage will necessarily be unsatisfactory. The task is yet more d i f f i c u l t for this p a r t i c u l a r study owing to two major factors which affected the results of my second experiment, but which have not been noted in other studies. F i r s t , the level of feed intake in Experiment 2 was considerably lower than desired and was certainly below maintenance l e v e l s . Secondly, the rise in f i b r e d i g e s t i b i l i t y associated with heating was completely unexpected and casts serious doubts on the v a l i d i t y of experiments which seek to a r t i f i c i a l l y induce heat damage in already fermented material. Natural heat damage is i n t e r n a l l y generated and is caused by respiration of carbohydrates by micro-organisms in the presence of oxygen. Heat damage in the present study was caused a r t i f i c i a l l y by an external heat source working on a sealed anaerobic and e s s e n t i a l l y b i o l o g i c a l l y inert material. The two methods may well not be compatible. Nevertheless, results from Experiment 1 showed highly s i g n i f i c a n t (P<.01) negative correlations between ADIN%N and the apparent d i g e s t i b i l i t y c o e f f i c i e n t s of DM, OM, N, ADF and NDF. These results taken with those from Experiment 2 show that nitrogen d i g e s t i b i l i t y is the factor most severely depressed by increased heat - 95 damage, although the nitrogen balance results from Experiment 2 suggest that moderate levels of heating may lead to increased e f f i c i e n c y of u t i l i z a t i o n of digested nitrogen thus negating the depression in nitrogen di ges t i b i 1 i ty. These results confirm that ADIN%N should be a part of any routine analysis of grass sil a g e , p a r t i c u l a r l y when heat damage is suspected. - 96 -BIBLIOGRAPHY A.D.A.S. 1974. Ensiling Grass. Ministry of Agriculture, Fisheries and Food, Pinner, England, p. 1. Bechtel, H.E., F.W. Atkeson and J.S. Hughes. 1943. 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