UBC Theses and Dissertations

UBC Theses Logo

UBC Theses and Dissertations

The biological half-life of inorganic mercury in the Dungeness crab Cancer magister Dana Sloan, John Peter 1974

Your browser doesn't seem to have a PDF viewer, please download the PDF to view this item.

Item Metadata

Download

Media
831-UBC_1974_A6_7 S59_6.pdf [ 2.06MB ]
Metadata
JSON: 831-1.0093043.json
JSON-LD: 831-1.0093043-ld.json
RDF/XML (Pretty): 831-1.0093043-rdf.xml
RDF/JSON: 831-1.0093043-rdf.json
Turtle: 831-1.0093043-turtle.txt
N-Triples: 831-1.0093043-rdf-ntriples.txt
Original Record: 831-1.0093043-source.json
Full Text
831-1.0093043-fulltext.txt
Citation
831-1.0093043.ris

Full Text

T H E B I O L O G I C A L H A L F - L I F E -OF I N O R G A N I C M E R C U R Y IN T H E D U N G E N E S S CRAB,  C a n c e r magister  DANA  by  John Peter Sloan B.Sc,  U n i v e r s i t y of B r i t i s h C o l u m b i a ,  1971  A thesis submitted in p a r t i a l fulfilment of the r e q u i r e m e n t s for the degree of M a s t e r of Science in the Department of Zoology  We accept this thesis as c o n f o r m i n g to the r e q u i r e d standard  T H E UNIVERSITY O F BRITISH C O L U M B I A April  1974  In  presenting  this  an a d v a n c e d  degree  the  shall  I  Library  f u r t h e r agree  for  scholarly  by  his  of  this  written  thesis at  the U n i v e r s i t y  make  that  it  purposes  for  freely  permission may  representatives. thesis  in p a r t i a l  financial  is  Columbia,  British  by  for  gain  Columbia  shall  the  that  not  requirements I  agree  r e f e r e n c e and copying  t h e Head o f  understood  of  The U n i v e r s i t y o f B r i t i s h V a n c o u v e r 8, Canada  of  for extensive  permission.  Department  of  available  be g r a n t e d  It  fulfilment  of  this  or  that  study. thesis  my D e p a r t m e n t  copying  for  or  publication  be a l l o w e d w i t h o u t  my  ABSTRACT The b i o l o g i c a l h a l f - l i f e of inorganic m e r c u r y in the Dungeness crab,  C a n c e r magister Dana, was determined e x p e r i m e n t a l l y to be  about 25 d a y s .  C r a b s were exposed to m e r c u r y solutions,  after varying periods of time,  sacrificed  and m e r c u r y determinations of whole  body homogenates made with an atomic absorption  spectrophotometer.  The simple and widely used negative exponential equation for calculating b i o l o g i c a l h a l f - l i f e was not entirely adequate for d e s c r i b i n g the m e r c u r y elimination process.  A better d e s c r i p t i o n was obtained using a n o n -  l i n e a r l e a s t - s q u a r e s fit of an equation d e s c r i b i n g elimination at different speeds f r o m two types of t i s s u e s .  A further model allowed for r e c y c l i n g  of m e r c u r y that was eliminated, and gave m a r g i n a l l y better d e s c r i p t i o n s in some c a s e s .  iii T A B L E OF CONTENTS Page ii  ABSTRACT T A B L E OF CONTENTS  iii  LIST O F FIGURES  iv  LIST O F T A B L E S  v  ACKNOWLEDGMENTS  vii  INTRODUCTION  1  METHODS  3  A.  G e n e r a l and P i l o t T e s t s  3  B.  Holding and D o s i n g  8  C.  M e r c u r y R e m o v a l by P r e c i p i t a t i o n  11  D.  D i s s e c t i o n and Weighing  11  E.  Digestion  12  F.  Determination  14  RESULTS  17  Biological H a l f - T i m e The A d e q u a c y of Simple Negative C l e a r i n g Rates  .  17  Exponential 23  DISCUSSION  28  L I T E R A T U R E CITED  31  APPENDIX  33  iv  LIST OF FIGURES Figure 1  Page Relation between log body content of Hg (ng/g) and time in crabs exposed to Hg solutions.  A to E, experiments  1 to 5 respectively 2  20  Relation between log body content of Hg (ng/g) and log time in crabs exposed to Hg solutions. ments 1 to 5 respectively  A to E, experi21  V  LIST OF  TABLES  Table IA  Page Pilot test. crab tissue.  Mercury content (|ig/g) of Hg in wet weight of Dose 480 L i g / g for two hours, freeze dry  and wet weight procedures, gill and muscle analyzed separately. A and B are replicate tissue samples IB  4  Pilot test. Mercury content (Lig/g) of Hg in wet weight of crab tissue.  Dose 580 |ig/g for two hours first animals,  three hours for last two animals. A and B are replicate tissue samples, repeated observations are replicate spectrophotometer IC  Pilot test.  determinations  5  Mercury content (Lig/g) of Hg in wet weight of  crab tissue. Dose 300 L i g / g for two hours.  A and B,  replicate tissues, repeated observations are replicate determinations ID  6  Pilot test. Mercury content (Lig/g) of Hg in wet weight of crab tissue.  Dose 68 L i g / g for two hours.  A and B,  replicate tissues; repeated observations are replicate determinations II  7  Results of standardization tests on mercury content of crab tissues  III  16  Dosing procedures in experiments on retention of Hg in crabs  IV  18  Body content Hg (|Jg/g) in crab tissue homogenates at time (hours) after dose  V  Experimentally determined biological half-life of inorganic mercury in Cancer magister  19 - 22  vi Table  VI  Page  Probability that there is no significant departure from linearity in regression  VII  Estimated parameters for concurrent processes model (Equation 2)  VIII  24  26  Estimated parameters for concurrent processes model with recycling (Equation 3)  27  ACKNOWLEDGMENTS  The r e s e a r c h r e p o r t e d here was done at the F e d e r a l Department of the E n v i r o n m e n t P a c i f i c E n v i r o n m e n t Institute to p a r t i a l l y fulfil the r e q u i r e m e n t s for a M a s t e r of Science degree f r o m the Department of Zoology,  U n i v e r s i t y of B r i t i s h C o l u m b i a .  The staff at P . E . I . ,  especially  D r . John D a v i s and M e s s r s , J . With and R . H a r b o , gave indispensably p r a c t i c a l assistance and encouragement. s u p e r v i s o r at P . E . I . ,  D r . J . A . Thompson,  my  is r e s p o n s i b l e for the topic of my project,  it  d i r e c t s u p e r v i s i o n , the maintenance and d i r e c t i o n of the l a b o r a t o r y in which I worked, and for underwriting the entire project f i n a n c i a l l y f r o m his r e s e a r c h grant.  He b o r e the e x t r a burden of a novice graduate  f r o m another f i e l d with, at times, Dr. P. A . Larkin,  student  i n c r e d i b l e patience and understanding.  as my M a s t e r ' s p r o g r a m s u p e r v i s o r , looked after my  p e r s o n a l f i n a n c i a l support and a d m i n i s t r a t i v e a r r a n g e m e n t s .  He  a r r a n g e d for the teaching assistantship at U . B . C . that paid my rent.  His  unfailingly positive point of view always seems to alleviate the most awkward situations I was able to manufacture.  Both he and D r . T h o m p s o n  g e n e r a l l y p r o v i d e d me with everything I needed to spend time learning, including advice, and it is understood that their experience and teaching ability are e x p r e s s e d i n any parts of this r e p o r t that are worthwhile. Its shortcomings are a l l my own.  1 INTRODUCTION  M a n ' s awareness of the effects of i n d u s t r i a l wastes on e c o s y s t e m s has  r e c e n t l y r e s u l t e d in surveys  of freshwater and marine fishes for  m e r c u r y and other toxic pollutants.  In p a r t i c u l a r , it was d i s c o v e r e d in  Canada that f i s h caught d o w n s t r e a m f r o m c h l o r - a l k a l i plants which utilize mobile cathods contained elevated concentrations of m e r c u r y et a l . , 1970;  Fimreite,  1970;  (Wobeser,  F i m r e i t e , et a l . , 1971).  A c h l o r - a l k a l i plant located on the Canadian west coast at the head of Howe Sound,  B . C . , uses about 275,000 pounds of m e r c u r y annually for  e l e c t r o l y t i c decomposition of b r i n e .  The chlorine and s o d i u m hydroxide  produced are then supplied to nearby pulp m i l l s .  In e a r l y 1970, specimens  of v a r i o u s m a r i n e fishes and invertebrates were obtained f r o m Howe Sound near the plant site,  and Dungeness crab,  C a n c e r magister,  contain up to 13ppm m e r c u r y (Bligh, 1971).  were found to  F l o u n d e r (Citharichthys spp.)  taken f r o m the same a r e a contained over one p p m of the m e t a l . r e s u l t of these findings,  fishing i n this a r e a was banned.  As a  In October, 1972,  l e v e l s of up to 3.3ppm i n C. magister were s t i l l present. Because c r a b s a r e f a i r l y omnivorous b o t t o m - f e e d e r s ,  and because  they also support a valuable West Coast f i s h e r y (1971 wholesale value i n B.C.,  $946,000), they have been included as one of s e v e r a l species used at  the P a c i f i c E n v i r o n m e n t Institute  ( F e d e r a l Department of the E n v i r o n m e n t ) ,  West V a n c o u v e r , B . C . , to study m e r c u r y b i o - a c c u m u l a t i o n .  P a r t of this  p r o g r a m i s d i r e c t e d toward d e t e r m i n i n g m e r c u r y l e v e l s i n water, sediments,  and organisms i n Howe Sound;  bottom  l e v e l s which must be p r e s u m e d  to be the r e s u l t (wholly or i n large part) of waste output f r o m the c h l o r a l k a l i plant. In the past five y e a r s ,  a number of studies of b i o - a c c u m u l a t i o n of  m e r c u r y by freshwater and m a r i n e o r g a n i s m s have been reported,  with  the bulk of the l i t e r a t u r e dealing with freshwater o r g a n i s m s (Hannerz, 1968; R u c k e r and A m e n d ,  1969; Uthe,  1972).  Of p a r t i c u l a r interest to r e s e a r c h e r s  2 i s determination of rates of a c c u m u l a t i o n and b i o l o g i c a l h a l f - l i v e s of different c h e m i c a l f o r m s of m e r c u r y .  It i s now w e l l known that the p r e -  dominant m e r c u r i a l found i n freshwater f i s h i s methyl m e r c u r y ;  its  existence in lake water has been demonstrated by Chau and Saitoh (1973) and i n estuariune sediments by A n d r e n and H a r r i s s (1973). a s s u m e d that m e t h y l m e r c u r y , obtained v i a the food c h a i n .  Thus,  which p o s s e s s e s an affinity f o r proteins,  is  B a c t e r i a l c o n v e r s i o n has been shown to be  the pathway of o r i g i n of m e t h y l m e r c u r y (Jensen and J e r n e l o v , et a l . ,  it i s  1969; Wood  1968), though this does not obviate the need to look into other,  possibly non-biological, mechanisms. In saline waters Anfalt,  et a l . , (1968) have shown that the p r e v a i l i n g  m e r c u r i a l species is the complex anion ( H g C l ^ , -  and since the existence of  the m e t h y l m e r c u r i e cation or d i m e t h y l m e r c u r y i n seawater has yet to be c o n c l u s i v e l y demonstrated, mercury,  it must be a s s u m e d that the important f o r m of  exclusive of feeding, to which o r g a n i s m s a r e exposed,  is inorganic.  The e x p l o r a t i o n of this aqueous solution avenue of contact with the metal i s the purpose of this study of the b i o l o g i c a l h a l f - l i f e of inorganic m e r c u r y . E x p e r i m e n t s were designed to explore three p r o b l e m s :  (1) the  c l e a r i n g rate (biological half-time) of inorganic m e r c u r y f r o m C . magister following a b r i e f exposure to r e l a t i v e l y high concentrations  of the ion;  (2) the r e l a t i o n of c l e a r i n g rate to concentration and duration of dose; and (3) evaluation of other possible c i r c u m s t a n c e s  that might lead to  further understanding of the physiology of m e r c u r y c l e a r i n g i n these organisms. A  pilot experiment on.LD^Q estimates for m e r c u r y toxicity is  r e p o r t e d i n Appendix 1.  3  METHODS A.  G e n e r a l and P i l o t T e s t s C r a b s were bought c o m m e r c i a l l y at C r e s c e n t Beach,  the n a t u r a l l y - o c c u r r i n g l e v e l of m e r c u r y i s low (<200ppb).  B . C . , where Injection of  m e r c u r y v i a the soft joints of c r a b s was rejected m a i n l y because it was an unnatural way of introducing the m e r c u r y ,  and exposure to m e r c u r y  d i s s o l v e d in seawater was chosen as a direct,  simple method,  s i m i l a r to  the situation in a polluted n a t u r a l environment. M e a s u r e m e n t of m e r c u r y was done using an atomic absorption spectrophotometer, killing animals.  though it gives only total m e r c u r y ,  and necessitated  Testing blood samples was not possible because of  instrument l i m i t a t i o n s . It was not known how m u c h m e r c u r y c r a b s would a s s i m i l a t e , or what effects the m e t a l would have on t h e m .  A c c o r d i n g l y , pilot tests were r u n  on a few specimens using s m a l l indoor a q u a r i a . m e r c u r y were t r i e d ,  always for a two-hour p e r i o d . D u r i n g the tests  s e v e r a l other p o s s i b i l i t i e s were considered, of v a r i o u s body parts,  V a r i o u s amounts of  including separate analysis  wet weight a n a l y s i s v e r s u s d r y weight a n a l y s i s ,  and evaluation of the v a r i a b i l i t y among c r a b s as a guide to suitable sample size for the m a i n e x p e r i m e n t s .  It seemed advisable to take m o r e frequent  readings i m m e d i a t e l y after dosing, since this was when fastest l o s s was expected (two readings in the f i r s t day, another the second day, a fourth the fourth day,  and so on).  The pilot tests (Table I) suggested a range of dosing of f r o m 10-500ppm d i g r a m s per g r a m or parts per m i l l i o n m e r c u r y ) .  T h i s amount  seemed to give readable levels of m e r c u r y in the a n i m a l s , but not enough to cause any apparent i l l effects.  T A B L E IA.  Pilot test. Mercury content (\lg/g) of Hg in wet weight of crab tissue. Dose 480 (ig/g for two hours, freeze dry and wet weight procedures, gill and muscle analyzed separately. A and B are replicate tissue samples.  Wet Animal  Muscle  1.  Control  2.  Immediately after 3  Freeze Dry Gill  98  43  272  , 1, 879  dosing 12 hours after dosing  (A) (B)  1,198 1, 86l  17, 475 15,233  4.  24 hours after dosing  (A) (B)  2, 140 721  819 1Z, 245  5.  12 days after dosing  (A) (B)  679 601  15,466 12, 878  3.  Muscle  Gill  353 0  __ 2, 052 c  -  658 '  _. . 21,014  -  TABLE  IB. P i l o t test. M e r c u r y content (|Jg/g) of H g in wet weight of crab tissues. Dose 580 Lig/g for two hours f i r s t animals, three hours for last two a n i m a l s . A and B are r e p l i c a t e tissue samples, repeated observations are replicate spectrophotometer determinations.  Animal 1.  Muscle  Control A  Gill  Homogenate of Remainder of Soft T i s s u e s  250  400 784 735 1, 650 1,500  B C  116 hours A after dosing  B C  A 12 days after dosing  B C  -  626 1, 011 1, 024 1, 170  30,200 31, 200 33, 631  . -  40, 444 43, 333  556 -  115 461 2, 560 2, 006  -  38, 38, 49, 46, 37, 33,  945 721 873 202 248 075  -  2, 745 2, 725 1, 937 2, 015  -  2, 516 3, 104 5, 322 4, 298 3, 063 2, 698  6  T A B L E  IC.  Pilot test. Dose  M e r c u r y content  300 Ug/g  observations  ftig/g)  f o r two h o u r s .  are  replicate  of H g i n w e t w e i g h t of c r a b  A and B,  replicate  tissues,  determinations.  Homogenate  of  Remainder Animal 1.  ;  Control  Muscle  A  Gill  -  -  B  -  -  C  -  -  Immediately after  A  dosing  B  6 hours after  A  dosing  B  18 h o u r s  5.  48 hours after  28 26 -  1, 8 3 0 1, 7 8 6  19 20 23  B  -  3, 4, 3. 3,  A  B  -  17  159 130  dosing  11  1, 7 3 6  -  869 020 985 985  Body  Homogenate  22 19  2 4 12  12,439  Whole  -  11. 1 4 2 11, 7 8 1 12, 6 2 1  A  dosing.  of S o f t T i s s u e s  13 13 34 42  8  after  tissues.  repeated  26  18 22 20 16  1, 2 6 6 1, 3 9 6 1, 2 7 3 1. 5 9 9  -  679  -  -  -  673  -  -  -  568 529  6.  120 h o u r s  A  after dosing  B C  -  -  -  278  -  '  -  -  256  -  -  -  -  -  •  228 -  209 222 247  7  T A B L E  ID.  P i l o t test. Dose  M e r c u r y content  (|Jg/g) o f H g i n w e t w e i g h t o f c r a b  68 ( i g / g f o r two h o u r s .  observations  are replicate  A and B,  replicate  of  Remainder  1.  Muscle  Control  Gill  ° f Sott.  Tissue  Whole  Body  Homogenate  A  -  B  -  -  -  100 39  C  -  -  -  39 26  -  -  -  22  100  40  D  •  22 2.  Immediately  A  after dosing  75  B  3.  6 hours  A  after dosing  4.  18 h o u r s  B  17  A  796 8 56  B  48 h o u r s  A  after dosing  B C D  6.  120 h o u r s after  13 11  after dosing  5.  22 14  A  dosing  B C  479 450  tissues.  repeated  determinations.  Homogenate Animal  tissues;  4, 4, 4, 4,  575 907 270 479  5 3  3,120 3, 2 6 2 2, 5 9 6 2,912  28 26 37 26  5,427 5,888 7,143 7, 1 4 3  75 76 3 37 670 693 628 628 592 566 555 555 397 339 355 368 304 294  8 U s i n g homogenized wet c r a b tissue, it was difficult to c o n t r o l the exact amount of seawater i n the b l e n d e r .  C r a b tissue is e x t r e m e l y soft,  and when the organs are dissected out of the shell,  a c e r t a i n amount of the  m a t e r i a l f o r m s a soup with water contained in the s h e l l . The a n a l y s i s of separate tissues f r o m every a n i m a l was given up after two pilot tests when it became obvious that it would be too time consuming.  One sample a n a l y s i s usually took 8-12 hours, and only about  half of that was free time while the samples were sitting undergoing reactions.  Although some steps were as e a s i l y done on f o r t y samples as  on twenty, there were s e v e r a l " r a t e - l i m i t i n g " steps where each sample had to be handled separately,  e.g.,  weighing tissue samples and running the  spectrophotometer. F r e e z e - d r y i n g seems to concentrate m e r c u r y by about a factor of ten, indicating the high water content in c r a b t i s s u e .  In one instance i n  which separate tissues were analyzed (test 1), there was an inconclusive difference between g i l l and m u s c l e concentrations.  G i l l concentration i n  f r e s h l y - d o s e d animals, on the other hand, was r e l a t i v e l y large (a factor of f r o m ten to a hundred over m u s c l e s ,  and slightly l e s s ,  on the average,  over r e m a i n i n g tissue). C o m p a r i n g average m e r c u r y contents for tests 3 and 4, it might be suspected that a l a r g e dose collects heavily in g i l l tissue, but does not a c c e l e r a t e m e r c u r y accumulation i n muscle over a s m a l l dose.  B.  Holding and Dosing F i v e c l e a r i n g rate experiments were p e r f o r m e d during the winter  and spring months of 1972.  M a t u r e specimens of C . m a g i s t e r were  obtained c o m m e r c i a l l y f r o m Boundary Bay in the extreme south-west c o r n e r of B r i t i s h C o l u m b i a .  F o r each experiment,  sixty c r a b s were  maintained in captivity, exposed simultaneously i n a tank to inorganic  9 m e r c u r y d i s s o l v e d in a q u a r i u m water,  returned to unpolluted seawater i n  the o r i g i n a l a q u a r i u m , and then analyzed for total m e r c u r y content i n groups of five (one as a control) at v a r i o u s time intervals after  exposure.  S e v e r a l large (1.8m d i a . , 1.2m depth) c y l i n d r i c a l f i b r e g l a s s aquaria were used to a c c o m p l i s h t h i s . E a c h a q u a r i u m used for holding animals before and after dosing was fitted with a c e n t r a l double concentric  standpipe with outlets in the  outer pipe at the bottom (for m a x i m u m circulation), inner pipe (approximately 0.5m)  and length of the  set to maintain constant depth.  Water  taken f r o m B u r r a r d Inlet entered f r o m a pipe trained obliquely to the a q u a r i u m at a rate thatrprovided a complete water change every 10 to 15 minutes.  Water flow also caused a slow, clockwise c i r c u l a t i o n in each  aquarium; ments,  aeration was supplemented by an a i r s t o n e .  D u r i n g the e x p e r i -  salinity and t e m p e r a t u r e showed only s m a l l seasonal  variations.  When dosing was done, a c l o s e d s y s t e m was maintained in which the outlet at the centre of a separate a q u a r i u m and another near its p e r i meter were each fitted with a rubber hose through which dosing water was drained,  and r e c y c l e d over the b r i m of the a q u a r i u m by two e l e c t r i c p u m p s .  A l l c r a b s were left undisturbed throughout the p r o g r a m and were fed to satiation with h e r r i n g . A standard procedure was followed throughout the p r o g r a m . b a s i c routine was s i m p l e : a week,  The  (1) maintain about sixty c r a b s in captivity for  which was presumably sufficient to a c c l i m a t i z e t h e m to the a q u a r i u m  and to any conditions of water that may have d i f f e r e d f r o m Boundary Bay; (2) m a r k ten of these as controls,  not to be exposed to m e r c u r y ;  (3) place  the r e m a i n i n g fifty for two hours in a separate a q u a r i u m containing a m e a s u r e d amount of d i s s o l v e d m e r c u r y ; r e t u r n them to the holding a q u a r i u m ; i n t e r v a l s following exposure,  (4) remove treated c r a b s and  (5) capture,  at specified time  four dosed and one c o n t r o l a n i m a l ;  and d i s s e c t a l l tissue f r o m these five;  (7)  (6) k i l l ,  digest tissue samples for  10 anal/sis; tion  (8)  measure total m e r c u r y concentration on an atomic a b s o r p -  spectrophotometer. Dosing was always done e a r l y on a Saturday morning, and the  weekend used to do the f i r s t two or three sets of m e r c u r y d e t e r m i n a t i o n s . E a c h dosing a q u a r i u m had two outlets at the bottom, hooked up by a r u b b e r hose to two e l e c t r i c water pumps.  The output lines f r o m the pumps  fed over the r i m of the a q u a r i u m so that water would c i r c u l a t e around the tank.  When two aquaria were used, the hoses c r o s s e d ,  their four pumps as a single s y s t e m .  functioning v i a  C l a m p s on one of the hoses  regulated any flow d i s c r e p a n c y . The dosing tank contained 1930 l i t r e s of seawater.  A calculated  amount of m e r c u r y salt (HgNO-j) was d i s s o l v e d in a few l i t r e s three) of w a r m e d seawater, aquarium,  (usually  and then added to the water c i r c u l a t i n g in the  letting it m i x for about one-half hour.  Just p r i o r to dosing t i m e  50 e x p e r i m e n t a l c r a b s were c a r e f u l l y piled in a large plastic tub beside the dosing a q u a r i u m . A t the start of dosing time,  c r a b s were put quickly into the dosing  tank, c a r e f u l l y enough to avoid disturbance and fighting, and this took about three minutes.  Next, a s m a l l sample of a q u a r i u m water was taken, to which  6N, H_.SC) was added. 2 4  (The a c i d stops loss of m e r c u r y to the air and sides  of the plastic container.) to monitor the dose.  A d d i t i o n a l samples were taken every h a l f - h o u r  T o w a r d the end of dosing (within three minutes),  the  e x p e r i m e n t a l c r a b s were caught and r e t u r n e d to the c i r c u l a t i n g holding aquarium,  which took about six to eight minutes.  Next, four 0 - t i m e e x p e r i m e n t a l crabs and one c o n t r o l were and caught,  and a f i n a l water sample was taken;  selected  then the pumps were shut  off and a m e r c u r y - p r e c i p i t a t i n g mixture was added to the a q u a r i u m .  11 . C.  M e r c u r y r e m o v a l by p r e c i p i t a t i o n About 30g each, of F e f N O - ^ - j and Na2S were used to c o - p r e c i p i t a t e  m e r c u r i c ion i n the exposure water before it was d i s c h a r g e d to B u r r a r d Inlet.  A f t e r two days settling,  c l e a n seawater, hole.  it was possible to siphon off the 2000 Z of  and then wash the precipitate into buckets through the plug  These buckets could then be left to settle some m o r e and the  precipitate c o u l d finally be f i l t e r e d and d r i e d . tested on the spectrophotometer,  Siphoned seawater was  which indicated total m e r c u r y below  0.5|ig/g. D o s i n g water always showed m e r c u r y l o s s .  F r o m the l e v e l c a l c u -  lated f r o m the weighed amount added, the start of each dosing p e r i o d , total m e r c u r y dropped f a i r l y steadily.  In establishing a dose for each  experiment the mean of five readings was taken. E x p e r i m e n t 1: 10-15|ig/g; 410-460Llg/g;  D.  Ranges of readings w e r e :  E x p e r i m e n t 2: 443-557|jg/g;  E x p e r i m e n t 4: 482-541|ig/g;  E x p e r i m e n t 3:  E x p e r i m e n t 5: 7-12|ig/g.  D i s s e c t i o n and Weighing  C r a b s were k i l l e d by hitting t h e m with a h a m m e r handle at the apex of the triangular v e n t r a l p a n e l . c r a b and counting its l i m b s ,  A f t e r m e a s u r i n g the length of each  soft tissue was r e m o v e d -(except the m u s c l e  f r o m the outer two leg segments) and placed in a W a r i n g b l e n d e r . A s p e c i a l 100ml plastic bottle attachment was used for this since it was an i d e a l size (later, a S o r v a l l stainless steel blender was used that worked well). Between each use, the bottle was r i n s e d with  HNO3 and d i s t i l l e d water.  homogenate was poured into weighing boats and c o v e r e d and m a r k e d . Weighing developed to a routine using plastic disposable type s y r i n g e s . test tube,  straw-  About a g r a m of homogenate was added to a weighed  and weighed by d i f f e r e n c e .  The  12 E.  Digestion D i g e s t i o n of tissue for total m e r c u r y determination was done  following the method of A r m s t r o n g and Uthe (1971).  The development of  the method r e p o r t e d here to the point where it was giving results,  satisfactory  was perhaps the most f o r m i d a b l e p r o b l e m encountered in the  project.  B a s i c a l l y , the digestion involves two steps,  one using s u l f u r i c  and n i t r i c acid, and a second oxidation step using permanganate or peroxide. F o r tissue-bound mercury,  c e r t a i n manipulations are needed to get  the m e r c u r y into its free atomic state, H g ° . found bound to p r o t e i n s .  M e r c u r y in tissue is usually +2  In this f o r m it is a divalent cation, Hg  , and is  usually attached by a covalent bond to protein sulfur, or to one or two methyl groups as methyl or d i m e t h y l m e r c u r y . A strong oxidizing agent (8ml of concentrated H2SO4 and 2 m l of concentrated HN.O3 per sample) was used as a f i r s t step in breaking these bonds.  Concentrated acid, over time,  mercury,  with heat, y i e l d s free divalent  some methyl and proteinated m e r c u r y ,  proteoses and amino a c i d s .  and a solution of  A f t e r thorough a c i d digestion (one hour) this  tissue had become a clear solution. The proteinated and methyl m e r c u r y can be b r o k e n down further by using a second oxidizing agent (6 per cent M n O ^  , and later 50 p e r cent  H2O2) which, with incubation for s e v e r a l hours, w i l l break up n e a r l y a l l +2 of these compounds so that n e a r l y a l l the m e r c u r y exists as free Hg D u r i n g permanganate (MnC>4 ) oxidation, manganese  undergoes  reduction to M n G ^ (MnlV) w h i c h appears as a brown p r e c i p i t a t e . reduction of the m e r c u r y ,  Before  it is f i r s t n e c e s s a r y to r e m o v e the precipitate  by further reduction to soluble Mn(II).  Reduction is best obtained by  addition of a solution of an hydroxylamine salt or hydrogen p e r o x i d e .  13 Once this step i s finished, cleared,  as shown by the last bit of sludge being  adding a couple of drops of the o r i g i n a l M n 0 4  w i l l r e t u r n the +2  solution to being safely " o x i d i z i n g " , and p r e s e r v e the m e r c u r y as Hg but none of the sludge of the intermediate M n O ^ is p r o d u c e d .  ,  When H ^ O ^  is used, no sludge is p r o d u c e d . F o r m e a s u r i n g m e r c u r y quantity i n this solution, either a flame is +2 used to atomize the H g is added.  , or,  in the f l a m e l e s s technique a strong  T o a c c o m p l i s h this,  reductant  an addition f u n n e l f u l l of reductant is fitted  to the v e s s e l containing the sample solution, and c l o s e d underneath with a stopcock.  In this way, an airtight s y s t e m can be set up so that air or  n i t r o g e n can be introduced. the reductant funnel above it,  A r e a c t i o n v e s s e l containing the solution with s t i l l closed, is included in the airtight  so that the a i r or nitrogen w i l l bubble into the solution. s t i r r e r is p l a c e d under the r e a c t i o n v e s s e l ,  A magnetic  which opens at the top both to  the c l o s e d reductant funnel and to a second a i r l i n e . leads to a g l a s s c e l l which has been lined up i n the beam,  system  T h i s second a i r l i n e spectrophotometer  and which is open v i a a m e r c u r y trap containing n i t r i c acid, to the  atmosphere. To r u n a determination, reaction vessel,  some reductant is f i r s t r e l e a s e d into the  and the s t i r r e r is started.  A t this point, the reduction  to H g ° takes place, but the s y s t e m is airtight and static, Hg° escapes.  A f t e r the r e a c t i o n is complete,  so that no volatile  air or nitrogen is bubbled  through the sample v e s s e l at a constant rate, and a f r a c t i o n of the total H g ° i s swept out of solution and up into the c e l l where it i s exposed to the spectrophotometer  beam.  The gas flow d r i v e s the H g ° on into the trap,  and finally c l e a r s the s y s t e m after the H g ° has been r e l e a s e d and r e c o r d e d .  14 F.  Determination T o t a l m e r c u r y was d e t e r m i n e d by f l a m e l e s s atomic a b s o r p t i o n  spectrophotometry, Ott (1968).  using a v a r i a t i o n of a method d e s c r i b e d by H a t c h and  The g l a s s c e l l (quartz windows) used was a c y l i n d e r , 15cm  long and 2 2 m m in d i a m e t e r .  Standards were made up daily f r o m a 100ppm  solution of m e r c u r i c c h l o r i d e i n 6N, H C l .  Some p r o b l e m s were encountered  i n achieving satisfactory r e c o v e r y of m e r c u r y , but the r e s u l t s r e p o r t e d here were taken using a method that gave 70-90 per cent r e c o v e r y . The weighed amount of tissue, volume,  i n solution, was made up to a known  so that the concentration of m e r c u r y r e a d by the spectrophotometer  (against standards) would be meaningful.  The spectrophotometer d e l i v e r y  apparatus was fitted with s p e c i a l 50ml p e a r - s h a p e d sample v e s s e l s ,  and i n  the e a r l y tests the sample was made up to 100ml i n v o l u m e t r i c f l a s k s and then 50ml was pipetted into sample v e s s e l s .  Later,  50ml digestion tubes  w e r e used to make up to volume, and p o u r e d d i r e c t l y into the sample vessels.  Although the quantity i n the sample v e s s e l was a c r i t i c a l issue i n  the same way as the volume of the a i r tubes,  it did not s e e m to affect  r e s u l t s when the v e s s e l s got 49+ m l , poured out of the tubes, 50+_.02 m l f r o m the pipettes.  instead of  The concentration was a c c u r a t e i n either  case. Reduction of m e r c u r y i n the r e a c t i o n v e s s e l was done with 2 m l of SnCl^i  with the s t i r r e r r u n for sixty second.  The c a r r i e r gas was  r e s e a r c h grade nitrogen, d e l i v e r e d using a needle valve at 3 0 0 m l / m i n .  A  plastic d r y i n g tube was fitted onto the gas line just before the g l a s s c e l l ; this tube contained m a g n e s i u m p e r c h l o r a t e and was changed once i n 20 to 30 d e t e r m i n a t i o n s . Standardization of the spectrophotometer was done f r o m a stock of lOOOppm HgQ.2 i n 6N, H C l .  Once a week a substock of lOOppm was made  f r o m the lOOppm solution, i n 0.2 N H C l .  It has to be a s s u m e d ,  and i s  15  a l m o s t certain, that these solutions, when f r e s h , were good to _+ the tolerance m a r k e d on the v o l u m e t r i c g l a s s w a r e . The r e s u l t s r e p o r t e d here are a l l quantities m e a s u r e d against e x t e r n a l standards.  T h i s means that they must a l l be low, c o m p a r e d to  the r e a l m e r c u r y contents of the c r a b s ,  since some m e r c u r y w i l l have  been lost i n p r e p a r a t i o n of the s a m p l e s . The i n f o r m a t i o n here r e p o r t e d was used for m e a s u r i n g b i o l o g i c a l half-time, content.  and the f o r m of the line relationship between time and body Neither of these is affected by inaccurate r e s u l t s ,  the r e s u l t s are consistently inaccurate, of different content. recoveries,  by the same p r o p o r t i o n i n samples  F r o m running internal standards and per cent  late in the project,  another laboratory,  as long as  and also having samples analyzed at  the r e s u l t s probably r e p r e s e n t e d 40 to 60 per cent  of the r e a l m e r c u r y contents. Table II shows per cent r e c o v e r y r e s u l t s ,  and also a c o m p a r i s o n  of some values using external standards, with some internal standards on the same s a m p l e s .  F r o m these n u m b e r s it is t h e o r e t i c a l l y possible  to r e w o r k a l l the previous r e s u l t s so that they would be e x p r e s s e d in t e r m s (1) of i n t e r n a l standards,  and (2) c o r r e c t e d for per cent r e c o v e r y .  The mean per cent r e c o v e r y of about 77 per cent,  shown in part B  of Table II, is meaningful only in t e r m s of percentage r e c o v e r i e s r e p o r t e d in the m a j o r i t y of a n a l y t i c a l c h e m i s t r y l i t e r a t u r e that deals with tissue analysis.  Percentage r e c o v e r y is there (e.g., A r m s t r o n g and Uthe,  1970)  treated as that per cent of a spiked blank which is shown in a spiked tissue s a m p l e . of tissue,  In other words, a standard solution is added to a sample  as i n the t h i r d p r o c e d u r e d e s c r i b e d p r e v i o u s l y , and the r e c o v e r y  c o m p a r e d to an i n t e r n a l standard,  or reagent blank with standard solution.  T A B L E II.  A.  Results of standardization tests on m e r c u r y content of crab tissues.  Comparison of internal and external standards (from test 5).  B.  P e r c e n t recovery, relative to external standards and internal standards.  »  Internal Standard (as fraction of External Standard)  Spiked T i s s u e (as fraction of E x t e r n a l Standard)  Experiment/ Animal  Internal Standard E x t e r n a l Standard  Experiment/ Animal  Percent Recovery (External)  5/1  .335  2/7  34.0  58.7  1  .910  .522  5/2  .710  3/1  45.0  77.7  2  .868  .785  5/3  .640  3/7  56.0  96.7  3  .923  .614  5/4  .532  3/8  48.4  83.5  4  .843  .810  5/5  .680  3/9  31.3  54.0  5  .843  .586  3/9  57.4  99.1  6  .873  .560  4/2  33.9  58.5  7  .991  .881  4/3  30.7  53.0  8  .872  .564  4/6  58.3  100.0  4/7  50.2  86.7  Mean  .579  Standard Deviation  .152  Percent Recovery (Internal)  Comparison of external standards to internal standard, and to spiked tissue.  Animal  Mean Standard Deviation  Mean  76.7  Standard Deviation  19.3  .881 . 564  17  T h e r e could be other f a c t o r s than poor technique involved i n low percentage r e c o v e r i e s .  P e r h a p s the m a t r i x effect of tissue on r e l e a s e of  m e r c u r y causes t i s s u e sample standards always to be low r e l a t i v e to inorganic blank standards.  RESULTS  F i v e experiments were conducted, the c i r c u m s t a n c e s being d e s c r i b e d in Table III.  The data i n Table IV show m e a s u r e d body content  of inorganic m e r c u r y at v a r i o u s t i m e s following dosing. When graphed, these data a r r a y themselves as scattered points w h i c h indicate a d r o p i n body content with time ( F i g u r e s 1 and 2). A n a l y s i s of the data a d d r e s s e d itself to two questions: roughly,  (1) what,  i s the time r e q u i r e d for c l e a r i n g of half of the m e r c u r y taken  up at dosing (biological h a l f - t i m e ) ;  and (2) i s the assumption of a simple  negative exponential c l e a r i n g rate a valid one and, if not, what other m a t h e m a t i c a l m o d e l would more adequately d e s c r i b e the way i n which inorganic mercury is cleared f r o m C . magister?  A t h i r d question of  whether c l e a r i n g rate v a r i e s with concentration or duration of dose may be answered,  somewhat i n c o n c l u s i v e l y , by r e f e r r i n g to Table III.  Biological H a l f - T i m e The data indicate that the f i r s t question may be answered as shown in Table V .  H a l f - t i m e s were calculated using the f o r m u l a  where T ^ is b i o l o g i c a l h a l f - t i m e ; 2  18 T A B L E III.  Experiments 1  D o s i n g p r o c e d u r e s i n experiments on retention of Hg i n c r ab s.  Dose Hg (lig/g) 12  ,  Remarks Hg ( N 0 ) 3  2  Cages used  478  Hg ( N O ) s  2  Cages used  433  H g O (in acid solution) No cages  500  Hg ( N O ) s  2  No cages  5  7-2  Hg  (N0 ) 3  No cages  2  T A B L E  IV.  B o d y content after  Hg  Time  Body  (hours) Experiment 1  1 12 36 144 816  2  1 12 24 48 552 912  1536  512, 27 5, 275, 195, U5.  479, 362, 225, 239, 229.  c ontrol:  532, 788, 547, 303, 147, 120, 186.  3  1 12 24 48 72  665. 1020, 834,  96 168  649, 753, 314. 422,  384  387,  576 744  329, 388, mean  Experiment  4  5  617, 565, 502, 473, 550, 381, 482. .  840  314,  1 17.2 41.8 94.6 144 672  at t i m e  (hours)  Content  152, 172,  .119.  Deviation  445, 226. 268, 171. 162,  305 329, 271 199 161  435 298 260 201 172  91 59 23 28 40  6 4 ( + 22) 511 666  621 607  115 155  364,  249, 121. 180, 127.  249, 169, 263, 200,  466 221 206 318 147  449 256 161 254 161  77 40 30 69 84  741 770  983 759  593 205  555 626 704 434 657  711 660 725 418 510  189 27 46 87 114  348  404  44  259 282  287 282  68 97  758 611 535 566 484 377 447  228 141 103 65 107 87 59  268  291  70  296  - 2 9 7 321 223 175 175 145  1871. 724. 804, 677. 670, 397. 416, 439. 204, 306.  980, 550, 686, 598, 343, 499. 475, 207.  control:  269, 318, 221,  Standard Mean  Ug/g)  724. 548,  control:  1 18 42 72 165 264 408  mean Experiment  homogenates  718. 426, 420.  mean control: Experiment  in crab tissue  (4 a n i m a l s ;  mean Experiment  (ug/g)  dose.  375. 370, 224, 204, 156, 131.  mean control:  7 8 ( + 13)  655, 521, 652, 687. 772, 526. 546, 440, 3 54, 152, 9 9 (+ 14  512, 507, 501, 624, 584, 307, 3 58, 374,  981 820 451 567 460 321 473  ~  7 4 (+ 3 6  247, 308, 176, 197, 174, 156. 53 (+ 21)  286 271 196 205 174  •  55 35 38 29 22 24  20  F i g u r e 1.  R e l a t i o n between log body content of Hg (ng/g) and time i n c r a b s exposed to Hg solutions. A to E , experiments 1 to 5 r e s p e c t i v e l y .  21  TIME. (HOURS)  5  10 TIME  Figure 2 .  30  100  1000  (HOURS)  R e l a t i o n between log body content of Hg (ng/g) and log time in c r a b s exposed to H g solutions. A to E , experiments 1 to 5 respectively.  22  TABLE  V.  E x p e r i m e n t a l l y d e t e r m i n e d b i o l o g i c a l h a l f - l i f e of inorganic m e r c u r y in Cancer magister.  Arithmetic plot (days)  S e m i - l o g plot (days)  1  27.1  34.5  2  32.9  25.8  3  18.3  15.4  4  24.4  26.7  5  21.7  21.7  Experiment  Average Standard Deviation  24.9  24.8  5.5  7.0  23  R is the rate of r e l e a s e ,  as d e t e r m i n e d by a l e a s t - s q u a r e s line of  best fit to the data; C  Q  is the i n i t i a l body content after dosing, as d e t e r m i n e d by the  Y - i n t e r c e p t of the line of best fit; B is the average background body content,  i.e.,  the mean body  content of a l l c o n t r o l a n i m a l s .  The A d e q u a c y of Simple Negative Exponential C l e a r i n g Rates  The second question (whether a simple negative exponential relationship is qualitatively adequate for d e s c r i b i n g the r e l e a s e of m e r c u r y f r o m C_. magister),  was approached by evaluation of linear and n o n - l i n e a r  components of l e a s t - s q u a r e s lines of best fit.  This procedure was used to  evaluate as w e l l as the negative exponential line f o r m , both an a r i t h m e t i c time v e r s u s arithmetic body content line f o r m , and a log time v e r s u s log content f o r m .  The graphs of two of these t r a n s f o r m a t i o n s are shown in  F i g u r e s 1 and 2.  Table VI shows the r e s u l t s of this analysis e x p r e s s e d as  p r o b a b i l i t i e s that the line of best fit is an adequate d e s c r i p t i o n of the r e l a t i o n s h i p between time and body content. The inadequacy of any of the three simple line f o r m s in d e s c r i b i n g the r e l a t i o n s h i p led to the idea that the c l e a r i n g rate might be the r e s u l t of multiple events, Specifically,  but that these events might not i n themselves be c o m p l e x .  it seemed possible that two or more sources of m e r c u r y  existed in the a n i m a l and that the c l e a r i n g rate f r o m each source might be different.  Further,  the r e l a t i v e l y fast initial c l e a r i n g r a t e s and r e l a t i v e l y  slow subsequent ones suggested two concurrent p r o c e s s e s , simple negative exponential c l e a r i n g r a t e . conceived to test this idea.  each with a  Equation (2) shows a m o d e l  24 . TABLE  V I . P r o b a b i l i t y that there i s no significant departure f r o m linearity in regression. (Legend:  axes a r e indicated as follows: A: B: C:  Axes  a r i t h m e t i c body content v e r s u s arithmetic time l o g a r i t h m i c body content v s . arithmetic time l o g a r i t h m i c body content v s . l o g a r i t h m i c time) Experiment  Probability  A  1 2 3 4 5  .00043 .00001 .15201 .09665 .00013  B  1 2 3 4 5  .00128 .00005 .10295 .18025 .00029  C  1 2 3 4 5  .93413 .01884 .02484 .15076 .00710  25  H  ST  =  H  g  (  0  P  l  e  "  K  l  T  +  (  1  -  P  l  )  e  "  K  2  T  )  ( 2 )  where H g i s i n i t i a l body content Hg; Q  P j i s the p r o p o r t i o n of the total Hg in the f i r s t concurrent  process;  is the rate of r e l e a s e f r o m the f i r s t c o n c u r r e n t p r o c e s s ; K 2 i s the rate of r e l e a s e f r o m the second concurrent  process.  A p r o g r a m ( M A R Q D = M a r q u a r d t , 1963)  parameters  to estimate  for n o n - l i n e a r systems such as this one was applied, for equation (2), the data f r o m a l l five e x p e r i m e n t s .  Table VII shows the best  to  parameters  d e r i v e d by t h i s p r o g r a m , and the per cent v a r i a n c e explained by each set of p a r a m e t e r s .  E x c e p t for the data of experiment 4, the l e a s t - s q u a r e s fit for equation (2) explained significantly more of the total v a r i a b i l i t y than s i m p l e linear r e g r e s s i o n with either a r i t h - l o g or l o g - l o g axes.  There  thus seems to be r e a s o n to believe that the p r o c e s s of c l e a r i n g is more complex than i s suggested by s i m p l e linear m o d e l s .  A modification to equation (2),  which would allow a p r o p o r t i o n Q  of the m e r c u r y r e l e a s e d by the f i r s t concurrent p r o c e s s to be r e c y c l e d into the second concurrent p r o c e s s and then r e l e a s e d at a rate, K £ , (equation (3)) was also tested. Hg  T  = Hg  Q  P  i  e  "  K  l  T  + (l-P  l ) e  " 2 K  T  +QP )l- " l )e" 2 _ 1  e  K  T  K  T  (3)  The p a r a m e t e r s d e r i v e d for this model by two c a l l s to the M A R Q D program,  are shown i n Table VIII.  It is c l e a r ,  f r o m the large r e s i d u a l v a r i a b i l i t y for the s i m p l e  negative exponential r e l a t i o n s h i p of body content to time, that values for b i o l o g i c a l h a l f - t i m e (calculated using that line form) are useful only as crude a p p r o x i m a t i o n s .  TABLE  VII. Estimated parameters for concurrent processes mod (Equation 2) Percent variance Experiment P K K explained J.  X  w  1  .51481  .00040  .09044  77.69  2  .74733  .02696  .00003  81.35  3  .65209  .00128  .06527  53.34  4 5  convergence not achieved .43778  .00006  .01408  73.12  27  TABLE  VIII.  Experiment 1  *  E s t i m a t e d p a r a m e t e r s for concurrent p r o c e s s e s m o d e l with r e c y c l i n g (Equation 3) Percent Variance Call Explained 1 2  .51480 37.830*  .00040 .00023  .09017 .000 58  .58392 1.8443*  77.69 59.86  1 2  .83745 .83768  .00084 .00084  .00559 .00559  20.487* 20.478*  83.92 83.92  1 2  .65207 .65209  .00128 .00128  .06460 .06467  .27364 .26844  53.34 53.35  1 2  .01169 .01234  .06732 .06730  .00077 .00077  23.407* 22.124*  59.09 59.09  1 2  .45727 .00000 .01055 959.67* CONVERGENCE NOT ACHIEVED  73.17  indicates nonsense  estimate  28 The c o n c u r r e n t p r o c e s s e s models (equations (2) and (3)) r e m a i n unsubstantiated by the statistical tests applied to them,  also although  a f a i r l y large percentage of the v a r i a t i o n is explained by those models in many c a s e s .  The r e c y c l i n g model showed nonsense values for best  e s t i m a t e s of some of its p a r a m e t e r s ,  and without independent evidence as  a basis for constraining some of the v a r i a b l e s ,  it was not possible to  evaluate the v a l i d i t y of the m o d e l assumption.  DISCUSSION  The r e s u l t s indicate that C a n c e r magister a s s i m i l a t e s e n v i r o n mental inorganic m e r c u r y so that its body content approaches or that of the surrounding water.  surpasses  Whole body levels of between 100 and 600  (ig/g (up to 1000 if method i n a c c u r a c i e s are allowed for) are found after a two-hour exposure to m e r c u r y ,  and these body l e v e l s appear to exist even  when the dose is appreciably reduced (see E x p e r i m e n t s 1 and 5).  Little  e x p e r i m e n t a l data exist dealing with c o m p a r a b l e exposure t i m e s to inorganic m e r c u r y in other species, in f i s h (Wobeser,  et a l . , 1970;  but l e v e l s found in the environment  Fimreite,  environmental l e v e l s in c r a b s (Bligh, 1971) between 50 and 150 l i g / g ) .  1970)  are comparable both to  and to my c o n t r o l l e v e l s (all  It was found i n these experiments,  expected f r o m the negative exponential c l e a r i n g rate concept,  as might be that at the  end of most experiments r e s i d u a l m e r c u r y r e m a i n e d w e l l above c o n t r o l levels. C l e a r i n g rates are also s i m i l a r to those found in other (Rucker and A m e n d ,  1969).  species  It i s important, though, to d i s t i n g u i s h  between work on inorganic m e r c u r y where c l e a r i n g r a t e s produce a h a l f time of the order of a few weeks to a few months, mercury,  and work on methylated  in w h i c h as m u c h as two to seven years may be r e q u i r e d for  halt the compound to be c l e a r e d . M e r c u r y l e v e l s , both organic and inorganic, as found even near to s o u r c e s of pollution in m a t u r a l waters,  are far below any used in this  29 study.  The r e s i d u a l l e v e l i n n a t u r a l l y - o c c u r r i n g a n i m a l s of about 50 to  150 l i g / g r e s u l t s f r o m water l e v e l s of total m e r c u r y about one-tenth of those found i n the a n i m a l s .  T h i s indicates,  of c o u r s e ,  that l o n g - t e r m  exposure tends to i n c r e a s e m e r c u r y concentration i n t i s s u e s . The r e l e v a n c e of this study is perhaps then l e s s i n its quantitative application to present n a t u r a l l y - o c c u r r i n g levels of m e r c u r y , than i n its conclusions that l o n g - t e r m effects r e s u l t even f r o m b r i e f m e r c u r y exposure and in the i m p l i c a t i o n that,  far l o n g e r - t e r m effects might be  expected f r o m the m o r e toxic organic m e r c u r i a l s and f r o m c h r o n i c expo s u r e . The m a t h e m a t i c a l models of c l e a r i n g rates were conceived to explain the r e s u l t s on the following b a s i s :  it was thought that two c o n c u r r e n t  s y s t e m s could exist in one or m o r e of s e v e r a l p h y s i c a l situations. of m e r c u r y f r o m two different tissues (e.g., at different but i n t r i n s i c a l l y consistent rates, in protein content i n the t i s s u e s .  Release  l i v e r and muscle) might occur perhaps due to differences  F u r t h e r models could be a l m o s t  infinitely extended to include other t i s s u e s .  F o r example, two c h e m i c a l  f o r m s of m e r c u r y might e x p r e s s their different affinities for tissue i n different rates of r e l e a s e .  In either case, m e r c u r y could leave the pool of one faster c o n c u r r e n t p r o c e s s to enter that of the slower one. slower p r o c e s s ,  It is conceivable that the  whether a tissue or a f o r m of m e r c u r y ,  might hold a  greater amount of m e r c u r y at some time after dosing than it does i m m e diately following exposure. Finally,  another hypothesis (not tested by a model here), that would_  explain a fast i n i t i a l c l e a r i n g rate and a slow later one, is degenerative pathological change.  Destruction, or blocking of enzyme pathways, in  e x c r e t o r y organs could lead to p r o g r e s s i v e l y reduced ability to e l i m i n a t e free m e r c u r y ,  which would then r e c y c l e into the o r g a n i s m . - •  30 The doubts r a i s e d by the r e s u l t s obtained here as to the validity of the w i d e l y - u s e d negative exponential " b i o l o g i c a l h a l f - t i m e " i n C . magister has led to the speculation that s i m i l a r doubts could be r a i s e d if other species and other f o r e i g n m a t e r i a l s were examined c l o s e l y .  It seems  reasonable to assume that the interface of l i v i n g tissue and n o n - l i v i n g , external m a t e r i a l , may often be c o m p l i c a t e d by v a r i a t i o n i n f o r m , whether of the tissue or the m a t e r i a l , or by pathological changes caused by the material.  It may also be that i n studying the m e c h a n i s m s of uptake,  binding, and r e l e a s e ,  where these are of interest,  hypotheses could be  conveniently tested with models and line f o r m analysis s i m i l a r to those used h e r e .  31 LITERATURE  A n d r e n , A . W . , and R . C . H a r r i s s . sediments. Anfalt,  Nature,  CITED  1973.  M e t h y l m e r c u r y i n estuarine  245:256-257.  T . , D . D r y s s e n , E . Ivanova and D . J a g n e r . divalent m e r c u r y i n natural w a t e r s .  1968.  The state of  Svensk k e m i s k t i d s k r i f t 80:  340-342. Armstrong,  F . A . J , and J . F . Uthe.  of m e r c u r y i n f i s h t i s s u e .  1972.  Semi-automated  determination  A t o m i c A b s o r p t i o n Newsletter  10:101-  104. Bligh,  E . G . 1971.  M e r c u r y l e v e l s i n Canadian f i s h .  R o y a l Society of  Canada International S y m p o s i u m on M e r c u r y in M a n ' s E n v i r o n m e n t , Ottawa, Chau,  p. 73.  Y - K and H . Saitoh.  1970.  D e t e r m i n a t i o n of s u b m i c r o g r a m  quantities of m e r c u r y in lake water.  Environ. S c i . & Technol.,  4:839. Fimreite,  N.  1970.  M e r c u r y uses i n Canada and their possible hazards  as s o u r c e s of m e r c u r y contamination. Fimreite,  N . , W . N . H o l s w o r t h , J . A . Keith,  1971.  L.  P . A . P e a r c e and I . M . G r u c h y .  M e r c u r y i n f i s h and f i s h - e a t i n g b i r d s near sites of i n d u s t r i a l  contamination in Canada. Hannerz,  E n v i r o n . P o l l u t . 1:119-131.  1968.  C a n a d . F i e l d - N a t . 85:212-220.  E x p e r i m e n t a l investigations on the accumulation of  m e r c u r y i n water o r g a n i s m s . Drottningholm.  Rept. Inst. F r e s h w . R e s . ,  48:120-175.  Hatch, W . R . and W . L . Ott.  1968.  D e t e r m i n a t i o n of s u b - m i c r o g r a m  quantities of m e r c u r y by atomic absorption A n a l . C h e m . 40:2085.  spectrophotometry.  32 Jensen,  S. and A . J e r n e l o v .  1969.  the aquatic environment. M a r q u a r d t , D . W . 1963.  B i o l o g i c a l methylation of m e r c u r y i n  Nature (G.B.) 223:5207.  A n algorithm for least-squares  non-linear parameters. R u c k e r , R . R . and D . F . A m e n d .  estimation of  J . S o c . Ind. A p p l . M a t h . 11:431-441. 1969.  A b s o r p t i o n and retention of organic  m e r c u r i a l s by rainbow trout and chinook and sockeye s a l m o n . P r o g . F i s h . C u l t . 31:197-201. Uthe, J . E . (ed.)  MS 1972.  M e r c u r y i n the aquatic environment:  a  s u m m a r y of r e s e a r c h c a r r i e d out by the F r e s h w a t e r Institute 19701971. Wobeser,  F i s h . R e s . B d . Canada M S Rept. S e r i e s 1167: 163pp.  G . , N . O . N i e l s e n and R . H . Dunlop.  1970.  M e r c u r y concentra-  tions i n tissues of f i s h f r o m the Saskatchewan R i v e r .  J. Fish. Res.  B d . Canada 27:830-834. Wood,  J . M . , F . S . Kennedy and C . G . R o s e n .  1968.  The synthesis of  m e t h y l - m e r c u r y compounds by extracts of methanogenic Science 220:173-4.  bacteria.  33  APPENDIX - L D  50  Experiment  P a r t of my Biology 101 teaching duties was running a two-week elective p r o g r a m for some of the students, test f o r  and I took the opportunity to  LD50 of H g (i.e., the dose of a toxic m a t e r i a l r e q u i r e d to k i l l half  of a population). S m a l l shore c r a b s were used r a t h e r than C . m a g i s t e r , expense and convenience of handling.  because of  The elective group made a f i e l d t r i p  to a l o c a l beach and c o l l e c t e d about 300 C a n c e r productus, H e m i g r a p s u s oregonensis, grams.  and H e m i g r a p s u s nudis, each weighing between one and two  The species were m i x e d and the m a l e - f e m a l e r a t i o was a p p r o x i -  mately equal i n each tank. F o u r students were r e s p o n s i b l e for each of nine t e s t s . was set up to show how many c r a b s a given dose would k i l l , i n c r e a s i n g i n eight steps f r o m 10.4 to 3400ppb m e r c u r y , tank.  with doses  and one c o n t r o l  E a c h 40 l i t r e tank was f i l l e d to 28 l i t r e s with seawater,  to an air supply, and c o m m e r c i a l f i s h food was added. dosed f r o m a r o u g h l y - m e a s u r e d solution of m e r c u r y , concentrations r e s u l t i n g were d e t e r m i n e d .  E a c h test  hooked up  The water was and the m e r c u r y  The c r a b s were added  i m m e d i a t e l y after the m e r c u r y . The experiment was c o m p l i c a t e d by the fact that (1) it took some time for even the strongest dose to k i l l c r a b s , tions decayed.  and (2) the m e r c u r y s o l u -  The decaying solutions were monitored three t i m e s - - o n  the second, fifth and sixteenth days.  O n the sixteenth day there was no  m e a s u r a b l e m e r c u r y in any of the tanks.  The average of the f i r s t two  readings was c o n s i d e r e d to be a guess of the average dose over the p e r i o d of the experiment. T a b l e A - l shows the average doses and the estimated t i m e s r e q u i r e d to k i l l half the c r a b s at each d o s e . extrapolated.  M o s t of the estimates were p e r f o r c e  34 TABLE  A-I  D o s e (|ig/ml)  0 9 43 73 500 725 1300 1200 3250  T i m e (days) 370 144 28 16 11.2 (not completed) 9.8 5.3 4.3  These r e s u l t s seemed to be i n t e r n a l l y consistent. A p p l i e d to the h a l f - t i m e experiment, they suggest the p o s s i b i l i t y of damage to the crabs, e s p e c i a l l y at the higher d o s e s .  

Cite

Citation Scheme:

        

Citations by CSL (citeproc-js)

Usage Statistics

Share

Embed

Customize your widget with the following options, then copy and paste the code below into the HTML of your page to embed this item in your website.
                        
                            <div id="ubcOpenCollectionsWidgetDisplay">
                            <script id="ubcOpenCollectionsWidget"
                            src="{[{embed.src}]}"
                            data-item="{[{embed.item}]}"
                            data-collection="{[{embed.collection}]}"
                            data-metadata="{[{embed.showMetadata}]}"
                            data-width="{[{embed.width}]}"
                            async >
                            </script>
                            </div>
                        
                    
IIIF logo Our image viewer uses the IIIF 2.0 standard. To load this item in other compatible viewers, use this url:
http://iiif.library.ubc.ca/presentation/dsp.831.1-0093043/manifest

Comment

Related Items