Frozen fish muscle—Chemical changes and organoleptic quality

I,'" I I I O 11! O l , { ) ( l ' i "

Vol. 3, No. I, 1967

FROZEN FISII MUSCLE-, CItEMICAL CHANGES AND ORGANOLEPTIC QU&LITY W. J. DYER Fisheries l?c.searrh lJor,,',l os C,m,d,,. llaliSax Lat)<,ratm'll, llaliSaz, A'o t'o Scotia. Canada |;'rozcll fish l)r(Jl)erl.v 1)r(Jee.-,sed :lnlOl"l{e life llltlV I)(, (Ir:lslieally re~hlro(i. 11 is well to reeolzllize the gre:lt variety of spoeio-~ of tisll :lll~] s}lellfisli differing widely in ('ontlms~ti(~n. I,il)id eont(,~lt lilgiV vgirv from h'ss than I% for the eoi,unervially imlmrtarit h':lll <~i'()llllilfish tyllifio(1 tiv ('o(l and h/ill(leek to lil~)i'o lh;ili 25% in .-~OlilO f:illv species, diffcr-e l l ( ' e s wlii('li :il'O roftool(,d in (,el(1 .-.forage elmr-

judged ll5, n ta.~tc p'mel (analytical). The j,ulges did not find it easy to discriminate The early slates of spoilage. It had previously been observed 1hat there was ;l sharp drop in grade as the rejeetion level was approached." On lhe other hand, samples of the same three lois of fish held for 4 weeks under good (-2(VC) or p o o r (-9°C) condilions of frozen Morage or ull(ler intermediate condi-

lions (--12°C for '2 weeks followed by slornge at --2(VC for "2 weeks) showed a sharply differenl l~allern (Fi,, 1) There was no change X

showe
]a;efoi't, ,lisrlis,,-iilg

t"]l:lll~(.'.', r('l:llillg tO (Ill'lilly ill lilt' llrolcins, lil)ids, :lilt] ol]lo.r coilsiil~ieili.-;i, well as" ihc llliysienl strucliire of the n-lu.~(.h, li:.~uo, licrh;iI)S ;iii illuslralion of tim imlmri:iliCO of qualily l,lss on frozen .storage to tile I1.'-(' of I]1o ti-~silo ;is 1"OO(1 wollld l)e liell~rid.

l"llozi,:X S'roItAGI,; QUALITY ~OSS

* !)r,,.-(:nlod nl lho T h i r d Annual M o e l i n g of (lie Socl(;ty for Cryol)ioh)gy, BosltOII, ~']ass,, AuglisL S, 19(.16, :is 1)girt, of :l synlposilllli, UOr.vool;cn~lislry of

Biological ]rood Sysl('nls: Interdiseiplingiry Considerlll ions."

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,~t --_o w, l)~;t lhose stored under poor conditions h:,d re:..hcd rejection levels in both lexture and taste. The halermc(liate samples

:it'l eri,-,i il's.

Sillee :l l)oor ~l':ide ill fish is gener:lllv :lssoriatetl wilh lmelerial -q)oilage prior to freezillg r : i l l i o r lh/ill with deterioration durillg frozen storage, ex]~erilnellls were sel up 1o lest file rehilive effects of the lwo deterioralive palhw:iys wilh mosl inleresiing restills. ''~ ('od fillols ()f grade I, very fresh: grnde lI, showing ilwil~leni spoilage: and Grnde III, of defil~telv spoiling but still edible quality, dM liol sllow sl~llislie'illv signifieanl, ({ifferenees when fro.-.hl.v frozen s:iinples were cooked by eilller lmldng, stoliming, or frying m~d

,"2"~"

fROZEN ~'I'OIt-~,(;E Qu.xLl'rY CHANiiES,

What h:tl~pens in tile tissue to result in lhese findings? The Izeneral changes in the proteins ummlly referred to ns dennlur:ftion and hydrolytic and oxidalive raneidity have been reviewed elsewhereY ' ' ~ ' ' In several speeies, the decrease during frozen storage i n taste lmnel neeepiability, espeein}lv tex/~lre, has 1)een found to be accompanied by a parallel exponential decrease in the proteins of the aelomyosin grotlp r,xlraelable in salt solution of relatively high ionic strengtli, usli<'llly 0.5 to 0.6 /,. Thi.~ decrease is assumed to be due to a denaturation, aeeompanied by aggregation, of 1he aelomyosin fe)rmed in lhe postrigor mus-

297

298

W . a . DYER

stable, primary covalent, bonds were formed during frozen storqge of cod. Secondary bondTEXTURE TASTE ing by disulfide, amide, or interprotein C---C 80 }nm(ls, possibly resulting from eouple(l oxida.,..:¢ ~io1~ of unatturated lipids, was implie'lted. 60 Thus modification of the structure (unfoldb.,I ing?) of ~he individtial proteins was apparrr" 0 40 ent ly small. L) Attempts to ehar:lcterize the component U') proteins in the myofibvillar (..,."alt-extr:mtable) 2O I)roiein extrae(s of cod muscle in (ho :.,nalytical ullraemltrifugc showed (we large 1;e'~ks in o ,A'IBjC AEBjC A'B'C A,B C the s('hlieren patfern of posirigor muscle, tenBAKED FRIE D B A K E D FRIED tativelv identified as ae(omyosin, peak ]'I, and Fro. 1. t.';ffecLof frozen storage deterioration on as myosin, peak III, together wilh smallei' taste lmnel acceptability in 1)'d
FRO,ZEN F I S H M U S C L E

tim liquid l)hase. No evidence was found for t,he presence of .qctomyosin G in unfrozen muscle apparently tiffs aI)l)Caz., on]y when the ionic slrengih has been raised by freezing (F. King, personal eommm~ieation). It does not, appear to have been established Amn, wlmther "~etin F appears in the gel fra e" which ol)viously needs more aiiention. In cod extra(,/.% the ~fitraeenirifuge patterns often show broa~lene(l peaks indie:,,tive of gre~:ter heterogenc,fi~y or degradation thnn indiealed above"' and Ilms perlmps other reactions will have to be considered and incorporated into a further nmditied model. Likewise the situation in prerigor frozen stored muscle is not clear .as yet. ]'{It(OR ~]~OWI'IS AND ]{I'JLAXATION

Major changes in the proteins and their interactions oec'ur as the muscle proceeds from the prerigor to the rigor mortis stage followed by subseq~wnt relaxation or tenderiz:ttion as it is sometimes termed. One illustration of these changes is the Dcmicke effect well known in fish muscle as in other qnir ~ 17 ma].~. Another eonsiderabh; diminution of myofibrill.'~r l)rotein extrnel:lble in s:dt solution occurs during the rigor phase, as the hydrolysis of adenosine lriphosphate promotes inerv'tsingly strong at laehment of the sliding aetin and myosin filaments. On resoltltion of rigor, however, extractability increases to nearly quant, ita tivc levels in fish, usually higher tll,'ul Jn ~he prerigor extract, TM the cxtr'mt now eontainh:g an aetomyosin and not simply a mixture of acfin and myosin. Thus the release of actin from ~l~c basic structural network, the z line;~: or possibly retieulin, seems an essential feature of the relaxation -,1 c,~b. Depolymerization prct, of actin could also be involved, and indeed it, has been suggested that two aetinins are present, "~ end tending ~o l)olymerize aeiin, the other to depolymerizc il. There remains the possibility that the I)oor extractability could be related to a, slower penelralion of peptizing solution into the eol]l ratted fibres during the b]ending. Temperature during p e l a g e into rigor strongly intluences the Deutieke effect. For ins{mine, in cod fillets, no reduetion in .,;altextrae(able protein o(,curred when lhe muscle went through rigor at, 0°C but a~ 25°C a , -j~<-,

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fillets fi'om rested cod. Box indicates approximate duration of rigor mortis stiffening. strong reduction to about 57%, of the initial value occurred at maximum rigor (Fig. 2) foliowe.d by a rapid return to the initial level on resblulion (W. ,1. Dyer and D. I. Fraser, unpulflished). Shortening was markedly greater at 25°C than at 0°C, 35% as compared to only 15% and less at, 0°C (Fig. 3). A similar shri~lkage Was observed in samples of muscle from prerigor fish soaked in water at room tempcr'~lureY The marked contraction occurs r,ather sh:trply at temperaiures above 16°C. '''''~ It seems that the physical na,ture of ~he eontr,tction in situ ought to receive more attention. I t is observed that prerigor fish muscle is pliable and tim surface is dry. WitA~ lhe onset of rigor and the contraction which occurs with inleraetion of the actin and myosin filaments, there is a force acting to shorten-the ce]ls, which results in a measurable thickening of the tissue mass. The cells become turgid and presumably the contractile, force of the filaments is opposed by the stretching of the cell walls and/or the collagen network surrounding the cells. As the energy sources, ereatine phosphgte and adenosine /riphosphate, become exhausted, the cell membranes become permeable to water and salts, and relatively suddenly the tissue mass becomes wet with the escape of water from the pressurized cell. The immediate result is a loss of turgidity and of stiffness, and a degree of relaxation. A contributing factor-is probably the decreased moisture-binding capacity of the protein duc to ,~ decrease in pH as glyeol)@s proceeds. This development of stiffness to ma,ximum rigor followed by a rapid

W'. J. DYER

300

muscle .stored 7 days than from muscle s(ored 2 days or samplt:d after slaugh(er, the concen|raiion of adenosine tril)lmsplm.le neeessary 1)eing 0.1,0.2, and 0.6 m~r. Much more. work is necessary to tel'tie th(,se changes to qualily in frozen meat during storage, tlmwil}g, and cooking. TEMPEI{ATURE [FI~UCTUATION,

T]1aWlNC,,

AND ]{ EFREEZING

Fro. 3. Contraction at, 25°C (left fillet) vm;s.,s 0°C (right fillet) in two fillets cut from "t freshly killed cod, each 30 n-fin, after placing in a polyethylene bqg and imm(.'rsing ih writer at 25°C or 0°C. but only purtial relaxation in an experiment with cod muscle is illustraied in Fig. 4 (W. J. Dyer, unpublished results). The rather sharp increase in contraction above about. 16°C s,,,~,~,~ -~t~...~_~,,,, a specific cause, perlmps a we:tkening of the structure or a change in the properties of lhe cell wall or its surrounding connective tissue. Collagen undergoes' such a change 'in state which in isolated collagen from cod ,skin occurs rather sharply at 13 -+- 0.5°C, tlm actual temperature being somewhat dependent on the method of isolation."' Solubilization of collagen in beef and rabbit muscle a t 370C has been observed but only when the samples were hea,ted prior to storage? 2 That changes in tim actomyosin m a y Mso ,occur in the postrigor please is noted, r)~rabbltsactomyosm becoming flflIy d~ssomgted a{~'ai much lower concentration of adenosine tr]ishosphate when prepared from . . . .

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The queslion of relative eft'eel on slor'tge chgmges and qualit,y of product during s(or:tge at thletualing temperatures as compared to relatively constant storage lemperatures is receiving renewed "tttenlion. Early |ilernlure, reviewed by Woodroof nnd Shelor?' indicates that some authors foun(l a difference while ollmrs concluded that fluclualion is not, delriment'll. ])robably the difl]c~flty of maintaining constant le)nperature (.onditions for ]on~ periods alld the problem of accurate measuremeats of small eb:)nges in quality param.,,ters and of standardization of evaluation proeedures have prevented sufl~eienlly accurate assessment. .Most subsequent results using storage temperatures fluctuating from *he "tve)'ttge temtmrature ht~ve shown no significant effec.t wilh severn[ fish and meat products?" ~'" '~ However, most of the experiments were eonducled with an avernge temperature in the vieini(,y of --18 1o --15°C; since fish in particular may be snbjee(ed to much higher iemperatures (up to - 8 ° C or higher), during transport or loading, '~':'s experiments were conducted with cod exposed temporarily (o temperatures up to - 9 ° C . '-''' -~" Considerable deterioration in quality as measured 1)3' lnste 30b-

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:Fie,. 4. Deformation, percentage of thickness, measured by a "precision" penetrometer with a square }mad, 0.5 era"-, pressure 100 g pro" cm ~, aciing for I rain a,t the head end of fillets cut fl'om resting cod stored at 25°C and 0.5°C.

FROZEN FISH MUSCLE

panel sad protein extractability occurred in fillets subjected to a 2-week period at -90C, after storage for either 1, 3, 6, or 9 months at --IS°C, (luring the subsequent storage a.t -1S°C. The quali~y loss was greater with doubie exposure and less with shorter times, 1 week 'rod 3 days. Statistical analysis of these and of more recent experiments (W. J. Dyer and D. I. Fraser, unpublished) indicated that the decreases, dur.ing subsequent storage, in t'tste scores and in ttie chemical changes, extractable prolein, and free fatty acids were for the most part significant (P = 0.05) when material stored a~, --]S°C or at --23°C for various periods was exposed to tempera4ures of --9 or --12°C for a 2-week period. The expected loss due to storage 'It the higher temper'tture did not account eoml)letely for the change observed, though more accurate measm'ements would be desirable. The most i~teresting and i,nporlant observation was tha~ the quality decrease occurred in the storage period of a month or so subsequent to the return of the samples to the original slor,qge temperature. A parlial explanation for this finding in cod in contrast to the previous work with other materials is provided by at, examination of the freezing out. curve for water in fish muscle (Fig. 5). Because of the shape,"' a temperature variation occurring in lhe region o f - - 5 to --10°C will result in a muel~ greater thawing of ice or freezing out of I

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Fin. 6. Quality, taste panel scores, of round ~rap caught cod and of rcfrozen fillets st0rcd at --18°C (aftx:r 20). o, round, gutted cod stored at --23 to -26°C: x, refrozen fillets, unstorcd; O, RFO; +, RF2; ~, RF4; [3, RF6; refrozen fillets stored at -18°C, cut from round fish thawed after being held 0, 2, 4, and 6 months at" --23 to -26°C (95% confidence intervals of mean scores are shown). water than the same variation occurring in a. lower temperature range, --20 or --30°C. This rehydration and dehydration of the protein and change in sqlt concentration would be expected to cause damage to the protein with consequent loss in quality. ]~.ecent studies on refrozen cod fillets appear related to the above findings. No immediate change in quality usually results in refrozen fish/' ~' :~''~o.= but few authors report storage quality of the refrozen product. With Newfoundland trap-caught cod likewise, no immediate loss of quality after thawing a n d refreezing of whole frozen cod 'stored for various periods up to 6 months was found as meas~red by taste panel though increased thaw drip levels and slightly decreased ' protein extractability indicated flint some change in quality had occurred." O~(storage Of the~ refrozen fillets, however,, ia.rge decreases (20 to 80 points) in taste panel scores occurred wiih]n 2 months following refreezing (Fig. 6). The decreases were'larger at --18°O than'at - 2 3 ° C but were similar whether thawing was carried out after a few days, or after storage of the round fish at a b o u t . - 2 3 ° C for 2 or 4 months. Accompanying decrease2~ in protein extractability and increases in free fatty acid also occurred. Similar changes in refrozen trawler-caught offshore cod were observed/"

302

W.J. DYER

t,hough other results" indicate lhat such rapid changes may not occur in all lots of fish. This increased rate of quality loss following refreezing or temporary exposure of frozen tissue to higher temperatures (short of {hawing) may bc influenced by several fact,ors. The solution of salts, protein, and org,,mie materials remaining unfrozen in the tissue is very viscous at, low temperatures, thus greatly reducing diffusion and restricting the availability of substrate to "the various enzymeaetive sites and for chemical and physical reactions. Itaising the temperature even for short periods wmlld promote diffusion and redistribution of metabolites, allowing reactions to proceed even though ~he temperature was again reduced. Another contributing factor could be the effect of lhe melting and iefreezing of ice erystgds in stretching and bre~'~king down the various membranes normally separating the cell constituents, resulting in liberation of enzymes and metal)elites from the initoehondria and oit'er cell inclusions. Various enzymes remain active in frozen tissue," ~' and evidence of release of the enzymes glutamate dehydrogenase and 3-hydroxybutyrale dehydrogenase, following damage by freezing and thawing to rat liver mitochondria has been obtained.'" ~'~ One should also remembm" the maximum activity of many enzymes at ~emperalures just below freezing in the region of --2 to ~4°C in fi'ozen tissue (reviewed by Love and Elerian"°). Also in refrozen cod, fatty acid formation is rapid following the refreezing;TM '" and the increased free fatty acids could promote insolubilizat,ion of. aetomyosin-or of actpmyosin F. The increased volume of less viscous tissue fluid with a. salt concentration of 0.5 .~r or higher consequent on the rise in temperature would promote dispersion'of the myofibrillar protein away from the myofilaments into the liquid phase. Hare it wouhI be in a much less, organized state and much less stable than in situ. ~' ~'~ LIPID

HYDROLYSIS

AND

LIPID-~ItOTEIN

]INTERACTION

A few years ago it was. observed thato extensive lipid hycrolysis with liberation" of free fatt, Y;."aeids):aceompanied frozen.storage deterioration{~ in" many fish species=~,o,....~ though not in all." Further in cod at --12°C

extensive lipid hydrolysis preceded lhe decrease in protein extractal)ility while at --_,~ ~ 1}o clmnge in taste scores or protein extractabilit,y lind occurrecl tl 1) to 12 mo,nths even though there was some hydrolysis.'", ~ Sm'prisingly, oxidation of the highly unsaturated N tty acids ]i!Jera1ed '~1)pea red 1o be very limited, a fact al..o, s noted in the.fatty a(:ids liberated during salting and drying of cod." ens~t~zaThe original suggestions, first, the ,.~, "".~ tion of protein to denaturation by liloe,'ated free fatty acid and, secondly, possible stabilization of prolein by unhydrolyzed lipid or a lipoprotein complex are supported by the experimental findings'~ that dispersed actomyosin was insolubilized by added fati.y acid, thai the amount of f,tity acid salts necess:ti'~to reduce prolein extractability was related to the total lipid present in nmscle from a series of seven species, and that neutral lipid proteete(1 againsl, free fatty acid."'":" Fatty • "1 and :l:~onfailv species aPl.ear to behave somewhat differently, depending on localization of lipid occurrence within the tissue, nnd pH may also affect (he free fatty add-protein interaction": lhese observations may account "" ' 1'" .q For for some of the "q~parent chscrep,t).e~e.. instance, lean fish may be denatured fairly rapidly, having no neutral lipid to counteract free fatty, acid formation; simila,rly, fatty fish h a v e /heir li]fid present as globules hal ' O.rots, while an in eomaet, with the muscle filam intermediate, slighliy fatty group of fish qre denatured only slowly, having available neutral lipid to eounter'~et, *he fatty adds. Just recently, no obvious trends in the composition of the free fatty acids or of 1)hospholipid in relation ~o protein stability were found.'"' '~ If one returns to eonsideralion of mechanisms, it, seems lha, only a start, has been made in seeing how the lipid or free fatty acid is bound with the protein fractions and modifies their l~hy,.moehcmlca -.~', , ", 1 properties and • , ~ ' ,* tion or polymerization. the degree of ,~,~,te=a Anderson, Steinberg, and King "° have recently contributed signifieanl new data supporting the hypothesis that lipid hydrolysis" is related to frozen storage-induced denat~urat,ion of lho protein. Maximum binding and ii~solubilization of added fatty acid and protein occurred at about, 0.5 Ix, and the reaction w~~s very much slower, comparable to rates observed in situ,

FROZEN FISH MUSCLE

:it ptI 6.4 than at 7.2. Since in frozen muscle the rqlo of denaturation is maximum in the region of - 1 . 5 ° C where the toni:, concentration will be about ().5 t~, this very nearly eoincicles with the point 'of nmximum preciI)itt~},ility of protein with fatty acid and 'l!so is ne'er the optimum temperature reported for lipid hydrolysis, --4°C? ~ Freezing amy" also affect the pH of lhe unfrozen plmse in the tissue, ~ llms further modifying the free fatty aeid-I.',rotein inleraetion. The weaker combination of protein and fatty acid at 0.8 as compared to 0.o l~ can be related to the dissociation of aetomyosin lo myosin and rmtin at tim higher ionic strength as indi('ated by ,Htraeentrifugal an'dysis:'~ Thus, the dissociated myosin requires much more fatty :wid for insolubiliz:ttion ihan does actom3"osin. This co, rid also mean tha, t muscletreated with s:~lts such as polyphosphate in combina.tion with brine at concentrqtions suffieienlly high ~o promote actomyosin dissociation'" should be less subject to fred fi, tty acid-induced insolubilization. Any accompanying ineree, se in pI-I, however, dtm to the alkalioe 1)holyI)hosphate added would result in the opposite effecl. The possible role of aetomyosin G formation in the frozen tissue must. also be considered. This fatty acid insolubilization of actomyosin and of myosin fits in with Connell's finding !hn,t 'mtin and ,'~ considerable part of the myosin can be recovered from a den'ttured tissue..Presumably the fatty acidprotein complex is brought into sotu}ion by ~he solvents used in the isolation procedure. Much more data is needed on the effect of pH and ionic strengttl in the muscle tissue •rod in (he cell fluid at frozen storage temperatures, and on the site of release of fatty acid and of damage to cell inclusions and membranes. It, wiI[ now be obvious that the many edible species of fish as well as the various classes of shellfish may differ greatly from one ano(her and from the vario~ls mammalian and avi'.ln species. While there are m a n y similarilies, there are also many differences lo be considered before the frozen storage changes occurring in the v'lrious m~lsele tissues and their relation to quality can be brought together in a theory or theories encompassing fisla, fowl,

303

and meat which will enable us to make more significant, progress in the prevention of qualit,y det,."rioration in the stored frozen product. REFERENCES 1. Anderson, M. L., King, F. J., and Steinberg, M. A. Effect of linolcnie, iinoleic~ and oleic acids on measuring protein extractability h'om cod skeletal muscle with the solubility test. J. Food Set., 28: 286-288, 1963. 2. Anderson, M. L., Steinberg, M..A., and King, F. J. Some physieal" effects of freezing fish muscle and their relation to protein-fatty acid inter'tetion. [n The technology of fish utilization, R. Kreuzer, ed., pp. 105-110. Fishing News (Books), London, 1965. 3. Banks, A. Recent developments in the freezing of fish at, sea. II, The. quality of seafrozen .cod. Chem. Industr., ~8: 1360-1362, 1955. 4. Bendall, J. 12. Meat proteins. In Proteins and their reactions, H. W. Schult,z and A. F. Anglemier, ed.. Chapt. 11, pp. 225-254. Avi Publislfing Co., Inc., Westport, Conn., 1964. 5. Bul,tkus, H. Preparation and properties of trout myosin. J. Fish. Res. Bd. Canada, 28: 563-573, 1966. 6. Cardin, A.; and Bordeleau, M. A. Changes occurring in (,he faL during tim processing of salt fish. Fish. ltes. Bd. Canada, Prog. Ilept. At,I. Coasl Sta.. 6'6: 16-20, 1957. 7. Castell, C. H., Moore. B. A., Janga,trd, P. M., and NeM, W. E. Oxidative rancidity in frozen stored cod fillets. J. Fish. Res. Bd. Canada, 23: 1385-1401, 1966. 8. Connell, J. J. Changes in the aetin of cod flesh during storage at, - f 4 ° c . J. Set. Food Agrie., 11: 515-519, 1960. 9. Connell, J. J. The relative stabilities of, the skeletal-muscle myosins of some. animals. Bioehem. J.~ 80: 503-509, 1961. 10. Con'n.ell, J. J. Changes in mnount of myo~n ext,'aek'tble from cod flesh during storage al; ~-t4 °. J. Set. Food Agric., 13: 607-617, 1962. 11. Connell, J. J, Fish muscle proteins and some effects on them of processing. I n P r o t e i n s and gheir reactions, Tf. W. Schult,z and A. F. Anglemier, eds., Chapt. 12, pp. 255-293. Avi Publishing Co., In(':, Westport,. Conn., 1964. 12. Cqnnel/, J. J. the use of ~odmm dodeeylsulfate in the study of protein interactions during (.he storage of cod flesh a~ --14 °. J. Sci. Food Agrie., 16: 769-783, 1965. 13. Dingle, J. IL, Ellis, D. G., Hines, J. A., and Lander, J: T. Proteins "in fish m:mcle. 18. Sedimentation patterns of myosih-B extracts of prerigor cod muscle. Canad. J. Biochem., /d-: 1915-1926, 1963. 14. Dyer, F. E., and Dyer, W. J. Changes in the palalabili},y of eod fillets. J. Fish. Res. Bd. Canada, 7; 44.9-460; 1949. 15. Dyer, W. J. Protein denaturation in frozen and stored fish. Food Res., I6: .522-527, 1951.

3,04

W. J. I.)YER

lira st ruetl)ro of nnl ural :jnd .,,vnl hp! i,' In'el eh~ 16. ])yer, W. ,l., B,'o('kerlmff, H., Hoyle, l(. J., and l"ra,-i'r, ]). I. Polyphosphnte tr(.alnlent of filanlpnls j'ronl slrh)lod musvl,.. J. Melee. Biol.. ," : 281-308, 1963. fi'oz(,n ('od. 1. l)n)(ein exlr'telibility antl lipid 33. Johnson, P., Napl)vr, D. ]1., m~! Rowe, A. J. hy(h'oly.-is. J. Ft.-h. Res. Bd. C:an,ula, gI: 101S('di/lw/llali(m .',lu(lh's on l)()lynwrizo(l :,olin 106, 196-1. soh/li,m.,.. Bio,'hiz,). Bi()pl)ys..\('l:~, 7~: 36517. ]3yer, W. ,1., and Dingle, . l . R . Fish proteins 373, 1903. witl~ si)e('ial ri,feron('(, 1o freezing, hi ]"i.-h 1is 3,1..l()tn,s, 'N. 17., Burl, J. R.. M iIrrav, J., "ln(l food, G. I.))org.-tronu ed., Chapl. 9. pp. 275327.-\va(h,mi¢' Press. New York, 1961. Strol/d, (.;. D. Nm'h'o(uh's an(l lh,. :(naly/u.al 18. Dym'. W. ,1., ami Iq'a,.,r, 13. I. P)-o(eins in frozen al)pr()nch 1o )he rigor nm)'tis l)rotJlenl. In Th(, fisl), la. l,il)i(! l)y~h'olysis. J. Fish. lies. Ihl. (o('hnohJgy of fish utilization, I{. Krellzor, od,. Fislfing News (Books), l,ondon. 1965. Canada, 16: ,13-52. 1959. 19. Dyer. W. J.. and Igrnse/", 1). I. 5l()J,-.lur(" in ti,-I~ 3,'-5. King, F. d. Ultraeon(rifug:ll antlly.-i:- ()f clvlnges in (he ('omt)osi(ion ¢)f myofil)rilhw pr¢)(('ii, (,xl)lo('ks l))'oeess('(i froln vc,ry fresh fi,d~. Canad. traels ol)tnin(,d from fresh and fri)z('n ('od ];l:,hern~al), .iS(8) : 17-19, 1961. m~z.-('h'. J. Foo(l 8ci., 31 : 6,19-663, 1966. 20. ])yer, W. J., Falser, D. I., Ellis. D. G., [diet', 36. King, F. J.. An,h,rson, iM. I,., an(l S((,inl)(,rg, D. R., 5l:)eC:illuln, W. A.. an(! lmisl)h'y, l';. M. A. ]iea('tion ill" eo(l 'wlo)nyo.-itl wi(l~ I/noQuality elm/Ig('s in stovo(l refrozon cod /illels. let(' and linoh, ni(' a('ui.-,. J. l"o()(1 S('i.. 27: 3(i3Bull. lnsl. lnlern, l"roi({ Ann,,x, l: 515-52,1, 366, 1962. 1962. 37. Klose, A. :\.. |%ol, .'%1. F., an(l Lint,woav,,r. ]I. 21. l)yer, W. ,}., Fra.-.'er. ]%. I., Ellis, D. (;., an(1 Effect ()f /lu¢'tuttti)Ig teml)t,)':lluv(,s ~)n frozen ~la¢'C'all¢~tn, W . . \ . In/l~a,~we of intorntittr, nt, lurk('ys. Foo(i Te('h., O: 372-376. 1955. short slora.u.e p,'riods a~, 15°1:, ~Ls encountered 38. ],('n(z, C. l'.. an(l Rooke, l':. A. Teml,(,ratur,.~ in during refri~(,ratt()r (':it' (ran.-l)ortation, on the frozen pouIlry, frui(, an(l vegetabh,s shipl,ed quality of frozen veal stor(,(l at 0°F. J. Fish. l)v ro'id in r(,fi'igeral(,d tr:dh:rs. I,'¢)od I,~ ('auReS. J:i([, (Tan:l(ia, 1 J: 627-635. 1957. aihl, 2I(12) : 3S-1 I, 1961. 22. Dyer, W. J., Fraser, D. I., and MacCallum, :19 1,on'ntz('n, (I. N,)('n forsok nwd ",l~flg)(,ItW. A. l"urlhe: s~u(ly of (hi, it~ihtenr'(' of slmrt fi'ysin~" av lorsk, l(uhte (l)('nm:u'k), 19(,1) " sit, rat:(, perio({s, 3 days to 2 w,,eks at IS°F, 53-58. 1965. on tho ilu;~HIy of frozen rod stored at 0"F. J. l:i.-h, lies. Bd. Cana(la, I.;: 925-929. I957. ,10. 1.eve, R. hl., :rod Elorvm. hi. K. Proi¢,m ,.Irn:lturaiion in frozen fi.-h. VIII.-1"Ira ten> 23. Dy('r, W. J., Fras(,r. D. I., ~[aclnlosh, R. G., Iwrature of ~nztximum demdur:ttion in cod. and ~lyer, 5I. Cookintz methoil and l)ahtt:thiliiy of f|'ozer~ t'ot{ filleis of varioll.., (lualitivs. J. Set. Foo(l Agri('., 15: 805-809, 196,1. ,1. Fish. Re.-. Bd. Cana(ia. 21 : 577-589. 1964., 41. l,ow,ru, ,I. A., and Olley, J. Iuhibilion an,t lw()2,1. Dyer, W. J., an(1 Morton. M. L. ,Storage of motion of l)osl-xnorten~ lipid hy~h'ol.vsis in frozen plaice fillets. J. Fish. Res. Bd. Canada, Lho flosll of fish. ,1. Food Sci., :?7: 551-559, tS: 129-13-1.1956. 1962. 25. Dyer, W. J., ,Morion, M. L., Fraser. D. 1.. :rod 42. ],us(,na, C. V., anti ].)as:, C. M. ,qtru('l~l~al Bligh. F,. (3. Slorage of frozen rosefish fillets. uhang(,s a.%ociated with (hi, r~,h,a.~e of (,nJ. l,'isll. Re.~. I3(1. Canad'i, 13: 569-579, 1956. zymes from ral-liv(,r miloel~on(h'm lw fret,z26. t':bt~shi, S., and ,:kIaruyama, 1£. Preparation and ing. Cana(l. J. Bioehem., .~ ~: 775-781. 1966. some properties of =-aetinin-f,'ee aetin. J. 43. Lu,-.ena, C. Y., nn(l l')el>o<':is, F. iletm'ogeneity Biochem., 58: 20-26, 1965. uml tlifferenliat fragilily of rat liv('r mito27. Ellis, ]). G.. and Win(:hester, P. hi. I'roleins it: dmndria. Cannd. J. Bioel~,n~...~.~: ,I!)7-508. fish mu.~ele. 12. l-ltraeenirifuge studies on 1<,)66. post-rigor extraets of structural "protein, J. .t4. Ma('Calh~m, W. A., Laisl)lc, y, E. J., Dy(:r, W. J., Fish. Res. ]3(1. Can:t(l'~, 16: 33--41. 195!). and Idh,r, D. R. Taste pnn(,l n.-s(.s.-nmn(, of 28. Fujimaki, 5I. N., Arakawa, .\., Okitani, A.. and co(l fill(:|s nf)er single nntl dent)l(, fre~,zing. Takagi, O. The changes of myosin 13 ("At'to,1. ]"is]). ]~e,,-. Bd. Ca/m(la. 23: 1063-1081, myosin") dtu'ing storage of rai)i)it )nusch,. 11. 1966. The dissociation of "5lyosin B" into myosin •15. Nog)/ehi, ]':. S))/'inkage of fi.-h Inus('lo ,-c):~ko(l i)) .\ aml at.tin and its interacimn with ATP. J. w:tler. ]n '1"11(, loe}moJogy of t}:.1~ ~,i])zalion, Food ,q('i., 30: 937-9-13. 1965. R. ](rettzer, e,l., t31). 70-73. ],'isl~ng Nr.ws (Book.-s'). London, 1965. 29. Gould. E. Testing the freshn(,ss of frozen lisb. Fishing News (Books), London, 1965. -16. Olh'y, J., alld Dtlll(';itl, ~V. 1)~. Lip)(l.'. :)nd pro30. 'Gutschmidt. ,l. 0 h e r das Gefri'~'ren yon Seelein donaiuration iu fish muscle. J. Svi. Food AgrJe., 16: 99-10.1, 1965. fisehen. Kiilteteelmik, 18: 210-225, 1961. 31. IIan.-on, S. W. F.. nml Olley, J. Obs(,rv:~tio))s on 47. Olle3", J., and I,overn, J. A. PhospholiI)id i~ythe relationship betweei~ lipids and p/'otein drol.vsi.~ in cod flesh stored at various tem(tete/-ioration. In The technology of fish utiliperatures. J. Set. Food Agric., II: 644-652, zation: R. Kreuzer, ed., pp. II1-115. Fishing 1960. News (Books). London, 1965. 48. Olley,.J., Pirie, R., and Watson, I-[. l,ipase and 32. Huxley, El. E. Eleet/'on microscol)e studies on l)hosldmlil)ase activity in fish skeletal muscle

F l t O Z E X FISH MI:'SCLE :m~l its relationship to protein denaturation. .l. Svi. iVootl Agric., I.J: 501-516, 1902. .19. Poters, J., MacC'dlum, W. A., Dyer, W. J., Idiot, D. It., Slavin, ,l., Lane, J. P., Fraser, D. 1., and Laishh,y, E: Storage ~luality of refrozen cod fillets from prerigor and postrigor fish, brine m" plate frozen, and thawe~l in wato," willl microwave energy and a preliminary study of tlm mim'owave thawing process. J. Fisll. l{(,s. Bd. Cuna(la. in lm"ss. 50. P,,t,,l'~. J.. :m~i Slavin, J. l"r~,ezin~, of fi.-h ai sea. ]"ish, Gazette, ?7: '32-136, 1960. 51. Poltinger, S. 1~. Effect~ of flucluating storage tomporai,ures on ¢tuality of frozf,n tislt fillets. Comm. li'i;h, l{ov., 13(2): 19-27, 1951. 52. Slmrl~, J. G. Aseptic autolysis in rabbii al]d bovine nmscle duz'iz~g storage at, 37 °. J. Sci. l,'oo
55. :56.

57.

Bull. Jap. Soc. Ski. Fish., o0. "' • 1022--1036, 196't. T~tylor, D. H., an([ Harrison, J. S. M. Itefrig~,rafor ear test~. Fish. Res. Bd. Canada, Pat. Prog. Ropt., 7.~" 23-25, 1948. Tonllin.'.on, N., ,Ion,is, R. E., aml Gciger, S. E. C,l.~'o[5.,q~ in lingcod mu.-cle during frozen storage. J ] Ml. Res. Bd. Canada, 20" 11451I52, 1963. Van dea Berg, ]~. Changes in p H of some frozen foods during storage. Food Tech., 15: 43.t-,to ~, 1961. \Vldte Fish Aui, hority, l{eport on an experiment into freezing of fish at sea. Tilbury House, Petty France, London, 1957. \Vint~,r, J. D., Hustrulid. A., Noble, I., and Ito~.. F. S. The <,fleet of fluctuating storage t~,ml)eraturc' on the quality of stored frozen foods. Food I ech., 6 : 311-3t8, 1952. Woodroof, J. G., and She lot, E. Isffect of freezing sioragc on sirawberries, blackberries, ra.~pberries and peaches. Food :Freezing, 2: 206-209,223, 1947. Ymm~z, E. G., and Lorimcr, J. W. The acidsolubk, eollalzen of cod skin. Arch. Biochem., SS" 373-331, 1960. c

58. 59.

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