Chemical changes in freeze-dried foods and model systems

CIt YOII I O L O G Y Vol. 3, No. ,1, 1967

CHEMICAL

C t l A N G E S IN F R E E Z E - D R I E D MODEL SYSTESIS~ "

FOODS

AND

MAIICUS KAREL, T£IEODOI~E P. LABUZA, AND J 0 t ] N l:'. MAI~ONEY Department o/,Vutrition and Food Science, 3f assachusetts l~mtit ule oJ Tech,nology, Camb ridge, Alassachusetls The potentially high organoleptie and nu~ritioaal qu~llity of freeze-dehydrated foods is limited by several chemical and physical deteriorative processes. These processes may occur during processing or in storag(' and are s~r(mgly influenced by vario~s fi~ctors such as ten~pcrature, moisture content, and the presence of oxygen. This paper discusses some recent work pertaining to oxidation of ]ipids, a deteriorative mechanism of primary import',nee in the storage stability of dried foods. Iff the case of foods, oxidation of lipids is usually initiated by peroxidation of unsalurated fatty .,.,;ds" it results in a munber of undesirable changes in flavor, color, and mltritm.ml value, as indie'~ted in TaMe 1. In addition, Iipi~t oxidation products participate iv. reqelions with nonlipid constituents and can produce in. foods more complicated effects th:m ~hose observed in pure lipid systems. For instt~nee, hydrol)eroxides and other oxidation products may react with proteins to produce, mMesirable textural properties by causing protein aggregation t'~ or seission (A. Zirlin and M . t(nrel, unpublished results). Oxidation products of fa.tty acids nmy also parlieipate in nonenzymatic browning, as indicated in Table 2. This typ.e of nonem, ymati,: browning in dehydrated foods is I)a t'tieularlv.~ m~por.ant " ~' because it can occur at moisture contents well below those optimal L, II*1 .~ 1

tlt.~

°

I

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* Presented at the Symposium on Freeze-Drying, Sooiety for Cryobiology, AugusL 8, 1966, Bo~ ton, Massachusetts. t'This study has been supported in part by U. S. Public I-Iealth Service Research Grant EF 00376-04 from the Division of Environmental Engineering and Food Protection and by Pro~ect No. 41-609-66-46, Aerospace Medical Division, Air Force Systems Command, United States Air Force, for Milch the authors are grateful. This manuscript is eontribu~t.ionNo. 1031 from the Department of Nut,rition a%d ]Yood Science, Mass:tchusei.ts Institute of Te~ihnology.

for sugar-amino browning, which predominates at high moisture contents. ~ One of ~he interesting features of lipid oxidation in dehydrated foods is the oeeeleratibn of the reaction at low m'oisture contents. Water is known ~o ret~rd lipid ext.. dation in m'Lny dehydrated and low moisture food I)ro¢luetsY"" '~ k number of hypotheses have been advanced to explain the protective effect of water in retarding lipid oxidation. The most imporl~lnt :ire: 1) thai water has a prole " "~ effect, due to rctardatmn of oxygen .~el~¢. diffusion;:' 2) that water lowers the effectivehess of metal (;,:t(alysts suelt ~s ('OPlJer and iron, o) that oxygen is attached to sites on the surface, t.hereby excluding oxygen from these si~es; ~'' 4) thai wt~ler promotes nonenzymatic browning, and browning can restilt iu the formalion of ant ioxidant~ com.pounds;" and 51 lhat water forms hydrogen bonds with hydroperoxides and re~ards hyd roperoxide decomposition. Recently, we have reported on studies condmqed on freeze-dried model systems based on cell,lose and ]inoleate.. The results of these studie.s supported the conclusion that water inhibits oxidation by the followillg mechanisms" 1) hydrogen bonding between water and hydroperoxides resulting in changes in the normal mecha,isn~ of initiation of the chain reaction and 2) deactiv:~Iion of metnI ea~alysls by hydration of their coordination shells?' u This paper presents some additional evidence for the proposed mechanisms in the cellulose-based model system, as well as resulks obtained with a protein-based model system and wi~h foods. , N)

O

I~'L~TERIALS AND

~'[ETHODS

Three model systems and one food material were used in the s.ud-,t.' of eflee.s" l. of water 288

289

CItEI~IICAI, CH-~NGES IN FIIEI,]ZE-DI:tlED FOODS

v~illlt'il{, on oxi(la'ion of lil)ids. The conll)osilioli of lhe lnatori,Iv] = [li slil'l]lll:irv, tilt'so procedures consist of higil .-.-l)~,ed lnixing of oonll)ol~eilts, t, xlrti:loll lille re:iclioll flask% freezing in liquid ,illro~(.ii, l'r(.ezt>-. Th(, ,-orl)tion isoliiol'll~.s for each s.vsleiri wi,re (tolerlniiwd wiih lho USe Of melhods de.trilled }ix" lr(elr(,l and .;Niekerson.: Salm(m llS('~l iii seine of lhe experiments was knmvn to have been eaugh~ and frozen loss ll~:~l] I we~:k prior to arrival in our laboratory. Tll(..,:,lllioll w:t~ .-:lived into !,~-ili slices :lad frooz(,-~lried. From ~he dried slices all tile ~tark folly li.,..-,lio :tim liar: th,~lt :~round lhe "i'k BI.I,; 1 LIPID {v)XIIIA I'i~)N

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TABLE 3

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8"FUDil.:i) (l)itY Basis) 1

Cellulose Sy-_tem • ¢

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(,Jli

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%

78.0 21.9

1.11

lipid

.

l0 jlpm* !

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I

II 100 ppm I

* Ill col)all -eat :dyzed st tidies. "i"Nol (lelerinilwd. cenlra[ cavity were removed; tim reinaining lissue was br(Jkcn up and mixed to insure s:lml)le homogeneity. The samples were tlmn w(.igtied into rezletion ttasks and adjusted to lhe desired wa!er activity. Tlw. oxidation of all samples was followed by sl:ln(htrd manometric techniques." The loss of -~stneene l)igrnent of salmon was measured, using teehniqtles reported by Lusk, /.C-trel, and (.,oldblit' " n . '" 'the. extent of oxidation is reported as mieroliters of oxygen (STP) per vram of lipid. Some of the kinetic analyses ,ire based on ealeulalions of oxygen absorbed I)er gr:tm of the major reactive fatty acid, whicl, in all cases was linoleic acid. These (.:,Jc'uhttiot]s were made on the Ioasis of gas cl~romalogr'q)liic an:llysis of sample composition. in the case of systems, such as salmon

TABI, E 2

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()xi,lized lJh(.'tmlie. . . . . . . . . c~m~p(;ullds

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1

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Pr, Muets ~,f lipid oxidation

i

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[ (,ott-llavor) Brown polymers loss of solubilil,y loss of biological value of proteins

Oxidized ascorbic acid

290

M. KAREL, T. P. LABUZA, AND J'. ~F. MALONEY

steaks, in which secondary reactions result in production of carbon dioxide, this gas was measured I)3" gas chromatographic techniques,; and the oxygen nbsorption data vrere corrected accordingly. P~ESUI,TS

All of the systems oxidized at different water contents at 3T°C were adjusted (o the desired water contents by equilibration with

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suitable relative humidities. The air used for oxidation w'~s also maintained at the equilibrium humidilies. Tile adjustment was made on the bnsis of sorption isolherms. Sorption k.0therms TM for each of the systems were oht.,ined at (he temperature of the oxidation experiments. These isotherms are presonic(1 in Figure 1. The sorption data were also used to (h:termine tim relntire humidities and the moisture contents corresponding to monolayer values. This was done using the Brunnuer-Emmett-Teller method. ~ The monolaver v'~lues for each of the systems are shown in Figure 1. Oxidation studies on methyl ]inoleate-eellulose systems were carried out. a! ,)~ v.. with manometric techniques. Typical results with highly purified systems are shown in Figure 2. It is evident that rates of oxidation decrease with increasing water activity and that some inhibition of oxidation may be observed at moisture contents below the monolayer coverage (equilibrium relative humidit3', 199~ ; moisture content, 2.69~, dry basis). Similar results for the cellulose-linoleate systems conlaining 10 ppm (on linoleate basis) of cobalt are presented in Figure 3.

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EQUILIBRIUM RELATIVE HUMIDITY

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CHEI~JlICAL CHANGES IN FP, EEZE-DRIED FOODS

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It was observed that the metal had a strong catalytic effect on oxidation in the dry state at 37°C but that humidific:~tion to45% relative humidity completely eliminated the prooxidant activity of cobalt. ]~esults obtained with a model system conlaining egg alblunin and methyl ]inoleate are shown in Figure 4. The same general trend of effect of humidification is apparent, in spite of a diffcreut support system (protein instez,d of cellulose) and a higher oxidation temperature (55°C). In an attempt to apply conclusions based on model systems to actual foods, experiments

were conducted on freeze-dehydrated salmon steaks maintained in air at 3 7 " C and at controlled water contents. The oxidation extent of these samples (calculated as microliters of oxygen per gram of total lipid) is presented in Figure 5. The oxidation experiments were conducted on dry samples and on samples maintained at two different water activities, one below and one above the monoIayer value. As can be seen from the results, increasing the water content was cffecuve in inhibiting oxid:~tion of the lipid, especially when water content was increased above the monolayer value.

292

M. KAIREI,, q'. P I,ABUZA AND J. F. MAI,ON],~

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During the same oxidatiml experiments, analyses were made of the deterioration of astacene, a. pigment responsible for the pink color of sahnon. The results, shown in Figure 6, indie'~ted that the deterioration, which is of an oxidative nature, ~° is retarded considerably when lipid oxidation is minimized by increasing the waier content above the monolayer value. DISCUSSION

Analysis of "(he results of the oxidation experiments showed that the curves of oxygen absorbed vs. time showed n curve with the typical autocatalytic shape which results, from the kinetics of a free radical reaction.:' = The progress of ox-i&l.tion is characterized-by three distinct periods. These are: 1) an initial

inductitm period in the samples low in coneentration of enl:llysls (in the ease of results presented here, induction periods are either very short or nonexistent for samI)les conraining col)nlt or myoglobin): 2) a subsequent period dtlring which the hydroperoxides lhnt have been t'ormed decompose by a monomolecular reaction occurring either autocatalytically or, as prol'.osed bv Uri, -~° catalyzed by trace amounts of the heavy metal transition elements such as cobalt, copper, and iron; and ,3) a period during which the hydroperoxides decompose by a bimolecular reae'i tion. 'Ih.s occurs at high hydroperoxide concentrations, probably because of bimoleeular assoei':tion through hydrogen bonding. Existence of association is supported by the speetrophotometrie evidence of Bnteman* for

CHEMICAL II

CHANGES

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Fro. 7. Mouomolecular rate period. Run 12- effect of humidification on model system at

37°C. ethyl linoleale and by that of Wailing and I:[c:tton -"~ for t-bury] bydroperoxide. Semenov ~' concluded on the basis of thermodynamic considcrations that hydrogen bonding is a probaisle step in bimoleeular decomposition. The bimo/ccul, r reaction c,n ~/so occur in a metal-cqialyzed process.' Heavy metals catalyze both types of decomposition and increase the orer-~lll rate of oxidation. The rezlction rate constant for the bimolecular initiation is higher and the energy of activation lower than the corresponding quantities for the monomolceular decomposition. Conditions which promote the onset o.f himolecular decomposition, therefore, arc in effect-prooxidant, and those which retard the change from monomoleeular to bimolecular decomposition are in effect antioxidant.

When. the kinetic equations governing the oxid:ttion renciion are integrated, it is evident that, during the monomolecular period, the .-.'quare root of oxygen absorbed plotted vs. time gives a straight line.// The slope of the lines is an indication of the magnitude of tim initiation rate constant,. The steeper the slope, the faster the reaction. For instance, addition of catalysts increases this slope and de:lctir:ltion of these catalysts decreases it? A subscquent change in the slope indicates a change in kinetics, usually a change to bimolecular initiat ion. Figures 7 and 8 and the curves labeled "control" in Figure 9 show kinetic plots for lhe monomolecular decomposition for the cellulose system without added catalysts; Figure 9 also shows a plot for the cobMt-cellulose

294

~ M. k A l l g d3I , , T. P. I,ABUZA, AND ,i. F. MAI,ONEY l

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system; Fig,,re 10 for the high protein, high fal system; and Figure 11 for salmon. Tim results shown in Figures 7, S, and 9 show thai nddition of water in a system con-

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FIo. 9. Monomolccular rate period. Cellulose systcra. Run 9" humidification to 45% relative humidity. Cobalt catalyst added. 37°C.

CHEMICAL CHANGES IN FREEZE-DRIED FOODS

Figllre 11 shows that monomolecular initiations seem to predominate in both dry and

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FI,~. 11. Freoze-(h'ied snhnon. Run 2. monomolocul:tr rate plot. of water evident from data shown in Figures 2 and :~. Ill the presence of added cobalt, the predominant effe(,t of water is the deactivation of oobalt, :~s evidenced by the magnitude of the slOl)eS of (lie plots for the cobalt-catalyzed :.:,'~mples in 1he presence and in the absence of w:~ter (Iqu. 9). An increase in the level :~! which tho transition occurs is not clearly evident in tim cobalt-eatal.vzed samples, and the :tntioxidant. effect due to hydrogen bonding of h.vdroperoxides is prol)ably less im1)or~nt in ~his case than is metal inactivation. The progress of oxidation in the high protein, high fat system and in salmon is more complicated. In addition to oxygen absorption l)v fatty acids, there is some (lire(,t oxidation of proteiusf' TM and secondary reactions betwe.c~ oxidizing lipids an(1 proteins :t re' likely. Nevertimless, certain fe'ltures observed in the simple model system based on cellulose are also aplmrent in the high protein malerials. Himfi(lifieation to levels above the monolayer results in substnntial reduction in the oxidalton-ra:es (Fig. 4), and it, is apparent from llw kinetic plot for the monomoleeulnr rate period tlmt addition of waler extends this perio(l suhstantially :is compared v:ith samples ox{dized in the dry state (Ft,-. 10). In ~he case of salmon, wa~er addition to a level above the monolayer value affords consideral)le protection against oxid:ltion (Fig. 5). An inspection of lhe kinetics plot in

humidified samples throughout the oxidation experiment. This is probably due to the low ~bsolute ooneentration of hydroperoxides. The absolute concentration of the reactive fatty acid, linoleic, is low with respect to the total lipid. Furthermore, sitice most of the fatty acid in salmon is in mixed triglycerides, hydrogen bonding between hydroperoxides is probably prevented by sterie factors, even when there is no interference due to hydrogen bonding with water. The predominant effect of water in this ease is probably due to inqetivation of mclal catalysts, including the iron in the heine group of myoglobin which is present in salmon tissue. The protective effect of water is also evident in the oxidative deterioration of the pigment astacene. In this case, as is evident from the results presented in Figure 6, the protective effect becomes strong when humidification is carried out to water content above the monolayer value. It appears likely that the oxidation of ast'tcene is due to interaction with oxidation products of fatty ,acids: thus, the inhibition of fatty acid oxidation results in the observed protection of astncene. Water may also proteet the pigment by inactivation of metal catalysts which have :~ direct catalytic effect on astaeene deterioration. Svs~rAaY

Lipid oxidation is one of tile major reaction,a limiting the stability of freeze-dried foods. Water has an inhibitory effect on the reaction. Studies conducted in model systems eont:~ining celhflose, methyI linoleate, and cobalt resulted in c'onfirmation of the" h.vpothesis that the effects of water are due to: 1) hydrogen botlding of hydroperoxides with a resultant delay in the onset of the rapid bimoleeular decomposition of hydroperoxides and 2) inactivati~)n, through hydration, of metal eat:dysts, slmh as cobalt. Studies were also conducted on a more ('omplicaied model system consisting of egg :~lbumin, linoleate, and myoglobin, and on freeze-dried sahnon. The results obtained are in agreement with those observed in the simpler model systems.

M. K A l t E I , , T. ]'. LABUZA, A N D 5. F. M A L O N E Y

296

li EI,~EI{I+NCI~,S 1. l:latel,mn, 1.,. Oh,fin oxidnlion. Qunrl. Rex'. (l,ondon). ,~,': 1.t7-1(17, 1!)5"t. 2. Bolland, J. I,. Kinolies of oh.'lin oxidalion. (~tmrt. J{r,v. (l,t)ndon), 3: 1-2I, 19-19. 3. lialton, P., an~] leis('h+'r, F,. A. Stu(tio.- on the stor~ige of ",vh(,alon flours. II. The' ab.-.orption of oxygen })y lh)m' .,~/o)'ed trader diffm'(,nt condi(ions. Cm'o.ll Chore., l,}: 267-271, 1937. ,!. Ingoht, 1{7. U. In ]Apids nn+l tllt,ir oxiclalion, 11. Se}ltlllz, O(l., Clmpter 5. Avi Publishing Co,, Wos( port, Conn., 1.9t~2. ,5. Karel, M,, Issenborg, P., Ronsivalli. l,., anti Jurin, V. Apl)lieation of gas cbromnloKr'll)hy (o (,Iw moasuremenl, of gg)s l)erm(,:)l)ilily of l)a('kag, in~ m.tterials. I:ood Fcvl)nol., 17" 327330, 1963. 6. [(arc'l, M., ,+ald l,al)ttza, T. P. 32oc}l:t~lisz)l+<'+ oJ" deterioration and f<:)rmtthtlion of spttce (lio(s. M. 17. T. reports on oolltrtt('t, roso;tr('h w~th tht' U. S. A h' f o r 'c..:X.('r()si)a.,'<' M<,(li<'al Division, Con( racI..I 1-609-66-46, 1966. 7. Kart, l, M., and Nicker.-,on, J. T. 1~. Effoc(s of rolaiive hun,idity, air, and vttf, titllll O11 I)l'()Wllin.u of dehy(h'ated oi'ango juico. Foot| T<,<'hnol.. IS: 10,1-108, 1964. 8. Imbuza, T. P.. M.~loney, J. F., .tml .Karel, M. Autoxid'ttion of methyl tinoh, ate in freeze
•

of warm" on tho aut
,

T h e nuthm's wish to acknowledge the assist.qnce of Mr. F e r n a n d o ~ l a r t i n e z in some of the work presented in this I)aper.