Gelatinization Mechanism of STARCH GRANULES RICHARD S . BEAR1 AND EDWARD G. SAMSA Iowa State College, Ames, Iowa
The true nature of the gelatinization mechanism of starch granules is disclosed by observations on certain gas bubbles which develop in granules enlarging under the influence of concentrated electrolyte swelling agents. These bubbles change in volume remarkably, rising from zero volume to maximum size and then disappearing. In initial stages tangential enlargement of the expanding granule layers takes place more rapidly than fluid can penetrate through them, with the result that lowpressure cavities are developed in the inte-
rior. In time the infiltering water weakens the swelling layers, which are then on the periphery to form the sac wall and eventually present the familiar appearance of collapsed balloons. The increase in area of the swelling layers is very great, and this change is exceedingly vigorous ; therefore it seems likely that actual contraction and thickening of radially oriented SIarch molecules is responsible for the dilatation, rather than that the phenomenon is caused by ordinary osmotic or hydration effects alone.
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H E remarkable swelling power of starch granules is of considerable importance industrially, but probably few chemists have a clear conception of what occurs in each granule when starch is gelatinized. During a study of the swelling of individual starch granules, as induced by chemical agents and observed microscopically, it was noticed that a gas bubble frequently formed within a granule and changed remarkably in size during the granule dilatation. This paper describes quantitative observations which use such bubbles as indicators of changing conditions within the granules. Undoubtedly the bubbles have been observed many times in expanding starch granules. As early as 1849 Schleiden (IS) noted them under certain conditions. Many years later Zwikker (16) cited their appearance as an argument for a theory that the individual concentric layers of a granule enlarge primarily in a tangential direction, radial increase of the whole granule being only a secondary result (Figure 1). The gas bubbles should not be confused with the nuclear spaces (in which the bubbles swim) developed in dilating granules. These spaces, frequently illustrated in the literature, are normally occupied by fluid, but the cause for their development is probably the same one that gives rise to bubble formation. The fact that starch granules enlarge by tangential expansion, and not by development of internal radially directed pressure, has been known by various investigators for more than half a century. Recent reviews by Alsberg ( 1 ) and Badenhuizen ( 2 ) discuss the evidence and accept the conception of tangential swelling. However, it is doubtful whether the full significance of this view has yet been felt. This paper confirms the tangential nature of the swelling and also presents new ideas regarding the molecular alterations responsible. 1 Present address, Massachusetts Institute of Technology, Cambridge, Mass.
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The words “gelatinization” and “swelling” have taken on special significance in starch chemistry, and in the interest of clarity their usage in this paper will be explained. Ordinarily the former term is applied t o processes in which granules undergo many fold enlargements under the influence of appropriate temperature or chemical conditions; the latter word is often reserved for !he smaller volume Increases observed when dry granules are subjected to polar solvents at relatively lowtemperatures. This article does not discuss the collective action of a ,number of granules In forming a gel or viscous paste, but rather the events
Figure 1. Difference between Concepts of Radial and Tangential Swelling of Starch Granules A i s the original unswollen granule, with layered structure. B is an early stage of a granule supposed to be enlarging either by develo ment of internal pressure Posmotic), directedasshownbythearrow or by thickening of each layer: C is a granule whose layers have increased i n surface area through extension i n tangential directions, as shown by the arrows, with consequent development of a central space, 8 , which amounts for moat of the volume increase.
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Figure 2. Potato Starch Granules Swelling in 0.5 M Sodium Hydroxide Solution, with Relatively Large Bubbles Developing The particularly large granule in the center shows the phenomenon beet. The large black boundaries divide solution from air spaces, into which the solution was allowed to penetrate slowly to permit observation soon after contact of granule and fluid.
happening within individual granules during gelatinization. Because “swelling” is particularly convenient and suitable for the description of single-granule dilatation, it has seemed desirable to use this word here in the more general sense as denoting any type of granule enlargement whatever. Careful distinction between “gelatinization” and “swelling” becomes necessary, however, in referring to special expressions such as “heat of gelatinization” which is very different from the more common “heat of swelling”.
Expansion of Granules Under appropriate conditions, starch granules in the act of gelatinization develop bubbles of gaseous composition within their interiors. That these bubbles are not liquid phases is readily discerned under the microscope because of the marked refraction effects a t their surfaces. Their location within the granules is easily demonstrated through determination of their depths with respect to granule walls by focusing. Their internal position is also indicated by the observations that such bubbles are only associated with granules and never appear outside them, and that in many instances collapse of a granule wall during swelling results in a movement of the bubble within the granule without escape. Except in rare instances only one bubble is formed per granule, though all granules do not develop them. The experiments were performed chiefly with potato starch granules because of their large size and the ease and frequency with which bubble formation could be induced in the samples available. The phenomena to be described can, however, be observed with other granule types, such as the cereal starches. The following procedure was typical of the way the experiments were carried out: Granules sprinkled on a microscope slide were treated with approximately 2.0 M calcium nitrate solution, a cover slip was applied, and as quickly as possible the granules were brought into focus by observation through the side telescope of a camera attached to the microscope. Pictures were registered from time to time on roll film as swelling proceeded. Individual views were projected on a screen and the granules measured. Since potato granules are not spherical, major and minor diameters were recorded as well as diameters for the bubbles, which were usually round. On each roll of film the scale of a stage micrometer was photographed for calibration purposes. Observation of the granules between crossed Nicol prisms was done visually, since photographic registration with the
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reduced light available is too slow. Kith the same starch sample, observations of bubble formation are apt to vary somewhat from day to day, probably because of temperature and humidity inconstancy. Furthermore, small granules resist swelling agents to a greater degree than large ones. Using typical commercial potato starch samples, the concentration relations are a p t to be as follows: Dilatation proceeds smoothly without bubble formation in 1.5 M calcium nitrate and in 0.1 M sodium hydroxide. I n 2.0 M calcium nitrate the expansion is accompanied by the formation of small bubbles; in 0.5 M sodium hydroxide relatively enormous cavities filling much of the entire granule interiors are observable (Figure 2 ) . For the quantitative experiments described below the choice of swelling agent was 2.0 M calcium nitrate, since in this solution expansion proceeds a t a manageable rate and the bubbles are not se large as to be unspherical and distorted as they often are in sodium hydroxide swelling. The latter reagent was, however, useful in some of the polarized light experiments.
Origin of Bubbles Pictures of the same field of view a t different swelling times are presented in Figure 3. Figure 4 is a typical graph of the changes in volume with time, both for a whole granule and its included bubble. I n calculating granule volume, the potato granules were taken to be prolate spheroids with a volume of (4/3) 7r ab2, where a and b are major and minor semiaxes, respectively. I n Figure 4 ordinates for granule volume are given in units 100 times as large as those for the bubble volume. The consecutive pictures and the graph show clearly that the gas bubbles undergo remarkable alterations in volume during the gelatinization process. With proper interpretation of their origin, their behavior should furnish valuable clues to conditions existing within the granules a t various moments during dilatation. The several possible interpretations regarding the bubbles can be divided into two groups on the basis of what is assumed concerning the ability of the granule wall to shield the interior from the constant exterior pressure of one atmosphere. If the wall is strong and compact during initial stages of granule expansion, the size of the bubble will be a function of amount of gas available and the pressure in the interior. If the wall is permeable or weak, the bubble volume will depend only on the quantity of gas, since the pressure within the granule 15 ill remain essentially constant a t one atmosphere. Sources for gas (at atmospheric pressure) are (a) air entrapped in fissures or other interstices of the original unswollen granule, or (b) gases dissolved in the solution and liberated as it passes into the granule. It is clear from Figure 4 that, in the case of this typical granule which underwent only moderately extensive bubble formation, the initial bubble volume is negligible compared to that developed as swelling progresses. Therefore, assuming atmospheric pressure a t all times, one mould be compelled to look for considerable evolution of gas during the process of enlargement. Evolution of gas might be brought about either by actual chemical production (from impurities) of a gas such as carbon dioxide or ammonia, or by decrease of solubility of oxygen or nitrogen in the swelling solution, caused by local temperature elevations due to heat liberated during the granule hydration or swelling process. Under the conditions of the present experiments the chemical evolution is improbable, since the effects were essentially the same either in the acid solution of calcium nitrate or in the basic solution of sodium hydroxide, and the bubbles always finally disappeared into the same solution in which they were produced. According to Mullen and Pacsu (11) the gelatinization of various starches in water and in aqueous pyridine solutions
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is a n endothermic process; this might be expected from the facts that most hydrated carbohydrates exhibit negative heats of solution, that gelatinization is greatly favored by increase of temperature, and that retrogradation (essentially the reverse of gelatinization) is brought about by decrease of temperature. Under conditions of swelling in calcium nitrate and sodium hydroxide solutions, however, qualitative tests have shown that moderate heat is produced, so that the possibility of gas evolution due to this cause has to be considered. It seems unlikely that the heating explanation is valid for the following reasons: Calculation indicates that the total amount of air dissolved in a volume of water equal to t h a t of the granule a t the time of maximum bubble development is not quite sufficient to account for bubble volume in the case of moderate development illustrated in Figure 4. The difficulty is even greater when relatively large bubbles are produced by concentrated swelling agents. I n addition, undoubtedly less gas is dissolved in the concentrated solutions employed, the temperature required t o release all the gas would seem difficult t o secure and maintain locally for the appreciable times observed, and it is difficult to understand why the bubble should disappear during the period of maximum swelling rate. Finally, it has been found that wellaerated calcium nitrate solutions and solutions which have been boiled to remove dissolved gases do not differ appreciably in their ability to induce bubble formation. If we assume a definite lowering of internal pressure, the phenomenon is more easily understood. The increase in bubble size is so great that, even before the most rapid rise (Figure 4) in bubble volume sets in, the pressure has un-
Figure 3.
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doubtedly been lowered to equal the aqueous tension of the solution. After this occurs, the increase in volume proceeds with water vaporizing to fill the cavity developed by the tangentially swelling granule. At first it may appear unbelievable t h a t an expandjng starch granule can develop and maintain for appreciable periods a pressure difference equal t o most of a n atmosphere between its interior and exterior. When it is noted that the phenomenon of limited swelling (dry granules in cold water) can develop pressures of the order of thousands of atmospheres ( l a ) , and that increase in bubble volume occurs only during early stages of granule dilatation (at 5 fold granule volume increase or a t 1.7 fold development of granule radius the bubble of Figure 4 reached a maximum size), the phenomenon does not seem too strange. Actually, in many instances while expansion is still progressing rapidly, the granule wall becomes weak a t local regions and proceeds to bend inward visibly, whereupon the bubble fades and disappears completely. The observation of wall collapse with accompanying bubble disappearance and the fact that bubbles always eventually vanish constitute the best direct evidence for the existence of internal low pressure and the supposition that most of the bubble content consists of water vapor. With the conception that the bubbles are low-pressure cavities, the volume of the bubble is most simply regarded as a measure of the lag of internal adjustments behind the essential enlargement process. The bubble volume is equal to the difference between the space enclosed by the tangentially expanding starch layers and the volume occupied by the fluid which has been able to filter from the outside through the developing layers.
Stages i n Granules Swelling, Included in Constant Field of View, with 2.0 M Calcium Nitrate Solution
Time progresses from a tof, with 30-second intervals between each.
Small black circles are gas bubbles; gray crements show regions of collapse.
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Events during Granule Dilatation With the bubbles interpreted as indicating lag of internal pressure adjustments, the stages occurring during chemically induced gelatinixation of starch can be cited. These statements summarize the observations of a large number of granules during enlargement, both from photographs and by visual study between crossed Nicol prisms. Reference to Figure 3 can be made for such phenomena as are readily reproduced: 1. At the outset the granule has the typical appearance: striations are frequently visible about the hilum as an eccentric center, the hilum often being marked by slight irregularities in its neighborhood. The polarization cross (positive) has its center at the hilum. 2. Initial expansion takes place tangentially rather than radially at all points in the granule. This condition tends to increase the periphery before fluid has been able to penetrate into the hilum or center. Pregumably small amounts of gas (air) or water vapor may be present in fissures of the granule, and these serve as nuclei for the development of the bubble observed. Swelling may proceed noticeably, however, before the bubble appears, and it is always found initially in the region of the hilum. 3. As dilatation progresses, the peripheral area is increased still further while the structure remains sufficiently intact to impede the flow of fluid to the interior from outside. Consequently the bubble continues to grom in volume, expanding at constant pressure equal to the aqueous vapor tension. During this period the birefringence of the thick peripheral swelling region is slowly reduced but remains readily visible; this confirms the fact that oriented structure persists. That the structure a t this stage still possesses considerable strength is shown by its ability to resist collapse under a considerable pressure difference between the exterior and the interior of the granule (almost a whole atmosphere). 4. In time the water beinq forced inward through the expanding peripheral layers weakens them by dissolving or maximally smelling their constituents. Influx of fluid now becomes more rapid than the enlargement demands. The bubble is then free to contract and proceeds to diminish in size until it disappears completely. During this period the double refraction becomes vanishingly small but never becomes negative, as it might be expected to do if the volume increase were caused purely by typical osmotic forces acting to distend a membrane. (The tension would produce double-r efraction positive with respect to the direction of tension, but negative with respect to granule radius.) Neither the swelling nor the final weakening of the swelling layers occurs, in general, uniformly throughout. In many instances the low internal pressure causes collapse of the wall at localized regions where weakening is evidenced by disappearance of double refraction, while vigorous expansion is yet in progress in other places whose birefringence is still visible. Concomitant n ith the caving in of the wall, the bubble rapidly fades away. 5 . Under the conditions of these experiments the granule continues to swell, though less rapidly, even after the bubble disappears. There is no need to regard this stage as any different from preceding ones, since it represents a situation in which iiiflux of water keeps pace with granule enlargement. The final completely quiescent granule presents the familiar picture of a wrinkled sack or a deflated balloon; according to the present view, this appearance results from localized collapse of the wall to relieve internal low pressure, rather than from shriveling brought about by loss of dissolved material from the granule. The final boundary material of a swollen granule should not be regarded as a true osmotic membrane. I t would seem to be a porous network of the more insoluble material drawn from the complete original granule rather than from any particular region, such as the external surface.
In addition to the above observations the following are also of interest: 6 . In many instances the accidental movement of a granule during swelling results in a displacement of the position of the bubble. This displacement happens in particular when a portion of the granule wall invaginates near the bubble. In such cases the bubble appears to move freely about within the granule; thus the central region, which appears isotropic in polarized light, is clearly a fluid. 7 . A granule swelling in sodium hydroxide often presents particularly well the phenomena under investigation, though photographic registration is difficult. Thus, even in 0.1 M sodium
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hydroxide, while a bubble is not developed, there is slight but sufficient difference between the refractive indices of the swelling layers and of the isotropic ctnter to distinguish visually the smooth boundary separating these two regions. Betwecn crossed Xicol prisms the swelling layers light up while the center remains dark. The slow decrease in double refraction can be followed to very small values by a technique found useful with weakly birefringent biological tissues (5)-namely, to insert in t,he tube slot of the microscope a Kohler l / ? ~wave-length rotating mica compensator. When the compensator is turned slightly either way from the dark-ficld position, alternate quadrants of the granule wall are, respectively, brighter or darker than the field; thus observation of sign and approximate magnitude of retardation is possible long after observation of a distinct polarization cross becomes difficult. 8. When swelling becomes too rapid, as in 0.5 J4 sodium hydroxide, a bubble may become very large (Figure 2) and sometimes irregular in shape, and may fill t,he central cavity of the granule almost completely. In such instances the swelling layers seem to pass fluid to the interior only reluctantly; and the bubble may remain for long periods, even after penetration of the granule by fluid has reduced its refractive index nearly t,o thc point of invisibility. The apparently isolated bubble of this t,ype then slowly but completely vanishes.
Interpretation in Terms of Granule Structure The experiments leave little room for doubt of the essential correctness of the tangential nature of starch granule swelling. Alsberg ( 1 ) favored this description, and the present observations support his general point of view. Since Alsberg discussed the question a t some length, we need only emphasize that his statements regarding radially oriented starch chains (or micelles), tangential thickening of these chains during swelling, the origin of the swollen granule sacks, and the absence of a preformed membrane, seem essentially correct. The exact nature of the thickening process remains to be determined. Recent discussions of granule structure have tended to emphasize either t'he starch chain as the important unit ( 1 , 7 ) or the knitting of these chains into interlocked micelles ( I O ) . While all have reached similar structural conclusioiis in spite of different emphasis, the view of Meyer and Bernfeld ( I O ) seems to neglect the t'angential type of smelling. A swollen granule is not a net'work of interlocked resistant' material except in the wall regions resulting from the tangential swelling. Ordinary osmotic effects through a semipermeable membrane are secondary to the aspiration effects which follow tangential expansion and cause fluid to enter the granule. The recognition of the true nature of starch gelatinization leads to a realization that the final volume attained by each granule is very dependent on swelling conditions. Thus, if a considerable portion of the relief of the internal reduced pressure is accomplished by caving in of the walls, the total fluid taken up will be sharply reduced. Slow swelling, which allows fluid to enter wit,hout much interior pressure reduction, will permit a maximum of fluid entry. A more constant result of complete granule swelling should be t'he surface developed. Estimates of the surface areas of a number of the granules measured during swelling in the above experiments indicat'e that the surface increases ten to sixteen fold. This is a conservative estimate of the power of potato starch granules to expand, as Furry's results (8) show for the same granules thermally enlarged. However, the rigid structure capable of withstanding almost an atmosphere of pressure difference between exterior and interior is maintained only up to about a twofold area increase (the point a t which the bubble begins to fade) under the conditions of the swelling in 2.0 A4 calcium nitrate; i t may perhaps he maintained farther in sodium hydroxide. The detailed description of a structure which is capable of such extensive lateral swelling requires a certain amount of imagination a t present. One might suppose, with Meyer and
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Bernfeld (IO), that small pockets within the swelling layers become filled with starch solution and are distended osmoticaliy ; expansion thus occurs most easily in tangential directions because of the radial orientation of fissures developed between chains or micelles. It is difficult t o believe that the relatively dilute solutions of starch of high molecular weight could successfully compete with the concentrated calcium nitrate solution to secure water rapidly enough for the vigorous expansion observed. If, as Alsberg ( 1 ) supposed, chains or micelles of more or less cylindrical shape take upon their surfaces sufficient water to cause a t least ten to twenty fold increases in cross-sectional area, then one must imagine greater than three to four fold increases in their diameters. One is tempted to conclude that the striking swelling manifestations of starch granules are a result of an active tendency of starch chains to coil or otherwise contract, and thus shorten in a radial direction and expand tangentially as they are placed in the granule. Only in such fashion is it easy to visualize the remarkable increases in cross-sectional area. Alsberg came to somewhat opposite conclusions. He regarded hydrated chains as fully extended and dehydrated ones as contracted, t o the extent allowed by granule structure, in order to explain the remarkable contraction of the diameters of dried granules which may amount to over 15 per cent. It would seem possible to suppose that in either extreme-of dehydration or of chemical or temperature swelling-the chains are more or less free to indulge in a natural tendency to shorten because certain critical cross linkages (hydrogen bridges?) have been broken. Precise evaluation of this hypothesis is difficult a t present. While the progressive radial thinning of the swelling layers during tangential expansion and the decrease of double refraction are in agreement with it, these observations could have other explanations, such as a deterioration in amount and organization of the starch brought about by the rush of water through the layer. It should be emphasized that the thinness of the final sack wall is not produced by pressure within the granule, but that the motivating forces come from within the original peripheral swelling layers themselves. The ability of the layers to remain intact during these alterations undoubtedly arises from the ramifications and interlacings of starch chains so frequently cited by Meyer and others. Acid solubilization destroys these links, and swelling phenomena are then abolished. While the suggestion of actively contracting starch chains indicates that starch molecules in freshly prepared solutions are in a contracted state, as viscosity studies (14) show they may be, it is not necessary to conclude that this is the equilibrium state under ordinary conditions. It is possible that at room temperature starch chains dissolved in nonswelling aqueous solutions may slowly become maximally extended (though perhaps never so fully as in cellulose, 6); the fullest possible hydration is thus possible. Recent x-ray diffraction studies (4) suggest that starch granules contain an organization of chains into crystalline regions whose structural repeating units (unit cells) resemble in dimensions those of cellulose and maltose; hence they do not call to mind a contracted configuration. Identical crystalline modifications are readily obtained by retrogradation from aqueous starch solutions at ordinary temperatures, and the reversion from the contracted state to an extended one may be an important phase of the retrogradation process. The picture of the molecular mechanism of swelling here presented is in some respects a denial that hydration of starch is the essential process in gelatinization, although considerable rearrangement and hydration of starch chains may take place. It is worth noting that the protein collagen in fibrous form (animal connective tissue, tendon, etc.) undergoes
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I
Figure 4. Typical Graph of Volume Changes in a Potato Starch Granule and Its Included Bubble during Swelling in 2.0 M Calcium Nitrate Solid circles apply to granule volumes, with the ordinate i n oubic centimeters X 107% open circles apply to bubble volume, with the ordinate in cubic centimeters X 10'.
extensive contraction under the influence of temperature increase or chemical treatment, much as do starch chains. Kuntzel and Prakke ( 9 ) concluded that the thermal and chemical shortening of tendon is not a hydration phenomenon, as visible appearances suggest, but rather depends primarily on an initial dehydration. Their paper contains arguments which might well be transferred to the case of starch gelatinization. The reader is also referred to a recent paper (3)describing relations between current concepts of contracted or helical models for starch chains, the iodine-staining ability of starch, the production of Schardinger dextrins, and the crystalline V (alcohol-precipitated) modification of starch. All of these relations add considerable credence to the view that the gelatinization process involves primarily a vigorous contraction of chains which in the granule were originally extended, and that retrogradation is a reversal of this change. While the experiments described in this paper bring little or no direct light to the amylose-amylopectin conception of starch, it has been suggested t o the writers by Eugene Pacsu, of Princeton University, that the straight-chain retrogradable amylose is probably the primary fraction involved in the molecular contraction process; the latter process is here postulated as chiefly responsible for gelatinization, and its reversal constitutes retrogradation. The nonretrogradable amylopectin fraction has been widely considered the chief constituent of the final insoluble residue forming the deflated sacs of the completely swollen granules; its branched-chain structure is not so suitable for extensive changes in molecular shape, except in so far as longer branches may be involved.
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The heats of gelatinization measured in Pacsu’s laboratory (11) are also of interest in connection with the present hypothesis. Since they are negative and of a magnitude consistent with the breaking of one or two hydrogen bridges per glucose residue, these measurements can be considered to agree with the conception that hydration is not the significant process in gelatinization; if it were, heat should be evolved. The problem is, however, not simple, since it is possible that the observed heat of gelatinization is the net effect of endothermic and exothermic processes involving the disruption and reconstitution of more than two hydrogen bridges per residue. Even if as many of these bridges exist after gelatinization as before, there may yet be differences in heat content arising from the nonequivalence of the hydroxyl groups involved.
Literature Cited (1) Alsherg, C. L., Plant Physio’., 13, 295 (1938). (2) Badenhuiaen, N. P., Rec. trav. botan. nEerZancl., 35,559 (1938).
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(3) Bear, R. S.,J . Am. Chem. Soc., 64,1388 (1942). (4) Bear, R. S.,and French, D., Ibid., 63, 2298 (1941). (5) Bear, R. S.,and Schmitt, F. O., J . CelZular Comp. Physiol., 9, 289 (1937). (6) Caesar, G. V., and Gushing, M. L., J . Phys. Chem., 45, 776 (1941). (7) Frey-Wyssling, A., Naturwissenschuften, 28, 78 (1940). (8) Furry, M . S.,U. S.Dept. Agr., Tech. Bull. 284 (1932). (9) Kuntael, A., and Prakke, F., Biochem. Z . , 267, 243 (1933). (10) hleyer, K. H., and Bernfeld, P., Helv. Chim. Acta, 23, 890 (1940). (11) Mullen, J. W., and Pacsu, Eugene, IKD,ENG.CHEX., 34, 807 (1942). (12) Samec, M.,“Kolloidchemie der StBrke”, p . 131, Dresden, Theodor Steinkopff, 1927. (13) Schleiden, J. M., “Principles of Scientific Botany”, t r . by Lankester, p. 12, London, Longmans, Green and Co., 1849. (14) Staudinger, H., and Eilers, H., Ber., A69, 819 (1936). (15) Zwikker, J. J. L., Rec. trav. botan. nderland., 18, 1 (1921). JOURNALPaper 5-921, Iowa Agricultural Experiment Station. Project 638. supported in part by a grant from Corn Industries Research Foundation.
Water Adsor tion By
Animal Glue CHARLES ill. MASON1 AND HERBERT E. SILCOX University of New Hampshire, Durham, N. H.
The adsorption of water on thin films of five animal glues and one gelatin have been investigated, the glues covering the range of physical properties commonly employed. Both adsorption and equilibrium moisture content have been studied. The former obeys Freundlich’s law, and the latter is found to be greater than represented by the moisture adsorbed on the surface. It is proposed that water is held by glue in two forms-by true adsorption on the glue surface and by some other mechanism in the voids of the glue itself.
A
S I D E from general interest in the field of colloidal science a knowledge of the water adsorption of animal glue is of great interest to all who use this material in manufacturing processes. Previous publications have been scanty and incomplete. Wilson and Fuwa (11) give some data on glue as part of an extensive study of many materials. Katz (4) and Sheppard, Houck, and Dittmar (7) studied gelatin, a similar material. The present investigation was undertaken to supply data for glues covering as wide a range of physical properties as possible. A t the same time a n investigation has been made of the adsorption of water on glue in relation to Freundlich’s law. 1
Present address, Tennessee Valley Authurity. Wilson Dam, Ala.
ADSORPTIOR OV T H I h FILMS
One bone glue, four hide glues, and a gelatin were chosen which seemed to cover the range of known physical properties of glues. The glues were obtained through the courtesy of a large industrial user who selected those most characteristic of available commercial glues. The gelatin was a sample sold as suitable for bacteriological work and was therefore supposed to be of unusual purity. No attempt was made to purify the samples further because this would have vitiated the results from a practical viewpoint. Table I gives physical characteristics as determined b y the firm which supplied the glues. HTJ?IIDITY CoxDIrIoxs. The basic problem involved in the pxperimental technique is tu obtain reproducible and controllable humidity and temperatures over long periods. Air was passed over saturated salt solutions in saturators of the type designed by Bichowsky and Storch ( 1 ) and then through a chamber containing the glue samples. The whole train was immersed in a water thermostat regulated t o 25 * 0.02’ C. The humidity produced by these salt solutions had previously been determined by extensive calibration at 25” C. The exposure of the glue samples to the humidity conditions was carried out in the form of thin films 0.008 t o 0.01 inch (0.2 to 0.25 mm.) thick. The films were prepared by first mixing equal parts of glue and water and alloving the mixture to stand for 2 hours. Water was then added t o the swollen product and the mixture heated t o 40” C. on the steam bath. When liquefied, the glue was drawn into films by the method of Kallender (S), using moistureproof regenerated cellulose (cellophane) as a supporting medium. The films were then allomed to cool and dry partially in the air. The glue n a s peeled from the support and cut into strips 1 inch vide and 15 or 18 inches long. They were rolled into loose cylinders and placed edgehise in glass weighing bottles, In the case of the glues of lomcr gel strength, which tended to melt down at high humidities, the films w-ere drawn on glass cloth instead of moistureproof regenerated cellulose. This
