INDUSTRIAL AND ENGINEERING CHEMISTRY
104
was consumed. Only 10% of the men took helpings of the darkened fruit, and 25% of that taken was discarded in the garbage. Similar laboratory tests were carried out on light and dark fruit of various sulfur levels. Here again normal appearing fruit xi-ith a moderately high SO2 level was preferred to lightly sulfured fruit that had darkened considerably during storage a t 49" C. From the edibility tests it may be concluded that apricots containing as much as 6500 p.p.m. of sulfur dioxide prior t o cooking are in no way objectionable to most persons. CONCLUSIONS
1. The influence of moisture on the rate of deterioration of apricots a t moisture contents greater than 10% is dependent upon the quantity of oxygen available to the fruit. Under anaerobic conditions the rate is decreased by increasing the moisture content. As the quantity of oxygen is increased, the beneficial effect of high moisture becomes progressively smaller; in the presence of very large amounts of oxygen the rate of deterioration may actually increase with moisture. 2 . Increasing the moisture content from 10 t o 25y0 causes a 15-30% increase in the storage life of apricots kept a t 49 ' C. i n an oxygen-free atmosphere. The effect of moisture a t 36.7" C. is somewhat smaller. 3. The influence of moisture is nearly independent of sulfur dioxide concentration when the latter is expressed on a dry weight basis. 4. Deterioration appears to be slower a t moisture contents below lyOthan a t 25%, but such low moisture contents are difficult to obtain without scorching the fruit. 5 . The storage life of apricots is directly proportional to the
WATE
Vol. 38, No. I
initial SO2 level, a t least over the range 1500 to 8000 p.p.ni. suifur dioxide. Higher SO2 levels (up t o 25,000 p.p.m.) cause a further increase in storage life, but the rate of increase gradually declines. The effect of sulfur dioxide on the percentage increase ~ I L storage life of apricots stored a t eit,her 36.7" or 49' C. is almost the same; the life is approximately doubled by increasing the SO,level from 2000 t o 7000 p.p.m. 6. Although sulfur dioxide retards deterioration, it, does not prevent it. Apricots can deteriorat,e to the point of inedibllitj even though the SO2 level never falls below 5000 p.p.m. 7. During storage at moderate or high temperatures, the SO2 content of apricots steadily declines until approximatelg 657, of t,he initial sulfur dioxide is lost by the time the fruit has reached the limit of edibility. The effect is independent of [E&ture and temperature between 36.7' and 49' 6. 8. Palatability test's indicate t h a t apricots containing as much as 6500 p.p.m. sulfur dioxide prior to cooking are not, ohjectionable to most people. LITERATURE CITED
(1) Nichols, P. F., Mrak, E. M.,a i d Bethel, R.,Food Research, 4 , 67-74 (1939). (2) Nichols, P. F., and Reed, H. M., IXD. ENG.CHEM.,ANAL.Ea ,, 4, 79-84 (1932). (3) Xichols, P. F., and Reed, H. M., Western Canner & Packer, 23, 5 , 11-13 (1931): Fruit Products J., 22, 206-8, 247-9 (1943) (4) Sorber, D. G., Ibid., 23, 8, 234-7 (1914). PRESEXTED on t h e program of t h e Division of Agricultural and Food Chemistry of the 1945 Meeting-in-Print, AXERIC.ASCHEMICAL SOCIETY.A report on a joint research project of the Qusrtermast,er General's Office, L , B .4rmy and University of California.
SISTANCE
Improvement through Chemical Modification HAROLD S. OLCOTT AND HEINZ FRAENICEL-CQNRAT Western Regional Research Laboratory, U . S. Department of Agriculture, Albany, CaliJ. Proteins were treated with a number of organic reagents and the products examined for water resistance by measurement of the uptake of water by pressed disks. Aryl and long-chain alkyl isocyanates and also aromatic acid anhydrides and chlorides proved most effective. A number of proteins yielded phenyl isocyanate derivatives showing 24-hour water absorption of 1 to 2Yo. Phthalic anhydride gave products of low w-ater absorption w-ith egg white and cattle hoof. Protein derivatives of low water absorption showed a tendency to plastic flow without the addition of water a s plasticizer.
T
HE use of proteins in plastics is seriously restricted by their
poor n-ater resistance ( 2 ) . Obviously the first step to render these materials more valuable for such use would be a modification to improve this property. If this could be accomplished, the next step would be modification t o render them compatible with water-repellent plasticizers. Possibly both ends could be attained by one reaction. The present paper, however, is concerned primarily with experiments designed t o produce protein derivatives with reduced affinity for water, through chemical reaction. Proteins n.ere treated with a number of substances known or believed to react with the various typeS of polar groups. I n many cases the extent of chemical modification was ascertained analytically. The modified proteins were then examined for evidence of a successful reaction by a water absorption technique. Disks pressed from the products were immersed in water for 24 hours, and the increase in weight was used as a quantitative
measure of their affinity for water. This method of approarlt m EL^ patterned after t h a t described by Brother and McKinney ,3j. Of a considerable number of reagents tested, organic m c j a nates and aromatic anhydrides produced. the most pronourised changes in the nature of several proteins. The best p r o d w t s flowed to translucent disks in the absence of water, and t h e 24hour water absorption of the disks was only 1 to 37,. Th'. q plicability of these derivatives in plastics is receiving f t i tliec study.
THE proteins used for most of the experiments were te.:ttril& products similar to those t,hat could be made available for iriiiustrial utilization. Hoof powder was prepared from catt'le hoof' which had been dried at 70" C., by grinding in a hammer niiil.. Feathers and hog hair were ext'racted with benzene and theri. ground in a lt7iley mill to pass a 60-mesh screen. The technical. egg white, gluten, acid casein, and zein were commercial sarnpies Peanut protein was supplied by the Southern Regional Resear 9;. Laboratory. Plastic disks were prepared by the following met'hod: The proteins or protein derivatives were conditioned in a n ~7::e~; ai; 50 C. for one day, a t which time the water contents ranged from 0.6 t o 2.6y0 for different preparations. Ten per cent water wa~i added to those that required it as a plasticizer immediataiy b e fore the pressing operation. A circular positive die 1 id^ Ir; diameter was held in the press until the temperature rearheu. 300" F. (149' C.). Two grams of sample were quickiy iiitroduced and pressed at 5 tons of total pressure for 6 minutes, at which t,ime the platens were usually at about 320" F. (160' C.) O
January, 1946
1os
I N D U S T R I A L AND E N G I N E E R I N G C'HEMISTRY
The die was cooled under pressure t o about 150" F. (66" C.) before the sample was removed. The disks, which were about I/* inch thick, were examined for translucency and surface smoothness as indications of plastic flow. Water absorption was determined by a modification of A.S.T.M. specification D45-33'(3), The disks were weighed, immersed in water at room temperature, and held there for 24 hours; at the end of that time they were wiped dry and reweighed. The gain in weight, expressed as percentage of the original disk weight, is referred t o as %-hour water absorption. Many reagents did not lower the water absorption of egg white and hoof powder, or produced insufficient changes t o encourage immediate further study (water absorption,greater than 10%). Among these were formaldehyde, acetaldehyde, glyoxal, ketenes, aliphatic acid anhydrides and acid chlorides, ethylene and propylene oxides, epichlorohydrin, nitrous acid, hydrogen peroxide, dimethyl sulfate, diazomethane, and diazobutane. The conditions for the reactions were those generally used with these reagents. Products obtained from cattle hoof by reaction with quinone and potassium dichromate had water absorptions of approximately 5%. They had the disadvantage of being dark colored; in addition, the reagents appeared to be continually leached during the attempted removal of the excess by washing procedures. The methods used in reacting proteins with phenyl isocyanate were described in detail elsewhere (6). The following procedures will serve as a n example: A sample of the protein was carefully dried. To the dry product were added a n equal amount of phenyl isocyanate and a 2.5 fold amount of dry pyridine. The mixture was heated at 70" C. for 1 t o 2 days. Solution was not effected. D r y toluene was added, and the protein derivative was separated by filtration or centrifugation, washed once with ethanol, and subsequently extracted with ethanol for 24 hours in a Soxhlet extractor t o remove traces of diphenylurea, a by.product of the reaction. The product was finally dried with ether and then in a n oven a t 50' C. The yields varied from 120 t o 150% of the weight of the protein used. Water absorptions ranged from 1 t o 5% (Table I). The properties of unextracted samples, retaining small amounts of diphenylurea, were not appreciably different. With some proteins, special pretreatment was needed to obtain preparations which would react extensively with phenyl isocyanate. Commercial egg white had t o be dialyzed from a solution at about p H 5 (6). The gluten, gliadin, and casein derivatives of lowest water absorption were obtained with samples of the proteins which had been lyophilized from 1 N acetic acid solution. Numerous other aromatic isocyanates were also prepared and reacted with proteins. They included p-biphenyl, o-biphenyl, obicyclohexyl, b-anthryl, and p-cetyl phenyl isocyanates, and t h e bi-isocyanates prepared from benzidine and p-phenylene diamine (6). I n no case were derivatives obtained with water absorption properties superior t o those of the proteins treated with phenyl isocyanate. Aliphatic isocyanates were synthesized according to the method described for undecyl isocyanate (1) from fatty acid chlorides and sodium azide. The conditions used for the reaction with proteins were the same as those described for phenyl isocyanate except that the protein-reagent ratio was 1 t o 2. T h e water absorptions ranged from 1.4 t o 3'% for egg white and keratins (Table I). The method used for reacting proteins with aromatic anhydrides can be illustrated by the following example: T o 250 grams of commercial egg white (as received) were added 500 grams of phthalic anhydride and 1250 ml. of technical pyridine. T h e mixture was held at 70 C. for 2 days, cooled, washed once with ether and six times with methanol, and air,dried. The product weighed 200 grams and had EL water absorption of 1.8%. The conditions described for the reaction were not critical.
TABLE I. ABSORPTION OF WATERDURING 24 HOURS BY DISKS PRESSED FROM PROTEINS AND PROTEIN DERIVATIVES Water Absorption, % ' of Weight of Disk HeptaBep- BenNo Phenyl Undecyl decyl Phthalic ZOIC 80 1 treatiso1so180anhyan?oh&Proteina ment cyanate cyanate cyanate dride dri e ride Cattlehoof 28a 1.7 2.0 3.0 4.4 4.6b 3.5 Feathers 20 1.0 1.8 Hog hair 36 4.0 41 ... Egg white 506 1.1 1.4 1.5 2.1 4.9 2.0 Gluten 75 2.6 22 30 10 Casein 676 1.6 14 10 3.8 Zein 17 1.0 13 Peanut 39b 4.8 a Egg white, gluten, and casein were pretreated as described in the text. b 10% water was added immediately before pressing. 7 -
...
... ...
... ... ... ... ...
...
...
...
... ...
... ... ...
...
When equal amounts of egg white and phthalic anhydride were used, the product had a slightly higher water absorption (about 3%). Longer reaction periods or higher temperatures resulted in somewhat lower water absorptions, b u t t h e products were darker i n color. Benzoic acid anhydride was. used i n similar fashion. Both aromatic acid chlorides gave dark products unless the precaution was taken of cooling the pyridine-protein mixture t o approximately 0 O C. before adding the reagent. Water absorptions are recorded i n Table I. Studies of the protein groups participating in the various reactions were faciliated by the availability of a dye technique for measuring the number of residual acid and basic groups ( 5 ) . Aromatic isocyanates were previously shown t o react with almost all basic and most of the acid groups of proteins (6). B y the same methods it was found that aromatic anhydrides also react with all of the basic groups-that is, the amino, imidagole, and guanidyl residues. They also decreased the acid groups, apparently through acylation of the phenol groups as well as through formation of anhydrides involving the carboxyl groups. However, when phthalic anhydride was used, new acid groups were introduced ihto the protein, leading t o increases in the total number of acid groups by 10 t o 30% over those originally present. Representative data of the effect of aromatic acylating agents on the acid and basic groups of egg white are given in Table 11. OF AROMATIC ACYLATING AGENTSON ACID TABLE 11. EFFECT AND
BASICGROUPSOF COMMERCIAL EGGWHITE
Equivalents per 10' Grams Protein" Acid Basic 11.3 8.4 None Benzoic anhydride 4.7 1.0 Benzo 1 chloride 1.5 0.0 Phthagp anhydride 15.0 0.5 Phenyl isocyanate 1.4 , 0.1 Average of two or more experiments with each reagent. Reagent
a
Although the above experiments were undertaken primarily t o study the water resistance of protein derivatiGes, some observations on their plastic flow are also of interest. It was found that the original proteins, with the exceptions of gluten and zein, required 10% of water as plasticizer in order t o obtain suitable disks for the water absorption measurements (Table I). Op the other hand, the phenyl isocyanate and most of the phthalic anhydride derivatives flowed t o homogeneous disks without added water. I n preliminary tests phenyl isocyanate and phthalic anhydride derivatives of egg white were examined for compatibility with some of the commoner synthetic resins. The protein derivative and resin were ground together in mortar and pestle and then tested for flow properties in a Tinius-Olsen flow tester. The phenyl isocyanate products were not compatible with the urea-formaldehyde, phenol-formaldehyde, or melamine prepara-
106
INDUSTRIAL AND ENGINEERING CHEMISTRY
tions used. On the other hand, the phthalic anhydride derivatives gave homogeneous, translucent products i n combination with these resins. It will be noted that products of low water absorp,tion were obtained from some proteins and reagents and not from others. I n the authors’ opinion, the unsuccessful experiments do not necessarily mean t h a t the materials used cannot be made to react. It appears possible that a thorough investigation of methods of pretreatment and treatment might lead to products of low water absorption from any protein by reaction with any of the reagents listed in Table I. Such products were obtained with all proteins reacted with phenyl isocyanate, which was the only reagent investigated in detail (6).
THE affinity of proteins for water, as measured in the present study, appears to be associated primarily with the polar groups of the amino acid side chains. When these are blocked by the introduction of hydrophobic groups, the water absorption is considerably decreased. It was shown else-ivhere that phenyl isocyanate reacts with most of the amino, guanidyl, imidazole, amide, carboxyl, and phenolic groups of proteins under the conditions described (6). It is therefore not surprising that this reagent can be used successfully t o reduce the water absorption of all the proteins tested. Phthalic anhydride treatment greatly reduced the number of basic but increased the acid groups of proteins; yet with some proteins this reagent gave derivatives of similar water absorption t o those of the phenyl isocyanate and benzoyl derivatives, which are low in acid groups. Although it is appreciated t h a t the carboxyl groups of phthalic acid may not correspond in their behavior t o those of proteins, nevertheless this observation
Specific
Vol. 38, No. 1
suggests that carboxyl groups do not play so important a role in determining the hydrophilic tendency of proteins as do their basic groups. Similar indications are obtained from comparisons of the water absorption of proteins, the carboxyl groups of which have been esterified. The influence of the various types of polar groups and of their replacement by different molecular structures on the water affinity of proteins is receiving continued study. The present findings suggest t h a t the introduction of hydrophobic residues into proteins may be necessary if the water absorption is t o be reduced to a practical extent. The lack of such water-repelling residues may explain in part the high water absorption of derivatives obtained with formaldehyde and epoxides, since these reagents combine with approximately as many protein groups as are available t o the acylating agents (4, 7 ) . ACKNOWLEDGMENT
The authors are indebted t o G. H. Brother for encouragement !I. G. Young, and B. Brandon for and advice, and to >Cooper, technical asssitance. LITERATURE CITED
(1) Allen, C. F. H., and Bell, A., Org. Syntheses, 24, 94 (1944). (2) Brother, G. H., in Sutermeister and Browne’s “Casein and Ite, Industrial Applications”, A.C.S. Monograph 30, 2nd ed, Chap. 7, New York, Reinhold Pub. Corp., 1939.
(3) Brother, G . H., and McKinney, L.
L.,IND.ENQ.CHEM.,30,
1236 (1938). (4) Fraenkel-Conrat, H., J . Bio2. Chem., 154, 227 (1944). (5) Fraenkel-Conrat, H., and Cooper, M., Ibid., 154, 239 (1944). (6) Fraenkel-Conrat, H., Cooper, M., and Olcott, H. S., J . Am. Chem. Soc., 67, 314 (1945). (7) Ibid., 67, 950 (1945).
eats of
evea, G
d GR-I Stocks W. H. HAMILL, B. A. MROWCA, AND R. L. ANTHONY University of Notre Dame, Ind.
CCURATE knowledge of the specific heats of various natural and synthetic rubber stocks is necessary for the thermodynamic study of rubberlike materials. For example, the correlation between the entropy component of the stress-strain curve and the reversible heat developed when rubber is stretched rapidly (6) requires knowledge of the specific heat of the rubber as a function of elongation and of temperature. Knowledge of the specific heat is also needed in correlating calculated energy losses with rise in temperature in flexometer experiments. For the thermodynamic studies mentioned above, cL, the specific heat a t constant length, is needed. The dependence of cL upon the elongation was measured by Ornstein, W70uda, and Eymers (IO)and by Boissonas (6) with conflicting results. Omstein et al. obtained a strong dependence of CL upon elongation. Boissonas found constancy of CL for a cured stock, while for crude Hevea CL increased very slightly with elongation. T h e present paper will point out t h a t for regions without crystallization the observed dependence of the stress upon the temperature necessarily leads t o a constancy of CL up t o the fourth decimal. Thus, for all practical purposes, C L may be considered identical with cg. Many workers have measured the specific heats of various types of rubber stocks. Results of earlier work are listed in standard treatises on the subject-for example, “Rubber,
Physical and Chemical Properties” (17): Bekkedahl and M a t h e son (9) gave a n extensive review of earlier work. The great majority of the published results are of little value for purposes of thermodynamic calculations. I n some cases the rubber stock employed was not sufficiently defined as to composition. I n others the values are definitely unreliable because of rayher dubious experimental techniques. Some recent values of considerable interest are those of Boissonas (6),those of Roth, Wirths, and Berendt (14) and those of Fouts ( 7 ) . These values will be discussed later in the paper. Recognizing the need for accurate thermodynamic data on rubber, under well-defined conditions, t h e National Bureau of Standards (8, 5,12) has for some years been carrying out a program of supplying such information for various unvulcanized rubbers. As a result of their work accurate values of specific heat at constant pressure are now available for purified natural rubber, Hycar OR-15, and GR-S over a continuous range of temperatures from absolute zero to well above room temperature. Most investigations of the equation of state of rubber, studies of stress-temperature relations, and studies of heat build-up employ vulcanized stocks containing all the usual ingredients, such as fillers, reinforcing agents, accelerators, etc. Specific heat values for such compounds are required for complete investigations of this nature. It is of interest t o see whether or not spe-
