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INDUSTRIAL AND ENGINEERING CHEMISTRY
DISCUSSION OF RESULTS It is apparent from the biological experiments with frozen orange juice that this juice can be frozen quickly and permitted to liquefy at room temperature with no appreciable loss of vitamin C, and under the conditions of this experiment there was no discernible advantage from the standpoint of vitamin C preservation in excluding atmospheric oxygen during the freezing process. Even after storage for 10 months the juice retained its vitamin C potency, irrespective of whether it was preserved in an atmosphere of oxygen or nitrogen. These facts showed that vitamin C in orange juice is quite resistant to oxidation a t low temperatures and in an acid medium. The variation in vitamin C content of oranges is worthy of further investigation. I n the present studies 1.5 cc. of fresh orange juice did not afford the same measure of protection from scurvy as in previous studies by Nelson and Mottern (3) on normal fruit. The difference in response cannot be attributed to experimental procedure in i o far as the animals, the basal diet, or laboratory technic are concerned. Titrations with 2,6-dichlorophenolindophenol show that there is a measurable difference in the reducing value of frozen orange juice that has been preserved in an atmosphere of nitrogen and air or oxygen. Assuming that this difference not be is due to a per cent loss in vitamin c' it detected by biological experiments. The reducing value as determined by 2,6-dichlorophenol-
Vol. 25, No. 2
indophenol is not necessarily an accurate measure of antiscorbutic potency. Szent-Gyorgyi @), Zilva (7, 8 ) ) and Svirbeley and Szent-Gyorgyi (4) have published several papers dealing with this point. There may be many substances present in plant material which have some similar reducing properties. While glucic acid and hexuronic acid have similar reducing properties, the former has none of the physiological properties that have recently been attributed to the latter. However, in attempting to eliminate undesirable effects on vitamin C in commercial processes used for the preservation of such products as orange juice, the colorimetric titration may be a very useful guide.
ACKNOWLEDGMENT The glucic acid used was kindly provided by E. K. Nelson.
LITERATURE CITED Mottern, H. H., Nelson, E. M., and Walker, Reed, J . Assoc. OficiaZ Agr. Chem., 15, 614 (1932). (2) Nelson, E. K., and Browne, C. A., J . Am. Chem. SOC.,51, 830 (1929). (3) Nelson, E. M.,and Mottern, H. H., Am. J . Pub. Health, 22, 587 (1932). (4) Svirbeley, J. L., and Saent-Gyorgyi, A., Nature, 129, 690 (1932). ( 5 ) Szent-Gyorgyi, A., Ibid., 129, 943 (1932). (6) Tillmans, J., 2. Untersuch. Lebensm., 64, 11 (1932). Zilva, S. S., Nature, 129, 690 (1932). Zilva, S. S.,Ibid., 129, 943 (1932). R E C ~ V EAugust D 10, 1932. Preaented before the Division of hgricultursl snd Food Chemistry at the 84th Meeting of the American Chemical Society,
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Denver, COIO., August 22 to 26, 1932.
Vitamin C Content of Frozen Orange and Grapefruit Juices LILLIANW. CONNAND ARNOLDH. JOHNSON Research Laboratories of National Dairy Products Corporation, Inc., Baltimore, Md.
D
URING the last few years there has been a large increase in the quantity of fruit and fruit juices which have been preserved by freezing. Of special importance in connection with the freezing of such products and t,heir subsequent storage in the frozen condition is the effect on the vitamin content. Frozen orange juice has been marketed for several seasons, but little has been reported concerning its vitamin C content. More recently grapefruit juice has also been marketed in the frozen condition. There are considerable data in the literature on the effects on the vitamin C content of storing oranges, lemons, and other fruits and vegetables a t temperatures slightly above the freezing point. Thus Davy (2) and Delf (3) showed that, for oranges and lemons stored at 2.5" to 5.4" C., the antiscorbutic value was retained unimpaired as long as the fruit remained in good condition. Delf also prepared frozen orange and lemon juice, these juices being stored a t -11" to -14' C. No measurable deterioration in vitamin C content occurred after storage of 17 months in the frozen condition, but after storage for 5 years the antiscorbutic value had decreased about 50 per cent. On the other hand, Koch and Koch (6) found that frozen orange juice lost practically its total antiscorbutic value during storage for 3 months.
MANUFACTURE OE FROZEN JUICE The washed and graded oranges or grapefruit., previously cooled to a temperature of 4 ' to 5" C.; were peeled by a mechanical peeling machine, after which the juice was ex-
pressed by a screw type press, the juice being filtered through a sieve in the bottom of the press and the pulp expelled a t the end. The fruit presses as well as the other equipment with which the juice came in contact were all made of 18-8 chromenickel steel, which metal was found to be least corroded by orange juice and to have the least effect on flavor and keeping quality. The juice immediately after expressing usually contained so much pulp that filtration was necessary in order to remove it. After filtration, the juice was drawn into a vertical direct-expansion ice cream freezer, operated under reduced pressure, where it was frozen, to a slushy consistency. The slushed juice was then discharged into an evacuated hopper connected with the filling machine. As a precautionary measure to prevent the juice from dissolving oxygen as far as possible, nitrogen was introduced to relieve the vacuum and to facilitate discharging the juice into the filling machine and into the containers from the tilling machine. While this measure undoubtedly did not result in completely preventing air from getting into the juice, the manufacturer felt that juice prepared in this way had a better chance of maintaining its quality and vitamin potency during storage than juice not so processed with nitrogen. The slushed juice was finally packed in paraffied cardboard containers which when sealed were conveyed to the sharp room maintained a t a temperature of -24' C. A blast of cold air was directed on the containers until the juice was completely frozen, after which it was stored a t a temperature of -12'to -15°C.
INDUSTRIAL AXD ENGINEERING CHEMISTRY
February, 1933
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INDUCED IN GUINEAPIGSBY FEEDINQ FRESH, FROZEN, AND CENTRIFUGED ORANGE AND GRAPEFRUIT TABLEI. DEGREEOF SCURVY JUICES AND ORANQE JVICE PULP KINDOF JUICETESTED
DAILYD O S A Q E
-
Whole juice
DEOREE OF SCURVY Juice from which pulp was removed
Unwashed pulp
Cc. ORANGE JlXCE
Fresh Porto Rican valencia Freah Florida valencia Frozen Porto Rican valencia, prepd. in lab., stored 42 days Frozen Florida pineapple, stored 123 days Frozen Florida seedling, stored 105 days Frozen Florida valencia, atored 115 days
5.0
None (2 pigs) None (1 pig) None (1 pig) Mild (2 pigs) None (3 pigs) None (3 pigs) Mild (3 pigs) None (3 pigs) None (2 pigs)
None (3 pigs) None (2 pigs) None (2 pigs) None (2 pigs) None (2 p k s ) None 3 pigs) None [3 pigs) None (3 pigs) None 3 piga) None 12 pigs) 1 none I m i l d (2 pigs) None (2 pjgs) None (2 pigs) Mild (3 pigs) None (3 pigs) None (2 . Dies) Mild to moderately 8evere (3 pigs) Very mild (3 pigs) None ( 3 pigs)
5 .0
Mild (3 pigs) None (3 pigs) None (3 pigs)
Very mild (3 pigs) None (2 pigs) None (3 pigs)
1.5 3.0 1.5 3.0 1.5 5.0 10.0 1.5 5 0 10.0 1.5 5 0 10.0 0.6 1.5 5 0 0.5 1.5
. .
.
Frozen Florida valencia, stored 159 days
Frozen Florida, storage period unknown
10.0 0.5 1.5
None None None None None None None None
(3 pigs) (3 pigs) (3 pigs) (3 pigs) (1 p!g) (3 pigs) (3 pigs) (3 pigs
........
........
........ ........ .......
........ . . . . . . . . . . .
........
........
........
2 moderate, 1 aevere ( 3 pigs) 1 moderate, 2 severe (3 pigs) Moderate (3 pigs)
.......
ORAPEFRUIT JUICE
...... Very mild (3 pigs) Very mild (3 pigs) ....... None (3 pigs) None (3 pigs) ........ None (3 pigs) None (2 pigs) 1 Mild (2 pigs) 2 none, 1 mild (3 pigs) ..... Frozen Florida, storage period unknown 2 None (3 pigs) ...... None (3 pigs) 3 None (2 pjgs) . ....... None (3 pigs) 4 ........ None (3 pigs) None (3 pigs) Positive control, one pig fed basal scurvy diet until scurvy 5 Negative control three pigs fed basal scurvy diet died of severe scurvy in 24 to 26 days. began t o develop, th& 5 cc. of fresh orange juice added to diet daily; autopsy after 83 days showed the only scurvy lesion remaining to be a very alight beading of the ribs. Fresh Florida
1
2 3
Except for the frozen Porto Rican juice, the frozen orange and grapefruit juices used in these experiments were prepared on a commercial scale in the manner which has been described. The Porto Rican juice was prepared in the laboratory, the juice from hand-peeled oranges being expressed by a small press of the same type used commercially. Within 5 minutes after expressing the juice, it was frozen to a slushed condition, then placed in half-pint paraffined containers, and stored in an ice cream hardening room a t -17" to -24' C. where freezing to a solid condition soon took place.
VITAMINC DETERMINATION The method used for the vitamin C assay was essentially that described by Sherman and Smith (8). Guinea pigs weighing 280 to 350 grams were fed the basal diet consisting of 59 parts of ground whole oats, 30 parts of skim milk powder which had been previously heated in open trays a t 110' C. until all the antiscorbutic vitamin was destroyed, 10 parts of filtered butter fat, and 1 part of sodium chloride. The pigs were fed this scurvy-producing diet plus a sufficient quantity of fresh orange juice for a preliminary period of a week, during which time unsuitable guinea pigs were discarded. When the test proper was started, the feeding of the known antiscorbutic was discontinued and was replaced with known quantities of the juices to be assayed. These juices were fed a t definite levels on the basis per 300-gram pig. The pigs were weighed twice each week. The feeding experiments were continued for 80 to 90 days, a t the end of which time the surviving pigs were chloroformed and autopsied. The pigs which died during the experimental period were also autopsied. The rate of growth, scurvy symptoms before death, survival period, and lesions found upon autopsy were all considered in determining the degree of scurvy. In recording the autopsy findings, the various lesions of scurvy were noted, including hemorrhagic conditions of muscles, joints, bones, ribs, and intestines; brittleness and looseness of teeth; fragility of bones; swollen joints; and beaded ribs.
Where the only lesions found were brittleness of teeth or slight fragility of bones, accompanied by little gain in weight through the experimental period, these were designated as very mild scurvy. It is obviously of importance to know the effect on the vitamin C content of variations in the extent to which the pulp has been removed. Among the test juices fed was a series in which the pulp had been completely removed, also a series in which the pulp alone was fed. Fresh orange juices, prepared from the Florida and the Porto Rican valencia oranges, were used as controls. The fresh juices were prepared daily, one or more oranges being squeezed into a glass reamer. Half the juice obtained was set aside for feeding. The other half was subjected to centrifugation in order to remove the pulp. The supernatant liquid was poured through coarse muslin to remove the few pulp particles which usually floated on the surface. The liquid was not entirely clear but was free from visible particles. Half-pint containers of the frozen juice manufactured as described were removed daily from the cold storage room and thawed either by placing in cold water for 1 to 2 hours or by keeping a t 12" C. for 24 hours. The container was shaken thoroughly and a portion of the melted juice was removed. The remainder was centrifuged as described for the fresh juice, perfectly clear centrifugates being obtained. In the instance of the stored frozen juice from Florida valencia oranges, the pulp mas resuspended in cold distilled water in amount equal t o original volume of juice taken. The juices were kept in stoppered bottles in an ice water bath until fed. The feeding was always finished within 2 hours after preparation of the juice. The juices used in the study were fresh juice prepared from Florida oranges of the valencia variety, frozen juices from the seedling, pineapple, and valencia varieties, fresh and frozen Porto Rican orange juices prepared from valencia oranges, a frozen orange juice prepared from an unknown variety of Florida oranges, and fresh and frozen Florida grapefruit juice. During the 90-day feeding experiment
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1KDUSTRIAL AND ENGINEERING CHEMISTRY
the storage period of the frozen juices increased. Thus the juice, which when first used was approximately 5 months old, had been in storage for approximately 8 months a t the end of the feeding experiment. Fresh juices were fed a t 1.5 and 3.0 cc. levels. Frozen juices were a t first fed in 1.5, 5, and 10 cc. quantities. Not knowing the extent to which reduction in vitamin C content might have occurred due to freezing and storage, the maximum dose of frozen orange juice (10 cc.) was rather high. Later the dosages of frozen juice were reduced to 0.5, 1.5, and 5.0 cc. Fresh grapefruit juice was fed in 1, 2, and 3 cc. quantities and frozen juice in 1, 2, 3, and 4 cc. quantities. Three guinea pigs were used on each feeding level. Where the pigs died early in the experiment from causes other than scurvy, they were replaced whenever possible. In all cases the juice was fed by pipet. Negative and positive controls were included in the experiment.
RESULTS The results in Table I indicate that the antiscorbutic value of the frozen juices was equal to that of the fresh juice. This result has been confirmed by Eddy (4) on frozen orange juices from the same source. Moreover, storage of the juice for 5 months in the frozen condition did not appreciably reduce the antiscorbutic value. Whole orange juice, either fresh or frozen when fed a t the rate of 1.5 cc. daily, was found to give complete protection against scurvy. However, the guinea pigs receiving juice from the seedling variety did not grow so well as pigs receiving juice from the other varieties of orange. It was very difficult to maintain the guinea pigs for any length of time on this juice. Thirteen of the pigs (not included in Table I) lost so much weight during the first month that they either died or were taken off the experiment. Those that died showed no symptoms of scurvy. It was later learned that the frozen seedling juice was not especially representative of commercial runs, having been produced from overripe oranges. In the case of grapefruit juice, complete protection against scurvy was afforded when 2 cc. of either the fresh or the frozen juices were fed. The minimum protective dose of 1.5 cc. of orange juice was also obtained by Davy ( 2 ) , Delf (S), and Willimott and Wokes (.9).l I n the case of grapefruit juice, the present results showed that a 1-cc. daily dose failed to protect, whereas 2 cc. gave complete protection. MacLeod and Booher (7) found that 1.5 grams were insufficient, whereas 2 grams gave protection. Eddy, Gurin, and Kohman (5) found that in all cases a 3-gram dosage was protective while only mild symptoms were in evidence with the 2-gram dosage. These symptoms were not present in all animals. 1 After this paper was submitted for publication, it came t o the authors’ attention t h a t work done by Nelson and Mottern [IND.ENG.CHBM.,25, 216 (1933)] showed that 1.5 cc. of orange juice were insufficient to protect guinea pi58 against scurvy. Previoua experiments by the same workers [ A m . J . Pub. Health, 22, 587 (1932)l showed t h a t 1.5 cc. of normal orange juice supported good growth and gave protection against scurvy. b u t that the vitamin C content of fruit from trees which had been sprayed with arsenical compounds was greatly reduced. Factors such as this one, a.hich are r.ot under the control of the laboratory worker, may be responsible for the differences found by various investigators in the vitamin content of citrus fruits. It is considered important from this point of view that specific statements be made as t o the source of fruit used in each study. The Porto Rican oranges, from which both the fresh and frozen juices were prepared, were picked prior t o April 16, 1931. The Florida orangespineapple, seedling, and valencia varieties-were grown near Tampa, Fla., a n d were picked during February, March, and April, 1931. respectively, with the exception of one sample of which the source and storage period were unknown. The fresh grapefruit juice was from Florida fruit bought in the open market in Baltimore in December, 1931. The frozen grapefruit juice wan prepared in Tampa from fruit grown in t h a t vicinity and picked in April. 1931.
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Willimott and Wokes (9) state that grapefruit juice is not quite so rich in vitamin C as orange juice. Browell and Zilva (I), on the other hand, found that certain varieties of grapefruit juice are slightly richer in vitamin C than orange juice. The centrifuged juices were as effective in protecting against scurvy as the whole juices, 1.5 cc. of orange juice and 2 cc. of grapefruit juice again affording complete protection except in two cases, one for the seedling variety where one pig receiving 1.5 cc. developed mild scurvy, and the other for the valencia juice which had been stored for 159 days when three pigs, on 1.5 cc. daily, developed very mild scurvy. In the case of the frozen Florida juice (valencia variety) which had been stored for 159 days, the unwashed pulp of the juice was also fed. The resuspended pulp was fed in such amounts that the guinea pigs received the pulp obtained from 1.5, 5.0, and 10 cc. of the juice. As this juice contained 6 per cent of pulp by volume, this represents, respectively, 0.09, 0.3, and 0.6 cc. of pulp. Scurvy developed in all cases where this pulp was used, and the animals died between the seventeenth and thirty-second day, presumably because of scurvy. Some of these animals were designated as having died of “moderate” scurvy as the lesions found were not extreme, and as it was not known definitely that scurvy was the cause of death, although no other cause was apparent. In view of such a short survival period the degree of severity of lesions may not have much significance. As the animals receiving 10 cc. developed practically as severe scurvy as those receiving only 1.5 cc., the slight protection they received is thought to be due to the small amount of adhering juice. On all other frozen juices, experiments were also conducted in which the pulp was washed once with water before being used. Severe scurvy developed on all feeding levels when this pulp was used. As some of the vitamin was probably removed by the washing, the importance attached to this is not so great as where the unwashed pulp was used. Before this work was done, it was feared that by depulping the juice the vitamin C content might be impaired. If the vitamin were concentrated in the pulp rather than in the clear juice, such amounts of pulp as were fed should have been fully protective inasmuch as they were derived from amounts of juice containing the minimum protective dose, or more, of vitamin C. Animals receiving 0.5 cc. of whole juice developed only mild scurvy; those receiving 0.6 cc. of pulp (pulp from 10 cc. of juice) developed scurvy to a more severe degree. As the juice completely depulped by centrifugation was equal in vitamin C potency to the whole juice, and as the pulp from as much as 10 cc. of juice was found to be practically nonantiscorbutic, it appears that the pulp may be removed from orange or grapefruit juice to any desired degree without reducing the vitamin C potency. The results which have been obtained in this study are in general agreement with those of Delf ( 3 ) who found that freezing did not reduce the vitamin C potency of orange juice, and, moreover, that storage of the frozen juice for even as long as 17 months did not operate to reduce the antiscorbutic value appreciably. Koch and Koch (6),on the other hand, found that during storage of the frozen juice for 3 months the loss in vitamin C potency was practically complete. The reason for the difference in results probably lies in the method of preparing the original juice. Koch and Koch (6) state that their juice was frozen within 48 hours after preparation. The commercially prepared juices used by the present authors were frozen within 2 hours after expressing from the fruit, and care was taken to prevent oxygen from entering the juice, both by keeping the juice under reduced pressure and by the use of nitrogen.
February, 1933
INDUSTRIAL AND ENGINEERING CHEMISTRY
COWLUSIONS The vitamin C contents of commercially prepared frozen orange and grapefruit$juices were equal to those of the fresh juices. Storage of orange juice for 5 months in frozen condition did not appreciably reduce the vitamin C potency. Orange and grapefruit juices from which the pulp was removed by centrifugation were practically equal iri vitamin C potency to the whole juices. Orange juice pulp obtained from 10 cc. of juice was found to be insufficient to protect against scurvy. Orange juice gave complete protection when 1.5 cc. of
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juice were fed per 300-gram pig, and grapefruit juice gave complete protection when 2 cc. were fed. LITERATURE CITED (1) Browell, M. F., and Zilaa, S.S., Biochem. J., 25, 1081 (1931). (2) Davy, A. J., I b i d . , 15, 83 (1921). (3) Delf, E. M., Ibid., 19, 141 (1925). (4) Eddy, W. H., private communication, 1931. (5) Eddy, IT. H., Gurin, C. Z., and Kohman, E. F., IND. ENQ. CHEM.,24, 457 (1932). (6) Koch, E. hl., and Koch, F. C., I h i d . , 24, 3.51 (1932). (7) MacLeod, G., and Booher, L., J . Home Econ., 22, 588 (1930). (8) Sherman, H. C., and Smith, S. L., "The Vitamins," pp. 148220, Chemical Catalog, 1931. (9) Willimott, S. G., and Wokes, F., Pliarm. J . , 118, 770 (1927).
RECEIVED .4ugust 29, 1932.
Development of Dakota Lignite VIII. Oil-Steam Atmosphere for Dehydrating Dakota Lignite A4.M. COOLEY,JR., AND IRVINLAVINE,University of North Dakota, Grand Forks, N. Dak. modification of this apparatus The use of small quantities of certain oils has BRIOUS methods for the consisted in a n oil reservoir and successful dehydration been found benejicial in dehydrating Dakota liginjector for drawing the oil into of lignite h a v e b e e n nite by steam. The nature as well as the quanthe steam. studied in this laboratory for the tity of oil has a n important bearing on the charpast few years. Lavine, Gauger, The injector was made of ordiacter of the dried fuel. nary pipe fittings and consisted and Mann (6) found t h a t certain of a l/p-inch (1.3-cm.) pipe, fitted Dehydrated Velzra lignite can be improved maDakota lignites can be dehyat one end with a perforated cap. drated with a minimum of disterially by 2he addition of a small quantity of a n This pipe was connected by a integration by the use of satusuitable union t o the steam line emulsijied asphalt to the steam. Details as to from the boiler, and the capped rated steam at pressures u p to the method .for incorporating the oil and quantiend was extended into a 3/4-inch 15 atmospheres. The work was (1.9-cm.) tee. A pi e then made ties desired are given. The physical charactercontinued later by Cooley and connection from t i e tee t o the istics of the Velva lignite dehydrated by the Lavine (a) who found that the drier, and the upper part of the tee was connected by suitable pipe deposit from Wilton, N. Dak., steam-oil atmosphere have been found to excel t o the oil reservoir. was suited especially for this the properties of Velva lignite dehydrated by The oil reservoir was made from process. The material was rea 6-inch (15.3-cm.) cast-iron pipe steam only. moved from the drier in a hard, equipped a t both ends with caps. The addition of a small quantity of oil has been The top contained a removable tough condition with but little plug for inserting oil and a small found not io increase materially the tendency evidence of checking. I n the steam pipe which was used to case of t h e d e p o s i t s f r o m towards spontaneous combustion. equalize the pressure above the oil. Soonan. Beulah. and Lehieh. A suitable pipe from the bottom contained a needle valve and exS . Dak., it was found &at steam-drying was accompanied by excessive disintegration tended directly to the top opening of the tee. Steam from the boiler in passing through the perforated cap drew oil from the of the lump. This investigation showed further that lignite reservoir, and the mixture then passed into the drier. The needle from Velva, Garrison, and Columbus, N. Dak., can be de- valve in the line from the reservoir qerved to regulate the rate of hydrated to a moisture content of about 15 per cent, but flow of oil. with these fuels the dehydration is accompanied by a less A further modification of the older set-up was an arrangesevere checking. The dried lignites from these deposits were found to withstand very well the ordinary conditions of ment of pipes F-hereby the steam-oil mixture could be inweathering and handling. The present investigation was serted either through the top or through the bottom of the undertaken with the view of minimizing the checking of the drier. I n the first experiments of this study the oil was injected lump lignite during dehydration. In considering means of accomplishing this end, it seemed directly with the steam from the yery start of the process that the presence of a n oil phase might prove of value. according to the procedure described in an earlier paper (6). Many colloidal materials, similar in character to lignite, Later this procedure was modified in that oil was used only exhibit a marked preferential adsorption for one component during the latter part of the drying process. Thus, saturated of a multicomponent system. Thus, it was thought that, steam was used until the release period and then the steamby incorporating a sniall quantity of a suitable oil with the oil mixture was used for a few moments prior to the release of steam before it came in contact with the coal, the lignite pressure. The lignites used in this work were obtained in sealed conwould adsorb sufficient oil to act as a cell binder. tainers from the mines listed in Tables I and 11. APPARATUS AND PROCEDURE The following oils were used: The apparatus used in this investigation was essentially 1. Iso-Vis motor oil KO.50 (Standard Oil Company) the one developed by Lavine, Gauger, and Mann (5). A 2 . Medium Texaco motor oil
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