H. D. R o y c e , The Southern Cotton Oil Company, Savannah, Ga.
T
HE relationship between o x id a t i v e in d u c t io n period and the fading time of methylene blue in oils exposed to r a d ia t io n was first pointed out by Greenbank and Holm (/}), who made it the basis of a photoelectric stability test.
The possibilities of adapting the m e th o d to r o u t in e keeping- indicator are presented. Photoelectric control o f the end p o in t has not been fo u n d satisfactory in testing hydrogenated fa ts, because o f secondary color changes that lake p la ce in the reaction m ix
while not detracting from the value of the method for certain applications, necessitated further changes in the apparatus.
Also the use of accelerated oxidation data to supplement in
dividual fading-time determinations will be pointed out, in conjunction with comparative figures on peroxide formation.
The increasing attention being centered on the develop
ment of improved stability test and evaluation of fat stabi
lizers becomes apparent in reviewing recent literature. Of especial interest in connection with the methylene blue method is the procedure recommended by Bruere and Fourmont (S), involving the use of a series of decolorized reduction indicators having graded reduction potentials. Depending on the stage of incipient rancidity in the fat under examination, certain of the indicators revert to the colored form. Probably a comparison of results by this method with the iodometric peroxide estimations of Lea (8) or Wheeler (18) would show that the color change depends upon the presence of peroxides in the sample. air thermostat of asbestos-lined wood c o n s t r u c t i o n 10 X 10 X 14 feet long. The sliding sample rack Z holds six American Oil Chemists’ Society oil color tubes 0 equidistant from the source of illumination L. This rack pulls out from the side of the box and facilitates the rapid change of samples with minimum temperature drop in the thermostat.
The magnesia block mounting is cut away under the tubes, so that the fading time may be checked by viewing the sample from above through TFj, as well as by direct transmitted light through W i. The blower heater unit B was adapted from a 400-watt relay and S3 in series with fan motor, best values for operation at 70° C. were Si, 50 watts, Si, 100 watts, and S t, 25 watts). When operating at other temperatures, or to maintain temperature at start of test before the irradiating lamp L is cut on, the necessary changes in resistance may be made easily. the mixture are then brought to 70° C. by holding for a with most vegetable oils and hydrogenated fats the color change is quite abrupt at the
July 15, 1933 I N D U S T R I A L A N D E N G I N E E R I N G C H E M I S T R Y 215 A few precautions are necessary to obtain consistent and
reproducible results. Moisture must be excluded in all opera
tions, and if the original sample contains more than traces of water it should be dried, care being taken to minimize oxi
dation in this operation. The methylene blue reagent should be prepared from aldehyde-free anhydrous alcohol, and can be kept satisfactorily for a few days in a stoppered bottle placed in a desiccator. A number of determinations have been made using aldehyde-free anhydrous propanol, but it has no apparent advantage over the lower alcohol. Newton (9) has emphasized the importance of thoroughly cleaning glassware which has been in contact with rancid fat, in con
nection with the incubation accelerated rancidity test, and it is likewise imperative to clean the color tubes carefully in the present method. It has been found that rinsing with a fat solvent, followed by a 30-minute boil in 20 per cent sodium hydroxide and overnight soaking in cleaning solution, serves the purpose very well. With most fat samples, the degree of accuracy of the method is determined by the personal factor in reading the end point. Colored oils rich in yellows and reds naturally give a green color with the reagent, and as the blue fades the green shades rather gradually to the original tint. However, visual observation of the end point has been found to be more satisfactory and reliable than the photo
electric method, for in spite of the use of color filters in
con-F iG u n E 2 . Ef f e c to f Te m p e r a t u r ea n d Me t h y l e n e Bl u e Co n c e n t r a t i o n o n Fa d i n g Tim e Teste made on freshly deodorized choice cottonseed oil.
1. Tem perature-fading tim e curve 2. Concentration-fading tim e curve
* 70° C. and 0.001 per cent m ethylene blue concentration adopted as standard for control work.
nection with the latter, many natural oils and all-hydrogenated fats which have been examined give secondary color changes that interfere with photoelectric control. Since the methylene blue reduction period for m ost refined edible fats and oils does not exceed one hour, and the average is less than 30 minutes, it is not particularly tedious or inconvenient to record the end points visually.
The duration of the methylene blue fading time may be varied within certain limits by changing the reaction tempera
ture, dye concentration, and light intensity. Figure 2 gives the values for freshly refined and deodorized cottonseed oil under the maximum range of conditions which have been found practical with the present apparatus. For control work on the estimation of stability of salad oils and shorten
ings, 70° C. and 0.001 per cent methylene blue concentra
tion were adopted as standard conditions.
Figure 3 shows the comparative stabilities of four types of hardened cottonseed oil shortenings, aged in friction top cans at 90° to 95° F. Organoleptic manifestation of rancidity is indicated at the points designated by R on the methylene blue fading-time curves. All-hydrogenated shortenings are
1. All-hydrogenated 3. Bulk vegetable compound
2. Package vegetable compound 4. Bulk oleo compound
generally conceded to have superior keeping quality in com
parison with compound types, and the fading-time curve 1 illustrates this point very well; 2 and 3 are two different grades of hydrogenated vegetable compound shortening, and having similar composition, the stability curves follow the same general trend. Curve 4 may be interpreted to show that the original sample was very close to a state of incipient rancidity, since the slope of the curve corresponding to the initial stage of the aging period resembles that of the other fats at a later stage in the test. While this inferior keeping quality of the oleo compound cannot be,attributed entirely to the oleostearin content, it has generally been found that strictly vegetable compounds show a higher initial fading time. If, however, the curves are to be interpreted for prac
tical purposes, it should be pointed out that the smaller slope and higher fading time during the later stages of the test on the oleo compound indicate that, although the original condi
tions of such fats may not be very good, the rate of change under ordinary storage conditions is slightly slower than the rate for vegetable compound.
Certain metals in oil-soluble form are known to have a marked effect on the stability of fats, and Figure 4 gives the fading time of methylene blue in cottonseed oil to which very low percentages of some of the common metals have been added. Copper is a powerful pro-oxidant for autoxidation reactions (i) and the accompanying curve shows that only 20 p. p. m. reduce the fading time from 24 to 8 minutes. Man
ganese in somewhat higher concentrations likewise has a strong pro-oxidant effect, but ferrous iron,1 divalent tin, and nickel at the low concentrations plotted on the graph have no effect on the fading time. Zinc apparently has a stabilizing action, but this point has not been checked by aging tests, and it is probable that the prolonged fading time in this case is due to some interaction of the zinc with methylene blue. Further work is being conducted on the applicability of the methylene blue reduction method in the presence of pro-oxidants and antioxidants.
Figure 5 presents keeping quality curves for a number of freshly refined and deodorized vegetable oils. Coconut oil
i Under oxidizing conditions, low concentrations of iron m ay shorten'the induction period of fats and oils considerably- In one experiment, 0,00025 per cent of ferric stearate was dissolved in cottonseed oil, and th e sam ple was aged b y aération and irradiation a t 100° C. T he rate of peroxide formation was accelerated greatly. In th e present experiment, it is thought th at oxida
tion of Fe + + to F e + + + doçs n o t occur in th e lim ited te st period (24 m in.).
Generally speaking, the m ethylene blue te s t is not always reliable when applied to fats containing substances having a high reduction or oxidation potential.
has a high original fading time, but this falls rapidly to a value only slightly higher than that for corn oil after aging 5 weeks in glass at room temperature. Soybean oil gives an aging curve similar to corn oil, while the remaining oils are grouped quite closely. Peanut oil, at least this particular sample, proved to be the most unstable of the group. Since the histories of these oils prior to refining and deodorizing are
Fi g u r e 4 . Ef f e c t o f Me t a l s o n Fa d i n g Tim ei n Co t t o n s e e d Oi l M etals added as freshly prepared soaps of
fully hydrogenated cottonseed fa tty acids
not known, it is not possible to generalize from the data given on the relative stabilities of these oils as a class. For example, some of the oils may have been prepared from old or damaged seed, and some may have been extracted with solvents, as against pressing for others. This graph is merely presented to show that methylene blue reduction affords a convenient method for following the progress of oxidative rancidity in various kinds of oils.
The relation of fading time to Kreis number and peroxide value is shown by Figure 6. A sample of cottonseed oil was aged rapidly by aerating at 100° C. under intense illumination,
bo that a rancid odor developed in 2 to 2.5 hours. Six hours under these conditions sufficed to complete a test, sending the peroxide value (millimoles of peroxide) to 250. (A compari
son of accelerated aging tests, with and without photocataly
sis, has been included in another paper.) It may be said
Fi g u r e 5 . St a b i l i t y Cu r v e s f o r Fr e s h l y Re f i n e d Ve g e t a b l e Oi l s f r o m Di f f e r e n t
So u r c e s
Samples aged at 9 0 ° to 95° F. in loosely stoppered glass bottles, exposed to diffused daylight.
1. Coconut 5, Cottonseed
2. Corn 6. Peanut
3. Soybean 7. Sunflower
4. Rape
here, however, that for pure cottonseed oil the shape of the aging curve is approximately the same in both cases, so that the light may be considered as a catalyst which does not markedly alter the type of reaction products which are opera
tive in bringing about the color change in methylene blue.
B y the use of a 200-watt lamp at 15 cm. from the oil, employ
ing a modification of Wheeler’s (IS) accelerated aging ap
paratus, the aging period has been reduced 100 per cent and more.
The peroxide value curve in Figure 6 shows the extent of the oxidative induction period, while the fading time and Kreis numbers follow a fairly constant inverse proportion.
This may be interpreted to indicate that the oxidative de
composition products causing the Kreis reaction are respon
sible for the methylene blue reduction, whereas the concentra
tion of peroxide as measured by Wheeler’s method is better adapted to register the termination of the induction period, when fat samples are aged by aerating at high temperature.
Like the Kreis test, the methylene blue method is most satis
factory when used on fresh nonrancid fats, or fats in a condi
tion of incipient rancidity, in which a rancid flavor has not become pronounced.
Interpretation of methylene blue fading time for practical application to problems of processing and storage of fats and oils may best be indicated by referring again to Figure 3.
All the vegetable shortenings used in this series were freshly bleached and deodorized at the beginning of the test. Thus, knowing the history of the fats, the initial fading times would
Fi g u r e 6 . Co m p a r a t i v e St a b i l i t y Cu r v e s o n Co t t o n s e e d Oi l Su b j e c t e d t o Ac c e l e r a t e d
Ox i d a t i o n a t 1 0 0 ° C . Irradiated b y 200-w att lam p at 15 cm. distance
be sufficient to classify the vegetable shortenings according to stability, in the order 1-2-3, with 1 standing considerably superior to 2 and 3. The oleostearin used in making the oleo compound had not been freshly deodorized, so that, although the initial fading time is the lowest of all, it would not be safe to conclude that this type of compound has the lowest sta
bility. Subsequent values on the aging curve tend to show that if the oleo compound had had the same treatment as the other samples, the initial fading time would have been equal to or higher than 2 and 3. For the estimation of stability when testing samples of known history, the methylene blue method ranks with direct oxygen absorption (IS), with the added advantage of a much shorter test period and simplicity of apparatus. Additional practical information on fat sta
bility may also be obtained by using the methylene blue test in conjunction with accelerated aging, or low-temperature aging, when time is not the dominant factor. Wheeler’s accelerated oxidation apparatus in a modified form is being used at present in this laboratory, and the reaction is followed by measuring the change in methylene blue fading time as well as increase in peroxide values.
Ac k n o w l e d g m e n t
The author wishes to express his appreciation to M. C. Kib- ler for assistance in the experimental work, and to W. S. Lovell for making Figure 1.
July 15, 1933 I N D U S T R I A L A N D E N G I N E E R I N G C H E M I S T R Y 247
Li t e r a t u r e Ci t e d
(1) Alyea, H . N ., and B ackstrom , H . L. J., J . A m . Chem. Soc., 51, 90 (1929).
(2) Bailey, H . S., and E b ert, H ., Cotton Oil Press, 7, 35 (1923).
(3) Brucre, P ., and Fourm ont, A., A n n . fals., 25, 91 (1932).
(4) G reenbank, G. R ., and Holm, G. E ., I n d . Eno. C h e m ., Anal.
E d ., 2, 9 (1930).
(5) Ibid., 17, G25 (1925).
(6) G rettie, D. P., and N ewton, R. C., Ibid., 8, 291 (1931).
(7) Issoglio, G., A n n . chim. applicata, 6, 1 (1916).
(8) Lea, C. H „ Soap, 7, 83 (1931).
(9) N ewton, R . C., Oil Soap, 9, 247 (1932).
(10) Pool, W. O., Oil Fat. Ind., 8, 331 (1931).
(11) Richardson, A. S., Eekey, E. W ., and Andrews, J. T . R ., Ibid., 8, 409 (1931).
(12) Royce, H. D., Soap. 7, 25 (1931).
(13) Wheeler, D. H ., M. M. A., Bull. 121 (Jan., 1932).
Re c e i v e d February 6 , 1 9 3 3 .