Tocopherols as Antioxidants for Vitamin A in Fish Liver Oils L. 0. BUXTON National Oil Products Company, Inc., Harrison, N . J .
Pure B- and y-tocopherols, alone and in association with vegetable oil lecithin, were studied for the inhibition of vitamin A destruction and peroxidation in soup-fin shark liver and halibut liver oils. Both forms of tocopherols were found to be effective antioxidants for retarding peroxidation, a characteristic not possessed by a-tocopherol. y-Tocopherol was more effective for inhibiting peroxidation in the oils studied than was B-tocopherol, especially during the later stages of vitamin A destruction. pand y-tocopherol at a 0.10% concentration markedly increased the stability of vitamin A in the various oils examined. Lecithin at a 1.0% level enhanced the effectiveness of both tocopherols for inhibiting vitamin A destruction and peroxidation. No difference was observed between the antioxidant characteristics of pure natural y-tocopherol and those of pure synthetic y-tocopherol.
T
HE inherent resistance of natural glyceride oils to oxidative
changes is due primarily to the presence in the oils of varying quantities of substances commonly termed antioxidants. Many natural oils, particularly fish liver oils, are notoriously easy prey to oxidative changes, regardless of the fact that they also contain some natural antioxidants. Fish liver oils are Our most valuable source of vitamin A; because of the lability of vitamin A to oxidative changes, i t is important that oxidation be inhibited to the fullest degree. Evidence to date has established that tocopherols and allied substances in association with phosphatidic materials are largely responsible for the stability exhibited by natural vegetable oils. Olcott and Emmerson (6) were the first to show that the tocoPherols are increasingly effective as antioxidants in lard in the order alpha, beta, and gamma* The gamma form was found to have about three times the antioxygenic index of a-tocoPherol, and the activity' of P-tocopherol was intermediate between the other two. Esters of the tocopherols were ineffective as antioxidants. Olcott and Mattill (7) showed that the antioxidants or inhibitols present in the lipid fractions of vegetables and in vegetable oils are so similar to vitamin E (tocopherols) that it was impossible t o fractionate them. I n a paper on the use of natural as well as synthetic a-tocopherol as an antioxidant in cottonseed oil, Swift, Rose, and Jamieson (11) demonstrated that the a-tocopherols function most effectively a t lower levels of concentration. These authors also showed that a cephalin fraction from cottonseed oil greatly enhanced the antioxygenic activity of a-tocopherol. Certain acids also increase the antioxygenic activity of tocopherols and of the corresponding quinones in autoxidizing fats (5). Published data on the antioxidant activity of phosphatides in oils as a whole is experience that, when ~t has been the tively pure phosphatides alone are added t o antioxidant-free fish liver oils, they have no antioxidant activity for inhibiting either peroxide formation or vitamin A destruction, even a t concentratiom as high as 5%. When relatively pure phosphatides are added to crude fish liver oil6 they may Or may not exhibit an antioxidant effect, depending upon whether the crude oils contain natural antioxidant principles which may act synergistically with
the added phosphatides to enchance the stability of the oils. Golumbic (4) reported that the antioxygenic o-quinones present in vegetable fats but not in animal fats retarded peroxide accumulation in the former type of oils after the complete disappearance of the tocopherols. Riemenschneider (8) showed that various vegetable oils added to lard in amounts of 1 to 10% appreciably increase its stability and that the increase is related to the tocopherol content of the oils. Robeson and Baxter (9) isolated a-tocopherol from Mangona shark liver oil and from soup-fin shark liver oil by distillation and concluded that the a-tocopherol found in the soup-fin shark liver oil was about 0.04%. The RIangona shark liver oil was found t o contain about 0.01% and was the major antioxidant present in the oil. Bird (W) concluded that the natural antioxidants in fish oils probably involve several types of compounds. Using halibut liver oil, he showed that exhaustive extraction with 80y0methano1 removed only part of the antioxidants, whereas with water or dilute aqueous alkali, complete extraction or destruction of the antioxidant fraction occurred, It was also reported that a-tocopherol a t high concentrations is an effective antioxidant for vitamin
A.
rn another report from these laboratories (3)i t was shown that the natural antioxidants present in various types of fish liver oils were either completely removed or destroyed when the oils were treated with activated carbon in the presence of a solvent for the of the pure oil, The presentinvestigation reports the effect p-, and 7-tocopherol isomers, alone and in combination with vegetable oil lecithin, on vitamin A destruction and peroxidation in soup-fin shark and halibut liver oils, It n.as postulated that such studies on the relation between the destruction of vitamin and the rate and extent of peroxidation, with and without the added lecithin, should bring to light tocopherols or tocopherols any similarity between the action of the latter type of antioxidants and those naturally occurring in the crude fishliver oils. Furthermore, to date the literature is lacking in information on the effectivenessof a-,p-, and 7-tocopherols as antioxidants for vitamin A as i t exists naturally in fish liver oils. This Rork is not concerned with the merits of tocopherols or tocopherols plus lecithin as antioxidants for the alcoholform of vitamin A. MATERIALS
oILS.
cRUDE The samples of crude soup-fin shark and halibut liver oils used in this work were of known origin and of the highest quality. They were filtered before using and were dry and free from foreign matter. It was established by numerous stability studies, under analogous storage conditions on other comparable lots of soup-fin shark and halibut liver oils, that the oils used in this investigation behaved in a manner quite representative of this type of oil, both as t o the rate and extent of peroxidation and the rate of vitamin A destruction. The crude oils used can be considered as being relatively stable, in so far as fish liver oils are concerned. Their analysis is given in Table 1. , CARBON-TREATED OILS. The samples of crude soup-fin shark and halibut liver oils were treated with 2001, (based on oil weight) of activated carbon (Nuchar XXX), using cyclohexane as the solvent (20% solution of oil in solvent) by the process reported previously (3). The cyclohexane used was a special grade sold by The Barrett Company and was devoid of unsaturates, as determined spectrophotometrically: The carbon-treated oil samples were 225
KNDUSTRIAL A N D E E G I N E E R I N G CHEMISTRY
226
4
/>
Figures P and 6
-3
'
1
Vol. 39, No. 2
Curve 1. Crude soup-fin shark liver oil C Curve 2. Carbon-treated shark liver oil C 0.1070 synthetic a-tocopherol Curvr 3. Same as 2 0:25% synthetic a-tocopherol Curve 4. Same Crime 5. Same 0.50% synthetic a-tocopherol 1.00% synthetic a-tocopherd Curve 6. Same Cur-le a. Same 4- 1.25% Veeol (40%)
+ +++
Figures 2 and 4 Curve Curve Curve Curve Curve Curve Curve Curve
E.
2. 3. 4.
5. 6. 7. 8.
Crude soup-fin shark liver oil C Carbon-treated shark liver oil C Same as 2 170 lecithin Same L 0.'50% lecithin and 0.05% synthetic a-tocopherol Same $ 0.50% lecithin and 0.10% synthetic a-tocopherol 0.50% lecithin and 0.50% synthetic a-tocopherol Same Same 5.0% leoithin nnd 0.50% Rynthetio a-tocopherol Same 0.50% lecithin and 1.25% Vegol (40%)
+
++ +
Figures 3 and 8 Curve Curve Curve Curve
1. Crude halibut liver oil A 2.
3. 4.
Carbon-treated halibut liver oil A Same as 2, 0.10% synthetic a-tocopherol Same 0.5070 synthetic a-tmopherol
+ +
221
INDUSTRIAL AND ENGINEERING CHEMISTRY
February 1941
200
0
0
200
400 0
200
HOURS A T 345°C:
400 0
200
+
used immediately after preparation or stored in completely filled, closed vials a t - 18 O C. until the peroxide and vitamin A stability .tests were conducted. From a vast accumulation of unpublished data on such carbon-treated oils, it can be assumed that the only difference between the crude and the carbon-treated oils, in so far as this investigation is concerned, is that the latter were devoid of natural antioxidants. Partial analytical data on the carbontreated oils are given in Table 11. TOCOPHEROLS. The synthetic 01-, 4-, and 7-tocopherols used were obtained from Merck & Company. The Vegol(40% grade) and the pure natural 01- and 7-tocopherols used were secured from Distillation Products, Inc. The E value of the Vegol sample a t 2965 A. in cyclohexane was 40.4. The tocopherol isomers were used without further purification and can be considered pure specimens, since the extinction coefficients checked favorably with reported values on similar types of materials.
$00
800
600
HOURS AT 34.YC.
Figures 4 and 9 Curve 1. Crude soup-fin shark liver oil C Curve 2. Same as 1 4-0.10% synthetic a-tocopherol Curve 3. Crude halibut liver oil A 0.1% synthetic a-tocopherol Curve 4. S a m e
600
400
Figures 5 and 10 Curve 1. Crude soup-fin shark liver oil B Curve 2. Same as 1, 0.1% natural y-tocopherol Curve 3. Same 0.1% natural y-tocopherol and 1.0% leoithin Curve 4. Same as 3, 0.5% natural y-tocopherol
+ ++
LECITHIN. The commercial lecithin product called Yelkin
BSS, obtained from the American Lecithin Company, Inc., was used without further treatment. This product is considered a refined grade of lecithin. Many different types of lecithin or phosphatide preparations were investigated in the course of this study; the differences were slight and merely a matter of degree. The samples of crude and carbon-treated shark liver and halibut liver oil, with and without the added tocopherols or tocopherols plus lecithin, were studied for percentage vitamin 9 destruction and peroxidation by the method outlined previously (IO). Placed in thoroughly cleaned open vials of uniform size, 2-mm. samples of the various oils were stored in a constant temperature oven maintained at 34.5" * 0.5' C. At definite intervals, as graphically recorded inFigures 6 to 15 and in Table 111,threevials of the same oil were separately tested for peroxide number and vitamin A. The average values of the individual determinations are re-
228 TABLE
INDUSTRIAL AND ENGINEERING CHEMISTRY I . 4SALYsIS O F CRUDES O U P - F I N SHARKAND HALIBUT LIVEROILS Soup-Fin Soup-Fin Soup-Fin Halibut Shark d Shark B Shark C A
Sp. gr. at 60' F. Color Red
Halibut B
0.912
0.927
0.930
0.931
0.925
4 1 18 0 0.65 177 0 139 4
3 7 23 0
4 0 36.0 0.2 181,7 175.9 4 3 105.000 5.3 29
2 1 30.0 0.3 170.5 139.0 11 0 94,200 1.2 10
3.6 70 0 0 6 173.0 142.5 8.4 60,000
Yellow Free fatty acid, "c Saponification value Iodine ralue a.75 Unsaponifiedmatter, Yc Vitamin A. units/eram 71.500 Peroxide number 0 138 Phosphorus, p.p.ni.
0 35 174.5 157 9 6.61 122.000 0 37
2.6 7
ANALYSISOF CARBOS-TREATED SOCP-FIX TABLE 11. PARTIAL SHARKAKD HALIBUT LIVEROILS Soup-Fin Shark C Color R Pil Red -..Yellow Free fatty acid, yo Titamin A , unitslgram Peroxide number Phosphorus, p.p,m.
0 4 10 0 0.2 103,000 4.0
0
Halibut A 0 . 33 10.0 0.25 93,500 1.1 0
Halibut B 0 .4
10.0
0 5 60,000 2.4 0
corded in Table 111 and in the various figures. The peroxide numbers viere established by the method already reported ( 1 0 ) . The vit,amin A determinations x-ere made in a spectrophotometric grade of 99% isopropanol by means of a Beckman spectrophotometer. The oil constants reported in Tables I and I1 TTere ascertained by official B.O.A.C. methods. The phosphorus coiitent of the oils was determined by the microcolorimetric method of Berenblum and Chain ( 1 ) . The data i n Table I show that tvio of the soup-fin shark liver oil samples possessed considerably higher iodine values than the samples of halibut liver oil. The iodine value of a fish liver oil is only a relatively minor element of the picture, however, in so far as peroxidation and vitamin A stability of the oil are concerned. Other factors, such as the content and types of antioxidants and synergists, the types of fatty acids, and the glycerol configuration, appear to play a major role in governing the resistance of an oil to oxidative changes. Crude halibut liver oils, even though more saturated than most lots of crude soup-fin shark liver oils, are generally less stable in regard t o vitamin A destruction than t,he latter type of oils. This is undoubtedly due mainly to :i higher concentration or to more potent vitamin .1antioxidants in the soup-fin shark liver oils. Many unpublished data accumulated in the course of this norlr establish that crude soiip-fin shark liver oils possess appreciably higher phosphorus contents (phosphatides) than do crude halibut liver oils. This may account for some of the stability differences, since the greater quant,ity of phosphatides would act synergistically with other natural antioxidants present in the soup-fin liver oils to further enhance their stability. The rate and extent of peroxide formation is, however, greater in the soup-fin shark liver oils. This behavior is unquestionably due, in part a t least, to the greater degree and type of unsaturation present in the soup-fin shark liver oils. -1 comparison of curves 1 and 3 of Figure 14 illustrates this point. The individual peroxide and percentage of vitamin A destruct,ion values obtained on the crude and carbon-treated fish liver oils exposed to air a t 34.5' C., n-ith and Tyithout the added tocopherols or tocopherols plus lecithin, are presented graphically in Figures 1 to 5 and in Table 111. RELATION OF VIT.4bIIN A DESTRUCTION TO PEROXIDATIOh-
Curves 1 and 2 of Figures 1 and 3 show the relation between the destruction of vitamin A and the rate and extent of peroxidation in the blank samples of crude and carbon-treated soup-fin shark C and halibut liver oil A, respectively. Here again, as reported previously, in fish liver oils treated with 20% carbon (3'1, the
Vol. 39, No. 2
percentage of vitamin A decrease is directly proportional t o the peroxide value, up t o a point where 60 to 70Ycof the viramin A is oxidized. This behavior is due t o the absence of active vitamin A antioxidants in the treated oils. A-umerically, the peroxide number a t a given percentage of vitamin A destruction in the samples of crude and carbon-treated soup-fin shark liver oil C is considerably higher than i n comparable samples of t,he halibut liver oil A. The curves on the samples of carbon-treated soup-fin shark C and halibut liver oil h containing added synt,hetic a-tocopherol and synthetic a-tocopherol plus lecithin (Figures 1, 2, and 3) clearly establish that, a t the same percentage of vitamin A destruction, a much higher peroxide number is reached in the 01tocopherol samples, the increase i n peroxide number a t a given percentage of vitamin A destruction being related to the quantity of a-tocopherol added. This is illustrated by curves f and 4 of Figure 1, in the comparison of the crude soup-fin shark liver oil C: and a samplr of the same oil carbon-treated and containing 0.2570 added synthetic a-tocopherol. This comparison is made because these tn-o oils exhibit about the same vitamin A stability. For example, a t a value of 19% vitamin A destruction, peroxide numbers of 58 and 133, respectively, xvere obserred. At approximately 64% vitamin A destruction the peroxide numbers were 110 and 180, respectively. Similar results are recorded in Figure 3 on halibut liver oil A and in Table 111 on halibut liver oil B. The data i n Table 111 show further that, i n crude halibut liver oil B, pure synthetic and pure natural a-tocopherol exhibit idcntical effects. Curves 5 and 7 in Figure 1 and curves 6 and 8 in Figure 2 show that a similar relation exists between peroxidation and vitamin A destruction i n samples of carbon-treated soup-fin shark liver oil C containing added synthetic a-tocopherol (0.50% by weight) ; this is compared x i t h a mixed tocopherol concentrate (Vegol, 40%, 1.25% by veight, or 0.50% tocopherol), with and without added lecithin. At the same percentage of vitamin -4. destruction, however: a somewhat lower peroxide value \vas noted in the Pamples containing the mixed tocopherol concentrate. Figure 4 shows that, when O , l O ~ oby rveight of synthetic atocopherol is added to crude soup-fin shark liver oil C and halibut, liver oil A, considerably higher peroxide numbers are reached than. in the blanks at similarlewls of vitamin A destruction. The data in Table I11 and Figure 5 establish that, a t a givcn percentage of vitamin A destruction, the peroxide number of the crude soup-fin shark liver oils A and B and the crude halibut liver oil B containing 0.10% of added 8- or ?-tocopherol arc not appreciably higher, and in the majority of instances are lov--cr,than under similar circumstances with the crude oils. This was ccrtainly the case when 0.10% y-tocopherol vias added to carbontreated halibut liver oil B (Table 111). When 0.50% $- and ytocopherol were added to crude soup-fin shark liver oil B (Figure 5) and to crude halibut liver oil B (Table 111),the peroxide values increased rapidly to a high maximum and then leveled off, essentially as observed 11-itha-tocopherol (Figure 1). At different percentages of vitamin A destruction in the various oils in Table 111, lon-er peroxide values were obPerved using y- as compared to 0tocopherol. I n all instances when p- and y-tocopherols werc used, alone or with lecithin, the peroxide values were lower a t a given loss of vitamin A than under comparable conditions n-ith 01tocopherol. Lecithin a t a 1.0% concentration in the sample of carbontreated soup-fin shark liver oil C (curves 2 and 3, Figure 2) had no effect on the relation betn-een peroxidation and vitami12 A destruction. However, Klien 0 , 5 0 ~ lecithin o was used in combination vc-ith synthetic 0-tocopherol in this same oil, R lo~\-erpc~roxide number was observed a t various stages of vitamin A destructioii. I n crude halibut liver oil B (Table 111),n-ith and vithout 0 . 1 5 added synthetic or natural a-tocopherol, lecithin a t a 17; conccntration lowered appreciably the peroxide number a t a given percentage of vitamin A destruction. Similar results were obtained with carbon-treated halibut liver oil B containing O.lnc
INDUSTRIAL AND ENGINEERING CHEMISTRY
February 1947
229
TABLE 111. STABILITY OF FISH LIVEROILS DAYSAT 34.5' C. Antioxidant None
1% Lecithin 0.1% syn. p-tocopherol
1% lecithin 0.1% syn. @-tocopherol 0.1% syn. y-tocopherol
1% lecithin 0.1% syn. 7-tocopherol
'0.1%nat. y-tocopherol
1% lecithin 0.1% nat. y-tocopherol
1
0
2
3
4
6
15
18
21
24
28
32
46.4 17.8 35.8 9.8 64.7 18.1 43.0 11.1 48.7 11.5 35.9 6.7 52.1 12.5 35.1 7.9
51 9 28.4 42.1 18.8 59.6 26.8 50.0 18.3 60.4 20.8 51.4 13,l 62.1 20.2 50.1 12.5
52.1 38.3 44.1 28.5 61.7 37.2 53.7 24.8 60.7 30.7 54.2 20.9 63.7 28.4 54.9 19.4
58.8 57.3 39.9 38.5 61.5 45.4 52.3 30.9 58.8 37.1 55.1 26.8 60.9 37.3 56.5 26.6
64.6 78.4 44.3 50.3 60.7 52.2 51.2 41.0 51.3 49.3 53.6 33.8 56.9 47.5 51.0 34.0
...
, . .
45.3 73.3 60.7 65.3 52.1 51.7 57.8 64.4 48.3 49.2 60.3 62 3
. . . . .
. . . .
11
7
13
SOUP-FINSHARKLIVER OIL A Peroxide No. % vit. A destr. Peroxide P.; 0. % vit. A destr. Peroxide No. % vit; A destr. Peroxide No. % vit. A destr. Peroxide No. % vit. A des%. Peroxide h o . vit. ' . A destr. eroxide No. % vit. A destr. Peroxide h 0 . % vit. A destr.
p
0
...
8:7
0
0
6'
..
..
24.4 . .
'
6.0
. . I
..
9.4 . . 3.5 19.4 . . 5.2 . . 5.0 , .
0
I .
0' 6' 6' 0'
0
17.4 1.2 3.8
'
0
, .
32.1 8.9 21.1 4.4 40.0 7.3 21.1 7.2 33.3 6.1 18.3 0.8 34.3 4.8 15.2 2.4
... ... ...
...
...
48.6 65.7
... .. .. . .
I
.
43.3 61.1
HALIBUTLIVEROIL B None
1% lecithin 0.1% syn. @-tocopherol 1% lecithin 0.1% syn. @-tocopherol 0.5% syn. @-tocopherol 0.1% nat. y-tocopherol
1% lecithin 0.17 nat. y-tocopherol 0.5d nat. y-tocopherol 0.5% nat. y-tocopherol 1% lecithin 0.1% syn. a-tocopherol
1% lecithin 0.1% syn. a-tocopherol 0.1% nat. a-tocopherol 1% lecithin 0.1% nat. a-tocopherol
Peroxide No. yo vit. A destr. Peroxide No. 70vit. A destr. Peroxide No. vit. ' . A destr. eroxide No. % vit. A destr. Peroxide No. vit. A destr. Peroxide h o . % vit. A destr. Peroxide No. 70vit. A destr. Peroxide No. % vit. A destr. Peroxide No. % vit. A destr. Peroxide No. yo vit. A destr Peroxide KO. % vit. A destr. Peroxide No. % vit. A destr. Peroxide No. vit. A destr.
p
12.4 16.3 4.4 8.7
2.5 ... 2.5 I
.
, .
..
.
2.5
...
2.5
... 1.9 ... 2.5 ...
.. ..
. . . . .
.. .. ..
51.4 , . 82.7 54.2 6i:3 38.7 53.2 33.1 , . 26.7 . . 73.2 . . 13.9 . . 45.2 , . 35.0 . . 28.4 . . 21.8 . . 42.2 . . 8.7 . . 13.5 . . 2.1 . . 77.6 . . 38.7 , . 45.4 , . 18.2 . . 71.7 . . 35.1 . . 48.9 , . 18.4 , .
.. ..
..
.. .. .. .. I .
..
2.5
, .
. . I
.. ..
1.9
...
2.1 .. : .5
,.
..
..
...
..
2.5
...
, .
..
.. .. ..
.. ..
..
..
2.5
..
,..
r0
2.5
Peroxide No. % vit. A destr. Peroxide No. vit. A destr. Peroxide No. 9%vit. A destr. Piroxide No. % vit. A destr. Peroxide No. % vit. -4destr. Peroxide No. % vit. A destr. Peroxide No. % vit. ii destr. Peroxide No. % vit. A destr.
2.4 22.6 22.9 i : 5 17.9 . . . 19.6 2.5 . .
...
. . . .
. . . .
. . . . 68.0 .
. . . . .
. . . . .
65.4 . . . 36.3 33.9 39.8 52.4 175.0 193.5 23.6 32.6 57.5 58.8 42.7 65.6 31.4 35.0 36.6 52.6 110.0 122.6 14.3 19.0 54.0 63.7 3.7 6.0 132,8 172.0 86.7 . . . 52.0 72.6 40.6 78.0 123,l 163.5 85.5 . . , 50.5 64.4 38.6 62.5
. . . . .
. . . . .
..
. . . . . . . . . .
,
.
. . . . . . . . . .
.,
. . . . . . . . . . , ,
.
, .
. . . .
. . . . .
.
. . . . . . . . .
. . . . .. ,.. ,..
2ij:o 229:o 2ii:o 2i6:o 39.0 49.0 59.8 . . .
. . . . . . . . .
. . . . . . . . .
.,
. .,
150:o 31.8 76.0 11.5
15s:o
,
,
.
.
159:o 17i:o 35.4 42.0 59.5 87.8 89.9 95.3 11.0 14.2 20.3
. . . . . . . . . ... . . . . . . . . . . . . . .
. . . . . . . . . . . . . . . . . .
.. ,
... ... ...
.,.
.,
... ... , . .
.
. . . . . . . . .
. . . . . . . . .
CARBON-TREATED HALIBUTLIVEROIL B None
Ir0lecithin 0.1% syn. p-tocopherol 1% lecithin 0.1% syn. @-tocopherol ~0.1%nat. -,-tocopherol
1% lecithin 0.1% nat. y-tocopherol
0.1% nat. a-tocopherol
1% lecithin 0.1% nat. a-tocopherol
...
2.5
..
..
... .. 2.5 . . ... .. 2.5 ... 2.5 . . ...
..
2.5
...
.
I
natural a-tocopherol and 1% lecithin. Lecithin also had a lowering effect, but to a greater degree than when a-tocopherol was used, on the peroxide number in the oils containing added pand ?-tocopherol a t different percentages of vitamin A loss (Figure 5 and Table 111). At a 5.0% concentration (curves 6 and 7 , Figure 2), lecithin, when tested in association with 0.5% added synthetic a-tocopherol i n carbon-treated soup-fin shark liver oil, also lowered t,he peroxide number a t the same percentage of vitamin A destruction, especially after about 10% of the vitamin A mas destroyed. EFFECT OF STORAGE ON VITAMIN A DESTRUCTION
The main object for adding antioxidants to vitamin L4materials such as fish liver oils is to inhibit vitamin A destruction t o the fullest practical extent. Figures 6-10 and Table I11 summarize the data on the percentage of vitamin A destroyed during storage at 34.5" C. of the various samples studied. The percentage of vitamin A destroyed in t,he carbon-treated oils is directly proportional to time up to the point where 60-70y0 of the potency is destroyed; this indicates the absence of active antioxidants. The crude soup-fin shark liver and the halibut liver oils were considerably more stable than the same oils after carbon treatment. The former were more stable than the crude halibut liver oils.
, .
...
.. ..
...
46:8 25.1 27.1 22.9 36.8 11.6 18.4 9.0 61.4 20.0 41,l 11.2
67.3 55.4 33.2 33.7 51.1 42.7 23.7 22.5 125.5 82.7 47.2 27.5
, . .
...
. . . . . . . . . . . . . . . . . .
. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
... 97.3 78.7 37.8 58.8 26.0 65.1 29.6 45.5
50.9 78.4 38.1 81.7 11.2 54.6
85.4
. . . . . . . . . . . . . . .
,..
, . .
.. .. ... ... ... , . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 72,4 . . . . . . . . . . . . . . . ,..
...
,..
,..
, . .
(
0
All the oils containing the added a->p-, and y-tocopherols or these tocopherols plus lecithin exhibited greater vitamin B stability than comparable blanks. KO appreciable difference (Table 111) was noted between the stabilizing effect of the synthetic and natural forms of CY- and ?-tocopherol, with and without 1%added lecithin, a t a 0.17, concentration in crude halibut liver oil B and in crude soup-fin shark liver oil A. In all the oils examined containing added a-, p-, or y-tocopherol in a concentration of 0.10% and greater, the tocopherols were effective antioxidants for vitamin 8. Under the condition employed, the gamma form was only slightly more effective than the beta form for inhibiting vitamin A destruction. Both forms are slightly more potent antioxidants than the alpha form for vitamin A. in fish liver oils. Lecithin enhanced the antioxidant effect,iveness of the pure tocopherols and of the mixed tocopherol concentrate in preventing vitamin A loss. This is particularly noticeable in the oils containing the lower percentage (0.10%) concentration of the pure tocopherols and also during the early stages of vitamin A destruction. Lecithin at 5% had a greater enhancing effect than 0.5% lecithin on 0.507, synthetic a-tocopherol in the carbon-treated soup-fin shark liver C oil (curves 6 and 7 , Figure 6). Additional unpublished data on the use of lecithin in combination with the different tocopherol isomers indicate that the enhancing or syn-
INDUSTRIAL AND ENGINEERING CHEMISTRY
230
I80
/
200
r
HOURS AT 34,5°C.
I
I
400
1
HOURS A T 345°C
HOURS AT 34.5"C. Figure 13
Figure 11 Curve 1. Curve 2. Curve 3. Curve 4. Curve 5. Curve 6. Curve 7.
Crude soup-fin shark liver oil C Carbon-treated shnrk liver oil C Same an 2, T 0.109'~Synthetic a-tocopherol Same 0.25% synthetic a-tocopherol Same 0.50% synthetic u-tocopherol 1.00$70 synthetic a-tocopherol Same 1.25% Vegol (40%) Same
+ ++ +
Figure 12 Curve 1. Crude soup-fin shark liver oil C Curve 2. Carbon-treated shark liver oil C Curve 3. Same as 2 1%lecithin 0'50% lecithin and 0.05% s y n t b t i c u-tocopherol Curve 4. Same 0.50% lecithin and 0.10% synthetic a-tocopherol Curve 5. Same Curve 6. Same 0.50% lecithin and 0.50% synthetic u-tocopherol 5.8% lecithin and 0.50% synthetic a-tocopherol Curve 7. Same 0.50% lecithin and 1.25% Vepol (40%) Same Curve 8.
+ +
+ + + +
Vol. 39, No. 2
Curve 1. Crude halibut liver oil A Curve 2. Carbon-treated halibut liver oil A 0.10% synthetic a-tocopherol Curve 3. Same as 2, 0.50% synthetic a-tocopherol Curve 4. Same
+ +
Figure 14 Curve 1. Crude soup-fin shark liver oil C 0.10% synthetic a-tocopherol Curve 2. Same as 1, Curve 3. Crude halibut liver oil A 0.1% synthetic a-tocopherol Curve 4. Same as 3,
+ +
Figure 15 Curve 1. Curve 2. Curve 3. Curve 4.
Crude soup-fin shark liver oil B Same as 1 0.1% natural y-tocopherol Same 0:1% natural y-tocopherol and 1.0% lecithin 0.5% natural y-tocopherol Same
+ + +
February 1947
INDUSTRIAL AND ENGINEERING CHEMISTRY
ergistic effect of lecithin is not proportional to its concentration and also depends upon the type of fish liver oil used. Lecithin alone exhibits no protective action for the vitamin A a t a 1.0% concentration in the carbon-treated soup-fin shark liver oil C (curves 2 and 3, Figure 6 ) and in the crude and carbon-treated halibut liver oil B (Table 111). However, a t a 1.0% concentration in crude soup-fin shark liver oil A, lecithin increased the vitamin A stability of crude soup-fin shark liver oil A, 50.3% for the lecithin-containing oil as compared to 78.4% vitamin A loss for the blank oil after 24 days of storage. These results indicate that this particular oil probably contains an appreciable amount of a natural antioxidant which is capable of acting synergistically with lecithin. If the oil does contain such an antioxidant, it is not predominsntly a-tocopherol, because of the slow rate and type of peroxidation displayed. The analysis of crude soup-fin shark liver oil A (Table I) shows that i t contains appreciably more phosphorus than the other oils studied. When 0.50% 7-tocopherol and l.0701ecithin were added to crude halibut liver oil B, the percentage loss of vitamin A after 32 days was 20.3 as compared to 66.3% for the original oil after 7 days (Table 111). All the oils containing added lecithin and either a-, p-, or 7-tocopherol exhibited greater vitamin A stability than analogous blanks containing only the tocopherols. A comparison of curves 1 and 4 in Figure 6 shows that vitamin A in the sample of carbon-treated shark liver oil containing O.%OJ, of added synthetic a-tocopherol still decreased slightly faster than i n the untreated crude oil. Curves 1and 3 of Figure 8 indicate that the addition of 0.1% synthetic a-tocopherol to carbon-treated halibut liver oil A stabilizes the vitamin A in the oil to about the same degree as it does in the crude oil. When 0.1% natural a-tocopherol is added to carbon-treated halibut liver oil B, it stabilizes the oil to a greater degree than the crude oil (Table 111). or-Tocopherol is a more effective antioxidant for vitamin A in the carbon-treated halibut liver oil A than in the carbontreated soupfin shark liver oil C. A concentration of 0.10% of p- and y-tocopherol in carbon-treated halibut liver oi' B (Table 111) stabilized the oil appreciably more than when 0.10% of atocopherol was used and t o a greater degree than the crude oil. Generally speaking, tocopherols alone or with lecithin are more effective as antioxidants for inhibiting vitamin A destruction and peroxidation in oils of low unsaturation than in oils of high iodine value. RATE AND EXTENT OF PEROXIDATION
Figures 11to 15 and Table I11 contain data on the rate of peroxidation. It has generally been thought that, in order to prevent vitamin A losses due to oxidation, peroxidation had to be inhibited nearly completely. The present data show that, when relatively high percentages (0.25% and up) of either a-, p-, or ytocopherol are added to soup-fin shark or halibut liver oils, relatively high peroxide values (100 and up) can be tolerated with only minor losses of vitamin A (around 10% and less). In nearly all crude fish liver oils and certainly in carbon-treated fish liver oils stored under the condit,ions of this study, when the peroxide value reaches a numerical figure of around 70 or 80 substantially all of the vitamin A content of the oils has been destroyed. Both @- and y-tocopherol at a 0.10% concentration are more efficacious antioxidants of the peroxide-inhibiting type for fish liver oils than a-tocopherol. The results on 7-tocopherol establish that i t is more effective than @-tocopherolfor inhibiting peroxidation. At a 0,50% concentration in the oils studied, p- and ytocopherol failed t o inhibit peroxidation even though they stabilized effectively the vitamin A contents of the oils, The three tocopherol forms were more potent in controlling peroxidation in the halibut liver oils than in the soup-fin shark liver oils. During the latter stages of vitamin A destruction, in the fish liver oils containing O . l O ~ oof added 7-tocopherol alone and i n combination with lecithin, some irregularities in peroxide values were observed (Table
231
111). A somewhat similar behavior was observed in soup-fin shark liver oil A containing 0.10% of added p-tocopherol alone and with lecithin. I n these instances the peroxidation proceeded smoothly until the peroxide value reached a rather low maximum -around 30 to 60 depending on the particular oil-and then i t either remains more or less static or decreases. This same behavior was observed in unpublished work on certain types of solvent extract fractions from shark liver oils particularly. No such peroxidation behavior was observed with a-tocopherol under analogous circumstances. Lecithin at 0.50% concentration exhibited a slight depressing effect on the peroxidation in carbon-treated soup-fin shark liver oil C containing added a-tocopherol. In crude and carbon-treated halibut liver oil B (Table 111)with and without added a-, @-, or y-tocopherol,lecithin a t a 1.0% concentrationdecreased theperoxidation rate appreciably. Lecithin also considerably lowered the peroxidation rate of crude soup-fin shark liver oil A. I n the oils containing added a-tocopherol alone, peroxides accumulated faster than in the respective crude oils. KO similarity was observed between the rate and extent of peroxidation i n the lots of crude and carbon-treated soup-fin shark and halibut liver oils and in the same oils containing added a-,p-, or y-tocopherol alone or with lecithin. SUMMARY
Stability studies of vitamin A in soup-fin shark and halibut liver oils, with and without added synthetic and natural 01- and ytocopherol and synthetic 8-tocopherol alone and plus lecithin, establish that the tocopherols are effective vitamin A antioxidants a t relatively high concentrations. However, a t the same concentration the tocopherols, either alone or associated with lecithin, are not effective antioxidants for inhibiting peroxidation in such oils. Lecithin alone a t 1.0% concentration in crude halibut liver, carbon-treated halibut liver, and soup-fin shark liver oils exhibited little or no activity for inhibiting vitamin A destruction. Under similar conditions andwhen associatedwith thepure tocopherols, lecithin had a depressing effect on the rate of peroxidation in such oils. The effectiveness of the pure tocopherols as antioxidants for vitamin Awas enhanced by using them in association with lecithin. In all instances much higher peroxide numbers were observed in oils containing synthetic or natural added or-tocopherol than in those without them a t the same percentage of vitamin A destruction. Pure 8- and y-tocopherol a t a 0.10% concentration in the oils studied decreased slightly the rate of peroxidation; however, a t a level of 0.50% the peroxidation rate was increased the same amount as when a-tocopherol was used. No appreciable difference could be noted in the oils examined between the antioxidant effectiveness of the pure synthetic and the natural forms of or- and y-tocopherol in inhibiting peroxidation and vitamin A destruction. When 0.1% synthetic a-tocopherol was added to samples of crude soup-fin shark and halibut liver oil, the rate of vitamin A destruction was decreased; however, the rate of peroxidation was markedly increased. Pure or-, 8-, and 7-tocopherols, alone or in combination with lecithin a t relatively high concentrations, do not act as antioxidants in preventing peroxidation in crude or carbon-treated soup-fin shark or halibut liver oils, whereas they are effective antioxidants for vitamin A in such peroxide-containing oils. The relations observed between peroxide formation and vitamin A destruction in crude and carbon-treated soup-fin shark and halibut liver oils, with and without a-,B-,and ytocopherols, indicate that the antioxidative behavior of the crude fish liver oils examined is not due predominantly to any tocopherols which may be present. ACKNOWLEDGMENT
The author expresses his thanks to H. J. Konen, now a t Schenley Distillers, Inc., to Jane A. Nagle for many helpful suggestions and assistance in accumulating the data presented in
232
INDUSTRIAL AND ENGINEFRING CHEMISTRY
Vol. 39, No. 2
(6) Olc!ott, H. S.,arid Plmmerson, 0. I{., I b i d . , 59,1008 (1937). (7) O l c O t t , H. 8., alld Mattill. H. A , , Ibid.. 58,1627 (1936). (8) Rienienschneider, 12. IT., Turcr, J., and A u l t , W. C., Oil h Soap, 21, 90 (1944). (9) Robeson, C. E., and Baster, J. G.. J . Am. Chem. Soc., 65, 940
this paper, and to Merck & Company, Inc., and Distillation ProdUCtS, Inc., for the samples of pure tocopherolsused in the course of this investigation.
(1943).
LITERATURE CITED
(10) Simons, E. J., B u s t o n , L. O., and Colman, H. B., IND.EXG. CHEX.,32, 706 (1940). (11) Sn-ift. C. E.. Rose, IT. G., and Janiieson, G. S . , Oil h Soap, 19, 176 (1942).
(1) Berenbluni, I., and C h a i n , E., Biochem. J . , 32, 295 (1938). (2) Bird, 0. D., Publication 298, Univ. Microfilms, .Inn Arbor, hIich., 1941. (3) Buxton, L. O.,IND. EXG.CHEX, 34, 1486 (1942). (4) Golumbic, C., Oil & Soap, 20, 105 (1943). (5) Golumbic, C., a n d Mattill, H.A., J . Am. Che77~.SoC., 63, 1279 (1941).
I’RESEXTED i n part before the Division of Biologioal ChemistrJr a t the I O D t h AIeeting of t h e . ~ M E R I C A \ TC H C ~ I I CSOCIETY, AL .Itlantic City, N.J.
BATCH DISTILL
APH I
MELVIN NORD Sord and Company, Inc., Keyport,
N T H E theory of simple batch distillation of binary solutions the following, k n o m as Rayleigh’s equation, occurs:
.Y.J . ,
I
whcre Li and L, arc the number of moles (or pounds) of original charge and residue, respectively, z1 and 2 2 are the mole fractions (or weight fractions) of light component in the original charge and t’he residue, and CY is the relative volatility of the light component, which is assumed to be constant over the concentration range considered’. In order to solve Equation I, lor rl or x2, trial-and-error solution is necessary. I t is possible, however, to construct a nomograph which will solve Equation 1 without trial and error. Rayleigh’s equation may be rewritten as follows:
L,
z=
or
(3)
If
Therefore, from a graph of lnf(z) against 2, for lines of constant one can subtract In j(z2) from in f(z1)nomographically to find the ratio L2/L1, or the fraction of the initial charge which remains in the still. This is demonstrated in Figure 1. As an example of the use of the nomograph, assume 21 = 0.80, N = 2.0, and LZ/Ll = 0.044; then z 2 is found as follows: At 0.80 on the z scale, go vertically up t o QI = 2.0, then horizontally over t o the scale marked 2 1 , scale 3. Connect,the point on the 21 scale Tvith point 0.044 on the &/L’ scale and continue t o scale 3, marked z2. Then draw a line from this point through the key point an$ continue t o scale 1, then horizontally to CY = 2.0 and vertically down to the x scale. The result is 0.36, which is the desired value of 22. This path is indicated by directed lines on the nomograph. The nomograph can be used in the reveme manner to find z‘, or it may be used to find & ~ / Lwhen I z1 and z~ are known. If more precise results are desired than can be obtained with a nomograph of moderate size, the values obtained in this way may be regarded as the first approximation in a trial-and-error solution. If it is desired to take into account variations in a,the nomograph may be applied over small enough int’ervals of concentration so that constant values of OL may be used over each interval. CY,
x,
X Z
0
.2
4
.6
.8
Figure 1. Simple Batch Distillation Nomograph
1
Perry, J. H., Chemical Engineers’ Handbook, 2nd ed., p. 1383 (1941).
