US2380410A - Fat-soluble vitamin ester concentration process - Google Patents

Description

Patented July 31, 1945 FAT-SOLUBLE vrrnm: ESTER OONCENTRATIGN PROCESS Loran 0. Button, Newark, N. 3., assignor to National Oil Products Company, Harrison, N. J a corporation of New Jersey 9 Claims.

This invntlon relates to a process for producing vitamin concentrates, and more particularly to an improved-process for preparing concentrates of fat-soluble vitamin esters.

One of the well known methods of preparing fat-soluble vitamin concentrates involves the saponiilcation of the fatty material present in fatsoluble vitamin containing oils and recovering Application July 13, 1942, erial No. 450,761

the unsaponlilable matter from the saponliled mass by a suitable solvent. This process has several disadvantages among which there may be. mentioned the concentration with the vitamins of many undesirable tastes and odors present in the original oil, thus producing a vitamin concentrate which is rather objectionable in these respects. Furthermore, such concentrates'consist primarily of vitamin alcohols, which are less stable towards oxidative deterioration than are the correspondingnatural vitamin esters.

In order to produce concentrates of the vitamins in their natural ester forms, the process which is disclosed and claimed in copending application of Dombrow, Serial No. 343,558, filed July 2, 1940, was developed. .In that process a vitamin-containing oil is contacted with an organic polar solvent which'is substantially miscible with fatty materials at temperatures substantially above room temperature and partially immiscible therewith at temperatures substantially below room temperature; and thereafter cooling the solventoil solution to a temperature substantially below room temperature whereby a substantial portion of the oil separates from the solution. The soluble fraction which remains in the polar solvent has a much higher fat-soluble vitamin content than the portion which is insoluble in the solvent. The vitamins present in the highly potent fraction are in their naturally occurring ester form. Such concentrates which are much more potent than the original oil, have found wide uses in the vitamin field. However, the concen-. trates which are produced by that process do have certain disadvantages. The solvent extraction, in addition to concentrating the fatsoluble vitamins. also concentrates therewith a large portion of the coloring matter originally present in the oil, as well as the free fatty acids and also many of the malodorous and ill-tasting constituents of the oil. A further disadvantage of this process is that many oils contain certain constituents, e. g., free fatty acids, saturated and slightly saturated triglycerides including stearins,

. etc, which tend to entrap the solvent which is being employed to extract the vitamins. As a result thereof, the eihciency of the extraction process is somewhat decreased and the yield of vitamin A is much lower than what it would be if no solvent were entrapped in the oil immiscible with the solvent at the low temperature. Also. because'of the great bulk of the material being treated, large volumes of solvent and much equipment are required. p

In my copending application, Serial No. 450,- 757, filed concurrently herewith, there is disclosed and claimed a process for producing highly'potent concentrates of esters of fat-soluble vitamins by selectively saponifying (hydrolyzing) a fat-soluble vitamin-containing marine oil. By carrying out the saponiflcation of a marine oil under the conditions disclosed in said application, it is possible to hydrolyze up to about 60% to 60% of v the glycerides in the oil without splitting substantial amount of the fat-soluble ylt esters, whereby the resulting unsaponifled iraction contains the vitamins in their ester form.

It is the object of this invention to provide an improved process for the preparation of fatsoluble vitamin ester concentrates.

A further object of the invention is to providean lmprovedprocess for the preparation oi con= centrates of vitamin A and D esters.

Another object of the invention is to provide concentrates of fat-soluble vi which are substantially devoid of free fatty acids and undesirable tastes and odors. 7

Another object of the invention is to provide highly potent concentrates of fat-soluble vitamins wherein the vitamins are substantially all present in the naturally occurring ester at.

Other objects of the invention will in part be obvious and .will in part appear hereinafter.

I have discovered that the above and other objects of the invention may be realized by saponifying 10% to of the glycerides in a fatsoluble vitamin-containing marine oil, separating the unsaponifled fraction from the saponified matter, contacting the unsaponlfled fraction with an organic aliphatic polar solvent which is characterized by being miscible withthe unsaponifled fraction at temperatures above room temperature and partially ble therewith at temperatures below room temperature separating the solvent solution of the vitamins thus extracted from the remainder of the unsaponiiled fraction.

Contrary to the general belief, I have found that in partially saponifying' a iish or fish liver oil the saturated and slightly unsaturated triglycerides, hereinafter referred to as "saturated glycerides," are selectively saponified, l. e. these saturated glycerides take precedence over unsaturated glycerides and other saponifiable matter, except free fatty acids, in order of reaction with alkali or alkaline agents. This .result may be due to the fact that in marine oils the unsaturated triglycerides have higher molecular weights than the more saturated glycerides present therein. Moreover, as shown in the curve comprising the drawing in the aforesaid application filed concurrently herewith, the vitamin esters do not react with alkali to any substantial degree until the oil has been saponified beyond about 75% or 80%.

The invention accordingly comprises the several steps and relation of one or more of such steps with respect to each of the others, and the product possessing the features, properties and the relation'of elements, which are exemplified in the following detailed disclosure, and the scope of the invention will be indicated in the claims.

In carrying out the process of this invention,

any fat-soluble vitamin-containing marine oil may be used includin inter alia, cod liver oil. shark liver oil, tuna liver oil, halibut liver oil, mackerel liver oil, ling cod liver oil, sole liver oil, spear fish liver oil, sword fish liver oil, herring oil, sardine oil, whale liver oil, seal liver oil, etc.

As aforementioned, the first step in the process of a fat-soluble vitamin-containing mafiilne oil. The degree of saponification of the oil ay be varied depending upon the type of concentrate desired. Good results are obtained when an amount of alkali sumcient to saponify from about 1 10% to about 60% of the glyceridespresent ,in the fatty material is employed, the preferred range, however, being from about 30% to about 5 "/6.

' The partial saponification eliminates from the unsaponified oil substantially all the free fatty acids and a large amount of the coloring matter and undesirable tastes and odors originally present therein and at least a major portion of the saturated glycerides. In carrying out' the parof the invention involves partial saponiilcation 1 tial saponification step by means of an alkaline agent it is preferred to employ potassium hydroxide, it being understood, however. that any suitable alkaline agent may be used in lieu of potassium hydroxide such as sodium hydroxide,

sodium carbonate, potassium carbonate. etc.

' In order to produce the most satisfactory and most highly potent ester concentrate, it is preferred to carry out the saponiflcation of the fatty material in such a manner that the glycerides will be selectively hydrolyzed, leaving the vitamin ester unattacked. The principal conditions which are necessary to obtain the desired selectivityare fully disclosed in my copending application, Serial 6 No. 450,157, referred to supra. However, in general, conditions which are conducive to selective saponiflcatton are relatively low temperatures,

e. 8.. room temperature, carrying out the saponifion the weight of the oil. In most saponlflcation procedures,- a saponification catalyst, e. g.. isopropanol, is employed to accelerate the reaction. Elimination of the use of such catalysts or a reduction of the amount thereof to about to 1% based on the weight of the oil when no nonpolar solvent is used will also aid in making the saponification more selective. It is to be understoodthat it is not necessary to carry out the saponification step in the presence of either a catalyst or solvent medium;

After the fatty material has been partially saponified, the unsaponified fraction, hereinafter termed the semi-concentrate," is separated from the soaps and glycerine by any suitable means, e. g., solvent extraction, centrifugation, etc. If the saponification has been carried out in the presence of an inert solvent, and it is desired to separate the unsaponified material from the soaps by means of solvent extraction, it is preferred to extract with the same solvent that was employed during the partial saponification step. The semiconcentrate may readily be recovered from the solvent solution thereof by distillation under reduced pressure. The semi-concentrate will be found to be devoid of free fatty acids and much lighter in color than the original oil and substantially free of undesirable tastes and odors.

The next step in the process of the invention is to contact the semi-concentrate with a solvent which is characterized by being miscible with the semi-concentrate at temperatures about room temperature, and partially immiscible therewith at temperatures below room temperature. The solvent employed in this step may be selected from a large number of aliphatic solvents found to be useful as a result of extensive experimentation; the choice of the solvent will depend to some extent upon the properties of the semiconcentrate to be treated, as will become more evident from the detailed description hereinafter .given. Results haveindicated that the solvents preferably employed are members of well recognized chemical classes; it has also been found that thenumber ofcarbon atoms in the solvent to be used is a particularly important factor in determining the availability thereof for use in the practice of this invention. The following table sets forths,the classes of solvents which have been found to be particularly useful in the practice of this invention:

Table Solvents falling in the classes above listed are all 7 liquid polar aliphatic organic compounds having me the properties of being miscible with marine oils or partially saponified fractions thereof at temperatures above room temperature, 1. e. 20 to 25 C., and partially immiscible therewith at temperatures substantially below room temperature, and it has been found that solvents falling within this class of compounds may be used in the practice of this invention. In addition it will be noted that the preferred solvents possess relatively low freezing points. i

min A at temperatures above'this point.

In order to more fully illustrate the nature of thesolvents which may be employed, a partial list thereof is herewith given; it is to be understood, however, ,that this list is not intended -to be complete, but is merely illustrative of the solbe used to a predetermined temperature at which the concentrate to be added will substantially completely dissolve in the solvent, and then add- '1 ing the concentrate to the solvent with agitation,

vents which may be employed. Thus it has been vents belong to that class of aliphatic organic compounds which have the properties of being miscible with marine oils and partially saponified marine oils at temperatures above room temperature and partially immiscible therewith at temperatures substantially below room temperature; furthermore, it will be noted that the majority of these solvents have relatively low freezing points. v

Occasionally it may be found that certain of the solvents hereinabove mentioned may be too miscible with some semi-concentrates whi may be treated bythis invention to effect a separation of, highly potent vitamin fractions therefrom; thus, for example, acetone is too miscible with some semi-concentrates to accomplish the purthe operation being carried out in an inert gas atmosphere.

The solution of the semi-concentrate in the solvent prepared as hereinabove described may then, in accordance with the process of the in F vention, be permitted to cool so as to effect a separation of the solution of the highly potent vita-v min extract from the remainder of the concentrate. Th temperature to which the solution is cooled may vary from about'room temperature to It has been found.

as low as 70 C. or lower. however, that it is preferable to cool the solution with agitation to temperatures somewhat below about 0 0;," e. g. invthe neighborhood of about -18 C. Upon cooling the solution separates into two layers. One layer consists chiefly of the po tion of thesemi-concentrate insoluble in the The vitamin consolvent at low temperatures.

a tent of this fraction is much less than that of the semi-concentrate, Thisfraction has a considerably lighter color than the semi-concentrate and also has lost the characteristic'odor possessed by many fat-soluble vitamin-containing oils.

The solvent layer obtained upon cooling the solution may be filtered and then treated to reposes of this invention; However, this condition I may be-easily corrected by diluting the solvent either with a small-amount of water or with a liquid aliphatic organic solvent relatively immiscible with marine. oils.' in general it may be; said that the effect of diluting any of the above more immiscible with marine oils or fractions thereof, so that if difficulty is encounteredin efmove the solvent therefrom, e. g. by vacuum dis- .tillation, whereby a concentrate is recovered having a vitamin content far in excess of the amount contained in the original semi-concentrate; the percentage increase in vitamin potency may be,

anywhere between about 100% and about 600%, the actual increase varying with the vitamin potency of the semi-concentrate.

The process of the invention may also be carried out by continuously contacting the semi-con- 40" centrate' with one ofthe above polar solvents at a solvents with water willbe to render the solvents fectingv proper separation of the highly potent In carrying. out the extraction of the semi-concentrate with the polar solvent. the-unsaponifled,

fraction is first mixed with the particular-solvent semi-concentrate to solv nt.in the mixture may vary widely; preferably th ratio of solvent to the concentrate should be greaterthan one and inmost cases mixtures containing between about 2% and about 25% of the concentrate are most suit-" able.- This mixture may then be heated until the semi-concentrate or the greater part thereof is dissolved in the solvent. The temperature to which the ture is .heated may vary widely de-,

pending upon the nature of the ingredients contained in the mixture; ingeneral it may be stated that it is inadvisable to hea fat-soluble vitamincontaining-materials to tem herftisirmes-in excess of 175 C.-because of the relative bility of vita- It is preferred to form the solution. of semi-concen trate in the solvent-by first heating the solvent to a to be employed.- The relative proportion of the 4 relatively low temperature, e. g. around 0 0.; this method of operation effects a continuous extraction of a highly potent vitamin fraction from the concentrate. However. it is preferred to employ th batch method hereinabove described.

The resulting high potency concentrate may, after the removal of solvent therefrom, be further extracted with methanol and/or ethanol to free the concentrate of the vitamin alcohols present therein thus providing a highpotency'concentrate substantially free of vitamin alcohols.

For a fuller understanding of the nature and objectsof the invention, reference should be had to the following examples which are given merely -to further illustrate the invention and are not to be construed ina limiting sense, all parts given being byweight, and all-vitamin potencies being expressed inU'. S, P. units:

Examzjle 200 parts of shark liveroil having a potency of 103,000 units of vitamin A per gram, 100 parts of ethylene-dichloride and 6 parts of isopropanol were admixed with sufficient 46% aqueous potassium hydroxide to saponify about 50% of the sharkliver oil. The mass was stirred at room temperature in the presence of N2 gas for about thirty minutes whereby a super-solvented soap,

emulsion was'formed. The mass'was allowed to stand overnight, and then 700 more parts of ethylene dichloride were added thereto, and the mass I heated to .a temperature of about 60 to C.

adjusted to about 25%.

The solvent-soap mass; was cooled to room'temperature and th moisture content of the soap The solvent separated out from the soap mass carrying with it the unsaponifled oil. The soap mass was extracted two more times with ethylene dichloride employing four parts of solvent to one part of soap. The

partially saponifled mass, the unsaponifled oil solvent extracts were combined, filtered, and the- H from the oil immiscible with the solvent at the low temperature. No difliculty was encountered in separating the solvent layer from the oil layer during any of the three extractions. The solvent extracts were combined and the solvent removed therefrom. A light golden yellow colored extract having a potency of 406,000 units of vitamin A per gram and containin over 60% of all the 'vitamin A which was present in the original oil was obtained. The taste and odor of the extract was greatly superior to the taste of extracts prepared bysolvent extracting the original oil.

A sample of the original oil was similarly ex; tracted three times with isopropanol. However, only about 37% of the vitamin A contained in the oil was recovered and the potency of the combined extracts was only about 215,000 units of vitamin A. These results clearly illustrate the great superiority of the process of the present invention.

ExampZeII A crude shark liver oil was partially selectively saponified similarly as in Example I except that about 60% of the oil was saponified. Th following analyses of the crude oil and the unsaponifled oil recovered from the partially saponified mass illustrate the improvement in quality obtained by means of the partial saponiflcation and also explain to a certain extent why the unsaponified oil may be so much more readily extracted with a polar solvent to produce improved concentrates.

' Partially Crude sapomfied oil on Percent free fatty acid 5. 1 0. 1 Color (Lovibond):

Red 49 10. 3 18 154 The oil recovered from the partially saponifled mass was much more satisfactory with regard to taste and odor than the crude oil, and, as may be seen by referring to the analyses, was much lighter-in color than the crude oil. The higheriodine value and lower cloud and pour points of.

the oil recovered from the partially saponifled mass show that stearins and other'high melting components which interfere with the solvent extraction have been eliminated- Furthermore the recovered oil is almost completely devoid of free fatty acids, which acids would tend to lessen the efliciency of the solvent extraction if they were present.

After recovering the unsaponifled oil from the 76 was extracted with isopropanol similarly as in Example I except that only two extractions were made. No dimculty at all was encounteredu'n making these extractions; only about 4% of the solvent was entrained in that part of the oil im- -miscible with thesolvent at the low tempera-- ture. A fraction having a potency ofv 268,000 units of vitamin A per gram was obtained. as compared with a potency of 103,000 units of vitamin A per gram when a fraction was prepared similarly by extracting the crude oil itself with isopropanol. Furthermore the total yield of vitamin A when the crude oil was extracted directly was only one-third that obtained by extracting the unsaponifled oil recovered from the partially 'saponified mass. Whereas about 96% of the solvent was recovered along with the extract when the unsaponified oil recovered from the partially saponified mass was extracted with solvent, only about 24% of the solvent was recovered with the extracts when the crude oil was extracted, the remaining 76% of the solvent being entrained in the residue of the crude oil.

Example m .Tuna liver oil having a potency of 73,000 units of vitamin A per gram was partially saponified about 50% similarly as in the previous examples.

Samples of the original oil and of the'oil recov- I ered from the partially saponiiied mass were extracted with isopropanol as in the other examples. In the first case, the extract recovered had a potency of only 137,000 units of vitamin A but in the latter case an extract having a potency of 239,000 units of vitamin A was obtained. The greatest dimculty of separation of the solvent from the oil and the poorest recovery of solvent accompanied by a consequently poorer vitamin recovery was encountered with the extraction of the original oil directly.

Example IV 200 parts of shark-liver oil containing 103,000 units of vitamin A per gram, 9% of the vitamin being in the alcohol form and the remaining 91% being in the ester form, were mixed with 100 parts of ethylene dichloride and 6' parts of isopropanol. While stirring in the presence of N2 gas, suflicient 45% aqueous KOH (39.4 gms.)

to saponify. about 50% of the oil was added and the stirring continued for about 15 minutes. The

thick soap mass was then insulated and left at room temperature for about 12 hours. 800 parts of ethylene dichloride were added and the solvent-soap mass heated to about 60 C. while stirring for about .15 minutes. Suflicient water was added at this stage to increase the moisture content of the soap to about 24%. The entire solvent-soap mass was then cooled to about 25 C. and the stirring ceased. After about 15 minutes, the lower solvent layer was drawn-oil. and the soap extracted 6 more times as before. The solvent extracts were combined, filtered and the solvent evaporated under reduced pressure at a low temperature in the presence'of N: gas. The recovered partially saponifled oil'contained 187,000 units of vitamin A per gram and was substantially. devoid of free fatty acids and was fighter in color and free of the original fishy taste and odor.

parts of the partially saponified oil con taining'18'7,000 units of vitamin A per gram were mixed with 360'parts of 99% isopropanol and the mixture warmed to-about 36' C. to give a clear solution. The isopropanol-oil solvent was then cooled gradually to about 18 C. After 1 2 hours, the upper solvent layer was removed and filtered. The insoluble solid oil fraction was then extracted again with a similar quantity of isopropanol. The clear filtrates were combined and the isopropanol removed under reduced pressure in the presence of N2 gas. The resulting concentrate contained 358,000 units of vitamin A per gram and possessed only a slightly flshy taste and odor. The free fatty acid content was 0.28% as compared to 1.5% for the original crude shark liver oil.

10 parts of the high P tency isopropanol soluble concentrate containing 358,000 units of vitamin A per gram were mixed with 40 parts of 95% ethanol and the mixture warmed to about 50 C..

The mixture was then cooled gradually to about '25 C. and filtered. The insoluble oil layer was removed and extracted similarly 3 more times. The combined ethanol filtrates were removed and the ethanol evaporated at a low temperature under reduced pressure in the presence of N2 gas. The recovered oil possessed a fishy taste and odor and contained 475,000 units of vitamin A per gram. The vitamin A was largely in the alcohol form and represented 10% of the amount present in the starting'material. The ethanol insoluble oil layer was freed of entrained solvent in the manner just described. The recovered oil was light yellow in color, completely without taste and odor, and contained 340,000 units of Vitamin A per gram, the vitamin A being in the ester form and comprising 90% of that present in the original isopropanol soluble concentrate.

It is evident from the above description and examples that my invention provides a highly improved process for producing highly potent ester concentrates of the fat-soluble vitamins. The vitamin ester concentrates produced by this process are much more potent than such concentrates produced by extracting the original oil directly and are also much more potent than the concentrates obtained merely by selectively saponiiying the oil. to 60% of the fatty material is eliminated be- Furthermore, since fro 3 fore carrying out the solvent extraction, a much u lesser quantity of fatty material has to be ex:

tracted than when treating the original oil. Not

only does this result in an increased saving in the amount of solvent which is required, but also the amount of eq pment necessary t prepare the concentrates is greatly reduced from that which would'be required if the concentrates were prepared by extracting the original oil. The fact that that portion of the semi-concentrate which is immiscible with the solvent at the low temperature does not tend to entrap the solvent, as in the case of the original oil, further lessens .the amount of solvent which is required and at the same time results in greatly increased efiiciency in extracting the vitamins. The highly desirable characteristics of the concentrates which are obtained as regards color, taste and odor, further add to the desirability of the process as compared to prior processes for producing yitamin concentrates.

.While the only specific method of eflecting' saponification (hydrolysis) of the esters con-' tained in marine oils has involved the use or an alkaline agent, other methods to effect splitting of the esters may be employed within the purview of this invention and particularly enzyme sa ponification (hydrolysis);

Since certain changes may be made in carrying out the above process without departing from the scope of the invention, it isintended that all matter contained in the abovedescription shall be interpreted as illustrative andnot in alimiting sense.

Having described my invention, what Iclaim as new and desire to secure by Letters Patent is:

. l. Aprocess of producing a fat-soluble vitamin ester concentrate, which comprises saponifying 10% to 60% of the glycerides present in afatsoluble vitamin-containing marine oil, separating the unsaponified fraction from the saponified;

matter, contacting the unsaponified fraction with an organic aliphatic polar solvent which is characterized by being miscible with the unsaponifled fraction at temperatures above room temperature and partially "immiscible therewith at temperatures below room temperature, cooling the mass to a temperature within the range of 0 C. to -70 C. to cause layer formations and separating the solvent layer containing the extracted vitamins from the solvent-insoluble portion of said fraction.

2. A process of producing a fat-soluble vitamin ester concentrate, which comprises saponii'ying 30% to 50% of the glycerides present in a-fatsoluble vitamin-containing marine oil, separating the unsaponified fraction from the saponified matter, contacting the unsaponifled fraction with an prganic aliphatic polar solvent which is characterized by being miscible with the unsaponified 10% to 60% of the glycerides present in a fatsoluble vitamin-containing marine oil, separating the unsaponified fraction from the saponified matter, contacting the unsaponified fraction with an organic aliphatic polar solvent which is characterized by being miscible with the unsaponifled fraction at temperatures above room temperature and partially immiscible therewith at temperatures below room temperature, cooling the mass to a temperature within the range of 0 C. to

70"- C. to cause layer formations, separating the solvent layer containing the extracted vitamins from the solvent-insoluble portion of said fraction, removing the solvent from the extracted vitamins and removing the vitamin alcohols therefrom by extraction with a solvent selected from the group consisting of methanol and ethanol.

4. A process of producing a fat-soluble vitamin ester concentrate, which comprises saponifying 10% to 60% of the glyceride's present in a fish liver oil, separating the unsapqnified fraction from the saponified matter, contacting the unsaponifled fraction with an organic aliphatic polar solvent which is characterized by being miscible with the unsaponified fraction at tem peratures above room temperature and partially immiscible therewith at temperatures below room temperature, cooling the mass to a temperature within the range of 0? C. to -70 C. to cause layer formations and separating the solvent layer containing the extracted vitamins from the sol- 5 vent-insoluble portion of said fraction.

5. A process of producing a fat-soluble vitamin ester concentrate, which comprises saponifying 30% to 50% of the glycerides present in a fish liver oil, separating the unsaponified fraction from the saponifled matter, contacting the un- I saponified fraction with an organic aliphatic polar solvent which is characterized by being miscible with the unsaponified fraction at temperatures above room temperature and partially immiscible therewith at temperatures below room temperature, cooling the mass to a temperature within the range of C. to -70 C. to cause layer formations and separating the solvent layer containing the extracted vitamins from the solvent-insoluble portion of said fraction.

6. A process of producing a fat-soluble vitamin ester concentrate, which comprises sapomfying 10% to 60% of the glycerides present in a fatsolu-ble vitamin-containing marine oil, separating the unsaponified fraction from the saponified matter, contacting the unsaponified fraction with isopropanol, cooling the mass to a temperature within the range of 0 C. to 'l0 C. to cause layer formations and separating the isopropanol layer containing the extracted vitamins from the moving the isopropanol'trom the extracted vitamins and removing the vitamin alcohols therefrom by extraction with a solvent selected from the group consisting of methanol and ethanol.

8. A process of producing a fat-soluble vitamin ester concentrate, which comprises saponfiying 10% to 60% of the glycerides present in a fatsoluble vitamin-containing marine oil, separating the unsaponified fraction from the saponifled matter, contacting the unsaponified fraction with isopropanol, cooling the mass to a temperature within the range of 0 C. to 70 C. to cause layer formations, separating the isopropanol layer containing the extracted vitamins from the isopropanol-insoluble portion of said fraction, re-

moving the isopropanol from the extracted vita mins and removing the vitamin alcohols therefrom by extraction with a solvent selected from the group consisting of methanol and ethanol.

9. A process of producing a fat-soluble vitamin ester concentrate, which comprises dissolving in a fish liver oil 25% to 75% (based on the weight of the oil) of a-non-polar solvent, saponifying 10% to of the glycerides present in said oil, separating the unsaponified fraction from the saponifled matter, contacting the unsaponified fraction with isopropanol, cooling the mass to a temperature within the range of 0 C. to C.

tracted vitamins and removing the vitamin alcohols therefrom by extraction with a solvent selected from the group consisting of methanol and 'ethanol.

LORAN O. BUXTON.