CA2031945A1 - Low-saturate edible oils and transesterification methods for production thereof - Google Patents
Low-saturate edible oils and transesterification methods for production thereofInfo
- Publication number
- CA2031945A1 CA2031945A1 CA002031945A CA2031945A CA2031945A1 CA 2031945 A1 CA2031945 A1 CA 2031945A1 CA 002031945 A CA002031945 A CA 002031945A CA 2031945 A CA2031945 A CA 2031945A CA 2031945 A1 CA2031945 A1 CA 2031945A1
- Authority
- CA
- Canada
- Prior art keywords
- fatty acid
- oil
- fatty acids
- unsaturated
- transesterification
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Abandoned
Links
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- 238000000034 method Methods 0.000 title claims abstract description 60
- 238000004519 manufacturing process Methods 0.000 title abstract description 16
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- FMMOOAYVCKXGMF-UHFFFAOYSA-N linoleic acid ethyl ester Natural products CCCCCC=CCC=CCCCCCCCC(=O)OCC FMMOOAYVCKXGMF-UHFFFAOYSA-N 0.000 description 1
- 239000000944 linseed oil Substances 0.000 description 1
- 235000021388 linseed oil Nutrition 0.000 description 1
- 150000002632 lipids Chemical class 0.000 description 1
- 239000007791 liquid phase Substances 0.000 description 1
- 150000004668 long chain fatty acids Chemical class 0.000 description 1
- 229910052751 metal Inorganic materials 0.000 description 1
- 239000002184 metal Substances 0.000 description 1
- 230000005012 migration Effects 0.000 description 1
- 238000013508 migration Methods 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- IJDNQMDRQITEOD-UHFFFAOYSA-N n-butane Chemical compound CCCC IJDNQMDRQITEOD-UHFFFAOYSA-N 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- JDLQRNRWXLXOMN-UHFFFAOYSA-N octadecanoic acid;urea Chemical compound NC(N)=O.CCCCCCCCCCCCCCCCCC(O)=O JDLQRNRWXLXOMN-UHFFFAOYSA-N 0.000 description 1
- 235000014593 oils and fats Nutrition 0.000 description 1
- 229940049964 oleate Drugs 0.000 description 1
- 239000003960 organic solvent Substances 0.000 description 1
- 239000001301 oxygen Substances 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- 150000002942 palmitic acid derivatives Chemical class 0.000 description 1
- SECPZKHBENQXJG-FPLPWBNLSA-N palmitoleic acid group Chemical group C(CCCCCCC\C=C/CCCCCC)(=O)O SECPZKHBENQXJG-FPLPWBNLSA-N 0.000 description 1
- 230000036961 partial effect Effects 0.000 description 1
- 150000002978 peroxides Chemical class 0.000 description 1
- 239000003208 petroleum Substances 0.000 description 1
- 229920001568 phenolic resin Polymers 0.000 description 1
- 239000005011 phenolic resin Substances 0.000 description 1
- 239000011148 porous material Substances 0.000 description 1
- 239000002244 precipitate Substances 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 238000005086 pumping Methods 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 230000036632 reaction speed Effects 0.000 description 1
- 230000035484 reaction time Effects 0.000 description 1
- 230000009257 reactivity Effects 0.000 description 1
- 238000004064 recycling Methods 0.000 description 1
- 230000001603 reducing effect Effects 0.000 description 1
- 238000007670 refining Methods 0.000 description 1
- 238000002407 reforming Methods 0.000 description 1
- 230000001105 regulatory effect Effects 0.000 description 1
- 238000003303 reheating Methods 0.000 description 1
- 230000000717 retained effect Effects 0.000 description 1
- 230000000630 rising effect Effects 0.000 description 1
- 235000012045 salad Nutrition 0.000 description 1
- 235000014438 salad dressings Nutrition 0.000 description 1
- 238000009738 saturating Methods 0.000 description 1
- 239000000344 soap Substances 0.000 description 1
- 239000007790 solid phase Substances 0.000 description 1
- 108010079522 solysime Proteins 0.000 description 1
- 238000001179 sorption measurement Methods 0.000 description 1
- 230000006641 stabilisation Effects 0.000 description 1
- 238000011105 stabilization Methods 0.000 description 1
- 238000006276 transfer reaction Methods 0.000 description 1
- 230000009261 transgenic effect Effects 0.000 description 1
- 125000005314 unsaturated fatty acid group Chemical group 0.000 description 1
- 238000005292 vacuum distillation Methods 0.000 description 1
- 238000009489 vacuum treatment Methods 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
- 239000010457 zeolite Substances 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P7/00—Preparation of oxygen-containing organic compounds
- C12P7/64—Fats; Fatty oils; Ester-type waxes; Higher fatty acids, i.e. having at least seven carbon atoms in an unbroken chain bound to a carboxyl group; Oxidised oils or fats
- C12P7/6436—Fatty acid esters
- C12P7/6445—Glycerides
- C12P7/6458—Glycerides by transesterification, e.g. interesterification, ester interchange, alcoholysis or acidolysis
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23D—EDIBLE OILS OR FATS, e.g. MARGARINES, SHORTENINGS, COOKING OILS
- A23D9/00—Other edible oils or fats, e.g. shortenings, cooking oils
-
- C—CHEMISTRY; METALLURGY
- C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
- C11C—FATTY ACIDS FROM FATS, OILS OR WAXES; CANDLES; FATS, OILS OR FATTY ACIDS BY CHEMICAL MODIFICATION OF FATS, OILS, OR FATTY ACIDS OBTAINED THEREFROM
- C11C3/00—Fats, oils, or fatty acids by chemical modification of fats, oils, or fatty acids obtained therefrom
- C11C3/04—Fats, oils, or fatty acids by chemical modification of fats, oils, or fatty acids obtained therefrom by esterification of fats or fatty oils
- C11C3/08—Fats, oils, or fatty acids by chemical modification of fats, oils, or fatty acids obtained therefrom by esterification of fats or fatty oils with fatty acids
-
- C—CHEMISTRY; METALLURGY
- C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
- C11C—FATTY ACIDS FROM FATS, OILS OR WAXES; CANDLES; FATS, OILS OR FATTY ACIDS BY CHEMICAL MODIFICATION OF FATS, OILS, OR FATTY ACIDS OBTAINED THEREFROM
- C11C3/00—Fats, oils, or fatty acids by chemical modification of fats, oils, or fatty acids obtained therefrom
- C11C3/04—Fats, oils, or fatty acids by chemical modification of fats, oils, or fatty acids obtained therefrom by esterification of fats or fatty oils
- C11C3/10—Ester interchange
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P7/00—Preparation of oxygen-containing organic compounds
- C12P7/64—Fats; Fatty oils; Ester-type waxes; Higher fatty acids, i.e. having at least seven carbon atoms in an unbroken chain bound to a carboxyl group; Oxidised oils or fats
- C12P7/6436—Fatty acid esters
- C12P7/6445—Glycerides
- C12P7/6472—Glycerides containing polyunsaturated fatty acid [PUFA] residues, i.e. having two or more double bonds in their backbone
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- Oil, Petroleum & Natural Gas (AREA)
- General Chemical & Material Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
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- Microbiology (AREA)
- Health & Medical Sciences (AREA)
- Biochemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
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- General Health & Medical Sciences (AREA)
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Abstract
LOW-SATURATE EDIBLE OILS AND
TRANSESTERIFICATION METHODS FOR PRODUCTION THEREOF
ABSTRACT
Transesterified vegetable oils having a very low saturated fatty acid content and batch, cocurrent and countercurrent enzymatic transesterification methods for preparing such oils.
TRANSESTERIFICATION METHODS FOR PRODUCTION THEREOF
ABSTRACT
Transesterified vegetable oils having a very low saturated fatty acid content and batch, cocurrent and countercurrent enzymatic transesterification methods for preparing such oils.
Description
CASE 47505 .~ Q3~ ~
LOW-8ATURATE ~D~BL~ OIL8 AND
TRAN~E8TERIFICATION METROD8 FOR PRODUCTION $HE~EOF
Backaround of the Inventio~
The present inventlon is directed to methods for preparing edible triglyceride vegetable oils having a very low level of saturated fatty acid components, and to enzymatic transesterification methods for producing such low-saturate, edible oil products.
Ed~ble oils and fats typically primarily comprise various fatty acid triesters o glycerol with the structure of the fatty acid moieties and their distribution on the glycerol backbone determining the physical characteristics of the oil or fat. The specific types of fatty acids also play an important role in diet and health. Fats and oils in general are a rich source of energy in the diet and are important in the synthesis of membranes and other essential cell components. Moreover, dietary fatty acid content may potentially be controlled to affect physiological characteristics such as serum cholesterol levels. For example, studies of normcholesterolemic men has shown that a dietary decrease in saturated fatty acids may have more of an effect in lowering serum cholesterol ~X~ys, "Prediction of .Serum Cholesterol R~ponse to Change in Fats in the Diet", Lancet, 2:959-962~ than an increa~e ln polyunsaturated fatty acids.
Natural vegetable oil triglycerides typically contain substantial amounts of esterified saturated fatty acids. For example, soybean oil may typically contain about 14-16 weight percent of esterified saturated fatty acids, and natural canola oil may contain about 5-8 weight percent of esterified saturated fatty acids. Intermediate carbon chain length (i.e., C12-C16) dietary satuxated fatty acids, notably lauric, myristic and palmitic acids, have been reported in the medical literature AS being a more ~ignificant factor in the increase of plasma cholesterol than stearic acid, which has been reported to ~ ~3~
have minimal or even reducing effects on cholesterol levels [nEffect of Dietary Stearic Acid on Plasma Cholesterol and Lipoprotein Levels~, Bonanome, et al., New England Journal of Medicine, Vol. 318, 1244-1271 (1988?]. Soybean oil and canola oil typically contain, respectively, over 10 percent and over 3 percent by weight of esterified intermediate chain length saturated fatty acids, primarily palmitic acid. Accordingly, it would be desirable to economically produce or manufacture triglyceride oils having very low levels of saturated fatty acids, and particularly low levels of lauric, myristic and palmitic acids. Most food products prepared from vegetable oils having less than about 3.5 weight percent of esterified saturated fatty acids may be regarded as substantially free of such fatty acids for regulatory purposes.
Potentially, the modification of vegetable oils to produce low-saturate oil products could be carried out by dehydrogenation, chemical transesterification, enzymatic transesterification or genetic selection and modification.
~owever, dehydrogenat~on processe~ are not available for selectively dehydrogenating esterified saturated fatty acids of vegetable oils. The use of somaclonal variation a~ a means of selection for genetic varlation in plant parts which may produce triglycerldes with specific desirable fatty acid~ may be used, but problems may arise in the stabilization of the desired trait in future generations. Recombinant DNA technigues might be used to lncrease the production o~ an oil of predetermined composition, but this is a very complex task in difficult or presently unknown areas of plant lipid biosynthesis.
tStumpf, "Biosynthesis and Function of Plant ~ipids", Am.
Soc. Plant Phy~iol., pp. 1-15, 1983]. For example, fatty acid synthesis has many potential rate limiting enzymes as well as proteins, such as acetyl-CoA carboxylase, ACP-acetyl transferase, 3-oxoacyl ACP synthase, ACP malonyl transferase and acyl carrier protein. Modifications of specific triglyceride synthesis entail changes in fatty ~ ~ ~ 3 ~
;
aeid eomposition dependent on acyl ACP thioesterase and various desaturase complexes, as well as aeyltransferase enzymes whieh attaeh the fatty aeid moieties to glyeerol.
Thus, to isolate the rate limiting enzymes/proteins and their eorresponding genes to ereate transgenic plants wh$eh specifically express them in tissues for storage purposes poses substantial technical problems.
Chemical and enzymatic transesterification are well known for modifying the fatty acid composition or distribution of triglyeeride oils. Chemical transesterification is based on tho use of a chemical catalyst sueh as sodium methoxide or a aodium metal to promote the migration of the fatty aeid moieties between glyceride moleeules, to produee a random distribution of the fatty aeid moieties.
Enzymatie transesterifieation of triglycerides may al80 be used to modify the characteristies and/or composition of triglycerides. Sueh processes may be used for selective interchange under rolatively mild reaction conditions. For example, vegetable oils may be transesterified with a fatty aeid or monoester to produee a variety of end produets as deseribed in U.S. Pàtents 4,268,527; 4,426,991: 4,275,011S 4,472,503 and U.X.
Applieation 2,199,397.
Extraeollular mierobial lipases aro generally of three type~, depending upon their speeifieity. One group of lipases is generally nonspeeifie, both as regards the position on the glyeerol moleeule wh$eh is hydrolyzed or ot-rifled, and the nature o~ the fatty aeid releasod or esterified. Dopending on tho reaetion eonditlons, sueh lipases eatalyze the nonseleetive hydrolysis, alcoholysis and/or esterifieation ~ineluding transesterification) of fatty aeid triglycerides. The lipases produeed by Candid~
çylindraeae, also known as ~ rugosa (Benzonana, G. and S.
Esposito, Bioehim~ Biophy. Aeta. 231:15 (1971)), Corynebaeterium aenes, (Hassing, G.S., Ik~l 242:381 ~d 3 ~ ~ L~ ,~
(1971)), and Staphylococcus ~y~y~, (Vadehra, D.V., ~ipids 9:158 (1974)), are examples of 8uch nonspecific lipases.
A second group of lipases preferentially acts on the primary, 1- and 3- positions of the glycerol or triglyceride molecule. When a 1-,3- positionally specific lipase is used to catalyze the transesterification of a mixture of triglycerides or a mixture of triglyceride plus free fatty acid or monoester, the action of the enzyme is substantially confined to the 1- and 3- positions of the glycerol. The lipases of Rhizopus dele~ma~ and Mucor miehei such as described in U.S. Patent 4,798,793, are examples of 1-,3- specific lipases.
A third group of lipase6 has substantial selectivity for certain long chain unsaturated fatty acids having a Çi~ double bond at the 9- position from the carboxylate group of the fatty acid. Long chain saturated fatty acids, and unsaturated fatty ~cid estars without a double bond in the 9- position, are only slowly hydrolyzed in the presence of such lipases. Thus the esters of oleic, palmitoleic, linoleic and linolenic acids, all of which have a ÇL~ double bond in the 9- position, are preferentially hydrolyzed, esterified or transesterfied.
The presence of an additional double bond between the carboxyl group and the double bond in the 9- position makes fatty acid esters re~istant to the action o~ this llpa~e.
Triglycerides containing medlum chain saturated C10 and C8 fatty acids may exhibit some, albiet reduced, reactivity with such enzymes. Examples of such delta-9 æpecific lipases which preferentially act on long chain fatty acids containing a cl~ double bond in the 9 position are the lipase produced by the mold Goetrichum candidum [Macrae, A.R., in ~icrobi~l ~nzy~es a~q ~otechnol~qy, edited by W.M. Fogarty, Applied Science Publishers, London, 1983, p. 225, Jensen, R.G., I~LDids 9:149 (1974), Jensen, R.G., and R.E. Pitas, in ~Dids, edited by R. Paolette, G.
Porcellati and G. Jacini, Raven Press, New York, 1976, ~ ~ 3 ~
Vol. 1, p. 141], and the lipase produced by ~enicilliun cyclopium ~Glyceride Synthesis by Four Kinds of Microbial Lipase, Tsujisaka, et al.; Biochim. Biophys. Acta 489;
415-422 (1977)]. Such lipases will activate transesterification of unsaturated delta-9 fatty acid groups of glyceride oils, but do not affect the saturated acid components of the oils.
Enzymatic methods which may be used to reduce the saturated fatty acid content of vegetable oils to levels below about 3.5 weight percent, and total levels of intermediate chain length fatty acids below about 2 weight percent, would be desirable, and it is an ob;ect of the present invention to provide 6uch low-saturate vegetable oils. It is a further ob~ect to provide such methods which may be used to provide low-saturate oils having specific unsaturated fatty acid distribution, as well as low saturate edible oils having specific, nutritionally desirable properties of oleic, linoleic and linolenic acids. These and other ob~ects will become apparent from the following detailed description and the accompanying drawings.
DescriDtion of the Drawing~
FIGURE 1 is a process flow diagram for an embodiment of a single step batch or cocurrent continuous enzymatic transe~teriflcatlon reactlon method for producing a low saturate trlglyceride in accordance wlth ths present invention having less than 3 weight percent e~terified saturated fatty acids:
FIGURE 2 is a process flow diagram for a continuous countercurrent sQlective enzymatic reaction method utilizing a supercritical countercurrent gas stream for producing a low saturate triglyceride oil having les6 than 3 weight percent of e~terified 6aturated fatty acids;
FIGURE 3 i6 a process flow diagram for another method of preparing a low-saturate edible oil by countercurrent enzymatic transesterification reaction, utilizing n countercurrent subcritical gas; and
LOW-8ATURATE ~D~BL~ OIL8 AND
TRAN~E8TERIFICATION METROD8 FOR PRODUCTION $HE~EOF
Backaround of the Inventio~
The present inventlon is directed to methods for preparing edible triglyceride vegetable oils having a very low level of saturated fatty acid components, and to enzymatic transesterification methods for producing such low-saturate, edible oil products.
Ed~ble oils and fats typically primarily comprise various fatty acid triesters o glycerol with the structure of the fatty acid moieties and their distribution on the glycerol backbone determining the physical characteristics of the oil or fat. The specific types of fatty acids also play an important role in diet and health. Fats and oils in general are a rich source of energy in the diet and are important in the synthesis of membranes and other essential cell components. Moreover, dietary fatty acid content may potentially be controlled to affect physiological characteristics such as serum cholesterol levels. For example, studies of normcholesterolemic men has shown that a dietary decrease in saturated fatty acids may have more of an effect in lowering serum cholesterol ~X~ys, "Prediction of .Serum Cholesterol R~ponse to Change in Fats in the Diet", Lancet, 2:959-962~ than an increa~e ln polyunsaturated fatty acids.
Natural vegetable oil triglycerides typically contain substantial amounts of esterified saturated fatty acids. For example, soybean oil may typically contain about 14-16 weight percent of esterified saturated fatty acids, and natural canola oil may contain about 5-8 weight percent of esterified saturated fatty acids. Intermediate carbon chain length (i.e., C12-C16) dietary satuxated fatty acids, notably lauric, myristic and palmitic acids, have been reported in the medical literature AS being a more ~ignificant factor in the increase of plasma cholesterol than stearic acid, which has been reported to ~ ~3~
have minimal or even reducing effects on cholesterol levels [nEffect of Dietary Stearic Acid on Plasma Cholesterol and Lipoprotein Levels~, Bonanome, et al., New England Journal of Medicine, Vol. 318, 1244-1271 (1988?]. Soybean oil and canola oil typically contain, respectively, over 10 percent and over 3 percent by weight of esterified intermediate chain length saturated fatty acids, primarily palmitic acid. Accordingly, it would be desirable to economically produce or manufacture triglyceride oils having very low levels of saturated fatty acids, and particularly low levels of lauric, myristic and palmitic acids. Most food products prepared from vegetable oils having less than about 3.5 weight percent of esterified saturated fatty acids may be regarded as substantially free of such fatty acids for regulatory purposes.
Potentially, the modification of vegetable oils to produce low-saturate oil products could be carried out by dehydrogenation, chemical transesterification, enzymatic transesterification or genetic selection and modification.
~owever, dehydrogenat~on processe~ are not available for selectively dehydrogenating esterified saturated fatty acids of vegetable oils. The use of somaclonal variation a~ a means of selection for genetic varlation in plant parts which may produce triglycerldes with specific desirable fatty acid~ may be used, but problems may arise in the stabilization of the desired trait in future generations. Recombinant DNA technigues might be used to lncrease the production o~ an oil of predetermined composition, but this is a very complex task in difficult or presently unknown areas of plant lipid biosynthesis.
tStumpf, "Biosynthesis and Function of Plant ~ipids", Am.
Soc. Plant Phy~iol., pp. 1-15, 1983]. For example, fatty acid synthesis has many potential rate limiting enzymes as well as proteins, such as acetyl-CoA carboxylase, ACP-acetyl transferase, 3-oxoacyl ACP synthase, ACP malonyl transferase and acyl carrier protein. Modifications of specific triglyceride synthesis entail changes in fatty ~ ~ ~ 3 ~
;
aeid eomposition dependent on acyl ACP thioesterase and various desaturase complexes, as well as aeyltransferase enzymes whieh attaeh the fatty aeid moieties to glyeerol.
Thus, to isolate the rate limiting enzymes/proteins and their eorresponding genes to ereate transgenic plants wh$eh specifically express them in tissues for storage purposes poses substantial technical problems.
Chemical and enzymatic transesterification are well known for modifying the fatty acid composition or distribution of triglyeeride oils. Chemical transesterification is based on tho use of a chemical catalyst sueh as sodium methoxide or a aodium metal to promote the migration of the fatty aeid moieties between glyceride moleeules, to produee a random distribution of the fatty aeid moieties.
Enzymatie transesterifieation of triglycerides may al80 be used to modify the characteristies and/or composition of triglycerides. Sueh processes may be used for selective interchange under rolatively mild reaction conditions. For example, vegetable oils may be transesterified with a fatty aeid or monoester to produee a variety of end produets as deseribed in U.S. Pàtents 4,268,527; 4,426,991: 4,275,011S 4,472,503 and U.X.
Applieation 2,199,397.
Extraeollular mierobial lipases aro generally of three type~, depending upon their speeifieity. One group of lipases is generally nonspeeifie, both as regards the position on the glyeerol moleeule wh$eh is hydrolyzed or ot-rifled, and the nature o~ the fatty aeid releasod or esterified. Dopending on tho reaetion eonditlons, sueh lipases eatalyze the nonseleetive hydrolysis, alcoholysis and/or esterifieation ~ineluding transesterification) of fatty aeid triglycerides. The lipases produeed by Candid~
çylindraeae, also known as ~ rugosa (Benzonana, G. and S.
Esposito, Bioehim~ Biophy. Aeta. 231:15 (1971)), Corynebaeterium aenes, (Hassing, G.S., Ik~l 242:381 ~d 3 ~ ~ L~ ,~
(1971)), and Staphylococcus ~y~y~, (Vadehra, D.V., ~ipids 9:158 (1974)), are examples of 8uch nonspecific lipases.
A second group of lipases preferentially acts on the primary, 1- and 3- positions of the glycerol or triglyceride molecule. When a 1-,3- positionally specific lipase is used to catalyze the transesterification of a mixture of triglycerides or a mixture of triglyceride plus free fatty acid or monoester, the action of the enzyme is substantially confined to the 1- and 3- positions of the glycerol. The lipases of Rhizopus dele~ma~ and Mucor miehei such as described in U.S. Patent 4,798,793, are examples of 1-,3- specific lipases.
A third group of lipase6 has substantial selectivity for certain long chain unsaturated fatty acids having a Çi~ double bond at the 9- position from the carboxylate group of the fatty acid. Long chain saturated fatty acids, and unsaturated fatty ~cid estars without a double bond in the 9- position, are only slowly hydrolyzed in the presence of such lipases. Thus the esters of oleic, palmitoleic, linoleic and linolenic acids, all of which have a ÇL~ double bond in the 9- position, are preferentially hydrolyzed, esterified or transesterfied.
The presence of an additional double bond between the carboxyl group and the double bond in the 9- position makes fatty acid esters re~istant to the action o~ this llpa~e.
Triglycerides containing medlum chain saturated C10 and C8 fatty acids may exhibit some, albiet reduced, reactivity with such enzymes. Examples of such delta-9 æpecific lipases which preferentially act on long chain fatty acids containing a cl~ double bond in the 9 position are the lipase produced by the mold Goetrichum candidum [Macrae, A.R., in ~icrobi~l ~nzy~es a~q ~otechnol~qy, edited by W.M. Fogarty, Applied Science Publishers, London, 1983, p. 225, Jensen, R.G., I~LDids 9:149 (1974), Jensen, R.G., and R.E. Pitas, in ~Dids, edited by R. Paolette, G.
Porcellati and G. Jacini, Raven Press, New York, 1976, ~ ~ 3 ~
Vol. 1, p. 141], and the lipase produced by ~enicilliun cyclopium ~Glyceride Synthesis by Four Kinds of Microbial Lipase, Tsujisaka, et al.; Biochim. Biophys. Acta 489;
415-422 (1977)]. Such lipases will activate transesterification of unsaturated delta-9 fatty acid groups of glyceride oils, but do not affect the saturated acid components of the oils.
Enzymatic methods which may be used to reduce the saturated fatty acid content of vegetable oils to levels below about 3.5 weight percent, and total levels of intermediate chain length fatty acids below about 2 weight percent, would be desirable, and it is an ob;ect of the present invention to provide 6uch low-saturate vegetable oils. It is a further ob~ect to provide such methods which may be used to provide low-saturate oils having specific unsaturated fatty acid distribution, as well as low saturate edible oils having specific, nutritionally desirable properties of oleic, linoleic and linolenic acids. These and other ob~ects will become apparent from the following detailed description and the accompanying drawings.
DescriDtion of the Drawing~
FIGURE 1 is a process flow diagram for an embodiment of a single step batch or cocurrent continuous enzymatic transe~teriflcatlon reactlon method for producing a low saturate trlglyceride in accordance wlth ths present invention having less than 3 weight percent e~terified saturated fatty acids:
FIGURE 2 is a process flow diagram for a continuous countercurrent sQlective enzymatic reaction method utilizing a supercritical countercurrent gas stream for producing a low saturate triglyceride oil having les6 than 3 weight percent of e~terified 6aturated fatty acids;
FIGURE 3 i6 a process flow diagram for another method of preparing a low-saturate edible oil by countercurrent enzymatic transesterification reaction, utilizing n countercurrent subcritical gas; and
2~3~ 3 FIGURE 4 is a process flow diagram of another embodiment of a transesterification method for transesterifying an unsaturated fatty acid monoester with saturated fatty acid-containing triglyceride vegetable oil to provide a triglyceride oil having very low saturated fatty acid content.
DescriDtion of the Invention Generally in accordance with the present invention, methods are provided for manufacturing a triglyceride oil having less than 3.5 and preferably less than 3 weight percent of esterified saturated fatty acid, and desirably less than 2, and more preferably less than 1 weight percent of intermediate chain length esterified saturated fatty acid.
Various aspects of the present invention are also directed to low saturate liquid vegetable oil products having less than 3.5 weight percent, and preferably less than 3 weight percent saturated fatty acid moieties, and specific unsaturated fatty acid distribution, which liguid vegetable oil products have de-irable properties for uso in food products such as mayonnaise, margarine, table spreads or salad dres~ings. Such low saturate vegetable oil products will al~o desirably have less than about 2 and preferably less than 1 weight percent of intermediate chain saturated fatty acids. By n intermediate ¢haln ~aturated fatty acid~ i8 meant, lauri¢, myri~tic and palmlti¢ acids having carbon chain l-ngths of C12, C14 and C16, rQspe¢tively. As us~d herein, the weight percent of ~aturated or unsaturated ~atty acid moieties in a margarine oil or vegetabl~ oil glyceride composition is calculated based on the total weight of the fatty acids contained in the margarine oil. Also, as used herein, when referring to weight percent of onQ or more fatty acid moiQtiQs of a composition ~such as the weight percentagQ of C12-C16 fatty acids), the weight percent is calculated based on all of tho fatty acids of the compo~ition being fully 2 ~ 3 ~
hydrolyzed to free fatty acid. The weight percent of one or more species of fatty acid i8 then calculated as the weight percent of such one or more species based on the total weight of free fatty acids. AOCS official method Ce 1-62 (81) may be used to determine the weight percent of respective fatty acids of an oil or fat composition.
In accordance with various method aspects of the present invention, a high-unsaturate vegetable oil, such as canola (low erucic acid rapeseed) oil, high oleic safflower oil, high oleic sunflower oil, high linoleic safflower oil, ~ '~
~5~ ~ high linoleic sunflower,~soybëa~ oil or m'ixtures th'ereof, R~
,"~ t.!having from about 4 weight percent to about ~ a~d ~ Jn~
~L ~ typ~c~lly fro~--bou~ to bout-} weight percent Of ~D ~ h' / ~ 5~ ~Itl~ saturated fatty acids with respect to the total fatty ac~d content of the vegetable oil is provided for transesterification reaction. Canola oil is a particularly desirable starting material because it has a relatively low amount of saturated fatty acid content. ~he vegetable oil should best be refined and bleached oil which is substantially free of proteinaceous and other nonglyceride components which might poison or otherwise interfere with transesterification enzymes.
Canola oil typical~y comprises in the range of from about 5 to about 7 weight percent of ~aturated fatty acid moietie~. However, th- ~aturat-d fatty acids of canola oil ~and other vegetable oil8) i8 not randomly distributed with respect to the 1-, 2- and 3- positions of the glyceride molecules of the oil. A major portion of the saturated fatty acid content i5 located in the 1- and 3-primarv positions of the glyceride molecules of the canola oil. In this regard, canola oil typically may have an overall stearic acid content of about 1.5 - 2 weight percent based on the total fatty acid content of the oil, with from about 2 to about 3 weight percent of the fatty acid moieties in the 1- and 3- positions being stearic acid and less than about 1 weight percent of the fatty acids in 2 ~ $ - ~
the ~econdary 2- position being stearie acid. Similarly, canola oil may typically have ~n overall palmitic acid content of about 4 weight percent, with about 6 weight pereent of the fatty acid groups in the 1- and 3- positions being palmitic acid and less than about l weight percent of the fatty acid groups in the 2- position being palmitic aeid.
In accordance with the present methods, the saturated fatty acids of the primary l- and 3- positions are selectively removed, and replaced with unsaturated fatty acids by selective transesterification reaction.
This may be accomplished without substantially affecting the fatty acid distribution of the starting material oil in the 2- position. 8y "transesterification" is meant an exehange of fatty acid moiety or aeyl radical at a glyeeride oxygen or hydroxyl group, whieh ineludes interesterification and intraesterification.
In order to earry out sueh methods, the high unsaturate vegetable oil sueh as eanola oil is eombined with an unsaturated fatty aeid transesterifleation component selected from the group consisting of free fatty acids, fatty aeid monoesters of lower alkyl monohydrie aleohols (e.g., methanol, ethanol and propane), and mixtures thereof, whieh has less than 2 weight pereent, and preferably le~i than l weight pereent of s8turated fatty aeids to provld- a transesteri~leation blend. The unsaturated fatty aeid transesterlfieation eomponent should eomprise at least about 98 weight pereent of unsaturated fatty aeids having a ehain length Or from 12 to about 22 earbon atoms, and less than 2 weiqht pereent of saturated fatty aeids having a earbon ehain length in the range of from 12 to 18 earbon atoms based on the total fatty aeid eontent of the transesterifieation eompQnent. Desirably, th- fatty acid transesterifieation eomponent eomprises less than 0.75 weight pereent and preferably less than 0.5 weight pereent of intermediate ehain saturated fatty aeids, 2 ~ ~3 ~
g by weight, ~ased on the fatty acid content of the transes~erification component.
The high-unsaturate vegetable oil such as canola oil and the unsaturated fatty ac~d are combined in a weight ratio of vegetable oil to unsaturated fatty acid transesterification component in the range of from about 1:10 to about 4:1 in batch reaction processes (which may utilize multiple reaction steps), and typically in the range of from about 1:2 to about 2:1 in single stage batch or cocurrent reactions to provide a transesterification mixture. The ratio of reactants may be selected to provide a desired degree of substitution under the reaction conditions, to provide a low-saturate glyceride product having a selective level of 6aturated fatty acid content below about 3.5 weight percent, and preferably below 3 weight percent. Moreover, the ratio of reactants and the composition of the unsaturated fatty acid transesteri-fication component ma-~ be selected to provide specific compositions of the transesterified low saturate oil, in terms of unsaturated fatty acid composition. For example, nutritionally deæirable unsaturated fatty acids in appropriate levels and ratios have been identified such that an increase in the omega-3 to omega-6 ratio in the average diet could yield distinct health benefits. Natural triglycerides do not typically contain theso unsaturated acids in such proportions, but ~uch compositions may be provided in accordance with the present invention. Further in accordance with methods of the present invention, the transesterification mixture is contacted with a transesterification enzyme, which iB desirably a 1-, 3-specific transesterification enzyme such as an immobilized lipase from Muco~ miehe1 as described in U.S. Patent 4,798,793 issued January 17, 1989, which i~ incorporated by reference herein. The transesterification mixture is contacted with the immobilized enzyme under time ~nd temperature conditions for 6ubstantially eguilibrating the ~3~
fatty acid content in the 1- and 3- positions of the glyceride component, with the fatty acid transesteri-fication components of the reaction mixture. The enzymatic transesterification reaction produces a transesterified triglyceride component and a transesterified fatty acid component. The reaction time may range from about 0.5 hour to about 100 hours, depending upon the concentration and activity of the lipase and the temperature of the reaction mixture. The transesterification reaction will desirably be carried out at a temperature in the range of from about 20- C. to about 80- C., and more preferably in the range of from about 40- C. to about 70- C. By "substantially eguilibrate" is meant that the transesterification reaction is at least about S0 percent complete, and preferably at least 90 percent complete. ~ower equilibrium transesterification conditions (e.g., 50-90 percent equilibrated) may be utilized to increase the reaction speed and or reduce the amount of enzyme used, but this increases the unsaturated fatty acid required and increases the separation step processing requirements.
Following enzymatic transesterification, a transesterified fatty acid component is separated from the transesterified glyceride component of the transesteri-fication mixturo. The transesterifled glyceride component has less than 3 woight percent esteri~led saturated fatty acid content, basod on the total woight o~ tho glyceride.
Depending upon the ratio of initial components and the extent of transesterification reaction, the transesterified glyceride component will comprise less than 3.5 weight percent of saturated ratty acids based on the total weight of fatty acids in the transesterified glyceride component, and may preferably have less than 3 weight percent of esterified saturated fatty acids, and for specific uses may desirably comprise less than two weight percent of saturated fatty acids.
Further, in accordance with the present invention, the transesterified fatty acid component separated from the ~&3~ 3 transesterification mixture is fractionated to separate the unsaturated fatty acids from saturated fatty acids, to provide a recycle unsaturated fatty acid ~ource material comprising less than 2 weight percent and preferably less than 1 weight percent of saturated fatty acids, based on the total weight of fatty acids in the source material.
The recycle fatty acid source material is combined in a recyclic manner with the high unsaturate vegetable oil such as canola oil as an unsaturated fatty acid transesterif~cation co~ponent, as previously described, to produce low-saturate triglyceride oils in accordance with the present invention. The recyclic use of the unsaturated fatty acid components is important to the economics of the process.
The fractionation of saturated fatty acids from the fatty acid mixture to provide such a low level of saturated fatty acid content is a difficult fractionation step and may be carried out by a variety of procedures, such as vacuum distillation, selective urea adduction and/or selective adsorption chromatography. The saturated fatty acids are difficult to remove from unsaturated fatty acids at low concentration levels. Solvent crystallization at low temperatures may be used to remove at lQast a portion of the saturated fatty acid content, although typically such ~olvent cry~tallization procedures do not reduce tho saturated fatty acid content to levels below about 2-3 weight percent o~ the fatty acid mixture.
However, such solvent crystallization fractionation procedures may be used to reduce the saturated fatty acid content for subsequent procedures such as molecular distillation, urea adduction or other selective absorption fractionation procedure~. It 18 important that the technique utilized ultimately provide a separation such that an unsaturated fatty acid recycle component having les~ than 2 weight percent by weight, based on the total woight of the fatty acids of the recycle component, and preferably less than 1 weight percent, is provided for recyclic transecterification use. Desirably, at least about gO weight percent of the unsaturated fatty acid component from the transesterification reaction will be recovered for recyclic use.
In accordance with the present methods, enzymatic transesterification is utilized to reduce the level of saturated fatty acids in a triglyceride vegetable oil by the addition of unsaturated fatty aeids in a ratio (oil:fatty aeid) such that at 6ubstantial equilibrium among the exchangeable fatty acids, the pereentage of unsaturates on the glyceride backbone is reduced from that of the starting material. By seleetively choosing the unsaturated fatty acids used for transesterification, the enzymatic transesterification reaction may be driven to a desired targeted fatty aeid eomposition in aeeordance with the present invention.
In accordance with such methods, enzymatic transesterification processes may produce oils with less than 3.5 weight percent of saturated fatty aeids and preferably less than 3 weight pereent saturated fatty acids, based on the total fatty acid content. For example, by using an immobilized Mueor miçhçi lipase and eanola oil, high oleie sunflower oil or high oleie safflower oil, respeetively, a~ the respeetiv- starting feedstoeks, $nteresterified olls having redueed levels of saturated fatty acid eontent6 of 1.6%, 1.0% and 0.9%, respeetively, may be readily provided. Moreover, sueh low ~aturate produets may be provided having a substantially 1:1 ratio of monounsaturate~ to polyunsaturates. Sueh starting feedstock materials may also readily be converted to oils having a 2:1:1 weight ratio of omega-9:omega-6:omega-3 un~aturated fatty aeid eontent. For example, eanola oil may also be readily interesterified to produee an oil with a 2:1:1 weight ratio of omega-9:omega-6:omega-3 esterified unsaturated fatty aeids, and a 6aturated fatty acid eontent of 2.1%.
In order to provide economical manufacturing methods, the free fatty acids which are used to drive the reaction are recycled by ~eparation of saturated fatty acids from unsaturated fatty acids. Separation of triglycerides from Sree fatty acids and~or separation of oleic, linoleic and linolenic acids from each othex may also be utilized in specific embodiments of such methods.
As previously discussed, the saturated fatty acid distribution of vegetable oils 6uch as canola oil is non-random, and is predominantly distributed at the 1-, 3-positions of the vegetable oil glycerides. Accordingly, by utilizing a 1-, 3- specific lipase such as previously described, the transesterification reaction may be limited to the 1-, 3- positions containing the predominant amount of saturated acids in the vegetable oil, without substantially affecting (or having a substantially reduced effect on) the fatty acids at the 2- position. In this manner, the amount of unsaturated fatty acid component utilized to achieve saturate reduction is decreased and effective transesterification efficiency is increased.
While 1-, 3- specific lipases are preferred for use with oils such as canola oil in which the saturated fatty acids are concentrated in the 1- and 3- positions, other lipases may al80 be used, particularly ln oils which do not signi~icantly ¢oncentrato ~aturated fatty acids in the 1-, 3- positions or which nevertheless have excessive saturated fatty acid amounts at every glyceride position.
For example, it may be more economical to use a non-specific lipase, such as the lipase from Çandida ruaosa to enable the reduction of stoichiometric amounts of free fatty acids in oils in which all three positions on the triglyceride will desirably be available for exchange.
Having generally described various aspects of the present invention, the invention will be more particularly described with the specific embodiment of FIGURE 1.
Illustrated in FIGURE 1 i3 a whematic diagram of a system 2 ~ 3 ~
100 for producing edible low saturata triglyceride oils in accordance with the present invention. As shown in FIGU~E 1, a variety of triglycerides may serve as fatty acid source materials for various unsaturated fatty acids, I to produce a desired end product. In this regard, the system 100 comprises a linseed oil storage vessel 102 to provide a source high in linolenic acid, a sunflower, safflower, corn or soybean oil storage vessel 104 to provide a source high in linoleic acid, and a hiqh oleic sunflower, high oleic safflower or olive oil storage vessel 106 to provide a source high in oleic acid.
Depending on the desired fatty acid content for the transesterification product, one or more of these high linolenic, high linoleic or high oleic source triglycerides is conducted through a conduit 108 to a hydrolysis reactor such as membrane reactor 110 for hydrolysis of the fatty acids of the oil. The reactor 110 may utilize a basic hydrolysis catalysts such as sodium hydroxide, sodium methoxide or an immobilized non-specific lipase to fully hydrolyze the triglyceride source material, or may utilize an immobilized unsaturated fatty acid-specific lipase, as previously described, to hydrolyze substantially only the unsaturated fatty acid components of the 60urce triglyceride.
The fatty acid components, which may include saturated ~atty acid component~ from the source triglycerid-, are ~eparated from tho glycerol component and conducted to a saturated fatty acid separator 120. The fatty acid separator 120 may utilize any appropriate separation technology, and, for example may be a separator such as a low temperature molecular distillation column, a urea adduction separator, ~nFatty Acids, Their Chemistry, Properties, Production and U8Q8~, Part 3, K.S. Markley, Ed., Interscience Publishers, 1964~, or a selective absorption elution column [Sorbex Separations, A Key to New Product Opportunities, Gembicki, et al., World Conference ¢ ~
on Emerging Technologies in the Fats and Oils Industry, A.R. Baldwin, Ed., American Oil Chemists Society (1985)], which is suitable for separating substantially all of the saturated fatty acids from the unsaturated fatty acid components. The fatty acids may be separated in a pretreatment step by low temperature ~olvent crystallization procedures to remove a substantial portion of the saturated fatty acid content prior to introduction into the separator 120, in order to reduce the cost of the separation step, if desired.
~ he separation step performed by the separator 120 is an important step in the method and, as will be described in more detail hereinafter, is also used to process recycled fatty acids and optionally diglycerides produced by the transesterifi~ation reaction. An unsaturated fatty acid transesterification mixture 122 comprising less than about 2 weight percent of saturated fatty acids, and preferably less than about 1 weight percent fatty acids is produced by the separator 120. A
saturated fatty acid stream 118 18 discharged from the separator, and may be utilized ~or other purposes such as the preparation of hard fats, ~oaps or chemical synthesis.
In the illustrated embodiment 100, the unsaturated transesterification fatty acid mixture 122 is combined with refined canola oil from storage ve~sel 130. The canola oil may typically have a ~aturated ~tty acid content which is concentrated in th- 1-, 3- position, as shown in the following table:
Typical Fatty Acid Distribution o~
Ç~nQl~ ~LI~YL~51d Fe~8tock~
Canola Oil Glyceride Position O~erall #1+#3 #2 Pal~itic 4.15 6.06 0.34 Stearic 1.81 2.72 less than 0.1 Oleic 56.~4 59.64 50.96 Linoleic 19.97 13.42 33 09 Linolenic 7.85 3.96 15 64 Other 9.48 14.20 le~s than 0.1 .
~ ~ 3 .~
The unsaturated fatty acid and canola oil are combined in a weight ratio in the range of from about 0.4 to about 3 of fatty acid to canola oil, to provide a canola oil - fatty acid transesterification reaction mixture 132 which is saturated or supersaturated with water by heat exchanger 134, and water saturator 136, as shown in FI~URE 1. The heat exchanger 134 heats the mixture 132 of canola oil and unsaturated fatty acids to approximately the desired reaction temperature of the transesterification reaction, which will preferably be in the range of from about 40- c. to about 70 C.
The heated transesterification mixture 135 is conducted into the water saturator 136, which desirably is a column of anionic resin such as the AmberLite IRA-900 anionic resin product of ~ohm and Haas, which is ~aturated with water. The anionic resin, in addition to rapidly saturating the canola oil-unsaturated fatty acid mixture with water, may also function to remove impurities which may poison or adversely affect the enzyme in the subsequent enzymatic transesterification reaction step.
The reaction mixture 138 which is discharged from the water saturator 136 is saturated or supersaturated with water, and in this regard, may typically comprise from about 0.2 to about 0.8 weight percent o~ water. The mixture 138 may be supersaturated by ~llghtly cooling a water-saturated reactlon mixture. The saturation or supersaturation of the mixture 138 with water is necessary to maintaln the utility of the resin-bound transesterification enzyme in the transesterification reaction, which will now be described in more detail.
The water-saturated or supersaturated reaction mixture 138 is introduced into an transesterification reactor 140, which ln the lllustrated embodiment 100 contains an immobilized 1-, 3- positionally specific transesterification lipase immobilized on or within an organopolymeric resin, ~uch às th~ Novo 3A lipase product 2 ~ 3 ,A ~ 3 , of Novo Industries, which is an immobil$zed 1-, 3-positlonally specific lipase derived from Mucor iehei, as described herein The fatty acids of the fatty acid source and the fatty acids of the 1-, 3- positions of the canola oil are substantially eguilibrated in the reactor 140 to provide an transesterification reaction product mixture 142 which is conducted to a triglyceride/fatty acid membrane separator 150 The mixture 142 generally contains from about 2 to about 8 weight percent of diglycerides produced by the action of the enzyme and water, but may contain up to about 15 weight percent of diglycerides, depending on factors including water content and reaction conditions The life time of the immob$1ized lipase system is important in commercial production, and may be monitored by means of an assay system to determine the half-life of the immobilized lipase in the canola oil reactor 140 As previously indicated, impurities in both the fatty acids and canola oil may accumulate on the resin and affect the enzymatic activity, and accordingly, pure materials should best be used in the reaction The illustrated triglyceride/fatty acid separator lSo may be a separator apparatus which separates the fatty acid components including those produced in the reaction from the di- and triglyceride components by any appropriate methods, such as by ~-lectiv- absorption rractionation processes, subcritical liguified ga~ (e g , propane) countercurrent extraction ~"Liguid-Liquid Extraction Employing Solvents in the Region of their Critical Temperaturesn, Hix~on, et al , American Institute of Chemical Engine-rs, ~oston, MA meeting May, 1942, pp 929~, wat-r washing, and/or vacuum deodorization, etc Optionally, at least a portion of the diglyceride components may be separated with the fatty acid component lf desired by 6elective adsorption fractionation processes, countercurrent selective solvent treatment processes, or other suitable procedure~, depending upon desired composition and food product utilization of the transesterified vegetable oil 160.
The transesterif~ed di- and triglyceride product 160 has an esterified saturated fatty acid conte~t of less than 3.5 weight percent, and may be used in a wide variety of food products, such as liquid margarine or cooking oils, mayonnaise and salad dressings, to provide products having extremely low levels of saturated fat. High levels of diglycerides together with very low levels of monoglycerides which may be provided in the procesR may be particularly desirable in the manufacture of certain emulsified focd products containing such low saturate content oils.
The free fatty acids in product stream 162 are separated from the fractionation solvent ~if used) by appropriate recovery apparatus 170, which is recycled to separator 150, to provide a recovered fatty acid stream 172 and recovered solvent 174. The recovered fatty acid stream 172 contains caturated fatty acids from the canola oil 130, and from the unsaturated fatty acid stream 122 as a result of the transesterification reaction. This fatty acid recycle stream, as previously indicated, is conducted to the separator 120 for separation of the fatty acid components and removal of saturated fatty acid components to permit recyclic USQ of the unsaturated fatty acid ; components. Amounts of saturated fatty acld recoverQd, and unsaturated fatty acld~ lost by thQ oeparator 120 or in other processing are made up by fatty acids from the source tanks 102, 104, 106 as needed to provide the proper proportion of fatty acid components, in the desired ratio, for the transesterification reaction.
Accordingly, it will be appreciated that low ~aturated oil may be produced by batch or continuous transesterification reaction m~thods to produce low-saturate oils with desired fatty acid compositions.
Such oils may be selected for fatty acid composition based ~3~
on the fatty acid composition of the starting oil and the fatty acid composition and relative proport$ons of the fatty acids used in the reaction.
While the production of low 6aturated edible vegetable oils using batch or continuous cocurrent reaction has been described, countercurrent systems may also be utilized to manufacture 6uch products. Countercurrent processes utilizing countercurrent supercr~tical or subcritical fluids which selectively extract and transport the fatty acid may desirably be utilized to provide efficient transesterification of the recycled stearic acid components. Countercurrent transesterification procedures may not only provide the reaction efficiencies of countercurrent operation, but also may facilitate ~eparation of reaction products.
In supercritical fluids ~solvents above their critical temperature and critical pressure) such as supercritical carbon dioxide, hydrocarbons such as ethane and ethane/propane mixtures, and fluorocarbons such as hexafluoroethane, solubility of fatty acid ester6 6uch as fatty acid methyl and ethyl esters are typically an inverse function of molecular weight of the fatty acid monoester under various condition~. Similarly, the solubility of fatty acids is inversely proportional to molecular weight of the fatty acid, although fatty acids ar- typlcally le88 soluble in supercrltlcal ga~es, than corresponding ratty acld lower alkyl monoesters of corresponding molecular weight because of the associative or hydrogen bonding characteristics of the fatty aclds.
Tho respective solubilities of fatty acids, fatty acid esters and triglyceride~ in carbon dioxide are also a function of temperature and partial pressure of C02 at relatively low 6upercritical pressures.
An embodiment Or continuou~ transesterification process which moves a fatty acid or fatty acid monoester component countercurrent to triglyceride flow, and which 2 ~
also removes such fatty acid transesterification reaction components from the transesterified glyceride, is illustrated in FIGURE 2.
As shown in FIGURE 2, a ~ource of canola oil 212 or other vegetable oil having a low (e.g., preferably less than 10 weight percent, and more preferably less than 7 weight percent) esterified saturated fatty acid content is provided as a starting material.
An unsaturated fatty acid (or fatty acid low alkyl monoester such as a mixture of unsaturated fatty acid methyl or ethyl esters~ derived from canola o$1 having less than 2 weight percent and preferably less than 1 weight percent of saturated fatty acids is provided as a transesterification reaction by selective enzymatic alcoholysis or esterification of unsaturated canola oil fatty acids components. Such methyl or ethyl esters may be provided by appropriate purification procedures such as fractional crystallization, solvent/solvent extraction, urea adduct fractionation, selective absorption fractionation, and/or sub- or supercritlcal solvent extraction, to 6eparate the unsaturated fatty acid or alcohol ester components from saturated components.
The canola oil 212 may be conducted through a column containing a water-saturated anionic exchange resin to remove non-triglyceride impuriti-- whlch might poison the enzym-, and conditlon th- oll 212 for the reactlon.
The rate o~ lntroductlon corr-~pond~ to th-tran~esterificatlon reactlon rate pormltted by the activity of the immobilized enzyme in the column 214. In this regard, the column is packed with an immobilized lipase enzyme, which is lmmobilized on an organic or lnorganic, high surface area ~ubstrat- such a~ porous ceramic rings or pellets, organic substrstes such a~ crosslinked ion exchange or phenolic resins (-.g., Novo 3A lipase product as described herein~ which are insoluble in the ~upercritical fluid, or diatomaceou- arth (e.g., C-litQ).
- 21 - 2 ~ ~s ~
The surface area of the column packing is very lar~e in order to promote interesterification reaction (e.g., more than 750 ~quare meters of 6urface area per cu~ic meter), and to promote equilibrium dissolution of the low molecular weight components in the ~upercritical fluid.
The unsaturated fatty acid (or preferably lower alkyl monoester) component 220 is introduced into a transesterification reactor 214 together with canola oil 212, for transesterification to produce a low saturate triglyceride. An immobilized 1-, 3- specific enzyme 222 such as previously described may be used for transesterification of canola oil, because the saturated acid content of canola oil is concentrated in the 1- and 3-positions. However, an immobilized nonspecific transesterification lipase such as the nonspecific enzyme of Candida cyl~dracae or Cand~ rugosa, may be used.
Moreover, dried lipase-containing microorgansism cells and mycelia may also be used as a transesterification enzyme, either retained in the reaction zone, or conducted therethrough with the triglyceride liguid pha~e flow te.g., see Gancet, et al., ~Catalysi~ by a Lipase-Bearing Rhizopus Arrhizus Mycellum in ~Halogeno)Fluorinated Hydrocarbons", Ann. N.Y. Acad. Sci., Vol. 542, pp. 213-218 (1988)].
~ he unsaturated fatty acid or preferably a lower alkyl unsaturated fatty acid monoester 220, such a~ a methyl or ethyl ester Or e.g., ethyl oleate, ethyl linoleate, ethyl llnolenate mixtur-, whioh i~ desired to be transesteri~led with the triglyceride 212, which may be saturated with water i8 introduced into the column 214 at a point 222 between the point 224 of introduction of triglyceride, and the lower outlet 218 at a rate which maximizes the desired transesteriflcation reaction.
B-cause this transester$ficatlon reactlon is conducted in a countercurrent manner, a lower ratio of unsaturated acid e~ters or acids 220 to canola oil 212 may be used.
In operatlon, supercritical carbon dioxide ~or another supercritical fluid such as an ethane-propane 2 ~
mixture of a fluorocarbon gas having a supercritical temperature for example in the range of from about 30- C.
to about 80- C.), is introduced at the bottom of the column 214 under pressure and temperature conditions at which relatively low molecular weight fatty acids or fatty acid esters are significantly dissolved, but at which the high molecular weight triglycerides are relatively not substantially dissolved. For example, carbon dioxide pressures in the range of from about 1100 psi to about 4500 (e.g., 2000-3000 psi for ethyl esters of oleic, linoleic and linolenic acids~, at a reaction temperature in the range of, for example, from about 30- C. to about 70- C., are particularly preferred to provide relatively high fatty acid and/or fatty acid monoester solubility, while providing relatively low triglyceride solubility in the upwardly moving supercritical carbon dioxide stream. The supercritcal fluid may contain a small amount of water vapor to maintain the catalyst and to facilitate fatty acid solubility. The temperature, o~ course, cannot exceed the operating temperature of the enzyme, which will be damaged at high temperatures. In this regard, at lower supercritical pressures, the solubility of the fatty esters and triglycerides i8 higher at lower temperatures. A
reaction temperature should be 6elected (e.g., 3S- -55- C.) which maximize~ throughput rate for countercurront transport of the fatty acid monoester, and the transesteri~ication reaction rate which i8 prov$ded by the enzyme to achieve transe~terification of the triglyceride and the fatty acid or fatty acid monoester. Fatty acid low-r alkyl monoesters are substantlally more soluble in the supercritlcal fluid than the corresponding acids, and accordingly are preferred reactants. The supercritical ga~
also serve~ as a diluent of the triglyceride phase to increase the reaction rate. If it is desired to operate the countercurrent column at a temperature higher than the lipase enzyme can tolerate, a plurality of enzyme reaction - 23 ~
zones 215 may be distributed along the column 214 which may be operated at a lower temperature, with appropriate heating and cooling means for fluid pumped therebetween.
In this manner, the transesterification reaction and countercurrent gradient functions may be separately maximized.
The 8upercritical carbon dioxide is less dense than the downwardly moving canola oil stream at pressures and temperatures used in the system of FIGURE 2 (e.g., 35-70- C., 1500-3500 psia). The pressure, temperature, column distances and flow rates of fatty acid or fatty acid monoester and carbon dioxide are selected so that in the zone 228 between the point of introduction of the carbon dioxide and the point 222 of introduction of the unsaturated fatty acid or monoester, the fatty acid or fatty acid monoester is progressively dissolved from the triglyceride into the upwardly moving supercritical CO2 stream; the acid or fatty acid monoester components (including the transesterified components) are substantially completely removed from the triglyceride stream 226 before it i~ discharged from the column at outlet 218. In this regard, the weight ratio of the flow rate of the carbon dioxide to the flow rate of the unsaturated acid or monoester component 220 introduced in the column 214 may de~irably be oelected to be in the range of from about 5:1 to about 50:1, under conditlons to maximize solubility o~ the ~atty acid or preferably fatty acid monoester component while minimizing the solubility of the triglyceride component phaso. In the zone 224, during the time o~ transit o~ the canola oil (e.g., .25 - 6 hours), the unsaturated acid component 220 undergoes transesterification with the triglyceride component.
Because the flow of triglyceride, and acid or monoester is cocurrent in this ~tripping zone, the enzymatic transesterification reaction will tend to approach the equilibrium condition of the unsaturated acid ~3~ $~
, monoester-triglyceride blend at the point 222 of introduction of the monoester 220. Accordingly, the composition of the fatty acid or fatty acid monoester which enters the countercurrent transesterification zone 226 from the monoester stripping zone 224 will be different from the composition of the fatty acid or monoester 220 introduced into the column 214 at least in part because of the transesterification which occurs in the stripping zone 224. The transesterified triglyceride product, which may have substantially all fatty acid and fatty acid monoester components removed therefrom, is withdrawn from outlet 218.
The ratio of triglyceride components to the unsaturated acid or monoester component 220 to achieve a desired degree of transesterification of the canola oil triglyceride is substantiàlly greater in the system of FIGURE 2 than the ratio of triglyceride to fatty acid or monoester utilized to achieve an equivalent degree of transesterification in a one or two step batch reaction.
In this regard, the unsaturated acid or monoester 220 is introduced into the bottom of the column at a rate compared to the rate of introduction of canola oil which ~ay, for example, be about half the proportion used in a batch reaction (e.g., 1:3 to 1:1 weight ratio of unsaturated acid component to canola oil).
The fatty acid or monoester component i5 dissolved in the upwardly movin~ supercritical C02 gas stream and carried into the transesterification zone 226, where lt approaches equilibrium with the countercurrent oil flow through the action of the immobilized enzyme in the coluLn. It is further transQsterified in a countercurrent manner with the liquid triglyceride ~tream a~ it is oonducted from its point of introduction 224 to the point 220 of introduction of the fatty acid monoester.
The triglyceride phase mixture continuously undergoes transesterification r~action as it moves 2 ~
downwardly in the zone 226 containing lipase enzyme countercurrent to the flow of supercritical gas, such that the mixture has an increasing concentration of the desired triglyceride components as it moves down the column. There is also an increasing concentration of transesterified fatty acid or monoester having fatty acid or monoester components derived from the triglyceride in the upwardly moving supercritical gas stream, in the direction toward the point of introduction of the triglyceride. Water vapor may be included in the carbon dioxide flow, the fatty acid ester flow and/or the triglyceride flow to accommodate the transesterification reaction, which may exceed the solubility of water in the triglyceride component, and to produce a desired level of diglyceride~, lf desired. Fatty acid components produced by hydrolysis reactions in the column 214 may also be removed by the supercritical carbon dioxide flow.
The transesterified fatty monoester or fatty acids dissolved in the supercritical C02 gas stream i8 carried from the column at outlet 216, through a pressurQ let-down system, where dissolved fatty acids or monoesters are taken out of supercritical solution as a result of pressure reduction. Th- tank 232 may alternatively be heated to further reduce the solubility of the fatty acid monoester.
The solubility reduction may al80 bo accompli~hed by a combination of a limlt-d pr-s~ur- r-duction (-.g., by 500-1000 p~i) and a temperatur- incr-a~ .g., to 70-lOC- C.) so that the work to rQcompress the C02 for recycle use may be reduced. The energy recovery sy~tem may comprise a piston or turbine ngine 232 in which the di~solved fatty acid or monoester 234 are collected in the rocovery system, 80 that the energy may be recovered for recompression of the carbon dioxide upon recyclic operation. ~he lower pressure C02, which is depleted in or froe of dissolved fatty acids or monoesters, is recompressed by compressor 238.
- 26 - 2~
The carbon dioxide 2~6 which is separated from the fatty acid or monoester is conducted to compressor/thermal conditioner 238 where it is recompressed and reintroduced at the preselected operating temperature as previously discussed A heat-pump or other thermal connector 240 may be used to transfer heat between the compressor 238 and the decompression engine 232 A portion of the extracted fatty acid components 234 may be recycled for reflux purposes to increase selectivity The flow rate of supercritical carbon dioxide (or other supercritical gas solvent) through the column 214tis correlated with the flow rate of fatty acid ester 220 so that it is adequate to transport and dissolve 6ubstantially all of the fatty acid monoester under the operating conditions, but dissolves a minimal amount of the initial canola oil and other triglyceride components The solubility of the fatty acid or fatty monoester components will desirably be greater than 1 weiqht percent, and preferably greater than 2 weight percent, while the solubility of triglycerides will be less than 0 5 weight percent and preferably d$ssolved triglycerides will be less than 10 percent of the dissolved fatty acid or monoester in the extracted product 234 The separated fatty acid or monoester 234 ~ay be purified in fractionation system 250 to separate purified unsaturated fatty acid- 220 from oth-r material~ 252 in any suitable mann-r, and the unsaturated transesterification component 220 may be recycled for transesterification use Another embodiment of a continuous countercurrent transesterification process iB illustrated in FIGURE 3, which moves a ~atty acid or fatty acid monoester component countercurrent to triglyceride ~low, utilizing a subcritical liquified ga~ solvent As 6hown in FIGURE 3, the vegetable canola oil 312 to be transesterified is introduced into a high pressure column 314 at a point 324 intermediate the upper outlet 316 and the lower outlet 318 The rate of introduction 2 ~ j L': ~
corresponds to the transesterification reaction rate permi~-ted by the activity of the immobil$zed enzyme in the column 314. The ~olumn may be packed with an immobilized lipase enzyme as previously described, which is immobilized ! on an inorganic, high surface area substrate ~uch as porous ceramic rings or pellets, or d$atomaceous earth (e.g., ~elite), or a suitable organic substrate such as an ion ? exchange resin substrate (e.g., Novo 3A lipase). Such packing may be in particulate film or fiber form, on plates, etc. Alternatively, as shown in FIGU~E 3, the column 314 may be used to establish concentration gradients and countercurrent flow in stages, and the immobilized I enzyme 3$3 may be located in corresponding stages in a separate pressure vessel 315. The surface area of the j column packing should be very large in order to promote I transesterification reaction (e.g., more than 750 square I meters of surface area per cubic meter), and to promote ! equilibrium dissolution of the low molecular weight ! compGnents in the supercritical fluid.
! A fatty acid or lower alkyl fatty acid monoester i 320, such as a methyl or ethyl ester of an unsaturated ¦ fatty acid component (e.g., a mixture of unsaturated fatty ! acids or unsaturated fatty acid esters such as methyl or ; ethyl esters), which is desired to be transesterified with the triglycerido 312, i8 introduced into tho column 314 at a point 322 intermediate the point 324 o~ introduction o~
triglyceride, and the lower outlet 318 at a rate which ! maxi~izes the desired transesterification reaction. For mnnufacture of the low saturated fat vegetable oil, a I mixture of unsaturated fatty acids or monoesters comprising I le~s than 2 weight percent of saturated fatty acids is used. The rate of introduction i8 about 0.25 to 1.0 the rate of vegetable oil input, lower ratios being used for higher purity ~e.g., less than 0.5 or 0.25 weight percent saturated fatty acid content). The transesterification reaction is conducted in a countercurrent manner, 80 that ~ 28 it is substantially more eff~cient than the batch reaction of the fatty acid and triglyceride component at the selected reaction ratios.
In operation, liquified propane 309 i8 introduced at a position 319 near the bottom of the column 314 under pressure, flow rate and temperature conditions at which relatively low molecular weight fatty acids and monoesters are significantly dissolved, but at which the high molecular weight triglycerides are less ~oluble and form a separate phase. The temperature, of course, cannot exceed the operating temperature of the enzyme, whlch will be damaged at high temperatures. In the illustrated embodiment, the propane may be introduced at a pressure of about 600 psi and a temperature of 70- C. at a rate of about 10 t~mes the rate of introduction of fatty acid 320 on a weight ratio basis. If a lower temperature is necessary, ethane or ethane/propane mixtures may be used.
The liquified propane gas also dissolves to some extent in the triglyceride phase and serves as a diluent to increase the reaction rate.
The liguified propane phase i6 less dense than the downwardly moving veqetable oil stream, having a density of 0.25-0.4 g/cm~, depending primarily on composition and temperature. ~he pressure, temperature, column distance~ and flow rate~ Or ratty acid~ ~or ~onoesters) and liquid propane are selected 80 that in the zone 3~4 between the point 319 of introduction of the pure liquified propane and the point 322 of introduction of the unsaturated fatty acid or monoester 320, the fatty ac~d or monoQster components are progressively dlssolved from the triglyceride into the upwardly moving liquid propane phase;
the fatty acid or monoester ~s substantially completely removed from the triglyceride ~tream 326 before it is discharged from the column at outlet 318. Because of countercurrent transesterification which occurs in the transesterlfication zone~ 313 (or in the column 314 in embodiments having lipase enzyme there~n), the fatty acids or monoesteræ in the upwardly rising liquid propane droplets will be different from the composition of the fatty acids 320 introduced into the column 314 because of the transesterification. The transesterified triglyceride product i8 withdrawn from outlet 318. It contains an amount of propane which is an inverse function of temperature (e.g., 30-60 weight percent propane), which is removed in propane stripping column 327 and returned for i recyclic use.
The ratio of triglyceride component 312 to the unsaturated fatty acid component 320 to achieve a desired degree of transesterification of the triglyceride will be substantially groater in the countercurrent system than the ratlo of triglyceride to fatty monoester necessary to achieve an equivalent degree of transesterification in a batch or cocurrent reaction, as previously discussed.
Moreover, high levels of transesterification substitution may be obtained with the proposed countercurrent process, which could otherwise only be achieved with multiple batch reaction, and multiple component ~eparation.
The fatty acid component is dissolved in the upwardly moving propane phase, and is continuously exchanged with the downwardly moving vegetable oil pha~e, creating a gradlent along th- column, and carried into th-tran~esterification zone, wher- it tends toward dynamic equilibrium with the countercurrent oil flow. It 18 transesterified ln a countercurrent manner with the liquid trlglyceride stream ~- it i~ conducted from the point of introduction 324 to the point 320 of introduction of the fatty acid monoe~ter.
A portion of the mixture is pumped from mixing ~tage~ in the column 314 to transesterification zones 313 containing a suitable transesterirication enzyme. The mixture continuously undergoes transe~terification reaction in the separate zones 313, and i~ returned to the
DescriDtion of the Invention Generally in accordance with the present invention, methods are provided for manufacturing a triglyceride oil having less than 3.5 and preferably less than 3 weight percent of esterified saturated fatty acid, and desirably less than 2, and more preferably less than 1 weight percent of intermediate chain length esterified saturated fatty acid.
Various aspects of the present invention are also directed to low saturate liquid vegetable oil products having less than 3.5 weight percent, and preferably less than 3 weight percent saturated fatty acid moieties, and specific unsaturated fatty acid distribution, which liguid vegetable oil products have de-irable properties for uso in food products such as mayonnaise, margarine, table spreads or salad dres~ings. Such low saturate vegetable oil products will al~o desirably have less than about 2 and preferably less than 1 weight percent of intermediate chain saturated fatty acids. By n intermediate ¢haln ~aturated fatty acid~ i8 meant, lauri¢, myri~tic and palmlti¢ acids having carbon chain l-ngths of C12, C14 and C16, rQspe¢tively. As us~d herein, the weight percent of ~aturated or unsaturated ~atty acid moieties in a margarine oil or vegetabl~ oil glyceride composition is calculated based on the total weight of the fatty acids contained in the margarine oil. Also, as used herein, when referring to weight percent of onQ or more fatty acid moiQtiQs of a composition ~such as the weight percentagQ of C12-C16 fatty acids), the weight percent is calculated based on all of tho fatty acids of the compo~ition being fully 2 ~ 3 ~
hydrolyzed to free fatty acid. The weight percent of one or more species of fatty acid i8 then calculated as the weight percent of such one or more species based on the total weight of free fatty acids. AOCS official method Ce 1-62 (81) may be used to determine the weight percent of respective fatty acids of an oil or fat composition.
In accordance with various method aspects of the present invention, a high-unsaturate vegetable oil, such as canola (low erucic acid rapeseed) oil, high oleic safflower oil, high oleic sunflower oil, high linoleic safflower oil, ~ '~
~5~ ~ high linoleic sunflower,~soybëa~ oil or m'ixtures th'ereof, R~
,"~ t.!having from about 4 weight percent to about ~ a~d ~ Jn~
~L ~ typ~c~lly fro~--bou~ to bout-} weight percent Of ~D ~ h' / ~ 5~ ~Itl~ saturated fatty acids with respect to the total fatty ac~d content of the vegetable oil is provided for transesterification reaction. Canola oil is a particularly desirable starting material because it has a relatively low amount of saturated fatty acid content. ~he vegetable oil should best be refined and bleached oil which is substantially free of proteinaceous and other nonglyceride components which might poison or otherwise interfere with transesterification enzymes.
Canola oil typical~y comprises in the range of from about 5 to about 7 weight percent of ~aturated fatty acid moietie~. However, th- ~aturat-d fatty acids of canola oil ~and other vegetable oil8) i8 not randomly distributed with respect to the 1-, 2- and 3- positions of the glyceride molecules of the oil. A major portion of the saturated fatty acid content i5 located in the 1- and 3-primarv positions of the glyceride molecules of the canola oil. In this regard, canola oil typically may have an overall stearic acid content of about 1.5 - 2 weight percent based on the total fatty acid content of the oil, with from about 2 to about 3 weight percent of the fatty acid moieties in the 1- and 3- positions being stearic acid and less than about 1 weight percent of the fatty acids in 2 ~ $ - ~
the ~econdary 2- position being stearie acid. Similarly, canola oil may typically have ~n overall palmitic acid content of about 4 weight percent, with about 6 weight pereent of the fatty acid groups in the 1- and 3- positions being palmitic acid and less than about l weight percent of the fatty acid groups in the 2- position being palmitic aeid.
In accordance with the present methods, the saturated fatty acids of the primary l- and 3- positions are selectively removed, and replaced with unsaturated fatty acids by selective transesterification reaction.
This may be accomplished without substantially affecting the fatty acid distribution of the starting material oil in the 2- position. 8y "transesterification" is meant an exehange of fatty acid moiety or aeyl radical at a glyeeride oxygen or hydroxyl group, whieh ineludes interesterification and intraesterification.
In order to earry out sueh methods, the high unsaturate vegetable oil sueh as eanola oil is eombined with an unsaturated fatty aeid transesterifleation component selected from the group consisting of free fatty acids, fatty aeid monoesters of lower alkyl monohydrie aleohols (e.g., methanol, ethanol and propane), and mixtures thereof, whieh has less than 2 weight pereent, and preferably le~i than l weight pereent of s8turated fatty aeids to provld- a transesteri~leation blend. The unsaturated fatty aeid transesterlfieation eomponent should eomprise at least about 98 weight pereent of unsaturated fatty aeids having a ehain length Or from 12 to about 22 earbon atoms, and less than 2 weiqht pereent of saturated fatty aeids having a earbon ehain length in the range of from 12 to 18 earbon atoms based on the total fatty aeid eontent of the transesterifieation eompQnent. Desirably, th- fatty acid transesterifieation eomponent eomprises less than 0.75 weight pereent and preferably less than 0.5 weight pereent of intermediate ehain saturated fatty aeids, 2 ~ ~3 ~
g by weight, ~ased on the fatty acid content of the transes~erification component.
The high-unsaturate vegetable oil such as canola oil and the unsaturated fatty ac~d are combined in a weight ratio of vegetable oil to unsaturated fatty acid transesterification component in the range of from about 1:10 to about 4:1 in batch reaction processes (which may utilize multiple reaction steps), and typically in the range of from about 1:2 to about 2:1 in single stage batch or cocurrent reactions to provide a transesterification mixture. The ratio of reactants may be selected to provide a desired degree of substitution under the reaction conditions, to provide a low-saturate glyceride product having a selective level of 6aturated fatty acid content below about 3.5 weight percent, and preferably below 3 weight percent. Moreover, the ratio of reactants and the composition of the unsaturated fatty acid transesteri-fication component ma-~ be selected to provide specific compositions of the transesterified low saturate oil, in terms of unsaturated fatty acid composition. For example, nutritionally deæirable unsaturated fatty acids in appropriate levels and ratios have been identified such that an increase in the omega-3 to omega-6 ratio in the average diet could yield distinct health benefits. Natural triglycerides do not typically contain theso unsaturated acids in such proportions, but ~uch compositions may be provided in accordance with the present invention. Further in accordance with methods of the present invention, the transesterification mixture is contacted with a transesterification enzyme, which iB desirably a 1-, 3-specific transesterification enzyme such as an immobilized lipase from Muco~ miehe1 as described in U.S. Patent 4,798,793 issued January 17, 1989, which i~ incorporated by reference herein. The transesterification mixture is contacted with the immobilized enzyme under time ~nd temperature conditions for 6ubstantially eguilibrating the ~3~
fatty acid content in the 1- and 3- positions of the glyceride component, with the fatty acid transesteri-fication components of the reaction mixture. The enzymatic transesterification reaction produces a transesterified triglyceride component and a transesterified fatty acid component. The reaction time may range from about 0.5 hour to about 100 hours, depending upon the concentration and activity of the lipase and the temperature of the reaction mixture. The transesterification reaction will desirably be carried out at a temperature in the range of from about 20- C. to about 80- C., and more preferably in the range of from about 40- C. to about 70- C. By "substantially eguilibrate" is meant that the transesterification reaction is at least about S0 percent complete, and preferably at least 90 percent complete. ~ower equilibrium transesterification conditions (e.g., 50-90 percent equilibrated) may be utilized to increase the reaction speed and or reduce the amount of enzyme used, but this increases the unsaturated fatty acid required and increases the separation step processing requirements.
Following enzymatic transesterification, a transesterified fatty acid component is separated from the transesterified glyceride component of the transesteri-fication mixturo. The transesterifled glyceride component has less than 3 woight percent esteri~led saturated fatty acid content, basod on the total woight o~ tho glyceride.
Depending upon the ratio of initial components and the extent of transesterification reaction, the transesterified glyceride component will comprise less than 3.5 weight percent of saturated ratty acids based on the total weight of fatty acids in the transesterified glyceride component, and may preferably have less than 3 weight percent of esterified saturated fatty acids, and for specific uses may desirably comprise less than two weight percent of saturated fatty acids.
Further, in accordance with the present invention, the transesterified fatty acid component separated from the ~&3~ 3 transesterification mixture is fractionated to separate the unsaturated fatty acids from saturated fatty acids, to provide a recycle unsaturated fatty acid ~ource material comprising less than 2 weight percent and preferably less than 1 weight percent of saturated fatty acids, based on the total weight of fatty acids in the source material.
The recycle fatty acid source material is combined in a recyclic manner with the high unsaturate vegetable oil such as canola oil as an unsaturated fatty acid transesterif~cation co~ponent, as previously described, to produce low-saturate triglyceride oils in accordance with the present invention. The recyclic use of the unsaturated fatty acid components is important to the economics of the process.
The fractionation of saturated fatty acids from the fatty acid mixture to provide such a low level of saturated fatty acid content is a difficult fractionation step and may be carried out by a variety of procedures, such as vacuum distillation, selective urea adduction and/or selective adsorption chromatography. The saturated fatty acids are difficult to remove from unsaturated fatty acids at low concentration levels. Solvent crystallization at low temperatures may be used to remove at lQast a portion of the saturated fatty acid content, although typically such ~olvent cry~tallization procedures do not reduce tho saturated fatty acid content to levels below about 2-3 weight percent o~ the fatty acid mixture.
However, such solvent crystallization fractionation procedures may be used to reduce the saturated fatty acid content for subsequent procedures such as molecular distillation, urea adduction or other selective absorption fractionation procedure~. It 18 important that the technique utilized ultimately provide a separation such that an unsaturated fatty acid recycle component having les~ than 2 weight percent by weight, based on the total woight of the fatty acids of the recycle component, and preferably less than 1 weight percent, is provided for recyclic transecterification use. Desirably, at least about gO weight percent of the unsaturated fatty acid component from the transesterification reaction will be recovered for recyclic use.
In accordance with the present methods, enzymatic transesterification is utilized to reduce the level of saturated fatty acids in a triglyceride vegetable oil by the addition of unsaturated fatty aeids in a ratio (oil:fatty aeid) such that at 6ubstantial equilibrium among the exchangeable fatty acids, the pereentage of unsaturates on the glyceride backbone is reduced from that of the starting material. By seleetively choosing the unsaturated fatty acids used for transesterification, the enzymatic transesterification reaction may be driven to a desired targeted fatty aeid eomposition in aeeordance with the present invention.
In accordance with such methods, enzymatic transesterification processes may produce oils with less than 3.5 weight percent of saturated fatty aeids and preferably less than 3 weight pereent saturated fatty acids, based on the total fatty acid content. For example, by using an immobilized Mueor miçhçi lipase and eanola oil, high oleie sunflower oil or high oleie safflower oil, respeetively, a~ the respeetiv- starting feedstoeks, $nteresterified olls having redueed levels of saturated fatty acid eontent6 of 1.6%, 1.0% and 0.9%, respeetively, may be readily provided. Moreover, sueh low ~aturate produets may be provided having a substantially 1:1 ratio of monounsaturate~ to polyunsaturates. Sueh starting feedstock materials may also readily be converted to oils having a 2:1:1 weight ratio of omega-9:omega-6:omega-3 un~aturated fatty aeid eontent. For example, eanola oil may also be readily interesterified to produee an oil with a 2:1:1 weight ratio of omega-9:omega-6:omega-3 esterified unsaturated fatty aeids, and a 6aturated fatty acid eontent of 2.1%.
In order to provide economical manufacturing methods, the free fatty acids which are used to drive the reaction are recycled by ~eparation of saturated fatty acids from unsaturated fatty acids. Separation of triglycerides from Sree fatty acids and~or separation of oleic, linoleic and linolenic acids from each othex may also be utilized in specific embodiments of such methods.
As previously discussed, the saturated fatty acid distribution of vegetable oils 6uch as canola oil is non-random, and is predominantly distributed at the 1-, 3-positions of the vegetable oil glycerides. Accordingly, by utilizing a 1-, 3- specific lipase such as previously described, the transesterification reaction may be limited to the 1-, 3- positions containing the predominant amount of saturated acids in the vegetable oil, without substantially affecting (or having a substantially reduced effect on) the fatty acids at the 2- position. In this manner, the amount of unsaturated fatty acid component utilized to achieve saturate reduction is decreased and effective transesterification efficiency is increased.
While 1-, 3- specific lipases are preferred for use with oils such as canola oil in which the saturated fatty acids are concentrated in the 1- and 3- positions, other lipases may al80 be used, particularly ln oils which do not signi~icantly ¢oncentrato ~aturated fatty acids in the 1-, 3- positions or which nevertheless have excessive saturated fatty acid amounts at every glyceride position.
For example, it may be more economical to use a non-specific lipase, such as the lipase from Çandida ruaosa to enable the reduction of stoichiometric amounts of free fatty acids in oils in which all three positions on the triglyceride will desirably be available for exchange.
Having generally described various aspects of the present invention, the invention will be more particularly described with the specific embodiment of FIGURE 1.
Illustrated in FIGURE 1 i3 a whematic diagram of a system 2 ~ 3 ~
100 for producing edible low saturata triglyceride oils in accordance with the present invention. As shown in FIGU~E 1, a variety of triglycerides may serve as fatty acid source materials for various unsaturated fatty acids, I to produce a desired end product. In this regard, the system 100 comprises a linseed oil storage vessel 102 to provide a source high in linolenic acid, a sunflower, safflower, corn or soybean oil storage vessel 104 to provide a source high in linoleic acid, and a hiqh oleic sunflower, high oleic safflower or olive oil storage vessel 106 to provide a source high in oleic acid.
Depending on the desired fatty acid content for the transesterification product, one or more of these high linolenic, high linoleic or high oleic source triglycerides is conducted through a conduit 108 to a hydrolysis reactor such as membrane reactor 110 for hydrolysis of the fatty acids of the oil. The reactor 110 may utilize a basic hydrolysis catalysts such as sodium hydroxide, sodium methoxide or an immobilized non-specific lipase to fully hydrolyze the triglyceride source material, or may utilize an immobilized unsaturated fatty acid-specific lipase, as previously described, to hydrolyze substantially only the unsaturated fatty acid components of the 60urce triglyceride.
The fatty acid components, which may include saturated ~atty acid component~ from the source triglycerid-, are ~eparated from tho glycerol component and conducted to a saturated fatty acid separator 120. The fatty acid separator 120 may utilize any appropriate separation technology, and, for example may be a separator such as a low temperature molecular distillation column, a urea adduction separator, ~nFatty Acids, Their Chemistry, Properties, Production and U8Q8~, Part 3, K.S. Markley, Ed., Interscience Publishers, 1964~, or a selective absorption elution column [Sorbex Separations, A Key to New Product Opportunities, Gembicki, et al., World Conference ¢ ~
on Emerging Technologies in the Fats and Oils Industry, A.R. Baldwin, Ed., American Oil Chemists Society (1985)], which is suitable for separating substantially all of the saturated fatty acids from the unsaturated fatty acid components. The fatty acids may be separated in a pretreatment step by low temperature ~olvent crystallization procedures to remove a substantial portion of the saturated fatty acid content prior to introduction into the separator 120, in order to reduce the cost of the separation step, if desired.
~ he separation step performed by the separator 120 is an important step in the method and, as will be described in more detail hereinafter, is also used to process recycled fatty acids and optionally diglycerides produced by the transesterifi~ation reaction. An unsaturated fatty acid transesterification mixture 122 comprising less than about 2 weight percent of saturated fatty acids, and preferably less than about 1 weight percent fatty acids is produced by the separator 120. A
saturated fatty acid stream 118 18 discharged from the separator, and may be utilized ~or other purposes such as the preparation of hard fats, ~oaps or chemical synthesis.
In the illustrated embodiment 100, the unsaturated transesterification fatty acid mixture 122 is combined with refined canola oil from storage ve~sel 130. The canola oil may typically have a ~aturated ~tty acid content which is concentrated in th- 1-, 3- position, as shown in the following table:
Typical Fatty Acid Distribution o~
Ç~nQl~ ~LI~YL~51d Fe~8tock~
Canola Oil Glyceride Position O~erall #1+#3 #2 Pal~itic 4.15 6.06 0.34 Stearic 1.81 2.72 less than 0.1 Oleic 56.~4 59.64 50.96 Linoleic 19.97 13.42 33 09 Linolenic 7.85 3.96 15 64 Other 9.48 14.20 le~s than 0.1 .
~ ~ 3 .~
The unsaturated fatty acid and canola oil are combined in a weight ratio in the range of from about 0.4 to about 3 of fatty acid to canola oil, to provide a canola oil - fatty acid transesterification reaction mixture 132 which is saturated or supersaturated with water by heat exchanger 134, and water saturator 136, as shown in FI~URE 1. The heat exchanger 134 heats the mixture 132 of canola oil and unsaturated fatty acids to approximately the desired reaction temperature of the transesterification reaction, which will preferably be in the range of from about 40- c. to about 70 C.
The heated transesterification mixture 135 is conducted into the water saturator 136, which desirably is a column of anionic resin such as the AmberLite IRA-900 anionic resin product of ~ohm and Haas, which is ~aturated with water. The anionic resin, in addition to rapidly saturating the canola oil-unsaturated fatty acid mixture with water, may also function to remove impurities which may poison or adversely affect the enzyme in the subsequent enzymatic transesterification reaction step.
The reaction mixture 138 which is discharged from the water saturator 136 is saturated or supersaturated with water, and in this regard, may typically comprise from about 0.2 to about 0.8 weight percent o~ water. The mixture 138 may be supersaturated by ~llghtly cooling a water-saturated reactlon mixture. The saturation or supersaturation of the mixture 138 with water is necessary to maintaln the utility of the resin-bound transesterification enzyme in the transesterification reaction, which will now be described in more detail.
The water-saturated or supersaturated reaction mixture 138 is introduced into an transesterification reactor 140, which ln the lllustrated embodiment 100 contains an immobilized 1-, 3- positionally specific transesterification lipase immobilized on or within an organopolymeric resin, ~uch às th~ Novo 3A lipase product 2 ~ 3 ,A ~ 3 , of Novo Industries, which is an immobil$zed 1-, 3-positlonally specific lipase derived from Mucor iehei, as described herein The fatty acids of the fatty acid source and the fatty acids of the 1-, 3- positions of the canola oil are substantially eguilibrated in the reactor 140 to provide an transesterification reaction product mixture 142 which is conducted to a triglyceride/fatty acid membrane separator 150 The mixture 142 generally contains from about 2 to about 8 weight percent of diglycerides produced by the action of the enzyme and water, but may contain up to about 15 weight percent of diglycerides, depending on factors including water content and reaction conditions The life time of the immob$1ized lipase system is important in commercial production, and may be monitored by means of an assay system to determine the half-life of the immobilized lipase in the canola oil reactor 140 As previously indicated, impurities in both the fatty acids and canola oil may accumulate on the resin and affect the enzymatic activity, and accordingly, pure materials should best be used in the reaction The illustrated triglyceride/fatty acid separator lSo may be a separator apparatus which separates the fatty acid components including those produced in the reaction from the di- and triglyceride components by any appropriate methods, such as by ~-lectiv- absorption rractionation processes, subcritical liguified ga~ (e g , propane) countercurrent extraction ~"Liguid-Liquid Extraction Employing Solvents in the Region of their Critical Temperaturesn, Hix~on, et al , American Institute of Chemical Engine-rs, ~oston, MA meeting May, 1942, pp 929~, wat-r washing, and/or vacuum deodorization, etc Optionally, at least a portion of the diglyceride components may be separated with the fatty acid component lf desired by 6elective adsorption fractionation processes, countercurrent selective solvent treatment processes, or other suitable procedure~, depending upon desired composition and food product utilization of the transesterified vegetable oil 160.
The transesterif~ed di- and triglyceride product 160 has an esterified saturated fatty acid conte~t of less than 3.5 weight percent, and may be used in a wide variety of food products, such as liquid margarine or cooking oils, mayonnaise and salad dressings, to provide products having extremely low levels of saturated fat. High levels of diglycerides together with very low levels of monoglycerides which may be provided in the procesR may be particularly desirable in the manufacture of certain emulsified focd products containing such low saturate content oils.
The free fatty acids in product stream 162 are separated from the fractionation solvent ~if used) by appropriate recovery apparatus 170, which is recycled to separator 150, to provide a recovered fatty acid stream 172 and recovered solvent 174. The recovered fatty acid stream 172 contains caturated fatty acids from the canola oil 130, and from the unsaturated fatty acid stream 122 as a result of the transesterification reaction. This fatty acid recycle stream, as previously indicated, is conducted to the separator 120 for separation of the fatty acid components and removal of saturated fatty acid components to permit recyclic USQ of the unsaturated fatty acid ; components. Amounts of saturated fatty acld recoverQd, and unsaturated fatty acld~ lost by thQ oeparator 120 or in other processing are made up by fatty acids from the source tanks 102, 104, 106 as needed to provide the proper proportion of fatty acid components, in the desired ratio, for the transesterification reaction.
Accordingly, it will be appreciated that low ~aturated oil may be produced by batch or continuous transesterification reaction m~thods to produce low-saturate oils with desired fatty acid compositions.
Such oils may be selected for fatty acid composition based ~3~
on the fatty acid composition of the starting oil and the fatty acid composition and relative proport$ons of the fatty acids used in the reaction.
While the production of low 6aturated edible vegetable oils using batch or continuous cocurrent reaction has been described, countercurrent systems may also be utilized to manufacture 6uch products. Countercurrent processes utilizing countercurrent supercr~tical or subcritical fluids which selectively extract and transport the fatty acid may desirably be utilized to provide efficient transesterification of the recycled stearic acid components. Countercurrent transesterification procedures may not only provide the reaction efficiencies of countercurrent operation, but also may facilitate ~eparation of reaction products.
In supercritical fluids ~solvents above their critical temperature and critical pressure) such as supercritical carbon dioxide, hydrocarbons such as ethane and ethane/propane mixtures, and fluorocarbons such as hexafluoroethane, solubility of fatty acid ester6 6uch as fatty acid methyl and ethyl esters are typically an inverse function of molecular weight of the fatty acid monoester under various condition~. Similarly, the solubility of fatty acids is inversely proportional to molecular weight of the fatty acid, although fatty acids ar- typlcally le88 soluble in supercrltlcal ga~es, than corresponding ratty acld lower alkyl monoesters of corresponding molecular weight because of the associative or hydrogen bonding characteristics of the fatty aclds.
Tho respective solubilities of fatty acids, fatty acid esters and triglyceride~ in carbon dioxide are also a function of temperature and partial pressure of C02 at relatively low 6upercritical pressures.
An embodiment Or continuou~ transesterification process which moves a fatty acid or fatty acid monoester component countercurrent to triglyceride flow, and which 2 ~
also removes such fatty acid transesterification reaction components from the transesterified glyceride, is illustrated in FIGURE 2.
As shown in FIGURE 2, a ~ource of canola oil 212 or other vegetable oil having a low (e.g., preferably less than 10 weight percent, and more preferably less than 7 weight percent) esterified saturated fatty acid content is provided as a starting material.
An unsaturated fatty acid (or fatty acid low alkyl monoester such as a mixture of unsaturated fatty acid methyl or ethyl esters~ derived from canola o$1 having less than 2 weight percent and preferably less than 1 weight percent of saturated fatty acids is provided as a transesterification reaction by selective enzymatic alcoholysis or esterification of unsaturated canola oil fatty acids components. Such methyl or ethyl esters may be provided by appropriate purification procedures such as fractional crystallization, solvent/solvent extraction, urea adduct fractionation, selective absorption fractionation, and/or sub- or supercritlcal solvent extraction, to 6eparate the unsaturated fatty acid or alcohol ester components from saturated components.
The canola oil 212 may be conducted through a column containing a water-saturated anionic exchange resin to remove non-triglyceride impuriti-- whlch might poison the enzym-, and conditlon th- oll 212 for the reactlon.
The rate o~ lntroductlon corr-~pond~ to th-tran~esterificatlon reactlon rate pormltted by the activity of the immobilized enzyme in the column 214. In this regard, the column is packed with an immobilized lipase enzyme, which is lmmobilized on an organic or lnorganic, high surface area ~ubstrat- such a~ porous ceramic rings or pellets, organic substrstes such a~ crosslinked ion exchange or phenolic resins (-.g., Novo 3A lipase product as described herein~ which are insoluble in the ~upercritical fluid, or diatomaceou- arth (e.g., C-litQ).
- 21 - 2 ~ ~s ~
The surface area of the column packing is very lar~e in order to promote interesterification reaction (e.g., more than 750 ~quare meters of 6urface area per cu~ic meter), and to promote equilibrium dissolution of the low molecular weight components in the ~upercritical fluid.
The unsaturated fatty acid (or preferably lower alkyl monoester) component 220 is introduced into a transesterification reactor 214 together with canola oil 212, for transesterification to produce a low saturate triglyceride. An immobilized 1-, 3- specific enzyme 222 such as previously described may be used for transesterification of canola oil, because the saturated acid content of canola oil is concentrated in the 1- and 3-positions. However, an immobilized nonspecific transesterification lipase such as the nonspecific enzyme of Candida cyl~dracae or Cand~ rugosa, may be used.
Moreover, dried lipase-containing microorgansism cells and mycelia may also be used as a transesterification enzyme, either retained in the reaction zone, or conducted therethrough with the triglyceride liguid pha~e flow te.g., see Gancet, et al., ~Catalysi~ by a Lipase-Bearing Rhizopus Arrhizus Mycellum in ~Halogeno)Fluorinated Hydrocarbons", Ann. N.Y. Acad. Sci., Vol. 542, pp. 213-218 (1988)].
~ he unsaturated fatty acid or preferably a lower alkyl unsaturated fatty acid monoester 220, such a~ a methyl or ethyl ester Or e.g., ethyl oleate, ethyl linoleate, ethyl llnolenate mixtur-, whioh i~ desired to be transesteri~led with the triglyceride 212, which may be saturated with water i8 introduced into the column 214 at a point 222 between the point 224 of introduction of triglyceride, and the lower outlet 218 at a rate which maximizes the desired transesteriflcation reaction.
B-cause this transester$ficatlon reactlon is conducted in a countercurrent manner, a lower ratio of unsaturated acid e~ters or acids 220 to canola oil 212 may be used.
In operatlon, supercritical carbon dioxide ~or another supercritical fluid such as an ethane-propane 2 ~
mixture of a fluorocarbon gas having a supercritical temperature for example in the range of from about 30- C.
to about 80- C.), is introduced at the bottom of the column 214 under pressure and temperature conditions at which relatively low molecular weight fatty acids or fatty acid esters are significantly dissolved, but at which the high molecular weight triglycerides are relatively not substantially dissolved. For example, carbon dioxide pressures in the range of from about 1100 psi to about 4500 (e.g., 2000-3000 psi for ethyl esters of oleic, linoleic and linolenic acids~, at a reaction temperature in the range of, for example, from about 30- C. to about 70- C., are particularly preferred to provide relatively high fatty acid and/or fatty acid monoester solubility, while providing relatively low triglyceride solubility in the upwardly moving supercritical carbon dioxide stream. The supercritcal fluid may contain a small amount of water vapor to maintain the catalyst and to facilitate fatty acid solubility. The temperature, o~ course, cannot exceed the operating temperature of the enzyme, which will be damaged at high temperatures. In this regard, at lower supercritical pressures, the solubility of the fatty esters and triglycerides i8 higher at lower temperatures. A
reaction temperature should be 6elected (e.g., 3S- -55- C.) which maximize~ throughput rate for countercurront transport of the fatty acid monoester, and the transesteri~ication reaction rate which i8 prov$ded by the enzyme to achieve transe~terification of the triglyceride and the fatty acid or fatty acid monoester. Fatty acid low-r alkyl monoesters are substantlally more soluble in the supercritlcal fluid than the corresponding acids, and accordingly are preferred reactants. The supercritical ga~
also serve~ as a diluent of the triglyceride phase to increase the reaction rate. If it is desired to operate the countercurrent column at a temperature higher than the lipase enzyme can tolerate, a plurality of enzyme reaction - 23 ~
zones 215 may be distributed along the column 214 which may be operated at a lower temperature, with appropriate heating and cooling means for fluid pumped therebetween.
In this manner, the transesterification reaction and countercurrent gradient functions may be separately maximized.
The 8upercritical carbon dioxide is less dense than the downwardly moving canola oil stream at pressures and temperatures used in the system of FIGURE 2 (e.g., 35-70- C., 1500-3500 psia). The pressure, temperature, column distances and flow rates of fatty acid or fatty acid monoester and carbon dioxide are selected so that in the zone 228 between the point of introduction of the carbon dioxide and the point 222 of introduction of the unsaturated fatty acid or monoester, the fatty acid or fatty acid monoester is progressively dissolved from the triglyceride into the upwardly moving supercritical CO2 stream; the acid or fatty acid monoester components (including the transesterified components) are substantially completely removed from the triglyceride stream 226 before it i~ discharged from the column at outlet 218. In this regard, the weight ratio of the flow rate of the carbon dioxide to the flow rate of the unsaturated acid or monoester component 220 introduced in the column 214 may de~irably be oelected to be in the range of from about 5:1 to about 50:1, under conditlons to maximize solubility o~ the ~atty acid or preferably fatty acid monoester component while minimizing the solubility of the triglyceride component phaso. In the zone 224, during the time o~ transit o~ the canola oil (e.g., .25 - 6 hours), the unsaturated acid component 220 undergoes transesterification with the triglyceride component.
Because the flow of triglyceride, and acid or monoester is cocurrent in this ~tripping zone, the enzymatic transesterification reaction will tend to approach the equilibrium condition of the unsaturated acid ~3~ $~
, monoester-triglyceride blend at the point 222 of introduction of the monoester 220. Accordingly, the composition of the fatty acid or fatty acid monoester which enters the countercurrent transesterification zone 226 from the monoester stripping zone 224 will be different from the composition of the fatty acid or monoester 220 introduced into the column 214 at least in part because of the transesterification which occurs in the stripping zone 224. The transesterified triglyceride product, which may have substantially all fatty acid and fatty acid monoester components removed therefrom, is withdrawn from outlet 218.
The ratio of triglyceride components to the unsaturated acid or monoester component 220 to achieve a desired degree of transesterification of the canola oil triglyceride is substantiàlly greater in the system of FIGURE 2 than the ratio of triglyceride to fatty acid or monoester utilized to achieve an equivalent degree of transesterification in a one or two step batch reaction.
In this regard, the unsaturated acid or monoester 220 is introduced into the bottom of the column at a rate compared to the rate of introduction of canola oil which ~ay, for example, be about half the proportion used in a batch reaction (e.g., 1:3 to 1:1 weight ratio of unsaturated acid component to canola oil).
The fatty acid or monoester component i5 dissolved in the upwardly movin~ supercritical C02 gas stream and carried into the transesterification zone 226, where lt approaches equilibrium with the countercurrent oil flow through the action of the immobilized enzyme in the coluLn. It is further transQsterified in a countercurrent manner with the liquid triglyceride ~tream a~ it is oonducted from its point of introduction 224 to the point 220 of introduction of the fatty acid monoester.
The triglyceride phase mixture continuously undergoes transesterification r~action as it moves 2 ~
downwardly in the zone 226 containing lipase enzyme countercurrent to the flow of supercritical gas, such that the mixture has an increasing concentration of the desired triglyceride components as it moves down the column. There is also an increasing concentration of transesterified fatty acid or monoester having fatty acid or monoester components derived from the triglyceride in the upwardly moving supercritical gas stream, in the direction toward the point of introduction of the triglyceride. Water vapor may be included in the carbon dioxide flow, the fatty acid ester flow and/or the triglyceride flow to accommodate the transesterification reaction, which may exceed the solubility of water in the triglyceride component, and to produce a desired level of diglyceride~, lf desired. Fatty acid components produced by hydrolysis reactions in the column 214 may also be removed by the supercritical carbon dioxide flow.
The transesterified fatty monoester or fatty acids dissolved in the supercritical C02 gas stream i8 carried from the column at outlet 216, through a pressurQ let-down system, where dissolved fatty acids or monoesters are taken out of supercritical solution as a result of pressure reduction. Th- tank 232 may alternatively be heated to further reduce the solubility of the fatty acid monoester.
The solubility reduction may al80 bo accompli~hed by a combination of a limlt-d pr-s~ur- r-duction (-.g., by 500-1000 p~i) and a temperatur- incr-a~ .g., to 70-lOC- C.) so that the work to rQcompress the C02 for recycle use may be reduced. The energy recovery sy~tem may comprise a piston or turbine ngine 232 in which the di~solved fatty acid or monoester 234 are collected in the rocovery system, 80 that the energy may be recovered for recompression of the carbon dioxide upon recyclic operation. ~he lower pressure C02, which is depleted in or froe of dissolved fatty acids or monoesters, is recompressed by compressor 238.
- 26 - 2~
The carbon dioxide 2~6 which is separated from the fatty acid or monoester is conducted to compressor/thermal conditioner 238 where it is recompressed and reintroduced at the preselected operating temperature as previously discussed A heat-pump or other thermal connector 240 may be used to transfer heat between the compressor 238 and the decompression engine 232 A portion of the extracted fatty acid components 234 may be recycled for reflux purposes to increase selectivity The flow rate of supercritical carbon dioxide (or other supercritical gas solvent) through the column 214tis correlated with the flow rate of fatty acid ester 220 so that it is adequate to transport and dissolve 6ubstantially all of the fatty acid monoester under the operating conditions, but dissolves a minimal amount of the initial canola oil and other triglyceride components The solubility of the fatty acid or fatty monoester components will desirably be greater than 1 weiqht percent, and preferably greater than 2 weight percent, while the solubility of triglycerides will be less than 0 5 weight percent and preferably d$ssolved triglycerides will be less than 10 percent of the dissolved fatty acid or monoester in the extracted product 234 The separated fatty acid or monoester 234 ~ay be purified in fractionation system 250 to separate purified unsaturated fatty acid- 220 from oth-r material~ 252 in any suitable mann-r, and the unsaturated transesterification component 220 may be recycled for transesterification use Another embodiment of a continuous countercurrent transesterification process iB illustrated in FIGURE 3, which moves a ~atty acid or fatty acid monoester component countercurrent to triglyceride ~low, utilizing a subcritical liquified ga~ solvent As 6hown in FIGURE 3, the vegetable canola oil 312 to be transesterified is introduced into a high pressure column 314 at a point 324 intermediate the upper outlet 316 and the lower outlet 318 The rate of introduction 2 ~ j L': ~
corresponds to the transesterification reaction rate permi~-ted by the activity of the immobil$zed enzyme in the column 314. The ~olumn may be packed with an immobilized lipase enzyme as previously described, which is immobilized ! on an inorganic, high surface area substrate ~uch as porous ceramic rings or pellets, or d$atomaceous earth (e.g., ~elite), or a suitable organic substrate such as an ion ? exchange resin substrate (e.g., Novo 3A lipase). Such packing may be in particulate film or fiber form, on plates, etc. Alternatively, as shown in FIGU~E 3, the column 314 may be used to establish concentration gradients and countercurrent flow in stages, and the immobilized I enzyme 3$3 may be located in corresponding stages in a separate pressure vessel 315. The surface area of the j column packing should be very large in order to promote I transesterification reaction (e.g., more than 750 square I meters of surface area per cubic meter), and to promote ! equilibrium dissolution of the low molecular weight ! compGnents in the supercritical fluid.
! A fatty acid or lower alkyl fatty acid monoester i 320, such as a methyl or ethyl ester of an unsaturated ¦ fatty acid component (e.g., a mixture of unsaturated fatty ! acids or unsaturated fatty acid esters such as methyl or ; ethyl esters), which is desired to be transesterified with the triglycerido 312, i8 introduced into tho column 314 at a point 322 intermediate the point 324 o~ introduction o~
triglyceride, and the lower outlet 318 at a rate which ! maxi~izes the desired transesterification reaction. For mnnufacture of the low saturated fat vegetable oil, a I mixture of unsaturated fatty acids or monoesters comprising I le~s than 2 weight percent of saturated fatty acids is used. The rate of introduction i8 about 0.25 to 1.0 the rate of vegetable oil input, lower ratios being used for higher purity ~e.g., less than 0.5 or 0.25 weight percent saturated fatty acid content). The transesterification reaction is conducted in a countercurrent manner, 80 that ~ 28 it is substantially more eff~cient than the batch reaction of the fatty acid and triglyceride component at the selected reaction ratios.
In operation, liquified propane 309 i8 introduced at a position 319 near the bottom of the column 314 under pressure, flow rate and temperature conditions at which relatively low molecular weight fatty acids and monoesters are significantly dissolved, but at which the high molecular weight triglycerides are less ~oluble and form a separate phase. The temperature, of course, cannot exceed the operating temperature of the enzyme, whlch will be damaged at high temperatures. In the illustrated embodiment, the propane may be introduced at a pressure of about 600 psi and a temperature of 70- C. at a rate of about 10 t~mes the rate of introduction of fatty acid 320 on a weight ratio basis. If a lower temperature is necessary, ethane or ethane/propane mixtures may be used.
The liquified propane gas also dissolves to some extent in the triglyceride phase and serves as a diluent to increase the reaction rate.
The liguified propane phase i6 less dense than the downwardly moving veqetable oil stream, having a density of 0.25-0.4 g/cm~, depending primarily on composition and temperature. ~he pressure, temperature, column distance~ and flow rate~ Or ratty acid~ ~or ~onoesters) and liquid propane are selected 80 that in the zone 3~4 between the point 319 of introduction of the pure liquified propane and the point 322 of introduction of the unsaturated fatty acid or monoester 320, the fatty ac~d or monoQster components are progressively dlssolved from the triglyceride into the upwardly moving liquid propane phase;
the fatty acid or monoester ~s substantially completely removed from the triglyceride ~tream 326 before it is discharged from the column at outlet 318. Because of countercurrent transesterification which occurs in the transesterlfication zone~ 313 (or in the column 314 in embodiments having lipase enzyme there~n), the fatty acids or monoesteræ in the upwardly rising liquid propane droplets will be different from the composition of the fatty acids 320 introduced into the column 314 because of the transesterification. The transesterified triglyceride product i8 withdrawn from outlet 318. It contains an amount of propane which is an inverse function of temperature (e.g., 30-60 weight percent propane), which is removed in propane stripping column 327 and returned for i recyclic use.
The ratio of triglyceride component 312 to the unsaturated fatty acid component 320 to achieve a desired degree of transesterification of the triglyceride will be substantially groater in the countercurrent system than the ratlo of triglyceride to fatty monoester necessary to achieve an equivalent degree of transesterification in a batch or cocurrent reaction, as previously discussed.
Moreover, high levels of transesterification substitution may be obtained with the proposed countercurrent process, which could otherwise only be achieved with multiple batch reaction, and multiple component ~eparation.
The fatty acid component is dissolved in the upwardly moving propane phase, and is continuously exchanged with the downwardly moving vegetable oil pha~e, creating a gradlent along th- column, and carried into th-tran~esterification zone, wher- it tends toward dynamic equilibrium with the countercurrent oil flow. It 18 transesterified ln a countercurrent manner with the liquid trlglyceride stream ~- it i~ conducted from the point of introduction 324 to the point 320 of introduction of the fatty acid monoe~ter.
A portion of the mixture is pumped from mixing ~tage~ in the column 314 to transesterification zones 313 containing a suitable transesterirication enzyme. The mixture continuously undergoes transe~terification reaction in the separate zones 313, and i~ returned to the
3 1~ ~
respective phase separation zones of the column 314, such that the mixture has an increasing concentration of the desired triglyceride as it moves down the column. It is noted that the columns 314 may be maintained at a higher temperature (e.g., 70-90- C.) at which phase separation is enhanced, while the lipase reaction zones 313 may be maintained at a lower temperature (e.g., 50-60- C.) at which the enzyme longevity and reaction are maximized, by appropriate heating and cooling of the streams conducted j there between. It is also noted that by cooling the liguid phases which increase mutual solubility (toward and including miscibility), and reheating them, which causes phase separation, extraction efficiency may be increased.
Accordingly, there is also an increasing concentration of transesterified fatty acid components derived from the triglyceride in the upwardly moving liquified gas stream, in the direction toward the point of introduction of the triglyceride. Water vapor may be included in the liquified gas phase flow, the fatty acid flow and/or the triglyceride flow to accommodate the transe~terification reaction, and which may equal or exceed the solubility of water in the triglyceride component, if desired, as previously discussed. Any fatty acid components, as well as monoglycerides (and diglycerides) produced by hydrolysis reactions in th- column 314 may al-o b- ~at l-ast partially) removed by the upwardly moving llquified propane phas- ~which may be facilitated by a small amount of water or ethanol in the propane phase).
~ he transesterified fatty acid component which is preferentially di~solved in the upwardly moving liquid propane phase, is carried from the column at outlet 316 at .g., 70-85- C., through heater/evaporator 330 where it is heated to 93- C. to evaporate pure propane 331 and then into separation tank 332, where dissolved fatty acid components 335 are taken out Or solution as a result propanQ evaporation and tomperature lncrea~e. In thl~
~ - - 31 - 2~
regard, the exiting propane phase is heated to further reduce the solubility of the fatty acid (and any triglycerides), without deactivatinq the enzyme, which is not present in the tank 332 or the upper reflux portion of the column 314. The liguid propane exiting the column may be heated to 94-95- C., to create two phases in the separation tank or column 332. Of course, all of the propane may be evaporated to recover the fatty acid components, but this is not necessary, particularly in view of the amount of propane utilized. A portion (e.g., 20-50 weight percent) of the separated fatty acid component may be reintroduced into the top of the column 314 into a roflux zone to enhance the ~electivity of the countercurrent transesterification reaction component separation. The remaining portion of the fatty acid material 33S, which contains 30-60 weight percent propane a~ well as some di and tr;glycerides (and perhaps very small amounts of monoglycerides) in addition to the fatty acid or monoester components, i~ conducted to a propane stripper 350 and from there the propane-free fatty acid components 352 (which may optionally first be hydrolyzed as previously described) are conducted to an appropriate separation system 352 for separation of the unsiaturated fatty acid components in relatively pure form for recycle u8e- Th- Baturated fatty a¢id components 354 are ~-parat-d from th- un~aturated fatty acid component~ ln a suitable separator 3S5. m e ~atty acids (and any mono, di- and triglyceridQs) may be further utilized as desir-d. The un~aturated fatty acid components may be esterified with ethanol and recycl-d.
Th- liguid propane which is separated from the fatty acids in the tank 332 is conducted to preheat the canola oil 312 and then to pump/thermal conditloner 334 where it i8 cooled to 70- - 75- C. and reintroduced at the preiselected operating temperature. Because it has a very small amount of product fatty acids, it is introduced at an intermediate position in the column 314. A portion of the propane is evaporated at heater/propane bo~ler 330 and recompres~ed to produce a pure propane stre~m for fully 6tripping the fatty acids and lower molecul~r weight components upon introduction at the bottom 319 of the reactor 314. The dissolved fatty acid or noester components may also be ~eparated by heating the propane stream in heater 330 to a temperature above the critical temperature (96-98- C.) of the propane (e.g., 100-110- C.) while maintaining the system at a pressure ~lightly above the critical pressure (42 atmospheres), such as 44-50 atmospheres. In this way, a substantial portion of the components 335 are separated in column 332, and the supercritical propane gas may be reliquified by cooling to a temperature below the critical temperature for reintroduction into the column 314. Such a substantially isobaric procedure minimizes heat and pumping expense.
The flow rate of liquified gas solvent through the column 314 is correlated with the flow rate of fatty acid ester 320 so that it i8 adequate to dissolve 6ubstantially all of the fatty acid or monoester under the operating conditions and remove it from the transesterified triglyceride 321.
Ag previously discus~Qd, urea adduction may be utilized to separate saturated fatty acid components from unsaturated fatty aclds or monoe~ter~ to pro~ide a substantially pure unsaturated fatty acid reaction component. Although urea typically crystallizes in a tetragonal form, it forms complexes with ~traight chain fatty acids and lower alkyl mono2sters in which the straight chain fatty compound i8 included within a hexagonal crystalline urea framework to form urea inclusion compounds having a weight ratio of approximately 3:1 of urea to the included compound. In general, saturated fatty acids form more 6table urea complexes than un~aturated fatty acids of the same carbon chain length. The stability constants decrease by an order of magnitude in the 6eries ~ ~ ~ 33 2 ~ 3 ~ v~ ~ ~
stearic, oleic and linoleic acids, respectively [Chapter XX
Techniques of Separation E. Urea Complexes, p. 2309, et seg., K.S. Markley, Ed., ~upra]. Saturated monoesters may have qreatly increased stability. The stability of fatty ac~d complexes also increases with the carbon chain length of the fatty acid, and saturated monoesters of lower alkyl alcohols may have enhanced stability.
The urea complexes may be readily decomposed by adding water or other solvent to dissolve the urea, leaving the saturated fatty acid inclusion compounds as an oil or solid, depending upon the temperature of decomposition. A
small quantity of an acid such as hydrochloric acid may be utilized to prevent formation of emulsions by traces of ammonia soaps. Conversely, heating the urea complex with a solvent such as hydrocarbon ~olvent in which the urea is insoluble may also be used to extract the included saturated fatty acid compound.
In order to separate saturated fatty acids (or monoesters) from unsaturated fatty acids (or monoesters), insufficient urea to combine with all the complex forming components of a mixture is added, ~uch that the saturated component will co~bine with the urea and preferentially precipitate with respect to the less stable unsaturated fatty acid complexes. Utilizing di~ferencQ6 in complex-~orming capacity, highly purlfled unsaturated ~atty acids may be isolated ~rom varlous natural ~ources for use in transesterification processes of the present invention as previously described.
Although urea complexes have the melting point of urea, which is approximately 133- C., urea adducts become less stable with increasing temperatures and decompose at a temperature below the melting point of urea, which is characteristic for a specific complex. The dissociation temperature for stearic acid/urea adduct in the absence of any solvent is about 126- C. The dissociation temperature of the palmitic acid/urea adduct in ths absence of any - 34 - ~ ?~
solvent is about 114- C., and the dissociation temperature of the oleic acid/urea compound in the absence of any solvent is about 110- C. These differences may be utilized in refining and separation processes.
The preferential activity of saturated fatty acids to form urea complexes may be used to remove substantially ¦ all of the stearic acid, and a substantial portion of the palmitic acid content of a saturated fatty acid stream in a liquid/solid countercurrent distribution method of separation. For example, the fatty acids and urea dissolved in appropriate solvent may be provided as a moving liquid phase, while the precipitated reaction products may serve as the stationary solid phase. The character of the distribution curve obtained for a given mixture of fatty acids depends on the differences in the I dlstribution coefficients for the individual acids when they are distributed between solid inclusion compounds and the organic solvent [W.N. Sumerwell, J. Am. Chem. Soc., 79, 1 3411-3415 (1957)].
Illustrated in FIGURE 4, a continuous system for economically providing a transesterified low saturate , vegetable oil in which the saturated fatty acids are ! removed by urea/unsaturated fatty acid inclusion reservoir compounds concomitantly with enzymatic acyl transfer reaction. In this regard, an unsaturated ~atty acld/urea reservoir complex may be prepared which ~erve~ as an ! exchange reservoir for removal of saturated ~atty acids.
¦ Such unsaturated exchange reservoir urea inclusion I compounds may be prepared in a suitable manner such as shown in FIGURE 4 by dissolving canola oil fatty acids or a fatty acid lower alkyl monoesters 402, (e.g., methyl or ethyl esters) in a suitable ~olvent. Methyl and ethyl esters are preferred because urea adducts of palmitic and stearic monoesters may have substantlally higher 6tability constants than oleic acid or oleic acid monoesters, thereby facilitating saturated component removal.
~ - 3S - 2~31~s~
The excbange reservoir inclusion compound crystals are formed by dis~olving the canola fatty acids or monoe~ters which comprise about 5-8 weight percent palmitic and stearic acids (or monoesters), in a ~uitable amount of a ~olvent such as ethanol or methanol 404 (and bexane if necessary), together with about 30-50 weigbt percent of urea 406, based on the weight of the fatty acids The solution is cooled in a crystallization vessel 408 to produce a flrst crop of urea inclusion crystal~ which comprise about lQ-12 weight percent of the fatty acid component The first crop of urea inclusion crystal~ are separated as first crop product 410 The first crop urea inclusion crystals are predominantly urea stearate and palmitate compounds, because of the tendency for the saturated fatty acid (or preferably monoester) to form urea inclusion compound6, leaving in solution at least 98-99%
pure unsaturated ~atty acids or monoesters in the crystallization vessel 601ution 408 An excess of urea, e g , a 7 1 or more weight ratio of urea to remaining canola unsaturated fatty acid~ in the ve~sel 408, is then added and dissolved at elevated temperature Upon cooling, a s-cond crop of urea inclusion compounds is formed which are predominantly urea oleate and linoleate inclusion compound crystals, which ~orm an exchange reservoir inclusion compound material 420 The~- exchange re~ervoir crystals 420 may b- drl-d to remov- ~olvent and u--d with an immobilized llpa-- nzym- 422 uch a~ 1-,3- ~pecific lipa-- from Mucor mlehei lmmobiliz-d on an ion exchang-rosin, such a~ the Novo 3A Lipaso product of Novo described in U S Patont 4,798,793, ln a weight ratio of 2 1 to about 10 1 unsaturat-d fatty acid uroa inclu~ion rosorvoir cry~tal6 to immobilized enzymo/ionic re~in component to form the pac~ing 424 of an intraesterification reaction column 426 The reservoir cry~tal~ are al~o placed, without an immobilized lipa~e component, a~ the packing into a column 430 Tho ur~a reservoir crystals may be !~ ' - 36 _ 2~3~ 9~
blended with, or alternated in layers with the transesterification lipase. Canola oil fatty acids or preferably lower al~yl monoesters 402 are conducted through the column 430 at a temperature in the range of 20~ C., preferably in the range of 40-60- C.
A temperature selected for maximum relative stability of the palmitic acid complex over the oleic acid complex may be selected if desired to maximize removal of the palmitic acid component. Because the urea complexes of stearic and palmitic acid or preferably monoester components of the mixture 402 have a higher stability than the oleic or linoleic complexes, the stearic and palmitic acids or monoesters exchange with these unsaturated fatty acids to form more stable inclusion compounds, thereby producing an unsaturated fatty acid or monoester stream 432 which is substantially free of saturated fatty acids or monoesters. The unsaturated discharge stream 432 may be blended with refined and bleached canola oil 440 in a weight ratio of from about 30:1 to about 1:1 canola oil to fatty acid component, and preferably $n a range of from about 10:1 to about 3:1, to produce a transesterification stream 436. The canola oil transesterification stream 436 may be conducted through a water saturated ionic exchange resin column 434 (e.g., at a temperature of 40-70- C.) to saturate the oil/Satty acld ~or monoester) blond 436 with water and remove impurities whlch might deactlvate the enzyme.
The saturated intraesterification strQam 436 accordingly may have a limited amount of fatty acid component and will require reduced separation treatment after transesterification.
A suitable solvent such as butane, pentane, hexane or pressurized propane 440 may be used to reduce the viscosity of the transesterification stream 436 which i8 introduced into the interesterification column 426. The stream 436 is conducted through the transesterification _ 3~ _ 2~
column 424 where the 1-, 3- fatty acid moieties of the canola oil are progressively released and exchanged with unsaturated fatty acid components of the stream 436 by the acyl transfer activity of the immobilized enzyme. Such unsaturated fatty acid components may be initially present in the stream 436, may be produced by a 6mall amount of hydrolysis from the water content of the stream, or may be derived from the unsaturated fatty acid urea adduct reservoir material 420. In this regard, as the saturated fatty acid moieties are released by the transesterification reaction, they undergo equilibration reaction with the reservoir urea inclusion compound crystals of the packing 424 and are exchanged with the unsaturated fatty acid components of the inclusion crystals. The efficiency of the overall production process i8 enhanced by close proximity of the enzyme and the inclusion compounds. In this manner, the saturated fatty components of the canola oil are progressively removed as the stream 436 is conducted through the column 426. An increase in diglyceride and fatty acid content will occur as a result of the small water content of the ~tream 432. When using unsaturated fatty monoesters, a vacuum may be periodically or continuously applied to the column 426 with a slow nitrogen bleed to remove monohydric alcohol, increase the triglyceride content and decrease the diglyceride content of the stream, if desired.
The 6tream 436 i9 conducted through the column 426 at a rate, combined with the transesterification rate and inclusion compound exchange rate, which produces an output stream 450 comprising less than 3 weight percent of saturated fatty acid content based on the total weight of the 6tream. The processing may be continued until the ~aturated fatty acid exchange capacity of the urea inclusion reservoir component 424 or the component 420 of the column 430 are exhausted. The output stream 450 may be refined in any appropriate manner such a8 countercurrent ~ 4~ ~ L~
solvent processing, to remove the fatty acid or monoester and any small amounts of urea. The low saturate triglyceride mixture may alternatively, or subsequently, be deodorized by conventional steam deodorization or supercritical carbon dioxide deodorization to produce a very low æaturate vegetable oil product 460.
The urea inclusion reservoir compohent column packings 420, 424 may be replaced or regenerated periodically. Such replacement and regeneration may be accomplished by redissolving the urea in a hydroxy solvent such as methanol to release and separate the saturated fatty acid inclusion compound, and recycling the urea in solvent-reformed unsaturated fatty acid reservoir crystals. However, lt is preferred to regenerate the hexagonal urea compound without dissolving it, using solvent or fatty acid or monoester components which are compatible with and/or easily separable from, the adduct.
In this regard, for example, the packing 420 may be regenerated by passing therethrough a stream of a solvent such as hexane, liqui~ied propane and/or unsaturated fatty acids 402 (which preferably may have had the saturated fatty acids removed therefrom) at a temperature sufficient to release the saturated fatty acid inclusion components, which may be up to a temperature above the decomposition temperature of the stearate ~nd palmltate inclu~ion compounds (e.g., about 126- C.), but below the melting temperature o~ the urea (133- C.1. An output ~tream high in stearic and palmitic acids i8 produced, which may be utilized or reprocessed as desired. The thermal stability limit for the enzyme should not be exceeded by such regeneration treatment if the enzyme is mixed or treated with the urea reservoir material. Layering as previously described permits separate regeneration treatment of the reservoir material. After releasing the saturated inclusion compounds from the packing 420 in this manner, it may be cooled in the presence of unsaturated fatty Acids ~31~
,.
compounds, thereby reforming the re6ervoir material as an unsaturated fatty acid complex Low molecular weight hydrocarbons such as hexane or petroleum ether, which form relatively unstable inclusion compounds, may be used to extract the 6aturated fatty acid component in such regeneration, or solvents which do not form an inclusion compound, such as liguified propane may be used It may be desirable to remove all such solvent following regeneration by methods such as vacuum treatment Regeneration may be carried out on a continuous countercurrent basis with the "spent" inclusion compound packing 424 being continuously or periodically withdrawn from the bottom (countercurrent inlet) and regenerated material added at the top transesteri~ied oil outlet While described for urea/unsaturated fatty acid adducts, other suitable adsorbent or adduct forming materials may be used which preferentially adsorb saturated fatty acids or monoesters over un6aturated fatty acids or monoesters For example, X and Y type zeolites and/or 6ilicalite having appropriate pore 6tructures which preferentially absorb saturated fatty acids may be regenerated or ~recharged~ with purified unsaturated fatty acids or monoesters, and us-d in admixture with the immobilized enzyme Countercurrent moving bed ~yst-m~ or simulated moving countercurr-nt bed systems may be used to facilitate continuous operation Whil- the countercurrent methods disclosed herein have been spocifically described with respect to low-~aturate vegetable oil manufacture, 6uch methods may be generally applied to enzymatic and base-catalyzed transesterification reaction~ of vegetable, marine and animal fats and oils Having generally described various aspects of the present low saturate vegetable oil products and methods which may be utilized to prepare such products, the invention will now be more particul~rly described with respect to the following specific examples .
Example ~
~ $~1 and 2 - Two different lots of canola o$1 were transesterified with oleic and linoleic acids.
Transesterification reactions were carried out in 2.5 ml of hexane, per gram of reactants. Novo 3A Lipase, an ~ immobilized 1-, 3- positionally specific lipase from Mucor ¦ miehei as previously descr~bed, was used at a level of 1 0.375 gram, per gram of oil. The transesterification ¦ reactions were run in a water bath at 40- C., under 250 rpm ¦ agitation for 6 hours. The reactions were stopped by removing the immobilized lipase by filtration. The hexane was removed by distillation. The reaction mixture at this point was deodorized under a vacuum of O.lmm Hg to a maximum temperature of 500- F. The product was then analyzed for fatty acid distribution (FAD).
The FAD may be resolved into the five ma~or fatty acids, palmitic (P), stearic (S), oleic (O), linoleic (L), linolenic (Ln) and all the remaining fatty acids designated as "othern. A typical FAD of the fatty acid components is set forth in the following Table 1.
Table 1 Oleic Linoleic Acid Acid _ P 0.51 ~~~
', S 0.17 0.04 r O 98.46 0.67 L 0.27 9B.85 Ln --- 0.06 Other 0.59 0.38 ! The weight of reactants used in these runs was as follows:
! Table 2 ¦ Weight of Percent Reactants ~a~ by weig,h,~
Canola oil lS7.2 13.58 ' Llnoleic acid 600.0 51.8S
j Oleic acid 400.0 34.57 I ThesQ amounts were ~elected based on a "target"
transesterified composition having a weight ratio of ~3~
esterified monounsa~urated fatty acids to polyunsaturated fatty acids of about 1:1. Following the reactions, the fatty acid distribution (FAD) of the products was determined, as set forth $n Table 3:
Table 3 Starting Material Canola Target % Oil~ Run 1 for Run 1 P 4.15 2.78 0.89 S 1.81 1.39 0.35 o 56.74 53.78 45.71 L 19.97 30.83 45.53 Ln 7.85 6.73 5.67 Other 9.39 4.49 1.82 Total Sats 6.03 4.26 1.26 (C12 -18 ) ¦ Starting Material Canola Target*
% Qil~ Run 2 Run 3 2 & 3 ! P 4.15 2.91 2.55 0.60 S 1.81 1.26 1.22 0.22 O 56.74 52.61 51.63 44.71 L 19-97 33.60 35.34 47.83 Ln 7.85 6.49 6.32 5.47 Other 9.39 3.13 2.94 1.14 Total Sats 6.03 4.17 3.87 0.83 ( C12-18 ) * Starting FAD and t~rgeted FAD are data from one t experiment, but are representative of all three trials ¦ Neither reaction reached its targeted equilibrium. These results suggest that there is some property of canola oil which i8 affectlng the catalytic properties of the ¦ immobilized lipase.
Run 3 - A further reaction was conducted in the same manner with 0.02% TBHQ added as an antioxidant to prevent the formation of peroxides during the reaction which might interfere wlth the transesterification. Also, the lipase enzyme product concentrat~on was doubled (0.750 2 ~ 3 ;~
grams lipase per grhm of oil) to compensate for any other inhibition which might be occuring. Table 3 also illustrates the FAD of t~e product triglycerides. Again, the targeted eguilibrium was not achieved and only a slight decrease in the saturates was observed compared to Runs 1 - and 2.
Run 4 - Runs 1-3 indicated that there was some contaminant in the canola oil which affected the lipase l activity in such a way that the reaction could not reach j equilibrium under those conditions. Canola oil triglycerides were purified by Florisil column chromatography. Canola oil was purified on activated Florisil (dried at 100- C. for 18 hours, then equilibrated to 3 weight percent water at room temperature by the addition of distilled water). A 125 ml volume of activated Florisil was equilibrated in hexane in a column of 2.5 X 50 centimeters. Fifty-five grams of canola oil were eluted with 3 column volumes of hexane. This fraction was collected and was found to be the triglyceride fraction.
This fraction was distilled to re~ove the hexane and used ! as a source of canola oil triglycerides. These purified ! triglycerides were used to determ~ne initial reaction ! conditions in Run 4. Run 4 was carried out in a manner substantially identical to Run 3 using the Flori~il purifled canola oil. Tablo 4 lllustrates thQ ~AD of the ' product triglycerid-~:
! Table I Starting I Material ¦ Canola Target*
% Qil* _ Run #4 Run #5for 4 & 5 P 4.15 1.05 1.02 0.60 S 1.81 0.58 0.63 0.22 0 56.74 48.16 47.33 44.71 L 19.97 43.48 42.95 47.83 Ln 7.85 4.98 5.67 5.47 Other 9.39 1.75 2.40 1.14 Total Sats 6.03 1.64 1.69 0.83 2 ~ 3 ~
Monounsat to Polyunsat 2:1 1:1 1:1 1:1 * See Table 3 Run 4 Florisil purified canola triglycerides with 0.02%
TBHQ - Run 5 canola oil with 0.02% TBHQ
Run 5 - Another lot of canola oil was also transesterified under substantially identical parameters as described for Run 3, including .02 weight percent TBHQ as an antioxidant. Table 4 also illustrates the FAD of this product. This lot did not demonstrate any lipase inhibition since the targeted equilibrium was nearly attained. Thus, tho contaminant causing this inhibition is variable from lot to lot.
Example 2 High oleic sunflower oil and high oleic safflower oil having low levels of saturated fatty acids were transesterified with substantially pure oleic and linoleic acids, substantially as described in Run 4 of Exa~ple 1.
Table 5 illustrates the starting FAD, and the FAD of the transesterified products:
Table 5 High Oleic,,,S,UnflOWer O~l Starting Transester$fied S ~9~Q~ oduct P 3.71 0.44 S 4.1S 0.54 O ' 80.86 47.46 ~ 8.92 49.43 Ln 0.10 0.21 Other 2.26 1.92 Total Sats 7.94 0.98 High Oleic Safflower Oil Startlng Transesterified ~ Material Product P 5.35 '0.55 S 2.26 0.33 O 73.15 47.49 L 17.25 50.99 Ln 0.11 0.05 Other 1.88 0.59 Total Sats 7.82 0.90 ~3~ ~afc-3 Neither oil appeared to demonstrate any inhibition of the lipase, and oils with <1 weight percent saturates were produced.
Example 3 An oil having very low saturated fatty acid content and a 2:1:1 weight ratio of omega-9:omega-6:omega-3 fatty acids is a nutritionally desirable product which is not naturally available. Canola oil was transesterified with purified oleic, linoleic and linolenic acids as previously described for Run 4 of Example 1, with the addition of linolenic acid to the reaction mixture to prepare such a product. Table 6 illustrates these results:
Table-Starting Transesterified %Ma~erial Product Taraet P 4.15 1.31 0.59 I S 1.81 0.79 0.20 56.74 49.14 48.02 L 19.97 21.81 24.49 ~n 7.85 22.05 25.31 ! Other9.39 4.90 1.38 Total Sats 6.03 2.10 0.80 Montunsat8 Polyunsats 2;1 1:1 1:1 , omega-9:
f omega-6:
! omega-37:2:1 2:1:1 2:1:1 I' ~X~!L~
¦ In the previously described runs, the I transesterified oils had a very low target amount of less than 1.3 weight percent saturated fats. To achieve this ¦ level of unsaturation in the ~inal product a ratio of fatty acids: oil of 6.4:1 in the reaction mixture was utilized, in which the reaction mixture contained only 13.5% oil.
Transesterified canola oils having about 3 weiqht percent saturated fats may be more economically produced using 2~3 ~
lower ratios of unsaturated fatty acids to oil in the react~on mixture. The calculations used to produce a canola oil with close to 3% saturates are shown in Table 7 Table 7 Total A~ount R E A C_ T A N__T ~ of Ex- Targeted Fatty Canola Lino- change- Acids at Oil leicOleic able Equilibrium, #1-#3 AcidAcid Fatty Weight Percent ~ 15?.2a 90 ~90 a Acids$1+#3Overall P 6.06 + ---- + 0.46 =6.522.33 1.~7 S 2.72 + 0.036 + 0.15 = 2.91 1.04 0.69 O 59.64 + 0.60 + 88.61 =148.85 53.16 52.42 L 13.42 + 88.97 + 0.24 =102.63 36.66 35.46 i Ln 3.96 + 0.06 + ---- = 4.02 1.44 6.17 Other14.09 + 0.34 + 0.53 ~ 14.96 5.34 3.~9 280.00100.00100.00 Weight of Percent Reactants (q~ by Weight ! Canola Oil 157.2 46.62 Linoleic Acid 90.0 26.69 Oleic Acid 90.0 26.69 100. 00 Product at equilibrium: 2.38% saturates (theoretical) 1 1.15:1 Fatty acids:Oil Theoretically, it should be possible to produce a canola oil having less than or about 3 weight percent 6aturates with a fatty acid:oil ratlo o~ 1.15:~, assuming the ~atty acid ~eedstock has a saturate level o~ 0.72 we$ght percent I or less.
¦ Canola oil (Kraft Food Ingredients Group) was transesterified using the procedure of Run 3, Example 1, with the proportion of reaction components as descr$bed hereinabove. The results ~rom two trials were tabulated in Table 8:
I
~ ~3 ~
T~ble ~
F~D'S~OF l~ansesterified Canola oll Products Starting Material Canola ~_ Oil Run #1 Run #2 Taraet P 4.15 1.86 1.71 1.67 S 1.81 1.14 1.07 0 69 O 56.74 53.12 52.58 52 42 L 19.97 33.35 34.27 35.46 Ln 7.85 5.58 5.03 6 17 Other 9.39 4.95 5.34 3 59 Total Sats 6.03 3.00 2.78 2.38 The FAD results show that the reactions approached their targeted equilibria, to produce a canola oil with less than or about 3 weight percent saturates using a fatty acid:oil reaction weight ratio of 1.15:1 in a batch reaction. The reaction mixture of this Example 4 contains 46.5% oil.
Instead of using a mixture of oleic and linoleic acids, canola oil may be transesterified with a fatty acid mixture I derived from canola oil, from which saturated fatty acids i have been removed. The transesterification reaction may be ¦ carried out using a canola oil un6aturated fatty acid:
¦ canola oil reaction weight ratio of 1.15:1 as described in I Runs 1 and 2 of this Example, to provide a transesterified ! canola oil having less than 3 weight percent esterified ; saturated fatty acids, and an o~ega-9:omega-6:omega-3 ~ esterified unsaturatsd ~atty acid welght ratlo o~ about r ~ 5~7'~~3~ 18S ~ t 0 ! Accordingly, it will be appreciated that in ¦ accordance with the present invention, novel low-saturate vegetable oils and methods for manufacturing such oils have been provided. Such methods can be used to produce oils with preselected unsaturated fatty acid compositions as a function of the composition of the starting oil and the fatty acids used in the reactions.
While the invention has been described with respect to certain 6pecific embodiments, various modifications and adaptations wlll be apparent from the present disclosure, whlch are intended to be within the scope of the following claims.
respective phase separation zones of the column 314, such that the mixture has an increasing concentration of the desired triglyceride as it moves down the column. It is noted that the columns 314 may be maintained at a higher temperature (e.g., 70-90- C.) at which phase separation is enhanced, while the lipase reaction zones 313 may be maintained at a lower temperature (e.g., 50-60- C.) at which the enzyme longevity and reaction are maximized, by appropriate heating and cooling of the streams conducted j there between. It is also noted that by cooling the liguid phases which increase mutual solubility (toward and including miscibility), and reheating them, which causes phase separation, extraction efficiency may be increased.
Accordingly, there is also an increasing concentration of transesterified fatty acid components derived from the triglyceride in the upwardly moving liquified gas stream, in the direction toward the point of introduction of the triglyceride. Water vapor may be included in the liquified gas phase flow, the fatty acid flow and/or the triglyceride flow to accommodate the transe~terification reaction, and which may equal or exceed the solubility of water in the triglyceride component, if desired, as previously discussed. Any fatty acid components, as well as monoglycerides (and diglycerides) produced by hydrolysis reactions in th- column 314 may al-o b- ~at l-ast partially) removed by the upwardly moving llquified propane phas- ~which may be facilitated by a small amount of water or ethanol in the propane phase).
~ he transesterified fatty acid component which is preferentially di~solved in the upwardly moving liquid propane phase, is carried from the column at outlet 316 at .g., 70-85- C., through heater/evaporator 330 where it is heated to 93- C. to evaporate pure propane 331 and then into separation tank 332, where dissolved fatty acid components 335 are taken out Or solution as a result propanQ evaporation and tomperature lncrea~e. In thl~
~ - - 31 - 2~
regard, the exiting propane phase is heated to further reduce the solubility of the fatty acid (and any triglycerides), without deactivatinq the enzyme, which is not present in the tank 332 or the upper reflux portion of the column 314. The liguid propane exiting the column may be heated to 94-95- C., to create two phases in the separation tank or column 332. Of course, all of the propane may be evaporated to recover the fatty acid components, but this is not necessary, particularly in view of the amount of propane utilized. A portion (e.g., 20-50 weight percent) of the separated fatty acid component may be reintroduced into the top of the column 314 into a roflux zone to enhance the ~electivity of the countercurrent transesterification reaction component separation. The remaining portion of the fatty acid material 33S, which contains 30-60 weight percent propane a~ well as some di and tr;glycerides (and perhaps very small amounts of monoglycerides) in addition to the fatty acid or monoester components, i~ conducted to a propane stripper 350 and from there the propane-free fatty acid components 352 (which may optionally first be hydrolyzed as previously described) are conducted to an appropriate separation system 352 for separation of the unsiaturated fatty acid components in relatively pure form for recycle u8e- Th- Baturated fatty a¢id components 354 are ~-parat-d from th- un~aturated fatty acid component~ ln a suitable separator 3S5. m e ~atty acids (and any mono, di- and triglyceridQs) may be further utilized as desir-d. The un~aturated fatty acid components may be esterified with ethanol and recycl-d.
Th- liguid propane which is separated from the fatty acids in the tank 332 is conducted to preheat the canola oil 312 and then to pump/thermal conditloner 334 where it i8 cooled to 70- - 75- C. and reintroduced at the preiselected operating temperature. Because it has a very small amount of product fatty acids, it is introduced at an intermediate position in the column 314. A portion of the propane is evaporated at heater/propane bo~ler 330 and recompres~ed to produce a pure propane stre~m for fully 6tripping the fatty acids and lower molecul~r weight components upon introduction at the bottom 319 of the reactor 314. The dissolved fatty acid or noester components may also be ~eparated by heating the propane stream in heater 330 to a temperature above the critical temperature (96-98- C.) of the propane (e.g., 100-110- C.) while maintaining the system at a pressure ~lightly above the critical pressure (42 atmospheres), such as 44-50 atmospheres. In this way, a substantial portion of the components 335 are separated in column 332, and the supercritical propane gas may be reliquified by cooling to a temperature below the critical temperature for reintroduction into the column 314. Such a substantially isobaric procedure minimizes heat and pumping expense.
The flow rate of liquified gas solvent through the column 314 is correlated with the flow rate of fatty acid ester 320 so that it i8 adequate to dissolve 6ubstantially all of the fatty acid or monoester under the operating conditions and remove it from the transesterified triglyceride 321.
Ag previously discus~Qd, urea adduction may be utilized to separate saturated fatty acid components from unsaturated fatty aclds or monoe~ter~ to pro~ide a substantially pure unsaturated fatty acid reaction component. Although urea typically crystallizes in a tetragonal form, it forms complexes with ~traight chain fatty acids and lower alkyl mono2sters in which the straight chain fatty compound i8 included within a hexagonal crystalline urea framework to form urea inclusion compounds having a weight ratio of approximately 3:1 of urea to the included compound. In general, saturated fatty acids form more 6table urea complexes than un~aturated fatty acids of the same carbon chain length. The stability constants decrease by an order of magnitude in the 6eries ~ ~ ~ 33 2 ~ 3 ~ v~ ~ ~
stearic, oleic and linoleic acids, respectively [Chapter XX
Techniques of Separation E. Urea Complexes, p. 2309, et seg., K.S. Markley, Ed., ~upra]. Saturated monoesters may have qreatly increased stability. The stability of fatty ac~d complexes also increases with the carbon chain length of the fatty acid, and saturated monoesters of lower alkyl alcohols may have enhanced stability.
The urea complexes may be readily decomposed by adding water or other solvent to dissolve the urea, leaving the saturated fatty acid inclusion compounds as an oil or solid, depending upon the temperature of decomposition. A
small quantity of an acid such as hydrochloric acid may be utilized to prevent formation of emulsions by traces of ammonia soaps. Conversely, heating the urea complex with a solvent such as hydrocarbon ~olvent in which the urea is insoluble may also be used to extract the included saturated fatty acid compound.
In order to separate saturated fatty acids (or monoesters) from unsaturated fatty acids (or monoesters), insufficient urea to combine with all the complex forming components of a mixture is added, ~uch that the saturated component will co~bine with the urea and preferentially precipitate with respect to the less stable unsaturated fatty acid complexes. Utilizing di~ferencQ6 in complex-~orming capacity, highly purlfled unsaturated ~atty acids may be isolated ~rom varlous natural ~ources for use in transesterification processes of the present invention as previously described.
Although urea complexes have the melting point of urea, which is approximately 133- C., urea adducts become less stable with increasing temperatures and decompose at a temperature below the melting point of urea, which is characteristic for a specific complex. The dissociation temperature for stearic acid/urea adduct in the absence of any solvent is about 126- C. The dissociation temperature of the palmitic acid/urea adduct in ths absence of any - 34 - ~ ?~
solvent is about 114- C., and the dissociation temperature of the oleic acid/urea compound in the absence of any solvent is about 110- C. These differences may be utilized in refining and separation processes.
The preferential activity of saturated fatty acids to form urea complexes may be used to remove substantially ¦ all of the stearic acid, and a substantial portion of the palmitic acid content of a saturated fatty acid stream in a liquid/solid countercurrent distribution method of separation. For example, the fatty acids and urea dissolved in appropriate solvent may be provided as a moving liquid phase, while the precipitated reaction products may serve as the stationary solid phase. The character of the distribution curve obtained for a given mixture of fatty acids depends on the differences in the I dlstribution coefficients for the individual acids when they are distributed between solid inclusion compounds and the organic solvent [W.N. Sumerwell, J. Am. Chem. Soc., 79, 1 3411-3415 (1957)].
Illustrated in FIGURE 4, a continuous system for economically providing a transesterified low saturate , vegetable oil in which the saturated fatty acids are ! removed by urea/unsaturated fatty acid inclusion reservoir compounds concomitantly with enzymatic acyl transfer reaction. In this regard, an unsaturated ~atty acld/urea reservoir complex may be prepared which ~erve~ as an ! exchange reservoir for removal of saturated ~atty acids.
¦ Such unsaturated exchange reservoir urea inclusion I compounds may be prepared in a suitable manner such as shown in FIGURE 4 by dissolving canola oil fatty acids or a fatty acid lower alkyl monoesters 402, (e.g., methyl or ethyl esters) in a suitable ~olvent. Methyl and ethyl esters are preferred because urea adducts of palmitic and stearic monoesters may have substantlally higher 6tability constants than oleic acid or oleic acid monoesters, thereby facilitating saturated component removal.
~ - 3S - 2~31~s~
The excbange reservoir inclusion compound crystals are formed by dis~olving the canola fatty acids or monoe~ters which comprise about 5-8 weight percent palmitic and stearic acids (or monoesters), in a ~uitable amount of a ~olvent such as ethanol or methanol 404 (and bexane if necessary), together with about 30-50 weigbt percent of urea 406, based on the weight of the fatty acids The solution is cooled in a crystallization vessel 408 to produce a flrst crop of urea inclusion crystal~ which comprise about lQ-12 weight percent of the fatty acid component The first crop of urea inclusion crystal~ are separated as first crop product 410 The first crop urea inclusion crystals are predominantly urea stearate and palmitate compounds, because of the tendency for the saturated fatty acid (or preferably monoester) to form urea inclusion compound6, leaving in solution at least 98-99%
pure unsaturated ~atty acids or monoesters in the crystallization vessel 601ution 408 An excess of urea, e g , a 7 1 or more weight ratio of urea to remaining canola unsaturated fatty acid~ in the ve~sel 408, is then added and dissolved at elevated temperature Upon cooling, a s-cond crop of urea inclusion compounds is formed which are predominantly urea oleate and linoleate inclusion compound crystals, which ~orm an exchange reservoir inclusion compound material 420 The~- exchange re~ervoir crystals 420 may b- drl-d to remov- ~olvent and u--d with an immobilized llpa-- nzym- 422 uch a~ 1-,3- ~pecific lipa-- from Mucor mlehei lmmobiliz-d on an ion exchang-rosin, such a~ the Novo 3A Lipaso product of Novo described in U S Patont 4,798,793, ln a weight ratio of 2 1 to about 10 1 unsaturat-d fatty acid uroa inclu~ion rosorvoir cry~tal6 to immobilized enzymo/ionic re~in component to form the pac~ing 424 of an intraesterification reaction column 426 The reservoir cry~tal~ are al~o placed, without an immobilized lipa~e component, a~ the packing into a column 430 Tho ur~a reservoir crystals may be !~ ' - 36 _ 2~3~ 9~
blended with, or alternated in layers with the transesterification lipase. Canola oil fatty acids or preferably lower al~yl monoesters 402 are conducted through the column 430 at a temperature in the range of 20~ C., preferably in the range of 40-60- C.
A temperature selected for maximum relative stability of the palmitic acid complex over the oleic acid complex may be selected if desired to maximize removal of the palmitic acid component. Because the urea complexes of stearic and palmitic acid or preferably monoester components of the mixture 402 have a higher stability than the oleic or linoleic complexes, the stearic and palmitic acids or monoesters exchange with these unsaturated fatty acids to form more stable inclusion compounds, thereby producing an unsaturated fatty acid or monoester stream 432 which is substantially free of saturated fatty acids or monoesters. The unsaturated discharge stream 432 may be blended with refined and bleached canola oil 440 in a weight ratio of from about 30:1 to about 1:1 canola oil to fatty acid component, and preferably $n a range of from about 10:1 to about 3:1, to produce a transesterification stream 436. The canola oil transesterification stream 436 may be conducted through a water saturated ionic exchange resin column 434 (e.g., at a temperature of 40-70- C.) to saturate the oil/Satty acld ~or monoester) blond 436 with water and remove impurities whlch might deactlvate the enzyme.
The saturated intraesterification strQam 436 accordingly may have a limited amount of fatty acid component and will require reduced separation treatment after transesterification.
A suitable solvent such as butane, pentane, hexane or pressurized propane 440 may be used to reduce the viscosity of the transesterification stream 436 which i8 introduced into the interesterification column 426. The stream 436 is conducted through the transesterification _ 3~ _ 2~
column 424 where the 1-, 3- fatty acid moieties of the canola oil are progressively released and exchanged with unsaturated fatty acid components of the stream 436 by the acyl transfer activity of the immobilized enzyme. Such unsaturated fatty acid components may be initially present in the stream 436, may be produced by a 6mall amount of hydrolysis from the water content of the stream, or may be derived from the unsaturated fatty acid urea adduct reservoir material 420. In this regard, as the saturated fatty acid moieties are released by the transesterification reaction, they undergo equilibration reaction with the reservoir urea inclusion compound crystals of the packing 424 and are exchanged with the unsaturated fatty acid components of the inclusion crystals. The efficiency of the overall production process i8 enhanced by close proximity of the enzyme and the inclusion compounds. In this manner, the saturated fatty components of the canola oil are progressively removed as the stream 436 is conducted through the column 426. An increase in diglyceride and fatty acid content will occur as a result of the small water content of the ~tream 432. When using unsaturated fatty monoesters, a vacuum may be periodically or continuously applied to the column 426 with a slow nitrogen bleed to remove monohydric alcohol, increase the triglyceride content and decrease the diglyceride content of the stream, if desired.
The 6tream 436 i9 conducted through the column 426 at a rate, combined with the transesterification rate and inclusion compound exchange rate, which produces an output stream 450 comprising less than 3 weight percent of saturated fatty acid content based on the total weight of the 6tream. The processing may be continued until the ~aturated fatty acid exchange capacity of the urea inclusion reservoir component 424 or the component 420 of the column 430 are exhausted. The output stream 450 may be refined in any appropriate manner such a8 countercurrent ~ 4~ ~ L~
solvent processing, to remove the fatty acid or monoester and any small amounts of urea. The low saturate triglyceride mixture may alternatively, or subsequently, be deodorized by conventional steam deodorization or supercritical carbon dioxide deodorization to produce a very low æaturate vegetable oil product 460.
The urea inclusion reservoir compohent column packings 420, 424 may be replaced or regenerated periodically. Such replacement and regeneration may be accomplished by redissolving the urea in a hydroxy solvent such as methanol to release and separate the saturated fatty acid inclusion compound, and recycling the urea in solvent-reformed unsaturated fatty acid reservoir crystals. However, lt is preferred to regenerate the hexagonal urea compound without dissolving it, using solvent or fatty acid or monoester components which are compatible with and/or easily separable from, the adduct.
In this regard, for example, the packing 420 may be regenerated by passing therethrough a stream of a solvent such as hexane, liqui~ied propane and/or unsaturated fatty acids 402 (which preferably may have had the saturated fatty acids removed therefrom) at a temperature sufficient to release the saturated fatty acid inclusion components, which may be up to a temperature above the decomposition temperature of the stearate ~nd palmltate inclu~ion compounds (e.g., about 126- C.), but below the melting temperature o~ the urea (133- C.1. An output ~tream high in stearic and palmitic acids i8 produced, which may be utilized or reprocessed as desired. The thermal stability limit for the enzyme should not be exceeded by such regeneration treatment if the enzyme is mixed or treated with the urea reservoir material. Layering as previously described permits separate regeneration treatment of the reservoir material. After releasing the saturated inclusion compounds from the packing 420 in this manner, it may be cooled in the presence of unsaturated fatty Acids ~31~
,.
compounds, thereby reforming the re6ervoir material as an unsaturated fatty acid complex Low molecular weight hydrocarbons such as hexane or petroleum ether, which form relatively unstable inclusion compounds, may be used to extract the 6aturated fatty acid component in such regeneration, or solvents which do not form an inclusion compound, such as liguified propane may be used It may be desirable to remove all such solvent following regeneration by methods such as vacuum treatment Regeneration may be carried out on a continuous countercurrent basis with the "spent" inclusion compound packing 424 being continuously or periodically withdrawn from the bottom (countercurrent inlet) and regenerated material added at the top transesteri~ied oil outlet While described for urea/unsaturated fatty acid adducts, other suitable adsorbent or adduct forming materials may be used which preferentially adsorb saturated fatty acids or monoesters over un6aturated fatty acids or monoesters For example, X and Y type zeolites and/or 6ilicalite having appropriate pore 6tructures which preferentially absorb saturated fatty acids may be regenerated or ~recharged~ with purified unsaturated fatty acids or monoesters, and us-d in admixture with the immobilized enzyme Countercurrent moving bed ~yst-m~ or simulated moving countercurr-nt bed systems may be used to facilitate continuous operation Whil- the countercurrent methods disclosed herein have been spocifically described with respect to low-~aturate vegetable oil manufacture, 6uch methods may be generally applied to enzymatic and base-catalyzed transesterification reaction~ of vegetable, marine and animal fats and oils Having generally described various aspects of the present low saturate vegetable oil products and methods which may be utilized to prepare such products, the invention will now be more particul~rly described with respect to the following specific examples .
Example ~
~ $~1 and 2 - Two different lots of canola o$1 were transesterified with oleic and linoleic acids.
Transesterification reactions were carried out in 2.5 ml of hexane, per gram of reactants. Novo 3A Lipase, an ~ immobilized 1-, 3- positionally specific lipase from Mucor ¦ miehei as previously descr~bed, was used at a level of 1 0.375 gram, per gram of oil. The transesterification ¦ reactions were run in a water bath at 40- C., under 250 rpm ¦ agitation for 6 hours. The reactions were stopped by removing the immobilized lipase by filtration. The hexane was removed by distillation. The reaction mixture at this point was deodorized under a vacuum of O.lmm Hg to a maximum temperature of 500- F. The product was then analyzed for fatty acid distribution (FAD).
The FAD may be resolved into the five ma~or fatty acids, palmitic (P), stearic (S), oleic (O), linoleic (L), linolenic (Ln) and all the remaining fatty acids designated as "othern. A typical FAD of the fatty acid components is set forth in the following Table 1.
Table 1 Oleic Linoleic Acid Acid _ P 0.51 ~~~
', S 0.17 0.04 r O 98.46 0.67 L 0.27 9B.85 Ln --- 0.06 Other 0.59 0.38 ! The weight of reactants used in these runs was as follows:
! Table 2 ¦ Weight of Percent Reactants ~a~ by weig,h,~
Canola oil lS7.2 13.58 ' Llnoleic acid 600.0 51.8S
j Oleic acid 400.0 34.57 I ThesQ amounts were ~elected based on a "target"
transesterified composition having a weight ratio of ~3~
esterified monounsa~urated fatty acids to polyunsaturated fatty acids of about 1:1. Following the reactions, the fatty acid distribution (FAD) of the products was determined, as set forth $n Table 3:
Table 3 Starting Material Canola Target % Oil~ Run 1 for Run 1 P 4.15 2.78 0.89 S 1.81 1.39 0.35 o 56.74 53.78 45.71 L 19.97 30.83 45.53 Ln 7.85 6.73 5.67 Other 9.39 4.49 1.82 Total Sats 6.03 4.26 1.26 (C12 -18 ) ¦ Starting Material Canola Target*
% Qil~ Run 2 Run 3 2 & 3 ! P 4.15 2.91 2.55 0.60 S 1.81 1.26 1.22 0.22 O 56.74 52.61 51.63 44.71 L 19-97 33.60 35.34 47.83 Ln 7.85 6.49 6.32 5.47 Other 9.39 3.13 2.94 1.14 Total Sats 6.03 4.17 3.87 0.83 ( C12-18 ) * Starting FAD and t~rgeted FAD are data from one t experiment, but are representative of all three trials ¦ Neither reaction reached its targeted equilibrium. These results suggest that there is some property of canola oil which i8 affectlng the catalytic properties of the ¦ immobilized lipase.
Run 3 - A further reaction was conducted in the same manner with 0.02% TBHQ added as an antioxidant to prevent the formation of peroxides during the reaction which might interfere wlth the transesterification. Also, the lipase enzyme product concentrat~on was doubled (0.750 2 ~ 3 ;~
grams lipase per grhm of oil) to compensate for any other inhibition which might be occuring. Table 3 also illustrates the FAD of t~e product triglycerides. Again, the targeted eguilibrium was not achieved and only a slight decrease in the saturates was observed compared to Runs 1 - and 2.
Run 4 - Runs 1-3 indicated that there was some contaminant in the canola oil which affected the lipase l activity in such a way that the reaction could not reach j equilibrium under those conditions. Canola oil triglycerides were purified by Florisil column chromatography. Canola oil was purified on activated Florisil (dried at 100- C. for 18 hours, then equilibrated to 3 weight percent water at room temperature by the addition of distilled water). A 125 ml volume of activated Florisil was equilibrated in hexane in a column of 2.5 X 50 centimeters. Fifty-five grams of canola oil were eluted with 3 column volumes of hexane. This fraction was collected and was found to be the triglyceride fraction.
This fraction was distilled to re~ove the hexane and used ! as a source of canola oil triglycerides. These purified ! triglycerides were used to determ~ne initial reaction ! conditions in Run 4. Run 4 was carried out in a manner substantially identical to Run 3 using the Flori~il purifled canola oil. Tablo 4 lllustrates thQ ~AD of the ' product triglycerid-~:
! Table I Starting I Material ¦ Canola Target*
% Qil* _ Run #4 Run #5for 4 & 5 P 4.15 1.05 1.02 0.60 S 1.81 0.58 0.63 0.22 0 56.74 48.16 47.33 44.71 L 19.97 43.48 42.95 47.83 Ln 7.85 4.98 5.67 5.47 Other 9.39 1.75 2.40 1.14 Total Sats 6.03 1.64 1.69 0.83 2 ~ 3 ~
Monounsat to Polyunsat 2:1 1:1 1:1 1:1 * See Table 3 Run 4 Florisil purified canola triglycerides with 0.02%
TBHQ - Run 5 canola oil with 0.02% TBHQ
Run 5 - Another lot of canola oil was also transesterified under substantially identical parameters as described for Run 3, including .02 weight percent TBHQ as an antioxidant. Table 4 also illustrates the FAD of this product. This lot did not demonstrate any lipase inhibition since the targeted equilibrium was nearly attained. Thus, tho contaminant causing this inhibition is variable from lot to lot.
Example 2 High oleic sunflower oil and high oleic safflower oil having low levels of saturated fatty acids were transesterified with substantially pure oleic and linoleic acids, substantially as described in Run 4 of Exa~ple 1.
Table 5 illustrates the starting FAD, and the FAD of the transesterified products:
Table 5 High Oleic,,,S,UnflOWer O~l Starting Transester$fied S ~9~Q~ oduct P 3.71 0.44 S 4.1S 0.54 O ' 80.86 47.46 ~ 8.92 49.43 Ln 0.10 0.21 Other 2.26 1.92 Total Sats 7.94 0.98 High Oleic Safflower Oil Startlng Transesterified ~ Material Product P 5.35 '0.55 S 2.26 0.33 O 73.15 47.49 L 17.25 50.99 Ln 0.11 0.05 Other 1.88 0.59 Total Sats 7.82 0.90 ~3~ ~afc-3 Neither oil appeared to demonstrate any inhibition of the lipase, and oils with <1 weight percent saturates were produced.
Example 3 An oil having very low saturated fatty acid content and a 2:1:1 weight ratio of omega-9:omega-6:omega-3 fatty acids is a nutritionally desirable product which is not naturally available. Canola oil was transesterified with purified oleic, linoleic and linolenic acids as previously described for Run 4 of Example 1, with the addition of linolenic acid to the reaction mixture to prepare such a product. Table 6 illustrates these results:
Table-Starting Transesterified %Ma~erial Product Taraet P 4.15 1.31 0.59 I S 1.81 0.79 0.20 56.74 49.14 48.02 L 19.97 21.81 24.49 ~n 7.85 22.05 25.31 ! Other9.39 4.90 1.38 Total Sats 6.03 2.10 0.80 Montunsat8 Polyunsats 2;1 1:1 1:1 , omega-9:
f omega-6:
! omega-37:2:1 2:1:1 2:1:1 I' ~X~!L~
¦ In the previously described runs, the I transesterified oils had a very low target amount of less than 1.3 weight percent saturated fats. To achieve this ¦ level of unsaturation in the ~inal product a ratio of fatty acids: oil of 6.4:1 in the reaction mixture was utilized, in which the reaction mixture contained only 13.5% oil.
Transesterified canola oils having about 3 weiqht percent saturated fats may be more economically produced using 2~3 ~
lower ratios of unsaturated fatty acids to oil in the react~on mixture. The calculations used to produce a canola oil with close to 3% saturates are shown in Table 7 Table 7 Total A~ount R E A C_ T A N__T ~ of Ex- Targeted Fatty Canola Lino- change- Acids at Oil leicOleic able Equilibrium, #1-#3 AcidAcid Fatty Weight Percent ~ 15?.2a 90 ~90 a Acids$1+#3Overall P 6.06 + ---- + 0.46 =6.522.33 1.~7 S 2.72 + 0.036 + 0.15 = 2.91 1.04 0.69 O 59.64 + 0.60 + 88.61 =148.85 53.16 52.42 L 13.42 + 88.97 + 0.24 =102.63 36.66 35.46 i Ln 3.96 + 0.06 + ---- = 4.02 1.44 6.17 Other14.09 + 0.34 + 0.53 ~ 14.96 5.34 3.~9 280.00100.00100.00 Weight of Percent Reactants (q~ by Weight ! Canola Oil 157.2 46.62 Linoleic Acid 90.0 26.69 Oleic Acid 90.0 26.69 100. 00 Product at equilibrium: 2.38% saturates (theoretical) 1 1.15:1 Fatty acids:Oil Theoretically, it should be possible to produce a canola oil having less than or about 3 weight percent 6aturates with a fatty acid:oil ratlo o~ 1.15:~, assuming the ~atty acid ~eedstock has a saturate level o~ 0.72 we$ght percent I or less.
¦ Canola oil (Kraft Food Ingredients Group) was transesterified using the procedure of Run 3, Example 1, with the proportion of reaction components as descr$bed hereinabove. The results ~rom two trials were tabulated in Table 8:
I
~ ~3 ~
T~ble ~
F~D'S~OF l~ansesterified Canola oll Products Starting Material Canola ~_ Oil Run #1 Run #2 Taraet P 4.15 1.86 1.71 1.67 S 1.81 1.14 1.07 0 69 O 56.74 53.12 52.58 52 42 L 19.97 33.35 34.27 35.46 Ln 7.85 5.58 5.03 6 17 Other 9.39 4.95 5.34 3 59 Total Sats 6.03 3.00 2.78 2.38 The FAD results show that the reactions approached their targeted equilibria, to produce a canola oil with less than or about 3 weight percent saturates using a fatty acid:oil reaction weight ratio of 1.15:1 in a batch reaction. The reaction mixture of this Example 4 contains 46.5% oil.
Instead of using a mixture of oleic and linoleic acids, canola oil may be transesterified with a fatty acid mixture I derived from canola oil, from which saturated fatty acids i have been removed. The transesterification reaction may be ¦ carried out using a canola oil un6aturated fatty acid:
¦ canola oil reaction weight ratio of 1.15:1 as described in I Runs 1 and 2 of this Example, to provide a transesterified ! canola oil having less than 3 weight percent esterified ; saturated fatty acids, and an o~ega-9:omega-6:omega-3 ~ esterified unsaturatsd ~atty acid welght ratlo o~ about r ~ 5~7'~~3~ 18S ~ t 0 ! Accordingly, it will be appreciated that in ¦ accordance with the present invention, novel low-saturate vegetable oils and methods for manufacturing such oils have been provided. Such methods can be used to produce oils with preselected unsaturated fatty acid compositions as a function of the composition of the starting oil and the fatty acids used in the reactions.
While the invention has been described with respect to certain 6pecific embodiments, various modifications and adaptations wlll be apparent from the present disclosure, whlch are intended to be within the scope of the following claims.
Claims (11)
PROPERTY OR PRIVILEGE IS CLAIMED ARE DEFINED AS FOLLOWS:
1. A transesterified low-saturate liquid vegetable oil comprising less than 3.5 weight percent esterified saturated C12-C18 fatty acids and at least about 96 weight percent esterified unsaturated C12-C22 fatty acids, based on the total fatty acid content of said oil, said esterified C12-C22 fatty acids having a weight ratio of esterified monounsaturated fatty acids to esterified polyunsaturated fatty acids in the range of from about 10:1 to about 1:2.
2. A low-saturate liquid vegetable oil in accordance with Claim 1 wherein said oil comprises less than about 2 weight percent of esterified intermediate C12-C16 saturated fatty acids.
3. A low-saturate vegetable oil in accordance with Claim 1 wherein said oil comprises from about 2 to about 15 weight percent of diglycerides and from about 85 to about 98 weight percent of triglycerides based on the total weight of the oil.
4. A low-saturate vegetable oil in accordance with Claim 1 wherein said oil has a weight ratio of esterified omega-9 unsaturated fatty acids to omega-3 unsaturated fatty acids in the range of from about 9:1 to about 1:1, a weight ratio of omega-9 unsaturated fatty acids to omega-6 unsaturated fatty acids in the range of from about 4:1 to about 1:1 and a weight ratio of omega-6 -unsaturated fatty acids to omega-3 fatty acids in the range of from about 1:3 to about 3:1.
5. A low-saturate vegetable oil in accordance with Claim 2 having a weight ratio of esterified omega-9 omega-6 omega-3 unsaturated fatty acids of about 5-7:2-3:1.
6. A low-saturate vegetable oil in accordance with Claim 2 having a weight ratio of esterified omega-9:omega-6:omega 3 unsaturated fatty acids of about 2:1:1.
7. A low-saturate vegetable oil in accordance with Claim 1 wherein said oil is a transesterified canola oil, high oleic sunflower oil, corn oil, olive oil, peanut oil, high oleic safflower oil, soybean oil or mixtures thereof.
8. An enzymatic transesterification method for preparing a low-saturate vegetable oil in accordance with Claim 1, comprising the steps of providing an unsaturated fatty acid source material selected from the group consisting of unsaturated C12-C22 fatty acids, unsaturated C12-C22 fatty acid monoesters of low molecular weight monohydric alcohols, and mixtures thereof, comprising less than about 2 weight percent of saturated C12-C18 fatty acids, based on the total weight of fatty acids in said source material, providing an edible liquid vegetable oil selected from the group consisting of canola oil, corn oil, olive oil, peanut oil, high oleic safflower oil, high oleic sunflower oil and mixtures thereof, comprising at least about 92 weight percent of esterified unsaturated C12-C22 fatty acids and less than about 8 weight percent of saturated C12-C18 fatty acids, based on the total fatty acids content of the oil, transesterifying said unsaturated fatty acid source material and said liquid vegetable oil with a transesterification enzyme at a weight ratio of unsaturated fatty acid source material to liquid vegetable oil in the range of from about 10:1 to about 1:3 to provide a transesterification mixture, separating transesterified free fatty acid components from the qlyceride components of the transesterification mixture to provide a transesterified liquid vegetable oil oil product comprising less than 3.5 weight percent of esterified C12-C18 saturated fatty acids and a fatty acid component mixture comprising unsaturated fatty acids, unsaturated fatty acid monoesters or mixtures thereof released from said vegetable oil, and separating the fatty acid mixture by removing saturated fatty acid components therefrom to provide an unsaturated fatty acid source material comprising less than 2 weight percent of saturated C12 -C18 fatty acids based on the total fatty acid content thereof for recyclic reaction with said vegetable oil triglyceride.
9. A countercurrent method for preparing a low-saturate liquid vegetable oil comprising the steps of providing a transesterification reaction zone containing a lipase transesterification enzyme, introducing a vegetable oil into the transesterification reaction zone to provide a triglyceride reaction stream through the reaction zone, introducing an unsaturated fatty acid source material selected from the group consisting of unsaturated C12-C22 fatty acids, unsaturated C12-C22 fatty acid lower alkyl monoesters, and mixtures thereof into the transesterification reaction zone to provide a fatty acid or fatty acid monoester reaction stream, conducting a supercritlcal gas or subcritical liquified gas which preferentially dissolves fatty acids and fatty acid monoesters over triglycerides under two-phase conditions through said zone countercurrent to the flow of the triglyceride reaction stream, at a rate and under pressure and temperature conditions to maintain a separate phase of countercurrent fluid containing fatty acid or fatty acid monoester through the reaction zone in intimate contact with the triglyceride reaction stream, carrying out transesterification reaction of the triglyceride stream with the fatty acid or fatty acid monoester stream in the reaction zone, withdrawing a transesterified low-saturate liquid vegetable oil stream which has been transesterified with the unsaturated fatty acid source material from the reaction zone, withdrawing a countercurrent fluid phase from said reaction zone countercurrent to the triglyceride oil reaction stream having dissolved therein transesterified fatty acids or fatty acid monoesters produced by transesterification of the unsaturated fatty acid source material with the liquid vegetable oil, separating saturated fatty acid components from the transesterified fatty acids or fatty acid monoesters to provide a recycle unsaturated fatty acid source material, and introducing the recycle source material into the reaction zone.
10. An enzymatic transesterification method for preparing a low-saturate vegetable oil comprising the steps of providing an unsaturated fatty acid source material selected from the group consisting of unsaturated C12-C22 fatty acids, unsaturated C12-C22 fatty acid monoesters of low molecular weight monohydric alcohols, and mixtures thereof, providing an edible liquid vegetable oil comprising at least about 85 weight percent of esterified unsaturated C12-C22 fatty acids, based on the total fatty acids content of the oil, transesterifying said unsaturated fatty acid source material and said liquid vegetable oil with a transesterification enzyme in a transesterification reaction mixture while selectively adducting saturated fatty acid components from said reaction mixture into a fatty acid reservoir material which preferentially adducts saturated fatty acid components at a weight ratio of unsaturated fatty acid source material to liquid vegetable oil in the range of from about 1:50 to about 1:1 to provide a transesterified mixture, and separating transesterified free fatty acid components from the glyceride components of the transesterified mixture to provide a transesterified liquid vegetable oil oil transesterification product comprising less than 3.5 weight percent of esterified C12-C18 saturated fatty acids.
11. A method in accordance with Claim 10 wherein said reservoir material is a urea adduct of an unsaturated fatty acid or fatty acid lower alkyl monoester.
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US45555589A | 1989-12-18 | 1989-12-18 | |
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CA002031945A Abandoned CA2031945A1 (en) | 1989-12-18 | 1990-12-11 | Low-saturate edible oils and transesterification methods for production thereof |
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KR (1) | KR910011144A (en) |
AU (1) | AU6785490A (en) |
CA (1) | CA2031945A1 (en) |
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WO (1) | WO1991008676A1 (en) |
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EP0519561A1 (en) * | 1991-06-18 | 1992-12-23 | Unilever N.V. | Enzymic triglyceride conversion |
US6262022B1 (en) | 1992-06-25 | 2001-07-17 | Novartis Ag | Pharmaceutical compositions containing cyclosporin as the active agent |
CY1958A (en) * | 1991-06-27 | 1992-06-24 | Sandoz Ltd | Transesterified corn oil products |
GB9113872D0 (en) * | 1991-06-27 | 1991-08-14 | Sandoz Ag | Improvements in or relating to organic compounds |
SG45449A1 (en) | 1992-05-13 | 1998-01-16 | Sandoz Ltd | Ophthalmic compositions |
CN1326982C (en) * | 1992-06-12 | 2007-07-18 | 诺瓦蒂斯有限公司 | Transesterification products of corn oil and glycerol and its producing process |
ES2168271T3 (en) | 1992-09-25 | 2002-06-16 | Novartis Ag | PHARMACEUTICAL COMPOSITIONS CONTAINING CYCLOSPORINS. |
DE4420733A1 (en) | 1994-06-15 | 1995-12-21 | Sueddeutsche Kalkstickstoff | Process for enzymatic transesterification |
FR2731015B1 (en) * | 1995-02-24 | 1997-05-30 | Sci Sartone | PROCESS FOR THE ENZYMATIC ENRICHMENT OF OILS OF MARINE ORIGIN AND THE TRIGLYCERIDES OF POLYUNSATURATED FATTY ACIDS THUS OBTAINED |
CZ295207B6 (en) | 1997-01-30 | 2005-06-15 | Novartis Ag | Hard gelatin capsule containing cyclosporin A |
EP1510133A1 (en) * | 2003-09-01 | 2005-03-02 | Belovo S.A., Egg Science & Technology | Balanced oil composition |
CA2803477C (en) | 2010-06-25 | 2019-04-30 | Epax As | Process for separating polyunsaturated fatty acids from long chain unsaturated or less saturated fatty acids |
EP3496545A4 (en) * | 2016-08-12 | 2020-04-01 | Cargill, Incorporated | Speciality low saturates canola oil |
KR102444926B1 (en) * | 2022-05-04 | 2022-09-21 | 주식회사 삼광켐 | Vegetable recombinant mimic mink oil and cosmetic composition comprising same |
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GB1577933A (en) * | 1976-02-11 | 1980-10-29 | Unilever Ltd | Fat process and composition |
US4420560A (en) * | 1981-11-17 | 1983-12-13 | Fuji Oil Company, Limited | Method for modification of fats and oils |
JPS63112694A (en) * | 1986-10-30 | 1988-05-17 | 日本油脂株式会社 | Separation and concentration of arachidonic acid and isomyristic acid |
JPH0198494A (en) * | 1987-10-09 | 1989-04-17 | Agency Of Ind Science & Technol | Continuous reaction process with immobilized lipase |
JPH01108990A (en) * | 1987-10-20 | 1989-04-26 | Nippon Oil & Fats Co Ltd | Production of oil and fat |
US4948811A (en) * | 1988-01-26 | 1990-08-14 | The Procter & Gamble Company | Salad/cooking oil balanced for health benefits |
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1990
- 1990-12-06 AU AU67854/90A patent/AU6785490A/en not_active Abandoned
- 1990-12-11 CA CA002031945A patent/CA2031945A1/en not_active Abandoned
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