JPH0225447A - Production of highly unsaturated fatty acids - Google Patents
Production of highly unsaturated fatty acidsInfo
- Publication number
- JPH0225447A JPH0225447A JP63172691A JP17269188A JPH0225447A JP H0225447 A JPH0225447 A JP H0225447A JP 63172691 A JP63172691 A JP 63172691A JP 17269188 A JP17269188 A JP 17269188A JP H0225447 A JPH0225447 A JP H0225447A
- Authority
- JP
- Japan
- Prior art keywords
- acid
- ester
- docosahexaenoic acid
- docosahexaenoic
- fatty acid
- 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.)
- Pending
Links
- 235000021122 unsaturated fatty acids Nutrition 0.000 title claims abstract description 8
- 150000004670 unsaturated fatty acids Chemical class 0.000 title claims abstract description 8
- 238000004519 manufacturing process Methods 0.000 title claims description 4
- MBMBGCFOFBJSGT-KUBAVDMBSA-N all-cis-docosa-4,7,10,13,16,19-hexaenoic acid Chemical compound CC\C=C/C\C=C/C\C=C/C\C=C/C\C=C/C\C=C/CCC(O)=O MBMBGCFOFBJSGT-KUBAVDMBSA-N 0.000 claims abstract description 171
- 235000020669 docosahexaenoic acid Nutrition 0.000 claims abstract description 87
- 229940090949 docosahexaenoic acid Drugs 0.000 claims abstract description 86
- JAZBEHYOTPTENJ-JLNKQSITSA-N all-cis-5,8,11,14,17-icosapentaenoic acid Chemical compound CC\C=C/C\C=C/C\C=C/C\C=C/C\C=C/CCCC(O)=O JAZBEHYOTPTENJ-JLNKQSITSA-N 0.000 claims abstract description 69
- 235000020673 eicosapentaenoic acid Nutrition 0.000 claims abstract description 69
- 229960005135 eicosapentaenoic acid Drugs 0.000 claims abstract description 69
- JAZBEHYOTPTENJ-UHFFFAOYSA-N eicosapentaenoic acid Natural products CCC=CCC=CCC=CCC=CCC=CCCCC(O)=O JAZBEHYOTPTENJ-UHFFFAOYSA-N 0.000 claims abstract description 68
- 229930195729 fatty acid Natural products 0.000 claims abstract description 57
- 239000000194 fatty acid Substances 0.000 claims abstract description 57
- 235000014113 dietary fatty acids Nutrition 0.000 claims abstract description 56
- 150000004665 fatty acids Chemical class 0.000 claims abstract description 37
- 238000000034 method Methods 0.000 claims abstract description 34
- 125000005456 glyceride group Chemical group 0.000 claims abstract description 26
- XSQUKJJJFZCRTK-UHFFFAOYSA-N Urea Chemical compound NC(N)=O XSQUKJJJFZCRTK-UHFFFAOYSA-N 0.000 claims abstract description 25
- 239000004202 carbamide Substances 0.000 claims abstract description 25
- 235000021323 fish oil Nutrition 0.000 claims abstract description 20
- 239000000203 mixture Substances 0.000 claims abstract description 20
- 238000004811 liquid chromatography Methods 0.000 claims abstract description 12
- 239000004367 Lipase Substances 0.000 claims abstract description 11
- 102000004882 Lipase Human genes 0.000 claims abstract description 11
- 108090001060 Lipase Proteins 0.000 claims abstract description 11
- 235000019421 lipase Nutrition 0.000 claims abstract description 11
- CURLTUGMZLYLDI-UHFFFAOYSA-N Carbon dioxide Chemical compound O=C=O CURLTUGMZLYLDI-UHFFFAOYSA-N 0.000 claims abstract description 8
- 241000222120 Candida <Saccharomycetales> Species 0.000 claims abstract description 7
- 125000005907 alkyl ester group Chemical group 0.000 claims abstract description 5
- 239000001569 carbon dioxide Substances 0.000 claims abstract description 4
- 229910002092 carbon dioxide Inorganic materials 0.000 claims abstract description 4
- 238000000605 extraction Methods 0.000 claims abstract 2
- 150000002148 esters Chemical class 0.000 claims description 43
- 238000000199 molecular distillation Methods 0.000 claims description 11
- 241000894006 Bacteria Species 0.000 claims description 4
- 235000020777 polyunsaturated fatty acids Nutrition 0.000 claims 1
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 abstract description 26
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 abstract description 24
- 239000002994 raw material Substances 0.000 abstract description 14
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 abstract description 8
- 238000000354 decomposition reaction Methods 0.000 abstract description 8
- 241000251468 Actinopterygii Species 0.000 abstract description 4
- 230000003301 hydrolyzing effect Effects 0.000 abstract description 4
- 239000003054 catalyst Substances 0.000 abstract description 3
- 239000003814 drug Substances 0.000 abstract description 3
- 235000019688 fish Nutrition 0.000 abstract description 3
- 239000003153 chemical reaction reagent Substances 0.000 abstract description 2
- 235000019512 sardine Nutrition 0.000 abstract description 2
- 239000004615 ingredient Substances 0.000 abstract 4
- 241000624562 Cololabis saira Species 0.000 abstract 1
- 241001125046 Sardina pilchardus Species 0.000 abstract 1
- 235000001497 healthy food Nutrition 0.000 abstract 1
- VLKZOEOYAKHREP-UHFFFAOYSA-N n-Hexane Chemical compound CCCCCC VLKZOEOYAKHREP-UHFFFAOYSA-N 0.000 description 102
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 description 33
- -1 15.9% Chemical compound 0.000 description 24
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 23
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 21
- 238000004817 gas chromatography Methods 0.000 description 16
- KWYUFKZDYYNOTN-UHFFFAOYSA-M Potassium hydroxide Chemical compound [OH-].[K+] KWYUFKZDYYNOTN-UHFFFAOYSA-M 0.000 description 15
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 15
- 239000002904 solvent Substances 0.000 description 10
- 239000007788 liquid Substances 0.000 description 9
- 239000000047 product Substances 0.000 description 9
- 238000004458 analytical method Methods 0.000 description 7
- 125000001495 ethyl group Chemical group [H]C([H])([H])C([H])([H])* 0.000 description 7
- 229920000642 polymer Polymers 0.000 description 7
- 238000005406 washing Methods 0.000 description 7
- 238000003756 stirring Methods 0.000 description 6
- 238000003815 supercritical carbon dioxide extraction Methods 0.000 description 6
- TYLNXKAVUJJPMU-DNKOKRCQSA-N Docosahexaenoic acid ethyl ester Chemical compound CCCCCCCCC\C=C\C=C\C=C\C=C\C=C\C=C\C(=O)OCC TYLNXKAVUJJPMU-DNKOKRCQSA-N 0.000 description 5
- 229920000064 Ethyl eicosapentaenoic acid Polymers 0.000 description 5
- 230000000052 comparative effect Effects 0.000 description 5
- 239000003480 eluent Substances 0.000 description 5
- SSQPWTVBQMWLSZ-AAQCHOMXSA-N ethyl (5Z,8Z,11Z,14Z,17Z)-icosapentaenoate Chemical compound CCOC(=O)CCC\C=C/C\C=C/C\C=C/C\C=C/C\C=C/CC SSQPWTVBQMWLSZ-AAQCHOMXSA-N 0.000 description 5
- 238000001914 filtration Methods 0.000 description 5
- HMZGPNHSPWNGEP-UHFFFAOYSA-N octadecyl 2-methylprop-2-enoate Chemical compound CCCCCCCCCCCCCCCCCCOC(=O)C(C)=C HMZGPNHSPWNGEP-UHFFFAOYSA-N 0.000 description 5
- 238000010992 reflux Methods 0.000 description 5
- 102000004190 Enzymes Human genes 0.000 description 4
- 108090000790 Enzymes Proteins 0.000 description 4
- 230000032050 esterification Effects 0.000 description 4
- 238000005886 esterification reaction Methods 0.000 description 4
- 239000000706 filtrate Substances 0.000 description 4
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 description 3
- 230000005526 G1 to G0 transition Effects 0.000 description 3
- WMFOQBRAJBCJND-UHFFFAOYSA-M Lithium hydroxide Chemical compound [Li+].[OH-] WMFOQBRAJBCJND-UHFFFAOYSA-M 0.000 description 3
- 239000002253 acid Substances 0.000 description 3
- 239000012141 concentrate Substances 0.000 description 3
- 238000009826 distribution Methods 0.000 description 3
- 238000004460 liquid liquid chromatography Methods 0.000 description 3
- 239000000463 material Substances 0.000 description 3
- 238000000746 purification Methods 0.000 description 3
- 239000000243 solution Substances 0.000 description 3
- 239000000126 substance Substances 0.000 description 3
- CHRJZRDFSQHIFI-UHFFFAOYSA-N 1,2-bis(ethenyl)benzene;styrene Chemical compound C=CC1=CC=CC=C1.C=CC1=CC=CC=C1C=C CHRJZRDFSQHIFI-UHFFFAOYSA-N 0.000 description 2
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 2
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- IMNFDUFMRHMDMM-UHFFFAOYSA-N N-Heptane Chemical compound CCCCCCC IMNFDUFMRHMDMM-UHFFFAOYSA-N 0.000 description 2
- WQDUMFSSJAZKTM-UHFFFAOYSA-N Sodium methoxide Chemical compound [Na+].[O-]C WQDUMFSSJAZKTM-UHFFFAOYSA-N 0.000 description 2
- 239000000061 acid fraction Substances 0.000 description 2
- 239000007864 aqueous solution Substances 0.000 description 2
- 238000006243 chemical reaction Methods 0.000 description 2
- HVYWMOMLDIMFJA-DPAQBDIFSA-N cholesterol Chemical compound C1C=C2C[C@@H](O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@H]([C@H](C)CCCC(C)C)[C@@]1(C)CC2 HVYWMOMLDIMFJA-DPAQBDIFSA-N 0.000 description 2
- MAMJNXVNGGBHFN-UHFFFAOYSA-N docosa-1,3,5,7,9,11-hexaene Chemical compound CCCCCCCCCCC=CC=CC=CC=CC=CC=C MAMJNXVNGGBHFN-UHFFFAOYSA-N 0.000 description 2
- 229940079593 drug Drugs 0.000 description 2
- 238000010828 elution Methods 0.000 description 2
- 230000005484 gravity Effects 0.000 description 2
- 235000013402 health food Nutrition 0.000 description 2
- 150000002632 lipids Chemical class 0.000 description 2
- 238000000622 liquid--liquid extraction Methods 0.000 description 2
- 238000012856 packing Methods 0.000 description 2
- 229920006395 saturated elastomer Polymers 0.000 description 2
- 238000000926 separation method Methods 0.000 description 2
- 239000011550 stock solution Substances 0.000 description 2
- OMPIYDSYGYKWSG-UHFFFAOYSA-N 2-(2-ethoxy-2-oxoethyl)-2-hydroxybutanedioic acid Chemical compound CCOC(=O)CC(O)(C(O)=O)CC(O)=O OMPIYDSYGYKWSG-UHFFFAOYSA-N 0.000 description 1
- NIONDZDPPYHYKY-UHFFFAOYSA-N 2-hexenoic acid Chemical compound CCCC=CC(O)=O NIONDZDPPYHYKY-UHFFFAOYSA-N 0.000 description 1
- 206010008132 Cerebral thrombosis Diseases 0.000 description 1
- 241000588881 Chromobacterium Species 0.000 description 1
- 241001062872 Cleyera japonica Species 0.000 description 1
- 241000555825 Clupeidae Species 0.000 description 1
- 241001149724 Cololabis adocetus Species 0.000 description 1
- 241000270281 Coluber constrictor Species 0.000 description 1
- 208000035150 Hypercholesterolemia Diseases 0.000 description 1
- 206010020772 Hypertension Diseases 0.000 description 1
- DGAQECJNVWCQMB-PUAWFVPOSA-M Ilexoside XXIX Chemical compound C[C@@H]1CC[C@@]2(CC[C@@]3(C(=CC[C@H]4[C@]3(CC[C@@H]5[C@@]4(CC[C@@H](C5(C)C)OS(=O)(=O)[O-])C)C)[C@@H]2[C@]1(C)O)C)C(=O)O[C@H]6[C@@H]([C@H]([C@@H]([C@H](O6)CO)O)O)O.[Na+] DGAQECJNVWCQMB-PUAWFVPOSA-M 0.000 description 1
- 201000001429 Intracranial Thrombosis Diseases 0.000 description 1
- NHTMVDHEPJAVLT-UHFFFAOYSA-N Isooctane Chemical compound CC(C)CC(C)(C)C NHTMVDHEPJAVLT-UHFFFAOYSA-N 0.000 description 1
- 241000277331 Salmonidae Species 0.000 description 1
- 241000269851 Sarda sarda Species 0.000 description 1
- 241000269821 Scombridae Species 0.000 description 1
- 150000001218 Thorium Chemical class 0.000 description 1
- 239000013543 active substance Substances 0.000 description 1
- 239000000654 additive Substances 0.000 description 1
- 230000000996 additive effect Effects 0.000 description 1
- 150000001298 alcohols Chemical class 0.000 description 1
- 125000000217 alkyl group Chemical group 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 238000009835 boiling Methods 0.000 description 1
- 210000004556 brain Anatomy 0.000 description 1
- 229920006026 co-polymeric resin Polymers 0.000 description 1
- 238000004440 column chromatography Methods 0.000 description 1
- 238000001816 cooling Methods 0.000 description 1
- JVSWJIKNEAIKJW-UHFFFAOYSA-N dimethyl-hexane Natural products CCCCCC(C)C JVSWJIKNEAIKJW-UHFFFAOYSA-N 0.000 description 1
- 238000004821 distillation Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- AKSGKEXSAPMKPZ-UHFFFAOYSA-N ethane-1,2-diol;methyl 2-methylprop-2-enoate Chemical compound OCCO.COC(=O)C(C)=C AKSGKEXSAPMKPZ-UHFFFAOYSA-N 0.000 description 1
- OQZCSNDVOWYALR-UHFFFAOYSA-N flurochloridone Chemical compound FC(F)(F)C1=CC=CC(N2C(C(Cl)C(CCl)C2)=O)=C1 OQZCSNDVOWYALR-UHFFFAOYSA-N 0.000 description 1
- 238000005194 fractionation Methods 0.000 description 1
- 235000021588 free fatty acids Nutrition 0.000 description 1
- 238000010438 heat treatment Methods 0.000 description 1
- 229930195733 hydrocarbon Natural products 0.000 description 1
- 150000002430 hydrocarbons Chemical class 0.000 description 1
- 230000007062 hydrolysis Effects 0.000 description 1
- 238000006460 hydrolysis reaction Methods 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-M hydroxide Chemical compound [OH-] XLYOFNOQVPJJNP-UHFFFAOYSA-M 0.000 description 1
- 235000020640 mackerel Nutrition 0.000 description 1
- 239000012046 mixed solvent Substances 0.000 description 1
- 208000010125 myocardial infarction Diseases 0.000 description 1
- 230000007935 neutral effect Effects 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- TVMXDCGIABBOFY-UHFFFAOYSA-N octane Chemical compound CCCCCCCC TVMXDCGIABBOFY-UHFFFAOYSA-N 0.000 description 1
- 239000003921 oil Substances 0.000 description 1
- 235000019198 oils Nutrition 0.000 description 1
- 235000014593 oils and fats Nutrition 0.000 description 1
- 229940094443 oxytocics prostaglandins Drugs 0.000 description 1
- 238000002953 preparative HPLC Methods 0.000 description 1
- 150000003180 prostaglandins Chemical class 0.000 description 1
- 239000012264 purified product Substances 0.000 description 1
- 210000001525 retina Anatomy 0.000 description 1
- 229910052708 sodium Inorganic materials 0.000 description 1
- 239000011734 sodium Substances 0.000 description 1
- 238000000638 solvent extraction Methods 0.000 description 1
- 208000010110 spontaneous platelet aggregation Diseases 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 238000005809 transesterification reaction Methods 0.000 description 1
Classifications
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
- Y02P20/00—Technologies relating to chemical industry
- Y02P20/50—Improvements relating to the production of bulk chemicals
- Y02P20/54—Improvements relating to the production of bulk chemicals using solvents, e.g. supercritical solvents or ionic liquids
Abstract
Description
【発明の詳細な説明】
（産業上の利用分野）
本発明は、高純度のエイコサペンタエン酸、ドコサヘキ
サエン酸またはそれらの低級アルキルエステル類の新し
い濃縮分離方法に関する。DETAILED DESCRIPTION OF THE INVENTION (Industrial Application Field) The present invention relates to a new method for concentrating and separating highly pure eicosapentaenoic acid, docosahexaenoic acid, or lower alkyl esters thereof.
（従来の技術）
エイコサペンタエン酸やドコサヘキサエンは、生体内の
重要な生理活性物質である■型のプロスタグランジンな
どへの変換が予想され、各種の生理活性能を発現するこ
とが知られている。特にエイコサペンタエン酸には、血
小板の凝集を阻害し、血中コレステロール値、中性脂質
値を下げる働きがあり、高血圧症、高コレステロール血
症、脳血栓、あるいは心筋梗塞などに効果があることが
知られている。又ドコサヘキサエン酸にもエイコサペン
タエン酸と同様の働きがあることが知られているが、そ
のほかにドコサヘキサエン酸は脳や目の網膜に多（含ま
れており、その機能が注目されている。(Prior art) Eicosapentaenoic acid and docosahexaene are expected to be converted into ■-type prostaglandins, which are important physiologically active substances in living organisms, and are known to exhibit various physiologically active abilities. . In particular, eicosapentaenoic acid has the ability to inhibit platelet aggregation and lower blood cholesterol and neutral lipid levels, and is known to be effective against hypertension, hypercholesterolemia, cerebral thrombosis, and myocardial infarction. It is being It is also known that docosahexaenoic acid has the same function as eicosapentaenoic acid, but docosahexaenoic acid is also abundant in the brain and retina of the eye, and its functions are attracting attention.
このようにエイコサペンタエン酸やドコサヘキサエン酸
は重要な脂質成分であり、その利用も医薬、健康食品な
どとして色々考えられ、高濃度の精製品の開発が期待さ
れている。As described above, eicosapentaenoic acid and docosahexaenoic acid are important lipid components, and their use is considered in various ways as medicines, health foods, etc., and the development of highly concentrated purified products is expected.
エイコサペンタエン酸やドコサヘキサエン酸の生化学的
価値が認められているにもかかわらず、これらの化学合
成は極めて困難であるため、天然の原料から抽出、精製
することが行われている。Despite the recognized biochemical value of eicosapentaenoic acid and docosahexaenoic acid, their chemical synthesis is extremely difficult, so they are extracted and purified from natural raw materials.
例えば尿素付加法、分子蒸留法、超臨界炭酸ガス抽出法
、液体クロマトグラフィー法等が個々に実施されている
。For example, urea addition method, molecular distillation method, supercritical carbon dioxide extraction method, liquid chromatography method, etc. have been individually implemented.
（発明が解決しようとする課題）
魚油はエイコサペンタエン酸およびドコサヘキサエン酸
を多く含む天然油脂であるが、魚油中にはその他の脂肪
酸類も多く含まれているため、エイコサペンタエン酸と
］Ｓコリ°ヘキサエン酸とを高純度で分取するには、か
なりの技術を要する。その理由として、魚油中にはエイ
コサペンタエン酸およびドコサヘキサエン酸の類似物が
含まれており、それらがエイコサペンタエン酸やドコサ
ヘキサエン酸と類似挙動を起こすため、これらの分取が
困難であった。(Problems to be Solved by the Invention) Fish oil is a natural oil containing a large amount of eicosapentaenoic acid and docosahexaenoic acid, but since fish oil also contains many other fatty acids, eicosapentaenoic acid and S coli A considerable amount of technology is required to separate hexaenoic acid with high purity. The reason for this is that fish oil contains analogues of eicosapentaenoic acid and docosahexaenoic acid, which exhibit similar behavior to eicosapentaenoic acid and docosahexaenoic acid, making it difficult to separate them.
また、エイコサペンタエン酸とドコサヘキサエン酸も互
いに似た挙動を示すため、従来からある分取型高速液体
クロマトグラフィーでは、魚油を原料としてエイコサペ
ンタエン酸とドコサヘキサエン酸とを大量に高濃度で濃
縮することは、原料組成が複雑なため、コスト的に問題
があった。Furthermore, since eicosapentaenoic acid and docosahexaenoic acid behave similarly to each other, conventional preparative high-performance liquid chromatography cannot concentrate eicosapentaenoic acid and docosahexaenoic acid in large quantities at high concentrations using fish oil as a raw material. However, due to the complicated raw material composition, there was a problem in terms of cost.
また、従来からの蒸留法などでも、同様に原料組成が複
雑なため、収率の低下が生じていた。In addition, even in conventional distillation methods, the raw material composition is similarly complicated, resulting in a decrease in yield.
本発明は、上記課題を解決するもので、安価で入手しや
すい天然原料である魚油から、簡単な操作によりエイコ
サペンタエン酸またはドコサヘキサエン酸あるいはそれ
らのエステル類を濃縮分離する方法を提供することを目
的としている。The present invention solves the above problems, and aims to provide a method for concentrating and separating eicosapentaenoic acid, docosahexaenoic acid, or their esters by simple operations from fish oil, which is a natural raw material that is inexpensive and easily available. It is said that
（課題を解決するための手段）
本発明は、魚油をギャンディダ（Ｃａｎｄｉｄａ）族菌
由来のリパーゼにより加水分解し、得られた分解混合物
を脂肪酸とグリセリドとに分離し、これらの各成分につ
いて低級アルキルエステル化し、ついで尿素付加法によ
り高度不飽和脂肪酸成分を濃縮し、さらに分子蒸留法、
超臨界炭酸ガス抽出法または液体クロマトグラフィー法
のいずれかの方法を用いて濃縮精製することにより、上
記脂肪酸成分からエイコサペンタエン酸またはそのエス
テルを、上記グリセリド成分からドコサヘキサエン酸ま
たはそのエステルを得ることを特徴とする高度不飽和脂
肪酸類の製造方法である。(Means for Solving the Problems) The present invention hydrolyzes fish oil with lipase derived from Candida family bacteria, separates the resulting decomposition mixture into fatty acids and glycerides, and converts each of these components into lower alkyl After esterification, highly unsaturated fatty acid components are concentrated by urea addition method, and then molecular distillation method,
Eicosapentaenoic acid or its ester is obtained from the fatty acid component and docosahexaenoic acid or its ester is obtained from the glyceride component by concentration and purification using either supercritical carbon dioxide extraction method or liquid chromatography method. This is a unique method for producing highly unsaturated fatty acids.
本発明において原料に用いる魚油は特に制限はないが、
俗に青みの魚といわれているイワシ、サバ、サンマ、カ
ツオ、マグロなどが好ましい。The fish oil used as a raw material in the present invention is not particularly limited, but
Preferable fish are sardines, mackerel, saury, bonito, and tuna, which are commonly referred to as blue-colored fish.
般にこれらの油脂の脂肪酸組成はＣ１，４：０ 、５．
９％、Ｃ１６：０ 、１５．９％、Ｃ１８：１ ； ６
．６％、Ｃ１８：ＯＦ２．５％、Ｃ１，８：Ｉ ； １
３．６％、Ｃ１８：２；１．２％、Ｃ１８：４ ；２．
５％、Ｃ２０：１ ； ８．６％、Ｃ２０：４ 、０．
９％、エイコサペンタエン酸； １２．９％、Ｃ２２：
１ ； ７．８％、Ｃ２２：５ 、１．９％、ドコサヘ
キサエン酸；８．８％、Ｃ２４：１ ； １．０％、そ
の他約１０％であるが、この数値は魚種、季節、産地に
よりいろいろと異なる。Generally, the fatty acid composition of these oils and fats is C1,4:0, 5.
9%, C16:0, 15.9%, C18:1; 6
．． 6%, C18:OF2.5%, C1,8:I; 1
3.6%, C18:2; 1.2%, C18:4;2.
5%, C20:1; 8.6%, C20:4, 0.
9%, eicosapentaenoic acid; 12.9%, C22:
1: 7.8%, C22:5, 1.9%, docosahexaenoic acid: 8.8%, C24:1: 1.0%, others about 10%, but these numbers vary depending on the fish species, season, and production area. It varies depending on the situation.
本発明者らは、このような組成を持つ魚油からエイコサ
ペンタエン酸、またはドコサヘキサエン酸あるいはそれ
らのエステルを効率よく取り出すために、以下のような
手段を見出した。The present inventors have discovered the following means for efficiently extracting eicosapentaenoic acid, docosahexaenoic acid, or their esters from fish oil having such a composition.
ます魚油を、ドコサヘキサエン酸を加水分解しにくい特
性を持つキャンディダ（Ｃａｎｄｉｄａ）族由来リパー
ゼで加水分解すると、ドコサヘキサエン酸を高濃度に含
有するグリセリドと、エイコサペンタエン酸を多く含有
する脂肪酸の混合物を得ることができる。ついでこの分
解混合物を脂肪酸成分とグリセリド成分とに分離する。When trout fish oil is hydrolyzed with lipase derived from the Candida family, which has the property of not easily hydrolyzing docosahexaenoic acid, a mixture of glycerides containing a high concentration of docosahexaenoic acid and fatty acids containing a large amount of eicosapentaenoic acid is obtained. be able to. This decomposition mixture is then separated into fatty acid components and glyceride components.
すなわち、混合物にエタノールとヘキサンと水を加え分
層させ、これに水酸化ナトリウムの水溶液を滴下して、
水層側へエイコサペンタエン酸を高濃度に含む脂肪酸を
すトリウム塩として溶かし出し、後に塩酸等で脂肪酸に
戻す。他方、ヘキサン層から回収したドコサヘキサエン
酸を高濃度に含有するグリセリド画分を、水酸化ナトリ
ウムによりケン化分解し、塩酸で脂肪酸に戻した脂肪酸
混合物を得る。That is, ethanol, hexane, and water are added to the mixture to separate the layers, and an aqueous solution of sodium hydroxide is added dropwise to this.
Fatty acids containing a high concentration of eicosapentaenoic acid are dissolved into the water layer as thorium salts, and later converted back to fatty acids with hydrochloric acid. On the other hand, a glyceride fraction containing a high concentration of docosahexaenoic acid recovered from the hexane layer is saponified and decomposed with sodium hydroxide, and a fatty acid mixture is obtained which is returned to fatty acids with hydrochloric acid.
ついで上記の各々の成分について低級アルコール中で少
量の触媒（塩酸、硫酸等）の存在下でエステル化して脂
肪酸低級アルキルエステルを得る。Next, each of the above components is esterified in a lower alcohol in the presence of a small amount of a catalyst (hydrochloric acid, sulfuric acid, etc.) to obtain a fatty acid lower alkyl ester.
また、上記のグリセリド画分を低級アルコール中で少量
の触媒（ナトリウムメチラート、水酸化ナトリウム、水
酸化カリウム等）の存在下でアルコリシスして同様に脂
肪酸低級アルキルエステルを得る。Furthermore, fatty acid lower alkyl esters are similarly obtained by alcoholysing the above glyceride fraction in a lower alcohol in the presence of a small amount of a catalyst (sodium methylate, sodium hydroxide, potassium hydroxide, etc.).
ついで尿素付加法により、これらの成分から高度不飽和
脂肪酸成分を濃縮する。尿素付加法は尿素と結晶性付加
物を形成させることによって直鎖状炭化水素、脂肪酸、
アルコール類を分離、分別する方法である。一般に油脂
化学の分野では、飽和またはモノエンの脂肪酸が尿素と
結晶性付加物を形成することを利用し、高度不飽和脂肪
酸の濃縮などに用いられる。尿素（−１加物を生成させ
るには、例えば試料に対して２〜４倍量（Ｗ：Ｗ）（７
）尿素を、６〜１０倍量メタノールに加熱上溶解してお
き、これに試料を加え攪拌する。５〜３０℃まで冷却し
て飽和またはモノエンの脂肪酸を結晶化させ、濾過、洗
浄の後、目的とする高度不飽和脂肪酸を分離する。The highly unsaturated fatty acid components are then concentrated from these components by the urea addition method. The urea addition method forms crystalline adducts with urea to produce linear hydrocarbons, fatty acids,
This is a method of separating and classifying alcohols. Generally, in the field of oleochemicals, saturated or monoene fatty acids form crystalline adducts with urea, which is used to concentrate highly unsaturated fatty acids. To generate urea (-1 additive), for example, 2 to 4 times the amount (W:W) (7
) Urea is heated and dissolved in 6 to 10 times the amount of methanol, and the sample is added to this and stirred. The saturated or monoene fatty acid is crystallized by cooling to 5 to 30°C, and after filtration and washing, the target highly unsaturated fatty acid is separated.
本発明においては上記の工程の後、さらに分子蒸留法、
超臨界炭酸ガス抽出法または液体クロマトグラフィー法
のいずれかの方法を用いて濃縮精製工程を実施する。In the present invention, after the above steps, further molecular distillation method,
The concentration and purification step is performed using either a supercritical carbon dioxide extraction method or a liquid chromatography method.
分子蒸留法は、低級アルコールのエステルと成した脂肪
酸エステルを、ｌ×１０″３〜３ Ｘｌ０−２ｍｍ１ｇ
に減圧下、１００〜１５０℃に加熱し、各脂肪酸エステ
ルの沸点の差を利用して分離する方法である。In the molecular distillation method, a fatty acid ester formed with an ester of a lower alcohol is distilled into a 1g
In this method, the fatty acid esters are separated by heating to 100 to 150° C. under reduced pressure and utilizing the difference in boiling point of each fatty acid ester.
この工程を複数回くりかえすと、より高純度の製品が得
られる。By repeating this process multiple times, a product with higher purity can be obtained.
超臨界炭酸ガス抽出法は、炭酸ガスを温度３１．１〜１
００°Ｃ１圧カフ５．２〜２００ ｋｇ／ｃｌで超臨界
状態にし、各脂肪酸エステルの超臨界炭酸ガスへの溶解
度の差により抽出分別する方法である。The supercritical carbon dioxide extraction method uses carbon dioxide at a temperature of 31.1 to 1
This is a method in which fatty acid esters are brought into a supercritical state at a pressure cuff of 5.2 to 200 kg/cl at 00° C. and extracted and fractionated based on the difference in solubility of each fatty acid ester in supercritical carbon dioxide.
液体クロマトグラフィー法は、通常の液体クロマトグラ
フィーあるいはオープンカラムのクロマトグラフィー等
があり、溶離液としてはヘキサン、ヘキサン／アセトン
、ヘプタン、オクタン、イソオクタン等が使用でき、カ
ラム充填材としてはステアリルメタクリレートポリマー
ゲル、スチレンジビニルベンゼンポリマーゲル、メタク
リル酸メチル−エチレングリコールジメククリレートボ
リマーゲル等が使用できる。他にカラム充填材を用いな
い遠心液々クロマトグラフィーがある。Liquid chromatography methods include regular liquid chromatography or open column chromatography, and hexane, hexane/acetone, heptane, octane, isooctane, etc. can be used as the eluent, and stearyl methacrylate polymer gel can be used as the column packing material. , styrene divinylbenzene polymer gel, methyl methacrylate-ethylene glycol dimecrylate polymer gel, etc. can be used. Another method is centrifugal liquid-liquid chromatography, which does not use column packing material.
遠心液々クロマトグラフィーは、液体混合物の原液に比
重の異なる２種の溶剤を作用させて、混合物の中のある
特定の物質を他の物質から分離する方法である。液々抽
出の特徴は原液と溶剤の二層を形成して、この二層を比
重の差により分離することであって、遠心力を用いて短
時間で希望する物質を選択的に溶剤層に移動分離するこ
とができる。魚油の分解物の精製には、ｎ−ヘキサンを
移動相、アセトニトリルまたは９０％エタノールを固定
相とし、移動相で溶出する成分を正溶出画分とし、送液
方向を反転し固定相を送液して固定相内に残留する成分
を反転溶出画分とする。なお、本発明の実施例では、遠
心液々分配クロマトグラフモデルＣＰＣ−ＬＬＮ（三鬼
エンジニアリング（（榊）を用い、分配カートリッジ２
５０Ｗ型を使用して回転数７００〜９００、送液速度１
．０〜３．Ｏｍβで分画した。Centrifugal liquid-liquid chromatography is a method in which two types of solvents with different specific gravities are applied to a stock solution of a liquid mixture to separate a specific substance from other substances in the mixture. The feature of liquid-liquid extraction is that it forms two layers, the stock solution and the solvent, and separates these two layers based on the difference in specific gravity.Centrifugal force is used to selectively transfer the desired substance to the solvent layer in a short period of time. Can be moved and separated. For the purification of fish oil decomposition products, use n-hexane as the mobile phase and acetonitrile or 90% ethanol as the stationary phase.The components eluted with the mobile phase are the positive elution fraction.The direction of liquid feeding is reversed and the stationary phase is fed. The components remaining in the stationary phase are used as the inversion elution fraction. In the embodiment of the present invention, a centrifugal liquid-liquid distribution chromatograph model CPC-LLN (Miki Engineering (Sakaki)) was used, and the distribution cartridge 2
Using 50W type, rotation speed 700-900, liquid feeding speed 1
．． 0-3. It was fractionated with Omβ.
（発明の効果）
本発明のように魚油を原料として、特定のリパーゼでエ
イコサペンタエン酸とドコサヘキサエン酸を選択加水分
解し、分離処理を行うことにより、直接加水分解して得
られた原料と比較して、エイコサペンタエン酸またはド
コサヘキサエン酸の高純度濃縮に有利な中間原料が得ら
れるようになり、また、従来では同一の原料からエイコ
サペンタエン酸とドコサヘキサエン酸を濃縮するには複
雑な分離工程が必要であったが、本発明のように特定の
濃縮工程を組み合わせることにより、高純度エイコサペ
ンタエン酸とドコサヘキサエン酸とを別々に大量に得る
ことが可能となった。本性により安価な魚油より高純度
エイコサペンタエン酸またはドコサヘキサエン酸を大量
に簡単に得られることができるので、試薬、医薬、健康
食品などの素材として、安価に供給することができる。(Effects of the invention) By selectively hydrolyzing eicosapentaenoic acid and docosahexaenoic acid with a specific lipase and performing separation treatment using fish oil as a raw material as in the present invention, comparison with raw materials obtained by direct hydrolysis is achieved. As a result, an intermediate raw material that is advantageous for high-purity concentration of eicosapentaenoic acid or docosahexaenoic acid can be obtained, and in the past, complicated separation processes were required to concentrate eicosapentaenoic acid and docosahexaenoic acid from the same raw material. However, by combining specific concentration steps as in the present invention, it has become possible to separately obtain large amounts of highly purified eicosapentaenoic acid and docosahexaenoic acid. Due to its nature, high-purity eicosapentaenoic acid or docosahexaenoic acid can be easily obtained in large quantities from inexpensive fish oil, so it can be supplied at low cost as a material for reagents, medicines, health foods, etc.
（実施例） 以下、実施例に基づいて本発明を具体的に説明する。(Example) The present invention will be specifically described below based on Examples.
実施例１
魚油５ｋｇと水２．５ｋｇを３０４容量の反応釜に入れ
、５０００ユニツトのキャンディダ・シリンドラセ（Ｃ
ａｎｄｉｄａ ｃｙｌｉｎｄｒａｃｅａ ）由来リパー
ゼを５００ ｍｌ。Example 1 5 kg of fish oil and 2.5 kg of water were placed in a 304 capacity reactor, and a 5000 unit Candida cylinder racer (C
500 ml of lipase derived from andida cylindracea.
の水に溶かしたものを加え、３７℃に保温しながら回転
攪拌し２４時間反応させた。分解物にヘキサン１１１１
を加え分層させ、下層を抜き去った後、ヘキサン層を温
水で洗い酵素を洗い落とした。これにエタノール５．２
βと水２１を加え、さらにＩＮの水酸化ナトリウム水
溶液１１．５ Ａを滴下攪拌後、静置して上層からグリ
セリド画分８８０ｇを得た。下層は塩酸を加えて脂肪酸
を遊離させ、３５５０ ｇの脂肪酸画分を得た。The mixture was dissolved in water and stirred rotatably while keeping the temperature at 37°C, and reacted for 24 hours. Hexane 1111 in the decomposition product
was added to separate the layers, the lower layer was removed, and the hexane layer was washed with warm water to remove the enzyme. Add this to ethanol 5.2
β and water 21 were added, and 11.5 A of an IN aqueous sodium hydroxide solution was added dropwise and stirred, and the mixture was allowed to stand to obtain 880 g of a glyceride fraction from the upper layer. Hydrochloric acid was added to the lower layer to liberate fatty acids, yielding 3550 g of fatty acid fraction.
それぞれの両分中のエイコサペンタエン酸、ドコサヘキ
サエン酸濃度は、ガスクロマトグラフィーによる分析の
結果、グリセリド画分がエイコサヘンクエン酸５．２％
、ドコサヘキサエンＭ４０．７％、脂肪酸画分がエイコ
サペンタエン酸１５．６％、ドコサヘキサエン酸１．３
％であった。The concentration of eicosapentaenoic acid and docosahexaenoic acid in each fraction was analyzed by gas chromatography, and the glyceride fraction was found to be 5.2% eicosahencitric acid.
, docosahexaenoic M40.7%, fatty acid fraction is eicosapentaenoic acid 15.6%, docosahexaenoic acid 1.3
%Met.
グリセリド画分５００ｇに５００ｇのエタノールと５ｇ
の水酸化カリウムを加え、リフランクス下２時間エステ
ル交換し、塩酸で中和し水洗後、脂肪酸エステルを１゜
５１のヘキサンで抽出した。ロータリーエバポレータで
脱溶媒後、４６０ｇの脂肪酸エステルを回収した。その
中から脂肪酸エステル２００ｇ、尿素８００ｇおよびヘ
キサン１６００ｍｐ、メタノール４８ｍ２を攪拌機の付
いたフラスコに仕込み、２時間室温で攪拌しながら反応
させた。付加体を濾別、ヘキサンで洗浄し、濾液と洗浄
液をロータリーエバポレータで減圧上脱溶剤し、１０４
ｇの濃縮エステルを得た。500g of glyceride fraction with 500g of ethanol and 5g
of potassium hydroxide was added thereto, transesterification was carried out under reflux for 2 hours, neutralized with hydrochloric acid, washed with water, and the fatty acid ester was extracted with 1°51 hexane. After removing the solvent using a rotary evaporator, 460 g of fatty acid ester was recovered. From there, 200 g of fatty acid ester, 800 g of urea, 1600 mp of hexane, and 48 m2 of methanol were charged into a flask equipped with a stirrer, and reacted with stirring at room temperature for 2 hours. The adduct was separated by filtration, washed with hexane, and the filtrate and washing liquid were desolventized under reduced pressure using a rotary evaporator.
g of concentrated ester was obtained.
濃縮エステルのエイコサペンタエン酸、ドコサヘキサエ
ン酸濃度はガスクロマトグラフィーによる分析の結果、
エイコサペンタエン酸９．１％、ドコサヘキサエン酸７
２．７％、その他１８．２％であった。The concentration of eicosapentaenoic acid and docosahexaenoic acid in the concentrated ester was determined by gas chromatography analysis.
Eicosapentaenoic acid 9.1%, docosahexaenoic acid 7%
2.7%, others 18.2%.
更にこのうちの５２ｇを分子蒸留（３Ｘ １０−’＋ｕ
＋Ｈｇ、９５℃）にかけ、それを２回繰り返すことによ
り、２９．２ｇの高純度ドコサヘキサエン酸エチルエス
テル（エイコサペンタエン酸１．３％、ドコサヘキサエ
ン酸９７．８％、その他０．９％）を得た。このうちの
Ｌｏｇをとり、メタノール５０艷、ＫＯＨ３ｇ、水０．
５ｇを加え、リフラックス１３時間加水分解し、塩酸で
中和後、５００ ｍｌ、のヘキサンで抽出し、８．５ｇ
の高純度ドコサヘキサエン酸を得た。Furthermore, 52g of this was subjected to molecular distillation (3X 10-'+u
+Hg, 95°C) and repeated twice to obtain 29.2 g of high purity docosahexaenoic acid ethyl ester (1.3% eicosapentaenoic acid, 97.8% docosahexaenoic acid, 0.9% others). . Take the Log of these, methanol 50g, KOH 3g, water 0.
5g was added, refluxed, hydrolyzed for 13 hours, neutralized with hydrochloric acid, extracted with 500ml of hexane, and 8.5g
High purity docosahexaenoic acid was obtained.
また、脂肪酸画分５００ｇに５００ｇのエタノールと硫
酸５ｇを加え、リフランクス下２時間エステル化し、水
洗後指肪酸エステルを１．５１のヘキサンで抽出した。Further, 500 g of ethanol and 5 g of sulfuric acid were added to 500 g of the fatty acid fraction, and esterification was carried out under reflux for 2 hours. After washing with water, the fatty acid ester was extracted with 1.51 hexane.
ロータリーエバポレータで脱溶媒後、４４０ｇの脂肪酸
エステルを回収した。その中から脂肪酸エステル２００
ｇ、尿素８００ｇおよびヘキサン１６００ｍＢ、メタノ
ール４８ｍ１を攪拌機のついたフラスコに仕込み２時間
室温で撹拌しながら反応させた。付加体を濾別、ヘキサ
ンで洗浄し、濾液と洗浄液をロータリーエバポレータで
減圧上脱溶剤し、５０ｇのン農縮エステルを得た。After removing the solvent using a rotary evaporator, 440 g of fatty acid ester was recovered. Among them, 200 fatty acid esters
g, 800 g of urea, 1600 mB of hexane, and 48 ml of methanol were placed in a flask equipped with a stirrer and allowed to react with stirring at room temperature for 2 hours. The adduct was separated by filtration and washed with hexane, and the filtrate and washing liquid were removed from the solvent under reduced pressure using a rotary evaporator to obtain 50 g of ester of nitrogen.
濃縮エステルのエイコサペンタエン酸、ドコサヘキサエ
ン酸濃度はガスクロマトグラフィーによる分析の結果、
エイコサペンタエン酸６０．２％、ドコサヘキサエン酸
５．１％、その他３４．７％であった。The concentration of eicosapentaenoic acid and docosahexaenoic acid in the concentrated ester was determined by gas chromatography analysis.
The content was 60.2% for eicosapentaenoic acid, 5.1% for docosahexaenoic acid, and 34.7% for others.
更にこのうちの２５ｇを分子蒸留（３Ｘ１０−３１１１
ｇ１９５℃）にかけ、それを２回繰り返すことにより、
脂肪酸画分から１２．３ｇの高純度エイコサペンタエン
酸エチルエステル（エイコサペンタエン１９４．７％、
ドコサヘキサエン酸２．６％、その他２．７％）を得た
。このうちの１０ｇをとり、メタノール５０ｍＬＫＯＨ
３ｇ、水０．５ｇを加え、リフラックス１３時間加水分
解し、塩酸で中和後、１００ｍｆのヘキサンで抽出し、
７．９ｇの高純度エイコサペンタエン酸を得た。Furthermore, 25g of this was subjected to molecular distillation (3X10-3111
g195℃) and repeating it twice,
12.3g of high purity eicosapentaenoic acid ethyl ester (eicosapentaene 194.7%,
2.6% of docosahexaenoic acid and 2.7% of others) were obtained. Take 10g of this, methanol 50mL KOH
Add 3g and 0.5g of water, reflux and hydrolyze for 13 hours, neutralize with hydrochloric acid, extract with 100mf hexane,
7.9 g of high purity eicosapentaenoic acid was obtained.
実施例２
実施例１で得た尿素付加後のドコサヘキサエン酸濃縮エ
ステル（エイコサペンタエンｔ１９．１％、ドコサヘキ
サエン酸７２．７％、その他１８．２％）５２ｇを液体
クロマトグラフィー（ステアリルメタクリレートポリマ
ーゲルカラム、溶離液ヘキサン）で分画し、８．３ｇの
高純度ドコサヘキサエン酸エステル（エイコサペンタエ
ン酸０．８％、ドコサヘキサエン酸９８．５％、その他
０．７％）を得た。Example 2 52 g of docosahexaenoic acid concentrated ester (eicosapentaene t 19.1%, docosahexaenoic acid 72.7%, others 18.2%) obtained in Example 1 after addition of urea was subjected to liquid chromatography (stearyl methacrylate polymer gel column, The mixture was fractionated with hexane eluent) to obtain 8.3 g of high purity docosahexaenoic acid ester (0.8% eicosapentaenoic acid, 98.5% docosahexaenoic acid, 0.7% others).
また、実施例１で得た尿素付加後のエイコサペンタエン
酸濃縮エステル（エイコサヘンクエン酸６０６２％、ド
コサヘキサエン酸５．１％、その他３４．７％）２５ｇ
を液体クロマトグラフィー（ステアリルメタクリレート
ポリマーゲルカラム、溶離液ヘキサン）で分画し、５．
６ｇの高純度エイコサペンタエ】 ４
ン酸エチルエステル（エイコサペンタエン酸９７．７％
、ドコサヘキサエン酸１．１％、その他１．２％）を得
た。In addition, 25 g of concentrated eicosapentaenoic acid ester obtained in Example 1 after addition of urea (6062% eicosahencitric acid, 5.1% docosahexaenoic acid, 34.7% others)
5. was fractionated by liquid chromatography (stearyl methacrylate polymer gel column, eluent: hexane).
6g of high-purity eicosapentaenoic acid ethyl ester (97.7% eicosapentaenoic acid)
, 1.1% of docosahexaenoic acid, and 1.2% of others).
実施例３
実施例１と同様に魚油をリパーゼで分解した後、４００
ｇの分解物にヘキサン８８０ ｍｌを加え分層させ、下
層を抜き去った後、ヘキサン層を温水で洗い酵素を洗い
落とした。これにエタノール４２０ ｍＢと水１６０ｍ
ｊ！を加え、更にＩＮの水酸化ナトリウム水溶液９．２
０ ｒｄを滴下攪拌後静置して上層からグリセリド画分
７０ｇを得た。下層は塩酸で中和し、遊離した脂肪酸画
分３０５ｇを得た。Example 3 After decomposing fish oil with lipase in the same manner as in Example 1, 400
880 ml of hexane was added to the decomposition product of g to separate the layers, the lower layer was removed, and the hexane layer was washed with warm water to remove the enzyme. Add to this 420 mB of ethanol and 160 m of water.
j! and then add IN aqueous sodium hydroxide solution 9.2
After dropping 0 rd and stirring, the mixture was allowed to stand, and 70 g of glyceride fraction was obtained from the upper layer. The lower layer was neutralized with hydrochloric acid to obtain 305 g of free fatty acid fraction.
それぞれの両分中のエイコサペンタエン酸、ドコサヘキ
サエン酸濃度はガスクロマトグラフィーによる分析の結
果、グリセリド画分がエイコサペンタエン酸４．８％、
ドコサヘキサエン酸３８．８％、脂肪酸画分がエイコサ
ペンタエン酸１４．７％、ドコサヘキサエン酸２．０％
であった。The concentration of eicosapentaenoic acid and docosahexaenoic acid in both fractions was analyzed by gas chromatography, and the glyceride fraction was found to be 4.8% eicosapentaenoic acid and 4.8% eicosapentaenoic acid.
Docosahexaenoic acid 38.8%, fatty acid fraction is eicosapentaenoic acid 14.7%, docosahexaenoic acid 2.0%
Met.
グリセリド画分５０ｇに５０ｇのエタノールと０．５ｇ
の水酸化カリウムを加え、リフラックス下２時間エステ
ル交換し、塩酸で中和し水洗後、脂肪酸エステルを１５
０ｍβのヘキサンで抽出した。ロータリーエバポレータ
で脱溶媒後、４５ｇの脂肪酸エステルを回収した。その
中から脂肪酸エステル１０ｇ、尿素４０ｇおよびヘキサ
ン８０ｍ１、メタノール２．４ｍｌを攪拌機の付いたフ
ラスコに仕込み、２時間室温で攪拌しながら反応させた
。付加体を濾別、ヘキサンで洗浄し、濾液と洗浄液をロ
ータリーエバポレータで減圧上脱溶剤し、５．１ｇの濃
縮エステルを得た。50g of glyceride fraction, 50g of ethanol and 0.5g
of potassium hydroxide was added, transesterified for 2 hours under reflux, neutralized with hydrochloric acid, washed with water, and the fatty acid ester was
Extracted with 0 mβ hexane. After removing the solvent using a rotary evaporator, 45 g of fatty acid ester was recovered. Among them, 10 g of fatty acid ester, 40 g of urea, 80 ml of hexane, and 2.4 ml of methanol were charged into a flask equipped with a stirrer, and reacted with stirring at room temperature for 2 hours. The adduct was separated by filtration and washed with hexane, and the filtrate and washing liquid were removed from the solvent under reduced pressure using a rotary evaporator to obtain 5.1 g of concentrated ester.
濃縮エステルのエイコサペンタエン酸、ドコサヘキサエ
ン酸濃度はガスクロマトグラフィーによる分析の結果、
エイコサペンタエン酸８．７％、ドコサヘキサエン酸６
９．９％、その他２１．４％であった。The concentration of eicosapentaenoic acid and docosahexaenoic acid in the concentrated ester was determined by gas chromatography analysis.
Eicosapentaenoic acid 8.7%, docosahexaenoic acid 6%
9.9%, others 21.4%.
これを更に遠心液々クロマトグラフィー（分配液ｎ−ヘ
キサン：９０％エタノール−１：１、操作温度２０°Ｃ
１回転数８００ｒｐｍ）で分画し、２．８ｇの高純度ド
コサヘキサエン酸エチルエステル（エイコサペンタエン
酸０．９％、ドコサヘキサエン酸９５．８％、その他３
．３％）を得た。This was further subjected to centrifugal liquid-liquid chromatography (distribution liquid n-hexane:90% ethanol-1:1, operating temperature 20°C).
2.8g of high purity docosahexaenoic acid ethyl ester (eicosapentaenoic acid 0.9%, docosahexaenoic acid 95.8%, and 3 others)
．． 3%).
脂肪酸画分５０ｇに５０ｇのエタノールと０．５ｇの硫
酸を加え、リフランクス下２時間エステル化し、水洗後
、脂肪酸エステルを１５０ｍｆのヘキサンで抽出した。50 g of ethanol and 0.5 g of sulfuric acid were added to 50 g of the fatty acid fraction, esterification was carried out for 2 hours under reflux, and after washing with water, the fatty acid ester was extracted with 150 mf of hexane.
ロータリーエバポレータで脱溶媒後、４０ｇの脂肪酸エ
ステルを回収した。その中から脂肪酸エステル１０ｇ１
尿素４０ｇおよびヘキサン８０ｍ２、メタノール２．４
慴βを攪拌機の付いたフラスコに仕込み、２時間室温で
攪拌しながら反応させた。付加体を濾別、ヘキサンで洗
浄し、濾液と洗浄液をロータリーエバポレータで減圧上
脱溶剤し、２．６ｇの濃縮エステルを得た。After removing the solvent using a rotary evaporator, 40 g of fatty acid ester was recovered. Fatty acid ester 10g1
40g of urea and 80m2 of hexane, 2.4m of methanol
The β-β was charged into a flask equipped with a stirrer, and reacted with stirring at room temperature for 2 hours. The adduct was separated by filtration and washed with hexane, and the filtrate and washing liquid were removed from the solvent under reduced pressure using a rotary evaporator to obtain 2.6 g of concentrated ester.
濃縮エステルのエイコサペンタエン酸、ドコサヘキサエ
ン酸濃度は、ガスクロマトグラフィーによる分析の結果
、エイコサペンタエン酸５８．６％、ドコサヘキサエン
酸４．５％、その他３６．９％であった。これを更に遠
心液々クロマトグラフィー（分配液ｎ−ヘキサン：９０
％エタノール−１＝１、操作温度２０°Ｃ１回転数８０
Ｏｒｐｍ）で分画し、１．７ｇの高純度エイコサペンタ
エン酸エチルエステル（エイコサペンタエン酸９３．３
％、ドコサヘキサエン酸４．９％、その他１．８％）を
得た。As a result of analysis by gas chromatography, the concentration of eicosapentaenoic acid and docosahexaenoic acid in the concentrated ester was 58.6% for eicosapentaenoic acid, 4.5% for docosahexaenoic acid, and 36.9% for others. This was further subjected to centrifugal liquid chromatography (distribution liquid n-hexane: 90%
% ethanol-1 = 1, operating temperature 20°C 1 rotation speed 80
1.7 g of high purity eicosapentaenoic acid ethyl ester (eicosapentaenoic acid 93.3
%, docosahexaenoic acid 4.9%, others 1.8%).
実施例４
魚油５ ｋｇと水２．５ｋｇを３０！容量の反応釜に入
れ、１０００ユニツトの実施例１と同じリバーゼヲ！１
ｉｏｏ ｍｐ。Example 4 5 kg of fish oil and 2.5 kg of water for 30! Put it in a reaction vessel with a capacity of 1,000 units, and use the same reversing method as in Example 1! 1
ioo mp.
の水に溶かしたものを加え、３７℃に保温しながら回転
攪拌し、６時間反応させた。分解物にヘキサン１１βを
加え分層させ、下層を抜き去った後、ヘキサン層を温水
で洗い酵素を洗い落とした。これにエタノール５．２１
と水２１を加え、更にＩＮの水酸化すｌ−ＩＪウム水溶
液７．５１を滴下攪拌後、静置して、上層からグリセリ
ド画分１８００ ｇを得た。The mixture was dissolved in water and stirred while keeping the temperature at 37°C, and reacted for 6 hours. Hexane 11β was added to the decomposition product to separate the layers, and after removing the lower layer, the hexane layer was washed with warm water to remove the enzyme. To this, ethanol 5.21
and 21 g of water were added thereto, and 7.51 g of an aqueous IN sulfur-IJium hydroxide solution was added dropwise. After stirring, the mixture was allowed to stand, and 1800 g of a glyceride fraction was obtained from the upper layer.
下層は塩酸を加えて脂肪酸を遊離させ、２２５０ ｇの
脂肪酸画分を得た。Hydrochloric acid was added to the lower layer to liberate fatty acids, yielding 2250 g of fatty acid fraction.
それぞれの両分中のエイコサペンタエン酸、ドコサヘキ
サエン酸濃度はガスクロマトグラフィーによる分析の結
果、グリセリド画分がエイコサペンタエン酸１６．２％
、ドコサヘキサエン酸２１．５％、脂肪酸画分がエイコ
サペンタエン酸９．８％、ドコサヘキサエン酸１．９％
であった。The concentration of eicosapentaenoic acid and docosahexaenoic acid in each fraction was analyzed by gas chromatography, and the glyceride fraction was found to be 16.2% eicosapentaenoic acid.
, docosahexaenoic acid 21.5%, fatty acid fraction is eicosapentaenoic acid 9.8%, docosahexaenoic acid 1.9%
Met.
このうちグリセリド画分５００ｇを実施例１と同様］
８
の方法でエチルエステル化し、４４０ｇのエステルを得
た。このエステル１０ｈを実施例１と同様の方法で尿素
付加し、濃縮エステルを４３ｇ回収した。濃縮エステル
のエイコサベンクエン酸、ドコサヘキサエン酸濃度はガ
スクロマトグラフィーによる分析の結果、エイコサペン
タエン酸３３．２％、ドコサヘキサエン酸４６．１％、
その他２０．７％であった。更にこれを超臨界炭酸ガス
抽出法（５５℃、１２０ ｋｇ／ｃｆｆｌ）を用いて分
画し、１９．５ ｇの高純度ドコサヘキサエン酸エチル
エステル（エイコサペンタエン酸７．５％、ドコサヘキ
サエン酸８８．２％、その他３．３％）を得た。Of these, 500 g of glyceride fraction was the same as in Example 1]
Ethyl esterification was performed using the method described in Section 8 to obtain 440 g of ester. This ester 10h was added with urea in the same manner as in Example 1, and 43 g of concentrated ester was recovered. The concentrations of eicosaben citric acid and docosahexaenoic acid in the concentrated esters were analyzed by gas chromatography, and were found to be 33.2% eicosapentaenoic acid, 46.1% docosahexaenoic acid,
Others accounted for 20.7%. This was further fractionated using supercritical carbon dioxide extraction method (55°C, 120 kg/cffl) to obtain 19.5 g of high purity docosahexaenoic acid ethyl ester (eicosapentaenoic acid 7.5%, docosahexaenoic acid 88.2%). %, others 3.3%).
また、脂肪酸画分５００ｇを実施例１と同様の方法でエ
チルエステル化し、４３０ｇのエステルを得た。Furthermore, 500 g of the fatty acid fraction was ethyl esterified in the same manner as in Example 1 to obtain 430 g of ester.
このエステル１００ｇを実施例１と同様の方法で尿素付
加し、濃縮エステルを１９．６ｇ回収した。濃縮エステ
ルのエイコサペンタエン酸、ドコサヘキサエン酸濃度は
ガスクロマトグラフィーによる分析の結果、エイコサペ
ンタエン酸４５．９％、ドコサヘキサエン酸８．７％、
その他４５．４％であった。更にこれを超臨界炭酸ガス
抽出法（５５°Ｃ１１２０ｋｇ ／ ｃｎｔ ）を用い
て分画し、９．５ｇの高純度エイフサベンクエン酸エチ
ルエステル（エイコサペンタエン酸８２．３％、ドコサ
ヘキサエン酸３．４％、その他１４．３％）を得た。Urea was added to 100 g of this ester in the same manner as in Example 1, and 19.6 g of concentrated ester was recovered. The concentration of eicosapentaenoic acid and docosahexaenoic acid in the concentrated ester was analyzed by gas chromatography and found that eicosapentaenoic acid was 45.9%, docosahexaenoic acid was 8.7%,
Others accounted for 45.4%. This was further fractionated using a supercritical carbon dioxide extraction method (55°C, 1120 kg/cnt), and 9.5 g of high-purity eifsaben citrate ethyl ester (82.3% eicosapentaenoic acid, 3.4% docosahexaenoic acid) was obtained. %, others 14.3%).
実施例５
実施例４と同様に魚油を分解した後、４００ｇの分解物
にヘキサン８８０ ｍｌを加え分層させ、下層を抜き去
った後、ヘキサン層を温水で洗い酵素を洗い落とした。Example 5 After decomposing fish oil in the same manner as in Example 4, 880 ml of hexane was added to 400 g of the decomposed product to separate the layers, the lower layer was removed, and the hexane layer was washed with warm water to remove the enzyme.
これにエタノール４２０ｍ１と水１６０ｍｆｆを加え、
更にＩＮの水酸化す１〜リウム水溶液９２０ ｍｌを滴
下攪拌後、静置して、上層からグリセリド画分１４０ｇ
を得た。下層は塩酸で中和し、Ｍ離した脂肪酸画分１６
０ｇを得た。それぞれの両分中のエイコサペンタエン酸
、ドコサヘキサエン酸濃度はガスクロマトグラフィーに
よる分析の結果、グリセリド画分がエイコサペンタエン
酸１５．５％、ドコサヘキサエン酸２０．８％、脂肪酸
画分がエイコサペンタエン酸１０．６％、ドコサヘキサ
エン酸１．５％であった。Add 420ml of ethanol and 160mff of water to this,
Furthermore, 920 ml of an aqueous solution of IN sodium to lithium hydroxide was added dropwise, stirred, and left to stand, and 140 g of the glyceride fraction was extracted from the upper layer.
I got it. The lower layer is neutralized with hydrochloric acid and separated from M fatty acid fraction 16.
Obtained 0g. The concentrations of eicosapentaenoic acid and docosahexaenoic acid in both fractions were analyzed by gas chromatography and found that the glyceride fraction was 15.5% eicosapentaenoic acid and 20.8% docosahexaenoic acid, and the fatty acid fraction was 10.8% eicosapentaenoic acid. 6% and docosahexaenoic acid 1.5%.
このうちグリセリド画分５０ｇを実施例３と同様の方法
でエチルエステル化し、４３ｇのエステルを得た。この
エステル１０ｇを実施例３と同様に尿素付加し、４．２
ｇのドコサヘキサエン酸濃縮エステルヲ得た。濃縮エス
テルのエイコサペンタエン酸、ドコサヘキサエン酸濃度
はガスクロマトグラフィーによる分析の結果、エイコサ
ペンタエン酸３１．５％、ドコサヘキサエン酸４４．８
％、その他２３．７％であった。全量の濃縮エステルを
、２００ｍβのスチレンジビニルベンゼン共重合体樹脂
を充填したオープンカラムでヘキサン／アセトン−８７
２の混合溶媒を用いて精製し、０．８ｇの高純度ドコサ
ヘキサエン酸エチルエステル（エイコサペンタエン酸１
．３％、ドコサヘキサエン酸９３．３％、その他５．４
％）を得た。Of these, 50 g of the glyceride fraction was ethyl esterified in the same manner as in Example 3 to obtain 43 g of ester. 10 g of this ester was added with urea in the same manner as in Example 3, and 4.2
g of docosahexaenoic acid concentrated ester was obtained. The concentration of eicosapentaenoic acid and docosahexaenoic acid in the concentrated ester was analyzed by gas chromatography, and the concentration was 31.5% for eicosapentaenoic acid and 44.8% for docosahexaenoic acid.
%, and others 23.7%. The entire concentrated ester was converted into hexane/acetone-87 in an open column packed with 200 mβ styrene divinylbenzene copolymer resin.
0.8g of high purity docosahexaenoic acid ethyl ester (eicosapentaenoic acid 1)
．． 3%, docosahexaenoic acid 93.3%, others 5.4
%) was obtained.
また、脂肪酸画分５０ｇを実施例３と同様の方法でエチ
ルエステル化し、４１ｇのエステルを得た。Furthermore, 50 g of the fatty acid fraction was ethyl esterified in the same manner as in Example 3 to obtain 41 g of ester.
このエステル１０ｇを実施例２と同様の方法で尿素付加
し、２．３ｇのエイコサベンクエン酸濃縮エステルヲ得
た６濃縮エステルのエイコサペンタエン酸、ドコサヘキ
サエン酸濃度はガスクロマトグラフィーによる分析の結
果、エイコサペンタエン酸４３．９％、ドコサヘキサエ
ン酸７．７％、その他４８．４％であった。全量の濃縮
エステルを、１．００ ｍｌのスチレンジビニルベンゼ
ン共重合体樹脂を充填したオープンカラムでヘキサン／
アセトン−８／２の混合溶媒を用いて精製し、０．３ｇ
の高純度エイコサペンタエン酸エチルエステル（エイコ
サペンタエン酸８５．２％、ドコサヘキサエン酸２．５
％、その他１２．３％）を得た。10 g of this ester was added with urea in the same manner as in Example 2 to obtain 2.3 g of eicosaben citric acid concentrated ester. The eicosapentaenoic acid and docosahexaenoic acid concentrations of the 6 concentrated esters were analyzed by gas chromatography. The contents were 43.9% for icosapentaenoic acid, 7.7% for docosahexaenoic acid, and 48.4% for others. The entire amount of the concentrated ester was diluted with hexane/
Purified using acetone-8/2 mixed solvent, 0.3g
High purity eicosapentaenoic acid ethyl ester (eicosapentaenoic acid 85.2%, docosahexaenoic acid 2.5%)
%, others 12.3%).
比較例１
魚油５００ｇをエタノール５００ｇ、 Ｋ ＯＨ５００
ｇで実施例１と同様の方法でエチルエステル化し、４７
０ｇのエステルを得た。そのうちの１００ｇを実施例１
と同様の方法で尿素イ」加し、濃縮エステルを２８．１
ｇ回収した。ｆｉ縮エステルのエイコサペンタエン酸、
ドコサヘキサエン酸濃度はガスクロマトグラフィーによ
る分析の結果、エイコサペンタエン酸４２．９％、ドコ
サヘキサエン酸２５．３％、その他３２．７％であった
。Comparative Example 1 500g of fish oil, 500g of ethanol, KOH500
g was ethyl esterified in the same manner as in Example 1, and 47
Obtained 0 g of ester. 100g of that was used in Example 1
Add urea in the same manner as above to obtain a concentrated ester of 28.1
g was collected. fi-condensed ester of eicosapentaenoic acid,
As a result of analysis by gas chromatography, the concentration of docosahexaenoic acid was 42.9% for eicosapentaenoic acid, 25.3% for docosahexaenoic acid, and 32.7% for others.
これを実施例１と同条件で２回繰り返し分子蒸留し、９
．５ｇのドコサヘキサエン酸両分（エイコサベンクエン
酸２８．４％、ドコサヘキサエン酸４６．４％、その他
２５．２％）を得た。This was subjected to molecular distillation twice under the same conditions as in Example 1, and 9
．． 5 g of docosahexaenoic acid (28.4% eicosaben citric acid, 46.4% docosahexaenoic acid, 25.2% others) was obtained.
また、実施例１と同条件下で２回繰り返し分子蒸留し、
１１．３ｇのエイコサペンタエン酸画分（エイコサベン
クエン酸７４．３％、ドコサヘキサエン酸１８．６％、
その他７．１ ％）を得た。In addition, molecular distillation was repeated twice under the same conditions as in Example 1,
11.3 g of eicosapentaenoic acid fraction (eicosaben citric acid 74.3%, docosahexaenoic acid 18.6%,
7.1%).
キャンディダ族菌由来のリパーゼによる処理をしないの
で、実施例１に比して目的物の純度が低いことがわかる
。It can be seen that the purity of the target product is lower than in Example 1 because the treatment with lipase derived from Candida group bacteria is not performed.
比較例２
角ｊ［ｌｌ ５ ｋｇと水２．５ｋｇを３０β容量の反
応釜に入れ、５０００ユニツトのクロモバクテリウム由
来リパーゼを５００ ｍＲの水に溶かしたものを加え、
３７℃に保温しながら回転撹拌し２４時間反応させた。Comparative Example 2 Put 5 kg of water and 2.5 kg of water into a reaction pot with a capacity of 30β, add 5000 units of Chromobacterium-derived lipase dissolved in 500 mR of water,
While keeping the temperature at 37°C, the mixture was rotated and stirred to react for 24 hours.
分解物を実施例１と同様の方法で脂肪酸とグリセリドに
分画し、グリセリド画分７９０ｇと脂肪酸画分３６８０
ｇを得た。The decomposition product was fractionated into fatty acids and glycerides in the same manner as in Example 1, and 790 g of glyceride fraction and 3680 g of fatty acid fraction were obtained.
I got g.
それぞれの両分中のエイコサベンクエン酸、ドコサヘキ
サエン酸濃度は、ガスクロマトグラフィーによる分析の
結果、グリセリド画分がエイコサペンタエン酸１０．５
％、ドコサヘキサエン酸１．０．３％、脂肪酸画分がエ
イコサベンクエン酸１６．３％、ドコサヘキサエン酸７
．６％であった。The concentration of eicosaben citric acid and docosahexaenoic acid in each fraction was analyzed by gas chromatography, and the glyceride fraction was found to be 10.5% eicosapentaenoic acid.
%, docosahexaenoic acid 1.0.3%, fatty acid fraction is eicosaben citric acid 16.3%, docosahexaenoic acid 7
．． It was 6%.
グリセリド画分７００ｇを実施例１と同様の方法でエチ
ルエステル化し、６６４ｇの脂肪酸エステルを回収した
。その中から脂肪酸エステル４００ｇを取り、実施例１
と同様の方法で尿素付加し、１１７ｇの濃縮エステルを
得た。濃縮エステルのエイコサベンクエン酸、ドコサヘ
キサエン酸濃度はガスクロマトグラフィーによる分析の
結果、エイコサベンクエン酸３０．３％、ドコサヘキサ
エン酸２７．２％、その他４２．５％であった。このう
ち５０ｇを実施例Ｉと同条件下で２回繰り返し分子蒸留
し、１５．２ ｇのドコサヘキサエン酸両分（エイコサ
ベンクエン酸３１．２％、ドコサヘキサエン酸３５．４
％、その他３３．４％）ヲ得た。700 g of the glyceride fraction was ethyl esterified in the same manner as in Example 1, and 664 g of fatty acid ester was recovered. 400g of fatty acid ester was taken from it and Example 1
Urea was added in the same manner as above to obtain 117 g of concentrated ester. As a result of analysis by gas chromatography, the concentrations of eicosaben citric acid and docosahexaenoic acid in the concentrated ester were 30.3% for eicosaben citric acid, 27.2% for docosahexaenoic acid, and 42.5% for others. Of this, 50 g was subjected to repeated molecular distillation twice under the same conditions as in Example I to obtain 15.2 g of both docosahexaenoic acid (eicosaben citric acid 31.2%, docosahexaenoic acid 35.4%).
%, others 33.4%).
また脂肪酸画分１０００ ｇを実施例１と同様の方法で
エチルエステル化し９１６ｇの脂肪酸エステルを回収し
た。その中から脂肪酸エステル４００ｇを取り、実施例
１と同様の方法で尿素付加し１０８ｇの濃縮エステルを
得た。濃縮エステルのエイコサベンクエン酸、ドコサヘ
キサエン酸濃度は、ガスクロマトグラフィーによる分析
の結果、エイコサペンタエン酸３８．３％、ドコサヘキ
サエン酸３２．４％、その他２９．３％であった。この
うち５０ｇを実施例１と同条件下で２回繰り返し分子蒸
留し、１２．３ ｇのエイコサペンタエン酸画分（エイ
コサベンクエン酸３６．８％、ドコサヘキサエン酸３２
．４％、その他３０．８％）を得た。キャンディダ族菌
由来のリパーゼによる処理をしないので、実施例１に比
して目的物の純度が低いことがわかる。Further, 1000 g of the fatty acid fraction was ethyl esterified in the same manner as in Example 1, and 916 g of fatty acid ester was recovered. From there, 400 g of fatty acid ester was taken and added with urea in the same manner as in Example 1 to obtain 108 g of concentrated ester. As a result of analysis by gas chromatography, the concentrations of eicosaben citric acid and docosahexaenoic acid in the concentrated ester were 38.3% for eicosapentaenoic acid, 32.4% for docosahexaenoic acid, and 29.3% for others. Of this, 50 g was subjected to repeated molecular distillation twice under the same conditions as in Example 1, and 12.3 g of eicosapentaenoic acid fraction (36.8% eicosaben citric acid, 32% docosahexaenoic acid) was obtained.
．． 4%, others 30.8%). It can be seen that the purity of the target product is lower than in Example 1 because the treatment with lipase derived from Candida group bacteria is not performed.
比較例３
比較例２で得た尿素付加後のドコサヘキサエン酸濃縮エ
ステル５０ｇを液体クロマトグラフィー（ステアリルメ
タクリレートポリマーゲルカラム、溶離液ヘキサンで分
画し、２．８ｇのドコサヘキサエン酸エチルエステル（
エイコサペンタエン！１２．５％、ドコサヘキサエン酸
６２．３％、その他２５．３％）を得た。Comparative Example 3 50 g of docosahexaenoic acid concentrated ester obtained in Comparative Example 2 after addition of urea was fractionated by liquid chromatography (stearyl methacrylate polymer gel column, eluent hexane) to obtain 2.8 g of docosahexaenoic acid ethyl ester (
Eicosapentaen! 12.5%, docosahexaenoic acid 62.3%, others 25.3%).
また、比較例２で得た尿素付加後のエイコサベンクエン
酸濃縮エステル５０ｇを、液体クロマトグラフィー（ス
テアリルメタクリレートポリマーゲルカラム、溶離液ヘ
キサン）で分画し、１．２ｇのエイコサペンタエン酸エ
チルエステルくエイコサペンタエン１１６５．８％、ド
コサヘキサエン９２０．０％、その他１４．２％）を得
た。In addition, 50 g of eicosabentaenoic acid concentrated ester obtained in Comparative Example 2 after addition of urea was fractionated by liquid chromatography (stearyl methacrylate polymer gel column, eluent: hexane), and 1.2 g of eicosapentaenoic acid ethyl ester was obtained. (1165.8% of eicosapentaene, 920.0% of docosahexaene, and 14.2% of others) were obtained.
Claims (1)
ーゼにより加水分解し、得られた分解混合物を脂肪酸と
グリセリドとに分離し、これらの各成分について低級ア
ルキルエステル化し、ついで尿素付加法により高度不飽
和脂肪酸成分を濃縮し、さらに分子蒸留法、超臨界炭酸
ガス抽出法または液体クロマトグラフィー法のいずれか
の方法を用いて濃縮精製することにより、上記脂肪酸成
分からエイコサペンタエン酸またはそのエステルを、上
記グリセリド成分からドコサヘキサエン酸またはそのエ
ステルを得ることを特徴とする高度不飽和脂肪酸類の製
造方法。Fish oil is hydrolyzed by lipase derived from Candida family bacteria, the resulting decomposed mixture is separated into fatty acids and glycerides, each of these components is converted into lower alkyl esters, and then polyunsaturated fatty acids are obtained by a urea addition method. By concentrating the components and further concentrating and purifying them using any of the methods of molecular distillation, supercritical carbon dioxide gas extraction, or liquid chromatography, eicosapentaenoic acid or its ester is converted from the fatty acid component into the glyceride component. A method for producing highly unsaturated fatty acids, which comprises obtaining docosahexaenoic acid or its ester from.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP63172691A JPH0225447A (en) | 1988-07-13 | 1988-07-13 | Production of highly unsaturated fatty acids |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP63172691A JPH0225447A (en) | 1988-07-13 | 1988-07-13 | Production of highly unsaturated fatty acids |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| JPH0225447A true JPH0225447A (en) | 1990-01-26 |
Family
ID=15946567
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP63172691A Pending JPH0225447A (en) | 1988-07-13 | 1988-07-13 | Production of highly unsaturated fatty acids |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPH0225447A (en) |
Cited By (24)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5013443A (en) * | 1989-01-23 | 1991-05-07 | Nihon Bunko Kogyo Kabushiki Kaisha | Extraction and separation method and apparatus using supercritical fluid |
| WO1991018105A1 (en) * | 1990-05-21 | 1991-11-28 | Martek Corporation | Labeled compounds for use in a diagnostic breath test |
| JPH0595792A (en) * | 1991-10-03 | 1993-04-20 | Agency Of Ind Science & Technol | Production of oil and fat containing concentrated highly unsaturated fatty acid |
| JPH05306979A (en) * | 1992-05-01 | 1993-11-19 | Zensuke Ota | Method for preparing sample of transmission electron microscope |
| JPH0762385A (en) * | 1993-08-27 | 1995-03-07 | Chikyu Kankyo Sangyo Gijutsu Kenkyu Kiko | Extracting method for fatty acid |
| WO1996037587A1 (en) * | 1995-05-24 | 1996-11-28 | Loders Croklaan B.V. | Production of materials high in long chain polyunsaturated fatty acids |
| WO1996037586A1 (en) * | 1995-05-24 | 1996-11-28 | Loders Croklaan B.V. | Production method for fats with long chain polyunsaturated fatty acids |
| EP0784694A1 (en) * | 1995-02-24 | 1997-07-23 | Goemar S.A. | Enzymatic methods for polyunsaturated fatty acid enrichment |
| JPH09510091A (en) * | 1994-03-08 | 1997-10-14 | ノルスク・ヒドロ・アクシェセルスカープ | Essential oil composition |
| JPH107618A (en) * | 1996-06-25 | 1998-01-13 | Tama Seikagaku Kk | Production of eicosapentaenoic acid ester |
| JPH11236590A (en) * | 1990-06-04 | 1999-08-31 | Nippon Suisan Kaisha Ltd | High-concentration eicosapentaenoic acid or its ester |
| WO2002027013A1 (en) * | 2000-09-27 | 2002-04-04 | Ikeda Food Research Co., Ltd. | Process for producing edible sterol fatty acid esters |
| WO2003089399A1 (en) | 2002-04-22 | 2003-10-30 | Industrial Research Limited | Improvements in or relating to separation technology |
| JP2006506483A (en) * | 2002-11-14 | 2006-02-23 | プロノヴァ・バイオケア・アーエス | Lipase catalyzed esterification of marine oil |
| JP2006508307A (en) * | 2002-11-26 | 2006-03-09 | ウーデ・ハイ・プレッシャー・テクノロジーズ・ゲゼルシャフト・ミト・ベシュレンクテル・ハフツング | High pressure device that closes the container in the clean room |
| JP2009504588A (en) * | 2005-08-10 | 2009-02-05 | ブルツツエーゼ，テイベリオ | Composition of n-3 fatty acids having a high concentration of EPA and / or DHA and containing n-6 fatty acids |
| US7619002B2 (en) | 2001-11-12 | 2009-11-17 | Pro Aparts Investimentos E Consultoria LDA. AV. Arriaga | Use of polyunsaturated fatty acids for the primary prevention of major cardiovascular events |
| CN102277004A (en) * | 2011-07-12 | 2011-12-14 | 泉州兴源塑料有限公司 | Strengthening and toughening master batch used for vinyl polymer and method for preparing same |
| JP2012523823A (en) * | 2009-04-17 | 2012-10-11 | セラビスタ ファーマシューティカルズ リミテッド | Composition with low phytanic acid content and rich in omega-3 fatty acids |
| JP2012528901A (en) * | 2009-06-02 | 2012-11-15 | ゴールデン オメガ エス．エー． | Method for obtaining a concentrate of esters of eicosapentaenoic acid and docosahexaenoic acid |
| JP2015063653A (en) * | 2013-08-30 | 2015-04-09 | 備前化成株式会社 | Method for producing high purity omega 3-based fatty acid ethyl ester |
| CN108208685A (en) * | 2017-12-18 | 2018-06-29 | 丁玉琴 | A kind of preparation method of fish oil soft capsule |
| JP2019108340A (en) * | 2009-12-30 | 2019-07-04 | ビーエイエスエフ ファーマ（コーラニッシュ）リミテッド | Simulated moving bed chromatographic separation process |
| US11414622B2 (en) | 2015-08-31 | 2022-08-16 | Nippon Suisan Kaisha, Ltd. | Free polyunsaturated fatty acid-containing composition and manufacturing method therefor |
-
1988
- 1988-07-13 JP JP63172691A patent/JPH0225447A/en active Pending
Cited By (33)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5013443A (en) * | 1989-01-23 | 1991-05-07 | Nihon Bunko Kogyo Kabushiki Kaisha | Extraction and separation method and apparatus using supercritical fluid |
| WO1991018105A1 (en) * | 1990-05-21 | 1991-11-28 | Martek Corporation | Labeled compounds for use in a diagnostic breath test |
| US5164308A (en) * | 1990-05-21 | 1992-11-17 | Martek Corporation | Preparation of labelled triglyceride oils by cultivation of microorganisms |
| US5376540A (en) * | 1990-05-21 | 1994-12-27 | Martek Corporation | Triglyceride oils labelled with 13 C prepared by cultivation of microorganisms |
| US5466434A (en) * | 1990-05-21 | 1995-11-14 | Martek Corporation | Method of diagnosing fatty acid metabolism or absorption disorders using labeled triglyceride oils produced by cultivation of microorganisms |
| JPH11236590A (en) * | 1990-06-04 | 1999-08-31 | Nippon Suisan Kaisha Ltd | High-concentration eicosapentaenoic acid or its ester |
| JPH0595792A (en) * | 1991-10-03 | 1993-04-20 | Agency Of Ind Science & Technol | Production of oil and fat containing concentrated highly unsaturated fatty acid |
| JPH05306979A (en) * | 1992-05-01 | 1993-11-19 | Zensuke Ota | Method for preparing sample of transmission electron microscope |
| JPH0762385A (en) * | 1993-08-27 | 1995-03-07 | Chikyu Kankyo Sangyo Gijutsu Kenkyu Kiko | Extracting method for fatty acid |
| JPH09510091A (en) * | 1994-03-08 | 1997-10-14 | ノルスク・ヒドロ・アクシェセルスカープ | Essential oil composition |
| US5945318A (en) * | 1994-03-08 | 1999-08-31 | Norsk Hydro A.S. | Refining oil compositions |
| EP0784694A1 (en) * | 1995-02-24 | 1997-07-23 | Goemar S.A. | Enzymatic methods for polyunsaturated fatty acid enrichment |
| WO1996037586A1 (en) * | 1995-05-24 | 1996-11-28 | Loders Croklaan B.V. | Production method for fats with long chain polyunsaturated fatty acids |
| WO1996037587A1 (en) * | 1995-05-24 | 1996-11-28 | Loders Croklaan B.V. | Production of materials high in long chain polyunsaturated fatty acids |
| JPH107618A (en) * | 1996-06-25 | 1998-01-13 | Tama Seikagaku Kk | Production of eicosapentaenoic acid ester |
| JP2002171993A (en) * | 2000-09-27 | 2002-06-18 | Ikeda Shokken Kk | Method of manufacturing fatty acid ester of sterol for foods |
| WO2002027013A1 (en) * | 2000-09-27 | 2002-04-04 | Ikeda Food Research Co., Ltd. | Process for producing edible sterol fatty acid esters |
| US7619002B2 (en) | 2001-11-12 | 2009-11-17 | Pro Aparts Investimentos E Consultoria LDA. AV. Arriaga | Use of polyunsaturated fatty acids for the primary prevention of major cardiovascular events |
| US8648061B2 (en) | 2001-11-12 | 2014-02-11 | Pro Aparts Investimentos E. Consultoria LDA. | Use of polyunsaturated fatty acids for the primary prevention of major cardiovascular events |
| US7709668B2 (en) | 2002-04-22 | 2010-05-04 | Industrial Research Limited | Separation technology |
| EP1501782A4 (en) * | 2002-04-22 | 2006-11-08 | Ind Res Ltd | Improvements in or relating to separation technology |
| EP1501782A1 (en) * | 2002-04-22 | 2005-02-02 | Industrial Research Limited | Improvements in or relating to separation technology |
| WO2003089399A1 (en) | 2002-04-22 | 2003-10-30 | Industrial Research Limited | Improvements in or relating to separation technology |
| JP2006506483A (en) * | 2002-11-14 | 2006-02-23 | プロノヴァ・バイオケア・アーエス | Lipase catalyzed esterification of marine oil |
| JP2006508307A (en) * | 2002-11-26 | 2006-03-09 | ウーデ・ハイ・プレッシャー・テクノロジーズ・ゲゼルシャフト・ミト・ベシュレンクテル・ハフツング | High pressure device that closes the container in the clean room |
| JP2009504588A (en) * | 2005-08-10 | 2009-02-05 | ブルツツエーゼ，テイベリオ | Composition of n-3 fatty acids having a high concentration of EPA and / or DHA and containing n-6 fatty acids |
| JP2012523823A (en) * | 2009-04-17 | 2012-10-11 | セラビスタ ファーマシューティカルズ リミテッド | Composition with low phytanic acid content and rich in omega-3 fatty acids |
| JP2012528901A (en) * | 2009-06-02 | 2012-11-15 | ゴールデン オメガ エス．エー． | Method for obtaining a concentrate of esters of eicosapentaenoic acid and docosahexaenoic acid |
| JP2019108340A (en) * | 2009-12-30 | 2019-07-04 | ビーエイエスエフ ファーマ（コーラニッシュ）リミテッド | Simulated moving bed chromatographic separation process |
| CN102277004A (en) * | 2011-07-12 | 2011-12-14 | 泉州兴源塑料有限公司 | Strengthening and toughening master batch used for vinyl polymer and method for preparing same |
| JP2015063653A (en) * | 2013-08-30 | 2015-04-09 | 備前化成株式会社 | Method for producing high purity omega 3-based fatty acid ethyl ester |
| US11414622B2 (en) | 2015-08-31 | 2022-08-16 | Nippon Suisan Kaisha, Ltd. | Free polyunsaturated fatty acid-containing composition and manufacturing method therefor |
| CN108208685A (en) * | 2017-12-18 | 2018-06-29 | 丁玉琴 | A kind of preparation method of fish oil soft capsule |
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