JP2008247876A - Absorption promoter for functional substance containing ether type lipid - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide an absorption promoter containing an ether type lipid having an ether bond represented by an Archaea-derived ether type lipid promoting absorption of functional substances into living body cells. <P>SOLUTION: The absorption promoter promotes the absorption a functional substance into the living body cells and contains an ether type lipid, and the ether type lipid is a specific ether type lipid represented by formula (II) (wherein R<SB>1</SB>and R<SB>2</SB>are saturated or unsaturated hydrocarbons which may contain a branched chain and R<SB>1</SB>and R<SB>2</SB>may be the same or different and Z is a polar group). <P>COPYRIGHT: (C)2009,JPO&INPIT

Description

  The present invention relates to an absorption promoter for a functional substance that can exhibit a certain function in a living body.

  The fat-soluble substance is dispersed in the digestive tract by the action of bile and lipid hydrolyzate, and further forms micelles with complex bile acids such as taurocholic acid, and is absorbed into the small intestinal epithelial cells. At this time, the complex bile acid has a polar part and a non-polar part, and forms micelles in a form in which a fat-soluble substance is dissolved therein together with a lipid such as phospholipid. As the phospholipid, a phospholipid having an ester bond such as phosphatidylcholine (PC) or lysophosphatidylcholine (lysoPC) is used.

  Archaea are a group of organisms that are thought to have separated from eubacteria and eukaryotic cells at the beginning of life, and are known to be able to grow in extremely harsh environments such as the Dead Sea and submarine hydrothermal vents. Yes. Due to adaptation to harsh environments, archaea have unique characteristics that are not recognized by other organisms and are expected to be useful genetic resources. The use of ether type lipids contained in archaea as vaccine compositions (see Patent Documents 1 and 2) and the use of ether type lipids for the treatment of cancer and the like (see Patent Document 3) have been reported, There are few studies on the use of ether type lipids as drug components.

Special Table 2000-502087 Special Table 2003-511421 JP 10-507450 gazette

  An object of the present invention is to provide an absorption promoter containing an ether type lipid having an ether bond typified by an archaeal ether type lipid that promotes absorption of a functional substance into a living cell.

  The present inventors have searched for new functionality of archaeal-containing components, and examined the influence of membrane lipids derived from archaea inhabiting a high heat environment on absorption of fat-soluble substances from the digestive tract. The present inventors extracted lipids from cell membrane fractions of Aeropyrum camini and Aeropyrum pernix K1 strains that are hyperthermophilic archaea inhabiting submarine hydrothermal vents, and lipid components Using the obtained membrane lipids, micelles of cholesterol and fucoxanthin, a carotenoid peculiar to marine organisms, were subjected to gastrointestinal absorption model experiments using Caco-2, and lyso-PC was used as a control. Compared to the case of micelles.

  The main components of the membrane lipids of Aeropyrum camini and Aeropyrum pernix K1 are archeetidyl glucosyl inositol, which is an ether type lipid having an ether bond. It was archietidyl inositol. From the results of gastrointestinal absorption model experiments using Caco-2 cells, micelles using membrane lipids derived from archaebacteria have the effect of further promoting the absorption of cholesterol and fucoxanthin compared to those using lyso-PC. Admitted.

  As described above, the present inventors have found that ether type lipids promote absorption of functional substances such as fat-soluble substances, and have completed the present invention.

[式中、R₁及びR₂は、分岐鎖を有していてもよい、飽和又は不飽和炭化水素であり、R₁及びR₂は同じであっても、異なっていてもよく、Ｚは、極性基である。]
[８] エーテル型脂質が古細菌由来のジエーテル型脂質である[１]〜[７]のいずれかのエーテル型脂質を含む吸収促進剤。
[９] ジエーテル型脂質が古細菌由来のアーキチジルグルコシルイノシトール又はアーキチジルイノシトールである[８]のエーテル型脂質を含む吸収促進剤。
[１０] 機能性物質がビタミン、色素物質、脂質、多不飽和脂肪酸、ホルモン、タンニン酸、アミノ酸、テルペン、ポリフェノール、殺菌剤物質からなる群から選択される脂溶性物質である[１]〜[９]のいずれかのエーテル型脂質を含む吸収促進剤。
[１１] [１]〜[１０]のいずれかのエーテル型脂質を含む吸収促進剤と機能性物質とを含む、前記機能性物質の生体細胞への吸収を促進する組成物。
[１２] 吸収が経粘膜吸収、経腸吸収又は経皮膚吸収である[１１]の機能性物質の生体細胞への吸収を促進する組成物。
[１３] 吸収が経腸吸収である、[１２]の機能性物質の生体細胞への吸収を促進する組成物。
[１４] 組成物がミセル組成物である[１１]〜[１３]のいずれかの機能性物質の生体細胞への吸収を促進する組成物。
[１５] 機能性物質がビタミン、色素物質、脂質、多不飽和脂肪酸、ホルモン、タンニン酸、アミノ酸、テルペン、ポリフェノール、殺菌剤物質からなる群から選択される脂溶性物質である[１１]〜[１４]のいずれかの機能性物質の生体細胞への吸収を促進する組成物。 That is, the present invention is as follows.
[1] An absorption promoter containing an ether type lipid that promotes absorption of a functional substance into living cells.
[2] The ether type lipid according to [1], wherein the ether type lipid has a basic skeleton structure in which glycerol and a hydrocarbon chain are bonded, and at least one of the bonds between glycerol and the hydrocarbon chain is an ether bond. An absorption enhancer.
[3] The ether type lipid has a polar head, and the polar head is composed of a hydroxyl group, a phosphate group, an amino group, a quaternary ammonium ion, a carboxyl group, a sulfate group, a sugar or a derivative thereof, and an amino acid or a derivative thereof. The absorption promoter containing the ether type lipid of [1] or [2], which is an ether type lipid comprising at least one polar part among the selected polar parts.
[4] An absorption promoter comprising the ether type lipid according to any one of [1] to [3], wherein the functional substance is a fat-soluble substance.
[5] An absorption enhancer comprising an ether type lipid according to any one of [1] to [4], wherein absorption is transmucosal absorption, enteral absorption, or transdermal absorption.
[6] An absorption enhancer comprising the ether type lipid according to any one of [1] to [5], wherein absorption is enteral absorption and the ether type lipid forms a micelle in which the functional substance is solubilized.
[7] An absorption enhancer comprising the ether type lipid of any one of [1] to [6], wherein the ether type lipid is represented by the following general formula (II).

[Wherein R ₁ and R ₂ are branched or saturated or unsaturated hydrocarbons, R ₁ and R ₂ may be the same or different, and Z is A polar group. ]
[8] An absorption enhancer comprising the ether type lipid according to any one of [1] to [7], wherein the ether type lipid is an archaeal diether type lipid.
[9] The absorption enhancer comprising the ether type lipid according to [8], wherein the diether type lipid is archidyl glucosyl inositol or archidyl inositol derived from archaea.
[10] The functional substance is a fat-soluble substance selected from the group consisting of vitamins, pigment substances, lipids, polyunsaturated fatty acids, hormones, tannic acid, amino acids, terpenes, polyphenols, and fungicide substances [1] to [ 9] An absorption enhancer comprising the ether type lipid according to any one of
[11] A composition for promoting absorption of a functional substance into living cells, comprising an absorption promoter containing the ether type lipid according to any one of [1] to [10] and a functional substance.
[12] A composition that promotes absorption of a functional substance of [11], wherein absorption is transmucosal absorption, enteral absorption, or transdermal absorption into living cells.
[13] A composition for promoting absorption of a functional substance of [12] into living cells, wherein absorption is enteral absorption.
[14] A composition for promoting absorption of a functional substance according to any one of [11] to [13] into a living cell, wherein the composition is a micelle composition.
[15] The functional substance is a fat-soluble substance selected from the group consisting of vitamins, pigment substances, lipids, polyunsaturated fatty acids, hormones, tannic acid, amino acids, terpenes, polyphenols, fungicide substances [11] to [ 14] The composition which accelerates | stimulates absorption of the functional substance in any one to a biological cell.

  As shown in the Examples, the ether type lipid promotes absorption of a functional substance such as a fat-soluble substance in cells, and can be used as an absorption accelerator for the functional substance from the intestine or the like.

  Hereinafter, the present invention will be described in detail.

  The ether type lipid of the present invention is a lipid having at least one ether bond. The ether type lipid has a basic skeleton structure in which glycerol and at least one hydrocarbon are bonded, and at least one of the bonds between glycerol and the hydrocarbon chain is an ether bond. There are 1, 2 or 3 hydrocarbon chains attached to glycerol. Examples of the ether type lipid of the present invention include a monoether type lipid having one ether bond and a diether type lipid having two ether bonds with respect to a bond between glycerol and a hydrocarbon chain. In addition, there are hetero compounds having both an ether bond and an ester bond, for example, one having an ether bond and one ester bond. Furthermore, as will be described later, caldoarchol is also included.

The ether type lipid of the present invention has the following characteristics.
(1) The bond between the hydrocarbon chain and glycerol contains one or two ether bonds.
(2) The hydrocarbon chain is a saturated or unsaturated hydrocarbon chain which may have a branched chain. For example, an alkyl group, alkynyl group, alkenyl group or isoprenoid which may have a substituent. The number of carbon atoms is C ₁₀ -C _40, preferably C ₂₀ -C _25.

For isoprenoids, a C ₁₀ -C _40, preferably C ₂₀ -C _25, more preferably C ₂₀ or C _25.
The ether type lipid of the present invention includes a core lipid in which a hydrocarbon chain is bound to glycerol, and may further have a polar head, the polar head is composed of a polar group, and a hydroxyl group is used as the polar group. Polar parts selected from ionic groups such as phosphate groups, amino groups, carboxyl groups, sulfate groups, quaternary amines containing quaternary ammonium ions, sugars or derivatives thereof, and amino acids or derivatives thereof, At least one of these polar parts is included. Sugars include monosaccharides, disaccharides or more oligosaccharides, and sugar derivatives. That is, the ether type lipid of the present invention includes a lipid having a polar head, such as phospholipid, glycolipid or phosphoglycolipid.

As described above, the ether type lipid of the present invention includes a diether type lipid in which two molecules of hydrocarbon chains are bonded to glycerol through an ether bond. In addition, two diether type lipids face each other and areoprenoids. Tetraether type lipids having a structure in which hydrocarbons are bonded to each other are also included. In this case, the carbon number of the isoprenoid per molecule is C _{20 to} C ₅₀ .

  The ether type lipid of the present invention has a structure represented by the following general formula (I), for example.

In general formula (I), A and B represent an ether bond (—O—) or an ester bond (—O—CO—), at least one of A and B is an ether bond, and the other is an ether bond. Or an ester bond may be sufficient. R _a and R _b are saturated or unsaturated hydrocarbon chains which may have a branched chain. For example, an alkyl group, alkynyl group, alkenyl group or isoprenoid which may have a substituent. The number of carbon atoms is C ₁₀ -C _40, preferably C ₂₀ -C _25. R _a and R _b may be the same or different. The number of carbon atoms is a C ₁₀ -C _40. For example, an isoprenoid, isoprenoids are C ₁₀ -C _40, preferably C ₂₀ -C _25, more preferably from C ₂₀ or C _25.

  Z is a polar head, and the polar head is composed of a polar group, and includes a ionic group such as a hydroxyl group, a phosphate group, an amino group, a carboxyl group and a sulfate group as a polar group, and a quaternary ammonium ion Examples include polar parts selected from secondary amines, sugars or derivatives thereof, and amino acids or derivatives thereof, including at least one of these polar parts. Sugars include monosaccharides, disaccharides or more oligosaccharides, and sugar derivatives. For example, the polar head includes a group bonded via a phosphate ester or a sugar chain bonded via a glycosidic bond. That is, for example, as Z, sugar, phosphoric acid, or phosphoric acid bonded with sugar, serine, ethanolamine or other groups can be mentioned.

  An example of the ether type lipid is one in which both A and B are ether bonds, for example, one having a diether type structure represented by the following general formula (II).

In the general formula (II), R ₁ and R ₂ are an unsaturated hydrocarbon chain such as a saturated hydrocarbon or a polyene hydrocarbon, which may have a branched chain. For example, an alkyl group, alkynyl group, alkenyl group or isoprenoid which may have a substituent. The number of carbon atoms is C ₁₀ -C _40, preferably C ₂₀ -C _25. R ₁ and R ₂ may be the same or different. Isoprenoids, isoprene units of 5 carbon atoms _{(-CH 2 -CH 2 = (CH} = CH 3) -CH 2) a is a compound having a basic skeleton and 10 have the isoprene units from 4. The isoprenoid is represented by the following formula (III).

For example, if both R ₁ and R ₂ are isoprenoids, both R ₁ and R ₂ are C ₂₀ isoprenoids, both R ₁ and R ₂ are C ₂₅ isoprenoids, and _one of R ₁ and R ₂ is C ₂₀ isoprenoids, the other being C ₂₅ isoprenoids. R ₁ and R ₂ may be bonded at the ends and have a cyclic structure.

  Z is a polar head, and the polar head is composed of a polar group, and includes a ionic group such as a hydroxyl group, a phosphate group, an amino group, a carboxyl group, and a sulfate group as a polar group, and a quaternary ammonium ion. Examples include polar parts selected from secondary amines, sugars or derivatives thereof, and amino acids or derivatives thereof, including at least one of these polar parts. Sugars include monosaccharides, disaccharides or more oligosaccharides, and sugar derivatives. For example, the polar head includes a group bonded through a phosphate ester or a sugar chain bonded through a glycosidic bond. That is, for example, Z includes sugar, phosphoric acid, or phosphoric acid in which sugar, serine, ethanolamine or other groups are bonded.

  Examples of the sugar include those in which 1 to 4 molecules of sugar selected from the group consisting of glucose, galactose, mannose, and fructose are bonded, such as gentiobiose (D-Glc- (β1-6) -D-Glc). Examples of the group bonded via a phosphate ester include hydrogen, glucosyl myo-inositol, myo-inositol, ethanolamine, serine, N-acetylglucosamine (N- acetylglucosamine), glucosaminyl myo-inositol, dimethylaminopentanetetrol, trimethylaminopentanetetrol, glycerol, glycerol phosphate, glycerol sulfate, glycerol sulfate ), Glycerol-1,2-cyclic phosphate, glycerol phosphate monomethylester, and the like. Z is preferably phosphoric acid, or a combination of phosphoric acid and sugar. In this case, the ether type lipid of the present invention is an ether type phospholipid having phosphoric acid in the polar head.

  When Z is a hydroxyl group, the compound represented by the general formula (I) is 2,3-di-o-alkyl sn-glycerol diether.

  In the ether type lipid represented by the general formula (II), the hydrocarbon chain is bonded to the sn-2 and sn-3 positions with respect to glycerol. When phosphoric acid is included, it has a structure of sn-glycerol-1-phosphate type.

  The ether type lipid of the present invention further has a tetraether type structure represented by, for example, the following general formula (IV) or (V).

  When Z is a hydroxyl group, the compound represented by the general formula (I) is 2,3-di-o-alkyl sn-glycerol diether.

R ₁ , R ₂ and Z are as described above, Y is the same as Z, and Y and Z may be the same or different.

The present invention also includes derivatives of the above-mentioned diether type lipids. Examples of the derivatives include those in which H of R ₁ and / or R ₂ isoprenoid is substituted with OH.

  Examples of the ether type lipid of the present invention include those derived from archaea. An archaea is a prokaryote that does not have a nucleus, and is a microorganism having a molecular biological characteristic that is 0.5 to 1.5 μm in diameter and 1 to several μm in length, which is different from a true bacterium. Archaea is characterized by the base sequence of small ribosomal subunit RNA. Archaebacteria include aerobic, anaerobic, thermophilic, hyperthermophilic, thermoacidophilic and halophilic microorganisms. The archaebacteria are described in detail in, for example, Yosuke Koga et al., Archaeological Biology, University of Tokyo Press, August 24, 1998.

  The nomenclature has been proposed for archaeal ether-type lipids (Nishihara, M. et al., J. Biochem., 101: 1007-1015; Yoga Koga et al., Archaeological Biology, University of Tokyo publication) Society, August 24, 1998) According to the nomenclature of the ether type lipid of the present invention, the core lipid is archaeol, which is a diether type lipid represented by the above general formula (II), Caldarchaeol, which is a tetraether type lipid represented by the general formulas (IV) and (V), is included. What is represented by the above general formula (IV) is called isocardioalkyl. Furthermore, these include glycolipids with sugar chains, phospholipids with only phosphate groups, and phospholipids (phosphoglycolipids) with both sugar chains and phosphate groups. For example, those with a sugar chain are called with the name of the sugar chain in front of archiol or cardiochiol. For example, there are glycosyl archiol, glycosyl cardiochiol and the like. In addition, archiol and cardiochiol monophosphate having only one phosphate group are called archidicic acid and cardiochidic acid. Further, those having a phosphate group and a sugar chain, for example, when the phosphate group is a core lipid and further a sugar chain is attached to the phosphate group, that is, archidic acid, cardiochidic acid, serine, ethanolamine, When sugars such as inositol and glycerol are bonded via a phosphodiester bond, they are called as arcidylserine, cardoachitidylserine, architidyl-myo-inositol. Furthermore, when one of the two glycerol parts of cardiochiol has a phosphate group and a sugar (eg, serine), and the other glycerol part has a sugar, like glucosylcardiochitidylserine, be called.

  The ether type lipids of the present invention include, for example, Aeropyrum pernix, Aeropyrum camini, Halobacterium cutirubrum, Methanococcus mazei, Methanococcus mazei, Methanospirillum hungatei), Methanococcus jannaschii, Methanosphaera stadtmanae, Methanobrevibacter smithii, Methanococrum volum (Methanococrum volum) Those derived from archaea such as Methanobacterium espanolae can be used.

  For example, from aeropyrum pernix and aeropyrum camini, you can obtain archaetidyl glucosyl inositol and archaetidyl inositol. It can be suitably used as the ether type lipid of the invention.

  The chemical formulas of archietidyl glucosyl inositol and archietidyl inositol are shown in FIG.

  The ether type lipid of the present invention can be extracted and purified from archaea, and can also be produced by chemical synthesis. In the case of extraction / purification from archaea, it can be obtained, for example, by extracting total lipid from various cells of archaea and then recovering the total polar lipid (TPL) as an acetone-insoluble fraction. Furthermore, it can refine | purify using the thin layer chromatography for isolation | separation, For example, the purity can be confirmed by negative ion fast atom bombardment mass spectrometry. In the case of producing by chemical synthesis, for example, the same lipid as the ether type lipid possessed by the archaebacteria described above or a structurally similar ether type lipid can be synthesized and used as the ether type lipid of the present invention.

  The ether type lipid of the present invention can promote absorption of a functional substance into living cells. Here, the functional substance means a substance capable of exhibiting functions as a nutritional component, a pharmaceutical component, and a cosmetic component in a living body. Examples of functional substances include organic and inorganic substances such as vitamins, medicines, and ions. Moreover, a hydrophobic substance, a fat-soluble substance, a lipophilic substance, etc. are mentioned. Absorption occurs through mucous membranes and skin, and examples of mucous membranes include intestinal mucosa such as small intestine and large intestine, and nasal mucosa. The ether type lipid of the present invention is particularly effective for promoting absorption in the intestine. These functional substances form a complex with the ether type of the present invention and are introduced into living cells. At this time, the ether type lipid of the present invention does not enter the living cell and only the functional substance may be introduced into the living cell, or the ether type lipid and the functional substance may be introduced into the living cell together. There is also. That is, regardless of whether or not the ether type lipid itself is introduced into the living body, the ether type lipid of the present invention has a function of promoting absorption of the functional substance into living cells.

  The ether type lipid of the present invention can function alone as an absorbent for a functional substance into living cells, and may be administered to a living body separately from the functional substance, for example. Moreover, you may administer by mixing with a functional substance. Accordingly, the present invention includes a functional substance absorption promoter for living cells containing the ether type lipid of the present invention as an active ingredient, and further comprises the ether type lipid of the present invention and a functional substance to be absorbed by living cells, A mixture of Although the administration method is not limited, it can be administered orally, for example, and can also be administered by application to mucous membranes or skin, spraying, spraying, or the like. When the ether type lipid is used alone for administration, 0.1 mg to 1000 mg may be administered per 1 kg body weight of the animal.

  For example, when the functional substance is a fat-soluble substance, the target fat-soluble substance is not limited, and any fat-soluble substance having a physiological function for animals is targeted. Examples of the physiological function include a function as a nutritional component, a function as a cosmetic, and a function as a medicine. Preferably, the subject fat-soluble substances are vitamins such as fat-soluble vitamins A, D, E and K; chlorophyll, xanthophyll, carotenoid (β-carotene, astaxanthin, apocarotenal, canthaxanthin, apoester, citranaxanthin, zeaxanthin Pigments such as cholesterol; lipids such as cholesterol; polyunsaturated fatty acids such as arachidonic acid, docosahexaenoic acid (DHA) or eicosapentaenoic acid (EPA); hormones; tannic acid; amino acids; diterpenes, triterpenes, sesquiterpenes Terpene such as: naphthol dianthracene-hypericin; polyphenol (saponin); bactericidal substance (allysine); coenzyme Q; cosmetic agent and the like.

  For example, the ether type lipid of the present invention forms a micelle containing the above fat-soluble substance, and promotes transmucosal, enteral or transdermal absorption of the fat-soluble substance. Examples of micelles include spherical micelles, cylindrical micelles, flexible bilayers, vesicles, planar bilayers, reverse micelles, and the like, and liposomes may be formed. For example, polar lipids form a bimolecular layered micelle with the polar group on the outside, or the micelle closes to a spherical vesicle to form a liposome. At this time, the fat-soluble substance is dissolved in the micelle and solubilized. Micelle formation is in addition to the ether type lipids and fat-soluble substances of the present invention, bile acids or conjugates thereof (eg conjugates with taurine or glycine), bile salts or semi-synthetic or synthetic analogues, preferably , Taurocholic acid, ursodeoxycholic acid, sodium deoxycholate, tauro-deoxycholate, glyco-deoxycholate, sodium ursodeoxycholic acid, glyco-ursodeoxycholate or tauro-ursodeoxycholate, etc. Formed in the presence of Furthermore, fatty acids such as oleic acid, monoacylglycerols such as monooleoylglycerol, phospholipids, phosphatidylcholines and the like may be included. In particular, lysophosphatidylcholine (J. Lipid Res. 39, 1197, 1998) may be included. For example, the ether type lipid of the present invention, a fat-soluble substance, sodium taurocholate, 1-monoolein and oleic acid can be mixed to obtain a micelle in which the fat-soluble substance is solubilized.

  Therefore, the micelles may be administered after the micelles are formed in a form in which the fat-soluble substance to be absorbed and administered in advance is included in the micelles containing the ether type lipid of the present invention as a constituent component. Moreover, you may administer a fat-soluble substance, the ether type lipid of this invention, and another micelle formation component separately. That is, the ether type lipid of the present invention can be used as a fat-soluble substance absorption promoting composition in the form of a composition mixed with a fat-soluble substance, or can be used alone as a fat-soluble substance absorption promoter. Further, it may be used as an absorption accelerator for fat-soluble substances in combination with lysophosphatidylcholine (J. Lipid Res. 39, 1197, 1998).

  Also, in the small intestine, the micelle-forming components present in the intestine at the time of absorption associate to form micelles, so the fat-soluble substance to be absorbed and the ether type lipid of the present invention are mixed, or They may be administered separately.

  For example, the ether type lipid of the present invention and a fat-soluble substance and other micelle forming components are mixed or dissolved in an organic solvent, for example, an organic solvent such as methylene chloride, ethanol and / or methanol, and then the solvent is removed with a rotary evaporator or the like. And the residue can be dissolved in water or an aqueous solution of a pharmaceutical additive such as a buffer or isotonic agent to form micelles.

  The mixing ratio of each component in the case of forming a micelle is not limited, but includes, for example, 5 to 30% (weight%) of ether type lipid. For example, a mixture of ether type lipid (final concentration 200μM), fat-soluble substance (final concentration 100μM), sodium taurocholate (final concentration 5mM), 1-monoolein (final concentration 500μM), oleic acid (final concentration 500μM) Can be mentioned. In addition, when the ether type lipid is used for administration alone, 0.1 mg to 1000 mg may be administered per 1 kg body weight of the animal.

  The ether type lipid of the present invention can be used as an additive for promoting absorption of components of enteral nutrients and pharmaceuticals.

  The present invention will be specifically described by the following examples, but the present invention is not limited to these examples.

Cholesterol absorption experiment Membrane lipids were extracted from the cell membrane lipid fraction of Aeropyrum pernix or Aeropyrum camini with chloroform / methanol (2: 1, v / v) by the Blight-Dyer method. . Two-dimensional development was performed on HPTLC plates to analyze lipids. TLC developed in two dimensions, and the developing solution used for the first vertical development was chloroform / methanol / 7M ammonia water (60: 35: 8), which was used for the second horizontal development. The developing solution was chloroform / methanol / acetic acid / water (80: 25: 10: 5). TLC used 25HPTLC Plates 10 × 10 Silicagel 60 (MERCK). A developed TLC is shown in FIG. FIG. 1A is a TLC development view of lipid derived from Aeropyram pernicus, and FIG. 1B is a TLC development view of lipid derived from Aeropyram kamini. Both mainly contained architidyl glucosyl inositol (AGI) and architidyl inositol (AI). Lipids were identified with reference to Biochim Biophys. Acta 1436, 426-436, 1999. 2A and 2B show the structural formulas of architidyl glucosyl inositol (AGI) and architidyl inositol (AI), respectively. Hereinafter, the cell membrane lipid fraction was used as archaeal membrane lipid. The archaeal membrane lipid was used in the experiment after the amount of phosphate was quantified by the Bartlett method (J. Biol. Chem., 234, 466, 1959).

Cholesterol was micellized for addition to the cells. Cholesterol (final concentration 100 μM), sodium taurocholate (final concentration 5 mM), 1-monoolein (final concentration 500 μM), oleic acid (final concentration 500 μM) It was added to become. Further, lysophosphatidylcholine (J. Lipid Res. 39, 1197, 1998) or archaeal membrane lipid, which is known to have an effect of promoting absorption of cholesterol, was added to a final concentration of 200 μM, and the solvent was removed with N ₂ gas. Dulbecco's modified Eagle medium (DMEM) containing 0.7% bovine serum albumin was added to the dried product, and the mixture was stirred with a Vortex mixer to make micelles. The micelle solution was sterilized through a 0.22 μm filter and then added to the cells.

Caco-2 cells (RIKEN GENEBANK) are 10cm dish (NUNC) using 10% FCS, 0.1mmol / L non-essential amino acids, 1MEM units / mL penicillin and 1OOμg / mL streptomycin in DMEM, 5% CO ₂ at 37 ° C. The cultures were cultured under the above conditions, and when they reached 70% confluence, they were passaged and maintained by trypsin treatment. Caco-2 cells were seeded on a 12-well plate at a concentration of 1.2 × 10 ⁵ cells / mL to 1 mL / well and cultured in an environment of 5% CO ₂ and 37 ° C. for 21 days. The medium was changed every 2-3 days. After culturing, the cells were washed twice with 1 mL of serum-free medium per 1we11, and 1 mL of serum-free medium containing micellized cholesterol was added to each we11. After incubation for 24 hours, the cells were washed with a PBS solution containing 10 mmol / L sodium taurocholate to remove cholesterol or carotenoid adhering to the cell surface, and further washed twice with PBS. The cells were collected with 1 mL of PBS and centrifuged at 1000 × g and 4 ° C. for 5 minutes to remove the supernatant. To the remaining cell pellet, 0.5 mL of PBS was added and ultrasonically homogenized to disrupt the cells. The amount of protein was quantified by Lowry method (J. Biol. Chem., 193, 265, 1951) using a part of the homogenate. Chloroform and methanol were added to the remaining homogenate to extract lipids, and the amount of cholesterol was quantified using cholesterol E-Test Wako (Wako Pure Chemical Industries). The obtained value was corrected with the amount of protein, and the amount of cholesterol taken up into the cells was calculated by subtracting the amount of cholesterol in untreated cells.

  FIGS. 3 and 4 show the calculated amounts of cholesterol when using membrane lipid fractions derived from Aeropyrum pernix and Aeropyrum camini, respectively. The vertical axis is nmol / mg protein. As shown in FIGS. 3 and 4, cholesterol absorption was promoted when the archaeal membrane lipid fraction was mixed with micelles.

Absorption experiment of fucoxanthin Fucoxanthin (final concentration 5 μM) was micellized according to the cholesterol method. A membrane lipid fraction derived from Aeropyrum camini was used. After incorporation into the cells in the same manner as above, fucoxanthin was extracted from the collected cells using a dichloromethane / methanol solution (2: 1, v / v). The contents of fucoxanthin incorporated into cells and fucoxanthinol generated from fucoxanthin were quantified using HPLC (J. Nutr. 132, 946, 2002).

The HPLC analysis conditions were as follows.
・ Flow rate 1.OmL / min
・ Column temperature 40 ℃
・ Mobile phase Acetonitrile: Methanol: Water = 75: 15: 10 (v / v / v, 0.1% ammonium acetate included)
・ Detection 444nm
The results are shown in FIG. The vertical axis is nmol / mg protein. The result of FIG. 5 shows the total amount of fucoxanthin and fucoxanthinol. As shown in FIG. 5, when the archaeal membrane lipid fraction was mixed with micelles, absorption of fucoxanthin was promoted.

It is a figure which shows the result of the TLC expansion | deployment of the cell membrane lipid fraction of Aeropyrum pernix (FIG. 1A) or Aeropyrum camini (FIG. 1B). It is a figure which shows structural formula of an architidyl glucosyl inositol (AGI) (FIG. 2A) and an architidyl inositol (AI) (FIG. 2B). It is a figure which shows absorption of cholesterol in the presence of archaeal membrane lipid derived from Aeropyram pernicus in Caco-2 cells. It is a figure which shows absorption of the cholesterol in the presence of the archaeal membrane lipid derived from Aeropyram kamini in Caco-2 cell. It is a figure which shows absorption of the fucoxanthin in the presence of the archaeal membrane lipid derived from Aeropyram kamini in Caco-2 cell.

Claims (15)

  An absorption enhancer containing an ether type lipid that promotes absorption of functional substances into living cells.   The ether type lipid according to claim 1, wherein the ether type lipid is an ether type lipid having a basic skeleton structure in which glycerol and a hydrocarbon chain are bonded, and at least one of the bonds between glycerol and the hydrocarbon chain is an ether bond. Absorption enhancer.   The ether type lipid has a polar head, and the polar head is selected from the group consisting of hydroxyl group, phosphate group, amino group, quaternary ammonium ion, carboxyl group, sulfate group, sugar or derivative thereof, and amino acid or derivative thereof. The absorption promoter containing an ether type lipid according to claim 1 or 2, which is an ether type lipid comprising at least one polar part among selected polar parts.   The absorption promoter containing an ether type lipid according to any one of claims 1 to 3, wherein the functional substance is a fat-soluble substance.   The absorption promoter containing an ether type lipid according to any one of claims 1 to 4, wherein the absorption is transmucosal absorption, enteral absorption, or transdermal absorption.   The absorption promoter containing an ether type lipid according to any one of claims 1 to 5, wherein absorption is enteral absorption, and the ether type lipid forms a micelle in which a functional substance is solubilized. The absorption promoter containing an ether type lipid according to any one of claims 1 to 6, wherein the ether type lipid is represented by the following general formula (II).

[Wherein R ₁ and R ₂ are branched or saturated or unsaturated hydrocarbons, R ₁ and R ₂ may be the same or different, and Z is A polar group. ]   The absorption promoter containing an ether type lipid according to any one of claims 1 to 7, wherein the ether type lipid is an archaeal diether type lipid.   The absorption enhancer containing an ether type lipid according to claim 8, wherein the diether type lipid is archidyl glucosyl inositol or archidyl inositol derived from archaea.   The functional substance is a fat-soluble substance selected from the group consisting of vitamins, pigment substances, lipids, polyunsaturated fatty acids, hormones, tannic acid, amino acids, terpenes, polyphenols, and bactericidal substances. An absorption enhancer comprising the ether type lipid according to item 1.   The composition which accelerates | stimulates the absorption to the biological cell of the said functional substance containing the absorption promoter containing the ether type lipid of any one of Claims 1-10, and a functional substance.   The composition for promoting absorption of a functional substance into living cells according to claim 11, wherein the absorption is transmucosal absorption, enteral absorption, or transdermal absorption.   The composition which accelerates | stimulates absorption to the biological cell of the functional substance of Claim 12 whose absorption is enteral absorption.   The composition which accelerates | stimulates the absorption to the biological cell of the functional substance of any one of Claims 11-13 which is a micelle composition.   The functional substance is a fat-soluble substance selected from the group consisting of vitamins, pigment substances, lipids, polyunsaturated fatty acids, hormones, tannic acid, amino acids, terpenes, polyphenols, and bactericidal substances. A composition that promotes absorption of the functional substance according to item 1 into living cells.

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