Title of Invention	METHOD OF CRYSTALLIZING REDUCED COENZYME Q10 FROM AQUEOUS SOLUTION.
Abstract	The present invention provides the excellent crystallization method suitable for production on the industrial scale for obtaining the reduced coenzyme Q10 crystal. The high-quality reduced coenzyme Q10 can be obtained in improving the slurry property and/or the crystallization yield by the method of present invention in which reduced coenzyme Q10 is crystallized in an aqueous solution, in particular, the method of crystallizing the reduced coenzyme Q10 by substituting an organic solvent solution containing the reduced coenzyme Q10 by water, the method of crystallizing the oily reduced coenzyme Q10 in an aqueous solution, or the like.

Title of Invention

METHOD OF CRYSTALLIZING REDUCED COENZYME Q10 FROM AQUEOUS SOLUTION.

Abstract

The present invention provides the excellent crystallization method suitable for production on the industrial scale for obtaining the reduced coenzyme Q10 crystal. The high-quality reduced coenzyme Q10 can be obtained in improving the slurry property and/or the crystallization yield by the method of present invention in which reduced coenzyme Q10 is crystallized in an aqueous solution, in particular, the method of crystallizing the reduced coenzyme Q10 by substituting an organic solvent solution containing the reduced coenzyme Q10 by water, the method of crystallizing the oily reduced coenzyme Q10 in an aqueous solution, or the like.

Full Text	DESCRIPTION METHOD OF CRYSTALLIZING REDUCED COENZYME Q10 FROM AQUEOUS SOLUTION TECHNICAL FIELD The present invention relates to a method of crystallizing reduced coenzyme Q10. Reduced coenzyme Q10 shows a higher level of oral absorbability as compared with oxidized coenzyme Q10 andis a compound useful as an ingredient ingoodfoods, functional nutritive foods, specific health foods, nutritional supplements, nutrients, drinks, feeds, animal drugs, cosmetics, medicines, remedies, preventive drugs, etc. BACKGROUND ART It is known that reduced coenzyme Q10 can be prepared by producing coenzyme Q10 in the conventional manner, for example by synthesis, fermentation, or extraction from natural products, and concentrating a reduced coenzyme Q10-containing eluate fraction resulting from chromatography (JP-A-10-109933). On that occasion, as described in the above-cited publication, the chromatographic concentration may be carried out after reduction of oxidized coenzyme Q10 contained in the reduced coenzyme-Q10 with a reducing agent such as sodium borohydride or sodium dithionite (sodium hyposulfite), or reduced coenzyme Q10 may be prepared by reacting the reducing agent mentioned above with an existing highly pure grade of coenzyme Q10 (oxidized form) . However, the thus-obtained reduced coenzyme Q10 is not always easy to be crystallized preferably but tends to occur as a low-purity crystalline, semisolid, or oily product containing such impurities as oxidized coenzyme Q10. Moreover, even when crystallization could be achieved somehow, some troubles are occurred due to its poor slurry properties, etc. For example, poor slurry fluidity causes stirring trouble or difficulty in brushing away from a crystallization container, poor filterability, and the crystallization yield which is not always high. In addition, it is very uneconomical to use a large amount of organic solvents in crystallization. Moreover, these organic solvents are brought into products and tend to give unfavorable characteristics to the products which humans take in. In order to decrease the residual amount of the organic solvent in products to a level lower than the trace amount, too much time and expensive manufacture equipment are necessary for removal of the organic solvent, drying, etc. SUMMARY OF THE INVENTION In view of the above, the present invention has its object for providing an excellent crystallization method for obtaining the reduced coenzyme Q10 crystal suitable for the industrial-scale production. As a result of intensive investigations, the present inventors found that the solubility and fluidity of the reduced coenzyme Q1o may be preferably controlled by using water, and that the high-quality reduced coenzyme Q10 crystal may be obtained by crystallizing the reduced coenzyme Q10 in an aqueous solution for improving the slurry property, yield, etc., and thereby completed the present invention. That is, the present invention relates to a method for crystallizing the reduced coenzyme Q10 which comprises a crystallization of the reduced coenzyme Q10 in an aqueous solution. DETAILED DESCRIPTION OF THE INVENTION Hereinafter, the present invention is described in detail. The reduced coenzyme Q10 which can be used in the present invention canbe obtained in the conventional manner, for example by synthesis, fermentation, or extraction from naturalproducts . Preferably, it can be obtained by reducing oxidized coenzymes Q10 such as an existing high-purity coenzyme Q10, or a mixture of the oxidized coenzyme Q10 and the reduced coenzyme Q10 using a common reducing agent. Firstly, a method for reducing the oxidized coenzyme Q10 is explained. Since reduced coenzyme Q10 is readily oxidized by molecular oxygen to give oxidized coenzyme Q10 as a byproduct, a solvent high in protective effect against oxidation is preferably used as the solvent in the step of reduction. Preferably used as such solvent is at least one species selected from among hydrocarbons, fatty acidesters, ethers, andnitriles. Hydrocarbons are most preferred. The hydrocarbons are not particularly restricted, but there may be mentioned, for example, aliphatic hydrocarbons, aromatic hydrocarbons, halogenated hydrocarbons, etc. Preferred are aliphatic hydrocarbons and aromatic hydrocarbons, and more preferred are aliphatic hydrocarbons. The aliphatic hydrocarbons are not particularly restricted, and may be cyclic or acyclic, or saturated or unsaturated. However, generally they contain 3 to 20 carbon atoms, and preferably 5 to 12 carbon atoms. As specific examples, there may be mentioned, for example, propane, butane, isobutane, pentane, 2-methylbutane, cyclopentane, 2-pentene, hexane, 2-methylpentane, 2,2-dimethylbutane, 2,3-dimethylbutane, methylcyclopentane, cyclohexane, 1-hexene, cyclohexene, heptane, 2-methylhexane, 3-methylhexane, 2,3-dimethylpentane, 2,4-dimethylpentane, methylcyclohexane,1-heptene, octane, 2,2,3-trimethylpentane, isooctane, ethylcyclohexane, 1-octene, nonane, 2,2,5-trimethylhexane, 1-nonene, decane, 1-decene, p-menthane, undecane, dodecane, etc. Among them, saturated aliphatic hydrocarbons having 5 to 8 carbonatoms are more preferred, andpreferably used are pentane, 2-methylbutane and cyclopentane, which have 5 carbon atoms (referred to as "pentanes"); hexane, 2-methylpentane, 2,2-dimethylbutane, 2,3-dimethylbutane, methylcyclopentane, cyclohexane, which have 6 carbon atoms (referred to as "hexanes"); heptane, 2-methylhexane, 3-methylhexane, 2, 3-dimethylpentane, 2, 4-dimethylpentane, methylcyclohexane, which have 7 carbon atoms (referred to as "heptanes") ; octane, 2,2,3-trimethylpentane, isooctane, ethylcyclohexane, which have 8 carbon atoms (referred to as octanes); and a mixture of these. In particular, the above heptanes are particularly preferred since they have a tendency to show a very high protection effect against oxidization, and heptane is most preferred. The aromatic hydrocarbons are not particularly restricted, but generally they contain 6 to 20 carbon atoms, preferably 6 to 12 carbon atoms, and more preferably 7 to 10 carbon atoms. As specific examples, there may be mentioned, for example, benzene, toluene, xylene, o-xylene, m-xylene, p-xylene, ethylbenzene, cumene, mesitylene, tetralin, butylbenzene, p-cymene, cyclohexylbenzene, diethylbenzene, pentylbenzene, dipentylbenzene, dodecylbenzene, styrene, etc. Preferred are toluene, xylene, o-xylene, m-xylene, p-xylene, ethylbenzene, cumene, mesitylene, tetralin, butylbenzene, p-cymene, cyclohexylbenzene, diethylbenzene and pentylbenzene. More preferred are toluene, xylene, o-xylene, m-xylene, p-xylene, cumene and tetralin, and most preferred is cumene. The halogenated hydrocarbons are not particularly restricted, and may be cyclic or acyclic, or saturated or unsaturated. However, acyclic halogenated hydrocarbons are preferably used. More preferred are chlorinated hydrocarbons and f luorinated hydrocarbons, and chlorinated hydrocarbons are still more preferred. Additionally, ones containing 1 to 6 carbon atoms, preferably 1 to 4 carbon atoms, and more preferably 1 to 2 carbon atoms are used. As specific examples, for example, there may be mentioned dichloromethane, chloroform, carbon tetrachloride, 1,1-dichloroethane, 1,2-dichloroethane, 1,1,1-trichloroethane, 1,1,2-trichloroethane, 1,1,1,2-tetrachloroethane, 1,1,2,2-tetrachloroethane, pentachloroethane, hexachloroethane, 1,1-dichloroethylene, 1,2-dichloroethylene, trichloroethylene/ tetrachloroethylene, 1,2-dichloropropane, 1,2, 3-trichloropropane, chlorobenzene, 1,1,1,2-tetrafluoroethane, etc. Preferred are dichloromethane, chloroform, carbon tetrachloride, 1,1-dichloroethane, 1,2-dichloroethane, 1,1,1-trichloroethane, 1,1,2-trichloroethane, 1,1-dichloroethylene, 1,2-dichloroethylene, trichloroethylene, chlorobenzene and 1,1,1,2-tetrafluoroethane. More preferred are dichloromethane, chloroform, 1,2-dichloroethylene, trichloroethylene, chlorobenzene and 1,1,1,2-tetrafluoroethane. The fatty acid esters are not particularly restricted, but there may be mentioned, for example, propionates, acetates, formates, etc. Preferred are acetates and formates, and more preferred are acetates. Ester functional groups thereof are not particularly restricted, but alkyl esters having 1 to 8 carbon atoms, aralkyl esters having 1 to 8 carbon atoms are used, preferred are alkyl esters having 1 to 6 carbon atoms, and more preferred alkyl esters having 1 to 4 carbon atoms. As the propionates, there may be mentioned, for example, methyl propionate, ethyl propionate, butyl propionate, isopentyl propionate, etc. Preferred is ethyl propionate. As the acetates, there may be mentioned, for example, methyl acetate, ethyl acetate, propyl acetate, isopropyl acetate, butyl acetate, isobutyl acetate, sec-butyl acetate, pentyl acetate, isopentyl acetate, sec-hexyl acetate, cyclohexyl acetate, benzyl acetate, etc. Preferred are methyl acetate, ethyl acetate, propyl acetate, isopropyl acetate, butyl acetate, isobutyl acetate, sec-butyl acetate, pentyl acetate, isopentyl acetate, sec-hexyl acetate and cyclohexyl acetate. More preferred are methyl acetate, ethyl acetate, propyl acetate, isopropyl acetate, butyl acetate and isobutyl acetate. Most preferred is ethyl acetate. As the formates, there may be mentioned, for example, methyl formate, ethyl formate, propyl formate, isopropyl formate, butyl formate, isobutyl formate, sec-butyl formate, pentyl formate, etc. Preferred are methyl formate, ethyl formate, propyl formate, butyl formate, isobutyl formate and pentyl formate, and most preferred is ethyl formate. The ethers are not particularly restricted, and may be cyclic or acyclic, or saturated or unsaturated. But saturated ones are preferably used. Generally, ones containing 3 to 20 carbon atoms, and preferably 4 to 12 carbon atoms and more preferably 4 to 8 carbon atoms are used. As specific examples, there may be mentioned, for example, diethyl ether, methyl tert-butyl ether, dipropyl ether, diisopropyl ether, dibutyl ether, dihexyl ether, ethyl vinyl ether, butyl vinyl ether, anisol, phenetole, butyl phenyl ether, methoxytoluene, dioxane, furan, 2-methylfuran, tetrahydrofuran, tetrahydropyran, ethylene glycol dimethyl ether, ethylene glycol diethyl ether, ethylene glycol dibutyl ether, ethylene glycol monomethyl ether, ethylene glycol monoethyl ether, ethylene glycol monobutyl ether, etc. Preferred are diethyl ether, methyl tert-butyl ether, dipropyl ether, diisopropyl ether, dibutyl ether, dihexyl ether, anisol, phenetole, butyl phenyl ether, methoxytoluene, dioxane, 2-methylfuran, tetrahydrofuran, tetrahydropyran, ethylene glycol dimethyl ether, ethylene glycol diethyl ether, ethylene glycol dibutyl ether, ethylene glycol monomethyl ether and ethylene glycol monoethyl ether. More preferred are diethyl ether, methyl tert-butyl ether, anisol, dioxane, tetrahydrofuran, ethylene glycol monomethyl ether and ethylene glycol monoethyl ether. More preferred are diethyl ether, methyl tert-butyl ether, anisol, etc., and most preferred is methyl tert-butyl ether. The nitriles are not particularly restricted, and may be cyclic or acyclic, or saturated or unsaturated. However, saturated ones are preferably used. Generally, ones containing 2 to 20 carbon atoms, preferably 2 to 12 carbon atoms, and more preferably 2 to 8 carbon atoms are used. As specific examples, there may be mentioned, for example, acetonitrile, propiononitrile, malononitrile, butyronitrile, isobutyronitrile, succinonitrile, valeronitrile, glutaronitrile, hexanenitrile, heptylcyanide, octylcyanide, undecanenitrile, dodecanenitrile, tridecanenitrile, pentadecanenitrile, stearonitrile, chloroacetonitrile, bromoacetonitrile, chloropropiononitrile, bromopropiononitrile, methoxyacetonitrile, methyl cyanoacetate, ethyl cyanoacetate, tolunitrile, benzonitrile, chlorobenzonitrile, bromobenzonitrile, cyanobenzoic acid, nitrobenzonitrile, anisonitrile, phthalonitrile, bromotolunitrile, methyl cyanobenzoate, methoxybenzonitrile, acetylbenzonitrile, naphthonitrile, biphenylcarbonitrile, phenylpropiononitrile, phenylbutyronitrile, methylphenylacetonitrile, diphenylacetonitrile, naphthylacetonitrile, nitrophenylacetonitrile, chlorobenzylcyanide, cyclopropanecarbonitrile, cyclohexanecarbonitrile, cycloheptanecarbonitrile, phenylcyclohexanecarbonitrile, tolylcyclohexanecarbonitrile, etc. Preferred are acetonitrile, propiononitrile, butyronitrile, isobutyronitrile, succinonitrile, valeronitrile, chloropropiononitrile, methyl cyanoacetate, ethyl cyanoacetate, tolunitrile and benzonitrile. More preferred are acetonitrile, propiononitrile, butyronitrile and isobutyronitrile, and most preferred is acetonitrile. In selecting the solvent to be used from among the solvents mentioned above, such properties as boiling point and viscosity are preferably taken into consideration; for example, the solvent should have a boiling point which allows appropriate warming for increasing the solubility and facilitates a solvent removal from wet masses by drying and solvent recovery from crystallization filtrates (about 30 to 150°C at 1 atm) , a melting i point such that solidification hardly occurs in handling at room temperature as well as upon cooling to room temperature or below (not higher than about 20°C, preferably not higher than about 10°C, still more preferably not higher than about 0°C), and a low viscosity (not higher than about 10 cp at 20°C) . From the industrial operation viewpoint, a solvent which is scarcely volatile at ordinary temperature is preferred; for example, one having a boiling point of not lower than about 80°C is preferred, and one having a boiling point of not lower than about 90°C is more preferred. Among the solvents in the reduction reactionmentioned above, a solvent having low compatibility with water is particularly preferred. The solvent in the reduction reaction promotes purifying and obtaining a reduced coenzyme Q10 efficiently, by extracting the reducing agent to be described below and/or impurities from the reducing agent and removing the same. As such solvents having low compatibility with water, there may be mentioned, for example, the above-mentioned hydrocarbons and fatty acid esters among the solvents mentioned above. Reduced coenzyme Q10, when in a dissolved state, tends to become more resistant to oxidation as the concentration thereof increases. Reduced coenzyme Q10 is highly soluble in the solvents mentioned above and, in this respect, too, the above solvents are suitable for the protection from oxidation. The concentration of reduced coenzyme Q10 which is preferred from the viewpoint of protection thereof from oxidation may vary depending on the solvent species, among others, hence cannot be absolutely specif ied. Generally, however, the concentration of reduced coenzyme Q10 in the above solvents is generally not lower than 1 w/w%, preferably not lower than 2 w/w%. The upper limit is not particularly restricted but, from the practical operability viewpoint, it is 400 w/w% or below, preferably 200 w/w% or below, more preferably 100 w/w% or below, still more preferably 50 w/w% or below. Thus, when such a solvent as mentioned above is used, it is possible to minimize the undesirable oxygen-involving side reaction via the step of reduction reaction. Additionally, the reduced coenzyme Q10 used for crystallization can be also obtained by reducing oily oxidized coenzyme Q10 in an aqueous solution. In this method, the reduced coenzyme Q10 can be synthesized without using an organic solvent, additional operations such as extraction to an organic phase, concentration, etc. are not required, the operation time can be shortened and subgeneration of the oxidized coenzyme Q10 can be minimized. The reduction reaction can be carried out, in the above solvent, using, as a reducing agent, a metal hydride compound, iron (metallic iron or iron in a salt form) , zinc (metallic zinc) , dithionous acid or a salt thereof, or an ascorbic acid or a related compound, for instance. The metal hydride compound is not particularly restricted but includes, amongothers, sodiumborohydride, lithium aluminum hydride, etc. The amount to be used of the metal hydride compound may vary depending on the species thereof, hence cannot be absolutely specified. Generally, however, the reduction can be favorably carried out by using it in an amount of 1 to 3 times the theoretical hydrogen equivalent. The reduction using iron or zinc is generally carried out using an acid. The acid to be used is not particularly restricted but includes, among others, fatty acids such as acetic acid, sulfonic acids such as methanesulfonic acid, inorganic acids_ such as hydrochloric acid and sulfuric acid, etc. Inorganic acids are preferred, and sulfuric acid is more preferred. The amount of iron to be used is not particularly restricted but, for example, an amount of about 1/5 by weight or larger based on the charged weight of oxidized coenzyme Q10 is appropriate for carrying out the reaction. The upper limit is not particularly restricted but, from the economical viewpoint, it is about twice the weight of the above charged weight or lower. Iron may be used not only in the form of metallic iron but also in the form of a salt, for example iron(II) sulfate, etc. The amount of zinc to be used is not particularly restricted but, for example, an amount of about 1/10 by weight or larger based on the charged weight of oxidized coenzyme Q10 is appropriate for carrying out the reaction. The upper limit is not particularly restricted but, from the economic viewpoint, it is about twice the weight of the above charged weight or lower. The dithionous acid or a salt thereof is not particularly restricted but a salt form of dithionous acid is generally used. The salt of dithionous acid is not particularly restricted but includes, as preferred species, alkali metal salts, alkaline earth metal salts, ammonium salt and the like. Alkali metal salts such as the lithium salt, sodium salt, and potassium salt are more preferred, and the sodium salt is most preferred. The amount to be used of the dithionous acid or salt is not particularly restricted but it is generally not smaller than about 1/5 by weight, preferably not smaller than about 2/5 by weight, and more preferably not smaller than about 3/5 by weight, based on the charged weight of oxidized coenzyme, Q10. Larger amounts may be used without causing any particularly trouble. From the economical viewpoint, however, the amount to be employed is not larger than about twice the weight of the above-mentioned charged weight, preferably not larger than the charged weight. Thus, the reaction can be more favorably carried out with employing ah amount within the range of about 2/5 by weight of the above-mentioned charge to a weight roughly equal to that of the charged weight. The ascorbic acid or a related compound are not particularly restricted, and include, for example, not only ascorbic acid, but also rhamno-ascorbic acid, arabo-ascorbic acid, gluco-ascorbic acid, fuco-ascorbic acid, glucohepto-ascorbic acid, xylo-ascorbic acid, galacto-ascorbic acid, gulo-ascorbic acid, allo-ascorbic acid, erythro-ascorbic acid, 6-desoxyascorbic acid, and the like ascorbic acid-related compounds, and may be ester forms or salts of these. Furthermore, these may be L-form, D-form or racemic form. More specifically, there may be mentioned, for example, L-ascorbic acid, L-ascorbyl palmitate, L-ascorbylstearate,D-arabo-ascorbicacid,etc. In producing the reduced coenzyme Q10, any of the above-mentioned ascorbic acidor related compounds may be suitably used. However, the water-soluble ones are suitably used in particular among the above-mentioned ascorbic acid or related compounds in view of separatability with the generated reduced coenzyme Q10, etc. And most preferred is a free form of L-ascorbic acid, D-arabo-ascorbic acid, and the like in view of the ready availability, price, etc. The amount to be used of the ascorbic acid or related compounds mentioned above is not particularly restricted but may be an amount effective in converting oxidized coenzyme Q10 to reduced coenzyme Q10. Generally it is not smaller than 1 mole, preferably not smaller than 1.2 moles, per mole of oxidized coenzyme Q10. The upper limit is not particularly restricted but, from the economical viewpoint, it is generally not higher than 10 moles, preferably not higher than 5 moles, and more preferably not higher than 3 moles, per mole of the oxidized coenzyme Q10. Among the reducing agent species mentioned above, zinc, dithionous acid or salts'thereof, and ascorbic acidor related compounds are preferred from the viewpoint of reducing ability, yield and/or quality, among others. In particular, dithionous acid or. salts thereof (specifically dithionous acidsalts) andascorbic acidor related compounds (particularly, free-form or salt thereof) are preferred from a viewpoint that they bring the reducing agent or impurities derived from the reducing agent into the reduced coenzyme Q10 crystal in only the trace amount or lower. In carrying out the reduction reaction, an alcohol and/or water are/is suitably used singly or in combination, as to be mentioned below. Water is preferred in particular when iron, zinc, or dithionous acid or a salt thereof is used as the reducing agent. When a metal hydride compound or an ascorbic acid or a related compound is used as the reducing agent, an alcohol can be used in combination. The combined use of water and an alcohol exhibits the characteristics of both water and the alcohol and contributes to improvements in reaction rate and yield, among others. In the following, a preferred method of reduction is described in detail. The reduction using dithionous acid or a salt thereof is preferably carried out using water in combination, namely in a mixed solvent system composed of at least one organic solvent selected from among the above-mentioned hydrocarbons, fatty acid esters, ethers, and nitriles, with water. On that occasion, the reaction is preferably carried out generally at a pH of not higher than 7, preferably at pH 3 to 7, more preferably at pH 3 to 6, from the viewpoint of yield, etc. The pH can be adjusted using an acid (e.g. an inorganic acid such as hydrochloric acid or sulfuric acid) or a base (e.g. an alkali metal hydroxide such as sodium hydroxide). In the reduction using dithionous acid 1 or a salt thereof, the amount of water is not particularly restricted but may be an amount of water such that an appropriate amount of the reducing agent, namely dithionous acid or a salt thereof, can be dissolved therein. Thus, for example, it is advisable that the amount of the dithionous acid or a salt be adjusted generally to not more than 30 w/w%, and preferably not more than 20 w/w%, relative to the weight of water. From the productivity viewpoint, among others, it is advisable that the amount be adjusted generally to not less than 1 w/w%, preferably not less than 5 w/w%, and more preferably not less than 10 w/w%. The reduction using the ascorbic acid or a related compound mentioned above can.be carried out using a solvent especially highly miscible with water as selected from among the above-mentioned hydrocarbons, fatty acid esters, ethers, and nitriles, in particular ethers and nitriles, which are highly miscible with water, and more specifically tetrahydrofuran, dioxane, acetonitrile or the like. It is particularly preferred to use the alcohols and/or ketones to be mentioned below (preferably alcohols and/or ketones, which are highly miscible with water (in particular, monohydric or dihydric alcohols (preferably monohydric ones) having 1 to 5 carbon atoms, preferably 1 to 4 carbon atoms, more preferably 1 to 3 carbon atoms, and/or, ketones such as acetone, methyl ethyl ketone or the like)). Namely, in the reduction using the ascorbic acid or a related compound, it is preferable to use alcohols and/or water-soluble organic solvents (the above ethers, nitriles, ketones and the like, which are highly miscible with water, for example). Furthermore, from the viewpoint of reactionpromotion (e.g. reaction temperature lowering or reaction time shortening) in the production of reduced coenzyme Q10, it is also possible to carry out the reduction using the ascorbic acid or a related compound in the presence of an additive having a reaction promoting effect, such as a basic substance or a hydrogensulf ite . The basic compound is not particularly restricted but may be either an inorganic compound or an organic compound. The inorganic compound is not particularly restricted but includes, among others, the hydroxides, carbonates and hydrogencarbonates of metals (preferably alkali metals, alkaline earth metals, and the like), ammonia, etc. As typical examples thereof, there may be mentioned alkali metal hydroxides such as sodium hydroxide, alkali metal carbonates such as sodium carbonate, alkali metal hydrogencarbonates such as sodium hydrogencarbonate, and alkaline earth metal carbonates such as magnesium carbonate. The organic compound is not particularly restrictedbut includes, among others, amines such as triethylamine, etc. Among the basic substances specifically mentioned above, weakly basic substances (weak bases or weak alkalis) such as the carbonates and hydrogencarbonates of metals (preferably alkali metals, alkaline earth metals, etc.), ammonia, and like inorganic compounds; amines such as triethylamine, and like organic compounds are preferably used. More preferred are the weakly basic inorganic compounds mentioned above. Preferred as the hydrogensulfite are, for example, alkali metal hydrogensulfites such as sodium hydrogensulfite, etc. The amount of the additive mentioned above is not particularly restricted but may be such that the reaction promoting effect of the additive can be produced to a desired extent (effective amount). From the economical viewpoint, however, the amount is generally not more than 20 moles, preferably not more than 10 moles, more preferably not more than 5 moles, and still more preferably not more than 2 moles, per mole of the ascorbic acid or a related compound. The lower limit is not particularly restricted but, generally, it is not less than 0.01 moles, preferably not less than 0.05 moles, more preferably not less than 0.1 moles, and still more preferably not less than 0.2 moles, per mole of the ascorbic acid or a related compound. The reduction reaction is preferably carried out under forced flowing. The power required for stirring to cause such flowing per unit volume is generally not less than about 0.01 kW/m3, preferably not less than about 0.1 kW/m3, and more preferably not less than about 0.3 kW/m3. The above forced flowing is generally causedby the turningof a stirringblade (s) . The use of a stirring blade (s) is not always necessary if the above flowing can be otherwise obtained. For example,a method based on liquid circulation may be utilized. The reduction temperature may vary depending on the reducing agent species and/or amount, hence cannot be absolutely specified. In the reduction using dithionous acid d or a salt thereof, for instance, the reduction is generally carried out at 100°C or below, preferably at 80°C or below, more preferably at 60°C or below. The lower limit is the solidification temperature of the system. Thus, the reduction can be favorably carried out generally at about 0 to 100°C, preferably at about 0 to 80°C, more preferably at about 0 to 60°C. In the reduction using an ascorbic acid or a related compound, the reduction is carried out generally at 30°C or higher, preferably at 40°C or higher, more preferably at 50°C or higher. The upper limit is the boiling point of the system. Thus, the reduction can be favorably carried out generally at about 30 to 150°C, preferably about 40 to 120°C, more preferably at about 50 to 100°C. The reduction of the oily oxidized coenzyme Q10 is generally carried out at 45°Cormore, preferablyat 48°Cormore, andmorepreferably at 50°C or more, although it is dependent on the purity, etc., then the oily reduced coenzyme Q10 can be obtained. The reaction concentration is not particularly restricted but the weight of oxidized coenzyme Q10 relative to the solvent weight is generally not less than about 1 w/w%, preferably not less than 3 w/w %, more preferably not less than 10 w/w%, and still more preferably not less than 15 w/w%. The upper limit is not particularly restricted but generally is not higher than about 60 w/w%, preferably not higher than 50 w/w%, more preferably not higher than 40 w/w%, and still more preferably not higher than 30 w/w%. Thus, the reaction can be favorably carried out at a reaction concentration of about 1 to 60 w/w%, preferably about 3 to 50 w/w%, and more preferably about 10 to 40 w/w%. The reduction reaction time may vary depending on the reducing agent species and/or the amount thereof, hence cannot be absolutely specified. Generally, however, the reaction can be driven to completion within 48 hours, preferably within 24 hours, more preferably within 10 hours, and still more preferably within 5 hours. After the reduction reaction, an organic phase containing the product reduced coenzyme Q10 or the oily reduced coenzyme Q10 is recovered (e.g. recovered by separation, extraction, concentration, etc.), and if necessary (preferably) , the organic phase is further washed repeatedly using water, brine, or the like to achieve contaminant elimination and, then, it can be subjected to crystallization as such or after dissolution to or substitution by other desirable solvents. As the other solvents, there may be mentioned, for example, hydrocarbons, fatty acid esters, ethers, alcohols, fatty acids, ketones, nitrogen compounds (inclusive of nitriles and amides) , sulfur compounds, etc. mentioned above or below. They were found that, the above-mentioned series of processes from the reduction reaction to the aftertreatment is particularly preferably carried out under a deoxygenated atmosphere, and, in the reduction reaction using dithionous acid or a salt thereof, in particular, such atmosphere greatly contributes to an improvement in reduction reaction yield and a reduction in reducing agent amount. The deoxygenated atmosphere can be attained by substitution with an inert gas, pressure reduction, boiling, or a combination of these. It is preferable to carry out at least the substitution with an inert gas, namely to use an inert gas atmosphere. As the inert gas, there may be mentioned, for example, nitrogen gas, helium gas, argon gas, hydrogen gas, and carbon dioxide gas. Nitrogen gas is preferred, however. The crystallization of reduced coenzyme Q10 of the invention is now described. The reduced coenzyme Q10 to be subj ected to crystallization can be obtained in the conventional manner, for example, by synthesis, fermentation, or extraction from a natural source. Preferred is the product obtained by reduction of oxidized coenzyme Q10. More preferred is a solution or oil, of the reduced coenzyme Q10 obtained by carrying out the reduction reaction in accordance with the present invention, as described above. Still more preferred is the one obtained by reducing the oily oxidized coenzyme Q10 in an aqueous solution using dithionite , or the one obtained by reducing the oxidized coenzyme Q10 using alcohols and/or water-soluble organic solvents. While the method of crystallization according to the invention can be applied also to products containing oxidized coenzyme Q10 in relatively large amounts, the method is particularly effective in crystallizing high-purity reduced coenzyme Q10 prepared by the reduction method described above. In the present invention, the reduced coenzyme Q10 is crystallized in an aqueous solution. Use of water contributes to the economical efficiency obviously, and to the safety on industrial operation and product safety as well. Moreover, the use of water also contributes to improvement of the slurry property and the yield of the reduced coenzyme Q10, and can promote isolation of the reduced coenzyme Q10 efficiently by leaving a reducing agent used in the reduction reaction or impurities derived from the reducing agent in amother liquor. Furthermore, the above-mentioned use of water, or preferably the use of water containing salts can protect the reduced coenzyme Q10 from oxidization by molecular oxygen. The above salts are not particularly restricted but there may be mentioned, for example, salts constituted from alkaline metals such as lithium, sodium and potassium; alkaline earth metals such as magnesium and calcium with halogen atoms such as fluorine, chlorine and bromine; a residue obtained by removing a proton from inorganic acids such as sulfuric acid and organic acids such as formic acid, acetic acid andpropionic acid. Among these, preferred are inorganic salts, and more preferred are sodium chloride, potassium chloride, sodium sulfate, etc. The concentration of the above salts is preferably high, and it is generally 3 w/w% or more, preferably 5 w/w% or more, more preferably 10 w/w% or more. Still more preferred is that the above salts are dissolved in water at saturation or close to saturation. As the method of crystallizing the reduced coenzyme Q10 mentioned above, general crystallization methods such as crystallization by cooling, crystallization by concentration and crystallization by solvent substitution can be used. It is preferred to use the crystallization by cooling singly or in combination. As particularly preferable embodiments, the following two methods may be mentioned. (1) A crystallization method which comprises substituting an organic solvent solution containing the reduced coenzyme Q10 (e.g. reaction solution, extraction solution, etc.) by water (for increasing the composition ratio of water). (2) A method of crystallizing the oily reduced coenzyme Q10 in an aqueous solution. In the above-mentioned two methods, it is more preferable to use the crystallization by cooling singly or in combination. Firstly, the method (1) is explained. The organic solvent containing the reduced coenzyme Q10 is not particularly restricted but there may be mentioned, for example, hydrocarbons, fatty acid esters, ethers, alcohols, fatty acids, ketones, nitrogen compounds (inclusive of nitriles and amides), sulfur compounds, etc. As the hydrocarbons, fatty acidesters, ethers andnitriles, those exemplified as the reaction solvent in the fore-mentioned explanation about reduction of the oxidized coenzyme Q10 can be preferably used. The alcohols are not particularly restricted but may be cyclic or acyclic, or saturated or unsaturated. Saturated ones are preferred, however. Generally, they contain 1 to 20 carbon atoms, preferably 1 to 12 carbon atoms, more preferably 1 to 6 carbon atoms. Still more preferred are monohydric alcohols containing 1 to 5 carbon atoms, dihydric alcohols containing 2 to 5 carbon atoms, and the trihydric alcohol containing 3 carbon atoms. As the monohydric alcohol, there may be mentioned, for example, methanol, ethanol, 1-propanol, 2-propanol, 1-butanol, 2-butanol, isobutyl alcohol, tert-butyl alcohol, 1-pentanol, 2-pentanol, 3-pentanol, 2-methyl-l-butanol, isopentylalcohol, tert-pentyl alcohol, 3-methyl-2-butanol, neopentyl alcohol, 1-hexanol, 2-methyl-l-pentanol, 4-methyl-2-pentanol, 2-ethyl-l-butanol, 1-heptanol, 2-heptanol, 3-heptanol, 1-octanol, 2-octanol, 2-ethyl-l-hexanol,1-nonanol, 1-decanol, 1-undecanol, 1-dodecanol, allyl alcohol, propargyl alcohol, benzyl alcohol, cyclohexanol, 1-methylcyclohexanol, 2-methylcyclohexanol, 3-methylcyclohexanol, 4-methylcyclohexanol, etc. Preferred are methanol, ethanol, 1-propanol, 2-propanol, 1-butanol, 2-butanol, isobutyl alcohol, tert-butyl alcohol, 1-pentanol, 2-pentanol, 3-pentanol, 2-methyl-l-butanol, isopentyl alcohol, tert-pentyl alcohol, 3-methyl-2-butanol, neopentyl alcohol, 1-hexanol, 2-methyl-l-pentanol, 4-methyl-2-pentanol, 2-ethyl-l-butanol and cyclohexanol. More preferred are methanol, ethanol, 1-propanol, 2-propanol, 1-butanol, 2-butanol, isobutyl alcohol, tert-butyl alcohol, 1-pentanol, 2-pentanol, 3-pentanol, 2-methyl-l-butanol, isopentyl alcohol, tert-pentyl alcohol, 3-methyl-2-butanol and neopentyl alcohol. Still more preferred are methanol, ethanol, 1-propanol, 2-propanol,1-butanol, 2-butanol, isobutylalcohol, 2-methyl-l-butanol and isopentyl alcohol. Most preferred is ethanol. As the dihydr ic alcohol, there may be mentioned, for example, 1,2-ethanediol, 1,2-propandiol, 1,3-propandiol, 1,2-butanediol, 1,3-butanediol, 1,4-butanediol, 2,3-butanediol, 1,5-pentanediol, etc. Preferred are 1,2-ethanediol, 1,2-propandiol and 1,3-propandiol, and most preferred is 1,2-ethanediol. As the trihydric alcohol, glycerol, etc. may be preferably used, for example. As fatty acids, there maybe mentioned, for example, formic acid, acetic acid, propionic acid, etc. Preferred are formic acid and acetic acid, and most preferred is acetic acid. The ketones are not particularly restricted, and ones having 3 to 6 carbon atoms are preferably used in general. As specific examples, there may be mentioned, for example, acetone, methyl ethyl ketone, methyl butyl ketone, methyl isobutyl ketone, etc. Preferred are acetone and methyl ethyl ketone, and most preferred is acetone. As the nitrogen compounds other than nitriles, there may be mentioned, for example, nitromethane, triethylamine, pyridine, formamide, N-methylformamide, N, N-dimethylformamide, N,N-dimethylacetoamide, N-methylpyrrolidone, etc. As the sulfur compounds, there may be mentioned, for example, dimethyl sulfoxide, sulfolane, etc. In selecting the solvent to be used from among the solvents mentioned above, such properties as boiling point and viscosity (for example, the solvent should have a boiling point which allows appropriate warming for increasing the solubility and facilitates a solvent removal from wet masses by drying and solvent recovery from crystallization filtrates (about 30 to 150°C at 1 atm) , a melting point such that solidification hardly occurs in handling at room temperature as well as upon cooling to room temperature or below (not higher than about 20°C, preferably not higher than about 10°C, still more preferably not higher than about 0°C) , and a low viscosity (not higher than about 10 cp at 20°C) ) are preferably taken into consideration. From the industrial operation viewpoint, a solvent which is scarcely volatile at ordinary temperature is preferred. Among the above solvents, in particular, hydrocarbons, fatty acid esters, ethers and nitriles can be preferably used in view of the oxidization protection of the reduced coenzyme Q10mentioned above. Additionally, in view of obtaining the high yield while preferably decreasing the solubility of the reduced coenzyme Q10, alcohols, fattyacids, ethers, ketones and nitriles can be preferably used. From a viewpoint of an industrial applicability, for example, hydrocarbons and alcohols can be preferably used. In the above method (1) , as a means for substituting the organic solvent solution containing the reduced coenzyme Q10 by water, there maybe mentioned, for example, a method comprising increasing the water ratio while concentrating and removing the organic solvent. Furthermore, if necessary, operations such as cooling and seed crystal addition, as to be described below, can be combinedly used properly. Next, the method (2), i.e., the method of crystallizing the oily reduced coenzyme Q10 in an aqueous solution is explained. According to this method, the reduced coenzyme Q10 crystal having a large particle diameter can be obtained, and filterability can be remarkably improved. The crystallization may be carried out by adding water to the oily reduced coenzyme Q10, for example, or on the contrary, by adding the oily reduced coenzyme Q10 to water. Moreover, the crystallization may be also carried out by cooling a mixture of the oily reduced coenzyme Q10 and water. More preferred is a method comprising removing an organic solvent from a mixed solvent solution consisting of an organic solvent containing the reduced coenzyme Q10 and water at a temperature not lower than the melting point of the reduced coenzyme Q10 or of a concentrate comprising the reduced coenzyme Q10 as a main component, to obtain oil in the system, and cooling to crystallize the oil. The above-mentioned temperature not lower than the melting point is generally 45°C or more, preferably 48°C or more, and more preferably 50°C or more although it is dependent on the purity, etc. of the reduced coenzyme Q10. The upper limit is not particularly restricted but generally 100°C or less is preferred, 80°C or less is more preferred, and 60°C or less is still more preferred. In the crystallization method of the present invention as described above, the crystallization temperature of the reduced coenzyme Q10 (cooling temperature at the time of crystallization) is generally 48°C or less, preferably 45°C or less, more preferably 40°C or less, and still more preferably 30°C or less in view of the yield, etc., although it is difficult to uniformly define since it is dependent on the purity of the reduced coenzyme Q10. The lower limit is a solidification temperature of the system. Therefore, the crystallization can be especially preferably carried out at the crystallization temperature of about 0 to 30°C. It is preferable to control the amount of crystallization per unit time in crystallizing, i.e. the rate of crystallization. The preferable amount of crystallization per unit time is, for example, not higher than the rate of crystallization which causes crystallization of about 50%, per unit time, of the whole amount of crystals to be obtained (i.e. at most 50%/hour), preferably not higher than the rate of crystallization which causes crystallization of about 25%, per unit time, of the whole amount of crystals to be obtained (i.e. at most 25%/hour). The rate of cooling in the crystallization by cooling is generally not higher than about 40°C/hour, and preferably not higher than about 20°C/hour. The crystallization is preferably carried out under forced flowing in order to prevent the state of supersaturation from occurring and thereby allowing the nucleation and crystal growth to proceed smoothly, in order to obtain crystals with uniform particle diamater and, furthermore, from the viewpoint of obtaining high-quality products. The flowing is generally brought about by a stirring power per unit volume of not weaker than about 0.01 kW/m3, preferably not weaker than about 0.1 kW/m3, and more preferably not weaker than about 0.3 kW/m3. The forced flowing is generally provided by the turning of a stirring blade (s) . However, the use of a stirring blade (s) is not always necessary if the above flowing can be otherwise obtained. For example, it is possible to utilize a method based on liquid circulation. In carrying out the crystallization, seed crystals are preferably added so that the state of supersaturation may be prevented from occurring and the nucleation and crystal growth may be allowed to proceed smoothly. The crystallization concentration, when expressed in terms of the weight of reduced coenzyme Q10 relative to the total weight of the crystallization solvent at the time of completion of crystallization, it is not higher than about 15 w/w%, preferably not higher than about 13 w/w%, more preferably not higher than 10 w/w%. From the productivity viewpoint, the lower limit to the crystallization concentration is generally not lower than about 1 w/w%, preferably not lower than about 2 w/w%. The thus-obtained crystals of reduced coenzyme Q10 can be recovered as a wet product, for example, by such a solid-liquid separation technique as centrifugation, pressure filtration, or vacuum filtration, if necessary followed by cake washing. They can be recovered also as a dry product by further charging the wet product in a reduced pressure drier (vacuum drier) internally purged with an inert gas and drying the same under reduced pressure. The recovery in a dry form is preferred. The crystallization of the invention, when it is carried out in a deoxygenated atmosphere, can increase protective effect against oxidation. The deoxygenated atmosphere can be attained by inert gas substitution, pressure reduction, boiling, or a combination of these. It is preferable to carry out at least the substitution with an inert gas, namely to use an inert gas atmosphere. As the inert gas, there maybe mentioned, for example, nitrogen gas, helium gas, argon gas, hydrogen gas, and carbon dioxide gas. Nitrogen gas is preferred, however. In accordance with the present invention, high-quality reduced coenzyme Q10 can be obtained with excellent workability and economical efficiency. The crystals of reduced coenzyme Q10 as obtained in accordance with the present invention are of very high quality and can be expected to have a reduced coenzyme Q10/oxidized coenzyme Q10 weight ratio of not lower than 96/4, preferably not lower than 98/2, more preferably not lower than 99/1. BEST MODE FOR CARRYING OUT THE INVENTION The following examples illustrate the present invention in further detail. These examples are, however, by no means limitative of the scope of thepresent invention. In the examples, the purity of reduced coenzyme Q10 and the reduced coenzyme Q10/oxidized coenzyme Q10 weight ratio were determined by the HPLC analysis specif ied below. The reduced coenzyme Q10 purity values as determined, however, are by no means indicative of the limit purity value attainable in accordance with the present invention. Likewise, the reduced coenzyme Q10/oxidized coenzyme Q10 weight ratio values obtained never indicate the upper limit to that ratio. (HPLC conditions) Column: SYMMETRY C18 (product of Waters), 250 mm (in length), 4.6 mm (in inside diameter); mobile phase: C2H5OH:CH3OH = 4:3 (v/v); detection wavelength: 210 nm; flow rate: 1 ml/min; retention time of reduced coenzyme Q10: 9.1 min; retention time of oxidized coenzyme Q10: 13.3 min. (Example 1) Oxidized coenzyme Q10 (100 g; purity 99.4%) was melted with stirring at 48°C. While stirring (power required for stirring: 0.3kW/m3), an aqueous solution prepared by dissolving 100 gof sodiumdithionite (purity: atleast 75%), as the reducing agent, in 1000 ml of water was gradually added to the oil and the reduction reaction was carried out at 48°C and at pH 4 to 6. The aqueous phase was removed from the reaction mixture containing the oil, and the oil was washed 6 times with 1000 g of deaerated saturated brine at 48°C. After that, by removing the aqueous phase, oily reduced coenzyme Q10 was obtained. 1500 g pf deaerated water, at 48°C was added to this oil, and while stirring (power required for stirring: 0.3 kW/m3) , the mixture was cooled to 2°C to obtain white slurry (fluidity of the slurry was good). All the above operations were carried out under a nitrogen atmosphere. The obtained slurry was filtered under reduced pressure, the wet crystal was washed with cold ethanol, cold water and cold ethanol in this order (the temperature of the cold solvent used for washing was 2°C) , and further the wet crystal was dried under reduced pressure (20 to 40°C, 1 to 30 mmHg), 97 g of dry white crystal was obtained (isolated product yield: 97 mol %) . The weight ratio of the reduced coenzyme Q10/oxidized coenzyme Q10 of the obtained crystal was 99.4/0.6, and the purity of the reduced coenzyme Q10 was 99.2%. (Example 2) The oxidized coenzyme Q10 (100 g; purity 99.4%) was dissolved in 1000 g of heptane at 25°C. While stirring (power required for stirring: 0.3 kW/m3) , an aqueous solution dissolving 100 g of sodiumdithionite (purity 75% or more) , as a reducing agent, in 1000 ml of water was gradually added and a reduction reaction was carried out at 25°C and at pH 4 to 6. 2 hours later, an aqueous phase was removed from the reaction solution, and a heptane phase was washed 6 times with 1000 g of deaerated saturated brine. 1000 g of water was added to this heptane phase, and while stirring (power required for stirring: 0.3kW/m3), heptane was removed by reducing pressure at 3 0°C to obtain white slurry. This slurry had good fluidity and was capable to be easily brushed away from the crystallization container. All the above operations were carried out under a nitrogen atmosphere. The obtained slurry was filtered under reduced pressure, and the wet crystal was washed with cold ethanol, cold water and cold ethanol in this order (the temperature of the cold solvent used for washing was 2°C) . The wet crystal was further dried under reduced pressure (20 to 40°C, 1 to 30 mmHg), and a dry white crystal was obtained ("isolated product yield: 97 mol %) . The weight ratio of the reduced coenzyme Q10/oxidized coenzyme Q10 of the obtained crystal was 99.5 / 0.5, and the purity of the reduced coenzyme Q10 was 99.2%. (Example 3) To 1000 g of ethanol, 100 g of the oxidized coenzyme Q10 (purity 99.4%), 60 g of L-ascorbic acid and 30 g of sodium hydrogencarbonate were added, and the mixture was stirred at 78°C and subjected to a reduction reaction. 3 hours later, the mixture was cooled to 50°C, and 1000 g of heptane and 1000 g of deaerated water were added while holding said temperature. After the mixture was cooled to 25°C, an aqueous phase was removed, and further washed 6 times with 1000 g of saturatedbrine to remove the aqueous phase. Heptane was removed from this heptane solution at 48°C to obtain the oily reduced coenzyme Q10. To this oil, 1500 g of deaerated water at 48°C was added, and while stirring (power required for stirring: 0.3 kW/m3), the mixture was cooled to 2°C to obtain white slurry (fluidity of the slurry was good as in Example 1) . All the above operations were carried out under the nitrogen atmosphere. The obtained slurry was filtered under reduced pressure, and the wet crystal was washed with cold ethanol, cold water and cold ethanol in this order (the temperature of the cold solvent used for washing was 2°C) . The wet crystal was further dried under reduced pressure (20 to 40°C, 1 to 30 mmHg) to obtain 97 g of dry white crystal (isolated product yield: 97 mol%). The weight ratio of the reduced coenzyme Q10/oxidized coenzyme Q10 of the obtained crystal was 99.4/0.6, and the purity of the reduced coenzyme Q10 was 99.2%. (Comparative Example 1) A heptane phase of the reduced coenzyme Q10 after being washed with deaerated saturated brine was obtained in the same manner as Example 2. While stirring this heptane phase (power required for stirring: 0.3 kW/m3) , the mixture was cooled to 2°C to obtain white slurry. This slurry was poor in fluidity, and more difficultly brushed away from the crystallization container as compared with Example 1. All the above operations were carried out under the nitrogen atmosphere. The obtained slurry was filtered under reduced pressure, the wet crystal was washed with cold ethanol, cold water and cold ethanol in this order (the temperature of the cold solvent used for washing was 2°C) . The wet crystal was further dried under reduced pressure (20 to 40°C, 1 to 30 mmHg) to obtain 93 g of dry white crystal (isolated product yield: 93 mol %) . The weight ratio of the reduced coenzyme Q10/oxidized coenzyme Q10 of the obtained crystal was 99.6/0.4, and the purity of the reduced coenzyme Q10 was 99.3%. (Reference Example 1) One gram of the reduced coenzyme Q10 (the weight ratio of the reduced coenzyme Q10/oxidized coenzyme Q10 is 99.6/0.4) was dissolved in 20 g of various solvents shown in Table 1 at 25°C. In the atmosphere, the weight ratio of the reduced coenzyme Q10/oxidized coenzyme Q10 in the solution after stirring for 24 hours at 25°C, and the result is shown in Table 1. (Reference Example 2) One gram of the reduced coenzyme Q10 (the weight ratio of the reduced coenzyme Q10/oxidized coenzyme Q10 is 99.6/0.4) was dissolvedin 100 gof various solvents shown in Table 2 at 35°C. In the atmosphere, the weight ratio of the reduced coenzyme Q10/oxidized coenzyme Q10 in the solution after stirring for 24 hours at 35°C was measured, and the result is shown in Table 2. INDUSTRIAL APPLICABILITY The invention, which has the constitution described above, is a method excellent in workability and economical efficiency on the industrial scale and can give high-quality reduced coenzyme Q10 in a convenient and efficient manner. WE CLAIM: 1. A method of crystallizing a reduced coenzyme Q10 which comprises crystallizating the reduced coenzyme Q10 in an aqueous solution. 2. The method as claimed in claim 1, wherein the reduced coenzyme Q10 is crystallized by substituting an organic solvent such as herein described containing the reduced coenzyme Q10 by water. 3. The method as claimed in claim 1, wherein oily reduced coenzyme Q10 is crystallized in an aqueous solution. 4. The method as claimed in claim 3, wherein the crystallization is carried out by cooling a mixture of the oily reduced coenzyme Q10 and water. 5. The method as claimed in claim 4, wherein an organic solvent such as herein described is removed from a solution of a mixed solvent comprising an organic solvent containing the reduced coenzyme Q10 and water at a temperature not lower than the melting point of the reduced coenzyme Q10 or of a concentrate comprising the reduced coenzyme Q10 as a main component, to obtain oily reduced coenzyme Q10 in the system, and cooled to crystallize the oily reduced coenzyme Q10 6. The method as claimed in any one of claims 1 to 5, wherein the crystallization by cooling is used singly or in combination. 7. The method as claimed in any one of claims 1 to 6, wherein the crystallization temperature is 48°C or less. 8. The method as claimed in any one of claims 1 to 7, wherein the crystallization is carried out under forced flowing with a stirring power per unit volume of not weaker than 0.01 kw/m3 . 9. The method as claimed in any one of claims 1 to 8, wherein the seed crystal is added in the crystallization. 10. The method as claimed in any one of claims 1 to 9, in which the rate of crystallization is not higher than the rate of crystallization which causes crystallization of 50%, per unit time, of the whole amount of crystals. 11. The method as claimed in any one of claims 1 to 10, wherein the crystallization is carried out under a deoxygenated atmosphere. 12. The method as claimed in any one of claims 1 to 11, wherein the reduced coenzyme Q10 used for the crystallization is obtained by reducing an oxidized coenzyme Q10. 13. The method as claimed in any one of claims 1 to 12, wherein the reduced coenzyme Q10 used for the crystallization is obtained by reducing oily oxidized coenzyme Q10 in an aqueous solution using dithionites. 14. The method as claimed in any one of claims 1 to 12, wherein the reduced coenzyme Q10 used for the crystallization is obtained by reducing the oxidized coenzyme Q10 using alcohols and/or water-soluble organic solvents. 15. The method as claimed in any one of claims 1 to 14, wherein a reducing agent used in the reduction reaction or impurities derived from the reducing agent is removed to a mother liquor in crystallizing the reduced coenzyme Q10. The present invention provides the excellent crystallization method suitable for production on the industrial scale for obtaining the reduced coenzyme Q10 crystal. The high-quality reduced coenzyme Q10 can be obtained in improving the slurry property and/or the crystallization yield by the method of present invention in which reduced coenzyme Q10 is crystallized in an aqueous solution, in particular, the method of crystallizing the reduced coenzyme Q10 by substituting an organic solvent solution containing the reduced coenzyme Q10 by water, the method of crystallizing the oily reduced coenzyme Q10 in an aqueous solution, or the like.

Full Text

DESCRIPTION
METHOD OF CRYSTALLIZING REDUCED COENZYME Q10
FROM AQUEOUS SOLUTION
TECHNICAL FIELD
The present invention relates to a method of crystallizing
reduced coenzyme Q10. Reduced coenzyme Q10 shows a higher level
of oral absorbability as compared with oxidized coenzyme Q10
andis a compound useful as an ingredient ingoodfoods, functional
nutritive foods, specific health foods, nutritional supplements,
nutrients, drinks, feeds, animal drugs, cosmetics, medicines,
remedies, preventive drugs, etc.
BACKGROUND ART
It is known that reduced coenzyme Q10 can be prepared by
producing coenzyme Q10 in the conventional manner, for example
by synthesis, fermentation, or extraction from natural products,
and concentrating a reduced coenzyme Q10-containing eluate
fraction resulting from chromatography (JP-A-10-109933). On
that occasion, as described in the above-cited publication, the
chromatographic concentration may be carried out after reduction
of oxidized coenzyme Q10 contained in the reduced coenzyme-Q10
with a reducing agent such as sodium borohydride or sodium
dithionite (sodium hyposulfite), or reduced coenzyme Q10 may
be prepared by reacting the reducing agent mentioned above with
an existing highly pure grade of coenzyme Q10 (oxidized form) .
However, the thus-obtained reduced coenzyme Q10 is not
always easy to be crystallized preferably but tends to occur
as a low-purity crystalline, semisolid, or oily product
containing such impurities as oxidized coenzyme Q10. Moreover,
even when crystallization could be achieved somehow, some
troubles are occurred due to its poor slurry properties, etc.
For example, poor slurry fluidity causes stirring trouble or
difficulty in brushing away from a crystallization container,
poor filterability, and the crystallization yield which is not
always high.
In addition, it is very uneconomical to use a large amount
of organic solvents in crystallization. Moreover, these
organic solvents are brought into products and tend to give
unfavorable characteristics to the products which humans take
in. In order to decrease the residual amount of the organic
solvent in products to a level lower than the trace amount, too
much time and expensive manufacture equipment are necessary for
removal of the organic solvent, drying, etc.
SUMMARY OF THE INVENTION
In view of the above, the present invention has its object
for providing an excellent crystallization method for obtaining
the reduced coenzyme Q10 crystal suitable for the
industrial-scale production.
As a result of intensive investigations, the present
inventors found that the solubility and fluidity of the reduced
coenzyme Q1o may be preferably controlled by using water, and
that the high-quality reduced coenzyme Q10 crystal may be obtained
by crystallizing the reduced coenzyme Q10 in an aqueous solution
for improving the slurry property, yield, etc., and thereby
completed the present invention.
That is, the present invention relates to a method for
crystallizing the reduced coenzyme Q10 which comprises a
crystallization of the reduced coenzyme Q10 in an aqueous
solution.
DETAILED DESCRIPTION OF THE INVENTION
Hereinafter, the present invention is described in detail.
The reduced coenzyme Q10 which can be used in the present
invention canbe obtained in the conventional manner, for example
by synthesis, fermentation, or extraction from naturalproducts .
Preferably, it can be obtained by reducing oxidized coenzymes
Q10 such as an existing high-purity coenzyme Q10, or a mixture
of the oxidized coenzyme Q10 and the reduced coenzyme Q10 using
a common reducing agent.
Firstly, a method for reducing the oxidized coenzyme Q10
is explained. Since reduced coenzyme Q10 is readily oxidized
by molecular oxygen to give oxidized coenzyme Q10 as a byproduct,
a solvent high in protective effect against oxidation is
preferably used as the solvent in the step of reduction.
Preferably used as such solvent is at least one species selected
from among hydrocarbons, fatty acidesters, ethers, andnitriles.
Hydrocarbons are most preferred.
The hydrocarbons are not particularly restricted, but
there may be mentioned, for example, aliphatic hydrocarbons,
aromatic hydrocarbons, halogenated hydrocarbons, etc.
Preferred are aliphatic hydrocarbons and aromatic hydrocarbons,
and more preferred are aliphatic hydrocarbons.
The aliphatic hydrocarbons are not particularly
restricted, and may be cyclic or acyclic, or saturated or
unsaturated. However, generally they contain 3 to 20 carbon
atoms, and preferably 5 to 12 carbon atoms.
As specific examples, there may be mentioned, for example,
propane, butane, isobutane, pentane, 2-methylbutane,
cyclopentane, 2-pentene, hexane, 2-methylpentane,
2,2-dimethylbutane, 2,3-dimethylbutane, methylcyclopentane,
cyclohexane, 1-hexene, cyclohexene, heptane, 2-methylhexane,
3-methylhexane, 2,3-dimethylpentane, 2,4-dimethylpentane,
methylcyclohexane,1-heptene, octane, 2,2,3-trimethylpentane,
isooctane, ethylcyclohexane, 1-octene, nonane,
2,2,5-trimethylhexane, 1-nonene, decane, 1-decene, p-menthane,
undecane, dodecane, etc.
Among them, saturated aliphatic hydrocarbons having 5 to
8 carbonatoms are more preferred, andpreferably used are pentane,
2-methylbutane and cyclopentane, which have 5 carbon atoms
(referred to as "pentanes"); hexane, 2-methylpentane,
2,2-dimethylbutane, 2,3-dimethylbutane, methylcyclopentane,
cyclohexane, which have 6 carbon atoms (referred to as
"hexanes"); heptane, 2-methylhexane, 3-methylhexane,
2, 3-dimethylpentane, 2, 4-dimethylpentane, methylcyclohexane,
which have 7 carbon atoms (referred to as "heptanes") ; octane,
2,2,3-trimethylpentane, isooctane, ethylcyclohexane, which
have 8 carbon atoms (referred to as octanes); and a mixture of
these. In particular, the above heptanes are particularly
preferred since they have a tendency to show a very high protection
effect against oxidization, and heptane is most preferred.
The aromatic hydrocarbons are not particularly restricted,
but generally they contain 6 to 20 carbon atoms, preferably 6
to 12 carbon atoms, and more preferably 7 to 10 carbon atoms.
As specific examples, there may be mentioned, for example,
benzene, toluene, xylene, o-xylene, m-xylene, p-xylene,
ethylbenzene, cumene, mesitylene, tetralin, butylbenzene,
p-cymene, cyclohexylbenzene, diethylbenzene, pentylbenzene,
dipentylbenzene, dodecylbenzene, styrene, etc. Preferred are
toluene, xylene, o-xylene, m-xylene, p-xylene, ethylbenzene,
cumene, mesitylene, tetralin, butylbenzene, p-cymene,
cyclohexylbenzene, diethylbenzene and pentylbenzene. More
preferred are toluene, xylene, o-xylene, m-xylene, p-xylene,
cumene and tetralin, and most preferred is cumene.
The halogenated hydrocarbons are not particularly
restricted, and may be cyclic or acyclic, or saturated or
unsaturated. However, acyclic halogenated hydrocarbons are
preferably used. More preferred are chlorinated hydrocarbons
and f luorinated hydrocarbons, and chlorinated hydrocarbons are
still more preferred. Additionally, ones containing 1 to 6
carbon atoms, preferably 1 to 4 carbon atoms, and more preferably
1 to 2 carbon atoms are used.
As specific examples, for example, there may be mentioned
dichloromethane, chloroform, carbon tetrachloride,
1,1-dichloroethane, 1,2-dichloroethane,
1,1,1-trichloroethane, 1,1,2-trichloroethane,
1,1,1,2-tetrachloroethane, 1,1,2,2-tetrachloroethane,
pentachloroethane, hexachloroethane, 1,1-dichloroethylene,
1,2-dichloroethylene, trichloroethylene/ tetrachloroethylene,
1,2-dichloropropane, 1,2, 3-trichloropropane, chlorobenzene,
1,1,1,2-tetrafluoroethane, etc.
Preferred are dichloromethane, chloroform, carbon
tetrachloride, 1,1-dichloroethane, 1,2-dichloroethane,
1,1,1-trichloroethane, 1,1,2-trichloroethane,
1,1-dichloroethylene, 1,2-dichloroethylene,
trichloroethylene, chlorobenzene and
1,1,1,2-tetrafluoroethane. More preferred are
dichloromethane, chloroform, 1,2-dichloroethylene,
trichloroethylene, chlorobenzene and
1,1,1,2-tetrafluoroethane.
The fatty acid esters are not particularly restricted, but
there may be mentioned, for example, propionates, acetates,
formates, etc. Preferred are acetates and formates, and more
preferred are acetates. Ester functional groups thereof are
not particularly restricted, but alkyl esters having 1 to 8 carbon
atoms, aralkyl esters having 1 to 8 carbon atoms are used,
preferred are alkyl esters having 1 to 6 carbon atoms, and more
preferred alkyl esters having 1 to 4 carbon atoms.
As the propionates, there may be mentioned, for example,
methyl propionate, ethyl propionate, butyl propionate,
isopentyl propionate, etc. Preferred is ethyl propionate.
As the acetates, there may be mentioned, for example, methyl
acetate, ethyl acetate, propyl acetate, isopropyl acetate, butyl
acetate, isobutyl acetate, sec-butyl acetate, pentyl acetate,
isopentyl acetate, sec-hexyl acetate, cyclohexyl acetate,
benzyl acetate, etc. Preferred are methyl acetate, ethyl
acetate, propyl acetate, isopropyl acetate, butyl acetate,
isobutyl acetate, sec-butyl acetate, pentyl acetate, isopentyl
acetate, sec-hexyl acetate and cyclohexyl acetate. More
preferred are methyl acetate, ethyl acetate, propyl acetate,
isopropyl acetate, butyl acetate and isobutyl acetate. Most
preferred is ethyl acetate.
As the formates, there may be mentioned, for example, methyl
formate, ethyl formate, propyl formate, isopropyl formate, butyl
formate, isobutyl formate, sec-butyl formate, pentyl formate,
etc. Preferred are methyl formate, ethyl formate, propyl
formate, butyl formate, isobutyl formate and pentyl formate,
and most preferred is ethyl formate.
The ethers are not particularly restricted, and may be
cyclic or acyclic, or saturated or unsaturated. But saturated
ones are preferably used. Generally, ones containing 3 to 20
carbon atoms, and preferably 4 to 12 carbon atoms and more
preferably 4 to 8 carbon atoms are used.
As specific examples, there may be mentioned, for example,
diethyl ether, methyl tert-butyl ether, dipropyl ether,
diisopropyl ether, dibutyl ether, dihexyl ether, ethyl vinyl
ether, butyl vinyl ether, anisol, phenetole, butyl phenyl ether,
methoxytoluene, dioxane, furan, 2-methylfuran, tetrahydrofuran,
tetrahydropyran, ethylene glycol dimethyl ether, ethylene
glycol diethyl ether, ethylene glycol dibutyl ether, ethylene
glycol monomethyl ether, ethylene glycol monoethyl ether,
ethylene glycol monobutyl ether, etc.
Preferred are diethyl ether, methyl tert-butyl ether,
dipropyl ether, diisopropyl ether, dibutyl ether, dihexyl ether,
anisol, phenetole, butyl phenyl ether, methoxytoluene, dioxane,
2-methylfuran, tetrahydrofuran, tetrahydropyran, ethylene
glycol dimethyl ether, ethylene glycol diethyl ether, ethylene
glycol dibutyl ether, ethylene glycol monomethyl ether and
ethylene glycol monoethyl ether. More preferred are diethyl
ether, methyl tert-butyl ether, anisol, dioxane,
tetrahydrofuran, ethylene glycol monomethyl ether and ethylene
glycol monoethyl ether. More preferred are diethyl ether,
methyl tert-butyl ether, anisol, etc., and most preferred is
methyl tert-butyl ether.
The nitriles are not particularly restricted, and may be
cyclic or acyclic, or saturated or unsaturated. However,
saturated ones are preferably used. Generally, ones containing
2 to 20 carbon atoms, preferably 2 to 12 carbon atoms, and more
preferably 2 to 8 carbon atoms are used.
As specific examples, there may be mentioned, for example,
acetonitrile, propiononitrile, malononitrile, butyronitrile,
isobutyronitrile, succinonitrile, valeronitrile,
glutaronitrile, hexanenitrile, heptylcyanide, octylcyanide,
undecanenitrile, dodecanenitrile, tridecanenitrile,
pentadecanenitrile, stearonitrile, chloroacetonitrile,
bromoacetonitrile, chloropropiononitrile,
bromopropiononitrile, methoxyacetonitrile, methyl
cyanoacetate, ethyl cyanoacetate, tolunitrile, benzonitrile,
chlorobenzonitrile, bromobenzonitrile, cyanobenzoic acid,
nitrobenzonitrile, anisonitrile, phthalonitrile,
bromotolunitrile, methyl cyanobenzoate, methoxybenzonitrile,
acetylbenzonitrile, naphthonitrile, biphenylcarbonitrile,
phenylpropiononitrile, phenylbutyronitrile,
methylphenylacetonitrile, diphenylacetonitrile,
naphthylacetonitrile, nitrophenylacetonitrile,
chlorobenzylcyanide, cyclopropanecarbonitrile,
cyclohexanecarbonitrile, cycloheptanecarbonitrile,
phenylcyclohexanecarbonitrile, tolylcyclohexanecarbonitrile,
etc.
Preferred are acetonitrile, propiononitrile,
butyronitrile, isobutyronitrile, succinonitrile,
valeronitrile, chloropropiononitrile, methyl cyanoacetate,
ethyl cyanoacetate, tolunitrile and benzonitrile. More
preferred are acetonitrile, propiononitrile, butyronitrile and
isobutyronitrile, and most preferred is acetonitrile.
In selecting the solvent to be used from among the solvents
mentioned above, such properties as boiling point and viscosity
are preferably taken into consideration; for example, the solvent
should have a boiling point which allows appropriate warming
for increasing the solubility and facilitates a solvent removal
from wet masses by drying and solvent recovery from
crystallization filtrates (about 30 to 150°C at 1 atm) , a melting
i point such that solidification hardly occurs in handling at room
temperature as well as upon cooling to room temperature or below
(not higher than about 20°C, preferably not higher than about
10°C, still more preferably not higher than about 0°C), and a
low viscosity (not higher than about 10 cp at 20°C) . From the
industrial operation viewpoint, a solvent which is scarcely
volatile at ordinary temperature is preferred; for example, one
having a boiling point of not lower than about 80°C is preferred,
and one having a boiling point of not lower than about 90°C is
more preferred.
Among the solvents in the reduction reactionmentioned above,
a solvent having low compatibility with water is particularly
preferred. The solvent in the reduction reaction promotes
purifying and obtaining a reduced coenzyme Q10 efficiently, by
extracting the reducing agent to be described below and/or
impurities from the reducing agent and removing the same. As
such solvents having low compatibility with water, there may
be mentioned, for example, the above-mentioned hydrocarbons and
fatty acid esters among the solvents mentioned above.
Reduced coenzyme Q10, when in a dissolved state, tends to
become more resistant to oxidation as the concentration thereof
increases. Reduced coenzyme Q10 is highly soluble in the solvents
mentioned above and, in this respect, too, the above solvents
are suitable for the protection from oxidation. The
concentration of reduced coenzyme Q10 which is preferred from
the viewpoint of protection thereof from oxidation may vary
depending on the solvent species, among others, hence cannot
be absolutely specif ied. Generally, however, the concentration
of reduced coenzyme Q10 in the above solvents is generally not
lower than 1 w/w%, preferably not lower than 2 w/w%. The upper
limit is not particularly restricted but, from the practical
operability viewpoint, it is 400 w/w% or below, preferably 200
w/w% or below, more preferably 100 w/w% or below, still more
preferably 50 w/w% or below.
Thus, when such a solvent as mentioned above is used, it
is possible to minimize the undesirable oxygen-involving side
reaction via the step of reduction reaction.
Additionally, the reduced coenzyme Q10 used for
crystallization can be also obtained by reducing oily oxidized
coenzyme Q10 in an aqueous solution. In this method, the reduced
coenzyme Q10 can be synthesized without using an organic solvent,
additional operations such as extraction to an organic phase,
concentration, etc. are not required, the operation time can
be shortened and subgeneration of the oxidized coenzyme Q10 can
be minimized.
The reduction reaction can be carried out, in the above
solvent, using, as a reducing agent, a metal hydride compound,
iron (metallic iron or iron in a salt form) , zinc (metallic zinc) ,
dithionous acid or a salt thereof, or an ascorbic acid or
a related compound, for instance.
The metal hydride compound is not particularly restricted
but includes, amongothers, sodiumborohydride, lithium aluminum
hydride, etc. The amount to be used of the metal hydride compound
may vary depending on the species thereof, hence cannot be
absolutely specified. Generally, however, the reduction can
be favorably carried out by using it in an amount of 1 to 3 times
the theoretical hydrogen equivalent.
The reduction using iron or zinc is generally carried out
using an acid. The acid to be used is not particularly restricted
but includes, among others, fatty acids such as acetic acid,
sulfonic acids such as methanesulfonic acid, inorganic acids_
such as hydrochloric acid and sulfuric acid, etc. Inorganic
acids are preferred, and sulfuric acid is more preferred.
The amount of iron to be used is not particularly restricted
but, for example, an amount of about 1/5 by weight or larger
based on the charged weight of oxidized coenzyme Q10 is appropriate
for carrying out the reaction. The upper limit is not
particularly restricted but, from the economical viewpoint, it
is about twice the weight of the above charged weight or lower.
Iron may be used not only in the form of metallic iron but also
in the form of a salt, for example iron(II) sulfate, etc.
The amount of zinc to be used is not particularly restricted
but, for example, an amount of about 1/10 by weight or larger
based on the charged weight of oxidized coenzyme Q10 is appropriate
for carrying out the reaction. The upper limit is not
particularly restricted but, from the economic viewpoint, it
is about twice the weight of the above charged weight or lower.
The dithionous acid or a salt thereof is not
particularly restricted but a salt form of dithionous acid
is generally used. The salt of dithionous acid is not
particularly restricted but includes, as preferred species,
alkali metal salts, alkaline earth metal salts, ammonium salt
and the like. Alkali metal salts such as the lithium salt, sodium
salt, and potassium salt are more preferred, and the sodium salt
is most preferred. The amount to be used of the dithionous
acid or salt is not particularly restricted but it is generally
not smaller than about 1/5 by weight, preferably not smaller
than about 2/5 by weight, and more preferably not smaller than
about 3/5 by weight, based on the charged weight of oxidized
coenzyme, Q10. Larger amounts may be used without causing any
particularly trouble. From the economical viewpoint, however,
the amount to be employed is not larger than about twice the
weight of the above-mentioned charged weight, preferably not
larger than the charged weight. Thus, the reaction can be more
favorably carried out with employing ah amount within the range
of about 2/5 by weight of the above-mentioned charge to a weight
roughly equal to that of the charged weight.
The ascorbic acid or a related compound are not
particularly restricted, and include, for example, not only
ascorbic acid, but also rhamno-ascorbic acid, arabo-ascorbic
acid, gluco-ascorbic acid, fuco-ascorbic acid,
glucohepto-ascorbic acid, xylo-ascorbic acid, galacto-ascorbic
acid, gulo-ascorbic acid, allo-ascorbic acid, erythro-ascorbic
acid, 6-desoxyascorbic acid, and the like ascorbic acid-related
compounds, and may be ester forms or salts of these. Furthermore,
these may be L-form, D-form or racemic form. More specifically,
there may be mentioned, for example, L-ascorbic acid, L-ascorbyl
palmitate, L-ascorbylstearate,D-arabo-ascorbicacid,etc. In
producing the reduced coenzyme Q10, any of the above-mentioned
ascorbic acidor related compounds may be suitably used. However,
the water-soluble ones are suitably used in particular among
the above-mentioned ascorbic acid or related compounds in view
of separatability with the generated reduced coenzyme Q10, etc.
And most preferred is a free form of L-ascorbic acid,
D-arabo-ascorbic acid, and the like in view of the ready
availability, price, etc.
The amount to be used of the ascorbic acid or related
compounds mentioned above is not particularly restricted but
may be an amount effective in converting oxidized coenzyme Q10
to reduced coenzyme Q10. Generally it is not smaller than 1 mole,
preferably not smaller than 1.2 moles, per mole of oxidized
coenzyme Q10. The upper limit is not particularly restricted
but, from the economical viewpoint, it is generally not higher
than 10 moles, preferably not higher than 5 moles, and more
preferably not higher than 3 moles, per mole of the oxidized
coenzyme Q10.
Among the reducing agent species mentioned above, zinc,
dithionous acid or salts'thereof, and ascorbic acidor related
compounds are preferred from the viewpoint of reducing ability,
yield and/or quality, among others. In particular,
dithionous acid or. salts thereof (specifically dithionous
acidsalts) andascorbic acidor related compounds (particularly,
free-form or salt thereof) are preferred from a viewpoint that
they bring the reducing agent or impurities derived from the
reducing agent into the reduced coenzyme Q10 crystal in only
the trace amount or lower.
In carrying out the reduction reaction, an alcohol and/or
water are/is suitably used singly or in combination, as to be
mentioned below. Water is preferred in particular when iron,
zinc, or dithionous acid or a salt thereof is used as the
reducing agent. When a metal hydride compound or an ascorbic
acid or a related compound is used as the reducing agent, an
alcohol can be used in combination. The combined use of water
and an alcohol exhibits the characteristics of both water and
the alcohol and contributes to improvements in reaction rate
and yield, among others.
In the following, a preferred method of reduction is
described in detail.
The reduction using dithionous acid or a salt thereof
is preferably carried out using water in combination, namely
in a mixed solvent system composed of at least one organic solvent
selected from among the above-mentioned hydrocarbons, fatty acid
esters, ethers, and nitriles, with water. On that occasion,
the reaction is preferably carried out generally at a pH of not
higher than 7, preferably at pH 3 to 7, more preferably at pH
3 to 6, from the viewpoint of yield, etc. The pH can be adjusted
using an acid (e.g. an inorganic acid such as hydrochloric acid
or sulfuric acid) or a base (e.g. an alkali metal hydroxide such
as sodium hydroxide).
In the reduction using dithionous acid 1 or a salt thereof,
the amount of water is not particularly restricted but may be
an amount of water such that an appropriate amount of the reducing
agent, namely dithionous acid or a salt thereof, can be
dissolved therein. Thus, for example, it is advisable that the
amount of the dithionous acid or a salt be adjusted generally
to not more than 30 w/w%, and preferably not more than 20 w/w%,
relative to the weight of water. From the productivity viewpoint,
among others, it is advisable that the amount be adjusted
generally to not less than 1 w/w%, preferably not less than 5
w/w%, and more preferably not less than 10 w/w%.
The reduction using the ascorbic acid or a related compound
mentioned above can.be carried out using a solvent especially
highly miscible with water as selected from among the
above-mentioned hydrocarbons, fatty acid esters, ethers, and
nitriles, in particular ethers and nitriles, which are highly
miscible with water, and more specifically tetrahydrofuran,
dioxane, acetonitrile or the like. It is particularly preferred
to use the alcohols and/or ketones to be mentioned below
(preferably alcohols and/or ketones, which are highly miscible
with water (in particular, monohydric or dihydric alcohols
(preferably monohydric ones) having 1 to 5 carbon atoms,
preferably 1 to 4 carbon atoms, more preferably 1 to 3 carbon
atoms, and/or, ketones such as acetone, methyl ethyl ketone or
the like)). Namely, in the reduction using the ascorbic acid
or a related compound, it is preferable to use alcohols and/or
water-soluble organic solvents (the above ethers, nitriles,
ketones and the like, which are highly miscible with water, for
example).
Furthermore, from the viewpoint of reactionpromotion (e.g.
reaction temperature lowering or reaction time shortening) in
the production of reduced coenzyme Q10, it is also possible to
carry out the reduction using the ascorbic acid or a related
compound in the presence of an additive having a reaction
promoting effect, such as a basic substance or a hydrogensulf ite .
The basic compound is not particularly restricted but may
be either an inorganic compound or an organic compound. The
inorganic compound is not particularly restricted but includes,
among others, the hydroxides, carbonates and hydrogencarbonates
of metals (preferably alkali metals, alkaline earth metals, and
the like), ammonia, etc. As typical examples thereof, there
may be mentioned alkali metal hydroxides such as sodium hydroxide,
alkali metal carbonates such as sodium carbonate, alkali metal
hydrogencarbonates such as sodium hydrogencarbonate, and
alkaline earth metal carbonates such as magnesium carbonate.
The organic compound is not particularly restrictedbut includes,
among others, amines such as triethylamine, etc. Among the basic
substances specifically mentioned above, weakly basic
substances (weak bases or weak alkalis) such as the carbonates
and hydrogencarbonates of metals (preferably alkali metals,
alkaline earth metals, etc.), ammonia, and like inorganic
compounds; amines such as triethylamine, and like organic
compounds are preferably used. More preferred are the weakly
basic inorganic compounds mentioned above.
Preferred as the hydrogensulfite are, for example, alkali
metal hydrogensulfites such as sodium hydrogensulfite, etc.
The amount of the additive mentioned above is not
particularly restricted but may be such that the reaction
promoting effect of the additive can be produced to a desired
extent (effective amount). From the economical viewpoint,
however, the amount is generally not more than 20 moles,
preferably not more than 10 moles, more preferably not more than
5 moles, and still more preferably not more than 2 moles, per
mole of the ascorbic acid or a related compound. The lower limit
is not particularly restricted but, generally, it is not less
than 0.01 moles, preferably not less than 0.05 moles, more
preferably not less than 0.1 moles, and still more preferably
not less than 0.2 moles, per mole of the ascorbic acid or a related
compound.
The reduction reaction is preferably carried out under
forced flowing. The power required for stirring to cause such
flowing per unit volume is generally not less than about 0.01
kW/m3, preferably not less than about 0.1 kW/m3, and more
preferably not less than about 0.3 kW/m3. The above forced
flowing is generally causedby the turningof a stirringblade (s) .
The use of a stirring blade (s) is not always necessary if the
above flowing can be otherwise obtained. For example,a method
based on liquid circulation may be utilized.
The reduction temperature may vary depending on the
reducing agent species and/or amount, hence cannot be absolutely
specified. In the reduction using dithionous acid d or a salt
thereof, for instance, the reduction is generally carried out
at 100°C or below, preferably at 80°C or below, more preferably
at 60°C or below. The lower limit is the solidification
temperature of the system. Thus, the reduction can be favorably
carried out generally at about 0 to 100°C, preferably at about
0 to 80°C, more preferably at about 0 to 60°C. In the reduction
using an ascorbic acid or a related compound, the reduction is
carried out generally at 30°C or higher, preferably at 40°C or
higher, more preferably at 50°C or higher. The upper limit is
the boiling point of the system. Thus, the reduction can be
favorably carried out generally at about 30 to 150°C, preferably
about 40 to 120°C, more preferably at about 50 to 100°C. The
reduction of the oily oxidized coenzyme Q10 is generally carried
out at 45°Cormore, preferablyat 48°Cormore, andmorepreferably
at 50°C or more, although it is dependent on the purity, etc.,
then the oily reduced coenzyme Q10 can be obtained.
The reaction concentration is not particularly restricted
but the weight of oxidized coenzyme Q10 relative to the solvent
weight is generally not less than about 1 w/w%, preferably not
less than 3 w/w %, more preferably not less than 10 w/w%, and
still more preferably not less than 15 w/w%. The upper limit
is not particularly restricted but generally is not higher than
about 60 w/w%, preferably not higher than 50 w/w%, more preferably
not higher than 40 w/w%, and still more preferably not higher
than 30 w/w%. Thus, the reaction can be favorably carried out
at a reaction concentration of about 1 to 60 w/w%, preferably
about 3 to 50 w/w%, and more preferably about 10 to 40 w/w%.
The reduction reaction time may vary depending on the
reducing agent species and/or the amount thereof, hence cannot
be absolutely specified. Generally, however, the reaction can
be driven to completion within 48 hours, preferably within 24
hours, more preferably within 10 hours, and still more preferably
within 5 hours.
After the reduction reaction, an organic phase containing
the product reduced coenzyme Q10 or the oily reduced coenzyme
Q10 is recovered (e.g. recovered by separation, extraction,
concentration, etc.), and if necessary (preferably) , the organic
phase is further washed repeatedly using water, brine, or the
like to achieve contaminant elimination and, then, it can be
subjected to crystallization as such or after dissolution to
or substitution by other desirable solvents.
As the other solvents, there may be mentioned, for example,
hydrocarbons, fatty acid esters, ethers, alcohols, fatty acids,
ketones, nitrogen compounds (inclusive of nitriles and amides) ,
sulfur compounds, etc. mentioned above or below.
They were found that, the above-mentioned series of
processes from the reduction reaction to the aftertreatment is
particularly preferably carried out under a deoxygenated
atmosphere, and, in the reduction reaction using dithionous
acid or a salt thereof, in particular, such atmosphere greatly
contributes to an improvement in reduction reaction yield and
a reduction in reducing agent amount. The deoxygenated
atmosphere can be attained by substitution with an inert gas,
pressure reduction, boiling, or a combination of these. It is
preferable to carry out at least the substitution with an inert
gas, namely to use an inert gas atmosphere. As the inert gas,
there may be mentioned, for example, nitrogen gas, helium gas,
argon gas, hydrogen gas, and carbon dioxide gas. Nitrogen gas
is preferred, however.
The crystallization of reduced coenzyme Q10 of the
invention is now described.
The reduced coenzyme Q10 to be subj ected to crystallization
can be obtained in the conventional manner, for example, by
synthesis, fermentation, or extraction from a natural source.
Preferred is the product obtained by reduction of oxidized
coenzyme Q10. More preferred is a solution or oil, of the reduced
coenzyme Q10 obtained by carrying out the reduction reaction in
accordance with the present invention, as described above.
Still more preferred is the one obtained by reducing the oily
oxidized coenzyme Q10 in an aqueous solution using dithionite ,
or the one obtained by reducing the oxidized coenzyme Q10 using
alcohols and/or water-soluble organic solvents.
While the method of crystallization according to the
invention can be applied also to products containing oxidized
coenzyme Q10 in relatively large amounts, the method is
particularly effective in crystallizing high-purity reduced
coenzyme Q10 prepared by the reduction method described above.
In the present invention, the reduced coenzyme Q10 is
crystallized in an aqueous solution. Use of water contributes
to the economical efficiency obviously, and to the safety on
industrial operation and product safety as well. Moreover, the
use of water also contributes to improvement of the slurry
property and the yield of the reduced coenzyme Q10, and can promote
isolation of the reduced coenzyme Q10 efficiently by leaving
a reducing agent used in the reduction reaction or impurities
derived from the reducing agent in amother liquor. Furthermore,
the above-mentioned use of water, or preferably the use of water
containing salts can protect the reduced coenzyme Q10 from
oxidization by molecular oxygen.
The above salts are not particularly restricted but there
may be mentioned, for example, salts constituted from alkaline
metals such as lithium, sodium and potassium; alkaline earth
metals such as magnesium and calcium with halogen atoms such
as fluorine, chlorine and bromine; a residue obtained by removing
a proton from inorganic acids such as sulfuric acid and organic
acids such as formic acid, acetic acid andpropionic acid. Among
these, preferred are inorganic salts, and more preferred are
sodium chloride, potassium chloride, sodium sulfate, etc.
The concentration of the above salts is preferably high,
and it is generally 3 w/w% or more, preferably 5 w/w% or more,
more preferably 10 w/w% or more. Still more preferred is that
the above salts are dissolved in water at saturation or close
to saturation.
As the method of crystallizing the reduced coenzyme Q10
mentioned above, general crystallization methods such as
crystallization by cooling, crystallization by concentration
and crystallization by solvent substitution can be used. It
is preferred to use the crystallization by cooling singly or
in combination. As particularly preferable embodiments, the
following two methods may be mentioned.
(1) A crystallization method which comprises substituting an
organic solvent solution containing the reduced coenzyme Q10
(e.g. reaction solution, extraction solution, etc.) by water
(for increasing the composition ratio of water).
(2) A method of crystallizing the oily reduced coenzyme Q10 in
an aqueous solution.
In the above-mentioned two methods, it is more preferable to
use the crystallization by cooling singly or in combination.
Firstly, the method (1) is explained.
The organic solvent containing the reduced coenzyme Q10
is not particularly restricted but there may be mentioned, for
example, hydrocarbons, fatty acid esters, ethers, alcohols,
fatty acids, ketones, nitrogen compounds (inclusive of nitriles
and amides), sulfur compounds, etc.
As the hydrocarbons, fatty acidesters, ethers andnitriles,
those exemplified as the reaction solvent in the fore-mentioned
explanation about reduction of the oxidized coenzyme Q10 can be
preferably used.
The alcohols are not particularly restricted but may be
cyclic or acyclic, or saturated or unsaturated. Saturated ones
are preferred, however. Generally, they contain 1 to 20 carbon
atoms, preferably 1 to 12 carbon atoms, more preferably 1 to
6 carbon atoms. Still more preferred are monohydric alcohols
containing 1 to 5 carbon atoms, dihydric alcohols containing
2 to 5 carbon atoms, and the trihydric alcohol containing 3 carbon
atoms.
As the monohydric alcohol, there may be mentioned, for
example, methanol, ethanol, 1-propanol, 2-propanol, 1-butanol,
2-butanol, isobutyl alcohol, tert-butyl alcohol, 1-pentanol,
2-pentanol, 3-pentanol, 2-methyl-l-butanol, isopentylalcohol,
tert-pentyl alcohol, 3-methyl-2-butanol, neopentyl alcohol,
1-hexanol, 2-methyl-l-pentanol, 4-methyl-2-pentanol,
2-ethyl-l-butanol, 1-heptanol, 2-heptanol, 3-heptanol,
1-octanol, 2-octanol, 2-ethyl-l-hexanol,1-nonanol, 1-decanol,
1-undecanol, 1-dodecanol, allyl alcohol, propargyl alcohol,
benzyl alcohol, cyclohexanol, 1-methylcyclohexanol,
2-methylcyclohexanol, 3-methylcyclohexanol,
4-methylcyclohexanol, etc.
Preferred are methanol, ethanol, 1-propanol, 2-propanol,
1-butanol, 2-butanol, isobutyl alcohol, tert-butyl alcohol,
1-pentanol, 2-pentanol, 3-pentanol, 2-methyl-l-butanol,
isopentyl alcohol, tert-pentyl alcohol, 3-methyl-2-butanol,
neopentyl alcohol, 1-hexanol, 2-methyl-l-pentanol,
4-methyl-2-pentanol, 2-ethyl-l-butanol and cyclohexanol.
More preferred are methanol, ethanol, 1-propanol, 2-propanol,
1-butanol, 2-butanol, isobutyl alcohol, tert-butyl alcohol,
1-pentanol, 2-pentanol, 3-pentanol, 2-methyl-l-butanol,
isopentyl alcohol, tert-pentyl alcohol, 3-methyl-2-butanol and
neopentyl alcohol. Still more preferred are methanol, ethanol,
1-propanol, 2-propanol,1-butanol, 2-butanol, isobutylalcohol,
2-methyl-l-butanol and isopentyl alcohol. Most preferred is
ethanol.
As the dihydr ic alcohol, there may be mentioned, for example,
1,2-ethanediol, 1,2-propandiol, 1,3-propandiol,
1,2-butanediol, 1,3-butanediol, 1,4-butanediol,
2,3-butanediol, 1,5-pentanediol, etc. Preferred are
1,2-ethanediol, 1,2-propandiol and 1,3-propandiol, and most
preferred is 1,2-ethanediol.
As the trihydric alcohol, glycerol, etc. may be preferably
used, for example.
As fatty acids, there maybe mentioned, for example, formic
acid, acetic acid, propionic acid, etc. Preferred are formic
acid and acetic acid, and most preferred is acetic acid.
The ketones are not particularly restricted, and ones
having 3 to 6 carbon atoms are preferably used in general. As
specific examples, there may be mentioned, for example, acetone,
methyl ethyl ketone, methyl butyl ketone, methyl isobutyl ketone,
etc. Preferred are acetone and methyl ethyl ketone, and most
preferred is acetone.
As the nitrogen compounds other than nitriles, there may
be mentioned, for example, nitromethane, triethylamine,
pyridine, formamide, N-methylformamide, N, N-dimethylformamide,
N,N-dimethylacetoamide, N-methylpyrrolidone, etc.
As the sulfur compounds, there may be mentioned, for
example, dimethyl sulfoxide, sulfolane, etc.
In selecting the solvent to be used from among the solvents
mentioned above, such properties as boiling point and viscosity
(for example, the solvent should have a boiling point which allows
appropriate warming for increasing the solubility and
facilitates a solvent removal from wet masses by drying and
solvent recovery from crystallization filtrates (about 30 to
150°C at 1 atm) , a melting point such that solidification hardly
occurs in handling at room temperature as well as upon cooling
to room temperature or below (not higher than about 20°C,
preferably not higher than about 10°C, still more preferably
not higher than about 0°C) , and a low viscosity (not higher than
about 10 cp at 20°C) ) are preferably taken into consideration.
From the industrial operation viewpoint, a solvent which is
scarcely volatile at ordinary temperature is preferred.
Among the above solvents, in particular, hydrocarbons,
fatty acid esters, ethers and nitriles can be preferably used
in view of the oxidization protection of the reduced coenzyme
Q10mentioned above. Additionally, in view of obtaining the high
yield while preferably decreasing the solubility of the reduced
coenzyme Q10, alcohols, fattyacids, ethers, ketones and nitriles
can be preferably used. From a viewpoint of an industrial
applicability, for example, hydrocarbons and alcohols can be
preferably used.
In the above method (1) , as a means for substituting the
organic solvent solution containing the reduced coenzyme Q10
by water, there maybe mentioned, for example, a method comprising
increasing the water ratio while concentrating and removing the
organic solvent. Furthermore, if necessary, operations such
as cooling and seed crystal addition, as to be described below,
can be combinedly used properly.
Next, the method (2), i.e., the method of crystallizing
the oily reduced coenzyme Q10 in an aqueous solution is explained.
According to this method, the reduced coenzyme Q10 crystal having
a large particle diameter can be obtained, and filterability
can be remarkably improved.
The crystallization may be carried out by adding water to
the oily reduced coenzyme Q10, for example, or on the contrary,
by adding the oily reduced coenzyme Q10 to water. Moreover, the
crystallization may be also carried out by cooling a mixture
of the oily reduced coenzyme Q10 and water. More preferred is
a method comprising removing an organic solvent from a mixed
solvent solution consisting of an organic solvent containing
the reduced coenzyme Q10 and water at a temperature not lower
than the melting point of the reduced coenzyme Q10 or of a
concentrate comprising the reduced coenzyme Q10 as a main
component, to obtain oil in the system, and cooling to crystallize
the oil.
The above-mentioned temperature not lower than the melting
point is generally 45°C or more, preferably 48°C or more, and
more preferably 50°C or more although it is dependent on the
purity, etc. of the reduced coenzyme Q10. The upper limit is
not particularly restricted but generally 100°C or less is
preferred, 80°C or less is more preferred, and 60°C or less is
still more preferred.
In the crystallization method of the present invention as
described above, the crystallization temperature of the reduced
coenzyme Q10 (cooling temperature at the time of crystallization)
is generally 48°C or less, preferably 45°C or less, more
preferably 40°C or less, and still more preferably 30°C or less
in view of the yield, etc., although it is difficult to uniformly
define since it is dependent on the purity of the reduced coenzyme
Q10. The lower limit is a solidification temperature of the
system. Therefore, the crystallization can be especially
preferably carried out at the crystallization temperature of
about 0 to 30°C.
It is preferable to control the amount of crystallization
per unit time in crystallizing, i.e. the rate of crystallization.
The preferable amount of crystallization per unit time is, for
example, not higher than the rate of crystallization which causes
crystallization of about 50%, per unit time, of the whole amount
of crystals to be obtained (i.e. at most 50%/hour), preferably
not higher than the rate of crystallization which causes
crystallization of about 25%, per unit time, of the whole amount
of crystals to be obtained (i.e. at most 25%/hour).
The rate of cooling in the crystallization by cooling is
generally not higher than about 40°C/hour, and preferably not
higher than about 20°C/hour.
The crystallization is preferably carried out under forced
flowing in order to prevent the state of supersaturation from
occurring and thereby allowing the nucleation and crystal growth
to proceed smoothly, in order to obtain crystals with uniform
particle diamater and, furthermore, from the viewpoint of
obtaining high-quality products. The flowing is generally
brought about by a stirring power per unit volume of not weaker
than about 0.01 kW/m3, preferably not weaker than about 0.1 kW/m3,
and more preferably not weaker than about 0.3 kW/m3. The forced
flowing is generally provided by the turning of a stirring
blade (s) . However, the use of a stirring blade (s) is not always
necessary if the above flowing can be otherwise obtained. For
example, it is possible to utilize a method based on liquid
circulation.
In carrying out the crystallization, seed crystals are
preferably added so that the state of supersaturation may be
prevented from occurring and the nucleation and crystal growth
may be allowed to proceed smoothly.
The crystallization concentration, when expressed in terms
of the weight of reduced coenzyme Q10 relative to the total weight
of the crystallization solvent at the time of completion of
crystallization, it is not higher than about 15 w/w%, preferably
not higher than about 13 w/w%, more preferably not higher than
10 w/w%. From the productivity viewpoint, the lower limit to
the crystallization concentration is generally not lower than
about 1 w/w%, preferably not lower than about 2 w/w%.
The thus-obtained crystals of reduced coenzyme Q10 can be
recovered as a wet product, for example, by such a solid-liquid
separation technique as centrifugation, pressure filtration,
or vacuum filtration, if necessary followed by cake washing.
They can be recovered also as a dry product by further charging
the wet product in a reduced pressure drier (vacuum drier)
internally purged with an inert gas and drying the same under
reduced pressure. The recovery in a dry form is preferred.
The crystallization of the invention, when it is carried
out in a deoxygenated atmosphere, can increase protective effect
against oxidation. The deoxygenated atmosphere can be attained
by inert gas substitution, pressure reduction, boiling, or a
combination of these. It is preferable to carry out at least
the substitution with an inert gas, namely to use an inert gas
atmosphere. As the inert gas, there maybe mentioned, for example,
nitrogen gas, helium gas, argon gas, hydrogen gas, and carbon
dioxide gas. Nitrogen gas is preferred, however.
In accordance with the present invention, high-quality
reduced coenzyme Q10 can be obtained with excellent workability
and economical efficiency. The crystals of reduced coenzyme
Q10 as obtained in accordance with the present invention are
of very high quality and can be expected to have a reduced coenzyme
Q10/oxidized coenzyme Q10 weight ratio of not lower than 96/4,
preferably not lower than 98/2, more preferably not lower than
99/1.
BEST MODE FOR CARRYING OUT THE INVENTION
The following examples illustrate the present invention
in further detail. These examples are, however, by no means
limitative of the scope of thepresent invention. In the examples,
the purity of reduced coenzyme Q10 and the reduced coenzyme
Q10/oxidized coenzyme Q10 weight ratio were determined by the
HPLC analysis specif ied below. The reduced coenzyme Q10 purity
values as determined, however, are by no means indicative of
the limit purity value attainable in accordance with the present
invention. Likewise, the reduced coenzyme Q10/oxidized
coenzyme Q10 weight ratio values obtained never indicate the
upper limit to that ratio.
(HPLC conditions)
Column: SYMMETRY C18 (product of Waters), 250 mm (in length),
4.6 mm (in inside diameter); mobile phase: C2H5OH:CH3OH = 4:3
(v/v); detection wavelength: 210 nm; flow rate: 1 ml/min;
retention time of reduced coenzyme Q10: 9.1 min; retention time
of oxidized coenzyme Q10: 13.3 min.
(Example 1)
Oxidized coenzyme Q10 (100 g; purity 99.4%) was melted
with stirring at 48°C. While stirring (power required for
stirring: 0.3kW/m3), an aqueous solution prepared by dissolving
100 gof sodiumdithionite (purity: atleast 75%), as the reducing
agent, in 1000 ml of water was gradually added to the oil and
the reduction reaction was carried out at 48°C and at pH 4 to
6. The aqueous phase was removed from the reaction mixture
containing the oil, and the oil was washed 6 times with 1000
g of deaerated saturated brine at 48°C. After that, by removing
the aqueous phase, oily reduced coenzyme Q10 was obtained. 1500
g pf deaerated water, at 48°C was added to this oil, and while
stirring (power required for stirring: 0.3 kW/m3) , the mixture
was cooled to 2°C to obtain white slurry (fluidity of the slurry
was good). All the above operations were carried out under a
nitrogen atmosphere. The obtained slurry was filtered under
reduced pressure, the wet crystal was washed with cold ethanol,
cold water and cold ethanol in this order (the temperature of
the cold solvent used for washing was 2°C) , and further the wet
crystal was dried under reduced pressure (20 to 40°C, 1 to 30
mmHg), 97 g of dry white crystal was obtained (isolated product
yield: 97 mol %) . The weight ratio of the reduced coenzyme
Q10/oxidized coenzyme Q10 of the obtained crystal was 99.4/0.6,
and the purity of the reduced coenzyme Q10 was 99.2%.
(Example 2)
The oxidized coenzyme Q10 (100 g; purity 99.4%) was dissolved
in 1000 g of heptane at 25°C. While stirring (power required
for stirring: 0.3 kW/m3) , an aqueous solution dissolving 100
g of sodiumdithionite (purity 75% or more) , as a reducing agent,
in 1000 ml of water was gradually added and a reduction reaction
was carried out at 25°C and at pH 4 to 6. 2 hours later, an
aqueous phase was removed from the reaction solution, and a
heptane phase was washed 6 times with 1000 g of deaerated saturated
brine. 1000 g of water was added to this heptane phase, and
while stirring (power required for stirring: 0.3kW/m3), heptane
was removed by reducing pressure at 3 0°C to obtain white slurry.
This slurry had good fluidity and was capable to be easily brushed
away from the crystallization container. All the above
operations were carried out under a nitrogen atmosphere. The
obtained slurry was filtered under reduced pressure, and the
wet crystal was washed with cold ethanol, cold water and cold
ethanol in this order (the temperature of the cold solvent used
for washing was 2°C) . The wet crystal was further dried under
reduced pressure (20 to 40°C, 1 to 30 mmHg), and a dry white
crystal was obtained ("isolated product yield: 97 mol %) . The
weight ratio of the reduced coenzyme Q10/oxidized coenzyme Q10
of the obtained crystal was 99.5 / 0.5, and the purity of the
reduced coenzyme Q10 was 99.2%.
(Example 3)
To 1000 g of ethanol, 100 g of the oxidized coenzyme Q10
(purity 99.4%), 60 g of L-ascorbic acid and 30 g of sodium
hydrogencarbonate were added, and the mixture was stirred at
78°C and subjected to a reduction reaction. 3 hours later, the
mixture was cooled to 50°C, and 1000 g of heptane and 1000 g
of deaerated water were added while holding said
temperature. After the mixture was cooled to 25°C, an aqueous
phase was removed, and further washed 6 times with 1000 g of
saturatedbrine to remove the aqueous phase. Heptane was removed
from this heptane solution at 48°C to obtain the oily reduced
coenzyme Q10. To this oil, 1500 g of deaerated water at 48°C
was added, and while stirring (power required for stirring: 0.3
kW/m3), the mixture was cooled to 2°C to obtain white slurry
(fluidity of the slurry was good as in Example 1) . All the above
operations were carried out under the nitrogen atmosphere. The
obtained slurry was filtered under reduced pressure, and the
wet crystal was washed with cold ethanol, cold water and cold
ethanol in this order (the temperature of the cold solvent used
for washing was 2°C) . The wet crystal was further dried under
reduced pressure (20 to 40°C, 1 to 30 mmHg) to obtain 97 g of
dry white crystal (isolated product yield: 97 mol%). The weight
ratio of the reduced coenzyme Q10/oxidized coenzyme Q10 of the
obtained crystal was 99.4/0.6, and the purity of the reduced
coenzyme Q10 was 99.2%.
(Comparative Example 1)
A heptane phase of the reduced coenzyme Q10 after being
washed with deaerated saturated brine was obtained in the same
manner as Example 2. While stirring this heptane phase (power
required for stirring: 0.3 kW/m3) , the mixture was cooled to
2°C to obtain white slurry. This slurry was poor in fluidity,
and more difficultly brushed away from the crystallization
container as compared with Example 1. All the above operations
were carried out under the nitrogen atmosphere. The obtained
slurry was filtered under reduced pressure, the wet crystal was
washed with cold ethanol, cold water and cold ethanol in this
order (the temperature of the cold solvent used for washing was
2°C) . The wet crystal was further dried under reduced pressure
(20 to 40°C, 1 to 30 mmHg) to obtain 93 g of dry white crystal
(isolated product yield: 93 mol %) . The weight ratio of the
reduced coenzyme Q10/oxidized coenzyme Q10 of the obtained crystal
was 99.6/0.4, and the purity of the reduced coenzyme Q10 was
99.3%.
(Reference Example 1)
One gram of the reduced coenzyme Q10 (the weight ratio of
the reduced coenzyme Q10/oxidized coenzyme Q10 is 99.6/0.4) was
dissolved in 20 g of various solvents shown in Table 1 at 25°C. In
the atmosphere, the weight ratio of the reduced coenzyme
Q10/oxidized coenzyme Q10 in the solution after stirring for 24
hours at 25°C, and the result is shown in Table 1.
(Reference Example 2)
One gram of the reduced coenzyme Q10 (the weight ratio of
the reduced coenzyme Q10/oxidized coenzyme Q10 is 99.6/0.4) was
dissolvedin 100 gof various solvents shown in Table 2 at 35°C. In
the atmosphere, the weight ratio of the reduced coenzyme
Q10/oxidized coenzyme Q10 in the solution after stirring for 24
hours at 35°C was measured, and the result is shown in Table
2.
INDUSTRIAL APPLICABILITY
The invention, which has the constitution described above,
is a method excellent in workability and economical efficiency
on the industrial scale and can give high-quality reduced
coenzyme Q10 in a convenient and efficient manner.
WE CLAIM:
1. A method of crystallizing a reduced coenzyme Q10
which comprises crystallizating the reduced coenzyme Q10 in an aqueous
solution.
2. The method as claimed in claim 1,
wherein the reduced coenzyme Q10 is crystallized by substituting an organic
solvent such as herein described containing the reduced coenzyme Q10 by
water.
3. The method as claimed in claim 1,
wherein oily reduced coenzyme Q10 is crystallized in an aqueous solution.
4. The method as claimed in claim 3,
wherein the crystallization is carried out by cooling a mixture of the oily
reduced coenzyme Q10 and water.
5. The method as claimed in claim 4,
wherein an organic solvent such as herein described is removed from a solution
of a mixed solvent comprising an organic solvent containing the reduced
coenzyme Q10 and water at a temperature not lower than the melting point of
the reduced coenzyme Q10 or of a concentrate comprising the reduced
coenzyme Q10 as a main component, to obtain oily reduced coenzyme Q10 in
the system, and cooled to crystallize the oily reduced coenzyme Q10
6. The method as claimed in any one of claims 1 to 5,
wherein the crystallization by cooling is used singly or in combination.
7. The method as claimed in any one of claims 1 to 6,
wherein the crystallization temperature is 48°C or less.
8. The method as claimed in any one of claims 1 to 7,
wherein the crystallization is carried out under forced flowing with a stirring
power per unit volume of not weaker than 0.01 kw/m3 .
9. The method as claimed in any one of claims 1 to 8,
wherein the seed crystal is added in the crystallization.
10. The method as claimed in any one of claims 1 to 9,
in which the rate of crystallization is not higher than the rate of crystallization
which causes crystallization of 50%, per unit time, of the whole amount of
crystals.
11. The method as claimed in any one of claims 1 to 10,
wherein the crystallization is carried out under a deoxygenated atmosphere.
12. The method as claimed in any one of claims 1 to 11,
wherein the reduced coenzyme Q10 used for the crystallization is obtained by
reducing an oxidized coenzyme Q10.
13. The method as claimed in any one of claims 1 to 12,
wherein the reduced coenzyme Q10 used for the crystallization is obtained by
reducing oily oxidized coenzyme Q10 in an aqueous solution using
dithionites.
14. The method as claimed in any one of claims 1 to 12,
wherein the reduced coenzyme Q10 used for the crystallization is obtained by
reducing the oxidized coenzyme Q10 using alcohols and/or water-soluble
organic solvents.
15. The method as claimed in any one of claims 1 to 14,
wherein a reducing agent used in the reduction reaction or impurities derived
from the reducing agent is removed to a mother liquor in crystallizing the
reduced coenzyme Q10.
The present invention provides the excellent
crystallization method suitable for production on the industrial
scale for obtaining the reduced coenzyme Q10 crystal.
The high-quality reduced coenzyme Q10 can be obtained in
improving the slurry property and/or the crystallization yield
by the method of present invention in which reduced coenzyme
Q10 is crystallized in an aqueous solution, in particular, the
method of crystallizing the reduced coenzyme Q10 by substituting
an organic solvent solution containing the reduced coenzyme Q10
by water, the method of crystallizing the oily reduced coenzyme
Q10 in an aqueous solution, or the like.

Documents:

55-kolnp-2004-granted-abstract.pdf

55-kolnp-2004-granted-assignment.pdf

55-kolnp-2004-granted-claims.pdf

55-kolnp-2004-granted-correspondence.pdf

55-kolnp-2004-granted-description (complete).pdf

55-kolnp-2004-granted-examination report.pdf

55-kolnp-2004-granted-form 1.pdf

55-kolnp-2004-granted-form 13.pdf

55-kolnp-2004-granted-form 18.pdf

55-kolnp-2004-granted-form 2.pdf

55-kolnp-2004-granted-form 3.pdf

55-kolnp-2004-granted-form 5.pdf

55-kolnp-2004-granted-gpa.pdf

55-kolnp-2004-granted-reply to examination report.pdf

55-kolnp-2004-granted-specification.pdf

55-kolnp-2004-granted-translated copy of priority document.pdf

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Patent Number

223876

Indian Patent Application Number

55/KOLNP/2004

PG Journal Number

39/2008

Publication Date

26-Sep-2008

Grant Date

23-Sep-2008

Date of Filing

16-Jan-2004

Name of Patentee

KANEKA CORPORATION

Applicant Address

2-4, NAKANOSHIMA 3-CHOME, KITA-KU, OSAKA-SHI, OSAKA

Inventors:

#	Inventor's Name	Inventor's Address
1	KITAMURA SHIRO	10-36-601, AIOCHO 1-CHOME, AKASHI-SHI, HYOGO 673-0882
2	UEDA TAKAHIRO	31-17-2018, SHIOYACHO 6-CHOME, TARUMI-KU, KOBE-SHI HYOGO 655-0872
3	UEDA YASUYOSHI	140-15, WAKU, ABOSHI-KU, HIMEJI-SHI, HYOGO 671-1227

PCT International Classification Number

C07C 43/23

PCT International Application Number

PCT/JP02/07146

PCT International Filing date

2002-07-15

PCT Conventions:

#	PCT Application Number	Date of Convention	Priority Country
1	2001-214477	2001-07-13	Japan
2	2002-114874	2002-04-17	Japan