Chemistry: molecular biology and microbiology – Micro-organism – tissue cell culture or enzyme using process... – Preparing compound containing saccharide radical
Reexamination Certificate
2001-04-25
2002-07-16
Lilling, Herbert J. (Department: 1651)
Chemistry: molecular biology and microbiology
Micro-organism, tissue cell culture or enzyme using process...
Preparing compound containing saccharide radical
C435S074000, C435S105000
Reexamination Certificate
active
06420142
ABSTRACT:
The invention relates to a process for the enzymatic cleavage of rutinosides to obtain rhamnose and/or the corresponding glucopyranosides, the reaction being carried out in the presence of a solvent mixture of water and one or more organic solvents.
In the context of the present invention, rutinosides are designated as those compounds which contain a sugar-free constituent, to which a radical of the
is bonded via a glycosidic bond. For example, the rutinosides are flavonoids having the bisglycosidic unit shown in formula I. Rhamnose and/or the corresponding glucopyranosides are produced from the rutinosides by the process according to the invention. The glucopyranosides are derived from the rutinosides in that, instead of the radical of the formula (I), they contain a radical of the formula (I*)
bonded to the sugar-free constituent. For example, both rhamnose and isoquercetin can be obtained from rutin by the process according to the invention.
Rhamnose is a monosaccharide which is of widespread occurrence in nature, but usually only in small amounts. An important source of rhamnose is, for example, the glycosidic radicals of natural flavonoids such as rutin, from which the rhamnose can be obtained by glycoside cleavage. Rhamnose, for example, plays an important role as a starting substance for the preparation of synthetic aromatic substances such as furaneol.
Isoquercetin is a monoglycosidated flavonoid of the following structural formula (II)
Flavonoids. (lat. flavus=yellow), which are widespread colorants in plants, are designated as being, for example, glycosides of flavones, to which the parent structure of flavone (2-phenyl-4H-1-benzopyran-4-one) is common.
The sugar-free constituent of the flavonoids is the so-called aglycone. Isoquercetin is, for example, a glycoside of the aglycone quercetin (2-(3,4-dihydroxyphenyl)-3,5,7-trihydroxy-4H-1-benzopyran-4-one), which differs from flavone by the presence of five hydroxyl groups. In isoquercetin, the carbohydrate radical glucose is bonded to the hydroxyl group in position 3 of the quercetin. Isoquercetin is designated, for example, as quercetin 3-O-&bgr;-D-glucopyranoside or 2-(3,4-dihydroxyphenyl)-3-(&bgr;-D-glucopyranosyloxy)-5,7-dihydroxy-4H-l-benzopyran-4-one. However, it is also known, for example, under the name hirsutrin.
Flavonoids and flavonoid mixtures are used, for example, in the foodstuffs and cosmetics industries and are increasingly gaining importance there. Particularly monoglycosidated flavonoids such as isoquercetin are distinguished by a good absorption capacity in the human body.
An example of a naturally occurring flavonoid having a bisglycosidic unit is rutin, which has the following structural formula (III):
Rutin, like isoquercetin, is likewise a glycoside of the aglycone quercetin where the carbohydrate radical rutinose is bonded to the hydroxyl group in position 3 of the quercetin. The carbohydrate radical in rutin consists of a glucose unit linked in the 1 and 6 positions and a terminally bonded rhamnose or 6-deoxymannose unit. Rutin is designated, for example, as quercetin 3-O-&bgr;-D-rutinoside or 2-(3,4-dihydroxy-phenyl)3-{[6-O-(6-deoxy-&agr;-mannopyranosyl)-&bgr;-D-gluco-pyranosyl]oxy}-5,7-dihydroxy-4H-1-benzopyran-4-one. However, it is also known, for-example, under the names sophorin, birutan, rutabion, tarutin, phytomelin, melin or rutoside.
Rutin, with three molecules of water of crystallization, forms pale yellow to greenish needles. Anhydrous rutin has the properties of a weak acid, becomes brown at 1250° C. and decomposes at 214-2150° C. Rutin, which occurs in many plant species—frequently as an associate of vitamin C —, e.g. in citrus species, in yellow pansies, forsythia and acacia species, various Solanum and Nicotiana species, capers, lime blossom, St. John's wort, tee etc. was isolated from the common rue (Ruta graveolens) in 1842. Rutin can also be obtained from the leaves of buckwheat and of the east-asiatic pagoda tree Wei-Fa (Sophora japonica, Farbaceae), which contains 13-27% of rutin.
For the abovementioned reasons, it is desirable to prepare both rhamnose and monoglycosidated flavonoids from natural raw materials, for example from flavonoids having a bisglycosidic unit. In this connection, for example, the cleavage of rutinosides to rhamnose and the corresponding glucopyranosides is of interest.
Enzymatically catalyzed preparations of rhamnose are described in the literature. For example, EP 0 317 033 describes a process for the preparation of L-rhamnose, with the rhamnosidic bonding of glycosides which contain rhamnose bonded in the terminal position being achieved by enzymatic hydrolysis. However, cleavages of this type carried out in aqueous media of glycosides having a bisglycosidic structure of the carbohydrate radical usually proceed with low selectivity. For example, on account of the bisglycosidic structure of the carbohydrate radical in rutin, a mixture of the two monosaccharides glucose and rhamnose usually results. Moreover, high proportions of the aglycone quercetin and other undesired by-products usually occur.
In addition, enzymatically catalyzed cleavages of rutin are also described, for example, in JP 01213293. However, reactions of this type carried out in aqueous media usually likewise proceed with low selectivity.
The object was therefore to develop a process for the enzymatic cleavage of rutinosides to obtain rhamnose and/or the corresponding glucopyranosides which avoids or at least diminishes the disadvantages of the known processes and in particular makes possible a preparation of rhamnose and the glucopyranosides which is as selective as possible, so that these products can be prepared in high yield.
Surprisingly, it has now been found that this object is achieved if the process for the enzymatic cleavage of rutinosides to obtain rhamnose and/or the corresponding glucopyranosides is carried out such that the reaction takes place in the presence of a solvent mixture of water and one or more organic solvents.
The process according to the invention is distinguished in particular in that the cleavage of rutinosides to rhamnose and the corresponding glucopyranosides takes place with high selectivity. Rhamnose and the glucopyranosides are preferably obtained by suitable work-up after the process according to the invention. Furthermore, however, either only rhamnose or only the glucopyranosides can also be obtained by suitable work-up after the process according to the invention.
The present invention makes available an advantageous process for the enzymatic cleavage of rutinosides to obtain rhamnose and/or the corresponding gluco-pyranosides. According to this process, the rutinoside is contacted with a catalytic amount of an enzyme in a solvent mixture of water and one or more organic solvents. Preferably, the reaction is carried out with thorough mixing, e.g. by stirring.
The reaction is preferably carried out under a nitrogen atmosphere.
Suitable rutinosides for the process according to the invention are, for example, rutinosides which, as a sugar-free constituent or aglycone, contain a 2-phenyl-4H-1-benzopyran-4-one parent structure which carries a radical of the formula (I) in position 3 and whose phenyl groups, apart from position 3, can also be mono- or polysubstituted by —OH or —O—(CH
2
)
n
—H, where n is 1 to 8.
n is preferably 1.
The substitution of the 2-phenyl-4H-1-benzopyran-4-one parent structure by —OH and/or —O—(CH
2
)
n
—H preferably occurs in positions 5, 7, 3′ and/or 4′.
Particularly preferred rutinosides correspond to formula (IV)
in which R is H (kaempferol rutinoside), OH (rutin) or OCH
3
(isorhamnetin rutinoside). Rhamnose and kaempferol glucoside can be obtained from kaempferol rutinoside by the process according to the invention, rhamnose and isoquercetin from rutin, and rhamnose and isorhamnetin glucoside from isorhamnetin rutinoside. The rutinoside rutin is particularly preferably used.
The invention also relates to the use of kaempferol glucoside, isoquercetin and/or isorhamnetin
Buchholz Herwig
Koppe Thomas
Schleehahn Michael
Lilling Herbert J.
Merck Patent Gesellschaft
Millen White Zelano & Branigan P.C.
LandOfFree
Method for enzymatic splitting of rutinosides does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with Method for enzymatic splitting of rutinosides, we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Method for enzymatic splitting of rutinosides will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-2911417