CS209571B1 - Process for preparing (U-14c) D-glucosamine - Google Patents

Abstract

The invention relates to a process for the preparation of (U-14C) labeled D-glucosamine. The essence of the invention lies in the fact that whole cells of the algae Chlorella cultured in a carbon dioxide environment labeled with the L^C radionuclide are treated with a solution of the enzymes chitinase and lysozyme buffered to pH 4.5 to 6, while the released (U-14C) D-glucosamine is obtained from the solution by paper chromatography. The invention is of importance for the preparation of (U-14C) labeled glucosamine, which is interesting from the point of view of carbohydrate chemistry and for bacteriological and clinical practice.

Description

(54) Preparation of (U-O) D-glucosamine

The present invention relates to a process for preparing labeled (U-C) D-glucosamine,

SUMMARY OF THE INVENTION The principle of the present invention is to treat whole cells of the Chlorella algae cultured in the environment of ^L 2 C-labeled carbon dioxide with a radionuclide solution of chitinase enzymes and lysozyme buffered to pH 4.5-6. chroinatografiou.

The invention is of importance for the preparation of (U-C) labeled glncosamine of interest in the field of carbohydrate chemistry and for bacteriological and clinical practice.

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The present invention relates to a method for the preparation of labeled (U - * C) D-glucosamine by hydrolytic digestion of cell shades of Chlorella by the action of the enzyme chitinase (EC 3.2.1.14, poles- / 5-1,4- (2-acetamido-2-deoxy)). - D-glucoside (glycanohydrolase) and lysozyme (EC 3.2.1.17, N-acetylmuramy mucopeptide ± hydrolase) after previous cultivation of Chlorella algae in a 4 C-labeled carbon dioxide medium.

Among the biochemical methods for the preparation of (U- ¹⁴ C) labeled mono- and oligosaccharides, the preparation of (U- ¹⁴ C) D-glucose is described. 882), (U-D-gluconic acid (No. AO No. 188 626)).

Chlorella, cultivated in a labeled carbon dioxide environment. However, cell walls of microorganisms are also a suitable source of mono- and oligosaccharides, and the mono- and oligosaccharides can be released from them by the action of suitable enzymes without substantially disrupting the rest of the cell.

SUMMARY OF THE INVENTION The invention provides that whole cells, algae, Chlorella cultured in a radionuclide-labeled carbon dioxide environment are treated with a solution of chitinase enzymes and lysozyme buffered to a pH of 4.5 to 6, releasing (U-C) D-glucosamine. The solution is obtained by paper chromatography.

An advantage of the proposed process for the preparation of (D-C) labeled D-glucosamine is that it uses unprocessed material (waste); allows processing of the remaining cellular material (intracellular content, nucleic acids, etc.) without changing the established technological procedure; The whole preparation process is in technological! Conversion is very simple and the enzymes required for the hydrolytic cleavage of the cell wall of the Chlorella alga are commercially available, both isolated and mixed. . ,.

Example 1

To 0.5 ml of a 20 kbq Chlorella alga suspension was added 0.5 ml of a 50 mM citrate phosphate solution (ph - 4.5) with dissolved chitinase (1 mg, 0.55 U) with lysozyme (5 mg, 3, 7 U) and the reaction mixture is allowed to shake for 6 hours at 30 ° C. After hydrolysis reaction e is carried out, the mixture is loaded onto Chromatographic Paper 1 and the chromatographic separation is carried out in acetone-n-butanol-water, 16 hours, zone. corresponding to the mobility of D-glucosamine is excised and eluted with a 50% aqueous solution of ethanol. The radiochemical yield of D-glucosamine was 0.63 kBq (3%) ·.

Example 2

The procedure was as in Example 1 except that the hydrolytic cleavage of the cell wall was transferred with 0.5 ml of a diluted enzyme system (1: 5) of a snail (IJelix pomatia) prepared from the digestive tract, with 50 mM citrate-phosphate buffer pH 6, 0. After a reaction time of 8 hours at 30 ° C, the reaction mixture is treated as described in Example 1. The radiochemical yield of D-glucosamine is as in Example 1.

Example 3

The procedure was as in Example 1, except that the reaction was carried out in distilled water. To obtain the same yields as in Example 1, the reaction must be run for 24 hours.

The invention is of importance for the preparation of (U-C) labeled glucosamine, of interest for ihemia and carbohydrate biochemistry, and for bacteriological and clinical practice.

Claims (1)

PB E D MET EXPLOITED A process for the preparation of (U-D-glucosamine) characterized in that the whole cell algae Chlorella cultured in a radionuclide labeled carbon dioxide medium is treated with a solution of chitanase enzymes and lysozyme buffered to a pH of 4.5 to 7, whereby the liberated (U-D-glucosamine) is recovered from the solution by paper chromatography.