CS209572B1 - A method of preparing (U-14C) D-galactose - Google Patents

Abstract

Invention of a method for the preparation of (U - ^4C) labeled D-galactose. The essence of the invention is that whole cells of Chlorella algae cultivated in a carbon dioxide environment labeled with a radionuclide are treated with a solution of the enzyme /J-galactosidase, buffered at pH 4.5 to 6, while (U-14C) D-galacto'se is isolated by chromatography on paper. The invention is important for the preparation of (U - 14C) labeled sugar D-galactose obtained from the hl'adiek ohemia and biochemistry of carbohydrates and for bacteriological and clinical practice.

Description

The present invention relates to a process for the preparation (U- ¹⁴ C) of D-galactose by hydrolytic non-digestion of Chlorella algal cell walls by the action of β-galactosidase enzyme (EG 3.2.1.23, β-D-galactoside galactohydrolase after previous cultivation of Chlorella algae) carbon dioxide.

Among the biochemical methods for the preparation of (U-C) labeled mono- and oligosaccharides, the preparation of (U- ¹⁴ C) D-glucose (U. AO 189 471), (U- ¹⁴ C) meltose (U. AO 187 882) is described. ), (U - f C) D-gluconic acid (No. AO No. 188 626) and (U - ^ C)--D-glucopyranosyl phosphate Xcs. AO č. 194 587), wherein the basic starting material for the preparation of these saccharides was intracellular <RTI ID = 0.0> (1,4) </RTI> glucan (U-C) isolated from Algae Chlorella cultivated in labeled carbon dioxide. However, a suitable source of mono- and oligosaccharides are the cell walls of the microorganisms, and the mono- and oligosaccharides can be released therefrom by appropriate enzyme systems without substantially disrupting the rest of the cell. .

SUMMARY OF THE INVENTION The whole of the cells of the Chlorella algae cultured in a radionuclide-labeled carbon dioxide environment are treated with a solution of the enzyme β-galactosidase (β-D-galactoside galacto hydrolase, EC 3.2.1.23), buffered to a pH of 4.5 to 6, wherein the (U -? - C) D-galactose is isolated by paper chromatography.

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The advantage of the proposed method of preparation (U - C) of D-galectose is that it: utilizes hitherto unprocessed wall material (waste), allows the processing of the remaining cellular material (intracellular content, nucleic acids, etc.) without changing the technological procedure used The preparation is very simple in technical conversion and all the enzymes required for the hydrolytic cleavage of the cell wall of the Chlorella algae 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 β-galactosidase (3 mg, 4.2 U) and the reaction mixture was shaken for a period of time. 6 hours at 30 ° C. After the hydrolytic resection is complete, the mixture is applied to Vfhatman 1 chromatography paper and chromatographic separation is carried out in an aeeton-n-butanol-water system for 16 hours. The zone corresponding mobility D-galacto ^# of the cut out and eluted with 50% aqueous ethanol. The radiochemical yield of D-galactose was 0.4 kBq (2%).

Example 2

The procedure was as in Example 1, except that bydrolytic cleavage of the cell wall was carried out with 0.5 ml of the diluted enzyme system (1: 5) of a snail (Helix pomatle) freshly prepared from the digestive tract with 50 mM citrate-phosphate buffer pH 6 , 0th After 8 hours of reaction at 30 ° C, the reaction mixture was worked up as described in Example 1. The radiochemical yield of D-galactose was as in Example 1.

Example 3 '' '

The procedure was as in Example 1, except that the reaction was carried out in distilled water. However, 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-labeled D-galactose sugar of interest in the field of carbohydrate chemistry and biochemistry and for bacteriological and clinical practice.

Claims (1)

Method of preparation (U-D-gapactose) characterized in that whole cells of the alga Chlo rella cultured in a radionuclide-labeled carbon dioxide medium are treated with an enzyme / β-galactosidase solution, buffered to a pH of 4.5 to 6, releases D-galactose (U-C) into solution, which is obtained from the solution by paper chromatography.