DD239209A1 - PROCESS FOR THE PREPARATION OF O-ALKYL SUBSTITUTED GLYCERO AND DESOXYGLYCEROPHOSPHO SERINES - Google Patents

Abstract

The aim of the invention is to develop a process for the preparation of 0-alkyl-substituted glycero- or deoxyglycerophospho-L-serines. According to the invention, these compounds are prepared by reaction of 0-alkyl-substituted glycerol or deoxyglycerophosphoric acid esters with L-serine under the action of phospholipase D.

Description

in which R ^{1 is} an alkyl radical having 5 to 30 C atoms and R ^{2 is} a hydrogen atom or an alkoxy group having 5 to 30 C atoms, and of their salts, characterized in that O-alkyl-substituted glycerol or Desoxyglycerophorsäureester the general formula II, in which R ¹ and R ^{2 have} the already known meanings and X is a methyl radical or an alkyl radical having 2 to 6 C atoms, which is also represented by OH, Cl, Br, NH ₂ , NHCH ₃ , N (CH ₃ I ₂ or N (CH ₃ I ₃ may be substituted.

CH ₀ -OR ^{1 2}

II

ii

CH ₂ -OPOX 0 "

means reacted with L-serine in the presence of phospholipase D and the resulting phosphoserine derivative is isolated and purified by conventional methods.

Field of application of the invention

Process for the preparation of O-alkyl-substituted glycero- or deoxyglycerophospho-L-serines of the general formula I,

CH ₉ -OR ^{1 1} 2 CH-R ² ₀

I ii

CH ₀ -OPO-CH ₀ -CH-COOH

OH NH ₂

in which R ^{1 is} an alkyl radical having 5 to 30 C atoms and R ^{2 is} a hydrogen atom or an alkoxy group having 5 to 30 C atoms, and of their salts.

These compounds are alkyl analogs of naturally occurring phosphatidylserines and lysophosphatidylserines, respectively. Due to the presence of an ether moiety instead of an ester group, these substances are not cleavable by the phospholipases Α _Ί and A ₂ . Because of this increased biostability, these compounds are interesting objects of study, for example, for a number of biological processes involving natural phosphatidylserines. Field of application of the invention is the pharmaceutical or chemical industry.

Characteristic of the known technical solutions

It is known that phosphatidylserines can be obtained by reaction of phosphatidic acids or their derivatives with serines whose amino and carboxyl groups are generally protected, or by reacting O-acylglycerohalohydrins with protected phosphoserines (H.Eibl, Chem. Phys. Lipids, 26 [1980] 40 &42; AJSIotb'oom et al., Chem. Phys.

Lipids, 5 [1970] 301-397; Hermetter et al., Chem. Phys. Lipids, 30 [1982] 35-45).

Another possibility for the synthesis of phosphatidylserines is the enzymatically catalyzed transfer of the phosphatidyl radical, for example phosphatidylcholine, to L-serine (P. Comfurius et al., Biochim. Biophys. Acta, 488 [1977]

Processes for the synthesis of the O-alkyl-substituted glycerol and deoxyglycerophospho-L-serines of the general formula I have hitherto not been described in the literature.

Object of the invention

The aim of the invention is to develop a process for the preparation of the compounds of general formula I.

Explanation of the essence of the invention

According to the invention, the 0-alkyl-substituted glycerol or deoxyglycerophospho-L-serines of the general formula I are obtained by reacting the O-alkyl-substituted glycerol or deoxyglycerophosphoic acid esters of the general formula II

CH ₉ -OR ¹

¹ * "2 CU-P, 0

CH ₀ -OPOX 0 "

in which R ¹ and R ^{2 have} the already known meanings and X is a methyl radical or an alkyl radical having 2 to 6 C atoms, which is also represented by OH, Cl, Br, NH ₂ , NHCH ₃ , N (CH ₃ I ₂ or N (CH ₃ ) ₃ may be substituted with L-serine in the presence of phospholipase D and the isolated Phosphorserin derivative isolated by conventional methods.In the individual implementation by reacting a solution or suspension of a phospholipid of formula II in a highly concentrated solution of L-serine in an aqueous buffer system, for example acetate and Tris buffer, at pHs between 4.8 and 8.5 in the presence of a 0.01 to 0.1 M solution of a Calcium salt, for example calcium chloride, under the action of Phcspholipase D react at temperatures between 10 and 50 ° C, wherein the reaction mixture diethyl ether or a mixture of diethyl ether with other organic solvents, for example chloroform, is added The implementation takes place within a period of 0.5 to 24 hours. The course of the reaction can be monitored by thin-layer chromatography. In order to avoid losses of yield due to hydrolysis during the work-up, a 0.1 M EDTA solution is added to the reaction mixture to deactivate the enzyme, and the resulting phosphoserine derivative is then isolated in the usual way. The process according to the invention enables the synthesis of the racemates and the optical stereoisomers of type I (in the case of R ² Φ H).

The compounds according to the invention have physiological activity. They are cancerostatic and inhibit, for example, the growth of Ehrlich ascites cells in vitro even at very low concentrations. The invention is illustrated by the following example:

embodiment example 1

2-deoxy-1-0-hexadecylglycero-3-phospho-L-serine

Formula I: R ¹ = C ₁₆ H ₃₃ , R ² = H)

A mixture of 1 g L-serine, 1.9 ml 1.0M acetate buffer (pH 5.6) with 0.1 M CaCl ₂ , 40 mg 2-deoxy-iO-hexadecylglycerophosphorsäureethylester (II: R ¹ = Ci ₆ H ₃₃ , R ² = H, X = C ₂ H ₅ ), 2 ml of ether / chloroform 9: 1, v / v) and 100 mg of a phospholipase D preparation obtained from about 500 g of white cabbage is stirred intensively at 40 ° ^{C. for} 4 hours. After cooling to room temperature, 4.35 ml of 0.1 M EDTA solution are added. The organic solvents are removed by introducing nitrogen. The mixture is stirred with 4.3 times the volume of chloroform / methanol (5: 8, v / v) for 30 minutes and the resulting unreacted serine is removed by suction.

The filtrate is stirred with 1 volume of water and 3.7 volumes of chloroform for 10 minutes, the organic phase separated and concentrated. The residue obtained is separated by column chromatography on 20 g of carboxymethylcellulose (Servacel CM 52), eluting successively with 75 ml of chloroform (fraction 1), 500 ml each of chloroform / methanol (9: 1.8: 2.7: 3.1: 1 , v / v, fractions 2 to 5). The phosphoserine derivative is obtained from fraction 5 in pure form (15 mg). TLC (Merck Kieselgel 60, finished plate):

Rf 0.13 CHCl ₃ ZCH ₃ OHZH ₂ O, 50: 25: 4, v / v / v).

Claims (1)

Invention claim: 1. A process for the preparation of O-alkyl-substituted glycerol and deoxyglycerophospho-L-serines of the general formula I, CH ₂ -OR ¹ PW-R I ii CH ₀ -OPO-CH ₀ -CH-COO !! I ² I ² I OH 'MH ₂