CS197162B1 - Aminoacids esters hydrochlorides with chloramphenicol and process for preparing them - Google Patents

Description

The present invention relates to hydrochloramides of amino acid esters with chloramphenicol and to a process for their preparation.

The antibiotic chloraufenicol is currently being studied in terms of its mechanism of action. It is known that the effect of this antibiotic is to inhibit transpeptidation in a cell of a microorganism.

A particularly potent inhibitor of proteosynthesis is the antibiotic puromycin, which is a chemical analogue of aminoacyladenoain. In studying the effect of chloramphenicol on the formation of acetyl-phenylalanylpuromycin, it was found that chloramphenicol acts as a competitive inhibitor of puromycin.

A suitable chloramphenicol derivative bearing a free amino group in a suitable configuration and resembling as closely as possible the terminal arrangements of aminoacyladenosine (reap. Aminoacyl.cytosine) could be an effective acceptor in the peptidyltransferase reaction. Based on these structural assumptions, it was possible to approximate the structure of aminoacyladenosine, resp. aminoacylcytosine by binding an appropriately selected amino acid. Therefore chloramphenicol esters with amino acids of both steric representations (ie D- and L-) were prepared.

197 162

As shown in the figure, the structure is the largest in chloraafenicol erythroitoners. Therefore, not only B-threochlosantphenicol, but also pharecologically inactive L-threo and raceeiekéhe erythreoeaer were prepared.

Fourteen amino acid esters of ehloraaphenicol were prepared and examined for acceptor activity in a system containing 70 S riboaoay and N-acetyl-L-leu cinylhaxanuclaotide and 14 ° C labeled leucine as donor. Accelerator activity was equal to the β activity of puroaycin in the same system. Significant activity was found for D, L-erythrochloramphenicol esters, with no significant difference between the D- and L-amino acid ethers, as shown in the table:

D, L-chloramphenicol sterol acceptor activity in comparison and purpmycin (%) L-Loucinet 57 L-phenylalanine 30 L-σ-methyl-thyrosinate 45 D-valinate , ⁶⁵ '

Another problem was to find a suitable method for esterifying chloramphenicol with amino acids. In the chemistry of chloramphenicol several analogous preparations are described, such as glyednate, succinate, palmitate, in which the chloramphenicol is treated with chlorides of the appropriate acid with simultaneous addition of alkali to the neutralization of the leaving hydrogen chloride. However, in the case of amino acids, this method has not been applied due to the low stability of the amino acid chlorides β protected by the amino group. The method utilizing: amino acid condensation with chloramphenicol in the presence of N, N-dimethylformamide-dineopentylaetal was also unsuccessful. Success was achieved with the aid of diocyclohexylcarbodiimide (DCCI), in which the reaction of the amino-protected amino acid with chlorophenicol in the pyridine medium proceeds according to the equation:

Esterification takes place on the primary alcoholic group, ohloramphenicol, as evidenced by elemental analysis.

The commonly used benzyloxycarbonyl protecting groups could not be used for removal. conventional hydrogenation methods due to the presence of the nitro group and the dichloroacetyl residue. Also when using hydrogen bromide in acetic acid, resp. Trifluoroacetate, the reagents used for this purpose, destroys the molecule. Therefore, an o-nitrosulfenyl group has been chosen to protect the free amino group in amino acids, which can be removed under very mild conditions. By reacting the amino acids thus protected with chloramphenicol in the presence of dicyclohexylcarbodiimide, a number of esters have been obtained.

Accordingly, the invention relates to chloramphenicol ester of hydrochloride amino acid esters of formula 1 wherein R is a group

isopropyl, 2-methylpropyl, benzyl and 4-methoxybenzyl.

(1)

Furthermore, the invention relates to a process for the preparation of hydroehlorides of amino acid esters with ohloramphenicol. It is based on the fact that amino acids with protected amino group of formula 11

B-CH-COOO

NH · (II) wherein O is nitro-sulfenyl and the chloramphenicol is treated with dioyclohexylcarbodiimide in the presence of a solvent, preferably pyrimidine, at a temperature of from 0 to 50 ° C and then removed from the formed intermediate by acidic hydrolysis with hydrogen chloride in ether at -10 up to 35 ° C protecting the o-nitrosulfenyl group.

According to further features of the invention, L-leucine, L-phenylalanine, D-phenylalanine, L-methylthyrosine, D-valine are used as amino acids, and D-threo-chloramphenicol, L-threo-chloramphenicol and D, L-erythro-chloramphenicol are used as chloramphenicol. different combinations.

Thus, according to the invention, a new type of acceptor substrate has been prepared, which has not been described so far and which exhibits significant acceptor activity. The compound of the invention interferes efficiently in the proteosynthesis of microorganisms and has the potential for effective pharmacological use, particularly with respect to the use of L-threo and L-erythroisomer.

The compound of the invention and the process for its preparation are described in more detail below with reference to several exemplary embodiments.

Example 1 mmol of the dicyclohexylammonium salt of an amino acid was finely triturated and shaken in a separator in 75 mL of ethyl acetate. Then 20 ml of 0.2 N sulfuric acid was added and the mixture was shaken vigorously. The suspension is dissolved. The aqueous layer was discarded, the organic washed with 50 mL of water and dried over anhydrous sodium sulfate. After the desiccant was suctioned off, the ethyl acetate was evaporated in vacuo. Upon cooling, the oily residue usually crystallized.

3 mmol of chloramphenicol and 3.2 mmol of dicyclohaxylcarbodiimide were added to the N-NPS amino acid thus prepared. The substances were dissolved. in 30 ml dry pyridine. The solution thus prepared was left for 2 hours at room temperature. The precipitated dioyclohexylurea was aspirated and washed with chloroform. The solvent was evaporated from the combined filtrates on a vacuum evaporator at 50 ° C. The pyridine residue was removed by evaporation several times with xylene. The residue was subjected to preparative thin layer chromatography. Fractions of JRp. about 0.5 was washed into ether. The ethereal solution was dried over sodium sulfate and evaporated to dryness. The residue was poured over 20 ml dry ether. A white crystalline material precipitated while the yellow N-NPS amino acid ester with chloramphenicol went into solution. The crystalline white solid was filtered off with suction and washed with 5 ml of dry ether. To the combined ethereal solutions was added 3 mL of 2 N hydrogen chloride in anhydrous ether. After 3-4 minutes, the amino acid chloride chloride precipitate with chloramphenicol was aspirated and washed with dry ether until the filtrate was colorless. The ester thus obtained was dried under vacuum. All operations after the addition of the hydrogen chloride solution were carried out.

Mey β by excluding air humidity.

íítóiaO

From 2.80 g of N-nitroalphenyl-L-leucine dicyclohexylammonium salt and 1.00 g of L-threo-chloroeaphenicol, analogously to Example 1, 0.75 g of L-leucinate hydrochloride] threochloroamphenicol hydrochloride was obtained, m.p. Mp 92-97 ° C.

From 80 g of N-NPS-L-leucine dicyclohexylammonium salt and 1.00 g of D-threo-chlorophenicol, analogously to Example-1, 0.65 g of D-threo-chloramphenicol L-leucinate hydrochloride was obtained, m.p. Mp 114-118 ° C.

Example 4

Analogously to Example 1, 0.65 g of L-threo-chloroaphenicol L-phenylalanate was obtained from 2 g of L-threo-chloramphenicol and 1.5 g of dicyclohexylammonium salt of N-NPS-L-phenylalanine and 0.70 g of DCCI. Mp 95-100 ° C.

ISÍfiaáL2

Analogously to Example 1, 0.45 g of L-O'-methylthyrosinate D-threo-chlorosphenicol was obtained from 1.05 g of dicyclohexylammonium salt of N-NPS O-methylthyroeine, 0.65 g of chloremphenicol and 0.70 g of DCCI, m.p. 165-172 ° C,

Example <

Analogously to step 1, 0.5 g of dicyclohexylammonium salt was N-NPS-D-valine. 0.40 g of L-chloramphenicol and 0.40 g of DCCI obtained 0.30 g of D-valinate hydrochloride of D-chloramphenicol m.p. 104 to 111 ®tt.

Example 7 \

Analogous to the procedure of 1 and the weighing of 6, 0.20 g of L-chloroaphenicol D-valinate hydrochloride was obtained, m.p. Mp 168-175 ° C.

Claims (8)

1. Amino acid ester hydrochloride with chloramphenicol of formula I: OH NO, B-CH-CO-O-CH-CH-CH · (I) NH? .HCl NHCOCHC1 ₂ wherein B stands for isopropyl, 2-methylpropyl, benzyl and 4-methoxybenzyl. and. A process for the preparation of hydrohalides of amino acid esters and of the chloramphenicol of formula (I), characterized in that it is an amino acid and a protected amino group of formula (II), B-CH-COCH} SB-% (ΪΙ) where it represents an o-nitro-sulfenyl group, and the chloramphenicol acts with dioyl cyclohexylcarbimide in the presence of a roe-surfactant, preferably pyrimidine, at temperatures of from 9 to 99, and the resulting intermediate is removed by acid hydrolysis with hydrogen chloride in ether at 10 to 35 ° C of the o-nitro-sulfanyl group. 3. The method of claim 2, wherein L-leuine is used as the amino acid. D-threo-chloramphenicol is used as ohloramphenicol. 4. The process according to claim 2, wherein ee is L-threo-chloramphenicol as the chloramphenicol. 5. The method according to claim 2 or 3, characterized in that L-phenylalanine is used as aminol. also, two pressurized feed vessels according to claim ¹ and 3, characterized in that the amino acid used as the AE D íenylalanin. 7. The method of claim 2, wherein ee is L-o-methylthyrosium as the amino acid. 8. The method of claim 2, wherein the ee is D-valine. 9. The method of claim 2 wherein the chloramphenicol is D, L-arythro-chloramphenicol.