Microbial Oxidation of KE-298 Metabolites by Rhizopus sp. and Rhodococcus sp. Strains

The metabolites of the antirheumatic agent KE-298 in humans, (-)-(2R)-M-4 [(-)-(2R)-4-(4-hydroxymethylphenyl)-2-methylthiomethyl-4-oxobutanoic acid], (-)-(2R)-M-5 [diastereomers of (-)-(2R)-4-(4-hydroxymethyl-phenyl)-2-methylsulfinyl-methyl-4-oxobutanoic acid], (-)-(2R)-M-6 [(-)-(2R)-4-(4-carboxyphenyl)-2-methylthio-methyl-4-oxobutanoic acid], and (-)-(2R)-M-7 [di- astereomers of (-)-(2R)-4-(4-carboxyphenyl)-2-methyl-sulfinylmethyl-4-oxobutanoic acid] were synthesized based on microbial transformation. The substrate KE-748 (racemic form of (-)-(2R)- and (+)-(2S)-4-(4-methyl-phenyl)-2-methylthiomethyl-4-oxobutanoic acid: 7.5 g) was converted to (-)-(2R)-M-4 (1.84 g) using Rhizopus sp. TF0040 in a 50-l jar fermentor. Specific cytochrome P-450 inhibitors, SKF-525-A and metyrapone strongly inhibited the hydroxylation reaction. It was suggested that cytochrome P-450 is responsible for the microbial reaction. Furthermore, (-)-(2R)-M-4 (200 mg) was transformed to (-)-(2R)-M-6 (144 mg) by co-oxidation with n-hexadecane as a carbon source using Rhodococcus sp. TA0250 in a 1.4-l jar fermentor. Starting from (-)-(2R)-M-4 and (-)-(2R)-M-6 obtained as above, (-)-(2R)-M-5 and (-)-(2R)-M-7, respectively were chemically synthesized by m-chloroperoxybenzoic acid oxidation.