サイクロセリンによるβ-アミノイソ酪酸およびβ-アラニン代謝阻害-2-β-アミノオキシアラニンおよびα-ケト酸結合物質について
スポンサーリンク
概要
- 論文の詳細を見る
In the preceding paper, it was reported that the increase in the amount of urinary D-β-aminoisobutyrate (β-AIB) and β-alanine was resulted from the inhibition of β-AIB: pyruvate aminotransferase and β-alanine: α-ketoglutarate aminotransferase by D-cycloserine (CS) or its metabolites. A maximal concentration of CS in the liver of rats which were administered intraperitoneally at a dose of 20mg of the drug per 100g of body weight showed only a slight inhibitory effect on the enzyme reactions. The concentration of β-AIR and β-alanine in rat liver reached to maximal level 4 hours after the injection of CS when. CS was barely defectable in the liver. This suggested that the inhibition is due to a metabolite of CS rather than CS itself. Presence of the inhibitory substance was studied in the present paper.<BR>For the isolation of the inhibitors, rats were used and male mice were used for toxicity test of the drugs. Determination method of β-AIB: pyruvate and β-alanine: α-ketoglutarate aminotransferase was reported previously. Rabbit liver was used for the study of enzymatic conjugation of β-AOAL and α-keto acids. CS was determined using a high voltage electrophoresis.<BR>An inhibitory metabolite of CS was detected in the rat liver after CS injection in a preliminary experiment, and the same inhibitory substance was found in urine of rats which were injected CS intraperitoneally. The inhibitor was purified from the rat.urine using combination of ion-exchange chromatographies, and was identified as β-aminoxy-alanine from its behavior on paper chromatography, paper electrophoresis and ion exchange chromatography. In order to confirm the identification, the isolated compound was conjugated with pyridoxal phosphate and compared with β-aminoxyalanine-pyridoxal phosphate conjugate on the chromatographies and UV-spectrum. The results verified the above identification. The concentration of β-aminoxyalanine required for 50% inhibition of β-AIB pyruvate and β-alanine: α-ketoglutarate aminotransferases was 10<SUP>-6</SUP>M while that of CS was 10<SUP>-3</SUP>M. The above results support hypothesis that a metabolite of CS, β-aminoxyalanine, is the major substanceswith inhibitory effect on the aminotransferases.<BR>During the isolation experiment of inhibitory substances derived from CS, another fraction of ion exchange chromatography different from those of CS and β-aminoxyalanine was also found to contain an inhibitory substance. The substance was acidic. When CS or β-aminoxyalanine was incubated with substrates of transamination reaction, enzymic formation of a conjugated compound of β-aminoxyalanine and keto acids was found. β-aminoxyalanine and glyoxalate was incubated with rat liver extract, and the enzyme product was purified using ion exchange chromatography to isolate a crystalline material. Elementary analysis matched that of C<SUB>5</SUB>H<SUB>8</SUB>N<SUB>2</SUB>O<SUB>5</SUB>. This was a shiff base of β-aminoxy-alanine and glyoxalate. The concentration of the base required for 50% inhibition of β-AIB: pyruvate aminotransferase was roughly 3×10<SUP>-5</SUP>M.<BR>It is known that CS has various side effects in clinical use including headache, anxiety and convulsion. Experiment for acute toxicity of β-aminoxyalanine was carried out by using mice. LD 50 was 2.6g per kg of body weight which is slightly lower than that of CS. It was interesting that convulsion was consistently observed with the lethal dose of β-aminoxyalanine, while none with CS.
- 一般社団法人 日本結核病学会の論文