1.Determination of the decomposition products of Usal in model systems and determination of dioxopiperazine in soft drinks by HPLC.
Prudel M, Davídková E. Nahrung. 1985;29(4):381-9.
A HPLC method for the determination of Usal (Aspartame hydrochloride, L-aspartyl-L-phenylalanine methyl ester hydrochloride) and its decomposition products was elaborated. Aspartic acid, phenylalanine, phenylalanine methyl ester, aspartyl-phenylalanine, phenylalanyl-aspartic acid, 5-benzyl-3,6-dioxo-2-piperazineacetic acid (DOP) and Usal were separated on Separon SI C-18. The mobile phase was: 0.5 M NaH2PO4 (pH 2.1) and methanol (85:15 v/v). The detection was carried out at 200 nm. The method for DOP determination was tested by the analysis of 10 types of soft drinks to which DOP was added. In two newly developed sorts of soft drinks sweetened with Usal the formation of DOP was followed during storage. The DOP increment after 34 days of storage reached 0.7 and 6 mg/l at 7 and 20 degrees C, resp. The method is also suitable for DOP determination in the sweetener itself.
2.Partial purification, and some properties and reactivities of cetraxate benzyl ester hydrochloride-hydrolyzing enzyme.
Kuroda H, Miyadera A, Imura A, Suzuki A. Chem Pharm Bull (Tokyo). 1989 Nov;37(11):2929-32.
Debenzylating enzyme from Aspergillus niger enzyme (commercial crude cellulase) catalyzes the hydrolysis of cetraxate benzyl ester hydrochloride (2), a precursor of the antiulcer agent (1). The enzyme was highly purified by three kinds of chromatographies (hydrophobic, ion exchange, gel filtration) with a recovery of 36%. The content of the debenzylating enzyme was about 0.1% in the crude cellulase, but the enzyme showed no cellulase activity. The purified enzyme was inactivated by Hg2+, and diisopropyl phosphorofluoridate (DFP). It was a monomer with a molecular weight of about 35,000, and its isoelectric point was estimated to be 5.3. It showed a debenzylating activity for the phenylpropionic acid benzyl ester moiety of various benzyl ester derivatives, and the benzyl ester of phenylalanine or that of tyrosine was also well hydrolyzed.