Identification of secretin diastereoisomers produced during synthesis

• Published in: Journal of pharmaceutical sciences Vol. 78; no. 2; p. 91
• Main Authors: Tsuda, T, Uchiyama, M, Sato, T, Yoshino, H, Tsuchiya, Y, Ishikawa, S, Ohmae, M, Watanabe, S, Miyake, Y
• Format: Journal Article
• Language: English
• Published: United States 01.02.1989
• Subjects: Amino Acids - analysis; Chromatography, Gas; Chromatography, High Pressure Liquid; Chymotrypsin; D-Amino-Acid Oxidase; Hydrolysis; Indicators and Reagents; Secretin - analysis; Secretin - chemical synthesis; Secretin - isolation & purification; Stereoisomerism; Trypsin
• ISSN: 0022-3549
• DOI: 10.1002/jps.2600780203

The byproducts P-1 and P-2, which were produced during the synthesis of porcine secretin, were isolated in pure form from the crude secretin by HPLC. These were identified by a combination of amino acid analysis, enzymatic digestion, and isocratic or linear gradient reversed-phase (RP)-HPLC. The amino acid compositions of P1 and P2, determined by amino acid analysis after acid hydrolysis, were found to be the same as those of porcine secretin without distinction between L-and D-amino acids. But, HPLC of their digestive fragments with trypsin and alpha-chymotrypsin differed from that of secretin. The fragments, S7-12 of P-1 and S13-21 of P-2 were determined to be different from the corresponding fragments obtained from secretin by HPLC analysis of their digestive fragments. The amino acid composition of each acid hydrolysate, following digestion with D-amino acid oxidase, was found to have less leucine or alanine content than secretin. The HPLC analysis of the fragments from P-1 and P-2 by tryptic and alpha-chymotryptic digestion showed that they are the same as those from synthetic D-Leu10 secretin or D-Ala17 secretin, respectively. Consequently, P-1 and P-2 are concluded to be the secretin diastereoisomers, D-Leu10 and D-Ala17 secretin, respectively.