IDENTIFICATION OF 2 NEW INHIBITORS OF THE HEPATIC GLUCOSE-6-PHOSPHATASE SYSTEM

A high-throughput screening assay aimed at the detection of inhibitors of the translocase components of the hepatic glucose-6-phosphatase (G 6Pase) system was set up in a microplate Format using untreated and Tr iton X-100(TM)-disrupted rat liver microsomes. The assay measured the phosphate released from glucose-6-phosphate (G6P) using a standard cal orimetric method. Mie identified two struct urally unrelated compounds , 2-hydroxy-5-nitrobenzaldehyde and chlorogenic acid, which inhibited the hydrolysis of G6P in untreated microsomes, each with IC50 values o f 338 mu M and 226 mu M, respectively, but were devoid of activity in disrupted microsomes. Thus, the two compounds exhibited a high degree of specificity for translocase components. The effects of 2-hydroxy-5- nitrobenzaldehyde bear a resemblance to the effects of pyridoxal phosp hate. Studies with compounds structurally related to 2-hydroxy-5-nitro benzaldehyde suggest that both a phenolic OH-group in ortho position t o the aldehyde group and a suitable electron-withdrawing group in posi tion 3 or 5 of the aromatic ring are indispensable for the activity of this class of inhibitors. The inhibition pattern of chlorogenic acid is distinct from that of phloretin and is dependent on a free carboxyl group. The products of chlorogenic acid hydrolysis, quinic acid and c affeic acid, are inactive. 2-Hydroxy-5-nitrobenzaldehyde type inhibito rs and chlorogenic acid are potent new inhibitors for investigating th e structure and function of the translocase components of the G6Pase s ystem. (C) 1998 Wiley-Liss, Inc.