Anti-inflammatory Active Compound From Metabolites Of Microorganisms

Inflammation is the complex biological response of vascular tissues to harmful stimuli, such as pathogens, damaged cells, or irritants which protect against foreign substances or injury, but it also was an important pathophysiology of several deseases including arthritis, asthma, diabetes, allergies, asthma and atherosclerosis.Human leucocute elastase is a lysosomal Proteinase stored in high quantities in the azurophilic granules of polymorphonuclear leucocytes. The enzyme has been the subject of intensive study because of its potential for tissue destruction in many disease states, such as pulmonary emphysema and rheumatoid arthritis. For decades administration of exogenous elastase inhibitors has been proposed as a possible means of protecting tissues from proteolytic attack.Interleukin 1_β-converting enzyme also called caspase 1, is an autocatalytic cysteine protease, and is essential in the proteolytic processing of cytokines IL-1 and IL-18, which regulate many cellular processes, including immune and inflammation. ICE inhibitors were used to prevent many kind of inflammation.Soil microorganism have been recognized as a prolific source of biologically active natural products. Screening inhibitors of HLE and ICE from metabolites of microorganism is worth and meaningful for the discovery of anti-inflammatory lead compound. In this paper we describe the details of the screening, fermentation, isolation and structure elucidation of HLE and ICE inhibitors.In this paper high throughout screening methods were first established for screening HLE and ICE inhibitors from a large number of metabolites of microorganism. The positive strains picked out as candidate were cultured in large scale and the culture broth were further isolated by solvent extration, silica gel chromatography, ODS reverse phase chromatography, Sephadex LH-20 and Sephadex G-10 gel filtration, preparative TLC and ODS HPLC to get pure compounds. The inhibitory activity of the compounds against HLE and ICE were evaluated.In the course of screening for HLE inhibitors, N01WA-735 was picked out from 2141 culture broth as candidates for further isolation. 5 compounds named N01WA-735A,B,C,D and E respectively were isolted from the extracts of broth. On the basis of analysis various modern spectroscopic data (UV, NMR and ESI-MS) and physicochemical properties, the structures of the 5 compounds were identified as Nanaomycinα-E ,β-sitosterol, BE-52440A , BE-52440Bå’ŒNanaomycin A. N01WA-735E showed a strong inhibitory activity against HLE with with IC50 of 3.02μmol/L.From 1694 culture broth, the culture broth of N03WB-1134 was proved to have inhibitory activity against ICE. 2 compounds named N03WB-1134A and B were obtained from the extracts of broth. Based on physico-chemical properties and spectroscopic data, the two compounds were eluciated to be the known compounds benzene alcohol and patulim. Among them, N03WB-1134B showed best inhibitory activity on ICE with IC₅₀ of 6.02μmol/L.