Construction of a microbial metabolite fraction library with NPPlot for discovery of novel metabolites

We have constructed a microbial metabolite fraction library combined with Natural Products Plot (NPPlot), which is an original and unique database based on physicochemical information of microbial metabolites, for efficient discovery of novel metabolites.  A fraction library has been prepared by a systematic separation based on a basic chromatographic technique including middle pressure liquid chromatography and HPLC.  Each fraction in the library is submitted to DAD-LC/MS analysis to obtain physicochemical data including UV and mass spectra of metabolites in the fraction.  Each metabolite is plotted in a two-dimensional area based on the retention time and molecular weight for X- and Y-axis to generate the NPPlot for each strain.  The NPPlot is a distribution map and shows characteristic pattern for the strain.  Therefore, we can use the NPPlot for screening of novel metabolites by searching a characteristic distribution pattern and comparison of NPPlots between strains.

On the screening for structurally unique metabolites from the fraction library by the NPPlot search, two classes of novel compounds, verticilactam and spirotoamides, and new quinomycin derivatives were discovered and isolated from the fraction libraries.  Verticilactam had unprecedented beta-keto-amide moiety in 16-membered lactam ring and spirotoamides had 6,6-spiroacetal moiety with a carboxamide group.

These results showed that our strategy constructing the fraction library combined with NPPlot was a powerful tool to discover and isolate novel metabolites from microorganisms.  Also the method can be applied to other natural sources.