Mass spectrometric determination of ergosterol in a prairie natural wetland

Fungi are the main decomposers of plant material in an aquatic system. Levels of ergosterol, a compound generally specific to the cell membranes of fungi can be used as an indirect measure of their presence and biomass. Described is a procedure utilising reversed-phase liquid chromatography with positive-ion atmospheric pressure chemical ionization tandem mass spectrometry (LC-APCI-MS-MS) for full quantification and confirmation of ergosterol in various wetland matrices. Solid and liquid samples (0.2-1 g dry weight and 10 ml) were subjected to alkaline saponification followed by serial extraction using pentane (3 x 10 ml). The procedure was applicable to quantitative analysis of wetland samples with little or no clean up. Under low energy (CID) collision induced dissociation conditions the major product-ion formed from m/z 379.4 [M + H-H2O]+ was m/z 69.4 [(CH3)2CHCH=CH]+. Selected reaction monitoring (SRM) of this transition along with the retention time were used to confirm that ergosterol was widely distributed at ppm levels (2.4 to 303 microg ash free dry mass (AFDM)) in matrices of decaying willow leaves, Scirpus stems (living and dead) and sediment collected at the water-sediment interface. Comparison between LC-APCI-MS-MS (SRM), LC-APCI-MS using selected ion monitoring (SIM) and liquid chromatography with ultraviolet absorption detection (LC-UV) indicated that SRM analysis was the most selective technique.