Abstract: GC-MS was applied to the determination of phospholipid fatty acids in soil. The sample was added into KOH solution with methanol as solvent. After methyl esterification, fatty acid methyl esters were extracted with hexane. The extract was separated on an Agilent DB-5MS column. Full-scaning mode and SIM were adopted in MS. Linear relationships were found between the peak area and the mass concentration of the 26 fatty acids methyl esters in definite ranges, with detection limits (3S/N) in the range of 0.001 2-0.295 9 mg·L-1. Recovery rates measured by standard addition method were in the range of 108%-115%, and RSDs (n=7) were in the range of 3.2%-6.4%.