We have quantified ribose, rhamnose, arabinose, xylose, fucose, mannose, glucose, and galactose in soil by gas chromatography (GC) simultaneously after converting to aldononitrile acetate derivatives. A recommended single-hydrolytic step by 4M trifluoroacetic acid (TFA) at 105°C for 4h was more effective for releasing soil neutral sugars from non-cellulosic carbohydrates and better suited to our purification procedure compared with the sulphuric acid hydrolysis. Linearity of the GC detection for each neutral sugar was in the range of 10–640μgml −1 and the recovery of neutral sugars from the spiked soil samples ranged from 76% to 109.7%. The coefficients of variation of the neutral sugars in four soils were lower than 2.0% for the instrument and 4.6–7.6% for the whole determination procedures. Compared with the trimethylsilyl (TMS) derivatization, the recovery of our newly modified method was more satisfactory and the reproducibility of ribose was improved significantly. Moreover, the aldononitrile acetate derivative was more stable than TMS derivative. Therefore, it is a promising approach suitable for a routine use in the quantitative analysis of soil neutral sugars, since it is fast, sensitive, and reproducible.