Excitation–emission matrix fluorescence spectroscopy coupled with parallel factor analysis was employed for characterisation of chloroform fumigation-extractable soil organic matter, commonly used for soil microbial biomass estimation. This allowed, for the first time, to discriminate between humic-like (i.e. noncellular) and microbial protein-like, fumigation-extractable components, challenging the presumption that only microbial constituents contribute to the fumigation flush of C serving as a proxy measure for soil microbial C. A Vertisol was assayed under increasing K₂SO₄ extractant molarity (0–0.5 M), which allowed increasing organic matter extractability levels and the association of these increases with relative contributions from microbial versus humic sources. Expectedly, protein-like fluorescence was found negligible in the nonfumigated soil extracts while comprising the bulk of fluorescence of the material becoming K₂SO₄-extractable due to fumigation. Nevertheless, fumigation also led to an increase in extractable concentrations of humic-like components, showing that not only microbial constituents were fumigation-extractable. Humic-like fluorescence in the fumigation flush generally increased with decreasing K₂SO₄ molarity, being minimal at 0.25 M K₂SO₄. Considering also the preference for maximal flush of extractable soil organic matter, indicative of maximal fumigation efficiency, the use of 0.25 M K₂SO₄ seems preferable for extraction of microbial biomass with minimal interference from humic substances, for the investigated Vertisol. The presented working framework for assessment and alleviation of interference from humic substances in microbial biomass estimation is recommended to be applied specifically to any soil type before routine monitoring.

激发-发射矩阵荧光光谱结合并行因子分析用于表征氯仿熏蒸可提取的土壤有机质，通常用于土壤微生物生物量估算。这首次允许区分腐殖质样（即非细胞）和微生物蛋白样，可熏蒸的成分，从而挑战了这样的假设，即仅微生物成分会导致C的熏蒸潮红作为土壤的替代指标。在增加的K ₂ SO ₄下测定了Vertisol萃取剂的摩尔浓度（0-0.5 M），可以提高有机物的萃取率，并且这些增加与微生物和腐殖质来源的相对贡献有关。预期地，发现在非熏蒸的土壤提取物中微不足道的蛋白质样荧光，而包括由于熏蒸而变为可提取的K ₂ SO ₄的物质的荧光的大部分。然而，熏蒸还导致腐殖质样成分的可提取浓度增加，这表明不仅微生物成分是可熏蒸提取的。熏蒸中的腐殖质荧光通常随K ₂ SO ₄摩尔浓度的降低而增加，在0.25 MK ₂ SO _4时最小。。还考虑到对最大可萃取土壤有机物冲洗的偏好，这表明最大的熏蒸效率，对于研究的Vertisol ，使用0.25 MK ₂ SO ₄似乎对于从腐殖质干扰最小的微生物生物质的萃取似乎是优选的。建议在常规监测之前，将所提出的评估和减轻微生物生物量估计中腐殖质干扰的工作框架专门应用于任何土壤类型。