To investigate the effect of longterm fertilization on lignin accumulation and clarify its influencing factors in subtropical agricultural upland soils, alkaline CuO oxidation and gas chromatography was performed to quantify the amount of lignin and its monomers components (V, S and C). The soil samples were collected from the fertilization treatments of NPK and NPKS (NPK combined with straw) in Huanjiang County, Guangxi Province (limestone soil) and Taoyuan County, Hunan Province (red soil). The results showed that NPK had no significant effect on the lignin content (Sum_VSC) of limestone soil, whereas the content in red soil significantly increased by (55±1)%. For the NPKS treatment, the lignin content in limestone and red soil increased by (328±4)% and (456±9)%, respectively. After the same fertilization treatment, the proportion of cinnamyl (C)type significantly increased in red soil, while a significant increase of vanillyl (V)type monomers occurred in limestone soil, indicating that lignin degradation in agricultural soils was monomer specific. Furthermore, the acidtoaldehyde ratios of syringyltype \[(Ac/Al) _S\] or vanillyltype \[(Ac/Al) _V\] monomers tended to decrease after long-term fertilization with the higher value for limestone soil, suggesting the degree of lignin degradation in limestone was higher than that in red soil. Soil organic matter and total nitrogen were not correlated with lignin content, but were significantly correlated with the composition of VSC monomers. Meanwhile, the available nutrient content in the soil (available nitrogen, phosphorus, and potassium) was closely related to the contents and components of V, S, and Ctype monomers (P<0.05). It indicated that the availability of soil nutrition should be considered as a key factor for the accumulation of lignin.