AbstractIt has been suggested that the Klason method overestimates the lignin content of non-wood tissues of plants. To evaluate the effect of pre-extraction treatments on lignin determination in leaves, nine kinds of pre-extraction treatment were applied to ginkgo leaves and zelkova leaves. The apparent lignin contents (lignin determination by the Klason method) of ginkgo and zelkova leaves without pre-extraction treatments were 30.7% and 42.6%, respectively. After the various pre-extraction treatments, the apparent lignin contents were still high. On the other hand, the yield of nitrobenzene oxidation products (NPs) per apparent lignin content was maintained at a very low level before and after pre-extraction treatments (maximum value was 6% for ginkgo leaf and 11% for zelkova leaf after extraction treatment) compared with the value from wood (25 to 60%). These results suggested that the Klason method overestimates the lignin content of leaves even after the pre-extraction treatments examined in this study. In addition, a considerable part of the sample from which NPs or neutral sugars originate was lost during these pre-extraction stages. These results implied that some parts of the cell wall components were also removed by these pre-extraction treatments.