Challenges in quantification of ligninolytic enzymes from Phanerochaete chrysosporium cultivation for pretreatment of cotton stalks

Enzymes play an important role in the breakdown of lignin during microbial pretreatment of lignocellulosic feedstocks. However, quantification of the various enzyme activities with assays developed for enzyme extracts from pure cultures can be challenging. In this study, spectrophotometric assays used for the quantification of peroxidases in enzyme extracts from submerged (SmC) and solid-state (SSC) cultivation of P. chrysosporium on cotton stalks during 14 days pretreatment failed to detect lignin peroxidase (LiP) and manganese peroxidase (MnP) activities. However, results from sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) suggested presence of protein bands with molecular weights corresponding to MnP and LiP in the enzyme extracts from fungal pretreatment cultures. Addition of crude enzyme extracts from SmC and SSC treated samples to fresh cotton stalks showed 3.42% and 7.45% increase in lignin content, respectively. This slight increase may be attributed to components within crude extracts that polymerize the phenolic compounds instead of resulting in delignification. It can be inferred from this study that although qualitative methods for ligninolytic enzyme estimation provide useful information, it is essential to investigate alternative approaches to quantify ligninolytic enzymes during cultivation on natural lignocellulosic materials to overcome the limitations of existing assays.