Reconciling apparent variability in effects of biochar amendment on soil enzyme activities by assay optimization

We studied the effects of a biochar made from fast pyrolysis of switchgrass on four soil enzymes (β-glucosidase, β-N-acetylglucosaminidase, lipase, and leucine aminopeptidase) to determine if biochar would consistently modify soil biological activities. Thus, we conducted a series of enzyme assays on biochar-amended soils. Inconsistent results from enzyme assays of char-amended soils suggested that biochar had variable effects on soil enzyme activities, thus we conducted a second experiment to determine if biochar reacts predictably with either enzyme or substrate in in vitro reactions. Both colorimetric and fluorescent assays were used for β-glucosidase and β-N-acetylglucosaminidase. Seven days after biochar was added to microcosms of 3 different soils, fluorescence-based assays revealed some increased enzyme activities (up to 7-fold for one measure of β-glucosidase in a shrub-steppe soil) and some decreased activities (one-fifth of the unamended control for lipase measured in the same shrub-steppe soil), compared to non-amended soil. In an effort understand the varied effects, purified enzymes or substrates were briefly exposed to biochar and then assayed. In contrast to the soil assays, except for β-N-acetylglucosaminidase, the exposure of substrate to biochar reduced the apparent activity of the enzymes, suggesting that sorption reactions between substrate and biochar impeded enzyme function. Our findings indicate that fluorometric assays are more robust to, or account for, this sorption better than the colorimetric assays used herein. The activity of purified β-N-acetylglucosaminidase increased 50–75% following biochar exposure, suggesting a chemical enhancement of enzyme function. In some cases, biochar stimulates soil enzyme activities, to a much greater degree than soil assays would indicate, given that substrate reactivity can be impeded by biochar exposure. We conclude that the effects of biochar on enzyme activities in soils are highly variable; these effects are likely associated with reactions between biochar and the target substrate.