Water-based enzyme assays are more sensitive than buffer-based assays to tillage and site in the U.S. Corn Belt

• Sensitivity of enzyme activities to tillage and site compared across assay methods. • Water-based assays were 2-fold more sensitive to tillage and site than buffer-based. • Lower P- but higher C-, N- and S-hydrolytic enzyme activities by buffer-based assays. • Differences between water- and buffer-based activities not explained by assay pH. • Tillage effects on soil enzyme activities driven by soil organic carbon. Assay methodology can influence the sensitivity of soil enzyme activities to agroecosystem management practices. Using soils from six long-term tillage experiments encompassing maize-based agroecosystems and three soil orders (e.g., Mollisols, Alfisols, Ultisols) across the U.S., we quantified the sensitivity of acid phosphomonoesterase (AC-PME), alkaline phosphomonoesterase (AK-PME), β-glucosidase (BG), N-acetyl-β-D-glucosaminidase (NAG), and arylsulfatase (SUL) activities to tillage practices (conventional vs none) and site measured by water-based assays versus buffer-based assays. To explain soil enzyme activity sensitivity differences between assay methodologies, we assessed (1) relative differences in activities measured with buffer- rather relative to water-based assays, (2) differences between soil and assay pH, and (3) activities on a soil versus soil organic carbon (SOC) basis. On a soil mass basis, enzyme activities were ≈2-fold more sensitive to tillage and to site using water- relative to buffer-based assays. Water-based NAG activity was most sensitive to tillage and site, whereas buffer-based SUL activity was least sensitive to tillage and buffer-based AK-PME activity was least sensitive to site. The difference between assay pH and soil pH were 1.5- to 6-fold lower for water-based assays, though pH divergences did not fully explain differences between water- and buffer-based activities. Buffer-based assays produced lower AC-PME (−32%) and AK-PME (−53%) activities but higher BG (+23%), NAG (+43%) and SUL (+87%) activities relative to water-based assays. Tillage effects on activities were primarily driven by SOC, except for buffer-based AK-PME activities. However, water-based activities maintained an enhanced sensitivity to site. We recommend the use of water-based assays to maximize the sensitivity of soil enzyme activities to tillage and to site.