The objectives were to develop an assay method for detection and quantification, and determine kinetic parameters of β-D-fucosidase activity in soil. The optimal activities of this enzyme were at approximately pH 6.0 and 55 °C, respectively. The Km values, based on hydrolyzing p-nitrophenyl-β-D-fucopyranoside, in five soils ranged from 2.40 to 2.89 mM, and the Vmax values spanned from 206 to 1060 μmol p-nitrophenol released kg−1 soil h−1. At reaction temperatures from 10 to 40 °C, the mean temperature coefficient (Q10) and activation energy (Ea) ranged from 1.67 to 1.90, and 44.22 to 56.39 kJ mol−1, respectively. Employing a chromogenic p-nitrophenyl substrate, the developed method detects as little as 3.5 μmol p-nitrophenol released kg−1 soil h−1 (limit of detection or LOD), quantifies ≥ 10.7 μmol p-nitrophenol released kg−1 soil h−1 (limit of quantitation or LOQ), and had coefficients of variance (CV) of < 5.6%. β-D-fucosidases purified from some sources also hydrolyze β-D-galactoside, β-D-glucosides, and/or α-L-arabinosides. This method offers reproducibility and precision that are comparable with bench-scale assays, but also has the advantage of using a smaller amount of soil, reducing costs on reagents, supplies, labor, and waste management.

中文翻译：

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量化土壤中 β-D-岩藻糖苷酶的活性

目的是开发一种检测和定量分析方法，并确定土壤中 β-D-岩藻糖苷酶活性的动力学参数。该酶的最佳活性分别在大约 pH 6.0 和 55 °C。Km 值基于水解对硝基苯基-β-D-吡喃岩藻糖苷，在五种土壤中的范围为 2.40 至 2.89 mM，Vmax 值的范围为 206 至 1060 μmol 对硝基苯酚释放 kg-1 土壤 h-1。在 10 至 40 °C 的反应温度下，平均温度系数 (Q10) 和活化能 (Ea) 的范围分别为 1.67 至 1.90 和 44.22 至 56.39 kJ mol-1。采用显色对硝基苯基底物，开发的方法检测到释放的 3.5 μmol 对硝基苯酚 kg−1 土壤 h−1（检测限或 LO​​D），量化≥ 10。7 μmol 对硝基苯酚释放 kg−1 土壤 h−1（定量限或 LO​​Q），变异系数 (CV) < 5.6%。从某些来源纯化的 β-D-岩藻糖苷酶也可水解 β-D-半乳糖苷、β-D-葡萄糖苷和/或 α-L-阿拉伯糖苷。这种方法提供了可与实验室规模化验相媲美的重现性和精确度，但还具有使用较少量土壤、降低试剂、供应、劳动力和废物管理成本的优势。