We set up a protocol for the assay of the arylesterase activity, using p-nitrophenyl acetate (p-NPA) as substrate, dimethylsulfoxide as solvent, modified universal buffer at pH 7.5, and determination of the reaction product (p-nitrophenol) after separation of non-hydrolysed p-NPA after reaction, and tested it using eight soils with a wide range of characteristics. Various incubation temperatures and times, pH values and substrate concentrations were also used to find the optimal conditions for the enzyme activity and to determine characteristics and kinetic parameters of soil arylesterase. Arylesterase activity was significantly correlated with total organic C, total N, and soil ATP content. Soil arylesterase activity showed a pH optimum at 7.5, optimal temperature between 55 and 65 °C and linear increase with incubation time. The Km values ranged from 4.3 to 8.5 mM, the Vmax values from 326 to 803 μmol p-NP g-1 h-1, with higher Km values observed in soils with higher organic matter content. We conclude that the proposed assay protocol is suitable to determine the arylesterase activity in a wide range of soils.
A protocol for the assay of arylesterase activity in soil
Renella G.
Conceptualization
2009
Abstract
We set up a protocol for the assay of the arylesterase activity, using p-nitrophenyl acetate (p-NPA) as substrate, dimethylsulfoxide as solvent, modified universal buffer at pH 7.5, and determination of the reaction product (p-nitrophenol) after separation of non-hydrolysed p-NPA after reaction, and tested it using eight soils with a wide range of characteristics. Various incubation temperatures and times, pH values and substrate concentrations were also used to find the optimal conditions for the enzyme activity and to determine characteristics and kinetic parameters of soil arylesterase. Arylesterase activity was significantly correlated with total organic C, total N, and soil ATP content. Soil arylesterase activity showed a pH optimum at 7.5, optimal temperature between 55 and 65 °C and linear increase with incubation time. The Km values ranged from 4.3 to 8.5 mM, the Vmax values from 326 to 803 μmol p-NP g-1 h-1, with higher Km values observed in soils with higher organic matter content. We conclude that the proposed assay protocol is suitable to determine the arylesterase activity in a wide range of soils.
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