Micellar electrokinetic chromatography after precapillary enzyme assay for the determination of phosphatase activity in semiarid soil

Abstract

Inorganic phosphorous (P) is an essential plant nutrient that must be added to most soils to maintain plant growth and sustain crop yields. Phosphatase enzymes play a major role in the processes of mineralization of organic P substrates. In the present study we propose a precise and accurate method to determine the alkaline and acid phosphatase activity in soil using adenosine monophosphate (AMP) as a substrate. The adenosine released by the enzymatic reaction was quantified by micellar electrokinetic chromatography (MEKC) method. The background electrolyte (BGE) consisted in 20 mM phosphate buffer, 10 mM sodium dodecyl sulphate (SDS) and 10% acetonitrile (ACN). The LOD and LOQ for adenosine were 2.27 × 10-4 and 6.4 × 10-4 mg mL-1, respectively. This sensitive method allowed the use of natural compound as enzymatic substrate, which is commonly found in soil. Moreover, we improve the detection method by diminishing the amount of waste and using a harmless substrate.