John Randall

Major: Water Resources Science and Management

Faculty Advisor: Inna Popova

Project Title:

Measuring Mustard Seed Myrosinase Activity in Soil

Abstract:

Myrosinase, an enzyme present in plants of Brassicaceae family, hydrolyses endogenous glucosinolates to biologically active isothiocyanates as a part of plant defense mechanism. Myrosinase-glucosinolates system can be utilized for development of biopesticides from agricultural crops such as mustard meal. This is increasingly important as the increasing reliance on synthetic pesticides has resulted in the adaptation of pesticide resistant species. This necessitates constant modification, or increased use of existing pesticides to combat increasingly resilient pests. While biopesticides, such as isothiocyanates, may prove to be a safer and more sustainable alternative to combating pests, their fate in soil is not well understood due to the lack of quantification assays. Thus, the objective of the study was to provide methodology for evaluation of myrosinase-glucosinolates system in soil. To measure myrosinase activity, a colorimetric assay based on the reaction between ferric ions and thiocyanate ions was used. Sinalbin isolated from S. alba seed meal was used as a substrate. When myrosinase reacts with sinalbin the thiocyanate that is produced will react with iron to produce iron (III) thiocyanate, which forms a dark, blood red color that absorbs light at 450 nm. Substrate and products of enzymatic reaction were quantified by high performance liquid chromatography and ion chromatography, respectively. The activity of myrosinase in the solution was determined by the change in absorbance over time at 450 nm using a UV/Vis spectrophotometer. The concentration and production of biopesticides throughout the soil column can then be measured by performing this assay with soil solution from varying depths.

Funding: USDA NIFA (2017-51102-27271).