IMMOBILIZATION OF MYROSINASE ON MEMBRANE FOR DETERMINING THE GLUCOSINOLATE CONTENT OF CRUCIFEROUS MATERIAL

A novel detection system for the direct determination of the glucosinolate content in crude aqueous extracts from cruciferous material is described. In the proposed system the myrosinase from Sinapis alba (thioglucoside glucohydrolase, EC 3.2.3.1) is immobilized with high yield on a nylon 6.6 membrane. The optimized immobilization and experimental parameters are also described. The enzymatic assay, based on the initial-rate method, uses the pH-stat technique. The biosensor was evaluated for linearity using sinigrin, progoitrin, and increasing amounts of crude extracts and showed a linear correlation (near l) up to 3.5-mu-mol of glucosinolate. The sinigrin recovery from a crude rapeseed extract ranged between 99 and 103%. Although the reproducibility appeared to be comparable to that of other time-consuming and more expensive techniques, it was affected by a certain difficulty of restoring the initial condition of the system after each measurement. A comparison of the data obtained by the biosensor with those of HPLC shows a good correlation (y = -0.2158 + 0.9989 + r = 0.999, n = 12). In addition, immobilized myrosinase, if stored wet at 4-degrees-C, appears to be very stable, retaining nearly all of its activity for more than 15 months and after about 1000 assays.