Fig. 1.

Characterization of MSP. (A) Four different preparations (from four commercial sources) of MSP, including MSP-4, MSP-3, MSP-2 and MSP-1, were each incubated with exogenous myrosinase in PBS (1 mg powder per ml with 0.1 U myrosinase) for 30 min at room temperature (longer incubation time did not lead to further increase in ITC yield). MSP-1 was also incubated in PBS at room temperature for 0.5–4 h without exogenous myrosinase. At the end of incubation, total ITC levels in each solution were measured by the cyclocondensation assay. Each value is a mean ± SEM (n = 3). The result at the 0 time point was obtained by mixing MSP-1 with DADW, so that the endogenous myrosinase was inactivated and potential conversion from sinigrin to AITC was blocked. (B) MSP-1 was either mixed with DADW or incubated with exogenous myrosinase in phosphate buffer for 30 min before high-performance liquid chromatography. The arrows point to sinigrin and two potential minor glucosinolates of unknown identity. The compound or compounds representing the peak around 4 min has not been characterized, but it is not a glucosinolate, because myrosinase treatment had no effect on the peak. (C) Sinigrin was mixed in water with or without myrosinase in phosphate buffer for 30 min before high-performance liquid chromatography. The results are representative of at least three experiments.