Constitutive and inducible bentazon hydroxylation in shattercane ( Sorghum bicolor) and Johnsongrass ( S. halapense)

Abstract

Bentazon hydroxylation was studied in excised shoots and microsomal preparations from shattercane ( Sorghum bicolor) and Johnsongrass ( S. halapense). Studies in vivo were conducted using excised shoots from untreated (constitutive) and naphthalic anhydride (NA)-treated (induced) etiolated seedlings. Both shattercane and Johnsongrass rapidly metabolized [ 14C]bentazon forming two metabolites: A minor metabolite of 6-OH-bentazon and a major metabolite, tentatively identified as the glycosyl conjugate of 6-OH-bentazon. Pretreating the seeds of both species with naphthalic anhydride increased bentazon metabolism in etiolated shoots by twofold. The cytochrome P450 inhibitor tetcyclacis (50 μ M) reduced bentazon metabolism in shoots from both NA-treated and untreated seedlings greater than 90%. Studies in vitro were conducted using microsomal preparations derived from NA-treated and untreated shattercance and Johnsongrass seedlings. Bentazon metabolism in microsomal preparations from both treated and untreated shattercane and Johnsongrass seedlings produced a single product: 6-OH-bentazon. Pretreatment of either shattercane or Johnsongrass with NA increased bentazon hydroxylation in vitro approximately twofold. Bentazon hydroxylation was nearly linear for 30 min in microsomal assays with both treated and untreated shattercane and Johnsongrass. Tetcyclacis (10 μ M) and PBO (100 μ M) inhibited bentazon hydroxylation in microsomes from treated and untreated seedlings from both species. Tridiphane (50 μ M) did not affect bentazon hydroxylation. Substrate saturation assays demonstrated that bentazon hydroxylation in vitro was saturable. Kinetic constants calculated for bentazon hydroxylation with NA-treated and untreated shattercane were K m = 234 and 317 μ M, respectively, and V max = 3.54 and 2.00 nmol · (mg protein · min) −1, respectively. Kinetic constants calculated for bentazon hydroxylation with NA-treated and untreated Johnsongrass were K m = 160 μM and 235 μ M, respectively, and V max = 1.25 and 0.68 nmol · (mg protein · min) −1, respectively. These data suggest that bentazon hydroxylation in NA-treated and untreated shattercane and Johnsongrass is mediated by constitutive and inducible cytochrome P450 monooxygenase(s).