title: Characterization of the interaction of 2-[2-nitro-4-(trifluoromethyl)benzoyl]-4,4,6,6,-tetramethyl- cyclohexane-1,3,5-trione with rat hepatic 4-hydroxyphenylpyruvate dioxygenase.
abstract: The synthetic beta-triketones are a novel family of chemicals, developed as herbicides that have activity on grass and broadleaf weeds and are selective in corn. Toxicological evaluation of a number of these chemicals has established that they interfere with rat hepatic tyrosine catabolism in vivo by inhibiting the enzyme 4-hydroxyphenylpyruvate dioxygenase (HPPD). This paper describes the kinetics of inhibition of rat hepatic HPPD in vitro by the representative beta-triketone 2-[2-nitro-4-(trifluoromethyl)benzoyl]-4,4,6,6- tetramethylcyclohexane-1,3,5-trione (1). A marked inhibition of rat hepatic HPPD was observed when 1 was incubated with the enzyme for 3 min at 37 degrees C prior to the initiation of the enzyme reaction by the addition of substrate. In this system, a concentration of 200 nM 1 resulted in a > 90% loss of HPPD activity, and an apparent IC50 was established at approximately 50 nM. The rate constant for the inactivation of HPPD by 1 was (1.5 +/- 0.2) x 10(-5) s-1 nM-1 as determined by progress curve data of oxygen consumed by HPPD with time. This inhibition is reversible in that the enzyme-inhibitor complex slowly dissociates, with approximately 5.5 +/- 0.6% of the enzyme activity being recovered by 6 h at 25 degrees C (t1/2, 25 degrees C, estimated at 101 +/- 14 h). In short, our studies establish 1 to be a tight-binding inhibitor of rat hepatic HPPD in vitro. This inhibition is characterized by the rapid inactivation of HPPD by the formation of an enzyme-inhibitor complex that dissociates extremely slowly with recovery of enzyme activity.
authors.affiliation: Zeneca Central Toxicology Laboratory, Cheshire, UK.
authors.forename: M K
authors.initials: MK
authors.lastname: Ellis
authors.forename: A C
authors.initials: AC
authors.lastname: Whitfield
authors.forename: L A
authors.initials: LA
authors.lastname: Gowans
authors.forename: T R
authors.initials: TR
authors.lastname: Auton
authors.forename: W M
authors.initials: WM
authors.lastname: Provan
authors.forename: E A
authors.initials: EA
authors.lastname: Lock
authors.forename: D L
authors.initials: DL
authors.lastname: Lee
authors.forename: L L
authors.initials: LL
authors.lastname: Smith
mesh_terms: D015111:4-Hydroxyphenylpyruvate Dioxygenase / Q000037:antagonists & inhibitors* / Q000187:drug effects*; D000818:Animals; D003510:Cyclohexanes / Q000737:chemistry / Q000494:pharmacology*; D003600:Cytosol / Q000187:drug effects / Q000201:enzymology; D004347:Drug Interactions; D006540:Herbicides / Q000494:pharmacology*; D006713:Homogentisic Acid / Q000138:chemical synthesis; D007659:Ketones / Q000737:chemistry / Q000494:pharmacology*; D008099:Liver / Q000187:drug effects / Q000201:enzymology*; D051381:Rats
chemical_list: C098916:2-(2-nitro-4-(trifluoromethyl)benzoyl)-4,4,6,6-tetramethylcyclohexane-1,3,5-trione; D003510:Cyclohexanes; D006540:Herbicides; D007659:Ketones; D015111:4-Hydroxyphenylpyruvate Dioxygenase; D006713:Homogentisic Acid