M11 PM Internal Exposure
Monday, 14 November 2005: 1:50 PM - 5:30 PM in 343-344

(NIC-1117-740096) Use of on-line injection microdialysis to determine the rate of phenol glucuronidation in unanesthetized rainbow trout.

Nichols, J¹, Hoffman, A¹, Fitzsimmons, P¹, Lien, G¹, ¹ US EPA/ORD/NHEERL/MED, Duluth, Minnesota, USA

ABSTRACT- Phenol (PH) and its principal phase II metabolite, phenylglucuronide (PG), were measured in vivo in arterial blood of unanesthetized rainbow trout using on-line injection microdialysis. In vitro studies were initially conducted to evaluate retrodialysis calibrators and characterize plasma protein binding. Subsequent in vivo studies were conducted using [¹⁴C] PH and p-nitrophenyl glucuronide (PNPG) as internal calibrators for PH and PG, respectively. In the first set of in vivo experiments, trout were dosed continuously with PG for 24 h using a ventral aortic cannula. Fish were then allowed to depurate PG for an additional 48 h. Mass-balance calculations showed that 93% of infused PG was eliminated in urine during the depuration period. Based on peak values, the mean concentration of PG in urine was 3.1 times that in blood, suggesting that PG was actively secreted by the kidney. The kinetics of PG in blood were adequately described by a one-compartment renal clearance model. The mean fitted clearance constant was 13.5 ml/kg/h, which is twice the estimated GFR and about 2% of total renal blood flow. The apparent volume of distribution ranged from 200 to 300 ml, which approximates the estimated extracellular water volume. In a second set of experiments, trout were exposed to aqueous solutions of PH. In vivo rate constants for PH glucuronidation were estimated by fitting predicted PG concentrations in blood to measured values using a one-compartment clearance-volume model. Expressed on a whole-fish basis, the rate constant for PH glucuronidation was approximately 0.1/kg/h. The techniques developed in this study offer significant advantages over periodic blood sampling for kinetic studies with fish and provide a relatively simple method for estimating in vivo biotransformation rates. This abstract does not necessarily reflect EPA policy.

Key words: biotransformation, microdialysis, glucuronidation