Comparison and in vivo Relevance of Two Different in vitro Head Space Metabolic Systems: Liver S9 and Liver Slices

Abstract

Ahstruct:In vitro systems of high biological organization. e.g. containing intact hepatocytes. have been considered as more reliable for metabolic studies and in vivo predictions of toxicokinetics than subcellular systems. For this reason, the kinetics and metabolism of low-molecular-weight volatile chemicals, i.e. head space elimination of substrate and formation of metabolites, were compared in liver S9 and in liver slices. Two substrates. toluene and n-hexane, were used as they represent differences in metabolic pathways and physical-chemical properties. The two systems responded similarly to diethyldithiocarbamate inhibition and acetone inhibitiodinduction of cytochrome P-450 mediated metabolism. In contrast to liver S9. liver slices did not respond adequately to phenobarbital induction raising the question of substrate availability for P-450 enzymes in liver slices. Liver slices appeared to be superior to liver S9 when specific metabolic pathways involving phase I1 enzymes were investigated. However, liver SY seemed to be at least as good as liver slices for estimation of total metabolic rate constants (K,,,, V,,,) as a basis for in vivo predictions. As the head space liver S9 system is faster and easier to operate than liver slices, it is a promising screening tool for the metabolism of volatile compounds and metabolic interactions.

Evaluation of physiologically based pharmacokinetic models for use in risk assessment

“…In particular, in vitro data for humans can then be extrapolated to in vivo human predictions by assuming that the same relationship that successfully describes the in vitro to in vivo relationship in animals effectively converts the human in vitro data to the in vivo situation. There are several examples of successful application based on appropriate in vitro-in vivo scaling methods (Reitz et al, 1989(Reitz et al, , 1996bKedderis and Held, 1996;Mortensen et al, 1997;Mortensen and Nilsen, 1998;Lipscomb et al, 1998Lipscomb et al, , 2003Cole et al, 2001;Hissink et al, 2002;U.S. EPA, 2006b), although the extrapolation of in vitro data to the intact animal is not always straightforward (Haddad et al, 1997(Haddad et al, , 1998.…”

In vitro Screening for Metabolic Interactions among Frequently Occurring Binary Mixtures of Volatile Organic Chemicals in Norwegian Occupational Atmosphere

“…Calculations were based on the integrated peak areas. All head space concentrations were calculated on the basis of a minimum of three freshly prepared calibration standards covering the linear range of each analytical series (Mortensen et al 1997). Linearity (r2>0.996) was obtained for all solvents up to 5000 ppm.…”

“…Metabolic rates (nmol/g liver wet weight/ min.) were calculated on the basis of disappearance of the parent chemical (substrate) from head space or liver S9 after correction for the disappearance rate from a concomitant incubation with heat inactivated liver S9 according to Mortensen et al (1997). The metabolic rate represented the amount substrate metabolized after incubation for 30 min.…”

“…Preparation and incubation of liver S9 were performed as described earlier (Mortensen et al 1997). Single incubations were performed for each solvent and animal with liver S9 taken from five induced or five non-induced individual animals.…”

“…This was done by introducing chemicals either alone or as binary mixtures to a vial equilibration head space rat liver S9 metabolic system, containing livers from untreated rats or livers from rats pretreated in vivo with the investigated binary mixture. The in vitro metabolic system to be used has proven well fitted for studies of metabolism of organic solvents in general (Nakajima & Sat0 1979a;Sat0 & Nakajima 1979;Mortensen et al 1997), for studies on metaAuthor for correspondence: Bjarte Mortensen, Section for Environmental Technology, Norsk Hydro ASA, Research Centre, PO. Box 2560, N-3901 Porsgrunn, Norway (fax +47 35563384). bolic interactions between organic solvents (Mortensen et al 1998), for studies of metabolism of solvents with different solubility (Mortensen & Nilsen 1998a) and for extrapolation to man (Mortensen & Nilsen 1998b).…”