Cytochrome carbon tetrachloride metabolized

On the other hand, microsomes may also directly oxidize or reduce various substrates. As already mentioned, microsomal oxidation of carbon tetrachloride results in the formation of trichloromethyl free radical and the initiation of lipid peroxidation. The effect of carbon tetrachloride on microsomes has been widely studied in connection with its cytotoxic activity in humans and animals. It has been shown that CCI4 is reduced by cytochrome P-450. For example, by the use of spin-trapping technique, Albani et al. [38] demonstrated the formation of the CCI3 radical in rat liver microsomal fractions and in vivo in rats. McCay et al. [39] found that carbon tetrachloride metabolism to CC13 by rat liver accompanied by the formation of lipid dienyl and lipid peroxydienyl radicals. The incubation of carbon tetrachloride with liver cells resulted in the formation of the C02 free radical (identified as the PBN-CO2 radical spin adduct) in addition to trichoromethyl radical [40]. It was found that glutathione rather than dioxygen is needed for the formation of this additional free radical. The formation of trichloromethyl radical caused the inactivation of hepatic microsomal calcium pump [41]. 

The metabolism of carbon tetrachloride proceeds via cytochrome P-450-dependent dehalogenation (Sipes et al. 1977). The first step involves cleavage of one carbon-chlorine bond to yield Cl- and a trichloromethyl free radical, which is then oxidized to the unstable intermediate trichloromethanol, the precursor of phosgene. Hydrolytic dechlorination of phosgene yields C02 and HC1 (Shah et al. 1979). Although there are similarities in the metabolism of chloroform and carbon tetrachloride, metabolic activation of chloroform produces primarily phosgene, whereas the level of phosgene production from... 

The relative importance of hepatic microsomal lipid peroxidation versus covalent binding of carbon tetrachloride-derived radicals has been the subject of considerable debate. Since cytochrome P-450 loss has been shown to be related to lipid peroxidation and to covalent binding, each in the absence of the other, both of these early consequences of carbon tetrachloride metabolism may contribute to P-450 destruction. Nevertheless, it is still not clear how these initial events are related to subsequent triglyceride accumulation, polyribosomal disaggregation, depression of protein synthesis, cell membrane breakdown and eventual death of the hepatocytes. Carbon tetrachloride... 

Gruebele, A., Zawaski, K., Kaplan, D. Novak, R.F. (1996) Cytochrome P45()2E1- and cyto-clrrome P4502Bl/2B2-catalysed carbon tetrachloride metabolism. Drug Metah. Dispos., 24, 15-22... 

In animal studies acetone has been found to potentiate the toxicity of other solvents by altering their metabolism through induction of microsomal enzymes, particularly cytochrome P-450. Reported effects include enhancement of the ethanol-induced loss of righting reflex in mice by reduction of the elimination rate of ethanol increased hepatotoxicity of compounds such as carbon tetrachloride and trichloroethylene in the rat potentiation of acrylonitrile toxicity by altering the rate at which it is metabolized to cyanide and potentiation of the neurotoxicity of -hexane by altering the toxicokinetics of its 2,4-hexane-dione metabolite.Because occupationally exposed workers are most often exposed to a mixmre of solvents, use of the rule of additivity may underestimate the effect of combined exposures. ... 

A single dose oral LDso value of approximately 13,000 mg/kg was reported for mice, and 14 daily doses of 625 mg/kg were lethal for 6 of 20 exposed male mice (Hayes et al. 1986). In rats fed carbon tetrachloride in stock diets or protein-free diets, LDso values of 10,200 or 23,400 mg/kg, respectively were reported (McLean and McLean 1966). The authors attributed the difference in sensitivity in animals in this study to protein depletion which has reportedly afforded protection against carbon tetrachloride toxicity. This may result from protein depletion-induced reduction in cytochrome P-450 synthesis, with a consequent diminished metabolic activation of carbon tetrachloride to toxic metabolites. In other studies using rats, an LDso value of approximately 7,500 mg/kg was reported (Pound et al. 1973), while 17/20 animals were killed within 14 days of a single oral gavage exposure to 8,000 mg/kg (Thakore and Mehendale 1991). Doses as low as 400 mg/kg have resulted in the death of cats (Chandler and Chopra 1926). 

Cytochrome P-450 from rat or human liver microsome preparations is inactivated when incubated anaerobically with carbon tetrachloride in the presence of NADPH and an oxygen-scavenging system (Manno et al. 1988 1992). Inactivation involved destruction of the heme tetrapyrrolic structure, and followed pseudo first-order kinetics with fast and slow half lives of 4.0 and 29.8 minutes. When compared with rat liver microsomes, the human preparations were 6-7 times faster at metabolizing carbon tetrachloride, and only about one- eighth as susceptible to suicide inactivation (about 1 enzyme molecule lost for every 196 carbon tetrachloride molecules metabolized). 

Noguchi T, Fong K-L, Lai EK, et al. 1982a. Specificity of a phenobarbital- induced cytochrome P-450 for metabolism of carbon tetrachloride to the trichloromethyl radical. Biochem Pharmacol 31 615- 624. 

The converse is true of drugs requiring metabolic activation for toxicity. For example, paracetamol is less hepatotoxic to newborn than to adult mice, as less is metabolically activated in the neonate. This is due to the lower levels of cytochromes P-450 in neonatal liver (Fig. 5.30). Also involved in this is the hepatic level of glutathione, which is required for detoxication. Although levels of this tripeptide are reduced at birth, development is sufficiently in advance of cytochrome P-450 levels to ensure adequate detoxication (Fig. 5.30). The same effect has been observed with the hepatotoxin bromobenzene. (For further details of paracetamol and bromobenzene see chap. 7.) Similarly, carbon tetrachloride is not hepatotoxic in newborn rats as metabolic activation is required for this toxic effect, and the metabolic capability is low in the neonatal rat. 

The cytochrome P-450 destroyed may be a specific isoenzyme, as is the case with carbon tetrachloride and allylisopropylacetamide (Table 5.28). Indeed, with carbon tetrachloride the isoenzyme destroyed is the one, which is responsible for the metabolic activation (CYP1A2). With allylisopropylacetamide, it is the phenobarbital-inducible form of the enzyme, which is preferentially destroyed as can be seen from Table 5.25. It seems that it is the heme moiety, which is destroyed by the formation of covalent adducts between the reactive metabolite, such as the trichloromethyl radical formed from carbon tetrachloride (see chap. 7), and the porphyrin ring. 

The hepatocytes, or parenchymal cells, represent about 80% of the liver by volume and are the major source of metabolic activity. However, this metabolic activity varies depending on the location of the hepatocyte. Thus, zone 1 hepatocytes are more aerobic and therefore are particularly equipped for pathways such as the p-oxidation of fats, and they also have more GSH and GSH peroxidase. These hepatocytes also contain alcohol dehydrogenase and are able to metabolize allyl alcohol to the toxic metabolite acrolein, which causes necrosis in zone 1. Conversely, zone 3 hepatocytes have a higher level of cytochromes P-450 and NADPH cytochrome P-450 reductase, and lipid synthesis is higher in this area. This may explain why zone 3 is most often damaged, and lipid accumulation is a common response (see "Carbon Tetrachloride," for instance, chap. 7). 

Carbon tetrachloride causes centrilobular liver necrosis and steatosis after acute exposure, and liver cirrhosis, liver tumors, and kidney damage after chronic administration. The mechanism underlying the acute toxicity to the liver involves metabolic activation by cytochrome P-450 to yield a free radical (trichloromethyl free radical). This reacts with unsaturated fatty acids in the membranes of organelles and leads to toxic products of lipid peroxidation including malondialdehyde and hydroxynonenal. This results in hepatocyte necrosis and inhibition of various metabolic processes including protein synthesis. The latter leads to steatosis as a result of inhibition of the synthesis of lipoproteins required for triglyceride export. 

The effects of chromium(III) chloride and sodium chromate(VI) on the hepatotoxicity of carbon tetrachloride exposure to mouse hepatocytes were examined by Tezuka et al. (1995). Primary cultures of mouse hepatocytes were pretreated with 10 or 100 pM chromium for 24 hours followed by exposure to 1-5 mM carbon tetrachloride for up to 1 hour. Chromium(VI) pretreatment significantly reduced the cell toxicity as well as lipid peroxidation caused by carbon tetrachloride. Chromium(III) pretreatment did not have any effect on cell toxicity. About 50% of chromium(VI) was taken up and reduced in the cells by 90% to chromium(III) within 10 minutes. The initial uptake rate of chromium(HI) into cells was greater than 500-fold less than chromium(VI), and only about 5% was absorbed. The protection against carbon tetrachloride damage by chromium(VI) was attributed to its rapid uptake and conversion to chromium(III), and it was determined that chromium(III) acts as a radical scavenger for the free radicals generated by carbon tetrachloride within the cell. Furthermore, chromium(VI) pretreatment reduced the activity of NADPH cytochrome c reductase which metabolizes carbon tetrachloride to reactive species. 

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