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L R Pohl

Publications and source records attributed to L R Pohl.

120 records · Page 7Linked to original sources
PubMedDate not supplied

Potential metabolic basis for enflurane hepatitis and the apparent cross-sensitization between enflurane and halothane.

Clinical case reports of unexplained hepatic dysfunction following enflurane and isoflurane anesthesia led to the hypothesis that oxidative metabolism of these drugs by cytochromes P-450 produces immunoreactive, covalently bound acylated protein adducts similar to those implicated in the genesis of halothane-induced hepatic necrosis. Microsomal adducts were detected by enzyme-linked immunosorbent assay and immunoblotting techniques utilizing specific anti-trifluoroacetyl (TFA) IgG hapten antibodies in rat liver following enflurane, isoflurane, or halothane administration. Preincubation of the antibodies with microsomes from halothane-pretreated rats or with 500 microM TFA-lysine, markedly inhibited adduct recognition, while preincubation with 500 microM acetyllysine had no effect. The relative amounts of immunoreactive protein adducts formed were halothane much greater than enflurane much greater than isoflurane and correlates directly with the relative extents of metabolism of these agents. These results support the view that acyl metabolites of the volatile anesthetics may become covalently bound to hepatic proteins, thus serving as antigens, and thereby account for the apparent cross-sensitization and idiosyncratic hepatotoxicity reported for these drugs.

Acylation↗
PubMedDate not supplied

Production of a dimer of 2-acetylaminofluorene during the sulfation of N-hydroxy-2-acetylaminofluorene in vitro.

During the sulfation of N-hydroxy-2-acetylaminofluorene (NOH-2AAF) by rat liver 100,000 g supernatant fraction in vitro, an unidentified metabolite is produced which accounts for 22% of the N-OH-2AAF metabolized. This product has been characterized as the 2AAF dimer, 1-(N-2'-fluorenylacetamido-2-acetylaminofluorene) by comparing its TLC, HPLC, UV, and mass spectral properties with a synthetic standard which was prepared from the reaction of N-acetoxy-2-acetylaminofluorene (N-AcO-2AAF) with 2AAF. Increasing amounts of 2AAF added to the incubation mixture of N-OH-[acetyl-14C]2AAF and rat liver 100,000 g supernatant fraction decreased the irreversible binding of 14C to protein, and increased the formation of 2AAF dimer proportionately. This suggests that the 2AAF dimer is formed from the reaction of 2AAF and the electrophilic species produced from the sulfated N-OH-2AAF. In the presence of the 9,000 g fraction of rat liver, the dimer of 2AAF was aroximately 1/25 as active as 2AAF in producing mutations in the Salmonella mutagenesis test system.

2-Acetylaminofluorene↗
PubMedDate not supplied

Glutathione-dependent dechlorination of chloramphenicol by cytosol of rat liver.

The 100,000 g supernatant fraction (cytosol) of rat liver converts chloramphenicol (CAP, RNHCOCHCl2) into two products. Most of the enzyme activity is lost during dialysis of the enzyme preparation overnight, but is restored by addition of glutathione (GSH). Other thiols are not as effective as GSH in restoring the enzyme activity. The formation of the metabolites is not inhibited when incubations are performed under anaerobic conditions. The major metabolite was identified as CAP aldehyde (RNHCOCHO) whereas the minor metabolite was identified as an alkali-unstable derivative of CAP oxamic acid (RNHCOCOOH). Plausible pathways are discussed for the formation of these metabolites.

Animals↗
PubMedDate not supplied

3-Hydroxyacetaminophen: a microsomal metabolite of acetaminophen. Evidence against an epoxide as the reactive metabolite of acetaminophen.

3-Hydroxyacetaminophen has been isolated and identified as a microsomal metabolite of acetaminophen. Analysis of the metabolite by gas chromatography-mass spectrometry revealed that the metabolite had a molecular ion and fragmentation pattern identical to that of authentic 3-hydroxyacetaminophen. Glutathione and ascorbic acid blocked covalent binding of reactive metabolite(s) to protein but did not block the formation of 3-hydroxyacetaminophen. Moreover, epoxide hydrolase did not block covalent binding of the reactive metabolite(s) to protein. Thus, the reactive metabolite apparently is not an epoxide substrate of the hydrolase, nor are 3-hydroxyacetaminophen and the reactive metabolite(s) formed from a common intermediate.

Acetaminophen↗
PubMedDate not supplied

Mechanism of glutathione-dependent dechlorination of chloramphenicol and thiamphenicol by cytosol of rat liver.

Chloramphenicol (CAP, RNHCOCHCl2) has previously been shown to be dechlorinated to CAP aldehyde (RNHCOCHO) and CAP oxamic acid (RNHCOCO2H) by rat liver cytosol. In the present study we have further characterized these reactions and have found that several homogeneous rat liver GSH transferases, particularly transferases A, metabolize CAP to CAP aldehyde by an apparent hydrolytic dechlorination mechanism. The aldehyde is further metabolized to CAP oxamic acid by an aldehyde oxidizing enzyme(s) which does not require GSH, but can utilize either NAD+ or NADP+. Thiamphenicol, the p-methylsulfonylphenyl derivative of CAP, also appears to be metabolized through these pathways, but to a lesser extent than is CAP.

Animals↗
PubMedDate not supplied

Mechanism of defluorination of enflurane. Identification of an organic metabolite in rat and man.

Difluoromethoxydifluoroacetic acid (CHF2OCF2CO2H) has been identified as a metabolite of enflurane (CHF2OCF2CHCIF) in rat liver microsomes in vitro and in human urine by gas chromatography mass spectrometry. The formation of the metabolite in rat liver microsomes was dependent upon the presence of NADPH and O2, and was inhibited when SKF 525-A or CO/O2 (8:2, v/v) were present in the reaction mixture. When the C-H bonds of the CHCIF group of enflurane or of the CHCI group of isoflurane (CHF2OCHCICF3) were replaced with a C-CI bond, virtually no fluoride ion was produced from either derivative in rat liver microsomes. These results indicate that cytochrome P-450 catalyzes the oxidative dehalogenation of CHF2OCF2CHCIF at its CHCIF group to form CHF2OCF2CO2H and chloride and fluoride ions. In contrast, the CHF2 group does not appear to be appreciably susceptible to metabolic oxidative dehalogenation. These results can be used for the more rational design of new inhalation anesthetics that would not be appreciably metabolized to the potential kidney toxin, F-.

Animals↗
PubMedDate not supplied

The formation of diglutathionyl dithiocarbonate as a metabolite of chloroform, bromotrichloromethane, and carbon tetrachloride.

One hour after the intraperitoneal administration of CHCl3, CBrCl3, or CCl4 to phenobarbital (PB)-treated rats, hepatic GSH levels decreased to 30, 59, and 88% of control levels, respectively; after 4 hr, the GSH levels had returned to 46, 65, 99%, respectively, of control levels. When incubated for 15 min in air with rat liver microsomes from PB-treated rats, a NADPH-generating system, and GSH (5 mM), all of the compounds were converted to diglutathionyl dithiocarbonate (GSCOSG). The rate of conversion of CHCl3, CBrCl3, and CCl4 to GSCOSG was 180, 58, and 8 nmol per mg of protein per 15 min, respectively. The GSCOSG was also identified in bile by 13C-NMR spectroscopy and HPLC as an in vivo metabolite of CHCl(3), CBrCl3, and CCl4. After the administration of CHCl3, CBrCl3, and CCl4, 2.89, 0.64, or 0.11 mumol of GSCOSG, respectively, was excreted in 6 hr. These results suggest that CHCl3, CBrCl3, and CCl4 are metabolized in vitro and in vivo to phosgene (COCl2), which reacts with GSH to produce GSCOSG. The reaction of GSH with COCl2 may be responsible at least in part for the GSH-depleting properties of CHCl3, CBrCl3, and CCl4, inasmuch as the relative amounts of formation of GSCOSG in vitro and in vivo paralleled their relative GSH-depleting activities.

Animals↗
PubMedDate not supplied

Influence of the intestinal microflora on the elimination of warfarin in the rat.

The effect of the intestinal microflora on the half-life and elimination of warfarin in rats was examined. When the intestinal microflora was reduced with neomycin, bacitracin, and tetracycline, or was nonexistent as in germ-free animals, more radioactivity was found in feces and less in the urine after ip administration of 14C-warfarin. The ratio of conjugated to free metabolites in the feces was higher in germ-free rats compared to conventional or ex-germ-free animals. In addition, fecal beta-glucuronidase levels were markedly decreased in antibiotic-treated rats and in germ-free rats when compared to conventional and ex-germ-free rats. In a crossover study, a 30% decrease in warfarin half-life was observed in germ-free and antibiotic-treated animals compared to the same rats in an ex-germ-free state. The antibiotic treatment, however, had effects other than reduction of the microflora. A significant decrease in the volume of distribution of warfarin was noted in antibiotic-treated animals which may invalidate the use of this widely used mixture as a model for the study of intestinal microflora-drug interactions. To confirm enterohepatic recycling of warfarin, bile from donor rats administered 14C-warfarin ip was infused into the upper duodenum of recipient rats. Bile from the recipient rats was shown to contain 0.1-0.9% of the radioactivity administered to the donor rats. At 6-8 hr after injection, serum of the recipient rats contained 2-10% of the radioactivity present in the serum from donor rats, and contained mainly free warfarin. These data are consistent with an important role for the intestinal microflora in facilitating enterohepatic recycling of warfarin in the rat.

Animals↗
PubMedDate not supplied

A new pathway for the oxidative metabolism of chloramphenicol by rat liver microsomes.

When chloramphenicol was incubated with rat liver microsomes, four previously unidentified metabolites were detected and identified. They include chloramphenicol aldehyde (chloramphenicol with the primary alcohol group oxidized to an aldehyde group), p-nitro-benzyl alcohol, N-(2-oxoethyl)dichloroacetamide, and N-(2-hydroxyethyl)dichloroacetamide. The formation of these metabolites was dependent upon the presence of NADPH and O2 and was inhibited when SKF 525-A or CO/O2 (8:2, v/v) were present in the reaction mixture. Moreover, the metabolites were formed by liver microsomes from phenobarbital-treated rats but not by microsomes from untreated rats or rats treated with beta-naphthoflavone. The formation of these metabolites is consistent with a mechanism that involves an initial oxidation of chloramphenicol to chloramphenicol aldehyde by cytochrome P-450. Inasmuch as this metabolite is a beta-hydroxyaldehyde, it can chemically undergo a retro-aldol cleavage to p-nitrobenzaldehyde and N-(2-oxoethyl)dichloroacetamide. Enzymatic reduction of these aldehyde intermediates would yield p-nitrobenzyl alcohol and N-(2-hydroxyethyl)dichloroacetamide, respectively.

Animals↗
PubMedDate not supplied

The mechanism of formation of o-bromophenol from bromobenzene.

Both o-bromophenol and p-bromophenol are formed from bromobenzene in rat liver microsomes. It has been established that p-bromophenol is formed via bromobenzene-3,4-oxide, but o-bromophenol could conceivably arise via either the 2,3-epoxide or the 1,2-epoxide or by direct insertion of oxygen. As described in the present article, we have isolated and identified bromobenzene 2,3-dihydrodiol as a microsomal metabolite of bromobenzene. Identification of the dihydrodiol therefore indicates the formation of its obligatory precursor, bromobenzene-2,3-oxide. Moreover, using bromo(2,4,6-2H3)benzene, we have clarified the mechanism of formation of o-bromophenol from bromobenzene. The rate of formation of o-bromophenol from bromobenzene and bromo(2,4,6-2H3)benzene in liver microsomes from 3-methylcholanthrene-treated rats was 0.72 +/- 0.02 and 0.74 +/- 0.06 nmol/mg/min (kH/kD = 0.99), respectively. The lack of a significant isotope effect indicates that the hydroxylation of bromobenzene to o-bromophenol is not by a direct insertion mechanism. Furthermore, the mass spectrum of o-bromophenol isolated from a microsomal incubation with bromo(2,4,6-2H3)benzene indicated that 70% of the product retained all three deuterium atoms. These results are consistent with the view that o-bromophenol is formed from the 2,3-epoxide intermediate but do not preclude formation by the addition of oxygen to the 2-position carbons followed by an NIH shift and rearrangement before an epoxide is formed.

Animals↗
PubMedDate not supplied

Strain and sex differences in chloroform-induced nephrotoxicity. Different rates of metabolism of chloroform to phosgene by the mouse kidney.

It has been known for many years that there are species, strain, and sex differences in the incidence and severity of the nephrotoxicity caused by chloroform. However, the molecular basis for these differences has not been clearly understood. In this investigation, we have found that sensitivity to CHCl3 correlates with the capacity of the kidney to metabolize CHCl3 to the toxic metabolite phosgene (COCl2). For example, kidney homogenates of sensitive male DBA/2J mice metabolized CHCl3 to COCl2 more rapidly than did the less sensitive C57BL/6J mice. Similarly, kidney homogenates from male mice, which are sensitive to CHCl3-induced nephrotoxicity, metabolized CHCl3 to COCl2 at nearly an order of magnitude more rapidly than did those from female mice. Treatment of female mice with testosterone, however, reversed this trend. Cytochrome P-450 in the microsomal and mitochondrial fraction of the kidney appeared to catalyze the metabolism of CHCl3 to COCl2.

Animals↗