Metabolism Of Halothane Research Articles

Biotransformation and toxicity of inhalational anaesthetics

In summary, anaesthetics and drugs used perioperatively are all xenobiotics and can be metabolized mainly by microsomal enzyme systems, which have a high activity in the liver. These enzyme systems are induced by repeated pre-administration of drugs, such as barbiturates and others which are used during the preoperative period. However, according to some reports, aerobic and anaerobic metabolism is inhibited by the simultaneous administration of drugs, such as isoflurane and halothane, halothane and enflurane, and cimetidine and halothane. Hypoxia is also an important factor in hepatic disorders and it is well known that anaerobic metabolism of halothane is increased by hypoxia and its intermediate production produces a free radical. Theoretically, this free radical is involved in hepatic disorders. In practice, in order to prevent hepatic dysfunction before, during and after anaesthesia, hypoxia and repeated pre-administration of enzyme-inducing drugs should be avoided. However, the choice and combination of drugs which inhibit drug metabolism and prevent hepato and/or nephro toxicity should be examined by further investigation.

HALOTHANE METABOLISM IN CHILDREN

Halothane (1% v/v inspired) was administered for 60 min to six children of mean age 74 months (range 14-119 months). Uptake of halothane was measured from the difference in the concentration in inspired and expired gas and varied from 176 to 310 mg kg-1, depending on minute ventilation. After administration of halothane ceased, its elimination in expired gas was measured in four patients until the conclusion of anaesthesia; 32-37% of the absorbed halothane was expired 90 min after halothane administration ceased. Urinary excretion of trifluoroacetic acid, fluoride and bromide was measured for up to 1 week. Of the absorbed halothane, 11.4% (range 6.3-18.2%) was excreted in urine as trifluoroacetic acid and 0.37% (range 0.10-0.64%) as inorganic fluoride. The urinary half-life of trifluoracetic acid was 41.8 h (range 10.4-59.1 h). The quantitative and qualitative metabolism of halothane via the reductive and oxidative pathways in children are comparable to values found in adults. No differences in the metabolism of halothane by children were found which would explain the different incidence of halothane-associated hepatitis compared with adults.

Structure Identification of Free Radicals by Esr And Gc/Ms of Pbn Spin Adducts From theIn Vitroandin vivoRat Liver Metabolism Of Halothane

Free radicals were detected from the in vitro metabolism of halothane (rat liver microsomes) by the PBN spin trapping method. The detected radical species include the 1-chloro-2,2,2-trifluoro-1-ethyl radical (I), as determined by mass spectral analysis, and lipid-type radicals assigned by high resolution ESR spectroscopy with the use of d14-deuterated PBN. The lipid-derived radicals are a carbon-centred radical with the partially assigned structure CH2R and an oxygen-centred radical of the OR' type. From the mass spectral analysis of the spin adduct mixture there is also evidence for a halocarbon double adduct of PBN of the type I-PBN-I.

Free radical metabolism of halothane In vivo: Detection of radical adducts in bile

DECPO, a new analogue of EMPO was synthesized through a two-step synthetic pathway. Its structure and its application to trap superoxide were investigated. The ESR detection of the DECPO–OOH spin adduct is easy even at low concentration of superoxide. In comparison with DEPMPO, the trapping of superoxide with DECPO is faster and the detection of DECPO–OOH can be performed using a very low nitrone concentration (0.5 mM).

Halothane-Induced Liver Injury as a Consequence of Enhanced Microsomal Lipid Peroxidation in Guinea Pigs

We investigated the role of microsomal lipid peroxidation in halothane hepatotoxicity in guinea pigs. Animals were exposed to halothane, isoflurane or enflurane. Enhancement of microsomal lipid peroxidation was specific to halothane. The time-course of lipid peroxidation and hepatic damage following a single exposure to halothane was investigated by measuring the thiobarbituric acid (TBA)reactive products, serum transaminase activity, reduced glutathione (GSH) concentration and histopathological examination. Microsomal lipid peroxidation was enhanced most rapidly and preceded GSH depletion and hepatic injury. Metyrapone, an inhibitor of cytochrome P-450, and N-tert-butyl-alpha-phenylnitrone (BPN), a radical trapping agent, inhibited halothane-induced lipid peroxidation and the incidence and severity of liver injury. The metabolism of halothane was considered to be inhibited by metyrapone and the reactivity of radical intermediates was considered to be decreased by BPN. Microsomal lipid peroxidation was initiated by radical metabolites of halothane but did not result from GSH depletion. Inhibition of microsomal lipid peroxidation reduced the halothane hepatotoxicity. These results demonstrated that the halothane-induced liver injury was caused by microsomal lipid peroxidation in guinea pigs.

The role of oxidative biotransformation of halothane in the guinea pig model of halothane-associated hepatotoxicity.

The role of the oxidative pathway of halothane biotransformation in mediating the hepatotoxicity of halothane in the guinea pig was examined by utilizing the deuterated form of halothane (d-halothane), which is resistant to oxidative metabolism. Male outbred Hartley guinea pigs were exposed to either 1% v/v halothane or d-halothane, FIO2 = 0.21, for 4 h. Significant reductions in both oxidative and overall halothane biotransformation were observed with the use of d-halothane as indicated by decreased plasma levels of trifluoroacetic acid and bromide ion, respectively, immediately following exposure. Plasma fluoride ion, indicative of the reductive metabolism of halothane, was significantly increased with the use of d-halothane. These changes in metabolism were accompanied by a reduced hepatotoxic response as indicated by significantly decreased plasma ALT levels 24-96 h following exposure and a significantly lesser incidence of centrilobular necrosis. Thus, the oxidative biotransformation of halothane is implicated as a mechanism of injury in guinea pigs.

Halothane Hepatotoxicity and Reductive Metabolism of Halothane in Acute Experimental Liver Injury in Rats

Reductive metabolism of halothane was measured after acute liver injury induced by galactosamine (1.0 g/kg, IP) in rats. On the seventh day of liver injury, when previously elevated serum alanine aminotransferase levels had returned to near normal range, anaerobic release of fluoride from halothane by hepatic microsomes, which appears to reflect the reductive pathway of halothane metabolism, was still remarkably decreased (1.36 +/- 0.56 nmol/mg protein/h vs 5.88 +/- 0.58 in controls, P less than 0.001). In another set of experiments, rats (n = 8) given galactosamine 7 days earlier and saline-treated control rats were given halothane anesthesia (1.0%) under mildly hypoxic conditions (F1O2 0.14). In saline controls, halothane anesthesia resulted in a mild but statistically significant increase in serum alanine aminotransferase levels (32 +/- 4 vs 59 +/- 6 U/ml, P less than 0.001). In contrast, serum levels of this enzyme were not changed by halothane anesthesia in galactosamine-treated rats (45 +/- 3 vs 49 +/- 4 U/ml). Although care should be taken in extrapolating the importance of these animal data to humans, the results of this study suggest that halothane hepatotoxicity can be attenuated in the presence of minor liver injury as a result of decreased hepatic biotransformation of the anesthetic. The data support the view that halothane anesthesia is not necessarily contraindicated in subjects with impaired liver function.

Risk factors for halothane hepatitis.

Helothane hepatitis is a rare but sometimes fatal complication of halothane anaesthesia. Examination of case reports has pointed to a number of risk factors. Studies in animals and humans in the laboratory have provided evidence of a complex multifactorial basis for halothane hepatotoxicity, with the following factors playing a part: genetic predisposition; metabolism of halothane; repeated halothane anaesthetics; female sex; age of patient; intrahepatic hypoxia; and enzyme induction. Immunologic changes can be detected in a high percentage of cases of halothane hepatitis; however, studies establishing a cause-effect relationship are not available to determine if these changes cause, or result from, hepatic damage.

A gas chromatographic mass spectrometric method for the analysis of trifluoroacetic acid: application to the metabolism of halothane by in vitro preparations.

A selected ion monitoring gas chromatographic mass spectrometric assay for trifluoroacetic acid was developed for the study of the metabolism of the volatile anesthetic agent halothane by in vitro preparations. The assay uses a headspace sampling technique after formation of methyl esters with dimethyl sulfate and sulfuric acid. Pentafluoropropionic acid proved to be a suitable internal standard, although care is required in the preparation of the calibration standards so that they reflect the composition and treatment of the samples. Contamination of the samples, possibly with trifluoroacetic acid itself, was found to be the primary factor in limiting the sensitivity of the assay for the metabolism of halothane. Generally, trifluoroacetic acid could be determined at levels as low as 1 microM in 50-100 microliters of incubate.

Hepatic Protection from Chemical Injury by Isoflurane

Isoflurane inhibits oxidative metabolism of halothane. Because hepatotoxicity of chemicals may be associated with their metabolism, whether isoflurane can protect the liver against chemical injury was investigated. Hepatic injury was produced in female F344 rats by a 30-minute exposure to 250 ppm of carbon tetrachloride. In this and all other parts of the study, the inspired oxygen concentration was maintained at 21%. The injury was accompanied by elevated activity of liver enzymes in serum (SGOT, SGPT, and SDH), enlarged liver, fatty infiltration of the liver, and vacuolar degeneration of hepatocytes. These signs of toxicity were partly or completely suppressed by concurrent exposure to subanesthetic concentrations of isoflurane (0.2 or 0.038%, respectively). The protective effect was concentration-dependent. Enflurane was protective, but less so than isoflurane. Nitrous oxide and fentanyl had no protective effect.

Effects of piperonyl butoxide on halothane hepatotoxicity and metabolism in the hyperthyroid rat

A series of experiments were conducted to examine the potential role of phase I metabolism in halothane-induced liver injury in the hyperthyroid rat. The metabolism of halothane was determined in both hyperthyroid (triidothyronine, 3 mg/kg per day, for 6 days) and euthyroid rats and in animals pre-treated with the cytochrome P-450 inhibitor piperonyl butoxide (75–100 mg/kg, i.p.). It was found that the hyperthyroid state, which is associated with a substantial increase in sensitivity to the hepatotoxic effects of halothane, decreases both oxidative and reductive routes of halothane metabolism in the rat. The production of trifluoroacetic acid (TFA), an oxidative metabolite, as well as that of chlorodifluoroethylene CDF) and chlorotrifluoroethane (CTF), 2 reductive metabolites, was significantly reduced in hyperthyroid animals. Consistent with these findings serum and urinary bromide levels resulting from the formation of TFA, CDF or CTF were significantly reduced. The only route of halothane metabolism significantly increased by the hyperthyroid condition was the defluorination of halothane. Piperonyl butoxide administration did not render euthyroid animals sensitive to the halothane-induced hepatotoxicity and had no effect on the defluorination of halothane in euthyroid animals. However, piperonyl butoxide markedly increased the hepatotoxicity of halothane in hyperthyroid rats and, except for a modest increase in debromination reactions, decreased all measured indices of halothane metabolism including the defluorination of halothane. Thus, none of the observed changes in halothane metabolism produced by triidothyronine or piperonyl butoxide treatment could be consistently correlated to the increases in hepatotoxicity linked to these 2 treatments. Based on these studies we suggest that the halothane hepatotoxicity induced in the hyperthyroid rat results from effects produced by either the parent compound or an as yet unidentified metabolite. In addition, these studies further demonstrate that considerable mechanistic differences exist for halothane-induced hepatotoxicity when comparing euthyroid and hyperthyroid animal models.

I-653 Resists Degradation in Rats

The ability of rats pretreated with phenobarbital to metabolize a new volatile anesthetic, I-653, was compared with the metabolism of halothane, isoflurane, and methoxyflurane. Each anesthetic was administered for 2 hours at 1.6 MAC (inspired). Control rats were given phenobarbital but not exposed to an anesthetic. In rats pretreated with phenobarbital and exposed to I-653, fluoride ion concentrations in serum and excretion of fluoride ion and organic fluoride in the urine were almost indistinguishable from values measured in control rats. In contrast, rats pretreated with phenobarbital metabolized small but significant amounts of isoflurane. In rats pretreated with ethanol and exposed to I-653, the 24-hour excretion of urinary organic fluoride was nearly ten times greater than that observed in control rats. Marked increases in organic fluoride (as high as 1000 times control values) and/or fluoride ion were found in serum and/or urine after anesthesia of phenobarbital-pretreated rats with halothane or methoxyflurane. The relative stability of I-653 indicates that it may possess minimal toxic properties.

Reductive metabolism of halothane by purified cytochrome P-450

The reductive metabolism of halothane was determined using purified RLM2, PBRLM4 and PBRLM5 forms of rat liver microsomal cytochrome P-450. The metabolites, 2-chloro-1,1,1-trifluoroethane (CTE) and 2-chloro-1,1-difluoroethylene (CDE), were determined. All three forms of cytochrome P-450 produced CTE with relatively small differences in its production among the various forms. There were major differences, however, in the production of CDE, with PBRLM5 being the most active. PBRLM5 was also the only form to show the development of a complex between halothane and cytochrome P-450. This complex absorbed light maximally at 470 nm. The complex formation and the production of CDE by PBRLM5 were stimulated by the addition of cytochrome b 5. Cytochrome b 5 had no effect on CDE production by PBRLM4 and inhibited the production of both CTE and CDE by RLM2. These results show that the two-electron reduction of halothane by cytochrome P-450 was catalyzed by the PBRLM5 form and that cytochrome b 5 stimulated the transfer of the second electron to halothane through PBRLM5, but not RLM2 or PBRLM4.

Halothane Hepatotoxicity and Reductive Metabolism of Halothane in Acute Experimental Liver Injury in Rats

Reductive metabolism of halothane was measured after acute liver injury induced by galactosamine (1.0 g/kg, IP) in rats. On the seventh day of liver injury, when previously elevated scrum alanine aminotransferase levels had returned to near normal range, anaerobic release of fluoride from halothane by hepatic microsomes, which appears to reflect the reductive pathway of halothane metabolism, was still remarkably decreased (1.36 ± 0.56 nmol/mg protein/h vs 5.88 ± 0.58 in controls, P < 0.001). In another set of experiments, rats (n = 8) given galactosamine 7 days earlier and saline-treated control rats were given halothane anesthesia (1.0%) under mildly hypoxic conditions (FIO2, 0.14). In saline controls, halothane anesthesia resulted in a mild but statistically significant increase in serum alanine aminotransferase levels (32 ± 4 vs 59 ± 6 U/ml, P < 0.001). In contrast, serum levels of this enzyme were not changed by halothane anesthesia in galactosarnine-treated rats (45 ± 3 z>s 49 ± 4 U/ml). Although care should be taken in extrapolating the importance of these animal data to humans, the results of this study suggest that halothane hepatotoxicity can be attenuated in the presence of minor liver injury as a result of decreased hepatic bio trans formation of the anesthetic. The data support the view that halothane anesthesia is not necessarily contraindicated in subjects with impaired liver function.

Extrahepatic sites of metabolism of halothane in the rat.

Rats were given 14C-halothane intravenously and whole-body autoradiography with freeze-dried sections, or with sections extracted in trichloroacetic acid, water, and organic solvents, was carried out to trace tissues accumulating halothane metabolites. In vitro incubations of tissue homogenates were performed to examine the capacity by the various organs to form tissue-bound 14C from the 14C-halothane. Autoradiography of isolated organs after incubation with 14C-halothane was performed to study the tissue localization of halothane metabolites formed under in vitro conditions. A localization of halothane metabolites was observed in several extrahepatic tissues in vivo, and the in vitro experiments showed a capacity by the same tissues to transform 14C-halothane to metabolites that bind strongly to tissue components. In addition to the liver, the other tissues shown to have a marked halothane-metabolizing capacity were the nasal mucosa, lateral nasal gland, mucosa of the tongue, cheek, soft palate (but not the hard palate), pharynx, larynx, oesophagus, and the tracheo-bronchial mucosa. The in vivo data obtained indicated a diffusion of the halothane over the walls of the large intestine and the caecum, followed by the formation of apparently reductive metabolites by intestinal microbes and a binding of the metabolites to the intestinal contents. The localization of halothane metabolites in the upper alimentary and respiratory pathways is correlated to the presence of cytochrome P-450 at these sites.

Effects of treatment with phenobarbitone or isoniazid on hepatotoxicity due to prolonged subanaesthetic halothane inhalation.

Rats were exposed to halothane vapour, 50 p.p.m., or air for a period of four weeks. Within each exposure group, some animals drank plain water, some received water plus phenobarbitone, while some received water plus isoniazid. Halothane exposure resulted in increased serum bromide concentrations and liver injury evidenced by increased serum alanine aminotransferase activity, focal hepatocellular necrosis and fatty change. Administration of isoniazid reduced halothane metabolism by 33% as assessed by serum bromide concentrations, and completely blocked the injurious effects of halothane on the liver, suggesting that halothane metabolism plays a role in halothane hepatotoxicity under these conditions. Administration of phenobarbitone partially prevented the increase in serum alanine aminotransferase activity and hepatocellular necrosis due to halothane. In contrast to isoniazid, phenobarbitone led to a slight increase in halothane metabolism. However, phenobarbitone also caused an increase in liver size, such that the amount of halothane metabolised per gram of liver was reduced by phenobarbitone treatment. These results suggest that metabolism of halothane is an important factor in liver injury due to prolonged, subanaesthetic halothane exposure.

A Randomized Prospective Controlled Study of the Metabolism and Hepatotoxicity of Halothane in Humans

COUSINS, M. J.; GOURLAY, G. K.; KNIGHTS, K. M.; LA M. HALL, P. DE; LUNAM, C. A.; O'BRIEN, P. Author Information

Generation of Halothane-Induced Immune Response in a Guinea Pig Model of Halothane Hepatitis

A guinea pig model of halothane hepatitis was used to explore the humoral immune response induced by multiple halothane exposures and the potential role this response might play in contributing to liver damage. Three different strains of guinea pigs (Strain 2, Amana, and Hartley) were exposed to 1% halothane under either 21 or 80% oxygen for 4 hr at 2-week intervals. In each strain, halothane induced the appearance of an antibody cross-reactive with trifluoroacetylated guinea pig serum albumin (TFA-GSA). Three of six Strain 2 guinea pigs demonstrated an association between antibody titer and serum glutamate pyruvate transaminase levels. However, the possible cause and effect relationship between these two factors requires more investigation. Hartley guinea pigs had a 4- to 11-fold higher level of anti-TFA antibody than the other two strains because of either a "higher responder" genetic background or exposure conditions that favored oxidative metabolism of halothane. Immunization of Amana guinea pigs with TFA-GSA evoked a specific anti-TFA antibody response. However, the presence of this antibody before halothane exposure did not potentiate the transient liver damage induced by exposure. Thus, these results demonstrate that in guinea pigs multiple exposures to halothane induce the formation of an antibody that recognizes a reactive intermediate of halothane formed during the anesthetic's metabolism.

Modulation of the reductive metabolism of halothane by microsomal cytochrome b5 in rat liver

To study the modulation of the reductive metabolism of halothane (2-bromo-2-chloro-1,1,1-trifluoroethane) by microsomal cytochrome b 5, formation of 2-chloro-1,1,1-trifluoroethane (CTE) and 2-chloro-1,1-difluoroethylene (CDE), major reduced metabolites of halothane, was analyzed in vivo and in vitro. Rats were pretreated with both malotilate (diisopropyl-1,3-dithiol-2-ylidenemalonate) and sodium phenobarbital (malotilate-treated rats) or only with sodium phenobarbital (control rats). The microsomes of malotilate-treated rats had significantly more cytochrome b 5 than the controls, whereas the cytochrome P-450 content was not different between the two groups. At the end of 2-h exposure to 1% halothane in 14% oxygen, the ratio of CDE to CTE in arterial blood was significantly higher in malotilate-treated rats than in the controls. Under anaerobic conditions, the formation of CDE and the ratio of CDE to CTE were significantly greater in microsomal preparations of malotilate-treated rats than those of the controls. In a reconstituted system containing cytochrome P-450 PB purified from rabbit liver, addition of cytochrome b 5 to the system enhanced the formation of CDE and increased the ratio of CDE to CTE. These results suggested that cytochrome b 5 enhances the formation ratio of CDE to CTE by stimulating the supply of a second electron to cytochrome P-450, which might reduce radical reactions in the reductive metabolism of halothane.

Halothane metabolism in cirrhotic rats.

A rat model was used to determine whether the metabolism of halothane is changed in the presence of cirrhosis and whether exacerbation of liver dysfunction is correlated with such a change. Cirrhosis was produced by gavaging enzyme-induced male Wistar rats with carbon tetrachloride in corn oil once weekly for 12 weeks. Control rats received corn oil only. After a 3-week period without treatment, blood and urine were collected from each rat for determination of background levels of inorganic fluoride, bromide, and trifluoroacetic acid (halothane metabolites) and for assessment of liver function. Rats were then anesthetized with 1.05% halothane in 50% oxygen for 3 h. Following anesthesia, serial blood and urine samples were taken to monitor halothane metabolism and liver function. No differences were observed between cirrhotic and non-cirrhotic rats in serum levels and urinary excretion of halothane metabolites. However, serum levels of SGOT and SGPT were significantly increased about 1.5-fold in the noncirrhotic group and about 2.5-fold in the cirrhotic group after anesthesia. The increased levels observed in the cirrhotic group were significantly greater than in the noncirrhotic group. The results imply that the exacerbation of liver dysfunction after halothane anesthesia is most likely related to an indirect effect, such as change in liver blood flow, rather than to toxic metabolites.