New Aspects of Hexobarbital Metabolism: Stereoselective Metabolism, New Metabolic Pathway via GSH Conjugation, and 3-Hydroxyhexobarbital Dehydrogenases

Abstract

Hexobarbital, a short-acting hypnotic, is metabolized to 3'-hydroxyhexobarbital by cytochrome P450, and then to 3'-oxohexobarbital by liver cytosolic dehydrogenase. New methods of separation for hexobarbital and its metabolites by TLC have been developed and applied to study the metabolism of hexobarbital enantiomers and stereoselective metabolism of hexobarbital. (+)-Hexobarbital preferentially was transformed into beta-3'-hydroxyhexobarbital and the (-)-enantiomer preferentially transformed into alpha-3'-hydroxyhexobarbital by rat liver microsomes. Glucuronidation and dehydrogenation of 3'-hydroxyhexobarbital were also stereoselective and the S-configuration at the 3'-position was preferred. Alpha-3'-hydroxyhexobarbital from (-)-hexobarbital and the beta-isomer from (+)-hexobarbital were shown to be preferentially conjugated with glucuronic acid in rabbit urine, and to be preferentially dehydrogenated to form 3'-oxohexobarbital by rabbit and guinea pig 3-hydroxyhexobarbital dehydrogenases. A new metabolic pathway of hexobarbital was found in which 3'-oxohexobarbital reacts with glutathione to form 1,5-dimethylbarbituric acid and a cyclohexenone-glutathione adduct, a novel metabolite. 1,5-dimethylbarbituric acid was excreted into the urine and the cyclohexenone-glutathione adduct into the bile of rats dosed with hexobarbital. 3-hydroxyhexobarbital dehydrogenases that dehydrogenate 3-hydroxyhexobarbital into 3'-oxohexobarbital were purified from the liver cytosol of rabbits, guinea pigs, goats, rats, mice, hamsters, and humans and characterized. These enzymes were monomeric proteins and had molecular weights of about 34500-42000, and used NAD(+) and NADP(+) as cofactors, except for the human enzyme that had a molecular weight of about 58000 and used NAD(+) alone. Each enzyme exhibited its own characteristics. Substrate specificity demonstrated that 3-hydroxyhexobarbital dehydrogenases dehydrogenate not only alpha,beta-unsaturated cyclic and acyclic secondary alcohols but also some 17 beta-, 3 alpha-hydroxysteroids or both, except for the human enzyme. The amino acid sequence of the hamster enzyme indicated that it belongs to the aldo-keto reductase superfamily and hydroxysteroid dehydrogenase subfamily.

Substrates, Inducers, Inhibitors and Structure-Activity Relationships of Human Cytochrome P450 2C9 and Implications in Drug Development

“…(+)-Hexobarbital preferentially was transformed into -3'-hydroxyhexobarbital and the (-)-enantiomer preferentially transformed into -3'-hydroxyhexobarbital in rat liver microsomes [591]. Glucuronidation and dehydrogenation of 3'-hydroxyhexobarbital were also stereo-selective and the S-configuration at the 3'-position was preferred.…”

“…Glucuronidation and dehydrogenation of 3'-hydroxyhexobarbital were also stereo-selective and the S-configuration at the 3'-position was preferred. -3'-hydroxyhexobarbital from (-)-hexobarbital and theisomer from (+)-hexobarbital were preferentially conjugated with glucuronic acid in rabbits, and was preferentially dehydrogenated to yield 3'-oxohexobarbital by cytosolic dehydrogenases in rabbits and guinea pigs [591]. In addition, 3'-oxohexobarbital was conjugated with GSH to form 1,5-dimethylbarbituric acid and a cyclohexenone-glutathione adduct.…”

“…In addition, 3'-oxohexobarbital was conjugated with GSH to form 1,5-dimethylbarbituric acid and a cyclohexenone-glutathione adduct. 1,5-Dimethylbarbituric acid was excreted into the urine and the cyclohexenone-glutathione adduct into the bile of rats dosed with hexobarbital [591].…”

An In Vivo Assay for Elucidating the Importance of Cytochromes P450 for the Ability of a Wild Mammalian Herbivore (Neotoma lepida) to Consume Toxic Plants

“…Alternatively, perhaps Great Basin animals depend more on P450 enzymes to metabolize hexobarbital than the Mojave animals due to having a reduced amount of conjugation pathways for hexobarbital. Previous work has shown that the Great Basin animals have lower activity of the conjugation enzyme glutathione-S-transferase compared to the Mojave animals on both a control and creosote diet (Haley et al 2008), and there is evidence that glutathione conjugation is a method for hexobarbital biotransformation in laboratory rats (Takenoshita and Toki 2004). …”

Enhancing Throughput of Glutathione Adduct Formation Studies and Structural Identification Using a Software-Assisted Workflow Based on High-Resolution Mass Spectrometry (HRMS) Data

“…Ninety-five (95) commercially available compounds were used in this analysis. Aminopyrine [4,12], Amodiaquine [13], Benzbromarone, Ethinylestradiol, Ezetimibe, Tolcapone [8], Acetaminophen, Acetylsalicylic acid, Amitriptyline, Amlodipine, Aripiprazole, Atorvastatin, Bromfenac, Buspirone, Captopril, Carvedilol, Clopidogrel, Citalopram, Diazepam, Diclofenac, Donezepil, Enalapril, Felbamate, Fluoxetine, Furosemide, Gabapentin, Lisinopril, Lorazepam, Memantine, Nifedipine, Olanzapine [4], Quetiapine, Phenytoin, Procainamide, Ranitidine, Sertraline, Simvastatin, Sitagliptin, Sulfamethoxazole, Valsartan, Venlafaxine, Verapamil, Warfarin and Zolpidem [3], Carbamazepine [14], Cilazapril [15], Clozapine [16], Flutamide [17], Imiloxan [18], Moclobemide [19], Nefazodone, Haloperidol, Indomethacin [20], Nevirapine [4,21,22], Probucol [23], Rimonabant [24], Caffeine and Ritonavir [21], Ticlopidine [25], Tienilic-acid [26], Pioglitazone, Rosiglitazone and Troglitazone [27], Dilthiazem, Diphenhydramin, Erythromycin, Imipramine, Tacrine, Valproic-acid, Zafirlukast [28], Alprazolam [29], Baclofen [30], Desipramine [4,31], Hexobarbital [32], Risperidone [33], Gemfibrozil [34], Propranolol and Tamoxifen [2], Sumatriptan [35], Amantadine, Atropine, Chlorpromazine, Dextromethorphan, Fenofibrate, Levofloxacin, Lidocaine, Olmesartan, Pentobarbital, Pindolol, Pravastatin, Prazosin, Tocopherol acetate, Paroxetine [36] and Zomepirac [4].…”