氮烯乙茶在成年大鼠肝细胞中的生物转化

目的:建立成年大鼠肝细胞分离培养方法,探讨抗癌新药氮烯乙茶在肝细胞内生物转化。方法:用肝细胞活率、乳酸脱氢酶、能荷值变化观察肝细胞悬浮培养方法的稳定性,以高效液相色谱法测定肝细胞悬液中氮烯乙茶及其代谢产物的浓度。结果:氮烯乙茶在大鼠肝细胞内生成代谢产物7-乙基-8-氨基茶碱(7-ethyl-8-aminotheophylline,EAT) ,其最大反应速度Vm 为12-80 nmol·min^-1/106 cells,米氏常数Km 为640-9 μmol·L^-1 ,氮烯乙茶的固有清除率CLint为20-0 μL·min^-1/106 cells。结论:肝细胞培养的方法是考察药物生物转化的较好体外模型,同时氮烯乙茶具有降低肝细胞能荷值的作用。     

五味子醇甲在大鼠肝微粒体内的代谢动力学和性别差异

体外研究五味子醇甲(schizandrin，SZ)在大鼠肝微粒体内的代谢动力学和性别差异。制备正常雌、雄大鼠肝微粒体，与SZ共同温孵，以高效液相色谱法测定SZ及其代谢产物。SZ在雄鼠肝微粒体内代谢反应的最大速率V_max、米氏常数K_m和清除率Cl_int分别为(21.88±2.30) μmol·L^-1·min^-1·mg^-1(protein)，(389.00±46.26) μmol·L^-1和(0.056 3±0.000 7) min·mg^-1(protein)；在雌鼠肝微粒体内代谢反应的最大速率V_max、米氏常数K_m和清除率Clint_{分别为(0.61±0.07) μmol·L^-1·min^-1·mg^-1(protein)，(72.64±13.61) μmol·L^-1和(0.008 4±0.000 8) min·mg^-1(protein)，雌、雄鼠肝微粒体内SZ的主要代谢物不同，分别为7,8-顺二羟基五味子醇甲(M1)和7,8-顺二羟基-2-去甲基五味子醇甲(M2b)。酮康唑、奎尼丁和奥芬得林对SZ的在雌、雄大鼠肝微粒体内代谢均有不同程度的抑制作用，西咪替丁对其在雄鼠肝微粒体内的代谢也有一定的抑制作用。SZ在雌、雄大鼠肝微粒体中代谢动力学及代谢产物存在明显的性别差异，这种差异可能主要是由CYP3A和CYP2C11在大鼠肝微粒体内的性别差异引起的。}     

MN9202在Beagle犬肝微粒体酶中的代谢动力学

目的研究MN9202在Beagle犬肝微粒体酶中的代谢。方法差速离心法制备Beagle犬肝微粒体酶，0.4 μmol·L^-1的MN9202与1.0 g·L^-1的肝微粒体酶在37 ℃水浴中孵育30 min，加入0.5 mL碱化液终止反应，然后采用RP-HPLC法测定孵育液中MN9202原形药物的浓度。根据所测浓度与反应速度做Lineweave-Brurk双倒数曲线，推导出药物的米氏常数K_m和最大反应速度V_max,并计算机体内在清除率。同时观察不同浓度和不同种类的人肝微粒体酶(CYP450)抑制剂对MN9202代谢的影响。结果MN9202在Beagle犬肝微粒体酶中的K_m为(22.6±8.0) μmol·L^-1；V_max为(0.54±0.17) μmol·g^-1·min^-1；CL_int为(0.024 2±0.000 9) L·g^-1·min^-1。醋竹桃霉素(Tro)和酮康唑(Ket)能够显著抑制MN9202的代谢；反苯环丙胺(Tra)对MN9202的代谢也有一定的抑制作用，而其他CYP450抑制剂对MN9202的代谢无明显影响。结论CYP3A和CYP2C19参与了MN9202的代谢，人CYP3A和CYP2C19的抑制剂可能使MN9202的代谢受到抑制，造成药物的药效或毒性的增加。     

治疗黑色素瘤的新型二芳基衍生物04022a在不同种属肝微粒体中的稳定性及代谢产物分析

目的 采用SD大鼠、人、比格犬的肝微粒体酶,考察新型二芳基衍生物04022a在3个种属肝微粒体中的代谢稳定性,比较代谢的种属差异,并对产生的代谢产物进行筛选与结构表征。方法 将04022a溶解在由NADPH启动的肝微粒体孵育体系中,孵育0 、15 、30 、60 、90 、120 min后,加入等体积的冰甲醇终止反应。采用高效液相色谱法测定04022a的质量浓度,计算各个时间点的剩余药物百分比、体外半衰期（t_1/2）和固有清除率（CL_int）。采用超高效液相色谱串联飞行时间质谱法对04022a在肝微粒体中的代谢产物进行检测与分析。结果 孵育120 min时,04022a在大鼠、人、比格犬肝微粒体中的剩余药物百分比分别为88.30%、82.15%、98.63%。t_1/2分别为770、364、3 465 min,CL_int分别为0.001 8、0.003 8、0.000 4 mL•（min•mg）^-1。初步判断04022a在3个种属共有5个体外代谢产物。结论 04022a在各种肝微粒体中的代谢稳定性良好,主要代谢转化途径包括脱烷基化、羟基化、脱氢。     

细胞色素P450介导的氯沙坦钾对瑞格列奈在大鼠肝微粒体中代谢的影响

目的 研究瑞格列奈在大鼠肝微粒体中的酶促反应动力学，并考察氯沙坦钾对其在大鼠肝微粒体中代谢的影响。方法 建立大鼠肝微粒体体外孵育体系对瑞格列奈的代谢进行研究；以洛伐他汀为内标，应用UPLC测定大鼠肝微粒体中瑞格列奈的浓度。采用底物减少法，通过GraphPad Prism 5.0软件计算瑞格列奈的酶促反应动力学常数V_max和K_m；分别以系列浓度氯沙坦钾（2.5~50μmol·L^-1）与瑞格列奈（44 μmol·L^-1）于37℃水浴中共同孵育，并测定肝微粒体中瑞格列奈的减少量，考察氯沙坦钾对瑞格列奈的抑制作用。结果 瑞格列奈在大鼠肝微粒体的最佳孵育时间为40 min，最佳蛋白质量浓度为1 mg·mL^-1；瑞格列奈酶促反应动力学参数V_max=47.29μmol·min^-1·（mg·protein）^-1，K_m=51.41 μmol·L^-1；氯沙坦钾对瑞格列奈在体外肝微粒体抑制作用的IC₅₀值为17.89 μmol·L^-1。结论 氯沙坦钾对瑞格列奈在大鼠肝微粒体中的代谢具有较强的抑制作用，两药联合应用可能发生相互作用，具有诱发低血糖的风险。     

噻吩诺啡在人、比格犬和大鼠肝微粒体中体外代谢比较

采用体外肝微粒体孵育体系, 研究噻吩诺啡在大鼠、比格犬和人肝微粒体中酶代谢动力学及代谢产物差异。通过对噻吩诺啡浓度、微粒体蛋白含量和孵育时间等条件的考察优化噻吩诺啡与肝微粒体的反应体系; 应用LC-MS/MS定量检测孵育体系中的噻吩诺啡及代谢产物, 分析比较噻吩诺啡在3种肝微粒体中代谢产物种类和生成量的差异, 计算并比较相应的动力学参数。噻吩诺啡在人肝微粒体中代谢转化最慢, 其相应的动力学参数K_m = (4.00 ± 0.59) µmol·L⁻¹、V_max = (0.21 ± 0.06) µmol·L⁻¹·min⁻¹、T_1/2 = (223 ± 6.10) min、CL_int = (117 ± 3.19) mL·min⁻¹·kg⁻¹; 比格犬和大鼠肝微粒体中相应的参数K_m、V_max、T_1/2和CL_int分别为 (3.57 ± 0.69) 和 (3.28 ± 0.50) µmol·L⁻¹、(0.18 ± 0.04) 和 (0.14 ± 0.04) µmol·L⁻¹·min⁻¹、(244 ± 1.21) 和 (70.7 ± 1.05) min、(213 ± 1.06) 和    (527 ± 7.79) mL·min⁻¹·kg⁻¹。在3个种属肝微粒体中均观察到噻吩诺啡的6个I相代谢产物, 但6个产物的相对生成百分比在不同种属肝微粒体中有一定差异。实验结果表明, 噻吩诺啡在体外人、比格犬和大鼠肝微粒体中主要的I相代谢途径相同, 但是代谢产物的生成量及噻吩诺啡的代谢动力学性质存在着一定的差异。     

藏、汉族健康人对安替比林及醋氨酚代谢的种族差异

用随机交叉设计,研究了藏、汉族健康人安替比林和醋氨酚的代谢差异。16例汉族人安替比林T_1/2为18.8±4.89h,17例藏族人为15.0±3.02h(P<0.05),汉族人安替比林清除率是1.35±0.37L·h^-1,藏族人是1.71±0.44Lt·h^-1(P<0.05)。汉族人醋氨酚T_1/2是3.1±0.86h,藏族人是3.2±1.12h(P>0.05)。实验结果表明,藏、汉族健康人安替比林的代谢有显著差异,藏族人安替比林清除率比汉族人增加了20%。藏、汉族醋氨酚的代谢无明显差异。     

地非三唑的RP-HPLC法测定及其体外代谢研究

目的为研究抗早孕新药地非三唑(DL111-IT)的代谢作用机理和进一步开发利用,建立其体外代谢的RP-HPLC测定法。方法以Lichrospher ODS-C₁₈为色谱柱,甲醇-pH 7.5磷酸盐缓冲液(70∶30)为流动相,检测波长:235 nm,流速1.0 mL·min^-1,地西泮为内标,对鼠肝微粒体孵育液中DL111-IT的RP-HPLC法进行方法学研究。应用建立的方法对DL111-IT在不同来源的鼠肝微粒体中的体外代谢进行试验。结果DL111-IT浓度在1.01～101.0 μg·mL^-1呈良好的线性关系,在不同浓度下测得平均绝对回收率和相对回收率分别为(92±4)%和(100.3±1.9)%(N=5);DL111-IT在鼠肝微粒体中的代谢, β-萘黄酮组明显快于其他组。结论本法简便、准确,可用于DL111-IT的体外代谢研究。     

芫花主要黄酮成分对肝微粒体UGTs及UGT1A1活性的体外抑制作用

目的 考察芫花主要黄酮成分芫花素和芹菜素对尿苷二磷酸葡萄糖醛酸转移酶（UGTs）及UGT1A1活性的影响。方法 采用体外肝微粒体孵育模型,以4-硝基酚（4-nitrophenol,4-NP）为底物检测UGTs活性,胆红素为底物检测UGT1A1活性;利用UV及UPLC-MS/MS测定底物或代谢产物的含量。结果 对UGTs,在大鼠肝微粒体、小鼠肝微粒体以及人肝微粒体孵育体系中,芫花素和芹菜素均能不同程度地抑制UTGs活性;抑制强弱顺序：在大鼠肝微粒体温孵体系中,芫花素>芹菜素;在小鼠肝微粒体以及人肝微粒体温孵体系中,芹菜素>芫花素。对UGT1A1,在人肝微粒体孵育体系中,芫花素和芹菜素均表现为中等强度的竞争性抑制作用,抑制强弱顺序：芹菜素（IC₅₀=12.40 μmol·L^-1）>芫花素（IC₅₀=23.21 μmol·L^-1）。结论 芫花素和芹菜素对不同肝微粒体孵育体系中UGTs及UGT1A1均可产生显著抑制作用且存在种属差异。芫花素及芹菜素可能存在基于UGT酶的药物相互作用。     

蛇床子素在兔体内药物代谢动力学

目的研究蛇床子素在兔体内的药物代谢动力学。方法用高效液相色谱法,以丹皮酚为内标,以甲醇-水(80∶20)为流动相,测定兔血液中蛇床子素(iv,10 mg·kg^-1)的含量。采用3P87程序计算药物代谢动力学参数。结果蛇床子素iv药代动力学符合二房室开放模型,T_1/2α=5.81 min,T_1/2β=42.2 min,K21=0.036 0·min^-1,K₁₂=0.045 0·min^-1,K₁₀=0.054 0·min^-1,AUC=235 mg·min·L^-1,CLs=0.043 0 L·min^-1·kg^-1,V_C=0.780 L·kg^-1。结论蛇床子素在兔体内分布及消除较快     

The Formation of Desethyl-Piperacillin from Piperacillin by Human Liver S9 in vitro

Piperacillin (PIPC) has been used as one of the most useful β-lactam antibiotics over the past 10 years. The metabolism of PIPC has been thoroughly investigated and it has been recognized that PIPC gives few metabolites in laboratory species or humans. Recently, an active metabolite, desethyl-piperacillin (DEt-PIPC), was detected in human plasma and urine after PIPC administration. In the current study, human tissues were obtained from organ donors (n = 3) and subcellular fractions (S9) were prepared. The time course of metabolism by S9 mix from liver, kidney cortex, and kidney medulla was then determined using 0·5 mM PIPC. For comparative purposes, rat liver S9 were also prepared and incubated with PIPC under the same conditions. DEt-PIPC was formed by human liver S9 mix from all three specimens studied, with the rate varying approximately eightfold. No DEt-PIPC was detected in any of the incubations with rat liver S9 mix (n = 3) and kidney S9 mix (n = 3) prepared from either the cortex or medulla. In summary, these data suggest that the formation of the unique human metabolite, DEt-PIPC, can be predicted by in vitro studies with human tissues and that this metabolite is formed predominantly by the liver. © 1997 John Wiley & Sons, Ltd.     

Comparative in vitro cytotoxicity of cyclophosphamide,its major active metabolites and the new oxazaphosphorine ASTA Z 7557 (INN mafosfamide)

Summary Cyclophosphamide (CPA), the most commonly used alkylating agent in the treatment of a wide variety of hematologic and solid tumors, requires oxidation by hepatic microsomal enzymes to its active alkylating species. A number of alternative methods exist to simulate the in vitro cytotoxicity of CPA against animal and human tumors, including the co-incubation of CPA with the S-9 fraction of rat liver homogenates (S-9) and the use of either 4-hydroperoxy CPA (a stabilized form of a major blood-borne metabolite of CPA), phosphoramide mustard (PM, considered to be the ultimate intracellular alkylating metabolite of CPA), or ASTA Z 7557 [4-(2-sulfonatoethylthio)-CPA, a new oxazaphosphorine compound which after dissolution undergoes rapid spontaneous hydrolysis in vitro with liberation of 4-hydroxy-CPA]. Using a human tumor clonogenic assay (HTCA) we have quantitated the median molar inhibitory dose 50 (ID₅₀) concentrations of S-9 activated-CPA, 4-hydroperoxy-CPA, PM, and ASTA Z 7557 against 107 previously untreated tumors, as well as determining the in vitro biological stability of the former three CPA metabolite preparations. 4-Hydroperoxy-CPA proved the most consistently cytotoxic (median molar ID₅₀=5.77#x00D7;10^–5M) compound, followed by ASTA Z 7557, S-9 activated-CPA and PM in that order. Of additional interest S-9 activated CPA and PM proved relatively unstable biologically when frozen at -120°C, whereas 4-hydroperoxy-CPA lost none of its cytotoxicity over a 36 day period during freezing. On the basis of these data 4-hydroperoxy-CPA appears the compound of choice for use in vitro to evaluate the activity that CPA is likely to express clinically against solid tumors. Since 4-hydroperoxy-CPA is not available for clinical use, ASTA Z 7557, which was slightly less cytotoxic to ovarian cancers and a wide variety of other tumors in the HTCA, appears an attractive agent to develop further clinically, especially for regional chemotherapy (e.g., intraperitoneal and intra-arterial treatment) of solid tumors.     

Caffeine metabolism in liver slices during postnatal development in the rat

The metabolism of caffeine was investigated in liver slices of young and adult rats. Liver slices from adult male rats metabolized caffeine at an initial rate of 48.31 ± 3.71 nmoles · (g liver)^?1 · hr^?1 to four main metabolite fractions. By a combination of thin-layer radiochromatography and high performance liquid chromatography, theophylline, paraxanthine and 1, 3, 7-trimethyldihydrouric acid were identified as caffeine metabolites. Apparent V_max of the overall reaction was 83.30 nmoles caffeine metabolites formed · (g liver)^?1 · hr^?1. Theophylline competitively inhibited caffeine metabolism [the apparent K_m was 19.20, μM in the absence of theophylline, the apparent k_i was 36.50 μM in the presence of theophylline (100 μM)]. SKF 525-A inhibited caffeine metabolism; the formation of all of the metabolite fractions was inhibited to a similar extent. Allopurinol (100 μM) had no effect. The specific activity of the enzyme system was extremely low when liver slices of 2-day-old-rats were used [1.46 ± 0.08 nmoles caffeine metabolites formed · (g liver)^?1 · hr^?1]; the reaction velocity increased gradually with increasing age and reached a peak [52.26 ± 1.41 nmoles caffeine metabolites formed · (g liver)^?1 · hr^?1]at 30 days of age. Changes in the formation of the four metabolite fractions with age followed the pattern of the overall caffeine metabolism. These results demonstrate that the liver of the newborn rat has an extremely limited capacity to metabolize caffeine in vitro and are consistent with the proposed involvement of the liver microsomal cytochromes P-450 monooxygenase system in the metabolism of caffeine. N-Demethylation is the main pathway of in vitro caffeine metabolism in the rat liver at all ages.     

An in vitro approach to investigate ocular metabolism of a topical,selective β1-adrenergic blocking agent,betaxolol

Abstract

1. Topical glaucoma treatments have often been limited by poor absorption and bioavailability. Betaxolol, a selective β1-blocker, has been well studied for its pharmacokinetics and disposition. Limited ocular, betaxolol metabolism data is available despite a growing number of novel ocular treatments.

2. In vitro ocular fractions indicated the formation of an active metabolite, across rat, rabbit and human, which was only observed historically in the liver.

3. Ocular metabolic profiles of preclinical toxicology species, rat and rabbit, were not predictive of human in vitro ocular data. M1 was specific to human and only captured by the liver data.

4. Liver S9 over predicted the extent of ocular metabolism compared to ocular fractions. Rabbit liver S9 fractions demonstrated extensive glucuronidation and higher parent turn-over in 1?h as compared to other matrices.

5. This research assesses in vitro species and organ differences across preclinical species and human. The complex data set highlights the need for an in vitro ocular system to explore poorly documented ocular metabolism.     

Species Variation and Stereoselectivity in the Metabolism of Nicotine Enantiomers

1. High performance liquid radiochromatographic systems have been developed for the identification and quantification of 7 urinary metabolites of both S-(-)-[³H-N'-CH₃]nicotine and R-(+)-[³H-N'-CH₃] nicotine in guinea pig, hamster, rat and rabbit.

2. 3′-Hydroxycotinine was a major urinary metabolite of both S-(-)-nicotine and R-(+)-nicotine in guinea pig, hamster and rabbit. Cotinine was not generally a significant urinary metabolite of either nicotine enantiomer, except in rat, where it constituted 14·6 and 10·4%, respectively, of the total radiolabel in the urine after administration of [³H]-S-(-)-nicotine or [³H]-R-(+)-nicotine. Nicotine N'-oxide was an important urinary metabolite of both nicotine isomers in guinea pig and rat, but in both cases, was not observable in hamster and rabbit. No N-methylated urinary metabolite of S-(-)-nicotine could be detected in any of the species examined. In R-(+)-nicotine experiments, only guinea pig afforded N-methylated metabolites. Significant amounts of 2 unidentified polar, non-basic urinary metabolites of both S-(-)- and R-(+)-nicotine-treated animals were observed.

3. Analysis of the comparative metabolism of the nicotine enantiomers in the four animals species studied, showed that stereoselective differences in the formation of oxidative metabolites existed, particularly in the formation of 3′-hydroxycotinine and nicotine-N'-oxide. A clear stereospecificity was observed in the guinea pig, in that only the R-(+)-nicotine enantiomer was N-methylated in this species.

4. Sex differences appear to exist in the metabolism of nicotine enantiomers in the rat. Female rats excreted more of the unidentified polar metabolite B than male rats, whereas the converse was true for nicotine-N'-oxide. In experiments with R-(+)-nicotine, urinary levels of 3′-hydroxycotinine and R-(+)-nicotine in female rats were higher than in male rats. Conversely, higher amounts of nicotine-N'-oxide were observed in the urine of male rats compared to those in female rats.     

In vitro inhibition and enhancement of liver microsomal S-777469 metabolism by long-chain fatty acids and serum albumin: insight into in vitro and in vivo discrepancy of metabolite formation in humans

1.?It was previously demonstrated that 10% of S-777469, a cannabinoid receptor 2 selective agonist, is metabolized to its carboxylic acid metabolite (S-777469 5-carboxylic acid, 5-CA) in humans in vivo, while the formation of 5-CA is extremely low in human cryopreserved hepatocytes and liver microsomes (HLMs). In this study, factors causing the different metabolite formation rates of S-777469 in vitro and in vivo were investigated.

2.?Formation of 5-CA and S-777469 5-hydroxymethyl (5-HM), a precursor metabolite of 5-CA, was catalyzed by CYP2C9. Arachidonic acid, α-linolenic acid, oleic acid and myristic acid, which have been reported to exist in liver microsomes, inhibited S-777469 oxidation by CYP2C9, but serum albumin enhanced this reactions.

3.?The IC₅₀ values of these fatty acids for 5-CA formation from 5-HM were lower than those of 5-HM formation from S-777469. Serum albumin extensively enhanced 5-CA formation from 5-HM in comparison to 5-HM formation from S-777469.

4.?CYP2C9 was the enzyme responsible for S-777469 oxidation in human livers. The suppressive effects of several fatty acids and enhancing action of serum albumin in vitro are likely to be the causal factors for the apparently different rates of in vitro and in vivo metabolite formation of S-777469.     

Metabolism and disposition of a potent and selective JNK inhibitor [14C]tanzisertib following oral administration to rats,dogs and humans

Abstract

1. The disposition of tanzisertib [(1S,4R)-4-(9-((S)tetrahydrofuran-3-yl)-8-(2,4,6-trifluorophenylamino)-9H-purin-2-ylamino) cyclohexanol], a potent, orally active c-Jun amino-terminal kinase inhibitor intended for treatment of fibrotic diseases was studied in rats, dogs and humans following a single oral dose of [¹⁴C]tanzisertib (Independent Investigational Review Board Inc., Plantation, FL).

2. Administered dose was quantitatively recovered in all species and feces/bile was the major route of elimination. Tanzisertib was rapidly absorbed (T_max: 1–2?h) across all species with unchanged tanzisertib representing >83% of plasma radioactivity in dogs and humans, whereas <34% was observed in rats. Variable amounts of unchanged tanzisertib (1.5–32% of dose) was recovered in urine/feces across all species, the highest in human feces.

3. Metabolic profiling revealed that tanzisertib was primarily metabolized via oxidation and conjugation pathways, but extensively metabolized in rats relative to dogs/humans. CC-418424 (S-cis isomer of tanzisertib) was the major plasma metabolite in rats (38.4–46.4% of plasma radioactivity), while the predominant plasma metabolite in humans and dogs was M18 (tanzisertib-/CC-418424 glucuronide), representing 7.7 and 3.2% of plasma radioactivity, respectively. Prevalent biliary metabolite in rats and dogs, M18 represented 16.8 and 17.1% of dose, respectively.

4. In vitro studies using liver subcellular fractions and expressed enzymes characterized involvement of novel human aldo-keto reductases for oxido-reduction and UDP-glucuronosyltransferases for conjugation pathways.     

The Metabolism of Talampicillin in Rat,Dog and Man

1. After administration of [phthalidyl-¹⁴]talampicillin (Talpen® to rat. dog and man, radioactivity was excreted mainly in the urine (90%, 86% and 98% in rat, dog and man respectively).

2. After administration of [ampicillin-¹⁴C]talampicillin, radioactivity was excreted in the urine of rats and dogs to a lesser extent (35% in both species) and only a small proportion of the dose was excreted in the bile (6% in rats, less than 0·1% in dogs).

3. The pattern of radiometaboletes was very similar in extracts of the urines of rat, dog and man dosed orally with [phthalidyl-14C]talampicillin. The major metabolite was 2-hydroxymethylbenzoic acid.

4. Unchanged talampicillin was present in the hepatic portal vein blood of dog and thus reached the liver, whereas in rat, no parent compound could be detected in portal vein blood. This result may help to explain differences in toxicity of the compound in rat and dog.

5. Studies in vitro showed that the intestinal wall is an important site of hydrolysis of talampicillin in rat and dog.     

In vitro hepatic biotransformation of moclobemide (Ro 11-1163) in man and rat

1. Moclobemide, an inhibitor of monoamine oxidase, shows mixed MAO A/B inhibition in rat, but pure MAO A inhibition in man. This is attributed to a primary amine metabolite which inhibits MAO B in vitro, but which is not detected in human plasma in vivo. A secondary amine metabolite, also present in rat but not human plasma, inhibits MAO B in vivo but not in vitro.

2. We have studied the biotransformation of moclobemide in vitro, to investigate whether hepatocytes and hepatic subcellular fractions can reproduce the in vivo interspecies differences.

3. Moclobemide was more extensively metabolized by rat liver preparations. compared with man. For example, of an initial 100nmol, 78 and 25 nmol were metabolized within 24?h by rat and human hepatocytes in primary culture, respectively.

4. Substantial amounts of secondary amine (12.5 nmol) were found with the rat preparation, compared with low amounts (1.5 nmol) from human hepatocytes. Similarly, for the primary amine, 1.5 nmol were formed by the rat hepatocytes compared with trace amount in the human preparations.

5. Identities of the two amines were confirmed by?h.p.l.c. cochromatography and negative Cl GC-MS.

6. In conclusion, all the in vitro models, but particularly hepatocytes, reflected the metabolism of moclobemide in vivo. Consequently, liver preparations can be used prospectively to screen the selectivity of related development compounds.