细胞色素P450与兽药的研发应用

细胞色素P450（Cytochrome P450，CYP）是一种广泛存在于生物体内含亚铁血红素的超家族蛋白酶，由于其还原态与CO结合的复合物在波长450nm处有最大吸收峰，故名之。CYP决定着生物体内外源性化合物（如激素、药物、环境污染物等）的生物转化过程。研究CYP是新兽药研发、兽医临床合理用药的新课题，对兽药残留研究也具有重大的指导意义。     

Adverse drug interactions of importance in veterinary practice

Multiple drug therapy is often used in veterinary practice; occasionally, inappropriate combinations of drugs may be administered which interact to produce adverse responses. Such interactions may occur in vitro because of physically or chemically incompatible drugs having been mixed before administration or in vivo. A number of important in vitro incompatibilities concerning parenteral drugs are tabulated, consideration having been given both to problems attending the use of intravenous fluids as vehicles for their administration and to incompatibilities between the drugs themselves. Interactions which occur in vivo are also examined; these have been grouped according to the various mechanisms by which they occur.     

Research Progress on Cytochrome P450 2E1

Cytochrome P450 2E1 (CYP2E1) is one of the body's metabolite of drugs and low molecular weight compounds mainly enzymes, and the increasing of activity or expression of CYP2E1 is closely related with the clinical variety of diseases. This paper reviews the research progress of protein structure, function, positioning mechanism and other aspects of genetic polymorphisms of CYP2E1 at home and abroad. Thereby the paper could lay the foundation for further exploration of the biological effects of CYP2E1 and its relationship with clinical disease.     

Cytochrome P450 3A, NADPH cytochrome P450 reductase and cytochrome b5 in the upper airways in horse

Gene and protein expression as well as catalytic activity of cytochrome P450 (CYP) 3A were studied in the nasal olfactory and respiratory mucosa and the tracheal mucosa of the horse. We also examined the activity of NADPH cytochrome P450 reductase (NADPH P450 reductase), the amount of cytochrome b(5) and the total CYP content in these tissues. Comparative values for the above were obtained using liver as a control. The CYP3A related catalytic activity in the tissues of the upper airways was considerably higher than in the liver. The CYP3A gene and protein expression, on the other hand, was higher in the liver than in the upper airway tissues. Thus, the pattern of CYP3A metabolic activity does not correlate with the CYP3A gene and protein expression. Our results showed that the activity of NADPH P450 reductase and the level of cytochrome b(5) in the relation to the gene and protein expression of CYP3A were higher in the tissues of the upper airways than in the liver. It is concluded that CYP3A related metabolism in horse is not solely dependent on the expression of the enzyme but also on adequate levels of NADPH P450 reductase and cytochrome b(5).     

Cytochrome P450-mediated hepatic metabolism of new fluorescent substrates in cats and dogs

This study aimed to investigate the biotransformation of cat liver microsomes in comparison to dogs and humans using a high throughput method with fluorescent substrates and classical inhibitors specific for certain isozymes of the human cytochrome P450 (CYP) enzyme family. The metabolic activities associated with CYP1A, CYP2B, CYP2C, CYP2D, CYP2E and CYP3A were measured. Cat liver microsomes metabolized all substrates selected for the assessment of cytochrome P450 activity. The activities associated with CYP3A and CYP2B were higher than the activities of the other measured CYPs. Substrate selectivity could be demonstrated by inhibition studies with α-naphthoflavone (CYP1A), tranylcypromine/quercetine (CYP2C), quinidine (CYP2D), diethyldithiocarbamic acid (CYP2E) and ketoconazole (CYP3A) respectively. Other prototypical inhibitors used for characterization of human CYP activities such as furafylline (CYP1A), tranylcypromine (CYP2B) and sulfaphenazole (CYP2C) did not show significant effects in cat and dog liver microsomes. Moreover, IC50-values of cat CYPs differed from dog and human CYPs underlining the interspecies differences. Gender differences were observed in the oxidation of 7-ethoxy-4-trifluoromethylcoumarin (CYP2B) and 3-[2-(N, N-diethyl-N-methylamino)ethyl]-7-methoxy-4-methylcoumarin (CYP2D), which were significantly higher in male cats than in females. Conversely, oxidation of the substrates dibenzylfluorescein (CYP2C) and 7-methoxy-4-trifluoromethylcoumarin (CYP2E) showed significant higher activities in females than in male cats. Overall CYP-activities in cat liver microsomes were lower than in those from dogs or humans, except for CYP2B. The presented difference between feline and canine CYP-activities are useful to establish dose corrections for feline patients of intensively metabolized drugs licensed for dogs or humans.     

家蚕氟中毒后中肠NADPH-细胞色素P450还原酶和NADPH-细胞色素C还原酶活性的变化

为了探究家蚕对NaF的代谢机制,以家蚕耐氟品种T6和氟化物敏感品种734为材料,从5龄起蚕开始分别添食50、100、200、400 mg/kg NaF溶液处理后的桑叶,检测蚕体中肠微粒体酶液中的黄素蛋白NADPH-细胞色素P450还原酶(CPR)和NADPH-细胞色素C还原酶(CR)的活性变化。氟物化敏感品种734的4个NaF处理组第3天的中肠CPR活性低于对照组,其余时间几乎都高于对照组,400 mg/kg NaF处理组在第4天的CPR活性最高,且各NaF处理组的CPR活性差异显著(P<0.05);耐氟品种T6 NaF处理组和对照组的中肠CPR活性整体上呈先升后降的趋势,几乎都在第2天达到最高值,各组之间的差异不显著(P>0.05)。氟化物敏感品种734的4个NaF处理组的中肠CR活性呈现先升后降的趋势,而对照组呈下降趋势;耐氟品种T6的50、100、400 mg/kg NaF处理组在第1～2天的中肠CR活性呈明显下降趋势,之后的变化相对较小,对照组的CR活性仅在第3～4天略高于NaF处理组,而其余时间NaF处理组的CR活性较高。2个家蚕品种添食不同浓度NaF后的中肠CR活性差异均不显著(P>0.05)。试验结果显示,耐氟品种T6在NaF作用下中肠的CPR和CR活性变化范围(分别为对照组的0.4～2.0倍和0.3～2.9倍)远小于氟化物敏感品种734这2种酶的活性变化范围(分别为对照组的0.6～9.3倍和0.4～4.6倍)。初步推测CPR和CR与家蚕对氟化物代谢具有一定的关联。     

恩诺沙星对细胞色素P450酶系影响的研究进展

恩诺沙星是一种抗菌药物,广泛应用于畜禽养殖业和水产养殖业。随着恩诺沙星使用量的不断增加,在其对细胞色素P450酶系的影响方面已有广泛的研究。介绍了细胞色素P450酶的基本特征,综述了恩诺沙星对细胞色素P450酶系影响的研究进展,以期为进一步阐明恩诺沙星与细胞色素P450酶系的相互作用机制及恩诺沙星在畜禽体内的药理学与毒理学特征提供理论依据。     

Analysis of polymorphisms of canine Cytochrome P 450-CYP2D15

Cytochrome P450 (CYP) proteins constitute a large ancient family of oxidative enzymes essential for the efficient elimination of a wide variety of clinically used drugs. Polymorphic variants of human CYP2D6 are associated with the conversion rate and efficacy of several drugs such as antidepressants. Polymorphisms of the canine orthologue CYP2D15 are of interest because these antidepressants are also used in dogs with behavioral problems and the outcome of the treatment is variable. However, the annotated CYP2D15 gene is incomplete and inaccurately assembled in CanFam3.1, hampering DNA sequence analysis of the gene in individual dogs. We elucidated the complete exon–intron structure of CYP2D15 to enable comprehensive genotyping of the gene using genomic DNA. We surveyed variations of the gene in four diverse dog breeds and identified novel polymorphisms in exon 2 in border collies. Further investigation to establish the impact of these canine CYP2D15 polymorphisms on interindividual variability in expression and function of this metabolizing enzyme is now feasible. Further knowledge of CYP pharmacogenetics will help individualize therapy and thereby increase therapeutic efficacy, especially in the use of antidepressants in veterinary behavioral medicine.     

兽药GMP在企业实施应用中的思考

通过兽药GMP认证,并不意味着以后便高枕无忧了。兽药GMP认证只是农业部规范我国兽药企业的一道门坎。企业通过GMP认证后,更应该时时刻刻以GMP标准来严格要求自己,这样,才能达到规范我国兽药生产的真正目的。     

Novobiocin Inhibits Both UDP-Glucuronosyltransferase and Cytochrome P450-Mediated Enzyme Activities in Pig Liver Microsomes

The effects of novobiocin (range 0.0125–2 mmol/L) on the hydroxylation of testosterone, the N-demethylation of erythromycin, and the glucuronidation of -naphthol and paracetamol were studied using pig hepatic microsomes, pooled from five animals. The final concentrations of these substrates in the incubation mixtures were selected to meet V_max conditions. Novobiocin caused a concentration-dependent inhibition of the glucuronidation of paracetamol; the formation of -naphthol-glucuronide was reduced to a lesser degree. These results confirm and extend earlier findings in laboratory animal species that novobiocin inhibits UDP-glucuronosyltransferases (UDPGTs). Moreover, novobiocin strongly inhibited 6-hydroxylation of testosterone. The microsomal N-demethylation of erythromycin and hydroxylation of testosterone at the 15 position were less affected by novobiocin. These results suggest that novobiocin inhibits not only UDPGTs, but also cytochrome P450 (CYP) enzyme activities, probably those belonging to the CYP3A subfamily. More research is needed to reveal which CYPs and UDPGTs are affected by novobiocin in vivo, in order to improve the understanding and probably the predictability of potential drug interactions with this antibiotic.     

姜黄素对奶牛子宫内膜炎症细胞CYP450表达的影响

为了探讨姜黄素对奶牛子宫内膜炎症细胞的作用机理,分离奶牛的子宫内膜细胞,用细菌脂多糖(LPS)制备子宫内膜炎症细胞模型,ELISA法检测细胞上清液中IL-lβ和TNF-α的浓度变化。姜黄素处理子宫内膜炎症细胞24 h、48 h,Western Blot检测子宫内膜炎症细胞CYP450的表达。结果表明,100 ng/mL的LPS对子宫内膜细胞抑制率最接近10%,模型组IL-lβ和TNF-α的含量显著高于对照组(P<0.01),这一剂量为诱导炎症细胞模型的最佳浓度。姜黄素处理子宫内膜炎症细胞48 h后,随姜黄素浓度的升高,CYP450的表达呈升高的趋势。5 mg/mL的姜黄素为最佳作用剂量。     

The pharmacogenomics of P-glycoprotein and its role in veterinary medicine

Despite advancements in pharmacogenetics in human medicine, the incorporation of pharmacogenetics into veterinary medicine is still in its early stages of development. To date, efforts to understand the pharmacologic impact of genetic variation in veterinary species have largely focused on genes encoding for the membrane transporter, P-glycoprotein (P-gp). The emphasis on the role of P-gp is largely because of safety concerns associated with the use of some macrocyclic lactones in dogs. Because of the body of information available on this topic, we use P-gp as a platform for understanding the importance of population diversity in veterinary medicine. The impact of P-gp on drug pharmacokinetics and pharmacodynamics is considered, along with endogenous and exogenous factors that can modulate P-gp activity. The review includes discussion of how population diversity in P-gp activity can lead to susceptibility to certain diseases or alter patient response to environmental stress or pharmaceutical intervention. In addition, phenotypic diversity also needs to be considered, as demonstrated by the impact of P-gp up-regulation and drug resistance. The aim of this review was to set the stage for further exploration into the impact of genetic and phenotypic variability on drug pharmacokinetics, disease propensity, product formulation and drug response in both companion and food-producing animals.     

大观霉素的研究进展及其在畜牧业中的应用

大观霉素是一种氨基糖苷类抗生素,因毒副作用小,且在动物体内几乎不残留,成为兽医用药的首选品种之一.本文对大观霉素的菌种选育、生物合成、检测方法及其在畜牧业中的应用前景等方面进行了较为全面的综述.     

Characterization of cytochrome P450-mediated drug metabolism in cats

In this study we examined activities of cytochrome P450 (CYP)1A, 2C, 2D and 3A using hepatic microsomes from five male and five female cats. CYP1A, 2C, 2D and 3A activities were referred by ethoxyresorufin O-deethylation (EROD), tolbutamide hydroxylation (TBH), bufuralol 1'-hydroxylation (BLH) and midazolam 1'- and 4-hydroxylation respectively. The anti-rat CYP1A2 and CYP3A2 serum significantly inhibited EROD and midazolam 1'- and 4-hydroxylation, suggesting that EROD and midazolam 1'- and 4-hydroxylation were catalysed by CYP1A and 3A in cats respectively. Quinidine inhibited BLH in cats microsomes at quite low concentrations, suggesting that BLH was catalysed by CYP2D in cats. Tolbutamide hydroxylation activities were negligible in hepatic microsomes from both male and female cats, suggesting CYP2C activities of cats are extremely low. This suggests that CYP2C substrates should be carefully administered to cats. Although there is no sexual difference in CYP1A activities, there are differences in CYP2D and 3A activities of cats. CYP2D activities were higher (3-fold), but CYP3A activities were lower (one-fifth) in female cats. These results might suggest that CYP2D and 3A substrates should be prescribed for male and female cats using different dosage regimen.     

Cytochrome P450 enzymes mediated by DNA methylation is involved in deoxynivalenol-induced hepatoxicity in piglets

Deoxynivalenol(DON) is an inevitable contaminant in animal feed and can lead to liver damage, then decreasing appetite and causing growth retardation in piglets. Although many molecular mechanisms are related to hepatoxicity caused by DON, few studies have been done on cytochrome P450(CYP450)enzymes and DNA methylation. To explore the role of CYP450 enzymes and DNA methylation in DONinduced liver injury, male piglets were fed a control diet, or diet containing 1.0 or 3.0 mg/kg DON for4 weeks. DO...     

论中西兽药的结合与应用

兽用中草药是指天然的植物、动物和矿物类药物,它是我国的独特药物,在我国尤其是我省其资源丰富、品种繁多.长期以来,它是我国人民用以和畜禽疾病作斗争的重要武器.西兽药尤其是抗感染类西药,是一类作用强、见效快的药物,它被广泛用于兽医临床.     

Macrolide antibiotics, drug interactions and microsomal enzymes: implications for veterinary medicine

The macrolide group of antibiotics includes natural members, pro-drugs and semi-synthetic derivatives, thus named because they are composed of a large aglycone ring (from 14 to 16 carbon atoms), to which are attached several sugars. Some of them are amino-sugars, containing a diethylamino, tertiary amine function. A number of antibiotics, including erythromycin, oleandomycin, triacetyl-oleandomycin (troleandomycin), carbomycin, spiramycin, tylosin, rosamicin, azithromycin, clarithromycin, dirithromycin and others, are members of this group. On a comparative basis, erythromycin and oleandomycin are similar, with the same basic 14-carbon lactone ring and side chain sugars. The remaining compounds contain a basic 15- or 16-carbon lactone ring and one or two side-chain sugars. Most of the macrolides are produced by Streptomyces spp bacteria. An exception is rosamicin, which is produced by Micromonospora. Clarithromycin and azithromycin are new semi-synthetic derivatives of erythromycin.