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五种中药提取物对拟穴青蟹肝微粒体CYP1A的抑制作用   总被引:1,自引:0,他引:1  
利用肝微粒体体外孵育体系,通过酶促反应动力学研究黄芩、柴胡、连翘、吴茱萸和野菊花5种中药提取物对拟穴青蟹(Scylla paramamosain)肝微粒体CYP1A的抑制作用。方法:5种中草药提取物分别与拟穴青蟹肝微粒体共同孵育,荧光分光光度法测定探针药物7-乙氧基异吩唑酮代谢产物试卤灵的生成量,来反映中药提取物对CYP1A活性的影响,计算酶促反应动力学常数Vmax和Km,推算表示抑制程度的IC50。结果:5种中草药提取物对EROD的抑制程度为黄芩>野菊花>连翘>柴胡>吴茱萸,IC50依次为0.46、0.80、1.07、3.99和17.79 mg/mL,其中,黄芩、柴胡和吴茱萸对EROD的抑制为竞争性抑制,野菊花则是反竞争性抑制,而连翘的抑制类型则是混合型。酶促反应动力学常数Vmax和Km为31.25 pmol/(min.mg)和1.02μM。黄芩、柴胡、连翘、吴茱萸和野菊花的抑制常数分别为0.13、0.38、3.39、1.74和70 mg/mL。结论:5种中药提取物对拟穴青蟹CYP1A均有不同程度的抑制作用,当它们与一些化学药物作为处方药搭配使用时可能产生药物相互作用,引起药效的改变。  相似文献
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Cytochrome P450 (CYP) 1A1 participates in the activation as well as detoxification of environmental pollutants such as aromatic hydrocarbons. This CYP form is also efficiently induced by aromatic hydrocarbons. The presence of CYP 1A1 in the brain might thus be of physiological and toxicological importance. In the present investigation on rainbow trout, the distribution of 7-ethoxyresorufin-O-deethylase (EROD) activity, a cytochrome CYP 1A1 catalyzed reaction, was measured in whole tissue homogenates from brain parts. In control fish, a relatively high activity was found in the rainbow trout olfactory bulb compared to the other brain parts. Although an EROD induction (3 to 7-fold) by β-naphthoflavone (BNF) was recorded in all brain parts from the rainbow trout, the highest induced activity was measured in the olfactory bulbs. To ascertain the distribution of EROD activity in cells, whole brain tissue was subfractionated by differential centrifugation. The fractionation scheme separated mitochondria (P2 fraction) and microsomes (P3 fraction) as determined by marker enzymes and electron microscopy. In control rainbow trout, a low EROD activity could be measured in the P2 fraction. BNF induced the EROD activity in both P2 and P3 fractions. Western blotting showed the induction by BNF of a protein band in the P2 and P3 fractions with a molecular mass around 58,000 when highly specific anti-cod CYP 1A1 antibodies were used. ELISA measurements confirmed the induction of CYP 1A1 protein in the rainbow trout brain subcellular fractions.  相似文献
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Shyamal C  MAHATA  Ryoichi  MITSUO  Jun-Ya  AOKI  Hironori  KATO  Takao  ITAKURA 《Fisheries Science》2003,69(3):615-624
ABSTRACT:     The cytochrome P450 (CYP) represents a large group of microsomal monooxygenases that catalyze drugs as well as a host of lethal environmental contaminants such as dioxins, leading to either detoxification and excretion from the animal or generation of carcinogenic intermediates. In the present study two forms of cDNA were cloned (Eu MC1 and Eu MC2) for European eel CYP1A genes by polymerase chain reaction (PCR) techniques. The cDNA of Eu MC1 was 3368 bp long coding 521 amino acid residues, and that of Eu MC2 was 2464 bp long coding 517 amino acid residues. Identities of deduced amino acid sequences between Eu MC1 and Japanese eel CYP1A1 and that between Eu MC2 and the second form of Japanese eel CYP1A were 98% and 97%, respectively, showing decisively that Eu MC1 and Eu MC2 are orthologous to Japanese eel CYP1A1 and the second form of CYP1A, respectively. A striking difference between the two eel species was that the Eu MC1 peptide was two amino acid residues longer than that of the Japanese eel CYP1A1. Existence of two loci of CYP1A in Japanese and European eels may suggest that the two forms of CYP1A exist widely among the eel species, because the divergence between the two eel species has been shown to be close to the basal divergence among eels. The identities in CYP1A may help to estimate genetic distance between European and Japanese eels.  相似文献
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林茂  杨先乐  纪荣兴 《水产学报》2010,34(6):685-690
为了建立以CYP1A cDNA为探针的水环境毒理学细胞模型,以β-萘黄酮(BNF)作为诱导剂,通过半定量PCR技术研究CYP1A在草鱼不同细胞系及其对应组织中的诱导表达情况。在半定量PCR反应参数研究中,对关键的退火温度和循环次数进行了优化,结果显示退火温度为57 ℃,循环次数为30次较为合适,该条件下Gauss迹量的CYP1A/ACT比值能更准确的反映CYP1A的表达水平。对照组和诱导组草鱼细胞中ACT和CYP1A cDNA扩增结果表明,细胞中CYP1A的基础表达量较低,而BNF诱导使GCL、CIK和CO细胞中CYP1A的表达水平得到显著的提高。GCL、CIK和CO细胞三者之间诱导后CYP1A的表达水平有显著差异(P<0.05),诱导表达量大小顺序为GCL>CIK>CO。草鱼细胞系的相应组织中ACTCYP1A cDNA扩增以及电泳的结果与细胞中较为相似,组织中CYP1A诱导表达量大小顺序也与相应的细胞相同:肝>肾>卵巢。比较分析的结果表明,CYP1A在体内与体外的诱导表达水平具有一定的相关性。  相似文献
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本研究以中药黄芩苷为受试药物,研究其在不同剂量和作用时间下对牙鲆(Paralichthys olivaceus)肝CYP1A酶活性和基因表达的影响.采用体内实验,将黄芩苷按低(50 mg/kg·d)、中(100mg/kg·d)、高(200 mg/kg·d)3个剂量分别口灌牙鲆,连续给药,并分别于给药3d、6d、9d后取样,测定CYP1A酶活性.结果表明,较空白组而言,给药3d,黄芩苷各剂量组鱼肝中EROD酶(CYP1A标志酶)活性没有明显变化(P>0.05);给药6d,高剂量组的EROD酶活性极显著增加(P<0.01),中剂量组的则显著增加(P<0.05);给药9 d,高、中剂量组的EROD酶活性均极显著增加(P<0.01),低剂最组的EROD酶活性显著增加(P<0.05),且黄芩苷对该酶的诱导作用与剂量和药物作用时间均呈正相关.半定量RT-PCR结果表明,各药物剂量组的CYP1A基因表达水平都发生了上调,且这种表达上调的幅度同CYP1A酶的活性一样,随给药时间和剂量的增加而加强,提示黄芩苷作为诱导剂对CYP1A酶活性的作用机制可能与调控CYP1A的转录水平有关.  相似文献
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