长链烷基化物与牛血清白蛋白相互作用的共振光散射研究

应用荧光分光度计进行共振光散射测定,研究了水相介质中长链烷基化物十二烷基硫酸钠（SLS）与牛血清白蛋白（BSA）的结合平衡。实验表明,在微酸性条件下,SLS与BSA形成的复合物引起共振散射增强,根据470mm的光散射增强信号建立了一定酸度下SLS与BSA结合的数学关系式,随着pH值的变化,二者相互作用的结合平衡常数（Kf）及每分子BSA所结合的SLS个数（n）均不相同。     

Di(2-ethylhexyl)phthalate and mono(2-ethylhexyl)phthalate in media for in vitro fertilization

In vitro fertilization (IVF) is one of the most important treatments of infertility to provide a chance of conceiving. In IVF treatment, sperm are washed and motile sperm are isolated with sperm washing media (SWM) for the purpose of fertilization; fertilized ova are then incubated for a maximum of 5 or 6 d in media for IVF (IVFM). The exposure of fertilized ova to chemicals via such media has not been studied. We determined the concentrations of two contaminants; di(2-ethylhexyl)phthalate (DEHP) and its hydrolyzed product mono(2-ethylhexyl)phthalate (MEHP) in IVFM, SWM, and protein sources (PS: human serum albumin or serum substitute) for IVFM and SWM. The DEHP and MEHP in these media were extracted by a liquid-liquid extraction method and their concentrations determined by liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). Fifteen IVFM, nine SWM, and six PS obtained in Japan were examined. The concentrations of DEHP and MEHP in IVFM and SWM were <10-114 and <2.0-263 ng mL⁻¹, respectively. The concentrations of both DEHP and MEHP were higher in the media containing PS than in those without PS. Either MEHP alone or both DEHP and MEHP were detected in PS. The concentrations of DEHP and MEHP in PS were <10-982 and 47.0-1840 ng mL⁻¹, respectively. The DEHP and MEHP detected in these media were derived from PS. This is the first study on the chemical contamination of IVFM, SWM, and PS.     

荧光光谱法研究全氟辛酸与牛血清白蛋白的相互作用

在模拟生理条件下(pH7.4),采用荧光光谱研究环境污染物全氟辛酸(PFOA)与牛血清白蛋白(BSA)的相互作用.结果表明,PFOA对BSA有荧光静态猝灭作用.PFOA与BSA在温度277,298,310K的反应结合常数分别为1.06×105,7.12×104,5.68×104L/mol;在BSA上有1个PFOA的结合位点,该位点更接近于色氨酸残基.由反应的热力学可推断PFOA与BSA的相互作用力主要是静电引力和疏水作用力.与PFOA结合后,BSA分子构象发生变化,色氨酸残基附近疏水性增强,蛋白质分子结构趋于折叠状态;而酪氨酸残基附近亲水性增强,蛋白质分子结构趋于松散状态.     

In vitro fluorescence displacement investigation of thyroxine transport disruption by bisphenol A

Bisphenol A (BPA) is a chemical with high production volume and wide applications in many industries. Although BPA is known as an endocrine disruptor, its toxic mechanisms have not been fully characterized. Due to its structural similarity to thyroid hormones thyroxine (T4) and triiodothyronine (T3), one possible mechanism of BPA toxicity is disruption of hormone transport by competitive binding with the transport proteins. In this study, the binding interactions of BPA, T4, and T3 with three thyroid hormone transport proteins, human serum albumin (HSA), transthyretin (TTR), and thyroxine-binding globulin (TBG) were investigated by fluorescence measurement. Using two site-specific fluorescence probes dansylamide and dansyl-L-proline, the binding constants of BPA with HSA at drug site I and site II were determined as 2.90 104 and 3.14 104 L/mol, respectively. By monitoring the intrinsic fluorescence of tryptophan, a binding constant of 4.70 103 L/mol was obtained. Similarly, by employing 8-anilino-1-naphthalenesulfonic acid as fluorescence probe, the binding a nity of BPA with TTR and TBG was measured to be 3.10 105 and 5.90 105 L/mol, respectively. In general, BPA showed lower binding a nity with the proteins than T3 did, and even lower a nity than T4. Using these binding constants, the amount of BPA which would bind to the transport proteins in human plasma was estimated. These results suggest that the concentrations of BPA commonly found in human plasma are probably not high enough to interfere with T4 transport.     

Patterns of serum calcium fractions,magnesium, phosphates and albumin in non‐insulin dependent diabetics (Nidd)

Blood level of calcium fractions (ionized and bound), phosphates, magnesium and albumin were measured in non‐insulin dependent diabetic patients (NIDD). Results were compared with healthy controls to elucidate the pathogenesis of probable changes of these elements. Total ionized calcium and magnesium showed significant decrease (p<0.001, p<0.05 and p<0.05) respectively. Phosphates showed non‐significant decrease while albumin reported nearly similar results. Positive correlation between inorganic phosphates and ionized calcium (r = 0.48, p<0.05) was observed only in healthy control group, not in diabetics. This may be attributed to alteration of ions excretions under diabetic condition through unknown mechanism. Additive increase of heparin sodium units in vitro to normal blood resulted in progressive fall in ionized calcium. Significant decrease was only observed upon addition of higher unit (5IU/ml). This must be taken into consideration biochemically for plasma determination of ionized calcium.     

增塑剂DIDP对肝脏氧化应激损伤及VitE拮抗作用研究

研究增塑剂邻苯二甲酸二异癸酯(didecyl phthalate, DIDP)致雄性小鼠肝损伤作用及其机理。以雄性BALB/c小鼠为受试动物,随机分为7组,包括溶剂对照组(生理盐水)、4个DIDP染毒组(0.15、1.5、15和150 mg·kg~(-1))、维生素E(vitamin E, VitE)(100 mg·kg~(-1))处理组和DIDP+维生素E处理组(150 mg·kg~(-1)DIDP+100 mg·kg~(-1)VitE),连续灌胃14 d。以肝组织匀浆测定活性氧(reactive oxygen species, ROS)、还原型谷胱甘肽(glutathione, GSH)、丙二醛(malondialdehyde, MDA)和细胞凋亡因子半胱氨酸天冬氨酸蛋白酶3(cysteine aspartic proteinase 3, Caspase-3)水平。采用动物自动生化分析仪检测肝功能指标血清中丙氨酸氨基转移酶(alanine aminotransferase, ALT)、天门冬氨酸氨基转移酶(aspartate aminotransferase, AST)、白蛋白(albumin, ALB)水平,并同时观察肝组织的病理变化与荧光染色结果。随着DIDP染毒剂量的增加,小鼠肝组织ROS、MDA和Caspase-3含量逐渐上升,血清ALT和AST水平也逐渐上升,GSH含量逐渐降低,血清ALB水平也逐渐降低,差异具有统计学意义(P 0.05,P 0.01); VitE处理组ROS、MDA和Caspase-3含量相应降低,血清ALT和AST水平也相应降低,GSH含量逐渐上升,血清ALB水平也相应上升。小鼠肝组织形态观察结果表明,随着DIDP染毒剂量的增加,小鼠肝组织的病理损伤程度呈上升趋势。研究表明,较高剂量(≥15 mg·kg~(-1))的DIDP能造成小鼠的肝脏损伤与细胞凋亡,抗氧化剂VitE可使肝脏损伤与细胞凋亡减轻,对小鼠肝组织起保护作用,说明氧化应激介导了DIDP对机体的损伤。     

Toxic interaction mechanism of two fluoroquinolones with serum albumin by spectroscopic and computational methods

To evaluate the toxicity of two fluoroquinolones (FQs), ciprofloxacin (CPFX), and enrofloxacin (ENFX), at the protein level, their binding modes with bovine serum albumin (BSA) were characterized by multiple spectroscopic and molecular docking methods under simulated physiological conditions. On the basis of fluorescence spectra, we concluded that both FQs greatly quenched the fluorescence intensity of BSA, which was attributed to the formation of a moderately strong complex mainly through electrostatic interactions. Besides, CPFX posed more of an affinity threat than ENFX. The molecular docking methods further illustrated that both CPFX and ENFX could bind into the subdomain IIIA of BSA and interact with Arg 508 and Lys 437, the positively charged residues in protein. Furthermore, as shown by the synchronous fluorescence, UV-Visible absorption and circular dichroism data, both CPFX and ENFX could lead to the conformational and microenvironmental changes of BSA, which may affect its physiological function.     

槲皮素修饰玻碳电极(Qu/GCE)的制备及铜离子-牛血清白蛋白结合反应机制研究

通过滴涂法制备槲皮素修饰玻碳电极(Qu/GCE,Quercetin/Glassy Carbon Electrode),并对修饰电极进行电化学性能表征;利用槲皮素修饰电极研究了Cu2+离子与牛血清白蛋白(BSA,Bovine Serum Albumin)的结合反应机制.循环伏安法测试结果显示该修饰电极具有准可逆氧化还原反应;将槲皮素修饰电极应用于测试重金属离子-血清白蛋白的结合反应机制,在pH 5.0 HAc-NaAc缓冲溶液体系中,使用槲皮素修饰电极测定Cu2+-BSA相互作用,显示出良好的线性响应关系.根据文献报道结合实验数据,本文建议了结合参数计算理论方程,并利用建议的理论方程计算了Cu2+-BSA的结合常数及结合位点数,相关研究结果可为槲皮素修饰电极研究非共价相互作用提供有益参考.     

Cu~(2+)离子与蛋白质混合体系相互作用机制研究

应用荧光光谱法研究Cu2+离子与血清蛋白质混合组份体系的结合反应机制,测定了反应体系的结合常数K、结合位点数n,探讨了荧光猝灭机理。同时探索了Cu2+与蛋白质混合组份体系及单一组份体系相互作用的差异。结果表明:混合组份体系中,牛血清白蛋白(Bovine serum albumin,BSA)与牛乳铁蛋白(Bovine lactoferrin,BLF)分子间也存在着相互作用,从而影响混合组份体系中BSA/BLF与Cu2+的相互作用,Cu2+与蛋白质混合组份体系相互作用并非蛋白质单组份体系作用之和。分析实验数据,首次发现当BSA/BLF达到临界比值时,金属离子与混合蛋白质组份相互作用过程中出现了蛋白质分子的"契合"现象。文章利用生物酶反应机制中的酶活性部位柔性假说合理解释了Cu2+与血清蛋白质混合组份体系相互作用中的"量敏效应"。     

荧光光谱法研究木栓酮与牛血清白蛋白的相互作用

为了解三萜类化合物在体内的结合转运机制,采用荧光光谱法结合紫外吸收光谱研究从中药爵床中分离的有效成分木栓酮(Friedelin,FDL)与牛血清白蛋白(Bovine serum albumin,BSA)的结合反应.通过研究木栓酮对牛血清白蛋白内源性荧光的淬灭作用,发现木栓酮可以与牛血清白蛋白相互结合,且其作用是一个静态猝灭的过程.通过计算木栓酮与牛血清白蛋白的结合常数和结合位点,发现在生理弱碱性溶液中二者具有较强的结合作用,且以1:1结合.根据热力学参数推测木栓酮与牛血清白蛋白的作用力类型为氢键和范德华力.根据F rster非辐射能量转移机理,计算出木栓酮与牛血清白蛋白结合距离为2.86 nm.研究表明木栓酮在体内可以通过血清白蛋白进行结合和转运.