Study of biodistribution of methotrexate-loaded bovine serum albumin nanospheres in mice

Nanospheres made from natural hydrophilic polymers have been proved efficient in terms of better drug-loading capacity, biocompatibility, and possibility less opsonization by reticuloendothelial system (RES) through an aqueous stearic barrier. Hence, nanospheres containing methotrexate were prepared from bovine serum albumin (BSA) by a novel pH coacervation method. A drug-to-polymer ratio study was carried out to determine the carrier capacity. The batch with the highest drug loading was subjected to in vitro analysis. It was found to provide a slow release after an initial burst release. Biodistribution of nanosphere-bound drug was compared with that of free drug in mice. It was observed that the percentage increase in drug distribution to the lungs, liver, and spleen was markedly high from the nanosphere when compared to free drug.     

碳量子点的制备及与牛血清蛋白的相互作用

以木炭为碳源,分别采用回流、微波及超声等不同方法制备碳量子点(Carbon quantum dots,CQDs)。比较不同方法的优劣并优化反应条件、考察不同因素对其荧光量子产率的影响。得到最佳制备方法,制得粒径较小的荧光CQDs,用钝化剂PEG2000修饰后,提高其荧光寿命和量子产率。将修饰后的CQDs应用于与牛血清白蛋白(Bovine serum albumin,BSA)的相互作用,采用紫外吸收光谱法和荧光光谱法探讨其相互作用机理。结果表明,经回流法所制CQDs的荧光量子产率最高,其与BSA之间的荧光猝灭为静态猝灭过程。     

A study on the preparation and anti-tumor efficacy of bovine serum albumin nanospheres containing 5-fluorouracil

The therapeutic profile of many anti-cancer drugs has been improved by their modified distribution through a colloidal carrier system. Hence, bovine serum albumin nanospheres containing 5-fluorouracil were prepared by pH-coacervation methods. To select the most suitable cryoprotector for the formulated nanosphere system, a study on the effect of cryoprotectors in the prevention of particle agglomeration was done. Using glucose and mannitol at various concentrations during freeze drying, glucose at a concentration of 5% was observed to be relatively more effective in the prevention of particle agglomeration than the other cryoprotectors. The carrier capacity was determined through the drug-to-albumin ratio. The particle size of all the drug-loaded batches was analyzed before and after freeze drying. The batch of nanospheres with uniform size distribution, and highest drug loading, was used for other subsequent studies. The effect of surfactant in drug loading was estimated through various concentrations of sodium lauryl sulfate, and it was observed that the surfactant has no influence on drug loading at the selected concentrations. The batch of nanospheres with highest drug loading was evaluated for its in-vitro release, and the drug release was found to be in a bi-phasic pattern. To evaluate the efficacy of 5-fluorouracil-loaded nanospheres against cancer cells, an in vitro cytotoxicity study was carried out using HEp-2 cell lines. The nanosphere-bound drug was observed to produce a better cytotoxic effect than the free drug. The anti-tumor efficacy of drug-loaded nanosphere was investigated in DLA tumor-induced mice models, and the percentage tumor inhibition was relatively higher in animals treated with nanosphere-bound drug than with free drug.     

TiO2纳米管阵列膜对牛血清白蛋白的吸附与脱附

采用电化学阳极氧化法以含氟的乙二醇溶液为电解液阳极氧化纯钛制备出排列规则的高长径比TiO2纳米管阵列膜,并用扫描电镜(SEM)、比表面积仪表征了TiO2纳米管阵列膜的形貌和比表面积。结果表明,所制得的TiO2纳米管阵列的管径约180nm,管长可达230μm,比表面积约59.8m2/g。以牛血清白蛋白(BSA)为药物蛋白分子的模型,并研究了TiO2纳米管阵列膜对BSA的吸附和脱附行为,考察了溶液pH值、BSA初始浓度和溶液离子强度对BSA吸附的影响与吸附态的BSA在不同pH值的PBS溶液中的释放行为。结果表明,BSA分子在其等电点(pH值=4.8)附近较容易吸附到TiO2纳米管上,吸附量随着BSA初始浓度的增加而增加,较高的离子强度会降低BSA的吸附,碱性条件下吸附态的BSA容易从TiO2纳米管上脱附,并由于纳米管的扩散限制效益呈现一定程度的缓释。     

Enhanced antithrombotic effect of hirudin by bovine serum albumin nanoparticles

The aim of this study was to design hirudin-loaded bovine serum albumin (BSA) nanoparticles to control release and improve antithrombotic effect of hirudin. BSA nanoparticles were designed as carriers for delivery of hirudin. Hirudin–BSA nanoparticles were prepared by a desolvation procedure and cross linked on the wall material of BSA. The hirudin–BSA nanoparticles were characterised by particle size distribution, zeta potential, entrapment efficiency, differential scanning calorimetry (DSC), and powder X-ray diffractometry (PXRD). The in vitro release characteristics and pharmacological availability were investigated. The morphology of hirudin–BSA nanoparticles was approximately spherical. The mean particle size was 164.1 ± 5.40 nm and the zeta potential was ?20.41 ± 0.64 mV. The mean entrapment efficiency and drug loading were 85.14% ± 4.79% and 66.38% ± 3.54%, respectively. Results from DSC and PXRD revealed that hirudin in BSA existed in an amorphous state. The release behaviours of hirudin from BSA nanoparticles in phosphate buffer solution were fitted to the bioexponential model. The in vivo result obtained after intravenous injection of hirudin–BSA nanoparticles in normal rats demonstrated that BSA nanoparticles could prolong the antithrombotic effect of hirudin in comparison with hirudin solution. These results suggest that hirudin–BSA nanoparticles may be a promising drug delivery system for thrombosis and disseminated intravascular coagulation therapy.     

水相合成 CdS纳米晶标记牛血清白蛋白

利用水相中直接合成的CdS纳米晶，与牛血清白蛋白(BSA)进行偶连标记。通过分子筛层析对标记后的牛血清白蛋白进行纯化，在紫外灯下即可观察到标记蛋白的荧光。对CdS纳米晶标记后的牛血清白蛋白的荧光光谱的研究表明，标记蛋白后的CdS纳米晶其荧光无明显淬灭。     

荧光光谱法研究头孢孟多脂与牛血清白蛋白的相互作用

模拟生理条件下,用荧光光谱法和紫外-可见吸收光谱法研究头孢孟多酯和牛血清白蛋白(BSA)结合反应的特征。研究表明:头孢孟多酯与BSA形成复合物,从而猝灭BSA的内源性荧光,该过程为静态猝灭过程。根据SternVolmer方程得出不同温度下结合位点数n和结合常数Ka;结合位点位于BSA的亚结构IIA中。通过计算相应的热力学参数,确定头孢孟多酯与BSA之间的作用力主要为静电作用力。利用同步荧光光谱探讨了头孢孟多酯与BSA作用前后白蛋白的构型变化。Hill系数nH1,表明头孢孟多酯有弱的负协同作用。此研究不仅对于揭示体内药物动力学问题和指导临床合理用药具有一定意义,而且对药物分子设计及新药开发等也具有重要指导意义。     

Preliminary study of adsorption behavior of bovine serum albumin(BSA) protein and its effect on antibacterial and corrosion property of Ti-3Cu alloy

The adsorption behavior,antibacterial,and corrosion properties of a Ti-3Cu alloy were studied in a phosphate-buffered saline solution containing 0,1,3,and 6 g L-1 bovine serum albumin protein at 37 ℃and pH =7.4 (±0.2).The protein adsorption behavior was examined via cyclic voltammetry,secondary ions mass spectroscopy (SIMS),and angle-resolved X-ray photoelectron spectroscopy (ARXPS).The cor-rosion property was analyzed by the open circuit potential (OCP),potentiodynamic polarization (PD),and electrochemical impedance spectroscopy (EIS) examinations.The antibacterial test was conducted according to the GB/T 21510 China Standard.It was observed that the surface charge density (Q～s) was directly proportional to the amount of the adsorbed BSA protein,signifying that the protein adsorption was accompanied by the charge transfer,pointing to chemisorptions phenomena.BSA amino groups and other organic species were observed in the surface analysis examinations.It was shown that the formation of barrier complexes between the TiO2 oxide-layer and PBS solution resulted in decreasing the release of Cu-ions,which consequently reduced the antibacterial activity.On the other hand,these barrier complexes improved the corrosion resistance by increasing the charge transfer resistance and double-layer capacitance of the Ti-3Cu alloy.     

含硅羟基磷灰石粉体的合成及其与蛋白质的相互作用研究

将硅掺入羟基磷灰石(HA)晶格中能有效地提高HA的生物相容性.采用湿法合成了含硅羟基磷灰石微粉(Si-HA),用X射线荧光光谱(XRF)、X射线衍射(XRD)、红外光谱(FTIR)、扫描电镜(TEM)和荧光光谱等对Si-HA的晶相、结构、化学组成、形貌及其与牛血清白蛋白(BSA)的相互作用进行了表征和分析.结果表明:Si元素溶入磷灰石晶格中,使HA晶胞参数和结构发生变化,随着Si含量的增加,HA中-OH和PO3-4的吸收峰呈减弱趋势,表明SiO4-4可能部分取代了PO3-4.Si-HA与BSA作用后的PO3-4吸收峰发生变化,在Si-HA图谱中出现了蛋白质的酰胺Ⅰ带和酰胺Ⅱ带吸收峰,随着Si-HA与BSA作用时间由2h～3d的变化,BSA溶液酰胺I带位置出现蓝移,半峰宽减小且峰型变得尖锐,酰胺II带则向低频区移动.Si-HA与BSA作用3d后,开始在1139处出现PO3-4的吸收峰.荧光光谱显示:加入相同质量的HA和Si-HA后蛋白质内源荧光强度较纯BSA溶液下降,随着Si-HA中Si质量分数的增加BSA荧光强度下降幅度增大,发射峰也出现红移,且Si-HA与BSA作用较纯HA与BSA的作用强,说明Si-HA使蛋白质二级结构发生改变,Si-HA中的Ca、PO3-4、SiO4-4与BSA之间发生了相互溶解、吸附和键合等作用,证明了Si-HA与生命物质蛋白质之间具有较高的反应性能,对Si-HA的生物活性及其与BSA相互作用的机理进行了详尽的阐述.     

N-组氨酸壳聚糖/聚乳酸支架吸附牛血清白蛋白研究?

采用二次相分离制备7种 N-组氨酸壳聚糖/聚乳酸(NHCS/PLLA)支架,考察其在模拟体液中对牛血清白蛋白(BSA)的吸附性能,并研究其吸附等温式、吸附动力学和热力学行为等.结果表明,在37．0℃、BSA 的初始浓度为2．5 mg/mL 时,50 kD-NHCS-3与PLLA质量比为5/5制得的 NPs3支架对BSA 的吸附容量最大,达928．53 mg/g;吸附遵循Langmuir吸附等温式和准一级动力学方程,可为分离纯化BSA提供借鉴.     

Concurrent zero-dimensional and one-dimensional biomineralization of gold from a solution of Au3+ and bovine serum albumin

Abstract

A technique was developed for preparing a novel material that consists of gold nanoparticles trapped within a fiber of unfolded proteins. These fibers are made in an aqueous solution that contains HAuCl₄ and the protein, bovine serum albumin (BSA). By changing the ratio of gold to BSA in solution, two different types of outcomes are observed. At lower gold to BSA ratios (30–120), a purple solution results after heating the mixture at 80 °C for 4 h. At higher gold to BSA ratios (130–170), a clear solution containing purple fibers results after heating the mixture at 80 °C for 4 h. UV–Vis spectroscopy and light scattering techniques show growth in nanocolloid size as gold to BSA ratio rises above 100. Data indicate that, for the higher gold to BSA ratios, the gold is sequestered within the solid material. The material mass, visible by eye, appears to be an aggregation of smaller individual fibers. Scanning electron microscopy and transmission electron microscopy indicate that these fibers are primarily one-dimensional aggregates, which can display some branching, and can be as narrow as 400 nm in size. The likely mechanism for the synthesis of the novel material is discussed.     

钛表面自组装牛血清白蛋白以控制血小板粘附行为的研究

将牛血清白蛋白通过自组装方法组装到钛表面,以改善其血液相容性.首先,用氢氧化钠活化钛,得到有活性羟基且带负电荷的多孔表面,然后浸入带正电荷的聚赖氨酸溶液,最后浸入带负电荷的牛血清白蛋白溶液.通过扫描电子显微镜(SEM)和原子力显微镜(AFM)观察自组装前后表面形貌变化,通过傅立叶红外漫反射(FTIR)检测各步处理层表面基团变化,通过视频接触角观察各步处理后钛表面接触角变化,通过体外血小板粘附实验评价自组装前后表面血液相容性变化.实验结果表明,牛血清白蛋白组装到钛表面后,血小板的粘附行为得到有效控制,钛表面的血液相容性显著改善.     

硅掺杂羟基磷灰石生物活性微粉对人血清白蛋白的吸附特性

对含硅羟基磷灰石(Si-HA)与人血清白蛋白(HSA)的吸附特性及动力学特征进行了研究,计算了表观活化能,用等温吸附曲线进行拟合。结果表明,该吸附属于Langmuir型。Si-HA和HA吸附HSA的表观活化能分别为21.28和35.57 kJ.mol-1,20℃下吸附反应的速率常数分别为1.48和1.34 min-1,饱和吸附量分别为25.34和21.34 mg.g-1,Si-HA具有比纯HA更好的吸附优势。通过XRD、FTIR及荧光光谱分析,探讨了HSA在Si-HA和HA表面的吸附作用机制,表明Si-HA与蛋白表面吸附反应是以化学吸附为主、包含物理吸附的混合吸附过程。从分子结构的角度揭示了Si-HA和HA吸附蛋白质性能的差异,为进一步探讨Si-HA和HA生物相容性提供了新途径。     

汽巴蓝功能微球对人血清白蛋白的吸附性能研究

通过分散共聚制得了聚苯乙烯接枝聚醋酸乙烯酯(PSt-g-PVAc)微球,粒径控制在500～700nm之间.在碱性条件下将PSt-g-PVAc微球表面的PVAc链醇解为聚乙烯醇(PVA),得到了PSt-g-PVA微球,提高了其在水中的分散稳定性.使汽巴蓝(CB)与PSt-g-PVA表面的羟基进行亲核反应,制得了CB作为配体的新型吸附剂汽巴蓝功能微球(CB微球),元素分析测得CB微球表面的CB最大含量为139.22μmol/g,探讨了其在不同人血清白蛋白(HSA)浓度、pH值和吸附时间下对HSA的吸附性能.当HSA浓度为1.0mg/mL、pH为5.14时,CB微球对HSA的最大吸附量为40.9mg/g,并分析了CB微球与HSA的相互作用机理.由NaSCN进行解吸试验,发现可将吸附在CB微球上的HSA解吸92.11%以上,且CB微球的重复使用性能良好.     

Study on the interaction of phthalate esters to human serum albumin by steady-state and time-resolved fluorescence and circular dichroism spectroscopy

Phthalate esters (PAEs) are globally pervasive contaminants that are considered to be endocrine disruptor chemicals and toxic environmental priority pollutants. In this paper, the interactions between PAEs and human serum albumin (HSA) were examined by molecular modelling, steady state and time-resolved fluorescence, ultraviolet-visible spectroscopy (UV-vis) and circular dichroism spectroscopy (CD). The association constants between PAEs and HSA were determined using the Stern-Volmer and Scatchard equations. The binding of dimethyl phthalate (DMP) to HSA has a single class of binding site and its binding constants (K) are 4.08 × 10³, 3.97 × 10³, 3.45 × 10³, and 3.20 × 10³ L mol⁻¹ at 289, 296, 303, and 310 K, respectively. The Stern-Volmer and Scatchard plots both had two regression curves for HSA-butylbenzyl phthalate (BBP) and HSA-di-2-ethylhexyl phthalate (DEHP), which indicated that these bindings were via two types of binding sites: the numbers of binding site for the first type were lower than for the second type. The binding constants of the first type binding site were higher than those of the second type binding site at corresponding temperatures, the results suggesting that the first type of binding site had high affinity and the second binding site involved other sites with lower binding affinity and selectivity. The thermodynamic parameters of the binding reactions (ΔG°, ΔH° and ΔS°) were measured, and they indicated the presences of hydrophobic forces and hydrogen interactions in the PAEs-HSA interactions, which agreed well with the results from molecular modelling. The alterations of protein secondary structure in the presence of PAEs were confirmed by UV-vis and CD spectroscopy. The time-resolved fluorescence study showed that the lifetime of Trp residue of HSA decreased after the addition of PAEs, which implied that the Trp residue of HSA was the main binding site.     

Characterization of Alizarin Red S binding sites and structural changes on human serum albumin: a biophysical study

Alizarin Red S (ARS), is a water-soluble, widely used anthraquinone dye synthesized by sulfonation of alizarin. In this report, the binding of ARS to human serum albumin (HSA) was characterized by employing fluorescence, UV/vis absorption, circular dichroism (CD), and molecular modeling methods. The data of fluorescence spectra displayed that the binding of ARS to HSA is the formation of HSA-ARS complex at 1:1 stoichiometric proportion. Hydrophobic probe 8-anilino-1-naphthalenesulfonic acid (ANS) was employed and elucidated that the dye was located in subdomain IIIA. This phenomenon corroborates the result of site-specific probe displacement experiments, which demonstrate the dye is at indole-benzodiazepine site (Sudlow's site II); and it is also consistent with guanidine hydrochloride (GuHCl) induced HSA unfolding studies and molecular modeling simulations. The features of the dye, which led to structural perturbations of HSA, have also been studied in detail by methods of UV/vis, CD and three-dimensional fluorescence spectroscopy.