微透析探针的制备及其在Ni(Ⅱ)与人血清白蛋白的相互作用研究中的应用

介绍了一种自制微透析探针的制备方法,考察了灌流液流速、温度以及探针膜面积等因素对金属离子Ni(Ⅱ) 回收率的影响。实验结果表明:自制微透析探针透析金属离子的各种指标都与商品的微透析探针性能相当,可以替代商品化微透析探针研究金属离子与蛋白质的相互作用。利用微透析与高效液相色谱联用技术研究了Ni(Ⅱ)与人血清白蛋白(HSA)的相互作用。用自制的微透析探针测定Ni(Ⅱ)与HSA之间的第一结合位点数和结合常数分别为1.16和8.6×105 L/mol,采用商品的微透析探针测定的结果分别为1.23和7.5×105     

半胱氨酸对高效液相色谱法测定多西紫杉醇白蛋白纳米粒含量的影响

建立多西紫杉醇(docetaxel,DOC)人血清白蛋白纳米粒(human serum albumin nanoparticles,HSA-NP)中药物含量测定的方法.利用半胱氨酸(cysteine,Cys)还原人血清白蛋白中交联的二硫键,释放HSA-NP中包裹的药物,再利用HPLC测定药物含量.结果表明,在本实验条件下Cys及制剂中辅料对DOC的含量测定无干扰.1 μg/L Cys、37 ℃孵化30 min,最后用乙腈沉淀蛋白的方法可以完全测定出制剂中DOC含量.     

高效液相色谱法测定人血清中假尿嘧啶核苷的浓度

应用反相高效液相色谱法测定人血清中假尿苷（ＰＤ）的含量,色谱柱为Ｎｏｖａ－ＰａｋＣ１８３．９ｍｍ×１５０ｍｍ,流动相为０．０４ｍｏｌ／Ｌ磷酸二氢钾缓冲液（ｐＨ４．０）,检测波长为２６３ｎｍ,线性范围为０．７～６．８μｍｏｌ／Ｌ,回收率为９３．５０％,日间误差ＣＶ＝３．１１％（ｎ＝６）。同时测定了部队体检正常人血清中ＰＤ的浓度,并用于临床观察肝炎、肾病、肺癌等多种疾病以及Ｈｅ－Ｎｅ激光治疗前后患者血中ＰＤ含量的变化。正常人血中ＰＤ的浓度无性别差异,成年人的正常值与文献一致。     

光谱法研究牡荆素与蛋白质相互作用及共存金属离子的影响

利用荧光光谱法和紫外-可见光谱法研究了牡荆素(VT)与牛血清白蛋白(BSA)之间的相互作用.VT对BSA的荧光猝灭为动态猝灭过程,测定了不同温度下的猝灭常数;根据F(o)rster非辐射能量转移理论,计算出VT在BSA中的结合位置与色氨酸残基间的距离为2.675.nm;通过热力学参数推断出VT与BSA之间主要靠疏水作用...     

Determination of drug-protein interactions by combined microdialysis and high-performance liquid chromatography

Summary A simple and rapid method for determination of the parameters of the interaction between drugs and protein, including the association constant and the number of binding sites, has been developed by use of a microdialysis sampling technique combined with high-performance liquid chromatography. The drug and protein (carbamazepine (5H-dibenz[b,f]flazepine-5-carboxamide, CBZ) and human serum albumin (HSA) were used as examples) were mixed in different molar ratios in 0.067 M potassium phosphate buffer, pH 7.4, and incubated at 37°C in a water-bath. The microdialysis probe was the used to sample the mixed CBZ-HSA solution at a perfusion rate of 1 μL min⁻¹. The concentration of CBZ in the microdialysate was determined by reversed-phase high-performance liquid chromatography. Relative recovery (R), determined in vitro under similar conditions, was approximately 42.7%; theRSD ofR was approximately 1.85%. The estimated association constant (K) and the number of the binding sites,n, on one molecule of HSA were 1.06×10⁴ M⁻¹ and 0.880, respectively, which is in good agreement with the literature values determined by high-performance frontal analysis. The potential use of microdialysis is also discussed.     

Pharmacokinetics of timolol in aqueous humor sampled by microdialysis after topical administration of thermosetting gels

In order to develop a thermosetting gel-based formulation, the ocular pharmacokinetics of timolol was studied utilizing microdialysis sampling technique after topical administration. A linear microdialysis probe was characterized and implanted in the anterior chamber of a rabbit. Dialysate samples collected from the aqueous humor (AH) were directly injected into the HPLC system without any pre-treatment and no interference was observed in the blank sample. The measured in vitro recovery of the probe was 57.67%; however, the in vivo recovery significantly decreased to 16.78% when assessed by the retrodialysis method, which was used to calculate the timolol concentration in AH. Although in the initial 15 min the drug concentrations in AH were comparable to that of the timolol solution, increased Cmax and significantly improved ocular bioavailability were obtained for the gel. When sodium deoxycholate (DC) was incorporated in the gel as a penetration enhancer, a 2-fold increment in the ocular bioavailability was achieved with an increased Cmax and significantly suspended Tmax. The results demonstrated that microdialysis coupled to HPLC is a powerful tool to investigate the ocular pharmacokinetic, and hence facilitates the design of ophthalmic formulations.     

Trace analysis of heavy metal ions by ion chromatography

Summary An ion chromatographic separation technique for heavy metal ions is described. Using pressure-stable, silica-based, ion-exchange supports and standard HPLC equipment with post-column reaction detector high resolution is achieved as well as extremely high sensitivity in the parts per trillion (ppt)-range.     

Determination of the number of binding sites and binding constant between diltiazem hydrochloride and human serum albumin by ultrasonic microdialysis coupled with online capillary electrophoresis electrochemiluminescence

A simple, sensitive and selective determination of diltiazem hydrochloride (DLT) is described using capillary electrophoresis electrochemiluminescence (CE-ECL). The CE-ECL parameters that affect separation and detection were optimized. Under the optimized conditions, the linear range of DLT was from 0.02 to 100 μmol/L (r(2) = 0.9983), with the detection limit of 5.1 nmol/L (3σ). The relative standard deviations of ECL intensity and the migration time were <2% for 0.1 μmol/L and 22 μmol/L DLT (n = 11). A new technique for determining of the number of binding sites and binding constant between DLT and HSA was developed using ultrasonic microdialysis coupled with CE-ECL. The number of binding sites and binding constant were 5.9 and 6.3 × 10(4) L/mol, respectively. The time required for ultrasonic microdialysis was 10 times less than that for traditional dialysis. Compared with traditional dialysis, ultrasonic microdialysis is simple, rapid, and should be applicable to a wide range of interactions of drugs and biomacromolecules.     

高效液相色谱法测定人血清中5-氟尿嘧啶浓度

建立了测定人血清中5-氟尿嘧啶的反相高效液相色谱法。采用ZorbaxODS柱4.6mm×250mm,预柱YWG-C184.6mm×50mm;甲醇:水（20:80）为流动相,紫外检测波长为273um,喃氟啶为内标定量。回收率为96.85％,田间和日内变异系数分别为537％和438％,血清中最低检测浓度为10μg／L。在0.04～50mg／L浓度范围内线性良好,相关系数为0.9990。     

Determination of L-tryptophan-serum albumin binding by high-performance liquid chromatography

Summary Two high-performance liquid chromatography (HPLC) techniques were developed for the determination of binding constants in the interaction of serum albumin with L-tryptophan: internal calibration and external calibration. The results obtained were compared with those obtained by the classical method of equilibrium dialysis and by gel filtration. While all the methods are equally reliable, the internal and external calibration techniques seem to be superior in their simplicity, speed and convenience.     

微透析联用反相高效液相色谱对大鼠皮肤葛根素的药代动力学研究

采用在体微透析取样技术联用RP-HPLC的方法对大鼠皮肤中的葛根素进行了体内药代动力学的考察.将微透析探针进行体内外校正后,植入大鼠皮下,以磷酸盐缓冲溶液作为灌流液,收集的样品在以Hypersil-C18(150 mm×4.6 mm, 5 μm)为色谱柱,流动相为V(甲醇)∶ V(0.1% 醋酸)＝30∶ 70,流速1 mL/min,检测波长221 nm的HPLC色谱条件下进行分析,所得的大鼠皮下游离葛根素浓度用Kinetica软件进行处理.结果显示:微透析探针体内外校正结果显示葛根素适宜应用微透析技术,在所建立的HPLC条件下葛根素在0.1～100 mg/L范围内线性关系良好(r=0.9999, n=6),浓度为100、10和0.1 mg/L的葛根素日内RSD分别为0.38%、0.80%和1.21%(n=6);日间RSD分别为0.44%、0.73%和1.68%(n=6),所得数据经Kinetica软件拟合后,得到大鼠皮下非结合葛根素药-时曲线和主要药动学参数.所建立的微透析联用RP-HPLC方法能够在体、实时监测大鼠皮下葛根素浓度.     

高效液相色谱法测定人血清中头孢克罗

建立了测定人血清中头孢克罗的高效液相色谱法。分析柱为Ｓｈｉｍ－ｐａｃｋＣＬＣ－ＯＤＳ（５μｍ）,６．０ｍｍ×１５０ｍｍ;流动相为磷酸盐缓冲液（ｐＨ５．８）∶乙腈＝８７．５∶１２．５,流速为１．２ｍＬ／ｍｉｎ;紫外检测波长为２５４ｎｍ。血清中头孢克罗浓度在０．１２５～２４ｍｇ／Ｌ范围内,峰高与浓度呈良好的线性关系,回归方程为Ｈ＝－３．２００＋９２．２９Ｃ（ｎ＝９,ｒ＝０．９９９９）,日内、日间变异系数分别为４．１％和５．８％,平均回收率为９９．３％。     

卟啉在高效液相色谱－光度法测定痕量金属离子方面的应用

评述了当前卟啉类化合物在高效液相色谱－光度法测定痕量金属离子方面的应用研究，展望了此研究领域的发展前景，参考文献６５篇     

高效亲和色谱法测定2种中药成分与人血清白蛋白的结合率

采用高效亲和色谱技术(HPAC)对中药成分与人血清白蛋白(HSA)的相互作用进行了研究。首先采用点击化学的方法制备了表面键合有HSA蛋白的硅胶固定相并装填成亲和色谱柱,根据药物在该色谱柱上与空白硅胶柱上的保留时间差计算得到药物与蛋白的结合率。利用该方法测得模型化合物华法令与HSA的结合率与文献中采用超滤法测得的结果基本一致,表明该方法可用于测定药物与HSA的结合率。在此基础上用该方法测定了葛根素和告依春两种中药成分与HSA的相对结合率分别为10.26%和10.20%。同时用超滤的方法测定了葛根素与HSA的结合率为14.25%。结果表明,HPAC可以作为研究药物与蛋白相互作用的一种简便可行的方法,其测定结果与超滤方法一致。     

Zinc silicate-bonded diethyldithiocarbamate — a selective adsorbent for separating transition metal ions and preconcentration of palladium(II)

Summary A new chelating adsorbent, zinc silicate-bonded diethyldithiocarbamate (DTC) has been prepared for the separation and preconcentration of some transition metal ions. The distribution coefficients (Kd values) of 15 metal ions have been estimated in solution at various pH's and some quantitative separations have been achieved. The high selectivity for Pd(II) has been utilized for its preconcentration from dilute aqueous solutions; quantitative elution is effected with acidic 4% thiourea solution.     

高效液相色谱法测定人血清中阿西美辛和吲哚美辛

摘要：建立了测定人血清中阿西美辛及其活性代谢物吲哚美辛的高效液相色谱法。分析柱为Spherisorb-C8（5μm）,4.6mm×250mm,流动相为V（醋酸盐缓冲液,pH4.6）:V（乙腈）:V（甲醇）=55:40:5,流速1.0mL/min,检测波长254nm。血清中药物质量浓度为12.5μg/L～1.6mg/L时,阿西美辛、吲哚美辛峰高与内标甲苯磺丁脲峰高比值和质量浓度呈良好的线性关系;阿西美辛日内、日间变异系数分别为3.6％和5.6％,平均回收率为78.3％;吲哚美辛日内、日间变异系数分别为2.4     

高效液相色谱-电化学检测法测定鼠脑微透析液中谷胱甘肽和半胱氨酸

采用高效液相色谱法,用电化学检测手段成功地测定了沙土鼠脑纹状体微透析液内的谷胱甘肽（ＧＳＨ）和半胱氨酸（Ｃｙｓ）。检测前用电化学自清洗方式能有效地提高玻璃碳电极的检测水平,使其具有较好的重现性和回收率。Ｃｙｓ和ＧＳＨ的回收率分别为９１．４％和８７．３％。电化学检测的工作电压为０．９Ｖ,用外标法定量。由微透析探针获取的纹状体透析液Ｃｙｓ和ＧＳＨ的浓度分别为６８．５９和６５．５４ｎｍｏｌ／Ｌ,验证了方法的实用性和可靠性。     

高效液相色谱法测定人血清中鬼臼乙叉甙浓度

建立了测定血清中鬼臼乙叉甙浓度的反相高效液相色谱方法。血清样品以乙酸乙酯提取,加表鬼臼毒噻吩糖甙作内标,以甲醇∶水（５５∶４５）为流动相,紫外２５４ｎｍ检测,最低检测浓度０．１ｍｇ／Ｌ,在药物浓度为０．５～５０ｍｇ／Ｌ范围内有良好的线性关系,Ｙ＝０．０１６０＋０．０３４０Ｘ,ｒ＝０．９９９１,回收率达９５．０％。日内、日间精密度分别为４．１６％和４．７６％（ｎ＝５）。用所建立的方法测定了患者的血清浓度,结果表明,方法简便、灵敏。     

A review on solid phase microextraction-high performance liquid chromatography as a novel tool for the analysis of toxic metal ions

This paper reviews the practical applications of solid phase microextraction-High performance liquid chromatography in the analysis of toxic metal species as these are important contaminants and are carcinogenic. Their determination in formulations, in feed and food, and in complex environmental matrices (e.g., waste water and industrial effluents) often requires analytical methods capable of high efficiency, unique selectivity, and high sensitivity. Solid phase microextraction (SPME) requires low solvent consumption and is quick in use. SPME is used for extraction and online desorption of analytes with the mobile phase of HPLC and subsequent detection by UV, ICP-MS or ESI-MS as detectors. Different SPME-HPLC methods are summarized in this article to demonstrate the usefulness of this technique for metallic species of As, Cr, Pb, Hg and Se.     

高效液相色谱法测定血清中茶碱浓度

采用高效液相色谱,分析柱：３μｍ,３．３ｃｍ＊４．６ｍｍ,Ｉ．Ｄ（Ｐｅｒｋｉｎ Ｅｌｍｅｒ,ＵＳＡ）;预柱：１０μｍ,１ｃｍ＊２．１ｍｍ,Ｉ．Ｄ（Ｐｅｒｋｉｎ ＥＬｍｅｒ,ＵＳＡ）;以乙酰氨基酚为内标对氯仿－异丙醇（９５：５,Ｖ／Ｖ）提取样品进行了分析,流动相：０．１ｍｏｌ／Ｌ醋酸缓冲液（ＰＨ＝４．５）－甲醇（７０：３０,Ｖ／Ｖ）;检测波长：２７０ｎｍ;流速０．５ｍＬ／ｍｉｎ,３ｍｉｎ即完成一次茶