四种神经肽的反相高效液相色谱法分离

以反相高效液相色谱法（ＲＰ－ＨＰＬＣ）分离甲硫脑啡肽（ＭＥＫ）、亮脑啡肽（Ｌｅｕ－ＥＮＫ）、强啡肽Ａ１－１７（ＤｙｎＡ１－１７）、胆囊收缩素（ＣＣＫ－８ｓ）。色谱柱为ＺＯＲＢＡＸＣ１８（２．１ｍｍＩＤ×１５０ｍｍ）；流动相：乙腈／水／三氟乙酸（ＴＦＡ），梯度洗脱，紫外检测器检测：ＵＶ２１５ｎｍ。上述条件下，四种神经肽在微克水平得到较好分离。     

The Separation of Pre-Column O-Phthalaldehyde Derivatized Amino Acids by High Performance Liquid Chromatography

Abstract

A method is presented in which a pre-column fluorescent derivative of primary amino acids is formed using o-phthalaldehyde (OPA). After a one minute derivatization period, the OPA amino acids are separated by high performance liquid chromatography using a ternary gradient in the reversed phase mode and fluorometric detection. The complete analysis time, including the derivatization step is less than 40 minutes with an average minimum detectable quantity of 40 picograms.     

高效液相色谱手性固定相法直接拆分4种吲哚类衍生物

将纤维素-三(3,5-二甲基苯基氨基甲酸酯)(CDMPC)涂敷于自制的球形氨丙基硅胶上,制备成了纤维素-三(3,5-二甲基苯基氨基甲酸酯)手性固定相(CDMPC-CSP)。利用正相高效液相色谱,在该固定相上对新合成的4种吲哚类衍生物对映体进行了手性拆分。通过选择不同结构和浓度的醇类改性剂,优化了色谱分离条件,同时探讨了醇的结构和浓度对于对映体拆分和保留的影响。结果表明,适合Ⅱ~Ⅳ号样品拆分的醇类改性剂分别为正丁醇、乙醇和乙醇,而适合Ⅰ号样品的醇类改性剂为乙醇和正丙醇组成的混合体系。在优化的各流动相体系下,4种吲哚类衍生物的对映体都得到了很好的分离。在此基础上计算了它们的对映体过量值(e.e.值)。实验结果令人满意,表明高效液相色谱手性固定相法是拆分这类化合物的一种理想方法。     

Separation and Quantitation of O-Phthalaldehyde Derivatives of Taurine and Related Compounds in a High Performance Liquid Chromatography (HPLC) System

Abstract

A sensitive specific assay for taurine using high performance liquid chromatograpy and fluorescence measurement is described. The method employs precolumn derivatization with o-phthalaldehyde in the presence of ethanethiol. Taurine is clearly separated from other amino acids including its precursors hypotaurine and cysteine sulfinic acid. The fluorescence peak height is linear between 1 and 100 picomoles of taurine. There is clear separation of taurine from a contaminant of other taurine assays, α-glycerophosphoryl ethanolamine.     

Ion Exchange High Performance Liquid Chromatography of Leukotrienes

Abstract

A novel anion exchange liquid chromatographic system has been developed for isocratic separation of leukotrienes. Hydrophobic as well as ionic forces were found to influence the separation. By optimization of solvent strenght, ionic strenght and pH, amphoteric peptidoleukotrienes could be separated simultaneously with hydroxy fatty acids such as leukotriene B₄ and its ω-oxidized metabolites. To obtain a good buffering capacity of the mobile phase at optimum pH, a multicomponent buffer was developed.     

反相高效液相色谱法分离蛋白质的研究

采用反相高效液相色谱法考察了几种大孔硅胶烷基键合固定相在等度淋洗条件下进行蛋白质分离的色谱性能。研究了冲洗剂中有机溶剂异丙醇的浓度、离子对酸（ＴＦＡ）浓度对蛋白质保留时间的影响。探讨了蛋白质在ＲＰ－ＨＰＬＣ中的保留机理。结果表明,大孔硅胶（２０～３０ｎｍ）短链（Ｃ４和Ｃ８）烷基键合固定相适合蛋白质的分离。     

The Purification of Xanthene Dyes by Reverse Phase High Performance Liquid Chromatography

Abstract

A reliable method for the separation of fluorescein dyes from their impurities was developed using high performance liquid chromatography and involved a μBondapak C₁₈ reverse phase column and mixtures of methanol and ammonium acetate buffer. This technique was used to verify the purity of commercial products as well as to aid in the development of an empirical theory related to retention of halogenated fluorescein dyes by reverse phase columns.     

反相高效液相色谱法分离测定酪胺

建立了分离酪胺与酪氨酸及其它杂质的反相键合相高效液相色谱法 ,讨论了流动相添加剂对色谱分离的影响和离子相互作用的分离机理。在 C8烷基键合相分离柱上 ,以含 Tris-高氯酸盐 ( 2 0 mmol/L Tris,用 HCl O4调节 p H为 7.9,并添加 KCl O4,使总高氯酸盐浓度为 30 mmol/L )的甲醇 -水溶液 (体积比为 4 0∶ 60 )作为流动相 ,以对甲苯磺酰胺为内标物 ,测定了 p-酪氨酸脱羧工艺产物——酪胺的质量分数。酪胺样品质量分数测定的准确度和重现性数据为 ( 96.4 0± 0 .633) ( n=11,RSD=0 .66 ) ,加样回收率为 99.33～ 10 0 .38。方法可用于工艺条件的选择和酪胺产品质量的检测     

Separation of Optical Isomers as Diastereomeric Derivatives by High Performance Liquid Chromatography

Abstract

Optical isomers occuring widely in nature, display different biochemical and pharmacological effects against their enantiomeric isomers. Therefore, it is very important to distinguish or resolve enantiomers, especially in biochemical and pharmaceutical applications.     

反相键合相高效液相色谱法分离测定萘普生

建立了分离测定萘普生和溴代萘普生的反相键合相高效液相色谱法。采用ＯＤＳ柱，以添加５０ｍｍｏｌ／Ｌ乳酸并用高氯酸调节ｐＨ为２．５的８０％甲醇－水溶液作为流动相，以苯甲酸为内标物，测定了不对称合成工艺产物中萘普生和溴代萘普生的含量。方法的准确度分别为９９．８３％～１０２．０７％（萘普生）和９９．０％～１００．８３％（溴代萘普生），相对标准偏差分别小于２．５８％（萘普生）和３．６４％（溴代萘普生）。方法可用于工艺条件的选择和质量检测。     

高效液相色谱法拆分外消旋药物联苯双酯及其衍生物

合成并制备了纤维素-三（3,5-二甲基苯基氨基甲酸酯）手性固定相,利用高效液相色谱法首次直接拆分了5种外消旋联苯类药物,考察了流动相组成,样品结构对保留和拆分的影响。     

大环糖肽抗生素键合相高效液相色谱法拆分7种氨基带保护基的氨基酸对映体

采用高效液相色谱法在装有大环糖肽抗生素键合相的手性柱上拆分了7种氨基带有芴甲氧羰基(fluorenylmethoxycarbonyl,Fmoc)保护的氨基酸对映体。比较了Fmoc-缬氨酸和相应的不带保护基的缬氨酸对映体在不同流动相体系中的色谱保留行为;考察了甲醇-醋酸-三乙胺流动相体系中醋酸和三乙胺的浓度以及它们二者的浓度之比对N-Fmoc氨基酸对映体拆分效果的影响。实验结果表明分离温度及流动相流速的变化也会对分离结果产生影响。该法简便快速,已成功地用于这类氨基酸的光学纯度测定。     

Automated Analysis of O-Phthalal-Dehyde Derivatives of Amino Acids in Physiological Fluids by Reverse Phase High Performance Liquid Chromatography

Abstract

An automated method is described for the determination of free amino acids in biological fluids using precolumn deriva-tization with o-phthalaldehyde and reverse phase high performance liquid chromatography. Chromatographic separation of amino acids is accomplished in 24 minutes (cycle time 44 minutes). As little as 1.5 pmol of most commonly occurring amino acids can be accurately quantified. Accuracy and reproducibility are optimized by automating the derivatization-injection sequence and by correcting for variations in the fluorescence response of each amino acid in each run. A total of 31 analyses can be completed in 24 hours on a single column (7 standards and 24 unknowns). The method can be used in the general determination of free amino acids in biological fluids, or can be further accelerated and used for the quantitation of specific amino acids simply by altering the elution conditions.     

异丙甲草胺立体异构体在高效液相色谱手性固定相上的分离

在Pirkle“刷型”(S,S)-Whelk—O1手性柱和涂敷型纤维素三(3,5-二甲基苯基氨基甲酸酯)(CDMPC)手性柱上,对异丙甲草胺立体异构体进行了分离。在(S,S)-Whelk-O1手性柱上考察了不同体积比的正己烷．异丙醇流动相对分离的影响,发现当异丙醇含量降低时有利于异构体的分离;在CDMPC手性柱上考察了正己烷中醇类改性剂的种类和浓度对分离的影响,发现醇含量降低时和采用大体积的醇时有利于异构体的分离。异丙甲草胺的对映异构体之间在(S,S)-Whelk-O1手性柱上获得了分离,在CDMPC手性柱上分离出了异丙甲草胺4个异构体中的3个。同时测定了富S-异丙甲草胺异构体中S-异构体的过量值(EE),本方法可用于富S-异丙甲草胺的定量。     

The Purification of Amino Coumarin Dyes by Reverse Phase High Performance Liquid Chromatography

Abstract

A reliable method for the separation of amino coumarin dyes from associated impurities has been developed using reverse phase high performance liquid chromatography using an ODS-18 column and mixtures of methanol and water. Further, the relation between the retention times and structure of amino coumarines has also been studied.     

Purification of Commercially Prepared Bovine Trypsin by Reverse Phase High Performance Liquid Chromatography

Abstract

A reverse phase high performance liquid chromatographic method for the purification of milligram amounts of commercially prepared bovine trypsin is described. Increased specific enzymatic activity is observed in the purified material. The advantages of using purified trypsin in protein digestions as well as the potential application of reverse phase high performance liquid chromatography for enzyme characterization are described.     

Reverse Phase High Performance Liquid Chromatography of Human Bence Jones Proteins

Abstract

A high performance liquid chromatography system is presented for analytical and preparative separations of human Bence Jones proteins. The method utilizes 5–7 um macroreticular polystylene resin with bonded hydroxymethyl functional groups, and the proteins are eluted with a linear gradient of an increasing concentration of acetonitorile(10–60%, V/V) in 0.1 % (V/V) trifluoroacetic acid, pH 2.1. By this elution condition, seven X type Bence Jones proteins with molecular weights of 23,600 (monomer)-47,000(dimers) daltons(216–434 amino acids) were eluted within 80 min with the yields of 78%–98%. The method allows a rapid and sharp elution of Bence Jones proteins.     

反相高效液相色谱法测定苯醚甲环唑原药的含量

建立了反相高效液相色谱法测定苯醚甲环唑原药含量的方法,乙腈―水（体积比60∶40）为流动相,检测波长225 nm,流速1.0 m L/min,ODS C18色谱柱（250 mm×4.6 mm（i.d.）,2.5um）,紫外检测器。平均回收率为103.6%,标准偏差为0.37%,变异系数为0.38%;方法线性相关性系数0.999 7,适用于产品质量的检测分析。     

反相高效液相色谱分离纯化天然除虫菊酯

天然除虫菊酯含有6种有效成分,但结构相似,分离困难。本研究以除虫菊酯精油为原料,通过优化反相高效液相色谱分离条件,分离纯化了除虫菊酯6种有效成分,纯度均达到99%。然后用气相色谱-质谱定性分析确证6种成分。本研究所建立的天然除虫菊酯6种有效成分的反相高效液相色谱分离纯化方法,为天然除虫菊酯除虫菊酯残留检测提供了准确的判断标准,为阐明天然除虫菊酯6种单一有效成分的杀虫机理研究奠定了基础,也为除虫菊酯6种单一有效成分纯品的生产提供了重要的方法参考。     

高效液相色谱手性固定相对氨基酸衍生物对映体分离研究

利用高效液相色谱手性固定相对光学异构体分离分析日益受到人们的重视。高效液相色谱氨基酸酰胺型手性固定相对拆分对映异构体具有较好的效果。本文为了探讨该类手性固定相对氨基酸的拆分影响,用L-异亮氨酸叔丁酰胺型手性固定相,对6种DL氨基酸(丙氨酸、2-氨基-n-丁酸、缬氨酸、亮氨酸、苯丙氨酸和蛋氨酸)的18种衍生物进行了拆分,并讨论了其衍生物取代基对手性拆分的影响及从立体化学角度提出了手性拆分机理。实验结果表明,这种手性固定相对外消旋氨基酸衍生物有较好的拆分能力。 1 实验部分 1.1 仪器和试剂 Varian 2010型高效液相色谱仪(美国);2050型紫外检测器及HP3394型积分仪(美国);L-异亮氨酸叔丁酰胺型手性柱(TBILS,上海药物研究所),粒度10μm,柱长100mm、内径4mm的不锈钢柱。 异丙醇(分析纯)用前重蒸,石油醚(60～90℃)分析纯,重蒸收集69℃镏分,用前全经过0.5μm薄膜并脱气。DL-氨基酸衍生物[丙氨酸(Ala),2-氨基-n-丁酸(Buty),缬氨酸(Val),亮氨酸(Leu),苯丙氨酸(Phe),蛋氨酸(Met)]用相应DL-氨基酸参考文献方法进行N-乙酰化及酯化。衍生物经元素分析合格,配制成5mg/mL溶液。