氨基酸衍生物在毛细管区带电泳下的分离研究

采用新合成的荧光试剂咔唑-9-乙基氯甲酸酯作为柱前衍生试剂,利用毛细管区带电泳法对衍生氨基酸进行分离,考察了该试剂用于毛细管区带电泳法进行氨基酸分离的关键条件,实现了12种氨基酸的快速基线分离.     

烟草及烟用香精香料中氨基酸检测方法研究进展

烟草及烟用香精香料中氨基酸的含量对烟草品质有着直接而重要的影响，因此有必要进行准确定性定量分析.氨基酸检测方法可分为直接检测法和间接检测法两大类，间接检测法又可分为柱前衍生法和柱后衍生法.分析了每种方法的优缺点以及常用的衍生化试剂，以期为进行氨基酸分析的研究者提供参考.     

AcQ·Tag~(TM)法测定鱼粉水解液中的氨基酸

１前言鱼粉是动物性蛋白质饲料,过去鱼粉中氨基酸测定大多采用氨基酸分析仪以离子交换树酯分离,柱后衍生测定。但氨基酸分析仪价格昂贵,分析时间长。近年来,柱前衍生高效液相色谱法测定氨基酸的技术因其具有衍生步骤简单,速度快,剩余试剂不干扰测定等优点,而有逐渐...     

氨基酸的咔唑-9-乙基氯甲酸酯柱前衍生高效液相色谱分析

采用一种新型氯甲酸酯类荧光衍生试剂咔唑－9－乙基氯甲酸酯（CEOC）对氨基酸进行柱前衍生，并通过荧光检测的方法进行高效液相色谱（HPLC）分析；在Hypersil BDS C18柱上对19种氨基酸衍生物进行了分离，衍生物的激发和发射波长分别为λex=293nm,λem=365nm,当衍生化缓冲液pH为9．0，衍生试剂总量（摩尔数）为氨基酸总量的4倍时，衍生化产率最大，平均检出限可达22．8fmol。     

高效液相色谱法（硝基氟苯柱前衍生化和紫外吸收）测定猪血蛋白氨基酸组成

目前所采用的氨基酸分析法,在柱前或柱后衍生化后,大多数都用价格昂贵的荧光检测器进行测定。近年来已有人采用一些新的衍生化试剂和方法,改用紫外吸收直接测定,取得了很大成功。我们采用1-氟-2,4-二硝基苯(FDNB)柱前     

OPA柱前衍生反相高效液相色谱法测定氨基酸含量

 建立了邻苯二甲醛（ＯＰＡ）手动柱前衍生反相高效液相色谱法测定样品中氨基酸含量的方法。以邻苯二甲醛（ＯＰＡ）／３－巯基丙酸（３－ＭＰＡ）为衍生试剂进行衍生，ＯＤＳ柱分离，３４０ｎｍ检测，在４０ｍｉｎ内１８种氨基酸全部得到基线分离。测定牛血清白蛋白（ＢＳＡ）的氨基酸组成和小鼠血清中的游离氨基酸，取得满意的结果。     

新型柱前衍生试剂对甲氧基苯磺酰氟用于氨基酸毛细管电泳分离

大多数氨基酸不含有芳香环等生色团,在紫外可见光区无吸收,无法直接用紫外和荧光法检测,因此将氨基酸衍生为具有较强紫外或荧光吸收的衍生物进行检测是常用的分析方法.本实验采用常温下能与氨基酸快速反应的对甲氧基苯磺酰氟(MOBS-F)为新型柱前衍生试剂,建立了毛细管电泳分离常见的20种氨基酸的方法.     

反相高效液相色谱法测定深层培养含硒构菌菌丝体中的硒蛋氨酸

本文用苯异硫氰酸酯作柱前衍生化试剂，衍生的硒蛋氨酸及其它非硒代氨基酸用反相高交液相色谱进行分离分析，分析表明，加无机硒培养的构菌菌丝体中含一定量的硒蛋氨酸，且这一方法广泛适于生物物质中其它硒代氨基酸的分析。     

乙酰葡萄糖异硫氰酸酯柱前手性衍生化拆分氨基酸立体异构体的反相高效液相色谱法研究

本文用乙酰葡萄糖异硫氰酸酯(GITC)作柱前手性衍生化试剂,反相高效液相色谱法拆分氨基酸立体异构体。研究了不同构型氨基酸与GITC反应后所生成的衍生物的紫外吸收响应特点。探讨了16种氨基酸GITC衍生物的色谱分离情况。     

新型紫外衍生试剂用于D-氨基酸的毛细管电泳分离

采用一种新型紫外衍生试剂对乙酰氨基苯磺酰氟(PAABS-F)作为柱前衍生试剂,成功地对19种标准D-氨基酸和甘氨酸进行了衍生和毛细管电泳分离。详细研究了各种分离条件对毛细管电泳分离的影响。实验结果表明:采用20 mmol/L硼砂(pH 9.3),126 mmol/L十二烷基硫酸钠,8 mmol/Lβ-环糊精和20mmol/L NaC l时分离结果最佳,16 m in内实现了20种氨基酸的基线分离。     

Homocyclic o-dicarboxaldehydes: Derivatization reagents for sensitive analysis of amino acids and related compounds by capillary and microchip electrophoresis

Amino acids are essential compounds for living organisms, and their determination in biological fluids is crucial for the clinical analysis and diagnosis of many diseases. However, the detection of most amino acids is hindered by the lack of a strong chromophore/fluorophore or electrochemically active group in their chemical structures. The highly sensitive determination of amino acids often requires derivatization. Capillary electrophoresis is a separation technique with excellent characteristics for the analysis of amino acids in biological fluids. Moreover, it offers the possibility of precapillary, on-capillary, or postcapillary derivatization. Each derivatization approach has specific demands in terms of the chemistry involved in the derivatization, which is discussed in this review. The family of homocyclic o-dicarboxaldehyde compounds, namely o-phthalaldehyde, naphthalene-2,3-dicarboxaldehyde, and anthracene-2,3-dicarboxaldehyde, are powerful derivatization reagents for the determination of amino acids and related compounds. In the presence of suitable nucleophiles they react with the primary amino group to form both fluorescent and electroactive derivatives. Moreover, the reaction rate enables all of the derivatization approaches mentioned above. This review focuses on articles that deal with using these reagents for the derivatization of amino acids and related compounds for ultraviolet-visible spectrometry, fluorescence, or electrochemical detection. Applications in capillary and microchip electrophoresis are summarized and discussed.     

Complete separation of ephedrines by graphite-layer open-tubular (GLOT) column

Summary A reversed phase high performance liquid chromatographic (RP-HPLC) method for the analysis of amino acids in kelp, using precolumn derivatization, is described. Phenylisothiocyanate (PITC) was used as the reagent for derivatization. The kelp samples were prepared by microwave hydrolysis in only 5 min; seventeen PTC amino acids were separated after hydrolysis and derivatization within 12 min. The coefficients of variation were >1.94 and the correlation coefficients for concentration versus response were >0.999 for all derivatives. The ratio of branched amino acid (BAA) to aromatic amino acid (AAA) was also studied. The method has the advantage of shorter hydrolysis and analysis times with optimum separation. In addition, it gives high repeatability of retention times and peak areas for all the amino acids present.     

Rapid analysis of amino acids using pre-column derivatization

A new approach to the pre-column derivatization and analysis of amino acids is described. The method is based upon formation of a phenylthiocarbamyl derivative of the amino acids. The derivatization method is rapid, efficient, sensitive, and specific for the analysis of primary and secondary amino acids in protein hydrolyzates. The liquid chromatographic system allows for the rapid, bonded-phase separation with ultraviolet detection of the common amino acids with 12-min analysis time and a 1-pmol sensitivity.     

Measurement of plasma and urine amino acids by high-performance liquid chromatography with electrochemical detection using phenylisothiocyanate derivatization

The use of reversed-phase liquid chromatography (LC) with pre-column derivatization for the analysis of amino acid mixtures is becoming established as a possible cheaper alternative to commercial amino acid analysers. The available derivatization procedures all have disadvantages when applied to clinical samples, partly due to the interferences found with body fluids when ultraviolet or fluorescence detection is used. An LC method is described for the separation of amino acids in blood or urine, using pre-column derivatization with phenylisothiocyanate (PITC), gradient elution and electrochemical detection. The use of electrochemical detection of PITC derivatives virtually eliminates interferences and enables the secondary amino acids to be measured. Examples are shown of normal urine and plasma and samples from patients with cystinuria and maple syrup urine disease.     

Analysis of amino acids as formamidene butyl esters by electrospray ionization tandem mass spectrometry.

Amino acid formamidene butyl esters are optimally prepared by heating amino acids with dimethylformamide dimethylacetal (DMF-DMA) for 2 minutes at 65 degrees C and then with n-butanol/hydrogen chloride for 15 minutes at 65 degrees C. The formamidene butyl esters of simple alpha-amino acids and beta-amino acids afford between 1.1 and 20 times the signal intensity of the corresponding butyl esters during electrospray ionization tandem mass spectral analysis. Complex alpha-amino acids, such as ornithine, arginine and citrulline, and gamma-amino acids are better analyzed as butyl esters. Secondary alpha-amino acids, such as proline and sarcosine, give a mixture of two iminium salts with the DMF-DMA derivatization method. A derivative incorporating two molecules of the amino acid predominates at higher derivatization temperatures. Contrary to a previous report, N-formylamino acids were not detected. The presence of secondary amino acids in amino acid mixtures, derivatized as formamidene butyl esters, affords derivatives that incorporate two different amino acids. The new formamidene butylation method is unlikely to replace the butylation procedure used by neonatal blood spot screening programs for amino acid disorders, since a much poorer response was obtained with formamidene butyl esters of arginine and citrulline, important in the diagnosis of arginase deficiency and citrullinaemia.     

Amino acid analysis using DABS-CL

Summary Pre-column derivatization followed by reversed-phase HPLC has become widely used as a sensitive and speedy method of amino acid analysis.The technique has been improved by the development of simple and reproducible strategies for derivatization of the amino acids with DABS and for the HPLC analysis. Results obtained indicate that the method has a high sensitivity is well and suited to generate precise amino acid composition information from peptides and proteins.     

Gas chromatography-mass spectrometry analysis of amino acid enantiomers as methyl chloroformate derivatives: application to space analysis

This work describes a GC-MS method for enantioselective separation of amino acids. The method is based on a derivatization reaction which employs a mixture of alkyl chloroformate-alcohol-pyridine, as reagents to obtain the N(O,S)-alkyl alkoxy carbonyl esters of amino acids. Various reaction parameters are investigated and optimized to achieve a reproducible derivatization procedure suitable for separation of amino acid enantiomers on Chirasil-L-Val chiral stationary phase. In particular, the following topics are investigated for 20 proteinogenic amino acids: (i) the proper reagent and reaction conditions to obtain the highest derivative yield; (ii) the amino acid reactivity and the MS properties of the obtained derivatives; (iii) the linearity and sensitivity of the analytical method; (iv) the retention behavior of the derivatives and their enantiomeric separation on the Chirasil-L-Val chiral stationary phase. By combining the resolution power of the Chirasil-L-Val column and the high selectivity of the SIM MS detection mode, the described procedure enables the enantiomeric separation and quantification of 16 enantiomeric pairs of amino acids. The procedure is simple and fast and reproducible. It displays a wide linearity range at ppb detection limits for quantitative determinations: these properties make this derivatization method a suitable candidate for amino acid GC-MS analysis on board of the spacecrafts in space exploration missions of solar system body environments.     

Determination of Amino Acids in an Individual Erythrocyteby Capillary Electrophoresis with Intracellular FITC-derivatization and Laser-induced Fluorescence Detection

A novel approach for analysis of amino acids in individual erythrocytes was established.In this method, the derivatization reagent was in~oduced into the living cells by electroporation.After derivatization, the amino acids in a single cell were determined by capillary electrophoresis with laser-induced fluorescence detection.     

Laser-induced fluorescence detection in liquid chromatography after preliminary derivatization of carboxylic acid and primary amino groups

The development of selective derivatization for the determination of carboxylic acids, amino acids and peptides in aqueous solutions is described as a preliminary study for the determination of these compounds in biological materials. The derivatization reactions are completed before the liquid chromatographic separation and laser-induced fluorescence detection for which a continuous-wave argon-ion gas laser is used in the ultraviolet or visble mode. Carboxylic acid groups arre derivatized with 9-hydroxymethylathracene and primary amino groups are derivatized with fluorescein isothiocyanate. Detection limits, in aqueous solutions, for the carboxylic acid derivatives are ca. 190 fg (ultraviolet mode). In the visible mode, the detection limits are ca. 1 fg for the primary amino derivatives of amino acids and peptides. In al the chromatographic analyses, the derivatization mixtures are injected onto a standard reversed-phase or reversed- phase ion-pair system and conventional flow cells are used without expensive photon counting or optical systems.