Analysis of Amino Acids in a Single Human Red Blood Cell by Capillary Zone Electrophoresis with Intracellular NDA—derivatization and Electrochemical Detection

A novel method for determination of amino acids in individual red blood cells has been developed. In this method, the derivatization reagents (NDA and CN^-) are introduced into living cells by electroporation. After completion of derivatization,the amino acids in a single cell is determined by capillary zone electrophoresis with end-column amperometric detection.     

Rapid Determination of Amino Acids in Ginseng by High Performance Liquid Chromatography and Chemometric Resolution

Ginseng is one of the most important traditional Chinese medicines and functional foods.A method for the fast determination of amino acids in ginseng samples using high performance liquid chromatography（HPLC） was developed,in which strong isocratic elution was employed for simplifying the separation and speeding up the analysis.All amino acids were eluted within 3 min with the chromatogram composed of overlapped peaks from the interferences.Then,non-negative immune algorithm（NNIA） was adopted to resolve the chromatographic signals of the components from the chromatogram measured.The results show that the signals of the amino acids can be correctly extracted by NNIA and the signal extracted can be used for the quantitative analysis.The method was validated via determining six amino acids of four different samples of ginseng.The recoveries of the spiked samples are in a range of 96.6％-106.3％.     

碳纳米管为基质对氨基酸的MALDI-FT MS分析

Twenty common amino acids have been analyzed successfully by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) using carbon nanotubes as matrix. From the spectra, little or no background interference or fragmentation of the analytes has been observed. This method was also applied to the analysis of amino acid mixture successfully. Carbon nanotubes have some features such as large surface area to disperse the analyte molecules sufficiently and prevent the sample aggregation and strong ultraviolet absorption to transfer energy easily to the analyte molecules. The present method has potential application for the rapid and sensitive analysis of amino acids and their mixture.     

Sensitive analysis of amino acids in injection liquor by reversed-phase HPLC

A convenient derivatization method of amino acids with l-fluoro-2,4-dimtrobenzene as reaction reagent and a separation system were described. The derivative amino acids were separated on a specific chemically bonded phase column with a simple linear gradient elution consisting of aqueous buffer and methanol. The eluate was detected by common ultraviolet absorption detector at 360 nm. The detection limits of amino acids were as low as 10 picomole. This method has been successfully applied to assay amino acid injection liquor used in hospital. It has good repro-ducibility and precision. The procedures avoid the requirements of particular derivative equipment and analyzer employed in conventional amino acid analysis.     

Characterization on the Mean Molar Absorptivity of Amino Acids in Microbial Lipopeptides

The molar absorption coefficients of each of 14 kinds of amino acids were determined by the spectrophotometric method, and the mean molar absorption coefficients of 37 different mixtures of each with amino acid composition exactly equivalent to that of the peptide chain of the corresponding lipopeptide were determined based on calculation or experimental. The significance of the results is that the mean molar absorption coefficients strongly demonstrate the regular patterns, though different amino acids bear quite different molar absorption coefficients.     

Determination of D- and L-amino acids in biological samples by two-dimensional column liquid chromatography

Summary A two-dimensional, column liquid chromatographic system is used for the determination of the D- and L-enantiomers of amino acids in biological samples. Separation of the amino acids is first on ion-exchange column by gradient elution with a sodium citrate-sodium chloride buffer. Enantioseparation is by subsequent injection of 3 μl heart-cuts of the individual amino acids onto a second column with a chiral crown ether stationary phase. Finally, fluorescence detection is after post-column labelling of the amino acids using ano-phthalaldehyde-2-mercaptoethanol reagent solution. The high separation power and selectivity of the system allow processing of complex samples with hardly any additional treatment and the determination of small quantities of D-amino acids in the presence of excess L-form. Applicability of the system is illustrated by the determination of D- and L-aspartate, serine, glutamate and alanine in various complex biological samples, such as protein hydrolysates, urine and biotechnological and food samples. Data are given on detectability, repeatability and linearity.     

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.     

Total synthesis of cyclic heptapeptide euryjanicin B

The first synthesis of the naturally occurring cyclic peptide euryjanicin B has been achieved.A general method was described to synthesize the cyclic peptide by a two-step solid-phase/solution synthesis strategy.All the amino acids in this study are L-configuration. The linear heptapeptide was assembled by standard Fmoc chemistry on solid-phase and subsequently cyclization was carried out by solution method.     

Helical secondary structures and supramolecular tilted chirality in N-terminal aryl amino acids with diversified optical activities

Helix structures at atomic/molecular level have not been found in self-assembled peptide seque nce with less than three residues.As β-sheet supramolecular secondary structures have been discovered in solidstate amino acids,we here report the conjugation of simple N-terminal aryl protecting group could give rise to helical supramolecular secondary structures in solid-state,which determines the optical activities of the adjacent aryl groups.The carboxylic acid-involved asymmetric H-bonds in N-te rminal aryl amino acids induce the emergence of super-helical structures of amino acid residues and aryl groups.In most cases,supramolecular tilted chirality of aryl groups is opposite to that of amino acid sequences,of which handedness and helical pitch are determined by the H-bond modalities.Determining correlation between supramolecular tilted chirality of aryl segments and their chiroptical activities is firstly unveiled,which was verified by the computational results based on density functional theory.Most aryl amino acids self-assembled by nanoprecipitation method via crystallization induced self-assembly into rigid one-dimensional microstructures with ultra-high Young's modulus.This study reveals the generic existence of chiral supramolecular structure s in aggregated amino acid derivatives and gives an in-depth investigation into the structural-property relationships,which could guide the rational design and screening of chiroptical supramolecular materials.     

Chemiluminescence determination of 20 amino acids and the mechanism study

<正>It was found that 20 amino acids could inhibit the intensity of the luminol-H_2O_2-CuSO_4 chemiluminescence system.Using this character,a rapid and sensitive method for the determination of 20 amino acids was developed with flow injection coupled with chemiluminescence detection.Under the optimal conditions,the detection limits of 20 amino acids were in the range of 4.5×10~(-7)- 4.3×10~(-10) mol/L,and the relative standard deviations were less than 3.2%.The proposed method was successfully applied to drug analysis.The possible mechanism was also discussed.     

Quantitative analysis of 17 amino acids in tobacco leaves using an amino acid analyzer and chemometric resolution

A method was developed for quantifying 17 amino acids in tobacco leaves by using an A300 amino acid analyzer and chemometric resolution. In the method, amino acids were eluted by the buffer solution on an ion‐exchange column. After reacting with ninhydrin, the derivatives of amino acids were detected by ultraviolet detection. Most amino acids are separated by the elution program. However, five peaks of the derivatives are still overlapping. A non‐negative immune algorithm was employed to extract the profiles of the derivatives from the overlapping signals, and then peak areas were adopted for quantitative analysis of the amino acids. The method was validated by the determination of amino acids in tobacco leaves. The relative standard deviations (n = 5) are all less than 2.54% and the recoveries of the spiked samples are in a range of 94.62–108.21%. The feasibility of the method was proved by analyzing the 17 amino acids in 30 tobacco leaf samples.     

Chromatographic amino acid analysis of protein hydrolysed in polyacrylamide after removal of ammonia

Summary A simple method for the removal of NH₃ from amino acids is presented. The method is based on a cation-exchange resin from which amino acids are eluted with NH₄OH. The eluent is then removed under reduced pressure. The method allows the ninhydrin-based detection of amino acids after hydrolysis of stained protein bands in polyacrylamide gels. This was previously not possible since NH₃ produced by the hydrolysis of polyacrylamide interferes with the ninhydrin-detection of basic amino acids. The method should also be applicable to the detection of amino acids with o-phthalaldehyde.     

中国梨木虱分泌物中氨基酸的分离与分析

利用离子交换色谱法分离了中国梨木虱分泌物中的氨基酸;利用毛细管气相色谱法对梨木虱分泌物中氨基酸的三氯乙酰了酯衍生物进行了分析。用标准样对照定性,内标法定量。分泌物中共检出13种氨基酸。     

Chiral separation and determination of the optical purity of thiamphenicol-related chiral compound by capillary zone electrophoresis with cyclodextrins as chiral selectors

Summary One classical method for quantitation of amino acids in proteins is hydrolysis of the proteins and determination of the free amino acids. Although the drastic experimental conditions necessary for complete hydrolysis always cause degradation of some of the amino acids, if mild hydrolysis conditions are used, a mixture of amino acids and oligopeptides is obtained. If these conditions are adequately tuned, the oligopeptides are almost exclusively dipeptides. For this reason we have initiated a study to find a derivatizing agent suitable for the analysis of amino acids and dipeptides by an absolute method of quantitation already tested for amino acids. FMOC-Cl was found to be a suitable derivatizing agent for this purpose.     

Determination of the specific radioactivity of amino acids by a combination of thin-layer chromatography and quantitative autoradiography

A new method for the analysis of the specific activity of amino acids is described. The analysis is carried out by thin-layer chromatography of the dansylated amino acids, computerized fluorescence evaluation and activity measurement by quantitative autoradiography. Quantitative evaluation of the autoradiographs is achieved by careful calibration of the X-ray film blackening. As shown for 14C-labelled phenylalanine and tyrosine, the method allows the simultaneous determination of the specific activity of 22 amino acids. About 10(-13) mol of an amino acid with a specific activity of less than 5 GBq/mmol can be detected and measured by this method.     

HPLC Analysis of Amino Acids with Ion Exchange Chromatography and O-Phthalaldehyde Post-Column Derivatization: Applications to the Assay of Endogenous Free Amino Acids and Their Transport in Human Placental Villus

Abstract

A method using high performance liquid chromatography (HPLC) for the analysis of primary amino acids in human placenta is described. This method involves separation of primary amino acids by high performance ion-exchange chromatography followed by post column derivatization using O-phlthalaldehyde (OPA) and 2-mercaptoethanol and fluorescence (excitation 340 nm and emission 410 nm) detection of derivatives. Waters 840 HPLC Amino Acid System was used for this purpose.

For analysis, villus tissue was extracted with acetonitrile, and the recovered amino acids were reconstituted in a sodium diluent (pH 2.2). The complete profile of the primary amino acids in the sample could be constructed in about 90 minutes. Up to 44 samples can be analyzed without special attention. Using this method, essential amino acids (threonine, valine, methionine, isoleucine, leucine, tyrosine, phenylalanine, lysine, histidine) and nonessential amino acids (aspartic acid, serine, glutamic acid, glycine, alanine, arginine) were detected and quantified in human placental villus in pmol quantities. Plots of peak heights (or areas) were linear for several amino acids. The same method was also used for (a) the assay of free primary amino acids in umbilical bloods, (b) the efflux of amino acids from isolated human placental villus, and (c) to study the uptake of α-aminoisobutyric acid (AIB), a non-metabolizable amino acid, by the isolated placental villus.     

GasChromatography of Amino Acids as their N-Trifluoroacetyl Methyl Esters and Comparison with the Ion-exchange Method in Maize and Oats

A method for the preparation of N-trifluoroacetyl methyl esters of amino acids has been developed, permitting the separation and determination of 20 amino acids by temperature-programmed gas chromatography on two columns. The method was applied to the analysis of the amino acids in hydrolyzed maize and in hydrolyzed oats, where a direct comparison was made with the ion-exchange method.     

Development and validation of a hydrophilic interaction ultra‐high‐performance liquid chromatography with triple quadrupole MS/MS for the absolute and relative quantification of amino acids in Sophora alopecuroides L.†

Amino acids are naturally occurring compounds in many edible or medicinal plants, which possess a variety of pharmacological effects on humans. The aim of this study is to develop and validate a hydrophilic interaction LC coupled with MS/MS method for the absolute and relative quantification of amino acids without derivatization. The application of this method has been proven through 20 naturally occurring amino acids in 21 samples from different parts and phenological growth stage of Sophora alopecuroides. The method was performed on an ultra‐high performance LC separation system coupled with ESI‐MS on a triple quadrupole mass spectrometer. The proposed absolute quantitative method was fully validated in terms of linearity, sensitivity, precision, repeatability as well as recovery. The analysis results showed that S. alopecuroides is rich in free amino acids. In addition, relative quantitative determination of amino acids with several amino acids selected for the best accuracy was investigated. The accuracies of relative quantitative method for amino acids determinations suggest that it is feasible to quantify amino acids by the proposed relative quantitative determination method, which contributes to breaking through the choke point of lack of standards.     

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.     

PICO－TAG反相色谱法测定鸡蛋中氨基酸含量

采用ＰＩＣＯ－ＴＡＧ反相色谱法测定了氨基酸，方法的相对标准偏差为０．７５％，平均回收率为９７．１２％。对鸡蛋样品进行了测定，发现不同产地不同皮色鸡蛋中氨基酸含量无显著差异，样品测定相对标准偏差为１．６７％。结果表明测定方法快速、灵敏、准确、重现性好。