Multiple gradient development TLC in analysis of complex phenolic acids fromLycopus europeaus L.

Summary Phenolic acids fromLycopus europaeus L. (Lamiaceae) were chromatographed on thin layers of precoated silica, LiChrospher silica, propylamine, diol layers using stepwise multiple development and, additionally, on an octadecyl layer with an isocratic mobile phase. The eluent contained decreasing concentrations of formic acid in a mixture of diisopropyl ether-cyclohexane for silica gel plates or methyl-ethylketone inn-hexane for diol plates and increasing concentrations of acetic acid in acetone for the propylamine layer. Extracts were separated into 25 components, 14 of which were identified using a mixture of standards. 2,4-Dihydroxybenzoic, gentisic, protocatechuic and 3,5-dihydroxybenzoic acids were ascribed to this genus for the first time. The adsorbent-eluent systems described were very efficient and rapid.     

TLC Resolution of Amino Acids in a New Solvent and Effect of Alkaline Earth Metals

Abstract

A new solvent system for the resolution of 15 component mixture of amino acids and the effect of alkaline earth metals on the resolution is reported, the TLC was carried out using plain and impregnated silicagel plates and, solutions of amino acids in tris buffer and pretreated with metal hydroxides.     

The application of gradient saturation in unidimensional planar chromatography of polar lipids. Part 1: Presentation of the HPTLC system

A thermostated HPTLC system was assembled to test the influence of gradient chamber saturation upon resolution of polar lipids chromatographed on boric acid-impregnated silica gel plates. Inexpensive mixtures of nonacidic (14 component peaks) and acidic (8 component peaks) lipids were prepared from soya lecithin by a simplified procedure involving DEAE-Sephadex liquid column chromatography to monitor performance of the system using a readily available reference material. The system requires ca. 5 min chamber presaturation after which the polar lipids are chromatographed in less than 30 min. Retention times (t_R) as well as peak area % values of all component peaks and measured by scanning photodensitometry to demonstrate the influence of different modes of chamber saturation upon the resolution of polar lopids.     

TLC Separations of Amino Acids on Silica Gel Impregnated Layers

Abstract

Copper sulphate and polyamide were tried as impregnants for improving the separation of twenty amino acids on silica gel ‘G’ layers using a new solvent system MeOH-BuOAc-AcOH-Pyridine(20:20:10:5). Tables are presented to illustrate the improvement in resolution of amino acids on silica gel plates.     

Identification of subnanomole amounts of PTH-amino acids by high performance thin-layer chromatography

Summary High performance thin-layer chromatography (HPTLC) of PTH-amino acids on 5×5 cm silica gel plates precoated with a fluorescence marker gives 10–20 fold increase in sensitivity compared to ordinary silica gel plates. Separation of PTH-Leu from PTH-Ile is easily achieved in contrast to chromatography on polyamide sheets. Only two solvent systems are required and as many as 12 samples can be chromatographed on each plate. However, if the sample is contaminated with N-phenylthiourea a third solvent system is necessary.     

A comparative study of HPLC and TLC separation of amino acids using Cu(II) ion

Separation of amino acids using Cu(II) by RP-HPLC and impregnated TLC is reported. For HPLC the sample mixture of amino acids containing copper (II) was injected into the column while for TLC the silica gel plates were impregnated with Cu(II). The mobile phase used in HPLC was acetate buffer (0.3 M , pH 6.0)-acetonitrile (9:1, v/v), and that in TLC was acetate buffer (0.3 M , pH 6.0):acetonitrile-n-butanol (12:5:10, by vol.). The results have been compared and discussed.     

Ion Exchange Application of Overpressured Thin-Layer Chromatography

Abstract

The application of overpressured thin-layer chromatography introduced into the field of ion exchange chromatography. The basic differences between overpressured thin-layer chromatography and classical thin-layer chromatography are discussed including the distinction between the separations performed on thin-layer plates containing silica gel and a mixture of ion exchanger material and silica gel. The basic increase of flow velocity of solvent front with the aid of a pressurized ultra-micro chamber and the effect of flow velocity on the height equivalent of the theoretical plates are also presented. For basic amino acids, the flow velocity vs plate height curves show optima at a moderately high rate of development.     

Resolution of enantiomers of DL-amino acids on silica gel plates impregnated with optically pure (-)-quinine.

TLC resolution of enantiomers from racemic amino acids was achieved on silica gel plates impregnated with optically pure (-)-quinine. The successful solvent systems were butanol-chloroform-acetic acid (3:7:5, v/v) for DL-methionine; 6:8:4, v/v for alanine; 10:1:4; v/v for threonine; and ethyl acetate-carbon tetrachloride-propionic acid (10.5:6.5:3.5, v/v) for valine. Minimum detection limits were found to be different for each of the amino acid, ranging between 0.9 and 3.7 microg. The effects of concentration of impregnating reagent, temperature and pH on resolution of enantiomers have been studied in details.     

Separation of DABS derivatives of amino acids by multiple gradient development (MGD) in thin-layer chromatography

Summary A mixture of 13 DABS-amino acids has been chromatographed on high-performance silica gel layers developed with eluents containing increasing concentrations of ethyl acetate in heptane + chloroform, using a modification of stepwise multiple development MGD described in a earlier paper. Densitograms were obtained at 485 nm. The MGD method was very efficient, separating all 13 DABS-amino acids, and rapid, owing to the use of a non-aqueous mobile phase.     

用铜（Ⅱ）—L—精氨酸配体交换薄层色谱拆分氨基酸对映体

报道了一种新的配体交换薄层色谱拆分氨基酸对映体的方法。以醋酸铜－Ｌ－精氨酸的络合物为配体交换剂，用浸渍的方法吸附在硅胶薄层板上，制成配体交换薄层，用ＰＲＩＳＭＡ优化法选择出展开剂的最佳组成为：甲醇／乙腈／四氢呋喃／水＝８０：８．２：５．８：６，在此色谱条件下，十对氨基酸对映体得到良好的分离，Ｄ－和Ｌ－氨基酸的相对比移值在１．０９－２．４０之间。文中对配体交换薄层的制备方法，样品的分子结构及色谱行为     

Direct TLC Resolution of the Enantiomers of Three β-Blockers by Ligand Exchange with Cu(II)–l-Amino Acid Complex, Using Four Different Approaches

TLC experiments have been performed to resolve the enantiomers of three β-blockers by complexation chiral TLC with Cu(II) cation and five l amino acids. Loading was carried out by using the Cu(II)–l amino acid complex as chiral mobile phase additive with untreated silica gel plates; by mixing the Cu complex with silica gel before preparing the TLC plates; by ascending development of untreated plates with solutions of the Cu complex; and by using a solution of Cu(II) acetate as mobile phase additive for plates prepared by mixing the l amino acid with silica gel. Spots were located by use of iodine vapour.     

Determination of Free Fatty Acids by Diphasic-Two Dimensional TLC-Fluorescence Spectrodensitometry

Abstract

A sensitive method for the determination of fatty acids is presented. Free fatty acids (20pg-lμg), were first covalently reacted in a displacement reaction with a 70 μM solution of the fluorescent probe, 4-bromomethy 1–6, 7-dimethoxy-coumarin (Br- Mdmc) and the fluorescence-labelled fatty acid esters were separated in a diphasic-two dimensional high performance thin layer chromatographic system (diphasic-2D-TLC). This system consisted of a reversed phase C18 layer (2 × 10 cm) interfaced with a AgNO3-modified silica gel layer (10 × 10 cm). Aliquots of the reeaction mixture were streaked onto the C18 layer, and the plates developed in the first dimension using acetonitrile-acetone-methanol-water (60:20:10:10, v/v/v/v) (solvent 1) as the mobile phase. Development in this dimension gave separation based on the number of carbons. Following the first development, the plates were dried and the silica gel layer impregnated with a saturated solution of AgNO3 in methanol. The plates were then predeveloped in the second dimension in chloroform-ethyl acctate-acetonitrile (90:8:2, v/v/v) (solvent 2) to the plate interphase, and developed in solvent 2 in the second dimension. Development in the AgNO3-modified silica gel, allowed separation based on the number of double bonds. The fluorescent bands were scanned with a Shimadzu CS-9000 spectrodensitometer in the fluorescent mode, using 352 nm as the excitation wavelength and a cut-off filter at 400 nm. Baseline resolution was obtained for all 15 fatty acids tested. The lower linear detector response extended to 0.13 pmol. This method provids a sensitive means of analysis of fatty acids present in biological systems.     

Determination of amino acids in human cerebrospinal fluid by gas chromatography

In the present study, the use of gas chromatography (GC) for the determination of amino acids in human cerebrospinal fluid (CSF) is described. Although some amino acids may be determined using a packed column following the removal of glucose, the major interfering component, the inadequate resolution of other amino acids from remaining unidentified components results in poor quantitation. The use of wide bore columns improves reproducibility considerably, but still it does not provide sufficient resolution to enable quantitative determination of all amino acids in human CSF. Good reproducibility data, with CV values for all amino acids of 7% or less and recoveries generally between 80% and 100%, can only be obtained using the fused silica open tubular (FSOT) column. Normal amino acid levels are presented for 10 samples of human CSF, which compare well with data previously reported in the literature.     

Detection of gangliosides with the fluorochrome NBD dihexadecylamine and its application for preparative high performance thin layer chromatography.

A new method for the detection of gangliosides based on the lipophilic fluorescence agent 4-(N,N-dihexadecyl)amino-7-nitrobenz-2-oxa-1,3-diazole (NBD dihexadecylamine) and its application for preparative high performance thin layer chromatography is described. Brain gangliosides were chromatographed on silica gel coated thin layer plates and located with non-destructive fluorochrome under longwave ultraviolet light. The fluorescent zones were scraped off and the gangliosides were extracted with a mixture of chloroform/methanol/water (30/60/8; v/v/v). The gangliosides were separated from uncharged NBD dihexadecylamine by anion exchange chromatography and impurities were removed by Iatrobeads chromatography. The method described offers a simple and successful preparative thin layer chromatographic strategy to obtain pure gangliosides in microgram and milligram quantities.     

A Comparison of Amino Acid Separations on Silica Gel,Cellulose, and Ion Exchange Thin Layers

Abstract

Separations of amino acids on silica gel, high performance silica gel, preadsorbent silica gel, microcrystalline and fibrous cellulose, and Fixion strong acid cation exchange layers were compared under standardized conditions using a mixture of the nine essential acids and a more complex 18-component mixture. Fixion layers were evaluated using three different development conditions, and loading studies of raw urine were carried out on all of the layers. Tables and figures are presented illustrating which amino acids can be resolved in each of the chromatographic systems.     

Chitin as Stationary Phase in Thin-Layer Chromatography–Application of Chromatographic Process Optimization Theory for Chitin Layers

Abstract

This paper describes initial of two basic theories of adsorption thin-layer chromatography potimization namely thermodynamic adsorption theory and theory based on mass action law, so called Snyder s theory in chromatographic systems containing chitin as stationary phase on wchich various amino acids were chromatographed. The results obtained were comparable to those obtained for analogous chromatographic systems containing silica gel as stationary phase. Comparison of applicability of these theories for well investigated silica gel and for not enough investigate chitin will permit to draw introductory conclusions about the processes carrying in the chromatographic systems containing chitin as stationary phase.     

Simultaneous Determination of Carbohydrates and Amino Acids by Capillary Zone Electrophoresis with Constant Potential Amperometric Detection at a Copper Electrode

Capillary zone electrophoresis with end-column amperometric detection at a copper microdisk electrode (100 μm in diameter) was successfully applied for simultaneous determination of carbohydrates and amino acids. Under the separation voltage of 27 kV and the separation electrolyte of 80 mM NaOH in a 75 cm fused silica capillary (10 μm i.d. × 375 μmU o.d.), complete separation of a standard mixture containing 9 carbohydrates and 12 amino acids was achieved in less than 25 min. With the electrokinetical injection of 5.4 s at 27 kV and the detection potential of 0.62 V vs. Ag/AgCl, the detection limits (S/N = 3) were 0.22–0.65 ppm (1.2–1.9 μM) for carbohydrates and 0.31–6.5 ppm (2.3–39 μM) for amino acids, respectively. The calculated numbers of theoretical plates were between 41,000 and 190,000. The use of this method for analysis of carbohydrates and amino acids in a urine sample was demonstrated.     

Determination of amino acids in the hepatic and brain tissue of the rat by gas chromatography

A procedure is described for the quantitative determination of amino acids in hepatic and brain tissue samples from the rat. Because the presence of certain matrix components in the tissue material led to interference with chromatographic analysis they were removed by a prechromatographic “clean-up” step. Quantitative analysis of amino acids, as their N-heptafluorobutyryl iso-butyl ester derivatives, was achieved by high resolution gas chromatography on an apolar fused silica open tubular column. Reproducibility data from the complete procedure are presented; coefficients of variation for arginine and histidine in hepatic tissue varied between 7.1 and 10.1% whereas those for most other amino acids were better than 5%, with a mean recovery of 90%.     

LC Separation of Co-Existing Cetylpyridinium Chloride, Tetradecyltrimethylammonium Bromide and Dodecyltrimethylammonium Bromide on Silica TLC Plates with Aqueous-Organic Eluents

Thin-layer chromatography (TLC) of three cationic surfactants was performed on silica TLC plates with various solvent systems. The mutual separation of cetylpyridinium chloride (CPC), tetradecyltrimethylammonium bromide (TTAB) and dodecyltrimethylammonium bromide (DTAB) was achieved on silica TLC plates with ethanol: 1% aqueous ammonium chloride (4:6, v/v) as an eluent. Effects of cations and anions in the mobile phase on mobility and separation of CPC, TTAB and DTAB were examined. The interference due to the presence of metal cations as impurities on the resolution in the mixture of CPC, TTAB and DTAB was also examined. The limits of detection of CPC, TTAB and DTAB estimated were 0.015, 0.031 and 0.062 μg zone⁻¹, respectively. The developed method was utilized to identify these surfactants in different spiked water samples after their preliminary separation.     

Resolution of multicomponent mixture of amino acids using environmentally benign eluents: A green chromatographic approach

A new green TLC has been used for identifying and monitoring the migration behavior of amino acids through silica and kieselguhr static flat bed in contact of n-butyl alcohol, ethyl acetate or ethylene glycol and their mixtures. From the point of view of chromatographic performance, a mixture of n-butyl alcohol-70% aqueous ethylene glycol-ethyl acetate ratio 5:3:2 by volume proves to be more efficient than the individual components for separation of amino acids from their binary, ternary and quaternary mixtures and the chromatographic parameters like ΔR(F) , separation factor (α) and resolution (R(S) ) for the separation were calculated. Effect of the presence of foreign substances such as metal cations, anions, vitamins and pesticides as impurities in the sample on the separation was also examined. Effect of substitution of butanol by various alcohols has been examined to assess the impact of hydrophobicity of alcohols on the separation of amino acids. The limits of detection for tyrosine, tryptophan, alanine, isoleucine, methionine and serine were found to be 0.10?μg/spot, whereas for lysine it is 0.05?μg/spot. Application of the selected TLC system for the identification and separation of amino acids present in drugs/pharmaceuticals has been performed.