Optimized separation of pesticide enantiomers by chiral high-performance liquid chromatography

Summary A computer-assisted method is described for optimization of multi-component, mobile phase selection for separating enantiomers of four pesticides in normal-phase HPLC. The method is based on the triangle, solvent-selection concept using a statistical scanning method. The optimization of the separation over the experimental region is based on a special polynomial estimation from seven experimental runs, and resolution (R_s) is used as the selection criterion. Excellent agreement was obtained between predicted and experimental data.     

Micropreparative separation of transfer ribonucleic acids by high-performance liquid chromatography

A method was developed for the micropreparative separation of individual species of tRNA using reversed-phase high-performance liquid chromatography on large pore spherical silica bonded with C3 alkyl chains. Columns were eluted with linear gradients of decreasing sodium chloride and increasing methanol concentrations. The decreasing salt gradient gradually abolished hydrophobic interactions and a significantly higher selectivity was thus obtained when compared with increasing gradients of salts usually employed in reversed-phase separations of tRNA. The acceptance of tRNA fractions was tested by charging them with fifteen different amino acids. Significantly different separations were obtained with tRNA from Escherichia coli and from rat liver. tRNAGlu and tRNATyr from E. coli were obtained in a pure form, all other tRNAs were more or less contaminated by adjoining tRNAs for other amino acids. Rechromatography under suitable isocratic conditions was required to obtain pure tRNA species from rat liver. Isoaccepting tRNAs for several amino acids were separated from rat liver. The method described seems suitable for preliminary fractionations of complex mixtures of tRNA and for a simple purification of isoaccepting species if the presence of tRNAs for other amino acids is not an hindrance.     

Separation of porphyrin isomers by high-performance liquid chromatography

A high-speed reversed-phase high-performance liquid chromatographic method using an octadecylsilyl 3 cm long (3 microns particle size) column to separate the free acids of uroporphyrins I and III and coproporphyrins I and III from each other, and from the type I isomers of several other porphyrin carboxylic acids, is described. Separation of the porphyrins was achieved in less than 8 min, and injections were possible every 12 min. The detection limits of uroporphyrin, coproporphyrin, and mesoporphyrin were 75, 45, and 35 fmol (at a signal-to-noise ratio of 2), respectively. Application of the method to the determination of urinary and liver porphyrin patterns is shown.     

Purification of CIS-trimedlure isomers by high-performance liquid chromatography

Summary High-performance liquid chromatographic procedures were developed to make it possible to obtain the fourcis-trimedlure isomers (V, W, X, and Y) in pure form. Trimedlure-V and-Y were each readilt separated from the four-componentcis-trimedlure mixture through high-performance liquid chromatographic analysis on 5-μm and 3-μm silica, but trimedlure-W and-X were not adequately resolved. Chromatography of 5-μm silica of the mixtures obtained by epimerization of thetrans-trimedlure isomers, C and B₂, yielded the respective epimers, trimedlure-W and-X, in pure form.     

Application of correlation high-performance liquid chromatography to the reverse-phase separation of traces of chlorinated phenols

A correlation high-performance liquid chromatography instrument operating under online microprocessor control is described. Pentachlorophenol (PCP) and 11 other polychlorinated phenols are separated on a reverse-phase system. With an ultraviolet detector, the detection limit is about 2.5 × 10^-7 absorption unit (about 6 μg PCP l^-1 at 254 nm).     

Direct separation of regio- and enantiomeric isomers of diacylglycerols by a tandem column high-performance liquid chromatography

A novel HPLC-based method for direct separation of the three isomers of mono-acid diacylglycerols (DAGs), i.e., 1,2-DAG, 2,3-DAG and 1,3-DAG, has been established. The method employs a tandem column system, in which two different columns (a conventional silica gel column and a chiral stationary phase column) are connected in series. Two isomeric mixtures of DAGs (i.e., dicapryloylglycerol and dioleoylglycerol) and lipase-catalyzed reaction mixtures were successfully resolved on the tandem column HPLC system without any derivatization prior to the analysis. According to the established analytical method, stereoselectivity of two lipases toward mono-acid triacylglycerols in ethanolysis reaction was investigated. The tested enzymes were immobilized Candida antarctica lipase B (CALB) and Rhizomucor miehei lipase (RML). Analyses of the enantiomeric purity of 1,2-DAG and 2,3-DAG, generated as intermediates during the reaction, revealed that CALB and RML have sn-3 and sn-1 stereopreference, respectively.     

Chiral separation of atropine by high-performance liquid chromatography

The separation and quantitation of the enantiomers and also the determination of the enantiomeric purity are now current and indispensable tasks for the pharmaceutical analysis. Among the various techniques, liquid chromatography remains the best modality owing to several advantages. High speed, sensitivity, and reproducible results make LC the method of choice in almost all laboratories. Phases that contain alpha1-acid glycoprotein as chiral selector are suitable for separation of charged and uncharged enantiomers with widely different structure. Atropine is widely used as parasympatolytic, anticholinergic and antiemetic drugs. It is one of the preferred antidote for immediate management of toxicity associated with nerve agents. Stereoselective separation was achieved with a prepacked alpha1-acid glycoprotein column without any derivatization procedure. The liquid chromatography system is coupled to mass spectrometry with an atmospheric pressure chemical ionization interface in the positive-ion mode. The chromatographed analytes are detected in selective ion monitoring after optimisation using factorial experimental design. Small amount of enantiomeric composition can be evaluated either by MS or by UV spectrometry (less than 5%).     

Semi-preparative high-performance liquid chromatographic separation of trimedlure isomers

Summary Semi-preparative high-performance liquid chromatographic procedures on 5 m silica were developed for the isolation of gram quantities of eight trimedlure isomers (trans: A, B₁, B₂, and C;cis: V, W, X, and Y) for comparative biological evaluation and NMR studies. Isolations were made from an eight-component 21cis: trans-trimedlure mixture, a four-componentcis-trimedlure mixture, trimedlure-B₂:X and trimedlure-C:W epimerization merization mixtures, a trimedlure-B₁:Y:B₂ mixture, and a trimedlure-A concentrate.     

Separation and study of corrinoid cobalt-ligand isomers by high-performance liquid chromatography

Vitamin B12 belongs to a group of complex organo-cobalt compounds, the corrinoids. "Complete" corrinoids, such as B12, contain a nucleotide as the cobalt alpha-(lower)ligand. This nucleotide is also connected to the periphery of the corrin ring. The "incomplete" corrinoids, in contrast, contain simple cobalt alpha-ligands, such as water or cyanide. Using analytical reversed-phase and anion-exchange high-performance liquid chromatography (HPLC), we have been able to study the behavior of several aquocyano-"incomplete" corrinoids: three isomeric cobinic acid pentaamides, cobinamide, and cobyric acid, all of which exist as thermally unstable isomers, which separate during HPLC. All but one of these corrinoids gave isomer mixtures of 1:1, the exception giving mixtures of 2:1 to 3:1. The separated stereoisomers had different retention times and were collected from analytical columns for further study.     

Resolution of triacylglycerol positional isomers by reversed-phase high-performance liquid chromatography

The ability of reversed-phase high-performance liquid chromatography (RP-HPLC) to separate some positionally isomeric disaturated and monounsaturated triacylglycerols (TAGs) as intact species is demonstrated for the first time. Mobile phases of acetonitrile modified with methanol, ethanol, 2-propanol, 1-propanol, 1-butanol, acetone, or dichloromethane were tested for the separation of POP-PPO, PLP-PPL, PEP-PPE, and PDP-PPD (P-palmitic, O-oleic, L-linoleic, E-eicosapentaenoic, D-docosahexaenoic acid residue) on a single RP-HPLC column. The resolution improved with increasing number of double bonds in the acyl residues. While POP and PPO were only partially resolved, PDP and PPD were fully separated with all tested mobile phases, except those containing methanol. Also separated were the four TAGs having the same equivalent carbon number (ECN = 42), PEP, PPE, PDP, and PPD, on a single RP-HPLC column with mobile phase acetonitrile-2-propanol (70:30, v/v) at 0.8 mL/min. In all cases the isomer with the unsaturated acyl residue in either 1- or 3-position was retained more strongly than the respective 2-isomer.     

Resolution of nucleic acids by high-performance liquid chromatography

High-performance liquid chromatography is a very powerful technique for the separation and isolation of nucleic acids. Nucleic acids can be generally characterized as long, negatively charged polymers with hydrophobic nucleobases. Chromatographic processes which use electrostatic interactions (anion-exchange), hydrophobic interactions (reversed-phase or salting-out) or both types of interactions (mixed-mode) are most effective for resolution of these materials.     

Direct enantiomeric separation of saterinone by high-performance liquid chromatography

A new method is described for the direct liquid chromatographic separation and determination of the enantiomers of saterinone, 1-[(4-cyano-1,2-dihydro-6-methyl-2-oxopyridinyl-5)-phenoxy]-3-[4-( 2- methoxyphenyl)piperazinyl-1]-propan-2-ol, using a Chiralcel OD column with methanol as eluent and a temperature set to 10 degrees C. Several lots of synthesized enantiomers were analysed for their enantiomeric purity of more than 99%, because of the excellent resolution (Rs = 2.2). Furthermore the enantiomeric ratio was determined in plasma samples after oral and intravenous administration of racemic saterinone.     

冠硫醚的高效液相色谱分离

由于王冠化合物冠硫醚的结构中含有软硷硫原子,因此选择它来配合作为软酸的过渡金属离子以分离或分析这些金属离子.过去,以冠硫醚为配体与铂系金属的配合作用的报道很少,我们为此进行了冠硫醚的合成,但发现它们难以用常法分离纯化,因而采用了高效液相色谱法(HPLG)分离.用HPLC法分离冠醚化合物的研究报道很少,近年来仅见Bij等对二苯并18-冠-6和二苯并24-冠-8在C_8和C_(18)烷基键合固定相及硅胶上,用水-甲醇等作移动     

Derivatization, identification and separation of carboxylic acids in wines and beverages by high-performance liquid chromatography

A high-performance liquid chromatographic method for the derivatization, identification and separation of carboxylic acids in beverages such as wines and other commercial drinks or natural fruit juices has been developed. The accuracy and precision of the method are discussed with reference to specific methods for the determination of single acids. Applications to the analysis of different wines and beverages are demonstrated.     

Composition analysis of positional isomers of phosphatidylinositol by high-performance liquid chromatography

An HPLC-based method has been developed for composition analysis of six positional isomers of phosphatidylinositol (PI), of which the phosphatidyl group was connected to different positions of the myo-inositol moiety. The method employed a combination of two types of HPLC analyses. One was direct separation of the six PI isomers into four peaks of 1(3)-PI, 2-PI, 4(6)-PI and 5-PI on a normal-phase silica gel column. The second method was for the separations of 1-PI from 3-PI and 4-PI from 6-PI, which were not separable on the normal-phase column. This method involved conversion of PI isomers into pentakis-(R)-1-phenylethylcarbamate (PEC) derivatives, which were separated on a reversed-phase column. Using the established method, positional specificity of several engineered phospholipases D in enzymatic synthesis of PI from myo-inositol and phosphatidylcholine was investigated. This was performed by analyzing the isomeric composition of PIs synthesized by the mutant enzymes. Among five mutant enzymes tested, two showed strong specificity to 1-OH, one showed moderate preference to 1-OH, one preferred 3-OH, and one showed broad specificity towards 1-, 3-, 4- and 6-OH.     

Preparation and separation of metallobacteriochlorophylla by reversed-phase high-performance liquid chromatography

Summary Semi-preparative high-performance liquid chromatography has been used for the preparation of copper(II) bacteriochlorophylla [Cu(II)-BChl-a] and zinc(II) bacteriochlorophylla [Zn(II)-BChl-a]. Both compounds are separated on a reversed-phase Inertsil ODS-2 column using a mobile phase of acetone-methanol (2575, v/v). The fractionated metallobacteriochlorophylls (M-Bchl-a) are identified by fast atom bombardment mass spectrometry. The spectroscopic parameters such as the wavelength of absorption maxima and the molar extinction coefficients are determined using pure M-Bchl-a obtained by preparative HPLC. The HPLC method proposed here has been demonstrated to be useful for the purification and determination of components of M-Bchl-a.     

The use of microparticulate guard columns in reverse-phase high-performance liquid chromatography

Summary The use of mciroparticulate guard columns in conjunction with high efficiency reverse-phase columns has been evaluated in terms of system efficiency and peak symmetry using three standard test mixtures. The effects of linking tube dimensions, particle size of guard column packing and the use of low or zero dead volume couplings have been investigated. Both valve and stopped-flow injection techniques have been used. Recommendations are given for the most efficient use of guard columns.     

Influence of temperature in reverse-phase high-performance liquid chromatography with gradient elution

The influence of temperature on the retention of several species separated by reverse-phase liquid chromatography by gradient elution is shown to be of enough importance to warrant careful control of temperature if reproducible results are to be obtained. The smaller the particle size in the column, the greater the effect of temperature, and therefore the control should be greater. Likewise, it has been verified that for a given solvent gradient, independent of its complexity, there is a linear relation between ln k′ and 1/T, which also occurs in separations by isocratic elution. Dufek's equation can be adjusted perfectly to the experimental data obtained from gradient elutions, and may be used in the simulation and optimization of gradient chromatographic processes.