High-performance ligand-exchange chromatography of amino acids on chiral stationary phases

Three chiral stationary phases, obtained by grafting silica gel with (-)-trans-1,2-cyclohexanediamine, were studied for the resolution of α-amino acids by ligand-exchange chromatography. The packings were prepared by bonding the chiral ligand to silica gel via different hydrocarbon spacers. Separation of the optical isomers was accomplished by eluents containing a constant concentration of copper(II) acetate (0.05mM). The elution sequence of amino acids was found to be dependent on the grafting reaction selected to prepare the chiral packings.     

Purfication of synthetic cardiotoxin by affinity chromatography

A polypeptide containing 60 amino acids with 4 disulphide bonds, synthesized by the solid-phase method, was highly purifed by anticardiotoxin-Sepharose affinity chromatography following gel filtration and CM-cellulose chromatography. The identification of the final product as cardiotoxin was confirmed by thin-layer chromatography on silica gel, polyacrylamide gel electrophoresis, amino acid analysis, circular dichroism spectra, N-terminal analysis and four biological tests.     

增强型绿色荧光蛋白的色谱分离和纯化

增强型绿色荧光蛋白(EGFP)是生物领域常用的标记物。在前期成功克隆表达EGFP的基础上,本实验建立了两步分离纯化EGFP的色谱方法,并验证其分离纯化效果,检验EGFP的活性。首先用金属螯合亲和色谱柱HisTrap HP对EGFP的重组菌体破碎上清液进行初步分离,再用葡聚糖凝胶排阻色谱柱Sephadex G-10 HR对其进行脱盐纯化。采用丙烯葡聚糖凝胶排阻色谱柱Sephacryl S-300 HR和十二烷基磺酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)检测分离纯化后的EGFP纯度。最后通过荧光分光检测器和非变性聚丙烯酰胺凝胶电泳(Native-PAGE)验证分离纯化后的EGFP是否具有荧光活性。结果表明该方法可以简便快速地分离纯化EGFP,纯度超过98%,同时保持了EGFP的荧光活性。     

Isolation from fetal bovine serum of a peptide similar to the alpha chain of thrombin by reversed-phase high-performance liquid chromatography

During the preparation of erythrotropin from fetal bovine serum, a group of peptides co-eluted with this erythroid cell stimulating factor on semi-preparative reversed-phase high-performance liquid chromatography. They could be subsequently separated by a combination of reversed-phase high-performance liquid chromatography in the presence of heptafluorobutyric acid as ion-pairing reagent and gel permeation high-performance liquid chromatography. One of these peptides has been extensively purified. Partial amino acid sequence analysis indicated that fourteen of the seventeen N-terminal amino acids are identical with the N-terminal sequence of the alpha chain of bovine thrombin. The same isolation procedure could be useful for the identification of other major peptides of fetal bovine serum.     

Gel permeation chromatography of oxygenated components of cigarette smoke condensate

Oxygenated neutral constituents of a tumor-inhibiting fraction of cigarette smoke condensate were separated from interfering phenolic compounds by a combination of silicic acid and gel chromatography. Silicic acid adsorption and Bio-Beads gel chromatography separated aliphatic from aromatic ketones, and gel chromatography on Sephadex LH-20 resolved aromatic ketones from phenols. Identifications were achieved by studying individual fractions by g.c. and g.c.—m.s.     

Gel permeation chromatography of branched polyethylene

The effect of long-chain branching must be considered in gel permeation chromatography to evaluate the molecular weight polydispersity of branched polyethylenes. Osmotic molecular weights of fractions of branched polyethylene were correlated with elution volumes; weight-average and number-average molecular weights of a branched polyethylene were determined. Molecular weight changes on crosslinking polyethylene by ionizing radiation are accompanied by branching and cannot be simply interpreted by gel permeation chromatography.     

Effects of sequence and structure in the separation of nucleic acids using ion pair reverse phase liquid chromatography

Ion pair reverse phase high performance liquid chromatography on non-porous alkylated poly(styrene-divinylbenzene) particles enables the high resolution separation of double stranded DNA fragments. To further understand the separation mechanisms involved in ion pair reverse phase liquid chromatography we have analysed the effects of curved or "bent" DNA fragments with respect to their separation using both gel electrophoresis and ion pair reverse phase liquid chromatography. Size dependent separations of curved DNA fragments that migrate anomalously during gel electrophoresis were observed using ion pair reverse phase liquid chromatography. To further study the sequence effect and resulting changes in hydrophobicity of the duplex DNA, PCR fragments were generated that contain uracil in place of thymine. The resulting fragments were shown to elute with shorter retention times, demonstrating that sequence-specific effects can alter the retention of duplex DNA. The study was extended to the investigation of non-canonical B-DNA structures (Holliday junctions) under various chromatographic conditions, demonstrating that the coaxial stacking of the helices in such structures, in the presence of magnesium causes a change in retention.     

Polymer studies by gel permeation chromatography

Samples of polystyrene and poly(methyl methacrylate) have been prepared by bulk polymerization under conditions previously investigated in the literature. The molecular weight distributions were determined for early-conversion samples by gel permeation chromatography, and the experimental distribution curves were compared with calculated curves based on kinetic data. The agreement between the experimental and calculated curves was such as to indicate the utility of gel permeation chromatography in the investigation of polymer reaction kinetics and mechanisms.     

Analysis of the negative peaks in gel permeation chromatography

The negative peak usually encountered in gel permeation chromatography has been resolved into three distinct peaks through selection of appropriate columns. The three peaks are associated with water, nitrogen, and oxygen. Separation of these simple molecules was found to follow the fundamental theory of gel permeation chromatography, that is, separation according to straight-chain length.     

Separation of naphthoquinones and lipophilic vitamins by gel and thin-layer chromatography.

A separation of naphthoquinones on silica gel and on silica gel impregnated with polyethylene glycol 200 by thin-layer chromatography was compared with gel permeation chromatography (GPC) on styrene-divinylbenzene copolymer S-832-gel using tetrahydrofuran as mobile phase. Factors affecting the separations attainable are discussed, and it is concluded that GPC is a suitable method for the determination of K vitamins in natural materials.     

Multi-grams scale purification of xanthanolides from Xanthium macrocarpum. Centrifugal partition chromatography versus silica gel chromatography

Plants of the Asteraceae family are known as a source of sesquiterpene lactones with interesting biological activities. The purification of several xanthanolides (xanthathin, 4-epi-xanthanol and 4-epi-isoxanthanol) was realized in one step, directly from the crude chloroformic extract of the leaves of X. macrocarpum by silica gel chromatography and for the first time by liquid/liquid chromatography (counter-current chromatography/centrifugal partition chromatography), using a FCPC 5L (Fast centrifugal partition chromatograph, Kromaton Technologies, Angers, France). Recovery, purity of xanthathin and solvent consumption were improved with the liquid/liquid chromatography compared to solid/liquid chromatography.     

High-performance liquid chromatography of membrane proteins

Various modes of high-performance liquid chromatography, gel filtration, ion-exchange chromatography, hydrophobic interaction chromatography, reversed-phase chromatography and metal chelate affinity chromatography, were investigated for the separation of membrane proteins. All were found applicable to membrane proteins, although the usefulness of each mode differed. For satisfactory results it was important to select appropriate elution conditions. The type and concentration of detergent was of special importance. The effects of other conditions, flow-rate, gradient steepness, type of buffer and salt, eluent pH, etc., were similar to those observed for soluble proteins.     

国内气相色谱近年的进展

本文对近三年国内学者在气相色谱方面的研究和应用作了综述 ,国内学者的研究基本和国际上气相色谱方面的研究类似 ,在全二维气相色谱、快速气相色谱 ,微型气相色谱仪、新型气相色谱固定相和色谱柱的溶胶 凝胶涂渍技术领域的研究方面作出了贡献。有关气相色谱的应用研究中 ,介绍了大量在药物分析、食品分析、环境分析、石油和石化分析和化工产品及高聚物分析等领域中应用的题目和摘要     

Separation and characterization of octylphenol ethoxylate surfactants used by reversed-phase high-performance liquid chromatography on branched fluorinated silica gel columns.

The separation and characterization of octylphenol ethoxylate surfactants were carried out by reversed-phase high-performance liquid chromatography on branched fluorinated silica gel columns. For Triton X-100, simultaneous separation of octylphenol ethoxylate oligomers, positional isomers of octylphenyl group and butylphenol ethoxylate oligomers was achieved. These oligomers were completely separated and identified by means of MS spectra. Ethoxylated oligomers are eluted in the sequence from small to large oligomers. Fifty-five oligomers of Triton X-405 could be separated by using gradient elution. To separate octylphenol ethoxylate surfactant, non-end-capped branched fluorinated silica gel columns were superior to end-capped columns. The relationship between ln k' and methanol concentration was linear, indicating that branched fluorinated silica gel columns were operating in the reversed-phase mode. As Van 't Hoff plots of capacity factor for all oligomers gave straight lines, the equilibrium of conformation for the ethylene oxide chain might lay to one side of either zigzag or meander conformers.     

Separation of phospholipids using a diethylaminoethyl-silica gel column and thin-layer chromatography

Phospholipids derived from egg yolk are readily separated by DEAE-silica gel column chromatography using stepwise gradient elution. The overall recovery of phospholipids from the column is 85-95% at a loading capacity of 120 mg of lipids per 10 g of DEAE-silica gel. The complete separation of eight phospholipids (phosphatidylcholine, sphingomyelin, lysophosphatidylcholine, phosphatidylethanolamine, phosphatidylglycerol, phosphatidylinositol, phosphatidylserine and lysophosphatidylserine; 5 micrograms each) is also achieved by one-dimensional DEAE-silica gel thin-layer chromatography with the solvent system chloroform-methanol-water-pyridine-58% ammonia solution (130:55:8:4:4, v/v).     

Development of silica-based stationary phases for high-performance liquid chromatography

Stationary phases are the basis of the development and application of high-performance liquid chromatography (HPLC). In this review we focused on the development of silica-based stationary phases, including the synthesis of silica gel and the application of silica in hydrophilic interaction chromatography (HILIC), reversed-phase liquid chromatography (RPLC), chiral separation chromatography, and ion chromatography. New stationary phases, advances in ionic liquid-modified silica, silica-based core-shell materials, and silica-based monolithic columns for HPLC are introduced separately.     

气相色谱法测烟草中拟除虫菊酯和有机磷类农残

建立了一种测定烟草中拟除虫菊酯类农药和有机磷类农药残留量的气相色谱法.用普通的聚苯乙烯凝胶柱代替昂贵的凝胶渗透色谱仪;层析柱填料用弗罗里硅土代替硅胶;用工作标准溶液过柱曲线作为标准曲线;有机磷类农药气相检测器用PFPD代替FPD.方法的检测量为0.01μg/g～0.04 μg/g,平均加标回收率为70.18%～95.86%,RSD为4.36﹪～9.96﹪,对样品进行了拟除虫菊酯类农药和有机磷类农药残留量的测定,结果满意.     

Gel chromatography of steroid oestrogens on sephadex LH-20.

The behavior of 23 steroid oestrogens Sephadex LH-20 using six solvent systems was investigated. The fractions eluted by gel column chromatography were analysed by thin-layer chromatography and spectrometry. The method was used for the radiochemical purity of labelled compounds by isotopic dilution.     

Isolation and purification of uremic middle molecules by multi-step liquid chromatography

Isolation and comparison of uremic sera and urine and normal sera and urine were performed by gel permeation chromatography, anion exchange chromatography and re-versed-phase high performance liquid chromatography. Two uremic middle molecular fractions (A and B) were obtained from uremic sera and urine and normal urine by gel permeation chromatography, but not from normal sera. The anion exchange chromatographic results of fraction A from different origins demonstrate that subfraction A-3 could be excreted in urine by healthy subject, but accumulated in uremic serum for renal failure of patient with uremia. After desalinization subfraction A-3 was analyzed by MALDI-TOF-MS. The results show that subfraction A-3 consists of six compounds with molecular weight 839, 873, 1007.94, 1106, 1680 and 2015 respectively. Finally, by reversed-phase high performance liquid chromatography, subfraction A-3 was further resolved into six independent fractions. Thus, the isolation and purification of six middle molecular c     

Salt promoted adsorption chromatography of malted barley amylases

Summary A sequence for the fractionation of the amylasic components from a malted barley extract is proposed using two salt-promoted, adsorption processes: thiophilic interaction chromatography (TIC) and hydrophobic interaction chromatography (HIC).Two fractions containing -amylase activity were recovered during the thiophilic chromatography; the first was resolved in to -amylase I and -amylase I by HIC on a phenyl-sepharose column; an enrichment factor of 32 was achieved for -amylase I. The other amylasic component eluted from the thiophilic gel was characterized as -amylase II. Although the adsorption of malt amylases on phenylsepharose and the thiophilic adsorbent is salt promoted, the interactions involved in each case are clearly distinguished by the different behaviour and disparate salt effects.