Glass capillary gas chromatography-mass spectrometry at high temperatures. direct analysis of free base porphyrins and metal porphyrin complexes extracted from the serpiano oil shale

This contribution describes GC/MS analysis of natural petroporphyrin extracts containing alkylporphyrins either as vanadyl complexes or as demetalated free bases. The combination of high temperature glass capillary gas chromatography with mass spectrometry allows, for the first time, direct determination of electron impact mass spectra of separated alkylporphyrins, making additional purification and derivatization unnecessary. The separation is carried out on glass capillary columns coated with the high temperature-stable medium polar, OH-terminated, polysiloxane phases PS 086 and OV-225-OH. The paper gives detailed working directions for the preparation of the high temperature GC/MS-interface, and of the high temperature stable OV-225-OH columns (max. working temperature 390°C).     

Gas chromatographic separation of triglycerides according to their degree of unsaturation on glass capillary columns

Triglycerides are separated according to their molecular weight and their degree of unsaturation on glass capillary columns by use of the on-column injection technique. The columns employed are silylated SE-30 columns of 4.5, 8, and 10m length with a film thickness of ca. 0.15 μm. The paper gives several examples of chromatograms of natural and hydrogenated triglycerides as well as of mixtures of pure, known triglycerides before and after interesterification.     

Fast high performance liquid chromatography analysis in lipidomics: Separation of radiolabelled fatty acids and phosphatidylcholine molecular species using a monolithic C18 silica column

HPLC procedures using conventional C₁₈ columns are usually used to separate simple and complex lipid mixtures but these methods of separation remain often laborious and very slow. Here, monolithic columns were successfully applied to separate lipids - radiolabelled fatty acid mixtures and individual phosphatidylcholine (PC) molecular species. For that, isocratic elution was performed using two Chromolith™ Performance RP-18e columns connected in series. Detection was achieved by online measurement of radioactivity for radiolabelled fatty acids and by UV absorbance at 205 nm for PC molecular species. The performances of such silica rods were compared to conventional reverse-phase silica columns. Monolithic stationary phase separated radiolabelled fatty acids and PC molecular species two times and four times faster, respectively. In each analysis, monolithic columns allowed better separation efficiency per unit of time, with lower inlet pressure. The main advantages of this method for lipid separation are that, under isocratic conditions, it is simpler and much faster, while remaining accurate and selective when compared to conventional methods. Therefore, monolithic columns may represent a powerful tool for the near future in the field of lipidomics.     

Gas chromatographic separation of sugars by on-column injection on glass capillary columns

Sugars were separated gas chromatographically on short apolar glass capillary columns by using cold, on-column injection (OCI) techniques. After silylation, oligomers up to the hexasaccharides could be efficiently separated in resonable retention times. Response factors of silylated sugars were determined as a function of varying sample amounts and concentrations. The optimum injection amount was found to be 1 μl in heptane as solvent.     

Preparation of inert and high-temperature stable apolar and medium polar glass-capillary columns using OH-terminated polysiloxane stationary phases

Coating intensively leached silica surfaces with OH-terminated phases provides a new way of producing, by simple means, columns with substantially increased inertness and thermostability. In addition, their separation efficiency is found to be typically higher than that of columns with traditional coatings. The underlying basic effect is a condensation process between terminal silanol groups of the phase and residual silanols, of the glass surface, thus producing the mentioned inertness. Moreover, the surface-bonded molecules are immobilized without addition of a radical generator. If required, crosslinking can also be effected using a volatile azo compound. No vinyl groups are required for this additional immobilization process. The paper discusses all processes involved, and gives detailed working directions for the following medium polar phases. OV-1701-OH, OV-31-OH (new, 17% cyanopropyl), OV-61-OH, and OV-17-OH, and the apolar phases PS-347.5 and PS-086. There is no doubt so far that the principle of terminal silanol groups is applicable to all silicone phases, and may replace the traditional endcapped stationary phases in the future.     

分子印迹整体柱的制备、分类与应用

分子印迹整体柱在分离科学领域获得广泛应用,能够用于萃取分离有机小分子、特异性识别金属离子和蛋白质分离。本文综述了分子印迹整体柱的印迹策略、材料分类与应用,重点介绍了新型的分子印迹整体柱,并从特异性、分离效率、快速检测等方面讨论了分子印迹整体柱目前面临的挑战,展望了其发展前景和方向。     

Automated sample preparation method for suspension arrays using renewable surface separations with multiplexed flow cytometry fluorescence detection

In this paper, we describe a new method of automated sample preparation for multiplexed biological analysis systems that use flow cytometry fluorescence detection. In this approach, color-encoded microspheres derivatized to capture particular biomolecules are temporarily trapped in a renewable surface separation column to enable perfusion with sample and reagents prior to delivery to the detector. This method provides for separation of the biomolecules of interest from other sample matrix components as well as from labeling solutions. After sample preparation, the beads can be released from the renewable surface column and delivered to a flow cytometer for direct on-bead analysis one bead at a time. Using mixtures of color-encoded beads derivatized for various analytes yields suspension arrays for multiplexed analysis. Development of this approach required a new technique for automated capture and release of the color-encoded microspheres within a fluidic system. We developed a method for forming a renewable filter and demonstrate its use for capturing microspheres that are too small to be easily captured in previous flow cells for renewable separation columns. The renewable filter is created by first trapping larger beads in the flow cell, and then smaller beads are captured either within or on top of the bed of larger beads. Both the selective microspheres and filter bed are automatically emplaced and discarded for each sample. A renewable filter created with 19.9 μm beads was used to trap 5.6 μm optically encoded beads with trapping efficiencies of 99%. The larger beads forming the renewable filter did not interfere with the detection of color-encoded 5.6 μm beads by the flow cytometer fluorescence detector. The use of this method was demonstrated with model reactions for a variety of bioanalytical assay types including a one-step capture of a biotinylated label on Lumavidin beads, a two-step sandwich immunoassay, and a one-step DNA binding assay. A preliminary demonstration of multiplexed detection of two analytes using color-encoded beads was also demonstrated. The renewable filter for creating separation columns containing optically encoded beads provides a general platform for coupling renewable surface methods for sample preparation and analyte labeling with flow cytometry detectors for suspension array multiplexed analyses.     

A problem in etching Pyrex glass capillary columns

Recent attempts to reproduce a literature method for etching of Pyrex glass capillary columns with ?etching ether”? resulted in reduction of the columns to dust through a violent explosion. Although modifications of the method produced etched columns, we found that less than satisfactory results were achieved. Introduction of etching ether by simply coating the column with a thin film of the ether produced an evenly etched column with well defined whiskers. Several observations made during the etching process will also be discussed.     

Modification of a packed GC injector to a versatile capillary injector

A modification of a packed GC injector to a capillary injector is presented using a simple device which is connected to the GC column by an adsorption-free connector. It permits injections of large sample volumes up to 500 μl hexane solutions on normal 0.3 mm i.d. capillary columns. No special requirements for the stationary phase are necessary. Involatile residues remain inside the device which can be exchanged easily. High performance of separation and quantification is achieved.     

High temperature (GC)2 and (GC)2/CI-MS of nucleosides: Analysis of purinyl nucleoside isomer composition

The analysis of low-volatile polar nucleosides by (GC)²/FID and (GC)²/CI-MS, employing HMDS-deactivated glass capillary columns and on-column injection, is described. The four linkage isomers of a specific nucleoside, formed in the synthetic procedure, are in each case sufficiently well separated in the chromatogram; GC/CI-MS allows for a differentiation of N-7 and N-9 linkage isomers.     

Determination of biogenic quaternary ammonium compounds on fused silica capillary columns with a splitless injector as a chemical reactor/pyrolysis chamber

A conventional splitless injector is used as a pyrolysis chamber or chemical reactor for the N-demethylation of acetylcholine and other choline esters. The novel uses of 2-aminoethanol as a N-demethylation reagent in splitless injection and bonded-phase fused silica capillary columns in the separation of the tertiary amine derivatives of choline esters are described. A comparison is made between non-polar and moderately polar fused silica capillary columns in the separation of choline esters.     

High resolution gas chromatography of tobacco smoke: The contributions of kurt grob

The contributions made by Kurt Grob to tobacco science are reviewed with emphasis on the application of high resolution gas chromatography to tobacco smoke analysis. Highlighted are his pioneering achievements in the preparation and use of glass capillary columns, GC-MS, and splitless injection.     

Gas chromatographic method for simulated distillation up to a boiling point of 750°C using temperature-programmed injection and high temperature fused silica wide-bore columns

It is demonstrated that linear injection characteristics are obtained for a wide boiling point range sample using a temperature-programmed injector in combination with wide-bore fused silica columns. The applicability of the described combination for high temperature simulated distillation is described. The method, using external standardization, gives accurate and repeatable results for different types of samples in the boiling range between 50 and 750°C. The lifetime of the fused silica wide-bore columns was found to be acceptable, viz. over 80 temperature-programmed cycles between ambient and 430°C. Some comments are made on the accuracy of boiling points for normal alkanes.     

A comparative study of microbore reverse-phase columns

A procedure for packing reverse-phase microbore columns using moderate pressure is described. Columns packed from different manufacturer's tubing are compared with commercially available columns. A method for coupling the column to the injection valve is suggested. The effect of detector cell volume on overall efficiency of the system is also examined.     

剖析液相色谱柱技术的复杂状况(英文)

 Column packings continue to evolve as the needs of users for high efficiency, high resolution and highly sensitive high performance liquid chromatographic (HPLC) analysis drive further developments. In comparing and contrasting modern HPLC columns technologies, diameters of column packings and particle materials are covered. Some products and applications of modern HPLC columns are provided. Future directions in packing developments are predicted in this introductory article.     

A comparative study of injection techniques in high-performance liquid chromatography

Summary The performance of injection valves with both curtain- and single-flow column couplings has been evaluated using reverse-phase octadecylsilane columns and a standard test mixture. The results are compared with those obtained from septum and stopped-flow syringe injections.Stopped-flow valve injection has been examined similarly. Configurations and couplings which give optimum efficiency and reproducibility are recommended.     

Development and validation of a direct plasma injection LC-MS method for YM087 and YM440 and metabolites in human plasma

Summary A fully automated, direct plasma-injection technique using an LC coupled to a quadrupole mass spectrometric detector with an on-line, sample clean-up procedure for two new pharmaceutical products, YM087 and YM440, has been developed. Plasma samples containing YM440 were mixed with internal standard or plasma containing YM087 and were injected into a chromatographic system based on two coupled columns. An extraction column packed with alkyl-diol-silica (ADS) was used for online sample cleanup. Using a back-flush technique analytes were subsequently transferred to an analytical column, for separation. Detection was based on tandem mass spectrometry either in electronspray or in APCI mode. Despite the relatively small volumes of plasma injected, reasonably low limits of quantitation were achieved. Validation of both assays was performed using guidelines concerning method validation. All parameters studied were within acceptance criteria. The methods were successfully applied to clinical pharmacokinetic studies.     

The preparation of thick-film glass capillary columns

A detailed method for the routine preparation of glass capillary columns is presented. The method consists of coating a glass tube with quartz powder prior to pulling the tube into a capillary. The inner surface of the capillary consists of an even distribution of quartz particles fused to the walls. This surface has been found readily deactivated by standard procedures and ideal for the preparation of thick-film glass capillary columns. The method has been thoroughly tested in two independent laboratories to ensure that the procedures described are reproducible.     

Characterization of fatty oils by pattern recognition of triglycerides profiles

The triglyceride profiles for corn, soybean, sunflower, grapestone, olive and olive husks oils were obtained by high temperature capillary gas chromatography on HT-FS capillary columns coated with OV-17-OH. The data were treated by methods of computerized pattern analysis, involving principal component analysis, discriminant analysis, and hierarchical clustering. A graphical representation of results as “star symbol plots” allows prompt visual recognition of the characteristic profiles and straightforward qualitative identification of unknowns as well as recognition of adulterations.     

Determination of the threshold odor concentration of main odorants in essential oils using gas chromatography-olfactometry incremental dilution technique

In dual-flow counter-current chromatography (DF-CCC), the two immiscible liquids are flowing in opposite directions in the coil. The method allows for the continuous separation of two solutes. In this study a numerical model was developed to allow for the detailed investigation of flow in such columns. The mesh model of the presented DF spiral column was developed in line with an existing experimental model. The paper presents results during the early filling stages for different rotation directions. These clearly illustrate the performance of the developed model by (1) confirming the importance of flowing the lighter phase from tail to head and the heavier phase from head to tail and (2) by visualising mixing waves and the recognised back and forth "swish-swash" motion as present in CCC in that operating mode.