Chromatography gas liquid

Gas-liquid chromatography (GLC), which is also called gas chromatography (GC), is a technique that may be used to separate mixtures of volatile compounds whose boiling points may differ by as little as 0.5 °C. It can also be applied as an analytical tool to identify the components of a mixture or in preparative applications when quantities of the pure components are desired. 

The retention time of a component is the elapsed time required for the compound to pass from the point of injection to the detector, and it may be used for purposes of identification. The retention time of a component is independent of the presence or absence of other components in the sample mixture. There are four experimental factors that influence retention time of a compound (1) the nature of the stationary phase, (2) the length of the column, (3) the temperature of the column, and (4) the flowrate of the inert carrier gas. Thus, for a particular column, temperature, and flowrate, the retention time will be the same for a specific compound. 

Although a large number of stationary liquid phases are available, only a few are widely used (Table 6.1). Each liquid phase has a maximum temperature limit above which it cannot be used. This temperature depends upon the stability and volatility of the liquid phase at higher temperatures, the liquid phase will vaporize and bleed from the column with the mobile phase. 

The differences in the partition coefficients (Secs. 5.2 and 5.3) of the individual components of a mixture in GLC depend primarily upon the differences in solubility 

Liquid Phase Type Property Maximum Temperature Limit, °C 

Gas-liquid chromatography has been widely used for the identification of reaction mixtures and for the separation of heterocycles. Some typical conditions are shown in Table 34. 

Pyrazolines Imidazoles Thiazoles Oxadiazoles 1 -Phenylpyrazoles Purines Dioxolanes Ox azolines 10% Cyanethylated mannite on Celite 545. 5% OV-17 on Chromosorb W, AW-DMCS (H.P.). Carbowax 4000, dioleate on firebrick, 190 °C. Silicone grease on Chromosorb P. Apiezon L on firebrick C-22, 220 °C. 15% Hallcomid M-18 on firebrick. Carbowax 20M on Gas-Chrom P. Ethyleneglycol succinate on Diatoport-S. 

Gas-Liquid Chromatography.—Liquid-liquid partition data on O-alkyl-O-aryl phenylphosphonothioates was obtained by g.l.c. There have also been several reports on the determination of inositol phosphates.  

Gas-liquid Chromatography (g.l.c.).—Letcher has reviewed the use of g.l.c. to obtain activity coefficients in non-poiymer systems. The method is claimed - to be an accurate means of obtaining thermodynamic quantities in binary solutions when the two components differ considerably in volatility. Clearly this applies to many polymer-solvent systems and then the pol3rmer is conveniently made to form the stationary (liquid) phase in standard equipment. The solvent of interest is introduced into the mobile (gas) phase and its specific retention volume measured, from which heats of mixing are calculated in the limit of zero concentration of solvent (a limit of interest in connection with the removal of volatiles from polymeric materials - ). 

The method has been applied to several systems and improvements 

methods the influence of surface free energies has apparently not been considered although it might be expected to be significant. The work of Calderon and St.-Pierre should be useful in this connection. 

A variant of the technique devised by Ishizawa et al examines the transmission pattern of an He-Ne laser beam to study pressure dependence of LCST with high accuracy . Coupled with titration methods (that is varying composition by addition of solvent or solution), turbidimetry has been used to good effect to study cloud point curves in polymeri-polymerj-solvent and ionic polymer systems and to obtain solubility parameters for copolymers in mixed solvents.  

An adaptation of the method studies phase separation in polymer blends from measurements of light transmission through films. 

Gas-liquid chromatography, when combined with ultraviolet analysis 556) and additionally with nuclear magnetic resonance spectroscopy (557), has been developed into a powerful tool for the characterization of sub-milligram quantities of coumarins from plant extracts. Free hydroxycoumarins are better analyzed as their trimethylsilyl ethers 203) though a new technique 206) has permitted the gas chromato-graphic-mass spectrometric analysis of coumarins avoiding the necessity of forming derivatives. 

Gas-Liquid Chromatography.—K two-part review by Dutton has discussed the g.I.c. of sugars and their derivatives.  

Mixtures of O-methyl ethers obtained by partial methylation of a number of methyl glycopyranosides have been acetylated and the products identified by g.l.c.-m.s. The use of partially ethylated alditol acetates for the analysis by g.I.c. of the components of polysaccharides has been described. The ethyl analogues are eluted before the corresponding methyl derivatives, and this often allows the separation of many polysaccharide components by g.l.c. that are not separable as their partially methylated alditol acetates. The use of molar flame-responses calculated on the basis of effective carbon responses has been advocated for the accurate determination of partially methylated and partially ethylated alditol acetates by g.I.c.  

Mixtures of common aldoses have been analysed by g.l.c.-m.s. of their 0-iso-propylidene derivatives, which, in most cases, give rise to clearly different mass spectra. Gas-liquid chromatographic data for the butaneboronic esters and TMS derivatives of some monosaccharides and alditols have been reported butaneboronic esters are useful for the qualitative identification of certain monosaccharides, and are of particular value in the quantitative analysis of mixtures of D-glucose and D-fructose.  

TMS derivatives of methyl (methyl 4-deoxy-0-methyl-j8-L-rAreo-hex-4-eno-pyranosid)uronates and of the monosaccharides obtained on methanolysis of glycoproteins and glycopeptides have been examined by g.l.c. 

Variation of the counter-ion attached to a polystyrene-based, strong cation-exchange resin has been shown to affect the chromatographic behaviour of sugars and other polyhydroxy-compounds. The separation of mixtures of sugars can often be significantly improved by choosing a suitable counter-ion, and, in certain cases, oc- and j3-anomers of monosaccharides can be separated. 

Gas-Liquid Chromatography.—Phthalic esters - which are extensively used as plasticizers in the formulation of plastics used to make containers for solvents, etc. - have been found as contaminants during the analysis of polysaccharides by gas chromatography of the alditol acetate derivatives for example, dibutyl phthalate behaves as a tetritol tetra-acetate on g.l.c. and di(2-ethylhexyl) phthalate has a retention time close to that for xylitol penta-acetate. The contamination by phthalic esters can be minimized by distilling all solvents used in the analytical steps or by using smaller volumes of potentially contaminated solvents. 

and tri-saccharides can be separated satisfactorily by g.l.c. of their trimethylsilylated oxime derivatives, and an investigation has been conducted into the most selective stationary phases for the separation of mixtures of pentoses and hexoses as their TMS derivatives. Traces of D-glucose and other related impurities in samples of o-glucitol can be detected by g.l.c. of the O-acetyl derivatives.  

Partially methylated derivatives of 2-deoxy-2-methylamino-D-glucose have been separated and identified as the corresponding alditol acetates by combined 

Optimum reaction conditions for the silylation of nucleosides with bis(tri-methylsilyl)trifluoroacetamide and the behaviour of the silylated nucleosides on g.l.c. have been investigated.  

Gas-Liquid Chromatography.—Much structural information can be derived from the chromatographic behaviour of long-chain esters, and this topic has been reviewed (see ref. 153c). Useful retention data are presented in several 

Chadha, and A. P. Davies, Biochim. Biophys. Acta, 1970, 218, 102 R. Aneja and J. S. Chadha, Ghent. andPhys. Lipids, 1970, 4, 60 Biochim. Biophys. Acta, 1971, 248, 455. 

Stanadev, L. Gospodid, and M. Prostenik, Ghem. and Phys. Lipids 1970, 7, 135. 

The basic design of gas chromatograph can be fitted with a range of specific injectors, columns and detectors to optimise the separation of components and aid their identification. Recent developments in computer control, the use of robotic autosamplers and the trend to couple instruments together for sequential procedures have led to increased automation for routine analytical tasks performed by GCs in research, factory and quality control environments. 

Many specialized injection systems are used with the GC in the modern laboratory to deliver the complete sample to the column without alteration. The conventional injection method is simply to inject a small volume (about 1 /d) of a dilution of the sample using a syringe with a fine needle, which pierces a silicon rubber septum and delivers the sample into a heated chamber where it is vaporized and carried on to the column in the stream of carrier gas. Some of the carrier gas containing vaporized sample may be split from the main column flow and vented to reduce the amount of sample delivered to the column. This is known as a split injection and is suitable for almost all liquid or 

The high temperature of the injector, typically 250 °C, means that this method is not suitable for analytes that are subject to thermal degradation. For these materials, an on-column method is preferable, in which the solution of sample is injected directly into the narrow capillary column with a fine needle. On-column injection techniques are also more suitable for extremely dilute samples, as more sample is delivered to the column, but are less suitable for dirty samples containing non-volatile contaminants, which accumulate on the column. 

The use of a headspace injection technique may be preferable if the analytes are contained within a non-volatile or corrosive matrix that cannot be injected directly (perfume in a washing powder, for example). The basic principle of headspace injection is the delivery of a volume of vapour from the space above the sample material to the GC column. This can be achieved in several ways  

Coupling the column from the GC to a mass spectrometer provides a very powerful combination, GC-MS, which can identify and quantify almost all the compounds in a complex mixture, such as an essential oil or perfume, by reference to libraries of mass spectra of known compounds. Careful investigation of the mass spectrum can be used deductively to determine a possible structure for an unknown material using fragmentation theories to identify sub-structural components of the molecule. Recent developments in benchtop mass spectrometers have brought a range of specialized MS techniques into the realm of GC-MS machines techniques such as chemical ionization and MS-MS are now available, which provide more information on individual sample components and allow better identification of unknown compounds. 

The prindple behind gas-liquid chromatography is the differential distribution of compounds between a stationary liquid phase in the column and a mobile gaseous phase flowing through the column - hence the name gas-liquid chromatography. 

The stationary phase consists of a thin layer (ca. 0.2 pm) of a non-volatile liquid, such as methylsilicone or methylphenylsilicone, tightly adsorbed to the inert surfaces of the column. The liquid stationary phases are classified according to their polarity, non-polar phases are used most often because they are easier to handle and are stable over a wide temperature range. The mobile phase is an inert carrier gas such as nitrogen, helium or hydrogen which flows at a constant, but adjustable rate through the column. Current capillary columns, with internal diameters between 0.1-0.6 mm, are constructed of fused silica, specially prepared from pure silicon tetrachloride, and formed into spirals up to 100 m long. Injectors, columns and detectors are located in separately thermostatable compartments. 

Other detectors in use include the thermal conductivity detector (TCD) and the phosphorus-nitrogen detector (PND) which is much used in toxicology because of its selectivity for nitrogen-containing compounds. It is a feature of gas-liquid chromatography that spectrophotometric detectors can be coupled readily to the outflow for detection this includes IR, NMR and, particularly, mass spectrometers, in combined GC-MS analysers. Spectroscopic analysis allows structural information to be 

It is often the case that a peak in a gas chromatogram can be identified from its relative retention index (or time). This provisional identification can be confirmed by spiking the unknown sample with an authentic sample which should co-chromatograph with the unknown. However, unambiguous identification of an unknown compound demands structural analysis, which can either be done on-line (e.g., using coupled GC-MS equipment) or off-line. 

The main area of application of gas-liquid chromatography is for quantitative analysis of compounds in mixtures of volatile substances in organic media. If the compounds are not sufficiently volatile, as is the case with carbohydrates, amino acids, steroids etc., they can usually be converted into suitable volatile compounds by derivativisation by methylation, acetylation or trimefhylsilylation and other similar treatments. 

This technique can be used to determine the unreacted or free TDl present in urethane prepolymer as a result of incomplete reaction or the use of non-stoichiometric quantities. The free TDI in the urethane prepolymer is thermally expelled within the vaporizing chamber of a gas chromatograph, but below the decomposition temperature of the parent or matrix material. Subsequently, on a column coated with silicone gum rubber, it is 

Detector flame ionization, complete with electrometer. 

Column pressure and flow 11-5 psig nitrogen, restricted to provide a flow of 27-5 ml min through the column. 

Column 100 ft of 3 mm copper tubing coated with a 2-5% solution of silicone gum rubber. 

Standards to cover the desired range are prepared by adding known amounts of TDI to approximately 4-5 g portions of prepolymer. One sample should have no additional TDI added so as to form a control. Then to each standard is added 0 5 g of trichlorobenzene as an internal standard and 10 ml of dry ethyl acetate (AR grade) samples for analysis are prepared by adding 0-5 g of trichlorobenzene to approximately 4 5 g of prepolymer and diluting with 10 ml of dry ethyl acetate. A 1 fil portion of the standards and samples is injected into the chromatograph. 

On nonpolar columns, the compounds of a homologous series separate as a function of their boiling points, and linear relationships have been established between the logarithms of the retention volumes and the number of carbon atoms in the 2-, 4-, and 5-positions (see Fig. III-l). 

A comparison of the molar volumes of 2-, 4-, and 5-alkylthiazoles with their relative retention volumes shows that these values also vary in the same direction (see Fig. III-2). 

The Kovats indices (173) of thiazole on various columns are given in Table 10-19. 

The Kovats indices of various alkyl, dialkyl, and other 2-substituted thiazoles are given in Table III-20 (174, 175). 

Comparison between molar volumes and retention volumes for 1-. 4-. and S-alkylthiazoles. 

Typical operating conditions for the GLC separation of azines are shown in Table 26. 

15% Hallcomid M-18 on firebrick Apiezon M and N or Reoplex 400 Flexol 8N8 on firebrick SE-30 on Chromosorb W Ethylene glycol adibatic on glass beads 

More recently, capillary or open tubular columns have in part replaced packed columns in GLC analysis because of their high efficiency (100000 theoretical plates overall) and their high resolving power. A number of methods for the separation of fatty acid methyl-esters have been reviewed (Christie, 1982a). 

The method by which the absolute configuration of aldoses may be determined by conversion to glycosides of (—)-butan-2-ol followed by g.c. (see Vol 12, p. 207) has been successfully applied to the hydrolysates of several polysaccharides and glycoproteins.  

Relative to methyl deoxycholate unless otherwise indicated. 

Retention times given relative to the bis(trimethylsilyl) ether of methyl deoxycholate. 

These compounds often give two peaks. The retention time of the major peak, which may be a degradation product, is given. Column temperature 220°C. 

The separation of bile acids by gas-liquid chromatography is determined by the choice of stationary phase and bile acid derivative. Data that permit the selection of stationary phase and bile acid derivative to suit most separation problems are shown in Tables XII-XIV. As in the original papers relative retention times have been used in the tables instead of the more acceptable retention index (112) which permits better interlaboratory comparisons. Relative retention times are subject to variation, mainly due to temperature differences and, to a lesser extent, to differences in column preparation. In our experience the temperature-dependence is most pronounced with trimethylsilyl ether derivatives on Hi-Eff-8B (cyclohexanedimethanol succinate) columns. Temperature differences do not affect relative retention times on QF-1 (a trifluoropropyl, methyl siloxane) to the same extent. 

Improved separations of bile acid methyl esters can be observed with increasing amounts of phenyl substituents in the stationary phase (5% in SE-52, 20% in PhSi-20, and 50% in OV-17). Positional and configurational isomers are better resolved than on SE-30 or OV-1 columns. For instance the valuable separation of the trimethylsilyl ethers of 3,6,7-substituted methyl cholanoates from the corresponding cholic acid derivative on OV-17 should be noted. The pronounced effect of a 7/3-hydroxy substituent on retention times on columns of SE-52 and PhSi-20 is noteworthy. The large separation factors between the diacetate derivatives of chenodeoxy- and ursodeo.xycholic acids may be most useful in work with bile acids of biological origin. 

Other reports have appeared on the g.l.c. of partially methylated alditol acetates, partially methylated mannononitrile peracetates, TMS ethers of methyl glycosides derived from mucopolysaccharides, and TMS ethers and butylboronate TMS ethers of polyhydroxyalkylpyrazines (note that several of these papers include reports of mass spectral studies). 

Special attention has been paid to the protection of N- and O-acyl substituents against methanolic HCl. Yu and Ledeen (1970) proposed a mild methanolysis 

Terminal sialic acids are released in a good yield under mild acidic conditions like formic acid, pH 2 (1 h, 70 X), 0.1 N H2SO4 or 0.1 N HCl (1 h, 80 C) (Schauer 1978). These conditions do not lead to N-deacylation however, 0-deacylation occurs to an extent of about 50%. Mild acid hydrolysis has frequently been chosen to release N,0-acylneuraminic acids from biological materials for identification by g.l.c./m.s. Derivatization procedures are presented in sections II. 3 and II.4. For g.l.c./m.s., see section III.2. Quantitative g.l.c. procedures after pertrimethyl-silylation (section II.4) have been worked out by Casals-Stenzel et al (1975). Because of partial 0-deacylation during hydrolysis and subsequent isolation, quantitative analysis of the various N,0-acylneuraminic acids present in native biological material is still a serious problem. The nonavailability of pure N,0-acylneuraminic acids as standard compounds makes it also difficult to determine reliable molar adjustment factors for g.l.c. analysis. 

SuGiTA (1979 a) developed a different approach for the combined determination of Neu5Ac and Neu5Gc, by applying solvolysis with l.ON -butanolic HCl. The formed A2-butyl acetate and -butyl glycolate are analyzed by g.l.c. (section II. 5). 

From the outset the biochemist must decide on the range of acids which he requires to measure in a particular physiological fluid. This may be simple in some cases where only a few acids are of interest which perhaps can be selectively extracted and run under specific GC conditions. Other workers may require to look at a larger, but single, group of acids (for example short-chain volatile fatty or aromatic acids), where specific extraction, derivatization and GC conditions will give an adequate analysis. However, many have adopted a comprehensive approach involving the attempt to extract acids as completely 

In an attempt to rationalize the many different structural types comprising acidic extracts which require to be included, the acids have been divided into convenient groups. There is overlap between some of these groups, and a separate section has been devoted to profile analysis in which all acids are considered together. 

A wide variety of gas-chromatographic systems is available commercially and although technical improvements and developments continue to be made in 

These columns are usually of length 0.3-4.0 m and internal diameter 2-4 mm and are packed with solid granules (the solid support ) on which is coated the liquid stationary phase. The ideal material as the support medium in a packed column should be of uniform particle size of large specific area and be chemically, thermally and mechanically stable. The frequently used diatomite supports are prepared from diatomaceous earths by a calcination process and differ from each other in their particle size, level of inertness and available surface areas. The surfaces are usually deactivated by acid washing and are silanized prior to coating with liquid phaste. 

The majority of analyses of biological compounds up to the present time have been carried out on packed columns owing mainly to the wide variety of stationary phases available, their high sample-loading capacities and the all round simplicity in their preparation and use. 

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Note 4. Gas-liquid chromatography showed complete conversion into the bis-ether. 

These values are practically temperature independant, and they are very close to those found for the Apiezon L column. Comparison with the values of a series of alkybenzenes shows that the 5-position of thiazole possesses behavior analogous to that of a benzenic position in gas-liquid chromatography. 

Analytical separations may be classified in three ways by the physical state of the mobile phase and stationary phase by the method of contact between the mobile phase and stationary phase or by the chemical or physical mechanism responsible for separating the sample s constituents. The mobile phase is usually a liquid or a gas, and the stationary phase, when present, is a solid or a liquid film coated on a solid surface. Chromatographic techniques are often named by listing the type of mobile phase, followed by the type of stationary phase. Thus, in gas-liquid chromatography the mobile phase is a gas and the stationary phase is a liquid. If only one phase is indicated, as in gas chromatography, it is assumed to be the mobile phase. 

Solution Polymers. Acryflc solution polymers are usually characterized by their composition, solids content, viscosity, molecular weight, glass-transition temperature, and solvent. The compositions of acryflc polymers are most readily determined by physicochemical methods such as spectroscopy, pyrolytic gas—liquid chromatography, and refractive index measurements (97,158). The solids content of acryflc polymers is determined by dilution followed by solvent evaporation to constant weight. Viscosities are most conveniently determined with a Brookfield viscometer, molecular weight by intrinsic viscosity (158), and glass-transition temperature by calorimetry. 

Epichlorhydrin (ECH) detection starts with detecting epoxide cycle using hydrochloric acid in combination with sodium chloride the reaction product - 1,3-dichlorhydrin - is extracted in diethyl ether and concentrated by removing the latter. Gas-liquid chromatography with a flame-ionization detector is used to detect glycerin 1,3-dichlorhydrin. The sensitivity of the method is 0.01 mg/dm. ... 

The method of detecting dimethylterephthalate (DMTP), dibuthyl-phthalate (DBP) and diocthylphthalate (DOP) in aqueous extract is based on their extraction with an organic solvent (hexane) and subsequent concentration using gas-liquid chromatography and an electron-absorbing detector. The detection limit is 0.05 mg/dirf for DMTP and DBP, and 0,01 mg/dm for DOP. 

Dissolved in water and extracted with -heptane to remove ethylene glycol dimethacrylate (checked by gas-liquid chromatography and by NMR) and distilled twice under reduced pressure [Strop, Mikes and Kalal J Phys Chem 80 694 1 976]. 

Now, in gas/liquid chromatography, very small concentrations of solute are employed and linear absorption isotherms are to be expected. However, in LC the detectors have much lower sensitivities and as a result, significantly larger charges... 

The system proposed by Freund et al. contained all the essential properties of the modern moving bed, or pseudo moving bed chromatographic systems. The procedure was extended by Scott [6], in 1958, to gas/liquid chromatography and... 

Exchange of active hydrogens can be effected by both column and gas-liquid chromatography when the absorbant is pretreated with heavy water. It should be kept in mind, however, that other activated hydrogens are also exchanged under these conditions (see section Il-B). 

The modern electronic industry has played a very important role in the development of instrumentation based on physical-analytical methods As a result, a rapid boom in the fields of infrared, nuclear magnetic resonance (NMR), Raman, and mass spectroscopy and vapor-phase (or gas-liquid) chromatography has been observed. Instruments for these methods have become indispensable tools in the analytical treatment of fluonnated mixtures, complexes, and compounds The detailed applications of the instrumentation are covered later in this chapter. 

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