Gas Chromatography
Chromatography
is the separation of a mixture of compounds (solutes) into separate
components. By separating the sample into individual components, it is
easier to identify (qualitate) and measure the amount (quantitate) of
the various sample components. There are numerous chromatographic
techniques and corresponding instruments. Gas chromatography (GC) is one
of these techniques. It is estimated that 10-20% of the known
compounds can be analyzed by GC. To be suitable for GC analysis, a
compound must have sufficient volatility and thermal stability. If all
or some of a compound's molecules are in the gas or vapor phase at 400 -
450ºC or below, and they do not decompose at these temperatures, the
compound can probably be analyzed by GC. The introduction of
chemically bonded fused silica capillary columns was a very recent
innovation in gas chromatography, more commonly referred to high
resolution gas chromatography (HRGC). Hybrid techniques, particularly
using a mass spectrometer as a detector, GC-MS, have added a further
dimension to GC analyses enabling separated compounds to be readily
identified.  Although direct analysis is limited to compounds with a molecular weight up to 400 - 500, derivatives of non-volatile materials and metal complexes can readily be made which have some measure of volatility without thermal degradation. How does gas chromatography work? In gas chromatography (GC), the sample is vaporized and injected onto chromatographic columns and then separated into many components. The elution is brought about by the flow of an inert gaseous mobile phase. The general principle of chromatography is applicable to gas chromatography. The carrier gas serves as the mobile phase that elutes the components of a mixture from a column containing an immobilized stationary phase. In contrast to most other types of chromatography, the mobile phase dose not interact with molecules of the analytes. Carrier gases, the mobile phase of GC include helium, hydrogen and nitrogen which are chemically inert. The stationary phase in gas-solid chromatography is a solid that has a large surface area at which adsorption of the analyte species (solutes) take place. In gas-liquid chromatography, a stationary phase is liquid that is immobilized on the surface of a solid support by adsorption or by chemical bonding. Gas chromatographic separation occurs because of differences in the positions of adsorption equilibrium between the gaseous components of the sample and the stationary phases. In GC the distribution ratio is dependent on the component vapor pressure, the thermodynamic properties of the bulk component band and affinity for the stationary phase. The equilibrium is temperature dependent. Hence the importance of selecting the stationary phase of column and column temperature programming in optimizing a separation. Inside a Capillary GC Column; Capillary GC column is comprised of two major parts - tubing and stationary phase. A thin film (0.1-10.0 mm) of a high molecular weight, thermally stable polymer is coated onto the inner wall of small diameter (0.05-0.53 mm I.D.) tubing. This polymer coating is called the stationary phase. Gas flows through the tubing and is called the carrier gas or mobile phase. The rate at which each sample band moves through the column depends on the structure of the compound, the chemical structure of the stationary phase and the column temperature. The width of the sample band depends on the operating conditions and the dimensions of the column. The proper column and operating conditions are critical in obtaining no, or the least amount of, peak co-elution. It is important to select the appropriate stationary phase of columns in optimizing gas chromatographic separation. The stationary phase of column system is chosen after considering the polar characteristics of the analytes, their volatility range and column temperature program. Detector The purpose of a detector is to monitor the carrier gas as it emerges from the column and to generate a signal in response to variation in its composition due to eluted components. The requirements of a detector for GC are listed below. Detection devices for GC must respond rapidly to minute concentration of solutes as they exit the column (fast speed of response, high sensitivity). Other desirable properties of a detector are linear response, good stability, ease of operation, and uniform response to a wide variety of chemical species or, alternatively predictable and selective response to one or more classes of solutes. Gas chromatography is often coupled with the selective techniques of spectroscopy or electrochemistry. The resulting so-called hyphenated methods (for examples, GC-MS and GC-IR) provide the chemist with a powerful tool for identifying the components of complex mixtures. Flame Ionization Detector (FID) The FID is one of the most widely used and generally applicable detectors for gas chromatography and hence is used for routine and general purpose analysis. It is easy to use but destructive of the sample. FID´s consist of a hydrogen/air flame and a collector plate. FID´s are normally heated independently of the chromatographic oven. Heating is necessary in order to prevent condensation of water generated by the flame and also to prevent any hold-up of solutes as they pass from the column to the flame. The way a FID works: The effluent from the column is mixed with hydrogen and air and then ignited electrically at a small metal jet. Most organic compounds produce ions and electrons that can conduct electricity through the flame. There is an electrode above the flame to collect the ions formed at a hydrogen/air flame. The number of ions hitting the collector is measured and a signal is generated. The organic molecules undergo a series of reactions including thermal fragmentation, chemi-ionization, ion molecule and free radical reactions to produce charged-species. The amount of ions produced is roughly proportional to the number of reduced carbon atoms present in the flame and hence the number of molecules. Because the flame ionization detector responds to the number of carbon atoms entering the detector per unit of time, it is a mass-sensitive, rather than a concentration-sensitive device. As a consequence, this detector has the advantage that changes in flow rate of the mobile phase has little effects on detector response. Functional group, such as carbonyl, alcohol, halogen, and amine, yield fewer ions or none at all in a flame. In addition, the detector is insensitive toward noncombustible gases such as H2O, CO2, SO2 and NOx. Selectivity: Compounds with C-H bonds. A poor response for some non-hydrogen containing organics (e.g., hexachlorobenzene). Sensitivity: 0.1-10 ng Linear range: 105-107 Gases: Combustion - hydrogen and air; Makeup - helium or nitrogen Temperature: 250-300ºC; 400-450ºC for high temperature analyses Electron Capture Detector (ECD) The ECD is one of a family of detectors invented by Lovelock around the late 1950s and early 1960s. The ECD uses a radioactive Beta emitter (electrons) to ionize some of the carrier gas and produce a current between a biased pair of electrodes. When organic molecules that contain electronegative functional groups, such as halogens, phosphorous, and nitro groups pass by the detector, they capture some of the electrons and reduce the current measured between the electrodes. The ECD is as sensitive as the FID but has a limited dynamic range and finds its greatest application in analysis of halogenated compounds. The ECD is extremely sensitive to molecules containing highly electronegative functional groups such as halogens, peroxides, quinones, and nitro groups. It is therefore a popular detector for trace level determinations of chlorinated insecticides and halocarbon residues in environmental samples. But it is insensitive toward functional groups such as amines, alcohols, and hydrocarbons. Sensitive and selective for halogenated and other electronegative compounds, the electron capture detector (ECD) remains one of the most widely used GC detectors. The way a ECD works: Electron-capture detector (ECD) operates in much the same way as a proportional counter for measurement of X-radiation. Here the effluent from the column passes over a beta-emitter, such as 63Ni or tritium (absorbed on platinum or titanium foil). An electron from the emitter causes ionization of the carrier gas (often nitrogen) and the production of a burst of electrons. In the absence of organic species, a constant standing current between a pair of electrodes results from this ionization process. The current decreases, however, in the presence of those organic molecules that tend to capture electrons. The response is non-linear unless the potential across the detector is pulsed. Selectivity: Halogens, nitrates and conjugated carbonyls Sensitivity: 0.1-10 pg (halogenated compounds); 1-100 pg (nitrates); 0.1-1 ng (carbonyls) Linear range: 103-104 Gases: Nitrogen or argon/methane Temperature: 300-400ºC Mass Selective Detector (MS) The introduction in recent years of low cost bench-top mass spectrometers which can readily be combined with high resolution gas chromatography, justifies the inclusion of the mass spectrometer as a detector. Mass spectrometry is based upon the ionization of solute molecules in the ion source and the separation of the ions generated on the basis of their mass/charge ratio by an analyzer unit. This may be a magnetic sector analyzer, a quadruple mass filter, or an ion trap. Ions are detected by a dynode electron multiplier. The way a MS works: The detector is maintained under vacuum. Compounds are bombarded with electrons (EI) or gas molecules (CI). Compounds fragment into characteristic charged ions or fragments. The resulting ions are focused and accelerated into a mass filter. The mass filter selectively allows all ions of a specific mass to pass through to the electron multiplier. All of the ions of the specific mass are detected. The mass filter then allows the next mass to pass through while excluding all others. The mass filter scans stepwise through the designated range of masses several times per second. The total number of ions are counted for each scan. The abundance or number of ions per scan is plotted versus time to obtain the chromatogram (called the total ion chromatogram, TIC). A mass spectrum is obtained for each scan which plots the various ion masses versus their abundance or number. Operating as a simple detector, in acquision mode, the mass spectrometer scans the total mass range, typically 30 - 600 atomic mass units (amu), every few seconds, sums all the ions detected and then produces a trace on the control system PC screen. This is called a total ion chromatogram and is analogous to the trace we might obtain from any other detector. In selected ion monitoring mode (SIM), during the acquisition, the appearance of a specific compound can be traced by selecting an ion which is characteristic of that compound; either the molecular ion or the characteristic ion of a group of compounds for example: Sensitivity: 1-10 ng (full scan); 1-10 pg (SIM) Linear range: 105-106 Gases: None Temperature: 250-300ºC(transfer line); 150-250ºC(source) EI ionization method is suitable for non thermolabile compounds. The volatility of the sample is required. Sample molecules in vapor state are bombarded by fast moving electrons, conventionally 70 eV energy. This results in ion formation. One electron from the highest orbital energy is dislodged, and as a consequence molecular ions are formed. Some of these molecular ions decompose and fragment ions are formed. The fragmentation of a given ion is due to the excess of energy that it acquires within the ionization. Fragment ions can be odd electron or even electron. Molecular ions formed in electron impact ionization are odd electron ions. Odd electron fragment ions are formed by direct cleavage(e.g. direct cleavage of a C-C bond). Even electron fragment ions are often formed by rearrangement (e.g. proton transfer). Sample can be introduced to the EI source via a gas chromatography device, for example in the case of mixtures, or directly via a solids probe device. The quantities needed for an experiment is usually less than a microgram of material. EI mass spectra, in most of cases, contain intense fragment ion peaks and much less intense molecular ion peak. When the molecular ion peak is not observed in the mass spectrum, chemical ionization can be used in order to get molecular ion information. Chemical Ionization: For organic chemists, Chemical Ionization (CI) is especially useful technique when no molecular ion is observed in EI mass spectrum, and also in the case of confirming the mass to charge ratio of the molecular ion. Chemical ionization technique uses virtually the same ion source device as in electron impact, except CI uses tight ion source, and reagent gas. Reagent gas (e.g. ammonia) is first subjected to electron impact. Sample ions are formed by the interaction of reagent gas ions and sample molecules. This phenomenon is called ion-molecule reactions. Reagent gas molecules are present in the ratio of about 100:1 with respect to sample molecules. Positive ions and negative ions are formed in the CI process. Depending on the setup of the instrument (source voltages, detector, etc...) only positive ions or only negative ions are recorded. In CI, ion molecule reactions occur between ionized reagent gas molecules (G) and volatile analyte neutral molecules (M) to produce analyte ions. Pseudo-molecular ion MH+ (positive ion mode) or [M-H]- (negative ion mode) are often observed. Unlike molecular ions obtained in EI method, MH+ and [M-H]- detection occurs in high yield and less fragment ions are observed. | |||
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