Gas Chromatography (GC)
Gas Chromatography : Introduction
Gas Chromatography is a process of separating component(s) from a compound/mixture by using a gaseous mobile phase.
It involves a sample being vaporized and injected onto the head of the chromatographic column. The sample is transported through the column by the flow of inert, gaseous mobile phase, The column itself contains a liquid stationary phase which is adsorbed onto the surface of an inert solid
Principle of Separation : Partition Chromatography
Two major types
- Gas-solid chromatography (stationary phase: solid)
- Gas-liquid chromatography (stationary phase: immobilized liquid)
Because separation of compound mixtures on the column occurs while they are in the gaseous state, solid and liquid samples must first be vaporized.
GC is limited to the study of thermostable and sufficiently volatile compounds
Characteristic of separation:
- Volatility – must be volatile (boiling point)
Better for organic compounds and very sensitive even in pico grams
Interaction in GC
GC is the only form of chromatography that does not utilize a mobile phase for interacting with the analyte
When the stationary phase is a solid adsorbent, the process is termed
gas–solid chromatography (GSC), and when it is a liquid on an inert support, the process is termed gas–liquid chromatography (GLC)
Components of GC
Gas Flow in the GC:
The mobile phase that transports the analytes through the column is a gas referred to as the carrier gas (as it only carries the analyte but don't react with it)
Analysis starts when a small quantity of sample is introduced as either liquid or gas into the injector, which has the dual function of vaporizing the sample and mixing it with the gaseous flow at the head of the column
The column is usually a narrow-bore tube which coils around itself with a length that can vary from 1 to over 100 m, The column, which can serve for thousands of successive injections, is housed in a thermostatically controlled oven. At the end of the column, the mobile phase (carrier gas), passes through a detector before it exits to the atmosphere
Example of component Seperation in GC
Assume two components (green and orange) need to be seperated using GC. Orange runs fast in GC which shows its low relative affinity with the stationary phase and more affinity to be in gas phase; thus it must be a low boiling point analyte however green component prefer to stay in stationary phase thus have high relative affinity with liquid stationary phase and thus is a high boiling point component.
The component with low boiling point / high volatility shows the first in detector as move fast in column; also it Boiling Point and volatility same than the lighter one i.e. smaller size component can travel fast.
- First peak is solvent in which we dissolved our component, very low boiling point thus pushed faster
- Interaction with stationary phase...orange like interact with gas; low boiling point
- Green : high Boiling point; prefer to stay in liquid phase
- Can also see difference in polarity and volatility
- If similar Boiling Point then can be also separated by size
Operational parameters of GC
For a given stationary phase:
- L, length of the column,
- u, velocity of the mobile phase
- T, temperature of the column
- Phase ratio which affects the retention factor k
The operating conditions of the chromatograph allows modifications in terms of T and u and therefore affects both the efficiency of the column and the retention factors
Compression Factor (J)
The pressure at the head of the column is stabilized either mechanically or through an electronic pressure control (EPC) in order that the flow rate remains constant at its optimal value
The injector and the detector have dead volumes (hold-up volumes) which are counted in the total retention volume. In GC, since the mobile phase is a gas, the flow rate measured at the outlet of the column should be corrected by a compression factor J, which compensates for the higher pressure at the head of the column
Pressure gradient correction factor (in gas chromatography) A factor that corrects for the compressibility of the carrier gas. The values of the measured quantities obtained after multiplication by the factor j are independent of the pressure drop in the column. If pi, po are respectively the pressures of the carrier gas at the inlet and outlet of the column, then J is given by:
Carrier gas and flow regulation
The mobile phase is a gas (helium, hydrogen or nitrogen), carrier gas must be free of all traces of hydrocarbons, water vapour and oxygen, because all of these may deteriorate polar stationary phases or reduce the sensitivity of detectors
Carrier gas system includes filters containing a molecular sieve to remove water and a reducing agent for other impurities
Types of GC Columns
- Packed columns
- Capillary columns
For packed columns the stationary phase is deposited or bonded by chemical reaction onto a porous support. 1.5 – 10m in length and 2 – 4mm internal diameter. These are generally made of stainless steel or glass
For capillary columns a thin layer of stationary phase is deposited onto, or bound to the inner surface of the capillary columns are 0.1 – 0.5 mm id and can be 10 – 100m long
Three types of capillary columns are commonly used in gas chromatography:
- Wall Coated Open Tubular (WCOT)
- Support Coated Open Tubular (SCOT)
- Porous Layer Open Tubular Column (PLOT)
- Fused Silica Open Tubular (FSOT)
Wall-Coated Open Tubular Column
Wall-Coated Open Tubular Column the internal wall is directly coated with the very thin stationary-phase layer at a film thickness of 0.05–3 μm. The process is done by passing a solution of liquid S.P. (dissolved in an organic solvent), blowing the column dry with a stream of inert gas.
Support-Coated Open Tubular Column
Capillary tube wall is lined with a thin layer of very fine solid support (such as Celite) on to which liquid phase is adsorbed. The separation efficiency of SCOT columns is more than WCOT columns because of increased surface area of the stationary phase coating
Porous Layer Open Tubular Column
Columns contain a porous layer of a solid adsorbent such as alumina, molecular sieves. Porosity can be achieved by either chemical means (e.g., etching) or by the deposition of porous particles on the wall from a suspension. The porous layer may serve as a support for a liquid stationary phase or as the stationary phase itself. PLOT columns are well suited for the analysis of light, fixed gases, and other volatile compounds
Fused Silica Open Tubular Column
The fused silica tubes have much thinner walls than glass capillary columns, and are strengthened by the polyimide coating. These columns are flexible and can be wound into coils. They offer the advantages of physical strength, flexibility, and low reactivity
The difference can be seen clerly in the picture given below
Inert materials is used, including glass, nickel, fluorocarbon polymer (Teflon), and steel covered with glass or Teflon. The packing is an inert support impregnated with 5–20% stationary phase
The solid support holds the liquid stationary phase which
- should have a large surface area,
- be chemically inert,
- have low sorptive activity toward common analytes,
- and have good mechanical strength to prevent the fracture of the coated particles during loading and handling
Diatomaceous earth, composed of hydrous silica with impurities, has been used as a solid support under the brand name Chromosorb
Packed versus Capillary Column
Capillary columns offer certain advantages relative to packed columns. Capillary columns are coated with a thin, uniform liquid phase. Because of the smooth, inert surface of fused silica, high efficiency can be achieved, typically 3000–5000 theoretical plates per meter. In contrast, packed columns have thicker, often non-uniform films, and generate only 2000 plates per meter
Due to the small pressure drop associated with open tube capillary columns, long columns of up to 60m can easily be used. However, packed columns are tightly filled with solid support and suffer from greater pressure drops; thus, it is impossible to use packed columns much longer than 2m
Resolution is proportional to the square root of the column length
Stationary phase in GC
Selectivity in gas chromatography is influenced by the choice of stationary phase
Elution order in GLC is determined primarily by the solute’s boiling point and, to a lesser degree, by the solute’s interaction with the stationary phase. Solutes with significantly different boiling points are easily separated. On the other hand, two solutes with similar boiling points can be separated only if the stationary phase selectively interacts with one of the solutes
The main criteria for selecting a stationary phase are that it should be chemically inert, thermally stable, of low volatility, and of an appropriate polarity for the solutes being separated
Liquid Stationary phases
Types of Detectors used in GC
Non-selective/Universal Detector – Responds to all compounds present in carrier gas stream except the carrier gas itself
Selective Detector – Responds to range of compounds with a common physical or chemical characteristic
Specific Detector – Responds to a single specific compound only
Detectors can also be grouped into concentration or mass flow detectors
Concentration Dependent – The response of such Gas Chromatography detectors is proportional to the concentration of the solute in the detector such as TCD. Dilution of sample with makeup gas will lower detector response
Mass Flow Dependent – Signal is dependent on the rate at which solute molecules enter the detector such as FID. Response of such detectors is not affected by makeup gas flow rate changes
Desirable characteristics of detectors
Flame Ionization Detector (FID)
- FID makes use of an oven, wherein a flame is produced by burning hydrogen gas in presence of oxygen or air; Effluent from the column is directed into a air/hydrogen flame
- A definite potential difference is maintained between the two electrodes with the help of a series of batteries
- Amplifier and recorder record chromatograms
The operation of the FID is based on the detection of ions formed during combustion of organic compounds in a hydrogen flame. The generation of these ions is proportional to the concentration of organic species in the sample gas stream.
- A portion of eluate coming from the column is directed into the furnace through the wire loop
- Solvent evaporates and organic compounds pyrolyses and forms ions
- These ions are attracted towards the respective electrodes
- This changes the potential difference between the electrodes and hence the current in the circuit
- As electrical resistance of flame is high and resulting current is small, an electrometer is employed
Advantages and Disadvantages
Minute amount of solute can be detected gives linear response
As it responds to the number of C- atoms entering the detector per unit time, it is mass sensitive rather than concentration sensitive
It is selective towards compounds containing sulphur and phosphorous (P at 526nm filter and S at 394nm filter)
It requires a combustion chamber to house the flame, gas lines for hydrogen (fuel) and air (oxidant), an exhaust chimney to remove combustion products, thermal (bandpass) filter to isolate only the visible and UV radiation emitted by the flame
Thermal Conductivity Detector (TCD)
Also known as Katharometer. This detector senses changes in the thermal conductivity of the column effluent and compares it to a reference flow of carrier gas.
Since most compounds have a thermal conductivity much less than that of the common carrier gases of helium or hydrogen, when an analyte elutes from the column the effluent thermal conductivity is reduced, and a detectable signal is produced
Non-destructive detector , inexpensive but low in sensitivity
It works on the principle of wheatstone’s bridge - Out of four resistances in the circuit, the magnitude of three resistances remains constant. But that of fourth resistance varies as per change in the temperature. This change is because of the difference in the capacity of the solute and the carrier gas to absorb heat (thermal conductivity differences). The change in the temperature changes the resistance and hence the current in circuit
Electron Capture Detector (ECD)
The ECD uses a radioactive beta particle (electron) emitter in conjunction with a so-called makeup gas flowing through the detector chamber. Usually, nitrogen is used as makeup gas, because it exhibits a low excitation energy, so it is easy to remove an electron from a nitrogen molecule. The electrons emitted from the electron emitter collide with the molecules of the makeup gas, resulting in many more free electrons
The analyte concentration is thus proportional to the degree of electron capture.
Photo Ionization Electrode (PID)
PID design uses a 10.6eV lamp with a high voltage power supply. Sample laden carrier gas flows from the analytical column into the PID sample inlet. When sample molecules flow into the cell, they are bombarded by the UV light beam. Molecules with an ionization potential lower than 10.6eV release an ion when struck by the ultraviolet photons. These ions are attracted to a collector electrode, then sent to the amplifier to produce a signal
Mechanism: Compounds eluting into a cell are bombarded with high energy photons emitted from a lamp. Compounds with ionization potentials below the photon energy are ionized. The resulting ions are attracted to an electrode, measured, and a signal is generated.
Selectivity: Depends on lamp energy. Usually used for aromatics and olefins (10 eV lamp)
Nitrogen Phosphorous Detector (NPD)
NPD uses a Hydrogen/Air flame through which the sample is passed.
It uses a rubidium/cesium bead which is heated by a coil, over which the carrier gas mixed with Hydrogen
The hot bead emits electrons by which are collected at the anode and provides the background current
When a component that contains N/P exits the column, the partially combusted N/P materials are adsorbed on the surface of the bead; this then increases the emission of electrons
NPD is used for Herbicides analysis
Mass Spectrometry (MS)
Mass Spectrometry comes as a detector associated with GC known as GC MS. It is widely used because of its precise qualitative as well as quantitaive measurements.
Mechanism: 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. A mass spectrum is obtained for each scan which plots the various ion masses versus their abundance or number.
Selectivity: Any compound that produces fragments within the selected mass range. May be an inclusive range of masses (full scan) or only select ions (SIM)
Atoms and molecules can be deflected by magnetic fields - provided the atom or molecule is first turned into an ion. Electrically charged particles are affected by a magnetic field.
Steps involved in MS analysis
Ionisation : The atom or molecule is ionised by knocking one or more electrons off to give a positive ion. Most mass spectrometers work with positive ions
Acceleration : The ions are accelerated so that they all have the same kinetic energy
Deflection : The ions are then deflected by a magnetic field according to their masses. The lighter they are, the more they are deflected. The amount of deflection also depends on the number of positive charges on the ion - in other words, on how many electrons were knocked off in the first stage. The more the ion is charged, the more it gets deflected.
Detection : The beam of ions passing through the machine is detected electrically
The need for a vacuum - It's important that the ions produced in the ionisation chamber have a free run through the machine without hitting air molecules
The vaporised sample passes into the ionisation chamber. The electrically heated metal coil gives off electrons which are attracted to the electron trap which is a positively charged plate. The particles in the sample (atoms or molecules) are therefore bombarded with a stream of electrons, and some of the collisions are energetic enough to knock one or more electrons out of the sample particles to make positive ions.
Most of the positive ions formed will carry a charge of +1 because it is much more difficult to remove further electrons from an already positive ion. These positive ions are persuaded out into the rest of the machine by the ion repeller which is another metal plate carrying a slight positive charge
Flow chart for working of MS
Environmental Applications of GC
Gas Chromatography is now a days widely used in Environmental Analysis:
- Volatile organic compounds (VOCs);
- polycyclic aromatic hydrocarbons (PAHs);
- pesticides; and, halogenated compounds.
Include polychlorinated dibenzo-p-dioxins and dibenzofurans, polychlorinated biphenyl, terphenyls, naphthalenes and alkanes, organochlorine pesticides, and the brominated flame retardants, polybrominated biphenyls and polybrominated diphenylethers