INSTRUMENTATION AND APPLICATIONS OF GAS CHROMATOGRAPHY

instrumentation Instruments; Carrier Gas Flow regulators and meters Sample injection system Columns & ovens Detectors

Schematic diagram of Gas Chromatograph

Gas Chromatograph Components top view Flame Ionization Detector Injection Port Column Oven front view

Carrier gas The mobile phase gas is called the carrier gas and must be chemically inert. Sample componet  column  detector mobile phase gas Helium ,argon ,nitrogen , carbon dioxide and hydrogen also used. Selection of the best crrier gas very important , because it effects both the column separation and detector performance . The ratio of viscosity of diffusion coefficient should be minimum for rapid analysis that’s why H, He are prepared for a carrier gas .

Impurities in the carrier gas such as air water vapour and trace gaseous hydrocarbons can cause sample reaction, column character and affect the detector performance. The carrier gas system should contains a molecular sieve to remove water and other impurities. These gases are available in pressurized tanks. presure regulateres and flow meters are required to control the flow rate of the gas. The gases are supplied from the high pressure gas cylinder , being stored at pressure up to 300psi carrier gas should be better then 99.99%and 99.999% is often used

H2 inlet (detector) N2 inlet(make-up gas) Air inlet (detector)) He inlet (carrier gas)

Process Flow Schematic Detector (flame ionization detector or FID) Sample injection Carrier gas (nitrogen or helium) Air Hydrogen Long Column (30 m)

Carrier Gas(mobile phase) Requirements: It should be inert and available at low cost High purity Easily available Less risk of explosion or fire hazards Pressure: -Inlet  10 to 50 psi -packed column  25 to 150 mL/min. - capillary column  1 to 25 mL/min.

Flow regulatores & meteres Flow regulators are used to deliver the gas with uniform pressure or flow rate Flow rates of carrier gas: – Linear flow rate (cm/s): u = L/tr – Volumetric flow rate (mL/min): u (π r2) L is length of column, tr is retention time, r is the internal radius of column Flow rate depends on type of column – Packed column: 25-100 mL/min – Capillary column: 1 to 25 mL/min Flow rate will decrease as column T increases

Soap bubble meter soap bubbles formed indicates the flow rate. Glass tube with a inlet tube at the bottom. Rubber bulb-----store soap solution When the bulb is gently pressed of soap solution is converted into a bubble by the pressere of a carrier gas &travel up. Aqueous solution of soap or detergent Soap bubble flow meter

inlet tube

Sample injection port Calibated Microsyringes are used to inject liquid sample Purge :volatile components are removed from sample by gentle heating Rubber or silicone diaphragm(septum) Sample port T: 50°C Packed C: sample sizes-1 to 20 μL Capillary C: 10 to 30 mL splitter is used to deliver a fraction of injection(1:50 to 1:500) Avaid over loading Slow injection & oversized samples cause band spreading & poor resolution

Micro syringe

1. Wash a syringe with acetone by filling the syringe completely and ejecting the waste acetone onto a paper towel. Wash 2-3 times. 2. Remove air bubbles in the syringe by rapidly moving the plunger up and down while the needle is in the sample. 3.Usually 1-2 mL of sample is injected into the GC.

Column Configurations Two types of columns are used in gas chromatography, packed and open tubular or capillary. Packed column length from less than 2 m to 5 m Capillary columns from few m to 100 m They are constructed of stainless steel, glass, fused silica, or Teflon.

Column ovens Column temperature is very important in GC The column is ordinarily housed in a thermostated oven. they are usually formed as coils having diameters of 10 to 30 cm. The optimum column temperature depends upon the boiling point of the sample and the degree of separation required. Roughly, a temperature equal to or slightly above the average boiling point of a sample results in a reasonable elution time (2 to 30 min).

Column Types of columns 1.packed columns 2. Open tubular or capillary. Capillary column- 30m Packed column-3m

Packed columns Packed columns are fabricated from glass, metal (stainless steel, copper, aluminum), or Teflon tubes that typically have Lengths------ 2 to 3 m Inside diameters ------- 2 to 4 mm. These tubes are densely packed with a uniform, finely divided packing material, or solid support, that is coated with a thin layer (0.05 m) of the stationary liquid phase. In order to fit in a thermostating oven, the tubes are formed as coils having diameters of roughly 15 cm.

Older packed columns Older packed columns – uniform silica particles (150-250 μm) required to ensure uniform path lengths usually 1/8” (3.2 mm OD, 2.2 mm I.D.) diameter, 1 – 2 m length max flow rate about 1 mL/min or 8 cm/min The columns themselves were either glass or stainless steel

capillary (or)Open tubular Columns 1.Wall-coated open tubular (WCOT) Capillary tubes coated with a thin layer of stationary phase Old: stainless steel, Al, Cu, plastic, glass. 2.Support-coated open tubular (SCOT) Inner surface of the capillary is lined with a thin film (~30μm) of a support materials, like diatomaceous earth Lower efficiency than WCOT, higher than packed column 3.Fused-silica open tubular column (FSOT): Physical strength, low reactivity, flexibility. 0.32 to 0.25 mm

Column Stationary Phases: Packed liquid coated silica particles (<100-300 mm diameter) in glass tube best for large scale but slow and inefficient Capillary/Open Tubular wall-coated (WCOT) <1 mm thick liquid coating on inside of silica tube support-coated (SCOT) 30 mm thick coating of liquidcoated support on inside of silica tube best for speed and efficiency but only small samples

The Stationary Phase requirements are: Low vapor pressure Thermal stability Low viscosity (for fast mass transfer) High selectivity for compounds of interest

detectors Use: Detect the difference between a pure carrier gas &eluted compound Ideal detector: High sensitivity to even small concentrtion linerity, ie, less response to low concentration &proportional response to high concentration Large linear dynamic range Useful at a range of temperatures Good stability and reproducibility Rapid response time Easy to use Stable, Predictable response Inexpensive operation from RT to 400 oC

Types of detectors Thermal Conductivity Detector(TCD) Flame Ionization Detector(FID) Atomic Emission Detector(AED) Electron Capture Detector(ECD) Nitrogen Phosphoroes Detector(NPD) Photo Ionization Detector(PID) Flame Photometric detector(FPD) Electrolytic conductivity detector (Hall detector) Absolute Mass Detector(AMD) Thermionic Detector(TD)

Flame Ionization Detector(FID) Most widely used, Air-H2 flame Number of ions depends on number of reduced (methylene) carbons in molecule The positive ions will be attracted to the cylindrical cathode. Negative ions and electrons will be attracted to the jet anode. Organic compounds  Produces ions and electrons  pyrolyzed(temp of flame)  burner tip and electrode.(fhv power) Ions &electrons move to ward the collector less sensitive for non-hydrocarbon groups Insensitive to noncombustible gases(CO2, SO2, NO2 and H2O) Insensitive to functional group (carbonyl, alcohol, halogen and amine)

AD: High sensitivity, low noise ,wide lenear range, easy to use, fast response DA: Destroy the sample

Thermal Conductivity Detector(TCD) Element(platinum, gold or tungsten wire) is electrically heated at constant power – Temperature depends on thermal conductivity (He & H)of surrounding gas. Hydrogen and helium have higher thermal conductivity and carrier gas provide best sensitivity Six times greater than the Organic compounds Poorer sensitivity than FID, but more universal Advantages: simplicity, large range, inexpensive,linearity is excelent. organic & inorganic species DA: low sensitivity ng/mL Thermal conductivity detector cell Arrangement of the twin detectors

Electron Capture Detectors (ECD) The sample elute from a column is passed over a radioactive β-emitter(nickel-63) Selectively to halogen-containing organic sample ,like pesticides and, polychlorinated biphenyls Ni-63: radioactive β-emitter-- electron -- ionization of carrier gas (N2) High electronegative group (halogen, peroxide, quinones and nitro group) in the sample capture the electron Highly selective and sensitive, nondestructive Insensitive to amines, alcohols and hydrocarbons AD: High sensitivity, analyse the polyhalogenated organic compounds Small linear range

Thermionic detector(nitrogen phosphorus detector) N or P containing organic compounds phosphorus atom is approximately ten times greater than to a nitrogen atom and 104 to 106 larger than a carbon atom. Compared with the FID , the thermionic detector is approximately 500 times more sensitive to phosphorus-containing compounds and 50 times more sensitive to nitrogen bearing species. Column effluant + H2 +air(hot gas)electrically hearted Rb2SiO4 (rubidium silicate)bead at 180 V  plasma (600 – 800°C ) ions  to determine compounds useful for detecting and determining the many phosphorus-containing pesticides.

Atomic emission detector Eluent(column)  hellium(carrier)  water cooled microwave cavity  hellium plasma(high temp)  characterstic atomic emission spectra  gratting  diode arry optical emmision spectrometer  detect the element .

Six elements detect simultaneously . Determine the hetero atoms(H,P,S,O),silicon , heavy metals(Pb , Hg),tin, arsenic ,copper ,iron.

PHOTO IONIZATION DETECTOR(PID) UV light (10.2 eV H2 or 11.7 eV Ar lamp) photoionization of molecular  current to flow between based electrodes Most sensitive for Aromatic and S, P easily photoionized molecules Linear range is high

Flame photometric detector (FPD): S and P – compounds photomultiplier to view light of 394 nm for sulphur (H2 + air  S2) measurement or 526 nm for phosphorus (H2 + air  HPO species) Filteres are used to isolate the appropriate bands Intensity is recorded photometrically X-, N-, Sn , Cr, Se and Ge filteres photomultiplier H2 + air Column effluent

Qualitative analysis: Retention time data should be useful for identification of mixtures Comparing the retention time of the sample as well as the standard Checking the purity of a compound: compar the standard and sample Additional peaks are obtained…..impurities are present….compound is not pure

Quantitative analysis: Direct comparison method: -comparing the area of the peak, peak height, width of peak. Calibration curves: -standards of varying concentration are used determine peak areas . Internal standard method: -A known concentration of the internal standard is added separately to the standard solution -The peak area ratio of sample and internal standard….unknown concentration is easily determined .

Elemental analysis Determination of C,H ,O ,S and N . Determination of mixture of drugs Isolation and identification of drugs Isolation and identification of mixture of components(amino acids ,plant extracts ,volatile oils)