Gas Chromatography (GC; GLC; GSC) 1 “Basic Gas Chromatography” by McNair, Wiley, 1997 “Modern Practise in GC” by Grob, pp. 900 “Gas Chromatography” by Willett, pp. 250 (Wiley) Martin and Synge – 1941 idea; 1952 instrument – 1969 Nobel prize Manufacturers – Perkin Elmer, Hewlett Packard, Shimadsu, Phillips, Carlo Erba, Varian, etc – price? inexpensive; many per laboratory Separation technique — pure n’ simple – partitioning between two phases Schematic GC apparatus Liquid sample of ca. 0.1ml volume injected via a syringe into heated injector port where it is rapidly volatilised and swept by a stream of flowing (carrier) gas thru a column & out via a detector. 2 INJECTOR PORT D SIGNAL COLUMN OVEN CARRIER GAS Impurities in styrene 60m Innowax, 2ml/min He, 1ml split 80:1, 80C (9min), 5C/min to 150C 3 Schematic gas chromatograph Carrier gas (high purity, unreactive, cheap): N2, He, H2 Flow control » constant, reproducible flow rate Injection port (sample inlet, microlitre syringe) Oven — thermostatted at constant T or linear rate Column Detector Data processing » retention time (volume) » peak area » recorder, integrator, microprocessor, computer, etc 4 Powerful analytical tool — why? Very large separating power – para (138.4C), ortho (144.4C) & meta (139.1C) xylene High speed of analysis – no sample pre-treatment usually Quantitative analysis – excellent High sensitivity – 58 ppm of phenylacetylene (#11) in styrene sample Qualitative analysis the Achilles heel! – Just because peak at 18 min is labelled a-methylstyrene Simple to use and operate – unskilled, automatic, low cost 5 Qualitative analysis Identification based on retention times (volumes) Not conclusive even by comparing two or more different columns If analytes known then reasonable supposition Unambiguous? – GC + MS or – GC + IR 6 Mixture of unknown alcohols n-amyl knowns Quantitative analysis 7 5.00 ml of a soln containing an internal standard, S, of concentration 100 mg/ml were added to a soln of unknown, X. Chromatography of the mixture gave an area ratio of (AX / AS) = 0.81 ± 0.01. Calibration of known weight ratio mixes gave: – weight ratio, W = (WX / WS) 0.80 – area ratio, A = (AX / AS) 0.20 0.40 0.23 0.46 0.91 Calculate weight of X in unknown. » By least-squares: A = 1.132 W + 0.005 so if A = 0.81 then W = 0.711 but WS = 500 mg \ WX = 356 mg Detectors — key components Flame ionization family – the parent FID — workhorse, quasi-universal, reliable – flame photometric FPD — #6 sulphur/phosphorus detector – alkali flame AFID or nitrogen/phosphorus NPD or TID Electron capture – ECD — #5 halothane in blood analysis – very high sensitivity, very selective Thermal conductivity – TCD or HWD or katharometer – robust, universal, low sensitivity Mass spectrometer MSD — expensive but worth it – excellent for identification 8 Flame ionisation detector(s) FID (basic design) – mix H2 and carrier, burn in clean dust-free air – collect ions formed – current eluting cpds COLLECTOR ELECTRODE AFID (N/P sensitive) – surround jet by alkali salt – surface catalysed reactions 9 FPD (collect photons emitted) – Sulphur mode 394 nm – Phosphorus mode 526 nm AIR SIGNAL FLAME HYDROGEN CARRIER Flame ionization detector MDQ — 5 picograms / second Response — quasi-universal Linearity — excellent (over 106) Stability — flow and temperature insensitive Temperature limit — 400 C Carrier gas — Nitrogen, helium or hydrogen Summary – – – – Rugged non-responsive to water and air (“inorganics”) destructive and very widely used 10 Flame photometric detector MDQ — 1 nanogram S (394 nm); 0.1 ng P (526 nm) Response — effectively only S and P compounds Linearity — moderate (104) Stability — good Temperature limit — 400 C Carrier — nitrogen Summary – very selective – flame needs clean hydrogen/air supply – expensive but invaluable for pesticide and air pollution work 11 Flame photometric detector Sulphur mode; 394 nm – large solvent peak – small hydrocarbon peak (pentadecane) for 4,000 ng – dodecanethiol (IS) 20 ng – methyl parathion 20 ng Phosphorus mode; 526 nm – tiny solvent peak – tributyl phosphate (IS) 20 ng – methyl parathion 20 ng Same sample in both cases 12 Hot Wire Detector (TCD) Tungsten-rhenium filaments COLUMN GAS FLOW CURRENT 13 CARRIER GAS FLOW – Current of 0.3 A at 16 V Temperature of filament? 350 C but depends on thermal conductivity of gas flowing over hot wire Resistance of wire changes as T changes – Pre & post column detection SIGNAL Thermal conductivity detector MDQ — 10 nanograms (about 50 ppm) Response — universal (all except the carrier) Linearity — moderate (104) Stability — flow and temperature sensitive Carrier — hydrogen or helium Temperature limit — 400 C Summary – non-destructive and simple to operate (portable) – moderate stability and sensitivy – used for fixed gas analysis, eg, H2, N2, O2, CO2, Ar, etc 14 Electron capture detector Radioactive source emits b-particles (fast electrons) which are converted into slow electrons by collision with N2 carrier gas These are captured by molecules to form a slower moving anions Reduction in current as compound flows through detector 15 + amplifier 63 Ni or carrier gas signal 3 H ECD: organohalogen pesticides Column DB-210+ 15 m x 0.53 mm id; film 1.0 mm He carrier; 100-220C at 3C/min. 600pg each – 2-lindane; 4-aldrin; 9-dieldrin; 13-DDT 16 Electron capture detector MDQ — very high sensitivity (picogram range) Response — very selective (halogenated compounds only) Linearity — Poor ( 500 to 104) Stability — fair Temperature limit — 220 C (3H) or 350 C (Ni) Carrier — nitrogen or argon + 10% methane Summary – easily contaminated, carrier must be dry – non-destructive – requires license for radioactive source 17 ECD; biphenyls at 30 ppb each MDQ: 10 fg lindane in 2ml injection 18 The column 19 Two kinds – capillary (WCOT: 0.2 to 5 mm film thickness, PLOT) » 0.3mm id 50m 300,000 plates 0.01ml 2 ml/min – packed » 3mm id 2m 3,000 plates 10ml 40 ml/min Liquid phase – low vapour pressure over operating range & thermally stable – chemically inert to solutes – good solvent for solutes used and low viscosity Temperature – isothermal – programmed (linear, reproducible) Packed columns (SS, glass) ¼ or ½“od; coiled, U-shaped Solid support – – – – uniform pore diameter (10mm or less) large inert surface area (AW, treated with DMCS) regularly shaped, uniformly sized (mesh nos.) eg Chromosorb W/AW/DMCS 100-120 mesh Preparation (5% X on Y): – slurry 5g liquid phase X with 100g solid support Y in small quantity of suitable solvent – Rotovap off solvent, pack column – Leave overnight at highest safe temperature in oven with flow of carrier 20 Effect of column temperature Increasing the column temperature reduces retention times – biggest effect on longest times Conflict: analysis time versus resolution Temperature programming sidesteps problem – initial, final, rate of climb and timings 21 Solute classes Based on H-bonding capability – Weak bond I Polyalcohols, amino alcohols, etc II Alcohols III Ethers, ketones IV Aromatics, olefins, halocarbons V Saturated hydrocarbons 22 “Liquid” phase — the heart of the GC ‘Polarity’ Squalane — the standard phase with zero polarity Silicone gum SE30/OV-1 Dexsil 300 Di-nonylphthalate OV-210 silicone 100-300C 220 50-400C 470 0-150C 790 20-275C 1500 Polyethylene glycol (CarboWax) 60-225C 2300 OV-275 silicone 4200 100-275C 23 RTX-200 (trifluoropropylmethyl polysiloxane) 24 Stabilwax (Carbowax PEG 20M) 25 Specialised applications Pyrolysis Headspace analysis Multicolumn techniques Hyphenated Preparative GC – brake lining dust – black peppercorns or cola can – dual – back-flushing – heart-cutting – GC + MS – GC/FTIR 26 Pyrogram Headspace analysis of 0.1% cpd in water 27 Multi-column techniques A B C D SV B D C 28 A D B A* D Vent D+C A B C D SV D C B DC Vent most of A A B C D SV D+C D B A D C Backflushing to vent » speeding up analysis of A, B by not bothering with C, D Heart-cutting » analyse for B in the presence of large amounts of interfering A Dual column for difficult separations » 1st column can separate A & B but not C & D; 2nd col vice-versa