GC and GC-MS

Gas Chromatography
• • • • • Function Components Common uses Chromatographic resolution Sensitivity

Function
• Separation of volatile organic compounds • Volatile – when heated, VOCs undergo a phase transition into intact gas-phase species • Separation occurs as a result of unique equilibria established between the solutes and the stationary phase (the GC column) • An inert carrier gas carries the solutes through the column

Components
• • • • • Carrier Gas, N2 or He, 1-2 mL/min Injector Oven Column Detector

Syringe Injector Detector Gas tank Column Oven .

Injector • A GC syringe penetrates a septum to inject sample into the vaporization camber • Instant vaporization of the sample. 280 C • Carrier gas transports the sample into the head of the column • Purge valve controls the fraction of sample that enters the column .

Splitless (100:90) vs. Split (100:1) Syringe Syringe Injector Injector He He Purge valve closed GC column GC column Purge valve open .

refocusing to a more narrow sample plug is possible with temperature programming . which means the sample plug is 30-60 seconds • As we will see.Split or splitless • Usually operated in split mode unless sample limited • Chromatographic resolution depends upon the width of the sample plug • In splitless mode the purge valve is close for 3060 s.

Open Tubular Capillary Column 0.32 mm ID Mobile phase (Helium) flowing at 1 mL/min Liquid Stationary phase 0.1-5 mm 15-60 m in length .

FSOT columns • Coated with polymer. crosslinked – Polydimethyl soloxane (non-polar) – Poly(phenylmethyldimethyl) siloxane (10% phenyl) – Poly(phenylmethyl) siloxane (50% phenyl) – Polyethylene glycol (polar) – Poly(dicyanoallyldimethyl) siloxane – Ploy(trifluoropropyldimethyl) siloxane .

C18 saturated methyl ester .Polar vs. olefins. fatty acids) retention time increases with chain length (or molecular weight) • Polar columns retain polar compounds to a greater extent than non-polar – C18 saturated vs. • Within a homologous series (alkanes. nonpolar • Separation is based on the vapor pressure and polarity of the components. alcohol.

C16:0 C18:1 C18:0 C18:2 C16:1 Polar column RT (min) C18:2 C18:1 C16:0 C16:1 RT (min) C18:0 Non-polar column .

run at one constant temperature • Temperature programming .Start at low temperature and gradually ramp to higher temperature – More constant peak width – Better sensitivity for components that are retained longer – Much better chromatographic resolution – Peak refocusing at head of column .Oven • Programmable • Isothermal.

Typical Temperature Program 220C 160C 50C 0 Time (min) 60 .

Detectors • Flame Ionization Detectors (FID) • Electron Capture Detectors (ECD) • Electron impact/chemical ionization (EI/CI) Mass spectrometry .

FIDs • Effluent exits column and enters an air/hydrogen flame • The gas-phase solute is pyrolized to form electrons and ions • All carbon species are reduced to CH2+ ions • These ions collected at an electrode held above the flame • The current reaching the electrode is amplified to give the signal .

FID • • • • • A general detector for organic compounds Very sensitive (10-13 g/s) Linear response (107) Rugged Disadvantage: specificity .

ECD • Ultra-sensitive detection of halogencontaining species • Pesticide analysis • Other detectors besides MS – IR – AE .

Mass Spectrometry .

What kind of info can mass spec give you? • Molecular weight • Elemental composition (low MW with high resolution instrument) • Structural info (hard ionization or CID) .

How does it work? • Gas-phase ions are separated according to mass/charge ratio and sequentially detected .

) .high vac Detector (electron multiplier tube) .Parts of a Mass Spec • • • • Sample introduction Source (ion formation) Mass analyzer (ion sep.

CI Involatiles • Direct infusion/electrospray (ESI) • HPLC/ESI • Matrix Assisted Laser Adsorption (MALDI) Elemental mass spec • Inductively coupled plasma (ICP) • Secondary Ion Mass Spectrometry (SIMS) – surfaces .Chemical ionization (CI) • GC/EI.Sample Introduction/Sources Volatiles • Probe/electron impact (EI).

EI. CI • EI (hard ionization) – Gas-phase molecules enter source through heated probe or GC column – 70 eV electrons bombard molecules forming M+* ions that fragment in unique reproducible way to form a collection of fragment ions – EI spectra can be matched to library stds • CI (soft ionization) – Higher pressure of methane leaked into the source (mtorr) – Reagent ions transfer proton to analyte .

EI Source Under high vacuum filament 70 eV e- To mass analyzer GC column anode repeller Acceleration slits .

M will fragment in the same pattern every time using a 70 eV electron beam .EI process • M + eM+* f1 f2 f3 f4 This is a remarkably reproducible process.

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Ion Chromatogram of Safflower Oil .

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usually not enough to give extensive fragmentation .CI/ ion-molecule reaction • 2CH4 + e. CH5+ and C2H5+ • CH5+ + M  MH+ + CH4 • The excess energy in MH+ is the difference in proton affinities between methane and M.

EI spectrum of phenyl acetate .

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Mass Analyzers • Low resolution – Quadrupole – Ion trap • High resolution – TOF time of flight – Sector instruments (magnet) • Ultra high resolution – ICR ion cyclotron resonance .

Resolution • • • • R = m/z/Dm/z Unit resolution for quad and trap TOF up to 15000 FT-ICR over 30000 – MALDI. Resolve 13C isotope for a protein that weighs 30000 – Resolve charge states 29 and 30 for a protein that weighs 30000 .

Quad – Unit resolution . ICR – complete separation of the isotope peaks of a +3 charge state peptide – Ion abundances are predictable – Interferences can be recognized and sometimes eliminated • Ion trap.High vs low Res ESI • Q-TOF.

0 m/z 901.7 595.2 891.594.3 Q-TOF 595.3 594.7 602.0 601.88 m/z .3 601.6 R = 0.3 892.0 MVVTLIHPIAMDDGLR C78H135N21O22S2+3 601.7 902.4 100 95 90 85 80 75 70 65 60 55 50 45 40 35 30 25 20 15 10 5 0 LCQ 891.6 900.

Quadrupole Mass Ion Filter .

Ion Trap .

Time of Flight -TOF .

6022x10-19 C) .Where: •mi = mass of analyte ion •zi = charge on analyte ion •E = extraction field •ti = time-of-flight of ion •ls = length of the source •ld = length of the field-free drift region •e = electronic charge (1.

TOF with reflectron http://www.html .rmjordan.com/tt1.

chem.edu/mass/tutorials/magnetmovie.Sector instruments http://www.html .harvard.

pdf .edu/chemistry/chem5 181/MS_FT-ICR_Huffman_Abraham.FT-ICRMS • http://www.colorado.

1011 (only 1 possibility) • Sector instruments.Mass accuracy • Mass Error = (5 ppm)(201.0991 to 201. TOF mass analyzers • How many possibilities with MA = 50 ppm? with 100 ppm? .1001)/106 =  0.0010 amu • 201.

Exact Mass Determination • Need Mass Spectrometer with a high mass accuracy – 5 ppm (sector or TOF) • C9H15NO4.0010 amu .1001 (mono-isotopic) • Mass accuracy = {(Mass Error)/FM}*106 • Mass Error = (5 ppm)(201. FM 201.1001)/106 =  0.

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