GAS CHROMATOGRAPHY AND MASS SPECTROMETRY (GC-MS) BY P.RAVISANKAR.
The use of a mass spectrometer as the detectorin gas chromatography was developed duringthe 1950s by Roland Gohlke and FredMcLafferty.GC = separationMS = identificationWhen GC is combined with MS, a powerfulanalytical tool is created.
Desired characters: To be suitable for GC analysis, a compound musthave sufficient volatility and thermal stability.(All or some of compound molecules are in the gasor vapor phase at 250-350 C or below, do notdecompose at these temperatures.) Organic compounds must be in solution for injectioninto the gas chromatograph. The solvent must bevolatile and organic (for example, hexane ordichloromethane).GAS CHROMATOGRAPHY
PRINCIPLE: The principle of separation in GC is partition. Gas is used as a mobile phase & liquid which is coatedon a solid support is used as a stationary phase. The component which is more soluble in stationaryphase travels slower & eluted later: Hence the components are separated according to theirpartition coefficient.
Injection port – One micro liter ( 0.000001 L) ofsolvent containing the mixture of molecules isinjected into the GC and the sample is carried by inert(non-reactive) gas through the instrument, usuallyhelium. The inject port is heated to 300 C to causethe chemicals to become gases. Column – column is one of the important part of GCwhich decides the separation efficiency. In GCMSsupport coated open tubular columns are mostlypreferredINSTRUMENTATION
Incompatibility of GC and MS GC operate at atmospheric pressure and the MS ion sourceat 10-5 Torr.108 fold pressure difference.Need of the interface. The carrier gas must be removed and GC peak componentstransferred to the MS ion source.
PRINCIPLE:- Mass spectra is also called positive ion spectra. In this electron bombardment is used to convert a neutralmolecule in to positively charged one. Obtaining mass spectra consists 2 steps: Conversion of neutral molecule in to positively chargedone. Separation of positively charged fragments formed basedupon their masses using electrical & magnetic field:
Ion Source – After passing through the GC, thechemical pulses continue to the MS. The samplemolecules must be ionized& converted in tocharged particles by the ion source before theycan be analyzed. The most commonly usedmethod for this purpose are : Electron ionization. Chemical ionization.
In GCMS QUADRAPO-LE ION TRAP mass analyzer ispreferredMass anylyzer--sample has been ionized the beam of ions isaccelerated by an electric field& then passes into the massanalyzer, the region of the mass spectrophotometer where theions are separated according to their m/z ratio.
Detector – A detector counts the number of ions with aspecific mass. The mass spectrum is a graph of the number ofions with different masses that traveled through the massanalyzer. ELECTRON MULTIPLIER CELL is the detectorused in GCMS
The GC works on the principle that a mixture willseparate into individual substances when heated. Sample introduced into GC inlet vaporized at 250 C, swept onto the column by He carrier gas &separated on column. Sample components emerge from column, flowinginto the capillary column interface connecting theGC column and the MS (He removed). Identification of a compound based on its massspectrum relies on the fact that every compoundhas a unique fragmentation pattern.HOW GC-MS WORKS
The computer drives the MS, records the data, andconverts the electrical impulses into visual displays andhard copy displays. As each solute exits the GC column, it is diverted into amass spectrometer which is capable of both monitoringthe amount of and identifying the chemical nature of thesolute.In this way, both quantitative and qualitative informationabout the mixture can be obtained. The sequence and relative intensity of the mass peaksgive information about the chemical identity of the solute. The absolute intensity of the peaks provides informationabout the amount of substance present.
Inject the specimen into the septum rapidly andsmoothly to attain good separation of the components ina specimen. If the technician injects the specimen too slowly, thepeak may be broad or overlap. A twin peak may resultfrom the technician hesitating during the injection..
Through GC – a chromatogram is obtained. Through MS – a spectrum is obtained. GC-MS gives a 3D graph which has bothchromatogram and a spectrum to each separatedcomponent in chromatogram.
While the instrumentruns, the computergenerates a graphfrom the signal. Thisgraph is called achromatogram. Each of the peaks inthe chromatogramrepresents the signalcreated when acompound elutesfrom the GC columninto the detector. x-axis shows the RT y-axis shows theintensity (abundance)of the signal.
The computer records agraph for each scan.This graph is referredto as a mass spectrum. The mass spectrum isessentially a fingerprintfor the molecule andcan be used to identifythe compound. The library comparesthe mass spectrumfrom a samplecomponent andcompares it to massspectra in the library.
It has both the chromatogram and spectrum on Y-axis it represents intensity/abundance, on X-axis it isretention time. The graph between retention time and abundance is achromatogram. The graph between m/z and retention time is massspectrum of the individual peaks of the chromatogram.
Environmental Monitoring andCleanup Criminal Forensics Law Enforcement Security Food, Beverage and Perfume Analysis Medicine
To analyze small molecule metabolite identifications indiabetic versus non diabetic urine samples Analysis of Anabolic Steroids in Biological materials. Quantitation of pollutants in drinking and wastewaterusing official U.S. Environmental Protection Agency (EPA)methods. Quantitation of drugs and their metabolites in blood andurine for both pharmacological and forensic. Determination of furans in food beverages
The separation and identification of degradationproducts of organic and organometallics making theelucidation of their structures. The routine analysis of substances present in minutequantities The identification of noxious and toxic compoundsand their quantitation in emergency cases. The diagnosis of inborn errors of metabolismespecially in new borns Analysis of butylated hydroxytoulene in food
General Only compounds with vapor pressures exceedingabout 10 torr Many compounds with lower pressures can beanalyzed if they are chemically derivatized . Determining positional substitution on aromaticrings is often difficult. Certain isomeric compounds cannot bedistinguished by mass spectrometry.
Dow gas chromatography and Bendix TOF massspectrometer in the Dow Spectroscopy Laboratory, 1957.
Instrumental methods of chemical analysis, Gurudeep RChatwal, Sham K. Anand , Pages 2.699 Practical Pharmaceutical Chemistry fourth edition-Parttwo, Edited by A.H.Beckett, J.B.Stenlake, Pages 474-477 Gas Chromatography Mass Spectrometry, Ronald A.Hites, Indiana University School of Public andEnvironmental Affairs and Department of Chemistry,Chapter 31, Pages 609-626 Journal of the American Society for Mass SpectrometryVolume 4, Issue 5, May 1993, Pages 367-371 http://www.labcompare.com/Mass-Spectrometry/154-Gas-Chromatograph-Mass-Spectrometer-GC-MS-Instrument/ http://www.chem.agilent.com/enUS/Products/Instruments/ms/7000triplequadrupolegcms/Pages/default.aspx http://www.chemistry.nmsu.edu/Instrumentation/GC_MS.html