GAS CHROMATOGRAPHY AND MASS SPECTROMETRY (GC-MS) BY P.RAVISANKAR.
1.
2. The use of a mass spectrometer as the detector
in gas chromatography was developed during
the 1950s by Roland Gohlke and Fred
McLafferty.
GC = separation
MS = identification
When GC is combined with MS, a powerful
analytical tool is created.
3.
4. Desired characters:
To be suitable for GC analysis, a compound must
have sufficient volatility and thermal stability.
(All or some of compound molecules are in the gas
or vapor phase at 250-350 C or below, do not
decompose at these temperatures.)
Organic compounds must be in solution for injection
into the gas chromatograph. The solvent must be
volatile and organic (for example, hexane or
dichloromethane).
GAS CHROMATOGRAPHY
5. PRINCIPLE:
The principle of separation in GC is partition.
Gas is used as a mobile phase & liquid which is coated
on a solid support is used as a stationary phase.
The component which is more soluble in stationary
phase travels slower & eluted later:
Hence the components are separated according to their
partition coefficient.
6. Injection port – One micro liter ( 0.000001 L) of
solvent containing the mixture of molecules is
injected into the GC and the sample is carried by inert
(non-reactive) gas through the instrument, usually
helium. The inject port is heated to 300 C to cause
the chemicals to become gases.
Column – column is one of the important part of GC
which decides the separation efficiency. In GCMS
support coated open tubular columns are mostly
preferred
INSTRUMENTATION
7. Incompatibility of GC and MS
GC operate at atmospheric pressure and the MS ion source
at 10-5 Torr.
108 fold pressure difference.
Need of the interface.
The carrier gas must be removed and GC peak components
transferred to the MS ion source.
10. PRINCIPLE:-
Mass spectra is also called positive ion spectra.
In this electron bombardment is used to convert a neutral
molecule in to positively charged one.
Obtaining mass spectra consists 2 steps:
Conversion of neutral molecule in to positively charged
one.
Separation of positively charged fragments formed based
upon their masses using electrical & magnetic field:
11. Ion Source – After passing through the GC, the
chemical pulses continue to the MS. The sample
molecules must be ionized& converted in to
charged particles by the ion source before they
can be analyzed. The most commonly used
method for this purpose are :
Electron ionization.
Chemical ionization.
12. In GCMS QUADRAPO-LE ION TRAP mass analyzer is
preferred
Mass anylyzer--sample has been ionized the beam of ions is
accelerated by an electric field& then passes into the mass
analyzer, the region of the mass spectrophotometer where the
ions are separated according to their m/z ratio.
13. Detector – A detector counts the number of ions with a
specific mass. The mass spectrum is a graph of the number of
ions with different masses that traveled through the mass
analyzer. ELECTRON MULTIPLIER CELL is the detector
used in GCMS
14. The GC works on the principle that a mixture will
separate 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, flowing
into the capillary column interface connecting the
GC column and the MS (He removed).
Identification of a compound based on it's mass
spectrum relies on the fact that every compound
has a unique fragmentation pattern.
HOW GC-MS WORKS
15. The computer drives the MS, records the data, and
converts the electrical impulses into visual displays and
hard copy displays.
As each solute exits the GC column, it is diverted into a
mass spectrometer which is capable of both monitoring
the amount of and identifying the chemical nature of the
solute.
In this way, both quantitative and qualitative information
about the mixture can be obtained.
The sequence and relative intensity of the mass peaks
give information about the chemical identity of the solute.
The absolute intensity of the peaks provides information
about the amount of substance present.
18. Inject the specimen into the septum rapidly and
smoothly to attain good separation of the components in
a specimen.
If the technician injects the specimen too slowly, the
peak may be broad or overlap. A twin peak may result
from the technician hesitating during the injection.
.
19. Through GC – a chromatogram is obtained.
Through MS – a spectrum is obtained.
GC-MS gives a 3D graph which has both
chromatogram and a spectrum to each separated
component in chromatogram.
20. While the instrument
runs, the computer
generates a graph
from the signal. This
graph is called a
chromatogram.
Each of the peaks in
the chromatogram
represents the signal
created when a
compound elutes
from the GC column
into the detector.
x-axis shows the RT
y-axis shows the
intensity (abundance)
of the signal.
21. The computer records a
graph for each scan.
This graph is referred
to as a mass spectrum.
The mass spectrum is
essentially a fingerprint
for the molecule and
can be used to identify
the compound.
The library compares
the mass spectrum
from a sample
component and
compares it to mass
spectra in the library.
22. It has both the chromatogram and spectrum
on Y-axis it represents intensity/abundance, on X-axis it is
retention time.
The graph between retention time and abundance is a
chromatogram.
The graph between m/z and retention time is mass
spectrum of the individual peaks of the chromatogram.
23. Environmental Monitoring and
Cleanup
Criminal Forensics
Law Enforcement
Security
Food, Beverage and Perfume Analysis
Medicine
24. To analyze small molecule metabolite identifications in
diabetic versus non diabetic urine samples
Analysis of Anabolic Steroids in Biological materials.
Quantitation of pollutants in drinking and wastewater
using official U.S. Environmental Protection Agency (EPA)
methods.
Quantitation of drugs and their metabolites in blood and
urine for both pharmacological and forensic.
Determination of furans in food beverages
25. The separation and identification of degradation
products of organic and organometallics making the
elucidation of their structures.
The routine analysis of substances present in minute
quantities
The identification of noxious and toxic compounds
and their quantitation in emergency cases.
The diagnosis of inborn errors of metabolism
especially in new borns
Analysis of butylated hydroxytoulene in food
26. General
Only compounds with vapor pressures exceeding
about 10 torr
Many compounds with lower pressures can be
analyzed if they are chemically derivatized .
Determining positional substitution on aromatic
rings is often difficult.
Certain isomeric compounds cannot be
distinguished by mass spectrometry.
27. Dow gas chromatography and Bendix TOF mass
spectrometer in the Dow Spectroscopy Laboratory, 1957.
29. Instrumental methods of chemical analysis, Gurudeep R
Chatwal, Sham K. Anand , Pages 2.699
Practical Pharmaceutical Chemistry fourth edition-Part
two, Edited by A.H.Beckett, J.B.Stenlake, Pages 474-477
Gas Chromatography Mass Spectrometry, Ronald A.
Hites, Indiana University School of Public and
Environmental Affairs and Department of Chemistry,
Chapter 31, Pages 609-626
Journal of the American Society for Mass Spectrometry
Volume 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/Instrumen
ts/ms/7000triplequadrupolegcms/Pages/default.aspx
http://www.chemistry.nmsu.edu/Instrumentation/GC_M
S.html