PRESENTED BY SAIRA FATIMA SABAHAT MEHMOOD SANA USMAN MSc 4 (2018-2020) Department of MicroBiology & Molecular Genetics University of the Punjab Lahore, Pakistan

1. GAS CHROMATOGRAPHY
AND MASS
SPECTROMETRY
PRESENTED BY
SAIRA FATIMA
SABAHAT MEHMOOD
SANA USMAN
Department of MicroBiology & Molecular Genetics
University of the Punjab
Lahore, Pakistan
MSc
2018-2020

2. Introduction:
 Gas chromatography-mass spectroscopy (GC-MS) is a hyphenated
analytical technique.
 It is sensitive but also specific and reliable.
 GC can separate volatile and semi-volatile compounds with great
resolution, but it cannot identify them.
 MS provide detailed structural information on most compounds such that
they can be exactly identified, but can’t readily separate them.
 Therefore, both instruments have been proposed shortly after the
development of GC in the mid-1950s.
 We obtain both qualitative and quantitative information of our sample in a
single run within the same instrument.
 Today computerized GC/MS instruments are widely used in environmental
monitoring ,in the regulation of agriculture and food safety , and in the
discovery and production of medicine.

3. It is used in identification of unknown samples, including that
of material samples obtained from planet Mars during probe
missions as early as the 1970s.
It can also be used in airport security to detect substances in
luggage or on human beings. Additionally, it can identify trace
elements in materials that were previously thought to have
disintegrated beyond identification.
Like liquid chromatography–mass spectrometry, it allows
analysis and detection even of tiny amounts of a substance.

4. History
 The use of mass spectrometer as a detector in gas chromatography was
developed during the 1950s by Roland.
 Miniaturized computers has helped in the simplification of instrument
 In 1968, the Finnigan Instrument Corporation delivered the first
quadrupole GC/MS
 By the 2000s computerized GC/MS instruments using quadrupole
technology had become essential

5. Principle of GC-MS
 The sample solution is injected into the GC inlet where it is vaporized and swept
onto a chromatographic column by the carrier gas (usually helium).
 The sample flows through the column and the compounds comprising the
mixture of interest are separated by virtue of their relative interaction with the
coating of the column (stationary phase) and the carrier gas (mobile phase).
 The latter part of the column passes through a heated transfer line and ends at
the entrance to ion source where compounds eluting from the column are
converted to ions and detected according to their mass to charge m/z ratio

6. Instrumentation
 The GC-MS is composed of two major building blocks, the gas chromatograph and
the mass spectrometer.
 The gas chromatograph utilizes a capillary column which depends on the column's
dimensions (length, diameter, film thickness) as well as the phase properties (e.g.
5% phenyl polysiloxane).
 The difference in the chemical properties between different molecules in a
mixture and their relative affinity for the stationary phase of the column will
promote separation of the molecules as the sample travels the length of the
column.
 The molecules are retained by the column and then elute (come off) from the
column at different times (called the retention time), and this allows the mass
spectrometer downstream to capture, ionize, accelerate, deflect, and detect the
ionized molecules separately. The mass spectrometer does this by breaking each
molecule into ionized fragments and detecting these fragments using their mass-
to-charge ratio.

7. GC-MS

8. Steps:-
1. Inject sample into Gas Chromatograph (GC) – The sample is injected into a port
which is heated to up to 300° C where the material is then volatilized.
2. Separation of gaseous components as they flow through the column – The column
is wound within a special oven which controls temperatures from -20° to
320°. The column surface is coated with a material which will separate the
various chemical compounds in the sample based on size and/or polarity. Sample
components that are more volatile and smaller in size will travel through the
column more quickly than others.
3. Analysis in the Mass Spectrometer (MS) – The separated components flow directly
out of the column and into the MS which has three internal steps:
a) Ionization source – components are blasted with electrons, causing them to
break up and turn into positively charged ions

9. Filter – the ions pass through an electromagnetic
field and filter through based on mass. Analysts set a
predetermined range of masses to be allowed
through as they pass from the ionization source.
Detector – counting the number of filtered ions,
the information is sent to a computer and a mass
spectrum, a distribution of ions of different sizes, is
generated.

10. Gas chromatography
1. Technique used for the separation of volatile compounds that were
easily vaporized at room temperature.
Components
1. Column
2. Mobile phase
3. Stationery phase
4. Detector

11. Column:-
Used in gas chromatography very long and arranged in a coil
1. Two types of column
A. Packed column
Consist of glass and stainless steel having Length 1 to 3 meters and
internal diameter 2 to 4 millimeter
A. Capillary column
It made of fused quartz having a length 10 to 100 meters and internal
diameter 0.1 to 1 mm
Column is placed in a chamber so that uniform temperature can be
maintained

12. Stationary phase
 Stationary phase is packed in the inner wall of the column
 Stationary phase is made of silicon grease or wax which can stand at
high temperature.

13. Mobile Phase
 Inert gas helium usually is used as a mobile phase
 Non reactive gas is also used as a mobile phase
 The mobile gas is packed in a cylinder which connected to the column
via molecular sieve which is used to remove unwanted gasses such as
hydrocarbon, water vapours and oxygen

14. Detector
 Detector present at the end of column, used to detect test samples
 Flamed ionization detector is commonly used in gass chromatography
 FID have three inlet
1. Attached with column
2. With hydrogen cylinder
3. Attached with oxygen cylinder
Hydrogen oxygen ignite produces flame when sample molecule reached
ionized and detect on detector in the form of electric current

15. Diagram

16. Working of gas chromatography
 Separated sample mixed with appropriate volatile such as heptane,
acetone and methanol
 Just before the column septum is present which is used to inject sample
 Temperature of the injection region is kept 20 to 50 degree centigrade
higher than the column
 Allowed the rapid volatilization of the sample
 This sample pass through the column where separation occurs
 During analyses temperature of the column is 150 to 300 degree
centigrade.

17. Separation principles
 Separation occurs on the base of interaction of the molecule with
stationary phase and volatile phase
 More volatile molecule less interact with stationary phase and move fast
 Less volatile molecule more interact with stationary phase and move
slowly
 When the separation is completed , we can seen results on detector.

18. Diagram

19. Mass spectrometry
 The mass spectrometer is an instrument which help in separating the
individual atoms or molecules because of difference in their masses.
 A mass spectrometer generates multiple ions from the sample under
investigation
 It then separates them according to their specific mass-to-charge ratio
(m/z), and then records the relative abundance of each ion type
 Produce ions from the sample in the ionization source.
 Separate these ions according to their mass-to-charge ratio in the mass
analyzer.

20. Components:-
 The instrument consists of three major components:
 Ion Source: For producing gaseous ions from the substance being
studied.
 Analyzer: For resolving the ions into their characteristics mass
components according to their mass-to-charge ratio.
 Detector System: For detecting the ions and recording the relative
abundance of each of the resolved ionic species.

21. Coupling of Gc-Ms
 Gas chromatograph and Mass spectrometry GC-MS Separates mixture
of components into individual Identifies molecules based on their mass
A chemical analysis technique combining two instruments to provide for
powerful separation & identification
 Interface play important role in GCMS

22. Types of interface
 Capillary direct interface
 Jet separator (packed column)
 Watson & Biermann effusion separator

23. Capillary direct interface
 Today most GC-MS systems use capillary columns & fused silica
tubing
 Permits an inert, high efficiency, direct transfer between the 2 systems.
 Fused silica tubing permits an inert, high efficiency, direct transfer
between the 2 systems.
 Flow rates is 5ml/min

24. Jet separator (packed column)
 The separator consist of two glass tubes aligned with a Small distance
between them.
 Carrier gas entering from the GC column is pumped away by a
separate vacuumed system.
 The larger sample molecules maintain their momentum and pass
preferentially in to the second capillary.
 Sample enrichment occurs & the initial atmospheric pressure is
reduced.

25. Watson & Biermann effusion separator
 It consists of a sintered glass tube .
 The carrier usually Helium, passes preferentially through the sintered
glass tube & the effluent in concentrated by a factor of up to 100.
 The gas flow rates in the order of 20-60ml/min.

26.  Applications
 Advantages
 Disadvantages
 Summary

27. Applications
 Criminal forensics
 Law enforcement
 Spot anti doping analysis
 Chemical warfare agent detection
 Chemical engineering
 Food, beverage and perfume analysis
 Astro chemistry
 Medicine

28.  GC-MS can analyze the particles from a human body in
order to help link a criminal to a crime
 there is even an established American Society for Testing
and Materials (ASTM) standard for fire debris analysis
 GCMS/MS is especially useful here as samples often
contain very complex matrices and results, used in court,
need to be highly accurate.
Criminal forensics

29. Law enforcement
 GC-MS is increasingly used for detection of illegal
narcotics, and may eventually supplant drug-sniffing dogs
 GC-MS is also commonly used in forensic toxicology to find
drugs and/or poisons in biological specimens of suspects,
victims, or the deceased.

30. Sports anti-doping analysis
 GC-MS is the main tool used in sports anti-doping
laboratories to test athletes' urine samples for prohibited
performance-enhancing drugs, for example anabolic
steroids security

31. Chemical warfare agent detection:-
 As part of the post-September 11 drive towards increased capability in
homeland security and public health preparedness, traditional GC-MS
units with transmission quadrupole mass spectrometers, as well as
those with cylindrical ion trap (CIT-MS) and toroidal ion trap (T-ITMS)
mass spectrometers have been modified for field portability and near
real-time detection of chemical warfare agents (CWA) such as sarin,
soman, and VX
 These complex and large GC-MS systems have been modified and
configured with resistively heated low thermal mass (LTM) gas
chromatographs that reduce analysis time to less than ten percent of
the time required in traditional laboratory systems.

32. Chemical engineering
 GC-MS is used for the analysis of unknown organic
compound mixtures.
 One critical use of this technology is the use of GC-MS to
determine the composition of bio-oils processed from raw
biomass.

33. Food, beverage and perfume analysis
 Foods and beverages contain numerous aromatic
compounds, some naturally present in the raw materials
and some forming during processing.
 GC-MS is extensively used for the analysis of these
compounds which include esters, fatty acids, alcohols,
aldehydes, terpenes etc.

34. Astro chemistry
 Several GC-MS have left earth. Two were brought to Mars
by the Viking program. Venera 11 and 12 and Pioneer
Venus analyzed the atmosphere of Venus with GC-MS The
Huygens probe of the Cassini–Huygens mission landed one
GC-MS on Saturn's largest moon, Titan. The MSL Curiosity
rover's Sample Analysis at Mars (SAM) instrument contains
both a gas chromatograph and quadrupol mass
spectrometer that can be used in tandem as a GC-MS. The
material in the comet 67P/Churyumov–Gerasimenko was
analyzed by the Rosetta mission with a chiral GC-MS in
2014

35. ADVANTAGES

36.  It is very useful technique and have a very
advantage given following,
 It is important for identification of
compound.
 It can Provides sensitive response to most
analytes.
 It is important to provide information of
particular or specific class of compound.
 It can provide information of structure or
different structure of compound.
 It is having high resolution and separation
capacity.
 It is time saving technique, having a high
resolution capacity

37. Continue……..
 It is simple, rapid, reproducible technique.
 It play important role for determination of
fragmentation pattern of compounds such as protein,
peptides, amino acids and all biological or natural
compounds.

38. Disadvantages
 Higher capital cost (approx. $ >85 K vs. $15 K for GC).
 Higher maintenance (time, expertise and money).
 For optimum results requires analyst knowledgeable in
both chromatography and mass spectrometry
 Very complex process and tricky process

39. Summary
 GC/MS begins with the gas chromatograph, where the
sample is volatized. This effectively vaporizes the sample
(the gas phase) and separates its various component using
a capillary column packed with stationary (solid) phase.
The compounds are propelled by an inert carrier gas such
as argon, helium or nitrogen. As the components become
separated, they elute from the column at different times,
which is generally referred to as their retention time.

40. Continue….
 Once the components leave the GC column, they are
ionized by mass spectrometer using chemical or electron
ionization source. Ionized molecules are then accelerated
through the instrument’s mass analyzer, which quite often
is ion trap. It is here that ions are separated based on
their different mass-to-charge (m/z) ratios.
 The final step of the process involve ion detection and
analysis, with compound peaks appearing as a function of
their (m/z) ratios. Peak heights, meanwhile, are
proportional to the quantity of the corresponding
compound. A complex sample will produce several
different peaks, and the final readout will be a mass
spectrum. Using computer libraries of mass spectra for
different compounds, researchers can identify and
quantitate unknown compounds and analytes.

41. Reference:
 https://en.wikipedia.org/wiki/Gas_chromatography%E2%80%93mass_spectro
metry
 Subramani Parasuraman1*, Anish R2 , Subramani Balamurugan3 , Selvadurai
Muralidharan4 , Kalaimani Jayaraj Kumar5 and Venugopal Vijayan5
 Sloan, K. M.; Mustacich, R. V.; Eckenrode, B. A. (2001). "Development and
evaluation of a low thermal mass gas chromatograph for rapid forensic GC-
MS analyses". Field Analytical Chemistry & Technology. 5 (6): 288–301.
doi:10.1002/fact.10011
 http://cires1.colorado.edu/jimenez/CHEM-
5181/Labs/Gas_Chromatography.pdf

42. Continue…….
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43. THANK YOU