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GC-MS
Gas Chromatography-Mass Spectrometry
An Hybrid technique which couples the powerful
separation potential of gas chromatography with the
specific characterization ability of mass spectroscopy.

Overview

GC History
What is GC
Key Components
Separation Process
GC Theory
Carrier Gas
Injectors
Columns

GC History
Development of GC (1941) by Martin and Synge
Theory of Capillary GC (1957) by Golay
Capillary GC Instruments (1977)
Fused Silica Capillary Columns (1979)

What is GC?
GC is a Separation Technique
Sample is usually a complex mixture we
require to separate into constituent
components.
Why: usually to quantify some or all
components e.g. Pharmaceuticals,
Environmental pollutants, etc
Occasionally as a qualitative tool

What is the sample?

Usually a mixture of several components
Sample usually introduced as a liquid
Components of interest (analytes) usually in
low concentrations (<1% to ppb levels)
Samples dissolved in volatile solvent

Comaparison: GC & HPLC

HPLC

GC

non-volatile samples

volatile & thermally stable

thermally unstable compounds

rapid analysis

macromolecules

good resolution

inorganic and ionic samples

easily interfaced to Mass Spec
More complex interface to Mass
Spec .

Key components of GC
Hardware to introduce the sample
Technique to separate the sample into components
Hardware to detect the individual components.
Data Processing to process this information.

Basic Block Daigram!

Separation Process
Sample is introduced into system via hot, vaporising
injector.
Typically 1ul injected
Flow of Carrier Gas moves vaporised sample (i.e. gas)
onto column
Column is coated with wax type material with varying
affinity for components of interest
Components are separated in the column based on this
affinity.
Individual analytes are detected as they emerge from the
end of the column through the Detector.

Example Chromatogram (Capillary)

c:\star\examples\level4.run

File: c:\star\examples\level4.run
Channel: A =TCD Results
Last recalc: 25/07/1993 18:35

3.210

mVolts

2.853
1.474

4.463

0.754

500

5.320

2.038

0.541

Detector
Response

1.113

750

5.562

250

Inject
Point

-87

5
Minutes

Time

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18.

Analysis of Halogenated Pesticides

-HCH
-HCH
-HCH
Heptachlor
-HCH
Aldrin
Heptachlor epoxide
Endosulfan I
4,4-DDE
Dieldrin
Endrin
4,4-DDD
Endosulfan II
4,4-DDT
Endrin aldehyde
Endosulfan sulfate
Methoxychlor
Endrin ketone

10

11

6
8

12
13
9

14

1
5

15

16

2ppb in Water

17 18

Chromatogram

GC Step by Step
Carrier Gas
Injector
Column
Capillary
Stationary Phase

Detectors
Mass Spectrometer

Carrier Gas

Inert

Helium

Choice dictated by detector, cost, availability

Pressure regulated for constant inlet pressure

Flow controlled for constant flow rate

Chromatographic grade gases (high purity)

Column Types
Capillary Columns
Length: 10m to 100m
Diameter: 180um,
250um, 320um &
530um I.d

Packed Columns
Length: <2m
Diameter: 1/8 & OD

Typical column flow rates

Capillary Column Flow
250 um 1 ml/min
320 um 1.5 ml/min
530um up to 2.0 ml/min

Purpose of Injection
Deposit the sample into the column in the narrowest band
possible
The shorter the band at the beginning of the
chromatographic process - tall narrow peaks
Gives maximum resolution and sensitivity
Therefore type of injection method and operating
conditions is critical in obtaining precise and accurate
results

Splitless injector Design

Graphite/Viton Seal
Reduced Sample Contact

UniqueDual Split Vent design

Improved Precision
Large Internal
Volume
Minimum
Solvent Tailing

More Efficient Sweep

Shortened Capillary Guide

Minimal Cold Spots
Minimal Upswept Volume

Cross Section of PTV Injector

Modern
Temperature
Programmable
Injector
(Varian 1079)
Programmable
Temperature
Vapourising
Injector

Split & Splitless Injection

Most common method of Injection into Capillary
Columns
Most commonly misunderstood also!
Same injector hardware is used for both
techniques
Electronically controlled Solenoid changes Gas
Flow to determine Injector function.

Split Injection
Mechanism by which a portion of the injected solution is discarded.
Only a small portion (1/1000 - 1/20) of sample goes through the column
Used for concentrated samples (>0.1%)
Can be performed isothermally
Fast injection speed
Injector and septa contamination not usually noticed

Splitless Injection
Most of the sample goes through the column (85-100%)
Used for dilute samples (<0.1%)
Injection speed slow
Should not be performed isothermally
Solvent focusing is important
Controlled by solenoid valve
Requires careful optimisation

On Column Injection
All of the sample is transferred to the column
Needle is inserted directly into column or into insert directly
above column
o

Trace analysis

Thermally labile compounds e.g Pesticides, Drugs

Wide boiling point range

High molecular weight

Large Volume Injection

To enhance sensitivity in Envoirnmental applications.
Uses 100L syringe: Inject up to 70 l
Very slow injection with injector temperature a few degrees below
solvent boiling point, split open, flow at about 150 mls/ min
Solvent vents out of split vent, thus concentrating the analytes
Close split
Fast temperature ramp to top column temperature +20C
Column programming as per sample requirements

Columns

Material of Construction
Metal (1957)
Glass (1959)
Fused Silica (1979)
Aluminium Clad (1984)
Inert Metal (1990)

Capillary Column Characteristics

Length (10M - 50M)
Internal Diameter (0.1mm - 0.53mm)
Liquid Stationary Phase
Film Thickness (0.1um - 5um)
Polarity (Non-polar - Polar)

Stationary Phases
Choice of phase determines selectivity

Hundred of phases available

Many phases give same separation

Same phase may have multiple brand names

Stationary phase selection for capillary columns much simpler

Like dissolves like

Use polar phases for polar components

Use non-polar phases for non-polar components

Column Bleed

Bleed increases with film thickness

Polar columns have higher bleed

Bleed is excessive when column is damaged or degraded

Avoid strong acids or bases

Adhere to manufacturers recommended temperature limits

Avoid leaks

Choosing a Column
Internal Diameter
Film Thickness
Length
Phase

Internal Diameter, Smaller IDs

Good resolution of early eluting compounds
Longer analysis times
Limited dynamic range

ID Effects - larger IDs

Have less resolution of early eluting compounds
Shorter analysis times
Sufficient resolution for complex mixtures
Greater dynamic range

Film Thickness

Amount of stationary phase coating

Affects retention and capacity

Thicker films increase retention and capacity

Thin films are useful for high boilers

Standard capillary columns typically 0.25m

0.53mm ID (Megabore) typically 1.0 - 1.5m

Column Capacity

The maximum amount that can be injected without significant peak

distortion

Column capacity increases with :-

film thickness

temperature

internal diameter

stationary phase selectivity

If exceeded, results in :

peak broadening

asymmetry

leading

Length effects - isothermal analysis

Retention more dependant on length
Doubling column length doubles analysis times
Resolution a function of Square Root of Length
Gain 41% in resolution
Is it worth the extra time and expense?-

Length effects - programmed analysis

Retention more dependant on temperature
Marginally increases analysis times
Run conditions should be optimised

Summary - Effect of ID, Film

Thickness, and Length

ID
Film Thickness
Length
Choice based on
Gain in resolution is
capacity and resolution Thick film for low
not double
Use 0.25mm for MSDs
boilers
Isothermal: tR L
Use 0.32mm for
Thin film for high
Programmed: tR is
split/ splitless & DI
boilers
more dependent on
Use 0.53mm for DI &
Thicker films for larger
temperature
purge & trap
ID's

Detectors

Overview
Basic Mass Spectrometry Theory
Types of Ionisation
- Electronic Ionisation
- Chemical Ionisation
Interpretation of Mass Spectra
Ion Trap Theory
Components of the Ion Trap

Ion Trap Mass Spectrometry

Basic Mass Spec.Theory

Mass Spec. is a Microanalytical Technique used to obtain information
regarding structure and Molecular weight of an analyte
Destructive method ie sample consumed during analysis
In all cases some form of energy is transferred to analyte to cause
ionisation
In principle each Mass Spectrum is unique and can be used as a
fingerprint to characterise the sample
GC/MS is a combination technique that combines the separation
ability of the GC with the Detection qualities of Mass Spec.

Basic GCMS Theory(1)

Sample injected onto column via injector
GC then separates sample molecules
Effluent from GC passes through transfer line into
the Ion Trap/Ion source
Molecules then undergo electron /chemical
ionisation
Ions are then analysed according to their mass to
charge ratio
Ions are detected by electron multiplier which
produces a signal proportional to ions detected

Basic GCMS Theory(2)

Electron multiplier passes the ion current
signal to system electronics
Signal is amplified
Result is digitised
Results can be further processed and
displayed

Types of Ionisation
Electron impact ionisation
Chemical Ionisation

Definition of Terms
Molecular The ion obtained by the loss of an electron from
ion
the molecule
The most intense peak in the MS, assigned 100%
Base peak
intensity
M+

Symbol often given to the molecular ion

Radical
cation

+ve charged species with an odd number of

electrons

Lighter cations formed by the decomposition of

Fragment
the molecular ion.
ions
These often correspond to stable carbcations.

Electron Ionisation(1)
Sample of interest vaporised into mass spec
Energy sufficient for Ionisation and Fragmentation
of analyte molecules is acquired by interaction
with electrons from a hot Filament
70 eV is commonly used
Source of electrons is a thin Rhenium wire heated
electrically to a temp where it emits free electrons

Electron Ionisation

Electron Ionisation
The physics behind mass spectrometry is that a charged particle
passing through a magnetic field is deflected along a circular path on a
radius that is proportional to the mass to charge ratio, m/e.
In an electron impact mass spectrometer, a high energy beam of
electrons is used to displace an electron from the organic molecule to
form a radical cation known as the molecular ion. If the molecular
ion is too unstable then it can fragment to give other smaller ions.
The collection of ions is then focused into a beam and accelerated into
the magnetic field and deflected along circular paths according to the
masses of the ions. By adjusting the magnetic field, the ions can be
focused on the detector and recorded.

Chemical ionisation
Used to confirm molecular weight
Known as a soft ionisation technique
Differs from EI in that molecules are ionised by interaction
or collision with ions of a reagent gas rather that with
electrons
Common reagent gases used are Methane , Isobutane and
Ammonia
Reagent gas is pumped directly into ionisation chamber
and electrons from Filament ionise the reagent gas

Chemical Ionisation(2)
First - electron ionization of CH4:
CH4 + e- CH4+ + 2e Fragmentation forms CH3+, CH2+, CH+

Second - ion-molecule reactions create

stable reagent ions:
CH4+ + CH4 CH3 + CH5+
CH3+ + CH4 H2 + C2H5+
CH5+ and C2H5+ are the dominant methane CI
reagent ions

Chemical Ionisation(3)
Form Pseudomolecular Ions (M+1)
CH5+ + M CH4 + MH+
M+1 Ions Can Fragment Further to Produce a Complex CI
Mass Spectrum

Form Adduct Ions

C2H5+ + M [M + C2H5]+
C3H5+ + M [M + C3H5]+

M+29 Adduct
M+41 Adduct

Molecular Ion by Charge Transfer

CH4+ + M M+ + CH4

Hydride Abstraction (M-1)

C3H5+ + M C3H6 + [M-H]+
Common for saturated hydrocarbons

EI vs CI for Cocaine analysis

EI Spectrum of Cocaine
Extensive Fragmentation
Molecular Ion is Weak at m/z 303

Methane CI of Cocaine
Pseudomolecular Ion and Fragment Ions

Proton Affinity
Proton Affinity Governs CI Susceptibility
The higher the affinity the more tightly
bound the proton is to the parent species
The greater the difference in proton
affinities between the analyte and reagent
gas the more energy transferred to the
protonated molecule more fragmentation

Interpretation of Mass Spectra(1)

Intepretation of Mass Spectra(2)

The MS of a typical hydrocarbon, n-decane is shown above.
The molecular ion is seen as a small peak at m/z = 142.
Notice the series ions detected that correspond to fragments that
differ by 14 mass units, formed by the cleave of bonds at
successive -CH2- units

Interpretation of Mass Spectra(3)

Interpretation of Mass Spectra(4)

The MS of benzyl alcohol is shown above.
The molecular ion is seen at m/z = 108.
Fragmentation via loss of 17 (-OH) gives a common fragment
seen for alkyl benzenes at m/z = 91.
Loss of 31 (-CH2OH) from the molecular ion gives 77
corresponding to the phenyl cation.
Note the small peaks at 109 and 110 which correspond to the
presence of small amounts of 13C in the sample (which has
about 1% natural abundance).

Determining Isotope Patterns in Mass Spectra

Mass spectrometers are capable of separating and detecting
individual ions even those that only differ by a single atomic mass
unit.
As a result molecules containing different isotopes can be
distinguished.
This is most apparent when atoms such as bromine or chlorine are
present (79Br : 81Br, intensity 1:1 and 35Cl : 37Cl, intensity 3:1)
where peaks at "M" and "M+2" are obtained.
The intensity ratios in the isotope patterns are due to the natural
abundance of the isotopes.
"M+1" peaks are seen due the the presence of 13C in the sample.

Isotope Patterns 2,Chloropropane

Examples of haloalkanes with characteristic isotope patterns.

The first MS is of 2-chloropropane.
Note the isotope pattern at 78 and 80 that represent the M
and M+2 in a 3:1 ratio.
Loss of 35Cl from 78 or 37Cl from 80 gives the base peak a
m/z = 43, corresponding to the secondary propyl cation.
Note that the peaks at m/z = 63 and 65 still contain Cl and
therefore also show the 3:1 isotope pattern.

1,Bromopropane

 The second MS is of 1-bromopropane.

Note the isotope pattern at 122 and 124 that
represent the M amd M+2 in a 1:1 ratio.
Loss of 79Br from 122 or 81Br from 124 gives
the base peak a m/z = 43, corresponding to the
propyl cation.
Note that other peaks, such as those at m/z = 107
and 109 still contain Br and therefore also show
the 1:1 isotope pattern.

ION TRAP THEORY

 Ionize analytes within the ion trap

Use energetic electrons to ionize
Store ions and continue to ionize until the optimum trap
capacity is reached
Optimum ion time calculated by software
Increase the voltage on the Ring Electrode of the ion
trap to scan ions out in order from low to high mass
This voltage-time relationship called the EI/MS
Scan Function
Store the mass-intensity information as a mass
spectrum

Electron Ionization Happens Inside the Ion Trap

Filament
Gate

Ring electrode

Trapped
Ions

CARRIER
GAS

Mass Spectrum of Toluene

Mass Spectrum of Caffeine

Mass Spectrum of Glycerin

Mass Spectrum of Cholesterol

Mass Spectrum of Aspirin