Fundamentals of

Gas Chromatography

Customer Support Centre

Shimadzu Asia Pacific Pte. Ltd.
Singapore
Gas Chromatography (1)

Chromatography

Separated, individual compounds

Mixture of compounds
Data analysis

Identification & quantification

. Chromatography is an analytical technique for separating compounds in a mixture

. Chromatography uses two phases to separate the compounds:
. Mobile phase
. Stationary phase

2
Gas Chromatography (2)

Mixture Stationary
phase

Mobile
(moving)
phase

t1 t2 t3

. In Gas Chromatography (GC), a gas is used as the mobile phase

(normally called “carrier gas”)

3
GC Samples
. Samples suitable for GC analysis:
. Thermally stabile
. Sufficiently volatile (can be vaporized at 400-4500C or below)

. Examples of GC analyses:
. Analysis of pesticides in vegetables, drinking water, wastewater
. Analysis of alcohol in cooking sauces
. Analysis of hydrocarbons in petroleum products
. Analysis of solvents in packaging materials, paint, photographic film,
drugs
. Analysis of components in food flavor and fragrances
. Analysis of triglycerides in oils

. Etc.

4
Gas Chromatograph

Instrument used for gas chromatographic

analysis is known as
Gas Chromatograph

5
Construction of a Gas Chromatograph ( GC- 2010)
Detector gas(es) flow
Carrier gas flow controller
Sample in microsyringe controller (APC, 1 per
(AFC, 1 per injector)
detector)

Detector Carrier gas

supply (e.g.
Injector
Helium, Nitrogen,
Hydrogen)

Data System

Oven
Column

= flow of carrier gas Instrumentcontrol

Data acquisition
Data processing
Data storage 6
GC Analysis
. CARRIER GAS flows into the injector, through the column, and then into the
detector
. Sample is introduced into the INJECTOR
. Sample is turned into vapor and mixed with the carrier gas
. Sample vapor is transferred into the COLUMN by the carrier gas
. Analytes travel through the column at certain rates
. mainly determined by their physical properties, the temperature of the column, and the
composition of the column
. If analytes travel through the column at different rates, separation is achieved
. the fastest moving analyte exits (elutes) the column first, followed by the remaining
analytes in corresponding order
. When an analyte elutes from the column, it enters the DETECTOR
. Electric & electronic signal is generated if analyte interacts with the detector
. The size of the signal is recorded by the DATA SYSTEM and plotted against
elapsed time to produce a plot called chromatogram

7
GC Sample Introduction Techniques
. Liquid and gas samples can be introduced to gas chromatograph.

. Most common technique: liquid injection

Sample is diluted in solvent and injected to GC Injection Port by a
syringe (volume of injection normally 1-2 ul).

. Other techniques:
. Introduction of gas sample by gas tight syringe or gas
. Valve
. Headspace
. SPME (Solid Phase Microextraction)
. Pyrolysis

8
 Headspace (HS)

Headspace

Incubation / Injection
Equilibrium Sampling
Extraction 。
SPME

Vial Extraction Injection Desorption

Injection

Fiber

10
Pyrolysis
. For analysis of samples which cannot be vaporized by
GC injector, usually polymers

. How it works:
. Sample is decomposed by heating (pyrolyzed)
. Decomposition products are analyzed by GC

11
Automated Sampling

AOC-20i for liquid injection

12
Automated Sampling

AOC- 2 0 i + s for liquid injection ( higher throughput)

13
Automated Sampling
AOC-5000:
- Liquid
- Headspace
- SPME

14
Main GC components & Their Functions

. Gas flow controllers

. Injector
. Column
. Detector
. Data system (PC w/ GCsolution software)

15
Gas Flow Controllers
. Control the supply of gas(es) . Electronic flow controllers:
to the GC . GC-2010 & GC-2014 carrier gas

. GC-2010 detector gases

. Electronic flow controller
. GC-2014 detector gases (option)
. Convenient, precise
. Manual flow controllers:
. Manual flow controller
. GC-2014 detector gases
. More economical (standard)

Detector Gas(es) Flow Controller

Carrier Gas Flow Controller
Detector
Injector

Hydrogen
Carrier

Make up
Gas

Air
Column
GC
Oven
16
Injection Port (Injector)
. Where sample is introduced to . Mixes sample vapor with carrier
GC gas
. Turns or keep sample in vapor
form

Injector
Sample

Flow Controller Detector

Carrier Gas
supply

Data System
Column
GC Oven

17
Column
. Separate mixture of compounds into individual components

Carrier Gas Column

Flow
Controller
Injection Port Detector Analyst
Gas supply
Carrier

Data System

GC Oven

18
 GC Column
Capillary Column Carrier gas

Carrier gas
Packed Column Stationary phase
Tubing
- Fused silica
- Metal

Glass or stainless
steel

Stationary phase (thin Solid support

film) material

19
 Retention Time =
time spent by a
Separation in column compound inside the
column

From
Injection To
Port Detector

Column

From To
Injection
Detector
Port

Column

From
To
Injection Detector
Port

Column
20
Detector
. Detects the compound(s) eluted . Signal is a function of time
from column . Signal is proportional to the
. Generates electric & electronic amount of the compound
signal
. Transmit signal to Data System
for recording & processing

Detector Gas(es)
Carrier Gas Detector Flow Controller
Flow Controller Injector

Hydrogen
Carrier

Make up
Gas

Air
Column
GC
Oven
21
Shimadzu GC Detectors
. FID (Flame Ionization Detector)
. Detects most organic compounds
. General detector

. TCD (Thermal Conductivity Detector)

. Detects inorganic gases and organic compounds
. General detector

. ECD (Electron Capture Detector)

. Selective detector
. For higher sensitivity detection of electrophilic compounds
. Example of analysis:
. analysis of PCBs ( polychlorinated biphenyls)
. analysis of PBBs ( polybrominated biphenyls)
. analysis of PBDEs (polybrominated diphenyl ethers)
. analysis of organochlorine pesticides (DDT, BHC, etc. )

22
Shimadzu GC Detectors
. FPD (Flame Photometric Detector)
. Selective detector
. For higher sensitivity of detection for sulfur, phosphorous, or tin containing
organic compounds
. Example of analysis:
. Analysis of organophosphorous pesticides (malathion, dichlorvos, chlorpyrifos,
etc.)
. analysis of H2S gas
. analysis of SO2 gas

. FTD 2010 (Flame Thermionic Detector)

. Selective detector
. For analysis of organic nitrogen and phosphorus containing organic
compounds
. Example of analysis:
. analysis of nitrosamines
. analysis of organophosphorous and nitrogen-containing pesticides

23
Data System
. Converts signal from detector into . Controls GC system (for the more
human-readable format advanced, software-based
. Process and analyze data systems)
. E.g.
. GCsolution Software (PC based)
. Chromatopac

Flow Controller

Injection Port
Detector Data System
Carrier Gas
supply

Column
GC Oven
24
GC Data : Chromatogram
uV(x10,000)
1.50
Chro matogram
1.25

1.00

0.75

0.50

0.25

0.00
1.0 2.0 3.0 4.0 5.0 6.0 7.0 8.0 9.0 min

. Time where peak is detected is used to identify the compound in the

sample
. Retention time: time required for a compound to travel through the
column
. Size (area or height) of peak is used to measure the amount of the
compound in the sample

25
Purpose of GC Analysis

Qualitative Analysis

Quantitative Analysis

26
Does this sample contain
pesticides???

27
Qualitative Analysis
. Retention times of compounds in
standard sample is compared to the Chromatogram of Standard Sample
retention times of peaks in unknown uV(x10,000)
sample. 1.50
Chro matogram
1.25 Stand ard Sa mple
1.00
. Under the same analysis condition, the 0.75
same compounds have the same retention 0.50

times. 0.25

0.00
1.0 2.0 3.0 4.0 5.0 6.0 7.0 8.0 9.0 min

Chromatogram of Unknown Sample

uV(x 10,000)
1.50
Chro mat ogram Unknown Sa mple
1.25

1.00

Same 0.75

Compound 0.50

0.25

0.00
1.0 2.0 3.0 4.0 5.0 6.0 7.0 8.0 9.0 min
28
How much
pesticides are
contained in this
sample???

29
Quantitative Analysis
. The peak areas or heights of compounds in standard sample (with known
concentration) are compared to that of the sample of unknown
concentration

Calibration curve

Y
(Area or height) Standard 3

Sample
Standard 2
Standard 1

X (Concentration)
30
Separation of compounds
. When analytes are introduced into
the column, the molecules
distribute between the stationary
and mobile phases
M
. The molecules in the mobile
phase are carried down the
S
column

. Those in the stationary phase are

temporarily immobile and do not
move down the column

M = mobile phase (carrier gas)

S = stationary phase 31
Separation of compounds
. The molecules in the mobile
phase are carried down the
column

M . Those in the stationary phase are

temporarily immobile and do not
move down the column
S

32
Separation of compounds
. Molecules in the mobile phase re-
enter the stationary phase when
they collide with the stationary
phase
M
. At the same time span, molecules
leave the stationary phase and
S enter the mobile phase

33
Separation of compounds
. The molecules in the mobile
phase are carried down the
column

M . The process is repeated many

many times inside the column

34
Separation of compounds
. All molecules of the same
compound travel through the
column at nearly the same rate
and appear as a band of
M molecules (called sample band)

. Compound which is less ‘soluble’

S in the stationary phase moves
faster

35
Separation of compounds
. Goal of gas chromatography :
. No overlap between adjacent sample bands as they exit the column

" Good
chromatography
Poor chromatography

Coeluting peaks

Analysis time Analysis time

Make each sample band travel at a different rate

Minimize the width of the sample band

36
Migration rates of compounds in column (1)

. Different migration rates of compounds can be achieved

if these compounds have different interaction strengths
with the stationary phase

Stronger interaction Weaker interaction

Stationary phase

37
38
Migration rates of compounds in column (2)

. Migration rate of compounds in column depend on:

. Compound chemical structure

. Stationary phase chemical structure

. Column temperature

39
Effect of compound chemical structure on
migration rate

M M

S S

GC Column GC Column
40
Effect of stationary phase on migration rate

M M

S S

GC Column 1 GC Column 2
41
Effect of column temperature on migration
rate

M M

S S
Lower T Higher T

GC Column GC Column

42
Sample Band Width

. Sample bandwidth depends on:

. Operating conditions

. Dimensions of the column

43
Basic GC parameters
. Retention time (tR)
. Retention time of unretained compound (tM)
. Retention factor (k)
. Distribution constant (K)
. Phase ratio (β)
. Separation factor (α )
. Resolution (R)
. Number of theoretical plates (N)
. Height equivalent to a theoretical plate (HETP)
. Carrier gas linear velocity (v)

44
Retention Time (tR)

. The time an analyte takes to travel through the column

.A measure of the amount of time an analyte spends in

the column

. Sum of the time spent in the stationary phase and the

mobile phase

45
Retention time of an unretained compound
(tM)

. The time an unretained compound to travel through the

column

. Unretained compound travels down the column at the

same rate as the mobile phase (carrier gas)

. Equivalent to the time a compound spends in the mobile

phase

46
Retention factor (k)
. Another measure of retention
. Ratio of the amount of time a compound spends in the
stationary and mobile phases
.A measure of retention by the stationary phase

. Previously called capacity factor, or partition factor

tR - tM
k=
tM

47
Distribution constant (K)
. Ratio of analyte concentration in the stationary phase
and mobile phase
. K is constant for a given compound, stationary phase,
and column temperature

cS
K=
cM

cS = concentration in stationary phase

cM = concentration in mobile phase

48
 Phase Ratio (β)
. The change in the phase ratio can be used to calculate the
change in a compound’s retention, provided that the same stationary
phase and column temperature (program or isothermal) are
maintained

. An increase in phase ratio results in a decrease in retention (k),

since K is constant; and vice versa

β= K=kβ

r = column radius (µ m)
df = film thickness (µ m)
49
Separation Factor (α)
. A measure of the time or distance between the maxima
of two peaks

. α = 1 means the two peaks have the same retention and

co-elute

k2 k1
α=

50
Resolution (R)
. A measure of overlap between two peaks; the higher the
resolution, the less the overlap
. Separation (α ) is only the distance between two peak maxima;
resolution takes both α and the width of the peaks into account
. Baseline resolution usually occurs at R = 1.50

tR2 – tR1 tR2 – tR1

R = 1.18 R= 2
wh1 + wh2 wb1 + wb2

wh = peak width at half peak height

wb = peak width at base

51
Number of theoretical plates (N) or Column
Efficiency
. Theoretical plates is a concept
. Theoretical plates numbers are an indirect measure of peak width
for a peak at a specific retention time
. Columns with high N are considered to be more efficient than those
with lower N
. A column with a high N will have a narrower peak at a given
retention time
. Column efficiency is a function of:
. Column dimensions
. Type of carrier gas and its average linear velocity
. Compound and its retention

tR tR
N = 5.545 N = 16
wh wb
52
Height equivalent to a theoretical plate
(HETP or H)

. Another measure of column efficiency

. Small plate heights indicate higher efficiency

L
H=
N

L = column length (mm)

N = theoretical plates number

53
Carrier Gas Linear Velocity (v)
. Affects the chromatographic
resolution (i.e. separation)

. For each gas there is a linear Van Deemter Curve

velocity where optimum
separation can be achieved
(minimum HETP)

Linear Velocity (cm/s)

54
55
56
 uV
7.5 Chromatogra m
(x10,000)

Analyses of Vegetable Oils

5.0

2.5

uV
7.5
(x10,000) 0.0

uV(x10,000) 12.5 15.0 17.5 20.0 min

7.5 Chromatogra m
5.0

Soya Bean Oil

2.5
5.0

0.0
2.5
12.5 15.0 17.5 20.0 min

Canola Oil 0.0

12.5 15.0 17.5 20.0 min

1.5 uV (x100,000)
Chromatogram

1.0 Sunflower Oil

0.5
Column: Rtx-65TG (30m x 0.25 mm x 0.1um)
Carrier gas: Helium
0.0
Linear velocity: 40cm/s (constant linear velocity)
12.5 15.0 17.5 20.0 min
Injector temp: 3650C

Olive Oil Injection method: Split (Split Ratio = 40)

Column temp: 500C(0.5min)>500 C/min> 3500 C(0min)> 10C/min> 3650 C(5min)
FID temp: 3650C
57
 Aromatic Components of Tea
by Headspace-GC

Analysis of Refined Green Tea Analysis of Coarse Green Tea Analysis of Black Tea

58
 Analysis of Spearmint Oil

Analysis of Peppermint Oil

59
60
Analyses of Kerosene and Light Oil

Kerosene Light Oil

61
62
63
Headspace-GC Analysis on Phenols in Electrical Wiring
Surfaces

64
65
66
 Chromatograms of two different swimming pools Chromatograms of a swimming pool
consisting of sea water
67
Determination of Trihalogenated Methanes by ECD Detector
68
Analysis of Anti-Epylepsy Drug

69
Analysis of Cold Medicine

70
 Analysis of Solvents by Headspace-GC

Instrument: GC equipped with Headspace Split Ratio: 3

Sampler Column: Rtx-624, 30m x 0.53mm x 3um

Injector temp:1800C Carrier gas: Helium

Pressure: 24.9 kPa FID temp: 2200C

Column flow: 4.89 ml/min Injection volume: 1 ml

Column temp:380C(2min)>10C/min>4000C(6min)>200C/min>2000C(7min)

uV( x 1 0 ,000)

T oluene
Ben zene

oethylene
Chromatogram
Hexane

T
richlo
2.0
Di chloromet hane

1.5

1.0
Dichlor omethane

Ch loroform

ol
Isopropa nol

0.5
Aceton e

n-Butan

0.0

5.0 7.5 10.0 12.5 15.0 min

71