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Philip Marriott
School of Chemistry, Monash University
Discovery Outstanding Researcher Award: Australian Research Council
4th MDC Workshop: TORONTO
Multidimensional Gas Chromatography: Past, Present &
Future
Australian Centre for Research on Separation Science
Thanks to … • My Monash ACROSS Group; all our
collaborators across many continents • Agilent Technologies • Sigma-Aldrich Supelco • Defence Science Technology Organisation • Other research sponsors:
ARC; Linkage Project supporters; Industry supporters
Has GC×GC spurred new developments ~ renaissance ~ in MDGC?
If GC×GC represented a new and exciting development in GC, and showed how enhanced separations benefits analysis, how should GC companies respond? The easiest is to use a method that is compatible with their software, but gives improved separation = MDGC
Landmarks in GC×GC John Phillips’ pioneering work LECO agrees to commercialise GC×GC-TOFMS ZOEX – GC Image GC×GC-qMS Cryogenic Modulation Diaphragm Modulation Flow Modulation The GC×GC Symposium series ~ now 10th …
.. And every one of you – for believing
Landmarks in GC×GC
Retention indices Retention prediction / modelling Chemometrics Novel interconversion processes Application of new detection systems Optimisation Modulation ratio and sampling theories ….
Definitions of MDGC / GC×GC
.. Blumberg and Klee: re-defined MDC as “n-dimensional analysis is one that generates n-dimensional displacement information”. (2010)
Marriott: MDGC – “the process of selecting a (limited) region or zone of eluted compounds from the end of one GC column, subjecting the zone to a further GC displacement”. (2005)
Giddings: two principal kinds of multidimensional systems (1) continuous two-dimensional (2D) operation (simultaneous zone displacement), and (2) coupled column assemblies (sequential displacement) [1984; 1987].
None of these definitions implicitly state nor require that the separation of compounds will be improved by MDGC.
… the guiding principle (3) is that a 2D separation can be called comprehensive if: 1. Every part of the sample is subjected to two different (independent) separations; 2. Equal percentages (either 100% or lower) of all sample components pass through both columns and eventually reach the detector; and 3. The separation (resolution) obtained in the first dimension is essentially maintained.
PJ Marriott; ZY Wu; P Schoenmakers. Nomenclature and Conventions in Comprehensive Multidimensional Chromatography – An Update. LC-GC Europe, 25 (2012) 266-275
And that was fine, until … along came flow modulated GC×GC – thanks to John Seeley. Now we had to change the concepts of how we implemented the procedure. Changes to the linear carrier flow rate from 1D to 2D now permitted the fast elution required on 2D, and to achieve adequate retention and resolution, we now have to use a longer – and often wider bore column, than in the more common thermal modulation interfaces.
The need for NOMENCLATURE Consistency amongst researchers Universal understanding of the meaning of symbols Clarity in definitions and terms
Use of 1X and 2X for symbols, vs 1X, 2X , or X1, X2 e.g. 1D and 2D for first and second dimensions, not 1D and 2D, or D1 and D2 (2D means two-dimensional) 1tR; 2tR, not 1tR; 2tR nor tR1, tR2 We can now define any symbol correctly: 2I; 2df; 2wb
… lingering confusion or inconsistency over the use of the words “comprehensive” and “orthogonal,” and of the multiplex sign (×) as a short notation for comprehensive two-dimensional (2D) separations.
Do NOT use ‘comprehensive GC GC’! Do NOT use GCxGC, GCXGC, GC X GC Do NOT use ‘normal phase’ column set And especially do NOT use reverse-phase column set
Are we at a stage where new-comers into GC×GC (and MDGC) do not have an adequate understanding of the technology, fundamentals and processes of these methods? Or maybe where some have little experience with classical capillary GC?
Is doing a GC×GC analysis because it is ‘new’ justified if the analysis does not require GC×GC, or .. If the method is not properly (or poorly) implemented?
High or low frequency Modulation / Sampling in MDGC The difference between classical MDGC and GC×GC is the sampling frequency and modes. Ú MDGC conventionally samples from 1D in a few discrete steps; normally a long 2D column is used. With or without cooling the oven. Ú GC×GC requires faster, and regular sampling, normally at a faster rate that 1wb. So this could be considered a ‘continuum’ of technology from MDGC thru GC×GC.
6 8 10 12 14 16 18 20 22 24 26 28 30 32 34 36 38 40
Res
pons
e (x
105 )
Retention time (min)
zone 1 1.0
3.0
4.0
2.0
0.0
GC – qMS (non-Modulated)
21 22 23
19 20 21
20 21 22
M’ N’
10’
10
M N
2.5
2.0
1.5
3.0
0.0
Retention time (min)
1.0 2.0 3.0 4.0
5.0
0.0 1.0 2.0 3.0 4.0 5.0
FID
(x 1
05)
qMS 1 min modln
qMS 1.5 min modln
Zone 10 = 1.5 min Zones M & N = 1.0
19
MDGC–qMS (Modulated) 1.0 / 1.5 min
8 10 12 14 16 188 10 12 14 16 18
MS
x 10
7
0.5
1.0
2.0
0.0
8 10 12 14 16 18
FID
x 1
06
1.0
3.0
6.0
8 10 12 14 16 18
1.0
3.0
6.0
1.5
0.5
1.0
2.0
0.0
1.5
Retention time (min)
1 2 3
4* 5
6
1(a)
1(b) 2
3
4*
5
6 1 2
3
4
5
6 2tR
1(a): 0.527 1(b): 0.508 2: 0.626 3: 0.774 4: 0.684 5: 1.779 6: 1.400
2tR 1: 0.519 2: 0.582 3: 0.706 4: 0.614 5: 1.542 6: 1.304
Retention time (min)
(B) 1 2
3 4*
5
6
FID
x 1
06 M
S x
107
Possible to get split peaks; Peaks are SHARP and TALL
1.0 Min 1.5 Min
2 DL M
S 2 D
S FID
1. 2. 3.
11.
. . . . . . .
Multidimensional sampling strategy
Sample 0.2 min, 2 min apart = 10 analyses to cover a full sample
& complete each heartcut in 2 min
Single (12 s) heart-cut; with (¾¾) & without (·····) cryotrapping. Without: can hardly recognize oxygenates: With: easy to recognize oxygenates
Another approach - Single heart-cut, cryotrap, reduce oven temp., release...
• Very high peak capacity (~600) • Would need ~150 runs to cover the whole 1D GC
Cut a 0.2 min section from the 1D GC run …
Single (12 s) heart-cut (see previous slide) PROCESS: (1) cryotrap; (2) cool the oven; (3) then elute on the long 2D column
Modulator, M Inj Det
1st Dimension; 1D
2nd Dimension;
2D (very short)
1+2 1+2
1 2
A B
C
GCxGC ~ a tutorial
Peaks are sliced (modulated); each slice is introduced into a short 2D column to provide further separation. Data presented as a 2D plot.
1D
2D
1D
2D
Movies of Modulation Operation
MODULATION DRIVE UNIT
CRYOTRAP MOVEMENT
REAL-TIME DATA ACQUISITION – as seen on chromatography data system …
How do we calculate the exact first dimension retention time? .. and is it important? Some commentators say that GC×GC loses information on the 1D column – Due to the modulation process, and loss of resolution. But what if we can re-construct the 1tR value? This will return to the 1D all of the primary information that we need from the GC process.
Reconstruct based on the modulated peak distribution, and the appropriate peak model
Modulation Phase (ΦM) in GC×GC
180o out-of-phase
Start time + 0.00
Start time + 0.02
Start time + 0.04
Start time + 0.06
In-phase
In-phase
3.10 min
3.08 min
3.04 min
3.00 min
3.02 min
3.06 min
Symmetric REAL 2D PEAKS
Adjust time of starting modulation
allows phase of modulation
to be readily altered In-phase
180o out-of-phase
Same phase
Modulation Phase PM = 6 s; 0.1min)
16.9 17.0 17.1 17.2 17.3 17.4 0
400
800
1200
16.9 17.0 17.1 17.2 17.3 17.4 0
200
600
1000
16.9 17.0 17.1 17.2 17.3 17.4 0
400
800
1200
Resp
onse
(pA
)
16.9 17.0 17.1 17.2 17.3 17.4 0
500
1000
1500
16.9 17.0 17.1 17.2 17.3 17.4 0
1000
2000
Retention Time (minutes)
A
B
C
D
E
Modulation period incremented A-E
2s
9.9s
6s
4s
3s
Start time 16.03min
A range of phases is observed
Modulation Period (PM) in GC×GC Effect of changing
modulation period on
pulse presentation - This affects
apparent resolution in
2D
Retention Time (minutes)
Resp
onse
(pA
)
24.5 24.6 24.7 24.8 24.9 25.0
100
200
16
16
16 16
16 16
17
17 17
17 17 17
24.5 24.6 24.7 24.8 24.9 25.0
100
200
17
17 17
17 17
16
16
16
16 16
24.5 24.6 24.7 24.8 24.9 25.0
100
200
300 16
16 16
17
17
17
24.5 24.6 24.7 24.8 24.9 25.0
100
200
300
17
17
16
16
16 17
?
How do we calculate the second dimension retention index in GC×GC?
FID1
Injector
1D columnSolGel Wax™
(30m x 0.25mm x 0.25mm)
first - 2D columnBPX5
(0.95m x 0.1mm x 0.1mm)
second - 2D columnBP10
(0.95m x 0.1mm x 0.1mm)
FID2
deactivated FS0.2m x 0.25mm
3 way splitter
longitudinally modulatedcryogenic system FID1
Injector
1D columnSolGel Wax™
(30m x 0.25mm x 0.25mm)
first - 2D columnBPX5
(0.95m x 0.1mm x 0.1mm)
second - 2D columnBP10
(0.95m x 0.1mm x 0.1mm)
FID2
deactivated FS0.2m x 0.25mm
3 way splitter
longitudinally modulatedcryogenic system
1I
2I1
2I2
(van den Dool) temp prog
(Kováts) isothermal
Slide 28
0 10 20 30 40 50 60
1
2
3
4
2 D re
tent
ion
time
[s] (
BPX5
col
umn)
1D retention time [min] (SolGel Wax™column)
C9
C10
C11
6
7
8
9
10
11
12
13 14
15
16
17 18
19 20
22
C 9-C
10
C 9-C
11
C 9-C
11
C 9-C
12
C 9-C
13
C 10-
C 14
C 10-
C 15
C 10-
C 16
C 11-
C 17
C 11-
C 18
C 12-
C 19
C 12-
C 20
C 13-
C 20
C 14-
C 20
C 15-
C 20
C12
C13
C14
C15 C16
C17
C18
C19
C20
C13
tr (z+1)
tr x
tr (z)
Make successive SPME injections of alkanes…
n-alcohols t=0 injection
Slide 29 0 10 20 30 40 50 60
0
1
2
3
4
5
C9
C10
C11 C12C13
C14C15 C16 C17 C18
C19
C20
1
2
3
8*
10
4
5a
5b
6
79 11
12
13 1615
18
20
14
19
17a
17b
21a
21b
22b22a
23
24 25
First retention index dimension
Sec
ond
rete
ntio
n in
dex
dim
ensi
on
A1
pA
2060
100140
A2
600800
1000 1200
1400 1600 1800 2000 2200
0 10 20 30 40 50 600
1
2
3
4
5
C9
C10
C11 C12C13
C14C15 C16 C17 C18
C19
C20
1
2
3
8*
10
4
5a
5b
6
79 11
12
13 1615
18
20
14
19
17a
17b
21a
21b
22b22a
23
24 25
First retention index dimension
Sec
ond
rete
ntio
n in
dex
dim
ensi
on
A1
pA
2060
100140pA
2060
100140
A2
600800
1000 1200
1400 1600 1800 2000 2200
?
?
?
Modulated alkanes
t =0 injection
2 D p
olar
1D nonpolar
How do we calculate the second dimension retention index in MDGC?
We have to get alkanes into the 2D column, and analyse them under identical conditions as the analyte. A number of ways are possible --- see later
Detection in GC×GC Apply classical GC detectors to the ‘fast’ peak profiles in GC×GC Ø FID Ø SCD Ø NCD Ø ECD Ø NPD Ø AED Ø FPD
Of prime concern: Is the chemistry and physical process of the detection transducer compatible with the fast data acquisition and peak flux needed for a GC×GC detector?
Often the answer is no ~ or at least, not quite.. ~ tailing; broadening; less than anticipated sensitivity
5.00 10.00 15.00 20.00 25.00 30.00 35.00 40.00 45.00 50.00
5.00 4.00 3.00 2.00 1.00 0.00
(B)
2 tR (s
)
1st Dimension Retention time (min)
Res
pons
e (p
A) (A)
1st Dimension Retention time (min)
2 tR (s
)
5.00 10.00 15.00 20.00 25.00 30.00 35.00 40.00 45.00
5.00 4.00 3.00 2.00 1.00 0.00
(D)
50.00
Res
pons
e (H
z)
(C)
Dual Detection NPD (A) & (B) and ECD (C) & (D)
C)
5.00 10.00 15.00 20.00 25.00 30.00
3.00
2.00
1.00
0.00
1
2
3 4
5
6
7
8 9 10
11
15.00 20.00 25.00 30.00
B)
5.00 10.00 15.00 20.00 25.00 30.00
3.50
2.50
1.50
0.50
1D Retention time (min)
2 D R
eten
tion
time
(s)
10
1
2
3 4
5
6 7
8 9
D)
11
5.00 10.00 15.00 20.00 25.00 30.00
4.00
3.00
2.00
1.00
7
11
7
11
GC×GC-NPD matrix
matrix + iprodione
matrix + all
22.5 25.0 27.5 30.0 32.5 35.0 37.5
4
3
2
1
0
m in
s
FPD/S - mode
22.5 25.0 27.5 30.0 32.5 35.0 37.5
5
4
3
2
1
2 tR
sec
FPD/P - mode
Diazinon
Methyl Chlorpyrifos
Methyl Pirimphos
Malathion
Ethyl parathion
Chlorfenvinphos
Fenthion ethyl
Methidathion
Bromophos ethyl
Carbophenothion
min
Good shape ~ same as FID
Broad, Tailing
Mass Spectrometry with GC×GC Why do we want / need MS?
Argument: If a peak’s position is equivalent to identity – and this is so much more certain in 2D space – then do we still need MS?
Implies need for / use of authentic standards; (and if only target analysis is wanted, for known compounds, maybe MS is less important) Need to give a name / tentative name to a spot … and the skeptic is likely to say – OK so you have lots of resolved peaks – what are they???
Mass Spectrometry with GC×GC
Slow scanning qMS – Frysinger Faster scanning qMS – Marriott, Shellie, Song, ~ reduced mass scanning range Fast acquisition TOFMS – Marriott (loan from LECO) Isotope ratio MS - Brenna Accurate mass MS – usually TOFMS systems - Patterson
Quantification vs qualification of components Use of GC×GC-FID integrated with GC×GC-MS
Mass Spectrometry with GC×GC
Some history .. Meeting in Park City, Utah ISCC (Milton Lee Symposium) LECO undertook commercialisation of fast TOFMS, and Jan Beens, Rene Vreuls and others (John Dimandja?) tried to convince LECO (Rick Parry) to invest in GC×GC, since ALL GC×GC users NEEDED fast MS. A focus on 1D GCMS means less of a demand for TOFMS. LECO obviously agreed!
Modulation & Modulators in GC×GC
See the presentation of Tadeusz Gorecki for more information ..
The Sweeper – originally we were told that this was a system that would end the need to innovate different modulation systems Cryogenic systems have proved to have longevity, established the present user-base, and much novel work has been performed using these. Diaphragm modulators Flow modulators
Modulation Ratio MR in GC×GC Mod Number nM vs Mod Ratio MR
“nM : Number of modulations per primary dimension peak”
Schure, et al. suggested ~ 3 – 4 modulations per peak
How do we decide nM?? It varies across peaks in the chromatogram; It varies with phase; It varies with amount injected; It varies with how small a peak we want to include in the “Number of Modulated peaks”!!!!!!!!
So define modulation ratio MR as the ratio of the
MR = (4 x 1wb) / PM Or more generally
MR = (4 / 2.54 1wh) / PM
25.6 25.7 25.8 25.9 26.0 26.1 26.2 26.3
20
40
60
80
100
Resp
onse
(pA)
Retention time (min)
PM = 7s
25.6 25.7 25.8 25.9 26.0 26.1 26.2 26.3
20
40
60
80
Resp
onse
(pA)
Retention time (min)
PM = 4s
25.6 25.7 25.8 25.9 26.0 26.1 26.2 26.3
20
40
60
80
100
Resp
onse
(pA)
Retention time (min)
PM = 6s
25.6 25.7 25.8 25.9 26.0 26.1 26.2 26.3
20
40
60
80
100
Resp
onse
(pA)
Retention time (min)
PM = 5s
25.6 25.7 25.8 25.9 26.0 26.1 26.2 26.3
10
20
30
40
50
60Re
spon
se (p
A)
Retention time (min)
PM = 3s
Symmetric REAL 2D PEAKS Alter PM
1wb ~ 18 s
MR ~ 6.0 MR ~ 4.5
MR ~ 3.7 MR ~ 3.0
MR ~ 2.5
Use Modulation Ratio to .. MR = (4 / 2.54 1wh) / PM
Approximately tells us (how many ‘large’) modulated peaks we will obtain. IF we require a MR of 3 …. If 1wh = 6 s, then PM ~ 1.6 s; So modulate at ~ 1.5 – 2 s What does this require of the METHOD? If we want no wraparound, then the desired PM above determines the max 2tR possible. This decides the column ID, L, 2df, etc required for the analysis. Note that all these parameters will alter 2tR, (or 2k I we know 2tM).
The Future … MDGC & GC×GC .. we will continue to investigate more integrated methods of analysis, incorporating GC×GC and MDGC, or using a combination of the two approaches. MDGC will see increasingly sophisticated implementation methods, improved prediction of separations, more chemometrics, and GC×GC will see more standard methods / protocols. More modulation methods?? Better detectors, more hyphenation approaches ….. All areas will benefits for MANY MANY MORE applications!
B Mitrevski; PJ Marriott. A novel hybrid comprehensive two-dimensional – multi-dimensional gas chromatography method for precise, high resolution characterisation of multicomponent samples. Anal Chem 84 (2012) 4837−4843.
PJ Marriott; S-T Chin; B Maikhunthod; H-G Schmarr; S Bieri. Multidimensional Gas Chromatography. TrAC, 34 (2012) 1-21.
S-T Chin; GT Eyres; PJ Marriott. System Design for Integrated Comprehensive and Multidimensional Gas Chromatography with Mass Spectrometry and Olfactometry. Anal. Chem. 84 (21) (2012) 9154–9162
Some of our recent Publications
M
VII 1DGC / MDGC with loop H/C & DS
DS
L
INJ DET 2 CT/SP DET 1
V Alternate GCxGC & MDGC
DS
INJ DET 1 DET 2
VI Simultaneous GCxGC / loop H/C
INJ DET 1 DET 2
VIII Simultaneous GCxGC/1DGC & MDGC
DS
INJ DET 1 SP DET 2 EPC EPC
EPC
M
M M
M
L
2
1
2
1
2
1
IX Dual parallel MDGC
XI Selective heart-cutting for semi-prep XII Multiple/repeated H/C screening
DS
INJ DET 1 DET 2 EPC
M
2
1
X
DS
INJ DET 1 DET 2 EPC
M
Alternate GCxGC & MDGC with CT
2
1
DS
INJ DET 1 EPC trapping capillary large bore
2
1
DS
INJ MS EPC
M
2
1
DET 2
Development of GC×GC/MDGC/FID/O/MS System sniff port
Deans switch
column unions
detectors
LMCS interface
.. various columns
cryotraps looped direct
INJ 2
MS
Selectable 1D or 2D GC system coupled to 3 simultaneous detections (MS, O / NPD / FPD)
K. Sasamoto & N. Ochiai (2010) J. Chromatogr. A 1217, p.2903
(a) 1D GC–MS analysis; (b) Heart-cutting; (c) 2D GC–MS analysis & 1D GC back flush.
DS P2
CT
MS
Inlet P1
LMCS CT
1D Polar
SolGel-WAX 30m*0.32mm*0.50um
2Dshort Apolar DB-5ms
1m*0.1mm*0.1um
2DLong Apolar
DB-5ms 30m*0.25mm*0.25um
FID
MDGC/GC×GC – O/FID/MS Configuration 2012
ES P3
S-T Chin et al. Anal. Chem. 84 (21) (2012) 9154
& 2I values
HYBRID – GC×GC-MDGC Results 1D GC-MS of JP-5 (TIC)
Dominant 57, 71 and 85 m/z
GC×GC-FID
WAX 5% p
heny
l
DS OFF: (GC×GC) DS ON: GC×GC-MDGC)
1D column: SolgelWax (30m x 0.32mm x 0.5µm) 2DM column: VF5 (5m x 0.15mm x 0.15µm) 3DL column: Rxi 17Sil (20m x 0.18mm x 0.18µm)
Two modes: Just Determined by Deans switch
FID1, without H/C – all cpds
On FID2, with H/C (wanted cpds)
FID1, with H/C (unwanted cpds.)
Full GC×GC run, PM = 30 s (20, 12 s also tested)
Cut the compounds in this region (lots of DS events!)
Only the cut compounds (FID2) appear; Separation improved..
After H/C, on FID1 only MATRIX cpds remain
GC×GC – FID1; Not heart-cut COFFEE volatile sample by using SPME:
DS scissors
With Heart-cut of the selected components
Targeted components are excised from the 2D plot!
GC×GC – FID1; Now heart-cut COFFEE volatile sample by using SPME:
9 10 110
100
200
300
400
500
600
Respon
se
Retention Time (min)
9 10 11 12
10
15
20
25
Resp
onse
Retention Time (min)
(R)E
(R)Z
(S)E
(S)Z
(S)Z & (R)Z
(B) (A)
(C) Z
E E
Chiral Interconversion Processes
ISCC and GC×GC 2013 Sunday, May 12, - Thursday, May 16, 2013
Renaissance Palm Springs Hotel
https://m360.casss.org/event.aspx?eventID=46811