multidimensional gas chromatography: past, present & future · zone of eluted compounds from...

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

No cryo

+ cryo 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