Time controlled Cryogenic Zone Compression (t- CZC) GC … · ppq 1 ul injected Attogram – ag!? Ultra sensitivity – What for? 6. Dried Blood Spot (DBS) analysis for POPs: •

Time controlled Cryogenic Zone Compression

(t- CZC) GC-HRMS:

- can we detect attogramm amounts of TCDD?

Dirk Krumwiede & Heinz Mehlmann

Thermo Fisher Bremen

Acknowledgements

Daniela Cavagnino

Brock Chittim

Jef Focant

Don Patterson Jr.

Wayman Turner

2

Wayman Turner

Andreas Sjodin

Jack Cochran

It is all about (instrumental) sensitivity!

3

DFS: magnetic sector HRMS

- Highest selectivity!

- Highest sensitivity!

- Highest……speed?

2005 �DFS High Resolution MS

Introduced atthe 2005 Dioxin Conference in

Toronto

4

It is all about (instrumental) sensitivity!

50

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10.83

ppb

ppt

Picogram - pg (/ul)

20 fg 2378-TCDD,

Femtogram

5

DFS: magnetic sector HRMS

- Highest selectivity!

- Highest sensitivity!

- Highest……speed?

7.5 8.0 8.5 9.0 9.5 10.0 10.5 11.0 11.5

Time (min)

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10.91 11.0910.299.09 11.609.268.81 11.268.78 10.04 10.539.749.338.478.137.68 7.92 9.638.407.59

ppt

ppq

20 fg 2378-TCDD,

1 ul injected

Attogram – ag!?

Ultra sensitivity – What for?

6

Dried Blood Spot (DBS) analysis for POPs:

• Very small sample (20 - 50 ul Serum)

• Extremely low analyte amounts for TCDD

• Sample extract volume = ca. 5 ul

� Sensitivity must come from instrument

DBS – the challenge: ultra sensitivity for some analytes

7

� Sensitivity must come from instrument

Newborn Screening Programs (NSPs), etc.:

• Very important samples for epidemiological studies

• Population exposure and trends

What can GC do to increase sensitivity?

8


RT: 21.69 - 22.12

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NL:3.10E4

m/z= 303.9419-303.9723 MS ICIS 5w_cryomix_11

9

21.7 21.8 21.9 22.0 22.1

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RT: 21.89AA: 10792


RT: 21.69 - 22.12

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RT: 21.89AA: 10539

NL:3.10E4


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RT: 21.95AA: 51.20

PCB 77 (13C, IS)

Produce narrow peaks = increase Peak Height

narrower peak

RT: 21.69 - 22.12

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RT: 21.89AA: 10539

NL:3.10E4


NL:3.10E4


0.6 sec pw (at 10%)

11

= higher peak

= better sensitivity

1/pw = ph

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RT: 21.89AA: 10792

RT: 21.95AA: 51.209.6 sec pw (at 10%)

Cryogenic (peak) Zone Compression

- from GCxGC to CZC

GCxGC – increased peak capacity / separation power

13

Modulation: several fast 2 dim. chromatograms over each first dim. Peak

GCxGC main feature: increased peak capacity

GCxGC secondary feature: increased sensitivity (very narrow 2d dim peaks)

How to maximize the sensitivity effect of GCxGC?

14


GCxGC

15

- modulation of 1 dim. Peak

- several 2 dim. chromatograms


GCxGC CZC

16

- modulation of 1 dim. Peak

- several 2 dim. chromatograms

- cryofocus complete 1 dim. Peak

- one single 2 dim. chromatogram

„Ion dilution“

Original idea for CZC – magnetic sector HRMS by….see paper

- R&D work at CDC Atlanta:

For Dioxin analysis…

…combine highest sensitive MS

with highest sensitive GC method

�� GCxGC + sector HRMS

17

�� GCxGC + sector HRMS

The entire peak is cryo-trapped

in one single event.

For only 2378 Cl containing…

- blood / serum samples

- dried blood spot analysis

Timed Cryo Zone Compression (t-CZC):

How does it work?

Injector

timed Cryogenic Zone Compression (t-CZC)

- one single standard column (no 2d column, no connection)

- only one jet is used

- jet on/off timed controlled

� Simple & flexible

19

(4m)56m

CO2 Jet

MS

2nd Dimension1st Dimension

Guideline: As complicated as necessary, as simple as possible!

Peak Zone Compression: Cryo Zone Compression (CZC)

20

CO2 Modulator of Thermo Scientific Trace GCxGC ultra

(4m)56m

MS

Injector


(t-CZC) Principle

21

CO2 Jet

RTnormaltime

ChromatogramNormal Peak – CZC inactive

(4m)56m

MS

Injector


At the time when the peak of interest reaches the Jet…

(t-CZC) Principle

22

CO2 Jet

RTJettime

Chromatogram

(4m)56m

MS

Injector


…the Jet is switched on to trap the entire peak of interest

(t-CZC) Principle

23

Jet on

CO2 Jet

RTJettime

Chromatogram

(4m)56m

MS

Injector


…and then reinject to the 2 d dimension (Jet off again)

(t-CZC) Principle

24

Jet off

CO2 Jet

RTCZCtime

ChromatogramReinject on 2d Dim.

c:\xcalibur\data\110622_cryo_01\cryo08 6/22/2011 12:00:10 PM CSL X1/10GC2: column: 60m x 0.25 mm x 0.1 um; 120(2)-10-220(0)-3-235(7)-4.6-310(5); He flow 1.0ml/min ; 45eV tcdd precool 23.10-24.3 tcdd trap I: 25.15-25.55 pcdd/f precool 29-29.5 and 30.5-31.07

RT: 19.90 - 50.42

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48.42

39.21 45.3338.0425.62 42.9337.81 40.5824.98 30.14 44.6132.01

32.50

NL:2.60E6

TIC - m/z= 314.4000-318.4000 MS Cryo09

„normal“ Dioxin/Furan chromatogram on 60 m col.

(t-CZC) Principle

25

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25.76

32.08

32.58

48.43

38.02 39.18 45.3242.9240.5837.7924.99 30.1340.02

NL:2.60E6

TIC - m/z= 314.4000-318.4000 MS cryo08


RT: 19.90 - 50.42

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48.42

39.21 45.3338.0425.62 42.9337.81 40.5824.98 30.14 44.6132.01

32.50

NL:2.60E6

TIC - m/z= 314.4000-318.4000 MS Cryo09

Selection of target analytes to becryofocused

(t-CZC) Principle

26

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32.58

48.43

38.02 39.18 45.3242.9240.5837.7924.99 30.1340.02

NL:2.60E6

TIC - m/z= 314.4000-318.4000 MS cryo08


RT: 19.90 - 50.42

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39.21 45.3338.0425.62 42.9337.81 40.5824.98 30.14 44.6132.01

32.50

NL:2.60E6

TIC - m/z= 314.4000-318.4000 MS Cryo09

Same scaling

(t-CZC) Principle

27

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32.08

32.58

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38.02 39.18 45.3242.9240.5837.7924.99 30.1340.02

NL:2.60E6

TIC - m/z= 314.4000-318.4000 MS cryo08

CZC focused peaks


RT: 19.90 - 50.42

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48.42

39.21 45.3338.0425.62 42.9337.81 40.5824.98 30.14 44.6132.01

32.50

NL:2.60E6

TIC - m/z= 314.4000-318.4000 MS Cryo09

(t-CZC) Principle

28

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48.43

38.02 39.18 45.3242.9240.5837.7924.99 30.1340.02

NL:2.60E6

TIC - m/z= 314.4000-318.4000 MS cryo08

Selected analytes with CZC2378-TCDD

1234-TCDD 12378-PeCDF

12378-PeCDD

23478-PeCDF Only selectedpeaks arerefocussed

standard GC-HRMS vs. CZC GC-HRMSpooled blood sample on DFS (HRMS) – 2378-TCDD ca. 10 fg

RT: 21.00 - 26.31

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RT: 25.97AA: 630SN: 8RMS

21.1725.3124.8724.3422.8422.51

23.52 24.14 24.5522.1121.77 23.47

RT: 25.99AA: 865SN: 16RMS

25.95

NL:1.87E2

m/z= 319.8810-319.9130 MS ICIS Cryo20

NL:2.58E2

m/z= 321.8779-321.9101 MS ICIS Cryo20

RT: 23.15 - 26.31

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RT: 26.08AA: 1023SN: 149RMS

25.18

RT: 26.08AA: 1292SN: 273RMS

NL:2.61E3

m/z= 319.8810-319.9130 MS ICIS Cryo18

NL:3.43E3

m/z= 321.8779-321.9101 MS ICIS Cryo18

AA: 866

Height: 258

SignoiseRMS: 16AA: 1292

Height: 3430

Qual mz

Quan mz

29

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25.7323.2224.2022.41 24.0621.51 22.19 22.82 25.6324.80

21.94

RT: 25.29AA: 254225SN: 3574

NL:5.68E4

m/z= 333.9173-333.9507 MS ICIS Cryo20

23.5 24.0 24.5 25.0 25.5 26.0

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RT: 26.07AA: 45.44SN: 13RMS

RT: 26.08AA: 260302SN: 70559

RT: 25.26AA: 275184SN: 6397

NL:7.05E5

m/z= 333.9173-333.9507 MS ICIS Cryo18

SignoiseRMS: 16 Height: 3430

SignoiseRMS= 273

non-cryofocused

1234-TCDD

Quan mz

13C IS mz

t-CZC paper at: http://pops.thermo-bremen.com

In Google:

- type „POPs“ & „Thermo“

- first hit is this page

30

Chromatography Today

February / March 2012

And…..? Can we detect ag amounts?

Goal: optimize method for as low iLOQs as possible

Wellington attogram standards

32

Simple goal:

Go as low as possible (absolute amount injected)

Thx, Brock!!

Lower & lower & lower

33

Lower & lower & lower …the Patterson LimboRT: 14.00 - 14.91

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RT: 14.64

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NL:

1.10E4

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_116

NL:

4.02E3

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_115

NL:

1.48E3

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_113

5 fg/ul ?

34

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Time (min)

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RT: 14.64

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NL:

9.74E2

m/z=

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321.9061 MS

ICIS

1209_CZC_TF_

LL_108

NL:

3.36E2

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_104

NL:

2.40E1

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_100

Lower & lower & lower …the Patterson limboRT: 14.00 - 14.91

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RT: 14.64

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m/z=

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321.9061 MS

ICIS

1209_CZC_TF_

LL_116

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4.02E3

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_115

NL:

1.48E3

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_113

5 fg/ul

2 fg/ul ?

35

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RT: 14.64

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NL:

9.74E2

m/z=

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321.9061 MS

ICIS

1209_CZC_TF_

LL_108

NL:

3.36E2

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_104

NL:

2.40E1

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_100


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RT: 14.64

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RT: 14.65

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NL:

1.10E4

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_116

NL:

4.02E3

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_115

NL:

1.48E3

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_113

5 fg/ul

2 fg/ul

1 fg/ul ?

36

14.0 14.1 14.2 14.3 14.4 14.5 14.6 14.7 14.8 14.9

Time (min)

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RT: 14.65

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RT: 14.64

AA: 104

NL:

9.74E2

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_108

NL:

3.36E2

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_104

NL:

2.40E1

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_100


0

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RT: 14.64

AA: 4727

RT: 14.64

AA: 1740

RT: 14.65

AA: 667

NL:

1.10E4

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_116

NL:

4.02E3

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_115

NL:

1.48E3

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_113

5 fg/ul

2 fg/ul

1 fg/ul

37

14.0 14.1 14.2 14.3 14.4 14.5 14.6 14.7 14.8 14.9

Time (min)

0

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RT: 14.64

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NL:

9.74E2

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_108

NL:

3.36E2

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_104

NL:

2.40E1

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_100

500 ag/ul ??


0

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RT: 14.64

AA: 4727

RT: 14.64

AA: 1740

RT: 14.65

AA: 667

NL:

1.10E4

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_116

NL:

4.02E3

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_115

NL:

1.48E3

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_113

5 fg/ul

2 fg/ul

1 fg/ul

38

14.0 14.1 14.2 14.3 14.4 14.5 14.6 14.7 14.8 14.9

Time (min)

0

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0

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RT: 14.65

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RT: 14.64

AA: 104

NL:

9.74E2

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_108

NL:

3.36E2

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_104

NL:

2.40E1

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_100

500 ag/ul

200 ag/ul ???


0

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RT: 14.64

AA: 4727

RT: 14.64

AA: 1740

RT: 14.65

AA: 667

NL:

1.10E4

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_116

NL:

4.02E3

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_115

NL:

1.48E3

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_113

5 fg/ul

2 fg/ul

1 fg/ul

39

14.0 14.1 14.2 14.3 14.4 14.5 14.6 14.7 14.8 14.9

Time (min)

0

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100

0

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0Rela

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bundance

RT: 14.65

AA: 300

RT: 14.64

AA: 104

NL:

9.74E2

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_108

NL:

3.36E2

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_104

NL:

2.40E1

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_100

500 ag/ul

200 ag/ul

????


0

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Rela

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bundance

0

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0

50

100

RT: 14.64

AA: 4727

RT: 14.64

AA: 1740

RT: 14.65

AA: 667

NL:

1.10E4

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_116

NL:

4.02E3

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_115

NL:

1.48E3

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_113

5 fg/ul

2 fg/ul

1 fg/ul

40

14.0 14.1 14.2 14.3 14.4 14.5 14.6 14.7 14.8 14.9

Time (min)

0

50

100

0

50

100

0

50

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0Rela

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bundance

RT: 14.65

AA: 300

RT: 14.64

AA: 104

NL:

9.74E2

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_108

NL:

3.36E2

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_104

NL:

2.40E1

m/z=

321.8739-

321.9061 MS

ICIS

1209_CZC_TF_

LL_100

500 ag/ul

200 ag/ul

blank

Wellington TF-LL-CS standards: 200 ag/ul to 5 fg/ul 2378-TCDD

2378-TCDD 5.0 fg/µl

triplicats of low levels

41

� 800 ag absolute amount for lowest level

�4 ul injected500 ag/µl

200 ag/µl

1.0 fg/µl

2.0 fg/µl


Wellington TF-LL-CS standards: 200 ag/ul to 5 fg/ul 23478-PCDF

23478-PCDF 5.0 fg/µl


42

500 ag/µl

1.0 fg/µl

2.0 fg/µl

200 ag/µl [fg / ul]

�4 ul injected

� More difficult than TCDD


800 ag 2378-TCDD (Wel. TF-LL-CS) – repeatability

RT: 13.64 - 14.97

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RT: 14.64AA: 85.11

RT: 14.64AA: 103

NL:1.87E2

m/z= 319.8810-319.9130 MS ICIS 1209_CZC_TF_LL_104

NL:3.36E2


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14.62

14.71 14.9013.99 14.07 14.3413.69 14.60

RT: 14.64AA: 49.65

NL:2.83E2


NL:1.38E2


1.) 800 ag 2.) 800 ag

RM

QM

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3.) 800 ag

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1.) 800 ag 2.) 800 ag

IS (13C)

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3.) 800 ag

4 ul Injection Volume (4 x 200 ag = 800 ag)

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RT: 14.64AA: 49.65

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NL:1.38E2


1.) 800 ag 2.) 800 ag

RM

QM

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RT: 14.65AA: 29.67

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3.) 800 ag

Sum Area:

188

Sum Area:

187Sum Area:

158

800 ag 2378-TCDD (Wel. TF-LL-CS) – repeatability

44

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1.) 800 ag 2.) 800 ag

IS (13C)

14.0 14.5

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RT: 14.22AA: 90.18

RT: 14.63AA: 150

NL:7.68E4


3.) 800 ag

PW = ca. 0.7 sec

(at 10%)

4 ul Injection Volume (4 x 200 ag = 800 ag)

CZC sensitivity

- We do not see nothing (!) at levels of 800 ag TCDD

- Statistics are getting worse the lower the level – no big surprise, is it?

- This is where we are – is it the optimum?

45

The challenges…

sensitivity

GC peak width MS dwell time

Why „only“ 0.6 – 1 sec wide peaks, why not narrower?

?

�Slow down GC

�Speed up HRMS

Find optimum settings

47

Time [sec]

? Find optimum settings

� Both sides - GC and MS - must

work at their optimum for best results!!

Method development – ion detection rateC:\Xcalibur\...\5w_cryomix_13 7/21/2011 4:25:22 PM CZC: 1 ul 2fg/ul pcb/pcdd/pcdf mix (WEL. TF-LL-CS5)

120(2)-10-220(10)-3-235(7)-4.6-310(8); He flow 1.0ml/min ; 45eV, 3 mass MID (+ L & C): cycle time 0.1 sec, ML ratio 10, eld 6 msec, 24 ms on nat, 12 on IS

RT: 21.80 - 21.98

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AA: 10792

NL:

2.66E3

m/z=

303.9419-

303.9723

MS ICIS

5w_cryomix

_11

standard pw >10 s

48

21.80 21.82 21.84 21.86 21.88 21.90 21.92 21.94 21.96 21.98

Time (min)

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0RT: 21.89

AA: 10539

NL:

3.10E4

m/z=

303.9419-

303.9723

MS ICIS

5w_cryomix

_13

CZC pw 0.6-0.8 s

Low level statistics

2378-TCDD

49

��We want to quantify where others hope to have their calculated LODs

RSD

e.g. ion ratios

(or RRFs, etc.)

+15%

Low level statistics – ion statistics

� Very few ions are detected at very low levels

50

amount

-15%

+15%

- Which minimum RSD value is required to quantify ?

How much variation is acceptable ?

CZC sensitivity – authors opionion

Statistics: ion ratios, RSDs, linearity, data points over peak, etc.??

Why do you need your prescription glasses in total darkness?

Before you are good you are not so good.

51

Standardization � Innovation

Potpourri of things…

PCDF: CZC vers. normal peak at 50 fg (CSL 1:10 dil) D:\Xcalibur\...\1209_CZC_pcddf_027 9/13/2012 3:11:07 PM

(used) TR-Dioxin5ms 30x0.25 (0.1), set jet on / off times, determin precool times without distorting non CZC targets, adapt MID times for CZC targets

RT: 16.03 - 17.47 SM: 5G

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RT: 17.41

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RT: 16.58

AA: 3891

RT: 17.41

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NL:

8.35E3

m/z=

339.8409-

339.8749

MS ICIS

1209_CZC_

pcddf_027

NL:

5.15E3

m/z=

341.8399-

341.8741

MS ICIS

1209_CZC_

50 fg

Quan mz

- PCDFs have very similar response

One analysis run / same injection:

- 1st PCDF � normal detection

- 2nd PCDF � cryofocussed

- compare directly

12378-PeCDF

23478-PeCDF

53

16.1 16.2 16.3 16.4 16.5 16.6 16.7 16.8 16.9 17.0 17.1 17.2 17.3 17.4

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AA: 2678

RT: 17.40

AA: 766176

RT: 16.56

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RT: 16.73

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1209_CZC_

pcddf_027

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m/z=

351.8824-

351.9176

MS ICIS

1209_CZC_

pcddf_027

Qual mz

13C IS mz

here: 30 m column

10 pg

PCDF: CZC vers. normal peak at 2.5 fg (CSL 1:200 dil) D:\Xcalibur\...\1209_CZC_pcddf_204 9/13/2012 9:17:06 PM

(used) TR-Dioxin5ms 30x0.25 (0.1), set jet on / off times, determin precool times without distorting non CZC targets, adapt MID times for CZC targets

RT: 16.03 - 17.47 SM: 5G

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RT: 16.58

AA: 18.00

RT: 17.42

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NL:

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m/z=

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339.8749

MS ICIS

1209_CZC_

pcddf_204

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1.78E2

m/z=

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341.8741

MS

1209_CZC_

2.5 fg

Quan mz

54

16.1 16.2 16.3 16.4 16.5 16.6 16.7 16.8 16.9 17.0 17.1 17.2 17.3 17.4

Time (min)

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RT: 16.57

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RT: 17.42

AA: 26236

RT: 16.57

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351.8824-

351.9176

MS ICIS

1209_CZC_

pcddf_204

here: 30 m column

Qual mz

13C IS mz

500 fg

RT: 23.00 - 28.00 SM: 9G

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RT: 23.96AA: 8378

RT: 26.44AA: 7718

RT: 25.56AA: 7655

RT: 25.99AA: 6939

RT: 27.07AA: 7184

RT: 25.67AA: 8420

RT: 25.96AA: 7725

RT: 26.41AA: 7406

RT: 27.04AA: 7006

RT: 26.11AA: 8740

NL:1.30E4

m/z= 321.7940-321.9940 MS ICIS hm_tczc_trap_01

NL:1.30E4


NL:1.30E4

m/z= 321.7940-321.9940 MS

1378-TCDD

1478-TCDD

1234-TCDD

2378-TCDD

• 1.) No Cryofocusing

• 2.) Only 1378 TCDD focused

Selective focusing of peaks in t-CZC

55

23.0 23.5

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25.0 25.5 26.0 26.5 27.0 27.5 28.0

Time (min)

RT: 25.57AA: 8813

RT: 26.42AA: 8281

RT: 27.07AA: 8408

RT: 26.54AA: 7486

RT: 25.58AA: 7826

RT: 27.06AA: 7248

RT: 26.02AA: 7369

RT: 27.17AA: 7371

RT: 25.97AA: 7885

RT: 25.55AA: 7543

RT: 26.45AA: 7351

321.9940 MS ICIS hm_tczc_trap_03

NL:1.30E4


NL:1.30E4





PCDFs separated on Dim 1 vers Dim 2

RT: 16.23 - 17.73 SM: 5G

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RT: 17.22AA: 112591RT: 16.47

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RT: 17.22AA: 69028

RT: 16.47AA: 66509

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m/z= 339.8409-339.8749 MS ICIS 1207_pcdd_cryo_dk623

NL:2.20E4


RT: 16.23 - 17.73 SM: 5G

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RT: 17.62AA: 1623

RT: 17.64AA: 70077

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RT: 17.51 - 17.83 SM: 5G

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Dim 1 Dim 2 Dim 2

PCDFs trapped

together and reinjected

56

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RT: 17.61AA: 105529

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17.6 17.7 17.8

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together and reinjected

on Dim. 2

• NTC resistor positioned in the cryo jet and connectet to a digital meter

• Jet temperature at the RTJet (peakfront) 195.25°C

Method development – temperature profiles

• Determination of the trapping-temperature

57

Jet on Jet off

Fast GC experiments for narrow peaks – here: 5 m column

RT: 4.34 - 8.16

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RT: 5.77AA: 25376796

RT: 6.21AA: 19945764

RT: 6.73AA: 12244407

RT: 7.38AA: 5598786

RT: 5.01AA: 26360860

RT: 5.45AA: 19065901

RT: 5.84

NL:2.90E7

TIC MS ICIS 1204_5m018_pcddf114

NL:3.61E7

TIC MS ICIS 1204_5m018_pcddf117

Ramp 30 ºC/min

Ramp 40 ºC/min

58

4.5 5.0 5.5 6.0 6.5 7.0 7.5 8.0

Time (min)

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RT: 5.84AA: 11185391

RT: 6.63AA: 4432709

RT: 4.91AA: 19089855

RT: 5.33AA: 11012244

RT: 5.67AA: 6168403

_pcddf117

NL:3.62E7

TIC MS ICIS 1204_5m018_pcddf121Ramp 50 ºC/min

Column: Restek RXI 5 Sil MS 5 m x 0.18 mm (0.18 um)

Thx for your help, Jack!

Fast GC – peak width “tuning”

RT: 5.70 - 5.86

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RT: 5.77

NL:2.90E7

m/z= 317.8390-318.0390 MS ICIS 1204_5m018_pcddf114

NL:

TCDF Column:

Restek RXI 5 Sil MS

5 m x 0.18 mm (0.18 um)

Ramp 30 ºC/min

PW (at 10 %) = 1.5 sec

59

5.70 5.72 5.74 5.76 5.78 5.80 5.82 5.84 5.86

Time (min)

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NL:3.62E7

m/z= 317.8390-318.0390 MS ICIS 1204_5m018_pcddf121

TCDF

Ramp 50 ºC/min

PW (at 10 %) = 1.08 sec

to overlay:

RT + 1.25

What next?

Serum samples from CDC – dilution of reference samples

61

- dilute contaminated serum with blank matrix (bovine serum)

- prepare defined TCDD, PCDF, PCDD serum sample levels this way

Serum sample TCDD - 1ul injectedRT: 14.20 - 14.59

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RT: 14.54MA: 173

14.4414.32

14.3414.2614.23 14.3814.57

14.27 14.4614.39

14.47

RT: 14.54MA: 151

14.53

14.3514.52

14.25

NL:3.71E2

m/z= 319.8810-319.9130 MS 1207_serum_1021

NL:3.94E2

m/z= 321.8779-321.9101 MS 1207_serum_1021

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14.47

14.53

14.27 14.41 14.5114.23 14.4314.29 14.4014.32

14.54

RT: 14.48MA: 474

NL:7.73E2

m/z= 319.8810-319.9130 MS 1207_serum_1022

NL:1.09E3

m/z= 321.8779-321.9101 MS 1207_serum_1022

8A – 1.25 fg/ul 6A – 5 fg/ul

62

14.3 14.4 14.5

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14.4114.3214.31

14.3914.46

14.22

RT: 14.54AA: 967532

NL:2.42E6

m/z= 333.9173-333.9507 MS ICIS 1207_serum_1021

14.3 14.4 14.5

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14.46 14.52 14.5314.24 14.35 14.4214.30

RT: 14.48AA: 1087219

RT: 14.50AA: 8814

NL:2.57E6

m/z= 333.9173-333.9507 MS ICIS 1207_serum_1022

Blanks – the real limitation?

Instrumental analysis push to improve

63

Sample preparationpush to improve

Conclusions & Outlook

Conclusions & Outlook:

In general: - t-CZC = sensitivity „booster“

- basically on any GC, preferable: selective detector

- selected applications / target analytes

Features & Advantages: - simplicity & flexibility

- low consumption of CO2

65







Current status (DFS): - sensitivity ca. 700 - 1000 ag (2378-TCDD, absolute amount)

66

Next step: - transfer method to customer lab (CDC Atlanta, US)







Current status (DFS): - sensitivity ca. 700 - 1000 ag (2378-TCDD, absolute amount)

67

Next step: - transfer method to customer lab (CDC Atlanta, US)

…other fields of application?

shortlong

CO2 Jet

MS

Injector


GCxGC – switch on/off different modes as needed?

Ability to switch on different modes just

as needed throughout a chromatogram:

1.) „normal“ sections – jet off

2.) CZC for increased sensitivity

3.) CZC for additional sep. on 2d dim

68

3.) CZC for additional sep. on 2d dim

4.) full GCxGC sectionsunpolar polar

(low bleed)

http://pops.thermo-bremen.com

69

http://pops.thermo-bremen.com

70

Thanks for your attention!!

?- suggestions?

- applications?

71

Documents

Time controlled Cryogenic Zone Compression (t- CZC) GC … · ppq 1 ul injected Attogram – ag!? Ultra sensitivity – What for? 6. Dried Blood Spot (DBS) analysis for POPs: •