Cameron George Technical Support Engineer 26 April 2001 An Optimized Dual Column System for the...

Cameron GeorgeTechnical Support Engineer26 April 2001

An Optimized Dual Column System for the Analysis of Chlorinated Pesticides, Herbicides and PCBs by GC-ECD

Cameron GeorgeApplications ChemistApril 26, 2001


11:00 a.m. ESTTelephone Number: 904-779-4715Chair Person: Lisa Lloyd




Starts in Five Minutes




Starts in One Minute

CLP Pesticides B-0447March 8, 2001

AREAS OF FOCUS

• Injector

• Detector

• Column (including guard column)


TRACE ANALYSIS INJECTION TECHNIQUES5-2000 pg On-Column

• Megabore direct

• Splitless

• PTV

• On-Column (cold & hot)

• Large volume


SAMPLE INJECTIONGoals

• Introduce sample into the column

• Reproducible

• No efficiency losses

• Representative of sample


ShortConcentrated

LongDiffuse

Solute Bands

Same column, same chromatographic conditions

INFLUENCE OF INJECTION EFFICIENCY


FIRST, SOME BASIC DEFINITIONS:

Regarding Inlets

• Backflash

• Discrimination


A FEW WORDS ABOUT BACKFLASH

Definition:

In a vaporization injection, a phenomenon wherein a portion of the sample expands beyond the boundary of the injection port liner towards the septum face and incoming gas line


BACKFLASHEffects

• Ghost peaks

• Erratic quantitation

• Poor accuracy


BACKFLASHCauses

• Highly volatile solvent

• Excessive inlet temperature

• Excessive injection volume

• Small liner volume


BACKFLASHPreventative Measures

• Lower inlet temperature

• Less volatile solvent

• 1-2 µL injection volume

• Chambered liner

• Using pulsed split or pulsed splitless


Yes NO!!!

INJECTION PORT LINERSSplitless


INLET DISCRIMINATION

• Injected sample Sample into the column

• Due to compound volatility differences

• Higher volatility = More into the column


INLET DISCRIMINATIONIn Pesticide Analyses

Generally inlet discrimination is not a concern for

pesticide analyses when utilizing appropriate GC

inlets such as Splitless, Pulsed Splitless and PTV


OTHER INLET CONSIDERATIONS

• Silylation

• Using glass wool

• Cleaning and reusing liners


A FEW WORDS ABOUT RESIDUES

• Semivolatile vs nonvolatile

• Worst offenders

• Sample prep


GUARD COLUMNSUse 0.53mm Or 0.32mm ID Deactivated Tubing

• Fit all inlets

• Enhanced deposition of residues

• Easy to work with

• Minimum 1 meter


Usually 1-5 meters long and same diameter as the column

Detector

DeactivatedFused SilicaTubing

UnionColumn

Injector

GUARD COLUMNTraps non-volatile sample residues


COLUMN CONNECTORSThermal Mass, Inertness, Seal Integrity

• Stainless steel

• VU-Tight

• Press-fits

• Integral guard columns (Duraguard)


Let’s Be SensitiveANALYTE DETECTION


The minimum amount of an analyte that can still be confidently identified as a peak (S/N > 4)

SENSITIVITYAnalytical Definition


SENSITIVITYAnalytical Definition

• Detector Sensitivity: No sample influence (standard)

• Method Sensitivity: Matrix influence (sample)


N

S

N

S x

"Normal" response

1. High noise, detector, background

2. Absorption, breakdown of analyte

3. Sample prep losses

"Improved" response

1. High quality circuit components and reagents

2. Inertness

3. Method optimization

RESPONSE TO A CONSTANT ANALYTE AMOUNT2 Cases

HP 6890 Series Micro-ECD Design

Comparison of HP 6890 ECDs

Standard

< 0.040

> 10^4

no spec.

5 - 80

5

Micro-ECD

< 0.008

> 5 x 10^5

> 5 x 10^4

1 - 500

50

pg/sec lindane

lindane

lindane

pg (ppb)

Hz

MDL

Dynamic range

Linear range

Linear Range, CLP pesticides

Maximum data rate


Break Number 1

For Questions and Answers

Press *1 on Your Phone to

Ask a Question


LETS GET TO THE CHROMATOGRAPHY


WHAT EXACTLY ARE WE TRYING TO ACHIEVE

• The best resolution possible

• Minimize analysis time

• Get MDLs as low as possible


RESOLUTION VS SEPARATION

• Separation: Time between the 2 peaks

• Resolution: Describes how well 2 peaks are separated with regard to their widths


10 11 12 13 14 15

10 11 12 13 14 15

11.24 12.72

(tm = 95.5)

(tm = 95.5)

11.6111.14

WHICH PAIR OF SOLUTES HAVE BETTER SEPARATION?


10 11 12 13 14 15

10 11 12 13 14 15

11.24 12.72

11.6111.14

Better Separation

= 1.17 Rs = 0.6

Better Resolution

= 1.05 Rs = 2.7

K = 7.00K = 6.07

K = 6.00 K = 6.30

RESOLUTION VS SEPARATION


RESOLUTION AND ANALYSIS TIME

• Improving resolution often results in the opportunity to shorten analysis times

• Many variables can affect resolution


N = (L, rc)

k = (T, df, rc)

= (T, phase)

RN kks =

4 11

RESOLUTION


The column must provide sufficient retention of the early eluting compounds without excessive retention of the late eluting compounds

IMPROVING RESOLUTION

Retention


IMPROVING RESOLUTIONFilm Thickness

Decreasing Film Thickness Results In:

• Increased efficiency

• Elution of analytes at lower temperatures

• Decreased analysis time

• Decreased bleed interference

• Increased column activity

• Decreased capacity


IMPROVING RESOLUTIONEfficiency

• High column efficiency is necessary to resolve large numbers of compounds

• Improperly operated injectors and/or improperly optimized carrier gas can result in efficiency losses


IMPROVING RESOLUTIONColumn Length

Increasing Column Length Results In:


• Increased analysis time

• Increased bleed

• Big increase in cost


IMPROVING RESOLUTION Column Inner Diameter

Decreasing Column Inner Diameter Results In:


• Increased head pressure

• Decreased capacity

• Decreased carrier gas flow rates


IMPROVING RESOLUTION Stationary Phase

• Stationary phase selectivity has the largest impact on separation, thus resolution

• Optimization of stationary phase selectivity should be approached cautiously


For Years Environmental Laboratories Have Suffered With Pesticide Analyses Using Non-

Optimal Stationary Phases


TRADITIONAL STATIONARY PHASES

•5% Phenyl-methylpolysiloxane

•35-50% Phenyl-methylpolysiloxane

•Trifluoropropyl-methylpolysiloxane

•14% Cyanopropylphenyl-methylpolysiloxane


PROBLEMS WITH TRADITIONAL PHASES

•Long analysis times (Over 30 minutes!)

•High bleed resulting in decreased sensitivity

•Poor resolution and confirmation capabilities

•Poor inertness of some phases


TRADITIONAL PESTICIDE COLUMNS

High Bleed at 280°C

0 10 20 30 40Time (min.)

0 10 20 30 40Time (min.)

608 Type Phase

1701 Type Phase

Coelution of peaks 10 & 11

40 min

Poor peak shape for Endrin aldehyde


EFFORTS TO IMPROVE PESTICIDE ANALYSES

Application Specific Phases

Stationary phases designed with a primary focus placed upon maximizing separation () for a specific group of target analytes


APPLICATION SPECIFIC PHASES

C2H4CF

CH3m n

Si SiO O

3

Phase 1Trifluoropropyl-dimethylpolysiloxane

3CH

CH3

m n

Si SiO O

C2H4CF

CH

3

3

Common CLP Pesticide Phases

Phase 2Trifluoropropyl-diphenyl-dimethylpolysiloxane

Dimethyl functionality of Phase 2 not shown


EFFECT OF PHASE POLARITY

Polarity

Thermal Stability


DRAWBACKS OF APPLICATION SPECIFIC PHASES

•Limited thermal stability resulting in longer analysis times and hampered sensitivity

•Excessive column conditioning times leading to increased column activity

•Decreased column lifetimes


PHASES DESIGNED WITH OPTIMUM PERFORMANCE IN MINDArylene Phase Technology


CH

CH3

CH3

CH3

CH3

CH3

CH3

Si

Si

Si

Si

O

O

O

OBiphenyl

CH3

CH3

CH3

CH3 CH

3

CH3 CH

3

3

Si

Si

Si

SiO

O

Arylene

LOW BLEED STATIONARY PHASESArylene Structure


DB-35ms

DB-35

8.00 10.00 12.00 14.00 16.00 18.00 20.00

8.00 10.00 12.00 14.00 16.00 18.00 20.00

320°C

300°C

CLP Pesticides Analysis

LOW BLEED STATIONARY PHASESDB-35ms vs. DB-35


STATIONARY PHASE AFFECT ON SENSITIVITY

Both 30 m x 0.25 mm I.D., x 0.25 µm5 ng Decachlorobiphenyl

S/N =10 S/N =3

DB-35ms, 320°C DB-35, 300°C


LOW BLEED STATIONARY PHASESSpectral Purity

m/z-->

51

78 96 119 144

207

197

214

250

253

286

315

346

356 377

405

428

455470

498

50 100 150 200 250 300 350 400 450 500

Primary IonDB-35ms

m/z-->

52

78

96 119

135156

197

207

249

253

281

315 331

356

377

405

428

451

470

498

50 100 150 200 250 300 350 400 450 500

Primary Ion

DB-35


BENEFITS OF LOW BLEED STATIONARY PHASES• Improved sensitivity

• Improved spectral purity

• Reduced analysis times

• Potential increase in column lifetimes

• Rigorous inertness testing

• Less Detector maintenance


Break Number 1

For Questions and Answers

Press *1 on Your Phone to

Ask a Question


DEMANDING PESTICIDE APPLICATIONS BENEFIT FROM ADVANCED PHASE TECHNOLOGY

DB-35ms and DB-XLB capillary columns contain second generation arylene stationary phases giving them enhanced thermal stability


DB-35MS PRIMARY COLUMNCLP Pesticides

•Should provide adequate resolution of all 22 CLP Pesticides and surrogates

•Short analysis time ideal

•Easily realized analysis conditions preferable

•Should give minimal column bleed


1. Tetrachloro-m-xylene (SS)2. -BHC3. -BHC4. -BHC5. Heptachl

or6. -BHC7. Aldri

n8. Heptachlor

epoxide9. -

Chlordane

10. -Chlordane

11. Endosulfan I12. 4,4’-DDE13. Dieldrin14. Endrin15. 4,4’-DDD16. Endosulfan II17

. 4,4’-DDT18

. Endrin

aldehyde19. Endosulfan

sulfate20. Methoxychlor21. Endrin

ketone 22.

Decachlorobiphenyl (SS)

CLP PESTICIDESColumn: DB-35ms

30 m x 0.32 mm I.D., 0.25 µmP/N: 123-3832

Carrier: Helium at 45 cm/sec (EPC in constant flow mode)Oven: 110°C for 0.5 min

110-320°C at 15°C/min320°C for 2 min

Injector: Splitless, 250°C30 sec purge activation time50 pg per component

Detector: µECD, 350°CNitrogen makeup gas (column + makeup flow = 30 mL/min constant flow)

min2 4 6 8 10 12 14 16

6 10 14 168 12

1

2

3

4

56

7

89 10

11

1213

14

15 16

17

18 19

20

21

22

Short Analysis Time

Low Bleed At 320ºC


DB-XLB CONFIRMATION COLUMNWith Regard To The Primary Column

•Should resolve any co-elutions

•Needs to provide adequate resolving power when run under the same analytical conditions

•Should possess at least equivalent thermal stability

•Should provide a different relative elution pattern

•Ideally, elution order changes should be realized


CLP PESTICIDESColumn: DB-XLB

30 m x 0.32 mm I.D., 0.50 µmP/N: 123-1236


110-320°C at 15°C/min320°C for 2 min



min2 4 6 8 10 12 14 166 10 14 168 12

11

2

3

4

6

5

7

88910

11

12

14

13

15

16

19

20

21

22

18

17

100% Confirmation In Under 16 Minutes

1. Tetrachloro-m-xylene (SS)2. -BHC3. -BHC4. -BHC5. Heptachl

or6. -BHC7. Aldri

n8. Heptachlor

epoxide9. -

Chlordane

10. -Chlordane

11. Endosulfan I12. 4,4’-DDE13. Dieldrin14. Endrin15. 4,4’-DDD16. Endosulfan II17

. 4,4’-DDT18

. Endrin

aldehyde19. Endosulfan

sulfate20. Methoxychlor21. Endrin

ketone 22.

Decachlorobiphenyl (SS)


OPTIMIZING FOR SPEED WITHOUT SACRIFICING ANALYTE RESOLUTION

•Use Hydrogen as the carrier gas

•Set linear velocity at 65 cm/sec (3.4 mL/min)

•Increase oven ramp rate from 15°/min to 25°/min


1. Tetrachloro-m-xylene (SS)2. -BHC3. -BHC4. -BHC5. Heptachlor6. -BHC7. Aldrin8. Heptachlor epoxide9. -Chlordane

10. -Chlordane11. Endosulfan I12. 4,4’-DDE13. Dieldrin14. Endrin15. 4,4’-DDD16. Endosulfan II17. 4,4’-DDT18. Endrin aldehyde19. Endosulfan sulfate20. Methoxychlor21. Endrin ketone

22. Decachlorobiphenyl

CLP PESTICIDES USING HYDROGEN CARRIER GASColumn: DB-35ms

30 m x 0.32 mm I.D., 0.25 µmP/N: 123-3832

3 4 5 6 7 8 9 1010

1

2

3

4

5 6

7

8 910

11

12

13

14

1516

17

19

20

21

22

4 85 6 7 9

18

Baseline Resolution In Under 9 Minutes!

Carrier: Hydrogen at 65 cm/sec (EPC in constant flow mode)Oven: 110°C for 0.5 min

110-320°C at 25°C/min320°C for 2 min




1. Tetrachloro-m-xylene (SS)2. -BHC3. -BHC4. -BHC5. Heptachlor6. -BHC7. Aldrin8. Heptachlor epoxide9. -Chlordane

10. -Chlordane11. Endosulfan I12. 4,4’-DDE13. Dieldrin14. Endrin15. 4,4’-DDD16. Endosulfan II17. 4,4’-DDT18. Endrin aldehyde19. Endosulfan sulfate20. Methoxychlor21. Endrin ketone

22. Decachlorobiphenyl

Carrier: Hydrogen at 65 cm/sec (EPC in constant flow mode)Oven: 110°C for 0.5 min

110-320°C at 25°C/min320°C for 2 min



3 4 5 6 7 8 9 10

1

2

3

4

6 5

7

8

9

10

11

12

14

13

15

16

19

20

2122

17

18

105 7 96 8

CLP PESTICIDES USING HYDROGEN CARRIER GASColumn: DB-XLB

30 m x 0.32 mm I.D., 0.5 µmP/N: 123-1236

Excellent Resolution And Confirmation In Under 10 Minutes


OTHER COMMON ECD APPLICATIONS THAT THIS DUAL COLUMN SHOULD BE ABLE TO PERFORM

•EPA 8151A (Phenoxy acid herbicides)

•EPA 508.1 (Pesticides)

•EPA 552.2 (Haloacetic acids)

•EPA 8082 (PCB congeners and Aroclors)


2 4 6 8 10 12 14 16Time (min)

EPA 8151A PHENOXY ACID HERBICIDES

DB-XLB

C785

2 4 6 8 10 12 14 16Time (min)

DB-35ms

C784

2 4 6 8 10 12 14 16

12

34

5

6 7

8

9 10

11

12 131415

16 17

18

19 20

min2 4 6 8 10 12 14 16

1 23 4

5

6 7

8

910

11

121314 15

16 17

1819 20


EPA 8151A PHENOXY ACID HERBICIDESDB-35ms

30m x 0.32 mm I.D., 0.25 µmP/N: 123-3832

DB-XLB30m x 0.32 mm I.D., 0.50 µmP/N: 123-1236


50-100°C at 25°C/min100-320°C at 12°C/min320°C for 2 min

Injector: Splitless, 250°C

30 sec purge activation time50 pg per component


1. Dalapon2. 3,5-Dichlorobenzoic acid3. 4-Nitrophenol4. Methyl-2,4-dichlorophenylacetate (SS)5. Dicamba6. MCPP7. MCPA8. 4,4´, Dibromooctafluorobiphenyl (IS)9. Dichloroprop

10. 2,4-D11. Pentachlorophenol12. 2,4,5-T,P13. 2,4,5-T14. Chloramben15. Dinoseb16. 2,4-DB17. Bentazone18. DCPA19. Picloram20. Acifluorofen


EPA 508.1 PESTICIDES

DB-XLB

DB-35ms

8 10 12 14 16 18 20 22 24Time (min)

min8 10 12 14 16 18 20 22 24

min8 10 12 14 16 18 20 22 24 8 10 12 14 16 18 20 22 24

Time (min)

11

2

3

4

5

6

7

8

9,10

11

12

14

13

1516

17

18

19

20

21,22

23

2425

2627

28

29

303132

33

34

35

3738

36

11

2

3

4

5

67

8,10

9,11

12

14

13

1516

17

18

19

2022

23

2425

262728

29

3031

32

33

34

35

3738

3621

C787

C786


EPA 508.1 PESTICIDES

DB-35ms30m x 0.32 mm I.D., 0.25 µmP/N: 123-3832



75-300°C at 10°C/min300°C for 2 min



1. Hexachloropentadiene2. Etridiazole3. Chloroneb4. Trifluralin5. Propachlor6. Hexachlorobenzene7. a-BHC8. Atrazine9. Pentachloronitrobenzene (IS)

10. Simazine11. g-BHC12. b-BHC13. Hepatachlor14. Alachlor15. d-BHC16. Chlorothalonil17. Aldrin18. Metribuzin19. Metolachlor20. DCPA

21. 4,4´-Dibromobiphenyl (SS)22. Heptachlor epoxide23. Cyanazine24. g-Chlordane25. a-Chlordane26. Endosulfan I27. 4,4´-DDE28. Dieldrin29. Chlorobenzilate30. Endrin31. 4,4´-DDD32. Endosulfan II33. 4,4´-DDT34. Endrin Aldehyde35. Endosulfan sulfate36. Methoxychlor37. cis-Permethrin38. Trans-Permethrin


min4 5 6 7 8

EPA 552.2 HALOACETIC ACIDS

DB-XLB

DB-35ms

4 5 6 7 8Time (min)

min4 5 6 7 8

14

25

6

78,9

10

11

12

4 5 6 7 8Time (min)

1 4

2,3

5

6

7

8

9

10

11

12

3

C789

C788


EPA 552.2 HALOACETIC ACIDS

1. Chloroacetic acid2. Bromoacetic acid3. Dichloroacetic acid4. Dalapon5. Trichloroacetic acid6. 1,2,3-Trichloropropane (IS)7. Bromochloroacetic acid8. Bromodichloroacetic acid9. Dibromoacetic acid

10. 2,3-Dibromopropionic acid (SS)11. Chlorodibromoacetic acid12. Tribromoacetic acid

DB-35ms30m x 0.32 mm I.D., 0.25 µmP/N: 123-3832



40-200°C at 15°C/min200°C for 2 min




EPA 8082 PCB CONGENERS

DB-XLB

DB-35ms

6 8 10 12 14 16Time (min)

min6 8 10 12 14 16

min6 8 10 12 14 16

6 8 10 12 14 16Time (min)

1

2

34

56

78

9

101112

13

1415

16

1718

19

20

21

1

2

3

4

56

78

9

101112

13

1415

16

1718

19

20

21

C791

C790


EPA 8082 PCB CONGENERS1. IUPAC 12. Tetrachloro-m-xylene (IS/SS)3. IUPAC 54. IUPAC 185. IUPAC 316. IUPAC 527. IUPAC 448. IUPAC 669. IUPAC 101

10. IUPAC 8711. IUPAC 11012. IUPAC 15113. IUPAC 15314. IUPAC 14115. IUPAC 13716. IUPAC 18717. IUPAC 18318. IUPAC 18019. IUPAC 17020. IUPAC 20621. Decachlorobiphenyl (IS/SS)

DB-35ms30m x 0.32 mm I.D., 0.25 µmP/N: 123-3832



110-320°C at 15°C/min320°C for 5 min




WHAT DOES THIS MEAN FOR ENVIRONMENTAL LABORATORIES?

•Improved productivity

•Increased sample throughput

•Superior results

•Total solution for ECD systems


CONCLUSION

•Excellent selectivity

•Exceptional inertness

•Superb thermal stability

DB-35ms and DB-XLB provide excellent resolution and confirmation for CLP Pesticides and other critical GC/ECD methods providing total performance:


CONCLUSION

When optimized columns are combined with effective injector setup and detector design improved testing results are realized while sample throughput and laboratory efficiency are improved dramatically


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