Chromatography Integration and Trouble-shooting

1. Chromatography Integration and Trouble-shooting Presented by: Marlene Moore mmoore@advancedsys.com October 2007

2. Agenda • Problems • Tools • Solutions

3. Problems • Baseline Disturbances • Irregular Peak Shapes, Sizes • Retention Time Shifts • Loss of Separation or Resolution • Quantitation Difficulties • Rapid Column Deterioration • Ghost Peaks • Broad Solvent Front

4. Baseline Disturbances spiking noise wander drift offset

5. Spiking • Particulate Matter Passing Through Detector • Loose Connections • Cables • Circuit boards

6. Noise • Contaminated Injector/column • Physical Defect Detector • Defective Detector Board • Carrier Gas Leak at Septum or Column • FID, NPD, FPD • Column in Flame • Incorrect Combustion Gases or Flow Rate/leak • ECD, TCD • Air Leak

7. Wander • Contaminated Carrier Gas • Isothermal Conditions • Contaminated Gas Chromatograph • Poor Control of Carrier Gas or Detector Gas Flows • Poor Thermal Control of Detector

8. Drift • Septum Bleed • Upward • GC or column contamination • Damaged stationary phase • Downward • Incomplete conditioning of column • Unequilibrated detector

9. Offset • Contamination: • Injector, column, detector • Carrier gas - change/install impurity traps • Column Inserted Into Flame (FID, NPD, FPD) • Malfunctioning or Improperly Set Recording Device

11. Peak Shape Peak Shape tells the story of your chromatography

12. Irregular Peak Shapes tailing fronting split reduced rounded negative

13. No Peaks • Plugged Syringe • Broken Column • Injecting Sample Into Wrong Injector • Column Installed in Wrong Detector • Integrator Reading Wrong Signal or Not Connected • Detector Not on • Temperature • Oven not heated • Column temperature too low • Injection port temperature too low • Detector Gases Improperly Set or Not on • Very Low or No Carrier Gas Flow

14. Reduced Peaks • Plugged Syringe • Change in Injection Technique • Large Leak - Injector • Split Ratio Too High • Too Short of Purge Activation Time for Splitless Injections • Very High Septum Purge Flow • Too Low Injector Temperature • Column Temperature Too Low • Sample Components Absorbed by Column or Inlet Liner

15. Reduced Peaks • Initial Temperature Column Too High for Splitless or On-column Injections • High Background Signal - • Contamination • Excessive column bleed • Check auto zero function and setting • Improperly Operated Detectors • Impurities Carrier Gas or Detector Gas • Detector Compound Mismatch • Excessive Attenuated Integrator Signal • Sample Concentration or Integrity Problems

16. Tailing Peaks • Active Injector Liner or Column • Contaminated • Injector liner • Column • Dead Volume Caused by Poorly Installed Liner, Column or Union • Poorly Cut Column End • Polarity Mismatch - Stationary Phase, Solute or Solvent • Cold Spot in the Flow Path

17. Tailing Peaks • Solid Debris in Liner or Column • Poor Injection Technique - Too Slow • Too Low of a Split Ratio • Overloading on a PLOT Column • Decrease sample amount • Amines and carboxylic acids tail on most columns • Need ph modified stationary phase • Some Peaks Always Exhibit Tailing

18. Rounded or Flat Topped Peaks • Overloaded Detector • Exceeding Range of Integrator

19. Split Peaks • Detector Overloaded • Poor Injection Technique • Poorly Installed Column Into Injector • Column Temperature Fluctuations • Coelution of Two or More Compounds • Mixed Sample Solvent for Splitless or On-column Injections.

20. Negative Peaks • All Peaks Negative - Polarity Check Connectors • TCD - Select Peaks • Thermal conductivity of compound greater than carrier gas • ECD - After Positive Peak • Dirty or old ECD cell

21. Retention Time Shifts • Different Column Temperature • Different Carrier Gas Flow or Linear Velocity • Leak in Injector - Septum • Contaminated Column • Change in Sample Solvent

22. Loss of Separation/Resolution • Contaminated Column • Damaged Stationary Phase • Different Column Temperature, Carrier Flow Rate, Column • Large Changes in Sample Concentrations • Improper Injector Operation

23. Quantitation Difficulties • Injection Techniques • Split Discrimination • Different Purge Activation Time for Splitless Injection • Baseline Disturbances • Improper Integrator Settings • Inconsistent Detector Gas Flows or Temperatures • Column or Liner Activity

24. Rapid Column Deterioration • Exposure of Column to Air at Elevated Temperatures • Exceeding Upper Temperature Limit of Column • Chemical Damage • Contamination of Column With High Molecular Weight Materials • Column Breakage

25. Ghost Peaks • Contamination • Injector • Column • Septum Bleed • Dirty Syringe • Sample Too Large • Previous Run Terminated Too Soon BOO!

26. Broad Solvent Front • Poorly Installed Column • Leak in the Injector • Solvent Interacting with Detector • Too Low • Split Ratio (Flow) • Injector Temperature • Too Long Purge Activation for Splitless Injections • Large Injection Volume • Low Column Temperature and High Boiling Solvent • High Column Temperature and Low Boiling Solvent

28. Tools • Flowmeter • Syringe (NEW) • Methane or Nonretained Compound • Septa, Ferrules, Injection Liners (NEW) • Leak Detector • Column Test Mixture (Reference) • Checkout Column • Instrument Manuals

29. Solutions Quickest Solution ---- Improve Chromatography --- Do Manual Integration --- Get Results out Fast !! NO! WRONG ANSWER!!!

30. Solutions • Preventive Maintenance • Clean detector • Clean injector • Clean and repair parts • Clean and tighten connections • Follow instrument manual suggestions • Allow Time for Optimizing System • Integration parameters • Programs

31. Does anyone have a question? Question ! Where do you document your trouble shooting?