ALEXYS system with poor reproducibility

Applies to

ALEXYS system, AS 100 autosampler

Summary

Poor reproducibility in peak height and peak area. Typically %RSD improves with concentration, however sometimes the %RSD is much worse than expected.

Cause

Possible causes:

1. Autosampler
   • most often: air in sampling tubing or syringe
   • leakage of injection loop, valve, syringe, needle or tubing
   • not enough sample in vial
   • air bubble on bottom of micro vial
   • contamination between injections
2. Data acquisition
   • baseline not correctly drawn
   • unstable baseline, large noise, small peaks
   • interfering peaks, shouldering peaks
3. Pump
   • leakage at connectors or seals
4. Detector, HPLC
   • contamination of working electrode
   • reference electrode contaminated, chloride concentration not constant
   • poor column performance, contamination of column
   • not optimised detector settings
   • carbohydrate analysis: carbonate in mobile phase increases ionic strength, constant shift in retention times

Remedy

1. Autosampler
   • remove air from tubing and syringe
   • apply maintenance to autosampler
   • needle wash ‘after each injection’ (in stead of ‘after each vial’)
2. Data acquisition
   • optimise acquisition parameters
3. Pump
   • apply maintenance, replace seals in case of leakage
4. Detector, HPLC
   • apply maintenance to flow cell
   • add constant chloride concentration to REF or mobile phase (ISAAC)
   • replace column (apply cleaning procedure first)
   • optimise detector settings
   • carbohydrate analysis: proper degassing and application of He to prevent carbonate from dissolving in mobile phase.