Different results from HPLC and MS

[viviane-lam]

Hello everyone,
I'm new on this forum so hello I'm having trouble quantifying a molecule by HPLC and MS. The results from HPLC (UV) are pretty stable but i have a lot of variations from MS quantification of the same molecule. For example while HPLC says 50 ppm after 2 times of analysis (different days but same procedure), MS says 50ppm once and 15 ppm the second time. I tend to think that HPLC is more reliable but I can't explain the differences obtained from MS. Do you have any ideas to explain that and how can I verify? Thank you in advance!

[JMB]

Part of the difference between the data can be explained by the instrumental difference between the two methods.

With an HPLC-UV method, the analyte passes through the flow-cell and leaves it clean as a whistle; you could run 50 or 100 replicates of the same sample and get only VERY minor differences in area counts. Over a longish period of time (some years) the deuterium lamp will age and area counts will decline. UV ( like NMR) is a non-destructive detection method.

Any method based on Mass Spec is destructive of the analyte, and every single analysis degrades the performance of the instrument. You COULD prove this to yourself by starting with a newly-cleaned source and rods and making replicate injections of a single analyte; you will see the area counts fall quite obviously.

50 ppm and 15 ppm are more different than you would expect to see if that was a single-analyte sample, run by MS back-to-back on the same day; I would expect only about 5% difference.
However, 50 and 15 ppm on DIFFERENT days—-how many other samples (each of which degraded the MS performance somewhat) went through in the interim?

If you set up to run say more than 5-10 samples at a time, you should think about including a check standard or calibration standard every so often; others can advise on the details.

Good Luck!

[H.Thomas]

Just as a side note: 50 ppm (mg/L) is probably way too much for the MS. By injecting such large concentrations you surely contaminate your ion souce.

And it's true, for quantitation UV is more reliable than MS, but only if you analyse high concentrations in pure solvent