HPLC/Radiolabeled Analyses
Posted: Wed Apr 30, 2014 6:16 pm
Hi,
We are doing analyses on a radiolabeled compound, utilizing HPLC-UV coupled with a radiometric detector. We run an HPLC “standard” of the radiolabeled parent material before and after each set of samples in order to confirm instrument and system stability. Typically, if our standard is 98% parent at the beginning of the run, we can expect it to be 98% plus or minus a percent or two at the end. Lately I have been having real trouble with the chromatographic integrity of the standard deteriorating over the course of a run; from 95% parent or better at the beginning to <80% at the end, with several preceding and trailing peaks around the parent. If I come in the next day, using the same standard solution in the same vial, I see the same pattern – OK at the beginning, garbage at the end. I thought that perhaps it was the test samples contaminating the column, but today I made several injections of the standard by itself, and I saw the same appearance of more and larger non-parent peaks as the day went on, although not as drastic as when running test samples. I have made fresh standards, fresh mobile phases – no help, same pattern. This might be a stupid question, but - is it possible that if the analytical column is not in the best of health, that it could “re-equilibrate” itself simply by sitting overnight, or when I condition it briefly in the morning before a run? Then, as a run progresses, it deteriorates again to the point at which it will no longer properly retain the analyte? Any ideas out there? Thanks much.
We are doing analyses on a radiolabeled compound, utilizing HPLC-UV coupled with a radiometric detector. We run an HPLC “standard” of the radiolabeled parent material before and after each set of samples in order to confirm instrument and system stability. Typically, if our standard is 98% parent at the beginning of the run, we can expect it to be 98% plus or minus a percent or two at the end. Lately I have been having real trouble with the chromatographic integrity of the standard deteriorating over the course of a run; from 95% parent or better at the beginning to <80% at the end, with several preceding and trailing peaks around the parent. If I come in the next day, using the same standard solution in the same vial, I see the same pattern – OK at the beginning, garbage at the end. I thought that perhaps it was the test samples contaminating the column, but today I made several injections of the standard by itself, and I saw the same appearance of more and larger non-parent peaks as the day went on, although not as drastic as when running test samples. I have made fresh standards, fresh mobile phases – no help, same pattern. This might be a stupid question, but - is it possible that if the analytical column is not in the best of health, that it could “re-equilibrate” itself simply by sitting overnight, or when I condition it briefly in the morning before a run? Then, as a run progresses, it deteriorates again to the point at which it will no longer properly retain the analyte? Any ideas out there? Thanks much.