Chromatography Forum

Hello to everyone,

I am facing a problem while quantification of Orotic acid in urine sample. I have prepared a calibration curve of orotic acid by running pure standard's different dilution.The Rt of orotic acid is at 17.280 min.Now when i run derivetized urine sample and when i quantify it ,it shows the Rt of orotic acid at 17.696 min in most cases and sometimes 17.823 min and the quality match shows only 1%.When i library search that peak it is not the orotic acid mass spectrum.This never happened with other compounds.

Please help

Chandra

What is your method of sample preparation? Do we assume correctly GGC/MS? Have you spiked orotic acid at several levels into urine to see if you can see a peak with increasing intensity corresponding the the increasing spiking level? Have you spiked water to see if the compound is recoverable with your method of preparation?

A shift in retention time would indicate that either there is a loading issue affecting retention times or that you are looking at some other compound. The fact that you see a spectrum that differs from that expected when you look at orotic acid indicates that there is something else present. This other thing may be masking the orotic acid or you may be following an unrelated peak.