Cogent Diol™ column is great for biological samples!

Many of you are analyzing blood samples for applications like clinical trials of pharmaceuticals, forensics testing, and metabolomics research. We have had a number of successful separations of these types of samples with the Diamond Hydride™ in ANP mode. You may be interested in seeing how the latest addition to our TYPE-C Silica™ line of HPLC columns, the Cogent Diol 2.o™, would work in these cases. We selected three test solutes that would be pertinent to blood sample testing: warfarin, hydroxybupropion, and codeine. Let’s take a look at each:

1) Warfarin: An anti-coagulant used for treatment of thrombosis. It does not have an amine group like the other two, which makes its retention lower. Warfarin does exhibit keto–enol tautomerism and its keto form is ionizable. In this form, it has a pKa of 5.0 due to an acidic hydrogen located between two electron-withdrawing carbonyl groups. Its retention was observed to be the lowest of the three.

2) Hydroxybupropion: The active metabolite of bupropion, an antidepressant and smoking cessation drug. Its main structural feature that influences retention in the ANP mode is a secondary amine. It eluted next, with baseline separation from codeine.

3) Codeine: An opiate used in many pharmaceutical formulations for its analgesic or antitussive properties. It has a tertiary amine which makes it amenable to ANP retention. Therefore it eluted the latest of the three compounds. The peak shape was highly symmetrical, which can sometimes be difficult to obtain for amines.

               We spiked the analytes in a real blood sample and separated them by LC-MS. Using extracted ion chromatograms, interferents from the complex sample matrix could be eliminated. What you will have left are three sharp, well-separated peaks corresponding to the analytes.

               The data you can obtain shows how ANP chromatography is not limited to the Diamond Hydride™ column. Any TYPE-C™ column can be used in the ANP mode. Another interesting feature we discovered is that acetone can be used instead of acetonitrile in the mobile phase. Acetone has the advantages to your laboratory of lower cost as well as lower toxicity.

              Click here for more information on this application.