¹³C labelled internal standards--a solution to minimize ion suppression effects in liquid chromatography-tandem mass spectrometry analyses of drugs in biological samples?

Abstract

Liquid chromatography-tandem mass spectrometry (LC-MS/MS) is frequently used to identify and quantify drugs in human biological samples due to the high selectivity and sensitivity of this technique. However, ion suppression effects caused by co-eluting compounds: drugs, metabolites, matrix components, impurities and degradation products, are a major concern. Stable isotope labelled internal standards (SIL ISs), usually deuterium ((2)H) labelled, are often used to compensate for these effects. In many LC separations the retention times of (2)H labelled ISs and their analogues will differ. Ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) is increasingly being used for bio-analysis. With the better chromatographic resolution provided with sub 2 μm particles, larger separation between analytes and their (2)H labelled analogues can be expected, which might reduce the benefits of the SIL IS. There is a greater difference in physico-chemical properties between hydrogen isotopes than between isotopes of other elements. (13)C, (15)N and (18)O labelled ISs are more similar to their analytes than (2)H labelled ISs and thereby expected to behave more similarly in chromatographic separations. In this study we have investigated the use of (13)C and (2)H labelled ISs for the determination of amphetamine and methamphetamine by UPLC-MS/MS. The (13)C labelled ISs were co eluting with their analytes under different chromatographic conditions while the (2)H labelled ISs and their analytes were slightly separated. An improved ability to compensate for ion suppression effects were observed when the (13)C labelled ISs were used. Furthermore, an UPLC-MS/MS method for determination of amphetamine and methamphetamine in urine using (13)C labelled ISs has been developed and validated. Unfortunately, there are few (13)C labelled ISs commercial available today. If more (13)C labelled ISs become commercial available they may well be the coming solution to minimize ion suppression/enhancement effects in LC-MS/MS analyses of drugs in biological samples.