A novel approach to simultaneous screening and confirmation of regulated pharmaceutical compounds in dietary supplements by LC/MS/MS with an information-dependent acquisition method

Abstract

The commercial success of synthetic phosphodiesterase-5 (PDE-5) inhibitors (viz. sildenafil, vardenafil and tadalafil) for erectile dysfunction (ED) has led to their widespread use as adulterants in dietary supplements (DSs). Reports on adulteration by ED drugs or their analogues in DSs suggest they may cause a serious threat to human health. The problem is becoming more complex as hidden and structurally modified analogues are continuously being reported. To analyse known drugs and their analogues, three commonly used PDE-5 inhibitors, naturally existing icariin and yohimbin, and their 19 analogues were analyzed in this study. They were identified using ion-spray liquid chromatography/tandem mass spectrometry (LC/MS/MS) using multiple reaction monitoring (MRM). This MRM procedure gave a limit of detection of less than 0.02 ng ml⁻¹ for the 24 compounds, selectivity of fragmentation using MRM for 2.5 – 8.5 min in a single run and peak height repeatability of coefficient of variation of 3.9 – 31.8%. An IDA method using the MRM scans to detect the presence of known analytes was set up and added to a built-in library for screening for PDE-5 inhibitors. These MRM experiments were used to trigger product ion scans using a hybrid quadrupole-linear ion trap instrument. The product ion scan was compared and confirmed by a library search of MS/MS spectra acquired from a reference standard. To search for new analogues of PDE-5 inhibitors, a precursor ion scan of an expected ion m/z 283, which was one of the mass fragments from the analogues of sildenafil or vardenafil, was performed and fragmentation of the precursor ion, by combining a precursor ion scan with automatic confirmation using EPI spectra, was acquired. Of the 37 DSs tested, two were eventually found to be adulterated with yohimbin and vardenafil, respectively. The approach proposed in this study would be valuable in characterizing chemical constituents of drug residues and their analogues with identical chemical substructures from complex natural and synthetic sources in DSs using an information-dependent acquisition-enhanced product ion (IDA–EPI) scan.

Keywords:

• chromatography, LC/MS
• LC/MS
• residues
• trace elements
• dietary supplements

Acknowledgements

This work was supported partly by the National Research Foundation of South Korea (NRF) grant funded by the South Korea Government (MEST) (No. 20100001707) and the South Korea Healthcare Technology R&D Project, Ministry for Health (A092006). The authors would like to thank the South Korea Food and Drug Administration and Korea Applied Biosystems for the providing facilities.