Comparison of electrospray and UniSpray™ ion sources with post-column hydrochloric acid infusion for 119 abuse relevant substances

Abstract

Comparing the sensitivity of the Waters UniSpray™ ion source (USI) with the Waters electrospray ion source (ESI) on a Waters Xevo TQ-XS mass spectrometer with and without additional post-column infusion of low concentrated hydrochloric acid (HCl). To reduce sample volumes or increase analytical sensitivity for drugs of abuse detection in human body fluids, technological advances in mass spectrometry often target an increase in ionization efficiency. One example for this development is the USI, which claims higher ion yields and therefore higher sensitivity due to a unique geometry. Another possibility to increase sensitivity through increased ion yield is the post-column infusion of low concentrated HCl. For sensitivity comparison, we choose the, in our opinion, 119 most relevant medications and drugs of abuse. These were prepared in eleven acetonitrile solutions containing 10 analytes each and one with 9 analytes at a concentration of 1 μg/mL for each substance. For analysis, every solution was diluted 1:1000 in mobile phase A/ethylene glycol (70:30 v/v) to a concentration of 1.0 ng/mL. Solutions were analysed in triplicate on the same day interrupted by the ion source change, to decrease inter-day variations in sensitivity. Chromatographic separation was performed on a Waters 2.1 × 150 mm, 1.8 μm BEH Phenyl column kept at 60 °C on an Acquity UPLC connected to a Xevo TQ-XS detector (Waters). The standard ESI ion source or the USI ion source was mounted on the detector. Mobile phase A consisted of 20 mM ammonium formate and 0.1% formic acid (pH 3) and mobile phase B was 0.1% formic acid in methanol. Gradient elution was conducted within 6.0 min at a flow rate of 0.5 mL/min with 85% A and 15% B at the beginning and 100% B at the end. Data were acquired with the ion sources operating in the positive ionization MRM mode. Two transitions for each analyte were monitored at a constant dwell time of 10 ms over the full chromatographic run time. In an additional experiment, both ion sources were compared with additional post-column infusion of 0.0037% HCl at a flow rate of 5.0 μL/min. Injection volume was 2.0 μL in all experiments. USI provided a general gain in signal intensity (peak area): 12% ( n = 14) of the analytes showed a peak area increase up to 2-fold, 85% ( n = 101) increased between 2- and 4-fold and 3% ( n = 4) increased above 4-fold in peak area. Post-column HCl infusion with the ESI source mounted, resulted in an analyte dependent additional gain in signal intensity for 15% ( n = 18) of the analytes ranging from 2- to 21-fold. 78% ( n = 93) ranged from 0.8-fold to 2-fold. In contrast 7% ( n = 8) showed a signal decrease between 0.4-fold and 0.8-fold. The combination of USI and post-column HCl infusion resulted in an increase in peak area from 2-fold up to 62-fold compared to ESI without infusion for 91% (n = 108) of the analytes. 8% ( n = 9) where between 0.8-fold and 2-fold and 1% ( n = 2) ranged from 0.4-fold to 0.8-fold. With USI, a general improvement in analytical sensitivity under identical conditions could be achieved for all of our 119 “favorite analytes” separated in one chromatographic run. Additional post-column infusion of HCl increased sensitivity for many substances. This approach can be used very selectively when applied to certain retention time windows only. Further experiments are needed to understand how different adjustments of ESI and USI and different matrices will affect ionization. USI, especially in combination with post-column HCl infusion, is a promising tool to increase sensitivity in toxicological multi-target LC-MS/MS analysis.