Enhancing detectability of anabolic-steroid residues in bovine urine by actively modulated online comprehensive two-dimensional liquid chromatography – high-resolution mass spectrometry

Abstract

In this study we describe an approach to enhance the sensitivity of an online comprehensive two-dimensional liquid chromatography (LC × LC) high-resolution mass spectrometry method for the separation and detection of trace levels of anabolic-steroid residues in complex urine matrices. Compared to one-dimensional liquid chromatography (1D-LC), LC × LC methods offer higher separation power, thanks to the combined effect of two different selectivities and a higher peak capacity. However, when using state-of-the-art LC × LC instrumentation, the price paid for the increase in separation power is a decrease in sensitivity and detectability of trace-level analytes. This can be ascribed to the sample dilution that takes place during each of the two chromatographic steps. The way in which fractions are collected and transferred from the first to the second column is also of paramount importance, especially the volume and the solvent composition of the fractions injected in the second column. To overcome the detection limitation, we present an active-modulation strategy, based on concentrating the fractions of the first-dimension effluent using a modulation interface that employs trap columns. We obtained a signal enhancement for anabolic-steroid compounds in a bovine-urine sample by a factor of 2.4-7.6 and an increase in the signal-to-noise ratio up to a factor of 7 in comparison with a standard loop-based modulation interface. In addition, thanks to the increased sensitivity of our method, a substantially larger number of peaks were detected (76 vs. 36). Moreover, we could reduce the solvent consumption by a factor of three (160 mL vs. 500 mL per run).

Advances in the Analysis of Veterinary Drug Residues in Food Matrices by Capillary Electrophoresis Techniques

“…In addition, there is a lack of regulation for veterinary drug residues (for example, for fluoroquinolones used as antimicrobials) in many foods, including baby foods [7]. EU guidelines suggest the use of the liquid chromatography (LC) technique; in particular, LC coupled with mass spectrometry (MS) was the most used approach to detect and analyze drug residues in complex matrices, such as milk and dietary products [8][9][10][11][12]. In addition, ion mobility spectrometry (IMS) coupled with MS represented a very promising powerful tool to detect analytes in traces [13].…”

Recent Developments in Two-Dimensional Liquid Chromatography: Fundamental Improvements for Practical Applications

“…76 analytes could be detected using SPAM due to the improvement in S/N, relative to 36 using passive modulation. 41 Reprinted from Anal. Chem.…”

“…; Schoenmakers, P.J., Enhancing detectability of anabolic-steroid residues in bovine urine by actively modulated online comprehensive two-dimensional liquid chromatography–high-resolution mass spectrometry, pp. 87–97 (ref (41)). Copyright (2018), with permission from Elsevier.…”

Hydrophilic Interaction Liquid Chromatography for LC-MS

“…Carbohydrates and other related compounds, which are difficult to detect with high sensitivity using other analytical instruments, have been subjected to LC-MS analysis [13,[23][24][25][26][27]. Furthermore, LC-MS analysis has been applied for diagnosis [12,[28][29][30] including biological amines [31][32][33][34], endogenous carboxylic acids [35,36], and lipids [13,[37][38][39]. LC-MS is also applied to toxicological analysis in the environment [40][41][42][43][44][45][46].…”