A High-throughput HPLC-MS/MS Assay for the Detection, Quantification and Simultaneous Structural Confirmation of 136 Drugs and Metabolites in Human Urine

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Abstract

Background:

Because of its superior sensitivity and specificity, multianalyte high-performance liquid chromatography coupled with tandem mass spectrometry has become the gold standard in clinical toxicology. Although several qualitative and quantitative liquid chromatography coupled with tandem mass spectrometry assays on various mass spectrometry platforms have been described in the literature, most methods either analyze only a limited number of compounds and/or require tedious and time-consuming sample preparation.

Methods:

A major challenge in urine toxicology screening is the minimization of false-negative and false-positive results. This was addressed by screening for a comprehensive panel of 136 compounds of importance for pain and drug addiction clinics, using high-end, high-sensitivity, fast-scanning mass spectrometry in combination with simultaneous structural confirmation based on ion ratios. The assay was validated and successfully participated in proficiency challenges.

Results:

The assay met all predefined acceptance criteria. The lower limit of quantifications ranged from 10 to 100 ng/mL. Interday trueness and imprecisions ranged from 73.8% to 116.2% and 2.4%–20.0%, respectively. The total assay run time was 10 minutes.

Conclusions:

We developed and successfully validated a robust, sensitive, and specific liquid chromatography coupled with tandem mass spectrometry-based urine toxicology screening platform that allows for the addition of drugs to quickly adjust to new clinical needs. To date, more than 3000 clinical samples have successfully been analyzed. Our results also indicated that such a quantitative multianalyte assay is pushing against the limits of current fast-scanning MS/MS instrumentation and that the number of analytes and their internal standards must be balanced with acceptable sensitivity, reproducibility, structural confirmation, and the ability to reliably quantify, all of which was achieved.

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