Quantitation using HRMS: A new tool for rapid, specific and sensitive determination of catecholamines and deconjugated methanephrines metanephrines in urine.

Urinary catecholamines and their methylated metabolites are biochemical indicators of pheochromocytoma, paraganglioma and neuroblastoma. A rapid and precise analytical method based on solid-phase extraction (SPE) and liquid chromatography separation coupled to high-resolution mass spectrometry (LC-HRMS) was developed and validated to measure urinary catecholamines (epinephrine (E), norepinephrine (NorE), dopamine (D)) and total methylated metabolites (normetanephrine (NorMN), metanephrine(MN) and 3-methoxytyramine (3-MT)) in a clinical setting. Results of 51 urine specimens measured using this LC-HRMS method were compared with a liquid chromatography assay with electrochemical detection (LC-EC). Urine samples (200 µL) were spiked with an internal standard solution followed by SPE purification. In the case of total methylated metabolites, urine was hydrolyzed before SPE purification. Separation was achieved on an Acclaim Mixed Mode WCX column, with an 8.5 min runtime. All compounds were detected in electrospray positive ionization mode with a parallel reaction monitoring acquisition and quantified with a linear regression (r2 > 0.998) between 2 and 200 µg/L (10.9-1090; 11.8-1182 nmol/L) for E and NorE respectively and between 10 and 1000 µg/L for others (65.2-6520; 50.7-5070; 54.5-5450 ; 59.8-5980 nmol/L for D, M, NorMN and 3-MT, respectively). Overall imprecision and bias did not exceed 15%. No significant matrix effect was observed. Correlation between the two assays was good except for epinephrine. Epinephrine concentrations measured by LC-EC method were slightly higher than values obtained with LC-HRMS method but without impact on clinical decision. This LC-HRMS assay provides a new tool for simultaneous quantitative catecholamine determination and was successfully applied in routine for the screening or follow up of pheochromocytoma, paraganglioma and neuroblastoma. LC-HRMS method offers significant advantages compared to LC-EC with good sensitivity, an unambiguous analyte determination and high sample throughput.

A simple ultra-high-performance liquid chromatography-high resolution mass spectrometry assay for the simultaneous quantification of 15 antibiotics in plasma.

Antibiotic (ATB) treatment of critically ill patients with pathophysiological injuries remains a challenge due to the constant increase in antimicrobial resistance. Therapeutic drug monitoring (TDM) is advised for ATB dose adjustments to avoid suboptimal concentrations and dose-related adverse effects. Therefore, a single and reliable analytical method for a broad selection of ATBs was developed using a high-resolution mass spectrometry (HRMS) platform for frequent use in intensive care units. An UHPLC assay coupled to high resolution accurate mass acquisition has been developed for the quantification of penicillins (amoxicillin, oxacillin, piperacillin, and ticarcillin), cephalosporines (cefepime, cefotaxime, ceftazidime, and ceftriaxone), carbapenems (ertapenem, imipenem, and meropenem), lincosamide (clindamycin), quinolones (ofloxacin and ciprofloxacin) and tazobactam. Plasma samples (100µL) were spiked with an internal standard solution followed by protein precipitation. Separation was achieved on an Accucore C18 column, which enabled sample analysis every 9min. All compounds were detected in electrospray positive ion mode and quantified with a linear regression between 0.5 and 32mg/L (r2>0.998). Overall precision and accuracy did not exceed 15%. No significant matrix effect was observed for the studied ATBs. Stored stock solutions at -20°C were stable for 6 months, except for amoxicillin and imipenem. Analytes in plasma were stable for 24h under ambient conditions as well as in post-preparation in an autosampler, except for amoxicillin and imipenem. This HRMS assay provides the simultaneous quantification of 15 ATB; it fulfills the usual quality criteria and was successfully applied for routine TDM of ATBs. The method is based on a full scan acquisition, and it would be easy to add other compounds to the present panel in the future, as this assay has already been proven to be efficient for different classes of compounds.