Semi-automated serum steroid profiling with tandem mass spectrometry.

Objectives: Highly selective and sensitive multi-analyte methods for the analysis of steroids are attractive for the diagnosis of endocrine diseases. Commercially available kits are increasingly used for this purpose. These methods involve laborious solid phase extraction, and the respective panels of target analytes are incomplete. We wanted to investigate whether an improvement of kit solutions is possible by introducing automated on-line solid phase extraction (SPE) and combining originally separate analyte panels. Methods: Sample preparation was performed using automated on-line SPE on a high-pressure stable extraction column. Chromatographic separation, including isobaric compounds, was achieved using a 0.25 mM ammonium fluoride-methanol gradient on a small particle size biphenyl column. Standard compounds and internal standard mixtures of two panels of a commercially available kit were combined to achieve an optimized and straightforward detection of 15 endogenous steroids. Validation was performed according to the European Medicines Agency (EMA) guidelines with slight modifications. Results: Validation was successfully performed for all steroids over a clinically relevant calibration range. Deviations of intra- and inter-assay accuracy and precision results passed the criteria and no relevant matrix effects were detected due to highly effective sample preparation. External quality assessment samples showed the applicability as a routine diagnostic method, which was affirmed by the analyses of anonymized clinical samples. Conclusions: It was found possible to complement a commercially available kit for quantitative serum steroid profiling based on isotope dilution LC-MS/MS by implementing automated on-line SPE, thereby improving the practicality and robustness of the measurement procedure.

MALDI mass spectrometry imaging: From constituents in fresh food to ingredients, contaminants and additives in processed food.

Mass Spectrometry imaging (MS imaging) provides spatial information for a wide range of compound classes in different sample matrices. We used MS imaging to investigate the distribution of components in fresh and processed food, including meat, dairy and bakery products. The MS imaging workflow was optimized to cater to the specific properties and challenges of the individual samples. We successfully detected highly nonpolar and polar constituents such as beta-carotene and anthocyanins, respectively. For the first time, the distributions of a contaminant and a food additive were visualized in processed food. We detected acrylamide in German gingerbread and investigated the penetration of the preservative natamycin into cheese. For this purpose, a new data analysis tool was developed to study the penetration of analytes from uneven surfaces. Our results show that MS imaging has great potential in food analysis to provide relevant information about components' distributions, particularly those underlying official regulations.

The Role of Non-Enzymatic Degradation of Meropenem-Insights from the Bottle to the Body.

Several studies have addressed the poor stability of meropenem in aqueous solutions, though not considering the main degradation product, the open-ring metabolite (ORM) form. In the present work, we elucidate the metabolic fate of meropenem and ORM from continuous infusion to the human bloodstream. We performed in vitro infusate stability tests at ambient temperature with 2% meropenem reconstituted in 0.9% normal saline, and body temperature warmed buffered human serum with 2, 10, and 50 mg/L meropenem, covering the therapeutic range. We also examined meropenem and ORM levels over several days in six critically ill patients receiving continuous infusions. Meropenem exhibited a constant degradation rate of 0.006/h and 0.025/h in normal saline at 22 °C and serum at 37 °C, respectively. Given that 2% meropenem remains stable for 17.5 h in normal saline (≥90% of the initial concentration), we recommend replacement of the infusate every 12 h. Our patients showed inter-individually highly variable, but intra-individually constant molar ORM/(meropenem + ORM) ratios of 0.21-0.52. Applying a population pharmacokinetic approach using the degradation rate in serum, spontaneous degradation accounted for only 6% of the total clearance.

An isotope-dilution LC-MS/MS method for the simultaneous quantification of meropenem and its open-ring metabolite in serum.

BACKGROUND: Therapeutic drug monitoring (TDM) of beta-lactam antibiotics and, among them, especially meropenem gains importance in the field of laboratory medicine. Meropenem is known to be unstable, resulting in a degradation product with an open beta-lactam ring. For a more comprehensive TDM of meropenem, the aim was to develop a LC-MS/MS method for the simultaneous quantification of meropenem and its main degradation product, the open-ring metabolite (ORM). METHODS: The method involves a protein precipitation followed by chromatographic separation using a formic acid-ammonium formate methanol gradient on a pentafluorophenyl column. Multiple reaction monitoring in the positive ion mode and stable isotope labeled internal standards were used for quantification. Validation was performed according to the European Medicines Agency guideline. RESULTS: Validation was successful performed within the linear drug concentration range of 1.0-100.0 mg/l for meropenem and 0.62-62.30 mg/l for the ORM. Investigation of selectivity, accuracy and precision showed good results and potential matrix effects were successfully compensated by the internal standards. The suitability of the method was shown by the comparison of 35 anonymized leftover serum samples from intensive care patients with routine analyses. CONCLUSION: For the first time, we herein describe a liquid chromatography tandem mass spectrometry (LC-MS/MS) method for the simultaneous quantification of meropenem and its ORM in human serum. The ratio of active to inactive compound provides valuable pharmaceutical and pharmacokinetic information, which may contribute to therapeutic efficacy.