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Anal Chim Acta ; 1177: 338760, 2021 Sep 08.
Artigo em Inglês | MEDLINE | ID: mdl-34482897


Beta-lactam antibiotics are of vital importance for the treatment of infections in a broad range of patients. Although most systemically administered antibiotics will be excreted renally, a fraction will reach the gastro-intestinal tract, affecting the intestinal microbiome by eradicating a wide range of bacterial species while facilitating the growth of antimicrobial-resistant species. A better understanding of the kinetics of beta-lactam antibiotics in the gastro-intestinal tract is essential to study their role in the development of antibiotic resistance in bacteria and to help develop future therapies to prevent damage to, or restore, the intestinal microbiome. Analysis of beta-lactam antibiotics in faeces is particularly challenging due to the heterogeneous nature of the matrix, rapid degradation of some beta-lactam antibiotics in faeces and very strong ion suppression when using mass spectrometry. Sample preparation was optimized using a sequential strategy of experimental designs. It resulted in lyophilization, a MOPS buffer system and the addition of the beta-lactamase inhibitor avibactam to minimize degradation of antibiotics allowing sensitive quantification. The developed liquid chromatography method with high-resolution mass spectrometric detection was successfully validated according to bioanalytical EMA guidelines and had a linear range of 1-200 µg g-1 lyophilized faeces for amoxicillin, piperacillin and meropenem; and 0.5-100 µg g-1 lyophilized faeces for tazobactam. Despite the highly complex and heterogeneous composition of faeces, the accuracy (0.1-15%) and precision (1.7-12.1%) were in line with those obtained for quantification methods of beta-lactam antibiotics in plasma, the golden standard matrix for therapeutic drug monitoring. The applicability of the method was illustrated by successful quantification of piperacillin and tazobactam in faeces from an intensive care unit patient receiving piperacillin/tazobactam in a continuous intravenous infusion. Both piperacillin and tazobactam were still present six days after discontinuation of the therapy.

Amoxicilina , Piperacilina , Antibacterianos , Cromatografia Líquida , Fezes , Humanos , Espectrometria de Massas , Meropeném , Projetos de Pesquisa , Tazobactam
Biomed Chromatogr ; 35(7): e5092, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-33590498


An ultra-high pressure liquid chromatography high-resolution mass spectrometric (UHPLC-HRMS) method was developed for the simultaneous and sensitive quantification of 10 ß-lactam antibiotics (cefepime, meropenem, amoxicillin, cefazolin, benzylpenicillin, ceftazidime, piperacillin, flucloxacillin, cefuroxime and aztreonam), linezolid and ß-lactamase inhibitors tazobactam and clavulanic acid in human plasma. Validation according to the EMA guidelines showed excellent within- and between-run accuracy and precision (i.e. between 1.1 and 8.5%) and high sensitivity (i.e. lower limit of quantification between 0.25 and 1 mg/L). The UHPLC-HRMS method enables a short turnaround time and high sensitivity and needs only a small amount of plasma, allowing appropriate routine therapeutic drug monitoring. The short turnaround time is obtained by speeding up the protocol on multiple levels, i.e. fast and workload-efficient sample preparation (i.e. protein precipitation and dilution), short (4 min) instrument run time, simultaneous measurement of all relevant ß-lactam antibiotics used in the intensive care unit and the use of the same instrument, column and mobile phases as for the other routine methods in our laboratory.

Monitoramento de Medicamentos/métodos , Linezolida/sangue , Inibidores de beta-Lactamases/sangue , beta-Lactamas/sangue , Cromatografia Líquida/métodos , Ensaios de Triagem em Larga Escala , Humanos , Modelos Lineares , Espectrometria de Massas/métodos , Reprodutibilidade dos Testes , Sensibilidade e Especificidade
Expert Rev Anti Infect Ther ; 18(11): 1155-1164, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-32597263


INTRODUCTION: Individualizing antibiotic therapy is paramount to improve clinical outcomes while minimizing the risk of toxicity and antimicrobial therapy. ß-lactam antibiotics are amongst the drugs most commonly prescribed in the Intensive Care Unit (ICU). The pharmacokinetics of ß-lactam antibiotics are profoundly altered in critically ill patients, leading to the failure of standard drug dosing regimens to result in adequate drug concentrations. Therapeutic Drug Monitoring (TDM) of ß-lactam antibiotics is a promising tool to help optimize ß-lactam antibiotic therapy. AREAS COVERED: The rationale behind TDM for ß-lactam antibiotics is explained, as well as some more practical aspects such as when to sample, what concentrations to strive for and how to use it in clinical practice. We also discuss microbiological and analytical considerations, knowledge gaps, and future perspectives of ß-lactam antibiotics TDM in ICU patients. EXPERT OPINION: TDM of ß-lactam antibiotics has been studied intensively in recent years. While TDM may not yet be widely available, and targets need to be further refined, TDM of ß-lactam antibiotics will help to optimize antibiotic therapy in the critically ill patient, as an integrated part of an antimicrobial stewardship program.

Antibacterianos/administração & dosagem , Monitoramento de Medicamentos/métodos , beta-Lactamas/administração & dosagem , Antibacterianos/farmacocinética , Gestão de Antimicrobianos , Cuidados Críticos/métodos , Estado Terminal , Humanos , Unidades de Terapia Intensiva , Medicina de Precisão , beta-Lactamas/farmacocinética
Anal Bioanal Chem ; 407(21): 6345-56, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-25893798


During the last decade, a significant increase in the occurrence of harmful algal blooms (HABs), linked to repetitive cases of shellfish contamination has become a public health concern and therefore, accurate methods to detect marine toxins in different matrices are required. In this study, we developed a method for profiling lipophilic marine microalgal toxins based on ultra-high-performance liquid chromatography coupled to high-resolution Orbitrap mass spectrometry (UHPLC-HR-Orbitrap MS). Extraction of selected toxins (okadaic acid (OA), dinophysistoxin-1 (DTX-1), pectenotoxin-2 (PTX-2), azaspiracid-1 (AZA-1), yessotoxin (YTX) and 13-desmethyl spirolide C (SPX-1)) was optimized using a Plackett-Burman design. Three key algal species, i.e., Prorocentrum lima, Protoceratium reticulatum and Alexandrium ostenfeldii were used to test the extraction efficiency of OA, YTXs and SPXs, respectively. Prorocentrum micans, fortified with certified reference solutions, was used for recovery studies. The quantitative and confirmatory performance of the method was evaluated according to CD 2002/657/EC. Limits of detection and quantification ranged between 0.006 and 0.050 ng mL(-1) and 0.018 to 0.227 ng mL(-1), respectively. The intra-laboratory reproducibility ranged from 6.8 to 11.7 %, repeatability from 6.41 to 11.5 % and mean corrected recoveries from 81.9 to 119.6 %. In addition, algae cultures were retrospectively screened for analogues and metabolites through a homemade database. Using the ToxID software programme, 18 toxin derivates were detected in the extract of three toxin producing microalgae species. In conclusion, the generic extraction and full-scan HRMS approach offers an excellent quantitative performance and simultaneously allows to profile analogues and metabolites of marine toxins in microalgae. Graphical Abstract Optimization of extraction, detection and quantification of lipophilic marine toxins in microalgae by UHPLC-HR Orbitrap MS.

Cromatografia Líquida de Alta Pressão/métodos , Lipídeos/química , Toxinas Marinhas/análise , Espectrometria de Massas/métodos , Microalgas/química , Limite de Detecção , Toxinas Marinhas/química