Search

Background: Mechanical ventilation injures lungs, but there are currently no reliable methods for detecting early injury. We therefore evaluated whether exhaled pentanal, a lipid peroxidation product, might be a useful breath biomarker for stretch-induced lung injury in rats., Methods: A total of 150 male Sprague-Dawley rats were investigated in 2 substudies. The first randomly assigned 75 rats to 7 hours of mechanical ventilation at tidal volumes of 6, 8, 12, 16, and 20 mL·kg-1. The second included 75 rats. A reference group was ventilated at a tidal volume of 6 mL·kg-1 for 10 hours 4 interventional groups were ventilated at a tidal volume of 6 mL·kg-1 for 1 hour, and then for 0.5, 1, 2, or 3 hours at a tidal volume of 16 mL.kg-1 before returning to a tidal volume of 6 mL·kg-1 for additional 6 hours. Exhaled pentanal was monitored by multicapillary column-ion mobility spectrometry. The first substudy included cytokine and leukocyte measurements in blood and bronchoalveolar fluid, histological assessment of the proportion of alveolar space, and measurements of myeloperoxidase activity in lung tissue. The second substudy included measurements of pentanal in arterial blood plasma, cytokine and leukocyte concentrations in bronchoalveolar fluid, and cleaved caspase 3 in lung tissue., Results: Exhaled pentanal concentrations increased by only 0.5 ppb·h-1 (95% confidence interval [CI], 0.3-0.6) when rats were ventilated at 6 mL·kg-1. In contrast, exhaled pentanal concentrations increased substantially and roughly linearly at higher tidal volumes, up to 3.1 ppb·h-1 (95% CI, 2.3-3.8) at tidal volumes of 20 mL·kg-1. Exhaled pentanal increased at average rates between 1.0 ppb·h-1 (95% CI, 0.3-1.7) and 2.5 ppb·h-1 (95% CI, 1.4-3.6) after the onset of 16 mL·kg-1 tidal volumes and decreased rapidly by a median of 2 ppb (interquartile range [IQR], 0.9-3.2), corresponding to a 38% (IQR, 31-43) reduction when tidal volume returned to 6 mL·kg-1. Tidal volume, inspiratory pressure, and mechanical power were positively associated with pentanal exhalation. Exhaled and plasma pentanal were uncorrelated. Alveolar space decreased and inflammatory markers in bronchoalveolar lavage fluid increased in animals ventilated at high tidal volumes. Short, intermittent ventilation at high tidal volumes for up to 3 hours increased neither inflammatory markers in bronchoalveolar fluid nor the proportion of cleaved caspase 3 in lung tissue., Conclusions: Exhaled pentanal is a potential biomarker for early detection of ventilator-induced lung injury in rats., Competing Interests: The authors declare no conflicts of interest., (Copyright © 2021 International Anesthesia Research Society.)

Fosfomycin is an antibiotic with a broad spectrum of activity against many multidrug-resistant bacterial strains. It is mainly excreted unchanged by the kidneys, and its half-life therefore depends on kidney function which varies considerably among individuals, and within individuals over time. Proper fosfomycin dosing thus depends on assaying blood concentration of the drug. We developed and validated a simple, sensitive and specific chromatography assay, which was coupled to electrospray ionization mass spectrometry for determination of fosfomycin. Separation of fosfomycin was based on the method of the hydrophilic interaction liquid chromatography; specifically, plasma and dialysate samples were acidified and the protein precipitated with acetonitrile. The calibration curves showed excellent coefficients of determination (R2 > 0.999) over the relevant concentration range of 25-700 μg/mL. Intraday precision was 1.1-1.2% and accuracy was -5.9% to 0.9% for quality control samples. Interday precision was 2.9-3.4% and accuracy was -3.7% to 5.5%. Extraction recovery was ≥87% and matrix effects ranged from 2.2% to 4.3%. After laboratory validation, the method was successfully applied to clinical samples., (© The Author(s) 2020. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

The combination of propofol, ketamine and rocuronium can be used for anesthesia of ventilated rats. However, reliable pharmacokinetic models of these drugs have yet to be developed in rats, and consequently optimal infusion strategies are also unknown. Development of pharmacokinetic models requires repeated measurements of drug concentrations. In small animals, samples must be tiny to avoid excessing blood extraction. We therefore developed a drug assay system using high-performance liquid chromatography coupled with quadrupole mass spectrometry that simultaneously determines the concentration of all three drugs in just 10 μL rat plasma. We established a plasma extraction protocol, using acetonitrile as the precipitating reagent. Calibration curves were linear with R 2  = 0.99 for each drug. Mean recovery from plasma was 91-93% for propofol, 89-93% for ketamine and 90-92% for rocuronium. The assay proved to be accurate for propofol 4.1-8.3%, ketamine 1.9-7.8% and rocuronium -3.6-4.7% relative error. The assay was also precise; the intra-day precisions were propofol 2.0-4.0%, ketamine 2.7-2.9% and rocuronium 2.9-3.3% relative standard deviation. Finally, the method was successfully applied to measurement the three drugs in rat plasma samples. Mean plasma concentrations with standard deviations were propofol 2.0 μg/mL ±0.5%, ketamine 3.9 μg/mL ±1.0% and rocuronium 3.2 μg/mL ±0.8% during ventilation., (© 2019 John Wiley & Sons, Ltd.)

Propofol concentration in human plasma can be quantified by liquid chromatography coupled mass spectrometry. Sample preparation usually requires solid phase extraction to remove matrix components and enrich the analyte. To facilitate user-independent measurements and speed extraction, we developed and validated a fully automated high throughput in-line sample preparation system with direct injection into liquid chromatography coupled mass spectrometry. We assessed linearity of each method over the clinically relevant concentration range from 0.5μg/mL to 8μg/mL plasma concentration. R 2 values were 0.99 for the automated process and 0.98 for manual sample preparation. The limit of detection was 6ng/mL and the lower limit of quantification was 18ng/mL for the automated method; for the manual process, the limit of detection was 1.58ng/mL and the lower limit of quantification was 4.79ng/mL. Intra-day precision for low, medium and high concentration range of the automated method was validated 4.14%, 9.68% and 3.04% relative standard deviation and 0.29%, 0.12% and 0.52% for the manual process. Carry over was 0.4% with the automated method, whereas there was no carry over with the manual method. Stability of plasma samples was tested with the manual method at concentrations of 1, 4, and 6μg/mL propofol and found to be stable over 150days at -20°C. The manual sample preparation method has successfully been transferred to a fully automated process with appropriate sensitivity and precision but the automatization failed with regard to trueness and working time due to lengthy sample preparation runtime. Therefore it is not suitable for daily use in a hospital laboratory e.g. for brain death diagnosis in the intensive care unit., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Breath analysis of rats using multi-capillary column ion-mobility spectrometry (MCC-IMS) revealed alterations in acetone and other ketones, including 3-pentanone, during inflammation. The alterations seem likely to result from oxidative branched-chain keto acid (BCKA) catabolism. We therefore tested the hypothesis that 3-pentanone arises during inflammation from increased BCKA oxidation in the liver with consequent accumulation of propionyl-CoA and its condensation products. Male Sprague-Dawley rats were anaesthetised and ventilated for 24 h or until death. Exhaled breath was analysed by MCC-IMS while rats were injected with low and high doses of lipopolysaccharide (LPS), tumour necrosis factor α (TNFα), or vehicle. The exhaled 3-pentanone peak was identified by pure substance measurements. Blood was collected 12 h after treatment for the determination of cytokine concentrations; transcription enzymes for BCKA catabolism and the activity of the BCKA dehydrogenase were analysed in liver tissue by quantitative real-time PCR and western blotting. Exhaled 3-pentanone decreased in all groups, but minimum concentrations with high-dose LPS (0.24  ±  0.31 volts; mean  ±  SD), low-dose TNFα (0.17  ±  0.10 volts) and high-dose TNFα (0.13  ±  0.04 volts) were lower than in vehicle animals (0.27  ±  0.12 volts). At 60% and 85% survival times (svt) concentrations of exhaled 3-pentanone increased significantly in all animals treated with low-dose LPS, (svt 60% 0.38  ±  0.18 volts, svt 85% 0.62  ±  0.15 volts) and high-dose LPS (0.26  ±  0.28 volts, 0.40  ±  0.22 volts), as well as low-dose TNFα, (0.20  ±  0.09 volts, 0.39  ±  0.17 volts) and high-dose TNFα (0.18  ±  0.06 volts, 0.34  ±  0.08 volts), but not in vehicle rats (0.27  ±  0.12 volts, 0.30  ±  0.09 volts). BCKA catabolism was seen in the liver, with increased expression and activity of the branched-chain alpha-keto acid dehydrogenase (BCKD), lower expression of the propionyl-CoA carboxylase (PCC) subunits, and altered expression levels of BCKD regulating enzymes. Exhaled 3-pentanone may arise from altered BCKA catabolism. Our results suggest that excessive propionyl-CoA is possibly generated from BCKAs via increased activity of BCKD, but may undergo unusual condensation reactions rather than being physiologically processed to methylmalonyl-CoA by PCC. The pattern of 3-pentanone during early and prolonged inflammation suggests that reuse of BCKAs for the synthesis of new proteins might be initially favoured. As inflammatory conditions persist, substrates for cellular energy supply are required which activate irreversible degradation of excessive BCKA to propionyl-CoA yielding increased levels of exhaled 3-pentanone.