Studies of the pharmacokinetics of morroniside and loganin in rat after oral administration of raw and wine-processed Corni fructus by UPLC-QqQ-MS/MS

Purpose: To study the pharmacokinetics of morroniside (MR) and loganin (LG) in rats after oral administration of raw and wine-processed Corni fructus by UPLC-QqQ-MS/MS. Methods: Arctiin (AT) was used as internal standard, and the plasma or tissue samples were extracted twice using ethyl acetate. Electrospray ionization (ESI) negative ion mode was used, and the multiple reaction monitoring mode (MRM) was set in acquisition mode. The extraction and detection method is supported by selectivity, sensitivity, precision, accuracy, stability, extraction, recovery and matrix effect. The non-atrioventricular model of das2.0 software was used to calculate the pharmacokinetic parameters. Results: The absorption rate of MR (PTmax=0.092) and LG (PTmax=0.092) in Corni Fructus after wine-processing was faster in rats. The mean residence time was longer, and distribution of MR (PMRT0~t = 0.294) and LG (PMRT0~t = 0.000) in wine-processed Corni Fructus group increased in liver and kidneys. Conclusion: The proposed method has been successfully validated and is suitable for studying the pharmacokinetics of the two analytes in rats.

A bioanalytical screening method for Enterococcus faecalis RNPP-type quorum sensing peptides in murine feces

Background: Bacteria coordinate their behavior as a group via communication with their peers, known as ‘ quorum sensing’. Enterococcus faecalis employs quorum sensing via RNPP-peptides which were not yet reported to be present in mammalian biofluids. Results: Solid phase extraction of murine feces was performed, followed by ultra high performance liquid chromatography (UHPLC–MS/MS) in multiple reaction monitoring (MRM) mode (in total <90 min/sample) for the nine known RNPP peptides. Limits of detection ranged between 0.045 and 52 nM. Adequate identification criteria allowed detection of RNPP quorum sensing peptides in 2/20 wild-type murine feces samples (i.e., cAM373 and cOB1). Conclusion: A fit-for-purpose UHPLC–MS/MS method detected these RNPP peptides in wild-type murine feces samples.

UPLC-MS/MS Method for Simultaneous Determination of 14 Antimicrobials in Human Plasma and Cerebrospinal Fluid: Application to Therapeutic Drug Monitoring

Pharmacokinetics/pharmacodynamics is the foundation for guiding the rational application of antibiotics in clinical practice, so it is necessary to establish quantitative methods for accurate drug concentration determination. This study aimed to develop a rapid and simple ultrahigh-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method for simultaneous quantification of 14 antibiotics (amikacin, etimicin, ceftazidime, cefepime, cefoperazone, ceftriaxone, daptomycin, latamoxef, linezolid, meropenem, biapenem, ampicillin, norvancomycin, and vancomycin) in human plasma and cerebrospinal fluid. Antibiotics were chromatographically separated on a Waters ACQUITY UPLC BEH C18 column (2.1 mm × 50 mm, 1.7 μm) via gradient elution within 3 minutes and were monitored using positive ion fitted with multiple reaction monitoring. The lower limit of quantification was 0.05–2.0 μg·mL−1. The method was verified according to the FDA bioanalysis method validation guidelines, which showed excellent accuracy (from 86.75% to 110.85%) and precision (from 0.46% to 10.97%). At last, this method was successfully applied to therapeutic drug monitoring in 113 patients under antibiotics treatment.

Qualitative analysis of 7- and 8-hydroxyzolpidem and discovery of novel zolpidem metabolites in postmortem urine using liquid chromatography–tandem mass spectrometry

Purpose Zolpidem (ZOL) is a hypnotic sometimes used in drug-facilitated crimes. Understanding ZOL metabolism is important for proving ZOL intake. In this study, we synthesized standards of hydroxyzolpidems with a hydroxy group attached to the pyridine ring and analyzed them to prove their presence in postmortem urine. We also searched for novel ZOL metabolites in the urine sample using liquid chromatography–triple quadrupole mass spectrometry (LC-QqQMS) and liquid chromatography–quadrupole time-of-flight mass spectrometry (LC-QqTOFMS). Methods 7- and 8-Hydroxyzolpidem (7OHZ and 8OHZ, respectively) were synthesized and analyzed using LC-QqQMS. Retention times were compared between the synthetic standards and extracts of postmortem urine. To search for novel ZOL metabolites, first, the urine extract was analyzed with data-dependent acquisition, and the peaks showing the characteristic fragmentation pattern of ZOL were selected. Second, product ion spectra of these peaks at various collision energies were acquired and fragments that could be used for multiple reaction monitoring (MRM) were chosen. Finally, MRM parameters were optimized using the urine extract. These peaks were also analyzed using LC-QqTOFMS. Results The presence of 7OHZ and 8OHZ in urine was confirmed. The highest peak among hydroxyzolpidems was assigned to 7OHZ. The novel metabolites found were zolpidem dihydrodiol and its glucuronides, cysteine adducts of ZOL and dihydro(hydroxy)zolpidem, and glucuronides of hydroxyzolpidems. Conclusions The presence of novel metabolites revealed new metabolic pathways, which involve formation of an epoxide on the pyridine ring as an intermediate.

Eco-Friendly, Simple, Fast, and Sensitive UPLC-MS/MS Method for Determination of Pexidartinib in Plasma and Its Application to Metabolic Stability

Pexidartinib is the first drug approved by the U.S. Food and Drug Administration specifically to treat the rare joint tumor tenosynovial giant cell tumor. In the current study, a validated, selective, and sensitive UPLC-MS/MS assay was developed for the quantitative determination of pexidartinib in plasma samples using gifitinib as an internal standard (IS). Pexidartinib and IS were extracted by liquid-liquid extraction using methyl tert-butyl ether and separated on an acquity BEH C18 column kept at 40 °C using a mobile phase of 0.1% formic acid in acetonitrile: 0.1% formic acid in de-ionized water (70:30). The flow rate was 0.25 mL/min. Multiple reaction monitoring (MRM) was operated in electrospray (ESI)-positive mode at the ion transition of 418.06 > 165.0 for the analyte and 447.09 > 128.0 for the IS. FDA guidance for bioanalytical method validation was followed in method validation. The linearity of the established UPLC-MS/MS assay ranged from 0.5 to 1000 ng/mL with r > 0.999 with a limit of quantitation of 0.5 ng/mL. Moreover, the metabolic stability of pexidartinib in liver microsomes was estimated.

Synovial and Systemic Pharmacokinetics of Florfenicol and PK/PD Integration against Streptococcus suis in Pigs

Florfenicol is a member of the phenicol group, a broad-spectrum antibacterial agent. It has been used for a long time in veterinary medicine, but there are some factors regarding its pharmacokinetic characteristics that have yet to be elucidated. The aim of our study was to describe the pharmacokinetic profile of florfenicol in synovial fluid and plasma of swine after intramuscular (i.m.) administration. In addition, the dosage regimen of treatment of arthritis caused by S. suis was computed for florfenicol using pharmacokinetic/pharmacodynamic (PK/PD) indices. As the first part of our investigation, the pharmacokinetic (PK) parameters of florfenicol were determined in the plasma and synovial fluid of six pigs. Following drug administration (15 mg/kgbw, intramuscularly), blood was drawn at the following times: 10, 20, 30, 40, 50 and 60 min, 2, 3, 4, 5, 6, 7, 8, 12, 24, 48 and 72 h; synovial fluid samples were taken after 1, 2, 3, 4, 6, 8, 12, 24, 48 and 72 h. The concentration of florfenicol was determined by a validated liquid chromatography-mass spectrometry (LC-MS/MS) method via multiple reaction monitoring (MRM) modes. As the second part of our research, minimum inhibitory concentration (MIC) values of florfenicol were determined in 45 S. suis strains isolated from clinical samples collected in Hungary. Furthermore, a strain of S. suis serotype 2 (SS3) was selected, and killing-time curves of different florfenicol concentrations (0.5 µg/mL, 1 µg/mL and 2 µg/mL) were determined against this strain. Peak concentration of the florfenicol was 3.58 ± 1.51 µg/mL in plasma after 1.64 ± 1.74 h, while it was 2.73 ± 1.2 µg/mL in synovial fluid 3.4 ± 1.67 h after administration. The half-life in plasma was found to be 17.24 ± 9.35 h, while in synovial fluid it was 21.01 ± 13.19 h. The area under the curve (AUC24h) value was 54.66 ± 23.34 μg/mL·h for 24 h in plasma and 31.24 ± 6.82 μg/mL·h for 24 h in synovial fluid. The drug clearance scaled by bioavailability (Cl/F) in plasma and synovial fluid was 0.19 ± 0.08 L/h/kg and 0.29 ± 0.08 L/h/kg, respectively. The mean residence time (MRT) in plasma and synovial fluid was 24.0 ± 13.59 h and 27.39 ± 17.16 h, respectively. The steady-state volume of distribution (Vss) in plasma was calculated from Cl/F of 0.19 ± 0.08 L/h/kg, multiplied by MRT of 24.0 ± 13.59 h. For the PK/PD integration, average plasma and synovial fluid concentration of florfenicol was used in a steady-state condition. The obtained MIC50 value of the strains was 2.0 µg/mL, and MIC90 proved to be 16.0 µg/mL. PK/PD integration was performed considering AUC24h/MIC breakpoints that have already been described. This study is the first presentation of the pharmacokinetic behavior of florfenicol in swine synovia as well as a recommendation of extrapolated critical MICs of S. suis for therapeutic success in the treatment of S. suis arthritis in swine, but it should be noted that this requires a different dosage regimen to that used in authorized florfenicol formulations.

Pharmacokinetics Study of Rabdosia Rubescens Drop Pills Based on UPLC-MS/MS

Background: Rabdosia rubescens drop pills have the effects of clearing away heat and toxin, detumescence, relieving pain. Objective: A simple and sensitive method for simultaneous determination of oridonin, ponicidin, and rosmarinic acid in rat plasma was developed based on ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). Methods: Chromatographic separation was performed on a Waters ACQUITY UPLC BEH C18 column (2.1 × 100 mm, 1.7 µm) with a mobile phase consisting of water containing 0.2% formic acid (mobile phase A) and methanol (mobile phase B) at a flow rate of 0.3 mL/min over a total run time of 3.8 min. All analytes were measured with optimized multiple reaction monitoring (MRM) in positive and negative ion ESI mode. Results: The transitions of oridonin, ponicidin, rosmarinic acid, diphenhydramine, and chloramphenicol were 365.3→347.3, 363.3→345.2, 359.0→160.9, 256.0→167.2, and 321.1→151.9, respectively. The linear ranges were 1-256 ng/mL for ponicidin and rosmarinic acid and 2-512 ng/mL for oridonin. The validated method wasstable and reliable. There was no significant difference in the half-life (t1/2) of the three analytes at three doses. The area under the curve (AUC0-t) and peak concentration (Cmax) of the three analytes decreased linearly in each dose range, and the linear correlation R2 of each analyte under the three doses was greater than 0.95. Conclusion: This method was successfully applied to pharmacokinetic studies of oridonin, ponicidin, and rosmarinic acid in rat plasma after intragastric administration of Rabdosia rubescens drop pills.

Simultaneous determination of rivaroxaban and sitagliptin in rat plasma by LC–MS/MS and its application to pharmacokinetic drug-drug interaction study

A sensitive and accurate LC-MS/MS method was developed and validated for the simultaneous quantification of rivaroxaban (RIV) and sitagliptin (SIT) in rat plasma using apixaban as internal standard (IS). An Agilent Eclipse plus C18 column (2.1 × 100 mm, 3.5 µm, Agilent) was used for chromatographic separation with isocratic elution. Multiple reaction monitoring (MRM) using positive-ion ESI mode to monitor ion transitions of m/z 436.8→144.9 for RIV, m/z 407.7→173.8 for SIT, m/z 459.8→442.8 for IS. The procedure of method validation included selectivity, linearity, precision, accuracy, matrix effect, extraction recovery and stability were conducted according to the guidelines of EMA and FDA. The results indicated that no obvious drug-drug interactions occurred might be owing to their differences in metabolic pathways.

A Lactic Acid Bacteria Consortium Impacted the Content of Casein-Derived Biopeptides in Dried Fresh Cheese

This study aimed to define a consortium of lactic acid bacteria (LAB) that will bring added value to dried fresh cheese through specific probiotic properties and the synthesis of bioactive peptides (biopeptides). The designed LAB consortium consisted of three Lactobacillus strains: S-layer carrying Levilactobacillus brevis D6, exopolysaccharides producing Limosilactobacillus fermentum D12 and plantaricin expressing Lactiplantibacillus plantarum D13, and one Enterococcus strain, Enterococcus faecium ZGZA7-10. Chosen autochthonous LAB strains exhibited efficient adherence to the Caco-2 cell line and impacted faecal microbiota biodiversity. The cheese produced by the LAB consortium showed better physicochemical, textural and sensory properties than the cheese produced by a commercial starter culture. Liquid chromatography coupled with matrix-assisted laser desorption/ionization-time of flight tandem mass spectrometry (LC-MALDI-TOF/TOF) showed the presence of 18 specific biopeptides in dried fresh cheeses. Their identification and relative quantification was confirmed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) using multiple reaction monitoring (MRM). The results also showed that their synthesis resulted mainly from β-casein and also α-S1 casein degradation by proteolytic activities of the LAB consortium. The designed LAB consortium enhanced the functional value of the final product through impact on biopeptide concentrations and specific probiotic properties.