Determination of camostat and its metabolites in human plasma - Preservation of samples and quantification by a validated UHPLC-MS/MS method.

OBJECTIVES: Camostat mesilate is a drug that is being repurposed for new applications such as that against COVID-19 and prostate cancer. This induces a need for the development of an analytical method for the quantification of camostat and its metabolites in plasma samples. Camostat is, however, very unstable in whole blood and plasma due to its two ester bonds. The molecule is readily hydrolysed by esterases to 4-(4-guanidinobenzoyloxy)phenylacetic acid (GBPA) and further to 4-guanidinobenzoic acid (GBA). For reliable quantification of camostat, a technique is required that can instantly inhibit esterases when blood samples are collected. DESIGN AND METHODS: An ultra-high-performance liquid chromatography-tandem mass spectrometry method (UHPLC-ESI-MS/MS) using stable isotopically labelled analogues as internal standards was developed and validated. Different esterase inhibitors were tested for their ability to stop the hydrolysis of camostat ester bonds. RESULTS: Both diisopropylfluorophosphate (DFP) and paraoxon were discovered as efficient inhibitors of camostat metabolism at 10 mM concentrations. No significant changes in camostat and GBPA concentrations were observed in fluoride-citrate-DFP/paraoxon-preserved plasma after 24 h of storage at room temperature or 4 months of storage at -20 °C and -80 °C. The lower limits of quantification were 0.1 ng/mL for camostat and GBPA and 0.2 ng/mL for GBA. The mean true extraction recoveries were greater than 90%. The relative intra-laboratory reproducibility standard deviations were at a maximum of 8% at concentrations of 1-800 ng/mL. The trueness expressed as the relative bias of the test results was within ±3% at concentrations of 1-800 ng/mL. CONCLUSIONS: A methodology was developed that preserves camostat and GBPA in plasma samples and provides accurate and sensitive quantification of camostat, GBPA and GBA by UHPLC-MS/MS.

Effect of chronic exercise in healthy young male adults: a metabolomic analysis.

AIMS: To examine the metabolic adaptation to an 80-day exercise intervention in healthy young male adults where lifestyle factors such as diet, sleep, and physical activities are controlled. METHODS AND RESULTS: This study involved cross-sectional analysis before and after an 80-day aerobic and strength exercise intervention in 52 young, adult, male, newly enlisted soldiers in 2015. Plasma metabolomic analyses were performed using liquid chromatography, tandem mass spectrometry. Data analyses were performed between March and August 2019. We analysed changes in metabolomic profiles at the end of an 80-day exercise intervention compared to baseline, and the association of metabolite changes with changes in clinical parameters. Global metabolism was dramatically shifted after the exercise training programme. Fatty acids and ketone body substrates, key fuels used by exercising muscle, were dramatically decreased in plasma in response to increased aerobic fitness. There were highly significant changes across many classes of metabolic substrates including lipids, ketone bodies, arginine metabolites, endocannabinoids, nucleotides, markers of proteolysis, products of fatty acid oxidation, microbiome-derived metabolites, markers of redox stress, and substrates of coagulation. For statistical analyses, a paired t-test was used and Bonferroni-adjusted P-value of <0.0004 was considered to be statistically significant. The metabolite dimethylguanidino valeric acid (DMGV) (recently shown to predict lack of metabolic response to exercise) tracked maladaptive metabolic changes to exercise; those with increases in DMGV levels had increases in several cardiovascular risk factors; changes in DMGV levels were significantly positively correlated with increases in body fat (P = 0.049), total and LDL cholesterol (P = 0.003 and P = 0.007), and systolic blood pressure (P = 0.006). This study was approved by the Departments of Defence and Veterans' Affairs Human Research Ethics Committee and written informed consent was obtained from each subject. CONCLUSION: For the first time, the true magnitude and extent of metabolic adaptation to chronic exercise training are revealed in this carefully designed study, which can be leveraged for novel therapeutic strategies in cardiometabolic disease. Extending the recent report of DMGV's predictive utility in sedentary, overweight individuals, we found that it is also a useful marker of poor metabolic response to exercise in young, healthy, fit males.

Pharmacokinetic behavior of peramivir in the plasma and lungs of rats after trans-nasal aerosol inhalation and intravenous injection.

Peramivir, a neuraminidase inhibitor, was approved globally and is indicated for the treatment of uncomplicated influenza in adults and children. However, the only approved intravenous formulation of peramivir limits its clinical application due to the need for the specialized dosing techniques and increases the risk of contracting influenza virus infection among healthcare professionals when dosing within a short distance to the patient. The purpose of this study was to investigate the pharmacokinetic profile of peramivir in plasma and the lung of rats and to compare the profiles following administration through trans-nasal aerosol inhalation (0.0888, 0.1776, and 0.3552 mg/kg) and intravenous injection (30 mg/kg). The plasma concentration reached the Cmax within 1.0 h (upon inhalation) and decreased at a t1/2 of 6.71 and 10.9 h after inhalation and injection, respectively. The absolute bioavailability of peramivir after inhalation was 78.2 %. Overall, the pharmacokinetic exposure of peramivir in the lungs was higher than that in the plasma after aerosol inhalation. After inhalation, the Cmax of peramivir in the lung was achieved within 1.0 h, and the elimination of the drug was slower than in the case of intravenous injection with t1/2 values 1.81 h for injection and 5.72, 53.5, and 32.1 h for low, middle, and high doses administered through inhalation. The Cmax and AUC0-t values for peramivir in the lungs increased linearly with the increased inhalation dose. The results elucidate the pharmacokinetic process of peramivir after trans-nasal aerosol inhalation to rats and provide useful information for further rational application of this drug formulation.

Combined exposure to dinotefuran and chronic mild stress counteracts the change of the emotional and monoaminergic neuronal activity induced by either exposure singly despite corticosterone elevation in mice.

Dinotefuran (DIN) belongs to the neonicotinoids (NNs), a class of globally applied pesticides originally developed to exhibit selective toxicity in insects. However, several reports have suggested that NNs also exert neurotoxic effects in mammals. We previously demonstrated neurobehavioral effects of DIN on mice under non-stressful conditions. For further toxicity assessments in the present study, we investigated the effects of DIN on mice exposed to stressful conditions. After subacutely administering a no-observed-effect-level (NOEL) dose of DIN and/or chronic unpredictable mild stress (CUMS) to mice, we conducted three behavioral tests (i.e., open field test [OFT], tail suspension test [TST] and forced swimming test [FST]). In addition, serotonin (5-HT) and tryptophan hydroxylase 2 (TPH2) of the dorsal raphe nuclei (DRN) and median raphe nuclei (MRN) and tyrosine hydroxylase (TH) of the ventral tegmental area and substantia nigra (SN) were evaluated immunohistochemically. A NOEL dose of DIN or CUMS alone increased of the total distance in OFT, decreased or increased the immobility time in TST or FST, respectively, and increased the positive intensity of 5-HT and TPH2 in the DRN/MRN, and TH in the SN. These changes were suppressed under the conditions of combined exposure to DIN and CUMS, though the blood corticosterone level was increased depending on the blood DIN values and the presence of CUMS. The present study suggests the multifaceted toxicity of the neurotoxin DIN.

Quantitative elucidation of maternal-to-fetal transfer of neonicotinoid pesticide clothianidin and its metabolites in mice.

Neonicotinoids (NNs), a widely used class of systemic pesticides, are regarded as exhibiting selective toxicity in insects. However, NNs are suspected of exerting adverse effects on mammals as well, including humans. To date, only adult male animal models have been subjected to general toxicity studies of NNs; fetuses have yet to be considered in this context. Here, we focused on the NN clothianidin (CLO) for the first quantitative LC-MS/MS analysis of maternal-to-fetal transfer and residual property of once-daily (single or multiple days), orally administered CLO and its metabolites in mice. The results revealed the presence of CLO and its five metabolites at approximately the same respective blood levels in both dams and fetuses. In the dams, CLO showed a peak value 1 h after administration, after which levels rapidly decreased at 3 and 6 h. In the fetuses of each group, levels of CLO were almost the same as those observed in the corresponding dams. The present results clearly demonstrated rapid passage of CLO through the placental barrier. However, metabolite-dependent differences observed in blood pharmacokinetics and residual levels. This is the first quantitative demonstration of the presence of CLO and its metabolites in fetal mouse blood.

Bioanalytical method development and validation of MES207, a neuropeptide FF receptor antagonist, and its application in preclinical pharmacokinetics.

The nonpeptide small molecule, MES207, exhibits 17-fold preferential binding to the neuropeptide FF receptor 1 (NPFFR1) over NPFFR2 and shows antagonist functionality at NPFF receptors. In order to further the development of MES207 as a NPFFR1 probe, an UPLC-MS/MS bioanalytical method was developed and validated to quantify MES207 in rat plasma for a linearity range of 3-200 ng/mL. The method was applied in the analysis of the plasma, brain, and urine samples collected during pharmacokinetic studies in healthy male and female Sprague Dawley rats. The animals were dosed through oral gavage (50 mg/kg) and intravenously (2.5 mg/kg). Test samples were analyzed using the validated bioanalytical method to generate plasma concentration-time profiles. The results were further subjected to non-compartmental analysis using Phoenix 6.4®. MES207 exhibits a large volume of distribution (1.2 ±â€¯0.6 L), high clearance (0.8 ±â€¯0.1 L/h), and a poor oral bioavailability (1.7 ±â€¯0.4%). The compound also showed a multiple peak phenomenon with a very short absorption phase. It appears that gender does not significantly influence the differences in pharmacokinetic parameters of this NPFF probe.

Dimethylguanidino Valerate: A Lifestyle-Related Metabolite Associated With Future Coronary Artery Disease and Cardiovascular Mortality.

Background Identification of lifestyle modifiable metabolic pathways related to cardiometabolic disease risk is essential for improvement of primary prevention in susceptible individuals. It was recently shown that plasma dimethylguanidino valerate (DMGV) levels are associated with incident type 2 diabetes mellitus. Our aims were to investigate whether plasma DMGV is related to risk of future coronary artery disease and with cardiovascular mortality and to replicate the association with type 2 diabetes mellitus and pinpoint candidate lifestyle interventions susceptible to modulate DMGV levels. Methods and Results Plasma DMGV levels were measured using liquid chromatography-mass spectrometry in a total of 5768 participants from the MDC (Malmö Diet and Cancer Study-Cardiovascular Cohort), MPP (Malmö Preventive Project), and MOS (Malmö Offspring Study). Dietary intake assessment was performed in the MOS. Baseline levels of DMGV associated with incident coronary artery disease in both the MDC (hazard ratio=1.29; CI=1.16-1.43; P<0.001) and MPP (odds ratio=1.25; CI=1.08-1.44; P=2.4e-3). In the MDC, DMGV was associated with cardiovascular mortality and incident coronary artery disease, independently of traditional risk factors. Furthermore, the association between DMGV and incident type 2 diabetes mellitus was replicated in both the MDC (hazard ratio=1.83; CI=1.63-2.05; P<0.001) and MPP (odds ratio=1.65; CI=1.38-1.98; P<0.001). Intake of sugar-sweetened beverages was associated with increased levels of DMGV, whereas intake of vegetables and level of physical activity was associated with lower DMGV. Conclusions We discovered novel independent associations between plasma DMGV and incident coronary artery disease and cardiovascular mortality, while replicating the previously reported association with incident type 2 diabetes mellitus. Additionally, strong associations with sugar-sweetened beverages, vegetable intake, and physical activity suggest the potential to modify DMGV levels using lifestyle interventions.

HPLC-MS/MS analysis of peramivir in rat plasma: Elimination of matrix effect using the phospholipid-removal solid-phase extraction method.

A simple HPLC-MS/MS method has been developed for the determination of peramivir in rat plasma in the present study. The analytes were separated on a C18 column (50 × 2.1 mm, 1.7 µm) and a triple-quadrupole mass spectrometer equipped with an electrospray ionization source was applied for the detection. A phospholipid-free cartridge solid-phase extraction was used to pretreat the plasma and eliminate the endogenous phospholipid. The in-source collision-induced dissociation approach showed that this pretreatment could result in negligible ion suppression from the extracted sample and could produce cleaner samples when compared with the protein precipitation. The method was linear over the concentration range of 0.12-1200.0 ng/mL for peramivir. The method was validated and successfully applied to a pharmacokinetic study after peramivir was orally and intravenously administered to Sprague-Dawley rats.

Dimethylguanidino valeric acid is a marker of liver fat and predicts diabetes.

Unbiased, "nontargeted" metabolite profiling techniques hold considerable promise for biomarker and pathway discovery, in spite of the lack of successful applications to human disease. By integrating nontargeted metabolomics, genetics, and detailed human phenotyping, we identified dimethylguanidino valeric acid (DMGV) as an independent biomarker of CT-defined nonalcoholic fatty liver disease (NAFLD) in the offspring cohort of the Framingham Heart Study (FHS) participants. We verified the relationship between DMGV and early hepatic pathology. Specifically, plasma DMGV levels were correlated with biopsy-proven nonalcoholic steatohepatitis (NASH) in a hospital cohort of individuals undergoing gastric bypass surgery, and DMGV levels fell in parallel with improvements in post-procedure cardiometabolic parameters. Further, baseline DMGV levels independently predicted future diabetes up to 12 years before disease onset in 3 distinct human cohorts. Finally, we provide all metabolite peak data consisting of known and unidentified peaks, genetics, and key metabolic parameters as a publicly available resource for investigations in cardiometabolic diseases.

The acute effect of beta-guanidinopropionic acid versus creatine or placebo in healthy men (ABC-Trial): A randomized controlled first-in-human trial.

AIMS: Increasing evidence indicates that the ATP-generating enzyme creatine kinase (CK) is involved in hypertension. CK rapidly regenerates ATP from creatine phosphate and ADP. Recently, it has been shown that beta-guanidinopropionic acid (GPA), a kidney-synthesized creatine analogue and competitive CK inhibitor, reduced blood pressure in spontaneously hypertensive rats. To further develop the substance as a potential blood pressure-lowering agent, we assessed the tolerability of a sub-therapeutic GPA dose in healthy men. METHODS: In this active and placebo-controlled, triple-blind, single-centre trial, we recruited 24 healthy men (18-50 years old, BMI 18.5-29.9 kg m-2 ) in the Netherlands. Participants were randomized (1:1:1) to one week daily oral administration of GPA 100 mg, creatine 5 g, or matching placebo. The primary outcome was the tolerability of GPA, in an intent-to-treat analysis. RESULTS: Twenty-four randomized participants received the allocated intervention and 23 completed the study. One participant in the placebo arm dropped out for personal reasons. GPA was well tolerated, without serious or severe adverse events. No abnormalities were reported with GPA use in clinical safety parameters, including physical examination, laboratory studies, or 12-Lead ECG. At day 8, mean plasma GPA was 213.88 (SE 0.07) in the GPA arm vs. 32.75 (0.00) nmol l-1 in the placebo arm, a mean difference of 181.13 (95% CI 26.53-335.72). CONCLUSION: In this first-in-human trial, low-dose GPA was safe and well-tolerated when used during 1 week in healthy men. Subsequent studies should focus on human pharmacokinetic and pharmacodynamic assessments with different doses.

[18F]Fluoro-Hydroxyphenethylguanidines: Efficient Synthesis and Comparison of Two Structural Isomers as Radiotracers of Cardiac Sympathetic Innervation.

Fluorine-18 labeled phenethylguanidines are currently under development in our laboratory as radiotracers for quantifying regional cardiac sympathetic nerve density using PET imaging techniques. In this study, we report an efficient synthesis of 18F-hydroxyphenethylguanidines consisting of nucleophilic aromatic [18F]fluorination of a protected diaryliodonium salt precursor followed by a single deprotection step to afford the desired radiolabeled compound. This approach has been shown to reliably produce 4-[18F]fluoro-m-hydroxyphenethylguanidine ([18F]4F-MHPG, [18F]1) and its structural isomer 3-[18F]fluoro-p-hydroxyphenethylguanidine ([18F]3F-PHPG, [18F]2) with good radiochemical yields. Preclinical evaluations of [18F]2 in nonhuman primates were performed to compare its imaging properties, metabolism, and myocardial kinetics with those obtained previously with [18F]1. The results of these studies have demonstrated that [18F]2 exhibits imaging properties comparable to those of [18F]1. Myocardial tracer kinetic analysis of each tracer provides quantitative metrics of cardiac sympathetic nerve density. Based on these findings, first-in-human PET studies with [18F]1 and [18F]2 are currently in progress to assess their ability to accurately measure regional cardiac sympathetic denervation in patients with heart disease, with the ultimate goal of selecting a lead compound for further clinical development.

A simple and selective UHPLC-MS/MS method for quantification of plantagoguanidinic acid in rat plasma and its application to a pharmacokinetic study.

A simple, sensitive and specific UHPLC-MS/MS method for quantification of plantagoguanidinic acid (PGA) in rat plasma was applied to investigate the pharmacokinetic behavior in vivo, using protopine as internal standard. The chromatography was separated on a Phenomenex® Luna-C18 column (2.1 × 150 mm, 3.0 µm) within 7.0 min using a mobile phase consisting of acetonitrile-0.1% formic acid solution under gradient elution at a flow rate of 0.4 mL/min. Prepared samples were monitored by multiple reaction monitoring mode, with the target fragmentions m/z 226.2 → 84.2 for PGA and m/z 354.2 → 188.9 for IS in positive electrospray ionization. The calibration curve of PGA was linear throughout the range 1-1000 ng/mL (r = 0.9962). The lower limit of quantitation in plasma for PGA was 0.1 ng/mL, and the recovery was >88.6%. Intra- and interday accuracy ranged from -8.6 to 4.9%. Furthermore, this validated method was successfully used for a pre-clinical pharmacokinetic study of PGA at a single dose of 20 and 5 mg/kg in rats via oral and intravenous administration. The study showed that PGA was absorpted rapidly and eliminated gradually with a greater absolute oral bioavailability of 70.1% in rats.

Pharmacokinetics and safety of intravenous peramivir, neuraminidase inhibitor of influenza virus, in healthy Japanese subjects.

BACKGROUND: Intravenous peramivir is a potent neuraminidase (NA) inhibitor with activity against influenza A and B viruses. The early use of NA inhibitors has been shown to reduce mortality in influenza patients. METHODS: To evaluate the pharmacokinetics of peramivir and confirm the safety and tolerability of multiple infusions of peramivir in healthy Japanese subjects, two Phase I, single-centre, randomized, double-blind and placebo-controlled studies consisting of a multiple-dose study and a high-dose study were conducted. RESULTS: Multiple intravenous infusions of peramivir were well tolerated up to 800 mg once a day and 400 mg twice daily for 6 days. Dose proportionalities for maximum plasma concentration (Cmax) and area under the plasma concentration-time curve (AUC) were established up to the 800 mg dose. Approximately 90% of unchanged peramivir was excreted into urine within 12 h after treatment with 800 mg of peramivir. The peramivir plasma and upper respiratory tract fluid levels were significantly higher than the 50% inhibition concentrations for NA enzyme activity (IC50) of epidemic influenza viruses, including those harbouring the H274Y mutation. CONCLUSIONS: The pharmacokinetic properties obtained here for intravenous peramivir are consistent with the previously reported clinical efficacy and safety of this antiviral.

Evaluation of the safety and efficacy of TY-51924 in patients with ST elevated acute myocardial infarction - Early phase II first in patient pilot study.

BACKGROUND: In myocardial ischemia-reperfusion injuries, the involvement of the Na(+)/H(+) exchanger (NHE) is considered to be one of the pathogenic mechanisms following reperfusion. TY-51924 is a novel hydrophilic NHE inhibitor with a lower risk of central neurotoxicity than previous NHE inhibitors. This open-label, dose-escalating study was undertaken to investigate the safety, efficacy, and pharmacokinetics of TY-51924 in patients with ST-elevation myocardial infarction (STEMI). METHODS: Consent was obtained from a total of 30 patients with first anterior STEMI. After 12 patients were determined to be ineligible, the remaining 18 patients, each of whom was undergoing primary percutaneous coronary intervention (pPCI), received TY-51924 intravenously up to 10, 20, or 30mg/kg as the low-, medium-, or high-dose groups, respectively (n=6 in each group). The primary endpoints were safety (up to 7 days) and plasma drug concentration. The myocardial salvage index (MSI) was measured by (201)Tl/(123)I-beta-methyl-p-iodophenyl pentadecanoic acid single photon emission computed tomography (SPECT) 3-5 days after pPCI. RESULTS: No side effects were observed. Plasma drug concentrations increased dose-dependently, and were subsequently eliminated rapidly. MSIs were 0.118, 0.335, and 0.192 in the low-, medium-, and high-dose groups, respectively. In additional analysis, the combined MSIs in the medium- and high-dose groups were significantly higher than those in the low-dose group, in patients with a longer time from symptom onset to reperfusion (p=0.0247). CONCLUSIONS: No side effects were observed even at the highest dose with this novel hydrophilic NHE inhibitor. Therefore, TY-51924 is thought to be safe in patients with STEMI, even at the highest dose. Potential cardioprotective effects of intravenous TY-51924 might be expected based on the results obtained for the MSIs using SPECT at 20-30mg/kg. However, further large-scale, double-blind, placebo-controlled clinical studies are required to confirm the efficacy and safety implied in the current study.

Population pharmacokinetics of peramivir in healthy volunteers and influenza patients.

UNLABELLED: Peramivir is an intravenous anti-influenza agent that inhibits viral growth by selectively inhibiting neuraminidase in human influenza A and B viruses. To characterize its pharmacokinetics, a population pharmacokinetic analysis of peramivir was performed using 3,199 plasma concentration data samples from 332 subjects in six clinical studies in Japan and the United States, including studies with renal impairment subjects, elderly subjects, and influenza patients. A three-compartment model well described the plasma concentration data for peramivir, and creatinine clearance was found to be the most important factor influencing clearance. Age and body weight were also found to be covariates for clearance and the volume of distribution, respectively. No difference in pharmacokinetics was found between genders or between Japanese and U.S. SUBJECTS: Small differences in pharmacokinetics were observed between uninfected subjects and influenza patients (clearance was 18% higher and the volume of distribution was 6% lower in influenza patients). Monte Carlo simulations indicated that single adjusted doses of 1/3- and 1/6-fold for patients with moderate and severe renal impairment, respectively, would give areas under the curve comparable to those for patients with normal renal function. The population pharmacokinetic model developed for peramivir should be useful for understanding its pharmacokinetic characteristics and for dose adjustment on the basis of renal function.

Simultaneous determination of praziquantel, pyrantel embonate, febantel and its active metabolites, oxfendazole and fenbendazole, in dog plasma by liquid chromatography/mass spectrometry.

A liquid chromatography-electrospray-mass spectrometry method (LC/MS) has been developed and validated for determination of praziquantel (PZQ), pyrantel (PYR), febantel (FBT), and the active metabolites fenbendazole (FEN) and oxfendazole (OXF), in dog plasma, using mebendazole as internal standard (IS). The method consists of solid-phase extractions on Strata-X polymeric cartridges. Chromatographic separation was carried out on a Phenomenex Gemini C6 -Phenyl column using binary gradient elution containing methanol and 50 mm ammonium-formate (pH 3). The method was linear (r(2) ≥ 0.990) over concentration ranges of 3-250 ng/mL for PYR andFEB, 5-250 ng/mL for OXF and FEN, and 24-1000 ng/mL for PZQ. The mean precisions were 1.3-10.6% (within-run) and 2.5-9.1% (between-run), and mean accuracies were 90.7-109.4% (within-run) and 91.6-108.2% (between-run). The relative standard deviations (RSD) were <9.1%. The mean recoveries of five targeted compounds from dog plasma ranged from 77 to 94%.The new LC/MS method described herein was fully validated and successfully applied to the bioequivalence studies of different anthelmintic formulations such as tablets containing PZQ, PYR embonate and FBT in dogs after oral administration.

A rapid LC-MS/MS quantification of peramivir using a simple and inexpensive sample precipitation: application to PK.

AIM: Peramivir is a newly approved selective neuraminidase inhibitor designed to inhibit influenza virus infection. METHODOLOGY/RESULTS: We report a robust and sensitive method utilizing simple precipitation extraction with LC-MS/MS for the high-throughput quantification. Addition of 0.06 M of ammonium formate and 0.1% formic acid in mobile phase could help reduce the matrix effect. This method uses 100 µl of plasma and covers a linear concentration range from 5 to 10,000 ng/ml. Other validation parameters are also evaluated and meet regulatory expectations by US FDA guidelines. CONCLUSION: The developed HPLC-MS/MS method has been successfully applied to support a clinical pharmacokinetic study. The strategy presented here can be applied elsewhere and may be useful for other amphiphilic drugs.

Pharmacokinetics of peramivir after single intravenous doses in healthy Chinese subjects.

1.The aim of the study was to evaluate the pharmacokinetics of peramivir after single intravenous (i.v.) doses in healthy Chinese subjects. 2.In a cross-over study, 12 subjects were given 300 and 600 mg peramivir by i.v. infusion. Blood and urine samples were collected at 17 designated time points and 7 designated intervals up to 36 h post-dose. Plasma and urine concentrations of peramivir were quantified by LC-MS/MS. 3.After single i.v. doses of 300 and 600 mg peramivir, Cmax and AUC0-t of peramivir were 21.4 ± 3.7, 41.1 ± 5.3 mgcL(-1) and 55.90 ± 10.62, 112.1 ± 13.2 mgch L(-1), respectively. Cmax and AUC increased in proportion to the dose. Within 12 h, accumulative urinary recoveries of peramivir after single i.v. doses of 300 and 600 mg peramivir were 84.31 ± 11.75% and 88.10 ± 7.39%, respectively. 4.In healthy Chinese subjects, peramivir displayed linear pharmacokinetics in the range of 300-600 mg, and was primarily excreted via urine as unchanged drug.

Determination of 12 potential nephrotoxicity biomarkers in rat serum and urine by liquid chromatography with mass spectrometry and its application to renal failure induced by Semen Strychni.

In previous nephrotoxicity metabonomic studies, several potential biomarkers were found and evaluated. To investigate the relationship between the nephrotoxicity biomarkers and the therapeutic role of Radix Glycyrrhizae extract on Semen Strychni-induced renal failure, 12 typical biomarkers are selected and a simple LC-MS method has been developed and validated. Citric acid, guanidinosuccinic acid, taurine, guanidinoacetic acid, uric acid, creatinine, hippuric acid, xanthurenic acid, kynurenic acid, 3-indoxyl sulfate, indole-3-acetic acid, and phenaceturic acid were separated by a Phenomenex Luna C18 column and a methanol/water (5 mM ammonium acetate) gradient program with a runtime of 20 min. The prepared calibration curves showed good linearity with regression coefficients all above 0.9913. The absolute recoveries of analytes from serum and urine were all more than 70.4%. With the developed method, analytes were successfully determined in serum and urine samples within 52 days. Results showed that guanidinosuccinic acid, guanidinoacetic acid, 3-indoxyl sulfate, and indole-3-acetic acid (only in urine) were more sensitive than the conventional renal function markers in evaluating the therapeutic role of Radix Glycyrrhizae extract on Semen Strychni-induced renal failure. The method could be further used in predicting and monitoring renal failure cause by other reasons in the following researches.

Quantification of peramivir in dog plasma by liquid chromatography/tandem mass spectrometry employing precolumn derivatization.

Peramivir is a novel influenza neuraminidase inhibitor used for anti-influenza. In this article, a novel method was developed to determine peramivir in dog plasma using a derivatization treatment step to increase the retention time and enhance the signal intensity. The sample preparation consisted of a protein precipitation extraction followed by derivatization with 10M hydrochloric acid-methanol (10:90, v/v) and determined by liquid chromatography coupled with tandem mass spectrometry. The selected reaction monitoring mode of the positive ion was performed and the precursor to the product ion transitions of m/z 343→284 and m/z 299→152 were used to measure the derivative of peramivir and Ro 64-0802 (internal standard, an active metabolite of oseltamivir). The chromatographic separation was achieved using a ZORBAX RX-C8 (2.0mm×150mm×5µm) analytical column with an isocratic mobile phase composed of acetonitrile-water-formic acid (30:70:0.1, v/v/v, 0.2mL/min). The method was linear over a concentration range of 0.25-250ng/mL. The average intra-day/inter-day precision values were 4.04-8.17% and 3.02-7.08%, respectively, while the average accuracy value was 93.99-106.48%. This method has been successfully applied to the preclinical dog research of peramivir following intragastric administration.