Future Pharmacokinetic Studies Research Articles

Hepatoprotective Study of Grewia Asiatica

Introduction: Plants have always been a very good source of drugs. Liver disorders are nearly one of the lethal ailments in the globe now a day. Hence, in the present study, we intend to evaluate the hepatoprotective study of Grewia asiatica. Materials and Methods: G. asiatica was collected from hills of Tirupati, dried, and powdered. Healthy male Wistar albino rats with paracetamol and CCl4-induced liver damage were studied on day 1 and 7 for the serum glutamate oxaloacetate transaminase (SGOT), serum alanine transaminase (SGPT), ALKP, and total protein levels after the extract was given at different concentrations. The results were compared histologically. Results: The hepatoprotective effect of the plant extract and combination also proved by measuring the liver related parameters such as SGOT, SGPT, ALKP, and total protein which were elevated in disease but significantly reduce in standard and plant extract treated groups. Conclusion: G. asiatica plant extracts were safe for treating the hepatic diseases and did not show any toxicities or drug-related interactions but future pharmacokinetic studies required to validate the given result.

A randomized trial of safety and pharmacodynamic interactions between a selective glucocorticoid receptor antagonist, PT150, and ethanol in healthy volunteers

PT150, a novel competitive glucocorticoid receptor (GR) antagonist, has proven safe in animal models, healthy volunteers, and people with depression. Our study is the first to investigate PT150’s safety with alcohol use. The primary objective of this study was to evaluate pharmacodynamic interactions between ethanol and PT150 in healthy subjects. This single-site, Phase I pilot trial consisted of community-recruited, healthy, alcohol-experienced participants aged 21–64 years. Of 32 participants screened, 11 were enrolled and randomized, one of which withdrew before intervention. PT150 (900 mg/day) was administered orally to all participants for five days. All participants received two beverage challenges on Day 1 (before PT150 administration) and Day 5 (after PT150 administration). On challenge days, they received both alcohol (16% ethanol) and placebo (1% ethanol) beverages in random order. Primary outcomes included breath alcohol level, blood pressure, heart rate, adverse events, and electrocardiogram changes. There were no statistically significant differences in vital signs or estimated blood alcohol concentrations between PT150 non-exposed and exposed groups during the ethanol challenge. There were no clinically significant abnormal electrocardiograms or serious adverse events. These data show that administration of PT150 with concurrent alcohol use is safe and well-tolerated. This study supports a future pharmacokinetic interaction study between PT150 and alcohol.Trial Registration ClinicalTrials.gov Identifier: NCT03548714.

Sequencing of the Canine Cytochrome P450 CYP2C41 Gene and Genotyping of Its Polymorphic Occurrence in 36 Dog Breeds.

Cytochrome P450 (CYP) drug metabolizing enzymes play an important role in efficient drug metabolism and elimination. Many CYPs are polymorphic and, thereby, drug metabolism can vary between individuals. In the case of canine CYP2C41, gene polymorphism was identified. However, as the first available canine genome sequences all were CYP2C41 negative, this polymorphism could not be clarified at the genomic level. The present study provides an exact characterization of the CYP2C41 gene deletion polymorphism at the genomic level and presents a PCR-based genotyping method that was used for CYP2C41 genotyping of 1,089 individual subjects from 36 different dog breeds. None of the Bearded Collie, Bernese Mountain, Boxer, Briard, French Bulldog or Irish Wolfhound subjects had the CYP2C41 gene in their genomes. In contrast, in the Chinese Char-Pei, Siberian Husky, Schapendoes and Kangal breeds, the CYP2C41 allele frequency was very high, with values of 67, 57, 43, and 34%, respectively. Interestingly, the site of gene deletion was identical for all CYP2C41 negative dogs, and all CYP2C41 positive dogs showed highly homologous sequence domains upstream and downstream from the CYP2C41 gene. CYP2C41 genotyping can now be routinely used in future pharmacokinetic studies in canines, in order to identify genetically-based poor or extensive drug metabolizers. This, together with more extensive in vitro drug screening for CYP2C41 substrates will help to determine the clinical relevance of CYP2C41, and to optimize drug treatment. Although the relative abundance of the CYP2C41 protein in the canine liver seems to not be very high, this CYP could substantially contribute to hepatic drug metabolism in dogs expressing CYP2C41 from both alleles and, when CYP2C41 shows higher catalytic activity to a given drug than other hepatic metabolic enzymes.

Development and validation of an LC-MS/MS method for the quantification of the anthelmintic drug moxidectin in a volumetric absorptive microsample, blood, and plasma: Application to a pharmacokinetic study of adults infected with Strongyloides stercoralis in Laos

Moxidectin is a promising candidate for addition to the lean repertoire of drugs against neglected tropical diseases (NTD) including strongyloidiasis. Pharmacokinetic (PK) and -dynamic studies are required to support its clinical development. Microsampling approaches enable PK studies in the challenging environments where NTDs are most prevalent, due to simplified collection and processing.We developed a liquid chromatography tandem mass spectrometry method for the sensitive quantification of moxidectin in human blood obtained by capillary sampling with the microsampling device Mitra® compared to blood and plasma obtained by venous sampling. Sample preparation consisted of protein precipitation, evaporation and reconstitution and also included phospholipid filtration for blood and plasma. Moxidectin was detected by multiple reaction monitoring (640.4 → 528.5 m/z) using a Luna C8(2) (30 × 2.0 mm, 3 µm particle size, 100 Å) analytical column with a gradient program of 6 min duration. Validation was performed with respect to accuracy, precision, sensitivity, selectivity, linearity, stability, recovery, and haematocrit influence with a limit of quantification of 0.5 and 2.5 ng/mL, for venous and capillary blood respectively. Moxidectin was stable up to 2 months at storage condition (blood and plasma: −20 °C, microsamples: room temperature), 3 cycles of temperature shift, for at least 4 h on the bench-top and 24 h in the autosampler (4 °C). Deviations of inter- and intra-assay accuracy and precision were smaller than 12.6% and recoveries were in the range of 80.7–111.2%.The method was applied to samples obtained from nine Strongyloides stercoralis-infected adults from northern Laos. A good agreement in the time-concentration profiles of moxidectin and a high consistency in PK parameters was found between the different matrixes and sampling strategies: e.g. identical time to reach maximal concentration of 4.0 h and a similar maximal concentration of 83.9–88.5 ng/mL of moxidectin.The simple and practical capillary procedure using Mitra® microsampling has been demonstrated to be suitable for PK studies of moxidectin and will pave the way for future PK studies.

Ibudilast (MN-166) in amyotrophic lateral sclerosis- an open label, safety and pharmacodynamic trial

Ibudilast (MN-166) is an inhibitor of macrophage migration inhibitory factor (MIF) and phosphodiesterases 3,4,10 and 11 (Gibson et al., 2006; Cho et al., 2010). Ibudilast attenuates CNS microglial activation and secretion of pro-inflammatory cytokines (Fujimoto et al., 1999; Cho et al., 2010). In vitro evidence suggests that ibudilast is neuroprotective by suppressing neuronal cell death induced by microglial activation. People with ALS have increased microglial activation measured by [11C]PBR28-PET in the motor cortices. The primary objective is to determine the impact of ibudilast on reducing glial activation and neuroaxonal loss in ALS, measured by PBR28-PET and serum Neurofilament light (NfL). The secondary objectives included determining safety and tolerability of ibudilast high dosage (up to 100 mg/day) over 36 weeks.In this open label trial, 35 eligible ALS participants underwent ibudilast treatment up to 100 mg/day for 36 weeks. Of these, 30 participants were enrolled in the main study cohort and were included in biomarker, safety and tolerability analyses. Five additional participants were enrolled in the expanded access arm, who did not meet imaging eligibility criteria and were included in the safety and tolerability analyses. The primary endpoints were median change from baseline in (a) PBR28-PET uptake in primary motor cortices, measured by standard uptake value ratio (SUVR) over 12–24 weeks and (b) serum NfL over 36–40 weeks. The secondary safety and tolerability endpoints were collected through Week 40.The baseline median (range) of PBR28-PET SUVR was 1.033 (0.847, 1.170) and NfL was 60.3 (33.1, 219.3) pg/ml. Participants who completed both pre and post-treatment scans had PBR28-PET SUVR median(range) change from baseline of 0.002 (−0.184, 0.156) , P = 0.5 (n = 22). The median(range) NfL change from baseline was 0.4 pg/ml (−1.8, 17.5), P = 0.2 (n = 10 participants). 30(86%) participants experienced at least one, possibly study drug related adverse event. 13(37%) participants could not tolerate 100 mg/day and underwent dose reduction to 60–80 mg/day and 11(31%) participants discontinued study drug early due to drug related adverse events.The study concludes that following treatment with ibudilast up to 100 mg/day in ALS participants, there were no significant reductions in (a) motor cortical glial activation measured by PBR28-PET SUVR over 12–24 weeks or (b) CNS neuroaxonal loss, measured by serum NfL over 36–40 weeks. Dose reductions and discontinuations due to treatment emergent adverse events were common at this dosage in ALS participants. Future pharmacokinetic and dose-finding studies of ibudilast would help better understand tolerability and target engagement in ALS.

Demonstrating Feasibility of an Opportunistic Sampling Approach for Pharmacokinetic Studies of β-Lactam Antibiotics in Critically Ill Children.

There has been increasing interest in incorporating β-lactam precision dosing into routine clinical care, but robust population pharmacokinetic models in critically ill children are needed for these purposes. The objective of this study was to demonstrate the feasibility of an opportunistic sampling approach that utilizes scavenged residual blood for future pharmacokinetic studies of cefepime, meropenem, and piperacillin. We aimed to show that opportunistic samples would cover the full concentration-versus-time profiles and to evaluate stability of the antibiotics in whole blood and plasma to optimize future use of the opportunistic sampling approach. A prospective observational study was conducted in a single-center pediatric intensive care unit, where pediatric patients administered at least 1 dose of cefepime, meropenem, or piperacillin/tazobactam and who had residual blood scavenged from samples obtained for routine clinical care were enrolled. A total of 138 samples from 22 pediatric patients were collected in a 2-week period. For all 3 antibiotics, the samples collected covered the entire dosing intervals and were not clustered around specific times. There was high variability in the free concentrations and in the percentage of drug bound to protein. There was less than 15% degradation for meropenem or piperacillin when stored in whole blood or plasma at 4°C after 6 days. Cefepime degraded by more than 15% after 3 days. The opportunistic sampling approach is a powerful and feasible method to obtain sufficient samples to study the variability of drug concentrations and protein binding for future pharmacokinetic studies in the pediatric critical care population.

Pharmacokinetics of ketamine and its major metabolites norketamine, hydroxynorketamine, and dehydronorketamine: a model-based analysis

BackgroundRecent studies show activity of ketamine metabolites, such as hydroxynorketamine, in producing rapid relief of depression-related symptoms and analgesia. To improve our understanding of the pharmacokinetics of ketamine and metabolites norketamine, dehydronorketamine, and hydroxynorketamine, we developed a population pharmacokinetic model of ketamine and metabolites after i.v. administration of racemic ketamine and the S-isomer (esketamine). Pharmacokinetic data were derived from an RCT on the efficacy of sodium nitroprusside (SNP) in reducing the psychotomimetic side-effects of ketamine in human volunteers. MethodsThree increasing i.v. doses of esketamine and racemic ketamine were administered to 20 healthy volunteers, and arterial plasma samples were obtained for measurement of ketamine and metabolites. Subjects were randomised to receive esketamine/SNP, esketamine/placebo, racemic ketamine/SNP, and racemic ketamine/placebo on four separate occasions. The time–plasma concentration data of ketamine and metabolites were analysed using a population compartmental model approach. ResultsThe pharmacokinetics of ketamine and metabolites were adequately described by a seven-compartment model with two ketamine, norketamine, and hydroxynorketamine compartments and one dehydronorketamine compartment with metabolic compartments in-between ketamine and norketamine, and norketamine and dehydronorketamine main compartments. Significant differences were found between S- and R-ketamine enantiomer pharmacokinetics, with up to 50% lower clearances for the R-enantiomers, irrespective of formulation. Whilst SNP had a significant effect on ketamine clearances, simulations showed only minor effects of SNP on total ketamine pharmacokinetics. ConclusionsThe model is of adequate quality for use in future pharmacokinetic and pharmacodynamic studies into the efficacy and side-effects of ketamine and metabolites. Clinical trial registrationDutch Cochrane Center 5359.

Systemic evaluation and localization of resistin expression in normal human tissues by a newly developed monoclonal antibody.

Resistin and resistin-like molecules are pleiotropic cytokines that are involved in inflammatory diseases. Our previous work suggested that resistin has the potential to be used as a biomarker and therapeutic target for human pulmonary arterial hypertension. However, data are limited on the distribution of resistin in healthy human organs. In this study, we used our newly developed anti-human resistin (hResistin) antibody to immunohistochemically detect the expression, localization, and intracellular/extracellular compartmentalization of hResistin in a full human tissue panel from healthy individuals. The potential cross reactivity of this monoclonal anti-hResistin IgG1 with normal human tissues also was verified. Results showed that hResistin is broadly distributed and principally localized in the cytoplasmic granules of macrophages scattered in the interstitium of most human tissues. Bone marrow hematopoietic precursor cells also exhibited hResistin signals in their cytoplasmic granules. Additionally, hResistin labeling was observed in the cytoplasm of nervous system cells. Notably, the cytokine activity of hResistin was illustrated by positively stained extracellular material in most human tissues. These data indicate that our generated antibody binds to the secreted hResistin and support its potential use for immunotherapy to reduce circulating hResistin levels in human disease. Our findings comprehensively document the basal expression patterns of hResistin protein in normal human tissues, suggest a critical role of this cytokine in normal and pathophysiologic inflammatory processes, and offer key insights for using our antibody in future pharmacokinetic studies and immunotherapeutic strategies.

Model-based analysis on systemic availability of co-administered cannabinoids after controlled vaporised administration.

The most important two medicinal cannabinoids are Δ9 -tetrahydrocannabinol (THC) and cannabidiol (CBD). Vaporised administration is superior due to its higher systemic availability, lower individual variability and faster drug delivery. Although it is common THC is co-administered with CBD, the influence of CBD on the pharmacokinetics, especially the systemic availability of THC after vaporised administration, is unknown. To investigate the influence of different doses of co-administered CBD on the systemic availability of THC, and to compare the availability of THC and CBD in a sample of frequent and infrequent cannabis users. The study used a randomised, double-blind, crossover placebo-controlled design. THC and/or CBD in ethanol was vaporised and inhaled. Plasma concentrations of THC and CBD were analysed. The THC data created in this study were pooled together with published THC pharmacokinetic data in order to cover all the phases of THC disposition. Population pharmacokinetic model of THC was developed based on the pooled data. The model of CBD was developed based on the data created in this study. Population pharmacokinetic models of THC and CBD were developed. With concomitant inhalation of high-dose CBD, the systemic availability of THC decreased significantly. Frequent cannabis users appeared to have higher systemic availability of THC and CBD when high-dose CBD was administered. The results observed in this study are useful for guiding future pharmacokinetic studies of medicinal cannabinoids, and for development of dosing guidelines for medical use of cannabis in the 'real-world' setting.

Prediction of Drug Exposure in Critically Ill Encephalopathic Neonates Treated With Therapeutic Hypothermia Based on a Pooled Population Pharmacokinetic Analysis of Seven Drugs and Five Metabolites.

Drug dosing in encephalopathic neonates treated with therapeutic hypothermia is challenging; exposure is dependent on body size and maturation but can also be influenced by factors related to disease and treatment. A better understanding of underlying pharmacokinetic principles is essential to guide drug dosing in this population. The prospective multicenter cohort study PharmaCool was designed to investigate the pharmacokinetics of commonly used drugs in neonatal encephalopathy. In the present study, all data obtained in the PharmaCool study were combined to study the structural system specific effects of body size, maturation, recovery of organ function, and temperature on drug clearance using nonlinear mixed effects modeling. Data collected during the first 5 days of life from 192 neonates treated with therapeutic hypothermia were included. An integrated population pharmacokinetic model of seven drugs (morphine, midazolam, lidocaine, phenobarbital, amoxicillin, gentamicin, and benzylpenicillin) and five metabolites (morphine‐3‐glucuronide, morphine‐6‐glucuronide, 1‐hydroxymidazolam, hydroxymidazolam glucuronide, and monoethylglycylxylidide) was successfully developed based on previously developed models for the individual drugs. For all compounds, body size was related to clearance using allometric relationships and maturation was described with gestational age in a fixed sigmoidal Hill equation. Organ recovery after birth was incorporated using postnatal age. Clearance increased by 1.23%/hours of life (95% confidence interval (CI) 1.03–1.43) and by 0.54%/hours of life (95% CI 0.371–0.750) for high and intermediate clearance compounds, respectively. Therapeutic hypothermia reduced clearance of intermediate clearance compounds only, by 6.83%/°C (95% CI 5.16%/°C–8.34%/°C). This integrated model can be used to facilitate drug dosing and future pharmacokinetic studies in this population.

The Interaction Between Rosuvastatin and Ticagrelor Leading to Rhabdomyolysis: A Case Report and Narrative Review.

Drug interactions are a common cause of morbidity and mortality and may require prompt discontinuation of therapeutic regimens due to harmful side effects. Patients with acute coronary syndromes are likely to be prescribed multiple medications that are metabolized through the cytochrome P450 system, increasing the probability for drug interaction. Atorvastatin and simvastatin are both well known to interact with the oral P2Y12 agent ticagrelor. The purpose of this paper is to describe the interaction of ticagrelor with rosuvastatin leading to rhabdomyolysis, which is less clearly defined in the literature. We report a case of a 74-year-old male who presented with bilateral lower extremity weakness and difficulty ambulating for one month after being prescribed ticagrelor for a drug eluting stent, in the setting of already being on rosuvastatin. His clinical picture and laboratory findings were consistent with a diagnosis of rhabdomyolysis. His medications were adjusted to a regimen of clopidogrel and alirocumab. One month later, he returned to his baseline status. The mechanism of interaction between rosuvastatin and ticagrelor appears to be multifactorial. It may be caused by CYP450-mediated metabolism from a small amount of crossover between isoenzymes. Ticagrelor may also cause acute kidney injury, increasing the concentration of rosuvastatin. Other mechanisms of interaction include genetic differences in the organic anion transporter polypeptides and transportation through p-glycoprotein. Future pharmacokinetic studies are warranted to better understand the interaction.

In Vitro Metabolic Profiling of Periplogenin in Rat Liver Microsomes and its Associated Enzyme-kinetic Evaluation

Background: Periplogenin, an active ingredient in Cortex Periplocae, is widely noted for its multiple biological activities; however, the metabolism of this compound has been scarcely investigated. The present report proposed the in vitro metabolic profiling and reaction pathways of periplogenin in rat liver microsomes. Method and Results: The metabolites of periplogenin in rat liver microsomes were analyzed. Two main metabolites, namely 14-hydroxy-3-oxo-14β-carda-4, 20 (22)-dienolide and 5, 14-dihydroxy-3-oxo-5β, 14β-card-20(22)-enolide were identified by HPLC-MSn, 1H-NMR and 13C-NMR. HPLC method was established for the simultaneous determination of periplogenin and its related metabolites (M0, M1 and M2), which was performed on Waters ODS column with a methanol-water solution (53:47, v/v) as mobile phase and descurainoside as an internal standard at 220 nm. The linearity ranges of M0, M1 and M2 were 0.64-820.51, 0.68-864.86 and 0.64-824.74 μM respectively with the regression coefficient (R2) above 0.9995. The limits of quantitation for these metabolites (M0, M1 and M2) were 0.18, 0.22 and 0.15 μM respectively. The developed method was also accurate (with relative errors of -3.6% to 3.2%) and precise (with relative standard deviations below 7.9%). The recoveries of the three analytes were above 85.7% with stability in the range of -2.4% to 3.6%. The enzyme-kinetic parameters of periplogenin including Vmax (6.08 ± 0.19 nmol/mg protein/min), Km (288.62 ± 14.54 μM) and Clint (21 ± 1.0 μL/min/mg protein) were calculated using nonlinear regression analysis. Conclusion: These findings significantly highlighted the metabolic pathways of periplogenin and also provided some reference data for future pharmacokinetic and pharmacodynamic studies.

Therapeutic Emergence of Rhein as a Potential Anticancer Drug: A Review of Its Molecular Targets and Anticancer Properties

According to the World Health Organization (WHO), cancer is the second-highest cause of mortality in the world, and it kills nearly 9.6 million people annually. Besides the fatality of the disease, poor prognosis, cost of conventional therapies, and associated side-effects add more burden to patients, post-diagnosis. Therefore, the search for alternatives for the treatment of cancer that are safe, multi-targeted, effective, and cost-effective has compelled us to go back to ancient systems of medicine. Natural herbs and plant formulations are laden with a variety of phytochemicals. One such compound is rhein, which is an anthraquinone derived from the roots of Rheum spp. and Polygonum multiflorum. In ethnomedicine, these plants are used for the treatment of inflammation, osteoarthritis, diabetes, and bacterial and helminthic infections. Increasing evidence suggests that this compound can suppress breast cancer, cervical cancer, colon cancer, lung cancer, ovarian cancer, etc. in both in vitro and in vivo settings. Recent studies have reported that this compound modulates different signaling cascades in cancer cells and can prevent angiogenesis and progression of different types of cancers. The present review highlights the cancer-preventing and therapeutic properties of rhein based on the available literature, which will help to extend further research to establish the chemoprotective and therapeutic roles of rhein compared to other conventional drugs. Future pharmacokinetic and toxicological studies could support this compound as an effective anticancer agent.

Quantifying Intranasally Administered Deferoxamine in Rat Brain Tissue with Mass Spectrometry.

Deferoxamine, a metal chelator, has been shown to be neuroprotective in animal models of ischemic stroke, traumatic brain injury and both subarachnoid and intracerebral hemorrhage. Intranasal deferoxamine (IN DFO) has also shown promise as a potential treatment for multiple neurodegenerative diseases, including Parkinson's and Alzheimer's. However, there have been no attempts to thoroughly understand the dynamics and pharmacokinetics of IN DFO. We developed a new high-performance liquid-chromatography electrospray-tandem mass spectrometry (HPLC/ESI-MS2) method to quantify the combined total levels of DFO, ferrioxamine (FO; DFO bound to iron), and aluminoxamine (AO; aluminum-bound DFO) in brain tissue using a custom-synthesized deuterated analogue (DFO-d7, Medical Isotopes Inc., Pelham NH) as an internal standard. We applied our method toward understanding the pharmacokinetics of IN DFO delivery to the brain and blood of rats from 15 min to 4 h after delivery. We found that IN delivery successfully targets DFO to the brain to achieve concentrations of 0.5-15 μM in various brain regions within 15 min, and decreasing though still detectable after 4 h. Systemic exposure was minimized as assessed by concentration in blood serum. Serum concentrations were 0.02 μM at 15 min and no more than 0.1 μM at later time points. Compared to blood serum, brain region-specific drug exposure (as measured by area under the curve) ranged from slightly under 10 times exposure in the hippocampus to almost 200 times exposure in the olfactory bulb with IN DFO delivery. These findings represent a major step toward future method development, pharmacokinetic studies, and clinical trials for this promising therapeutic.

Novel bioanalytical method for the quantification of rufinamide in mouse plasma and tissues using HPLC-UV: A tool to support pharmacokinetic studies

Rufinamide is an antiepileptic drug approved for seizures treatment associated with Lennox-Gastaut syndrome. To support future pharmacokinetic studies in rodents, this work aimed to validate for the first time a fast and simple high-performance liquid chromatographic (HPLC) method for rufinamide quantification in mouse plasma and brain, liver and kidney tissues. For that, aliquots (100 μL) of plasma or tissues homogenates were spiked with known amounts of rufinamide and chloramphenicol (internal standard). Compounds were extracted using a combination of protein precipitation and liquid-liquid extraction. Their separation was accomplished using a LiChroCART® Purospher Star column (C18, 55 mm × 4 mm; 3 μm) protected by a LiChroCART® Purospher Star pre-column (C18, 4 mm × 4 mm; 5 μm) at 35 °C. Mobile phase [water/acetonitrile (82:18, v/v)] was isocratically pumped at 1.0 mL min−1 and detection was performed using a diode-array detector set at 210 nm. A preliminary in vivo pharmacokinetic study was also performed by orally administering rufinamide (10 mg kg−1) to mice. The bioanalytical method herein developed was validated according to international bioanalytical guidelines and showed to be selective and linear (r2 ≥ 0.9918) over the concentration range of 0.1–30 μg mL−1. Regarding quality control samples, overall imprecision was lower than 14.5% and inaccuracy ranged between −14.6% and 15.0%. In all tested matrices, rufinamide recoveries varied between 73.1% and 85.2%. This method was successfully applied in a preliminary pharmacokinetic study, suggesting to be a useful bioanalytical tool to support further non-clinical pharmacokinetic-based studies involving rufinamide.

A review on analytical methods for natural berberine alkaloids.

Berberine alkaloids, a group of protoberberine alkaloids under the classification of isoquinoline alkaloids, include berberine, coptisine, palmatine, columbamine, dehydrocorydaline, jatrorrhizine, and epiberberine from natural sources. Studies have shown that berberine alkaloids have various pharmacological functions, such as antibacterial, antiviral, blood pressure-lowering, hypoglycaemic, antiarrhythmia, and anticancer effects. Therefore, it is worthwhile to develop analytical methods to investigate the pharmacokinetics and activity mechanisms of berberine alkaloids and to study berberine alkaloids more comprehensively. Current analytical methods for berberine alkaloids include liquid chromatography, thin-layer chromatography, ultraviolet spectroscopy, capillary electrophoresis, and gas chromatography. The most widely used detection method is mass spectrometry. In order to provide a systematic and comprehensive summary and to serve as a reference for the future pharmacokinetics studies and analysis of berberine alkaloids, analytical methods for natural berberine alkaloids that have been used in the past ten years are reviewed here.

Quantification of U-47700 and its metabolites in plasma by LC-MS/MS

Novel Synthetic Opioids (NSO) have caused a recent epidemic both nationally and globally. NSO have gained popularity in the illicit drug market and have brought about an increase in fentanyl and its derivatives, as well as other chemically unrelated opioid agonists. U-47700, a non-fentanyl analog analgesic opioid, was first developed by The Upjohn Company and has a reported potency of 7.5 times that of morphine. Like many NSO, U-47700 is usually sold as a research chemical that can be purchased online but can also be found in “Gray Death” which is a mixture of fentanyl(s), heroin, and U-47700. With the emergence of these NSO, there is a need for laboratories to be able to detect these drugs in various matrices. In this study, a liquid chromatography tandem mass spectrometry (LC-MS/MS) method was optimized and validated to detect and quantify U-47700 and its metabolites, N-desmethyl-U-47700 and N,N-didesmethyl-U-47700, in 100 μL human plasma using an optimized solid phase extraction procedure. A small sample size (100 μL) was utilized for a future pharmacokinetic study in rats. The method was validated according to SWGTOX guidelines, including: precision and bias, linearity, carryover, interferences, matrix effects, limit of detection (LOD), limit of quantification (LOQ), dilution integrity, and stability. The LOD were 0.05 ng/mL for U-47700 and N-desmethyl-U-47700 and 0.1 ng/mL for N,N-didesmethyl-U-47700. Linear ranges for U-47700 and N-desmethyl-U-47700 were 0.1–100 ng/mL and 0.5–100 ng/mL for N,N-didesmethyl-U-47700. Matrix effects were analyzed following the post-extraction addition approach and were <5%, indicating little ion suppression or enhancement. Extraction recovery was >79%. Analytes were stable in all conditions and no endogenous or exogenous interferences were detected. This method was cross-validated in rat plasma with acceptable bias (2.1–6.2%) and precision (−14.7–15.7%) within acceptable limits. Matrix effects and extraction efficiency was comparable to human plasma validation. Postmortem whole blood samples (n = 15) were analyzed with the validated method. U-47700, N-desmethyl-U-47700 and N,N-didesmethyl-U-47700 concentration ranges were 1.1–1367 ng/mL, 4.0–1400 ng/mL and 0.7–658 ng/mL, respectively.

Evaluation of apramycin against spectinomycin-resistant and -susceptible strains of Neisseria gonorrhoeae

The emergence of Neisseria gonorrhoeae resistant to all currently available antimicrobial therapies poses a dire public health threat. New antimicrobial agents with activity against N. gonorrhoeae are urgently needed. Apramycin is an aminocyclitol aminoglycoside with broad-spectrum in vitro activity against MDR Gram-negative pathogens and Staphylococcus aureus. However, its activity against N. gonorrhoeae has not been described. The activity spectrum of apramycin against a collection of MDR N. gonorrhoeae was assessed. Isolates tested included those susceptible and resistant to the structurally distinct aminocyclitol, spectinomycin. The modal MICs for apramycin and spectinomycin were 16 mg/L and 32 mg/L, respectively. The epidemiological cut-off (ECOFF) for apramycin was 64 mg/L. No strains among 77 tested had an MIC above this ECOFF, suggesting very low levels of acquired apramycin resistance. In time-kill analysis, apramycin demonstrated rapid bactericidal activity comparable to that of spectinomycin. Apramycin has broad-spectrum, rapidly bactericidal activity against N. gonorrhoeae. Future pharmacokinetic and pharmacodynamic studies will be needed to determine whether apramycin and/or apramycin derivatives hold promise as new therapeutics for N. gonorrhoeae infection.

Development and validation of an LC-MS/MS method for the determination of DPHB in rat plasma and its application in pharmacokinetic studies.

The aim of the present study was to develop a rapid, specific and sensitive LC-MS/MS method for the determination of DPHB [7-(4″-hydroxy-3″-methoxyphenyl)-1-phenyl-4-hepten-3-one] in rat plasma using yakuchinone A as an internal standard (IS). n-Hexane was used for the extraction of DPHB from rat plasma. Chromatographic separation of DPHB was achieved using a Kinetex XB-C18 column (2.10 × 50 mm, 2.6 μm) at 40°C. The mobile phase consisted of water (containing 0.1‰ formic acid, A) and acetonitrile (containing 0.1‰ formic acid, B) under a gradient elution at a flow rate of 0.3 mL min-1 . Positive electrospray ionization and multiple reaction monitoring mode were used for detection. The selected precursor ion to product ion pairs, m/z 311.3 → 137.0 for DPHB and m/z 313.1 → 137.0 for yakuchinone A, were monitored. Good linearity was observed over the concentration range from 2 to 2000 ng mL-1 (r = 0.9969). The recovery efficiency of DPHB from rat plasma was 54.8-69.7%, while the matrix effect ranged from 99.7 to 113%. Intra- and inter-day precision and accuracy values were within ±15% at three different quality control concentration levels. This validated method was successfully applied to pharmacokinetic studies in rats after a single p.o. or i.v. dose of DPHB solution. The route of administration significantly influenced systemic exposure to DPHB, and low bioavailability of DPHB was observed. The method developed here will be further improved and used in future pharmacokinetic studies.

Short communication: Use of an ultrafiltration device in gland cistern for continuous sampling of healthy and mastitic quarters of lactating cattle for pharmacokinetic modeling

Pharmacokinetic studies of the drugs in the milk are often limited due to infrequent sampling associated with milking. Alternatively, frequent sample collection with repeated milking may increase drug elimination. The objective of this study was to determine the feasibility of continuously sampling the udder using ultrafiltration. An ultrafiltration probe was placed into the gland cisterns through mammary parenchyma of normal and mastitic quarters of 6 mature mid-lactation Jersey cows with naturally occurring subclinical mastitis. An ultrafiltration probe was secured to the caudal or lateral aspect of the udder depending on the quarter being sampled. The timed interval samples were collected at 0, 2, 4, 6, 8, 12, 18, 24, 28, 32, 36, 48, 60, 72, 84, and 96 h after drug administration. Plasma samples were collected at the same time points. Each cow received 2.2 mg/kg of flunixin intravenously before milking at time 0. All cows were routinely milked by machine every 12 h. Flunixin concentrations in plasma, whole milk, and milk ultrafiltrates were analyzed by use of ultra-high-performance liquid chromatography with mass spectrometric detection. We found no significant effects on the appearance of the milk or the ability to milk the cows after implantation of the ultrafiltration probes. The concentration of flunixin collected from the ultrafiltration probes in the mastitic quarters tended to be greater than that of the healthy quarters. We concluded that collection of ultrafiltration samples from the mammary gland of cows provides a viable means to continuously assess drug concentrations in the milk while continuing to milk the cow normally. This study demonstrates the utility of continuous sampling of milk via ultrafiltration for future pharmacokinetic studies in cattle.