Development and validation of a chiral LC-ESI-MS/MS method for simultaneous determination of the enantiomeric metabolites isosorbide 2-mononitrate and isosorbide 5-mononitrate of isosorbide dinitrate in rat and human plasma.

A specific liquid chromatography-tandem mass spectrometry (LC-ESI-MS/MS) assay was developed and validated for simultaneous determination of two active metabolites of isosorbide dinitrate (ISDN), namely, isosorbide 2-mononitrate (IS 2-MN) and isosorbide 5-mononitrate (IS 5-MN). A simple protein precipitation extraction technique was employed using 13C6 isosorbide 5-mononitrate as the internal standard. The two isomers were separated on a chiral column and mass detection was carried out by electrospray ionization (ESI) in negative multiple reaction monitoring (MRM) mode (ESI -ve). As neutral organic nitrates do not ionize well in ESI ion source, adduct formation of IS 2-MN and IS 5-MN were evaluated. Acetate adduct ions of IS 2-MN and IS 5-MN were well ionized and fragmentable in the negative mode by liquid chromatography electrospray ionization/tandem mass spectrometry. These acetates adduct ions, of IS 2-MN and IS 5-MN were selected as parent mass for quantitation. The method was developed and validated in rat and human plasma with K2EDTA as an anticoagulant. This simultaneous quantitation method was shown to be linear over a working range of 25.0 ng/mL to 5050 ng/mL and 12.4 ng/mL to 2500 ng/mL for IS 2-MN (r2 > 0.99) and IS 5-MN (r2 > 0.99), respectively, in rat and human plasma. Sensitivity was determined as 25.0 ng/mL for IS 2-MN and 12.4 ng/mL for IS 5-MN in rat and human plasma. Inter- and intra-day accuracy and precision were within ±15% in both method validations. This validated method was subsequently applied to a pharmacokinetic (PK) study of ISDN in rat after oral administration.

A simple and sensitive LC-MS/MS method for quantification of Bepridil in rat plasma and its application to pharmacokinetic studies.

Bepridil is potent inhibitor of Na+, K+ and Ca+ channel in cardiomyocytes. It has demonstrated strong antianginal effect with type I antiarrhythmic and with minimum antihypertensive therapeutic effect. Till date, a specific LC-MS/MS method to quantify Bepridil concentrations in biological matrix have not been reported yet. In current study, a highly sensitive, specific and simple LC-MS/MS method for quantification of antianginal drug Bepridil in rat plasma is presented. The LC-MS/MS method was validated in terms of selectivity, specificity, sensitivity, accuracy and precision, matrix effect, extraction recovery and stability as per USFDA's bioanalytical method validation guideline. The validated assay was applied for quantification of Bepridil from pharmacokinetic study in rats following oral and intravenous administration. The lower limit of quantification (LLOQ) of Bepridil was 1 ng/mL. The calibration curve ranges from 1 ng/mL to 1000 ng/mL with desirable linearity and r2 > 0.99. The method exhibited 10-fold dilution integrity. The intra-day and inter-day accuracy were within 101.32-96.80% and 102.87-95.35% with coefficient of variation 10.11-2.89% and 10.45-3.97% respectively. No significant interference observed by endogenous peak at the retention time of Bepridil and IS. The assay was free from any matrix effect, precise recovery across the calibration curve range and samples were stable under all experimental conditions. The validated assay was successfully applied to analyze plasma samples of pharmacokinetic study in rat to determine concentrations of Bepridil. In summary, a novel method for analyzing Bepridil in rat plasma has been successfully validated and is now being utilized for quantification of Bepridil from pre-clinical studies.

In vitro metabolism, disposition, preclinical pharmacokinetics and prediction of human pharmacokinetics of DNDI-VL-2098, a potential oral treatment for Visceral Leishmaniasis.

The in vitro metabolism and in vivo pharmacokinetic (PK) properties of DNDI-VL-2098, a potential oral agent for Visceral Leishmaniasis (VL) were studied and used to predict its human pharmacokinetics. DNDI-VL-2098 showed a low solubility (10µM) and was highly permeable (>200nm/s) in the Caco-2 model. It was stable in vitro in liver microsomes and hepatocytes and no metabolite was detectable in circulating plasma from dosed animals suggesting very slow, if any, metabolism of the compound. DNDI-VL-2098 was moderate to highly bound to plasma proteins across the species tested (94-98%). DNDI-VL-2098 showed satisfactory PK properties in mouse, hamster, rat and dog with a low blood clearance (<15% of hepatic blood flow except hamster), a volume of distribution of about 3 times total body water, acceptable half-life (1-6h across the species) and good oral bioavailability (37-100%). Allometric scaling of the preclinical PK data to human gave a blood half-life of approximately 20h suggesting that the compound could be a once-a-day drug. Based on the above assumptions, the minimum efficacious dose predicted for a 50kg human was 150mg and 300mg, using efficacy results in the mouse and hamster, respectively.

Unilateral cerebellar hypoplasia.

Unilateral cerebellar hypoplasia is a relatively rare malformation. We report the case of a 7-year-old boy who presented with a history of a fall, which was followed by cerebellar signs. Imaging findings suggested a diagnosis of unilateral cerebellar hypoplasia. The child recovered with conservative management, probably because the cerebellar signs were due to the trauma and not the hypoplasia itself.

Dual effect of Toxicodendron pubescens on Carrageenan induced paw edema in rats.

BACKGROUND: Toxicodendron pubescens is the current botanical name of homeopathic Rhus toxicodendron (Rhus tox). Rhus tox drug is widely used in homeopathically diluted form in the treatment of inflammatory and edematous conditions. We studied the effect of crude form of this plant, after single and multiple doses in Carrageenan induced paw inflammation in rats. METHOD: We evaluated effects of single dose and multiple doses of orally administered Rhus tox on Carrageenan induced paw inflammation in rats. We tested 10 mg/kg, 20 mg/kg and 40 mg/kg doses of Rhus tox. In the single dose study, Rhus tox was administered 1 h prior to the subplantar injection of Carrageenan. In the multiple dose study, Rhus tox was administered twice daily for three days and Carrageenan was injected 1 h after the last dose. Paw volume was measured using a digital plethysmometer. RESULTS: Administration of a single dose of Rhus tox 1h prior to injection of Carrageenan significantly reduced the paw inflammation in a dose dependent manner. Administration of multiple doses of Rhus tox increased the intensity of inflammation induced by Carrageenan, but this was not statistically significant. CONCLUSION: Rhus tox, in crude form, exerts anti-inflammatory effects after a single dose and proinflammatory effect after multiple doses in Carrageenan induced paw inflammation in rats. Further study is needed to explain this dual effect.