Stereoselective pharmacokinetics and tissue distribution of levodropropizine after administration of pure levodropropizine and the rac-dropropizine to Sprague-Dawley rats.

Levodropropizine (LDP) is a non-opioid anti-tussive. The stereoselective pharmacokinetics and tissue distribution (TD) of LDP vs. dextrodropropizine (DDP) have been characterized after oral and intravenous (IV) administration of LDP and rac-dropropozine in rats.Oral/IV doses of 50/5.0 mg/kg and 25/2.5 rac-dropropizine and LDP were employed. TD study focused on tissues such as liver, lung and kidney. Blood samples were collected for pharmacokinetic and TD evaluation. Validated methods were used to quantitate LDP, DDP and rac-dropropizine.No stereoselectivity in pharmacokinetics was observed between LDP vs. DDP following rac-dropropizine. However, LDP pharmacokinetics after LDP administration (oral/IV) appeared to be different compared to LDP derived from rac-dropropizine.TD data were similar between the two enantiomers regardless of oral/IV rac-dropropizine administration. When LDP alone was administered, levels were comparable to those derived for LDP from rac-dropropizine after oral/IV. However, in the lung and kidney tissues, the exposure after oral dosing was higher for LDP alone as compared to LDP from rac-dropropizine.In summary, complete characterization of stereoselective pharmacokinetics and TD of rac-dropropizine has been reported after oral/IV routes. It was evident that the presence of DDP, increased the plasma/tissue exposure of LDP which was evident after oral rac-dropropizine dosing.

Enantioselective LC-ESI-MS/MS method for quantitation of (-)-lumefantrine and (+)-lumefantrine in mice plasma and application to a pharmacokinetic study.

We developed and validated a simple, sensitive, selective, and reliable LC-MS/MS-ESI method for the direct quantitation of lumefantrine (LFN) enantiomers [(-)-LFN and (+)-LFN] in mice plasma as per regulatory guideline. LFN enantiomers and carbamazepine (internal standard) were extracted from mice plasma using Strata X SPE (solid-phase extraction) cartridges. Good resolution between enantiomers was achieved on a Chiralpak IA-3 column using an isocratic mobile phase (0.1% of diethyl amine in methanol), which was delivered at a flow rate of 0.8 mL/min. Detection and quantitation were performed using multiple reaction monitoring mode following the transitions m/z 530.27 → 512.30 and 237.00 → 194.00 for LFN enantiomers and the internal standard, respectively, in the positive-ionization mode. The proposed method provided accurate and reproducible results over the linearity range of 2.39-895 ng/mL for each enantiomer. The intra- and inter-day precisions were in the range of 1.03-6.14 and 6.36-8.70 and 2.03-4.88 and 5.82-11.5 for (-)-LFN and (+)-LFN, respectively. Both (-)-LFN and (+)-LFN were found to be stable under different stability conditions. The method was successfully used to delineate stereoselective pharmacokinetics of LFN enantiomers in mice after an oral administration of rac-LFN (20 mg/kg). The pharmacokinetic results indicated that the disposition of LFN enantiomers was stereoselective in mice.

Enantioselective LC-ESI-MS/MS determination of dropropizine enantiomers in rat plasma and application to a pharmacokinetic study.

We developed and validated a simple, sensitive, selective and reliable LC-ESI-MS/MS method for direct quantitation of dropropizine enantiomers namely levodropropizine (LDP) and dextrodropropizine (DDP) in rat plasma without the need for derivatization as per regulatory guidelines. Dropropizine enantiomers and carbamazepine (internal standard) were extracted from 50 µL rat plasma using ethyl acetate. LDP and DDP resolved with good baseline separation (Rs = 4.45) on a Chiralpak IG-3 column. The mobile phase consisted of methanol with 0.05% diethylamine pumped at a flow rate of 0.5 mL/min. Detection and quantitation were done in multiple reaction monitoring mode following the transitions m/z 237 → 160 and 237 → 194 for dropropizine enantiomers and the internal standard, respectively, in the positive ionization mode. The proposed method provided accurate and reproducible results over the linearity range of 3.23-2022 ng/mL for each enantiomer. The intra- and inter-day precisions were in the ranges of 3.38-13.6 and 5.11-13.8 for LDP and 4.19-11.8 and 8.89-10.1 for DDP. Both LDP and DDP were found to be stable under different stability conditions. The method was successfully used in a stereoselective pharmacokinetic study of dropropizine enantiomers in rats following oral administration of racemate dropropizine at 100 mg/kg. The pharmacokinetic results indicate that the disposition of dropropizine enantiomers is not stereoselective and chiral inversion does not occur in rats.

Validation of a chiral LC-MS/MS-ESI method for the simultaneous quantification of darolutamide diastereomers in mouse plasma and its application to a stereoselective pharmacokinetic study in mice.

A simple, selective and reliable LC-MS/MS method was validated for simultaneous quantitation of darolutamide diastereomers in 50 µL mouse plasma using warfarin as an internal standard (IS) as per regulatory guidelines. Plasma samples were extracted by liquid-liquid extraction and the chromatographic separation was achieved on a Chiralpak IA column with an isocratic mobile phase 5 mm ammonium acetate-absolute alcohol (20:80, v/v) at a flow rate of 1.0 mL/min. Detection and quantitation was done in multiple reaction monitoring mode following the transitions m/z 397 → 202 and 307 → 250 for darolutamide diastereomers and the IS, respectively, in the negative ionization mode. The linearity range was 100-2400 ng/mL for each diastereomer. The intra- and inter-day precisions were in the ranges of 1.78-4.20 and 4.34-14.6, and 3.63-4.74 and 4.78-5.15 for diastereomer-1 and diastereomer-2, respectively. Both diastereomers were found to be stable under different stability conditions. The validated method was applied to a pharmacokinetic study in mice. Following oral administration of darolutamide at 10 mg/kg, maximum concentration in plasma was 4189 and 726 ng/mL for diastereomer-1 and diastereomer-2, respectively. The terminal half-life was found to be ~0.50 h for both the diastereomers. The AUC(0-t) was found to be 18,961 ng*h/mL for diastereomer-1 and 1340 ng*h/mL diastereomer-2.

Validation of an enantioselective LC-MS/MS method to quantify enantiomers of (±)-OTX015 in mice plasma: Lack of in vivo inversion of (-)-OTX015 to its antipode.

A highly sensitive, specific and enantioselective assay has been validated for the quantitation of OTX015 enantiomers [(+)-OTX015 and (-)-OTX015] in mice plasma on LC-MS/MS-electrospray ionization as per regulatory guidelines. Protein precipitation was used to extract (±)-OTX015 enantiomers and internal standard (IS) from mice plasma. The active [(-)-OTX015] and inactive [(+)-OTX015] enantiomers were resolved on a Chiralpak-IA column using an isocratic mobile phase (0.2% ammonia/acetonitrile 20 : 80, v/v) at a flow rate of 1.2 mL/min. The total run time was 6.0 min. (+)-OTX015, (-)-OTX015 and IS eluted at 3.34, 4.08 and 4.77 min, respectively. The MS/MS ion transitions monitored were m/z 492 → 383 for OTX015 and m/z 457 → 401 for IS. The standard curves for OTX015 enantiomers were linear (r2 > 0.998) in the concentration range 1.03-1030 ng/mL. The inter- and intraday precisions were in the range 2.20-13.3 and 8.03-12.1% and 3.80-14.4 and 8.97-13.6% for (+)-OTX015 and (-)-OTX015, respectively. Both the enantiomers were found to be stable in a battery of stability studies. This novel method has been applied to the study of stereoselective oral pharmacokinetics of (-)-OTX015 and unequivocally demonstrated that (-)-OTX015 does not undergo chiral inversion to its antipode in vivo in mice.

Pharmacokinetics of Darolutamide in Mouse - Assessment of the Disposition of the Diastereomers, Key Active Metabolite and Interconversion Phenomenon: Implications to Cancer Patients.

BACKGROUND: Darolutamide is recently approved for the treatment of non-metastatic castrate resistance prostate cancer. Hitherto, no stereoselective pharmacokinetic data have been published pertaining to darolutamide and its diastereomers in animals or humans. The key aims of the experiment were to examine darolutamide, S,S-darolutamide and S,R-darolutamide with respect to (a) assessment of in vitro metabolic stability and protein binding and (b) characterization of in vivo oral and intravenous pharmacokinetics in mice. METHODS: In vitro (liver microsomes stability and protein binding) and in vivo experiments (oral/intravenous dosing to mice) were carried out using darolutamide, S,S-darolutamide and S,Rdarolutamide. Besides, tissue levels of darolutamide, S,S-darolutamide and S,R-darolutamide were measured following oral and intravenous dosing. Appropriate plasma/tissue samples served to determine the pharmacokinetics of various analytes in mice. Liquid chromatography in tandem with mass spectrometry procedures enabled the delineation of the plasma pharmacokinetics, in vitro and tissue uptake data of the various analytes. RESULTS: Chiral inversion was absent in the metabolic stability study. However, darolutamide showed profound stereoselectivity (S,S-darolutamide greater than S,R-darolutamide) after either intravenous or oral dosing. S,R-darolutamide but not S,S-darolutamide showed conversion to its antipode post oral and intravenous dosing to mice. Regardless of oral or intravenous dosing, active keto darolutamide formation was evident after administration of darolutamide, S,S-darolutamide or S,R- darolutamide. Tissue data supported the observations in plasma; however, tissue exposure of darolutamide, S,Sdarolutamide and S,R-darolutamide was much lower as compared to plasma. CONCLUSION: In lieu of the human pharmacokinetic data, although the administration of diastereomeric darolutamide was justified, it is proposed to delineate the clinical pharmacokinetics of S,Rdarolutamide and S,S-darolutamide relative to darolutamide in future clinical pharmacology studies.

A prospective transmission electron microscopic study of muscle status in oral submucous fibrosis along with retrospective analysis of 80 cases of oral submucous fibrosis.

AIM AND OBJECTIVE: The present study is undertaken to study the ultra structural features of muscle tissue in moderate and advanced stages of oral submucous fibrosis along with retrospective analysis of 80 cases of oral submucous fibrosis ( osmf) 0 during the period of year 2002 to 2005. MATERIALS AND METHODS: Five patients with moderate and advanced stages of osmf0 were screened from outpatients department of oral diagnosis, sri Ramachandra dental college and hospital. After a detailed case history, they were subjected to incisional biopsy from an area of buccal mucosa with maximum palpable fibrotic bands.the specimens were cut into two halves, one half was fixed in 10% formalin for routine processing. Second half was fixed in 2.5% glutaraldehyde for electron microscopic examination. RESULTS: Prospective study of muscle fibres from moderate and advanced stages of osmf0 revealed varying changes in high proportion of muscle fibres which includes, irregularity of surface of fibre,sarcolemmal foldings, reduplicated basement membrane, loss and alterations in the myofilaments, hypercontraction of myofibrils, Z line abnormalities, internal nucleus, autophagic vacuoles. These features are suggestive of muscle atrophy and necrosis. CONCLUSION: Within the limitations of this study, it can be concluded that the ultra structural features In moderate and advanced stages of osmf0 were best studied. These muscle changes can be manifestation of disease, atrophy being secondary to the limited functional activity of the muscles which is brought about by fibrosis or it could be essential part of the disease process itself.