Pullulan based oral thin film formulation of zolmitriptan: Development and optimization using factorial design.

The goal of study was to formulate and characterize pullulan based oral thin film (OTF) of zolmitriptan by solvent casting method. Based on preliminary trials, glass, PEG 400 and sucralose were selected as casting surface, water-miscible plasticizer and sweetener for OTF, respectively. A 32 factorial design was used to study the effect of amount of PEG 400 (X1) and sucralose (X2) as independent variables on tensile strength (Y1), elasticity (Y2), % in-vitro drug release in phosphate buffer of pH 6.8 at 5min (Q5min, Y3) and overall taste of OTF (Y4) as responses. OTF of batch F4 (PEG 400, 200mg; sucralose, 12mg) was identified as an optimized batch showing in-vitro, in-vivo disintegration time 20.70 and 21.58s, respectively; 95.53% Q5min; satisfactory thickness, strength, % elongation, ease of handling, smooth mouthfeel, excellent overall taste; even distribution of all ingredients in pullulan OTF (SEM study); and stable film at specified conditions concluding that pullulan, PEG 400 and sucralose are used in combination to make palatable, stable OTF of zolmitriptan.

Determination of ergocalciferol in human plasma after Diels-Alder derivatization by LC-MS/MS and its application to a bioequivalence study.

An accurate, sensitive and selective method is developed for determination of ergocalciferol (vitamin D2) in human plasma using LC-MS/MS. After liquid-liquid extraction with n-hexane, ergocalciferol was derivatized by reacting with 4-phenyl-1,2,4-triazoline-3,5-dione (PTAD), a strong dienophile based on Diels-Alder reaction. Ergocalciferol and its deuterated internal standard, ergocalciferol-d6, were analyzed on X Select CSH C18 (100 mm×4.6 mm, 2.5 µm) column using acetonitrile and 0.1% (v/v) formic acid in water containing 0.14% methylamine within 6.0 min under gradient elution mode. Tandem mass spectrometry in positive ionization mode was used to quantify ergocalciferol by multiple reaction monitoring (MRM). Entire data processing was done using Watson LIMS™ software which provided excellent data integrity and high throughput with improved operational efficiency. The major advantage of this method includes higher sensitivity (0.10 ng/mL), superior extraction efficiency (≥83%) and small sample volume (100 µL) for processing. The method was linear in the concentration range of 0.10-100 ng/mL for ergocalciferol. The intra-batch and inter-batch accuracy and precision (% CV) values varied from 97.3% to 109.0% and 1.01% to 5.16%, respectively. The method was successfully applied to support a bioequivalence study of 1.25 mg ergocalciferol capsules in 12 healthy subjects.

Determination of ergocalciferol in human plasma after Diels-Alder derivatization by LC–MS/MS and its application to a bioequivalence study / 药物分析学报

An accurate, sensitive and selective method is developed for determination of ergocalciferol (vitamin D2) in human plasma using LC–MS/MS.After liquid-liquid extraction with n-hexane,ergocalciferol was derivatized by reacting with 4-phenyl-1,2,4-triazoline-3,5-dione (PTAD), a strong dienophile based on Diels-Alder reaction. Ergocalciferol and its deuterated internal standard, ergocalciferol-d6, were analyzed on X Select CSH C18 (100 mm×4.6 mm, 2.5μm) column using acetonitrile and 0.1% (v/v) formic acid in water containing 0.14% methylamine within 6.0 min under gradient elution mode. Tandem mass spectrometry in positive ionization mode was used to quantify ergocalciferol by multiple reaction monitoring(MRM).Entire data processing was done using Watson LIMS? software which provided excellent data integrity and high throughput with improved operational efficiency.The major advantage of this method includes higher sensitivity(0.10 ng/mL), superior extraction efficiency(≥83%)and small sample volume(100μL)for processing.The method was linear in the concentration range of 0.10–100 ng/mL for ergocalciferol.The intra-batch and inter-batch accuracy and precision (% CV) values varied from 97.3% to 109.0% and 1.01% to 5.16%, respectively. The method was successfully applied to support a bioequivalence study of 1.25 mg ergocalciferol capsules in 12 healthy subjects.

High-sensitivity simultaneous liquid chromatography-tandem mass spectrometry assay of ethinyl estradiol and levonorgestrel in human plasma.

A sensitive and simultaneous liquid chromatography-tandem mass spectrometry method was developed and validated for quantification of ethinyl estradiol and levonorgestrel. The analytes were extracted with methyl-tert-butyl ether: n-hexane (50:50, v/v) solvent mixture, followed by dansyl derivatization. The chromatographic separation was performed on a Kinetex C18 (50 mm×4.6 mm, 2.6 µm) column with a mobile phase of 0.1% (v/v) formic acid in water and acetonitrile in gradient composition. The mass transitions were monitored in electrospray positive ionization mode. The assay exhibited a linear range of 0.100-20.0 ng/mL for levonorgestrel and 4.00-500 pg/mL for ethinyl estradiol in human plasma. A run time of 9.0 min for each sample made it possible to analyze a throughput of more than 100 samples per day. The validated method has been successfully used to analyze human plasma samples for application in pharmacokinetic and bioequivalence studies.

Development and optimization of modified release IPN macromolecules of oxcarbazepine using natural polymers.

The study shows development and optimization of modified release interpenetrating polymer network (IPN) macromolecules (beads) of oxcarbazepine using sodium alginate-egg albumin prepared by ionotropic gelation method and CaCl2 as a cross-linker. Independent variables were identified based on preliminary study of investigation. The effect of amount of both polymers on drug entrapment efficiency (DEE,%), bead size (µm) and cumulative drug release at 8 h (Q8h, %) were optimized using 3(2) factorial design. The DEE, average size and Q8h were found in the range of 65.08-91.02%, 976-1084 µm and 73.50-94.06% respectively. The beads were also characterized by FTIR, DSC, SEM and XRD. The experiential responses were coincided well with predicted values obtained by Design-Expert(®) 8.0.6.1 software. The swelling of beads were influenced by the pH of a release medium. The in vitro drug release from IPN beads exhibited sustained release Hixson-Crowell pattern with anomalous non-Fickian diffusion mechanism concluding that the developed sodium alginate-egg albumin IPN composite beads are suitable for sustained delivery of oxcarbazepine for desired period.

Chromatographic separation of (E)- and (Z)-isomers of entacapone and their simultaneous quantitation in human plasma by LC-ESI-MS/MS.

A selective, sensitive and high throughput liquid chromatography-tandem mass spectrometry (LC-ESI-MS/MS) method has been developed and validated for the chromatographic separation and quantitation of (E)-entacapone and (Z)-entacapone in human plasma. Sample clean-up involved liquid-liquid extraction (LLE) of both the isomers and carbamazepine used as internal standard from 500microL of human plasma. Both the analytes were chromatographically separated with a resolution factor of 3.0 on a Gemini C18 (50mmx4.6mm, 5microm particle size) analytical column using 1% formic acid and methanol (50:50, v/v) as the mobile phase. The selectivity factor (alpha) of the column for the separation was 2.0, based on the capacity factors of 2.6 and 1.3 for (E)- and (Z)-isomers respectively. The parent-->product ion transitions for both the isomers (m/z 306.1-->233.0) and IS (m/z 237.3-->194.2) were monitored on a triple quadrupole mass spectrometer, operating in the multiple reaction monitoring (MRM) and positive ion mode. The method was validated over the concentration range of 24.3-6076ng/mL and 23.8-5960ng/mL for (E)-entacapone and (Z)-entacapone respectively. Matrix effect was assessed by post-column analyte infusion experiment and the process/extraction efficiency found was 94.3% and 89.3% for (E)- and (Z)-isomers respectively. The method was successfully applied to a pivotal bioequivalence study in 36 healthy human subjects after oral administration of 200mg (E)-entacapone tablet formulation under fasting conditions.

Isolation of Moraxella bovis from frozen bovine semen and determination of microbial load.

In the present study, isolation of Moraxella bovis was done from the microbiological examination of frozen semen straws from clinically healthy twelve Jersey and one Jersey-cross bull supplied by a semen laboratory. The organism was identified on the basis of colonial morphology and biochemical characteristics. Further, viable cell count of the bacterium was studied in detail in the semen sample.