Effects of lipopolysaccharide on follicular estrogen production and developmental competence in bovine oocytes.

Reproductive efficiency and female fertility is essential for productive and sustainable beef cattle operations. Gram-negative bacterial infections cause release of the endotoxin lipopolysaccharide (LPS) which initiates immune responses shown to alter ovarian steroidogenesis and impair oocyte development. The current study was designed to investigate the impact of varying levels of naturally occurring infection and follicular LPS on estradiol (E2) production and oocyte maturation. Bovine ovary pairs were harvested from a slaughterhouse, and oocytes were aspirated from small follicles and matured in vitro. Meiotic events were evaluated on nuclear maturation and spindle morphology to classify oocytes as normal or abnormal. Follicular fluid LPS concentrations were measured and subsequently separated into Low or High LPS groups. A marked difference was detected between the percent of abnormal oocytes matured from Low LPS follicles, compared to the percent of abnormal oocytes matured from High LPS follicles (P = 0.1). Follicular E2 concentrations tended to be greater for high LPS follicles (P = 0.1), however, relative abundance of mRNA transcripts for aromatase (P = 0.93) and beta-catenin (P = 0.63) were similar between groups. No changes were detected in Toll-like Receptor 4 (P = 0.15), Myeloid Differentiation Factor-2 (P = 0.61), or cluster of differentiation 14 (P = 0.46) mRNA transcript abundance in follicles with high LPS, compared to low. Therefore, even Low levels of follicular LPS indicating a subacute infection is capable of impacting the ovarian milieu and may represent an unappreciated factor leading to reduced female fertility and decreased cow retention.

Simple, economical, and reproducible LC-MS method for the determination of amoxicillin in human plasma and its application to a pharmacokinetic study.

A simple, economical, and reproducible high-performance liquid chromatography mass spectrometric (MS) method is developed and validated for the determination of amoxicillin in human plasma. The present method has been successfully used to determine bioequivalence between a test and innovator formulation of amoxicillin 500 mg capsules. The method is validated in terms of selectivity, precision/accuracy, recovery, dilution integrity, matrix effect, effect of anti-coagulant, and stability studies. Sample preparation is carried out by solid-phase extraction (HLB Oasis cartridges). The processed sample is chromatographed on Hypersil Gold (4.6 x 50 mm); 3 microm C18 column, using 10mM ammonium formate buffer (pH 5.0) and acetonitrile, (10:90, v/v) as mobile phase. Amoxicillin is detected by MS-MS detection with turbo-ion spray in positive ion mode. The weighed (1/X2) calibration curves were linear over the range of 0.17 to 17.0 microg/mL. The intra day precision is from 1.3% to 8.8% and intra day accuracy is 94.1% to 108.5%. The inter day precision is from 1.8% to 6.2% and inter-day accuracy is 95.1% to 105.9%. Mean recovery of 66.3% is observed for amoxicillin and 71.6% for internal standard (ampicillin). The stability of amoxicillin is studied at -15 degrees C and -50 degrees C using human plasma with different anti-coagulants (citrate, monobasic sodium phosphate, dextrose, and adenine-citrate, monobasic sodium phosphate, dextrose, and adenine and ethylene diamine tetraacetic acid-ethylene diamine tetraacetic acid). No significant degradation is observed for 60 days.

A bioequivalence study comparing two formulations of lopinavir/ritonavir capsules.

This investigation was carried out to evaluate the bioavailability of a new single fixed-dose combination formulation of lopinavir and ritonavir, relative to reference product, Kaletra (133.3 mg lopinavir/33.3 mg ritonavir) capsules, manufactured by Abbott Laboratories, Chicago, IL, USA. The bioavailability study was carried out on 72 healthy male and female volunteers who received a single dose of 3 capsules (133.3 mg lopinavir/33.3 mg ritonavir) of the test (T) and the reference (R) products in the fasting state, in a randomized, balanced, 2-way crossover design. After dosing, serial blood samples were collected for a period of 72 hours. Plasma harvested from blood was analyzed for lopinavir and ritonavir by a sensitive and validated simultaneous liquid-chromatographic and mass-spectrometric (LC-MS/MS) assay. Mean oral clearance (Cl/F) values of the FDC were 4.92 and 23.54 l/h for lopinavir and ritonavir, respectively, the maximum plasma concentrations (C(max)), area under the plasma concentration-time curve up to the last measurable concentration (AUC(0-t)), and to infinity (AUC(0-infinity)), were analyzed statistically under the assumption of a multiplicative model. The time to maximum concentration (t(max)) was analyzed assuming an additive model. The parametric confidence intervals (90%) were calculated by Schuirmann's two 1-sided t-test criteria. It was found that the test/reference (T/R) ratios for the pharmacokinetic parameters AUC(0-t), AUC(0-infinity) and C(max) (after initial log transformation) were well within the bioequivalence acceptance range of 80-125% as per international regulatory guidelines. Therefore, the two formulations were considered to be bioequivalent [Food and Drug Administration 2003].