Determination of Mirabegron in rat plasma by UPLC-MS/MS after oral and intravenous administration

SUMMARY Mirabegron is a kind of β3 adrenergic receptor agonist which is an effective drug for the treatment of overactive bladder. In this research, a UPLC-MS/MS method is developed and validated for the study of mirabegron pharmacokinetic in rats. A protein precipitation method is applied for sample preparation with acetonitrile. m/z 397.3→379.6, m/z 326.4→121.0 for mirabegron, tolterodine (IS), respectively in the positive ion mode was performed for quantitation. The method is reliable and reproducible in our study (intra-day precision≤11.06%, inter-day precision≤11.43%) with concentration curves linear from 5 to 2500 ng/mL(R2>0.999). Stability studies demonstrated that mirabegron was stable under a variety of storage conditions. This method was successfully applied for determining mirabegron in rats after oral and intravenous administration.

RESUMO Mirabegron é um tipo de agonista do receptor adrenérgico beta 3 que demonstra eficácia no tratamento de bexiga hiperativa. Nesta pesquisa, o método UPLC-MS/MS é desenvolvido e validado para o estudo da farmacocinética mirabegron em ratos. Um método de precipitação de proteínas é aplicado para a preparação de amostras com acetonitrilo. 397.3 → 379.6 M / Z, M / Z 326.4 → 121.0 para mirabegron, tolterodina (IS), respectivamente, para o íon positivo foi realizado para quantificação. O método é fiável e reprodutível em nosso estudo (precisão intradia ≤ 11,06%; precisão entredia ≤ 11.43%), com curvas de concentração linear de 5 a 2 ng/ml (R2 > 0,999). Estudos de estabilidade demonstraram que mirabegron permanece estável sob uma variedade de condições de armazenamento. Este método foi aplicado com sucesso para a determinação de mirabegron em ratos após administração oral e intravenosa.

Quantitation of meloxicam in the plasma of koalas (Phascolarctos cinereus) by improved high performance liquid chromatography

An improved method to determine meloxicam (MEL) concentrations in koala plasma using reversed phase high performance liquid chromatography equipped with a photo diode array detector was developed and validated. A plasma sample clean-up step was carried out with hydrophilic-lipophilic copolymer solid phase extraction cartridges. MEL was separated from an endogenous interference using an isocratic mobile phase [acetonitrile and 50 mM potassium phosphate buffer (pH 2.15), 45:55 (v:v)] on a Nova-Pak C18 4-microm (300 x 3.9 mm) column. Retention times for MEL and piroxicam were 8.03 and 5.56 min, respectively. Peak area ratios of MEL to the internal standard (IS) were used for regression analysis of the calibration curve, which was linear from 10 to 1,000 ng/mL (r2 > 0.9998). Average absolute recovery rates were 91% and 96% for MEL and the IS, respectively. This method had sufficient sensitivity (lower quantitation limit of 10 ng/mL), precision, accuracy, and selectivity for routine analysis of MEL in koala plasma using 250-microL sample volumes. Our technique clearly resolved the MEL peak from the complex koala plasma matrix and accurately measured MEL concentrations in small plasma volumes.

Quantitation of meloxicam in the plasma of koalas (Phascolarctos cinereus) by improved high performance liquid chromatography

An improved method to determine meloxicam (MEL) concentrations in koala plasma using reversed phase high performance liquid chromatography equipped with a photo diode array detector was developed and validated. A plasma sample clean-up step was carried out with hydrophilic-lipophilic copolymer solid phase extraction cartridges. MEL was separated from an endogenous interference using an isocratic mobile phase [acetonitrile and 50 mM potassium phosphate buffer (pH 2.15), 45:55 (v:v)] on a Nova-Pak C18 4-microm (300 x 3.9 mm) column. Retention times for MEL and piroxicam were 8.03 and 5.56 min, respectively. Peak area ratios of MEL to the internal standard (IS) were used for regression analysis of the calibration curve, which was linear from 10 to 1,000 ng/mL (r2 > 0.9998). Average absolute recovery rates were 91% and 96% for MEL and the IS, respectively. This method had sufficient sensitivity (lower quantitation limit of 10 ng/mL), precision, accuracy, and selectivity for routine analysis of MEL in koala plasma using 250-microL sample volumes. Our technique clearly resolved the MEL peak from the complex koala plasma matrix and accurately measured MEL concentrations in small plasma volumes.

Improved method of total antioxidant assay.

Commercially available analytical kits for the estimation of total antioxidant status are expensive and time-consuming. Most of the commercially available kits for total antioxidants estimation are based on the principle of suppression of ABTS radical cation formation by antioxidant in the serum sample. The method requires stringent assay conditions, like exact incubation time and the temperature (37 degrees C) of the reaction and on an average not more than 40 samples can be analyzed on a day. We have adapted the assay to a microplate, thereby allowing more number of samples to be analyzed per day. Further, the reagent volume required is one fourth than that for the original procedure thereby cutting cost. Thirty samples were analyzed by original method on spectrophotometer and our adapted microplate assay. The values of total antioxidant obtained by the two methods correlated well. Thus, total antioxidant can be estimated reliably using the microplate method.

Letrozole compared to clomiphene citrate in patients with unexplained infertility undergoing IUI

To compare the efficacy of Ietrozole as ovulation induction agent for patients with unexplained infertility as regards is mean number of follicles resulting, endumetrial thickness and pregnancy rate [PR] in comparison to CC alone and in combination with gonadotropines [HMG] and Ietrozole in combination with HMG for COH and IUI A prospective randomized comparative study. Infertility outpatient clinic, Faculty of Medicine. Consequtive patients who agreed to participate were randomized on an alternating basis to receive either 2.5 mg bid letrozole or 50 mg bid CC on days 3-7, or letrozole followed by HMG, or CC followed by HMG 75IU/day until the day of HCG [15 patients in each group].Folliculometry starting D7 was done till the Ieading follicle[s] reached 18mm ovulation was triggered by HCG 10000 IU and endometrial thickness was measured on the day of HCG administration. A single IUI was done 36 hours after HCG administration. The luteal phase was supported by vaginal micronized progesterone 600 mg/day. Pregnancy was established by blood beta-subunit detection 2 weeks after IUI. A total of 60 patients [15 patients in each group] meeting the inclusion criteria were included All patients had primary unexplained infertility of less than 3 years and there was no statistically significant difference in age nor BMI between the 4 groups. The outcome measures we examined were endometrial thickness, mean number of mature follicles > 18 mm on day of HCG administration and pregnancy rate. The mean endometrial thickness was: 8.8 mm in the letrozole group, 8.3 mm in the CC group, 8.5 mm in the letrozole + HMG group. 9.9 mm in the CC + HMG group i.e. the showed the least mean endometrial thickness but with no statistically significant difference between the 4 groups. mean number of mature follicles >18 mm on the day of HCG was: 3.2 in the letrozole group. 2 6 in the CC group, 2.4 in the letrozole + HMG group .3.4 in the CC + HMG group with no statistically significant difference between the 4 groups. Pregnancy rate per cycle for the 4 groups was: 20% in the letrozole group, 40% in the CC' group, 13.3% in the letrozole + HMG group, 26.7% in the CC + HMG group with no statistically significant difference between the 4 groups. In conclusion we tailed to prove that there is clinical preference of Ietrozole or combined protocols over CC alone in COH + IUI or women with unexplained infertility, this could be attributed to the small sample size, bigger study is needed before Ietrozole is prescribed as a first- line induction drug being more expensive than CC