Search

Background: The number of validated quantification methods for rifampicin, a prototypical Oatp inhibitor, in biological rat samples is limited. Objective: This study was conducted to validate a modified reversed-phase liquid chromatographic method for the determination of rifampicin in rat liver tissue according to the current ICH M10 Bioanalytical Method Validation Draft Guideline (2019) for application to samples of in situ rat liver perfusion studies. Methods: Liver tissue samples were obtained from recirculatory in situ rat liver perfusion studies. The analysis was performed on a C18 column with a mobile phase composed of 0.05 M phosphate buffer (pH 4.58): acetonitrile (55:45, v/v). The assay was validated for selectivity, calibration curve and range, matrix effect, carry-over, accuracy and precision, reinjection reproducibility, and stability. Results: The method was considered selective and stable, without having carry-over and matrix effects. The calibration curve was linear (R2: 0.9983) within the calibration range (0.5-60 ppm). Accuracy and precision values fulfilled the required limits. Liver concentrations of rifampicin in liver tissue, obtained after 60 min perfusion with 10 μM and 50 μM of rifampicin, were 45.1 ± 11.2 and 313.4 ± 84.4 μM, respectively. Conclusion: The bioanalytical method validation was completed and the method was successfully applied for the determination of rifampicin in rat liver tissue.

Background: Metabolomics is one of the main areas to understand cellular process at molecular level by analyzing metabolites. In recent years metabolomics has emerged as a key tool to understand molecular basis of diseases, to find diagnostic and prognostic biomarkers and develop new treatment opportunities and drug molecules. Objective: In this study, untargeted metabolite and lipid analysis were performed to identify potential biomarkers on premature ovarian insufficiency plasma samples. 43 POI subject plasma samples were compared with 32 healthy subject plasma samples. Methods: Plasma samples were pooled and extracted using chloroform:methanol:water (3:3:1 v/v/v) mixture. Agilent 6530 LC/MS Q-TOF instrument equipped with ESI source was used for analysis. A C18 column (Agilent Zorbax 1.8 μM, 50 x 2.1 mm) was used for separation of the metabolites and lipids. XCMS, an “R software” based freeware program, was used for peak picking, grouping and comparing the findings. Isotopologue Parameter Optimization (IPO) software was used to optimize XCMS parameters. The analytical methodology and data mining process were validated according to the literature. Results: 83 metabolite peaks and 213 lipid peaks were found to be in semi-quantitatively and statistically different (fold change >1.5, p <0.05) between the POI plasma samples and control subjects. Conclusion: According to the results, two groups were successfully separated through principal component analysis. Among the peaks, phenyl alanine, decanoyl-L-carnitine, 1-palmitoyl lysophosphatidylcholine and PC(O-16:0/2:0) were identified through auto MS/MS and matched with human metabolome database and proposed as plasma biomarker for POI and monitoring the patients in treatment period.

Background: The aim of this study was to develop HPLC methods for individual determination of 20 active pharmaceutical ingredients (amoxicillin sodium, antipyrine, atenolol, caffeine, carbamazepine, cimetidine, enalapril, furosemide, hydrochlorothiazide, ibuprofen, ketoprofen, metoprolol tartrate, methyldopa, naproxen sodium, pindolol, piroxicam, propranolol HCl, ranitidine, theophylline, and verapamil HCl) to be used for determination of their intestinal permeabilities across Ussing-Chamber. Method: Two different stationary phases (Waters X-Bridge C18- 150 x 4.6 mm, 5 μm; and ACE 5 C18- 150 x 4.6 mm, 5μm) were used for the separation of the compounds. Three different aqueous phases (20 mM phosphate buffers pH 3.0, pH 6.0 and water) and two different organic phases (methanol and acetonitrile) were used to prepare the mobile phases. Total analysis time was shorter than 7 minutes for all applications. Result: The developed methods were validated according to the ICH guideline and found to be linear, sensitive, selective, precise and accurate. The developed methods could be applied for analyses of these compounds not only for Ussing-Chamber studies but also for other permeability studies.

The main objective of this study was to investigate the effect of pH on the solubility and intestinal permeability of ketoprofen by using HPLC. Ketoprofen is a slightly soluble acidic drug, and its solubility in aqueous phase is affected by pH changes dramatically. However, there is no data in the literature to support whether this pH dependent water solubility of ketoprofen will influence its absorption/ bioavailability. In this study, the distribution-coefficient (log D) of ketoprofen at various pH values between 4.5 and 7.5 were investigated using HPLC, and then it was made an attempt to correlate the log D values with the intestinal permeability values. For this reason, in vivo intestinal permeability studies were performed at pH 4.5 and pH 7.5. The concentrations of model and reference compounds and also the blank buffers passed though the rat intestine were analyzed by means of a validated HPLC method. A nonlinear relationship was found between the results of in vitro and in vivo studies indicating that the diffusion of ketoprofen was not only related with passive diffusion, but also could be related with active transport.