HPLC method development for the simultaneous analysis of amlodipine and valsartan in combined dosage forms and in vitro dissolution studies

A simple, rapid and reproducible HPLC method was developed for the simultaneous determination of amlodipine and valsartan in their combined dosage forms, and for drug dissolution studies. A C18 column (ODS 2, 10 μm, 200 x 4.6 mm) and a mobile phase of phosphate buffer (pH 3.6 , 0.01 mol L-1):acetonitrile: methanol (46:44:10 v/v/v) mixture were used for separation and quantification. Analyses were run at a flow-rate of 1 mL min-1 and at ambient temperature. The injection volume was 20 μL and the ultraviolet detector was set at 240 nm. Under these conditions, amlodipine and valsartan were eluted at 7.1 min and 3.4 min, respectively. Total run time was shorter than 9 min. The developed method was validated according to the literature and found to be linear within the range 0.1 - 50 μg mL-1 for amlodipine, and 0.05 - 50 μg mL-1 for valsartan. The developed method was applied successfully for quality control assay of amlodipine and valsartan in their combination drug product and in vitro dissolution studies.Desenvolveu-se método de HPLC rápido e reprodutível para a determinação simultânea de anlodipino e valsartana em suas formas de associação e para os estudos de dissolução dos fármacos. Utilizaram-se coluna C18 (ODS 2, 10 μm, 200 x 4,6 mm) e fase móvel tampão fosfato (pH 3,6, 0,01 mol L-1):acetonitrila: metanol para a separação e a quantificação. As análises foram efetuadas com velocidade de fluxo de 1 mL min-1 e à temparatura ambiente O volume de injeção foi de 20 μL e utilizou-se detector de ultravioleta a 240 nm. Sob essas condições, anlodipino e valsartana foram eluídas a 7,1 min e 3,4 min, respectivamente. O tempo total de corrida foi menor que 9 min. O método desenvolvido foi validado de acordo com a literatura e se mostrou linear na faixa de 0,1-50 μg mL-1 para anlodipino e de 0,05-50 μg mL-1 para valsartana. O método desenvolvido foi aplicado com sucesso para ensaios de controle de qualidade de associações de anlodipino e valsartana e nos estudos de dissolução in vitro

Gıda kaynaklı polifenollerin K562 lösemi hücreleri üzerine in vitro çoğalmayı önleyici etkisinin karşılaştırmalı proteomik çalışmalarla incelenmesi

[Objective]: The lack of success with classical targeted therapies in cancer treatment has forced researchers to employ either combined therapies or agents that interfere with multiple pathways. In this study, a proteomic evaluation was performed to understand the in vitro antiproliferative effect of dietary compounds against leukemia cells in proteome level. Typical human cell models of leukemia (i.e., wild K562 cells and multi-drug resistant leukemia cells K562/R) were treated with rosemary (Rosmarinus officinalis L.) extracts rich in polyphenols and a comparative proteomic study was performed to identify up- or down-regulated proteins by in vitro antiproliferative effect of polyphenols. [Methods]: The protein fraction from the polyphenol-treated and control K562 and K562/R cells were separated through two dimensional polyacrylamide gel electrophoresis (2DE) and their proteomic profiles compared by image analysis. The proteins identified to be overexpressed or underexpressed in K562 and K562/R cells after the polyphenol treatment were identified by MALDI-TOF/TOF MS. [Results]: The results in this study were evaluated to understand the mode of action of these dietary constituents against leukemia cell proliferation. Dietary polyphenols extracted from rosemary bring about the up regulation or down regulation of different proteins on leukemia cells. These proteins are related to tumorigenesis, cancer proliferation and antioxidant activity. [Conclusion]: Our results revealed that the studied rosemary extract rich in polyphenols induced the down regulation of adenine phosphoribosyl transferase and annexin A1 in K562/R cell lines and tubulin alpha-1C chain in K562 cell lines. According to the expression proteomics results in this study, it could be concluded that the rosemary polyphenols shows in vitro antiproliferative activity in K562 and K562/R leukemia cell lines.Mustafa Çelebier thanks to The Council of Higher Education, Turkey for his grant. This work was supported by AGL2008-05108-C03-01 (Ministerio de Ciencia e Innovación, Spain).Peer Reviewe

Simultaneous determination of olmesartan medoxomil and amlodipine besylate in pharmaceutical formulations by capillary zone electrophoresis

A simple, efficient and reliable capillary zone electrophoresis method with diode array detection was developed and validated for the simultaneous determination of olmesartan medoxomil and amlodipine besylate in their binary mixtures. The optimum separation for these compounds was achieved with a fused - silica capillary column (i.d. 75.0 µm, total length 48.5 cm and effective length 40.0 cm) and 40.0 mM citrate buffer at pH 6.0 as the running buffer. The samples were injected hydrodynamically for 3 s at 50 mbar and applied voltage was + 15 kV at 30 °C capillary temperature. Detection wavelength was set at 235 nm. Valsartan was used as internal standard. The method was validated with respect to stability, linearity, sensitivity, precision, accuracy, recovery and selectivity. The linear calibration range was found to be 2.00 - 30.00 µg/mL for olmesartan medoxomil and amlodipine besylate. The limits of detection (LOD) were 0.05 µg/mL for both compounds. The relative standard deviations (RSD) of the proposed method ranged between 1.51 and 2.49 % for intra-day precision and 1.51 and 3.73 % for inter-day precision. The developed and validated method successfully applied for the simultaneous determination of olmesartan medoxomil and amlodipine besylate in their pharmaceutical formulations.Colegio de Farmacéuticos de la Provincia de Buenos Aire

Ht29 Ve K562 Kanser Hücrelerinde Protein ve Metabolitlerin Analizi Için Çeşitli Analitik Yöntemlerin Geliştirilmesi

Cancer is one of the most dangerous fatal diseases threating the human being both economically and socially. Therefore, the studies are performed on molecular level to understand cancer on molecular level and prevent the people from cancer. There are mainly two different objectives in this thesis. The first objective is to investigate the molecular mechanism of the anticancer activity of Rosmarinus officinalis L. on HT29 column cancer cells and K562/R and K562/wt leukemia cells. For this purpose, the enrichment extracts of Rosmarinus officinalis including polyphenols were appiled on in vitro cultures. The proteins were separated through two dimensional gel electrophoresis and the over- or under-expressed proteins through extracts treatment on cell lines were compared. The proteins found to be changed quantitatively on treated cells and control group were analyzed by matrixassisted laser desorption/ionization - time of flight - mass spectrometry and identified through using protein databases. The identified proteins seem to be associated with the anticancer activity according to the literature. The second objective is to adapt the XCMS, which is a metabolite profiling software working under ¨R¨ language for liquid chromatography - mass spectrometry studies, on capillary electrophoresis - mass spectrometry experiments. For this purpose, the metabolite of K562/R cell line were analyzed by using capillary electrophoresis - mass spectrometry and the electropherograms were used on metabolite profiling through the evalution of XCMS parameters for capillary electrophoresis. Some of the identified metabolites were tried to be associated with the cancer.Günümüzde tedavisi zor ve ölümcül hastalıklardan biri olan kanser, sosyal ve ekonomik anlamda insanlığı tehdit etmektedir. Kanserin önlenmesi ve tedavisi ile ilgili çözüm arayışları halen devam etmektedir ve bu amaçla moleküler düzeyde çalışmalar yapılmaktadır. Bu tez çalışması başlıca iki aşamadan oluşmaktadır; birinci aşamada HT29 kolon kanseri hücreleri ve K562/R ve K562/wt kan kanseri hücreleri üzerinde biberiye (Rosmarinus officinalis L.)' nin antikanser aktivitesi karşılaştırmalı proteomik çalışmalarla moleküler düzeyde incelenmiştir. Bunun için biberiyeden elde edilen polifenolce zengin ekstrelerin kanserli hücre hatlarına in vitro olarak uygulanması sonucu hücre büyümesinde azalmanın görüldüğü işlenmiş gruplar ile kontrol gruplarına ait proteinler iki boyutlu jel elektroforez ile ayrılarak biberiye ekstreleri ile işlem sonrası düşük veya yüksek miktarda ifade edilen proteinler tespit edilmiştir. Tespit edilen proteinlerden en üst düzeyde farklılaşmış olanlar matriks destekli desorpsiyon/ iyonlaşmalı - uçuş zamanlı kütle spektrometrisi ile analiz edilmiş ve protein veribankaları aracılığıyla tanımlanmıştır. Tanımlanan proteinler, biberiyenin antikanser aktivitesini proteom düzeyinde açıklamak amacıyla kaynaklardaki çalışmalarla ilişkilendirilmiştir. İkinci aşamada ise ¨R¨ programlama dili altında çalışan ve sıvı kromatografisi - kütle spektrometrisi verilerinin değerlendirilmesinde kullanılan bir metabolit profilleme yazılımı olan XCMS, kapiler elektroforez - kütle spektrometrisi çalışmalarına uyarlanmıştır. Bu amaçla K562/R hücre hattına ait metabolitler, kapiler elektroforez - kütle spektrometrisi kullanılarak analiz edilmiş ve analiz sonucu elde edilen elektroferogramlar XCMS için çeşitli parametrelerin değerlendirilmesiyle metabolit profillemede kullanılmıştır. Tespit edilen metabolitlerden bazıları kanser ile ilişkilendirilmeye çalışılmıştır

Application of capillary electrophoresis to investigate the degradation kinetic of olmesartan medoxomil

A simple capillary zone electrophoresis method was applied in order to investigate the degradation of olmesartan medoxomil in basic pHs. A kinetic approach was performed to enlighten its degradation process. Olmesartan medoxomil, solved in acetonitrile and diluted with 30 mM phosphate buffer, was injected six times within five hours by using running electrolytes of different pHs (30 mM phosphate buffer pH 7.5, 8.0 and 8.5). A fused silica capillary (i.d. 50.0 μm, total length 48.5 cm and effective length 40.0 cm) was employed for the analysis. The separation and best peak shape was achieved by applying 30 kV voltage at 30 °C capillary temperature. A diode array detector was used at 210 nm wavelength. Diflunisal was the internal standard. It was clarified that the degradation of olmesartan medoxomil progresses with a first order reaction kinetic. For pH 7.5, 8.0, and 8.5 the first order kinetic constants (k) were found to be 0.042, 0.092, and 0.171 respectively.Colegio de Farmacéuticos de la Provincia de Buenos Aire

Ultrafiltration-Based Extraction and HPLC Analysis of Naproxen Sodium in Human Plasma Samples: an Innovative Approach to Pharmaceutical Analysis

The purpose of this study was to assess the possibility of using ultrafiltration-based extraction for pharmaceutical analysis and develop an innovative, rapid and simple analysis technique for determination of naproxen sodium in human plasma. The proposed extraction technique can be adapted for wide range of active pharmaceutical ingredients including water-soluble nonsteroidal anti-inflammatory drugs. Naproxen sodium was isolated from human plasma by ultrafiltration-based extraction and analyzed by HPLC. Proteins in the plasma samples were precipitated with methanol and the supernatant was filtered through commercial centrifugal filters (<3 kDa). The filtered part was vacuum centrifuged, dried, and then dissolved in water and centrifuged again. The supernatant was injected into the HPLC system where an ACE 5 µm C18 100 Å LC column (150 × 4.6 mm) was used. The mobile phase was acetonitrile – phosphate buffer (pH 3.0, 20 mM) 55 : 45 v/v mixture, the injection volume was 20 µL, and the UV detection was performed at 240 nm. Using the developed extraction technique combined with HPLC method, plasma samples containing naproxen sodium were successfully analyzed (RSD 1.04–6.47, bias 2.33–6.00). © 2016, Springer Science+Business Media New York

A Foodomics approach: CE-MS for comparative metabolomics of colon cancer cells treated with dietary polyphenols

The potential of capillary electrophoresis-mass spectrometry (CE-MS) for metabolomics is demonstrated through the analysis of metabolites from human HT29 colon cancer cells treated and non-treated with dietary polyphenols. Prior to CE-MS analysis, four different metabolite purification strategies are investigated. Namely, the results obtained after methanol deproteinization, ultrafiltration, and two solid-phase extraction methods using C18 and polymer-based cartridges are described. These generic methods can have broad applications to analyze metabolites in a large variety of matrices and fields, including the new Foodomics area.This work was supported by AGL2008-05108-C03-01 (Ministerio de Ciencia e Innovación, Spain) and CSD2007-00063 FUN-CFOOD projects (Programa CONSOLIDER, Ministerio de Educacion y Ciencia, Spain). MC thanks The Council of Higher Education, Turkey for his grant. CI thanks Ministerio de Educacion y Ciencia, Spain for her FPI grant.Peer Reviewe

Analysis Of The Antiproliferative Effect Of Ankaferd Hemostat On Caco-2 Colon Cancer Cells Via Lc/Ms Shotgun Proteomics Approach

Ankaferd hemostat (ABS), a traditional herbal extract, is a hemostatic agent used for wound healing and bleeding treatment. A standardized form of plants contains many biomolecules. In recent years, previous studies have demonstrated the antineoplastic effect of ABS. In the present work, we focused on the mechanism of its antineoplastic effect over Caco-2 colon cancer cells. The LC/MS-based proteomics method was used to understand the effect of ABS at the protein level. The results were evaluated with gene ontology, protein interaction, and pathway analysis. As shown by our results, ABS altered glucose, fatty acids, and protein metabolism. Moreover, ABS affects the cell cycle machinery. Moreover, we found that ABS induced critical cancer target and suppressor proteins such as carboxyl-terminal hydrolase 1, 60S ribosomal protein L5, Tumor protein D52-like2, karyopherin alpha 2, and protein deglycase DJ-1. In conclusion, the proteomics results indicated that ABS affects various cancer targets and suppressor proteins. Moreover ABS has systematical effect on cell metabolism and cell cycle in Caco-2 cells, suggesting that it could be used as an antineoplastic agent.PubMedWoSScopu