Optimization of HPLC method using central composite design for estimation of Torsemide and Eplerenone in tablet dosage form

Abstract A simple, precise, accurate and robust high performance liquid chromatographic method has been developed for simultaneous estimation of Torsemide and Eplerenone in tablet dosage form. Design of experiment was applied for multivariate optimization of the experimental conditions of RP-HPLC method. A Central composite design was used to study the response surface methodology and to analyse in detail the effects of these independent factors on responses. Total eleven experiments along with 3 center points were performed. Two factors were selected to design the matrix, one factor is variation in ratio of Acetonitrile and the second factor is flow rate (mL/min). Optimization in chromatographic conditions was achieved by applying Central composite design. The optimized and predicted data from contour diagram comprised mobile phase (acetonitrile, water and methanol in the ratio of 50: 30: 20 v/v/v respectively), at a flow rate of 1.0 ml/min and at ambient column temperature. Using these optimum conditions baseline separation of both drugs with good resolution and run time of less than 5 minutes were achieved. The optimized assay conditions were validated as per the ICH guidelines (2005). Hence, the results showed that the Quality by design approach could successfully optimize RP-HPLC method for simultaneous estimation of Torsemide and Eplerenone.

Effect of light, oxygen and temperature on the stability of artepillin C and p-coumaric acid from Brazilian green propolis.

Brazilian Green Propolis (BGP) is an important bee product, which displays important biological activities, making it valuable in the international market. The major prenylated phenolic compound in BPG is (E)-artepillin C, along with its precursor (E)-p-coumaric acid, both contributing to the biological effects of BGP. Taking that into account, it was evaluated the effect of light, temperature and air oxygen in their content to establish the best storage and transport conditions for crude BGP and the pure compounds. For that, (E)-artepillin C and (E)-p-coumaric acid were initially submitted to degradation for five days under sunlight and high temperature (50 °C), furnishing three major (E)-Artepillin C isomers and one from (E)-p-coumaric acid. Then, it was developed and validated a Reverse Phase High Performance Liquid Chromatography (RP-HPLC) method for quantifying these compounds in crude BGP and in its extracts. In the stability studies, it was used a Full Factorial and Central Composite Design to establish the desirable storage conditions. (E)-Artepillin C, both pure and in BGP should be kept protected from light and storage below -2.5 °C. (E)-p-Coumaric acid can be stored at room temperature. Therefore, the best storage and transport conditions to keep the content of both compounds in BGP are protection from light at low temperatures.

Development and validation of a RP-HPLC method for the simultaneous detection and quantification of simvastatin's isoforms and coenzyme Q10 in lecithin/chitosan nanoparticles.

Hybrid nanocapsules constituted of phospholipids and polysaccharides have been proposed as colloidal systems for the delivery of drugs via non-parenteral administration routes, due their capacity of high drug loading, controlled drug release and targeted delivery to the specific organ. Moreover, nanoparticles systems offer the possibility of co-encapsulation of drugs in the same drug delivery system and, consequently, the simultaneous administration of compounds. Characterization of nanoparticles properties, specifically involves quantification of the active pharmaceutical ingredients and is pivotal in the development of innovative nanomedicines. Therefore, this study has proposed and validated a new RP-HPLC-UV method for the simultaneous determination of simvastatin and coenzyme Q10 in hybrid nanoparticles systems. A reversed phase (RP) C8 column and a gradient elution of water: methanol at flow rate of 1.5 ml/min was used. Simvastatin (SVT), simvastatin hydroxyacid isoform (SVA) and coenzyme Q10 were identified by dual wavelength-UV detection at 238 nm (statins) and 275 nm, respectively. The proposed method was selective and linear in the range of 0.5-25 µg/ml (r2 > 0.999), precise, with values of relative standard deviation (RSD) lower than 2%, robust and accurate (recovery values of 100 ±â€¯5%), satisfying FDA guidelines. Furthermore, low detection (LOD <0.2 µg/ml) and quantification limits (LOQ <0.4 µg/ml) were suitable for the application of the method for the in vitro study of release kinetics of simvastatin and coenzyme Q10 co-encapsulated in lecithin/chitosan nanoparticles. The proposed method represents, to our knowledge, the only method for the simultaneous quantification of simvastatin, coenzyme Q10 and of the hydrolysed hydroxyacid isoform of the statin in nanoparticles.

RP-HPLC analysis of manool-rich Salvia officinalis extract and its antimicrobial activity against bacteria associated with dental caries

In this paper we screened the dichloromethane extract from the aerial parts of Salvia officinalis L., Lamiaceae, against a representative panel of microorganisms that cause caries, conducted a bioassay-guided fractionation to establish themselves the most active metabolite (manool) and determined the Salvia officinalis fraction with the manool highest concentration to be used to activate an ingredient in oral care products such as toothpastes and mouthwashes. Both manool and S. officinalis extract showed very promising minimal inhibitory concentration values (between 6.24 and 31.36 µg.ml-1) and time kill curves against the primary causative agents of dental caries (Streptococcus mutans) revealed that, at twice its minimal bactericidal concentration (12.48 µg.ml-1), manool required 6 h to completely kill the bacteria. Salvia officinalis extract at twice its minimal bactericidal concentration (31.36 µg.ml-1 ) needed 12 h. The results achieved with Salvia officinalis extract motivated us to develop and validate an analytical RP-HPLC method to detect and determine manool in this extract. The validation parameters were satisfactorily met and evaluated allows us to consider the developed method suitable for use in different labs. In conclusion, our results evidenced that the manool-rich S. officinalis extract can be considered an analytically validated alternative to develop novel and effective antimicrobial agents against the main bacteria responsible for dental caries.