New Spectrophotometric Analytical Methods for Determination Some Cardiovascular Drugs in Pharmaceutical Formulations

Abstract

The efforts in this thesis have represented a big challenge to develop new and cost effective analytical spectrophotometric methods for the determination of five medications components belonging to numerous categories specializing in treating cardiovascular diseases (CVDs) involves; Atorvastatin Calcium (ATOR), Rosuvastatin Calcium (RUSV), Amlodipine Besylate (AMLO), Atenolol (ATEN), and Hydrochlorothiazide (HCTZ).The study, which included the estimation of the studied drugs individually and simultaneously in their pure forms, pharmaceutical formulations (tablets) serum and urine. In the individual estimation method developing a novel indirect reverse spectrophotometric method for determining mentioned drugs in their pure forms and pharmaceutical formulations was described. It was based on the oxidation of the drug with a fixed excess concentration of KIO4 in acidic HCl solution, consequently the residual unreacted oxidizing agent (KIO4) reacts with the p-ansidine reagent, as a result yellowish orange soluble color complex was formed with 538 nm λmax and measured spectrophotometrically. The indicated λmax absorption peak has been adopted as a baseline to construct the reverse estimate at 425nm as a new inverse peak for the determination of the drugs under study. The absorption intensity of the inverse peak was inversely related to the drugs concentrations. Three methods were used to find optimum conditions for the experiment; univariate and multivariate methods (Central Composite Design (CCD) and Box Benhken Design (BBD)). The influence of three factors has been studied and optimized for their effective impact on the course of the selected methods and these factors are: oxidant volume (mL), p-anisidine concentration (w/v) and acid volume (mL). The optimal conditions these parameters are 1.5 mL of (3 % w/v) KIO4, (0.08 % w/v) of p-anisidine and 0.6 mL of (1N) HCl for univariate method, 1.5 mL of (3 % w/v) KIO4, (0.07 % w/v) of p-anisidine and 0.7 mL of (1N) HCl by (CCD) method, and 1.525 mL of (3 % w/v) KIO4, (0.075 % w/v) of p-anisidine and 0.55 mL of (1N) HCl by (BBD) method. Calibration curves were constructed for all drugs by plotting absorbances signals against concentration (µg.mL-1 ) at 425 nm and found to be linear over the concentration range (5-170 µg.mL-1 ), (25-170 µg.mL-1 ), (8- 148 µg.mL-1 ), (12-150 µg.mL-1 ) and (3-210 µg.mL-1 ) for ATOR, RUSV, AMLO, ATEN, and HCTZ, respectively via univariate method, (5-185 µg.mL-1 ), (15-195 µg.mL-1 ), (5-158 µg.mL-1 ), (3-160 µg.mL-1 ) and (1-230 µg.mL-1 ) for ATOR, RUSV, AMLO, ATEN, and HCTZ, respectively via CCD method and (3-188 µg.mL-1 ), (8-208 µg.mL-1 ), (5-158 µg.mL- VII 1 ), (3-160 µg.mL-1 ) and (1-240 µg.mL-1 ) for ATOR, RUSV, AMLO, ATEN, and HCTZ, respectively via BBD method. The limit of detection (LOD) and limit of quantification (LOQ) for the mentioned drugs by those methods were found be in the range (0.14384 – 0.19306 µg.mL-1 ) LOD and (0.43590 - 0.58503 µg.mL-1 ) LOQ via univariate method, (0.11423 – 0.17973 µg.mL-1 ) LOD and (0.34615 - 0.54466 µg.mL-1 ) LOQ via CCD method, and (0.0991– 0.18615 µg.mL-1 ) LOD and (0.3003 – 0.56410 µg.mL-1 ) LOQ via BBD method. Validity of all methods were checked by measuring relative standard deviation (RSD %) relative error (RE %) and it was found RSD% < 5% while RE% < 4%,. The analysis results do not show any interference from the excipients by using these techniques. Therefore ATOR, RUSV, AMLO, ATEN and HCTZ were suitable for estimation in their pharmaceutical formulation. Under the title “simultaneous determination of ATOR, RUSV, AMLO, ATEN and HCTZ drugs” estimation of the five drugs in their binary, tertiary, quaternary and pentanary mixtures by six develop methods (first (D1) and second (D2) derivative spectrophotometric, derivative ratio spectrophotometry method (DRS), artificial neural network (ANN), partial least squares (PLS), and principal component regression (PCR) techniques) were described. Derivative spectrophotometric techniques (D1 and D2) were developed for the simultaneous determination of ATOR, RUSV, AMLO, ATEN and HCTZ drugs in their binary mixtures. It was found that all cited drugs could be determined by both modes of derivative. The zero crossing technique was employed in D1 measurements, using peak height to baseline at 294.6, 234, 253.6, 284.2 and 261.4 nm for ATOR, RUSV, AMLO, ATEN and HCTZ respectively, same procedure was applied in D2 by using the peak height to baseline at 311.6, 342.8, 257.7, 287.8 and 234 nm for ATOR, RUSV, AMLO, ATEN and HCTZ respectively. Derivative ratio spectrophotometry method (DRS) was applied for the simultaneous estimation of ATOR, RUSV, AMLO, ATEN and HCTZ in their tertiary, quaternary and pentanary mixtures. A 308 synthetic mixtures were prepared and distributed as 88, 140 and 80 from tertiary, quaternary and pentanary mixtures respectively, using Design Experts Software (DES) version (2019) according to Mixture Design (MD). The first derivative ratio of amplitude spectrum was applied for constructing the calibration graph for drugs in their pentanary mixtures. The calibration graphs of D1-amplitude spectrum were constructed in the range of (5 – 30 μg.mL-1 ) at 283.2 nm for ATOR, (5 – 35 μg.mL-1 ) at 318.2 nm for RUSV, (3 – 28 μg.mL-1 ) at 404.2 nm for AMLO, (5 – 30 μg.mL-1 ) at 233.2 nm for ATEN and (5 – 40 μg.mL-1 ) at 263.6nm for HCTZ. Quaternary mixtures were composed as a divisor, where half VIII of the linear concentrations of each drug were used in that composition. The result of relative standard deviation percentage (RSD %) and relative error percentage (RE %) were in the ranges (0.151% - 3.725%) and (-3.94% - 4.98%) for tertiary mixtures, (0.399% - 3.87%) and (-5.423% - 4.62%) for quaternary mixtures and (0.327% - 4.034%) and (-6.178% - 6.956%) for pentanary mixtures respectively. ANN, PLS and PCR techniques were used as chemometric technique for simultaneous spectrophotometric estimation of ATOR, RUSV, AMLO, ATEN and HCTZ in their tertiary, quaternary and pentanary mixtures. The first section in the methodology of the techniques includes the construction of a calibration (training) set of spectrophotometric data, obtained from zero orders calibration graph, composing of 60 mixtures chosen randomly by the design experts software (DES). The absorbance data matrices related to the weighted of concentration data matrices for each of the 60 mixtures were obtained by the measurements of absorbance in the range 200-400 nm with a scan speed of 10 nm.sec-1 , averaging of 0.2 nm, bandwidth of 1.8 nm, and data interval of 0.1 nm against solvent blank. The ANN, PLS and PCR regression models were built with the Assistance of Origin Pro software version (2015) program and the concentration of the five drugs were then predicted. The result of relative standard deviation percentage (RSD %) and relative error percentage (RE %) were in the ranges (0.019% - 1.219%) and (-0.542% - 0.644%) for ANN, (0.048% - 3.1%) and (- 3.158% 1.345%) for PLS and (0.087% - 2.642%) and (-4.612% - 4.817%) for PCR respectively. The proposed method was successfully applied in the simultaneous determination of ATOR, RUSV, AMLO, ATEN and HCTZ in their sample of laboratory synthetic mixtures, spike in serum, and urine.