High Performance Liquid Chromatographic Method on Rosuvastatin and Ezetimibe (Article Sample)

High Performance Liquid Chromatographic Method on Rosuvastatin and Ezetimibe
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Section 1: Introduction
Issues addressed
The article presents an accurate, reproducible, fast, and sensitive spectroscopic analytical technique known as the reversed-phase high performance liquid chromatographic method (RP-HPLC). The article addresses how this method can be used to simultaneously estimate the concentration of Rosuvastatin (RSV) and Ezetimibe (EZE) drugs from their combined dosage that is used to treat hypercholesterolemia condition (Beludari, Prakash, & Mohan, 2013, p. 205).
The article also confirms the validation of the technique such as the validation of the method by measuring the various parameters to ascertain its suitability. The article also addresses the concern about the accuracy of the method through a process known as a standard external addition. Other issues talked in the article include precision and robustness of the technique (Beludari, Prakash, & Mohan, 2013, p. 205).
Significance of HPLC to pharmaceutical analysis
The technique can be of importance in the field of analysis in pharmaceutical industries in ensuring the stability of test samples used and in control of the quality of dosage of drugs that in turn can lead to effective treatment. Another significance of this method of pharmaceutical exploration is that it doesn’t get interference from impurities, excipients, and degraded particles of the drug due to its inconstant stress settings and powerful resolving ability. During analysis process, the impurities that generated are detached from the principal peaks meaning pure chromatograms are only obtained (Chandrika, 2015, p. 150).
Section 2: Principles and Experiments
Principles of HPLC
The separation ability of the method is based on the hydrophobicity of the different drugs analyzed. The technique consists of the mobile phase and stationary phase. The stationary phase consists of a hydrophobic (non-polar) compound while the mobile phase consist of hydrophilic (polar) organic solvent such as acetonitrile combined with water. The stationary phase is composed of silica gel because of its functionalized ability, efficiency, and rigidity (Chandrika, 2015, p. 150).
The solvent used must be miscible in water. So far, acetonitrile has been the solvent of the choice of RP-HPLC technique due to its ultra-violet cut –off of 190nm that allows lower wavelengths to be measured. Acetonitrile is less viscous when compared to methanol hence it causes minimal pressure fluctuation during analysis. The significance of combining acetonitrile with water is that acetonitrile forms fewer bubbles giving the best recording. The solute used gets adsorbed in the stationary phase until the concentration of the solvent used is higher to allow elution of molecules from the hydrophobic phase. The rate of elution of solutes is related to the nature of the solute regarding its hydrophobicity (Beludari, Prakash, & Mohan, 2013, p. 206).
Experimental steps
The use of RP-HPLC technique involves the following experimental steps. The first step includes the correct choice of the column to be used and other instrumentation. In RP-HPLC, the commonly used column is C18 with dimension 250 x 4.6mm with particle size 5 µm, a degasser, and quaternary gradient system. The system also contains detectors especially photodiode and the auto sampler. The rate at which mobile phase flows should be 1.2 mL/min. The wavelength detection of EZE and RSV is carried at 230nm and 253nm absorbance respectively. The second step involves selection of reagents and chemicals. Pure compounds of RSV and EZE must be within a purity percentage of 98.5-99.9%. The buffer used must have a pH of 8. The third step entails preparing both standard and stock solutions. The standard solution to be prepared must be well analyzed and should contain 1 mg/mL of both drugs. The solution is made by dissolving acetonitrile, water, and the buffer in a ratio of 40:10:50 by V/V respectively. The stock solution of EZE and RSV should be made of a concentration of 30-90 mg/mL (Beludari, Prakash, & Mohan, 2013, p. 205).
The fourth step involves preparation of the test solution by weighing 20 tablets and then crashing them to form a powder. 10mg of both RSV and EZE were weighed and transferred into a flask. To the flask, 50mL of the mobile phase was added. The resulting solution is then filtered through a membrane filter of pore size 0.45µm to obtain a concentration of 60µm/mL of both RSV and EZE. Step five involves determining the stability of the technique and tests used by forced degradation of the drugs used. The forced degradation is carried out under oxidative stress, photolytic, thermolytic, and hydrolytic conditions. Step six involves validation of the technique used by following ICH rules. All parameters of the method were evaluated and measured. The precision of the method was evaluated through several injections involving standard solutions. In step seven, a graph of peak values against concentration is constructed, and the equation of the line calculated. This is to ascertain the linearity of the results obtained (Beludari, Prakash, & Mohan, 2013, p. 206).
In step 8, the accuracy of the method was tested using recovery studies by adding the known standard to the pre-analyzed trial at three altered levels. In step 9, the precision of the method used was assessed by dissolving standard preparations of EZE and RSV at 60µm/mL and then analyzing it three times for three days. In step 10, calculation of signal-to-noise ratio of the drugs was done to determine their limit of quantitation and limit of detection. In step 11, the robustness of the technique was carried out by subjecting test preparation to minimal change in pH and rate of flow of the solute. The pH was increased by 0.2 while flow rate by 0.1mL/min. The temperature of the column was maintained at 35oc (Beludari, Prakash, & Mohan, 2013, p. 207).
Section 3: Application
Application of HPLC
The technique is used in the pharmaceutical analysis to identify, purify, and quantification of molecules or compounds that are of interest. Due to its wid...