Development and Validation of New Analytical Method for Estimation of Drotaverine in Bulk and Pharmaceutical Dosage Form by UV Spectrophotometry

About Author: Khuntia  Bhabani  S.,  M.Pharm.,  B.P.U.T.,
Department  of  Pharmaceutical  Analysis  and  Quality  assurance,
Royal  College  of  Pharmacy  and  Health  Sciences,  
Berhampur, Odisha, India

Reference ID: PHARMATUTOR-ART-1056

Abstract
The present study includes a simple, sensitive and specific UV method development and validation for the quantitation of Drotaverine in bulk and pharmaceutical dosage form. The λmax was found to be 357nm by taking acetonitrile and water (50:50) as solvent. The validation of the proposed method was carried out as per ICH Guidelines. It was found that the drug was shown the linearity between the range 5-90µg/ml. The regression of the curve was y = 0.022x - 0.033. The developed method was found to be with %RSD 0.548499 for Drotaverine. The %RSD for intra-day and inter-day precision was lower than 2%. The percentage recovery values of pure drug from the reanalyzed solution of formulation were in between 95.68-99.5%. The ruggedness of the method was studied by taking in account of different analyst and by varying the temperature. Based on the performance characteristic the proposed UV method was found to suitable for the estimation of Drotaverine in bulk and pharmaceutical dosage form.

Introduction
Drotaverine is an antispasmodic drug, structurally related to papaverine. Drotaverine is a selective inhibitor of phosphodiesterase 4, and has no anticholinergic effects. It has been shown to possess dose-dependent analgesic effects in animal models. It is 1-[(3,4-diethoxyphenyl)-methylene]-6,7-diethoxy-1,2,3,4-tetrahydroisoquinoline with molecular formula C24H31NO4 presented in. It inhibits phosphodiesterases hydrolysing cAMP, thereby increasing cAMP concentration, decreasing Ca uptake of the cells and changing the distribution of calcium among the cells. An ideal stability indicating method is one that quantifies the drug and also resolves its degradation products.
Drotaverine is freely soluble in water, methanol, ethanol, acetonitrile, chloroform and slightly soluble in HCl, NaOH. The λmax was found to be at 357nm.

Literature review for Drotaverine analysis revealed several methods based on different technique such as HPLC with UV detection. Visible spectrophotometric methods, LC-ms and HPLC ESI/MS assay for its quantification in plasma and serum, Chiral Chromatography, Pulse Polarography, Chemiluminescence assay, LC with coulometric Detection.  However there is no method reported for the detection of Drotaverine in bulk and pharmaceutical formulation by UV spectrophotometry with such parameters.
The aim of present work is to find out a simple, sensitive, specific, spectrophotometric method developed for the detection of Drotaverine in bulk drug and pharmaceutical formulation.

Methods and Material: Simple UV Spectrophotometric Method is used. Drotaverine working standard was supplied by Khandelwal Remedies, Mumbai. Drotin-DS (Label claim: 80mg tablet) was manufactured by Martin & Harris Lab, Roorkee. All other chemicals used in the analysis were AR grade. A double – beam spectrophotometer (Shimadzu-1700) was used for the detection of absorbance, Electronic Balance (Sigma 200/A Super) and Sonicator (PCi-3.5L) were used for experimental purpose.

Subjects and Methods:
Preparation of Stock Solutions:
Standard stock solution was prepared by dissolving 100 mg of the drug in 100 ml of acetonitrile and water (50:50) to get concentration of 1mg/ml (1000 μg/ml) solutions.

Preparation of working solutions:
From the above stock solution 1ml of each were taken in a 10 ml volumetric flask and the volumes were made up to mark with the solvent to get concentration 0f 100μg/ml solutions. From this 1ml, 2ml, 3ml so on were transferred into 10ml volumetric flasks and volume was made up to the mark with acetonitrile and water (50:50) to make 10µg/ml, 20µg/ml, 30µg/ml respectively. Then the samples were scanned with UV-VIS Spectrophotometer against acetonitrile and water (50:50) as blank and the wavelength corresponding to maximum absorbance was noted which is its λ_max.

Preparation of Calibration Curve:
Different concentrations form 5-100µg/ml were prepared form the stock solution in different volumetric flasks. Then the absorbances were noted by scanning in UV. Then a calibration curve was plotted taking concentration on X-axis and absorbance on Y-axis.

Validation
Validation can be defined as (ICH) “Establishing documented evidence, which provides a high degree of assurance that a specific activity will consistently produce a desired result or product meeting its predetermined specifications and quality characteristics”.
Method validation is an integral part of the method development; it is the process of demonstrating that analytical procedures are suitable for their intended use and that they support the identity, quality, purity, and potency of the drug substances and drug products. Simply, method validation is the process of proving that an analytical method is acceptable for its intended purpose.
The validation for UV method development was performed using parameters like Linearity, Accuracy, Precision, Robustness, Ruggedness, Limit of detection (LOD), Limit of quantitation (LOQ).

Results:
Determination of Working Wave Length (λ_max):
In order to ascertain the wavelength of maximum absorption (λ max) of the drug, different solutions of the drugs (5μg/ml and 10μg/ml) in acetonitrile and water (50:50) was scanned using UV-VIS spectrophotometer within the wavelength region of 200 – 400 nm against acetonitrile and water (50:50) as blank. The resulting spectra are shown in figure below and the absorption curve shows characteristic absorption maxima at 357nm for Drotaverine.

Preparation of Calibration Curve:
For preparation of calibration curve of Drotaverine, a stock solution of 1000µg/ml was prepared. From it different concentrations ranging from 5 – 100 µg/ml was prepared. Then the samples were scanned through UV-VIS Spectrophotometer and their absorbances were noted, which were given in the table below.
A calibration curve was plotted using those readings taking Concentration on X-axis and Absorbance on Y-axis.

Calibration Curve of Drotaverine:
From the calibration curve it was found that it shows linearity in the range of 5– 90µg/ml with regression 0.999.

METHOD VALIDATION
Linearity:
Various aliquots were prepared form the stock solution (1000µg/ml) ranging 5-100µg/ml. The samples were scanned in UV-VIS Spectrophotometer against acetonitrile and water (50:50) as blank. It was found that the selected drug shows linearity between the ranges of 5-100µg/ml. The results are tabulated below.

Linearity Table of Drotaverine in Working Standard