U.V SPECTROPHOTOMETRIC METHOD FOR THE ESTIMATION OF MESALAZINE IN BULK AND ITS PHARMACEUTICAL DOSAGE FORMSHTML Full Text
U.V SPECTROPHOTOMETRIC METHOD FOR THE ESTIMATION OF MESALAZINE IN BULK AND ITS PHARMACEUTICAL DOSAGE FORMS
Rakesh Kumar Singh*1, Pankaj Singh Patel2 and Pragya Gupta3
College of Pharmaceutical Sciences *1, Mohuda, Berhampur, Orissa
Indira Gandhi Institute of Pharmaceutical science 2, Bhubaneswar, Orissa
Biotech Park, CIF Lab 3, Lucknow
Mesalazine is used, either orally or rectally, as an anti-inflammatory agent in treating ulcerative colitis and, to a lesser extent, Crohn's disease. Mesalazine is a white to pinkish crystalline powder. It is slightly soluble in cold water and alcohol; more soluble in hot water, soluble in hydrochloric acid. A simple UV spectrophotometric method was developed for the determination of Mesalazine (MEZ) in pure and its pharmaceutical formulations. Mesalazine exhibiting max absorbance at 210 nm in methanol and obeyed linearity in the concentration range of 0.2-50 mg/ml. The proposed method was statistically validated.
INTRODUCTION: The scope of developing and validating analytical methods is to ensure a suitable method for a particular analyte more specific, accurate and precise. The main objective for that is to improve the conditions and parameters, which should be followed in the development and validation. Chemically Mesalazine (MEZ) is 5-amino salicylic acid. It is an anti-inflammatory drug structurally related to salicylates and active in inflammatory bowel disease. Tablet formulations containing 250, 400 and 500 mg MEZ are available in the market. Literature survey revealed that Mesalazine is estimated by HPLC and micellar electrokinetic chromatography. No UV spectrophotometric methods have been reported for estimation of MEZ in single component formulation. Hence, an attempt has been made to develop new UV methods for its estimation in pharmaceutical formulations with good accuracy, simplicity, precision and economy.
5-Amino-2-hydroxybenzene-1 carboxylic acid [Mesalazine or 5-Amino salicylic acid (ASA)]
Instrumentation: Spectral and absorbance measurements were made on an ELICO SL-159 UV-Vis spectrophotometer by using 1 cm quartz cells. Dhona balance was used for weighing the samples. Commercially available tablets of Mesalazine were procured from the local market and estimated.
- Disodium hydrogen phosphate anhydrous (Merck)
- Citric acid
- Double distilled water
- Mesalazine, Sun pharma ltd., (India)
Optimization: Scanning and determination of maximum wavelength (lmax):
In order to ascertain the wavelength of maximum absorption (λmax) of the drug, different solutions of the drugs (10 μg/ml and 20 μg/ml) in methanol were scanned using spectrophotometer within the wavelength region of 200 – 380 nm against Phosphate buffer (pH- 3.6) as blank. The resulting spectra were shown in fig 1 and the absorption curve showed characteristic absorption maxima at 298 nm for Mesalazine.
Fig 1: UV Spectrum of Mesalazine in Phosphate buffer (pH- 3.6)
Sample: Mesalazine (10 μg/ml)
Reference: Phosphate Buffer (pH-3.6) Instrument: Elico SL 159 UV –Visible Spectrophotometer.
Preparation of Phosphate buffer solution (pH-3.6): 0.9g of anhydrous disodium hydrogen phosphate and 1.298g of citric acid monohydrate were weighed accurately, mixed and volume made with double distilled water (1000ml).
Preparation of Stock Solutions: Standard stock solution was prepared by dissolving 25 mg of each drug in 25 ml of Phosphate Buffer (pH-3.6) to get concentration of 1mg/ml (1000 μg/ml) solutions.
Preparation of Working Standard Solutions and construction of standard graph: The prepared stock solution was further diluted with Phosphate Buffer (pH-3.6) to get working standard solutions of 10 mg/ml and 100 mg/ml of mesalamine to construct Beer’s law plot for pure drug, different aliquots of Mesalazine were taken and diluted to 10 ml with Phosphate Buffer (pH-3.6). The absorbance was measured maximum at 245 nm, against Phosphate Buffer (pH-3.6) as blank. The results were shown in table (1). The standard graph was plotted by taking concentration of drug on x-axis and absorbance on y-axis and was shown in Fig. (b) The drug has obeyed Beer’s law in the concentration range of 0.2-50µg/ml.
Table 1: Linearity table of Mesalazine in Working Standard
|Concentration (mg /ml)||Absorbance|
Table 2: Optical characteristics
|Beer’s Law limit (µg/mL)||0.2-50|
|Sandell’s sensitivity(µg/cm2/0.001absorbance unit)||0.0390|
|Molar extinction coefficient (1 mole-1 c.m-1)||4.90 ´ 103|
|% Relative standard deviation||1.1083|
|95% Confidence limits||0.189209|
|99% Confidence limits||0.249058|
|Regression equation (Y*)|
* Y= a + bC where C is the concentration of mesalamine and Y is the peak area
Fig 2: Standard graph of Mesalazine
Table 3: Recovery from the formulation:
|Formulation||Labeled amount(mg)||UV- method*|
|Mean ± s. d (amount mg recovered)||%Drug recovered||% RSD|
* Each value is average of three determinations ± standard deviation.
Precision: The precision of the proposed method was ascertained by actual determination of eight replicates of fixed concentration of the drug within the Beer’s range and finding out the absorbance by the proposed method. From this absorbance, Mean, Standard deviation, % RSD was calculated. The readings were shown in Table 4.
Table 4: Precision readings:
|Concentrations (mg/ml)||Absorbance||Statistical analysis|
|10||0.246||Mean = 0.2456SD = 0.002722
%RSD = 1.1083
Accuracy: To determine the accuracy of the proposed method, recovery studies were carried out by adding different amounts (80%, 100%, and 120%) of bulk samples of mesalamine within the linearity range were taken and added to the pre-analyzed formulation of concentration 20mg/ml. From that percentage recovery values were calculated. The results were shown in Table 5.
RESULTS AND DISCUSSION: From the optical characteristics of the proposed method, it was found that mesalazine obeys linearity within the concentration range of 0.2-50 mg/ml. From the results shown in Table (4) it was found that the % RSD is less than 2, which indicates that the method has good reproducibility. From the results shown in accuracy Table 5, it was found that the percentage recovery values of pure drug from the preanalyzed solution of formulation were in between 98.676 – 101.397, which indicates that the proposed method is accurate and also reveals that the commonly used excipients and additives in the pharmaceutical formulations were not interfering in the proposed method.
Table 5: Accuracy table
|Sample ID||Concentration (mg/ml)||%Recovery of Pure drug||Statistical Analysis|
|S1 : 80 %||8||10||101.29||Mean||101.397|
|S2 : 80 %||8||10||101.52||SD||0.1159|
|S3 : 80 %||8||10||101.38||% RSD||0.1143|
|S4 : 100 %||10||10||99.046||Mean||99.3253|
|S5 : 100 %||10||10||99.57||SD||0.2637|
|S6 : 100 %||10||10||99.36||% RSD||0.2655|
|S7 : 120 %||12||10||98.819||Mean||98.6757|
|S8 : 120 %||12||10||98.328||SD||0.3026|
|S9 : 120 %||12||10||98.880||% RSD||0.3067|
CONCLUSION: The proposed method was simple, sensitive and reliable with good precision and accuracy. The proposed method is specific while estimating the commercial formulations without interference of excipients and other additives. Hence, this method can be used for the routine determination of Mesalazine in pure samples and pharmaceutical formulations.
The authors thank Sun Pharma Ltd. for providing the gift sample of Mesalazine. Authors are thankful to CIF Lab Biotech Park, Lucknow for providing necessary facilities for the work. Authors are also thankful to Dr. Pragya Gupta under whom this work was completed.
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Rakesh Kumar Singh*, Pankaj Singh Patel and Pragya Gupta
College of Pharmaceutical Sciences *, Mohuda, Berhampur, Orissa
20 January, 2010
15 February, 2010
24 February, 2010
01 March, 2010