ISSN- 22315640 (Print) www.asianpharmaonline.org

ISSN- 22315659 (Online)

 

 

RESEARCH ARTICLE

 

Development and Validation of Spectrophotometric Method for Determination of Azelaic Acid

 

Kadam T.V. 1, Darekar A.B.1, Gondkar S.B.1, Saudagar R.B.2

1Department of Pharmaceutics, R.G. Sapkal College of Pharmacy, Anjaneri, Nashik-422213 Maharashtra India

2Department of Pharmaceutical Chemistry, R.G. Sapkal College of Pharmacy, Anjaneri, Nashik-422213 Maharashtra India

*Corresponding Author E-mail:truptikadam91@gmail.com

 

ABSTRACT:

In these studies describes a simple, accurate, precise and cost effective UV-visible spectrophotometric method for the estimation of Azelaic acid in pure and pharmaceutical formulations. The method is based on the measurement of absorbance of Azelaic acid solution in Phosphate buffer pH 6.8 at 204nm in the wavelength range of 200-400nm. The method obeys Beers Lamberts law in the selected concentration range10-50 μg/ml in selected media. The slope, intercept and correlation coefficient were also calculated. Results of percentage recovery study shows that the method was not affected by the presence of common excipients in tablets. The parameters like linearity, precision, accuracy, sensitivity study i.e. limit of detection and limit of quantitation were studied according to International Conference on Harmonization (ICH) guidelines. The developed method was validated in terms of accuracy, precision, linearity, limit of detection and limit of quantitation which proves suitability of proposed method for routine estimation of Azelaic acid in pure and pharmaceutical formulations.

 

KEYWORDS: UV-visible spectrophotometer, method development, Azelaic acid.

 

 


1. INTRODUCTION:

Azelaic acid is dermatologic agent. Chemically it is known as 1, 7- heptanedicarboxylic acid with molecular weight 188.22. Freely soluble in phosphate buffer solution pH 6.8, 0.1N NaOH and elimination half-life 45 min (oral) and 12 hrs (topical).It is thought that Azelaic acid manifests its antibacterial effects by inhibiting the synthesis of cellular protein in anaerobic and aerobic bacteria, especially Staphylococcus epidermidis and Propionibacterium acnes.

 

In aerobic bacteria, Azelaic acid reversibly inhibits several oxidoreductive enzymes including tyrosinase, mitochondrial enzymes of the respiratory chain, thioredoxin reductase, 5-alpha-reductase, and DNA polymerases.

 

Received on 05.06.2015 Accepted on 30.06.2015

Asian Pharma Press All Right Reserved

Asian J. Res. Pharm. Sci. 5(2): April-June 2015; Page 83-85

DOI: 10.5958/2231-5659.2015.00014.4

 

In anaerobic bacteria, Azelaic acid impedes glycolysis.1,2 Literature survey revealed that most of the HPLC, GC methods used hyphenated techniques with detectors such as mass spectrometry, ultraviolet, Electrospray mass spectrometric techniques all these methods have high sensitivity, but most of them highly expensive and are not easily available in quality control laboratories The literature survey clearly indicates there is no spectrophotometric method is available for the quantification of Azelaic acid. The analytically useful functional groups in Azelaic acid have not been fully exploited for designing suitable, visible spectrophotometric methods and so still offer a scope to develop more visible spectrophotometric methods with better sensitivity, selectivity, precision and accuracy. The author has made an attempt in this direction and succeeds in developing methods.3-5 so authors objective is to develop accurate, simple, UV spectrophotometric method which is free from extraction techniques, and shorter time and highly sensitive technique.

 

 

 

EXPERIMENTAL MATERIALS:

Azelaic acid was received as a gift sample from Cadila Pharmaceuticals Ltd., Thane, India. All analytical grade chemicals and solvents were supplied by S.D. Fine Chemicals, Mumbai, India. Distilled water was used to prepare all solution. Freshly prepared solutions were always employed.

 

Instruments:

Jasco V-630 UV-visible spectrophotometer with data processing system were used. The sample solution were recorded in 1 cm matched quartz cells was used for spectral an absorbance measurements. Shimadzu AUX-220 electronic balance was used for weighing the sample.

 

Standard Solution Selection of common solvent:

Phosphate buffer solution (PBS) pH 6.8 was selected as a common solvent for developing spectral characteristics of drug. The selection was made after using different solvents and their different normalities.

 

Preparation of standard drug solution:

Standard stock solution containing Azelaic acid was prepared by dissolving 10 mg of Azelaic acid separately in 25 ml of PBS pH 6.8 shake for 5 min. and then final volume of the solutions was made up to 100 ml with same solvents to get stock solution containing 100 g/ml

 

Selection of λ max:

The standard stock solution was further diluted with PBS pH 6.8 to get a 10 μg/ml of concentration (1 ml to 10 ml). The solution was scanned between 200 and 400 nm using PBS pH 6.8 as blank. From the spectrum obtained, 204 nm was selected as λmax for the analysis of Azelaic acid. Standard stock solution was further diluted to obtain 10-50 g/ml with PBS pH 6.8. Calibration curve was plotted in the concentration range of 10-50 g/ml of Azelaic acid using PBS pH 6.8 as blank.

 

Validation of the proposed method 6, 7

The proposed method was validated according to the International Conference on Harmonization (ICH) guidelines.

 

Linearity (calibration curve)

The developed method validated as per ICH guidelines. The plot of absorbance verses concentration is shown in figure no.1 for PBS pH 6.8. It can be seen that plots are linear in the concentration range 10-50 g/ml with correlation coefficients (r2) is 0.994.

 

 

Precision (repeatability)

Intraday and interday precision was determined by measurement of the absorbance for three times on same day and on three different days. The relative standard deviation for replicates of sample solution was less than 2% which meet the acceptance criteria for established method.

 

Accuracy (recovery study):

Recovery study was carried out by adding a known quantity of pure drug to the preanalysed formulations and the proposed method was followed. From the amount of drug found, percentage recovery was calculated as per ICH guidelines.

 

Sensitivity:

Sensitivity study was carried out where limit of detection (LOD) and limit of quantification (LOQ) were determined using following equation.

LOD = 3.3* σ / S

LOQ = 10* σ / S

Where, σ = standard deviation of the response.

S = slope of calibration curve

 

RESULT AND DISCUSSION:

The maximum absorption for Azelaic acid in PBS pH 6.8 was observed at 204 nm. The high values of correlation coefficient in PBS pH 6.8 indicate linearity for Azelaic acid in PBS pH 6.8. Beers law was obeyed for PBS pH 6.8 in the range of10-50 g/ml the accuracy of method was determined by calculating mean percentage recovery and % relative error. The percentage recovery ranges from 98 g/ml and % relative errors was within 2% in PBS pH 6.8 and are presented in table no. 4. Precision was calculated as repeatability, inter and intraday variation for Azelaic acid, percentage RSD was found to be less than 1. The repeatability data are presented in table no. 2 and table no. 3. LOD and LOQ were found to be 0.099 g/ml and 0.30 g/ml respectively for detection of Azelaic acid in PBS pH 6.8. The proposed method was found to be simple, accurate, precise and rapid for the routine determination of Azelaic acid in pure and pharmaceutical formulation

 

Table no.1: Optical Parameters for Determination of azelaic acid

Precision:

Sr.no.

Data

Results

1

Wavelength (nm)

204

2

Slope

0.0022

3

Correlation coefficient

0.994

4

Intercept

0.067

5

LOD (g/ml)

0.099

6

LOQ(g/ml)

0.30

 


 

Figure no.1: Calibration Curve of azelaic acid in PBS pH 6.8

 

Table no.2: Intraday Variability

Conc. (g/ml)

Absorbance

Mean

Standard Deviation ()

RSD (%)

Trial I

Trial II

Trial III

10

0.0867

0.0875

0.0879

0.0873

0.00061

0.7061

20

0.1041

0.1052

0.1057

0.1050

0.00081

0.7795

30

0.1333

0.1342

0.1349

0.1341

0.00080

0.5988

40

0.1515

0.1523

0.1528

0.1522

0.00065

0.4308

50

0.1713

0.1719

0.1729

0.1720

0.00080

0.4705

 

Table no.3: Interday Variability

Conc. (g/ml)

Absorbance

Mean

Standard Deviation ()

RSD (%)

Trial I

Trial II

Trial III

10

0.0912

0.0919

0.0925

0.0918

0.00065

0.7143

20

0.1143

0.1151

0.1159

0.1151

0.00080

0.6950

30

0.1428

0.1437

0.1449

0.1438

0.0010

0.7326

40

0.1612

0.1628

0.1638

0.1626

0.00131

0.8065

50

0.1819

0.1833

0.1849

0.1833

0.00150

0.8201

 

Table no.4: Result of Assay

Formulation

Label claim

% amount found

RSD (%)

Conventional tablet

100mg

101.8%

0.4666

 

Table no.5: Recovery Study

Amount of drug taken from tablets (mg)

Amount of standard drug added (mg)

Total amount recovered (mg)

%recovery

Standard deviation ()

RSD (%)

10

5

14.75

98.34%

0.0017

0.4344

10

10

19.60

98%

0.0013

0.2654

10

15

24.65

98.61%

0.0016

0.2731

 

 


CONCLUSION:

Simple spectrophotometric method for determination of Azelaic acid have been developed and validated as per ICH guidelines. The proposed method is found to be simple, rapid, sensitive, accurate and reproducible also can be used for the routine quality control analysis of Azelaic acid in bulk and pharmaceutical formulations.

 

REFERENCES:

1.     Indian Pharmacopoeia 2014, Volume II, Government of India Ministry of Health And Family Welfare, Published by the Indian Pharmacopoeia Commission, Ghaziabad, pn.1113-14.

2.     http://www.drugbank.ca/drugs/DB00548.Accessed On 13 Aug. 2014 At 12.25pm.

3.     Medikondu Kishore, M. Jayaprakash, T. Vijayabhaskara Reddy. Spectrophotometric Determination of Azelaic Acid in Pharmaceutical Formulations.Journal of Pharmacy Research 2010; 3(12):3090-3092.

4.     Muhammed Alzweiri, Yusuf M. Al-Hiari, Talal Aburjai, Osama Abdel-Aldaem. On-Column Approach in the HPLC-UV Analysis of Non-chromophoric Compounds Using Azelaic Acid as a Model. Jordan Journal of Pharmaceutical Sciences 2012; 5(3): 243-50.

5.     Mahdi Garelnabi, Dmitry Litvinov, Sampath Parthasarathy. Evaluation of a gas chromatography method for azelaic acid determination in selected biological samples. North American Journal of Medical Sciences 2010; 2(9): 397-402.

6.     Q2A: Text On; Validation of Analytical Procedures. In: International Conference on Harmonization. Federal Register, 60(40):11260-11262 (1995)

7.     Q2B: Text On; Validation of Analytical Procedures. In: International Conference on Harmonization. Federal Register, 62(96):27463-27467 (1997)