Journal of Drug Delivery and Therapeutics

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Method development and validation for estimation of Gliclazide in bulk and tablet form by UV Spectrophotometer

Ravindra Hanwate *^{1, 3}, Bhupendra Singh ², Nilesh Khairnar ³, Sushmita Chavan ³

¹ Research Scholar, Teerthanker Mahaveer University, Moradabad UP India

² Professor, Teerthanker Mahaveer University, Moradabad UP India

³ Dept of Pharmaceutics, Valmik Naik College of Pharmacy, Telwadi Kannad Dist Aurangabad 431103

 

 

INTRODUCTION

Gliclazide is oral hypoglycemic drug which belong to second generation sulphonylureas. Chemically it is [1-(1-azabicyclo (3, 3, 0) octyl)-3-(p-tolylsulphonylurea)] (Figure 1) 

 

Figure 1: Chemical structure of Gliclazide

The mechanism of action of Gliclazide may reverse insulin resistance in type-II diabetic patients and improve defective insulin secretion. It is readily absorbed from the gastro-intestinal tract with peak concentrations in plasma occurring about 2-4 h and it is highly protein bound ^1-3. Various analytical methods have been reported for the estimation of the drug including HPLC, Gas chromatography, radioimmunoassay, and Evaporative Light Scattering Detection ^4-8 However, no UV spectrophotometric method is available for quantitative determination of Gliclazide in its pharmaceutical dosage forms. Some reported analytical methods involve time-consuming and laborious extraction steps ^{5, 6}, lengthy retention time or large volume of biological samples ^{5, 6}, use of mass spectroscopy for detection and identification of drug or solid phase extraction processes ^9-11. The present aim of the study is to develop a simple, rapid, accurate and specific UV spectrophotometric method for the estimation of Gliclazide in pharmaceutical dosage forms. The method was further validated for the parameters like precision, accuracy, sensitivity and linearity. The limit of detection (LOD) and limit of quantification (LOQ) were also determined. The results of the analysis were validated statistically and by recovery studies. The proposed method is recommended for routine analysis since it is simple, accurate, rapid, and sensitive.

 MATERIALS AND METHOD

Instruments and Reagents: 

Gliclazide was procured as a gift sample from J B Chemicals Thane. All the reagents used in the study were of analytical grade. Distilled water is used throughout the study. Different brands of tablets were procured from local medical shops. These are of brand glycogen by USV Ltd and glycol by Cedilla Pharmaceuticals. UV/VIS 1600 spectrophotometer with 1 cm matched quartz cells was used for the estimation manufactured by A Shimadzu, Japan

Selection of solvent: 

The solubility of Gliclazide is determined in a variety of solvents as per pharmacopoeia standard. Solubility test was carried out in different solvents like methanol, ethanol, and 0.1N sodium hydroxide. From the solubility studies, it was found that Gliclazide is soluble in methanol and 0.1N sodium hydroxide. In this study, methanol was selected as the solvent.

Preparation of standard stock solution of Gliclazide 

An accurately weighed 5 mg of Gliclazide was dissolved in 10 ml of methanol in a 50 ml volumetric flask and the volume was adjusted up to the mark with distilled water to obtain a stock solution of 100 µg/ml. The solution was filtered through Whatman’s filter paper No. 41.

Determination of λmax:

 The standard solution of Gliclazide (10µg/ml) was scanned in the UV region (200-400nm) and the spectrum was recorded. The solvent was used as blank. It was seen that at 232nm maximum absorbance was found, as shown in Figure 2. Therefore, 232nm was selected for this study.

 

 

Figure2: absorbance spectra of Gliclazide

 

 

RESULTS 

Method Validation: 

The objective of method validation is to demonstrate that the method is suitable for its intended purpose. The method was validated for linearity, precision, accuracy, robustness, ruggedness, LOD, LOQ, and specificity as per ICH guidelines

Linearity: 

Using the standard stock solution, the various dilutions in the concentrations of 2, 4, 6, 8, 10, 12, 14 16, 18, and 20 were prepared. (Table 1) The solutions were scanned at 232 nm and the absorbance was recorded, given in Table 1. From this calibration curve was obtained by plotting absorbance versus concentration of Gliclazide and the linearity graph was represented in Figure 3. The correlation coefficient was found to be 0.997

Table1: linearity parameter

Sr no.	Concentration(µg/ml)	Absorbance
1	2	0.057
2	4	0.1532
3	6	0.2512
4	8	0.3168
5	10	0.4355
6	12	0.501
7	14	0.616
8	16	0.6911
9	18	0.8255
10	20	0.9129

 

Figure 3: linearity graph of Gliclazide

Precision:

 Repeatability of method was checked by scanning 10µg/ml of solution for 6 times. Intra-day precision was determined by checking the absorbance of (10µg/ml) on the same day. Inter-day precision was determined by checking the absorbance of (10µg/ml) on three different days. The %RSD was found to be 0.74% for intra-day and 0.95% inter-day as shown in table 2.

Table2: Precision parameter for Gliclazide

Scan	Intraday	Interday
1	0.4350	0.4320
2	0.4392	0.4395
3	0.4364	0.4360
4	0.4396	0.4384
5	0.4350	0.4306
6	0.4320	0.4326
mean	0.4345	0.4327
SD	0.003212	0.00412
% RSD	0.73	0.92

 

Accuracy: 

The accuracy study was conducted by spiking at three different concentration levels (80%, 100%, and 120%). At each level samples were scanned and from the absorbance recovery percentage was determined. 

Limit of detection and Limit of quantification: 

Limit of detection is the lowest amount of an analyte in a sample that can be detected, but not necessarily quantified, under the stated experimental conditions. The limit of quantification is the lowest amount of analyze in a sample that can be quantified under stated experimental conditions. The LOD and LOQ for Gliclazide were found to be 0.15µg/ml and 0.50µg/ml.

 

Assay of Gliclazide tablets:

 Two different brands of Gliclazide were analyzed using the validated method. For the analysis, six replicates of each brand were assayed. 20 tablets of Gliclazide were weighed and finely powdered. An accurately weighed quantity of powder equivalent to 50mg of Gliclazide was taken in a 50-volumetric flask. 10ml of methanol was added to it followed by 20ml of solvent. The solution was sonicated for 15 min and then filtered through Whatman’s filter paper No.41 and volume was adjusted with the solvent. From this further dilution was made to get a final concentration of 10µg/ml. The absorbance of the sample solution was measured and the amount of Gliclazide was determined by referring to the calibration curve. The results were presented in

 table 3.

 

Table 3: assay of Gliclazide tablets

 

 

DISCUSSION 

The method was validated and developed as per ICH guidelines. The method was validated in terms of linearity, precision, accuracy, robustness, ruggedness, LOD, LOQ and specificity. Beer’s law is obeyed over the concentration range of 2-20 µg/ml, using regression analysis of the linear equation y = 0.047x-0.043 with a correlation coefficient of r2=0.0997. The precision results show a % RSD of less than 2 at each level, which indicates clearly that the method is precise enough for the analysis of Gliclazide. The accuracy of the method was checked by recovery studies. The high recovery values indicated the accuracy of the developed method. The robustness and ruggedness studies reveal that the method is enough robust and rugged. The LOD and LOQ values indicate the sensitivity of the method. There was no interference observed from the excipients present in the formulation, indicating that the method is specific. Determination of Gliclazide in tablet formulation of two brands showed content of Gliclazide was very close to the labeled amount. The %RSD values in all the parameters were within the acceptable limit. The optical characteristics of the method are represented in Table 4.

Optical characteristics 

Table represents the optical characteristics and precision of the proposed method for estimation of Gliclazide. 

Table 4: Validation parameters for standard Gliclazide

* Indicates mean of six determinations (n=6)

CONCLUSION:

An approved UV Spectrophotometric technique has been produced for the assessment of Gliclazide in mass as well as drug measurement’s structure. The created technique was viewed as simple, exact, precised, specific and reproducible and straight over the fixation range examined. The proposed technique can be utilized for routine examination of Gliclazide in mass as well as drug details.

Conflict of Interest

The authors affirm that they have no known financial or interpersonal conflict that might have looked to have an impact on the research performed in this paper.

Acknowledgement

Authors are thankful to JB Chemicals thane for providing gift sample of drug. The required resources to perform work provided by Valmik Naik College of Pharmacy, Telwadi Kannad, for which the authors are grateful.

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