INTRODUCTION

Dapagliflozin is a Category III antidiabetic drug under the Biologics Classification System (BCS) of the European Medicines Agency (EMA). These inhibitors are a new class of antidiabetic drugs called flozins, which are more soluble and nearly impermeable [1]. It is a sodium-glucose co transporter 2 (SGLT2) inhibitor that works largely by inhibiting glucose reabsorption from the liver, resulting in higher urine glucose excretion and, as a result, decreased blood sugar levels in type 2 diabetes patients. The medication has demonstrated an enhanced mode of action that is independent of insulin and only depends on plasma glucose and renal function. Dapagliflozin is a pill that is taken orally. It is particularly effective in the treatment of type 2 diabetes mellitus (DM) patients, both as a single agent and in combination with other anti-diabetic medications. Recent studies have shown that the fast action of Dapagliflozin decreased the fasting plasma glucose levels within one week of treatment [2].

This is a crystalline white powder that is readily soluble [3] in methanol, ethanol, dimethylformamide, and dimethylsulfoxide. Chemically, it is (1S)-1, 5-Anhydro-1-[4-chloro-3-(4-ethoxybenzyl) phenyl]-D-glucitol with a molecular weight of 408.98 and a molecular formula of C₂4H₃₃ClO₈. Figure 1 depicts the chemical structure of Dapagliflozin.

Literature review [4] and overview [5] deduce the drug from bulk and marketed formulations by UV spectroscopy [6-9] and RP-HPLC [10-18], UPLC [19, 20]. There have been only a couple of techniques published for UV spectrophotometric measurement of Dapagliflozin alone and in combination with other drugs [21, 22], utilizing commercially available solvents and buffers. This method was developed using methanol for solubilization and distilled water as the solvent for dilution. This is the most economical method of routine analysis and has not been previously reported based on a literature search performed before the work began. In addition, the newly proposed technique was validated for accuracy, precision, robustness, and linearity in accordance with ICH Q2 (R1) [23]. The results showed the reliability of the method.

MATERIALS AND METHODS

Equipment

The suggested study was conducted using a UV-1800 SHIMADZU and UV-3200 LAB INDIA UV-Visible spectrophotometer with 1 cm quartz-matched cells. Weighing was done on an electronic balance (Shimadzu-BL220H), sonicated with Sonica Ultrasonic Cleaner, Spincotech PVT LTD

Chemicals and reagents

Dapagliflozin-standard was acquired as a gift sample from Dr.Reddy Laboratories, Hyderabad, Dapagliflozin-tablets (Udapa*10) label claim 10 mg produced by MSN Laboratories was purchased from the local market, and Analytical grade solvent-methanol was obtained from Rankem, Maharashtra, India.

Standard preparation

Considering that methanol was discovered to make the medicines soluble, the standard stock solution was produced by dissolving 10 mg of Dapagliflozin in 1.5 ml of methanol and then increasing the volume to 10 ml with distilled water to reach a concentration of 1000 g/mL. A suitable dilution of the standard stock solution using distilled water was used to create the working standard solution, which contained 10 g/ml.

Determination of wavelength of maximum Absorption

The drug's maximum absorbance (max) was discovered to be 220 nm, as shown in Figure 2 when 10 g/ml of standard Dapagliflozin was scanned in a UV spectrophotometer between 190 and 300 nm.

Assay

The quantity of tablet powder equal to 10 mg of Dapagliflozin was precisely weighed, transferred to a 100-ml volumetric flask, and solubilized by dissolving in 15 mL of methanol, sonicating for 10 minutes, and then diluting with distilled water to the mark to obtain a concentration of 100 g/ml, followed by filtering through a Whatman filter paper. A 10 ml volumetric flask containing 1 ml of the filtrate was filled with the mixture, which was then diluted with distilled water to achieve the final concentration of 10 g/ml. The results of measuring the absorption using a selected wavelength are displayed in Table 1.

Weight of 10 Tablets = 2830 mg

10 Tablets average weight = 2830/10 =283 mg

Table 1. Percentage assay of Dapagliflozin

Method validation

The process of creating narrative proof that a system, procedure, or movement has been put into use or tested and maintained the necessary level of consistency across all phases is known as methodology validation. Approved scientific strategies are essential for improving diagnostic techniques and have been time and again tested for specificity, linearity, accuracy, precision, range, limits of detection, and quantization cutoffs. In summary, the development and approval of a systematic strategy confirm that accurate and reliable potency estimation of medicinal products has been performed.

Validation parameters

To prove in writing that a method's performance complies with the demands of the intended analytical application, validation parameters are utilized. The purpose of the verification is to demonstrate that the analytical results obtained using a particular method are suitable for that purpose and are as specified below.

Method validation

The goal of this work was to provide a novel, simple, and affordable method for measuring Dapagliflozin spectroscopically. In accordance with the ICH recommendations, the method's linearity, accuracy, precision, and dependability were evaluated.

Linearity

The Linear relationships must be evaluated using a variety of analysis methods. For linearity to be established, at least five concentrations are advised. By diluting the standard stock solution using the suggested technique, it may be directly identified with the API. The correlation coefficient, the point of intersection with the ordinate, and the slope of the regression line should be provided.

Accuracy

Recovery tests were carried out by combining known quantities of the standard medication with formulation samples in order to verify the validity of the aforementioned procedure. Three distinct levels—50%, 100%, and 150%—were used for recovery tests.

Precision

The accuracy of the process is determined by the degree of agreement between the individual test results when it is employed for several samples of a homogeneous sample. The performance of feature parameters and the statistical processing of analytical data for both intraday and Interday are both necessary for the validation of analytical procedures. These procedures establish the analytical data's tolerance for variance. Typically, the variance, standard deviation, or coefficient of variation of a collection of measurements is used to describe the analytical procedure's accuracy.

Reproducibility

Reproducibility is also referred to as intra-assay precision. Reproducibility is defined as short-term accuracy under identical conditions of use.

Robustness

Robustness evaluation relies on the type of approach being researched and should be taken into account throughout the design process. The reliability of the assay concerning intentional changes to method parameters should be demonstrated.

LOQ (Limit of detection) and LOQ (Limit of quantification)

Response standard deviation and determined by the linearity slope

The detection limit (DL) can be written as follows: DL = 3.3 σ/ S, where σ is the response's standard deviation and S is the calibration curve's slope. The limit of quantification (LOQ) can be written as LOQ = 10 σ/S, where σ is the response's standard deviation and S is the calibration curve's slope. The slope can be calculated using the analyte calibration curve.

RESULTS AND DISCUSSION

All validation parameters were carried out under the conditions mentioned in the ICH Q2 R (1) guidelines.

Linearity and range

The results from the linearity research were produced using five distinct aliquots of the reference solution and a chosen wavelength of 220 nm to test various concentrations (5, 10, 15, 20, and 25 g/mL). The limit of detection (LOD), limit of quantification (LOQ), and standard curve plot of the assay were also computed. The findings are presented in Table 2.

Table 2. Linearity of Dapagliflozin in working standards.

Acceptance criteria: correlation coefficient (r²) -0.999.

Dapagliflozin concentration was plotted on the X-axis, and absorbance was plotted on the Y-axis to create a calibration curve. Figures 3 and 4 of the r² correlation reveal that a linear association was seen in the concentration range of 5 to 30 g/ml (r²-0.999).

Accuracy

Recovery studies evaluated the proposed accuracy. A known amount of the drug substance Dapagliflozin (100 μg/ml) diluted from a stock solution was added to the pre-analytical tablet formulation (100 μg/ml) into a 10ml volumetric flask and volume made upto 10 ml with distilled water. Analysis of Dapagliflozin was performed at 50%, 100%, and 150% concentrations. The suggested procedure's recoveries were estimated. The results are displayed in Table 3.

Table 3. Accuracy readings of Dapagliflozin