Validated RP-HPLC Method Development for Estimation of Ertugliflozin and Sitagliptin in Bulk and Dosage Forms

A simple, accurate, precise method was developed for the simultaneous estimation of the Ertugliflozin and Sitagliptin in bulk and tablet dosage form. Chromatogram was run through Std Discovery C18150 x 4.6 mm, 5. Mobile phase containing Buffer 0.1% OPA (2.2ph): Acetonitrile taken in the ratio 60:40 was pumped through column at a flow rate of 1 ml/min. Buffer used in this method was 0.1% Ortho Phosphoric Acid (OPA). Temperature was maintained at 30°C. Optimized wavelength selected was 218 nm. Retention time of Ertugliflozin and Sitagliptin were found to be 2.228min and 2.880min. %RSD of the Ertugliflozin and Sitagliptin were and found to be 0.4 and 0.7 respectively. %Recovery was obtained as 99.23% and 100.01% for Ertugliflozin and Sitagliptin respectively. LOD, LOQ values obtained from regression equations of Ertugliflozin and Sitagliptin were 0.20, 0.61 and 0.46, 1.40 respectively. Regression equation of Ertugliflozin is y = 43997x + 2639, and y = 46501.x + 13112 of Sitagliptin. Retention times were decreased and that run time was decreased, so the method developed was simple and economical that can be adopted in regular Quality control test in Industries. Original Research Article Rajeswari et al.; JPRI, 34(20B): 22-26, 2022; Article no.JPRI.84234 23

Literature review reveals that very few analytical methods have been reported for the determination of Ertugliflozin and Sitagliptin by using various analytical techniques like HPLC, LCMS, LCMSMS [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15]. It was found that no suitable validated method was available from the literature for determination of bioavailability and bioequivalence of Ertugliflozin and Sitagliptin in biological samples. The main aim of the present study is to develop an accurate, precise, sensitive, selective, reproducible and rapid analytical technique for simultaneous estimation of Ertugliflozin, Sitagliptin in bulk ant tablet dosage form.

Chemicals and Reagents
Ertugliflozin and Sitagliptin pure drugs (API) were obtained as gift sample from SS Pharma, Combination Ertugliflozin and Sitagliptin tablets (QTERN) were purchased from local market, Distilled water, Acetonitrile, Phosphate buffer, Methanol, Potassium dehydrogenate ortho phosphate buffer, Ortho-phosphoric acid. All the above chemicals and solvents are from Rankem.

Instruments
Electronics Balance-Denver, p H meter -BVK enterprises, India, Ultrasonicator-BVK enterprises, WATERS HPLC 2695 SYSTEM equipped with quaternary pumps, Photo Diode Array detector and Auto sampler integrated with Empower 2 Software.UV-VIS spectrophotometer PG Instruments T60 with special bandwidth of 2 mm and 10mm and matched quartz cells integrated with UV win 6 Software was used for measuring absorbances of Ertugliflozin and Sitagliptin solutions.

Preparation of Solutions
Diluent: Based up on the solubility of the drugs, diluent was selected, acetonitrile and water taken in the ratio of 50:50. Preparation of Sample stock solutions: 5 tablets were weighed and the average weight of each tablet was calculated, then the weight equivalent to 1 tablet was transferred into a 10 ml volumetric flask, 5ml of diluents was added and sonicated for 25 min, further the volume was made up with diluent and filtered by HPLC filters (150µg/ml of Ertugliflozin and 1000µg/ml of Sitagliptin).

Method Development and Optimization
Method development was done by changing various method parameters like mobile phase ratios, column, flow rate, wavelength etc. Chromatographic separation was achieved with Mobile phase in the ratio of 60% 0.1% OPA buffer: 40% Acetonitrile. Water and Acetonitrile in the ratio 50:50 was used as Diluent. Chromatographic separation was achieved on Discovery C18 (4.6 x 150mm, 5µm) column with 1 ml/min flow rate at 218 nm wave length.
System suitability: All the system suitability parameters were within the range and satisfactory as per ICH guidelines.

Fig. 2. System suitability Chromatogram for Ertugliflozin and Sitagliptin
Discussion: According to ICH guidelines plate count should be more than 2000, tailing factor should be less than 2 and resolution must be more than 2. All the system suitable parameters were passed and were within the limits. Various parameters were tabulated.

CONCLUSION
The proposed research work is highly specific and prior over other developed methods reported previously. Chromatographic conditions were improved. Hence the proposed method has significant advantages over previously reported methods in-terms of selectivity, sensitivity, linearity, reproducibility, accuracy. Therefore the developed method can be used in quality control laboratories for estimation of Ertugliflozin and Sitagliptin in bulk and tablet dosage form.

DISCLAIMER
The products used for this research are commonly and predominantly use products in our area of research and country. There is absolutely no conflict of interest between the authors and producers of the products because we do not intend to use these products as an avenue for any litigation but for the advancement of knowledge. Also, the research was not funded by the producing company rather it was funded by personal efforts of the authors.

CONSENT
It is not applicable.

ETHICAL APPROVAL
It is not applicable.