AEGAEUM JOURNAL ISSN NO: 0776-3808
“ESTIMATION OF ANAGLIPTIN AND METFORMIN HCl IN BULK & PHARMACEUTICAL DOSAGE FORM USING STABILITY INDICATING HIGH PERFORMANCE LIQUID CHROMATOGRAPHY”
Ruchi H. Majithia*1 & Dr. Akruti Khodadiya2 *Assistant Professor, SAL Institute of Pharmacy, Ahmedabad, Gujarat- 380060 1Research Scholar, C. U. Shah College of Pharmacy and Research, Gujarat- 363030 1Mail ID: [email protected] 1Mobile No: +91-8155921801 2Associate Professor, C.U. Shah University, Wadhwan city, Gujarat- 363030 2Mail ID: [email protected] *Corresponding author
Volume 8, Issue 7, 2020 http://aegaeum.com/ Page No: 46 AEGAEUM JOURNAL ISSN NO: 0776-3808
ABSTRACT: A simple, accurate and precise stability indicating high performance liquid chromatography was developed and validated for estimation of Anagliptin and Metformin HCl in bulk and pharmaceutical dosage form. The mobile phase Potassium Dihydrogen Phosphate Buffer (pH 2.7): Acetonitrile solution in the ratio of (15:85% V/V) at detection wavelength 243 nm for Anagliptin and Metformin and flow rate 2 mL/min and the retention time for Anagliptin and Metformin HCl was found to be 11.23 and 6.02 respectively. The method was linear over the concentration ranges 10-60 μg/mL for Anagliptin and 25-150 μg/mL for Metformin. The LOD was 1.158 μg/mL for Anagliptin and 2.344 μg/mL for Metformin. The LOQ was 3.508 μg/mL for Anagliptin and 7.102 μg/mL for Metformin. During stress conditions, ANA degraded significantly under acidic, alkaline, oxidative and thermal stress conditions; degraded moderately under photolytic stress conditions; and showed negligible degradation under elevated temperature and humidity conditions. MET degraded significantly under acidic, alkaline and oxidative stress conditions; degraded moderately under thermal and humidity stress conditions and showed negligible degradation under photolytic stress condition.
KEYWORDS: Anagliptin (ANA), Metformin HCl (MET), High Performance/ Pressure Liquid Chromatography(HPLC), Simultaneous estimation. INTRODUCTION Anagliptin is used for type 2 diabetus mellitus and longer duration of action for treatment of type 2 non-insulin dependent diabetus mellitus disease as a Dipeptidyl Peptidase 4 inhibitor Anagliptin, chemically N-[2-[[2-[(2S)-2-Cyanopyrrolodin-1-yl]-2-oxoethyl] amino]-2- methylpropyl]-2-methylpyrazolo [1, 5-a] pyrimidine-6-carboxamide (Figure 1) is a Dipeptidyl Peptidase 4 (DPP 4) inhibitor which is used in treatment of type 2 NIDDM [01]. Dipeptidyl Peptidase 4 enzyme breaks down the incretins GLP-1 gastrointestinal hormones released in response to a meal. By preventing GLP-1 inactivation, they are able to increase the secretion of insulin and suppress the release of glucagon by the alpha cells of the pancreas. This drives blood glucose levels towards normal level [02-03]. This drug is not official in any of the pharmacopeia. Anagliptin is a very effective pill with minimum risk profile for type 2 diabetus mellitus and longer duration of action for treatment of type 2 non- insulin dependent diabetus mellitus disease [04]. A literature survey revealed that few methods are reported for determination of ANA, either alone or in combination, by spectrophotometric [05-07], HPLC [08-09], LC/MS [10].
Volume 8, Issue 7, 2020 http://aegaeum.com/ Page No: 47 AEGAEUM JOURNAL ISSN NO: 0776-3808
Figure 1: Chemical Structure of Anagliptin
.HCl
Figure 2: Chemical structure of Metformin HCl
Metformin is chemically a 1-Caramimidamido-N, N-Dimethylmethanimidamide [11] (Figure 2) and has pharmacological action based on Biguanides category [12-13]. It suppresses hepatic gluconeogenesis and glucose output from liver. This is the major action responsible for lowering blood glucose in diabetics. It is official in IP [14], BP [15], and USP [16]. Literature review revels that many spectrophotometric [17-18], HPLC [19-20], HPTLC [21] methods are reported for determination of Metformin hydrochloride (HCl), either alone or in combination[08]. The aim of the present work was to develop a stability indicating HPLC method for simultaneous estimation of ANA and MET in bulk and pharmaceutical dosage form. It is pertinent to note that, some of the published methods enabled estimation of drugs in combination products containing two drugs via spectrophotometric. However, so far, not any HPLC method was reported in pharmaceutical dosage form. Hence, HPLC chromatographic conditions were developed with forced degradation studies and the method was validated to establish selectivity with respect to excipients and degradation products.
MATERIALS AND METHOD:
Apparatus
Chromatographic separation of drugs was performed on a Shimadzu HPLC instrument (LC_2010 CHT) [software LC Solution, equipped with photo diode array detector (SPD-
Volume 8, Issue 7, 2020 http://aegaeum.com/ Page No: 48 AEGAEUM JOURNAL ISSN NO: 0776-3808
M20A), Auto-sampler]. The system contains a quaternary gradient pump, auto sampler, column oven and a PDA detector. Software referred to above was used to record and integrates the chromatograms. Among other instruments, analytical balance (Acculab ALC- 210.4, Huntingdon Valley, PA), Photostability chamber (TH-90S, Thermolab, Mumbai, India) , Hot air oven (TO-90S, Thermolab), pH meter (Thermo Electron Crop., Pune, India), Sonicator (EN 30 US, Enertech Fast Clean, Mumbai, India) were also used at different stages of development and validation of stability indicating assay method.
Chemicals and Reagents
The API of Anagliptin and Metfromin procured from Intas Pharmaceuticals LTD, Ahmedabad. Combination test product METOANA Tablets 100/250 mg (manufactured by Sanwa Kagaku Kenkyusho Co. LTD) was procured from the market. All the solvents like Methanol, Acetonitrile, Phosphate Buffer and Potassium dihydrogen phosphate were of HPLC grade from Finar Chemicals Ltd, Ahmedabad, India. All the chemical reagents were of analytical grade.
Preparation of Standard Stock Solution
The standard solution was prepared by weighing accurately about 10 mg of ANA and 25 mg of MET transferred into clean and dry 10 mL volumetric flask. Initially about 5 mL methanol was added to the flask respectively and sonicated. The volume was made up to the mark with the methanol to achieve 1000 µg/mL of ANA and 2500 µg/mL of MET standard stock solutions.
Preparation of working standard solution
From the above prepared Stock solutions, pipette out 1 mL of ANA and 1 mL MET solution and transferred into a clean and dry 10 mL volumetric flask, methanol was added upto the mark to get final concentration 100 µg/mL & 250 µg/mL respectively, in mixture.
Preparation of test solution
Twenty METOANA tablets were accurately weighed, their average weight was calculated. Amount of finely powdered tablet equivalent to 100 mg ANA and 250 mg MET was weighed and transferred into a 100 mL volumetric flask and the volume was adjusted to mark with methanol. The contents of the flask were sonicated for 30 min to dissolve the active ingredients completely. The solution was filtered through a Whatman filter paper no. 41.
Volume 8, Issue 7, 2020 http://aegaeum.com/ Page No: 49 AEGAEUM JOURNAL ISSN NO: 0776-3808
From this 1 mL aliquot was transferred into a 100 mL volumetric flask and the volume was made with methanol. This corresponds to 10 µg/ mL of ANA & 25 µg/ mL MET, in mixture was analyzed for assay determination.
Standardized Chromatographic conditions
The analyte drugs and degradation products were well separated with ZORBAX RX C8 (250 mm × 4.6 mm, 5 µm) as a stationary phase and gradient system of Potassium Dihydrogen Phosphate Buffer (pH 2.7): Acetonitrile solution in the ratio of (15:85% V/V). The flow rate was adjusted to 2 mL/min. Determination was done at wavelength 243 nm. Column temperature was set to 300 C. Total run time was 20 min. Injection volume was 10 µL. Methanol was used as a diluent.
METHOD VALIDATION The proposed method was validated as per ICH guidelines Q2 R1 [22].
System suitability test parameters System suitability tests are used to verify that the resolution and repeatability of the system were adequate for the analysis intended. The parameters used in this test were the chromatographic peak resolution (>2), theoretical plate number (>2000) and tailing factor (<1.8). The repeatability of these parameters was checked by injecting six solutions of ANA and MET.
Linearity From the standard stock solution containing 100 µg/mL ANA and 250 µg/mL MET, aliquots of 1 mL, 2 mL, 3 mL, 4 mL, 5 mL and 6 mL were transferred in clean and dry 10 mL volumetric flasks respectively. The volume was made up to the mark with diluent. This yielded solution of 10, 20, 30, 40, 50 and 60 µg/mL of ANA and; 25, 50, 75, 100, 125 and 150 µg/mL of MET respectively. An injection volume of 10 μL of each solution was injected under operating chromatographic condition. Six replicates of each concentration were performed and then calibration plots were determined by linear least – squares regression. The plate was developed on previously described mobile phase. The peak areas were plotted against the corresponding concentrations to obtain the calibration graphs.
Precision Repeatability was determined by applying six replicates of test solution (10 µg/mL ANA and 25 µg/mL MET). The intraday and interday precisions were determined by responses of six
Volume 8, Issue 7, 2020 http://aegaeum.com/ Page No: 50 AEGAEUM JOURNAL ISSN NO: 0776-3808
replicates on same and different days for the test concentration. The results were reported in terms of % RSD.
Accuracy Recovery study was carried out by standard addition method where known amount of standard concentration at 50%, 100% and 150% of the test solution were spiked in the test solution in triplicate. The amount of drugs was estimated by substituting values in regression equation. The % RSD of the recovery was calculated.
LOD and LOQ The LOD and LOQ of the developed method were calculated from the calibration curve using equations, LOD = 3.3 ×ϭ/S and LOQ = 10 × ϭ/S where ϭ is the standard deviation of y- intercept and S is the slope of the curve.
Robustness By introducing small changes in the detection wavelength (±2 nm) and saturation time for the chamber before plate development (±2 min), the effects on the results were determined. One factor at a time was changed and the effect on peak area of the drug was studied. Robustness of the method was done on single level for six replicates and %RSD was calculated.
FORCED DEGRADATION STUDIES: Force degradation study was intended to ensure the effective separation of ANA and MET and their potential degradation products which are generated under different stress conditions like acid, alkaline and neutral hydrolysis, oxidative degradation, thermal and photolytic degradation.
Acid Hydrolysis Accurately weighed 10 mg ANA and 25 mg MET were transferred in 10 mL volumetric flask individually and in combination. To this were added 5 mL methanol and 5 mL of 0.1 N HCl and it was kept at room temperature for 5 h. From that solution 1 mL was transferred into 10 mL volumetric flask, neutralized with 0.1 N NaOH and diluted to mark with methanol. This corresponds to 100 µg/mL ANA and 250 µg/mL MET were injected under operating condition. Alkaline Hydrolysis
Volume 8, Issue 7, 2020 http://aegaeum.com/ Page No: 51 AEGAEUM JOURNAL ISSN NO: 0776-3808
Accurately weighed 10 mg ANA and 25 mg MET were transferred in 10 mL volumetric flask individually and in combination. To this were added 5 mL methanol and 5 mL of 0.1 N NaOH and it was kept at room temperature for 1 h. From that solution 1 mL was transferred into 10 mL volumetric flask, neutralized with 0.1 N HCl and diluted to mark with methanol. This corresponds to 100 µg/mL ANA and 250 µg/mL MET were injected under operating condition. Oxidative Hydrolysis Accurately weighed 10 mg ANA and 25 mg MET were transferred in 10 mL volumetric flask individually and in combination. To this were added 5 mL methanol and 5 mL of 3% H2O2 and it was kept at RT for 3 h. From that solution 1 mL was transferred into 10 mL volumetric flask, neutralized with 0.1 N HCl and diluted to mark with methanol. This corresponds to 100 µg/mL ANA and 250 µg/mL MET were injected under operating condition. Thermal Degradation Accurately weighed 10 mg ANA and 25 mg MET were transferred in 10 mL volumetric flask individually and in combination in petridish. Those were kept at 700 C, 4 h for 4 h. After that those were dissolved in 10 mL methanol. From that solution 1 mL was transferred into 10 mL volumetric flask and diluted to mark with mobile phase. This corresponds to 100 µg/mL ANA and 250 µg/mL MET were injected under operating condition.
Photolytic degradation Accurately weighed 10 mg ANA and 25 mg MET were transferred in 10 mL volumetric flask individually and in combination in petridish. It was exposed in photo stability chamber (TH- 90S, Thermo lab, Mumbai, India) in UV light at 254 nm for 36 h to get 200 watt hours / m2 intensity. After that those were dissolved in 10 mL methanol. From that solution 1 mL was transferred into 10 mL volumetric flask and diluted to mark with mobile phase. This corresponds to 100 µg/mL ANA and 250 µg/mL MET were injected under operating condition.
RESULT AND DISCUSSION
RESULT AND DISCUSSION
OPTIMIZED CHROMATOGRAPHIC CONDITION
Volume 8, Issue 7, 2020 http://aegaeum.com/ Page No: 52 AEGAEUM JOURNAL ISSN NO: 0776-3808
For separation of ANA and MET, and their degradation peaks, different mobile phases with different solvents in different ratio were tried like (1) Water: ACN (50:50 %V/V) (2) Buffer: ACN (pH 3) (50:50 % V/V) (3) Acetonitrile: Buffer (pH 2.7) (70:30 %V/V). The mobile phase Potassium Dihydrogen Phosphate Buffer (pH 2.7): Acetonitrile solution in the ratio of (15:85% V/V) showed well resolved peak with better peak shape. The drugs were resolved with tR of 6.02 ± 0.41 min and 11.23 ± 0.37 min of MET and ANA, respectively. Determination of all three drugs was done at wavelength 243 nm with column temperature 300 C and adjusted flow rate 2.0 mL/min. Injection Volume was set 20 µL for a good peak shape. (Figure 3)
Figure 3: Chromatogram of STD ANA (100 µg/mL) and MET (250 µg/ mL)
Linearity
The method was found linear over the concentration range of 10-60 µg/ mL 25-150 µg/ mL for ANA and MET respectively. The calibration curve obtained by the least square regression analysis between average peak area and concentration showed linear relationship with a correlation coefficient of 0.9991 and 0.999 for ANA and MET respectively. The linear regression equation were y = 35.448x - 71.612, and y = 178.98x + 793.84 for ANA and MET respectively. (Table 1, Figure 4, Figure 5)
Table 1: Linearity range of ANA and MET ANA MET Sr Conc Conc No. Peak Area* ± SD %RSD Peak Area * ± SD %RSD (µg/ mL) (µg/ mL) 1 10 309.33 ± 3.20 1.03 25 5357 ± 66.15 1.23
Volume 8, Issue 7, 2020 http://aegaeum.com/ Page No: 53 AEGAEUM JOURNAL ISSN NO: 0776-3808
2 20 623.16 ± 7.54 1.21 50 9873 ± 132.89 1.34 3 30 971.66 ± 9.72 1.00 75 14161 ± 193.53 1.36 4 40 1345.33 ± 15.16 1.12 100 18349 ± 206.07 1.12 5 50 1687.5 ± 18.11 1.07 125 22751 ± 245.14 1.07 6 60 2077.33 ± 24.17 1.16 150 28153 ± 360.00 1.27 *Average of six determinations
Calibration curve of ANA at 243 nm 2500
2000
1500
1000 y = 35.448x - 71.612 500 R² = 0.9991
0 0 10 20 30 40 50 60 70
Figure 4: Calibration Curve of Linearity of ANA at 243 nm Calibration curve of MET at 243 nm 30000
25000
20000
15000
10000 y = 178.98x + 793.84 R² = 0.999 5000
0 0 20 40 60 80 100 120 140 160
Figure 5: Calibration Curve of Linearity of MET at 243 nm
Precision The % RSD for repeatability was found to be 1.23 for ANA and 1.14 for MET. The % RSD of Intraday Precision was found to be 1.37 for ANA and 1.21 for MET. The % RSD of
Volume 8, Issue 7, 2020 http://aegaeum.com/ Page No: 54 AEGAEUM JOURNAL ISSN NO: 0776-3808
Interday precision was found to be 1.41 for ANA and 1.39 for MET. Hence, confirming precision of the developed method. (Table 2)
Table 2: Repeatability, Intraday and Interday Precision of ANA and MET ANA MET Parameter Conc Conc Peak Area* ± SD %RSD Peak Area * ± SD %RSD (µg/ mL) (µg/ mL) Repeatability 10 372 ± 4.50 1.23 25 4603.19 ± 52.73 1.14 Intraday 10 376.66 ± 5.16 1.37 25 4632.80 ± 56.40 1.21 Precision Interday 10 371 ± 5.25 1.41 25 4626.13 ± 64.59 1.39 Precision *Average of six determinations
Accuracy The accuracy of the developed method was established by standard addition method by adding known standard concentration solutions to the pre-analysed samples. Recoveries were in between 99.69-101.69 % for ANA and 99.68-101.45 % for MET which is in accordance with ICH guidelines which proves method to be accurate. (Table 3)
Table 3: Recovery study data of ANA and MET Amount of Amount Total Recovered Test of Stand. Amount % % Drug Peak Area* ± SD amount Solution added Found Recovery RSD (µg/ mL) (µg/ mL) (µg/ mL) (µg/ mL) 10 0 425 ± 5.29 9.96 -- 99.69 1.49 10 5 612.36 ± 7.07 15.25 10.25 101.69 1.30 ANA 10 10 790.66 ± 10.40 20.28 10.28 101.42 1.44 10 15 959 ± 15.71 25.11 10.11 100.47 1.76 25 0 5252.46 ± 50.43 24.92 -- 99.68 1.13 25 12.5 7588.7 ± 88.09 37.98 25.48 101.28 1.29 MET 25 25 9849 ± 94.69 50.61 25.61 101.23 1.04 25 37.5 12137 ± 120.37 63.40 25.9 101.45 1.06 *Average of three determinations
Volume 8, Issue 7, 2020 http://aegaeum.com/ Page No: 55 AEGAEUM JOURNAL ISSN NO: 0776-3808
LOD and LOQ The LOD calculated by formulae was found to be 1.158 μg/mL for ANA and 2.344 μg/mL for MET. The LOQ calculated by formulae was found to be 3.508 μg/mL for ANA and 7.102 μg/mL for MET.
Robustness Slight change in the chromatographic condition of the developed method like small changes in the Flow rate (±0.1 mL), column temperature (± 50 C), wavelength (± 2 nm) and pH of buffer solution in mobile phase (± 0.2) did not affect the result significantly. The % RSD values were found below 2 indicated the method to be robust. (Table 4)
Table 4: Robustness study of ANA and MET
% Assay* ± SD % RSD# Condition Variation ANA MET ANA MET Normal N/A 100.53 ± 1.07 100.21 ± 1.20 2.1 101.10 ± 1.23 99.98 ± 1.24 Flow rate mL/min (± 0.1 mL/min) 1.9 101.04 ± 0.87 100.96 ± 1.35 mL/min
Detection 241 nm 100.32 ± 1.11 101.2 ± 1.74 Wavelength (243±2 1.26 1.31 nm) 245 nm 100.02 ± 0.83 101.67 ± 1.03 Column 250 C 101.23 ± 1.23 99.45 ± 1.34 temperature 350 C 101.86 ± 1.45 99.75 ± 1.26 (± 50 C) 2.5 100.85 ± 1.18 101.15 ± 1.21 pH (±0.2) 2.9 101.15 ± 1.25 101.69 ± 1.24 *RSD of original and modified conditions
Analysis of marketed formulation
Volume 8, Issue 7, 2020 http://aegaeum.com/ Page No: 56 AEGAEUM JOURNAL ISSN NO: 0776-3808
The developed method was applied to marketed tablet preparation. The assay results of ANA and MET were 100.53 ± 3.89 % and 100.21 ± 47.99 %, respectively of the labelled amount. (Table 5)
Table 5: Analysis of marketed tablet formulation Amount of drug % Label claimed Drug % RSD (% Assay* ± SD)
Labeled Estimated
ANA 100 100.53 100.53 ± 3.89 1.09 MET 250 250.43 100.21 ± 47.99 1.07 *Average of six determinations
Forced degradation studies The result of forced degradation studies are summarized in (Table 6). Under the optimized chromatographic conditions degradation products of analytes were well resolved and the percent degradation was calculated by comparing peak area with standard preparation.
During stress degradation experiments, it was observed that ANA degraded significantly under acidic, alkaline, oxidative and thermal stress conditions; and showed negligible degradation under photolytic stress conditions and neutral hydrolysis condition. MET degraded significantly under alkaline and oxidative stress conditions; degraded moderately under acidic, neutral and thermal stress condition and showed negligible degradation under photolytic stress condition.
The purity of the drug (analyte peak) was ascertained by analysing spectrum at peak start, peak end and peak apex position which showed no interference of any excipients or process impurities in analyte peak. The method is also deemed to be specific from potential degradation products as most of the degradation products are adequately resolved from both analyte peaks. The sample tablet was exposed to thermal and photolytic degradation conditions as per ICH Q1A R2 guidelines [23]. The method was therefore being considered to be stability indicating for tablet solid dosage form. (Figure 6, Figure 7, Figure 8, Figure 9, Figure 10 and Figure 11)
Volume 8, Issue 7, 2020 http://aegaeum.com/ Page No: 57 AEGAEUM JOURNAL ISSN NO: 0776-3808
Figure 6: Chromatogram of Acid hydrolysis of mixture of ANA and MET at 243 nm
Figure 7: Chromatogram of Alkali hydrolysis of mixture of ANA and MET at 243 nm
Figure 8: Chromatogram of Neutral hydrolysis of mixture of ANA and MET at 243 nm
Volume 8, Issue 7, 2020 http://aegaeum.com/ Page No: 58 AEGAEUM JOURNAL ISSN NO: 0776-3808
Figure 9: Chromatogram of Oxidative hydrolysis of mixture of ANA and MET at 243 nm
Figure 10: Chromatogram of Thermal degradation of mixture of ANA and MET at 243 nm
Volume 8, Issue 7, 2020 http://aegaeum.com/ Page No: 59 AEGAEUM JOURNAL ISSN NO: 0776-3808
Figure 11: Chromatogram of Photolytic (UV Light) degradation of mixture of ANA and MET at 243 nm
Table 6: Summary of Forced Degradation study of individual API in mixture of ANA and MET No. of tR of Degradation Degradation tR of Analyte degradati %Degradation peak condition on peaks ANA MET ANA ANA MET 0.1 N HCl at 1.903, 3.019, 7.511, 11.453 6.011 4 4.79 16.97 RT, 5 h 8.801 5.225, 6.617, 0.1 N NaOH 11.45 6.011 5 14.138, 15.057, 26.63 23.75 at RT, 1 h 15.561 Water, RT, 3 11.23 6.02 -- -- 1.02 1.17 h
3% H2O2, RT, 1.9, 3.1, 8.8, 14, 11.453 6.011 5 13.69 15.64 3 h 15.1 Thermal at 11.453 6.011 3 3.01, 7.5, 10.0 1.83 9.09 700 C, 4 h UV light, 254 11.23 6.02 2 1.9, 3.0 1.15 3.77 nm, 36 h
Table 7: Summary of Validation Parameters
Sr. Parameter ANA MET No. 1 Specificity Specific Specific 2 tR value 11.23 ± 0.37 6.02 ± 0.41 3 Linearity Range 10-60 μg/mL 25-150 μg/mL 4 Regression Line equation y = 35.448x - 71.612 y = 178.98x + 793.84 5 Correlation Coefficient 0.9991 0.999 Precision 6 Repeatability 1.23 1.14
Volume 8, Issue 7, 2020 http://aegaeum.com/ Page No: 60 AEGAEUM JOURNAL ISSN NO: 0776-3808
Intraday Precision 1.37 1.21 Interday Precision 1.41 1.39 7 Accuracy (% Recovery) 99.69-101.69 % 99.68-101.45 % 8 LOD (μg/mL) 1.15 2.34 9 LOQ (μg/mL) 3.50 7.10 10 Robustness Robust Robust
CONCLUSION The proposed HPLC method is precise, specific, linear and accurate for the estimation of ANA and MET in pharmaceutical dosage form without interference from the excipients and potential degradation products in various stress conditions like acid hydrolysis, alkaline hydrolysis, neutral hydrolysis, oxidative, thermal and photolytic degradation conditions. The developed method is validated as per ICH guidelines. The results showed the suitability of developed method for stability studies of the pharmaceutical dosage form.
AKNOWLEDGEMENT The authors are graceful to INTAS Pharmaceuticals, Ahmedabad for providing a gift sample of Anagliptin and Metformin. The authors are grateful to Global Analytical Laboratory, Ahmedabad for providing all the facilities to carry out research work.
CONFLICT OF INTEREST: Nil
REFERENCES 1. “Anagliptin Compound ” (Accessed on June, 2020) https://pubchem.ncbi.nlm.nih.gov/compound/44513473 2. H. Mohan, Textbook of Pathophysiology, 6th Edn, Medical publishers Pvt Limited, (2005) 818 - 828. 3. K. D. Tripathi, Essential of Medical Pharmacology, 6th Edn, Jaypee Brother Mesical Publishers Ltd, (2008) 254 - 274. 4. Y. Takeuchi, M. Namiki, Y. Kitahara, Safety, Tolerability, Pharmacokinetics and Pharmacodynamics of E3024, a Novel and Selective Dipeptidyl Peptidase-IV Inhibitor, in Healthy Japanese Male Subjects: Rash Development in Men and Its Possible Mechanism, Pharmacology & Pharmacy. 4 (2013) 663-678.
Volume 8, Issue 7, 2020 http://aegaeum.com/ Page No: 61 AEGAEUM JOURNAL ISSN NO: 0776-3808
5. P. Bhatti, H. Panchal, Stability Indicating Chromatographic Method Development And Validation for the Simultaneous estimation of Metformin HCl & Anagliptin in its synthetic mixture by HPLC, World J. of Pharmaceutical Res. 6 (2017) 956-973. 6. R. Majithia, J. Shah, Development and validation of analytical method for estimation of Anagliptin tablet dosage form by UV spectrophotometric method, International J. of Pharmacy and Technologies. 6 (2015) 7765–7771. 7. S. Shah, D. Maheshwari, First order Derivative Spectrophotometric method for Simultaneous Estimation of Anagliptin and Metformin HCl in Bulk and Synthetic Mixture, J. of Glob. Tr. in Pharmaceutical Sci. 6 (2015) 2925-2929. 8. A. Patil, A. Shirkhedkar, Development and Validation of Five Simple UV- Spectrophotometry Methods for Estimation of Anagliptin in Bulk and in-house Tablets, Pharm. Methods. 7 (2016) 127-131. 9. C. Pandya, S. Rajput, Stress Degradation Studies of Anagliptin, Development of Validated Stability Indicating Method, Degradation Kinetics Study, Identification and Isolation of Degradation Products, Chromatographia, An International J. of sep. sci. (2018) 1-21. 10. S. Baira, D. Sigalapalli, N. Bathini, LC/QTOF/MS/MS characterization, molecular docking and in silico toxicity prediction studies on degradation products of Anagliptin, J. of Pharmaceutical and Biomedical Anal. 159 (2018) 92-99. 11. “Metformin Hydrochloride Compound ” (Accessed on June, 2020) https://www.drugbank.ca/drugs/DB00331
12. W. Roger, Cate W, Clinical Pharmacy & Therapeutics, 4th Edn, Churchill Livingstone Elsevier, (2008) 685 - 710.
13. H. Rang, M. Dale, Rang & Dale’s pharmacology, 7th Edn, Elsevier Publication, (2012) 402 - 408 14. IP: The Indian Pharmacopoeia Commission, 2014, Ghaziabad, India. 15. BP: The British Pharmacopoeia, Medicine & Healthcare Products Regulatory Agency, 2010, London. 16. USP: The United States Pharmacopoeia Convention, 2010, Rockville, MD, USA. 17. C. Parle, A. Parle, Development and validation of UV spectrophotometric method for simultaneous estimation of metformin hydrochloride and Alogliptin benzoate in bulk drugs and combined dosage forms, Der Pharma Chemica. 6 (2014) 303-311.
Volume 8, Issue 7, 2020 http://aegaeum.com/ Page No: 62 AEGAEUM JOURNAL ISSN NO: 0776-3808
18. J. Chopade, S. Deshpande, S. Shah, Simultaneous estimation of Metformin HCl and Gliclazide by Q- Analysis Method, International J. for Pharmaceutical Res. Scho. 2 (2013) 66- 73. 19. P. Umapathi, J. Ayyappan, Quantitative Determination of Metformin Hydrochloride in Tablet Formulation Containing Croscarmellose Sodium as Disintegrant by HPLC and UV Spectrophotometry, Tropical J. of Pharmaceutical Res. 11 (2012) 107-116. 20. M. Kar, P. Chaoudhary, HPLC Method for Estimation of Metformin HCl in Formulated Microspheres and Tablet Dosage Form, Indian J. of Pharmaceutical Sci. 71 (2009) 318-320. 21. C. Reddy, G. Mubeen, M. Pal, HPTLC method for estimation of metformin hydrochloride, Biomed. & Pharmacol. J. 1 (2008) 445-448. 22. ICH Harmonized Tripartite Guideline, Validation of Analytical Procedures: Text and Methodology Q2 (R1), International Conference on Harmonization, Geneva, Switzerland, (2005) 1 – 12. 23. ICH Harmonized Tripartite Guideline, Stability Testing of New Drug Substances and Products Q1 A (R2), International Conference on Harmonization, Geneva, Switzerland, (2003) 1 – 20.
Volume 8, Issue 7, 2020 http://aegaeum.com/ Page No: 63