Analytical Method development and Validation of Teneligliptin by

RP-UFLC

 

Maruthi R¹, Chandan R.S*, Barath M¹, Mr. G Naveen Datta², Ms. Merryl D’silva²,

Ms. Kajal Kumari M³, Mr. Farhan Ahmad⁶, Ms. Geetha R⁷

¹Department of Pharmaceutical Chemistry, JSS College of Pharmacy, Mysuru, JSS Academy of Higher Education and Research, Sri Shivarathreeshwara Nagara, Mysuru- Karnataka, India 570015

²Department of Pharmaceutics, JSS College of Pharmacy, Mysuru, JSS Academy of Higher Education and Research, Sri Shivarathreeshwara Nagara, Mysuru- Karnataka, India 570015

³Department of Pharmacy Practise, JSS College of Pharmacy, Mysuru, JSS Academy of Higher Education and Research, Sri Shivarathreeshwara Nagara, Mysuru- Karnataka, India 570015

 

ABSTRACT:

To estimate Teneligliptin by Ultra-Fast Liquid Chromatographic (RP-UFLC), a fast, reliable, and cost-effective UFLC method was developed. The approximate flowing rate was 1.0 mL/minfor the Phenomenex Kinetex column C18 (250mmx 4.6mm. 5 Micron) with MeOH and ACN (60:40 v/v) as a portable stream. A PDA detector at 246 nm was used to track the eluent. Teneligliptin elutes at 4.938 minutes of the processing period. The proposed method has a linearity of 10 to 50μg/ml concentration range; the calibration diagram is continuous and 0.9942 percent for the correlations variable (r2). For the suggested procedure, both LOD and LOQ are measured as 0.097 and 1.023μg/mL, respectively. In the pharmaceutical product, the drug has been under the states of stress conditions acidic, alkaline, degradation, Ultraviolet, and heat stress. The degradation product from the Teneligliptin was well solved, and thus the stability of this method was demonstrated. The process was tested in compliance with ICH criteria of device suitability, linearity, reliability, consistency, and robustness.

 

 

INTRODUCTION:

Determination of the chemical composition of the material involves the classification of the product, verification of the substance structure, and quantitative analysis of its composition. The product analysis is used to isolate, assess, and identify the relative amounts of the materials. A chemical engineering research can be used for the mixture components in science and art to assess the structure of the material. The main area of analytical chemistry in drug research. Pharmaceutical tests are based on the principles of chemistry, microbiology, electrical engineering, and so on, in various fields of science.

 

Analytical approaches can be used to conduct qualitative and quantitative studies. Value analysis has shown the specimen organisms' chemical identification. A separation phase is generally necessary for qualitative as well as quantitative analyses. There are two dimensions of estimation of the usual effects of the quantitative analysis. One is the specimen weight and quantity to be evaluated, and the other is the calculation of certain amounts in comparison to the testing aspect of the analysis. As an antagonist of dipeptidyl peptidase-4 (DPP-4), Teneligliptin has recently emerged as a new antidiabetic category that has positive effects on improved Glycaemic Regulation. DPP-4 inhibitors increase the efficacy of GP-1 DPP-4 inhibitors, thus increasing the glucose-dependent hyperglycemia in patients with diabetes by raising Serum insulin blockers concentration and reducing serum glucagon concentration. Such drugs are effective for patients with diabetes and serve as the latest anti-diabetic group of associated agents such as DPP-4. Such drugs may reduce glucose spikes¹.

 

Teneligliptin:

Molecular structure:

 

Figure 1. Chemical structure of Teneligliptin

 

 

Dose and Administration:

Teneligliptin is normal for adults at 20mg once every day. When insufficient, the dosage can be lowered to 40 mg once daily during close medical monitoring.^5,6

 

MATERIALS AND METHODS^7-13

Instrumentation:

For the current study, UFLC with PDA Detector LC-2010 AD a make of Shimadzu was used.

 

Table 1: Optimized Chromatographic Conditions for Teneligliptin

 

Chemicals and reagents:

Teneligliptin standard was procured from Micro labs, Bengaluru. Teneligliptin marketed formulation manufactured by WellCare Pharmaceuticals was procured from local market. All chemicals used were analytical grade purchased from Merck pharmaceuticals. HPLC grade MeOH was used as the diluent for preparation of the solutions.

 

Preparation of diluent:

MeOH (HPLC grade) was the diluent used.

 

Preparation of Mobile Phase:

MeOH and ACN in the ratio of (60:40 v/v) were used as the mobile phase.

 

Preparation of STD Soln:

Stock Solution for Teneligliptin:

In a volumetric flask of 100ml, teneligliptin 100mg was collected. Dissolved with 50ml diluent, make it up to label (1000μg/ml), and sonicated. In a 10ml volumetric flask, pipette 1ml of the above solution and use the diluent (100μg/ml) to appropriate volume².

 

Preparing linearity solutions

Linear alternatives are established by diluting the stock solution diluent. Conc of 10, 20, 30, 40, 50μg/mL of teneligliptin are prepared In 10 ml volumetric flasks. pipette 1, 2, 3, 4, 5ml, and dissolved in the diluent to obtain these concentrations dimension of 10-50μg/mL³.

 

Preparation of Calibration Curve:

Aliquots of teneligliptin are made from the stock solution (100μg/mL) in a 10ml volumetric flask sequence. The HPLC grade diluent has been used to dilute the label and filters it through a membrane filter of 0.45μg/mL to achieve a concentration range of 10-50 μg/mL. Solutions injected 10μL and total areas reported. The calibration curve has been set. The law on beer applies to 10-50μg/mL range⁴.

 

RESULTS AND DISCUSSION:

Systematic work on the impacts of different parameters has been carried out in the development of the method and optimization of chromatographic conditions. For the Teneligliptin drugs, solubility was first assessed. Chromatographic conditions have been optimized to achieve a good peak in the RP-UFLC method. In the end, the compound was eluted by the different affinity towards the mobile phase; the highest parameters (Peak area, height, strength, theoretical plate tail, or asymmetry factor) are chosen for the selection of mobile phase and flow rate. With a very high fluid ratio of 60:40 (v/v) mobile Phase B (ACN) method with the flow rate of 1 ml/ min was used. It consists of Stage A (MeOH) portable. The optimum wavelength for measurement was 246nm, offering the drug a better response to the detector. Teneligliptin's retention period was 4.93 min. The figures 1, 2 and 3 shown in the blank standard and teneligliptin chromatogram (sample).

 

Figure 2: Chromatogram of Blank (Diluent)

 

 

Figure 3: Chromatogram for Teneligliptin (Standard)

 

 

Figure 4: Chromatogram for Teneligliptin (sample)

 

System suitability:

Appropriate tests are used to check chromatographic system reproductiveness and system suitability for their effectiveness in analyzing the drug

 

Table 3: Results for System Suitability for Teneligliptin by UFLC Method

 

Evaluation of data:

Table 3 shows that the system appropriateness criteria are part of the authorization specifications of the information presented above.

Linearity:

Table 4: Results for Linearity of Teneligliptin by UFLC method

 

 

Figure 5: Calibration curve of Teneligliptin

 

criteria for approval:

Ø The coefficient of correlation should be greater than 0.99.

 

Evaluation of data:

Table 4 of Teneligliptin shows the reliable of linearity data that the Teneligliptin reaction is between 10 to 50 μg/ml of working concentration.

 

Precision:

When the method is used continuously for distinct and homogenous samples, an analysis method is accurate.

 

System Precision:

The system's accuracy is to guarantee a correctly functioning analytical system.

 

Table 5: Results for System precision studies for Teneligliptin by UFLC method

 

Criteria of approval:

In six standardized injections, RSD percent of the area response should be less than 2.0% for teneligliptin

Evaluation of data:

Table 5 above shows that the area response is consistent, displaying the RSD.

 

Method precision:

The method's precision shows whether a particular substance achieves reliable results or not.

 

Table 6: Results for Method precision studies for Teneligliptin by UFLC method

 

Criteria of approval:

In 6 determinations, the percentage of RSD calculated should be 2.0% LESS THAN.

 

Evaluation of data:

It can be concluded from Table 6 above that the technique is precise.

 

LOD and LOQ

The LOD is the lowest sample measurement to be defined but not generally measured under the specified experimental conditions.

 

The LOQ test is the lowest number of analytes to be tested with reasonable precision and reliability under certain experimental conditions.

 

LOD and LOQ is determined on the basis of sloping, intercepting, and ratios, as well as the relative standard curve variance.

 

Table 7: Results for LOD and LOQ for Teneligliptin by RP-UFLC method

 

 

Table 8: Recovery results for Teneligliptin by UFLC method

 

 

 

 

 

Evaluation of data:

Table 7 above shows the observation of distinguishing peaks at the LOD and LOQ level.

 

Accuracy:

The exactness of an analytical method is that it is close to the actual value (normal value) of the test results gained by the method.

 

                                   Amount of drug recovered

% Recovery = ––––––––––––––––––––––––––––––––– × 100

                                    Amount of drug added

 

Criteria of approval:

Individual and average retrieval percent should vary between 98.0 and 102.0 percent at each level.

 

Evaluation of data:

The recovery could be achieved within the limit obtained from Table 8. Therefore, the process is accurate.

 

Robustness:

An analytical method's reliability is a measure of its capacity to remain unimpacted by minimal but deliberate methodology differences.

 

Table 9: Robustness for Teneligliptin by UFLC method

 

Evaluation of data:

Table 9 indicates that due to modifications in the chromatographic circumstances mentioned above, no important change has been observed to ensure that the process is robust.

 

Forced degradation studies:

Teneligliptin drug was studied at 50μg/mL. There were stress tests. In 0min., 30 min., 1 hour, 2 hours, 4 hours, 8 hours, 6 hours and 32 hours, bulk drug, the use of 0.1 M Hydrochloric acid, and 0.1M Sodium hydroxide in the drug solution induce an acidic pressure. The specific stress tests also included 2.0ml of 0.1 M of Sodium hydroxide and 2ml of 0.1 M Hydrochloric acid neutralization. For oxidation tests in bulk pharmaceutical items, an application of 2.0ml (3% H2O2), thermal testing heating trials (600C), and UV work (450C) respectively are carried out. Various volumetric flasks (10ml) are loaded with full specimens and dissolved in HPLC grade MeOH. The product was subsequently stored in MeOH for analysis and included in chromatograms. The growth of the degradable material was confirmed by the fact that the chromatogram for all these stability studies remained under ordinary, unstressed conditions.^10-13

 

Acid Stress:

Add 2ml 0.1N Hydrochloric acid to 2ml sample for 5 minutes and add 2.0ml of 0.1N Sodium hydroxide and inject the sample and run it for 32 hours at an interval of 30 minutes, 1 hr., 2 hrs.

 

 

Figure 6: Chromatogram for Acid Stress of Teneligliptin

 

Base Stress:

Add 2 ml of 0.1N Sodium hydroxide for 2 ml of sample for 5 min, then add 2 ml of 0.1N hydrochloric acid and inject the sample.

 

 

Figure 7: Chromatogram for Basic Stress of Teneligliptin

 

Peroxide Stress:

Add 1 ml of 3% peroxide solution for 2 ml of the sample and inject this sample

 

Figure 8: Chromatogram for Peroxide Stress of Teneligliptin

Heat Stress:

Add 2 ml of sample and heat at 80 c for 1 hr.

 

Figure 9: Chromatogram for Heat stress of Teneligliptin

 

Photolytic Stress:

Add a sample of 2 ml and position 1 hr in a UV room. UV-Lamp 450Inject the sample, respectively.

 

Figure 10: Chromatogram for Photolytic stress of Teneligliptin

 

TABLE 10: recovery studies results of Teneligliptin after the stress conditions (% recovery of the drug)

 

CONCLUSION:

A novel, simple, quick, and cost-effective RP-UFLC method was successfully developed in order to estimate Teneligliptin in its simple form. The proposed technique has been optimized and validated for the various experimental parameters. The impact of mobile phase pH, mobile phase ratio, and flow rate on Teneligliptin's evaluation was assessed. In less than 10 minutes, all the analytes were well solved and divided. The method used to determine the content of the Teneligliptin in routine and stabilization samples may conveniently be used by the quality control outfits. In pharmaceutical preparations, this method could be used to analyze medicines and laboratory routine evaluations. Overall, the proposed method provides excellent precision for the determination of Teneligliptin, precision, selectivity, and reproductivity.

 

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Accepted on 24.02.2020           © RJPT All right reserved

Research J. Pharm. and Tech 2020; 13(9):4035-4040.