Stress degradation studies on Telmisartan and development of a validated method by UV spectrophotometry in bulk and pharmaceutical dosage forms.

AIM AND
BACKGROUND: To develop and validate a simple, precise, accurate, and stability indicating a UV-method for estimation of Telmisartan (TELM). UV, HPLC, HPTLC, and many more experiments were carried out by taking single drug and also by combining with other drugs. However, Such type of studies was not reported.
MATERIALS AND METHODS: In both methods, TELM has the absorbance maxima at 296 nm. Method A involves method development and validation and Method B involves forced degradation study. In these methods, methanol was used as a solvent. Linearity was observed in the concentration range of 4-16 μg/ml. Validation experiments were performed to demonstrate system suitability, specificity, precision, linearity, accuracy, robustness, LOD, and LOQ as per International Conference on Harmonization guidelines. Furthermore stability studies of TELM were carried out under acidic, alkali, neutral, oxidation, photolytic, and thermal degradation as per stability indicating assay methods.
RESULTS: The results of analysis have been validated, and recovery studies were carried out using a standard addition method by adding specific drug amount (80%, 100%, and 120%) and show recovery studies in the range (99.26-101.26)%.
CONCLUSION: The proposed method can be successfully applied for method development, validation, and stability study of TELM.

INTRODUCTION

Telmisartan (TELM) is chemically known as 4′-[(1,4′-dimethyl-2-propyl [2,6′-bi-1H-benzimidazol]-1′-yl) methyl] [1,1′-biphenyl]-2-carboxylic acid.[1-3] TELM is an antihypertensive drug (angiotensin-II receptor antagonist) and prevents binding of Angiotensin-II to the AT1 receptor. This is used for treatment of hypertension and diabetic nephropathy with an elevated serum creatinine and proteinuria (>300 mg/day) in patients with type-2 diabetes and hypertension.[45]

The UV spectrophotometric method was developed and validated as per International Conference on Harmonization (ICH) guidelines.[6] Spectrophotometry is generally preferred especially by small-scale industries as the cost of the equipment is less and the maintenance problems are minimal. The method of analysis is based on measuring the absorption of a monochromatic light by colorless compounds in the near ultraviolet path of spectrum (200–380 nm). TELM is official in British Pharmacopoeia (2009) and Indian Pharmacopoeia (2010), which recommends UV spectrophotometry for its analysis. The active pharmaceutical ingredient is subjected to a number of forced degradation conditions to include acidic, basic, and oxidative conditions. Forced degradation should be one of the activities performed early in the development process to ensure that the method is discriminating before a lot of time, effort and money have been expended. Depending on the TELM not every stress agent may effect a degradation, but each agent has to be evaluated to determine whether degradation results.[7]

Literature review show that there are developed methods including UV,[8-14] visible spectrophotometric,[15] HPTLC,[1617] HPLC,[18] and UPLC[19] methods for estimation of TELM, with many drugs combined other than the single drug with stability study. However, no method has been reported till date for the method validation and stability study of TELM using the UV spectrophotometric method. This work deals with the method development, validation, and stability study of TELM by various UV spectrophotometric methods.

MATERIALS AND METHODS

Apparatus

Digital balance: Acculab (ALC 210.4)

Sonicator: Eneritech (Ultra Sonicator)

Photo stability chamber: Thermolab

Hot air oven: Hicon

A double beam UV-Visible spectrophotometer (Shimadzu-1800) with UV probe 2.31 software.

Material

Pure samples: TELM was kindly supplied by Zydus Cadila Healthcare Ltd., Ahmedabad, Gujarat, India.

Marketed formulation

TELSAR™ 40 (EmcureR Pharmaceutical Ltd., Hinjwadi, India) was purchased from an open market for this study which contains TELM IP 40 mg.

Reagent and chemical

Methanol was used as a solvent which was procured from Finar Chemicals Ltd., Ahmadabad, India. HCl, NaOH, and H2O2 are of analytical grade. Double distilled water was used throughout the analysis.

Stock solution

Accurately weighed 10 mg of TELM was transferred to a 100 ml volumetric flask, 50 ml of methanol was added and allowed to sonicate for 15 min and finally volume was made up to the mark by methanol. Standard stock solution of TELM (100 μg/ml) was prepared.

Absorption maxima method

For the selection of analytical wavelength, 10 μg/ml solution of TELM was prepared by appropriate dilution of standard stock solution and scanned in the spectrum mode from 400 to 200 nm. From the spectra of drug [Figure 1], λmax of TELM 296 nm was selected for the analysis.

Figure 1

UV spectrum showing Telmisartan at 296 nm.

Preparation of working standard solutions

From the stock solution of 100 μg/ml, working standard solutions of drug was prepared by appropriate dilution and were scanned in the entire UV range to determine the λmax. Standard solutions were prepared having concentration of 4, 6, 8, 10, 12, 14, and 16 μg/ml of TELM to construct Beer's law plot for TELM. The absorbance of each solution was measured at 296 nm against methanol as a blank [Table 1 and Figure 1].

Table 1

Linearity

Preparation of calibration curve

The standard graph/calibration curve for TELM was plotted by taking concentration of drug on the X-axis and absorbance on the Y-axis [Figure 2].

Figure 2

Linearity graph of Telmisartan

Preparation of sample solution

The proposed method was applied to analyze commercially available TELM tablet. Ten tablets were weighed and powdered. The amount of tablet powder equivalent to 10 mg of TELM was weighed accurately and transferred to a 100 ml volumetric flask. Then 50 ml of methanol was added and allowed to sonicate for 15–20 min and the volume was made up to mark with methanol. The solution was then filtered through a Whattman filter paper. This filtrate was diluted suitably with a solvent to get the solution of 10 μg/ml concentration. The absorbance was measured against blank solution. The drug content in each tablet was estimated by using the standard graph.

METHOD A: METHOD VALIDATION

Linearity

The linearity of measurement was evaluated by analyzing different concentrations of the standard solution of TELM. Beer–Lambert's concentration range was found to be 4–16 μg/ml for below methods [Table 2].

Table 2

Optical characteristics

Specificity

Specificity is a procedure to detect quantitatively the analyte in presence of component that may be expected to be present in the sample matrix. Commonly used excipients in tablet preparation were spiked in a pre weight quantity of drug and then absorbance was measured and calculation done to determine quantity of drugs [Figure 3].

Figure 3

UV spectrum showing Telmisartan standard, test sample, and placebo

Accuracy

To ascertain the accuracy of the proposed methods, recovery studies were carried out by the standard addition method at three different levels (80%, 100%, and 120%) of the bulk sample of TELM to the previously analyzed solution of formulation of concentration of 40 μg/ml. The percentage recovery for TELM by all the methods was found to be in the range of (99.26–101.26)% as shown in Table 3.

Table 3

Accuracy

Precision

The precision of the method was achieved by replicate (n = 6) analysis of tablet preparations [Table 4]. The precision was also studied in terms of intra-day changes in absorbance of drug solution on the same day and inter-day changes on three different days over a period of 1 week. The intra-day and inter-day variations were calculated in terms of percentage relative standard deviation and the results are given in Table 4 (a and b).

Table 4

Precision

Robustness

This procedure was carried out by changing the λmax ± 5%. Then, the results are given in Table 5.

Table 5

Robustness

METHOD B: METHOD FORCED DEGRADATION STUDY

Acid degradation

First 0.1 N HCl was taken in a 10 ml volumetric flask and then accurately weighed 10 mg bulk drug was dissolved in it. To make soluble the drug, few drops of methanol was added and then the volume is made by 0.1 N HCl. Then, this solution was refluxed for 5 h at 70 °C in water bath. Initially at 0 h take 0.1 ml of this solution and the volume was made up to 10 ml with methanol and then, withdrawing the specific amount of solution in every hour. After this, the absorbance was measured by scanning the prepared solution of required concentration in a UV spectrophotometer [Table 6 and Figure 4].

Table 6

Acid degradation

Figure 4

Acid degradation spectrum at 0 h and after 5 h

Alkali degradation

First 0.1 N NaOH solution was prepared. Accurately weighed 10 mg bulk drug was taken in a 10 ml volumetric flask. Then, the volume was made with 0.1 N NaOH. Then this solution was refluxed for 5 h at 70 °C in a water bath. Initially at 0 h take 0.1 ml of this solution and the volume was made up to 10 ml with methanol. The absorbance was measured in every hour by withdrawing the required amount of the sample. Then, scanning was performed with a UV spectrophotometer [Table 7 and Figure 5].

Table 7

Alkali degradation

Figure 5

Alkali degradation spectrum at 0 h and after 5 h

Neutral degradation

Accurately weighed 10 mg of bulk drug was taken in a 10 ml volumetric flask. Then, little amount of methanol was added to dissolve the drug. The volume was adjusted up to the mark with double distilled water. Then, that solution was refluxed for 5 h at 70 °C in a water bath. Initially at 0 h take 0.1 ml of this solution and the volume was made up to 10 ml with methanol. The absorbance was measured at one-hour interval by withdrawing the required amount of sample solution. Then, scanning was performed with a UV-spectrophotometer [Table 8 and Figure 6].

Table 8

Neutral degradation

Figure 6

Neutral degradation spectrum at 0 h and after 5 h

Thermal degradation

A specific amount of bulk drug was taken in a cleaned Petridis and dried, then the Petridis along with bulk drug was placed into the oven at 70 °C for 5 h, at every hour 10 mg of bulk drug was taken from the Petridis, and 1000 ppm solution with methanol was prepared. After this, the required concentration was made and the absorbance measured in the UV spectrophotometer and percentage of degradation was calculated [Table 9 and Figure 7].

Table 9

Thermal degradation

Figure 7

Thermolytic degradation spectrum at 0 h and after 5 h

Photolytic degradation

Accurately weighed 10 mg of bulk drug was taken in a 10 ml volumetric flask. The volume was adjusted up to the mark with methanol. Then, that solution was placed into the photostability chamber for 5 h Initially at 0 h take 0.1 ml of this solution and the volume was made up to 10 ml with methanol. The absorbance was measured at one-hour interval by withdrawing the required amount of sample solution. Then, scanning was performed with a UV-spectrophotometer [Table 10 and Figure 8].

Table 10

Photolytic degradation

Figure 8

Photolytic degradation spectrum at 0 h and after 5 h.

Oxidation with H2O2

Ten milligrams of bulk drug was weighed accurately, 2–3 drops of methanol were added to make the drug soluble. Then the volume was made up by 3% H2O2 and placed it in a cupboard for 5 h. At one-hour interval specified amount the sample was taken and the required concentration was prepared. It was scanned in a UV spectrophotometer [Table 11 and Figure 9].

Table 11

Oxidative degradation

Figure 9

Oxidative degradation spectrum at 0 h and after 5 h

RESULTS AND DISCUSSION

The main objective of this work was to develop and validate the stability indicating a UV method for TELM in the pharmaceutical dosage form. The absorbance maxima of TELM at 296 nm and linearity were observed in the concentration range of 4–16 μg/ml for all validated methods. A percent assay for TELM by above validated methods was found in the range of 98.8–99.60%. Standard deviation was found to be less than ±2.0 and the coefficient of variance was found to be less than ±1.0 indicating the precision of the methods. Accuracy of proposed methods was ascertained by recovery studies, and the results were expressed as % recovery. Percent recovery for TELM was found in the range of 99.26–101.26%. Values of standard deviation and coefficient of variation were satisfactorily low indicating the accuracy of all the methods.

For forced degradation studies, the absorbances in all stressed conditions were decreased for repeated times and percent degradation was found out. Two overly spectra indicate that the degradation shows at initial 0 h and total degradation after 5 h. Therefore, the drug TELM undergoes degradation in all stressed conditions. TELM gives more absorbance with alkali medium as compared to other medium, alkali degradation, chances of generating degradation with 0.1 N sodium hydroxide solution drug is degraded in specific time interval as compared to original drug. Based on the results obtained, it is found that the proposed methods are accurate, precise, reproducible, and economical and can be employed for routine quality control of TELM in its pharmaceutical dosage form [Table 12].

Table 12

Forced degradation study results for Telmisartan after 5 h

CONCLUSION

The proposed method is specific in estimating the commercial formulation without interference of excipients and the other additives. Hence, this method can be used for routine determination of TELM in the bulk sample and pharmaceutical formulation. The proposed method for stability study shows that there is appreciable degradation of TELM found in stress conditions. A new simple analytical method has been developed to apply for the evaluation of the stability of TELM to quantify TELM and its degradation products in a solid premix dosage forms.