Derivative UV Spectrophotometric Method for Validation of Metformin hydrochloride in Bulk and Pharmaceutical Dosage Form

 

Rajan V. Rele*, Prathamesh P. Tiwatane

Central Research Laboratory, D. G. Ruparel College, Matunga, Mumbai 400016

 

ABSTRACT:

Simple and precise derivative UV spectrophotometric method by first and second order have been developed and validated for the estimation of metformin hydrochloride in bulk and its tablet formulation. The standard and sample solutions of metformin hydrochloride were prepared in distilled water. Metformin hydrochloride was estimated by first and second order at wavelength 225nm and 252nm respectively. Beer’s law was obeyed in the concentration range of 1 to 14μg/ml with coefficient of correlation value 0.9999. These methods were tested and validated for various parameters according to ICH guidelines. The precision expressed as relative standard deviation were 0.6787% for first order and 0.2421% for second order derivative method respectively. The proposed method was successfully applied for the determination of metformin hydrochloride in pharmaceutical formulation. Results of the analysis were validated statistically and were found to be satisfactory. The proposed method is simple, easy to apply, low-cost and require relatively inexpensive instruments.

 

 

 

INTRODUCTION:

Metformin hydrochloride is a biguanide hypoglycemic agent and non-insulin-dependent diabetes mellitus. Chemically metformin hydrochloride is a 1,1-Dimethyl biguanide hydrochloride. The drug is official in USP¹. Literature survey reveals spectrophotometric and HPLC^2,3 and HPLC^4-16 for assay of drug. This proposed work presents simple, accurate and reproducible UV, area under curve spectrophotometric method for determination of metformin hydrochloride in tablet dosage form.  

 

 

MATERIALS AND METHOD:

Instrument and reagents:

Spectral scan was made on a Shimadzu UV-spectrophotometer, model 1800 (Shimadzu, Japan) with spectral band width of 0.5nm with automatic wavelength corrections by using a pair of 10mm quartz cells.

 

 

All spectral measurements were done by using UV-Probe 2.42 software.

 

Reference standard of metformin hydrochloride was obtained from reputed firm with certificate of analysis.

 

Preparation of standard drug solutions:

100mg standard metformin hydrochloride was weighed accurately and transferred to a 100ml volumetric flask and sonicated with 30ml of distilled water for 15 minutes. The volume was made up to the mark with distilled water to give a stock solution of metformin hydrochloride of concentration 1000μg/ml. From this solution, 10ml of solution was pipetted out and transferred into 100ml volumetric flask. The volume was made up to mark with distilled water to give a working standard solution of concentration 100μg/ml.

 

Estimation from tablets:

Twenty tablets were weighed accurately and average weight of each tablet was determined. Powder equivalent to 10mg of metformin hydrochloride was weighed and transferred in 100ml of volumetric flask. A 30ml of distilled water was added and sonicated for 15minutes and filtered. The filtrate and washing were diluted up to the mark with distilled water to give concentration as 100μg /ml. Such solution was used for analysis.

 

EXPERIMENTAL:

Method: First order derivative method:

For the selection of analytical wavelength, 10μg/ml solution of metformin hydrochloride was scanned in the spectrum mode from 400nm to 200nm by using distilled water as blank. The first order derivative spectrum was obtained by using derivative mode by UV probe 2.42 software. From the spectrum, the amplitude of the derivative spectrum was measured at 225 nm.

 

Method: Second order derivative method:

For the selection of analytical wavelength, 10μg/ml solution of metformin hydrochloride was scanned in the spectrum mode from 400nm to 200nm by using distilled water as blank. The second order derivative spectrum was obtained by using derivative mode by UV probe 2.42 software. From the spectrum, the amplitude of the derivative spectrum was measured at 252nm.

 

Preparation of calibration curves:

Series of solutions containing 0.5–10µg/ml of metformin hydrochloride were used to determine linearity of the proposed method respectively.  Solutions were scanned in the spectrum mode and absorbance spectra were converted to first order derivative spectra [Fig. 1(a), 1(b)]. 

 

Fig. 1(a): Overlay spectra of first order derivative of metformin hydrochloride in the concentration range of 2 – 14 µg/ml.

 

Fig. 1(b): Overlay spectra of second order derivative of metformin hydrochloride in the concentration range of 2 – 14 µg/ ml.

 

After observing the overlain first order derivative spectra of metformin hydrochloride, wave length selected was 225 nm, where metformin hydrochloride showed considerable absorbance. The calibration curves were plotted of dA/dλ against concentrations [Fig. 2 (a)].

 

After observing the overlain second order derivative spectra of metformin hydrochloride wave length selected was 252 nm, metformin hydrochloride showed considerable absorbance. The calibration curves were plotted of dA/ dλ against concentrations [Fig. 2(b)].

 

Fig.2 (a). Calibration curve for metformin hydrochloride by First order derivative method in the concentration range of 2-14 µg/ml.

 

Fig. 2 (b): Calibration curve of metformin hydrochloride by second order derivative method in the concentration range of 2-14 µg/ml.

 

Results of analysis are given in table 1.

Table 1: Values of results of optical and regression of drug

Parameter	First order derivative	Second order derivative
Detection Wavelength (nm)	225	252
Beer Law Limits (µg/ml)	2-14	2-14
Correlation coefficient(r2)	0.9999	0.9987
Regression equation (y=b+ac)
Slope (a)	0.002	0.0005
Intercept (b)	0.0001	0.0002

 

Estimation from tablets:

Twenty tablets were weighed accurately and average weight of each tablet was determined. Powder equivalent to 10 mg of metformin hydrochloride was weighed and transferred in 100 ml of volumetric flask. A 30 ml of 0.1N distilled water was added and sonicated for 15 minutes and filtered. The filtrate and washing were diluted up to the mark with distilled water to give concentration as 100 μg /ml of each drug. Such solution was scanned in the range of 200-400 nm against distilled water as blank. The absorbance spectra were converted to first order and second order derivative spectra. Calculations were done as per the equations. The concentrations of metformin hydrochloride present in tablets were calculated by substituting the values of absorbance in linearity equations.

 

(a) For first order derivative method, Y =   0.002x + 0.0001

(b)  For second order derivative method, Y = 0.0005x + 0.0002

 

Method Validation:

These methods were validated according to ICH guidelines.

 

Accuracy:

To ascertain the accuracy of proposed methods, recovery studies were carried out by standard addition method. Percent recovery for metformin hydrochloride was found as 100.32 % for first order derivative method and 101.022 % for second order derivative respectively. (Table 2(a) and (b)).

 

Table 2 (a):  Statistical evaluation of the data subjected to accuracy for first order derivative method

 

Table 2 (b):  Statistical evaluation of the data subjected to accuracy for second order derivative method 

 

Table 3: Statistical evaluation of the data subjected to method of precision

Sr. No.	Sample No.	% Assay
		First order derivative method	Second order derivative method
1	1	100.3636	100.18
2	2	99.63636	100.36
3	3	98.54545	99.818
4	4	99.63636	99.812
5	5	100.3636	100.181
6	6	100.001	99.814
Mean % assay		99.75773	100.0275
%R.S.D.		0.678759	0.242193

 

Linearity:

The linearity of measurement was evaluated by analyzing different concentration of the standard solutions of metformin hydrochloride in the range of 2-14 µg/ml for first order and second order derivative respectively.

Precision:

The method precision was established by carrying out the analysis of tablets powder blend containing metformin hydrochloride. The assay was carried out for the drugs by using proposed analytical method in six replicates. The values of relative standard deviation were well 0.6787% and 0.2421% for metformin hydrochloride respectively indicating the sample repeatability of the methods.  The results obtained are tabulated in table 3.

 

Table 4: Summary of validation parameter for intra-day and inter-day

Sr. No.	Parameters	First order derivative	Second order derivative
1	Intra-day precision  (N=3) amount found ± % R.S.D.	98.61%   0.6875	99.41%    0.2378
2	Inter-day precision  (N=3) amount found ± % R.S.D.	98.25   0.2457	98.31%   0.2975

Intra-day precision was estimated by assaying tablets powder blend containing metformin hydrochloride. The assay was carried out for the drugs by using proposed analytical method in six replicates. The results were average for statistical evaluation.

 

Inter-day precision was estimated by assaying tablets powder blend containing metformin hydrochloride for three consecutive days (i.e. 1st, 3rd and 5th days). The statistical validation data for intra and inter day precision is summarized in table 4.

 

Both intra- day and inter-day precision variation found to be less in % RSD values. It indicates high degree of precision of the method.

 

RESULT AND DISCUSSION:

The developed first and second order derivative spectrophotometric methods for determination of metformin hydrochloride in tablet formulation was found to be simple and convenient for the routine analysis of drug. The proposed method is accurate, precise and reproducible. It is confirmed from validation data as given in tables 1 to 4. The % RSD was found to be less than 1, which indicates validity of method. Linearity was observed by linear regression equation method for metformin hydrochloride in different concentration range. The correlation coefficient of these drugs was found to be close to 1.00, indicating good linearity figure 2 (a) and 2 (b).

 

The assay results obtained by proposed method is shown in table 2 are in good agreement. Hence proposed method can be used for routine analysis pharmaceutical dosage form. Methods are simple, accurate, precise, reliable, rapid, sensitive, reproducible and economical. It is validate as per ICH guidelines.    

 

The proposed methods are simple, precise, accurate and rapid for the determination of metformin hydrochloride pharmaceutical dosage form. The methods do not require any ratio of first and second order derivative. The amplitude of first and second order derivative can be directly used to assay of formulation. This method can be adopted as an alternative to the existing methods. It can be easily and conveniently adopted for routine quality control analysis.

 

ACKNOWLEDGMENT:

Authors express sincere thanks to the Principal, Dr. Tushar M. Desai of D. G. Ruparel College, Mumbai.

 

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Received on 19.08.2019         Modified on 07.09.2019

Accepted on 14.10.2019         © AJRC All right reserved