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Method Development and Validation For The Estimation of Metronidazole in Tablet Dosage Form by UV Spectroscopy and Derivative Spectros
Method Development and Validation For The Estimation of Metronidazole in Tablet Dosage Form by UV Spectroscopy and Derivative Spectros
Method Development and Validation For The Estimation of Metronidazole in Tablet Dosage Form by UV Spectroscopy and Derivative Spectros
[Research article]
Method development and validation for the estimation of
metronidazole in tablet dosage form by UV spectroscopy
and derivative spectroscopy
K.Narendra reddy and MK.Ranganath
Department of pharmaceutical analysis, Krupanidhi College of Pharmacy, Chikkabellandur,
Carmelaram, V arthur hobli, Bangalore- 560035.
ABSTRACT
Two simple, precise, rapid, sensitive and accurate spectrophotometric methods have been developed for the
estimation of metronidazole in pure form and in tablet dosage form. Metronidazole has absorbance maxima at
313 nm in methanol: water (80:20) for UV spectroscopy and absorbance minima at 298nm for first order
derivative spectroscopy. The linearity was obtained and obeyed Beer’s law in the concentration range of 4 - 12
μg/ml and correlation coefficients were 0.9995 and 0.9897 for UV spectroscopy and derivative spectroscopy
respectively. The precision values for UV and derivative spectroscopy were found to be 0.35-0.64 and 1.78-
1.92 with mean accuracies 98.60-99.70% and 97.55-103.80% respectively. Limit of detection values for UV
and derivative spectroscopy were found to be 0.23 and 1.01 respectively. Limit of quantification for both UV
and derivative spectroscopy were found 0.69 and 3.06 respectively. The methods were validated as per the
International Conference on Harmonization (ICH) guidelines. The proposed validated methods were
successfully used for the quantitative analysis of API and tablet dosage form.
Keywords: Metronidazole, Beer’s law, derivative spectroscopy, ICH.
228
*Corresponding author: K.Narendra reddy
E-mail address: reddy.narendra143@gmail.com
K. Narendra reddy, et al / Int. J. of Pharmacy and Analytical Research Vol-3(2) 2014 [228-232]
Structure of metronidazole
313nm
Beer’s law concentration range The absorbance of these solutions were measured
Aliquots of standard solution 2-3ml were against blank at the wavelengths of 313 nm and
transferred into a series of 10ml volumetric flasks 298 nm for UV spectroscopy and derivative
and makeup to volume with solvent up to the mark. spectroscopy respectively. The absorbance values
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K.Narendra reddy, et al / Int. J. of Pharmacy and Analytical Research Vol-3(2) 2014 [228-232]
against the concentrations were plotted in the which is accepted either as a conventional true
calibration curve (Fig.2,3). From the calibration value or an accepted reference value and the value
curve it was found metronidazole obeys Beer’s law found.
in the concentration range of 4-12µg/ml. The To study the accuracy of the proposed method,
regression analysis was carried out for the recovery studies were carried out at three different
regression line which estimates degree of linearity. levels (50%,100% and 150%). The results were
Preparation of sample solution represented in table. 01.
Twenty tablets were weighed accurately and
ground into a fine powder. An accurately weighed Method precision
quantity of tablet powder equivalent weight to The precision of an analytical procedure expresses
50mg of metronidazole was transferred to a the closeness of agreement (degree of scatter)
50mlvolumetric flask and dissolved and diluted to between a series of measurements obtained from
the mark with solvent to obtain sample stock multiple sampling of the same homogeneous
solution having concentration of 1000 μg/ml. The sample under the prescribed conditions. The results
solution was filtered through a filter paper were represented in table. 01.
(whatman. 41) to get the clear solution. From this
solution 1ml of solution was pipetted out into a
Limit of Detection (LOD)
50ml volumetric flask and diluted to the mark with
the solvent. Concentration of the solution was The detection limit of an individual analytical
20µg/ml. procedure is the lowest amount of analyte in a
sample which can be detected but not necessarily
Validation quantitated as an exact value. The limit of
Both the methods were validated in compliance detection (LOD) of the drug was derived by
with ICH guidelines. calculating the signal-to-noise ratio (S/N, i.e., 3.3
for LOQ) using the following equation designated
by International Conference on Harmonization
Accuracy (recovery study)
(ICH) guidelines. The results were represented in
The accuracy of an analytical procedure expresses
table. 01.
the closeness of agreement between the value
LOD = 3.3 × σ/S
Where,
σ = the standard deviation of the response and
S= slope of the calibration curve.
LOQ = 10 × σ/S
Where,
σ = the standard deviation of the response and
S= slope of the calibration curve.
Range
Linearity The range of an analytical procedure is the interval
The linearity of an analytical procedure is its ability between the upper and lower concentration
(within a given range) to obtain test results which (amounts) of analyte in the sample (including these
are directly proportional to the concentration concentrations) for which it has been demonstrated
(amount) of analyte in the sample. The results that the analytical procedure has a suitable level of
were represented in table. 01. precision, accuracy and linearity. The results were
represented in table. 01.
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K. Narendra reddy, et al / Int. J. of Pharmacy and Analytical Research Vol-3(2) 2014 [228-232]
1.8
0.6 R² = 0.9995
1.7
1.6
0.5
Absorbance
1.5
1.4
0.4 1.3
1.2
0.3 1.1
1.0
0.2 0.9
0.8
0.1 0.7
0.6
0 0.5
0.4
0 5 10 15 0.3
0.2
0.0 nm
200 210 220 230 240 250 260 270 280 290 300 310 320 330 340 350 360 370 380 390 400
0 0.035
-0.001 0 10 20 30 0.030
0.025
-0.002 0.020
-0.003
Absorbance
0.015
0.010
-0.004 0.005
-0.005 0.000
-0.005
-0.006 -0.010
-0.007 -0.015
y = -0.0004x + 0.0005
-0.008 -0.020
R² = 0.9897 -0.025
-0.009 -0.030
-0.035
-0.040 nm
200 210 220 230 240 250 260 270 280 290 300 310 320 330 340 350 360 370 380 390 400
Fig. 3: Linearity curve for metronidazole UV Fig. 5: First order derivative overlay spectra
first order derivative spectroscopy of metronidazole
Table. 01: Regression analysis data and summary of validation parameters for the proposed UV
spectroscopic method
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K.Narendra reddy, et al / Int. J. of Pharmacy and Analytical Research Vol-3(2) 2014 [228-232]
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