Development and Validation of UV Spectrophotometric Method of ...

Table No.1 - Calibration Curve parameter.
S. No. Concentration
( µg/ml)
Absorbance ±
SD
1 4 0.1965 ± 0.03 1.7
2 8 0.3349 ± 0.037 1.1
3 12 0.4708 ± 0.037 1.7
4 16 0.6114 ± 0.035 1.5
5 20 0.745 ± 0.023 1.3
6 24 0.889 ± 0.057 2.6
7 28 1.0288 ± 0.037 3.6
Asian J. Research Chem. 2(2): April.-June, 2009
,
%Relative
standard
deviation
Figure No.1 - Chemical structure of cefuroxime axetil 1 .
Figure No.2 - Determination of max of cefuroxime axetil
by UV scanning.
Preparation of standard stock solution:
Standard drug solution of cefuroxime axetil was prepared
by dissolving 10mg cefuroxime axetil in 20ml methanol
and transfered it to 100ml volumetric flask and volume
was made upto mark with phosphate buffer pH 7.0 to
obtain stock solution of 100µg/ml concentration. For
obtaining clear solution, solution was ultrasonicated.
Preparation of calibration curve:
Calibration curve was prepared in 2:8 (methanol: 7.0 pH
phosphate buffer) at max 281 nm using Perkin Elmer UVvisible
spectrophotometer. For this stock solution of 100
µg/ml was prepared. Serial dilution of 4, 8, 12, 16, 20, 24
and 28 µg/ml were prepared and absorbance was taken at
max 281 nm. Averages of such 8 sets of values were
taken for standard calibration curve, and solutions were
scanned in the range of 200-400 nm against blank. The
calibration curve was plotted. The optical characteristics
are summarized in Table no.1
Preparation of sample solution:
Ten tablets were weighed and powdered. The amount of
tablet powder equivalent to 10 mg of cefuroxime axetil
was weighed accurately and transfer to 20 ml methanol
and kept for 15 min with frequent shaking and volume
was made up to 100 ml mark with phosphate buffer pH
7.0.The solution was then filtered through whatmann
filter paper # 41. This filtrate was diluted suitably with
solvent (20% v/v methanol in phosphate buffer pH 7.0) to
get the solution of 16µg/ml concentration .The
absorbance was measured against blank. The drug content
of the preparation was calculated using standard
calibration curve. Amount of drug estimated by this
method is given in Table no.3
RESULT AND DISCUSSION:
Precision:
Assay of method precision (intra-day precision) was
evaluated by carrying out three independent assays of test
samples of cefuroxime axetil. The intermediate precision
(inter-day precision) of the method was also evaluated
using two different analysts, systems and different days in
the same laboratory. The relative standard deviation
(RSD) and assay values obtained by two analysts were
0.36, 99.50 and 0.31, 99.60 respectively (Table no.4).
Accuracy (Recovery Test):
Accuracy of the method was studied by recovery
experiments. The recovery experiments were performed by
adding known amounts to tablet. The recovery was
performed at three levels, 80,100and 120% of cefuroxime
axetil standard concentration. The recovery samples were
prepared in afore mentioned procedure. Three samples were
prepared for each recovery level. The solutions were then
analyzed, and the percentage recoveries were calculated
from the calibration curve. The recovery values for
cefuroxime axetil ranged from 99.97 ± 0.3969 (Table no.3).
Linearity:
The linearity of the response of the drug was verified at 2
to 40 g/ml concentrations,but linearity was found to be
between 4-28 g/ml concentration. The calibration graphs
were obtained by plotting the absorbance versus the
concentration data and were treated by linear regression
analysis (Table no.2). The equation of the calibration
curve for cefuroxime axetil obtained y = 0.0346x +
0.0566, the calibration curve was found to be linear in the
aforementioned concentrations. The correlation
coefficient (r 2 ) of determination was 0.9999.
Limit of Detection (LOD) and Limit of Quantification
(LOQ):
The LOD and LOQ of Cefuroxime axetil were
determined by using standard deviation of the response
and slope approach as defined in International Conference
on Harmonization (ICH) guidelines 7 .The LOD and LOQ
was found to be as in table no.2.
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