Development and Validation of UV Spectrophotometric Method of ...

Asian J. Research Chem. 2(2): April.-June, 2009
,
ISSN 0974-4169
www.ajrconline.org
RESEARCH ARTICLE
Development and Validation of UV Spectrophotometric Method of Cefuroxime
Axetil in Bulk and Pharmaceutical Formulation.
Santosh Shelke* 1 , Santosh Dongre 1 , Amit Rathi 2 , Dinesh Dhamecha 2 , Saifee Maria 2 and Mohd
Hassan G Dehghan 2
1
Yash Institute of Pharmacy, South city, Bajaj Nagar, Post Box no. 968,Waluj,Aurangabad-431134
2
Y.B.Chavan College of Pharmacy, Maulana Azad Education Trust, Dr. Rafiq Zakaria Marg,Aurangabad-431001
*Corresponding Author E-mail: santoshshelke24@yahoo.com
ABSTRACT
A simple, accurate, cost effective and reproducible spectrophotometric method has been developed for the estimation
of cefuroxime axetil in bulk and pharmaceutical dosage form. UV spectrophotometric method, which is based on
measurement of absorption at maximum wavelength 281nm.The percentage recovery of cefuroxime axetil ranged
from (99.97 ± 0.3969) in pharmaceutical dosage form. The developed method was validated with respect to linearity,
accuracy (recovery), precision and specificity. Beers law was obeyed in the concentration range of 4-28µg/ml having
line equation y = 0.0346x + 0.0566 with correlation coefficient of 0.9999. Results of the analysis were validated
statistically and by recovery study.
KEYWORDS: UV spectrophotometry, cefuroxime axetil
INTRODUCTION:
Chemically cefuroxime axetil is (RS)-1-hydroxyethyl (6R,
7R)-7-[2-(2-furyl) glyoxyl-amido]-3-(hydroxymethyl)-8-
oxo-5-thia-1-azabicyclo[4.2.0]-oct-2-ene-2-carboxylate,
72-(Z)-(O-methyl-oxime), 1-acetate 3-carbamate (figure
no.1) ,is semisynthetic ,broad spectrum cephalosporin
antibiotic for oral administration. It is indicated in case of
pharyngitis / tonsillitis, acute bacterial otitis media, acute
bacterial maxillary sinusitis, chronic bronchitis, urinary
tract infection, lyme disease 1 .
Analysis is an important component in the formulation
development of any drug molecule. A suitable and
validated method has to be available for the analysis of
drug(s) in the bulk, in drug delivery systems, from release
dissolution studies and in biological samples. If a suitable
method, for specific need, is not available then it becomes
essential to develop a simple, sensitive, accurate, precise,
reproducible method for the estimation of drug samples
The estimation of cefuroxime axetil by mercurimetric
method 2 , high performance liquid chromatography
[HPLC] 3 , high performance thin layer chromatography
[HPTLC] 4 and spectroflurimetric method 5 is reported in
literature.
Received on 25.04.2009 Modified on 30.04.2009
Accepted on 21.06.2009 © AJRC All right reserved
Asian J. Research Chem. 2(2): April.-June, 2009 page 222-224
Although simultaneous UV estimation of cefuroxime
axetil and probenecid has been reported by Chaudari et.
al. 6 , but single estimation of this drug has not been
reported in bulk and in pharmaceutical formulation. Thus
the present study was undertaken to develop and validate
a simple, sensitive, accurate, precise, and reproducible
U.V method for cefuroxime axetil.
MATERIALS AND METHOD:
Instrument and materials:
Instrument used were Perkin Elmer double beam
UV/Visible Spectrophotometer and schimadzu AX200
analytical balance. Cefuroxime axetil pure drug was
obtain from Aurobindo Pharma LTD. Hyderabad as gift
sample with 99.99% w/w assay value and was used
without further purification. All chemicals and reagents
used were of analytical grade.
Selection of media:
Main criteria for media selection are solubility and
stability, i.e. drug should be soluble as well as stable for
sufficient time in selected media. Though the official
reported media for this drug is 0.07 M pH 7.0 phosphate
buffer but in provided experimental condition drug is not
giving clear solution in pH 7.0 phosphate buffer, same
time drug is soluble in methanol, so for present work
combination of methanol and pH 7.0 phosphate buffer in
ratio of 2: 8 has been selected as analytical media.
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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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Asian J. Research Chem. 2(2): April.-June, 2009
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Table.2 - Validation parameters
Sr. No. Parameter Result
1. Absorption maxima(nm) 281
2. Linearity Range (µg/ml) 4-28
3 Standard Regression Equation y = 0.0346x + 0.0566
4 Correlation Coefficient (r2 ) 0.9999
5 Molar absorptivity 49647.9
6 A (1%, 1cm) 398.453
7 Accuracy (% recovery ±SD) 99.97 ± 0.3969
8 Precision 99.50% (Intra-day precision) and 99.60%(Inter-day precision)
9 Specificity A 16 µg/ml solution of candidate drug in 7.0-pH phosphate buffer at UV
detection λ of 281 nm will show an absorbance value of 0.6114 ± 0.035
10 Sandell’s Sensitivity
0.02534
(µg/cm 2 /0.001 absorbance unit) 8
11 LOD (µg/ml) 1.3669
12 LOQ (µg/ml) 4.1421
Table No.3 - Determination of Accuracy by percentage recovery method
Tablet amount Level of Amount Drug
Ingredient (µg/ml) addition (%) added (mg) (µg/ml)
found
% Recovery
cefuroxime 16 80 12.8 28.6918 99.6243
axetil* 16 100 16 31.9672 99.8976
16 120 19.2 35.3430 100.4064
*cefuroxime axetil having brand name Kefstar-250
Average %
recovery
99.97 ± 0.3969
Table No.4 - Determination of Precision
Sample Assay of cefuroxime axetil as % of labeled
number
amount
Analyst-I
(Intra-day precision)
1 99.42 99.70
2 99.63 99.23
3 99.58 99.57
4 99.10 99.88
5 100.12 99.98
6 99.20 99.25
Mean 99.50 99.60
RSD 0.36 0.31
Analyst-II
(Inter-day precision)
Determination of Active Ingredients in Tablets:
The validated method was applied to the determination of
cefuroxime axetil in Tablets. Six tablets were assayed and
the results are shown in (Table no. 3) indicating that the
amount of drug in tablet samples met with requirements
(99–102% of the label claim).
REFERENCES:
1. Dollery C. Therapeutic drugs .second edn. Churchill
Livingstone, London UK.1999;C135-C140.
2. Pospisilova B and Kubes J. Mercurimetric determination of
cephalosporin antibiotics. Pharmazie.1998; 43(4):246-248.
3. US Pharmacopoeia (XXIV) and National Formulary
(XIX). Asian Edn., US Pharmacopoeial
convection,Inc.Rockville,MD.2000;335.
4. Shaha NJ, Shaha SK, Patel VF, Patel NM. Development
and validation of HPTLC method for the estimation of
cefuroxime axetil. Ind J Pharm Sci.2007;69(1):140-142.
5. Murillo JA, Lemus JM, Garcia LF. Specctroflurometric
analysis of cefuroxime axetil in pharmaceutical dosage
form. J Pharm Biomed Analysis.1994;12(7):875-881.
6. Chaudari SV, Karnik A, Adhikary A, Tandale RS, Vavia
PR. Simultanious UV spectrophotometric method for
estimation of cefuroxime axetil and probenecid from solid
dosage form. Ind J Pharm Sci.2006;68(1):59-63
7. Validation of Analytical Procedures, Methodoly, ICH
Harmonised tripartite guidelines; 1996:1.
8. Khopkar SM, Basic concepts of analytical chemistry. New
age international publisher. 2008; 3 rd ed: 277-278.
CONCLUSIONS:
The developed method was found to be simple, sensitive,
accurate, precise, reproducible, and can be used for
routine quality control analysis of cefuroxime axetil in
bulk and pharmaceutical formulation
ACKNOWLEDGEMENT:
We are thankful to Aurobindo Pharma Ltd.. Hyderabad
for providing the gift sample of cefuroxime axetil.We
would also like to thank Dr.Subhash Devhde Patil
Director Yash Institute of Pharmacy for providing all the
facilities to complete our work successfully.
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