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Colloids and Surfaces B: Biointerfaces 87 (2011) 423– 426

Contents lists available at ScienceDirect

Colloids and Surfaces B: Biointerfaces

jou rn al h om epage: www.elsev ier .com/ locate /co lsur fb

oltammetric determination of cefpirome at multiwalled carbon nanotubeodified glassy carbon sensor based electrode in bulk form and pharmaceutical

ormulation

ajeev Jain ∗, Vikaschool of Studies in Chemistry, Jiwaji University, Gwalior 474011, M.P., India

r t i c l e i n f o

rticle history:eceived 18 April 2011eceived in revised form 27 May 2011ccepted 1 June 2011vailable online 12 June 2011

eywords:

a b s t r a c t

A new, simple and low cost voltammetric method for the determination of cefpirome in pharmaceuticalpreparations has been developed using multiwalled carbon nanotube modified glassy carbon electrode(MWCNT), which showed stable response with enhanced selectivity and sensitivity over the bare glassycarbon electrode. A multiwalled carbon nanotube (MWCNT) modified glassy carbon electrode (GCE) isused for the simultaneous determination of cefpirome by differential pulse voltammetry and squarewave voltammetry. Results indicated that cathodic peak of cefpirome is greatly improved at MWCNT

efpirome (CEF)ulti-walled carbon nanotube modified

lassy carbon electrode (MWCNT/GC)harmaceutical formulationensor

modified GC electrode as compared with the bare GC electrode showing excellent electrocatalytic activ-ity towards cefpirome reduction. Linear calibration curves are obtained over the concentration range100–600 �g mL−1 in Britton Robinson buffer at pH 4.51 with limit of detection (LOD) and limit of quan-tification (LOQ) are 0.647 �g mL−1 and 2.159 �g mL−1 using SWV and 5.540 �g mL−1 and 18.489 �g mL−1

using DPV, respectively. The described method is rapid and can be successfully applied for the determi-nation of cefpirome in bulk form and pharmaceutical formulations.

. Introduction

Cefpirome is a new C-3′ quaternary ammonium cephalosporinA) has been classified as a fourth generation cephalosporin. Its highly active against both gram negative organisms includingseudomonas aeruginosa and gram positive organisms includingtaphylococci. Cefpirome could be effectively used for the treatmentf upper and lower urinary tract; lower respiratory tract, skin andoft tissue infections [1,2].

There are few analytical methods for the identification and

uantification of the cefpirome and their metabolites in bulk form,harmaceutical formulation and biological fluids. These meth-ds include HPLC [3], UV-spectrometry and potentiometry [4,5].

∗ Corresponding author. Tel.: +91 751 2442766; fax: +91 751 2346209.E-mail address: rajeevjain54@yahoo.co.in (R. Jain).

927-7765/$ – see front matter © 2011 Elsevier B.V. All rights reserved.oi:10.1016/j.colsurfb.2011.06.001

© 2011 Elsevier B.V. All rights reserved.

Although spectroscopic and chromatographic methods are widelyused for the analysis of various pharmaceutical drugs, most ofthese methods require separation and pretreatment steps. Electro-chemistry has many advantages making it an appealing choice forpharmaceutical analysis. Electrochemical techniques have excel-lent detection limits with a wide dynamic range and are likelyto get preference when low analyte concentrations, small samplevolumes or complex sample is to be analyzed.

Electrode surface modification is a field of paramount impor-tance in the modern electrochemistry especially due to the variousapplication possibilities of modified electrodes [6–25]. The interestin developing electrochemical-sensing devices for use in clinicalassays is growing rapidly. MWCNTs are now used extensively inthe fabrication of novel nanostructure electrochemical sensors.MWCNT-modified electrodes have many advantages over otherforms of carbon electrodes due to their small size, high electricaland thermal conductivity, high chemical stability, high mechani-cal strength and high specific surface area which make them verypromising candidates in a wide range of applications. As MWCNTshave the ability to promote electron-transfer reactions and theypossess a high electrochemically accessible surface area they canbe used as a support material for various catalysts.

In the present work, a MWCNTs film-coated GCE has beendeveloped and tested to take advantage of the convenience ofelectrochemical methods and characteristics of MWCNTs. Theresults showed that the sensitivity for the determination of cef-

4 ces B: Biointerfaces 87 (2011) 423– 426

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24 R. Jain, Vikas / Colloids and Surfa

irome using the MWCNTs film coated GCE increased markedly.ome experimental conditions were optimized and a voltammetricethod for determination of cefpirome in bulk form and phar-aceutical formulation in the presence of surfactant without any

ime-consuming extraction or separation steps prior to analysis haseen developed.

. Experimental

.1. Reagents and materials

Cefpirome was generously provided by Lupin Pharmaceuti-al Company, Mumbai, India. Injection vials containing cefpiromeCef – 4) labeled 1.0 g cefpirome were obtained from commer-ial sources. Multiwalled carbon nanotubes with a 95% purity,.d. = 10–20 nm, i.d. = 5–10 nm and 0.5–50 �m tube length werebtained from Aldrich (USA). For the preparation of standardefpirome stock solution (10 mg mL−1), 500 mg cefpirome wasccurately weighed, dissolved in 1.0% cetyltrimethylammoniumromide and then adjusted to 50 mL with the same surfactant toive the appropriate concentration. Standard working solutionsere prepared by appropriate dilutions of the stock solution. Brit-

on Robinson buffers in the pH range 2.0–12.0 were prepared inistilled water by adding suitable amounts of 0.4 M NaOH solutionbasic solution) to a stock solution composed of a mixture of 2.14 mLhosphoric acid, 2.472 g boric acid and 2.3 mL of glacial acetic acidacidic solution). The ionic strength was kept constant by adjustingith 1.0 M KCl. All reagents and solvents were of analytical reagent

rade quality and were obtained from Aldrich. The square wave andifferential pulse voltammograms were then recorded. The con-ents of the drug in pharmaceutical preparations were determinedsing a calibration curve.

.2. Instrumentation

The voltammetric experiments were performed using-autolab type III Potentiostat/Galvanostat (Utrecht, Theetherlands) PGSTAT, Metohm 663 VA stand as electrochem-

cal cell, fitted with a PC provided the appropriate GPES 4.2oftware. A three electrode system composed of modified glassyarbon as working, Ag/AgCl as reference and Pt wire as auxiliarylectrode was used. Coulometric experiments were performed inhe potentiostatic mode using Pt foil with large surface area asorking electrode and Pt wire as counter electrode. All solutions

xamined by electrochemical techniques were purged for 5 minith purified nitrogen gas. All pH-metric measurements wereade on a Decible DB-1011 digital pH meter fitted with a glass

lectrode and a saturated calomel electrode as reference.

.3. Preparation of MWCNT modified glassy carbon electrode

The GCE was polished on a polishing micro-cloth with 0.5 �Mlumina powder then rinsed and sonicated for 5 min in an ultra-onic bath. The electrode was then transferred to the supportinglectrolyte and potential in the range of −0.1 to −1.2 V was appliedn a regime of cyclic voltammetry for 20 cycles until a stable voltam-

ogram was achieved. The glassy carbon electrode surface wasoated with 10 �L MWCNT suspension solution and the solventas evaporated at room temperature in vacuum drying [26].

.4. Characterization of the modified electrode

Surface morphological studies of the modified electrode werearried out by scanning electron microscopy (SEM) using PhilipsCI quanta 400 instrument. Scanning electron microscopy imagef the MWCNTs on the GCE surface is shown in Fig. 1. It can be seen

Fig. 2. Differential pulse voltammograms of 500 �g mL−1 cefpirome at (a) blank, (b)bare glassy carbon electrode and (c) MWCNT-GCE.

that the MWCNTs were seen in the form of tubes some of whichtwisted together.

3. Results and discussion

The electrochemical behaviour of cefpirome on multiwalledcarbon nanotube modified glassy electrode (MWCNT/GC) was stud-ied by using square wave voltammetry (SWV), differential pulsevoltammetry (DPV) and cyclic voltammetry (CV). In all electro-chemical methods cefpirome gave one well defined reduction peakin cetyltrimethylammonium bromide (CTAB), which is attributedto the reduction of exocyclic –C N– bond.

3.1. Comparison of square wave voltammetric behaviour ofcefpirome at GCE and MWCNT film-coated GCE

On comparing the voltammetric behaviour of cefpirome in GCEand MWCNTs film-coated GCE, it is observed that cefpirome showssubstantial increase in peak current and the limit of detection isalso found to be lower in MWCNTs film-coated GCE (Fig. 2).

3.2. Effect of pH and scan rate

Differential pulse (DP) and square wave (SW) voltammetry stud-ies were carried out to investigate the influence of solution pH

es B: Biointerfaces 87 (2011) 423– 426 425

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Table 1Analytical parameters for voltammetric determination of cefpirome using SWV andDPV modes.

Method SWV DPV

Conc. range (�g mL−1) 200–600 200–600Regression equations ip = 148.2 − 19.12 ip = 15.16 − 1.318r2 0.991 0.988Sa 0.0032 0.0028LOD (�g mL−1) 0.647 5.540LOQ (�g mL−1) 2.159 18.489

Sa Standard deviation of the intercept of regression line.

Fig. 3. The dependence of the differential pulse voltammetric peak current of cef-pirome of different concentrations in 1.0% CTAB at MWCNT-GCE; pH 4.51 (a) blank,

−1 −1 −1 −1 −1

of the calibration curve.

Table 2Application of the proposed voltammetric methods for the analysis of dosage form.

Parameters MWCNT-GCE

SWV DPV

Labeled amount (g mL−1) 1.0 1.0Amount found (g mL−1)a 0.987 0.962RSD (%) 0.74 0.48Amount added (mg mL−1) 1.0 1.0

R. Jain, Vikas / Colloids and Surfac

nd scan rate on the reduction of cefpirome at the MWCNT modi-ed glassy carbon electrode. The reduction peak current increasess pH value increases from 2.0 to 4.51. At pH value beyond 4.51,t decreased gradually with the further rise in pH value. There-ore, Britton Robinson buffer of pH 4.51 has been selected forhe determination of cefpirome in pharmaceutical formulations.n additional, the reduction peak potential (EP) shifts towards neg-tive potential with the increase of pH, indicating that proton isirectly involved in the reduction of cefpirome.

The reversibility of the reduction process was further studiedsing cyclic voltammetry with glassy carbon as working, Ag/AgCleference and platinum wire as auxiliary electrode. The cyclicoltammogram of cefpirome in Britton Robinson buffer in 1.0%etyltrimethylammonium bromide exhibits a single well definedathodic peak at all concentrations due to the reduction of –C N–roup. An anodic peak was obtained at far off positive potentialn the entire pH ranges indicating the irreversible nature of thelectrode process.

Useful information involving electrochemical mechanism usu-lly can be acquired from the relationship between peak currentnd scan rate. Therefore, the electrochemical behaviour of cef-irome at different scan rates of 10–50 mV/s was also investigatedn the surface of the MWCNTs/Nafion modified GCE by cyclicoltammetry. As the scan (�1/2) rate is increased from 10 to 50 mV/st a fixed concentration of cefpirome: (i) the peak potential shiftsathodically during forward scan and anodically during reversecan, (ii) the peak current function, ip/AC�1/2 exhibits almost con-tancy. A linear Randles–Seveik plot (plot of ip against �1/2) isbtained, a straight line is obtained with linear regression equa-ion; ip (�A) = 4.076�1/2 + 3.584, r2 = 0.984 for cathodic peak and ip�A) = 1.133�1/2 + 1.607, r2 = 0.968 anodic peak; indicating a diffu-ion controlled electrode process [27]. This finding is confirmed bylotting log ip against log �1/2; a straight line is obtained which cane expressed by the equation: log ip (�A) = 0.854 log �1/2 + 0.782,2 = 0.993 and log ip (�A) = 0.747 log �1/2 + 0.341, r2 = 0.968 withlope value 0.854 and 0.747, which is less than the theoretical valuef 1.0 that is expected for an ideal reaction of surface species. Theower experimental slope (0.854 and 0.747) than the theoreticalne may be attributed to the partial involvement of the diffusiverug molecules in the electrode reaction of the adsorbed ones.he overall electrode process is mainly diffusion-controlled withdsorption of the drug molecules at the electrode surface [28].

.3. Analytical application

In order to develop a voltammetric method for determininghe drug, the differential-pulse voltammetric mode and SWV waselected, because the peaks are sharper and better defined at loweroncentration of cefpirome than those obtained by cyclic voltam-etry, with a lower background current, resulting in improved

esolution. According to the obtained results, it is possible to applyhis technique to the quantitative analysis of cefpirome. The BRuffer solution of pH 4.51 was selected as the supporting elec-rolyte for the quantification as cefpirome gave maximum peakurrent at pH 4.51. The quantitative evaluation was based on theependence of the peak current on cefpirome concentration. Theeak currents increased linearly with increasing amounts of cef-irome by differential-pulse and square-wave voltammetry. Underhe optimum conditions the calibration plot of the peak currentersus the concentration was found to be linear over the range00–600 �g mL−1 in the square wave and differential pulse voltam-etric method and the linear regression equation is expressed as:

WV : [ip(�A) = 148.2 − 19.12], r2 = 0.991 (1)

PV : [ip(�A) = 15.16 − 1.318], r2 = 0.988 (2)

(b) 100 �g mL , (c) 200 �g mL , (d) 300 �g mL , (e) 400 �g mL , (f) 500 �g mLand (g) 600 �g mL−1.

The regression plots showed that there is a linear dependenceof the current intensity on the concentration in both SWV and DPVmodes over the range as given in Table 1. The table also showsthe detection limits and the results of the statistical analysis of theexperimental data such as slopes, intercept, the correlation coeffi-cients obtained by the linear least squares treatment of the resultsalong with standard deviation (SD) of intercept (Sa) on the ordinate(Fig. 3). The good linearity of the calibration graphs and the negli-gible scatter of the experimental points are clearly evident by thevalues of the correlation coefficient and SD The characteristics ofthe calibration plots are listed in Table 1. The LOD and LOQ wereestimated using the following equations [29]:

LOD = 3Sa

b, LOQ = 10Sa

b(3)

where Sa is the standard deviation of the intercept and b is the slope

Amount found (mg mL−1)a 0.992 0.985Recovery (%) 99.2 98.5RSD (%) 0.85 0.54

a Each value in the mean of five experiments.

426 R. Jain, Vikas / Colloids and Surfaces B: Biointerfaces 87 (2011) 423– 426

Table 3Quantification of cefpirome in injection by the proposed methods.

Technique Nominal conc. (�g mL−1) aFound conc. (�g mL−1) RSD (%) Average recovery (%) Bias (%)

SWV 400 397.15 1.28 98.28 0.712600 592.28 1.01 98.71 1.286

DPV 400 394.23 0.91 98.55 1.44

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.4. Specificity

For specificity test, voltammograms of standard solutions ofnjection were recorded under selected experimental conditions.esponse of analyte in this mixture was compared with theesponse of pure cefpirome. It was found that assay results wereot changed.

.5. Accuracy

The accuracy of developed method was carried out by spikingith accurately weighed amounts of cefpirome at concentration

f the commercial injection (1.0 g). The accuracy is expressed as mean relative error (measured conc. − nominal conc./nominalonc. × 100). The mean recoveries in injection were found to be8.99%, respectively and the values of mean relative error arecceptable (Table 2); this shows the best accuracy obtained by usinghese methods.

.6. Precision/reproducibility

The precision and reproducibility of these developed methodsSWV, DPV) for cefpirome were determined in five replicates anal-ses of 400, 600 �g mL−1 (Table 3). The precision of the proposedrocedure was estimated by analyzing cefpirome in injection assayolutions for five times in three successive days using SWV andPV. The results confirmed good precision of the proposed proce-ure and stability of the drug’s solution. The mean RSD% was foundo be 1.14% and 1.02% for SWV and DPV methods, respectively. Theariation coefficients were found less than 2% indicating that twoethods are precise and confidence.

. Pharmaceutical applications

The amount of cefpirome in injection was calculated by refer-nce to the appropriate calibration plots. The results obtained areiven in Table 2. The proposed techniques could be applied withreat success to cefpirome assay in injection without any interfer-nce.

As there is no official technique in the pharmacopoeias or otheriterature describing the determination of cefpirome in pharma-eutical dosage forms. For this reason, the proposed techniquesere checked by performing recovery tests. To determine whether

xcipients in the injection interfered with the analysis, the accu-acy of the proposed methods were evaluated by recovery testsfter addition of known amounts of pure drug to pre-analyzed for-ulations of cefpirome (Table 2). The results showed the validity of

he proposed techniques for the quantitative determination of cef-irome in injection. The proposed DPV and SWV techniques provedo be precise and accurate for reliable electro analytical analysis ofefpirome.

. Conclusions

A sensitive and selective electrochemical method for determi-ation of cefpirome has been developed. This method was based

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on the enhanced peak current responses of cefpirome reduc-tion at MWCNT modified electrode. Compared to the responseat bare glassy carbon electrodes, the reduction current responseat the modified electrode was greatly improved. Because of thevery rich electrochemical properties of MWCNTs, they act aselectron-mediators thus operating as electron relays for activa-tion of reduction of the analyte. This novel sensing system forcefpirome was convenient and showed excellent analytical char-acteristics such as significant lowering of the detection limit,high sensitivity and satisfactory selectivity. A differential pulse(DP) and square wave (SW) voltammetry method could be suc-cessfully applied to the quantification of cefpirome in bulk formand pharmaceutical formulation. The modified electrode has goodoperating characteristics like simplicity, sensitivity, stability, selec-tivity, low detection limit and wide linear working range. Aboveadvantages together with the very easy preparation and easyregeneration of electrode surface by simply polishing make thesystem useful in constructing sensor device for determination ofcefpirome.

Acknowledgment

The authors are grateful to the Director, Defence Research andDevelopment Establishment, Gwalior (M.P.), India for prosily SEMfacilities to carry out of this work.

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