Electrochemical Determination of Ascorbic Acid Using Modified Glassy Carbon Electrode by Multiwall Carbon Nanotube-Nafion in Chloroacetic Acid Media

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1 Asin Journl of Chemistry; Vol. 25, No. 1 (213), Electrochemicl Determintion of Ascorbic Acid Using Modified Glssy Crbon Electrode by Multiwll Crbon Nnotube-Nfion in Chlorocetic Acid Medi MEISSAM NOROOZIFAR 1,*, MOZHGAN KHORASANI-MOTLAGH 2 nd HAMED TAVAKKOLI 1 1 Anlyticl Reserch Lbortory, Deprtment of Chemistry, University of Sistn nd Bluchestn, Zhedn, P.O. Box , Irn 2 Inorgnic Reserch Lbortory, Deprtment of Chemistry, University of Sistn nd Bluchestn, Zhedn, Irn *Corresponding uthor: Fx: ; Tel: , E-mil: mnoroozifr@chem.usb.c.ir (Received: 4 October 211; Accepted: 12 July 212) AJC In this study, scorbic cid ws simply determined t lower potentil compred to bre glssy crbon by replcement phosphte buffer solution with chlorocette buffer solution t surfce of modified glssy crbon electrode. A film of Nfion with multi-wlled crbon nnotubes (MWCNT) ws prepred by rpidly csting solution onto the surfce of glssy crbon electrode nd drying under room temperture. For MWCNT-Nfion film ws not observed ny peks in the used potentil rnge in phosphte buffer solution. However, by replcing phosphte buffer solution with chlorocette buffer solution, due to interction between nfion film nd component of chlorocette buffer solution in positive scn, n oxidtion pek ws obtined t bout.442 V nd t reversl scn reduction pek ws observed t bout.282 V. Scn rte, ph nd interference effects for the determintion of scorbic cid were studied. It ws found tht glssy crbon electrode/mwcnt-nfion in chlorocette buffer solution ph = 2, increse significntly the cyclic voltmmogrm pek currents s well s decresing the oxidtion overpotentils of scorbic cid compred to bre glssy crbon. A liner rnge between mol/l nd mol/l nd detection limit (3σ) mol/l ws obtined for modified electrode. Finlly, the novel modified electrode ws successfully pplied to the determintion of scorbic cid in phrmceuticls nd foods in cette medium. Key Words: Electrode modifiction, Nfion, Multi-wlled crbon nnotubes, Chlorocette buffer, Ascorbic cid, Electroctlytic determintion. INTRODUCTION Ascorbic cid (AA) is used in lrge scle s n ntioxidnt in food, niml feed, beverges, phrmceuticl formultions nd cosmetic pplictions. Ascorbic cid is the mjor soluble ntioxidnt found in plnts nd is lso n essentil component of humn nutrition 1. In ddition, scorbte is importnt for neuronl mturtion nd function, s well s for protection of the brin ginst oxidnt stress, is well supported by previous work 2. In view of the gret importnce nd wide use of scorbic cid, numerous nlyticl techniques hve been proposed for its determintion in different mtrices nd t different levels. These include flow injection (FI) nlysis 3, chromtogrphy 4, spectrophotometry 5,6 nd voltmmetry 7-9. Ascorbic cid cn be mesured using voltmmetry methods in order to gin the dvntges of high sensitivity nd selectivity. Recently, polymer films modified electrodes hve been receiving extensive interest due to their wide pplictions in the fields of chemicl sensors nd biosensors. Polymermodified electrodes hve mny dvntges in the detection of nlytes becuse of its high selectivity, sensitivity nd homogeneity in electrochemicl deposition, strong dherence to electrode surfce nd chemicl stbility of the film 1. Glssy crbon electrodes hve been widely used for modifiction 11,12 compred with metl electrodes due to its biocomptibility with tissue, hving low residul current over wide potentil rnge nd miniml propensity to show deteriorted response s result of electrode fouling 13. Nnotubes, the lst in the focus of scientists in series of ll crbon mterils discovered over the lst severl decdes re the most interesting nd hve the gretest potentil 14. A huge, cylindricl surfce re nd grphene sheet crbon nnotube hve specil properties, such s high electricl conductivity, chemicl stbility nd ctlyst support, which re useful for n nlyticl biosensor 15. Crbon nnotube-modified electrodes promote electron trnsfer due to their conductivity nd mechnicl properties. Composites of conducting polymer nd crbon nnotubes show synergistic ffects nd hs been mde for different pplictions 16. Wrpping crbon nnotubes by polymeric chins is now new pproch for chieving the solubility without ny considerble impirment their physicl, chemicl nd electrochemicl properties 17.

2 12 Noroozifr et l. Asin J. Chem. Composite mterils bsed on solubilizing crbon nnotubes with vrious polymers especilly Nfion nd chitosn hve been reported Nfion is n nionic sulfonted fluorocopolymer of tetrfluoroethylene (Teflon) nd perfluoro-3,6-diox-4-methyl- 7-octene-sulfonic cid. Nfion is not only like Teflon highly resistnt to chemicl ttck, it lso exhibits highly selective bsorption nd trnsfer of compounds. It hs hydrophobic (-CF 2-CF 2-) nd hydrophilic (-SO 3H) regions in its polymeric structure (Fig. 1) nd its supercidity is ttributed to the electron-withdrwing effect of the perfluorocrbon chin cting on the sulfonic cid group. Nfion is ble to ctlyze vrious rections, such s lkyltion, disproportiontion nd esterifiction. Moreover, it hs the unique ion exchnge, discrimintive, good electricl conductivity, good biocomptibility, excellent film forming nd dhesion bility, high chemicl stbility nd bility to resist interferences from nions nd biologicl mcromolecules, which mke it good mtrix for bimoleculr immobiliztion 21. In consequence of this bilities modified electrodes bse on Nfion-crbon nnotube hve been investigted intensively for nlyticl ppliction vriety of bimoleculrs such s scorbic cid, dopmine, uric cid, methnol nd others nd pplied for the voltmmetric sensing in the electrochemicl sensors nd biosensors. Fig. 1. Chemicl structures of Nfion Herein, this work includes the fbriction of new modified glssy crbon electrode bsed on multiwlled crbon nnotube dispersed in Nfion for the determintion of scorbic cid. The electrochemicl behviour of this modified electrode hs been studied in chlorocette cid buffer (CAAB) solutions ph = 2. Anlyticl performnce of this sensor for determintion of this nlyte ws evluted nd showed stisfctory results. Finlly, this sensor hs been used for the determintion of this compound in phrmceuticls nd foods successfully. Esy electrode preprtion nd low detection limits for scorbic cid ws obtined due to the high electroctlytic properties of MWCNT-Nfion in cidic medi. It ws lso successfully used for the determintion of scorbic cid in presence of the common interference compounds. EXPERIMENTAL Ascorbic cid ws purchsed from Fluk-Sigm nd ws used s received. Multiwll crbon nnotubes (MWCNT), with nnotube dimeters, OD = 2-3 nm, wll thickness=1-2 nm, length =.5-2 µm nd purity of > 95 % ws purchsed from Aldrich. A 2.5 % Nfion solution (Aldrich, EW, 11, dissolved in lower liphtic lcohol nd wter) ws used for prepring MWCNT-Nfion film. H 3PO 4, NH 2PO 4 nd NOH, were purchsed from Merck. Chlorocetic cid (pk α = 2.87) ws used for preprtion of buffer solutions between 1.8 nd 3.8, it ws prepred by djusting it to desire ph with.1 mol L -1 NOH solution. In sme wy for cetic cid buffer solution, cetic cid (pkα = 4.76) for ph between 4. nd 5.6 ws used. Phosphte buffer solutions were prepred from H 3PO 4 nd NH 2PO 4 (.1 mol L -1 ) nd djusted the ph rnge over 2-1 with.1mol L -1 H 3PO 4 nd NOH solutions. All experiments were performed under nitrogen tmosphere t room temperture. All the solutions were freshly prepred with doubly distilled wter nd ll other chemicls used were of nlyticl regent grde. Electrochemicl mesurements were crried out with SAMA5 electronlyser (SAMA Reserch Center, Irn). All electrochemicl experiments were crried out in conventionl three-electrode cell t room temperture. A pltinum electrode nd sturted clomel electrode were used s the counter nd reference electrodes, respectively. A Metrohm 632 ph-meter with Metrohm double junction glss electrode ws used for monitoring ph djustment. Preprtion of working electrodes: Before being modified, the glssy crbon electrode ws refined successively with.5-µm Al 2O 3 slurry. After rinsing with doubly distilled wter, it ws sonicted in wter nd bsolute ethnol for bout 1 min ech. Next, the glssy crbon electrode ws trnsferred to the electrochemicl cell for further clening by using cyclic voltmmetry between -1. nd +1. V t scn rte of 1 mvs -1 in recently prepred deoxygented.5 mol L -1 H 2SO 4 until stble cyclic voltmmetric profiles were obtined. The treted electrode ws dried under nitrogen strem nd used instntly for modifiction. After tht, first mke 2.5 % solution of Nfion (diluted with ethnol: doubly distilled wter, 1:1) nd then dded 25 mg of MWCNT nd gittion with ultrsonic to give blck suspension. Then the glssy crbon electrode ws smered evenly with 6 µl of suspension solution by micro-syringe nd dried under n infionrred lmp for 1 min. This modified GC electrode ws denoted s GC/MWCNT-Nfion. Also, the GC/Nfion electrode ws prepred with the sme method. RESULTS AND DISCUSSION Optimiztion of buffer solution nd electrode composition: The first step of the work consisted of finding the optimum buffer solution nd electrode composition. In order to decide bout the optimum condition of the developed electrode, modified electrode were exmined by recording cyclic voltmmogrms of modified electrode in different buffer solution (.1 mol L -1 ) of the chlorocette cid buffered solution nd phosphte buffered solution. A typicl cyclic voltmmogrm re given in Fig. 2. Glssy crbon electrode/mwcnt-nfion electrode did not show ny nodic nd cthodic peks in phosphte buffer solutions (curve ). Bsed on cyclic voltmmogrm for glssy crbon electrode/mwcnt-nfion (Fig. 2, curve c), the potentil of the nodic (E p ) nd cthodic (E pc ) peks nd the hlf-wve potentil (E 1/2 ) re.442,.282 V nd.362 V, respectively. Also, the glssy crbon electrode/nfion electrode exhibited pir of redox peks nd the hlf-wve potentil (E 1/2 ) t.654,.65 nd.36 V (curve b) respectively, in

3 Vol. 25, No. 1 (213) Electrochemicl Determintion of Ascorbic Acid 121 Modified electrode TABLE-1 ELECTROCHEMICAL DATA FOR DIFFERENT MODIFIED ELECTRODES IN BOTH CHLOROACETATE BUFFER AND PHOSPHATE BUFFER SOLUTIONS (vs. SCE). SCAN RATE, ν =1 mv s -1, ph = 2. E p E/V E pc E 1/2 /V E p /V I p I /µa I pc I p / I pc Buffer type GCE/MWCNT-NAF PBS GCE/NAF CABS GCE/MWCNT-NAF CABS chlorocette cid buffered solution, which cn be ttributed to the electron trnsformtions of Nfion in chlorocette cid buffered solution. A summry of electrochemicl dt for the modified electrodes re shown in Tble-1. The increse in current vlue nd the decrese in pek potentil (curve b nd c) cn be ttributed to the electroctlytic ctivity of crbon nnotube, which cn medite the electron trnsfer rections. Lrge surfce re is nother importnt dvntge of crbon nnotube tht cn fcilitte the modifiction of the glssy crbon electrode/mwcnt-nfion with biologicl molecules (A) j c b ( B ) y =.4773x R 2 =.971 () I p (µa) Fig. 2. Cyclic voltmmogrms of () glssy crbon electrode/mwcnt- Nfion in.1 mol L -1 phosphte buffer solution, ph 2 (b) glssy crbon electrode/nfion nd (c) glssy crbon electrode/mwcnt- Nfion in.1 M chlorocette buffer solution (ph = 2). Scn rte ws 1 mv s -1 It ws lso concluded tht electrochemicl behviour of glssy crbon electrode/mwcnt-nfion ws more reversible compred to glssy crbon electrode/nfion, where Nfion ws coupled with multi-wll crbon nnotube nd leding to considerble improvement in the nlyticl sensitivity. The effect of the scn rte for glssy crbon electrode/ MWCNT-Nfion by using cyclic voltmmetry is shown in Fig. 3A. These cyclic voltmmogrms were used to exmine the vrition of the pek current versus the sweep rte. Fig. 3A shows tht the modified electrode hd chemiclly qusireversible redox couple in phosphte buffer solutions (ph 2.) solution nd tht pek currents were incresed due to the cyclic voltmmetric scn rte. As shown in Fig. 3B, I p nd I pc were linerly dependent on the scn rte, s expected for surfce confined-redox process. The rtio of the nodic pek current (I p) to cthodic pek current (I pc) is lmost equl to unity (I p:i pc = 1). These behviours re consistent with diffusionless system, reversible electron trnsfer process t low scn rtes y = x R 2 =.9655 (b) (mvs ) Fig. 3. (A) Cyclic voltmmogrms of GCE/MWCNT-/NAF in.1 mol L -1 CABS ph 2, t vrious scn rtes: () 25; (b) 5; (c) 75; (d) 1; (e) 125; (f) 15; (g) 25; (h) 3; (i) 4; (j) 5; (B) Plots of nodic nd cthodic pek currents of GCE/MWCNT-NAF vs. scn rte (ν) from cyclic voltmmogrms Influence of ph on the film behviour: The effect of the ph on the electrochemicl behviour of scorbic cid ws exmined over the ph rnge from 2 to 1 t the surfce of the modified electrode by cyclic voltmmetry. Bsed on the results, the nodic pek current of glssy crbon electrode/ MWCNT-Nfion decreses with n increse in the solution ph until with incresing ph, the nodic pek current ws disppered t ph vlue 6. The cthodic pek current of glssy crbon electrode/mwcnt-nfion lso decreses slightly compred to nodic pek current with incresing ph (Fig. 4). Hence, for glssy crbon electrode/mwcnt-nfion, the highest nodic pek current ws obtined t ph 2. It must be noted tht ll the nodic pek potentils for glssy crbon electrode/mwcnt-nfion were decresed liner with incresing ph from 2 to 6, showing tht protons hve tken prt in their electrode processes 28. Becuse of suitble pek currents, chlorocette cid buffered solution with ph 2 ws chosen s optiml ph.

4 122 Noroozifr et l. Asin J. Chem ph= (A) Fig. 5. Cyclic voltmmogrms of GCE/MWCNT/NAF in 5 consecutive potentil scn cycles in CABS (ph 2.) t scn rte of 1 mv s -1 Current (µa) Fig. 4. ( B ) ph Potentil (V) (A) Cyclic voltmmogrms of GCE/MWCNT-NAF t vrious ph 2.; 4.; 6.; 8. nd 1.. (B) The effect of ph on the nodic pek current (solid line) nd nodic pek potentil (dsh line) of GCE/ MWCNT-NAF t scn rte 1 mv s -1 Stbility nd reproducibility of the modified electrode: Prior to using glssy crbon electrode/mwcnt-nfion for the electroctlytic oxidtion of scorbic cid, reproducibility nd stbility of the modified electrode ws investigted. The rte of loss of electrochemicl ctivity for this electrode ws evluted by noting ny decrese in the nodic chrge (q), in five consecutive potentil scn cycles (Fig. 5). Bsed on obtined results, ny chnge in the hlf-wve potentil, E 1/2 nd ny decrese in the nodic nd cthodic pek currents of glssy crbon electrode/mwcnt-nfion were not observed. The long lifetime stbility of the sensor ws exceptionl. It ws notble tht fter 6 dys the sensor retined 97.3 % of the initil sensitivity to scorbic cid. Good response reproducibility nd stbility my be explined by the fct tht the Nfion-chlorocette cid buffered solution complexes re uniformly self-ssembled t surfce of glssy crbon electrode nd unble to exit from it during potentil sweep. Electroctlytic oxidtion of scorbic cid t the surfce of glssy crbon electrode/mwcnt-nfion: The min objective of the present work is to determine the concentrtion of scorbic cid using modified glssy crbon electrode in chlorocette buffered solution. Fig. 6 depicts the CVs obtined for bre glssy crbon nd glssy crbon electrode/ MWCNT-Nfion modified electrode in.1 M chlorocette cid buffered solution ph = 2 contining M scorbic cid. The bre glssy crbon nd glssy crbon electrode/ MWCNT-Nfion electrodes showed n oxidtion wve t.66 nd.455 V (Fig. 6) nd current difference ( I/µA) scorbic cid, 4.5 nd 34.4 µa for M, respectively. It is shown tht the cyclic voltmmogrm of the oxidtion pek potentil of scorbic cid t the glssy crbon electrode/ MWCNT-Nfion shifted by.25 V to the negtive direction compred with tht bre glssy crbon electrode nd the pek current t the glssy crbon electrode/mwcnt-nfion lso incresed. The results indicte tht the electroctlytic ctivity of the modified electrode cn be pplied to the determintion of scorbic cid. The pek potentil for the oxidtion of scorbic cid ws lowered by t lest.25 V compred with tht obtined t n unmodified glssy crbon electrode. In contrst to bre glssy crbon electrode, when glssy crbon electrode/ MWCNT-Nfion ws used s the working electrode, the detection sensitivity ws improved significntly nd effective incresing of the nodic pek of scorbic cid ws obtined c d Fig b Cyclic voltmmogrms of () bre glssy crbon, (b) s () in present of M AA, (c) GCE/MWCNT-NAF nd (d) s (c) in present of M AA in CABS (ph 2.) t scn rte 1 mv s -1 Evlution of the proposed modified electrode: Under vrious optimum experimentl conditions, determintion of

5 Vol. 25, No. 1 (213) Electrochemicl Determintion of Ascorbic Acid 123 scorbic cid ws crried out t the potentil rnge of. to 1. V using the cyclic voltmmetry mode. Cyclic voltmmogrms of glssy crbon electrode/mwcnt-nfion versus the scorbic cid concentrtion re shown in Fig. 7. The scorbic cid electro-oxidtion pek ws observed t the potentil of bout.445 V versus sturted clomel electrode. The electroctlytic pek current of scorbic cid oxidtion t the surfce of glssy crbon electrode/mwcnt-nfion ws linerly dependent on the scorbic cid concentrtion. These pek currents were liner up to.23 M nd ws described by the eqution = [scorbic cid] , r =.999, n = 16, where is the difference of the oxidtion pek currents glssy crbon electrode/mwcnt-nfion before nd fter ddition of scorbic cid, [AA] (M) is the scorbic cid concentrtion, r is the correltion coefficient nd n represents the number of determintions (Fig. 7). The detection limit (3σ) ws M. Reltive stndrd devition (RSD %) for 5 determintions of scorbic cid with M concentrtion ws.6 %. Current (µa) Fig (A) (B) y = x R 2 = AA (mol) -1 (A) Cyclic voltmmogrms of scorbic cid t vrious concentrtion (from to p):.;.5;.31;.69; 1.2; 1.9; 2.9; 4.4; 5.9; 7.2; 8.9; 1.5; 13.; 15.1; 17.4; 19.; 21.5 nd 23.2 mm t surfce of the GCE/MWCNT/NAF. Solution conditions were ph 2. (.1 M CABS), scn rte 1 mvs -1. (B) Plot of I p vs. c p Interference studies: The influence of other severl compounds such s mleic cid, mltose, sucrose, fructose, oxlic cid, succinic cid, trtric cid nd glucose on the electroctlytic pek current of scorbic cid oxidtion t the surfce of glssy crbon electrode/mwcnt-nfion ws exmined using cyclic voltmmetry. If the tolernce limit ws tken s the mximum concentrtion of the foreign substnces, which cused n pproximtely 5 % reltive error, for mol L -1 scorbic cid. The experimentl results showed tht the presence of these compounds (except glucose) did not significntly influence the determintion of scorbic cid under the experimentl conditions. Therefore, this result demonstrted the selectivity of this method for the voltmmetric determintion of scorbic cid. The results re listed in Tble-2. TABLE-2 EFFECT OF DIVERSE SUBSTANCES ON THE ANALYSIS OF mol L -1 ASCORBIC ACID Substnce [Interference substnce]/[aa] Mleic cid, mltose 1 Sucrose 5 Fructose, oxlic cid, succinic cid, trtric cid 1 Glucose 1 Rel smples: In order to demonstrte the ctlytic oxidtion of scorbic cid in the rel smple, we exmined this bility in the voltmmetric determintion of scorbic cid in some phrmceuticl preprtions nd foods. Ornge juice nd phrmceuticl smples such s scorbic cid tblets nd powder (purchsed from locl sources) were selected s rel smples for nlysis by the proposed method using the stndrd ddition method in order to prevent ny mtrix effect. The results were compred with those obtined using the officil iodometric titrtion method 29. The pired t-test eqution, pplied in sttistics is; d n t = (6) where, d nd sd re the men nd stndrd devition respectively of d (the difference between pired vlues). For the pirs of vlues in Tble-3, the men difference, d, is nd the stndrd devition of the differences, s d, is The clculted t ws found to be The criticl vlue of t (tbulted t) for n-1 degrees of freedom (in this work 3) is 3.18 (P =.5). Since the clculted vlue of t is less thn this the null hypothesis is not rejected: the methods do not give significntly different results for scorbic cid concentrtion 3. Smple TABLE-3 DETERMINATION OF ASCORBIC ACID IN REAL SAMPLES USING GCE/MWCNT/NAF ELECTRODE Climed preprtion (mg) s d Proposed method / mg (±SD) Iodine method (mg) (±SD) AA tblet 25 per tblet (± 1.5) (± 1.7) AA Powder 5 per schet 48.2 (± 1.6) (± 1.9) Bluchestn s (± 2.4) 84.9 (± 2.1) Ornge (1 ml) Results bsed on five replicte determintions per smple

6 124 Noroozifr et l. Asin J. Chem. Conclusion This work explore simple method for determintion of scorbic cid t lower potentil compred to bre glssy crbon electrode using modified glssy crbon electrode by esy chnging buffer solution. The modified glssy crbon electrode with mixture of MWCNT nd Nfion shows efficient electroctlytic ctivity for determintion of scorbic cid in chlorocette cid buffered solution (ph 2.). For glssy crbon electrode/mwcnt-nfion modified electrode ny peks were not observed in the used potentil rnge in phosphte buffer solutions. In contrst, becuse of interction between Nfion film nd component of chlorocette cid buffered solution (formtion complex t surfce of electrode) in positive scn n oxidtion pek ws obtined t bout.442 V nd t reversl scn reduction pek ws observed t bout.282. Modified electrode not only shifted the oxidtion potentil of scorbic cid towrds less positive potentil (t lest.25 V compred to bre glssy crbon) but lso enhnced its oxidtion pek. The proposed voltmmetric method is rpid, simple, strict, sensitive, suitble nd selective method for routine control nd cn be crried out directly without ny pretretment for the determintion of scorbic cid in rel smples. For study the cpbility of this modified electrode in the rel smples, some phrmceuticl preprtions nd foods were used nd compred with stndrd titrimetric method. There ws no significnt difference between the two methods. Very esy fbriction procedure, wide liner rnge, high stbility nd good reproducibility for repeted determintion, propose this electrode s good nd ttrctive pplicnt for prcticl pplictions. REFERENCES 1. S.A. Kumr, P.H. Lo nd S.M. Chen, Biosens. Bioelect., 24, 518 (28). 2. F.E. Hrrison nd J.M. My, Free Rdic. Biol. Med., 46, 727 (29). 3. M. Noroozifr nd M. Khorsni-Motlgh, Tlnt, 61, 173 (23). 4. A. Ti nd E. Gohd, J. Chromtogr. B, 853, 214 (27). 5. M. Noroozifr, M. Khorsni-Motlgh nd A. Frhmnd, Act Chim. Slov., 51, 717 (24). 6. M. Noroozifr nd M. Khorsni-Motlgh, Turk. J. Chem., 27, 717 (23). 7. T. Rohni nd M.A. Ther, Tlnt, 78, 743 (29). 8. M. Noroozifr, M. Khorsni-Motlgh nd H. Tvkkoli, Anl. Sci., 27, 929 (211). 9. M. Khorsni-Motlgh nd M. Noroozifr, Anl. Sci., 19, 1671 (23). 1. R. Zhng, G.D. Jin, D. Chen nd X.Y. Hu, Sens. Actutors B, 138, 174 (29). 11. A.A. Ensfi, S.D.-Tehrni nd B. Rezei, J. Serb. Chem. Soc., 75, 1685 (21). 12. S. Stevnoic, D. Tripkovic, A. Kowl, D. Minic, V.M. Jovnovic nd A. Tripkovic, J. Serb. Chem. Soc., 73, 845 (28). 13. G. Jin, F. Hung, W. Li, S.Yu, S. Zhng nd J. Kong, Tlnt, 74, 815 (28). 14. S.N. Mrinkovic, J. Serb. Chem. Soc., 73, 891 (28). 15. S. Young Ly, Bioelectrochemistry, 68, 227 (26). 16. R.E. Sbzi, K. Rezpour nd N. Smdi, J. Serb. Chem. Soc., 75, 537 (21). 17. M. O'Connell, P. Poul, L. Ericson, C. Huffmn, Y. Wng, E. Hroz, C. Kuper, J. Tour, D. Ausmn nd R. Smlley, Chem. Phys. Lett., 342, 265 (21). 18. K. Wng, H. Yng, L. Zhu, J. Lio, T. Lu, W. Xing, S. Xing nd Q. Lv, J. Mol. Ctl. B: Enzym., 58, 194 (29). 19. B. Ge, Y. Tn, Q. Xie, M. M nd S. Yo, Sens. Actutors B, 137, 547 (29). 2. Y. Cheng, Y. Liu, J. Hung, Y. Xin, W. Zhng, Z. Zhng nd L. Jin, Tlnt, 75, 167 (28). 21. W. M nd D. Tin, Bioelectrochemistry, 78, 16 (21). 22. W.S. Hung, C.H. Yng nd S.H. Zhng, Anl. Bionl. Chem., 75, 73 (23). 23. H.S. Wng, T.H. Li, W.L. Ji nd H.Y. Xu, Biosens. Bioelectron., 22, 664 (26). 24. K. Lee, J.W. Lee, S.I. Kim nd B.K. Ju, Crbon, 49, 787 (211). 25. S. Shhrokhin nd H.R.Z.-Mehrjrdi, Electronlysis, 19, 2234 (27). 26. M.D. Rubines nd G.A. Rivs, Electrochem. Commun., 9, 48 (27). 27. L. Fernández nd H. Crrero, Electrochim. Act, 5, 1233 (25). 28. M. Shrp, M. Petersson nd K. Edstrom, J. Electronl. Chem., 95, 123 (1979). 29. A.A. Izugie nd F.O. Izugie, J. Agric. Biol. Sci., 3, 367 (27). 3. J.C. Miller nd J.N. Miller, Sttistics for Anlyticl Chemistry, John Wiley & Sons, New York, edn. 2, p. 58 (1988).