Characterization and application of humic acid modifiedcarbon electrodes

Carlos D. Garcıa, Patricia I. Ortiz *

INFIQC, Facultad de Ciencias Quımicas, Departamento de Fısicoquımica, Universidad Nacional de Cordoba Ciudad Universitaria,

Cordoba, Argentina

Received 11 March 2003; received in revised form 9 May 2003; accepted 14 May 2003

Abstract

Four routes for the modification of carbon electrodes with humic acids and the determination of three divalent

metallic cations were studied. The determination of bound Fe2�, Cu2� and Ni2� was performed by cyclic and square

wave voltammetry using either a batch or flow analysis system. Using the FIA system and SWV, linear relationships

between the oxidation (or reduction) current and the cations concentration were obtained with the modified electrodes,

while no signals were obtained for the same conditions for bare carbon electrodes. The system can be used to study the

interaction between a wide range of electroactive cations and humic substances; however, the performance as an

analytical tool is limited due to the high limits of detection (mM). However, some advantages like simplicity, short

analysis time, inexpensive instrumentation needs and miniaturization capabilities are remarkable.

# 2003 Elsevier B.V. All rights reserved.

Keywords: Fe2�, Cu2� and Ni2� determination; Humic acid; Modified carbon electrodes

1. Introduction

Humic substances comprise a general class of

biogenic, refractory, yellowish-black organic sub-

stances that are ubiquitous, occurring in all

terrestrial and aquatic environments. These humic

substances constitute the major organic fractions

in soils and have been studied by soil scientists for

two centuries. One of the first difficulties encoun-

tered in the study of humic substances relates to

terminology, because these substances do not

corresponds to a unique chemical entity, and

accordingly, cannot be described in unambiguous

structural terms. There are three major fractions of

humic substances operationally defined in terms of

their solubility. By these criteria, humic acids

(HAs) are considered those humic substances

that are not soluble in water under acid conditions

(below pH 2), but becomes soluble at greater pH

values [1]. Humic (and also fulvic) acids are

natural, weak polyelectrolytes that are very active

in binding ions and organic molecules [2] and play

* Corresponding author. Present address: INFIQC,

Departamento de Fısicoquımica, Facultad de Ciencias

Quımicas, Universidad Nacional de Cordoba, Ciudad

Universitaria, Cordoba, Argentina. Tel.: �/54-351-4334169/80;

fax: �/54-351-4334188.

E-mail addresses: carlos.garcia@colostate.edu (C.D.

Garcıa), portiz@fisquim.fcq.unc.edu.ar (P.I. Ortiz).

Talanta 61 (2003) 547�/556

www.elsevier.com/locate/talanta

0039-9140/03/$ - see front matter # 2003 Elsevier B.V. All rights reserved.

doi:10.1016/S0039-9140(03)00321-7

an important role in the environment, being one ofthe major complexing components in soils and

water systems [3]. They can also affect diverse

phenomena such as transport mechanisms, toxicity

or bioavailability [4,5]. Citric acid groups, weakly

acid enols [6], malonate, salicylic and aromatic o-

dicarboxylic acids groups present in HA are as

responsible for the complexing properties between

HA and metallic cations such as H�, Ca2�, Cd2�,Cu2� [7], Pb2� [8], Al3� [9,10], Ni2�, Fe2� and

Fe3� [11].

However, the potential utility of HA for metal

ion analysis depends on the modifications that can

be performed on electrode surfaces using these

substances. Chemically modified electrodes have

been widely applied in electroanalytical chemistry

as a means of improving sensitivity and selectivity[12], in particular those based on carbon, have

gained much attention during the last few years.

Different preparation methods for these modified

electrodes have been described [12,13] and depend-

ing on the electrode material, there are various

methods for each modification. In the case of

carbon paste electrodes (CPEs), one of the most

commonly used preparations is the direct mixingof the modifier into the carbon, i.e. an insoluble

particulate component is mixed into the graphite

powder and then the pasting liquid is added [14]

allowing the preferential preconcentration of ana-

lytes on their surfaces [15,16]. In the case of glassy

carbon electrodes [17], HA cannot be included into

the base material, so a surface modification must

be performed. A film deposition can be obtainedby a combination of chemisorptive effects and low

solubility in the contacting solution [18] allowing

one to control the deposited amount or speed of

the process by adjusting the deposition time and/or

the applied potential, respectively.

As pointed out above, modification of carbon

surfaces with HAs can impart selectivity and

sensitivity to the electrochemical quantificationof cations. The main purpose of this paper is to

report some of the analytical possibilities of

carbon electrodes modified by HAs. Different

deposition methods and two modified carbon

electrodes were studied using standard solutions

of Ni2�, Cu2� and Fe2� as probes. For each

electrode, a different working procedure was used.

The first one is a batch arrangement, using cyclicvoltammetry (CV) and a modified carbon paste

electrodes (MCPEs). The second approach con-

sists of a flow injection analysis (FIA) system

coupled with square wave voltammetry (SWV)

detection and using modified glassy carbon elec-

trodes (MGCEs).

2. Experimental

2.1. Reagents

All the chemicals were of analytical grade. The

solutions were prepared with ultrapure water from

Millipore MilliQ System. Nickel, iron and copper

stock solutions were prepared daily and diluted as

required to prepare standard solutions from nickel

sulphate (Mallinckrodt), iron(II) sulphate (Ane-

dra) and copper(II) sulphate (Cicarelli), respec-tively. A 1.0 M HClO4 solution from Merck was

used for cleaning the electrode.

HA, provided by Fluka, was washed with

concentrated HCl and the precipitate was thor-

oughly washed with water to remove Cl� anions.

HA was then oven-dried at 40 8C, and the powder

was weighed and redissolved in NaOH (pH 12.0)

to prepare a stock solution of 4.766 g l�1.

2.2. Apparatus

Cyclic voltammetry experiments were per-

formed with a LyP M5 potentiostat coupled to a

LyP programmer and a Houston X-Y recorder

and carried out in a conventional three-electrode

cell. SWV experiments were performed with an

Autolab PGSTAT 30 and data collected with a PC

using GPES software (version 4.8). AgjAgClj3 MNaCl and Pt were used as reference and auxiliary

electrodes, respectively. Carbon paste electrodes,

described previously [19], and glassy carbon elec-

trodes (MF-1000 BioAnalytical Systems) were

used throughout. A Metrolab Spectrophotometer

was used for HA adsorption characterization.

C.D. Garcıa, P.I. Ortiz / Talanta 61 (2003) 547�/556548

2.3. Electrode preparation

Four different immobilization routes were ana-

lyzed for the HA immobilization on carbon

particles. For the first modified carbon paste

electrode (MCPE1), carbon particles were mixed

with the HA solution and gently stirred overnight.

The modified carbon particles were mixed with

light mineral oil (30%, w/w). Direct, solid HAmixing with the carbon and mineral oil was also

tested (MCPE2), but a visible loss of HA was

observed during the experiment. The third mod-

ification route was performed electrochemically

(MCPE3) on an already prepared carbon paste

electrode, applying 1.2 V vs Pt in a stirred alkaline

HA solution for 1 h. Finally, for comparative

purposes the carbon paste electrode was immersedin the HA solution without applying any potential,

up to 3 h (MCPE4); nevertheless, no HA adsorp-

tion was observed under these conditions. Accord-

ing to these previously obtained results, MGCEs

were prepared using the same scheme of MCPE3.

2.4. Analytical procedure

For the first batch procedure, a typical accumu-lation step was performed by immersing the

modified electrode in stirred solutions of different

HA concentration values for 5 min and applying

�/0.40 V. Further, the electrode was washed with

purified water and finally the corresponding cyclic

voltammogram was obtained in a new base solu-

tion between �/0.20 and 1.00 V at 0.10 V s�1.

For the FIA procedure, the accumulation wascarried out by flowing different HA concentration

solutions for 5 min, applying �/0.60 V. After that,

the corresponding SWV procedure was performed

in the same working solution from the accumula-

tion potential value (�/0.60 V) to 1.20 V (forward)

and from 1.20 to �/0.60 V. Fig. 1 shows the FIA

configuration used for this methodology.

For both methodologies, quantification wasobtained from the corresponding oxidation or

reduction peak current of the accumulated metal.

A cleaning step was performed after the determi-

nation by immersion or rinsing the electrodes with

0.1 M HClO4 for 10�/30 s. For all experiments, the

detection limit corresponds to the lowest measured

concentration with a signal-to-noise ratio of atleast 3.

3. Results and discussion

3.1. Adsorption isotherm

An adsorption isotherm was constructed inorder to determine the obtained amount of modi-

fier per gram of carbon powder. Experiments were

carried out in 0.10 M potassium hydrogen phtha-

late (KHPh) buffer and were evaluated spectro-

photometrically measuring the absorbance at 550

nm. Different HA aliquots were added to weighed

amounts of acetylene carbon black particles,

dispersed in buffer. After properly mixing, theywere allowed to stand overnight, and finally after a

centrifugation step the supernatant concentration

was analyzed by comparison with a previously

constructed calibration curve under the same

conditions.

The adsorption isotherm is shown in Fig. 2,

where the experimental data are compared to the

Langmuir equation. HA readily adsorbs on car-bon particles with an adsorption constant equal to

0.3 and a surface saturation value of 0.55 g HA

g�1 carbon powder.

3.2. Batch results using MCPE

Prior to use in an analytical determination, a

cyclic voltammetric study of the carbon paste

electrodes, either modified or unmodified, wasperformed in supporting electrolyte. Modified

electrodes showed a current increase over the

whole i /E profile when compared with the un-

modified ones. This adverse current increase was

higher for MCPE1. For that reason, and consider-

ing the other two routes have very low stability

(MCPE2) or current response (MCPE4), MCPE3

was used throughout. It is worthy to note thatneither anodic nor cathodic current peaks were

observed with the unmodified carbon paste elec-

trode when treated in the same way as the

modified ones for cation determination.

Fig. 3A shows the potentiodynamic i /E profiles

carried out with MCPE3 starting at �/0.2 V and

C.D. Garcıa, P.I. Ortiz / Talanta 61 (2003) 547�/556 549

progressing towards positive potential values in a

blank (0.10 M KHPh) and standard Fe2� solu-

tions (prepared in 0.10 M KHPh) after 5 min

accumulation. Two current peaks were observed;

the first anodic peak at 0.21 V can be attributed to

the oxidation of Fe2� to Fe3�, while the cathodic

peak obtained at ca. 0.05 V is related to the quasi-

reversible reduction process of the previously

formed Fe3�. Calibration curves were performed

in the same conditions and analyzed using the first,

anodic current peak (see Table 1). A linear

dependence of the current with the Fe2� concen-

tration was observed up to 1.0�/10�3 M with a

slope of 0.309/0.03 A cm�2 M�1 (regression

coefficient�/0.98). The limit of detection was

1�/10�5 M.

The potentiodynamic i /E profiles with MCPE3

from �/0.2 V towards positive potential values for

a blank (0.10 M KHPh) and different Cu2�

standard solutions in 0.10 M KHPh after 5 min

accumulation is shown in Fig. 3B. During the

positive potential scan, an anodic current peak at

Fig. 1. Diagram of the instrumental configuration used to perform the FIA-SWV analysis.

Fig. 2. Adsorption isotherm for HA on carbon powder (0.10 M KHPh buffer).

C.D. Garcıa, P.I. Ortiz / Talanta 61 (2003) 547�/556550

0.05 V that shifts to higher potential values as

Cu2� concentration increases is observed. During

the negative potential sweep, no peaks can be

observed. The anodic current peak-concentration

dependence shows a linear relationship up to

1.0�/10�4 M, with a slope of 43.49/0.9 A cm�2

M�1 (regression coefficient�/0.9998) and a limit

of detection of 1.0�/10�5 M.

Copper quantification was also tested by im-

mersing the electrode in different standard solu-

tions without potential application. Under these

conditions, an oxidation current peak at 0.09 V

(not shown) was obtained and a linear relationship

of the current peak with Cu2� concentration was

observed, covering also a wider concentration

range (1.0�/10�5�/1.0�/10�3 M). The slope of

Fig. 3. (A) Potentiodynamic i /E profiles in 0.10 M KHPh at 0.10 V s�1 after the electrode immersion in Fe2� solutions with different

concentration values: (---) blank, (- - -) 1.0�/10�3 M and (� � �) 1.0�/10�4 M. (B) Cyclic voltammograms in 0.10 M KHPh at 0.10 V

s�1 in different copper standard solutions, accumulation for 5 min at �/0.40 V: (---) blank, (- - -) 1.0�/10�5 M, (- �/- �/-) 5.0�/10�5 M,

(� � �) 1.0�/10 �4 M and (- �/ �/- �/ �/-) 5.0�/10�4 M. (C) Cyclic voltammograms in 0.10 M Na2SO4 at 0.10 V s�1 in different Ni2� standard

solutions, accumulation for 5 min at �/0.40 V (except for the blank only the negative scans are shown): (---) blank, (- - -) 8.0�/10�5 M,

(- �/- �/-) 2.0�/10�4 M and (� � �) 8.0�/10 �4 M.

Table 1

Analytical parameters of the calibration curves performed with an MCPE for Fe2� and Cu2� in 0.10 M KHPh and Ni2� in 0.10 M

Na2SO4

Cation Accumulation potential (V) Ep (V) Linearity range (M) Slope (A cm�2 M�1) R

Fe2� 0 0.21 1.0�/10�5�/1.0�/10�3 0.309/003 0.98

Cu2� 0 0.09 1.0�/10�5�/1.0�/10�3 0.7229/0.005 0.9996

�/0.4 0.05 1.0�/10�5�/1.0�/10�4 43.49/0.9 0.9998

Ni2� 0 No peak observed

�/0.4 0.50 1.0�/10�5�/8.0�/10�4 0.10339/0.007 0.994

C.D. Garcıa, P.I. Ortiz / Talanta 61 (2003) 547�/556 551

the curve was 0.7229/0.005 A cm�2 M�1, with aregression coefficient of 0.9996 and a limit of

detection of 1�/10�5 M. Although this method

can be used for copper quantification and the

concentration range is wider, the slope is consider-

ably lower than that obtained with a potential

accumulation step, so that the first procedure is

recommended.

For Ni2� determination, no response wasobtained in KHPh solutions using the conditions

previously described. For that reason, the accu-

mulation was studied in 0.10 M Na2SO4. Fig. 3C

shows the potentiodynamic i /E profiles with

MCPE3 starting at �/0.2 V towards positive

potential values for a blank (0.10 M Na2SO4)

and different Ni2� standard solutions after 5 min

accumulation. No current peaks were obtainedduring the positive sweep while in the negative

scan, a sharp cathodic current peak at 0.50 V was

observed. The peak shifts toward less positive

potential values as Ni2� concentration increases.

A linear relationship between the cathodic current

peak and Ni2� concentration was obtained up to

8.0�/10�4 M, with a slope of 0.10339/0.007 A

cm�2 M�1 (regression coefficient�/0.994) and thelimit of detection was 1.0�/10�5 M.

Analytical parameters corresponding to the

calibration curves for the three analyzed cations

in the above-mentioned conditions are summar-

ized in Table 1.

In all cases, neither an oxidation nor a reduction

current peak was observed with an unmodified

carbon paste electrode, under the same conditionspreviously described, indicating that Fe2�, Cu2�

or Ni2� cannot be deposited on the electrode

surface in the absence of HA.

3.3. FIA results using MGCE

Several parameters can affect the current peak

in the FIA-SWV method. Specifically, each para-

meter of the SWV method is able to modify, in acertain way, the magnitude of the current peak.

Fig. 4 shows the waveform and the assigned names

of the optimized parameters during the following

study. Note that according to the SWV technique,

the recorded curve corresponds to the difference

between the average currents in the forward and

the reverse pulse, sampled just before each flank

[20].

During the first step, the accumulation proce-

dure parameters were optimized recording the

positive scan (�/0.6 to 1.2 V) and the reverse

scan (1.2 to �/0.6 V) with no delay time. Fig. 5

shows the effect of deposition time on the electro-

chemical response evaluated from 100 to 750 s

using a 1.9�/10�4 M Cu2� solution in Na2SO4

and applying �/0.6 V. As can be observed, a clear

current increase can be obtained increasing the

accumulation time but, in order to obtain a good

relationship between current magnitude and ana-

lysis time, 300 s was found to be optimum.

Not only the time, but also the deposition

potential has an important effect on the electro-

chemical response of the cations. This effect was

studied from 0 to �/0.80 V using 0.10 V steps. As

the potential becomes more negative, a linear

current increase was observed for Cu2� until �/

0.60 V, where a current plateau was reached (Fig.

6). An analogous behavior was found for Ni2�.

Similar to the experiments performed in batch, it

was not necessary to apply potential to perform

the accumulation of Fe2�. However, in order to

analyze mixtures, the same procedure was applied

to all the analytes.

For all three cations, the forward and reverse

scans were recorded in order to characterize the

responses and analyze mixtures of them. SWV was

optimized by the variation of modulation time

(MT), modulation amplitude (MA), interval time

(IT) and step potential (SP) values that should be

adjusted according to the kinetics of the electro-

Fig. 4. Waveform and assigned names of the optimized

parameters during the electrochemical study, according to the

SWV technique.

C.D. Garcıa, P.I. Ortiz / Talanta 61 (2003) 547�/556552

chemical process. The variation of different para-

meters affects not only the magnitude of the signal,

but also the oxidation (or reduction) potential

value.

Fig. 5. Effect of deposition time on the electrochemical response from 100 to 750 s (1.9�/10�4 M Cu2� solution in Na2SO4 and

applying �/0.6 V).

Fig. 6. Effect of deposition potential (DP) on the electrochemical response from �/100 to �/800 V (1.9�/10�4 M Cu2� solution in

Na2SO4).

C.D. Garcıa, P.I. Ortiz / Talanta 61 (2003) 547�/556 553

For Ni2�, the reverse scan was used for thequantification process, where a clear cathodic

wave was obtained at around 0.60 V. In the case

of Cu2�, three peaks were observed in the positive

scan (�/0.6 to 1.2 V), but only the best defined one

at �/0.19 V was used for the quantification. A

well-defined peak was observed for iron solutions

during the positive scan at 0.030 V (not shown).

The obtained values during the optimizationprocess are summarized in Table 2, in which

MT�/0.05 s, MA�/0.025 V, IT�/0.5 s and

SP�/0.005 V were set as the best combination

for obtaining calibration curves for the three

cations.

Fig. 7 shows the dependence between peak

current and concentration for the analyzed cou-

ples. As can be observed, the best response wasobtained with Cu2� solutions for which not only a

higher slope, but also a better detection limit

(6.0�/10�7 M) was obtained. Fe2� and Ni2�

showed similar slopes and linear ranges, being

2.0�/10�6 and 6.0�/10�6 M, respectively, the

correspondent limits of detection (see Table 3).

In all cases, neither an oxidation nor a reduction

current peak was observed with the unmodifiedglassy carbon electrode, indicating that Fe2�,

Cu2� or Ni2� cannot be accumulated on the

carbon surface in the absence of HA.

4. Conclusions

Different procedures can be used to prepare

HA-modified carbon electrodes. Taking into ac-count the stability of the electrodes, the reprodu-

cibility of the results and the time needed for the

preparation, the most suitable methodology is

electrochemical immobilization on glassy carbon

surfaces. This can be achieved by applying 1.2 V

(vs Pt) in an alkaline-stirred HA solution for 1 h.

The complexation reactions between the immobi-

lized HA and Fe2�, Cu2� or Ni2� were enhancedby applying a negative potential to the electrode.

Compared to a batch procedure, FIA methodol-

ogy has proven to be faster and facilitates sample

handling procedures, but the detection limits were

not better than those obtained with standard CV.

To overcome this inconvenience, further studies Ta

ble

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7

C.D. Garcıa, P.I. Ortiz / Talanta 61 (2003) 547�/556554

using larger amounts of modifier have to be

performed. The electrodes showed very good

linear responses between the concentration and

the peak current for the three analyzed cations;

however, the analytical applications of these

electrodes are limited due to the limits of detection.

However, these can be overcome by increasing the

amount of HA immobilized on the surface by

increasing the electrodeposition time. The de-

scribed setup can also be used to study and

characterize the complexation between HA and

electroactive cations suggesting an enormous num-

ber of possible applications. Related studies are

currently being performed to characterize the

effect of different pollutants (surfactants and

organic solvents) on the complexation reaction

between heavy metals and humic substances.

Acknowledgements

This work was supported by CONICET from

Argentina, CONICOR from Provincia de Cordo-

ba, Argentina, and SECyT-UNC. Carlos D.Garcıa thanks CONICET for the scholarship

granted. The authors also wish to thank M.E.

Sosa for performing part of the experiments.

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Fig. 7. Calibration curves for (m) Cu2�, (') Ni2� and (j) Fe2� solutions (DP�/�/0.6 V, DT�/300 s, MA�/0.025 V, MT�/0.05 s,

Table 3

Analytical parameters of the calibration curves performed with an MGCE for Fe2�, Cu2� and Ni2� in Na2SO4 0.10 M

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