preparation of stock solutions -...

62

DESIGNING OF EXPERIMENTS

PREPARATION OF STOCK SOLUTIONS

All the reagents used were of Analytical grade (Sigma Chemicals). Graphite

powder (< 20 µm) and Mineral oil light, Nujol (density 0.838) were purchased from

Aldrich. The stock solutions of desired concentrations of Pb (II), Cd (II), Hg (I),

Cu (II) and Zn (II) were prepared by weighing requisite amount of Lead Nitrate

[Pb(NO3)2], Cadmium Nitrate [Cd(NO3)2], Mercurous Nitrate [Hg2(NO3)2.2H2O],

Copper Sulphate [CuSO4] and Zinc Nitrate [Zn(NO3)2] respectively and dissolving

them in distilled water (DW). The working solutions were freshly prepared from

stock solutions for each experimental run. Distilled water (ELGA PURELAB

option Q, R=18 MΩ) was used throughout the experiment. Mettler Toledo JB 1603-

C/FACT, single pan electronic balance was used for accurate weighing with

accuracy of four digits i.e. 0.1 mg.

PREPARATION OF PLANT MATERIALS

Plant material (modifier) preparation has been varied depending on the plant.

Different parts used for the preparation of the modified carbon paste electrodes are

as follows.

Water hyacinth: Leaves

Coconut: Outer shell

Rice: Straw (Prepared Cellulosic Nano fibers)

Lotus root: Root

Black rice: Grains (Extract)

Water hyacinth leaves were collected from Keetham Lake situated at west side of

Agra, in June 2010. Leaves were washed repeatedly with water to remove dust and

soluble impurities, dried in an oven at 50˚C, chopped in small pieces and powdered

in a grinder. Powdered material was passed through the 150 µm mesh copper sieve.

Prior to any experimentation, water hyacinth leaves powder was stirred in 0.01 M

HCl solution to remove any traces of metal impurity present previously in powder

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due to hyper accumulating nature of this plant. There after water hyacinth leaves

powder was washed twice with deionized (DI) water to remove any traces of acid.

Brown Coconut was purchased from the local market of Agra. The fibrous shell

(Meso carp) part was separated, washed with water to remove dust and soluble

impurities, dried in an oven at 50 ˚C for 24 hours, chopped in small pieces and

powdered in a grinder. The fraction of the particles with size less than 150 µm was

selected for electrode preparation. The coconut shell powder obtained by this

procedure is used as such for the preparation of electrodes.

Cellulosic nano fibers prepared from rice straw was received from Chemistry

Laboratory (Prof. M. M. Srivastava) D.E.I. These nano fibers were used for the

preparation of electrodes.

Lotus roots were collected from Keetham Lake. Similar to coconut shell, lotus roots

were washed with water to remove dust and soluble impurities, chopped in small

pieces, dried in an oven at 50 ˚C and powdered in a grinder. The particles with less

than 150 µm were separated. The lotus root powder obtained by this procedure is

used as such for the preparation of electrodes.

Black rice grains were obtained from China. Grains were washed with water to

remove dust and soluble impurities then dried and crushed. Ground rice particles

were used to extract anthocyanins. Ethanol with 0.1 % HCl was used as an

extracting medium. For 1 gm black rice, 30 ml extracting solvent was used. The

mixture was kept for sonication at 20K Hz frequency for 1 hour. The purple color

extract was filtered through Whatman No 1 filter paper. The extract was used for

the preparation of carbon paste electrodes (Rodriguez-Saona et al. 2001).

PREPARATION OF MODIFIED CARBON PASTE ELECTRODES

Unmodified carbon paste electrode (CPE) was prepared by mixing 0.3 gm graphite

powder to 100 µl mineral oil (80:20 w/w ratio). Modified carbon paste electrodes

(MCPEs) were prepared by substituting corresponding amounts of graphite powder

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(5%, 10%, 15 %, 20 %, 25 %, 30 %, 35 %, 40 % w/w) by modifier, followed by

addition of the mineral oil and thoroughly hand mixed for 15 minutes in mortar and

pestle. In case of black rice extract as a modifier, variable volume of the black rice

extract (5µl- 40µl) was mixed with fixed weight of graphite powder (0.3 g). The

homogenized paste was packed in a glass tube (id 3mm) where a copper wire was

inserted for electrical contact. The schematic diagram for the preparation of

modified CPEs and the corresponding images are shown in Figure 3.1 (Svancara et

al 2012). Image A shows the mixing of modifier and mineral oil with spatula, image

B shows the addition of graphite powder to it. Image C shows the mixing in mortar

and pestle and D shows the image of the prepared electrode.

Figure 3.1: Schematic diagram for the preparation of Modified carbon paste electrodes

Graphite Powder

Modifier

Mineral oil

Mixed in mortar and

pestle

Carbon (Graphite) paste as electrode

material

Filled in glass tube

Connection with a copper wire

Determination of metal ions

a) b)

c) d)

MORPHOLOGICAL CHARACTERIZATION OF PREPARED MODIFIED

CARBON PASTE ELECTRODES

Modified CPEs surface was analyzed by Scanning Electron Microscopy (SEM) and

Atomic Force Microscopy (AFM) to determine its morphological characteristic.

Energy Dispersive X-ray (EDX) analysis of water hyacinth MCPE was carried out

to check the accumulation of metals at electrode surface. It was done after open

circuit accumulation of metal ions for 10 min in stirring condition.

Scanning Electron Microscope instrument (Zeiss EV 040) [AIRF JNU]

Atomic Force Microscope

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CARBON PASTE ELECTRODES



ray (EDX) analysis of water hyacinth MCPE was carried out

accumulation of metals at electrode surface. It was done after open

circuit accumulation of metal ions for 10 min in stirring condition.


ic Force Microscope instrument (Nanosurf easy scan 2) [Department of

Chemistry, DEI]




ray (EDX) analysis of water hyacinth MCPE was carried out

accumulation of metals at electrode surface. It was done after open


instrument (Nanosurf easy scan 2) [Department of

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CHARACTERIZATION OF CELLULOSIC NANO FIBERS (PREPARED FROM

RICE-STRAW) AND BLACK-RICE EXTRACT

Cellulosic nano fibers prepared from the cellulosic content of agricultural waste

(rice straw) were characterized by Department of chemistry for their structure,

morphology, size and functional groups by SEM, TEM, XRD and FTIR. The

preparation of cellulose was confirmed by FTIR studies. Black rice extract was

characterized for the presence of different anthocyanins by Mass Spectroscopy.

Transmisson electron microscope instrument (JEOL 2100 F) [AIRF JNU]

X-ray diffractometer (Bruker AXS D8 Advance, Germany) [Dept of Chemistry, DEI]

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ELECTROCHEMICAL CHARACTERIZATION OF PREPARED

MODIFIED CARBON PASTE ELECTRODES

Cyclic-voltammetry study

Cyclic Voltammetry (CV) of unmodified and modified carbon paste electrode

(CPE) was carried out to check the changes of the electrode behaviour after

modification. The experiment was carried out in 1 mM K3[Fe(CN)6] solution with

0.1 M KCl as supporting electrolyte. The peak potential difference and current

values of anodic and cathodic peaks of modified electrodes were determined and

compared with unmodified CPEs (Books et al 2006).

Potentiostat (PGSTAT 302 N and µ Autolab III) [Remote Instrumentation Lab, USIC, DEI]

Effective Surface area measurement

The effective surface area of the unmodified and modified electrodes was

determined according to the Randles-Sevcik equation (Rezaei et al. 2008). For this

CV was carried out in 1 mM K3 [Fe(CN)6] with 0.1 M KCl solution with increasing

scan rate. The Randles-Sevcik equation is-

Ip = 0.4463 n F A C ( n F ν D / R T)1/2 eq 1

In this equation, n is the number of electrons appearing in the half-reaction for the

redox couple, ν is the rate at which the potential is swept (V/sec), F is Faraday’s

constant (96485 C/mol), A is the electrode area (cm2), C is the concentration of the

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probe molecule in the bulk solution (mol cm-3), R is the universal gas constant

(8.314 J/mol K), T is the absolute temperature (K), and D is the analyte’s diffusion

coefficient (cm2/sec). At temperature 25˚C (298.15 K), for a reversible reaction the

Randles-Sevcik can be written in a more concise form-

Ip = (2.69 × 105) n 3/2 ACD1/2 ν½ eq 2

Accordingly, the current is directly proportional to the concentration and increases

with the square root of the scan rate. To determine the effective surface area cyclic

voltammograms are recorded in 1 mM K3[Fe(CN)6] solution. The anodic or

cathodic current (Ip) values were plotted against the square root of the scan rate

(ν½). The slope of the relation Ip Vs ν½ will be equal to (2.65 x 105) n 3/2 ACD1/2.

For 1 mM K3[Fe(CN)6] in the 0.1 M KCl electrolyte: n=1 and D=7.6 x10-6 cm s-1

which are constant. By substituting the values of n, C and D, the effective surface

areas can be calculated.

Surface activation and Potential window

Pre-treatment was carried out to activate the surface of the prepared electrodes. The

pre-treatment of the modified CPEs was carried out by doing cyclic voltammetry in

0.1 M HCl within the range of -1 V to + 1 V at the scan rate 50 mV/S (Kalcher et al

2006). Cyclic Voltammetry was carried out for 10 cycles. In this constant current

response the potential range in which modified electrode didn’t show any oxidation

or reduction peak is termed as potential window.

ELECTROCHEMICAL IMPEDANCE STUDY

Electrochemical impedance study of the unmodified and modified CPEs were

carried-out in K3[Fe(CN)6] with 0.1 M KCl as electrolyte (Wang et al 2009). For

electrochemical impedance study AC amplitude of 10 mV at a frequency of

1-100000 Hz was applied and the corresponding Nyquist and Bode plot were

measured. Related parameters were measured with equivalent electrochemical

circuit fit.

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DETERMINATION OF METAL IONS

Electrochemical determination of metals was carried out by stripping voltammetry

using differential pulse as an input wave. The step by step procedure for the

determination of metals is as follows-

1. Ability to pre-concentrate metal ions at electrode

Ability to pre-concentrate metal ions at electrode surface was carried out by CV in

0.1 M HCl after accumulation of metal ions at electrode surface. A peak was

observed in the cyclic voltammogram depending upon the oxidation or reduction of

the accumulated metal ions. From the cyclic voltammograms it would be clear

whether the modified electrode is able to determine particular metal ion or not.

Depending on the CV study, direction of differential pulse study has been decided.

For black rice extract modified CPE accumulation was carried out at -1.2 V where

as for other modified CPEs accumulation was carried out at open circuit.

2. Anodic Stripping Voltammetry

Differential pulse anodic stripping voltammetry (DPASV) was used to determine

metal ions using MCPEs. Each DPASV run was made up of two steps- First

accumulation followed by stripping.

Determination of lead, cadmium, copper and zinc using Black rice extract

modified carbon paste electrodes

For black-rice extract modified carbon paste electrode accumulation was carried

out by keeping the electrode in metal solution at a negative potential (-1.2 V) so that

the desired metal ion get reduced and accumulated at the electrode surface.

Stripping was carried out in the same solution. For stripping a differential pulse

voltammograms was recorded from a negative potential (-1.2 V) to positive

potential (+0.5 V). In this scanning when the potential reaches to the oxidation

potential, the accumulated metal gets oxidized and strip-out from the electrode

surface. These voltammograms were used for qualitative and quantitative

determination of the metals.

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Figure 3.2 Schematic diagram for the determination of metal ions after reducing them

at negative potential

Determination of lead, cadmium and mercury using Water hyacinth, Coconut

shell, Cellulose nano fibers and lotus-root modified carbon paste electrodes

Open circuit accumulation and MEX approach (Svanca et al. 2012) was used for

these modified CPEs. Each DPASV run was made up of two steps: first open circuit

accumulation followed by medium exchange and stripping. For accumulation the

MCPEs was kept in stirred metal solution for a definite period of time called as

accumulation time/deposition time. In this step metal ions get reduced and

accumulated at the electrode surface. After accumulation the electrode was rinsed

with distilled water, followed by medium exchange for stripping analysis. In

stripping step a potential scan was applied to it. In this potential scan we obtain a

anodic peak corresponding to the oxidation of the accumulated metal. The

schematic diagram for the voltammetric determination of metals is given in Figure

3.3.

Figure 3.3: Schematic diagram of step by step procedure for the determination of

metal ions after open circuit accumulation

M+ M+ M+ M+

M+

M+

Activation Accumulation in metal solution

Washing Determination of metal ions in 3 electrode cell

Accumulation at -1.2 V Stripping

Equilibrium for 30 sec

M+ M+

M+

M+ M+

M+

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The range of potentials and the direction of scan were selected on basis of the

previous CV study. Since lead, cadmium, mercury, copper and zinc show anodic

peak in CV at the potential -0.45 V, -0.76 V, 0.1 V, -0.1 V and 1 V respectively for

Pb (II), Cd (II), Hg (I), Cu (II) and Zn (II) hence, the potential scan was carried out

in anodic direction from -1 V to + 0.5 V. In this voltammograms the peak was

observed at a particular potential which is characteristic of a particular metal. The

differential pulse voltammograms were recorded and used for qualitative and

quantitative determination.

OPTIMIZATION OF PARAMETERS

Different parameters for electrochemical determination of metals were optimized

(in terms of maximum current values) to get the most favorable experimental

conditions with respect to amount of modifier, different electrolytes, concentration

of electrolyte, pH of electrolyte, deposition potential and deposition time.

EXPERIMENTAL CONDITIONS AT A GLANCE

AMOUNT OF MODIFIER

METAL CONCENTRATION

TEMPERATURE

pH VALUE SUPPORTING ELECTROLYTE

ACCUMULATING SOLVENT ACCUMULATION TIME

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FTIR STUDIES

FTIR studies were carried out of the plant materials, native and after treatment with

these metal ions.

Fourier Transform Infra red spectrometer (Varian 7000) [AIRF JNU]

EFFECT OF INTERFERING IONS

To see the effect of other metal ions on the determination of lead, cadmium,

mercury, copper and zinc, accumulation of these metal ions were carried out in

presence of the increasing concentration of the other metal ion with keeping the

concentration of the determining metal ion constant. The decrease in the current

response for the metal ion with the increase in the other metal was determined.

EFFECT OF SURFACTANTS

Triton X-100, Sodium dodecyl sulfate (SDS) and Cetyl trimethylammonium

bromide (CTAB) were selected as representative of nonionic, anionic and cationic

surfactants to study the effect of surfactants on the determination of metal ions.

Metal ions of a fixed concentration was determined in the presence of increasing

amounts of surfactants. The effect of the increasing amount of surfactant on the

determination of particular metal ion was determined. The electrode surface was

renewed for each surfactant.

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EFFECT OF TEMPERATURE

The effect of temperature on the voltammetric determination of metals was

determined by varying the temperature of the accumulating metal solution within

the range from 7.5 ˚C to 50 ˚C.

ANALYTICAL CHARACTERISTICS

Analytical performance of all the plant modified CPEs were evaluated with the

increasing concentrations of metal ions under the optimum conditions determined

prevoiusly. The current values were plotted with the concentration of metal ions and

linear working range between the concentration and current value was determined.

For statistical analysis limit of detection and limit of quantification were

determined.

APPLICATION OF CNT AND GRAPHENE AS ELECTRODE MATERIAL

To see the effect of other carbonaceous materials carbon nanotubes and graphene,

20% of graphite powder was replaced with CNT and graphene separately for the

optimized amount of modifier for the MCPEs by the same procedure as used for the

preparation of modified carbon paste electrodes. Amount of mineral oil 150 µl was

used in place of 100 µl. Similarly the calibration curves were plotted for the MCPEs

with CNT and graphene for determination of metals and for the statistical analysis.

Limit of detection and limit of quantification were determined.

ELECTRODE STORAGE, SURFACE REGENERATION AND STABILITY

The electrodes were stored in desiccators for first 24 hours, and then the electrodes

were stored in the refrigerator. For a new experiment the electrode surface was

renewed by pushing the electrode material from back side, a small amount of paste

was removed and polishing the tip on a photo paper to get the surface smoothed.

After smoothing the surface of the electrode it becomes shiny. In order to do the

analysis with the same electrode, the electrodes were stored in 0.1 M HCl solution

during the experiment. Stability of the prepared MCPEs were determined by

repeated measurement of a fixed concentration of metals upto 9 months.

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References

Alemdar A and Sain M (2008) Isolation and characterization of nanofibers from agricultural residues – Wheat straw and soy hulls. Bioresource Technology, 99: 1664–1671.

Books LS, Harris C and Kasem KK (2006) Electrochemical Behavior of Hexacyano Iron (III/II) in Frozen Aqueous Electrolytes. American Journal of Undergraduate Research, 5(4): 25-32.

Kalcher K., I. Švancara , R. Metelka , K. Vytřas , A. Walcarius , in The Encyclopedia of Sensors, Vol. 4 (Eds: C. A. Grimes, E. C. Dickey, M. V. Pishko), American Scientific, Stevenson Ranch 2006, pp. 283–430.

Rezaei B and Damiri S (2008) Multiwalled carbon nanotubes modified electrode as a sensor for adsorptive stripping voltammetric determination of hydrochlorothiazide. Sensors 8(9):1523–1529

Rodriguez-Saona LE and Wrolstad RE (2011) Current Protocols in Food Analytical Chemistry, John Wiley & Sons, F1.1.1-F1.1.11.

Serrano N, Martin N, Diaz-Cruz JM, Arino C and Esteban M (2009) Bismuth Film Electrode in Metal Complexation Studies: Stripping Analysis of the Pb(II)-, Cd(II)-, and Zn(II)-Binding with Phthalate. Electroanalysis 21, (3-5):431 – 438

Svancara I, Kalcher K, Walcarius A and Vytras K (2012) Electroanalysis with Carbon Paste Electrode, Page No 20-22, 188-190.CRC Press. Wang B, Sain M (2007) Isolation of nanofibers from soya bean source and their reinforcing capability on synthetic polymers. Composites Science and Technology, 67: 2521-2527. Wang F, Liu J, Wu Y J, Gao Y and Huang XF (2009) Anodic Stripping Voltammetric Determination of Mercury(II) in Water Using a 4-tert-Butyl-1-(ethoxycarbonylmethoxy)thiacalix[4]arene Modified Glassy Carbon Electrode. Journal of the Chinese Chemical Society, 56:778-784.

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