American Journal of Anal yt ical Chemistry, 2011, 2, 697-709
doi:10.4236/ajac.2011.26080 PublishedOnline October 2011 (
Copyright © 2011 SciRes. AJAC
A New Thioamide Derivative for Separ atio n a nd
Preconcentration of Multi Elements in Aquatic
Environment by Cloud Point Extraction
Mohamed M. Hassanien1,2*, Ali M. Hassan3, Wael I. Mortada4, Ahmed A. El-Asmy5
1Chemistry Department, Industrial Education College, Beni-Suef University, Beni-Suef, Egypt
2Chemistry Department, Faculty of Science, Al-Jouf University, Saudia, Arabia
3Chemistry Department, Faculty of Science, Al-Azhar University, Cairo, Egypt
4Clinical Chemistry Department, Urology and Nephr ol o gy Ce nt er, Mansoura University, Mansoura, Egypt
5Chemistry Department, Faculty of Science, Mansoura University, Mansoura, Egypt
E-mail: *
Received May 27, 2011; revised July 20, 2011; accepted August 10, 2011
2-(pyridine-2-yl)-N-p-chlorohydrazinecarbothioamide (HCPTS) was synthesized, characterized and suc-
cessfully applied for the preconcentration of Cu(II), Ni(II), Zn(II), Cd(II), Co(II), Pb(II), Fe(II), and Hg(II) in
water, blood, and urine samples prior to graphite furnace atomic absorption determination (GFAAS); Hg was
determined by cold vapor technique. Under the optimum experimental conditions (i.e. pH = 8, 10–4 M of
HCPTS, 0.05% w/v of Triton X-114), calibration graphs were linear in the range of 0.02 to 200 ng·mL–1 for
Co(II), Cd(II), Pb(II) and Ni(II); 0.03 to 200 ng·mL–1 for Cu(II); 0.07 to 200 ng·mL–1 for Fe(II) and Zn(II)
and 0.02 to 150 ng·mL–1 for Hg(II). The enrichment factors were 43, 51, 41, 46, 54, 40, 45 and 52 for Cu(II),
Ni(II),Zn (II), Cd(II), Co(II), Pb(II), Fe(II), and Hg(II), respectively. The limit of detection were found to be
0.019, 0.094, 0.0514, 0.052, 0.0165, 0.047, 0.068 and 0.041 ng·mL–1 for Cu(II), Ni(II), Zn(II), Cd(II), Co(II),
Pb(II), Fe(II), and Hg(II), respectively. The developed method was applied to the determination of these
metal ions in water, blood and urine samples with satisfactory results.
Keywords: Heavy Metals, Preconcentration, Cloud Point Extraction,
1. Introduction
Heavy metals are of the great importance for the life.
Some of these such as Cu, Zn, Co, Fe are essential to
humans [1]. Others such as Hg, Cd and Pb are toxic fol-
lowing occupational and environmental exposure [2].
Due to the low concentration of these metals in the envi-
ronmental and biological samples and interfering effects,
a preconcentration/separation technique is generally ne-
cessary prior to the determination. For this purpose,
various analytical procedures have been used, such as
adsorption on activated carbon [3,4] co-precipitation [5,6]
column extraction [7,8] ion-selective electrode [9,10]
liquid–liquid extraction (LLE) [11] and cloud-point ex-
traction (CPE) [12-22].
CPE has attracted considerable attention mainly be-
cause it complies with the “Green Chemistry” principle
[23], as the amount of organic solvent is much less than
that of traditional liquid extraction. Moreover, It is sim-
ple, cheap, highly efficient, fast, and of lower toxicity
than those extractions that use organic solvents. This
method has been applied for extraction and preconcen-
tration of some metal ions such as Cu, Ni, Co and Zn in
water samples [12], Ag, Zn, and Pb in environmental
samples [13], Mo in sea water [14], Cd, Pb, Pd and Ag in
environmental samples [15], Cr, Pb, Cu, Ni, Bi, and Cd
in environmental samples [16], Cd, Cu, Co and Ni in
water [17], U, Th, Zr and Hf in aqueous samples [18], Be
in water [19], La, Eu and Lu [20], Rh [21] and Yb, Gd,
Eu, Sm, Sc, Ho in biological samples [22].
The cloud point is the temperature above which aque-
ous solutions of non-ionic and zwitter ionic surfactants
become turbid. Specifically, above the cloud point the
solution is separated into two phases: a rich phase con-
taining a high surfactant concentration in a small volume
and a poor phase with a surfactant concentration close to
the critical micelle concentration [24,25]. CPE enables to
avoid hazardous organic solvents and allows achieving a
much higher concentration of recovered metal ions than
in the case of liquid-liquid extraction, because the micel-
lar phase volume is about 10 - 100-fold less than the vol-
ume of an aqueous phase [24]. Accordingly, any metal
ions that either directly interact with micelles or after
prerequisite binding with hydrophobic chelating ligand,
can be extracted from the parent solution by CPE proce-
dure. Trace elements can be extracted to the surfac-
tant-rich phase usually after formation of a hydrophobic
complex with an appropriate chelating agent [25]. The
small volume of the surfactant-rich phase obtained with
this methodology permits the design of extraction schemes
that are simple, cheap, highly efficient, fast, and of lower
toxicity than those extractions that use organic solvents.
Thiosemicarbazides constitute an important class of
NS donors. The chemistry of these compounds was early
explored [26] for their variable donor properties, struc-
tural diversity and biological applications. The coordina-
tion chemistry of a number of metal ions and a wide va-
riety of complexes has been reported [27-29]. Thiosemi-
carbazides have been used for extraction and determina-
tion of some metal ions in biological and pharmacologi-
cal samples [30-32]. In recent studies, 4-Ethyl-1-(pyri-
din-2-yl)thiosemicarbazide and its Cu(II) complexes with
different anions have been synthesized and characterized
[28] and used as a chelating agent for separation, pre-
concentration, and determination of Cu(II) ions in satu-
rated saline solutions by a cloud point extraction tech-
nique [33]. In addition to, complexes of 4-ethyl and
4-(p-tolyl)-1-(pyridin-2-yl)thiosemicarbazides with Pd(II),
Pt(II) and Ag(I) were synthesized and characterizes [29]
These complexes show antibacterial activity to some
Gram positive and Gram negative bacterial strains.
In the present study, we have developed and optimized
a simple CPE-GF AAS methodology for multi-elements
determination in water, blood and urine samples, which
shows rapid and efficient performance. Copper(II), Ni
(II), Zn(II), Cd(II), Co(II), Pb(II), Fe(II), and Hg(II)
were preconcentrated using Triton X-114 in the presence
of 2-(pyridine-2-yl)-N-p-chlorohydrazinecarbothioamide
(HCPTS) (Structure 1) as a new chelating agent.
2. Experimental
2.1. Reagents and Solutions
All chemicals were of analytical-reagent grade and were
used without previous purification (A.R. from Fluka,
Aldrich or Merck). All solutions were prepared using
Structure 1. HCPTS.
double distilled water. Metal ion solutions (1000 mg/l)
were prepared by dissolving appropriate amounts of the
sulphates, chlorides or nitrates in double-distilled water.
Working reference solutions were prepared daily by
stepwise dilution from stock solution. Triton X-114 was
used without further purification. HCPTS solution (1 ×
10–3 M) was prepared by dissolving 0.0278 g in 100 mL
distilled water. Buffers were used to control the pH of
the solutions: hydrochloric acid-glycine (pH 1 - 3), hexa-
mine-nitric acid (pH 4 - 8), ammonium chloride-am- mo-
nium hydroxide (pH 9, 10). Solutions of alkali metal salts
(1%) and various metal salts (0.1%) were used in order
to test the interference of anions and cations, respectively.
2.2. Preparation of the Ligand
HCPTS (Structure 1) was prepared by heating under
reflux a mixture of 2-hydrazinopyridine (0.1mol) and 4-
chlorophenylisothiocyanate (0.1 mol) in 20 ml absolute
ethanol for 2 h. On cooling, white fine crystals were
formed, filtered off, washed with EtOH and Et2O and
recrystallized from EtOH (m.p. 175˚C; yield 96%). The
purity of the compound was checked by TLC. Elemental
analysis Calculated for C12H11N4SCl: C 51.70, H, 3.98;
N, 20.10.and the found: C, 51.23, H, 3.75; N, 19.92.
2.3. Apparatus
Elemental analysis was carried out using a Perkin Elmer
Elemental Microanalyser. Infrared spectrum of the ligand
was measured using KBr discs on Mattson 5000 FTIR
spectrometer. Calibration of the frequency reading was
made with polystyrene film. The electronic spectrum of
the ligand in DMF was recorded by Unicam UV-Vis
spectrometer UV2 using 1 cm stoppered silica cells.
Thermogravimetric analysis measurement (TG) of the
ligand was performed using an automatic recording
thermobalance type (951 DuPont instrument). Sample
was subjected to heat using a rate of 10˚C/min from
room temperature to 800˚C in nitrogen flow. The mass
spectrum of the ligand was recorded on 70 eV with a
Varian MAT 311 instrument. 1H NMR spectrum of
HCPTS, in d6-DMSO, was recorded on EM-390 (200
MHz) spectrometer.
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GFAAS measurements were carried out using A Per-
kin Elmer atomic absorption spectrophotometer (Model
Analyst 800) with a longitudinal Zeeman background
correction furnished with a transversely heated graphite
atomizer (THGA) was used for the determination of the
metal ions, except Hg. Sample solutions were injected
into the atomizer by using AS-800 autosampler. The
sample injection volume was 20 µL and the modifier “5
µg Pd/3 µg Mg(NO3)2” volume was 10 µL. The system
is equipped with winLab 32 software. Hg was deter-
mined by cold vapor technique (CVAAS). The operating
conditions are given in Tables 1 and 2.
The pH of the solution was adjusted using Hanna in-
strument model 8519 digital pH meter. A centrifuge with
a model of CH90-2 (Hinotek Technology Co., Ltd. China)
was used to accelerate the phase separation process.
2.4. Procedure of CPE
For the CPE, an aliquot of 50 mL of a solution contain-
ing metal ion buffered with pH 8, Triton X-114 (0.05%
w/v) and 10–4 M HCPTS were kept for 10 min in a ther-
mostatic bath at 40˚C. Subsequently, separation of the
phases was achieved by centrifugation for 10 min at
4000 rpm. The phases were cooled down in an ice bath
in order to increase the viscosity of the surfactant rich
phase. The bulk aqueous phase was easily decanted sim-
ply by inverting the tube and dried in water bath. The
surfactant-rich phase in the tube was made up to 1 mL by
adding absolute methanol/conc. HNO3 mixture (5:1).
2.5. Preparation of Real Samples
2.5.1. Water S ampl e
The water samples were filtered firstly through filter
paper to separate the coarse particles and suspended mat-
ter and secondly through a Millipore cellulose nitrate
membrane (pore size 0.45 µm), acidified to pH 2 with
HNO3 and stored in a refrigerator in a dark polyethylene
2.5.2. Blood a nd Uri ne S am pl es
One mL of blood or 5 mL of urine was digested with 6
mL [HNO3 (65%) + HClO4 (70%)] (2:1) in 50 mL
beaker covered with a watch glass. The content of the
beaker was heated gradually on a hot plate till dryness.
Table 1. Instrumental parameters and temperature program for metal ion analysis by GFAAS.
(a) Instrumental parameters:
Fe(II) Pb(II) Co(II) Cd(II) Zn(II) Ni(II) Cu(II) Parameter
248.3 283.3 242.5 228.8 213.9 232 324.8 Wavelength (nm)
0.7 0.7 0.7 0.7 0.7 0.2 0.7 Slit width (nm)
25 25 15 12 25 20 25 Lamp current (mA)
(b). Temperature program:
Time (s) Temperature (˚C) Argon flow rate ml/min
HoldRampFe(II)Pb(II)Co(II)Cd(II)Zn(II) Ni(II)Cu(II)
250 30 1 110 Drying 1
250 30 15 130 Drying 2
250 20 15 1400850 1400 500 700 11001200 Pyrolysis
0 6 0 21001600 2400 1500 1800 23002000 Atomization
250 5 1 2500 Cleaning
Table 2. Instrumental parameters for the determination of Hg by CVAAS.
Source Hg hollow cathode lamp.
Slit width (nm) 0.7
Carrier gas flow rate (ml/min) 1000
Reducing agent 1% m/v NaBH4 in 0.05 % m/v NaOH
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All heating steps were carried out under a hood with
necessary precautions. After increasing the pH to about 3
using 1 M NaOH, the volume was completed to 10 mL
volumetric flask. Aliquots of 5 mL of the digested sam-
ples were analyzed according to the prescribed proce-
3. Result and Discussion
3.1. Characterization of the Ligand (HCPTS)
The IR spectrum of HCPTS exhibits characteristic bands
at 3262, 3172 and 3131 cm–1 attributed to ν(N4H), ν(N1H)
and ν(N2H), respectively [28]. The ν(C=N)Py appearing
as a strong band at 1598 cm–1. The appearance of the
(C=N)Py and N2H at lower values suggests intramolecu-
lar hydrogen bonding [29]. Also, two bands at 1512 and
1450 cm–1 were observed and assigned to thioamide I
and III, respectively. Moreover, two sharp bands at 783
and 723 cm–1 were attributed to ν(C=S) and ρ(NH) [28].
The absence of any band due to ν(SH) in the range 2500 -
2600 cm–1 [28] indicated that the ligand exists in the
thione form.
The absorption spectrum of HCPTS in DMF showed a
band at 35,714 cm–1 and three shoulders at 31,250, 24,390
and 23,148 cm–1 attributing to (π→π*)Py, (π→π*)C=S,
(n→π*)Py and (n→π*)C=S [29] transitions, respectively.
The TGA curve of HEPTS indicates a thermal stability
till 167˚C coincident with its melting point (175˚C). The
curve showed two decomposition steps. The first from
167 to 219˚C is corresponding to the loss of C5H4N and
ClC6H4 (Found 68.2%; Calcd. 68%), the second from
219 to 347 is attributed to the loss of N3H3 (Found 15.9%,
Calcd. 16.1%), leaving a residue of C + S (Found 15.9%,
Calcd. 15.8%).
The 1H NMR spectrum of HCPTS in d6-DMSO sh-
owed signals due to the protons of the phenyl and pyri-
dine [29]. The three singlet signals at 8.54, 9.79 and 9.91
ppm are attributed to the N2H, N1H and N4H protons,
respectively. The N2H signal appears in the aromatic
region confirming an intramolecular H-bond. The other
NH protons are shifted up field due to the inductive ef-
fect of pyridyl and phenyl rings as electron donor groups
The mass spectrum of HCPTS shows the final peak at
280 amu [(C12H11N4SCl), calculated molecular weight
278.76 amu], and other peaks at 171, 111 and 77 amu
may correspond to various fragments. The peak described
at 171 amu is assigned to the fragment [C7H5NSCl]+,
corresponding to the loss of [C5H6N3]+. The peak at 111
is corresponding to the fragment [C6H4Cl]+, correspond-
ing to the loss of [CHNS]+ fragment. Peak at 77 repre-
sent the fragment C6H4 with the loss of Cl.
3.2. Effect of pH on CPE
The formation of metal complexes and its stability are
important factors for the CPE. The pH plays a unique
role on metal chelate formation and subsequent extrac-
tion. The extraction depends on the pH at which complex
formation occurs. The effects of the pH on the percent-
age extraction recovery of Co(II), Hg(II), Cd(II), Fe(II),
Pb(II), Ni(II), Zn(II) and Cu(II) were investigated at dif-
ferent pH (1 - 10). Where the percentage extraction re-
covery is the percentage of the concentration of metal
found in spiked sample to the actual concentration. The
results are shown in Figure 1. It was observed that Pb(II)
and Cu(II) were extracted quantitatively (>95%) at pH 5;
Co(II), Ni(II) and Zn(II) at pH 6; Hg and Fe(II) at pH 7;
Cd(II) pH 8. At lower pH values, the extraction is not
quantitative, and at higher pH values, the hydrolysis of
cations occurs except for Co and Cd(II) a result, pH 8
was chosen as the working pH for the multi determina-
tion of these metals.
3.3. Effect of HCPTS Concentration
The extraction recoveries of the metals depend on the
concentrations of the chelating agent. In order to select
the optimum concentrations of HCPTS, the effects of the
concentration of the HCPTS on the recoveries were in-
vestigated in the range of 1 × 10–5 to 1.6 × 10–4 M; the
results are shown in Figure 2. For all metals, the maxi-
mum recoveries were obtained at 10–4 M of HCPTS.
Therefore, these concentrations were selected as the op-
timum concentration of the chelating agent.
Recovery %
Figure 1. Influence of the pH on the recovery of extraction
for the metal ions. Conditions: 10 ng·mL–1 each metal; 50
mL cold aqueous solution; 10-4 mol·L–1 of the ligand; 0.05%
w/v Triton X-114; 40˚C.
0.00 0.02 0.04 0.06 0.08 0.10 0.12 0.14 0.16 0.18
Recovery %
Concentration of Li
and mM
Figure 2. Effect of the ligand concentration on the recovery
of extraction for the metal ions. Conditions: 10 ng·mL–1
each metal; pH, 8; 50 mL cold aqueous solution; 0.05% w/v
Triton X-114; 40˚C.
3.4. Effect of Triton X-114 Concentration
The non-ionic surfactant, Triton X-114, was chosen be-
cause of its commercial availability in a high purified
homogeneous form, low toxicological properties and cost.
Also, the high density of the surfactant rich phase facili-
tates phase separation by centrifugation. Additionally,
the cloud point (23˚C - 26˚C) of Triton X-114 permits its
use in the extraction and/or preconcentration of a large
number of molecules and chelates. The variation of the
recoveries as a function of the concentration of Triton
X-114 in the range of 0.01 to 0.1% (w/v) is shown in
Figure 3. A concentration of 0.05% (w/v) was chosen as
optimum concentration for the extraction of the all metal
ions. At lower concentrations, the extraction recovery of
the complex is low probably because of the inadequacy
of the assemblies to entrap the hydrophobic complex
3.5. Effect of the Equilibration Temperature and
the Centrifugation Time
In order to optimize the method, it was necessary to ex-
amine the effect of the temperature on cloud-point ex-
traction. It was desirable to employ the lowest possible
equilibration temperature and the shortest incubation
time of extraction for efficient separation. For this pur-
pose, the effect of temperature and equilibration time on
the extraction recovery was investigated in the range of
30˚C - 90˚C and 5 - 35 min, respectively, while keeping
all other parameters constant. It is desirable to employ
low equilibration temperature, as a compromise between
the completion of extraction and efficient separation of
phases. Excellent recovery was found at 40˚C - 60˚C.
Above this temperature, reduction of extraction effi-
ciency was noted. Therefore, 40˚C was chosen for the
0.00 0.02 0.04 0.06 0.08 0.10
Recovery %
Concentration of Triton X-114
(% w/v)
Figure 3. Effect of Triton X-114 concentration on the re-
covery of extraction for the metal ions. Conditions: 10
ng·mL–1 each metals; pH 8; 50 mL cold aqueous solution;
10–4 mol·L–1 of the ligand
equilibration temperature. The results also show that
incubation time of 10 min is adequate to achieve the
highest extraction efficiency.
3.6. Effect of Centrifugation Time and Rate
Generally, centrifugation time and rate ever affect mi-
celle formation but accelerate the separation. The effect
of the centrifugation time on the extraction efficiency
was studied in the time range of 5.0 - 25.0 min at 4000
rpm. A centrifugation time of 10.0 min was selected, since
the analyte extraction is almost quantitative. No appre-
ciable improvements were observed for longer times.
3.7. Effect of Ionic Strength
The cloud point of micellar solutions can be controlled
by addition of salts, alcohols, nonionic surfactants and
some organic compounds (salting-out effects). To date,
most of the studies conducted have shown that ionic
strength has no appreciable effect on the extraction effi-
ciency. An increase in the ionic strength in the cloud
point extraction does not seriously alter the efficiency of
extraction of the chemical forms. Moreover, the addition
of a salt can markedly facilitate the phase-separation
process, as demonstrated with some nonionic surfactant
system, since it alters the density of the bulk aqueous
phase [24]. In our study, it was observed that the addition
of KI within the interval of 0.1 - 1.0 mol/L had no sig-
nificant effect on the cloud point extraction efficiency.
3.8. Effect of Methanol
Since the surfactant-rich phase obtained after the cloud
point preconcentration contains a high concentration of
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Triton X-114 and, at the same time, the volume obtained
is rather small (0.5 mL), absolute methanol/conc. HNO3
mixture (5:1) was added to the surfactant rich phase after
phase separation. Moreover, it was necessary to decrease
the viscosity of the rich phase without excessive dilution
of the micelle to facilitate the introduction of the sample
into the atomizer of the spectrometer. There is an opti-
mum volume (1 mL) with respect to the metal ion recov-
ery. Larger volumes of acidified methanol dilution are
clearly predominated resulting in a gradual absorbance
reduction. One mL volume of methanol was therefore
used throughout the remaining experiments.
3.9. Analytical Characteristic of the Method
The characteristics of the proposed cloud point extraction
procedure for Co(II), Hg(II), Cd(II), Fe(II), Pb(II), Ni(II),
Zn(II) and Cu(II) are given in Table 3. Calibration
curves were obtained using 50 ml of the standard solu-
tion of the metal ion under the experimental conditions
specified in procedure. The GFAAS signals were found
to be a linear function of concentration range [0.02
ng·mL–1 for Co(II), Cd(II), Pb(II) and Ni(II); 0.03
ng·mL–1 for Cu(II); 0.07 ng·mL–1 for Fe(II) and Zn(II) to
200 ng·mL–1 for all ions], while the linear concentration
range of HGAAS was 0.02 to 150 ng·mL–1 for Hg(II).
Table 3 gives the parameters of the calibration curves
and the relative standard deviations (RSD) obtained for 3
replicates subjected to the complete procedure. The cor-
relation coefficient (r) of the calibration curves are
within 0.990 to 0.999, which indicates good linearity in
the studied concentration range.
The limits of detection (LOD) were calculated as three
times (3s) of standard deviation of blank signal by 7 rep-
licate measurements. All the metals achieved very low
detection limits in the range of 0.0165 - 0.094 and is bet-
ter than the previous procedures (Tabl e 4 ); indicating the
high sensitivity of HCPTS to the investigated metal ions.
The preconcentration factor for all metal ions were
calculated by dividing the aqueous phase volume to the
final volume of preconcentrated phase is 50, while the
enrichment factor as the ratio of slope of the calibration
curve obtained from preconcentrated samples to that ob-
tained without preconcentration for Cu(II), Ni(II), Zn(II),
Cd(II), Co(II), Pb(II), Fe(II), and Hg(II), were 43, 51, 41,
46, 54, 40, 45 and 52, respectively. This indicates that
the extraction using the proposed procedure is quantita-
tive for all metal ions. Furthermore, the determination of
Cu(II), Ni(II), Zn(II), Cd(II), Co(II), Pb(II), Fe(II), and
Hg(II) by the proposed method was compared with other
CPE-AAS or CPE-ICP methods [12,15,34-38] and the
results are summarized in Table 4 . As it can be seen the
proposed method has higher preconcentration factor and
lower LOD.
3.10. Effect of Interfering Ions
Under the optimized conditions determined for this study,
the percentage removal of 20 ng·mL–1 of Cu(II), Ni(II),
Zn(II), Cd(II), Co(II), Pb(II), Fe(II), and Hg(II) were
studied in the presence of high concentrations of various
cations and anions. All of the cations were used as their
chlorides or sulfates, whereas the anions were used as the
corresponding sodium salts. An ion was considered to be
an interferent when it caused a variation greater than
±5% in the absorbance of the sample. The corresponding
results obtained are listed in Table 5. The results ob-
tained indicates that the removal of Cu(II), Ni(II), Zn(II),
Cd(II), Co(II), Pb(II), Fe(II), and Hg(II) ions was quanti-
tative in all cases. Moreover, 1000 fold of (K+, Na+, Cl,
, 2
); 750 fold of (Citrate Thiocyanate Thiourea
Thiosulphate Acetate); 500 fold of (Ca2+, Mg2+, Ba2+,
Sr2+, oxalate, phosphate, I) and 50 fold of (Mn2+, B3+,
Al3+, Bi3+) have no effect on the extraction efficiency.
Table 3. Analytical characteristic of the method.
Hg(II) Fe(II) Pb(II) Co(II) Cd(II) Zn(II) Ni(II) Cu(II) Parameter
0.041 0.068 0.047 0.0165 0.052 0.0514 0.094 0.019 Limit of detection (ng·ml1)
2.76 4.1 3.61 2.19 1.64 3.14 2.61 3.35 Reliability (% RSD, n = 7)
0.02-150 0.07-200 0.02-200 0.02-200 0.02-200 0.07-200 0.02-200 0.03-200 Linearity
0.0141 0.0068 0.0027 0.0049 0.0624 0.0154 0.0087 0.0069 Slope of Calibration Curve
0.0002 10-6 9 X 10-5 7 X10-6 0.0038 3 X 10-5 3 X10-5 3 X 10-5 Intercept of Calibration curve
0.993 0.982 0.990 0.991 0.999 0.993 0.995 0.996 Correlation coefficient (r)
52 45 40 54 46 41 51 43 Enrichment Factor (EF)
Table 4. CPE applications for metal ions analysis of previous studies.
Ions Reagent Surfactant
system Matrix DL
(ng·mL–1)Linear range
(mL) EF Reference
Cu(II) 1.2 10-500 -
Zn(II) 1.1 10-700 -
Cd(II) 1.0 20-2000 -
4-(2-pyridylazo)-resorcinol Triton
X-114 ICP-OESwater
6.3 50-2500
Cd(II) 1.4 - 48
bis((1H-benzo [d] imidazol-
2yl)ethyl) sulfane
X-114 FAAS
radiology waste,
blood and urine 2.8 -
Zn(II) 8.8 8.8-80 13
Co(II) 4.9 4.9-3000 15.5
Ni (II) 7.8 7.8-2000 15
2-Guandinobenzimidazole Triton
X-114 FAAS water
11 11-6000
Cu(II) 1.4 10-250 35
Zn(II) 1.0 10-250 39
X-114 FAAS Blood, orange
juice and lotus tree
1.9 15-200
Cu(II) 0.45 2.5-25 18
Cd(II) 0.5 2.5-25 23
Co(II) 1.25 5-25 18
0.6 5-25 20
Cu(II) 0.44 2.5-25 20
Cd(II) 0.25 2.5-25 22
Co(II) 0.60 5-25 17
X-114 FAAS water
1.55 5-25
Ni(II) p-nitrophenylazoresorcinolTriton
X-114 FAAS Water and food 2.7 10-400 50 17[37]
1, 5-bis(di-2-
X-114 FAAS
water, food and
0.95 10-200 10 10.5[38]
Cu(II) 0.019 0.03-200 43
Ni(II) 0.094 0.02-200 51
Zn(II) 0.0514 0.07-200 41
Cd(II) 0.052 0.02-200 46
Co(II) 0.0165 0.02-200 54
Pb(II) 0.047 0.02-200 40
Fe(II) 0.068 0.07-200 45
HCPTS Triton
(CVAAS for
Water, blood
and urine
0.041 0.02-150
This work
Table 5. Tolerance limits of interfering ions.
Recovery %
Interfering ion Tolerance
limit Cu(II) Ni(II) Zn(II) Cd(II) Co(II) Pb(II) Fe(II) Hg(II)
K+, Na+, Cl, ,
SO 1000 99.7 ± 0.8 96.1 ± 2.197.2 ± 1.899.2 ± 1.598.4 ± 1.697.8 ± 2.4 98.1 ± 2.499.3 ± 3.2
Citrate, Thiocyanate, Thiourea,
Thiosulphate, Acetate 750 96.2 ± 1.2 99.4 ± 1.8100.2 ± 1.6100 ± 1.1100 ± 1.598.1 ± 0.8 96.9 ± 1.998.4 ± 1.8
Ca2+, Mg2+, Ba2+, Sr2+,
Oxalate, Phosphate, I 500 100.4 ± 0.699.2 ± 2.398.3 ± 2.4100.3 ± 2.198.2 ± 2.298.6 ± 1.6 96.2 ± 2.299.2 ± 2.1
Mn2+, B3+, Al3+, Bi3+ 50 97 ± 1.6 98.4 ± 0.997.4 ± 1.797.8 ± 3.1100.1 ± 2.296.7 ± 0.7 98.7 ± 1.997.6 ± 1.9
Copyright © 2011 SciRes. AJAC
Table 6. Recovery of trace metal ions from spiked real samples after application of presented procedure.
Sample Ion Added (ng·mL–1) Found (ng·mL–1) Recovery %
- 2.1
Cu 1.0 3.15
- 1.09
Ni 1.0 2.04
- 0.54
Zn 1.0 1.51
- 0.12
Cd 1.0 1.08
- 0.29
Co 1.0 1.32
- 1.7
Pb 1.0 2.66
- 32.6
Fe 10.0 43.1
- 0.2
Tap Water
Hg 1.0 1.16
- 924
Cu 100 1027
- 0.84
Ni 1.0 1.86
- 5,579
Zn 1000 6591
- 0.92
Cd 1.0 1.90
- 0.12
Co 1.0 1.11
- 55.6
Pb 10.0 65.8
- 556
Fe 100 660
- 2.43
Hg 1.0 3.44
- 34.8
Cu 10.0 45.1
- 2.45
Ni 1.0 3.46
- 375
Zn 100 478
- 0.84
Cd 1.0 1.85
- 1.21
Co 1.0 2.20
- 9.6
Pb 10.0 19.8
- 152
Fe 100 255
- 2.13
Hg 1.0 3.15
Copyright © 2011 SciRes. AJAC
Copyright © 2011 SciRes. AJAC
3.11. Accuracy and Precision
To check the applicability, reliability and accuracy of the
studied separation method, the proposed procedure has
been applied to determine Cu(II), Ni(II), Zn(II), Cd(II),
Co(II), Pb(II), Fe(II), and Hg(II) ions in spiked tap water,
blood and urine samples. The results are presented in
Table 6. The recovery values were around 100% in the
different samples. This indicates the capability of the
method in the determination of analytes in real samples
with high efficiency. In order to establish the validity of
the proposed method, the described procedure was ap-
plied for the determination of investigated ions in mul-
tielement standard solution 70006 (Fluka). The obtained
results assembled in Table 7 are in very good agreement
with the labeled values. The error of determination does
not exceed 1.5%.
3.12. Analysis of Real Water Samples
The CPE method was applied for the preconcentration
and separation of Cu(II), Ni(II), Zn(II), Cd(II), Co(II),
Pb(II), Fe(II) and Hg(II) in different water samples. Sur-
face water samples were collected from Nile River at
Mansoura, Miet Antar, Sherbin, and Faraskour; brackish
water at Damietta bridge, Elgerbi, Ezbet Elborg and El-
manzalah lake and Mediterranean Sea at Port Fouad and
Port Said. Table 8 shows the results of the application
process in comparison with the standard methods: direct
HGAAS in the case of Hg(II) and solvent extraction
(ammonium pyrrolidinedithiocarbamate in methyl iso-
butyl ketone) followed by ICP-MS determination in the
case of the other metal ions [39]. It can be concluded that
the concentration of the investigated metal ions increases
obviously in the region beginning from Damietta Bridge
to the River effluent and in Elmanzalah Lake. This may
be attributed to the domestic and anthropogenic activities
in these regions which are also shown in the water qual-
ity and nutrients measurements. It can be also concluded
that the concentrations of heavy metal ions are within the
permissible levels and are in agreement with those re-
ported previously [39-41].
3.13. Analysis of the Metals in Blood of
Hemodialysis Patients
Finally, the metal ions under investigation were deter-
mined in 13 hemodialysis patients and 11 matched
healthy persons (control group). For statistical calcula-
tions we used SPSS-PC software, version 8 (MAS
Medical & Scientific Eq. Co, IL, USA). Differences be-
tween groups were tested with the t-test for independent
samples. Correlation between variables was assessed
with Spearman’s rank correlation coefficient (r). A
probability value (p) less than 0.05 was considered to be
significant. The results summarized in Table 9 show that
the hemodialysis patients had normal blood levels of Co,
Ni and Hg. The blood levels of Cu, Zn and Fe were sig-
nificantly lower than the normal group. However, the
levels of Cd and Pb were higher than the control group.
These findings indicated that hemodialysis patients suf-
fer from deficiency of some essential metals as well as
they are at high risk to some toxic metals.
4. Conclusions
The developed CPE procedure is simple, rapid, safe and
provide high preconcentration factor and low detection
limits for the determination of ultra trace levels Cu(II),
Ni(II), Zn(II), Cd(II), Co(II), Pb(II), Fe(II), and Hg(II)
Table 7. Determination of metal ions in multielement standard solution 70006 (n = 3).
Metal ion Found by present method (mg·L–1) Certified value (mg·L–1)
Cu(II) 19.8 ± 1.1 20
Ni(II) 20.2 ± 0.9 20
Zn(II) 98.7 ± 2.2 100
Cd(II) 10.05 ± 0.4 10
Co(II) 9.87 ± 0.3 10
Pb(II) 40.2 ± 1.2 40
Fe(II) 100.5 ± 1.8 100
Hg(II) Not detected -
Table 8. Analysis of the metal ions (ng·mL-1)in natural water samples using CPE procedure in comparison with those obtained by ICP-MS after solvent extraction with
APDC/MIBK (direct HGAAS for Hg).
*BDL = below detection limit
Copyright © 2011 SciRes. AJAC
Copyright © 2011 SciRes. AJAC
Table 9. Metal ions levels (ng·mL1) in blood of hemodialysis (n = 13) and healthy persons (n = 11).
Metal Hemodialysis patients Normal persons
Cu(II) 715 ± 195* 895 ± 215
Ni(II) 0.69 ± 0.22 0.79 ± 0.19
Zn(II) 3954 ± 293* 5421 ± 452
*p < 0.05.
in water, blood and urine samples. This procedure agrees
with the Green Chemistry principles.
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