Research Journal of Environmental and Earth Sciences 4(7): 704-710, 2012
ISSN: 2041-0492
© Maxwell Scientific Organization, 2012
Submitted: May 08, 2012
Accepted: May 29, 2012
Published: July 25, 2012
Use of Sequential Extraction to Assess Copper Fractionation in Soil and Guinea
Corn from Kaduna Metropolis, Nigeria
S.S. Mohammed, M.B. Mohammed and N. Musa
Department of Applied Science, College of Science and Technology, Kaduna Polytechnic,
Kaduna, Nigeria
Abstract: In this study, sequential extraction technique was employed to determine the concentrations of copper in
Guinea corn and soil samples. The copper contents of guinea corn and soil were determined using Flame Atomic
Absorption Spectrometry (FAAS). The soil samples related to the guinea corn were digested and extracted using
different digestion and extraction reagents. The results revealed that the soil samples obtained from various locations
in Kaduna, contain varying amounts of Cu and was distributed between Residual, Oxide and Carbonate/Organic
fractions. The results also showed that in some of the sampling locations, the Cu concentration in the soil was above
the tolerable limit of 100 mg/kg and the ANOVA (p = 0.007<0.05), indicated a significant difference in the copper
concentrations across the various guinea corn crops. Similarly, the ANOVA (p = 0.114>0.05) showed that there is
no significant difference in the copper concentrations across the various guinea corn grown soils.
Keywords: Guinea corn, kaduna metropolis, sequential extraction, soil, type copper fractionation
research, the extractable Cu in soil samples was
determined by Flame Atomic Absorption Spectrometry
(FAAS) using sequential extraction technique. The soil
samples were extracted using the chemical reagents,
0.05 M EDTA, 1.0 M acetic acid and 1.0 M oxalic acid.
INTRODUCTION
Soils are receptacles for heavy metals released
from industrial activities, municipal wastes, water
sludge, urban composts, road traffic, atmospheric
deposits and chemicals used in agriculture (Phosphate
fertilizers, pesticides) and spread out into the
environment (Adriano, 1986). Heavy metals are
persistent in the environment; they are non-thermo
degradable and thus readily accumulate to toxic levels
(Sharma et al., 2007). Many soils especially those in
hazardous wastes sites are contaminated by heavy
metals such as Ni, Cu, Zn, Cu, etc. Heavy metal
contamination in arable soils through industrial and
anthropogenic activities is a serious problem in Nigeria.
Metals uptake by plants may pose risks to human health
when such plants are grown on or near contaminated
areas. Metals accumulation in plant depends on plant
species, growth stages, types of soil and metals, soil
conditions, weather and environment (Chang et al.,
1984; Petruzzelli, 1989).
Determination of metals in soil can be
accomplished via single reagent leaching, ion-exchange
resins and sequential extraction procedures. The
number of available extraction techniques developed
over the last three decades begs inquiry as to which
technique is preferable over another. A large number of
extracting solutions have been used to assess plant
available trace elements (Gupta and Aten, 1993; He and
Singh, 1993; Mohammed and Ayodele, 2011). In the
MATERIALS AND METHODS
A flame atomic absorption spectrophotometer
model 8010 Young Lin was used for the Cu
determination. In the extraction procedures, 1.0 M
oxalic acid, 0.05 M Na2EDTA and 1.0 M acetic acid
were used.
Preparation of samples: The research covered seven
agricultural sites in Kaduna, Nigeria. The sites are:
Nasarawa (NS), Sabon Tasha (ST), Unguwar Muazu
(UM), Tudun Wada (TW), Kakuri (KK), Mando (MD),
Kabala (KB) west and Kachia (KC). The samples were
collected during the harvest season (Oct-Nov., 2008,
2009 and 2010). The soil samples were collected from
the different areas enumerated at a depth of about 10
cm below the surface (Yaman et al., 2005). The cereal
samples were collected at each of the locations. Guinea
corn was chosen for the purpose of this research work
as it is the staple food being produced and consumed in
these areas. Kachia, a town situated about 130 km away
from Kaduna was taken as a control Fig. 1. The cereal
was thoroughly washed with water and allowed to drain
on a filter paper. Both the cereal and soil samples were
Corresponding Author: S.S. Mohammed, Department of Applied Science, College of Science and Technology, Kaduna
Polytechnic, Kaduna, Nigeria
704
Res. J. Environ. Earth Sci., 4(7): 704-710, 2012
Fig. 1: Map of Kaduna metropolis showing the sampling sites
705 Res. J. Environ. Earth Sci., 4(7): 704-710, 2012
dried at 85ºC. All the analyses were carried out in the
analytical laboratory of the department of Applied
Science, College of Science and Technology, Kaduna
Polytechnic, Kaduna-Nigeria
Wet ashing of cereal: Five (5) g of oven dried guinea
corn sample was accurately weighed into an
evaporating dish and ashed at 480ºC in an ashing
furnace for 4 h. Ten cm³ of a mixture of nitric acidhydrogen peroxide (2+1) was added to the ashed
sample and dried with occasional shaking on a hot plate
and cooled, 4 cm³ of 1.5 mol/L nitric acid was then
added and centrifuged. Sixty cm³ water was added to
the clear digest and was filtered. This was analysed for
Cu using FAAS model 8010 Young Lin. A blank digest
was carried out in the same way.
Digestion and extraction of soil: Soil pH was
measured (1:5, w/v) by digital pH meter. A modified
Tessier et al. (1979) extraction method developed by
Yaman et al. (2005) was used. Ten cm³ of a mixture of
nitric acid-hydrogen peroxide (2+1) was added to 5 g of
soil sample and dried with occasional shaking on a hot
plate and cooled. Four cm³ of 1.5 mol/L nitric acid was
added to the remainder, centrifuged and diluted to 60
cm³ with water and filtered. The clear digest was
analysed for Cu using FAAS model 8010 Young Lin. A
blank digest was carried out in the same way. Soil
extracts were obtained by shaking separately, 5 g of soil
samples with 10 cm³ of 0.05 mol/L Na2EDTA (for
carbonate and organically bound phases), 1.0 mol/L
oxalic acid (for oxide phases) and 1.0 mol/L acetic acid
(for carbonate phases). The mixture was evaporated
with occasional shaking on a hot plate. Four cm³ of 1.5
mol/L nitric acid was added to the remainder and
centrifuged. This is referred to as hot extraction. The
digest was diluted to 60 cm³ and analyzed for Cu using
FAAS model 8010 Young Lin. A blank digest was
carried out in the same way.
RESULTS
Copper content in guinea corn and soils: The copper
content for the samples collected from the sampling
locations is shown in Table 1 to 8.
The results of the Cu concentration in guinea corn
and soil vary from one location to another. The results
of the Cu concentrations of some sampling sites in
Kabala (KB) indicate higher concentrations of the metal
in soil than in the corresponding guinea corn (KB1,
KB2, KB3). This could be due to agricultural practices
and other human activities such as application of
fertilizer, pesticides and heavy traffic in the sampling
locations. Similar results were reported by
Urunmatsoma et al. (2010). A higher Cu concentration
was observed in guinea corn than the corresponding soil
samples in KB4, KB5, KB6, KB7 and KB8. This could be
attributed to the species of guinea corn grown and other
anthropogenic activities (Ana-Irina et al., 2008;
Urunmatsoma et al., 2010). The Cu concentration at
Nasarawa (NS) followed a similar pattern to that of
Kabala (KB) with NS1, NS3, NS5 Copper and NS2
Copper with higher Cu concentration in soil than the
corresponding guinea corn samples. The Cu
concentration at NS2 Copper and NS8 Copper is higher
in the guinea corn than the corresponding soil samples.
Similar observations were observed at other sampling
locations.
In some of these sampling locations, the Cu
concentration in the soil was above the tolerable limit
of 100 mg/kg (Lindsay and Norvell, 1978). The highest
concentration of Cu in the areas is not only a problem
to plant nutrition and food chain, it may as well
constitute a direct health hazard (Seward and
Richardson, 1990).
The ANOVA (p = 0.007<0.05) indicated a
significant difference in the Copper concentrations
across the various guinea corn sites. The real
differences of Copper concentrations can further be
Table 1: Results of Cu concentrations in maize and soil samples at Kabala
Hot extraction
-------------------------------------------------------------------------------------------------Metal conc in
Metal conc in soil
Oxalic
Acetic
G/corn sample
sample HNO3/H2O2
(2+1)
EDTA 0.05 M
acid 1.0 M
acid 1.0 M
Sample site
pH
HNO3/H2O2 (2+1)
KB1
5.16
nd
nd
1.15±0.6
1.73±0.6
0.63±0.5
KB2
5.46
12.69±0.6
2.88±0.6
6.92±0.6
1.73±0.6
3.46±1.2
5.36
13.85±0.6
2.88±0.6
7.50±1.2
2.31±0.6
2.88±0.6
KB3
5.26
12.12±0.6
10.96±1.2
1.73±1.2
2.31±0.6
1.73±0.6
KB4
5.66
13.27±0.6
3.46±0.6
7.50±0.6
2.31±0.6
4.04±0.6
KB5
6.12
13.85±0.6
4.04±0.6
7.50±1.2
2.88±0.6
4.62±0.6
KB6
6.12
13.85±0.6
3.46±0.6
8.08±1.2
2.31±0.6
4.04±1.2
KB7
6.12
13.85±0.6
3.46±0.6
8.65±0.6
2.88±1.2
3.46±0.6
KB8
Results of mean values (mg/kg) ± standard deviation (n = 3)
706 Res. J. Environ. Earth Sci., 4(7): 704-710, 2012
Table 2: Results of cu concentrations in maize and soil samples at Nasarawa
Hot extraction
----------------------------------------------------------------------------------------------------Metal conc in
Metal conc in soil
G/corn sample
sample HNO3/H2O2
Oxalic
Acetic
Sample site
pH
HNO3/H2O2 (2+1)
(2+1)
EDTA 0.05 M
acid 1.0 M
acid 1.0 M
NS1
5.49
nd
nd
3.46±0.6
3.46±0.6
2.31±0.6
NS2
6.12
5.77±0.6
6.92±0.6
4.62±0.6
5.19±0.6
4.62±0.6
NS3
5.33
5.77±0.6
6.92±0.6
4.04±1.2
4.62±0.6
4.62±1.2
NS4
5.92
5.77±0.6
4.62±0.6
4.04±1.2
3.46±0.6
2.88±1.2
NS5
6.12
6.35±0.6
7.50±0.6
5.19±0.6
5.77±0.6
5.19±1.2
NS6
6.45
6.92±0.6
8.08±1.2
5.77±0.6
6.35±0.6
5.77±1.2
NS7
6.45
6.35±0.6
8.08±0.6
5.19±1.2
6.35±0.6
5.19±1.2
NS8
6.45
7.50±0.6
8.65±1.2
5.77±0.6
6.35±0.6
5.77±0.6
Results of mean values (mg/kg) ± standard deviation (n = 3)
Table 3: Results of cu concentrations in maize and soil samples at Mando
Hot extraction
---------------------------------------------------------------------------------------------------Metal conc in
Metal conc in soil
G/corn sample
sample HNO3/H2O2
Oxalic
Acetic
Sample site
pH
HNO3/H2O2 (2+1)
(2+1)
EDTA 0.05 M
acid 1.0 M
acid 1.0 M
MD1
4.15
nd
nd
2.31±0.6
4.04±0.6
4.04±0.6
MD2
4.45
31.15±1.2
8.65±0.6
8.08±1.2
13.27±1.2
9.81±1.2
MD3
4.56
31.73±0.6
9.23±1.2
8.08±1.2
13.85±0.6
10.38±0.6
MD4
4.35
26.34±5.9
29.27±2.9
17.56±2.9
26.34±5.9
23.41±2.9
MD5
4.26
160.98±2.9
160.98±2.9
43.90±5.9
67.32±2.9
52.68±5.9
MD6
4.75
163.90±2.9
49.76±2.9
46.83±2.9
73.17±2.9
55.61±5.9
MD7
4.75
163.90±2.9
46.83±5.9
46.83±2.9
70.24±2.9
55.61±5.9
MD8
4.75
160.98±2.9
49.76±2.9
46.83±2.9
73.17±2.9
52.68±2.9
Results of mean values (mg/kg) ± standard deviation (n = 3)
Table 4: Results of Cu concentrations in maize and soil samples at Kakuri
Hot extraction
---------------------------------------------------------------------------------------------------Metal conc in
Metal conc in soil
G/corn sample
sample HNO3/H2O2
Oxalic
Acetic
Sample site
pH
HNO3/H2O2 (2+1)
(2+1)
EDTA 0.05 M
acid 1.0 M
acid 1.0 M
5.08
nd
nd
1.73±0.6
1.15±0.6
1.73±0.6
KK1
KK2
4.98
20.77±1.2
8.08±1.2
9.81±1.2
9.23±1.2
9.23±1.2
KK3
5.14
13.85±0.6
11.54±0.6
10.96±1.2
9.23±0.6
9.23±1.2
KK4
5.14
5.19±1.2
3.46±0.6
2.88±1.2
2.31±0.6
2.88±0.6
KK5
4.34
13.85±0.6
13.85±0.6
9.23±0.6
10.38±0.6
9.81±1.2
KK6
5.15
14.42±0.6
6.35±0.6
11.54±0.6
9.81±1.2
10.96±0.6
KK7
4.5
14.42±0.6
11.54±0.6
10.96±1.2
9.81±0.6
10.38±0.6
KK8
4.5
13.85±0.6
11.54±1.2
10.96±1.2
9.23±0.6
10.38±0.6
Results of mean values (mg/kg) ± standard deviation (n = 3)
Table 5: Results of cu concentrations in maize and soil samples at T/WADA
Hot extraction
---------------------------------------------------------------------------------------------------Metal conc in
Metal conc in soil
G/corn sample
sample HNO3/H2O2
Oxalic
Acetic
Sample site
pH
HNO3/H2O2 (2+1)
(2+1)
EDTA 0.05 M
acid 1.0 M
acid 1.0 M
5.07
nd
nd
1.73±0.6
1.15±0.6
1.73±0.6
TW1
TW2
5.12
20.77±1.2
8.08±1.2
9.81±1.2
9.23±1.2
9.23±0.6
TW3
5.25
21.92±0.6
8.65±0.6
10.38±0.6
9.23±1.2
9.23±0.6
TW4
5.27
5.77±0.6
4.62±0.6
2.88±0.6
2.31±0.6
2.88±0.6
TW5
5.17
21.35±0.6
21.35±0.6
10.38±0.6
9.81±0.6
9.81±0.6
TW6
5.49
21.92±1.2
9.23±0.6
10.96±0.6
10.38±0.6
10.38±1.2
TW7
5.49
21.35±0.6
9.23±0.6
10.38±1.2
10.38±0.6
9.81±1.2
TW8
5.49
21.35±0.6
9.23±0.6
10.96±0.6
9.81±0.6
9.81±1.2
Results of mean values (mg/kg) ± standard deviation (n = 3)
707 Res. J. Environ. Earth Sci., 4(7): 704-710, 2012
Table. 6: Results of cu concentrations in maize and soil samples at S/TASHA
Hot extraction
---------------------------------------------------------------------------------------------------Metal conc in
Metal conc in soil
G/corn sample
sample HNO3/H2O2
Oxalic
Acetic
(2+1)
EDTA 0.05 M
acid 1.0 M
acid 1.0 M
Sample site
pH
HNO3/H2O2 (2+1)
5.14
ND
ND
2.88±0.6
2.89±0.6
2.31±0.6
ST1
ST2
5.20
10.96±1.2
12.69±0.6
9.81±0.6
8.65±1.2
5.77±0.6
ST3
5.25
11.54±1.2
13.27±1.2
9.81±1.2
8.65±0.6
6.35±0.6
ST4
5.82
3.46±0.6
7.50±0.6
4.62±0.6
4.62±1.2
4.04±1.2
5.25
11.54±0.6
13.27±0.6
10.38±0.6
9.23±0.6
6.35±0.6
ST5
ST6
6.10
12.12±0.6
13.85±0.6
10.96±0.6
9.81±1.2
6.92±0.6
ST7
6.10
11.54±0.6
13.85±0.6
10.38±1.2
9.81±1.2
6.35±0.6
6.10
10.96±1.2
13.27±0.6
10.38±0.6
9.23±1.2
5.77±0.6
ST8
Results of mean values (mg/kg) ± standard deviation (n = 3)
Table 7: Results of cu concentrations in maize and soil samples at U/Muazu
Hot extraction
---------------------------------------------------------------------------------------------------Metal conc in
Metal conc in soil
G/corn sample
sample HNO3/H2O2
Oxalic
Acetic
Sample site
pH
HNO3/H2O2 (2+1)
(2+1)
EDTA 0.05 M
acid 1.0 M
acid 1.0 M
4.01
ND
ND
1.15±0.6
1.15±0.6
1.15±0.6
UM1
UM2
4.10
21.92±1.2
7.50±1.2
7.50±0.6
109.62±11.5
126.92±11.5
4.22
22.50±0.6
8.08±0.6
8.08±1.2
109.62±0.6
126.92±1.2
UM3
UM4
5.82
5.77±0.6
6.92±0.6
3.46±1.2
3.46±1.2
4.62±0.6
UM5
4.11
22.50±1.2
8.13±0.7
8.08±1.2
115.38±5.8
126.92±5.8
4.54
23.08±0.6
8.65±0.6
8.65±1.2
121.15±1.2
138.46±1.2
UM6
UM7
4.54
22.50±0.6
7.50±0.6
8.65±1.2
8.08±0.6
110.19±0.6
UM8
4.54
22.50±0.6
8.71±0.5
8.65±1.2
109.62±5.8
126.92±0.6
Results of mean values (mg/kg) ± standard deviation (n = 3)
Table 8: Results of cu concentrations in Maize and soil samples at Kachia
Hot extraction
---------------------------------------------------------------------------------------------------Metal conc in
Metal conc in soil
Oxalic
Acetic
G/corn sample
sample HNO3/H2O2
Sample site
pH
HNO3/H2O2 (2+1)
(2+1)
EDTA 0.05 M
acid 1.0 M
acid 1.0 M
KC 1
6.16
6.35±0.6
14.42±0.6
3.46±0.6
1.15±0.6
1.73±1.2
KC 2
6.24
2.88±1.2
3.46±0.6
2.31±0.6
2.88±0.6
2.31±0.0
KC 3
6.15
79.02±5.9
23.41±2.9
20.49±5.9
32.20±2.9
26.34±2.9
KC 4
6.08
7.50±0.6
5.19±1.2
4.62±1.2
4.04±1.2
4.62±0.6
KC 5
6.07
10.38±0.6
4.04±0.6
5.19±1.2
4.62±1.2
4.62±1.2
KC 6
6.14
4.62±0.6
6.35±0.6
5.19±1.2
4.62±1.2
2.88±1.2
KC 7
6.01
10.96±0.6
4.62±0.6
4.62±1.2
5.19±1.2
6.35±0.6
KC 1
6.16
6.35±0.6
14.42±0.6
3.46±0.6
1.15±0.6
1.73±1.2
Results of mean values (mg/kg) ± standard deviation (n = 3)
analyzed by a post-hoc test using the Duncan Multiple
range test where means of homogeneous subgroups are
displayed. Moreover, the mean plots that follow depict
the mean values of Copper concentrations across the
various guinea corn sites.
The Duncan multiple range tests showed that
Kachia and Nasarawa had the least Copper
concentration, while Sabon Tasha had the highest
Copper concentration as shown in Fig. 2.
Metal speciation: The Cu distribution in soil samples
collected from Kabala (KB), varied in all the sampling
sites. The metal existed in residual, oxide,
carbonate/organic phases.
The
metal
concentration
bound
to
carbonate/organic phase is higher in KB2, KB3, KB4
and KB6 Copper than in other fractions. The carbonate
fraction caused the release of the metal into the soil
solution, which is then available for the plant uptake
through movement of the element from soil solution to
the plant root. The copper in these soils was organically
bound and hence bioavailable and mobile (Hickey and
Kittrick, 1984; Urunmatsoma et al., 2010).
The concentration of the metal bound to residual
fraction is highest in KB1 Copper. The metal is said to
be bioavailable and mobile in the soil. This is in
agreement with the results reported by Hickey and
Kittrick (1984) and Urunmatsoma et al. (2010).
708 Res. J. Environ. Earth Sci., 4(7): 704-710, 2012
Fig. 2: Mean plot for copper concentration in maize crops
The copper concentration bound to oxide fraction
is highest in KB1, KB7, KB8 Copper, respectively. The
oxide fraction is most likely to cause release of the
metal into the soil solution. Thus, the metal is available
for plant uptake through movement of element from
soil solution to plant root. Similar results were reported
by other investigators (Chamon et al., 2005;
Urunmatsoma et al., 2010).
The metal concentration bound to carbonate is
highest KB5, Copper, respectively. Hence the metal is
said to be carbonate bound and available for plant
uptake (Hickey and Kittrick, 1984; Chamon et al.,
2005; Urunmatsoma et al., 2010).
The distribution of Cu in the soil samples from
Nasarawa (NS) indicates predominance of the
carbonate fraction (EDTA) in NS1, NS2, NS3, NS5, NS7
Copper and NS2, NS4, NS8 Copper. While the metal
concentration bound to residual fraction is highest in
NS1, NS5 Copper and that bound to oxide fraction is
highest in NS8 Copper, respectively. Similar results
were reported by other investigators (Hickey and
Kittrick, 1984; Chamon et al., 2005; Urunmatsoma
et al., 2010).
Similar observations were observed in MD, KK,
TW, ST, UM and KC. The HNO3/H2O2, EDTA,
CH3COOH and (COOH)2 extractable Cu are considered
as available Cu in these locations.
The pH of the soil samples from the various
locations is acidic. This could be attributed to the
various agricultural processes on the sampling sites
(Kashem et al., 2007; Chamon et al., 2005). The
bioavailability of copper, lead and zinc from soil
decreases with increasing pH (Moraghan and Mascani,
1991; Morel, 1997). The acidity of the soils increases
the solubility and mobility of the metal in the soils.
Sites
n ta
sh
Un
a
gw
an
mu
az
u
Ka
ch
ia
wa
da
la
we
st
10.00
Ka
ba
Ka
chi
a
nw
ada
n ta
Un
sh a
gw
an
mu
az u
Sites
Sa b
o
uri
Tu
du
Ka
k
Ma
n do
0.00
20.00
Sa
bo
20.00
30.00
Tu
du
n
40.00
40.00
Ka
kur
i
60.00
50.00
Na
s ar
aw
a
M
an d
o
Mean of copper concentration
in soil sample (mg/Kg)
80.00
Ka
bal
aw
es t
Na
s ar
aw
a
Mean of copper concentration
in maize sample ( mg/K g)
100.00
Fig. 3: Mean plot for copper concentration in maize grown
soil
Such increase in solubility of the metal is likely to lead
to increased availability of the metal for plant uptake
(Takac et al., 2009).
The ANOVA (p = 0.114>0.05) showed that there is
no significant difference in the Copper concentrations
across the various guinea corn sites. Moreover, the
mean plots that follow will clearly depict the mean
values of the metal concentrations across the various
guinea corn fields as shown in Fig. 3.
CONCLUSION
Total trace metal composition of soil is of little
importance in determining its uptake by plants and
consequently, in contaminating the food chain since the
different forms have different mobilities, bioavailabilities
and potential environmental contamination potential. The
results on heavy metal speciation in the study indicated
that the soil samples collected from various areas contain
varying amounts of the metal. The metal was distributed
between residual, oxide and carbonate fractions. An
increase of the metal concentration in some areas suggests
that heavy use of agrochemical materials for planting
activities could cause increase in the content of heavy
metals in the soil.
ACKNOWLEDGMENT
The authors show great appreciation to Kaduna
Polytechnic, Nigeria for providing facilities to analyze the
samples and to Kabiru Shehu and Yusuf AbdulRaheem,
for the help in sample collection, metal and statistical
analyses.
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