CADMIUM, NICKEL, AND LEAD ACCUMULATION IN CHAMOMILE, THYME AND
SAGE, GROWN ON HEAVY METAL-ENRICHED SOIL.
Melpomeni Skoula1, Monika Fabian1, and Nikos Lydakis – Simantiris1,2,*.
1
Mediterranean Agronomic Institute of Chania, Alsyllion Agrokepion, P.O. Box 85, 73100 Chania, Crete, Hellas, and
2
Technical and Educational Institute of Crete, Department of Natural Resources and the Environment, Chania, Crete,
Hellas.
*Corresponding author: lydakis@chania.teicrete.gr, tel. (+30)28210 23072
ABSTRACT
The effects of heavy metal, (Pb, Ni, and Cd), enrichment of soil on growth and quality of chamomile, sage, and thyme
planted in it, as well as their ability to accumulate these metals in their tissues, were investigated by a series of pot
experiments. Three parts of the plants were examined for heavy metal content: the roots, the leaves and the flowers. The
measured parameters were the growth, yield, chlorophyll content, essential oil composition, and heavy metal content in
the different plant tissues. The levels of the heavy metals in the soil used in this study were not lethal for the plants and
the effects on growth, as well as on the other quality parameters examined, were minor. The heavy metal content in
different tissues of the three plants varied from very low levels to levels quite higher than the maximum acceptable
concentrations set by WHO. From this study it is concluded that sage and chamomile can be considered as Cd excluders,
with sage showing a stronger capacity as a Cd excluding plant.
1. INTRODUCTION
During the last decade, spices and medicinal plants have gained a more important role in agricultural production because
of their increased use as raw materials in the food and pharmaceutical industry and their everyday consumption by
people. The heavy metal accumulation ability of these plants is of particular interest since in many cases they are
collected by consumers directly from areas potentially contaminated by toxic elements. In this study we examined the
accumulation of cadmium lead and nickel in three different parts of chamomile, thyme and sage (roots, leaves and
flowers), in relation to their growth, yield, chlorophyll content and essential oil content. Care was taken to keep the
heavy metal concentrations in the cultivation soil below lethal levels. Cadmium and lead are considered as the most
hazardous heavy metals and the main environmental contaminants [1]. Nickel, at low concentrations, is used by the
plants as a micronutrient, but at high concentrations it has toxic effects on vegetal growth [2]. For the presented study
we designed a pot experiment, where various amounts of heavy metal nitrate salts were added to cultivation soil in order
to achieve certain concentrations of the toxic metals in the soil. The plants were then planted and their quality
parameters were checked periodically whereas quantitative analysis for the chlorophyll essential oil and heavy metal
content was performed in the middle, (six weeks after planting), and at the end of the experiment (twelve weeks after
planting). Results show that, at the concentrations used, the accumulation of heavy metals in the examined three tissues
varied considerably, and in some cases exceeded the acceptable limits. On the other hand, the growth, yield, chlorophyll
and essential oil contend were not affected significantly by the presence of the heavy metals (data not shown).
2. MATERIALS AND METHODS
Plant Material: Matricaria recutita (annual), Salvia officinalis and Thymus vulgaris (perennials) were planted in soil in
which different amounts of Cd, Pb, and Ni, in the form of their nitrate salts, were added and homogenized. Each
experimental condition was repeated five times. The size of the pots were one litre. The experiment lasted from
November 2001 until May 2002. Samples were collected six and twelve weeks after planting in the contaminated soil.
After collection, fresh weight was measured and then the samples were dried at 65ºC for 40 hours. The dry weight was
measured and then they were stored at room temperature in air tight containers until analysis. Chlorophyll content
determination was carried out in leaves according to [3]. For quantitative and qualitative analysis of essential oils, head
space analysis by GC-FID and GC-MS techniques were used, respectively.
Heavy metal content: Cd, Pb, and Ni concentrations were determined in the cultivation soil (total amount and plantavailable amount) and in three different plant tissues: roots, leaves and flowers. For total amount of heavy metals in the
soil, the samples were first digested in a microwave digestor in the presence of concentrated HCl and HNO3. For plant
available amounts, a DTPA extraction was carried out [4]. For plant analysis, the samples were first dry ashed and then
the ash was dissolved in HCl 2N plus a small amount of HNO3 4N [5]. ICP spectroscopy was used for the determination
of the heavy metal concentrations in all samples.
3. RESULTS AND DISCUSSION
Table 1 shows the calculated concentrations of heavy metals after their addition to the soil, and the corresponding
available concentration as it was determined in the DTPA extractant. In all cases, the available amount of heavy metals
was a fraction of the total concentration. Each given value is the mean of three determinations. ICP determination of the
total amounts of heavy metals in the soil was performed and the results were comparable to the calculated concentrations
(data not shown).
TABLE 1. Relation between added and available amounts of heavy metals in the cultivation soil.
Metal
Added
Available
Metal
Added
Available
Metal
Added
(ppm)
(ppm)
(ppm)
(ppm)
(ppm)
0
0.167
0
6.95
0
Cd
Pb
Ni
1
0.875
60
13.7
20
Cd
Pb
Ni
3
1.35
180
28.7
60
Cd
Pb
Ni
10
2.57
600
36.2
200
Cd
Pb
Ni
30
10.1
1800
188
600
Cd
Pb
Ni
Available
(ppm)
0.643
8.43
22.7
77.2
153
ICP analyses were carried out on the acidic solutions of the dissolved ashes of the tissues collected twelve weeks after
planting (see Materials and Methods). In general, the concentrations of Cd, Pb, and Ni in the roots, leaves and flowers
followed the trend: [Cd, Pb, Ni]roots > [Cd, Pb, Ni]leaves > [Cd, Pb, Ni]flowers. In the following sections, the results of ICP
analyses of dried tissues of the three different plants are presented.
3.1 Matricaria recutita (chamomile)
Table 2 shows the results of the heavy metal content analysis of the three different tissues. Extra care was taken to
remove the soil remains from the roots before weighing and drying the tissue. Each value in the table is the mean of
three experiments. Figure 1 shows the results in Table 2 as bar graphs.
Control
(0 ppm Ni)
20 ppm Ni
60 ppm Ni
200 ppm Ni
600 ppm Ni
2.01
5.12
2.72
2.62
8.61
42.15
84.75
156.5
153
40
30
20
10
0
Flowers
Roots
Leaves
Flowers
Roots
Leaves
Flowers
Roots
Leaves
1.91
5.35
6.41
21.93
29.2
0.15
4.47
1.43
1.78
7.88
24.8
29.6
46.25
148
206
288
8.40
27.1
43.93
61.16
[Pb], ppm
1.27
50
180
160
140
120
100
80
60
40
20
0
350
300
250
[Ni], ppm
Control
(0 ppm Pb)
60 ppm Pb
180 ppm Pb
600 ppm Pb
1800 ppm Pb
60
[Cd], ppm
TABLE 2 Heavy metals uptake in flowers,
roots and leaves of chamomile
Supply to
Flowers Roots Leaves
the soil
ppm
ppm
ppm
0.541
1.32
0.30
Control
(0 ppm Cd)
1.96
1.57
1.88
1 ppm Cd
3.025
3.64
3.07
3 ppm Cd
1.79
12.75 7.63
10 ppm Cd
9.27
48.62 27.13
30 ppm Cd
200
150
100
50
Figure 1. Relative uptake of Cd, Pb, and Ni by flowers,
roots and leaves in chamomile.
0
The highest acceptable concentrations of Cd and Pb, set by WHO, [6], are 0.2 and 10 ppm, respectively, in all plant
parts. As can be seen in Table 2 and Figure 1, the roots of chamomile are able to accumulate heavy metals at much
higher concentrations compared to leaves and flowers, in agreement with what is already known about this tissue.
Cd was accumulated in all three tissues at higher than the acceptable limits even in the case of control, the soil of which
contained some Cd, as can be seen in Table 1. The presence of Cd in the control experiment is attributed to
contamination.
Pb was accumulated mainly in the roots. In the aboveground parts, Pb accumulation was higher than the acceptable
limits only in leaves and only when Pb concentration in the soil was above 180 ppm. In the flowers, even at the highest
soil concentration examined, Pb did not reach the WHO levels.
Ni was accumulated in the three tissues at concentrations a little higher than Pb.
3.2 Salvia officinalis (sage)
Table 3 shows the results of the ICP analysis of sage roots and leaves. Each value in Table 3 is the mean of three
measurements. In Figure 2, the results shown in Table 3 are presented as bar graphs, for easy comparison.
TABLE 3. Heavy metals uptake in
roots and leaves of sage.
Roots,
ppm
0.85
Leaves,
ppm
0.18
2.41
4.58
22.05
51.7
0.29
0.48
1.60
2.31
60
50
[Cd], ppm
Supply to
the soil, ppm
Control
(0 ppm Cd)
1 ppm Cd
3 ppm Cd
10 ppm Cd
30 ppm Cd
40
30
20
10
0
5.10
1.04
21.25
31.30
95.10
177
9.74
3.31
6.95
5.31
Leaves
Roots
Leaves
Roots
Leaves
200
180
160
140
[Pb], ppm
Control
(0 ppm Pb)
60 ppm Pb
180 ppm Pb
600 ppm Pb
1800 ppm Pb
Roots
120
100
80
60
40
20
2.05
0.57
29.80
56.15
74.65
248.5
11.95
11.28
16.16
29.53
0
300
250
[Ni], ppm
Control
(0 ppm Ni)
20 ppm Ni
60 ppm Ni
200 ppm Ni
600 ppm Ni
200
150
100
50
Figure 2. Relative uptake of Cd. Pb, and Ni by flowers,
roots and leaves in sage.
0
As in chamomile, the heavy metal accumulation in sage tissues was profoundly higher in the roots than in the leaves.
Again, the levels of Cd concentration were higher than the WHO acceptable limits, even in the case of the control
experiment.
In general, the accumulation of Pb in sage occurred in a lower extent, as compared to chamomile. The accumulation of
Pb in the roots was above the WHO limits (10 ppm), when the concentration of the metal in the soil exceeded 60 ppm.
However, in the leaves, even at the highest concentrations examined, Pb did not reach the WHO limits.
Ni was also accumulated at a lower extent in all three tissues as compared to chamomile.
3.3 Thymus vulgaris
Table 4 shows the accumulation of Cd, Pb and Ni in thyme roots and flowers, as resulted from ICP analyses of the dried
tissues. Figure 3 presents the same data as bar graphs. Each value in Table 4 is the mean of three ICP determinations.
As the data presented above show, the roots of thyme accumulated Cd at a lower level as compared to the roots of the
other two plants examined. Cd concentration in thyme leaves were in general in between the Cd concentration in leaves
of chamomile and sage, and above the WHO limits. For intermediate Pb concentrations in the soil, Pb accumulation in
the roots was higher as compared to sage, but considerably lower as compared to chamomile. Pb accumulation in leaves
was higher than WHO limits in all treatments, except control.
10
9
8
7
[Cd], ppm
TABLE 4. Heavy metals uptake in flowers,roots and leaves
of sage.
Supply to
Roots,
Leaves,
the soil
ppm
ppm
0.33
0.34
Control
(0 ppm Cd)
0.75
0.57
1 ppm Cd
1.68
0.65
3 ppm Cd
3.50
0.87
10 ppm Cd
8.55
8.65
30 ppm Cd
6
5
4
3
2
1
0
Roots
5.89
1.59
38.10
43.55
165
155.9
15.89
17.01
17.16
12.18
180
160
140
[Pb], ppm
Control
(0 ppm Pb)
60 ppm Pb
180 ppm Pb
600 ppm Pb
1800 ppm Pb
Leaves
120
100
80
60
40
2.63
0.55
21.35
42.55
64.40
154.5
27.86
33.52
57.50
46.53
20
0
Roots
Leaves
Roots
Leaves
180
160
140
[Ni], ppm
Control
(0 ppm Ni)
20 ppm Ni
60 ppm Ni
200 ppm Ni
600 ppm Ni
120
100
80
60
40
20
Figure 2. Relative uptake of Cd. Pb, and Ni by
flowers, roots and leaves in thymus.
0
Summarizing, this study shows that, in the concentrations used, the order for the accumulation of Cd, Pb, and Ni in the
examined plants was roots > leaves > flowers (measured only for chamomile). The soil concentrations of heavy metals
used in this study not only were not lethal for the plants, but didn’t seem to affect significantly the quality parameters we
measured (growth, yield, essential oil content, chlorophyll content, (data not shown). Finally, we can consider
chamomile and sage to be Cd-excluders as more Cd was detected in the roots than in the soil and this Cd was
translocated only at very low concentrations in the aboveground parts. Between the two plants, sage can be considered
as a stronger excluder than chamomile
4. REFERENCES
[1] Zheljazkov, V.D., and Fair, P., (1996) “Study of the Effect of Highly Heavy Metal Polluted Soils on Metal Uptake
and Distribution in Plants from Genera Artemisia, Draccocephalum, Inula, Ruta and Symphytum” Acta Horticultura,
426: 397-417.
[2] Palacios, G., Gomez, I., Moral, R., and Mataix, J., (1995) “Nickel Accumulation in Tomato Plants, Effect on Plant
Growth” Frecenius Envir. Bull. 4: 469-474.
[3] Harborne, J. B. (ed.) (1998). Phytochemical Methods: A Guide to Modern Techniques of Plant Analysis, 3rd ed.
Chapman and Hall, London.
[4] Amacher, M. C., (1996) “Nickel, Cadmium and Lead” in Methods of Soil Analysis, Part 3, SSSA, pp. 739-768.
[5] Yoong K. Soon (1998) “Determination of Cd, Cr, Co, Pb, and Ni in Plant Tissue” in Handbook of Reference
Methods for Plant Analysis (Y. P. Kalra ed) CRC Press, pp. 193-198.
[6] World Health Organization, (1998) “Determination of Arsenic and Heavy Metals”. In Quality Control Methods for
Medicinal Plant Material, Genova, p. 62-63.