Note: Descriptions are shown in the official language in which they were submitted.
CA 02354249 2001-11-13
1
METHOD OF SUSTAINING PLANT GROWTH IN TOXIC SUBSTRATES
POLLUTED W1TH HEAVY METAL ELEMENTS.
Activities in the industrial Age have resulted in the deposit of high levels
of many
metals in certain sites, to the point that human life is seriously threatened.
Metal-production activities, such as mining or smelting, as welt as the
ubiquitous
use of metals, have created many sites where toxic metals have become
concentrated in soils.
In recent years, efforts have been made to develop phyto-remediation methods,
i.e. the use of metal-accumulating plants called metallophytes to remove
contaminating metals from sites.
It has been known for some time that many plant species will concentrate
certain
metals in their leaves, stems and roots to a varying degree.
For heavy metals, two different types of phyto-remediation methods can be
distinguish:
- rhizofiltration, by concentration of heavy metals in plan roots;
- phyto-stabilisation, the roots of the plants limiting heavy metals
availability and
limiting mobility of said metals into the groundwater.
More than 400 phyto-remediator are known, most of them absorbing nickel. The
more rarely absorbed heavy metals include manganese, cadmium and lead.
Various metallophytes have been tested, such as Brassicaceae (Thlaspi
brachypetal, Thlaspi ochroleucum, Thlaspi caerulescens, Thlaspi rotundifolium,
Cardaminopsis halleri), Caryophyllaceae (Minuartia verna, Polycarpea
synandra),
Fabaceae (Astragalus pectinatus, Astragalus bisculatus) Myriophyllium
verticillatum, Pshychotrai douerrer, Vola calaminaria.
Document US-A-5.917.117 relates to a method by which hyperaccumulation of
metals in plant shoot such as Brassicaceae (e.g. Brassica, Sinapsis, Thlaspi,
Alyssum, Eruca) is induced by exposure to inducing phytotoxic agents such as
chelating agents (e.g. Roundup ~) and high concentrations of heavy metals. The
exposure to inducing agent is made after a period of plant growth, as metal
accumulation into plant shoots has dramatic negative effects on plant growth.
The use of phytotoxic inducing agents as described in document US-A-5.917.117
is non-ecological and potentially dangerous for the operator.
CA 02354249 2001-11-13
2
Document US-A-5.711.784 disclose a method of extracting nickel, cobalt and
other metals including the platinum palladium metal families from soil by
phytomining. The conditions include 1) lowering the soil pH by addition of
sulphur
and use of ammonium N fertilisers, 2) maintaining low Ca in the soil by
acidification of the soil with sulphur or sulphuric acid and, 3) applying
chelating
agents to the soil, such as NT,A.
The method described in document US-A-5.711.784 is complicated and non-
ecological.
Document US-A-5.927.005 relates to a method of removing heavy metals from
'10 soil using creosote plants (Lacrea tridentate). Again, to increase the
rate of metal
uptake in the plants, it is proposed to increase the acidity or to add
chelators to
the soil in which the creosote bushes are growing.
Other phyto-rernediation techniques are described in documents WO-A-00/28093,
WO-A-00/31308, WO-A-98/59080, WO-A-94/01357, EP-A-0 911 387, JP-A
57.000.190, DE-A-4100758, DE-A-39.21336, US-A-5 100 455, US-A- 5.320.663,
US-A-5 364 451, US-A-5 785 735, US-A-5 809 693, US-A-5 853 576, US-A-5 928
406, US-A-5.944.872, US-A-6 117 462.
Despite increasing interest and research, several problems associated with
phyto-
remediation remain. For example, some metals in contaminated areas may be
;?0 hardly reached via phyto-remediation because they lie beneath the
rhizosphere,
many of the known metal-accumulating plants being simply too small to
accumulate large quantities of metals. Additionally, many of the plants thus
far
identified as useful in phyto-remediation are from tropical regions.
One object of the invention i:> to provide means to increase plant root growth
in
;?5 toxic substrates polluted with heavy metal elements.
Another object of the invention is to provide means preventing surface
erosion,
especially for toxic substrates polluted with heavy metal elements.
Another object of the invention is to provide means of promoting growth of
metallophytes and other plants on toxic ground polluted by the presence of
heavy
30 metal elements.
Another object of the invention is to provide above mentioned means, said
means
being ecological and less expensive than most of known bio-remediation
methods.
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3
According to the invention, there is provided a method of sustaining plant
growth
in toxic substrates polluted with heavy metal elements, characterised in that
it
comprises amendment of the toxic substrates with an organo-zeolitic mixture.
The heavy metal element caa~ be zinc, copper, lead, cadmium or arsenic, the
orgo-zeolitic mixture being added to the said polluted substrate between 10 %
and
25 %.
The method of the present invention can be used to sustain the growth of
various
plants, especially plant root growth in toxic substrates polluted with heavy
metal
elements.
Normally, owing to the lack of available nitrogen and other essential nutrient
elements, ground containing high levels of toxic metals would not sustain
plant
growth to a level that will prevent surface erosion. By amending the ground
with
the organo-zeolitic fertiliser this condition can be overcome by growing
plants with
very dense root systems.
The method of the present invention can be manipulated to vary the shoot to
root
ratio of the plant species used. In tt-~is respect, plants which concentrate
heavy
metals such as Zn, Cd and Cu in their shoots can be grown successfully and
Groped to remove the metals from the rhizosphere.
The method of the present invention will enable the metal enriched plant
tissue,
on ashing, to be reduced to a small volume v~rhich can be disposed of easily
by
mixing with zeolite amended Portland cement and used in the production of
concretes that are known to have high compressive strengths.
More precisely, after harvesting and ashing the plant, the heavy metal cations
contained in the ash can be put into aqueous solution and ion-exchanged into a
2:5 zeolitic tuff. The resulting zeolitic material can be dried and used to
produce
blended cements which have improved compressive strength and are also known
to reduce the expansion caused by alkali-aggregate reactions.
It is known that natural zeolite minerals can be used as biological fertiliser
(see
for instance JP-A-10210855, ,JP-A-4197110, EP-A-444392, US-A-5 082 488, US-
~~0 A-5 451 242,US-A-5 900 38'l, RU-A-2 121 777, RU-A-2 132 122, RU-A-2 137
340). The preparation of an organic fertiliser incorporating zeolitic tuff is
described
r
in document US-4.559.073, the inclusion of the zeolitic component being
claimed
to lower the water content of the mixture to allow effective aerobic
fermentation.
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4
Document US-A-5.106.405 disclose the property of ion-exchanging ammonium
ions that, via soil microbioloc3ical reactions, would supply available
nitrogen to
plants growing in a substrate amended with a bio-fertiliser containing
zeolitic
component.
The inventor has discovered that natural zeolite materials could be used to
prepare a biological fertiliser which can be applied to ground contaminated
with
heavy metal cations to enable the sustainable growth of plants and to control
the
development of shoot to root ratio in such a way that plant morphology can be
adjusted to either maximise soil retention by dense root growth or increase
the
foliage uptake of toxic heavy metal ions.
If untreated, such ground will not support vegetation and becomes subject to
surface erosion by wind and rain. Toxic material transported by these agents
into
local drainage patterns is thus isolated and therefore uncontrollable.
Specific implementation of the invention will now be described, by way of
example, provided for illustrative proposes and not intended to limit the
scope of
the invention as claimed herein. Any variations in the exemplified
compositions
and methods which occur to the man skilled in the art are intended to fall
within
the scope of the present invention.
Example
A clay rich toxic soil containing : 2.87 % Organic matter, 1.1 iv Calcium
carbonate,
2.24 % total Iron, 28.9 mg.kg~' Copper, 915 mg.kg-' Zinc, 670 mg.kg-' Lead,
12.2
mg.kg~' Cadium and 18.9 mg.kg-' Arsenic has been amended with 16.7 % organo-
zeolitic fertiliser"
Organo-zeolitic fertiliser is prepared as follows.
Animal waste, e.g. chicken manure, is composted together with crushed zeolitic
tuff containing the zeolite Ca, K, Clinoptilolite in a ratio of 1.2 (by
volume) i.e. tuff
to manure. The materials are mixed together with enough water to make the pile
damp and choppen straw is added. Air is forced through the pile from a
perforated
plastic pipes) laid inside the pile during construction and the reaction is
carried
out under cover. This could prevent saturation of the pile with rain water.
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The pile reaches 50-70 °C and then the temperature drops to ambient
at which
stage the composted materi~ is dry, friable, odourless and ready for use as an
organo-zeolitic fertiliser.
Spring Wheat (Triticum aestivum L, cv. Red Fife) was sown in two kilograms
5 substrates. Wheat grown in 'the untreated soil was used for the comparison.
The
plants were grown in 255 mm diameter pots, replicated four times, under
ordinary
lighting conditions in a greenhouse. Watering, with de-ionised water, was by
weight to field capacity (180 ml per 2 kg substrate) and the pots were placed
in
shallow trays to retain leachate. Watering, generally on a daily basis,
prevented
1' 0 the plants from drying out and any water running from the dots was
returned to
the substrate surface with little loss. Plants were harvested on a regular
basis
each month and the shoot weights were recorded after drying to constant weight
at 70 °C.
One month altar germination the substrates were leached with 400 ml of de-
-I 5 ionised water (pH = 8.4) and after removal of fine colloidal particles
were analysed
chemically. Two further leachate collections were made at monthly intervals
over
a three month growth period.
.?0
Leachate chemistr~at the third harvest
Toxic substrate ~ N conc 0.23mg/I
Amended substrate N conc 178.00 mg/I
Toxic Substrate K conc ' 17.38 mg/I
Amended Substrate K conc 66.70 mg/I
Toxic Substrate Ca conc 20.40 mg/I
Amended Substrate Ca conc 253.00 mg/I
Toxic Substrate Mg conc 2.69 mg/I
~
Amended Substrate Mg conc 27.10 mg/1
Toxic Substrate pH 7.8 E.C. 149 ~tS/cm
Amended Substrate pH 7.2 E.C. 2077 ~S/cm
r
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6
These results demonstrate the degree of mobilisation of major cations in the
amended soil solution.
In case of the metal trace elements a general decrease is seen in the
leachates
between high concentrations in the toxic substrates and low concentrations in
the
amended substrates. An example is shown below for Znc.
Toxic substrate: Zn conc 0.65 mg/L
Amended Substrate: Zn conc 0.10 mg/L
Following the analysis of the leachates the chemical analyses of the plant
shoots
express the way in which nutrient and trace metal elements are taken up from
the
respective substrates.
Plant shoot chemistry at the third harvest
Amended substrate Toxic Substrate
Ncon ~~.l9wt~ 1.16wt%
K conc 35.33 mg/g 18.20 mg/g
Ca conc 5.82 mg/g 2.82 mg/g
Mg conc 1.24 mg/g 0.85 mg/g
Zn conc '124 ugJg 67 ~9~9
Pb conc 5 1~9~9
Cu conc 17 ~g/9 5 X9/9
The plant shoot chemistry can now be compared to the established nutrient
range
for Spring lNheat.
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7
dry wt . ~9I9
N P K Ca Mg Zn Cu
Adequate Range : 3.0-4.50.3-0.52.9-3.80.4-1.00.15-0.320-70 5-10
Toxic Substrate : 1.16 0.5 1.80 0.3 0.10 67 5
Amended Substrate : 2.190.3 3.50 0.6 0.12 124 17
Shoot dry weight recorded at monthly harvests
15' harvest Dry weight (g/plant)
Toxic Substrate: 0.26
Amended Substrate: 0.89
2"° harvest
Toxic Substrate: 0.52
Amended Substrate: 5.36
3'd harvest
Toxic Substrate: 1.03
Amended Substrate: 6.77
Comments on example
The type of organo-zeolic fertiliser of the present invention can be adapted
to
grow plants with a dense root system on toxic soils that cannot normally
supply
;25 sufficient plant nutrients to support such growth. This is achieved by
microbiological means only as no inorganic mineral salts have been added..
It can be seen from the chemistry of the plant shoots than when the available
nitrogen in the substrate is some 35 % below the adequate range a dense root
system, in the case of Spring Wheat, is formed. As the percentage of organo-
zeolitic material added to t:he toxic soil can be altered the concentration of
available nitrogen can be adjusted ,to suit the plant species concerned. If
maximum shoot growth is required then the percentage or organo-zeolitic
material can be adjusted upwards to put the nitrogen concentration into the
CA 02354249 2001-11-13
adequate range. T his would bs desired if maximum plant uptake was required in
order to remove heavy metals from the rhizosphere.
The trace element concentrations of zinc and copper in the plant shoots show
that
the mobilisation of cations in the soil solution, due to the microbial
activity of the
organo-zeolitic material, m~;ke these elements available to the plant. On
harvesting the soil will be partially depleted in these elements and in time
the
rhizosphere will become less polluted. The volume of plant material after
harvest
can be greatly reduced by asking and can be safely stored or possibly re-
cycled.
As it is known that the addition of finely crushed zeolitic tuff to Portland
Cement
'10 improves its physical and chemical properties the suggestion is made that
heavy
metal cations remaining in the plant ash could be exchanged into zeolitic tuff
which is afterwards used for such a purpose.
Possible explanation
'15 Proposed explanation of the above mentioned results is given below, the
detailed
mechanism still being the subject of research by the inventor. During organo-
zeolitic fertiliser preparation, ~~hoppen straw likely provides a source of
carbon to
support bacterial growth. Ammonifying bacteria as typified by Clostridium and
Penicillium, acting on the organic material, decompose protins, amino sugars
and
;?0 nucleic acids to ammonia. The ammonia in the cationic form NH4+ is ion-
exchanged into the zeolite where it is held, loosely bound, within the pore
space of
the crystal lattice. The bacterial activity causes the increase of temperature
up to
50-70 °C, the completion of the reaction being reached as the
temperature drops
to ambient.
;25 On addition of the fertiliser to a plant substrate the NH4+ ions held in
the zeolite
pore space diffuse at an expotential rate into the substrate. Nitrifying
bacteria
present in the organo-zeolitic component use the diffusing NH4+ to build very
large populations and in so doing oxidise NH4+ to produce a large source of
nitrate that is used in plant growth. As a consequence of t'~ bacterial
reactions
.30 free hydrogen ions (protons) are liberated and mobilise the soil solution
causing
the dissociation of metal cations present in the substrate.
The inventor has demonstrated how the organo-zeolitic fertiliser can be used
with
soils polluted with heavy mertals such as Zinc, Cadium, Copper and Lead. The
CA 02354249 2001-11-13
inventor has now found that when 16-7 7 % organo-zeolitic fertiliser is added
to
such soils similar effects occur which greatly increase nitrate concentration
and
mobilise metal rations in the soil solution.
Amending a toxic soil in this way slightly lowers the pH of the soil solution
but
increases its electrical conductivity by an order of magnitude. At the level
of
amendment quoted (16-17 °/o) twice the amount of available nitrogen
present in
the toxic soil is provided. This increase is 35 % below the adequate range for
Spring Wheat (Triticum ae;>tivum L, cv. Red Fife) and has the effect of
maximising the root/shoot ratio. In this way a dense root system can be
developed
by increasing the amount of organo-zeolitic fertiliser added to the soil the
root to
shoot ratio can be decreased, in which case shoot growth is favoured.
Cation mobilisation in the soil solution provides the growing plant with
nutrients
such as potassium, calcium, magnesium, and zinc and the plant's requirement
acts to buffer the system against concentration of these elements and
diffusion
from the rhizosphere. By increasing the level of plant nutrients in this way a
healthy plant can be grown and sustained on toxic soils polluted with heavy
metals. In the case of zinc and copper the inventor has observed that these
elements are taken up by the plant at a rate that can be tolerated and the
damage
occurring in the same plants grown in the toxic soil is not seen. This
property can
be used to remove heavy metal elements from the rhizosphere by harvesting the
plant. The plant material can be greatly reduced in volume by ashing without
loss
of the heavy metals and incorporated in a mixture of Portland Cement and
finely
crushed zeolitic tuff. In this vvay concrete of high compressive strength and
low
alkali reactivity can be made and used to store the heavy metal elements.
CA 02354249 2001-11-13
Inventor : Dr. Peter l.Leggo
Statement
Work on Ryegrass (Lolium perenne L.) has shown that amendment of the plant
substrate with
the zeolite bio-fertilizer will enhance plant growth in a similar way to that
seen with Spring
Wheat (T'riticum aestivt~m L., cv.Par2gon).
Dense root systems can be sustained in soils polluted with heavy metals. The
conraminated
substrate used was identical to than. in the spring wheat programme as
described in the above
patent
In the ryegrass programme un-ammoniated zeolitite (i.e. zeoliased volcanic
tuff containing
the zeolite mineral clinoptilolite) was added in varying proportions to the
initial amended
substrate containing 16 -17 (volume %) organo-zeolitic fertilizer (zeolite bio-
ferdlizer).
The exchangable rations in the clinoptilolite used are Calcium and Potassium.
Zeolite
minetaLs with appreciable quantities (> ca.2 Wt %) are undesirable as
reactions in the plant
substrate increase the sodium availability which can cause depression is
growth The typical
formula of the clinoptilolite used i<; : (K ,9Ca t~ Na a, Mg 0.,) al ~, Si :99
0 ;,. 23.1 H,0
It was found that the relationship between shoot dry weight and weight
°,o excess zeblitite
defined the (cut-aid limit of growth enhancement above which continued
addition of zeolitite
diminished plant growth.
This effect was found to vary according to plant density. A low density group
(2~ plancslpot)
having a higher cut-off limit than a high density group (ca. 600 plane /pot).
The low density plants also showed greater growth enhancement than the high
density group.
Analytical data: Shoot dry weights
(i) Low density group harvested at four months
Plant substrate Shoot weight (g)
Gp.l Toxic substrate 738
GpZ Amended substrate, (16 --17) vol % zeolite bio-fertilizer 31.46
Gp.3 Ditto + 75 wt % un-amoniatedzeolitite 32.73
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11
Gp.4 Ditto + 1 ~0 wt % un-amoniated zeolitite 33.36
Gp.S Ditto + 225 wt % un-ammo;niated zeolitite 33.98
Gp.6 Ditto + 300 wt % un-ammoniated zeolitite 28.87
(ii) High density group harvested at five months.
Plant substrate Shoot weight (g)
Gp. l Toxic substrate 6.76
Gp.2 Amended substrate (16-17) vol% zeolite bio-fercilizer 25.83
Gp.3 Ditto + ~,5 wt% ~_~o~ated zeolitite 26.49
Gp.4 Ditto + 150 wt% un-ammoniated zeolitite 25.46
Gp.S Ditto + 225 wt% un-ammoniated zeolitite 25.10
Gp.6 Ditto + 300 wt% un-ammoniated zeolitite 24.51
Statement
The root systems in the low density group show a decline in root mass above an
excess
addition of 150 wt% un-ammoniated zeolitite.
As in the case of the shoot mass a distinct cut-off is seen in root growth. In
the present case,
working with heavy metals in the specified range in neutral to slightly
alkaline conditions no
advantage is gained in increasing tire excess zeolitite above 150 wt %.
Whereas shoot growth
is maximised at 225 wt% excess zeolitite.
This again demonstrates that the tdo-fertilizer can be formulated to maacimise
either shoot or
root growth.
In order to formulate the bio-ferti'(izer for these effects it will be
necessary to conduct initial
laboratory trials as the biological factors (i.e. remedialplantspecies and
density), bacterial
population, soil properties, heavy metal species and concentrations will vary
independently at
specific sites. .
Analytical data: Root dry weights
(i) Low density group harvested at five months.
Plant substrate Root weieht (g)
Gp. l Toxic substirate 3.45
Gp.2 Amended substrate (16 -17) vol% zeolite bio-fertilizer 6.53
Gp.3 Ditto + 75 wt% un-ammoniated zeolitite 8.43
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12
Gp.4 Ditto + 150 wt% un-ammoniated zeolitite 11.80
Gp.S Ditto + 225 wt% un-ammoniated zeolitite 11.67
Gp.6 Ditto + 300 wt% un-ammoniated zeolitite 7_q.4
Statement
Harvested plant shoots in the high density group were analysed for trace
concentrations of
Zinc, Copper and Lead. By relating the trace metal chemistry to the shoot dry
weight of the
amended groups it was found that the cut-off limit corresponded to a
concentration of 138
ug.g't Zinc. Above this value shoot dry weight dropped dramatically.
Analytical data: Trace metal chemistry in shoots
Plant substrate pg. ~ t
Zn Pb Cu
Gp.Toxic subsuate . 74.2 6.7 16.1
l
Gp.2Amended substrate (16-17) vol% zeolite106.7 6.8 22.2
bio-fertilizer
Gp.3Ditto + 75 wt% un-ammoniated zeolitite138.1 6.1 26.0
Gp.4Ditto + 150wt% un-ammoniated zeolitite140.6 7.6 24.6
Gp.SDitto +'?25 wt% un-ammoniated zeolitite163.4 6.5 22.9
Gp.6Ditto + 300 wt% un-ammoniated zeolitite170.0 9.4 23.
Statement
A large increase in the elecorical conductivity is seen to occur between
leachate solutions
from the toxic and amended substrates; which is chazacteristic of soils
amended with the bio-
fertilizer.
This reaction is taken to infer that the nitrifying bacteria are boosted by
addition of the bio-
fertilizer. .... .
A relationship is seen between shoot total nitrogen and excess zeolitite. The
data shows that a
fluctuation occurs between the amended substrates throughout the range of
excess zeolitite.
This behaviour is not clearly understood but is apparently a function of the
bacterial
composition developed in the amended substrates.
CA 02354249 2001-11-13
13
Analytical data: Eleca-ical conductivity of substrate leachate solutions and
shoot total
nitrogen
Plant substrateE.C.(u Siemens/cmShoot total nitrosen
lwt,%1
(Low density) (High
decLSiry)
Gp.l 119 0.74 0.98
GP~2 472 1.=18 1.~~:
Gp.3 285 1.17 1.39
Gp.4 323 1.37 l.-~7
Gp.~ l89 1.09 1.38
Gp.6 488 1.13 1.32
Comments oa results
The plant weight results demoruu-ate the performance of ryegiass grown in
heavy metal
polluted soils of neutral to slightly alkaline pH that has been amended with
zeolite bio-
fertil izer.
Very large increases in plant growth occur in the amended substrates. The
increase obtained
in root growth is particularly important for the retention of soil particles
contaminated by
heavy metal residues.
However, limits are reached in the addition of un-ammoniated zeolitite above
which the
plants suffer deleterious effects.
As the zinc concentrations found in the shoots correlates to the behaviour of
the shoot dry
weights this element is thought to causing a major phytotoxic effect which is
limiting growth.:
Although the detailed interactions between the bio-fectilizer, soil chemistry
and bacterial
population are st~I under investigation it is clear that by studying the
effect of varying the
concentration of the un-ammoniated zeolitite the bio-fettilizer can be
formulated to provide
maximum growth enhancement .
r
