WO2012140623A1

WO2012140623A1 - Method for phytoremediation of heavy metal contaminated sites

Info

Publication number: WO2012140623A1
Application number: PCT/IB2012/051846
Authority: WO
Inventors: Claudia Ortiz; Marcela WILKENS; Sergio CASTRO; Raúl ESPINACE; Pamela VALENZUELA; Juan PALMA
Original assignee: Universidad De Santiago De Chile
Priority date: 2011-04-13
Filing date: 2012-04-13
Publication date: 2012-10-18
Also published as: AU2012241367A1; CA2833153A1; CA2833153C; PE20140857A1; CL2011000826A1; AU2012241367B2; BR112013026388A2

Abstract

The invention relates to a method for recovering substrates, mainly mine tailings, using plant species endemic to the location of the substrate to be treated. The substrate recovery method essentially comprises the following steps consisting in: selecting the endemic plant species capable of growing in substrates containing heavy metals; performing a tolerance test on said plant species, finishing with a new selection of the most tolerant species; vegetatively propagating the selected endemic plant species; acclimating the vegetatively propagated species and installing the plant species in the soil of the substrate to be treated. The present method is particularly suitable for use in substrates obtained from tailings, i.e., those containing a high concentration of metals.

Description

DESCRIPTIVE MEMORY

Field of application of the invention

The present invention relates to the recovery of substrates presenting industrial wastes, particularly mining, such as tailings.

Most mining tailings are transformed into lands that are abandoned and unused, with a low probability of being recovered. The present application shows a method for recovering said land, which is based on the use of endemic and non-endemic native plant species, which can grow on substrates containing industrial wastes, such as heavy metals such that the roots of said plants they can act as phytostabilizers of the land, and the trunk, stem and leaves, can act as phytoextractors. In addition, it is related to the use of said plant species to recover said substrates.

Comments of the prior art

Many of the industries, especially in the chemical and mining area, result in both organic and inorganic wastes, which are sent to conditioned land and dumps, these lands being mostly abandoned and unused, generating accumulation of toxic substances that are finally transform into pollution sources.

In the case of mining, it is common to find tailings that mostly correspond to toxic waste, by-products of mining processes, with a high concentration of minerals, which are usually mixed with soil, other minerals, water and rocks. At present, there have been several attempts to recover these lands with plant species. For example, in US 5,785,735 a process for removing metal ions from the soil and methods for effecting such removal is disclosed. This process is based on the manipulation of the culture medium, where effective amounts of chelating agents are added and the use of a crop of the Brassicaceae family of plants, so that the metals found in the soil are made more available for the plants. These plants absorb metals in their roots.

Also, US 6,159,270 discloses a process for removing metal ions from the soil using plants and methods to effect such removal. Said removal is performed using a chelating agent in the medium where the removal is carried out, and Brassicaceae plants are used. US 7,034,202 discloses a method and compositions used for phytoremediation of heavy metals, including plants that are genetically modified to overexpress glutamylcysteine synthetase (ECS) thereby increasing the accumulation of heavy metals in said plants.

EP 1 260 282 discloses a method for the phytoremediation of a medium contaminated with organic substances, heavy metals, radionuclides or a mixture thereof, which includes the cultivation in said contaminated medium of a plant and the recovery of the elements present in said plant.

US 2005/0198707 discloses methods, recombinant DNA molecules, recombinant host cells containing the molecules of DNA, plant tissue and plants that contain and express at least one sequence encoding biosynthetic phytokelatin, under the regulatory control of the ACT2 constitutive promoter or the light-inducible SRS1 promoter. Transgenic plants are tolerant of heavy metal ions and can accumulate them from contaminated media, so it is possible to perform phytoremediation of a solid substrate or contaminated liquid that contains them.

US 2007/0093388 discloses a method for phytoremediation of a contaminated site by at least one type of contaminant, which comprises cultivating at least one phytoremediation plant capable of fixing at least a portion of the contaminant and harvesting or destroying the site. plant that has fixed the contaminant.

US 2009/015591 1 discloses genetically modified plants capable of accumulating heavy metals in the stems, together with methods to remove and possibly recover said heavy metals, using said genetically modified plants.

US 2008/0289252 discloses a method of using genetically transformed plants in phytoremediation of lead by growing the transformed plants in the contaminated medium.

Although, there are documents in the prior art that describe the recovery of contaminated land, using planting of plant species that can grow under conditions of heavy metal substrates, among other pollutants, none of them describe decontamination with plant species from it zone, or endemic native plants and not endemic to the substrate to be decontaminated, where a previous and specific stage of acclimatization of the plant species in the substrate where they will definitely be planted is carried out.

Indeed, in the prior art, plant species whose mineral extraction properties are already known are used and then, a methodology for using them has been implemented. In other documents, genetic intervention is mentioned in certain plant species to improve extraction.

However, not all of these plants give good results for the treatment of substrates in tailings lands because the mining deposits are in inhospitable places, where plant species can have an extremely short life.

Although, there are publications that describe potential uses of native and / or endemic species to treat substrates contaminated with different metals, do not describe the methodology associated with the use, do not include selection processes of plant species and do not disclose stages of acclimatization of species vegetables. Nor do they disclose the ability of plant species to incorporate a metal into their tissues.

The objective of the method of the present invention is to recover substrates contaminated with industrial wastes, particularly mining wastes by selecting endemic and non-endemic native plant species that have grown on the substrate to be treated. Subsequently, a tolerance test for said plant species is carried out in the laboratory from which a new selection of species, which are reproduced vegetatively and then acclimatized to finally install them in the substrate to be treated.

Summary of the invention

The present invention relates to a method for the recovery of substrates contaminated with industrial wastes, mainly with mining tailings through the use of endemic and non-endemic native plant species from the location of the substrate to be treated.

The substrate recovery method basically comprises the steps of: selecting plant species that are capable of growing on substrates with the presence of industrial wastes, such as heavy metals; carry out a tolerance test with these plant species, which leads to a new selection of those most tolerant species; vegetatively reproduce said selected plant species; acclimatize vegetatively propagated species and install plant species in the soil of the substrate to be treated.

The method of the present invention is especially used in substrates contaminated with industrial wastes, such as tailings, that is, those containing a high concentration of metals, such as, for example, Cd, Pb, Zn and Cu, among others.

Brief description of the drawings

The accompanying drawings, which are included to provide a greater understanding of the invention, being incorporated and constituting part of this description, illustrate an embodiment of the invention, and together with the description, serve to explain the principles of the invention. Figure N ^Q 1 shows a diagram of the installation design of a phytostabilization plan with location of prets and water tanks on the surface to be treated.

Figure N ^Q 2 shows a diagram of the installation scheme of the irrigation lines, the plant species (Sp 1 -5) and the wind erosion and wind speed evaluation points in the phytostabilization plan of the surface to be treated.

Figure N ^Q 3 shows a detail of the installation scheme of the irrigation lines in the phytostabilization plan of the surface to be treated.

Description of the invention

The present invention relates to a method for the recovery of substrates with industrial wastes, particularly mining, such as tailings, using native endemic and non-endemic native plant species for said recovery.

Tailings are toxic wastes, byproducts of mining processes that therefore contain a high concentration of minerals. Usually these tailings substrates correspond to a mixture of earth, minerals, water and rocks. These lands, due to their characteristics, are often abandoned and unused, generating accumulation of toxic substances in certain physical spaces, which eventually become sources of pollution.

To eliminate these industrial wastes in the present invention, a method for recovering said substrates, such as tailings substrate, by using endemic and non-plant species is described endemic, those that can clearly grow in said substrates and those that can retain high concentrations of heavy metals in such a way that the roots of said plants can act as a phytostabilizer of the substrate and, the trunk, stem and leaves, can act as a phytoextractor of the substrate minerals.

The method comprises a first stage of selection of the endemic and non-endemic native plant species present in the substrate to be recovered with the presence of industrial wastes, such as heavy metals.

These plant species belong to species included in the Poaceae family such as Polypogon Australis and Stipa plumosa, the Chenopodiaceae family as Atríplex deserticola and the Asteraceae family such as Baccharis salicifolia. Of these, Polypogon australis is the species that exhibits the best performance in laboratory tests, since its growth is faster in time compared to the rest of the plant species. The species that have been evaluated are those that grow on tailings substrates. These species have rapid growth as they are herbaceous and shrubby plants. As an example, it is mentioned that the copper accumulation rate for P. australis is 0.48 mg / day according to results obtained at the laboratory level, using synthetic copper solutions at pH 5.0. The growth of the roots of these species, moreover, allows to mechanically stabilize the substrates of fine particulate material, which presents a risk of dispersion by wind action. The environmental plasticity exhibited by these species makes them particularly interesting for treatments in systems with low water requirements, high solar radiation and day / night temperature differences of more than 20 ° C. They do not have large nutrient requirements and are tolerant to saline soils. Also, it should be noted that the work with endemic and non-endemic native plants facilitates the adaptation of the species to acid substrates with high levels of copper, highly saline and with low organic matter content. In addition, minimizing the use of water resources in arid and semi-arid areas, minimizes the use of organic amendments for plant establishment and minimizes the environmental impact of the technological application, since they are not invasive plants that would affect ecosystems fragile as those in arid areas. The use of these plant species allows, therefore, to improve both the chemical and mechanical properties of the substrate to be treated.

To achieve this selection it is necessary to define the study substrate; to then characterize said substrate through its organic matter, NPK index, pH, electrical conductivity, exchangeable bases Na, Ca, Mg and K as a measure of the Cation Exchange Capacity, metal content; and through the mechanical characteristics of the substrate such as texture, structure, density and temperature. Next, a sampling and characterization of the plant species that have grown in the place is carried out, to then calculate the R index of these species. The R index corresponds to the ratio of accumulation of a sheet metal versus roots. Finally, a ranking of use is made for each metal and the plant species is chosen considering the R index and the treatment that you want to apply to the substrate. The R value is determined by the distribution of the metal (such as Cu) in aerial tissue: root tissue (leaves: roots). An R value greater than 1 is an indicator of a species with characteristics suitable for use in metal extraction (phytoextraction). A value of R less than 1 indicates a species with characteristics suitable for use in metal stabilization (phytostabilization). Therefore, according to the type of recovery that you want to implement in the substrate to be treated, it will be the value of R selected.

In a second stage, a tolerance test is carried out with these plant species, generating a new selection with those more tolerant species. For this, seeds of the selected species are sown in the first stage, in a liquid medium that is composed of vitamins such as Inositol in a concentration between 5 and 10 g / L, Nicotinic acid at a concentration between 0.01 and 0.05 g / L, HCI-Pyridoxine at a concentration between 0.01 and 0.05 g / L, Glycine at a concentration between 0.1 and 0.3 g / L, HCI-Thiamine at a concentration between 0.005 and 0.01 g / L; microelements such as MnS0 ₄ -H ₂ 0 at a concentration between 1 and 2 g / L, ZnS0 ₄ -7H ₂ 0, H ₃ B0 ₃ at a concentration between 0.5 and 0.7 g / L, Kl at a concentration between 0.05 and 0.1 g / L, Na ₂ MoO ₄ -2H ₂ 0 at a concentration between 0.02 and 0.03 g / L, CuS0 ₄ -5H ₂ 0 at a concentration between 2.5 x10 ^"4 and 3 x 10 ^"4 g / L, CoCI ₂ -6H20 at a concentration between 2.5 x10 ^{" 4} and 3 x 10 ^"4 g / L; macroelements such as NH ₄ N0 ₃ at a concentration between 15 and 17 g / L, KN0 ₃ at a concentration between 15 and 20 g / L, CaCI ₂ -2H ₂ 0 at a concentration between 4 and 5 g / L, MgS0 ₄ - 7H ₂ 0 at a concentration between 3 and 4 g / L, KH ₂ P0 ₄ at a concentration between 1 and 1.5 g / L; and a carbon source such as 2% sucrose. He The medium also contains increasing concentrations of the contaminating substance, that is, constant numbers of plants are subjected to different concentrations of the contaminating substance, until they reach a lethal concentration. To perform the test, the pH is controlled between 5.0 and 7, the temperature between 20 and 25 ^Q C, and the photo period, with a light regime of 14/12 (14 to 12 hours of light daily). Finally, a new ranking is generated with the species with the highest tolerance for the substrate, that is, after 2 weeks of cultivation, the growth of the roots is measured as well as that of the aerial part of the plant and the content is determined of chlorophyll of each plant. With these three parameters described above, the ranking is constructed and one or more plant species are chosen to be used in the treatment of the substrate to be treated.

As there are two stages of plant species selection, the technological risk of implementation is minimized. These selection stages allow working with genetically optimized individuals because of their tolerance characteristics to acid substrates with high levels of metals, such as copper, highly saline and with low organic matter content. These stages are the general cadastre, evaluation of potential use and a second stage of tolerance in the laboratory for the selection of more tolerant clones. It is also important to note that, for this same reason, no organic amendments or amendments are used to improve soil pH, given that the tolerance characteristics of the plants and their acclimatization favor the survival of the plant species in the selected substrate . That is, it is not necessary add any reagent that modifies the content of organic matter or modifies the pH value of the substrate.

In order to perform the installation and phytoremediation, it is necessary to vegetatively reproduce the plant species or species selected in the second stage. Therefore, in a third stage a vegetative reproduction is carried out according to two techniques to choose: by cloning by cuttings and / or by tissue culture "in vitro".

For the cloning technique by cuttings, use is made of a liquid substrate containing vitamins such as Inositol in a concentration between 5 and 10 g / L, Nicotinic acid at a concentration between 0.01 and 0.05 g / L, HCI- Pyridoxine at a concentration between 0.01 and 0.05 g / L, Glycine at a concentration between 0.1 and 0.3 g / L, HCI-Thiamine at a concentration between 0.005 and 0.01 g / L; microelements such as MnS0 ₄ -H ₂ 0 at a concentration between 1 and 2 g / L, ZnS0 ₄ -7H ₂ 0, H ₃ BO ₃ at a concentration between 0.5 and 0.7 g / L, Kl at a concentration between 0.05 and 0.1 g / L, Na ₂ MoO ₄ -2H ₂ 0 at a concentration between 0.02 and 0.03 g / L, CuS0 ₄ -5H ₂ 0 at a concentration between 2.5 x10 ^"4 and 3 x 10 ^"4 g / L, CoCI ₂ -6H20 at a concentration between 2.5 x10 ^{" 4} and 3 x 10 ^"4 g / L; macroelements such as NH ₄ N0 ₃ at a concentration between 15 and 17 g / L, KN0 ₃ at a concentration between 15 and 20 g / L, CaCI ₂ -2H ₂ 0 at a concentration between 4 and 5 g / L, MgS0 ₄ - 7H ₂ 0 at a concentration between 3 and 4 g / L, KH ₂ P0 ₄ at a concentration between 1 and 1.5 g / L; and a carbon source such as 2% sucrose; all of the above at a pH that varies between 5.0 and 7.0, and at a temperature that varies between 20 and 25 ° C. For the "in vitro" tissue culture technique, a nutrient medium consisting of vitamins such as Inositol is used in a concentration between 5 and 10 g / L, Nicotinic acid at a concentration between 0.01 and 0.05 g / L, HCI-Pyridoxine at a concentration between 0.01 and 0.05 g / L, Glycine at a concentration between 0.1 and 0.3 g / L, HCI-Thiamine at a concentration between 0.005 and 0.01 g / L; microelements such as MnS0 ₄ -H ₂ 0 at a concentration between 1 and 2 g / L, ZnS0 ₄ -7H ₂ 0, H ₃ BO ₃ at a concentration between 0.5 and 0.7 g / L, Kl at a concentration between 0.05 and 0.1 g / L, Na ₂ MoO ₄ -2H ₂ 0 at a concentration between 0.02 and 0.03 g / L, CuS0 ₄ -5H ₂ 0 at a concentration between 2.5 x10 ^"4 and 3 x 10 ^"4 g / L, CoCI ₂ -6H20 at a concentration between 2.5 x10 ^{" 4} and 3 x 10 ^"4 g / L; macroelements such as NH ₄ N0 ₃ at a concentration between 15 and 17 g / L, KN0 ₃ at a concentration between 15 and 20 g / L, CaCI ₂ -2H ₂ 0 at a concentration between 4 and 5 g / L, MgS0 ₄ - 7H ₂ 0 at a concentration between 3 and 4 g / L, KH ₂ P0 ₄ at a concentration between 1 and 1.5 g / L; and a carbon source such as 2% sucrose and a combination of hormones such as auxins greater than 0.05 μΜ and cytokinins greater than 1 mg / L, on a solid sterile support as agar at a pH that varies between 5.0 and 7, 0, and at a temperature that varies between 20 and 25 ° C.

Before installing the plant species in the substrate to be treated, it is necessary to acclimatize said vegetatively reproduced species, through a process that involves subjecting said plants that have roots of at least 15 cm in length, to a hardening treatment in variable proportions between 20 and 80% of substrate to be treated and inert substrate, for at least three weeks, at a temperature that varies between 20 and 25 ° C with a photo period of 14 light hours / 10 dark hours. Non-metallic minerals are used as an inert substrate, such as perlite.

Said acclimatization phase of the selected clones allows maximizing the success of the installation. This acclimatization phase is carried out with the same substrate to be treated so that the establishment of plant species is highly successful. We can mention, as an example that an establishment on land of up to 72% was obtained for species of the genus Atriplex in 13 months.

Finally, the field installation of each of the acclimatized species is carried out, for this purpose the soil of the substrate to be treated with stakes is delimited; then, the access roads are established and the substrate is homogenized. Samples of the homogenized soil are taken to take to the laboratory where the content of at least 5 metallic elements (copper, zinc, iron, cadmium and nickel) and their bioavailability are analyzed; the content of organic matter, phosphorus, nitrogen, pH and electrical conductivity, in addition to interchangeable bases (Na, Ca, Mg and K as a measure of Cation Exchange Capacity).

Then, the irrigation system is installed in the soil of the substrate to be treated, which consists of a comb drip system with main pipes between 0 and 1 "(0 to 2.54 cm) and secondary pipes between 0 and ½ "(0 to 1, 27 cm). This system is operated with an initial irrigation regime of between 0 and 1 liter per specimen, with intermittent regime and with pause between irrigation. The pause between irrigation is one day during the first month, and the irrigation is done with a volume that varies between 0.3 and 0.5 L per plant, between two to three times a day. Finally, the irrigation system includes constantly monitoring the regime and amount of water provided.

In addition, if the irrigation system is installed, instruments for measuring temperature, humidity, and wind direction and speed are located, such as thermometers and anemometers.

Next, the already acclimated species are planted, with a planting density between 1 and 3 plants / m ² , with a separation distance between plants of between 20 and 50 cm in a batch distribution.

For a better monitoring of the installation system, a georeferenced survey of the land and plantations is carried out (each plant species will be associated with a point), which will allow monitoring the terrain through GPS in order to locate the lots and species planted in them.

If there are plant species that suffered a catastrophic event, such as: frost, high temperatures, floods or winds, the missing species will be replaced. However, if all the plant species associated with a lot suffered a catastrophic event, these species are replaced by another group of plant species that have endured the catastrophic event.

In a systematic way, substrate samples are collected every 3 meters in an area that varies between 50 m ² and the total planted area. The samples are taken from up to a depth of between 10 and 20 cm, collecting a fresh substrate weight of up to 1.4 kg. The corresponding analyzes already described above are performed, which correspond to: analysis of content of at least 5 metallic elements, such as copper, zinc, iron, cadmium and nickel, and their bioavailability; analyze content of organic matter, phosphorus, nitrogen, pH and electrical conductivity; in addition to interchangeable bases (Na, Ca, Mg and K as a measure of Cation Exchange Capacity). Finally, it is determined how the soil conditions vary, after planting these species in the substrate.

As an alternative to the present application, it is possible to extract from the plant material the metal or metals that have been deposited both in the root and in the stems and leaves.

To improve the understanding of this application, the following example is presented.

Example 1

Nine species of vascular plants were collected in a 250 m "area of a substrate to be treated, in this case, in northern Chile. Five of these species were identified as native: Baccharis salicijolia, Schinus polygamus, Atriplex deserticota, Scirpus asper and Polypogon australis, and four were identified as non-native species: Casuarina equis etifolia, Acacia melanoxylon, Pennisetum clandestinum and Cynodon dactylon.Of the nine species young and adult plants were taken, of which four grasses were found under the tree canopy. Atriplex deserticola plants were found on the slope of the tailings deposit.

All these species correspond to the families of the species belonging to the Poaceae family such as Polypogon Australis, the Chenopodiaceae family as Atriplex deserticola and the Asteraceae family as baccharis salicijolia. Copper was the most abundant metal in the soil, where the level of this metal was determined at 1.0 to 2.0 g (fresh weight) of leaves and roots of the nine species mentioned. The amount of copper in the leaves and roots in mg kg ^"1 by weight of dried plant was used to calculate the ratio of copper leaves: roots (L: R). The copper content in the stem was not determined. In the Table 1 shows the determined L: R ratios, which ranged from 0.2 (C. equis etifolia) to 9.4 (B. salicifolia), according to the following table:

Table 1 Copper in sheets and

Relationship

Copper roots content

Vegetable species Leaves: roots

(mg kg ^-1 dry weight (mf kg ^"1 weight

(H: R) dry)

Root Leaves

Baccharis salicifolia

667.9 ± 2.4 71.3 ± 0.4 739.2 9.4 (PWS, As)

Schinus polygamas (P,

1213.5 ± 8.9 260 ± 0.9 1473.5 4.7 Ana)

Casuarina

470.2 ± 3.3 2923.3 ± 7.7 3393.5 0.2 equisetifolia (P, Ca)

Atriplex deserticota

1357.7 ± 3.5 2160 ± 71 3517.7 0.6 (PWS, Che)

Acacia melanoxylon

158.6 ± 1.6 484.0 ± 8.9 642.6 0.3 (P, Le)

Pennisetum

259.2 ± 6.2 866.4 ± 7.1 1125.6 0.3 clandestinum (G, Po)

Scirpus Asper

459.3 ± 5.0 205.7 ± 6.1 710 1.8 (G, Cy)

Cynodon dactylon (G,

246.5 ± 8.1 81.4 ± 0.6 327.9 3.0 Po)

Polypogon australis

669.5 ± 5.9 223 ± 16 892.5 3.0 (G, Po) From this Table it can be observed that those species that obtained an R less than 1, are those species that have a phytostabilizer behavior.

The following shows, in Table 2, the results obtained of copper content for a substrate treated with the method of the present application, where the recovery of the substrate from both chemical and mechanical points of view is clearly observed.

Table 2

In addition, since the electrical conductivity is an indicator of salinity, it can be seen that the plant species, once selected, can be used in acidic substrates, with high levels of copper, highly saline and with a low content of organic matter.

Claims

1. Method for recovering substrates with industrial wastes, particularly mining, CHARACTERIZED because it comprises:

to. select plant species that are capable of growing on substrates with the presence of industrial wastes, such as heavy metals; b. perform a tolerance test with these plant species, end with a new selection of those species more tolerant to the substrate;

C. vegetatively reproduce the plant species selected in stage b;

d. acclimatize vegetatively propagated species in stage c; e e. install acclimatized plant species in the field in stage d.

2. Method for recovering substrates with industrial wastes, particularly mining, according to claim 1, CHARACTERIZED because the selection of plant species is carried out following the steps of:

- define the study site;

determine on the study site the content of organic matter, NPK index, pH, electrical conductivity, interchangeable bases such as Na, Ca, Mg and K as a measure of Cation Exchange Capacity, metal content, and obtain soil characteristics, such as texture, structure, density and temperature;

sampling and determining the qualities of plant species located on the ground with the substrate to be treated; obtain the R index of the plant species collected in the soil, that is, the ratio of metal accumulation in leaves versus roots; make a ranking of use for each metal, using the R index; and choose the plant species considering the R index, according to the treatment process to be applied.

3. Method for the recovery of substrates with industrial wastes, particularly mining, according to claim 1, CHARACTERIZED because performing the tolerance test with said plant species by the following steps:

- sow seeds of the species selected in stage a, in a liquid medium that is composed of vitamins such as Inositol, Nicotinic, HCI-

Pyridoxine, Glycine, HCI-Thiamine; microelements such as MnS0 ₄ -H20, ZnS0 ₄ -7H ₂ 0, H ₃ BO ₃ , Kl, Na ₂ MoO ₄ -2H ₂ 0, CuS0 ₄ -5H ₂ 0, CoCI ₂ -6H20; macroelements such as NH ₄ N0 ₃ , KN0 ₃ , CaCI ₂ -2H ₂ 0, MgS0 ₄ -7H ₂ 0, KH ₂ P0 ₄ ; and a carbon source such as sucrose;

- incorporate increasing concentrations of the contaminating substance into the liquid medium, that is, sow constant numbers of plant seeds at different concentrations of the contaminating substance, until reaching a lethal concentration;

- control the pH between 5.0 and 7, the temperature between 20 and 25 ^Q C and the photo period with a light regime 14/12;

- measure the growth of the roots, the growth of the aerial part of the plant and determine the chlorophyll content after two weeks; Y - design a new ranking and choose the species with the highest tolerance to the substrate to be treated based on the three previous parameters.

Method for the recovery of substrates with industrial wastes, particularly iron, according to claim 1, CHARACTERIZED because the reproduction; etative, is carried out by the following mechanisms:

- clone by means of cuttings, using a liquid substrate containing vitamins such as Inositol in a concentration between 5 and 10 g / L, Nicotinic acid at a concentration between 0.01 and 0.05 g / L, HCl-Pyridoxine at a concentration between 0.01 and 0.05 g / L, glycine at a concentration between 0.1 and 0.3 g / L, HCl-Thiamine at a concentration between 0.005 and 0.01 g / L; microelements such as MnS0 ₄ H ₂ 0 at a concentration between 1 and 2 g / L, ZnS0 ₄ -7H ₂ 0, H ₃ BO ₃ at a concentration between 0.5 and 0.7 g / L, KI at a concentration between 0 , 05 and 0.1 g / L, Na ₂ MoO ₄ -2H ₂ 0 at a concentration between 0.02 and 0.03 g / L, CuS0 ₄ -5H ₂ 0 at a concentration between 2.5 xlO ^"4 and 3 x 10 ^"4 g / L, CoCl ₂ -6H20 at a concentration between 2.5 xlO ^{" 4} and 3 x 10 ^"4 g / L; macroelements such as NH ₄ N0 ₃ at a concentration between 15 and 17 g / L, KN0 ₃ at a concentration between 15 and 20 g / L, CaCl ₂ -2H ₂ 0 at a concentration between 4 and 5 g / L, MgS0 ₄ - 7H ₂ 0 at a concentration between 3 and 4 g / L, KH ₂ P0 ₄ at a concentration between 1 and 1.5 g / L; and a carbon source such as 2% sucrose; at a pH that varies between 5.0 and 7.0 at a temperature that varies between 20 and 25 °; I

- cultivate tissues "in vitro", using a nutritive medium consisting of vitamins such as Inositol at a concentration between 5 and 10 g / L, Nicotinic acid at a concentration between 0.01 and 0.05 g / L, HCl-Pyridoxine at a concentration between 0.01 and 0.05 g / L, glycine at a concentration between 0.1 and 0.3 g / L, HCI-Thiamine at a concentration between 0.005 and 0.01 g / L; microelements such as MnSO ₄ -H ₂ O at a concentration between 1 and 2 g / L, ZnSO ₄ -7H ₂ O, H ₃ BO ₃ at a concentration between 0.5 and 0.7 g / L, Kl at a concentration between 0.05 and 0.1 g / L, Na ₂ MoO ₄ -2H ₂ 0 at a concentration between 0.02 and 0.03 g / L, CuS0 ₄ -5H ₂ 0 at a concentration between 2.5 x10 ^"4 and 3 x 10 ^"4 g / L, CoCI ₂ -6H20 at a concentration between 2.5 x10 ^{" 4} and 3 x 10 ^"4 g / L; macroelements such as NH ₄ N0 ₃ at a concentration between 15 and 17 g / L, KNO ₃ at a concentration between 15 and 20 g / L, CaCI ₂ -2H ₂ 0 at a concentration between 4 and 5 g / L, MgS0 ₄ - 7H ₂ 0 at a concentration between 3 and 4 g / L, KH ₂ P0 ₄ at a concentration between 1 and 1.5 g / L; and a carbon source such as 2% sucrose and a combination of hormones such as auxins greater than 0.05 μΜ and cytokinins greater than 1 mg / L in a solid sterile support as agar at a pH that varies between 5.0 and 7.0 , and at a temperature that varies between 20 and 25 ° C.

5. Method for the recovery of substrates with industrial wastes, particularly mining, according to claim 1, CHARACTERIZED because the acclimatization process of vegetatively propagated species consists in subjecting vegetatively reproduced species in stage c, presenting roots of at least 15 cm in length, at a hardening treatment in varying proportions between 20 and 80% of the substrate to be treated and inert substrate, for at least three weeks at a temperature that varies between 20 and 25 ° C with a photo period of 14 Light hours / 10 hours dark.

6. Method for recovering substrates with industrial wastes, particularly mining, according to claim 5, CHARACTERIZED because in the acclimatization process of vegetatively propagated species, the inert substrate used corresponds to non-metallic minerals, such as perlite. .

7. Method for recovering substrates with industrial wastes, particularly mining, according to claim 1, CHARACTERIZED because the following steps are carried out in the field installation:

- delimit the land with the substrate to be treated, with stakes, establishing the access roads within the land;

- homogenize the substrate to be treated;

- take samples of the homogenized soil to analyze the content of at least 5 metallic elements, such as copper, zinc, iron, cadmium and nickel, and their bioavailability; the content of organic matter, phosphorus, nitrogen, pH and electrical conductivity, together with the following interchangeable bases such as Na, Ca, Mg and K as a measure of the Cation Exchange Capacity;

- install an irrigation system in the soil of the substrate to be treated;

- install instruments to measure wind speed and direction; temperature and humidity;

- plant the acclimatized species of stage d, with a planting density between 1 and 3 plants / m ² , with a separation distance between plant species between 20 and 50 cm, in a batch distribution; - conduct a georeferenced survey of the land and plantations, allowing each plant species to be associated with a point on a terrain plan;

- monitor the terrain through GPS, in order to locate the lots and the species planted in them;

- replace the missing plant species, if there are plants that suffered a catastrophic event, such as frost, high temperatures, floods or winds;

- replace the plant species of the group of species associated with a lot that suffered a catastrophic event, with another group of plant species that has endured the catastrophic event;

- take samples of the substrate every 3 meters, in an area that will vary between 50 m ² and the total planted area, up to a depth of between 10 and 20 cm, collecting a fresh substrate weight of up to 1.4 kg;

- perform analysis of the organic matter content, NPK index, pH, electrical conductivity, interchangeable bases such as Na, Ca, Mg and K as a measure of Cation Exchange Capacity, metal content, and obtain soil characteristics, such as texture , structure, density and temperature; Y

Determine how much the ground conditions varied.

8. Method for recovering substrates with industrial wastes, particularly mining, according to claim 7, CHARACTERIZED because the irrigation system consists of a comb drip system with main pipes between O and 1 "(O at 2.54 cm) and secondary pipes between O and ½" (O at 1, 27cm);

9. Method for recovering substrates with industrial wastes, particularly mining, according to claims 7 and 8, CHARACTERIZED because it comprises operating the irrigation system with an initial irrigation regime of between 0 and 1 L per specimen, with regime intermittent and with pause between risks.

10. Method for the recovery of substrates with industrial wastes, particularly mining, according to claim 7 to 9, CHARACTERIZED because it comprises making an increase in the pause of the irrigation regime of one day during the first month with a volume between 0, 3 and 0.5 L per plant two to three times per day.

eleven . Method for recovering substrates with industrial wastes, particularly mining, according to claims 10, CHARACTERIZED because it comprises watering the plants between 0 and 60 minutes at a time.

12. Method for recovering substrates with industrial wastes, particularly mining, according to claims 7 to 1, CHARACTERIZED because it comprises constantly monitoring the regime and amount of water provided in the soil of the substrate to be treated

13. Method for recovering substrates with industrial wastes, particularly mining, according to claim 1, CHARACTERIZED because it also comprises the step of extracting the metal recovered by the aerial part of the plant, that is, trunk, stem and leaves.

14. Use of plant species that grow on a substrate to be treated CHARACTERIZED because they serve for the recovery of substrates with industrial wastes, particularly mining, using the method of claim 1.