CN109574230B - MgCl2And KNO3Application of plant in improving cadmium enrichment capacity - Google Patents

Abstract

The invention discloses MgCl₂And KNO₃The new application of (1), namely the application of the cadmium-enriched powder in efficiently and rapidly improving the cadmium-enriching capacity of plants; the experimental results show that MgCl is used₂And KNO₃The treated plant has enhanced absorption of cadmium ion in the culture solution, and the plasma membrane H⁺Enhancement of ATPase Activity, MgCl₂And KNO₃The combined treatment promotes the absorption and enrichment of cadmium ions by plants.

Description

MgCl2And KNO3Application of plant in improving cadmium enrichment capacity

Technical Field

The invention belongs to the technical field of improving phytoremediation of heavy metal pollution, and particularly relates to an inorganic chemical reagent MgCl₂And KNO₃The new application of the cadmium-enriched plant extract in improving the cadmium enrichment capacity of plants.

Background

Density of 5 × 10^-3 kg/m³The above metals are called heavy metals, and there are about 45 kinds, such as cadmium (Cd), copper (Cu), mercury (Hg), zinc (Zn), and the like. In recent years, along with the rapid development of industry, the removal of a large amount of heavy metals leads to the continuous increase of the content of heavy metals in the environment, and has great harm to the health of people and animals; heavy metals in the environment cannot be decomposed and utilized by microorganisms, animals and plants absorb the heavy metals through air, water, soil and the like, the heavy metals are amplified and enriched into a human body through a food chain and a food net, and the health of the human body can be damaged by extremely low concentration; heavy metal contamination has become a worldwide environmental problem.

In recent years, sudden heavy metals have intensified environmental pollution events, so that the heavy metals in the environment are increased in a short time, and serious heavy metal pollution is generated. The harm of heavy metal pollution has the following two aspects: 1) the heavy metal harms the soil, and firstly, the heavy metal harms microorganisms in the soil, so that the number of the microorganisms in the soil is reduced; and secondly, the plants destroy tissues and functions after absorbing heavy metals, so that the plants are weakened in growth vigor, and meanwhile, the heavy metals can be enriched through a food chain and a food net. 2) Heavy metals are harmful to water environment, have high toxicity and are not easy to degrade, and after entering the water environment, people can accumulate the heavy metals in vivo through drinking water and can enrich the heavy metals through a food chain and a food net.

Researches show that the form of heavy metal absorbed by plants is mainly in a free ion state, and heavy metal Cd in soil has various forms such as a water dissolution state, an exchangeable state, a carbonate binding state, a silicate binding state and the like. Wherein the water soluble state and the exchangeable state are effective states for the enrichment of heavy metals by plants. In the process of absorbing metal ions by plants, there are two types; (1) passive transportation: the passive transportation includes two processes of cation exchange and diffusion. Heavy metal Cd exists in a root system environment, when the plant root system epidermal cells absorb Cd, the reversible exchange process with the external environment is cation exchange, and when Cd enters the epidermal cell walls and then enters cell membranes, the process is a diffusion process. (2) Active transportation: active transport requires carriers and energy to be part of heavy metals such as Cd²⁺An important pathway into cells.

Geranium (A) and (B)pelargonium sp.fresham) Is a perennial herb flower, has strong adaptability, easy growth, bright color and higher ornamental value; geranium is strongly resistant to cadmium, but is not a hyper-enriched plant.

The existing technology for treating heavy metal sewage mainly comprises a physical method, a chemical method, a biological method and the like. (1) The physical method, the adsorption method, is the most common physical method, and some porous substances are used as adsorbents to be put into heavy metal sewage to adsorb heavy metal anions and cations in the sewage. The activated carbon is the first and most widely used adsorbing substance at present, has large surface area and strong adsorption capacity, but is expensive in use cost and difficult to desorb, and the development of the activated carbon in heavy metal sewage treatment is limited due to the cost. (2) The biological method, the microbial flocculation method, is the most common method for biologically purifying sewage, and the microbial flocculation method utilizes microorganisms or bacterial organisms to carry out metabolism to generate certain floccules, and the floccules are subjected to flocculation precipitation to remove heavy metals. The microorganism flocculation method has good treatment effect on heavy metal sewage, is safe, convenient and nontoxic to use, cannot cause secondary pollution to water quality, but has general effect. (3) The chemical method, chemical precipitation, is based on the fact that heavy metals can react with certain compounds to produce heavy metal compounds that are insoluble in water. These water-insoluble compounds will precipitate in the reaction zone. The chemical precipitation method is characterized in that the principle is determined according to the chemical properties of heavy metals, heavy metal ions in the heavy metal sewage are changed into metal compounds from the original free state to precipitate after the heavy metal sewage passes through a series of chemical reaction zones, and the metal compounds are separated through further chemical reaction; this method is costly.

Disclosure of Invention

It is an object of the present invention to provide MgCl₂And KNO₃The application of the cadmium-enriched powder is an application in efficiently and rapidly improving the cadmium enrichment capacity of plants.

In order to achieve the above object of the present invention, the technical solution of the present invention is as follows:

(1) KNO with the molar concentration of 1mol/L is prepared₃Mother liquor of solution, 1mol/L MgCl₂Solution mother liquor;

(2) subjecting the water cultured Pelargonium Hortori growing in the nutrient solution to KNO treatment in the step (1)₃And MgCl₂Pretreating the mixed solution, and adding KNO in the mixed solution₃The final concentration of (A) is 20-70mmol/L, MgCl₂The final concentration of (b) is 100-500 mu mol/L;

(3) the pretreated plants were placed in CdCl containing 100. mu. mol/L₂In the solution, the absorption rate of plants to cadmium ions is observed and the plasma membrane H is determined⁺-activity of ATPase.

KNO provided by the invention₃And MgCl₂The treating agent is convenient to use and low in cost; the treatment agent remarkably increases the absorption of cadmium ions by plants; the invention opens up a new way for improving the cadmium enrichment capacity of plants and is beneficial to scientific and technological workers to KNO₃And MgCl₂The research on the molecular mechanism of stimulating the plant growth has wide prospect in agricultural production.

The invention has the beneficial effects that:

the KNO for improving the enrichment capacity of the cadmium in plants₃And MgCl₂The promoter has low investment, simple operation and high effectHigh yield, KNO at room temperature₃And MgCl₂The invention is an ideal treating agent for improving the cadmium enrichment capacity of plants, and can obviously improve H on plant cell membranes⁺The activity of ATPase, and the purpose of obviously improving the cadmium enrichment capacity of plants is achieved.

Drawings

FIG. 1 shows the growth of pelargonium in each experimental group after pretreatment with additives and cadmium treatment;

FIG. 2 is a differential pulse anodic stripping voltammetry curve of cadmium on a Nafion-carbon nanotube screen-printed electrode, wherein a-g are cadmium concentrations: 50. 100, 150, 200, 300, 400, 500 μmol/L;

FIG. 3 is a standard curve of the dissolution peak current of cadmium versus the corresponding cadmium concentration;

FIG. 4 is a differential pulse anodic stripping voltammogram of Pelargonium root enriched cadmium;

FIG. 5 is a differential pulse anodic stripping voltammogram for determining the concentration of cadmium ions remaining in solution after one week of cadmium ion treatment for each experimental group;

FIG. 6 shows the content of cadmium ions in the rhizome leaves of Pelargonium graveolens after one week of cadmium ion treatment;

FIG. 7 is a pelargonium root tissue plasma membrane H after pretreatment with additives⁺-ATPase activity scheme;

FIG. 8 is a photograph showing the plasma membrane H of geranium root tissue after pretreatment with an additive and treatment with cadmium⁺-ATPase activity scheme;

CK in the figure is a control group.

Detailed Description

The invention is explained in more detail below with reference to the figures and examples, without limiting the scope of the invention. In the examples, the procedures were carried out in accordance with the usual procedures unless otherwise specified.

Example 1: MgCl for improving cadmium enrichment capacity of geranium₂And KNO₃Application of treating agent

(1) By KNO₃KNO with the molar concentration of 50mmol/L is prepared₃Solution of KNO₃And MgCl₂KNO with the molar concentration of 50mmol/L is prepared₃And MgCl 200. mu. mol/L₂Mixing the solution;

(2) in the experiment, geranium is used as a test material; planting Pelargonium in water, culturing for 4 weeks, selecting small seedlings with uniform size, and adding MgCl₂Treatment of the solution and addition of MgCl₂With KNO₃Treating the mixed solution;

(3) after 72h of treatment with the two different solutions, the treated plants were transferred to a medium containing 100. mu. mol/L CdCl₂In a culture solution; simultaneously, a control group (CK) -experimental pelargonium was not KNO-treated₃Solution and MgCl₂Solution treatment; and measuring the relevant physiological indexes of the geranium on the absorption of cadmium ions in the culture solution.

Example 2: determination of cadmium content in geranium

(1) Collection of cadmium-treated Pelargonium samples

Taking out the pretreated geranium branches treated by cadmium, cleaning the tail ends of the stems for three times by double distilled water, and wiping the stems by filter paper; cutting stem of Pelargonium 0.2g away from stem end 0.5 cm; cutting yellow leaves and tender leaves of geranium 0.2g respectively, cutting root of geranium 0.2g, washing with double distilled water for three times, and wiping with filter paper; wrapping a sample with tinfoil paper, freezing the sample in liquid nitrogen, grinding the frozen sample into powder, putting the powder into a 1.5mL centrifuge tube, adding 500 mu mol/L BR buffer solution with pH4.5, centrifuging (5000 rpm, 2min, 20 ℃) and taking 200 mu L of supernatant for later use;

(2) preparation of screen printing electrode Nafion-carbon nano tube

Pretreating the disposable screen printing electrode: selecting a disposable silk-screen printing electrode, diluting 99% concentrated sulfuric acid into 10% dilute sulfuric acid, diluting 30% hydrogen peroxide into 3% dilute sulfuric acid with 10% dilute sulfuric acid, and shaking up to obtain a mixed solution; shaking and washing the screen-printed electrode for 20min, washing with 1mL of double-distilled water for three times each time, and drying at normal temperature;

preparing a screen printing electrode Nafion-carbon nanotube: adding 1mg of carboxylated multi-wall carbon nano-tube into 1mL of solution, and diluting the solution with absolute ethyl alcohol to obtain 0.05% Nafion-carbon nano-tube suspension; dripping 5 mu L of carbon nanotube suspension on the surface of a working electrode of the screen printing electrode, airing at normal temperature to form a Nafion-carbon nanotube film, and carrying out cation exchange with heavy metal and adsorbing on the surface of a modified electrode.

(3) Determination of cadmium Standard Curve

Preparing 50, 100, 150, 200, 300, 400 and 500 mu mol/L cadmium chloride solution by taking 0.1 mu mol/L of BR buffer solution with pH4.5 in a certain volume, and dropwise adding 100 mu L of the cadmium chloride solution on a disposable silk-screen printing electrode to ensure that the solution covers the whole electrode; setting the parameters of an electrochemical workstation thereof; the dissolution detection method comprises the steps of differential pulse anodic voltammetry, initial potential of-0.3V, termination potential of-1V, scanning speed of 50mV/s, deposition potential of-1V, and recording the electrochemical behavior after enrichment for 180s at-1V; carrying out parallel measurement for three times, taking the concentration of cadmium as an abscissa, and differentiating the ordinate of the pulse current value to make a standard curve;

(4) determination of cadmium content of geranium branches

100 μ L of the collected supernatant of the roots, stems and leaves of geranium were dropped onto a disposable screen-printed electrode, and the solution was allowed to cover the entire electrode. Setting the parameters of an electrochemical workstation thereof: the dissolution detection method is a differential pulse anodic voltammetry method, the initial potential is-0.3V, the termination potential is-1V, the scanning speed is 50mV/s, the deposition potential is-1V, the electrochemical behavior is recorded after enrichment for 180s at-1V, and the electrochemical behavior is measured in parallel for three times.

In FIG. 1, CK represents the growth of water-treated pelargonium; k + Mg + Cd is MgCl₂And KNO₃After the mixed solution is pretreated, the growth condition of the geranium is treated by Cd for one week; mg + Cd being MgCl₂After the mixed solution is pretreated, the growth condition of the geranium is treated by Cd for one week; cd is the growth of non-pretreated geranium after one week of Cd treatment, and it can be seen from the figure that geranium is subjected to MgCl₂And KNO₃The growth condition is good after the mixed solution is pretreated.

FIG. 2 is a differential pulse anodic stripping voltammetry curve of cadmium ion solutions with different concentrations on a Nafion-carbon nanotube modified disposable silk screen printing electrode, wherein the cadmium ion concentrations respectively corresponding to a, b, c, d, e, f and g are 50, 100, 150, 200, 300, 400 and 500 μ M/L; FIG. 3 is a graph showing the concentration of septa as the abscissa and the corresponding difference pulseA standard curve chart is drawn by the ordinate of the impulse current value; the linear regression equation is I_Cd=0.0222x-0.9863(μmol/L)，R²=0.9992，SD=4.935*10-7，P<0.0001, the linear range of detection is 0.002-50mmol/L, and the detection limit is 0.2 nmol/L.

The results in FIG. 4 show the passage through MgCl₂And KNO₃After treatment, the cadmium ion content in the pelargonium roots was significantly increased compared to the control group, and the results in fig. 5 show that MgCl is added₂And KNO₃After treatment, the amount of cadmium ions remained in the culture solution is minimum; the results in FIG. 6 show the passage through MgCl₂And KNO₃The treated geranium rhizome leaves have the highest content of cadmium ions.

Through MgCl₂And KNO₃The result of the treatment on the content of cadmium ions in each group of the geranium shows that MgCl is adopted₂And KNO₃After the treatment agent is used for treating the geranium, the content of cadmium ions in the geranium seedling roots is obviously higher than that of a control group CK, the content of the residual cadmium ions in the solution is obviously lower than that of the control group CK, and the enrichment capacity of the cadmium ions is obviously enhanced.

Example 3: MgCl₂And KNO₃Pretreatment of geranium plasma membrane H with activator⁺Effect of ATPase Activity

（1）MgCl₂Solution, KNO₃And MgCl₂After the mixed solution is pretreated for 72 hours, 0.5g of root tissues of the pelargonium under different treatments are taken, the root tissues are quickly frozen and stored by liquid nitrogen, and the root tissues are ground into powder by the liquid nitrogen and then added with a proper amount of uniform plasma membrane extracting solution;

（2）H⁺determination of ATPase Activity in a 0.5mL reaction containing 50mM BTP/MES, 5mM MgSO₄、50mM KCl、0.02% Brij (w/v)、50mM KNO₃、1mM（NH₄）₂MoO₄、1mM NaN₃、4mM ATP-Na₂Starting the reaction after adding 500. mu.g of plasma membrane protein; the reaction mixture was subjected to 30 ℃ water bath for 30min, and 1mL (containing 2% H) of a reaction stop solution was added₂SO₄ (v/v), 5% SDS (w/v) and 0.7% (NH)₄)₂MoO₄ (w/v)) immediately thereafter, 50. mu.l Vc were added and left at room temperature for about 20min, the measurement wavelength being 700nmThe absorbance value of (c).

(3) Calculating the release content of inorganic phosphorus according to the standard curve of inorganic phosphorus

1 Unit plasma Membrane H⁺-ATPase activity is defined as: mM inorganic phosphate released by catalytic ATP decomposition per mg of protein in 1 minute under reaction conditions of 30 ℃.

FIG. 7 is the plasma membrane H of geranium root tissue in each experimental group after pretreatment⁺ATPase Activity Via KNO₃Treatment of pelargonium membranes H⁺The results of the effect of-ATPase Activity show that MgCl is used according to the invention₂And KNO₃After the treatment agent treats the pelargonium, the pelargonium seedling root mesoplasm membrane H⁺The ATPase activity is obviously higher than that of CK in a control group.

Example 4: MgCl₂And KNO₃Activator treatment of root membrane H under geranium cadmium stress⁺Effect of ATPase Activity

(1) At KNO₃Solution, KNO₃And MgCl₂After the mixed solution was pretreated for 72 hours, the seedlings of geranium were transferred to a medium containing 100. mu. mol/L CdCl₂Performing intermediate treatment for 7 days, taking 0.5g of root tissues of the pelargonium graveolens under different pretreatment, quickly freezing and storing by using liquid nitrogen, grinding by using the liquid nitrogen into powder, and adding a proper amount of uniform plasma membrane extracting solution;

(2) determination of H + -ATPase Activity carried out in a 0.5mL reaction System

The reaction system contained 50mM BTP/MES, 5mM MgSO₄、50mM KCl、0.02% Brij (w/v)、50mM KNO₃、1mM （NH4）₂MoO₄ 、1mM NaN₃，4mM ATP-Na₂The reaction was initiated after the addition of 500. mu.g of plasma membrane protein. The reaction mixture was subjected to 30 ℃ water bath for 30min, and 1mL (containing 2% H) of a reaction stop solution was added₂SO₄(v/v), 5% SDS (w/v) and 0.7% (NH4)₂MoO₄(w/v)), immediately adding 50. mu.l Vc and standing at room temperature for about 20min, and measuring the absorbance at a wavelength of 700 nm;

(3) the release content of inorganic phosphorus, 1 unit of plasma membrane H, was calculated from the inorganic phosphorus standard curve⁺-ATPase activity is defined as: under the reaction condition of 30 ℃, within 1 minute per minuteMg protein catalyzes the decomposition of ATP to release inorganic phosphate.

FIG. 8 shows the plasma membrane H of geranium root tissue in each experimental group after one week of cadmium ion treatment after pretreatment⁺ATPase Activity, results show that MgCl according to the invention is used₂And KNO₃After the treatment agent treats the pelargonium, the pelargonium seedling root mesoplasm membrane H⁺The activity of ATPase is obviously higher than that of CK in a control group, and the cadmium enrichment capacity is obviously improved.

Claims (1)

1. MgCl₂And KNO₃The application of the method in improving the cadmium enrichment capacity of the geranium;

the specific treatment method comprises the following steps:

(1) KNO with the molar concentration of 1mol/L is prepared₃Mother liquor of solution, 1mol/L MgCl₂Solution mother liquor;

(2) subjecting the water cultured Pelargonium Hortori growing in the nutrient solution to KNO treatment in the step (1)₃And MgCl₂Pretreating the mixed solution, and adding KNO in the mixed solution₃The final concentration of (A) is 20-70mmol/L, MgCl₂The final concentration of (b) is 100-500 mu mol/L;

(3) the pretreated plants were placed in CdCl containing 100. mu. mol/L₂In the solution, the absorption rate of plants to cadmium ions is observed and the plasma membrane H is determined⁺-activity of ATPase.

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