RU2746764C1 - Method for determining air toxicity by the reaction of germination of seeds of higher plants - Google Patents

Abstract

FIELD: toxicology.

SUBSTANCE: method relates to toxicology and can be used to determine the toxic effects of organic and inorganic chemical compounds that make up the atmospheric air. The method can also be used for environmental assessment of the state of populated areas. The method for determining the toxicity of air by the germination of seeds of higher plants is carried out in several stages: at the first stage, leaves are collected from the same type of woody plants in dry weather on the experimental territory under study and in a conditionally clean zone; at the second stage, one-type water suspensions are prepared from selected leaves of woody plants. After that 100 leaves from the experimental area and 100 leaves from a conditionally clean zone are placed in beakers with distilled water for solution preparation. The volume of each beaker is 1000 ml. The leaves are mixed for 2-3 minutes. Then the plant leaves are taken from beakers with tweezers. At the third stage, sterilized Petri dishes with a diameter of 10 cm are taken for research. After that filter paper is laid on their bottom. An aqueous pulverized suspension from the experimental zone with a volume of 5 ml is poured into one of the Petri dishes, and a 5 ml aqueous pulverized suspension from a conditionally clean zone is poured into the other Petri dish. 50 seeds of plant test biocultures are carefully and evenly placed on filter paper in each Petri dish with tweezers. The Petri dishes are covered with coverslips and placed in a thermostat for germinating seeds. At the fourth stage, the degree of seed germination of test crops is determined by the length of the roots of germinated seeds and then the level of phytoxicity is calculated on the substrate of aqueous pulverized suspensions from the experimental territory according to the formula: T_ph = (Lc_av.-Le_av. / Lc_av.) ×100%, where Lc_av. is the average value of the length of the roots of seeds (mm), which grew on the substrate of aqueous pulverized suspensions from a conditionally clean zone, and Le_av. is the average value of the length of the roots of seeds (mm), which grew on the substrate of aqueous pulverized suspensions from the experimental zone.

EFFECT: invention is aimed at increasing the efficiency of the method for determining the toxicity of air by the reaction of germination of seeds of higher plants in aqueous pulverized suspensions containing organic and inorganic substances.

1 cl, 4 dwg, 1 tbl

Description

The inventive method relates to toxicology and can be used to determine the toxic effects of organic and inorganic chemical compounds that make up the atmospheric air and environmental assessment of the state of settlements.

A known method for determining the toxicity of air by the reaction of a test bioculture. The method is carried out by capturing and dissolving organic and inorganic toxic substances from the air using a saline solution of Lozin-Lozinsky. Infusoria Paramecium caudatum is used as a biological object to determine air toxicity. The toxicity of the obtained samples is evaluated on the basis of the ability of the ciliates Paramecium caudatum to respond to the presence in this solution of substances that pose a danger to their vital activity, and to move directionally along the concentration gradient of these substances (chemotactic reaction), avoiding their harmful effects, and by this reaction the nfusoria Paramecium caudatum is judged about the degree of air toxicity [RF Patent No. 2335770, published 10.10.2008-prototype].

The disadvantage of this method is the complexity of its implementation using various technical means and chemical reagents, aspiration equipment, Rykhter absorbers with a saline solution of Lozin-Lozinsky, a photometer, a 5% solution of polyvinyl alcohol, etc., as well as additional measures in the form of preparing samples of test cultures for carrying out experiment, namely, washing test samples from metabolic products and feed, testing them for sensitivity with a CuSO ₄ solution with a concentration of 0.1 mg / dm ³ , etc., which also requires certain professional skills.

The technical result is an increase in the efficiency of the method for determining the toxicity of air by the reaction of germination of seeds of higher plants in aqueous dust-like suspensions containing organic and inorganic substances in them.

The objective of the claimed invention is to increase the efficiency of the method for determining the toxicity of air by the reaction of germination of seeds of higher plants in aqueous pulverized suspensions containing organic and inorganic substances in them by simplifying the method, excluding measures for sample preparation of test biocultures using reagents and various technical means, while reducing labor costs. and the economic costs of implementing the method.

Solution of a technical problem

A method for determining air toxicity by germination of seeds of higher plants, including capturing organic and inorganic toxic substances from the air, dissolving them to obtain the investigated aqueous solutions and determining the assessment of air pollution by the reaction of test biocultures in the test solutions with determining the toxicity index of the air, in which for capturing organic and inorganic toxic substances from the air, leaf blades of woody plants of the same species growing in the experimental zone and in a conditionally clean zone are used, and seeds of higher plants are used as test biocultures, while the method is carried out in several stages: at the first stage in dry weather, in the experimental area under study and in a conditionally clean area selected as a control area, leaves are collected from the same species of woody plants with dust-like particles supposed to contain organic and inorganic natural substances, 10 leaves are selected from one woody plant and from at least 10 woody plants, each sample includes 100 leaves, both from the experimental territory and from the conditionally clean zone, at the second stage, aquatic plants are prepared from the selected leaves of woody plants of the same species suspensions - washing off dust from plant leaves, for preparing a solution, collected leaves in an amount of 100 pcs. from the experimental area and 100 pcs. leaves from a conditionally clean zone are placed in beakers with distilled water, 1000 ml in volume, mixed for 2-3 minutes and two aqueous suspensions containing dust-like particles are obtained, then plant leaves are taken from beakers with tweezers, at the third stage for studies, Petri dishes with a diameter of 10 cm are taken, which are sterilized in an oven at 130 ° C for 1 hour or by boiling in water for 40 minutes, after which filter paper is placed on the bottom of the Petri dishes, an aqueous pulverized suspension is poured into one of the Petri dishes. the experimental zone with a volume of 5 ml, and in the other - 5 ml of an aqueous pulverized suspension from the conditionally clean zone, in each Petri dish, 50 pieces of seeds of plant test biocultures are carefully and evenly placed on filter paper with tweezers, and the liquid level in the Petri dishes should be below the surface of the seeds, the Petri dishes are covered with coverslips and placed in a thermostat at a temperature of 20 ° C for germination of seeds, the time of germination of seeds in a thermostat is determined by the choice of test plant cultures of higher plants, at the fourth stage of testing, the degree of germination of seeds of test crops is determined by measuring the length of the roots of germinated seeds of test plant biocultures both in the control and in the experimental groups and then the level of phytoxicity on the substrate is calculated water dusty suspensions from the experimental territory according to the formula: T _f = (Lc _avg. –Le _cf. / Lc _avg .) X 100%, where Lc _avg. - the average value of the length of the roots of seeds in mm, which grew on the substrate of aqueous dusty suspensions from a conditionally clean zone, and Le _cf. - the average value of the length of the roots of seeds mm, which grew on the substrate of aqueous dust-like suspensions from the experimental zone, according to the level of phytotoxicity of dust-like suspensions, the ecological state of the air environment of the territory is estimated, with phytotoxicity - T _f = 90 -100% (5th level of phytotoxicity), which corresponds to very strong air pollution, with toxicity T _f = 75-90%, (4th level of phytotoxicity), which corresponds to severe air pollution, with phytotoxicity T _f = 50-75%, (3rd level of phytotoxicity), corresponds to the average air pollution, with phytotoxicity Т _ф = 25 -50%, (2nd level of phytotoxicity), corresponds to low air pollution and air toxicity and phytotoxicity Т _ф <25% - 1st level of phytotoxicity, which corresponds to very low pollution atmosphere and phytotoxicity of the air or the absence of pollutants in the air.

Implementation of the method

The method for determining the toxicity of air and assessing the ecological state of the air environment of the territory consists in the fact that it is carried out in several stages.

At the first stage, in dry weather, in the experimental area under study and in a conditionally clean area selected as a control, leaves are collected from the same species of woody plants with suspected dust particles and containing organic and inorganic substances on the surface of the leaf blades. 10 leaves are selected from one woody plant and from at least 10 woody plants of the same species, each sample includes 100 leaves, both in the experimental territory and in the conditionally clean zone. At the second stage, dust-like aqueous suspensions are prepared from the selected leaves of woody plants of the same species (washing away dust from plant leaves). To prepare the solution, the collected leaves in the amount of 100 pcs. from the experimental area and 100 pcs. leaves from a conditionally clean zone are placed in beakers with distilled water, 1000 ml each, mixed for 2-3 minutes and two aqueous suspensions are obtained containing dust-like particles containing organic and inorganic substances. Then the leaves of the plants are carefully selected from the beakers with tweezers and aqueous pulverized suspensions are obtained, ready for research. At the third stage, Petri dishes with a diameter of 10 cm are taken for research. Petri dishes are sterilized in an oven at 130 ° C for 1 hour or by boiling in water for 40 minutes. Then, filter paper is placed on the bottom of the Petri dishes. In one of the Petri dishes, an aqueous pulverized suspension is poured (washing away dust from plant leaves from the experimental area) with a volume of 5 ml, and into the other - 5 ml of an aqueous pulverized suspension from a conditionally clean zone. In each Petri dish, 50 pieces of test plant seeds are carefully and evenly placed on filter paper with tweezers. The liquid level in the Petri dishes should be below the surface of the seeds. Petri dishes are covered with coverslips and placed in a thermostat at 20 ° C for germination. The germination time of seeds in a thermostat is determined by the choice of test plant cultures of higher plants (Appendix No. 1 GOST 120 3 8 -8 4 C. 10). At the fourth stage of testing, the degree of germination of seeds of test crops is determined by measuring the length of the roots of germinated seeds of test crops both in the control and in the experimental groups and then the level of phytotoxicity is calculated (the ability of chemicals dissolved in water to inhibit (inhibit) the germination of seeds of higher plants) aqueous dust suspensions according to the formula: Тф = (Lк - Le / Lк) x 100%.

The level of phytotoxicity of dusty suspensions is used to assess the ecological state of the territory's atmosphere.

With phytotoxicity - Т _ф = 90 -100% or more (5th level of phytotoxicity), which corresponds to very strong air pollution, with toxicity Т _ф = 75 -90%, (4th level of phytotoxicity), which corresponds to severe air pollution air, with phytotoxicity T _f = 50-75%, (3rd level of phytotoxicity), corresponds to the average air pollution, with phytotoxicity T _f = 25 -50%, (2nd level of phytotoxicity), corresponds to low air pollution and air toxicity and if _{phytotoxicity T f} <25%, this is the 1st level of phytotoxicity, which corresponds to very low air pollution and air toxicity or the absence of pollutants and air toxicity.

An example of a specific execution

In accordance with the stated essence, an example of implementation of the method is given.

Phytotoxicity is the ability of chemicals, including surfactants (surfactants), dissolved in water, to inhibit (inhibit) the germination of seeds of higher plants.

Leaves of white willow (Salixalba L.), which have the greatest ability to adsorb organic and inorganic chemical compounds from the air, were used as samples for preparing aqueous suspensions. Sampling of leaves of white willow (Salixalba L) was carried out in the village of Kilyakovka in the Sredneakhtubinsky district of the Volgograd region, located on the banks of the Akhtuba river, the territory of tectonic faults characterized by a natural source of pollutants (experimental territory) and in the cottage village "Pribrezhny" (conditionally clean zone ), located on the banks of the Volga River. The method for determining the toxicity of air and assessing the ecological state of the atmosphere of the territory was carried out in several stages.

Examples of specific implementation of the method are illustrated in the photographs:

- Fig. 1 shows photographs of samples of a group of Petri dishes with a substrate of aqueous dusty suspensions from a conditionally clean zone with seeds of watercress DUKAT;

- figure 2 shows a photograph of seedlings of seeds of watercress DUCAT with a substrate from a conditionally clean zone;

- figure 3 shows a photograph of seedlings of seeds of watercress DUCAT with a substrate from the experimental zone;

- Fig. 4 shows a comparison of two seedlings along the length of the roots from the seeds of watercress DUCAT, which were cultivated with a substrate of aqueous dust-like suspensions from a conditionally clean zone and with a substrate from the experimental zone.

At the first stage, in dry weather, in the study areas, leaves were collected from the white willow (Salixalba L.) with putative dust particles on the surface of the leaf blades. 10 leaves were taken from one woody plant - white willow (Salixalba L.), each sample included 100 leaves from 10 woody plants growing in the experimental territory and 100 leaves from 10 woody plants of the same species growing in a conditionally clean zone.

At the second stage, from the leaves of the white willow (Salixalba L.) from the experimental territory (E) and from the conditionally clean zone (C), we prepared dust-like aqueous suspensions (washing off the dust from the leaves). For this, white willow leaves (Salixalba L.), in the amount of 100 pcs., From the experimental territory and leaves in the amount of 100 pcs. From a conditionally clean zone, they were placed in beakers with distilled water with a volume of 1000 ml, they were stirred in distilled water for 2-3 minutes, then the leaves were carefully taken from beakers with tweezers and two aqueous suspensions containing dust-like particles were obtained.

At the third stage of the study, 8 Petri dishes with a diameter of 10 cm were taken and sterilized by boiling in water for 40 min. Filter paper was placed on the bottom of the Petri dishes. Four Petri dishes were poured into prepared aqueous dust-like suspensions from the experimental area with a volume of 5 ml, and into the other four Petri dishes, 5 ml of an aqueous dust-like suspension from a conditionally clean zone were poured. In each Petri dish, 50 pieces of plant test bioculture seeds were carefully and evenly placed on filter paper with tweezers, while observing the condition that the liquid level in the Petri dishes was below the surface of the seeds. The seeds of watercress DUKAT (manufacturer: LLC "CENTER-OGORODNIK") were used as a test bioculture. In total, eight Petri dishes were used: 4 Petri dishes with a substrate of aqueous dust-like suspensions from a conditionally clean zone and 4 Petri dishes with a substrate from the experimental territory. Petri dishes were covered with coverslips and placed in a thermostat at a temperature of 20 ° C for 72 hours (GOST 120 38 - 84 C. 10 (Appendix No. 1).

At the fourth stage - after 72 hours, the length of the roots in mm of seedlings was measured both in the control and in the experimental groups, in which the seeds of watercress Dukat gave different lengths of seedling roots, and then the phytotoxicity level of aqueous dust-like suspensions was calculated using the formula: Тф = (Lк - Le / Lk) x 100%, where

Lk _Wed - the average value of the length of the roots of seeds in mm, which grew on the substrate of aqueous dust-like suspensions from the conditionally clean zone, Le _cf. - the average value of the length of the roots of seeds in mm, which grew on the substrate of aqueous dust-like suspensions from the experimental zone.

The results of measurements after 72 hours of the length of the roots of germinated seeds of watercress DUKAT in aqueous dust-like suspensions in Petri dishes (No. 1, No. 2, No. 3, No. 4) from the conditionally clean zone (K) and in Petri dishes (No. 5, No. 6, No. 7, No. 8) from the experimental territory (E) are presented in Table 1. In seedlings of seeds grown in a substrate from a conditionally clean zone, the length of the roots is much longer than in seedlings of seeds grown in a substrate from an experimental territory.

Table 1

Phytotoxicity index Average values of the phytotoxicity index of aqueous dusty suspensions from a conditionally clean zone
(L _to ) Average value of the indicator
from 4 samples
(Lk _Wed ) Average values of the phytotoxicity indicator of water dusty suspensions from the experimental territory
(L _uh ) Average value of the indicator in samples
(Le _Wed ) Phyto level
toxicity
(T _f ) Assessment of the ecological state of the atmosphere of the territory # 1 # 2 Number 3 # 4 No. 5 # 6 # 7 No. 8 Spine length (L _x ), mm 35.36 + 2.11 40.58 + 1.76 39.18 + 1.83 38.32 + 2.14 38.36 + 0.99 11.98 + 0.99 10.44 + 0.69 10.30 + 0.70 11.2 + 0.38 11.19 + 0.39 3 "Average pollution"

T _{f - the} level of phytotoxicity (%) of aqueous dusty suspensions was determined by the formula:

T _f = (Lc _avg. -Le _avg. / Lc _avg .) X 100%, where

Lk _Wed - the average value of the length of the roots of seeds, mm, which grew on the substrate of aqueous dust-like suspensions from a conditionally clean zone

Le _Wed - the average value of the length of the roots of seeds, mm, which grew on the substrate of aqueous dust-like suspensions from the experimental territory.

Using the above formula and the results of experimental data, the phytotoxicity in the experimental zone (E) in the village of Kilyakovka was determined, which was T _f = (38.36–11.19 / 38.36) x100% = 70.83%. The obtained value is in the range T _f = 50-75%, which corresponds to the 3rd level of phytotoxicity or average pollution and toxicity of the air of the territory, that is, dust particles on the leaves of white willow (Salixalba L.) contain substances - toxicants that can represent danger to the environment.

Claims (1)

A method for determining air toxicity by germination of seeds of higher plants, including capturing organic and inorganic toxic substances from the air, dissolving them to obtain test solutions and determining the assessment of air pollution by the reaction of test biocultures in test solutions with determining the toxicity index, characterized in that for For capturing organic and inorganic toxic substances from the air, leaf blades of woody plants of the same species growing in the experimental zone and in a conditionally clean zone are used, and seeds of higher plants are used as test biocultures, while the method is carried out in several stages: at the first stage in dry weather in the experimental area under study and in a conditionally clean area selected as a control area, leaves are collected from the same species of woody plants with supposed dust particles containing organic and inorganic substances TV, 10 leaves are selected from one woody plant and from at least 10 woody plants, each sample includes 100 leaves both in the experimental territory and in a conditionally clean zone, at the second stage, aqueous suspensions are prepared, for the preparation of solutions, collected leaves in the amount of 100 PC. from the experimental area and 100 pcs. leaves from a conditionally clean zone are placed in beakers with distilled water, 1000 ml each, mixed for 2-3 minutes, then plant leaves are carefully taken from beakers with tweezers and two aqueous suspensions containing dust particles are obtained, at the third stage for research, take Petri dishes with a diameter of 10 cm, which are sterilized in an oven at a temperature of +130 ° C for 1 hour or by boiling in water for 40 minutes, after which filter paper is placed on the bottom of the Petri dishes, water powder is poured into one of the Petri dishes. suspension - washing off dust from plant leaves from the experimental zone with a volume of 5 ml, and in the other - 5 ml of an aqueous pulverized suspension from a conditionally clean zone, in each Petri dish, 50 pieces are carefully and evenly placed on filter paper with tweezers. seeds of plant test biocultures, and the liquid level in the Petri dishes should be below the surface of the seeds, the Petri dishes are covered with coverslips and placed in a thermostat at a temperature of 20 ° C for germination of seeds, the time of germination of seeds in a thermostat is determined by the choice of test plant cultures of higher plants, on the fourth The test stages determine the germination of seeds of test biocultures by measuring the length of the roots of germinated seeds of test plant biocultures both in the control and in the experimental groups, and then calculating the level of phytoxicity on the substrate of aqueous dusty suspensions from the experimental territory according to the formula: T _f = (Lc _cf. - Le _av. / Lc _avg. ) × 100%, where Lc _avg. - the average value of the length of the roots of seeds, mm, which grew on the substrate of aqueous dust-like suspensions from a conditionally clean zone, Le _cf. - the average value of the length of the roots of seeds, mm, which grew on the substrate of aqueous dust-like suspensions from the experimental territory, according to the level of phytotoxicity of dust-like suspensions, the ecological state of the air environment of the territory is estimated, with phytotoxicity T _f = 90-100% - the 5th level of phytotoxicity, which corresponds to very strong air pollution, with toxicity T _f = 75-90% - the 4th level of phytotoxicity, which corresponds to strong air pollution, with phytotoxicity T _f = 50-75% - the 3rd level of phytotoxicity, corresponds to the average air pollution , with phytotoxicity T _f = 25-50% - the 2nd level of phytotoxicity, corresponds to low air pollution and air toxicity, and with phytotoxicity T _ph <25% - the 1st level of phytotoxicity, which corresponds to very low pollution of the atmosphere and phytotoxicity of air or the absence pollutants in the air.

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