Enzymatic activity of soils. The concept of the enzymatic activity of soils

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Introduction

1. Literature review

1.1 General understanding of soil enzymes

1.2 Enzymatic activity of soils

1.3 Methodological approaches to determining the enzymatic activity of soils

1.3.1 Allocation of experimental area

1.3.2 Features of selection and preparation of soil samples for analysis

1.4 Influence of various factors (temperature, water regime, selection season) on the enzymatic activity of soils

1.5 Change of communities of microorganisms in soils

1.6 Methods for studying the activity of soil enzymes

Conclusion

List of sources used

Introduction

Under the conditions of the increased anthropogenic load on the biosphere of the planet, the soil, being an element of the natural system and being in dynamic equilibrium with all other components, undergoes degradation processes. Fluxes of substances entering the soil as a result of anthropogenic activity are included in natural cycles, disrupting the normal functioning of the soil biota, and, as a consequence, the entire soil system. Among the various biological criteria for assessing the anthropogenic impact on soils, the most operational and promising are biochemical indicators that provide information on the dynamics of the most important enzymatic processes in the soil: synthesis and decomposition of organic matter, nitrification, and other processes.

Available information on enzymatic activity different types soils are currently insufficient and require further study. This makes it very important in theoretical and practical terms to study the issues raised in this work.

Object of study: activity of soil enzymes.

Target term paper: Study of soil enzymes and soil enzymatic activity.

In accordance with the purpose of the study, the following were set. tasks:

1. To give a general idea of ​​soil enzymes and the enzymatic activity of soils.

2. Consider methodological approaches to determining the enzymatic activity of soils.

3. Determine the influence of various natural factors on the enzymatic activity of soils

4. To study the issue of the presence and change of communities of microorganisms in soils

5. List and describe the methods for studying the activity of soil enzymes.

1 ... Literature review

1.1 General understanding of soil enzymes

It is difficult to imagine that enzymes, highly organized protein molecules, could be formed in soil outside of a living organism. It is known that the soil has a known enzymatic activity.

Enzymes are catalysts for chemical reactions of a proteinaceous nature, characterized by the specificity of action in relation to the catalysis of certain chemical reactions.

Enzymes are products of biosynthesis of living soil organisms: woody and herbaceous plants, mosses, lichens, algae, fungi, microorganisms, protozoa, insects, invertebrates and vertebrates, which are represented in nature by certain aggregates - biocenoses.

The biosynthesis of enzymes in living organisms is carried out due to genetic factors responsible for the hereditary transmission of the type of metabolism and its adaptive variability. Enzymes are the working apparatus by which the action of genes is realized. They catalyze thousands of chemical reactions in organisms, of which, as a result, cellular metabolism is composed. Thanks to enzymes chemical reactions in the body are carried out at great speed.

To date, out of two thousand known enzymes, more than 150 have been obtained in crystalline form. Enzymes are classified into six classes:

1. Oxy reductases - catalyze redox reactions.

2. Transferases - catalyze the reactions of intermolecular transfer of various chemical groups and residues.

3. Hydrolases - catalyze the reactions of hydrolytic cleavage of intramolecular bonds.

4. Lyases - catalyzing reactions of addition of groups to double bonds and reverse reactions of the removal of such groups.

5. Isomerases - catalyze isomerization reactions.

6. Ligases - catalyze chemical reactions with the formation of bonds due to ATP (adenosintri phosphoric acid).

When living organisms die off and decay, some of their enzymes are destroyed, and some of them, getting into the soil, retain their activity and catalyze many soil chemical reactions, participating in the processes of soil formation and in the formation of a qualitative feature of soils - fertility.

V different types Soils under certain biocenoses have formed their own enzymatic complexes, which differ in the activity of biocatalytic reactions.

An important feature of the enzymatic complexes of soils is the orderliness of the action of the existing groups of enzymes. It manifests itself in the fact that the simultaneous action of a number of enzymes representing different groups is ensured. Enzymes eliminate the accumulation of excess of any compounds in the soil. Surplus accumulated mobile simple connections(for example, NH 3), in one way or another, they temporarily bind and direct into cycles, ending with the formation of more complex connections... Enzymatic complexes can be represented as some kind of self-regulating systems. In this, microorganisms and plants play the main role, constantly replenishing soil enzymes, since many of them are short-lived.

The number of enzymes is indirectly judged by their activity in time, which depends on the chemical nature of the reacting substances (substrate, enzyme) and on the conditions of interaction (concentration of components, pH, temperature, composition of the medium, the action of activators, inhibitors, etc.).

Enzymes belonging to the classes of hydrolases and oxidoreductases are involved in the main processes of soil humification; therefore, their activity is an essential indicator of soil fertility. Therefore, let us briefly dwell on the characteristics of enzymes belonging to these classes.

Hydrolases include invertase, urease, phosphatase, protease, etc.

Invertase - catalyzes the hydrolytic cleavage of sucrose into equimolar amounts of glucose and fructose, also affects other carbohydrates (galactose, glucose, rhamnose) with the formation of fructose molecules - an energy product for the life of microorganisms, catalyzes fructose transferase reactions. Studies by many authors have shown that invertase activity reflects the level of fertility and biological activity of soils better than other enzymes 3, p. 27.

Urease - catalyzes the hydrolytic cleavage of urea into ammonia and carbon dioxide. In connection with the use of urea in agronomic practice, it should be borne in mind that the activity of urease is higher in more fertile soils... It increases in all soils during the periods of their greatest biological activity - in July-August.

Phosphatase (alkaline and acidic) - catalyzes the hydrolysis of a number of organophosphorus compounds to form orthophosphate. The phosphatase activity is the higher, the less mobile forms of phosphorus are in the soil, so it can be used as an additional indicator when determining the need for phosphorus fertilization in soils. The highest phosphatase activity is in the rhizosphere of plants.

Proteases are a group of enzymes that break down proteins to polypeptides and amino acids, which are subsequently hydrolyzed to ammonia, carbon dioxide and water. In this regard, proteases have critical importance in the life of the soil, since they are associated with a change in the composition of organic components and the dynamics of forms of nitrogen, which are easily assimilated by plants.

The class of oxidoreductases includes catalase, peroxidase and polyphenol oxidase, etc.

Catalase - as a result of its action, hydrogen peroxide is decomposed, which is toxic to living organisms:

H2O2> H2O + O2

Vegetation has a great influence on the catalase activity of mineral soils. Soils under plants with a powerful deeply penetrating root system are characterized by high catalase activity. The peculiarity of catalase activity is that it changes little down the profile, has an inverse dependence on soil moisture and a direct dependence on temperature.

Polyphenol oxidase and peroxidase in soils play the main role in the processes of humus formation.

Polyphenol oxidase catalyzes the oxidation of polyphenols to quinones in the presence of free atmospheric oxygen. Peroxidase catalyzes the oxidation of polyphenols in the presence of hydrogen peroxide or organic peroxides. Moreover, its role is to activate peroxides, since they have a weak oxidizing effect on phenols. Further, the condensation of quinones with amino acids and peptides can occur with the formation of a primary molecule of humic acid, which can be further complicated due to repeated condensations.

The ratio of the activity of polyphenol oxidase (S) to the activity of peroxidase (D), expressed as a percentage, has a connection with the accumulation of humus in soils; therefore, this value is called the conditional coefficient of humus accumulation (K):

Consider the varieties of soil enzymes.

The class of oxidoreductases includes catalyzing redox reactions.

In the overwhelming majority of biological oxidations, it is not the addition of oxygen to the oxidizing molecule that occurs, but the elimination of hydrogen from the oxidized substrates. This process is called dehydrogenation and is catalyzed by enzymes, dehydrogenases.

Distinguish between aerobic dehydrogenases, or oxidases, and anaerobic dehydrogenases, or reductases. Oxidases transfer hydrogen atoms or electrons from the substance being oxidized to atmospheric oxygen. Anaerobic dehydrogenases transfer hydrogen atoms and electrons to other acceptors, enzymes, or hydrogen carriers, without transferring them to oxygen atoms. Numerous organic compounds that enter the soil with plants and animals undergo oxidation: proteins, fats, carbohydrates, fiber, organic acids, amino acids, purines, phenols, quinones, specific organic matter such as humic and fulvic acids, etc.

In the redox processes in the cells of animals, plants, microorganisms, as a rule, anaerobic dehydrogenases are involved, which transfer the hydrogen cleaved from the substrate to intermediate carriers. In the soil environment, mainly aerobic dehydrogenases are involved in redox processes, with the help of which substrate hydrogen is transferred directly to atmospheric oxygen, i.e. oxygen is the acceptor of hydrogen. The simplest redox system in soils consists of an oxidizable substrate, oxidases, and oxygen.

A feature of oxidoreductases is that, despite a limited set of active groups (coenzymes), they are able to accelerate big number a wide variety of redox reactions. This is achieved due to the ability of one coenzyme to combine with many apoenzymes and form each time an oxidoreductase specific to a particular substrate.

Another important feature of oxidoreductases is that they accelerate chemical reactions associated with the release of energy, which is necessary for synthetic processes. Redox processes in the soil are catalyzed by both aerobic and anaerobic dehydrogenases. By their chemical nature, these are two-component enzymes, consisting of a protein and an active group, or coenzyme.

An active group can be:

NAD + (nicotinamide adenine dinucleotide),

NADP + (nicotinamide adenine dinucleotide phosphate);

FMN (flavin mononucleotide);

FAD (flavin adenine dinucleotide), cytochromes.

Found about five hundred different oxidoreductases. However, the most common oxidoreductases are those that contain NAD + as an active group.

By combining with protein and forming a two-component enzyme (pyridine protein), NAD + enhances its ability to repair. As a result, pyridine proteins become capable of taking away from substrates, which can be carbohydrates, dicarboxylic and keto acids, amino acids, amines, alcohols, aldehydes, specific soil organic compounds (humic and fulvic acids), etc., hydrogen atoms in the form of protons (H +) ... As a result, the active group of the enzyme (NAD +) is reduced, and the substrate goes into an oxidized state.

The mechanism of binding of two hydrogen atoms, i.e. two protons and two electrons, is as follows. The active group of dehydrogenases that accepts protons and electrons is the pyridine ring. When NAD + is reduced, one proton and one electron are attached to one of the carbon atoms of the pyridine ring, i.e. one hydrogen atom. The second electron is attached to the positively charged nitrogen atom, and the remaining proton is transferred to the environment.

All pyridine proteins are anaerobic dehydrogenases. They do not transfer the hydrogen atoms removed from the substrate to oxygen, but send them to another enzyme.

In addition to NAD +, pyridine enzymes may contain nicotinamide adenine dinucleotide phosphate (NADP +) as a coenzyme. This coenzyme is a derivative of NAD +, in which the hydrogen OH - group of the second carbon atom of adenosine ribose is replaced by a phosphoric acid residue. The mechanism of substrate oxidation with the participation of NADP * as a coenzyme is similar to that of NAD +.

After the addition of hydrogen, NADH and NADPH have a significant reduction potential. They can transfer their hydrogen to other compounds and reduce them, while they themselves are converted into an oxidized form. However, hydrogen attached to anaerobic dehydrogenase cannot be transferred to oxygen in the air, but only to carriers of hydrogen. These intermediate carriers are flavin enzymes (flavoproteins). They are two-component enzymes, in which phosphorylated vitamin B2 (riboflavin) can be found as an active group. Each molecule of such an enzyme has a molecule of riboflavin phosphate (or flavin mononucleotide, FMN). Thus, FMN is a compound of the nitrogenous base of dimethylisoalloxazine with the residues of the five-carbon alcohol ribitol and phosphoric acid. FMN is capable of accepting and giving up two hydrogen atoms (H) at the nitrogen atoms (N) of the isoalloxazine ring.

Transferases are called transfer enzymes. They catalyze the transfer of individual radicals, parts of molecules and whole molecules from one compound to another. Transfer reactions usually take place in two phases. In the first phase, the enzyme cleaves the atomic group from the substance participating in the reaction and forms a complex compound with it. In the second phase, the enzyme catalyzes the addition of a group to another substance participating in the reaction, and itself is released in an unchanged state. The class of transferases contains about 500 individual enzymes. Depending on which groups or radicals carry transferases, a distinction is made between phosphotransferases, aminotransferases, glycosyltransferases, acyltransferases, methyltransferases, etc.

Phosphotransferases (kinases) are enzymes that catalyze the transfer of phosphoric acid residues (H2PO3). ATP is usually the donor of phosphate residues. The transfer of phosphate groups is carried out to alcohol, carboxyl, nitrogen-containing, phosphorus-containing and other groups of organic compounds. Phosphotransferases include the ubiquitous hexokinase, an enzyme that accelerates the transfer of the phosphoric acid residue from the ATP molecule to glucose. With this reaction, the conversion of glucose into other compounds begins.

Glycosyltransferases accelerate the reactions of transfer of glycosyl residues to molecules of monosaccharides, polysaccharides or other substances. These are enzymes that provide reactions for the synthesis of new carbohydrate molecules, the coenzymes of glycosyltransferases are nucleoside diphosphate sugar (NDP-sugar). From them, during the synthesis of oligosaccharides, the glycosyl residue is transferred to the monosaccharide. About fifty NDF-sugars are currently known. They are widespread in nature, synthesized from phosphoric esters of monosaccharides and the corresponding nucleoside triphosphates.

Acyltransferases transfer residues of acetic acid CH3CO - as well as residues of other fatty acids to amino acids, amines, alcohols and other compounds. These are two-component enzymes, which include coenzyme A. The source of acyl groups is acylcoenzyme A, which can be considered as an active group of acyltransferases. When the residues of acetic acid are transferred, acetyl coenzyme A participates in the reaction.

The class of hydrolases includes enzymes that catalyze hydrolysis, and sometimes the synthesis of complex organic compounds with the participation of water.

The subclass of esterases includes enzymes that accelerate the hydrolysis reactions of esters, alcohols with organic and inorganic acids.

The most important subclasses of esterases are hydrolases of carboxylic acid esters and phosphatases. The hydrolysis reactions of fats (triglycerides), as a result of which glycine and higher fatty acids are released, are accelerated by the hydrolase of glycerol esters by lipase. There are simple lipases, which catalyze the release of higher fatty acids from free triglycerides, and lipoprotein lipases, which hydrolyze protein-bound lipids. Lipases are one-component proteins with molecular weights from 48 thousand to 60 thousand. Yeast lipase is well studied. Its polypeptide chain consists of 430 amino acid residues and is folded into a globule, in the center of which is the active center of the enzyme. The leading role in the active center of lipase is played by the radicals of histidine, serine, dicarboxylic acids, and isoleucine.

The activity of lipases is regulated by their phosphorylation-dephosphorylation. Active lipases are phosphorylated, inactive lipases are dephosphorylated.

Phosphatases catalyze the hydrolysis of phosphate esters. Phosphatases acting on esters of phosphoric acid and carbohydrates are widespread. Such compounds include, for example, glucose-6-phosphate, glucose-1-phosphatase, fructose-1,6-diphosphate, etc. The corresponding enzymes are called glucose-6-phosphatase, glucose-1-phosphatase, etc. They catalyze the cleavage of the phosphoric acid residue from phosphoric esters:

Phosphodiester phosphatases - deoxyribonuclease and ribonuclease catalyze the cleavage of DNA and RNA to free nucleotides.

The subclass of hydrolases includes glycosidases that accelerate the hydrolysis of glycosides. In addition to glycosides containing monohydric alcohol residues as aglycones, oligo- and polysaccharides are substrates on which glycosidases act. Of the glycosidases acting on oligosaccharides, maltose and sucrose are the most important. They carry out the hydrolysis of maltose and sucrose.

Of the glycosidases acting on polysaccharides, amylases are of the greatest importance. Salient feature amylase - lack of absolute specificity of action. All amylases are metalloproteins, contain Zn 2+ and Ca 2+. The active centers of amylases are formed by the radicals of histidine, aspartic and glutamic acids, and tyrosine. The latter performs the function of binding the substrate, and the former is tricatalytic. Amylases accelerate the hydrolysis reactions of glycosyl bonds in the starch molecule with the formation of glucose, maltose or oligosaccharides.

Of no small importance is cellulase, which catalyzes the breakdown of cellulose, inulase, which breaks down the polysaccharide inulin, and aglucosidase, which converts the disaccharide maltose into two glucose molecules. Some glycosidases can catalyze the transfer reactions of glycosyl residues, in which case they are called transglycosidases.

Proteases (peptide hydrolases) catalyze the hydrolytic cleavage of peptide CO-NH bonds in proteins or peptides to form peptides of lower molecular weight or free amino acids. Among peptide hydrolases, there are endopeptidases (proteinases), which catalyze the hydrolysis of internal bonds in a protein molecule, and exopeptidases (peptidases), which provide cleavage of free amino acids from the peptide chain.

Proteinases are divided into four subclasses.

1. Serine proteinases, the active center of these enzymes includes a serine residue. The sequence of amino acid residues in the region of the polypeptide chain for serine proteinases is the same: aspartic acid-serine-glycine. The hydroxyl group of serine is characterized by high processes. The second active functional group is the imidazole of the histidine residue, which activates serine hydroxyl as a result of hydrogen bonding.

2. Thiol (cysteine) proteinases have a cysteine ​​residue in the active center, sulfhydryl groups and an ionized carboxyl group have enzymatic activity.

3. Acid (carboxyl) proteinases, pH optimum<5, содержат радикалы дикарбоновых кислот в активном центре.

4. Metal proteases, their catalytic action is due to the presence in the active center of Mg 2+, Mn 2+, Co 2+, Zn 2+, Fe 2+. The bond strength of the metal with the protein part of the enzyme can be different. The metal ions included in the active center take part in the formation of enzyme-substrate complexes and facilitate the activation of substrates.

An important feature of proteinases is the selective nature of their action on peptide bonds in a protein molecule. As a result, an individual protein under the influence of a specific proteinase is always cleaved into a strictly limited number of peptides.

5. Peptide hydrolases that cleave amino acids from a peptide, starting from an amino acid with a free NH2 group, are called aminopeptidases, which have a free COOH group - carboxypeptidases. The hydrolysis of the protein is completed by dipeptidases, splitting the dipeptides into amino acids.

6. Amidases catalyze the hydrolytic cleavage of the bond between carbon and nitrogen: deamination of amines. This group of enzymes includes urease, which carries out the hydrolytic cleavage of urea. oxidative enzyme

7. Urease is a one-component enzyme (M = 480 thousand). The molecule is a globule and consists of eight equal subunits. Possesses absolute substrate specificity, acts only on urea.

It should be noted that in order to detect free enzymes in the soil, it is necessary first of all to free it from living organisms, that is, to carry out complete or partial sterilization. The ideal factor that sterilizes the soil for the needs of enzymology should kill living cells without disturbing their cellular structure, and at the same time, not affect the enzymes themselves. It is difficult to say whether all the currently used sterilization methods meet these requirements. Most often, the soil for the needs of enzymology is sterilized by adding toluene as an antiseptic, by treating the soil with ethylene oxide or, which is now increasingly practiced, by killing microorganisms of various kinds with ionizing radiation. The further technique for determining the catalytic properties of the soil does not differ from the methods for determining the activity of enzymes of plant or animal origin. A certain concentration of the substrate for the enzyme is added to the soil, and the reaction products are studied after incubation. Analyzes of many soils carried out by this method have shown that they contain free enzymes with catalytic activity.

1.2 Enzymatic activity of soils

Enzymatic activity of soils [from lat. Fermentum - ferment] -the ability of the soil to exert a catalytic effect on the processes of transformation of exogenous and its own organic and mineral compounds due to the enzymes present in it. When characterizing the enzymatic activity of soils, they mean the total indicator of activity. The enzymatic activity of different soils is not the same and is associated with their genetic characteristics and a set of interacting environmental factors. The level of enzymatic activity of soils is determined by the activity of various enzymes (invertase, protease, urease, dehydrogenase, catalase, phosphatase), expressed by the amount of decomposed substrate per unit time per 1 g of soil.

Biocatalytic activity of soils depends on the degree of their enrichment with microorganisms and on the type of soil. The activity of enzymes changes along the genetic horizons, which differ in humus content, types of reactions, redox potential and other parameters along the profile.

In virgin forest soils, the intensity of enzymatic reactions is mainly determined by the horizons of the forest litter, and in arable soils, by the arable layers. All biologically less active genetic horizons located under the A or Ap horizons have low enzyme activity. Their activity slightly increases with soil cultivation. After the development of forest soils for arable land, the enzymatic activity of the formed arable horizon decreases sharply in comparison with the forest litter, but as it is domesticated, it increases and in highly cultivated soils approaches or exceeds the values ​​of forest litter.

Enzymatic activity reflects the state of soil fertility and internal changes occurring during agricultural use and increasing the level of farming culture. These changes are found both when virgin and forest soils are involved in culture, and when they are used in various ways.

Throughout Belarus, up to 0.9 t / ha of humus is lost annually in arable soils. As a result of erosion, 0.57 t / ha of humus is irretrievably carried away annually from the fields. The reasons for dehumification of soils are an increase in the mineralization of soil organic matter, the lag of the processes of new formation of humus from mineralization due to insufficient supply of organic fertilizers to the soil and a decrease in the enzymatic activity of the soil.

Biochemical transformations of soil organic matter occur as a result of microbiological activity under the influence of enzymes. enzymatic activity soil microorganism

Enzymes play a special role in the life of animals, plants and microorganisms. Soil enzymes are involved in the decay of plant, animal and microbial residues, as well as in the synthesis of humus. As a result, nutrients from difficult-to-digest compounds are transferred into readily available forms for plants and microorganisms. Enzymes are distinguished by high activity, strict specificity of action and great dependence on various environmental conditions. Due to their catalytic function, they provide a fast course of a huge number of chemical reactions in the body or outside of it.

Together with other criteria, the enzymatic activity of soils can serve as a reliable diagnostic indicator for determining the degree of soil cultivation. As a result of research 4, p. 91, a relationship was established between the activity of microbiological and enzymatic processes and the implementation of measures that increase soil fertility. Soil cultivation, fertilization significantly change the ecological situation for the development of microorganisms.

Currently, several thousand individual enzymes have been found in biological objects, and several hundred of them have been isolated and studied. It is known that a living cell can contain up to 1000 different enzymes, each of which accelerates a particular chemical reaction.

Interest in the use of enzymes is also due to the fact that the requirements for increasing the safety of technological processes are constantly increasing. Being present in all biological systems, being at the same time products and instruments of these systems, enzymes are synthesized and function under physiological conditions (pH, temperature, pressure, presence of inorganic ions), after which they are easily eliminated, undergoing destruction to amino acids. Both products and wastes of most of the processes involving enzymes are non-toxic and easily degradable. In addition, in many cases, the enzymes used in industry are obtained in an environmentally friendly way. Enzymes are distinguished from nonbiological catalysts not only by their safety and increased biodegradability, but also by the specificity of action, mild reaction conditions and high efficiency. The efficiency and specificity of enzyme action allows obtaining target products in high yield, which makes the use of enzymes in industry economically profitable. The use of enzymes helps to reduce the consumption of water and energy in technological processes, reduces CO2 emissions into the atmosphere, and reduces the risk of environmental pollution by by-products of technological cycles.

The use of advanced agricultural technology can change in a favorable direction the microbiological processes of not only arable, but also subsurface soil layers.

With the direct participation of extracellular enzymes, decomposition of organic soil compounds occurs. Thus, proteolytic enzymes break down protein substances into amino acids.

Urease decomposes urea to CO2 and NH3. The resulting ammonia and ammonium salts serve as a source of nitrogen nutrition for plants and microorganisms.

Invertase and amylase are involved in the breakdown of carbohydrates. Enzymes of the phosphate group decompose organophosphorus compounds of the soil and play an important role in the phosphate regime of the latter.

To characterize the general enzymatic activity of the soil, the most common enzymes characteristic of the overwhelming majority of soil microflora are usually used - invertase, catalase, protease, and others.

In the conditions of our republic, a lot of research has been carried out 16, p. 115 on the study of changes in the level of fertility and enzymatic activity of soils under anthropogenic impact, however, the data obtained do not provide an exhaustive answer to the nature of the changes due to the difficulty of comparing the results in view of the difference in experimental conditions and research methods.

In this regard, the search for an optimal solution to the problem of improving the humus state of the soil and its enzymatic activity in specific soil and climatic conditions based on the development of resource-saving methods of basic soil cultivation and the use of soil-protective crop rotations that contribute to the preservation of the structure, prevention of soil overconsolidation and improvement of their qualitative state and restoration of soil fertility at minimal cost, very relevant.

1.3 Methodological approaches to the determination of enzymaticsoil activity

1.3.1 Isolation experimentallythplotand mapping

A test site is a part of the study area characterized by similar conditions (relief, uniformity of the soil structure and vegetation cover, the nature of economic use).

The test site should be located in a typical location for the study area. On an area of ​​100 sq. m, one test site with a size of 25 m is laid. In case of heterogeneity of the relief, the sites are selected according to the relief elements.

A preliminary plan for laying the main sections and half-sections is outlined so that they characterize the soils of all found landforms and differences in soil cover.

The loop method is used in areas with difficult terrain and a dense geographic network. With this method, the investigated area is divided into separate elementary sectors, taking into account the peculiarities of changes in the relief or hydrographic network. The sector is surveyed from one center by making loop-like routes in the radial direction.

Taking into account the peculiarities of the relief and the hydrographic network in one specific area, the survey routes can be planned in a combined way, i.e. part of the site is examined by the method of parallel intersections of the territory, and part by the method of loops.

Along the routes, the points of the sections are marked so that all the main differences in the relief and vegetation are covered, i.e. the distances between the sections are not limited, therefore, in some places, as a rule, difficult in relief, the sections may be thickened, while in other, relatively homogeneous areas, the location of the sections may be rare.

Further, work related to soil mapping and detailed study of soils begins with a reconnaissance survey of the site (quarter). During a reconnaissance survey, they familiarize themselves with the boundaries of the site and, in general, with the object of research, which is bypassed along clearings, sighting lines, and roads. In the most characteristic places, cuts are laid, the location of which is applied to the plan. According to the results of reconnaissance surveys, the routes and places of laying the soil sections are finally adjusted.

After the reconnaissance survey, they begin the actual survey, during which it is necessary to have a plan for the establishment of soil sections and a clean copy of the outline of the taxation description. A general idea of ​​the soil differences and the initial markings of the boundaries of the soil contours are obtained from the study of the main and control sections. Clarification of the boundaries of the distribution of the soil contour is carried out using pits. In this case, in the field diary for each section, a form for describing the soil section is filled out. The field study of the distribution of soils is carried out after the laying and binding of the sections to establish the classification of this soil. Based on the results of the field assessment of the soil cover and all other elements of the landscape, a separate, relatively homogeneous or uniformly variegated area is distinguished as a soil contour.

The basis for identifying the boundaries between the contours of different soils is to identify patterns between soils, relief and vegetation. Changes in soil formation factors lead to changes in soil cover. With a clear change in the relief, plant formations and parent rocks, the boundaries of the soil differences coincide with the boundaries on the ground. In turn, the ease of fixing the boundaries on the map and the accuracy of the selection of soil contours depend on the accuracy of the topographic base. However, in nature, most often one has to deal with unclear boundaries, a gradual transition. In this case, the establishment of the boundaries of soil contours requires the establishment of a large number of holes, as well as a wealth of practical experience and good observation. When performing the actual survey in the field, on the basis of the plan copied from the taxation plate, they make up the outline of the soils of the study area.

It should be remembered that there are no strict boundaries between soil differences in nature, since the replacement of one soil difference by another occurs gradually through the accumulation of some characteristics and the loss of others. Therefore, soil surveying allows only to a greater or lesser extent to convey the schematic outlines of the distribution of soil contours, and the accuracy of identifying their boundaries depends on the scale of the survey, the type of soil and other conditions. The minimum dimensions of soil contours that must be marked on a soil map are determined by technical standards.

1.3.2 Features of selection and preparation of soil samples for analysis

To correctly determine the content of a particular substance in the soil, all agrochemical analyzes must be performed with impeccable precision and accuracy. However, even a very thorough analysis will give unreliable results if soil sampling is incorrect.

Since a very small sample for analysis is taken, and the results of the determination should give an objective characterization of large quantities of material, attention is paid to the elimination of heterogeneity in the selection of soil samples. Averaging a soil sample is achieved by step-by-step selection of the initial, laboratory and analytical samples.

A mixed initial sample should be made up of separate samples (initial samples) taken within the same soil difference. If the site has a complex soil cover, then a single average sample cannot be taken. There should be as many as there are soil differences.

Depending on the configuration of the site, the location of the points for taking initial samples on it varies. On a narrow, elongated section, they can be placed along (in the middle) of it. On a wide, close to square, site, a staggered arrangement of sampling sites is better. On large areas, soil sampling is used along the length of the site in its middle, in an amount of up to 20 pieces.

Thoroughly mix the initial soil sample taken on a piece of tarpaulin, sequentially average and reduce to the required volume, then pour it into a clean bag or box. This is a laboratory sample, its mass is about 400 g.

Put a plywood or cardboard label written in pencil on top of the box with the laboratory sample, indicating:

1. Names of the object.

2. Site names.

3. Plot numbers.

4. Depths of sampling.

5. Sample numbers.

6. The names of the person who supervised the work or who took the sample.

7. Dates of the work.

The same entry is made simultaneously in the journal.

A soil sample delivered from the site is poured into the laboratory on thick paper or a sheet of clean plywood and all caked lumps are kneaded with your hands. Then they select foreign inclusions with tweezers, mix the soil well, grind it slightly. After such preparation of the laboratory sample, it is again scattered to bring it to an air-dry state, then crushed and passed through a sieve with 2 mm holes.

The soil drying room must be dry and protected from the access of ammonia, acid vapors and other gases.

To determine the enzymatic activity, usually take soil, dried in the open air; wet samples should be dried in the laboratory at room temperature. Care should be taken to ensure that the sample does not contain undecomposed plant debris. Lumps of soil are crushed and sieved through a 1 mm sieve. When studying the enzymatic activity of a fresh (wet) sample, even more attention should be paid to the complete removal of plant residues. Simultaneously with the study of activity, soil moisture is also determined, the result obtained is recalculated per 1 g of absolutely dry soil.

1.4 Influence of various factorson the enzymatic activity of soils

An important factor on which the rate of the enzymatic reaction depends (equal to the catalytic activity of the enzyme) is the temperature, the effect of which is shown in Figure 1. It can be seen from the figure that as the temperature rises to a certain value, the reaction rate increases. This can be explained by the fact that with an increase in temperature, the movement of molecules is accelerated and the molecules of the reacting substances have a greater opportunity to collide with each other. This increases the likelihood that a reaction between them will occur. The temperature that provides the fastest reaction is called the optimum temperature.

Each enzyme has its own optimum temperature. In general, for enzymes of animal origin, it lies between 37 and 40C, and for plant ones - between 40 and 50C. However, there are exceptions: b-amylase from sprouted grain has an optimal temperature at 60C, and catalase - within 0-10C. When the temperature rises above the optimum, the rate of the enzymatic reaction decreases, although the frequency of collisions of molecules increases. This happens due to denaturation, i.e. loss of the native state by the enzyme. At temperatures above 80C, most enzymes completely lose their catalytic activity.

The decrease in the rate of the enzymatic reaction at temperatures exceeding the optimum depends on the denaturation of the enzyme. Therefore, an important indicator characterizing the ratio of an enzyme to temperature is its thermolability, i.e. the rate of inactivation of the enzyme itself with increasing temperature.

Figure 1 - Influence of temperature on the rate of starch hydrolysis by amylase

At low temperatures (0C and below), the catalytic activity of enzymes drops to almost zero, but denaturation does not occur. As the temperature rises, their catalytic activity is restored again.

Also, moisture, the content of microorganisms, and the ecological state of soils affect the enzymatic activity of soils.

1.5 Change of communities of microorganisms in soils

Soil microorganisms are very numerous and varied. Among them are bacteria, actinomycetes, microscopic fungi and algae, protozoa and living creatures close to these groups.

The biological cycle in the soil is carried out with the participation of different groups of microorganisms. Depending on the type of soil, the content of microorganisms varies. In garden, vegetable garden, arable soils, there are from one million to several billion microorganisms in 1 g of soil. The soil of each garden plot contains its own microorganisms. They participate with their biomass in the accumulation of soil organic matter. They play a huge role in the formation of accessible forms of plant mineral nutrition. The importance of microorganisms in the accumulation of biologically active substances in the soil, such as auxins, gibberellins, vitamins, amino acids that stimulate the growth and development of plants, is extremely great. Microorganisms, which form mucus of a polysaccharide nature, as well as a large number of fungi filaments, take an active part in the formation of the soil structure, gluing dusty soil particles into aggregates, thereby improving the water-air regime of the soil.

The biological activity of the soil, the number and activity of soil microorganisms are closely related to the content and composition of organic matter. At the same time, such important processes of soil fertility formation as the mineralization of plant residues, humification, dynamics of mineral nutrition elements, the reaction of soil solution, the transformation of various pollutants in the soil, the degree of accumulation of pesticides in plants, the accumulation of toxic substances in the soil, and the phenomenon of soil fatigue. The sanitary and hygienic role of microorganisms is also great in the transformation and neutralization of heavy metal compounds.

A promising direction for the restoration and maintenance of fertility and biological intensification of agriculture is the use of organic waste processing products with the participation of earthworm vermicompost, which are in symbiosis with microorganisms. In natural soils, litter decomposition is carried out by earthworms, coprophages and other organisms. But microorganisms are also involved in this process. In the intestines of worms, more favorable conditions are created for them to perform any functions than in the soil. Earthworms, in alliance with microorganisms, transform various organic waste into highly effective biological fertilizers with a good structure, enriched with macro- and microelements, enzymes, active microflora, providing a prolonged (long-term, gradual) effect on plants.

So, ensuring the development of microorganisms in the soil, increases the yield and improves its quality. After all, microorganisms develop, i.e. divide every 20-30 minutes and, if sufficient nutrition is available, form a large biomass. If a bull weighing 500 kg per day forms 0.5 kg - 1 kg, then 500 kg of microorganisms per day is biomass, and 500 kg of plants create 5 tons of biomass. Why is this not observed in the soil? And because for this microorganisms need nutrition, and on the other hand, various factors, in particular pesticides, limit. On an area of ​​1 hectare, as a result of the vital activity of soil microbes, 7500 m3 of carbon dioxide are released during the year. And carbon dioxide is necessary both as a source of carbon nutrition for plants and for dissolving hard-to-reach salts of phosphoric acid and converting phosphorus into a form available for plant nutrition. Those. where microorganisms work well, there is no need for phosphorus fertilization. But the microorganisms themselves need organic matter.

In the balance of soil organic matter, the role of cultivated plants is great. The accumulation of humus in soils is facilitated by perennial grasses, especially legumes. After they are harvested, phytomass remains in the soil, which is enriched with nitrogen due to its fixation by nodule bacteria from the air. Tilled and vegetable crops (potatoes, cabbage, etc.) reduce the humus content in the soil, because they leave a small amount of plant residues in the soil, and the applied system of deep tillage ensures an intensive supply of oxygen to the arable layer and, as a consequence, provides a strong mineralization of organic matter, i.e. its loss.

When analyzing soils, the number of individual physiological groups of microorganisms is often taken into account. This is done by the so-called titer method, in which liquid selective (elective) nutrient media for certain groups of microorganisms are contaminated with different dilutions of the soil suspension. Establishing, after holding in a thermostat, the degree of dilution, which showed the presence of the desired group of microorganisms, one can then, by simple recalculation, determine the number of its representatives in the soil. In this way, they find out how rich the soil is in nitrifiers, denitrifiers, cellulose-decomposing and other microorganisms.

To characterize the type of soil and its condition, not only the indices of the number of different groups of microorganisms are important, but also the analysis of the condition in the soil of their individual species. With rare exceptions, even the physiological groups of microorganisms are very wide. The external environment can dramatically change the species composition of soil microorganisms, but little or no effect on the number of their physiological groups. Therefore, when analyzing the soil, it is important to strive to establish the state of certain types of microorganisms.

Among soil microorganisms, there are representatives of different systematic units capable of assimilating not only easily digestible organic compounds, but also more complex substances of an aromatic nature, which include such compounds characteristic of soil as humus substances.

All soils on Earth were formed from the very diverse rocks that emerge on the surface of the earth, which are usually called parent rocks. The soil-forming rocks are mainly loose sedimentary rocks, since igneous and metamorphic rocks come to the surface relatively rarely.

The founder of scientific soil science V.V.Dokuchaev considered the soil as a special body of nature, as distinctive as a plant, animal or mineral. He pointed out that different soils are formed under different conditions, and that they change over time. According to VV Dokuchaev's definition, soil should be called "daytime", or surface horizons of rocks, naturally changed by the influence of a number of factors. The type of soil is composed depending on: a) parent rock, b) climate, c) vegetation, d) relief of the country and e) age of the soil-forming process.

Developing the scientific foundations of soil science, V.V.Dokuchaev noted the enormous role of living organisms, and, in particular, microorganisms, in the formation of soil.

The period of creativity of V.V.Dokuchaev coincided with the time of the great discoveries of L. Pasteur, which showed the enormous importance of microorganisms in the transformation of various substances and in the infectious process. At the end of the last century and at the beginning of the current century, a number of important discoveries were made in the field of microbiology, which were of fundamental importance for soil science and agriculture. It was found, in particular, that the soil contains a huge number of different microorganisms. This gave reason to think about the essential role of the microbiological factor in the formation and life of the soil.

Simultaneously with V.V.Dokuchaev, another outstanding soil scientist P.A.Kostychev 24, p. 72. In the monograph "Soils of the chernozem region of Russia, their origin, composition and properties" (1886), he wrote that geology is of secondary importance in the question of chernozem, because the accumulation of organic matter occurs in the upper layers of the earth, geologically diverse, and chernozem is the question of the geography of higher plants and the question of the physiology of lower plants decomposing organic matter. PA Kostychev conducted a series of experiments to clarify the role of individual groups of microorganisms in the creation of soil humus.

A great contribution to the understanding of the role of a biological factor in the transformation of the Earth and in the process of soil formation was made by the student of V.V.Dokuchaev, Academician V.I. Vernadsky. He believed that organisms are the main factor in the migration of chemical elements in the upper part of the earth's crust. Their activity affects not only organic, but also minerals of the soil and subsoil layers.

Already from the initial stages of the transformation of rocks into soil, the role of microorganisms in the processes of weathering of minerals is very clear. Outstanding scientists V.I. Vernadsky and B. B. Polynov considered the weathering of rocks as a result of the activity of plant, mainly lower organisms. To date, this point of view has been confirmed by a large amount of experimental material.

Usually, the first settlers of rocks are crusty lichens, which form sheet-like plates, under which a small amount of fine earth accumulates. Lichens, as a rule, are in symbiosis with non-spore-forming saprophytic bacteria.

With regard to a number of elements, lichens act as their accumulators. In the fine earth under the lithophilic vegetation, the amount of organic matter, phosphorus, iron oxide, calcium and magnesium increases sharply.

Among other plant organisms that settle on parent rocks, microscopic algae should be noted, in particular blue-green and diatoms. They accelerate the weathering of aluminosilicates and also usually live in association with non-spore-forming bacteria.

Algae, obviously, play an essential role as autotrophic accumulators of organic substances, without which the vigorous activity of saprophytic microorganisms cannot proceed. The latter produce various compounds that cause the weathering of minerals. Many blue-green algae are nitrogen fixers and enrich the destroyed rock with this element.

The main role in the weathering process is probably played by carbon dioxide, mineral and organic acids produced by various microorganisms. There are indications that some keto acids have a strong dissolving effect. The possibility of participation in the weathering of humus compounds is not excluded.

It should be noted that many bacteria form mucus, which makes it easier for the microorganisms to come into close contact with the rock. The destruction of the latter occurs both under the influence of the waste products of microorganisms, and as a result of the formation of complex compounds between the substance of mucus and chemical elements that make up the crystal lattices of minerals. Weathering of rocks in nature should be considered as a unity of two opposite processes - the decay of primary minerals and the emergence of secondary minerals. New minerals can arise when microbial metabolites interact with each other.

...

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According to the type of catalyzed reactions, all known enzymes are divided into six classes:

1. Oxidoreductases that catalyze redox reactions.

2. Hydrolases that catalyze the reactions of hydrolytic cleavage of intramolecular bonds in various compounds.

3. Transferases that catalyze the reactions of intermolecular or intramolecular transfer of a chemical group and residues with a simultaneous transfer of energy contained in chemical bonds.

4. Ligases (synthetases), catalyzing the reaction of combining two molecules, coupled with the cleavage of the phyrophosphate bonds of ATP or other analogous triphosphate.

5. Lyases that catalyze the reactions of non-hydrolytic cleavage or addition of various chemical groups of organic compounds to double bonds.

6, Isomerases that catalyze the conversion of organic compounds into their isomers.

In soil, oxidoreductases and hydrolases, which are very important in soil biodynamics, are widespread and have been studied in detail.

Catalase

(H 2 O 2: H 2 O 2 -oxidoreductase)

Catalase catalyzes the decomposition reaction of hydrogen peroxide with the formation of water and molecular oxygen:

H 2 O 2 + H 2 O 2 O 2 + H 2 O.

Hydrogen peroxide is formed during the respiration of living organisms and as a result of various biochemical reactions of the oxidation of organic substances. The toxicity of hydrogen peroxide is determined by its high reactivity, which is manifested by singlet oxygen, * O 2. Its high reactivity leads to uncontrolled oxidation reactions. The role of catalase is that it destroys hydrogen peroxide, which is toxic to organisms.

Catalase is widespread in the cells of living organisms, including microorganisms and plants. Soils also exhibit high catalase activity.

Methods for determining the catalase activity of soil are based on measuring the rate of decomposition of hydrogen peroxide when it interacts with the soil by the volume of oxygen released (gasometric methods) or by the amount of undecomposed peroxide, which is determined by permanganatometric titration or by the colorimetric method with the formation of colored complexes.

Research by E.V. Dadenko and K.Sh. Kazeva found that during storage of samples, the activity of catalase of all enzymes decreases to the greatest extent; therefore, its determination must be carried out in the first week after sampling.

A.Sh. Galstyan

Analysis progress. To determine the activity of catalase, a device is used consisting of two burettes connected by a rubber hose, which are filled with water and balanced its level. Maintaining a certain water level in the burettes indicates the achievement of temperature equilibrium in the device. A weighed portion (1 g) of soil is introduced into one of the compartments of a double flask. In another compartment of the flask, 5 ml of a 3% hydrogen peroxide solution are poured. The flask is tightly closed with a rubber stopper with a glass tube, which is connected to the measuring burette by means of a rubber hose.

The experiment is carried out at a temperature of 20 ° C, since at a different temperature the reaction rate will differ, which will distort the results. In principle, it is not the temperature of the air that is important, but of the peroxide, it should be 20 0 C. If the air temperature is significantly higher than 20 0 C (in summer), it is recommended to carry out the analysis in the basement or in another cool room. The use of a water bath with a temperature of 20 ° C recommended in such cases is hardly effective.

The beginning of the experiment is marked by a stopwatch or hourglass at the moment when the peroxide is mixed with the soil, and the contents of the vessel are shaken. Shaking the mixture is carried out throughout the experiment, trying not to touch the flask with your hands, holding it by the stopper. The evolved oxygen displaces water from the burette, the level of which is noted after 1 and 2 minutes. The recommendation to determine the amount of oxygen every minute for 3 minutes due to the straightforwardness of the reaction of decomposition of peroxide only increases the time spent on analysis.

This technique allows one researcher to analyze the catalase activity of more than 100 samples per day. It is convenient to carry out the analysis together using 5-6 vessels. In this case, one person is directly involved in the analysis and monitors the level of the burette, while the second monitors the time, records data and washes the vessels.

The soil is sterilized by dry heat (180 ° C) as a control. Some soils, compounds and minerals have a high activity of inorganic catalysis of peroxide decomposition even after sterilization - up to 30-50% of the total activity.

Catalase activity is expressed in milliliters of O 2 released in 1 min from 1 g of soil.

Reagents: 3% solution of H 2 O 2. Perhydrol concentration must be checked periodically, the working solution is prepared immediately before analysis. To establish the concentration of perhydrol on an analytical balance, 1 g of H 2 O 2 is weighed in a 100 ml volumetric flask, the volume is brought to the mark and shaken. Place 20 ml of the resulting solution in 250 ml conical flasks (3 replicates), add 50 ml of distilled water and 2 ml of 20% H 2 SO 4. Then titrated with 0.1 N. solution of KMnO 4. 1 ml of KMnO 4 solution corresponds to 0.0017008 g of H 2 O 2. After the concentration of perhydrol is established, a 3% solution is prepared by dilution with distilled water. The KMnO 4 titration solution is prepared from a fixed channel and incubated for several days to establish the titer.

Dehydrogenase

(substrate: NAD (P) -oxidoreductase).

Dehydrogenases catalyze redox reactions by dehydrogenating organic substances. They run as follows:

AH 2 + B A + BH 2

In soil, the substrate for dehydrogenation can be nonspecific organic compounds (carbohydrates, amino acids, alcohols, fats, phenols, etc.) and specific (humic substances). Dehydrogenases in redox reactions function as hydrogen carriers and are divided into two groups: 1) aerobic, transferring mobilized hydrogen to oxygen in the air; 2) anaerobic, which transfer hydrogen to other acceptors, enzymes.

The main method for detecting the action of dehydrogenases is the reduction of indicators with a low redox potential, such as methylene blue.

To determine the activity of soil dehydrogenases, colorless tetrazolium salts (2,3,5-triphenyltetrazolium chloride - TTX) are used as hydrogen, which are reduced to red formazan compounds (triphenylformazan - TPP).

Analysis progress. A weighed portion (1 g) of the prepared soil is carefully placed through a funnel on the bottom of a 12-20 ml test tube and mixed thoroughly. Add 1 ml of 0.1 M solution of dehydrogenation substrate (glucose) and 1 ml of freshly prepared 1% TTX solution. The tubes are placed in an anaerostat or vacuum desiccator. The determination is carried out under anaerobic conditions, for which the air is evacuated at a vacuum of 10-12 mm Hg. Art. within 2-3 minutes and placed in a thermostat for 24 hours at 30 ° C. When incubating soil with substrates, toluene is not added as an antiseptic, since; it strongly inhibits the action of dehydrogenases. The control is sterilized soil (at 180 ° C for 3 h) and substrates without soil. After incubation, add 10 ml of ethyl alcohol or acetone to the flasks, shake for 5 minutes. The resulting colored TPP solution is filtered and colorimetric. With a very intense color, the solution is diluted with alcohol (acetone) 2-3 times. Use 10 mm cuvettes and a light filter with a wavelength of 500-600 them. The amount of formazan in mg is calculated from the standard curve (0.1 mg in 1 ml). The activity of dehydrogenases is expressed in mg of TTF per 10 g of soil for 24 h. The determination error is up to 8%.

Reagents:

1) 1% solution of 2,3,5-triphenyltetrazolium chloride;

2) 0.1 M glucose solution (18 g of glucose is dissolved in 1000 ml of distilled water);

3) ethyl alcohol or acetone;

4) triphenylformazan for the standard scale. To draw up a calibration curve, prepare a series of solutions in ethyl alcohol, acetone or toluene with a formazan concentration (from 0.01 to 0.1 mg formazan in 1 ml) and photocolorimetrate as described above.

In the absence of formazan, it is obtained by reducing TTX with sodium hydrosulfite (ammonium sulfite, zinc powder in the presence of glucose). The initial concentration of the TTX solution is 1 mg / ml. Crystalline sodium hydrosulfite is added to 2 ml of the TTX stock solution at the tip of the lancet. The formed precipitate of formazan is taken up in 10 ml of toluene. This volume of toluene contains 2 mg of formazan (0.2 mg / ml). Working solutions for the scale are prepared by further dilution.

Invertase

(β-fructofuranosidase, sucrase)

Invertase is a carbohydrase; it acts on the β-fructofuranosidase bond in sucrose, raffinose, gentianose, etc. This enzyme hydrolyzes sucrose most actively to form reducing sugars - glucose and fructose:

invertase

С 12 Н 22 О 11 + Н 2 О С 6 Н 12 О 6 + С 6 Н 12 О 6

sucrose glucose fructose

Invertase is widespread in nature and occurs in almost all types of soil. A very high activity of invertase was found in mountain meadow soils. Invertase activity clearly correlates with humus content and soil fertility. Recommended when studying the effect of fertilizers to assess their effectiveness. Methods for determining the activity of soil invertase are based on the quantitative account of reducing sugars according to Bertrand and on the change in the optical properties of the sucrose solution before and after the action of the enzyme. The first method can be applied when studying an enzyme with a very wide amplitude of activity and substrate concentration. Polarimetric and photocolorimetric methods are more demanding on the concentration of sugars and are unacceptable for soils with a high organic matter content, where colored solutions are obtained; therefore, these methods are of limited use in soil research.

Invertase - catalyzes the hydrolytic cleavage of sucrose into equimolar amounts of glucose and fructose, also affects other carbohydrates with the formation of fructose molecules - an energy product for the life of microorganisms, catalyzes fructose transferase reactions. Studies by many authors have shown that invertase activity reflects the level of fertility and biological activity of soils better than other enzymes. [...]

Analyzes of invertase after 1 year indicate a further decrease in it in all samples by 2-3 times, depending on the type of soil, which, apparently, is explained by the depletion of the soil with carbon-containing compounds. [...]

From the class of hydrolases, the activity of invertase, which hydrolyzes sucrose into glucose and fructose, and urease, which catalyzes the hydrolysis of urea, have been studied. The activity of these enzymes in the soil is very low, but when peat is applied, it increases in proportion to its doses and depends little on the amount of mineral fertilizers. It should be noted that the application of the highest dose (NPKTS, as well as CaCOe has no advantages over lower doses of fertilizers in stimulating the activity of both hydrolases and oxidoreductases. [...]

For the route airport - pos. Kangalassy no inverse relationship between the activity of urease, invertase, and protease and lead content was found. This indicates the absence of the inhibitory effect of lead at a dose not exceeding the MPC. There is a parallel increase in the activity of all enzymes and lead with distance from the source of pollution, which in this case is explained by an increase in soil humus content. It is known that soils with a high humus content accumulate HMs to a greater extent and are characterized by increased FA. [...]

Compounds of this group delay the growth of new shoots, temporarily reduce the activity of invertase in sugar beets and inhibit chlorophyll biosynthesis. Yet their primary action is to inhibit the biosynthesis of aromatic amino acids. Compounds of the N-phosphonomethylglycine type inhibit this synthesis by acting on the sites of conversion of dehydroquinic and prefenic acids. [...]

Apparently, the formation of sucrose occurs in the parenchymal cells of the phloem, from where it enters the sieve tubes, which are devoid of enzymes that decompose sucrose (invertases), which determines the safety of this compound along the entire path of its transport. [...]

The work performed allows us to conclude that the accumulation of mobile forms of lead and nickel in doses exceeding the MPC leads to a decrease in the activity of enzymes in soils. A decrease in the activity of protease, urease and invertase in soils leads to a corresponding inhibition of the processes of hydrolysis of proteins, urea and oligosaccharides, which generally leads to a decrease in the biological activity of soils. Changing FA is a promising method for diagnosing the ecological state of soils. Of the enzymes considered by us, urease exhibits the highest diagnostic properties. [...]

The state of the soils was assessed by two bioindication methods: by the enzymatic activity of soils and the mutational effect of soils on the test object. In urban soils, the activity of three enzymes was determined - invertase, catalase, and urease (Khaziev, 1990), of which the most variable was the activity of urease. For this reason, the indicators of this particular enzyme were selected for the integral assessment, the activity of which largely depended on the concentration of a wide range of pollutants in the soil. [...]

Histochemical analyzes made it possible to establish the commonality of the oxidative regime of pollen and pollen tubes in various representatives of angiosperms. It was found that the most intensive biochemical processes occur in the tip of the pollen tube. [...]

Another group of evocation changes is associated with the activation of energy processes necessary for the implementation of the morphogenetic program of reproductive development. [...]

With the introduction of large norms of HCBD in both liquid and granular form, the inhibition of the development of individual groups of microorganisms does not go away even by one and a half years after fumigation. The activity of soil enzymes (catalase and invertase) by this time, according to these (experimental variants, 70-80% of the enzyme activity in the control variant. 5 months after the introduction of large norms of HCBD (liquid and granular), the content of nitrates in the soil decreases, inhibition of the nitrification process. [...]

Agrochemical properties of soils were determined by conventional methods, pH of water and salt extracts - potentiometric, carbon content - by Tyurin's method, mobile nitrogen - according to Bashkin and Kudeyarov, mobile phosphorus - according to Chirikov, enzymatic activity of soils (invertase, urease and catalase) - according to Khaziev. [ ...]

In many representatives of radiant fungi, the amylase enzyme has been identified, with the help of which organisms break down starch with different intensities, depending on the type of culture. Some cultures decompose starch to dextrins, others to sugars. In some actinomycetes, the enzyme invertase is found, which breaks down sucrose into easily digestible sugars - glucose and fructose. It was noted that proactinomycetes can assimilate sucrose without decomposition. [...]

Such levels of pollution were reflected in the content of mobile, plant-accessible forms of heavy metal compounds. Their number also increased by 1.5-2 and even 5 times. These changes were reflected in soil biota, general soil properties and soil fertility. In particular, the activity of soil enzymes sharply decreased: invertase, phosphatase, urease, catalase; the production of CO2 decreased by about 2 times. Enzymatic activity is a good integral indicator of the ecological situation in the "soil - plant" system. On contaminated soils, the yield of various crops has also sharply decreased. So, the yield of tomatoes (c / ha) on average decreased from 118.4 to 67.2; cucumbers - from 68.3 to 34.2; cabbage - from 445.7 to 209.0; potatoes - from 151.8 to 101.3; apples - from 72.4 to 32.6 and peaches - from 123.6 to 60.6. [...]

Among the tundra soils of the floodplain, the potential of biochemical activity increases from the soils of the near-river floodplain to the central and near-terrace ones. In turn, the enzymatic activity in organogenic floodplain soils is higher than in mineral soils. In the humus horizons (0-13 cm) of the studied soils, there is a rather high activity of urease, invertase, phosphatase and dehydrogenase - enzymes involved in the metabolic processes of nitrogen, carbohydrates, phosphorus and redox. [...]

The phosphatase activity is low, and in most cases there is no phosphatase activity, which is associated with a very low content of mobile phosphorus against the background of a relatively high content in humus-peaty horizons of its bulk forms. Unlike the enzymes involved in the metabolic processes of nitrogen and phosphorus, the enzymes of hydrocarbon metabolism (invertase) show their activity up to the suprapermafrost horizons, which is determined by the humus content of the profile. [...]

The change in the enzymatic activity of soils for four years of the experiment is shown in table. 6.8. As can be seen from the results obtained, the activity of urease and phosphatase decreased, but the main regularities - a higher activity in the variants without the use of PPS when applying peat and mineral fertilizers and the absence of enzymatic activity in the control variants - remain. At the same time, the activity of invertase, which plays an important role in the carbon cycle in the biogeocenosis, increases in the fourth year in almost all variants of the experiment, including the addition of PPS, which also confirms the intensity of the mineralization processes of peat and Universines. [... ]

A very promising method of water purification from all kinds of polluting substances, especially synthetic ones, is the use of immobilized (fixed, insoluble) enzymes - "enzymes of the second generation". The idea of ​​fixing enzymes on a water-insoluble carrier and the use of such powerful catalysts in technological processes and medicine arose long ago. Back in 1916, the adsorption of invertase on activated carbon in freshly isolated aluminum hydroxide was carried out. Since 1951, protein-cellulose conjugation has been used to fractionate antibodies and isolate antigens. Until recently, there was only one method for fixing enzymes - ordinary physical adsorption. However, the adsorption capacity of the known materials with respect to proteins is clearly insufficient, and the adhesion forces are small, and the breaking of the bond between the enzyme and the adsorbent surface can occur from the slightest changes in the process conditions. Therefore, this method of immobilization has not found wide application, but since it is simple and can, apparently, help to elucidate the mechanism of action of enzymes in living systems, silts and soil, and in some cases apply in practice, some researchers are studying the adsorption of enzymes, searching for new, effective media, etc. [...]

Taking into account the pronounced and long-term physiological changes in the growth and development processes caused by ethylene, it will not seem surprising that there are also changes in the synthesis of RNA and protein and in the activity of enzymes. The possibility of a direct effect of ethylene on the activity of various enzymes, for example, glucosidase, a-amylase, invertase and peroxidase, was repeatedly tested, but negative results were obtained. At the same time, the synthesis of a number of enzymes is clearly increasing. Peroxidase is one of the enzymes relatively rapidly synthesized after exposure to ethylene. In citrus fruits, the synthesis of phenyl-alanine-ammonia-lyase is enhanced, and CO2 and transcription inhibitors block this process. Ethylene causes the formation of cellulase in the release tissue. The link between this effect and the stimulation of the separation process is obvious. True, the accelerated separation occurs even before the rise in the synthesis of cellulase, but this is probably due to the fact that ethylene also causes the release of cellulase from the bound form and its secretion into the intercellular spaces. The release of amylase from barley aleurone cells is also accelerated by the action of ethylene. Rapid "effects of ethylene, for example, suppression of cellular elongation, which manifests itself already after 5 minutes, are associated more with the effect on membranes than with changes in protein synthesis. [...]

As you know, one of the reasons for the toxicity of soils is their salinity. Used drilling fluids and drill cuttings contain, in some cases, a significant amount of mineral salts hazardous to soils. Therefore, it is of interest to identify the influence of this factor on the biological productivity of soils. The research results indicate that mineral spines in the amount of bottee 0 8-4.0 kt / m2 of soil sharply reduce the activity of invertase, and in an amount of more than 1.5-1.6 kg / m2 soils begin to significantly affect the yield of cultivated crops. them crops. [...]

Honey is a high-calorie product. Natural honey is a sweet, viscous and aromatic substance produced by bees from plant nectar, as well as from honeydew or honeydew. Honey can appear as a crystallized mass. The value of honey lies in the fact that it has bactericidal properties. Therefore, honey is not only a valuable food product, but also a remedy. The main constituents of flower honey are fruit and grape sugar, of which it contains about 75%. The calorie content of honey is over 3 thousand calories. It contains enzymes: diastase (or amylase), invertase, catalase, lipase. [...]

The studies were carried out in the valley of the lower reaches of the Sysola River (Komi Republic, subzone of the middle taiga). The biochemical parameters of soils were characterized by the level of activity of oxidoreductases (catalase), hydrolases (invertase), and the release of CO2 from the soil surface. During all sampling periods, the maximum values ​​of catalytic activity were noted in forest litters of the Adl soil (4.2–8.6 ml O2 / g soil), which is the “driest” in the studied series of soils. However, the Al soil was in the lead in terms of the invertase level at all sampling periods (11.9–37.8 mg glucose / g soil in the AO horizon). In the same soil, a maximum in the release of CO2 (0.60 ± 0.19) kg / ha-hour was recorded in July. Using the integral BAP index, which takes into account all the parameters of biological activity, it was shown that the most active biological processes at all sampling periods occur in the Al soil, which occupies an intermediate position in the hydrothermal regime between the Adl and Alb soils. [...]

The destabilization of the nitrification process disrupts the flow of nitrates into the biological cycle, the amount of which predetermines the response to a change in the habitat of the denitrifier complex. Enzyme systems of denitrifiers reduce the rate of complete recovery, weaker involving nitrous oxide in the final stage, the implementation of which requires significant energy costs. As a result, the content of nitrous oxide in the above-soil atmosphere of eroded ecosystems reached 79 - 83% (Kosinova et al., 1993). The alienation of some organic matter from chernozems under the influence of erosion affects the replenishment of the nitrogen fund in the course of photo- and heterotrophic nitrogen fixation: aerobic and anaerobic. At the first stages of erosion, anaerobic nitrogen fixation is suppressed at a rapid rate due to the parameters of the labile part of organic matter (Khaziev and Bagautdinov, 1987). The activity of the enzymes invertase and catalase in strongly washed out chernozems decreased by more than 50% in comparison with unwashed chernozems. In gray forest soils, as their washout increases, the invertase activity decreases most sharply. If in weakly washed out soils, a gradual attenuation of activity with depth is noted, then in strongly washed out soils already in the subsoil layer the invertase activity is very low or not detected. The latter is associated with the emergence of illuvial horizons with extremely low enzyme activity on the day surface. In terms of the activity of phosphatase and, especially, catalase, no clear dependence on the degree of soil washout was observed (Lichko, 1998). [...]

The primary substances in lichens are generally the same as in other plants. The hyphae shells in the lichen thallus are composed mainly of carbohydrates. Chitin is often found in hyphae (C30 H60 K4 019). A characteristic component of hyphae is the polysaccharide lichenin (C6H10O6) n, called lichen starch. A less common lichenin isomer, isolichenin, was found, in addition to hyphae membranes, in the protoplast. Of the high-molecular-weight polysaccharides in lichens, in particular in the membranes of hyphae, there are hemicelluloses, which are obviously reserve carbohydrates. In the intercellular spaces of some lichens, pectin substances are found, which, absorbing large quantities of water, swell and lick the thallus. Lichens also contain many enzymes - invertase, amylase, catalase, urease, zymase, lichenase, including extracellular ones. Of the nitrogen-containing substances in lichen hyphae, many amino acids were found - alanine, aspartic acid, glutamic acid, lysine, valine, tyrosine, tryptophan, etc. Phycobiont produces vitamins in lichens, but almost always in small quantities. [...]

During the experiments, it was found that semi-liquid and solid drilling waste has an extremely negative effect on the biological productivity of soils. It is known that the greatest negative impact is exerted by oil and oil products contained in waste. These pollutants significantly reduce the activity of redox and hydrolytic enzymes, which leads to the suppression of the microbiological activity of the soil. This effect is pronounced for waste containing more than 4-5% oil and oil products. With a lower content of this pollutant, the effect of reducing the biological productivity of the considered soil types is characteristic for the period from 3 to 6 months, and then there is an increased reproduction of nitrogen-fixing, denitrifying and sulfate-reducing bacteria, which use oil and its derivatives as a source of carbon and energy, in as a result of which there is a gradual oxidation and mineralization of oil. At the same time, the yield of agricultural crops and the activity of invertase naturally fall. If the waste contains more than 5% of oil and oil products, no visible activity of the hydrocarbon-oxidizing bacterial microflora is observed even after 1 year. The specified level of waste pollution is critical, and therefore requires the use of special agrotechnical and agrochemical methods that stimulate the biological productivity of soils (application of fertilizers containing nitrogen, phosphorus and potassium; intensive aeration of the oil pollution zone; sowing of special herbs that enhance the activity of hydrocarbon-assimilating bacterial microflora). [ ...]

To study the mechanism and nature of the effect of semi-liquid (waste drilling fluids) and solid (drill cuttings) drilling waste, i.e. of those types of waste that are backfilled with mineral soil in sludge pits during their elimination, for the biological productivity of soils and the development on this basis of a complex of agrotechnical measures for the restoration of contaminated lands, vegetation-field and field studies were carried out. The experiments were carried out according to standard methods. We experimented with drilling waste of various degrees of contamination for oil and petroleum products (OP), organic carbon (chemical oxygen consumption index - COD) and mineral salts (calcined residue index - PO), which were added to soils in a 1: 1 ratio. The range and level of contaminated waste are as follows: for NG1 - 1.0-12.0%; COD - 20.0 - 60.0 kg / m3; for PO (per unit area of ​​soil) - 0.4-1.6 kg / m2 of soil. Three types of soil were used in the studies, i.e. the most common types of soils on which drilling is carried out in areas of active agricultural land use. The integral indicators of the biological productivity of soils were the yield of standard barley of the "Courier" variety and the activity of invertase, which was determined according to a well-known method. [...]

However, despite the close relationship that exists between lichens and the substrate on which they settle, it is still not known with certainty whether lichens use the substrate only as a place of attachment or they extract from it some nutrients necessary for their vital activity. On the one hand, the ability of lichens to grow on substrates that are poor in nutrients suggests that they use the substrate only as a place of attachment. However, on the other hand, the selective capacity of lichens during dispersal, the strict confinement of most of them to a certain substrate, the dependence of the species composition of lichen vegetation not only on the physical, but also on the chemical properties of the substrate involuntarily suggests that lichens use the substrate and how additional power supply. This is confirmed by biochemical studies carried out in recent years. For example, it turned out that in the same species of lichen growing on different tree species, the composition of lichen substances may be different. Even more obvious evidence is the discovery of extracellular enzymes in lichens that are secreted into the environment. Extracellular enzymes, such as, for example, invertase, amylase, cellulase and many others, are widely represented in lichens and have a fairly high activity. Moreover, as it turned out, they are most active in the lower part of the thallus, by which the lichen is attached to the substrate. This indicates the possibility of active influence of lichen thallus on the substrate in order to extract additional nutrients from it.

Introduction ... 3

1. Literature review ... 5

1.1 The concept of the enzymatic activity of soils ... 5

1.2 Effect of heavy metals on enzymatic activity

1.3. Influence of agrochemical agents on the enzymatic activity of soils ... 23

2. Experimental part ... 32

2.1 Objects, methods and conditions of research ... 32

2.2. Influence of agrochemical backgrounds on the enzymatic activity of soddy-podzolic soil contaminated with lead ... 34

2.2.1. Agrochemical characteristics of soil contaminated with lead and its content in the soil of the experiment ... 34

2.2.2. Influence of agrochemical backgrounds on the yield of spring grain crops in the heading phase on soil contaminated with lead ... 41

2.2.3. Influence of agrochemical backgrounds on the enzymatic activity of soil contaminated with lead ... 43

2.3. Influence of agrochemical backgrounds on the enzymatic activity of soddy-podzolic soil contaminated with cadmium ... 54

2.3.1. Agrochemical characteristics of soil contaminated with cadmium and its content in the soil of the experiment ... 54

2.3.2. Influence of agrochemical backgrounds on the yield of spring grain crops in the heading phase on soil contaminated with cadmium ... 60

2.3.3. Influence of agrochemical backgrounds on the enzymatic activity of soil contaminated with cadmium ... 62

2.4. Influence of agrochemical backgrounds on the enzymatic activity of sod-podzolic soil contaminated with zinc ... 69

2.4.1. Agrochemical characteristics of soil contaminated with zinc and its content in the soil of the experiment ... 69

2.4.2. Influence of agrochemical backgrounds on the yield of spring grain crops in the heading phase on soil contaminated with zinc ... 75

2.4.3. Influence of agrochemical backgrounds on enzymatic activity

zinc contaminated soil ... 76

2.5. Influence of agrochemical backgrounds on the enzymatic activity of soddy-podzolic soil contaminated with copper ... 82

2.5.1. Agrochemical characteristics of soil contaminated with copper and its content in the soil of the experiment ... 83

2.5.2. Influence of agrochemical backgrounds on the yield of spring grain crops in the heading phase on soil contaminated with copper ... 89

2.5.3. Influence of agrochemical backgrounds on enzymatic activity

soil contaminated with copper ... 90

Conclusion ... 96

Conclusions ... 99

References ... 101

Appendix

Introduction

Introduction.

The use of agrochemicals in the agroecosystem is the most important condition for the development of modern agriculture. This is dictated by the need to maintain and improve the level of soil fertility, and, as a result, obtain high and stable yields.

Agrochemical agents perform a number of ecological functions in the agrocenosis (Mineev, 2000). One of the most important functions of agrochemistry is to reduce the negative consequences of local and global technogenic pollution of agroecosystems with heavy metals (HM) and other toxic elements.

Agrochemical agents reduce the negative impact of HMs in several ways, including by inactivating them in the soil and enhancing the physiological barrier functions of plants that prevent HM from entering them. If on the issue of HM inactivation in soil in the literature there is a lot of information (Ilyin, 1982, etc., Obukhov, 1992, Alekseev, 1987, etc.), then on the enhancement of the barrier functions of plants - isolated studies. Due to the enhancement of physiological barrier functions under the action of agrochemical agents, significantly less HMs enter plants at the same content on different agrochemical backgrounds (Solov'eva, 2002). Strengthening the barrier functions is accompanied by the optimization of plant nutrition, and as a result, an improvement in the biological situation in the soil.

This ecological function, namely, the improvement of the biological activity and the structure of the microbial community of soil contaminated with HM under the influence of agrochemical means, has not yet been sufficiently substantiated experimentally.

It is known that some indicators of biological activity when a stressful situation occurs in the soil change earlier than

other soil characteristics, for example, agrochemical (Zvyagintsev, 1989, Lebedeva, 1984). The enzymatic activity of the soil is one such indicator. Numerous studies have established the negative influence of heavy metals on the activity of enzymes. At the same time, it is known that agrochemical agents have a protective effect in relation to the enzymatic activity of the soil. We tried to consider this problem as a whole and to reveal whether the ecological protective properties of agrochemicals are manifested in relation to the enzymatic activity of the soil when contaminated with biogenic and abiogenic metals. This side of agrochemicals can be detected only if in different variants of the experiment there will be the same amount of HM, and this is possible only with the same indicators of soil acidity. We were unable to find such experimental data in the literature.

1. LITERATURE REVIEW

1.1. The concept of the enzymatic activity of soils.

All biological processes associated with the transformation of substances and energy in the soil are carried out with the help of enzymes that play an important role in the mobilization of plant nutrients, as well as determining the intensity and direction of the most important biochemical processes associated with the synthesis and decomposition of humus, hydrolysis of organic compounds and the redox regime of the soil (1976; 1979, etc.).

The formation and functioning of the enzymatic activity of the soil is a complex and multifactorial process. According to the system-ecological concept, it is a unity of ecologically determined processes of input, stabilization, and manifestation of enzyme activity in the soil (Khaziev, 1991). These three links are defined as the blocks of production, immobilization and action of enzymes (Khaziev, 1962).

Enzymes in the soil are metabolic products of the soil biocenosis, but opinions on the contribution of various components to their accumulation are contradictory. A number of researchers (Kozlov, 1964, 1966, 1967; Krasilnikov, 1958; and others) believe that the main role in the enrichment of the soil with enzymes belongs to the root secretions of plants, others (Katsnelson, Ershov, 1958, etc.) - to soil animals, the majority (Galstyan, 1963; Peive, 1961; Zvyagintsev, 1979; Kozlov, 1966; Drobnik, 1955; Hofmann, Seegerer, 1951; Seegerer, 1953; Hofmann, Hoffmann, 1955,1961; Kiss et al., 1958, 1964, 1971; Sequi, 1974; and others) are of the opinion that the enzymatic pool in the soil consists of intracellular and extracellular enzymes, mainly of microbial origin.

Soil enzymes are involved in the decay of plant, animal and microbial residues, as well as in the synthesis of humus. As a result of enzymatic processes, nutrients from difficult-to-digest

compounds pass into readily available forms for plants and microorganisms. Enzymes are distinguished by extremely high activity, strict specificity of action and great dependence on various environmental conditions. The latter feature is of great importance in the regulation of their activity in the soil (Khaziev, 1982 and

Enzymatic activity of soils (1979)

consists of:

a) extracellular immobilized enzymes;

b) extracellular free enzymes;

c) intracellular enzymes of dead cells;

d) intracellular and extracellular enzymes formed under artificial conditions of the experiment and not typical for this soil.

It has been established that each enzyme acts only on a completely specific substance or a similar group of substances and a completely specific type of chemical bond. This is due to their strict specificity.

By their biochemical nature, all enzymes are high molecular weight protein substances. The polypeptide chain of proteins - enzymes is located in space in an extremely complex way, unique for each enzyme. With a certain spatial arrangement of functional groups of amino acids in the molecule6).

Enzymatic catalysis begins with the formation of an active intermediate, an enzyme-substrate complex. The complex is the result of the attachment of a substrate molecule to the catalytically active center of the enzyme. In this case, the spatial configurations of the substrate molecules are somewhat modified. New oriented

the placement of reacting molecules on the enzyme ensures a high efficiency of enzymatic reactions that reduce the activation energy (Khaziev, 1962).

For the catalytic activity of the enzyme, not only the active center of the enzyme is responsible, but also the entire structure of the molecule as a whole. The rate of the enzymatic reaction is regulated by many factors: temperature, pH, enzyme and substrate concentration, the presence of activators and inhibitors. Organic compounds can act as activators, but more often various microelements (Kuprevich, Shcherbakova, 1966).

The soil is able to regulate the enzymatic processes occurring in it in connection with changes in internal and external factors through factorial or allosteric regulation (Galstyan 1974, 1975). Allosteric regulation occurs under the influence of chemical compounds introduced into the soil, including fertilizers. Factorial regulation is due to the acidity of the environment (pH), chemical and physical composition, temperature, humidity, water-air regime, etc. The influence of soil specificity, humus and biomass content and other factors on the activity of enzymes used to characterize the biological activity of soils is ambiguous. (Galstyan, 1974; Kiss, 1971; Dalai, 1975, McBride, 1989, Tiler, 1978).

The enzymatic activity of the soil can be used as a diagnostic indicator of the fertility of various soils, because the activity of enzymes reflects not only the biological properties of the soil, but also their changes under the influence of agroecological factors (Galstyan, 1967; Chunderova, 1976; Chugunova, 1990, etc.).

The main routes of entry of enzymes into the soil are intracellular extracellular enzymes of microorganisms and plant roots and intracellular enzymes that enter the soil after the death of soil organisms and plants.

The release of enzymes into the soil by microorganisms and plant roots is usually adaptive in the form of a response to the presence or absence of a substrate for the action of an enzyme or a reaction product, which is especially pronounced with phosphatases. With a lack of mobile phosphorus in the environment, microorganisms and plants sharply increase the release of enzymes. This relationship is the basis for the use of the value of soil phosphatase activity as a diagnostic indicator of plant availability with available phosphorus (Naumova, 1954, Kotelev, 1964).

Enzymes, getting from various sources into the soil, are not destroyed, but remain in an active state. It must be assumed that enzymes, being the most active component of the soil, are concentrated where the vital activity of microorganisms is most intense, that is, at the interface between soil colloids and soil solution. It has been experimentally proven that enzymes in soil are mainly in the solid phase (Zvyagintsev, 1979).

Numerous experiments carried out under conditions of suppression of enzyme synthesis in microbial cells using toluene (Drobnik, 1961; Beck, Poshenrieder, 1963), antibiotics (Kuprevich, 1961; Kiss, 1971) or radiation (McLaren et al., 1957) indicate that that the soil contains a large amount of "accumulated enzymes", sufficient to carry out the transformation of the substrate for some time. These enzymes include invertase, urease, phosphatase, amylase, etc. Other enzymes are much more active in the absence of an antiseptic, which means that they accumulate in the soil insignificantly (a - and P-galactosidase, dextranase, levanase, malatesterase, etc.). The third group of enzymes does not accumulate in the soil, their activity is manifested only during the outbreak of the vital activity of microbes and is induced by the substrate. Received so far

experimental data indicate a difference in the enzymatic activity of soils of different types (Konovalova, 1975; Zvyagintsev, 1976; Khaziev, 1976; Galstyan, 1974, 1977, 1978; and others).

The most well-studied enzymes in soil are hydrolases, which represent a wide class of enzymes that carry out hydrolysis reactions of various complex organic compounds, acting on various bonds: ester, glucoside, amide, peptide, etc. Hydrolases are widespread in soils and play an important role in their enrichment mobile nutrients sufficient for plants and microorganisms, destroying high-molecular organic compounds. This class includes the enzymes urease (amidase), invertase (carbohydrase), phosphatase (phosphohydrolase), etc., the activity of which is the most important indicator of the biological activity of soils (Zvyagintsev, 1980).

Urease is an enzyme involved in the regulation of nitrogen metabolism in the soil. This enzyme catalyzes the hydrolysis of urea to ammonia and carbon dioxide, causing the hydrolytic cleavage of the bond between nitrogen and carbon in organic molecules.

Of the enzymes of nitrogen metabolism, urease has been studied better than others. It is found in all soils. Its activity correlates with the activity of all the main enzymes of nitrogen metabolism (Galstyan, 1980).

In soil, urease is found in two main forms: intracellular and extracellular. The presence of free urease in the soil allowed Briggs and Segal (Briggs et al., 1963) to isolate the enzyme in crystalline form.

Part of the extracellular urease is adsorbed by soil colloids, which have a high affinity for urease. The connection with soil colloids protects the enzyme from decomposition by microorganisms and promotes its accumulation in the soil. Each soil has its own stable level of urease activity, determined by the ability of soil colloids,

mainly organic, exhibit protective properties (Zvyagintsev, 1989).

In the soil profile, the humus horizon exhibits the highest enzyme activity; further distribution along the profile depends on the genetic characteristics of the soil.

In connection with the widespread use of urea as a nitrogen fertilizer, issues related to its transformations under the action of urease are practically significant. The high urease activity of most soils prevents the use of urea as a universal source of nitrogen nutrition, since the high rate of urea hydrolysis by soil urease leads to a local accumulation of ammonium ions, an increase in the reaction of the medium to alkaline values, and as a consequence, nitrogen losses from the soil in the form of ammonia ( Tarafdar J. C, 1997). By splitting urea, urease prevents its isomerization into phototoxic ammonium cyanate. Although urea itself is partially used by plants, however, as a result of the active action of urease, it cannot persist in the soil for a long time. In the studies of a number of scientists, volatilization of urea nitrogen in the form of ammonia from the soil with high urease activity was noted, and when various urease inhibitors were introduced into the soil, the hydrolysis of urea was slowed down and the losses were less (Tool P. O., Morgan M. A., 1994). The rate of urea hydrolysis in the soil is influenced by temperature (Ivanov, Baranova, 1972; Galstyan, 1974; Cortez et al., 1972, etc.), soil acidity (Galstyan, 1974; Moiseeva, 1974, etc.). The saturation of the soil with carbonates has a negative effect (Galstyan, 1974), the presence of significant amounts of salts of arsenic, zinc, mercury, sulfate ions, copper and boron compounds, from organic compounds, aliphatic amines, dehydrophenols and quinones significantly inhibit urease (Paulson, 1970, Briggsatel ., 1951).

Invertase activity is one of the most stable indicators, revealing the clearest correlations with influencing factors. Research (1966, 1974) established the correlation of invertase with the activity of other soil carbohydrases.

The invertase activity has been studied in many soils and discussed in several review works (Aleksandrova and Shmurova, 1975; Kuprevich and Shcherbakova, 1971; Kiss et al., 1971, and others). The invertase activity in the soil decreases along the profile and correlates with the humus content (Pukhitskaya and Kovrigo, 1974; Galstyan, 1974; Kalatozova, 1975; Kulakovskaya and Stefankina, 1975; Simonyan, 1976; Toth, 1987; etc.). Correlation with humus may be absent if there is a significant content of aluminum, iron, sodium in the soil. The close relationship between invertase activity and the number of soil microorganisms and their metabolic activity (Mashtakov et al., 1954; Katsnelson and Ershov, 1958; Kozlov, 1964; Chunderova, 1970; Kiss, 1958; Hofinann, 1955, etc.) indicate an advantage in soil microbial invertases. However, this dependence is not always confirmed (Nizova, 1970), the invertase activity is a much more stable indicator and may not be directly related to fluctuations in the number of microorganisms (Ross, 1976).

According to a report (1974), soils with a heavy granulometric composition have a higher enzymatic activity. However, there are reports that invertase is noticeably inactivated upon adsorption by clay minerals (Hofmann et al., 1961; Skujins, 1976; Rawald, 1970), and soils with a high content of montmorillonite have low invertase activity. The dependence of invertase activity on soil moisture and temperature has been insufficiently studied, although many authors explain seasonal changes in activity by hydrothermal conditions.

The effect of temperature on the potential activity of invertase was studied in detail (1975), setting the optimum at a temperature of about 60 ° C, the threshold of enzyme inactivation after warming up the soil at 70 ° C, and complete inactivation after three hours of heating at 180 ° C.

Many authors considered the invertase activity of soils depending on growing plants (Samtsevich, Borisova, 1972; Galstyan, 1974, Ross 1976; Cortez et al., 1972, etc.). The development of the meadow process, the formation of a thick sod under the herbaceous cover promotes an increase in invertase activity (Galstyan, 1959). However, there are works in which the effect of plants on the activity of invertase has not been established (Konovalova, 1975).

In soils, there is a large amount of phosphorus in the form of organic compounds, which comes with dying remains of plants, animals and microorganisms. The release of phosphoric acid from these compounds is carried out by a relatively narrow group of microorganisms with specific phosphatase enzymes (Chimitdorzhieva et al., 2001).

Among the enzymes of phosphorus metabolism, the activity of orthophosphoric monophosphoesterases has been studied most fully (Aleksandrova, Shmurova, 1974; Skujins J. J., 1976; Kotelev et al., 1964). The producers of phosphatases are mainly the cells of soil microorganisms (Krasilnikov and Kotelev, 1957, 1959; Kotelev et al., 1964).

Phosphatase activity of the soil is determined by its genetic characteristics, physicochemical properties and the level of farming culture. Among the physicochemical properties of soil, acidity is especially important for phosphatase activity. Sod-podzolic and gray forest soils, which have an acidic reaction, predominantly contain acidic phosphatases; in soils with a weakly alkaline reaction, alkaline phosphatases predominate. It should be noted that the optimum activity of acidic

phosphatase is found in a weakly acidic zone, even when the soils have a strongly acidic reaction (Khaziev, 1979; Shcherbakov et al., 1983, 1988). This fact confirms the importance of liming acidic soils to accelerate the hydrolysis of complex organic phosphates and enrichment of the soil with available phosphorus.

The observed characteristic distribution of phosphatases in soils, depending on their acidity, is due to the composition of the microflora. Microbial communities adapted to certain environmental conditions function in the soil, which secrete enzymes that are active under these conditions.

The total phosphatase activity of the soil depends on the content of humus and organic phosphorus, which is a substrate for the enzyme.

Chernozems are characterized by the highest phosphatase activity. In sod-podzolic and gray forest soils, the phosphatase activity is low. The low activity of these acidic soils is due to the stronger adsorption of phosphatases by soil minerals. Due to the low content of organic matter in such soils, the adsorbing surface of minerals is more exposed in comparison with high-humus chernozems, where clay minerals are covered with humified organic matter.

Phosphatase activity is dynamic during the growing season. During the active phases of plant growth at high soil temperatures and sufficient moisture in the summer months, the phosphatase activity of soils is maximal (Evdokimova, 1989).

In some soils, a correlation was noted between phosphatase activity and the total number of microorganisms (Kotelev et al., 1964; Aliev, Gadzhiev, 1978, 1979; Harutyunyan, 1975, 1977; and others) and the number of microorganisms mineralizing organic phosphorus compounds (Ponomareva et al. , 1972), in others - the relationship of phosphatase activity with the number

microorganisms have not been established (Ramirez-Martinez, 1989). The effect of humus is manifested in the nature of changes in the activity of the enzyme along the profile, when comparing soils with different degrees of humus content and taking measures to cultivate the soil (Aleksandrova and Shmurova, 1975; Arutyunyan, 1977). Studies by many authors indicate a direct dependence of the phosphatase activity of soils on the content of organic phosphorus in the soil (Gavrilova et al., 1973; Harutyunyan and Galstyan, 1975; Harutyunyan, 1977; and others).

Let us consider in somewhat more detail the general regularities of the formation of the phosphatase pool of soils.

A significant part of the total phosphorus in the soil is made up of organophosphorus compounds: nucleic acids, nucleotides, phytin, lecithin, etc. Most of the organophosphates found in the soil are not directly assimilated by plants. Their absorption is preceded by enzymatic hydrolysis carried out by phosphohydrolases. Substrates of soil phosphatases are specific humic substances, including phosphorus of humic acids, as well as non-specific individual compounds represented by nucleic acids, phospholipids and phosphoproteins, as well as metabolic phosphates. The former accumulate in the soil as a result of the biogenesis of humic substances, the latter, as a rule, enter the soil with plant residues and accumulate in it as products of intermediate metabolic reactions.

The role of higher plants in the formation of the phosphatase pool of soils used in agriculture is lower than that of microorganisms and is mainly associated with the entry of crop residues and root exudates into the soil, which is confirmed by the data and (1994), which investigated the influence of various agricultural crops on activity of hydrolytic

and redox enzymes; phosphatase, invertase, protease, urease, catalase on shallow peat soil. The phosphatase activity was approximately the same under all crops: barley, potatoes, and black fallow, and only slightly more under perennial grasses, while the activity of other enzymes varied significantly depending on the nature of soil use.

, (1972) note an increase in phosphatase activity in the rhizosphere of wheat and legumes, which may be associated with both an increase in the number of microorganisms in the rhizosphere and with the extracellular phosphatase activity of roots. From an agrochemical point of view, the final result is important - the growth of the enzymatic pool of soils with an increase in the power of plant root systems.

The depletion of plants in agrocenoses leads to a decrease in the rhizosphere effect and, as a consequence, to a decrease in the activity of soil phosphatase. A significant decrease in the phosphatase activity of soils was noted during the cultivation of monoculture. The inclusion of soils in the crop rotation creates conditions for the improvement of hydrolytic processes, which leads to an increase in the metabolism of phosphorus compounds. (Evdokimova, 1992)

(1994) studied soddy-podzolic soils formed under natural (forest) vegetation of different composition and determined the distribution of phosphatase activity in the soil profile, the ratio between labile and stable forms of enzymes, and their spatial and temporal variability. It was found that in soils formed under natural forest vegetation, the genetic horizons differ in phosphatase activity, the distribution of which in the profile closely correlates with the humus content. According to the data, the highest phosphatase activity was observed in the litter layer, then decreased by times in the humus-accumulative layer and sharply decreased in the soil layer.

below 20 cm in the soil under the spruce (forest vegetation). Under meadow vegetation, the distribution is somewhat different: the maximum activity in the soddy horizon is 1.5-2 times lower in the humus-accumulative horizon, and a further significant decrease is observed only after 40-60 cm. Based on the above, it can be concluded that the maximum contribution to the formation of the phosphatase pool under natural vegetation, microorganisms and plant debris are introduced as a substrate, root exudates and intracellular enzymes entering postmorally play a somewhat lesser role.

The intensity of biochemical processes in the soil and the level of its fertility depends both on the conditions of existence of living organisms that supply enzymes to the soil, and on factors that contribute to the fixation of enzymes in the soil and regulate their actual activity.

1.2. The influence of heavy metals and microelements on the enzymatic activity of soils.

One of the promising directions of using enzymatic activity for diagnosing the biological properties of soils is to identify the level of soil contamination with HM.

Heavy metals, entering the soil in the form of various chemical compounds, can accumulate in it to high levels, which pose a significant danger to the normal functioning of the soil biota. A large amount of data has been accumulated in the literature, indicating the negative impact of soil pollution with HMs on soil biota. When the chemical balance in the soil is disturbed, a stressful situation arises. There is evidence that biological indicators react earlier than agrochemical indicators to changes in conditions that affect various soil properties (Lebedeva,

Bibliography

Enzymes are catalysts for chemical reactions of a proteinaceous nature, characterized by the specificity of action in relation to the catalysis of certain chemical reactions. They are products of the biosynthesis of all living soil organisms: woody and herbaceous plants, mosses, lichens, algae, microorganisms, protozoa, insects, invertebrates and vertebrates, represented in the natural environment by certain aggregates - biocenoses.

The biosynthesis of enzymes in living organisms is carried out due to genetic factors responsible for the hereditary transmission of the type of metabolism and its adaptive variability. Enzymes are the working apparatus by which the action of genes is realized. They catalyze thousands of chemical reactions in organisms, which ultimately constitute cellular metabolism. Thanks to them, chemical reactions in the body are carried out at high speed.

More than 900 enzymes are currently known. They are classified into six main classes.

1. Oxy reductases that catalyze redox reactions.

2. Transferases that catalyze the reactions of intermolecular transfer of various chemical groups and residues.

3. Hydrolases that catalyze the reactions of hydrolytic cleavage of intramolecular bonds.

4. Lyases catalyzing the reactions of addition of groups to double bonds and the reverse reactions of the removal of such groups.

5. Isomerases catalyzing isomerization reactions.

6. Ligases that catalyze chemical reactions with the formation of bonds due to ATP (adenosine triphosphoric acid).

When living organisms die off and decay, some of their enzymes are destroyed, and some of them, getting into the soil, retain their activity and catalyze many soil chemical reactions, participating in the processes of soil formation and in the formation of a qualitative feature of soils - fertility. In different types of soils, under certain biocenoses, their own enzymatic complexes have been formed, which differ in the activity of biocatalytic reactions.

VF Kuprevich and TA Shcherbakova (1966) note that an important feature of the enzymatic complexes of soils is the orderliness of the action of the existing groups of enzymes, which manifests itself in the fact that the simultaneous action of a number of enzymes representing different groups is ensured; the formation and accumulation of compounds present in the soil in excess are excluded; the excess of accumulated mobile simple compounds (for example, NH 3) are temporarily bound in one way or another and sent to cycles, ending with the formation of more or less complex compounds. Enzymatic complexes are self-balancing self-regulating systems. In this, microorganisms and plants play the main role, constantly replenishing soil enzymes, since many of them are short-lived. The amount of enzymes is indirectly judged by their activity over time, which depends on the chemical nature of the reacting substances (substrate, enzyme) and on the conditions of interaction (concentration of components, pH, temperature, composition of the medium, action of activators, inhibitors, etc.).

This chapter examines the participation in some chemical soil processes of enzymes from the class of hydrolases - the activity of invertase, urease, phosphatase, protease and from the class of oxidoreductases - the activity of catalase, peroxidase and polyphenol oxidase, which are of great importance in the conversion of nitrogen- and phosphorus-containing organic substances, substances of a carbohydrate nature and in the processes of humus formation. The activity of these enzymes is an essential indicator of soil fertility. In addition, the activity of these enzymes in forest and arable soils of various degrees of cultivation will be characterized using the example of sod-podzolic, gray forest and sod-calcareous soils.

CHARACTERISTICS OF SOIL ENZYMES

Invertase - catalyzes the hydrolytic cleavage of sucrose into equimolar amounts of glucose and fructose, also affects other carbohydrates with the formation of fructose molecules - an energy product for the life of microorganisms, catalyzes fructose transferase reactions. Studies by many authors have shown that invertase activity reflects the level of soil fertility and biological activity better than other enzymes.

Urease - catalyzes the reaction of hydrolytic cleavage of urea into ammonia and carbon dioxide. In connection with the use of urea in agronomic practice, it should be borne in mind that the activity of urease is higher in more fertile soils. It increases in all soils during the periods of their greatest biological activity - in July - August.

Phosphatase (alkaline and acidic) - catalyzes the hydrolysis of a number of organophosphorus compounds to form orthophosphate. Phosphatase activity is inversely related to the availability of mobile phosphorus to plants; therefore, it can be used as an additional indicator in determining the need for phosphorus fertilization in soils. The highest phosphatase activity is in the rhizosphere of plants.

Proteases are a group of enzymes, with the participation of which proteins are broken down to polypeptides and amino acids, then they are hydrolyzed to ammonia, carbon dioxide and water. In this regard, proteases are of great importance in soil life, since they are associated with changes in the composition of organic components and the dynamics of nitrogen forms assimilated for plants.

Catalase - as a result of its activating action, hydrogen peroxide, toxic to living organisms, is split into water and free oxygen. Vegetation has a great influence on the catalase activity of mineral soils. As a rule, soils under plants with a powerful deeply penetrating root system are characterized by high catalase activity. The peculiarity of catalase activity is that it changes little down the profile, has an inverse dependence on soil moisture and a direct dependence on temperature.

Polyphenol oxidase and peroxidase - they play an important role in the processes of humus formation in soils. Polyphenol oxidase catalyzes the oxidation of polyphenols to quinones in the presence of free atmospheric oxygen. Peroxidase, on the other hand, catalyzes the oxidation of polyphenols in the presence of hydrogen peroxide or organic peroxides. Moreover, its role is to activate peroxides, since they have a weak oxidizing effect on phenols. Further, the condensation of quinones with amino acids and peptides can occur with the formation of a primary molecule of humic acid, which can be further complicated due to repeated condensations (Kononova, 1963).

It was noted (Chunderova, 1970) that the ratio of the activity of polyphenol oxidase (S) to the activity of peroxidase (D), expressed as a percentage (), has a relationship with the accumulation of humus in soils; therefore, this value was called the conditional coefficient of humus accumulation (K). In the arable, poorly cultivated soils of Udmurtia, for the period from May to September, it was: in the soddy-podzolic soil - 24%, in the gray forest podzolized soil - 26% and in the soddy-calcareous soil - 29%.

ENZYMATIVE PROCESSES IN SOILS

Biocatalytic activity of soils is in significant agreement with the degree of their enrichment with microorganisms (Table 11), depends on the type of soil and changes along the genetic horizons, which is associated with the peculiarities of changes in the humus content, reaction, Red-Ox-potential and other parameters along the profile.

In virgin forest soils, the intensity of enzymatic reactions is mainly determined by the horizons of the forest litter, and in arable soils, by the arable layers. Both in some and in other soils, all biologically less active genetic horizons located under the A or A p horizons have low enzyme activity, which slightly changes in the positive direction during soil cultivation. After the development of forest soils for arable land, the enzymatic activity of the formed arable horizon in comparison with the forest litter is sharply reduced, but as it is domesticated, it increases and, in highly cultivated species, approaches or exceeds the values ​​of the forest litter.

11. Comparison of biogenesis and enzymatic activity of soils in the Middle Urals (Pukhidskaya, Kovrigo, 1974)

No. of section, name of soil	Horizon, depth of sampling, cm		The total number of microorganisms, thousand per 1 g of abs. dry soil (on average for 1962, 1964-1965)	Enzyme activity indicators (on average for 1969-1971)
				Invertase, mg glucose per 1 g of soil for the first day	Phosphatase, mg phenolphthalein per 100 g of soil for 1 hour	Urease, mg NH, per 1 g of soil for 1 day	Catalase, ml 0 2 per 1 g of soil for 1 min	Polyphenol oxidase		Peroxidase
								mg of purpurogallin per 100 g of soil
3. Sod-medium podzolic medium loamy (under the forest)
					Not defined
1.Soddy-medium-podzolic medium-loamy, poorly cultivated
10.Gray-forest podzolized heavy loamy, poorly cultivated
2. Sod-carbonate slightly leached light loamy, poorly cultivated

The activity of soil biocatalytic reactions changes. It is lowest in spring and autumn, and the highest is usually in July-August, which corresponds to the dynamics of the general course of biological processes in soils. However, depending on the type of soil and their geographic location, the dynamics of enzymatic processes is very different.

Test questions and tasks

1. What compounds are called enzymes? What are their production and significance for living organisms? 2. Name the sources of soil enzymes. What role do individual enzymes play in soil chemical processes? 3. Give the concept of the enzymatic complex of soils and its functioning. 4. Give a general description of the course of enzymatic processes in virgin and arable soils.