Soil Organic Matter

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Definition: Soil organic matter consists of a whole series of products which range from undecayed plant and animal tissues through ephemeral products of decomposition to fairly stable amorphous brown to black material bearing no trace of the anatomical structure of the material from which it was derived and it is the latter material that is normally defined as ‘humus’.

Humus: According to Buckman and Brady, Humus is a complex and rather resistant mixture of brown or dark brown amorphous and colloidal organic substances modified from the original tissues or synthesized by the various soil organisms.

Non- humified & humified organic matter: Soil organic matter consists of two major types of compounds, unhumified substances and the humified remains of plant and animal tissues. The non-humified organic matter is composed of compounds released during decomposition in the original or slightly modified form. Although numerous organic compounds are present in the plant tissue, only a few exist in soils in detectable amounts after their release in soils. They are primarilyi. carbohydrate ii. amino acids and proteins iii. lipids iv. nucleic acids v. lignin and vi. organic acids Humified organic matter or humic matter is a group of compounds that includes humic acids, fulvic acids, hymotomelanic acid and humans. This humified soil organic fraction is also known as ‘humus’ or currently as ‘humic compounds’. Today humic compounds are defined as amorphous, colloidal poly dispersed substances with yellow to brown –black color and high molecular weights.

Sources of organic matter: 1. The original/primary source of the soil organic matter is plant tissue. Under natural conditions, the tops & roots of tress, shrubs, grasses ad other native plants annually supply large quantities of organic residues. Even with harvested crops, one tenth to one third of the plat tops commonly fall to the soil and remain there or are incorporated into the soil.

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But these organic materials are decomposed and digested by soil organisms; they become part of the underlying soil by infiltration or by actual physical incorporation. Accordingly, the residues of higher plants provide food for soil organism, which in turn create stable compounds that help maintain the soil organic levels. 2. The secondary sources of organic matter are animals. As they attack the original plant tissues, they contribute waste products and leave their own bodies as their life cycles are consummated. Certain forms of animal life, especially the earthworms, termites, and ants also play an important role in the translocation of soil and plant residues. 3. Another source of organic matter is soil organisms. It is of ecological significance.

Types of materials present in organic matter: There are three types of material in soil organic matter. These are: 1. Fresh or undecomposed material Materials in which the anatomical structure of the plant substances is still visible. 2. Partially decomposed materials: Soft green portions are decomposed in this stage. This decomposition depends upon the composition of the tissues. 3. Completely decomposed material: In this stage, the materials are completely decomposed, and are called ‘humus’. The whole of the organic residue is not decomposed all at once or as a whole. Some of the constituents are decomposed very rapidly some less readily, and others very slowly. The speed with which the organic residues are decomposed depends on the nature and abundance of various constituents that make up the residues, and the environmental condition, viz, moisture supply, aeration, temperature and soil reaction, under which the microorganisms carry out their activities. As the original material is decomposed new ones are simultaneously synthesized in the forms of microbial bodies and tissues. It is evident that soil organic matter consists of plant, animal and microbial residues of various stages of decomposition and contains, at any given time, i. original residues,

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ii. iii.

Various products of their decomposition, both simple and complex; and Newly synthesized microbial bodies, both dead and alive.

Composition of plant tissues: On a volume basis, the moisture content of plant residues is high; varying from 60 to 90% and the rest 10 to 40% is solid matter. On a weight basis, 75% is H2O & 25% is solid matter, there are 11% C, 10% O, 2% H & the rest 2% is others (N, P, K, Mg, As, etc. i.e., ash). On an elemental basis (number of atoms of the elements), hydrogen predominates. In representative plant residues, there are 8 hydrogen atoms for every 3.7 carbon atoms and 2.5 oxygen atoms (H: C: O=8:3.7:2.5). These three elements dominate the bulk of organic tissue in the soil. Even though other elements are present only in small quantities, they play vital role in plant nutrition and in meeting microorganism body requirements. The actual organic compounds in plant tissue are many and varied and they can be grouped into a small number of classes. Representative percentages as well as ranges are shown in the following figure: Water 75% Dry matter 25%

45% cellulose 5%su 2% fat 8%pr 18%cellulose

Carbon42% Hydrogen 8%

8%ash

Figure: Typical composition of representative green-plant material

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The carbohydrates, which range in complexity from simple sugars and starches to cellulose, are usually the most prominent of the organic compounds found in plants. Fats and oils, which are somewhat more complex than carbohydrates and less, so than lignins, are found primarily in seeds & leaf coatings. Proteins contain- in addition to carbon, oxygen and hydrogen – about 16% nitrogen and smaller amounts of other essential elements such as sulfur, manganese, copper and iron. Proteins are primary sources of these essential elements. Simple proteins decompose easily while the complex crude proteins are more resistant to breakdown and release their nitrogen. Lignins which are complex compounds with multiple type or phenol structures are components of plant cell walls. The content of lignin increases as plants mature and is especially high in woody tissues. Lignins and polyphenols are notoriously resistant to decomposition. Rate of decomposition: Organic compounds may be limited in terms of ease of decomposition as follows: 1. Sugars, starches and simple proteins Rapid 2. Crude proteins decomposition 3. Hemicelluloses 4. Cellulose Very slow 5. Fats, Waxes & so forth decomposition 6. Lignin and phenolic compounds. Some general terms: *Glucose Glucose is the smallest unit of polysaccharide. It is an important monosaccharide. The molecular sign of glucose is C6H12O6; it has one aldehyde radical (-CHO), four secondary (-CHOH) & one primary (-CH2OH) alcohol radical. Another name of glucose is ‘dextrose’. There are two configurations: i. D-glucose & ii. L-glucose CHO CHO (CHOH) 3

(CHOH)3

H-C-OH

HO-C-H

CH2OH

CH2OH

D-Glucose

L-Glucose

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There are 2 stereo isomers of glucose: i. α- D- glucose ii. β- D- glucose H

OH C

H- C-OH OH-C-H O H- C-OH C CH2OH α- D- glucose CHO (CHOH) 3 H-C-OH CH2OH D-glucose H

OH C

H- C-OH - β-D-glucose OH-C-H O H- C-OH C CH2OH

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7

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The organic compounds (non- humified):

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The plant tissue is composed of C, H, O, N, S, P and a number of other elements. The organic part of the plant tissue is composed of numerous organic compounds, but only a few are present in detectable amounts in soils after decomposition. They are primarily i. Carbohydrates ii. Amino acids & proteins iii. Lipids iv. Nucleic acids v. Lignins vi. Humus • Carbohydrates Carbohydrates are, by definition, poly hydroxy aldehydes, ketons or substances that yield one of these compounds on hydrolysis. Glucose (C6H12O6) and fructose are examples of an aldose and a kelose, respectively.

H

O C

H- C-OH OH-C-H O H- C-OH C CH2OH Glucose (aldose)

H

OH

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C H- C=O OH-C-H O H- C-OH C CH2OH Fructose (Kelose) The term carbohydrate indicates that these compounds could be represented by hydrates of carbon: Cx(H2O)y. But several compounds exist with properties of carbohydrates which do not have the required ratio of hydrogen to oxygen 0f 2:1; e.g. the sugar deoxyribose (C5H10O4), which is a constituent of DNA, a component of every plant cell. Some of the carbohydrates may also contain N and S, and their formulas do not agree with Cx(H2O)y. Approximately 5 to 16% of the soil organic matter is in the form of carbohydrates. Sources: • Soil carbohydrates are derived from plant residues and from the remains of microorganisms and animals. • A major portion of the soil organic matter is derived from the added and residual vegetative material. • Animals contribute a smaller portion of soil carbohydrates. • Minerals contribute indirectly to soil carbohydrates. • Soil bacteria are capable of producing polysaccharides containing all sugars found in soils except arabinose and galactose. Classification: Carbohydrates can be divided into three groups: i. Monosaccharide ii. Oligosaccharides iii. Polysaccharides The properties of these carbohydrates change significantly with increasing molecular complexity.

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Monosaccharides are simple sugars that can’t be hydrolyzed into smaller molecules under reasoning mild conditions. According to the number of carbon atoms, monosaccharide may be trioses (C3H6O3), testroses, and so on up to octoses or nonoses. Hexoses-(glucose, galactose, mannose, fructose) Pentoses-(arabinose, xylose, ribose, fucose, rhammose). Oligosaccharides are compound sugars that, upon hydrolysis, yield two to six molecules of simple sugars. Disaccharides for example, hydrolyzes into two monosaccharide and upon hydrolysis, pentosaccharides yield five monosaccharide. Disaccharide (sucrose, cellobiose, gentibiose). Polysaccharides are group of compounds that yield many different monosaccharides upon hydrolysis. They include cellulose and hemicelluloses. Polysaccharides are very complex in structure and have high molecular weights. They are sometimes designated into: i. Homopolysaccharides ii. Hetaropolysaccharides • Amino acids & proteins Amino acids are the fundamental structural units of protein. The nitrogen in amino acids, occurs as an amino (-NH2) group, is attached to the C chain. The acid part consists of a terminal C linked to an O atom and an OH group, often written as –COOH which is called carboxyl group. These are the reasons for the name amino acids. The general formula of amino acids may be written as, NH2 R-CH-COOH Proteins are complex combinations of amino acids. 21 amino acids are usually found as protein constituents. The protein is formed by the linkage of many amino acids through the amino & carboxyl group. H O H H O H2N-C-C-OH+H-N-C-C-OH Glycerin amino acid HO H HO H2N-C-C-N-C-C-OH +H2O

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H

H

(Peptide bond) Both amino acids and proteins are major sources of nitrogen compounds in soils. They are more difficult to breakdown than carbohydrates because of their size and complexity in molecular structure. They are amphoretic in nature, and consequently react with acids and bases. At the isoelectric point, amino acids behave as zwilter ions; in other words, behave as cations & anions (Tan 1993). In acid soils, the amino acids are positively charged and behave as cations; whereas in basic soils they are negatively charged and behave as anions. O R-CH-C-O-H NH2 In acid medium:

In basic medium:

O R-CH-C-Oˉ NH3

O

O

R-CH-C-Oˉ + H+

R-CH-C-OH

NH3

NH3 O

R-CH-C-Oˉ + OHˉ

O R-CH-C-Oˉ +H2O

NH3 NH2 Decomposition of amino acids & proteins: The main reaction process for the decomposition of three compounds is hydrolysis. Hydrolysis of proteins, brought about by proteinases and peptides, of soil microorganisms, results in cleavage of peptide bonds, releasing in this way produces the amino acids. The latter compounds are broken down further into NH3 by the enzymes called amino acid dehydrogenases and oxidases. Schematically the main pathway of decomposition can be represented as follows: Proteins—Peptides---amino acids---NH3 The decomposition reaction of proteins as described above is frequently called deamination or putrefication (Gartner, 1949;

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Stevenson, 1986). Deamination reactions can take place in aerobic or anaerobic conditions, and are, therefore called oxidative and non-oxidative deamination respectively. The reaction for oxidative deamination can be written as: R-CH-COOH-O2---R-COOH+CO2+NH3 NH2 Anaerobic deamination may result in i. Deamination or reduction, and ii. decarboxylation, which can be written as follows: Deamination or reduction: R-CH-COOH+H2-------R-CH2-COOH+ NH3 NH2 Decarboxylation: R-CH-COOH-------R-CH2-NH2 +CO2 NH2 (Amino acid) (Amine) • Lipids: Lipids are heterogeneous compounds of fatty acids, waxes, and oils. The term lipid does not imply a particular chemical structure, as with amino acids. The name is used to describe substances that are soluble in fat solvents, such as ether, chloroform, or benzene. Lipids are usually classified into three groups: 1. Simple lipids: These include natural lipids, fats, oils and waxes. 2. Compound lipids: Phosphatides, glycolipids, sulfolipids and terpenoid lipids, including carotenoids, belong to this group. 3. Derived lipids: these are lipids derived from hydrolysis of simple & compound lipids. They include fatty acids, alcohols and sterols. The fatty acids can be unsaturated fatty acids (e.g. oleic acid C18H34O12) palm oil or coconut oil is rich in palmitic acids. Cholesterol is an example of a sterol, which upon UV radiation will form vitamin D. The basic component of lipids is glycerol, C3H8O3 or other alcohols; glycerol is a trihydroxy alcohol with the following structure: H2C-OH

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HC-OH H2C-OH Lipids have limited solubility in water and exhibit a hydrophobic character. Many of the lipids in plant and animals are associate with proteins and carbohydrates (e.g. glycolipids) • Nucleic acids Each plant and animal cell contains a discrete rounded or spherical body, called the nucleus, which contains nucleic acids. Nucleic acids, first isolated in 1869 by F. Miescher, are polymers with high molecular weights. Their repeating unit is a mononucleotide, rather than an amino acid. Two types of nucleic acids are generally recognized: i. deoxyribonucleic acid (DNA), a constituent of cell nuclei; and ii. Ribonucleic acid (RNA) located in the nucleolus and in the cytoplasmic membrane, called endoplasmic reticulum. • Lignin Lignin is a system of thermoplastic highly aromatic polymers, derived from coniferyl alcohol or guaiacyl propane monomers. Plant lignin can be divided into three types of basic monomers: i. Lignin from softwood- coniferyl alcohol, derived mostly from softwood or coniferyl plants. ii. Lignin from hardwood- sinapyl alcohol, derived from hard wood. iii. Lignin from grasses, bamboo or palm. Or grass lignincoumaryl alcohol, derived from grasses and bamboos. These basic monomers form large, complex polymer (polymeric molecules). A hypothesis of a more systematic arrangement of the basic monomers into lignin is presented by Tan (1993). Many of the C atoms are connected to the

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OH Radicals (phenolic hydroxyl groups) in which the behavior of the H is much the same as that in the carboxyl groups of organic acids. An example of a systematic linkage of softwood lignin monomers to form polymeric lignin is shown in the following figure:

Such a combination can also take place with the other types of monomers, whereas the linkages can continue in many directions. The ultimate source for formation of lignin is carbohydrates or intermediate products of photosynthesis related to carbohydrates. In the growth of woody plants, carbohydrates are synthesized first. The formation of lignin then begins, and the spaces existing between the cellulose fibers are gradually filled with lignified carbohydrates. This process is called lignification and serves several functions:

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a. it elements and anchors the fibers together b. It increases the resistance of the fibers against physical and chemical breakdown. c. It increases rigidity and strength of cell walls. It is believed that after lignification, the lignified tissue then no longer plays an active role in the life of plants, but serves only as a supporting structure. The bulk of the lignin occurs in the secondary cell walls where it is associated with cellulose & hemi cellulose in stems. The quantity of lignin increases with plant age & stem content. it is very important constituent of woody tissue and it contains the major portion of the methoxyl content of the wood. A large amount of lignin is also detected in the vascular bundles of plant issues. Lignin is insoluble in water, in most organic solvents and in strong sulphuric acid. It hydrolyzes into simple products, as do the complex carbohydrates and proteins. Non-lignified plant parts contain more moisture, & are soft & break more easily. Lignin is considered an important source for the formation of soil humus, or humic matter. The high resistance of lignin to microbial decomposition is perhaps why it accumulates in soil. It is believed that, depending on the conditions, this could result in the formation of peat, which in time, can be converted into lignite, leonardite, coal and ultimately oil (fossil fuel) deposits. Humified organic matter: Humified organic matter or humic matter is a group of compounds that includes humic acids, fulvic acids, hymotomelanic acids, and humans. This humified soil organic fraction is also known as humus, or currently as humic compounds. Today humic compounds are defined as amorphous, colloidal poly dispersed substances with yellow to brown black color and relatively high molecular weights. According to Buckman & Brady, ‘ Humus is a complex and rather resistant mixture of brown to dark brown amorphous and colloidal organic substances modified from the original tissues or synthesized by the various soil organisms’ The term ‘humic acid’ originated with Berzelius in 1830, who classified the soil humic fraction into 1. humic acid, the fraction soluble in bases;

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2. Crenic and apocrenic acid, the fraction soluble in water; and 3. Humin, the insoluble and inert part. Humic acid was also referred to as humic acid, whereas humin was also called ulmin by Mulder in 1840. In 1912, Oden proposed the use of the name fulvic acid replacing the terms crenic and apocrenic acids. • Although humus is heterogeneous it possesses properties that distinguish it sharply from original parent tissues of anatomical structure of the material from which it was derived. Types of humic matter: Humic compounds or humic matter exists not only in soils, but also in streams, lakes, rivers, oceans and their sediments. They can also occur in lignite or leonardite, coal and other geologic deposits. These deposits are the sources for the production of commercial humates (Labortini et al. 1991; Burdick, 1965) that are used as soil amendments. Consequently, three categories of humic matter can be distinguished. a. Terrestrial or terrigenous humic matter: This is humic matter in soils that comprises mainly lignoprotein (lignin +protein) complexes humic and fulvic acids are major constituents. From the type of the lignin monomers, the group can perhaps be studied into: i. Softwood terrestrial humic matter, which is structurally characterized by coniferyl alcohol. ii. Hardwood terrestrial humic matter, which is composed of sinapyl alcohol monomers. iii. Grass or bamboo terrestrial humic matter, which consists of coumaryl alcohol monomers. b. Aquatic Humic matter: This is humic matter in streams, lakes and oceans, and their sediments and is composed mostly of fulvic acids. Humic acid is only a minor constituent. It consists of carbohydrateprotein complexes. The group can be subdivided into: i. Allochthonous aquatic humic matter:  This humic matter is brought from the outside in water.  The humic matter is formed in soils and other formation, is leached or eroded into rivers, lakes and oceans.

20  Although physical and chemical changes may be

induced by the aquatic environment. The nature of the humic matter is still related to soil humic matter, which consists of lignoprotein complexes. ii. Autochthonous aquatic humic matter:  This humic matter is formed from cellular constituents of indigenous aquatic organisms.  In marine sediments, this kind of humic matter consists of carbohydrate-protein complexes (Jackson, 1975; degrees & Mopper, 1975)  The source is organic debris from plankton, seaweed and kelp.  # Allochthonous- not produced in H2O, Autochthonous- produced in H2O, native humic substance. C. Geologic Humic matter: This is humic matter in lignite or leonardite and other geologic deposits. It is composed mostly of humic acids. Because of the organic process, most if not all, of the fulvic acids have apparently polymerized into humic acids. Fraction of organic matter: Soil organic matter may be subdivided into three components that are readily dispersed and separated from the mineral particles, and that which can only be dispersed by aquatic chemical treatment. A suggested subdivision is as follows: i. Micro organic matter: it is the fresh fraction of soil organic matter. This consists of the most recently added plant & animal residues/debris, which due to the entrapment of air has a density near 1 gm/cm³ and can be separated from the heavier mineral particles by floatation in water. ii. Light fraction: this consists of partially humified plant & faunal remains, which can comprise up to 25% of the total organic matter in grassland soils, separation from the mineral particles is achieved after ultrasonic vibration of the dry soil, to disrupt the soil aggregates, and floatation of the light fraction in a heavy organic solvent (density 2gcm-³) with the aid of a surfactant. iii. Humified fraction: it is the product of completely decomposed soil organic matter, adheres strongly to the

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mineral particles, particularly to the clay to form a clayhumus complex. This organic matter can not be separated by density floatation, and is incompletely dissolved in metal-chelating solvents, such as acetyl acetone. It is most effectively dispersed by strong alkalis, such as sodium hydroxide, which acts both by hydrolyzing and depohymerizing the large organic molecules that are tightly adsorbed on the mineral surface. The humified fraction or humin is subdivided further according to its solubility in alkaline & acidic solutions. Fractionation of soil Humus: Soil humus consists of two major types of compounds, unhumified substances and the humified remains of plant and animal tissues. The humified material, which represents the most active fraction of humus, consists of a series of highly acidic, yellow to black colored, high molecular weight polyelectrolyte referred to by such names as humic acid, fulvic acid, and so on. The fractionation of humic compounds is based on solubility in acids and alkalis. Humic substances are normally recovered from the soil by extraction with caustic alkali (usually 0.10.5N NaOH), although in recent years use has been made of mild reagents, such as neutral sodium pyrophosphate. After treating with NaOH solution two fractions are obtainedi. Black solution (humic substances) ii. Insoluble residues of humic matter (humin) + nonhumic matter. Then the black solution is treated with HCl/H2SO4 and two other fractions are obtainedi. fulvic acid (acid soluble) ii. Humic acid (acid insoluble) When this acid insoluble humic acid is treated with alcohol, soluble hymotomelanic acid and insoluble humic acid are obtained. So, based on solubility characteristics, the humic compounds can be separated into the following humic fractions (Flaig et al. 1975): Fractions Solubility in Alkali Acid Alcohol Fulvic acid Soluble Soluble Humic acid Soluble Insoluble Insoluble

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Hymotomelanic acid Humin

Soluble

Insoluble

Soluble

Insoluble

Insoluble

Insoluble

According to the German workers, humic acid can be further separated with natural salt solutions into – i. Brown humic acid (braunhuminsaure) soluble in NaOH not coagulated by an electrolyte and are characteristic of humic acids in peat and spodosol. ii. Gray humic acid (Grauhuminsaure) - insoluble in NaCl, is easily coagulated and is characteristic of humic acids in mollisols. In addition to the major fractions, several authors have also reported the isolation of a green humic acid fraction (Kumada and Hurst, 1967). The acid soluble fulvic acid also has sub-fractions as it is adjusted to pH 4.8, such as i. soluble fulvic acid ii. Insoluble s-humus A complete fractionation scheme is given in the following page……………………………………………….. i) Fulvic acid gives two subtractions at pH 4.8, namely fulvic acid (soluble) & s-humus (insoluble). j) It is more susceptible to microbial attack. k) According to some investigations, fulvic acids are humic acids capable of dissolving in water. l) Hydrolysis of fulvic acids with 5% H2SO4 showed the presence in fulvic acids of 20-25% reducing substances. m) Fulvic aid has the ability to enter into exchange reactions (the capacity for NH4 absorption was 318.6 m eq for fulvic acids from podzolic soil and 324 m eq per 100g substance fro those from chernozen. n) Fulvic acid are found to be rich in contain nitrogen o) Investigations by infra-red spectroscopy have shown that components of an aromatic nature are present in fulvic acids (Kasatochkin; Kobo & Tatsukawa; Schwitzer et al) p) X-ray analysis shows that the net of aromatic carbon in fulvic acids is very weakly expressed and side radicals predominate.

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r)

Due to the weakly expressed aromatic structure, the ratio C: H is generally lower in fulvic acids than in humic acids. The hypothetical model structure of fulvic acid (Buffle’s model) contains both aromatic and aliphatic structures, both extensively substituted with oxygen- containing functional groups:

Humic acid: a. The conception of ‘humic acids’ at the

present time is a group of substances having a form of structure although not completely identical with one another. In the last century, humic acids were regarded as products of the oxidation and dehydration of single substances, mainly carbohydrates. Further investigations have shown that several substances participate in the formation of humic acids. b. It is high molecular weight fraction of humic matter.

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c. It is soluble in alkali, but insoluble in acid and water. d. Humic acids are intermediate in resistance to degradation. e. It has low oxygen, but high carbon content. f. It possesses less acidic functional groups. g. When humic acid is treated with alcohol, it gives hymotomelanic acid (soluble) and insoluble humic acid, from which again brown humic acid (soluble) and gray humic acid (insoluble) are obtained, when treated with salt. h. Humic acids are dark brown to black in color. i. Electron microscope observations revealed the humic acids to different soils to have polymeric structure, appearing in form of rings, chains and clusters. The size of their micro molecules can range from 60-500Aº. j. Humic acids are thought to be complex aromatic macromolecules with amino acids, amino sugars, peptides, aliphatic compounds involved in linkages between the aromatic groups. k. The hypothetical structure for humic acid, shown in the figure, contains free and bound phenolic OH groups, quinine structures, nitrogen and oxygen as bridge units and COOH groups variously placed on aromatic rings.

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26 l. The elementary composition of humic

acid: at present time, comprehensive information is available on the elementary composition of the humic acid isolated from different soils. Some investigators draw attention to a similarity in values of this criterion in humic acid of different origin. Gillian (1940) found no essential differences between the elementary composition of humic acid isolated from forest soil, meadow soil and manure soil. However there is similarity in the elementary composition of humic acids isolated from different soils. m. Functional groups of humic acids: Oden, on the basis of his investigations showed that a molecule of humic acid from peat contains four carboxyl groups and he reflected this in the approximate formula C60H52O24(COOH)4 with a molecular weight of 1350. Fuchs & Stenge (1929) determined that four carboxyl and three phenolic groups were present in humic acid from brown coal. In humic acid there may also be present alcoholic (-OH) groups. Besides carboxyl, phenolic and alcoholic groups, humic acids also contain methoxyl groups (-OCH3) up to 1-2% in amount. A quinod group present in humic acids it may also be assumed that in soil humic acids, double-bond carbon groupings [-CH=CH-] occur. the presence of functional carboxyl and phenolic hydroxyl (OH) groups explains one extermely important property of humic aicds- their participation in exchange reactions. The participation of humic acids in exchange reactions depends on the

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conditions of the soil medium: the higher the soil reaction, the greater the number of functional groups of humic acids participating on the exchange reactions. n. the aromatic ring of humic acids: Hopppe-Seylar was the first to demonastrate the presence of aromatic ring in humic acids obtained from peat & brown coal. Shumuk (1924) was the first to show the presence of the aromatic ring in humic acid from chernoze. Together, with humic acids of aromatic nature also occur humic acids with masked aromatic rings or without aromatic rings. o. A theoratical scheme for the decomposition products of humic acids compiled by Flaig(1958), is given below:

p. Reducing substances in the composition

of soil humic acids: it is not yet clear that whether or not the humic acid molecule

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contains reducing substances, although this information would be of considerable interest fron the point of view of the nature & origin of humic acid. Subsequent works (Drogunor, 1948, 1950) confirmed the presence of reducing substances in soil humic acids, their amount depending on the method of purification of the solution obtained by hydrolysis of the humic acid. At present, we can’t say for certain whether reducing substances are a contaminent or part of the molecule of humic acid. q. The nitrogen-containing part of humic acids: at present time there is no doubt that soil humic acids contain nitrogen, the amount being approximatly 3.5-5%. The availability of humic acids to microorganisms and subsequently to plants depends on the form of nitrogen linkage in the humic acids. This important fact should be taken into consideration when estimating the nitrogen reserves of soils. r. The structure of the humic acid molecule: from the data at present available, it has been established that the humic acid molecule consists of an aromatic ring, nitrogen containing compounds in cycle froms and in the form of peripheral chains’ and possible reducing substances. The humic acid molecules has therefore, a complex structure. humic acids do not posseses a clear crystalline structure; this has given rise to ontradictory interpretations. humic acid molecules are not compact but have a loose (spongy) structure with a large number of internal spaces is of great importance in soil processes. These characteristics of the structure determine

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the water-holding capacity and the sorptive properties of humic acids to a considerable degree. *Elemental composition of humic substances & several plant materials: Substances % dry ash free basis C H O N Fulvic acid 44-49 3.5-5.0 44-49 2.0-4.0 Humic acid 52-62 3.0-5.5 30-33 3.5-5.0 Proteins 50-55 6.5-7.3 19-24 15.0-19.0 Lignins 62-69 5.0-6.5 26-33 3. Hymatomelanic acid: a. hymatomelanic acid is the alcohol soluble part of humic acid b. in Tyrin’s opinion, hymatomelanic acid represents a complex mixture of substances, which are humic acid derivatives, some being more oxidized and others more reduced. c. hymatomelanic acid, contain methoxyl, carboxyl and hydroxyl groups. d. they have a characteristically high carbon content (more than 60%). e. according to Kukharenko, hymatomelanic acids can be formed both by synthesis from the products of decomposition of organic residues and also during the oxydative-hydrolytic destruction of humus substances by oxygen and moisture. f. like huic acids, hymatomelanic acids are heterogenous. Consequently, it is considered that hymatomelanic acids are not an independent group of humus substances but an alcohol-soluble fraction of humic acids. Soluble in alkali; insoluble in acid & water. 4.Humins : a. Humin is insoluble in water, alcohol, acids and alkali. b. It is highest in molecular weight. c. Darkest in color. d. Most resistant to microbial attack. e. Humus substances not extracted from decalcified soil during treatment with alkali solutions are placed in the humin group. f. In the majority of soils. The humin group is represented mainly by humic acids; their loss of the capacity for dissolving

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in alkali solutions is due mainly to their stable linkage with the mineral part of the soil. g. In soils, the humin group may be partly represented by combined plant residues. h. Humins participate in the formation of a water-stable structure. I. Humin constitutes a concentration of considerable amounts of nitrogen, phosphorus, sulfur and other elements, forming a reserve which is gradually drawn into cycles of mineralization and mobilized for plant nutrition. j. it is soluble in hot alkali. Functional groups in humic substances: The functional groups present in humic substances are given below: O i. Carboxyl (acid group) --- R-C-OH R-C-Oˉ + H+ ii. Hydroxyl (phenol or alcohol group) R-OH R-Oˉ +H+ [Ar-OH Ar-Oˉ +H+] iii. Carbonyl (keto or ester group) R=O R-Oˉ iv. Amide (ammonium group) R-NH2 R-NH3+ or, R-NHˉ (depends on pH value) R-NH3(low pH), R-NHˉ (high pH) Properties of humus: Humus possesses certain properties that differentiate it from undecomposed plant and animal residues. Some of its important properties are as under: a. humus shows colloidal properties b. Humus (organic colloid) is a non crystalline organic substance. c. It consists of very large organic molecules, whose chemical composition varies considerably, but generally contains 40 to 60% C, 30 to 50% O, 0.3 to 7% H, and 1 to 5% N. d. The molecular weight of humic acids, a major type of colloidal humus, ranges from 10, 0000 to 100,000 g/mol.

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e. The surface area of humic colloids per unit mass is very high, generally exceeding that of silicate clays. f. The colloidal surfaces of humus are negatively charged as a result of H+ dissociation from carboxide (-COOH) or phenolic (-OH) groups. The extent of negative charge is pH dependent. Humus colloidal can also retain cations, but the negative sites always outnumber the positive ones, and a very large net negative charge is associated with humus. g. It is of dark brown to black color. The black color of surface soils is usually due to the presence of humin. h. It has a great water absorbing power; the addition of humus to a soil increases its water holding capacity. i. It possesses the power of addition and cohesion. Humus is much less adhesive because of this property that it acts as a cementing agent in crumb formation. In sandy soil it helps to bind sand particles as its adhesive capacity is greater than sand. On the other hand it makes clayey soils less sticky by reducing the coherence of clay particles. Hence, the maintenance of crumb structure is useful for both sandy and clayey soils. j. It has a high ion adsorbing capacity nearly four to six times that of clay. k. The adsorbed cations undergo Base Exchange as they do in the case of colloidal clay. Hence it has a high Base Exchange capacity. Because of the highly heterogeneous nature of the material, its cation exchange capacity (CEC) varies widely, from 30 to 350 meq percent. l. It is insoluble in water. m. It dissolves readily in dilute alkali giving a dark colored liquid. It is reprecipitated to a large extent when the alkaline solution is neutralized with acid. n. It behaves like a weak acid and forms salts with bases. With alkalis it forms soluble salts such as potassium humates, while with alkaline earths it forms insoluble salts. Humus is electronegative like clay acid. It is usually present in the soil in combination with bases mainly calcium forming calcium humate.

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o. It acts as a buffering agent. It also acts as an oxidation-reduction buffer. p. It serves as a source of energy and food for the development of various microorganisms. q. It is comparatively more resistant to microbial attack, yet it undergoes slow decomposition. Hence it does not remain in the soil indefinitely. r. It is highly dynamic as it is constantly formed from plant and animal residues and is continuously decomposed by microorganisms. s. Humus is an important source of nutrients for higher plants. t. Although humus is transitional and does not remain in the soil forever, it has a certain permanency and disappears from the soil only slowly. These properties go to show that humus has a number of functions to perform in the soil. Physically it modifies soil color, texture, structure and water holding capacity. It improves aeration and drainage by making the soil more porus. Chemically it serves as a source of food nutrients for the growing crops, increases the buffering power of soil, and influences the solubility of certain soil minerals on account of the continuous liberation of CO2. It also serves as a source of food for soil microorganisms. Humus thus occupies a very prominent place in the increase and maintenance of soil fertility. Structural chemistry of humic acids: Several hypotheses have been reported on the structural chemistry of humic acids, but apparently the hypothesis lack desirable uniformity and much disagreement still exists. Hypothesis of Schnizer and Khan (1972) The concept is based on information obtained from chemical degradation of fulvic acids. Degradation reactions do not ensure that artifacts may not have been produced. Depending on the severe ness or mildness of the reactions applied, any breakdown product can be obtained, ranging from elemental C, H, O, benzene rings to heterocyclic rings. Schmitzer & Khan were of the opinion that humic substances must be broken down into smaller sub units to study their structural chemistry, thus, four basic types of degradation procedures have been used:

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1. Oxidation with alkaline permanganate, nitric acid, H2O and CUO-NaOH mixture: The degradation products were invariably benzene carboxylic acids.

2. Reduction with Na amalgam or with Zn dust: Fulvic acid also yielded benzene derivatives with this method. 3. Hydrolysis with hot water, with acids or bases: Fulvic acid yielded benzene derivative such as hydroxybenzene and vanillic acids.

4. Biological degradation: It is a natural and mild process. This is a method by which fulvic acid is decomposed with the aid of microorganisms (e.g. penicillium sp, Aspergillus sp, and Trichoderma sp- the most common 3 fungi found in soil).

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The compounds produced from fulvic acid by biological degradation were also identified as benzene derivatives.

On the basis of the predominant findings of benzene derivatives, Schnitzer and Khan (1972) assume that fulvic acid is composed of phenolic and benzene carboxylic acids, joined together by H-bonds to form a polymeric structure. The latter contains many voids or openings in which other organic compounds, such as amino acids and carbohydrates can be trapped. Those could be present as impurities, but not as structural part. Hypothesis of Kononova (1961)(depending on synthetic pathway) 1. Kononova is of the opinion that at least three basic steps are involved in formation of humic acids: i. formation of structural units from the decomposition of plant tissues; ii. Condensation of three units; and iii. Polymerization of the condensation products. 2. The result is a multi component system, called humic or fulvic acids. They show similar structural patterns but may differ in details of structural and chemical composition. For example, fulvic acid

35

has a less condensed nucleus, but has more highly developed peripheral components. 3. In Kononova’s opinion fulvic acid can be both predecessor and the decomposition product of humic acid. 4. The basic structural units of humic compounds are considered to be phenolic or quinoid, bonded to nitrogen containing compounds and carbohydrates, the latter chiefly polyuronides. The inclusion of N-containing compounds and carbohydrates as fundamental units of the humic molecule is a matter of much controversy. [ Several investigators regard the later as accidental contaminants trapped in the maze work of the humic structure(Burges, 1960; Schnitzer & Khan, 1972) but others show evidence for the necessary participation of carbohydrates and Ncompounds in the formation of humic acids (Kononova, 1961; Flaig et al, 1975).] 5. Kononova (1961) suggests that the following reaction occurs for the inclusion of N in the humic molecule:

Such a combination produces a stable condensation product of phenols and α-amino acids and increases the stability of N in acid hydrolysis.

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Hypothesis of Kononova(1961): The role of microorganisms as sources of polyphenols has been emphasized by kononova in 1966. He gives a detailed account of research in which histological microscopic techniques and chemical methods were used to study the decomposition of plant residues. Stages leading to the formation of humic substances were postulated to be as follows: Stage-1: Fungi attack simple carbohydrates and parts of the protein and cellulose in the medullary rays, cambium and cortex of plant residues. [ I.e. the interval structure of plant tissue is attacked by fungi.] Stage2: Cellulose of the xylem is decomposed by aerobic myxobacteria. Polyphenols synthesized by the myxobacteria, are oxidized to quinines by polyphenoloxidase enzymes, and the quinines subsequently react with Ncompounds to form brown humic substances. Stage-3: Lignin is decomposed, phenols released during decay also serve as source materials for humus synthesis. The relative importance of lignins and microorganisms as sources of polyphenols for humus synthesis is unknown; this may depend upon environmental conditions in the soil. Because lignins are relatively resistant to microbial decomposition and a major plant constituent, they are sometimes considered to be the major, if not the primary, source of phenolic units. Some of the microscopic fungi that decompose lignin in soil, produce humic acid like substances in which the phenolic units originate from both lignin and through biosynthesis by fungi. Modern view of humus formation: It is generally accepted that humic acid and fulvic acids are formed by a multiple stage processes that includes, 1. Decomposition of all plant components including lignin, into simpler monomers; 2. Metabolism of the monomers with an accompanying increase in the soil biomass’ 3. Repeated recycling of the biomass C (and N) with synthesis of new cells; and 4. Concurrent polymerization of reactive monomers into high molecular weight polymers. (Flaig & Kononova, 1966).

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According to present day concepts, polyphenols derived from plants (e.g. lignin), or synthesized by microorganisms, are enzymetically converted to quinos, which undergo self-condensation or combine with amino compounds to form N-containing polymers. Flaig’s concept of humus formation is as follows (1966): 1. Lignin, freed to its linkage with cellulose during decomposition of plant residues, is subjected to oxidative splitting with the formation of primary structure units (derivatives of phenyl propane). 2. The side chains of the lignin building units are oxidized, demethylation occurs, and the resulting polyphenols are converted to quinols by polyphenolixidase enzymes. 3. Quinons arising from lignin (as well as from other sources) react with N-containing compounds to form dark-colored polymers.

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Flaig’s Hypothesis (1975): Flaig and his co-workers (Flaig et al, 1975) suggest lignin to be the source, or starting point, for the formation of humic acid and fulvic acids. Lignin is assumed to be broken down by degradation or decomposition reactions into its basic units (i.e. coniferyl alcohol or gluaiacyl propane monomers). The se lignin basic units are then subjected to oxidation, followed by demethylation to substituted polyphenols and further oxidation to quinine derivatives. Consideration of the quinone groups with amino acids and polysaccharides may then yield humic acid like substances. Lignin degradation products have been detected in hydrolysis of humic acids. Then some microbiologists said, i. Microbial cell wall materials add extra cellular gums which are predominantly polysaccharide like materials. ii. Products of microbial metabolism include both polysaccharides & polyphenols. Bacteria are known to be an important source of polysaccharides and some fungi of polyphenols. Martin and Haider (1971): According to Martin and Haider, microscopic fungi of the imperfecti group play a significant role in the synthesis of humic substances in soil. Their studies have shown that such fungi as Aspergillus, Epicoccum nigrium, Hendersonula toruloida, Stachybotyrs atra, and S. chartarum degrade lignin as well as cellulose or other organic constituents and in the process, synthesize appreciable amounts of humic acidic polymers. Phenolic units making up the polymer originated from lignin, as well as theory synthesis by the fungi. In recent studies, Martin & Haider investigated the synthesis of humic acid-type substances from several fungi belonging to the imperfecti group. The quantities of humic acid synthesized by fungi can be appreciable. Tan’s Hypothesis (1993) The different opinions, existing on the origin and structure of humic matter, indicate that phenolic groups and nitrogen compounds are the building blocks fro humic matter. The latter concept is reflected in the lignoprotein theory, which differs only lightly from hypothesis derived from oxidation and degradation research on humic compounds.

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Oxidation of humic acids yields phenols and a series of phenol-carboxylic acids, which from humic acids by condensation and polymerization. (Schnitzer & Khan, 1972) The general opinion is that humic acid can not be described in definite chemical terms, but current research data point otherwise. Results of 13C-NMR (nuclear magnetic resonance) analysis suggest the presence of a consistent structural composition including aliphatic, aromatic, and carboxyl compounds. Infrared spectroscopy also provides evidence for only one basic structure related to the presence of aliphatic C-H, carboxyl, and phenolic –OH functional groups: In addition, the elemental composition is fairly consistent with C content varying only from 40 to 50%, H from 4 to 5%, N from 1 to 5% and S from 0.2 to 0.3%. For these reasons, Tan proposes the following basic structures for the smallest possible molecule of terrestrial humic acid and aquatic humic acid.

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In the 1st figure the aromatic nucleus can vary depending on the type of the lignin monomers. The figure also shows the possibility to add a carbohydrate molecule to the aromatic nucleus. The latter (2nd figure) is of importance for the structure of fulvic acids, since fulvic acids contain less N and more carbohydrates, as

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noticed from both NMR and infrared analysis. A complex structure can also be built by combining several units of the basic molecule, or adding other compounds to the aromatic nucleus. So, the two main opinions of Tan are, 1. Carbohydrates are directly attached to the structure, and 2. There is no existence of phenolic group. Colloidal nature of humus: The colloidal characteristics of humus are as follows: 1. The humic compounds are hydrophobic in nature. 2. The tiny colloidal humus particles (micelles) are composed of carbon, hydrogen, and oxygen (probably in the form of polyphenols, polyquinones, polyurinoids and polysaccharides). 3. The surface area of humus colloids per unit mass is very high, generally exceeding that of silicate clays. The surface area of humus colloid ranges from 500-800 m²/g. 4. The colloidal humus must possess charge either positive or negative. Mostly negative charges are dominant under prevailing condition of pH. 5. At high pH values the cation exchange capacity (C.E.C) of humus on a mass basis (150-300 c mol c /Kg) for exceeds that of most silicate clays. 6. The water-holding capacity of humus is very high and it, on a mass basis is 4-5 times greater than that of the silicate clays. 7. Humus has a very favorable effect on aggregate formation and stability. 8. The black color of humus tends to distinguish it from most of the other colloidal constituents in soils. 9. Cation exchange reactions with humus are qualitatively to those occurring with silicate clays. 10.Humus colloid acts as a buffer. Function/significance of humus: 1. Store house of nutrients: Humus provides a store house for the exchangeable and available cations- K+, Ca²+, Mg²+. Ammonium fertilizers are also prevented from leaching loss because humus holds ammonium in an exchangeable available form.

42 2. Increases water holding capacity: Humic acid is reported to 3. 4. 5.

6.

7.

8.

9.

increase soil water holding capacity. C.E.C: Humic acid is reported to increase C.E.C. Reduce Al toxicity: Humic acid reduces micronutrient toxicity in acid soils, especially Al toxicity. Improving seed germination: Humic acid is capable of improving seed germination, root initiation and elongation, respiration, uptake of nutrients and development of green mass. Economical production of commercial humates: Humic acids, occurred as natural geologic deposits, are the main sources for the economical production of commercial humates used as soil amendments. Direct influence on plant growth: Humus is an important source of nutrients for higher plants. Under natural conditions, humus is probably the only source for the supply of nitrogen to higher plants. Humus also supplies phosphorus and sulfur required by growing plants. By virtue of its high adsorptive capacity it is able to adsorb and retain large quantities of various bases like Ca²+, Mg²+, etc. on its surface, which are also available for the nutrition of higher plants. Humus thus supplies both basic & acidic nutrient ions for the growth and development of higher plants. Influence on the aquatic environment: The presence of humic acids and Fulvic acids in lakes & stream may stimulate the growth of phytoplankton. Present in large concentrations, humic acids may reduce the photosynthesis of many aquatic green plants because of the dark brown color that they impart to the water. Due to its enormous chelation capacity on the other hand humic acid is capable in detoxifying lakes that are affected by metal pollution. In the form of cation, the reductive metals can be adsorbed and chelated by humic acid and rendered immobile. Influence on soil physical properties: Humus tends to give surface horizons dark brown to black colors. Germination and aggregate stability are encouraged. The humic fractions help reduce the plasticity, cohesion, and stickiness of clayey soil, making these soil easier to manipulate soil water retention is also improved.

43 10. Influence on soil chemical properties: Humus generally

account for 50 to 90% of the cation adsorbing power of mineral surface soils. Like clays, humus colloids hold nutrient cation in easily exchangeable form. Through its cation exchange capacity and acid and base functional groups, humins also provides much of the pH buffering capacity in soils. Amphoretic nature of humic substances: The compounds which are capable of acting either as a base or an acid are termed as amphoretic, e.g. amino acids which contain both a basic group (-NH2) and an acidic group (-COOH). The humic compounds in soil contain both a carboxyl and an amino group. The presence of these two functional group gives them the ability to exist as cation or anion. In other words, the compounds which have the ability to exist either as cation or anion are called the substance amphoretic in nature. For example, at pH 7.0, humic compounds act as Zwilter ion. NH3+ amino group H3C-C-COOˉ

carboxyl group

When alkali is added, the excess proton on the amino group is neutralized (when pH=9) NH3 NH2 H3-C-COOˉ+OHˉ

H3C-C-COOˉ+H2O

H H That is, humic compound is acting as anion when acid is added, the dissociated carboxyl group (COOˉ) accepts the proton (when pH=3). NH3 H3-C-COOˉ+H+ H

NH3+ H3C-C-COOH H The humic compound is now acting as cation.

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so, humic substances show amphoretic nature due to the presence of functional groups such as carboxyl group, R-COOH; hydroxyl group, R-OH; keto or ester group R=O and amide group. And amide group R-NH2 * What is E4/E6 ratio? What does it indicate about the nature of humic substances? E4:E6 is the optical property of soil humus. It is used to identify the nature of humic substances. E4:E6 ratio determines the degree of condensation of humic substances at various wavelengths. E4 is the value of photometer reading at 446 nm wavelength and E6 is at 665 nm the color ratio is E4/E6 or Q4/6 = absorbance at 465 nm absorbance at 665 nm The color ratio is then used as an index for the rate of highest absorption in the visible range. Some investigations reported that the intensity of light adsorption was characteristic for the type of molecular weight of humic substances. Humic acids with high molecular weights (MW>30,000) have lower E4/E6 value (4.32-4.45) than humic acids with lower molecular weights (MW=15,000) the lower molecular weight factors exhibit E4/E6 value of 5.47-5.49. Humic acid Fulvic acid 1. It is soluble in alkali, but insoluble 1. It is soluble in alkali, acid & water. in acid & water. 2. It is high molecular weight 2. It is low molecular weight fraction fraction of humic matter. of humic matter. 3. It is intermediate in resistance to 3. It is most susceptible to microbial degradation. attack. 4. Dark brown to black in color. 4. Straw yellow to wine red in color. 5. It has low oxygen but high carbon 5. It has high oxygen but low carbon content. content. 6. It possesses less acidic functional 6. It possesses more acidic functional groups (-COOH). group (-COOH). 7. When it is treated with alcohol, it 7. Fulvic acid gives two sub fractions gives soluble hymotomelanic acid at pH 4.8, namely fulvic acid and insoluble humic acid fraction (soluble) & s-humus (insoluble). which again gives brown humic acid

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(soluble) & gray humic acid (insoluble) as treated with neutral salt. 8. At present it can’t be said whether 8. Reducing substances are present in reducing substances are a fulvic acid. containment or part of the molecule of humic acid. 9. The ratio C: H is generally higher. 9. The ratio C: H is generally lower. 10. Oxygen occurs as structural 10. Oxygen is present in functional component of nucleus (e.g. In ether groups (-COOH, -OH, -CHO). or ester linkages). 11. The half-life may be 10-50 years. 11. Depending on the environment the half-life is generally measured in centuries. Terrestrial Humus 1. This is humic matter in soils. 2. It comprises mainly lignoprotein (lignin + protein) complexes. 3. Humic acids and fulvic acids are major constituents. 4. It can be subdivided into i. Softwood terrestrial humus ii. Hardwood terrestrial humus ii. Grass or bamboo terrestrial humus

Aquatic Humus 1. This is humic matter in stream, lakes & oceans and their sediments. 2. It comprises carbohydrate protein complexes. 3. It is composed mostly of Fulvic acid, Humic acid is only a minor constituents. 4. it can be subdivided into i. Alochthonus aquatic humus ii. Autochthonus aquatic humus

Organic matter Humus 1. Soil organic matter is any 1. Humus is a complex & rather substance of organic region that resistant mixture of brown to dark consists of undecomposed, partially brown amorphous & colloidal decomposed & completely substances modified from the decomposed plant & animal residues. original tissues or synthesized by the various soil organisms. 2. It is subjected to decomposition. 2. It is resistant to further decomposition. 3. It is a non-colloidal substance. 3. It is a colloidal substance. 4. It is a complex compound. 4. It is even more complex.

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5. Plant & animal residues are the sources of this. 6. All the portions are not amorphous & crystalline. 7. All the portions have no adsorptive power. 8. Does not possess any functional group. 9. Does not have C.E.C. 10. C/N ratio is not constant.

5. Humus comes from the organic matter. 6. Amorphous & not crystalline. 7. It has adsorptive power.

11. It has definite recognizable structure. It containsCellulose-20-50% Hemicellulose-10-28% Lignin-10-36%

8. Possesses several functional groups. 9.Has high C.E.C. 10. C/N ratio is more or less constant. 11. It has no definite recognizable structure. It containsCellulose-4% Hemicellulose-7% Lignin-45%

Peat 1. An organic soil that contains more than 50% organic matter & the organic matter is partially decayed or non-decayed, is called peat. 2. The kind of plant in the peat can be identified. 3. Peat soils are coarse/fine textured depending on the nature of deposited plant residues.

Muck 1. An organic soil that contain 2050% organic matter & the organic matter is completely decomposed, is called muck. 2. The plant materials of muck cannot be identified. 3. Muck soils are quite fine textured as the original plant materials are broken down.