WO2001038454A2

WO2001038454A2 - Oxidative polymerization of natural organic matter and co-polymerization of organic contaminants catalysed in particular by water soluble metal-porphyrins

Info

Publication number: WO2001038454A2
Application number: PCT/EP2000/011719
Authority: WO
Inventors: Alessandro Piccolo
Original assignee: Universita' Degli Studi Di Napoli 'federico Ii'
Priority date: 1999-11-26
Filing date: 2000-11-24
Publication date: 2001-05-31
Also published as: IT1314238B1; AU2837101A; ITMI992483A1; WO2001038454A3; ITMI992483A0

Abstract

A process is described of oxidative polymerization of natural organic matter and co-polymerization of organic contaminants into the matrix of organic matter catalysed by mimetic compounds of the heme group, in particular water soluble metal-porphyrins. The reaction is carried out by mixing together the organic matter with or without an organic contaminant, an oxidizing agent and the catalyst, preferably at room temperature for more than 2 hours.

Description

OXIDATIVE POLYMERIZATION OF NATURAL ORGANIC MATTER AND CO- POLYMERIZATION OF ORGANIC CONTAMINANTS CATALYSED IN PARTICULAR BY WATER SOLUBLE METAL-PORPHYRINS State of the art The present invention refers to a process of oxidative polymerization of natural organic matter and co-polymerization of organic contaminants catalysed in particular by water soluble metal-porphyrins. More particularly the process of the invention favours the co-polymerization of organic contaminants (chlorophenols, PCBs, dioxins, dibenzofurans, etc.) into the matrix of natural organic matter. Prior art

The natural organic matter (also known as humus or composition of humic substances) is a chemically heterogeneous material that derives from the biological degradation of vegetable and animal matter and from microbial catabolism. It is made up of cell synthesized biomolecules, namely proteins, peptides, carbohydrates, fatty acids and, among others, products of lignin degradation such as: phenolic and benzencarboxylic acids. The natural organic matter is diffused in the environment in various compartments. In fact, it is found, in different amount and composition, in soils, marine sediments, carboniferous deposits and in sea and inland waters. Its importance in the biosphere in maintaining the ecological equilibrium of earth is comparable to photosynthesis and fixation of atmospheric nitrogen. In fact, it is the constant mineralization process of the natural organic substance which chiefly sustains the biogeochemical carbon cycle, returning the CO₂ fixed by photosynthesis into the atmosphere. In addition, the binding capacity of the natural organic matter to the inorganic soil components allows the optimal distribution of micro- and macro- aggregates in soil and, therefore, the physical structural stability which supports vegetable growth and biological activity in soil.

Recent studies [Piccolo, A., Nardi, S. and G. Concheh. 1996. Micelle-like conformation of humic substances as revealed by size-exclusion chromatography. Chemosphere, 33, 595-602; Piccolo, A., Nardi, S. and G. Concheri. 1996. Macromolecular changes of soil humic substances induced by interactions with organic acids. European Journal of Soil Science 47,319-328; Piccolo, A. and P. Conte 2000. Molecular size of humic substances. Supramolecular associations versus macromolecular polymers. Advances in Environmental Research, 3, 508- 521] have shown that the natural organic matter, even though characterised by an apparent high molecular weight, is however composed of heterogeneous simple molecules, which instead of being covalently bound to each other in polymeric macromolecules, are combined in self-assembling supramolecular associations stabilized by only weak intermolecular bonds. The conformational stability of such associations is only temporary, as shown by studies using High Pressure Size Exclusion Chromatography (HPSEC) . In fact, the addition of just acetic acid (or other organic acids) to the humic solutions so as to lower the solution pH from approx. 7 to approx. 3.5 is sufficient to move the elution of humic moieties of from retention volumes typical of material of apparently high molecular dimensions to those which are instead characteristic of small molecular dimensions [Piccolo, A., Conte, P., and A. Cozzolino. 1999. Molecular association of dissolved humic substances as affected by interactions with mineral and monocarboxylic acids European Journal of Soil Science 50, 687-694]. The same effect is noted when the pH of the HPSEC mobile phase slightly varies (from 7 to 5.6) by adding HCI or acetic acid without however varying the ionic strength of the eluent [Conte, P. and Piccolo A. 1999. Conformational arrangement of dissolved humic substances. Influence of solution composition on the association of humic molecules. Environmental Science and Technology 33. 1682 1690]. The repeated occurrence of this shift of chromatographic bands from low to high elution times is an evidence of the self-assembled nature of the apparent high molecular dimension of the natural organic matter. Moreover, the actual chemical characterization of natural organc matter is extremely difficult since its chemical and physical-chemical nature is heterogeneous and very complex. At present, there are not simple methods and single spetroscopic techniques capable to produce a complete chemical structure of the natural organic matter. Classes of compounds that contribute in varying degrees to form the natural organic matter, according to the present invention, are for example: fatty acids, carbohydrates, peptides, and other natural substances including aromatic nuclei or condensed nuclei functionalized in various ways. Since organic matter is not made of high molecular-weight polymers, its components can then be subjected to polymerization (as described, e.g., in Saccomandi, F., Conte, P., and Piccolo A. 1998. Use of oxidase enzyme to increase polymerization of soil organic matter. Fresenius Environmental Bulletin 7, 537-543) through an oxidative coupling reaction catalyzed by a peroxidase enzyme. However, the use of this oxyreductive enzyme for the polymerization reaction is not efficient since enzymes are easily denaturated and rapidly lose catalytic activity. The enzyme high molecular-weight combined with the presence of weak interaction sites can also alter the humic conformation by dispersing the humic molecules and stereochemically inhibiting polymerization, which therefore loses effectiveness. Summary of the invention. It has now been found, and forms the object of the present invention, a process that promotes the oxidative polymerization of natural organic substances by formation of carbon-carbon or carbon-oxygen covalent bonds among the heterogeneous humic molecules, and in which the catalyst is a synthetic molecule that is mimetic of the heme group active in the enzymatic catalysts. In particular the catalyst consists of metal-porphyrins which are functionalized to become water soluble.

Another object of the invention is the use of compounds mimetic of the heme group that is the active catalytic site in enzymes, in particular metal-porphyrins, which are functionalized in order to become water soluble, land function as catalysts for the above-cited polymerization. A further object of the invention is the use of the polymeriation process of humic substances for: 1. ecological purposes, for both the physical stabilization of soil and the reduction of CO₂ emission from soil and, 2. environmental purposes, the decontamination of soils and waters from contaminating organic substances. Further objects of the invention will be evident from the following detailed description of the invention.

Detailed description of the invention. By heme group is meant the prosthetic group of haemoproteins (cytochromes, haemoglobin, etc.); it consists of a metal-porphyrin. By a compound mimetic of the heme group is meant a metal-porphyrin that provides a reversible bond between the metal and oxygen, as in haemoglobin and myoglobin. It is necessary, for the aims of the present invention, to prevent the formation of a subsequent metal- dioxygen complex, that is the irreversible formation of a μ-oxo complex, through the reaction of the bound oxygen with a second molecule of metal-porphyrin. While in haemoproteins the bonding of a second molecule of metal-porphyrin is prevented by the steric hindrance of a proteic moiety (histidine), the synthetic metal-porphyrins are protected by various functionalizations on the pyrrole rings (R.A. Sheldon. 1994. Metal-porphyrins in Catalytic Oxidation. Marcel Dekker, New York, pp. 1-27).

It is assumed that the polymerization, according to the invention, occurs through the oxidative coupling of free radicals formed in the organic matter by an oxidizing agent which may be, for instance, UV radiation, hydrogen peroxide, potassium monopersulphate or other oxidizing compounds and provides the highly reactive oxygen activated by a phenoloxidase type catalyst. Catalysts for such oxidative coupling leading to polymerization of the organic matter can be the peroxidases. These enzymes, however, present the disadvantages already mentioned above and limit the efficient polymerization. Conversely, it has been now experimentally verified that metal complexes which can act as the heme group in peroxidases, e.g. metal-porphyrins, can replace the aforesaid enzymes in the catalytic activity during the polymerization of humic substances and the said metal complexes do not present the same disadvantages as the enzymes. The metal complexes of this invention must be water soluble in order to be used in suitable applications. The metal may be iron, copper, manganese or cobalt.

According to the invention, either metal-porphyrins or all other metal complexes similar to metalporphyrins can be used advantageously. Other metal complexes can be tetrasulphonate-phenylphthalocyanines or dimers and trimers of the metal-porphyrins which are made water soluble by functionalizations with hydrophilic groups. In fact, if the reaction takes place in an aqueous medium or in the soil solution, in which organic matter is either soluble or suspended it is necessary that the catalyst be also water soluble. In the case of metal-porphyrins and relative dimers and trimers, functionalization with chlorobenzensulphonic, ammoniummethyl-benzene, peptidic or carbohydratic groups can be advantageous.

The functionalization of the catalyst is a process known by the experts of the art and may be carried out according to procedures which are described in the chemical literature. The polymerization of the humic matter, according to the invention, can advantageously occur at room temperature, but is efficient up to 80°C. It can be carried out in the pH interval between 3 and 14. The polymerization preferably occurs at the following ratios in water:

- tens of mg/ml of natural organic matter, e.g. 1 -50 mg.

- hundreds of μl of oxidizing agent (e.g. 12M H₂O₂; potassium monopersulphate, 50 μmole/ml), e.g. 100-300 μl for tens of nanomoles of catalyst, e.g. 60-100 nmoles.

The proportion between the reactants and the catalyst in solution reported above should be intended as just an indication, since it may vary with the molecular composition (never similar) of the different natural organic materials. Also the reaction ratedepends on the chemical characteristics of the substrate. Generally, a maximum yield of polymerization is reached after a period between 2 and 100 hours.

The operational method is simple and involves only the mixing the reactants together: substrate/catalyst/oxidizing agent. In particular, the organic matter to be polymerized is defined as substrate while the organic contaminant to be co- polymerized in the organic matter is the co-substrate. The natural organic matter, within the scope of the invention, may be defined as "native" (that which is naturally present in the environment, such as soil, sludge, compost, or water, to be treated) or "exogenous" (that which is added to the environment to be treated). The occurred polymerization in solution may be assessed using a HPSEC system eluting the mixture of reactants before and after the polymerization reaction on a column with a pH 7 phosphate buffer and noting the elution volume and peak intensity in the respective chromatographic peaks. The treatment of solutions, before and after polymerization, with acetic acid to pH 3.5 before injection into the HPSEC system clearly shows whether the peaks (the area under the chromatographic peaks) are due to simple supramolecular association (shifting of peaks to high retention volumes) or, instead, to real polymeric macromolecules (the peaks remain unchanged after treatment with acetic acid). Similarly, the occurred co-polymerization of organic contaminants into the organic matter matrix in solution can be followed by the same HPSEC method. The chromatographic peak of the contaminant progressively disappears with the progress of co- polymerization. Moreover, the new chromatographic peaks obtained at the end of the co-polymerization process are not changed by the acetic acid treatment thereby confirming the occurred polymerization. Spectroscopic methods such as IR and NMR can also show the occurrence of polymerization. In the case of infrared spectrometry (FTIR transmittance or DRIFT), the humic matter subjected to oxidative coupling catalysed by metal-porphyrins shows bands at approx. 1250 cm^"1 and 1 150 cm^'1 which are indicative of aryl and alkyl ethers, respectively, and are not visible in the original humic matter. Soild-sate NMR spectroscopy (CPMAS-13C-NMR) can also be used to assess the occurred polymerization and co-polymerization reactions by measuring the relaxation times of protons in the solid matrix (T→ H) which are always much larger (from two to ten times higher according to humic composition) than the T→ H in the non polymerized material. Polymerization occurs with the same proportion as the aforesaid reactants also in situ in soil, on the account of the native organic matter and/or an exogenous organic matter added in concentrations ranging between 40 to 80 mg/g of soil. The occurred polymerization is detected, among different methods, by the soil fractionation into different size particles (Stemmer, M., Gerzabek, M.H. and Kandeler, E. 1998. Soil organic matter and enzyme activity in particle size moieties of soils obtained after low-energy sonication. Soil Biology and Biochemistry, 24, 849-855), by measuring differences in soil structural stability (Kemper, WD and Roseπau, RC. 1986. Size distribution of aggregates. In: A: Klute (Editor), Methods of Soil Analysis, Part 1 , 2nd edition, ASA Monograph 9, Madison, Wl, pp.425-442), by measuring the organic carbon content in the different soil size particles, and by extracting the humic matter before and after polymerization. The process of polymerization, according to the invention, has considerable importance in the fields of environmental and ecological control, agrochemstry, geochemistry and mining for its ability to control, by increasing molecular weight, the general chemical reactivity of the natural organic matter and its resistance to biological degradation. The polymerization reaction according to the invention can be performed:

- for ecological purposes for: (i) the physical stabilization of soil, to control landslides and erosion and to limit the process of desertification; (ii) the resistance of organic carbon against biological transformation. Enhanced sequestration of organic carbon leads to a reduction of its mineralization and consequent CO₂ emission into the atmosphere.

- for environmental purposes to permanently detoxify contaminating organic compounds, namely PCB, dioxins, polyphenols, polychlorophenols, chlorinated aromatic hydrocarbons and relative compounds, which are incorporated (co- polymerized) into the organic matter during the polymerization process. In the case of chlorophenols, it has been observed a decrease in the concentration of free contaminant ranging from 50 to 90% depending on the number of substituted chlorine atoms.

Therefore, given the simplicity of the polymerization process in terms of operating conditions and resilience of catalysts, the reaction can be performed either in situ to decontaminate the soil or the contaminated soil is first washed with an exogenous humic matter to extract the organic contaminants, and then the mixture subjected to co-polymerization according to the invention. The co- polymerized mixture can be safely put back into the environment. For example, the metal-porphyrins can be used to catalyse the degradation of chlorophenols or other aromatic hydrocarbons during the purification of polluted waters treated as by standard procedures known to the experts of the field. If the purification processes do not reach a quantitative elimination of the pollutants, the addition of an exogenous organic matter (e.g. very aromatic humic substances such as those from an oxidised coal) allows the residual contaminants to be co- polymerized in the humic matrix, thereby making them insoluble and inert. Another application that envisages the use of humic substances is that with soils that are highly contaminated by chlorinated hydrocarbons. This application can include: 1 ) a direct treatment of the soils with the suitable amounts of exogenous organic substance (e.g. from oxidised coal) and polymerization mixture (oxidizing agent and catalyst), or, 2) extraction of the soil contaminants, which is carried out using methods known to experts depending on the type of contaminant to be extracted (e.g. by surfactant solutions but also by dissolved humic substances) followed by an ex situ treatment with a natural organic matter as well as with a oxidizing agent/catalyst mixture to co-polymerize the contaminants in the humic matrix. The co-polymers containing the contaminants which have been inactivated because of the chemical modification induced by the free radical reaction can then be transferred once again in the soil.

The catalyst of the polymerization reaction can also be used if immobilized on the surface of specific carriers. These can be either the very same exogenous humic matter to be added to soils or waters or any synthetic resin (known to the man skilled in the art) that may be able to preserve the activity of the catalyst for many cycles of reaction. In the case of resins, the activity of the catalyst may be modulated at will according to the type of linkages (weak dispersive bonds, covalent bonds, etc.) ensuring immobilization and to the functionalization of the catalyst. The present invention will be illustrated by the following non limiting examples. Examples

Examples of catalyst preparation are reported together with examples of oxidative polymerization performed on three different humic acids, catalysed both by soluble iron- as well as manganese-porphyrins which are functionalized with chlorosulphonic groups.

The reactants used were reagent-pure products provided by Sigma-Aldhch, Milan. The HPSEC system consists of a HPLC pump (Perkin Elmer LC-200) that elutes the mobile phase (a pH 7 phosphate buffer solution) at a flow of 0.6 ml.min^"1 on a TSK 3000 size exclusion column (Toso Haas). The chromatographic peaks are detected with a UV detector (Perkin Elmer LC-295) at 260 nm and processed using a Perkin Elmer Turbochrom integrated software. Example 1 (preparation of the catalysts)

The metal-porphyrins are generally prepared according to the following general pattern: 1 . H₂TDCPP:[meso-tetra-2,6-dichlorophenylporphyrin] The meso-tetra-2,6-dichlorophenylporphyrin is obtained according to a method reported in literature: Traylor P. S., Dolphin D., Traylor T. G., J. Chem. Soc. Chem. Commun.1984, 279-280.

1 .1 H₂TDCPP (SO₃Η⁺)₄: [meso-tetra- (2,6-dichloro-sulphonatephenyl) porphyrin]. In a 25 cc single neck flask 200 mg of porphyrin are dissolved in 8 ml of fuming H₂SO₄ and left under agitation at 160°C for 12 hours under Argon. Afterwards, the contents are very carefully poured into a flask containing H₂O and ice, rinsing the flask with the min. amount of H₂O. The solution, cooled with ice, is neutralized, taking great care, with a saturated NaOH solution. It is carefully dried by rotovapor at 60-70 °C, at 20 mmHg. The residue is recovered with CH₃OH and the salts remaining in the flask are filtered and washed with CH₃OH. All the filtrate is dried by rotovapor, and is collected with the minimum amount of CH₃OH and is precipitated with ethyl ether. It is filtered and the residue is redissolved with CH₃OH and is passed on a strong cationic exchange resin column, previously conditioned with a 10% HCI solution and then washed with H₂O up to pH 6-7 and eluted with H₂O; the moiety containing the compound is vacuum evaporated and recrystallized by CH₃OH/Acetone. Yield: 75%. UV/Vis in methanol: 417, 522, 602, 659

1.2. [MnTDCPP(SO₃Η⁺)₄]C [meso-tetra- (2,6-dichloro-3-sulphonate-phenyl) porphyrinate of manganese (III) chloride)]. In a 100 cc twin-neck flask 200 mg of the aforesaid porphyrin: [meso-tetra- (2,6- dichloro-sulphonatephenyl) porphyrin] are dissolved with 100 mg of MnCI₂.2 H₂O, in 100 ml of DMF. The solution is degassed for 10 minutes with Argon and then left still under Argon for 12 hours. The solvent is vacuum evaporated and the residue, collected with water, is subjected to ionic exchange chromatography according to the aforesaid methods for the free base. The moiety containing the desired product is evaporated and the residue is crystallized by CH₃OH/Acetone. Yield 70%. UVΛ/is in methanol: 379, 470, 571 , 653

1 .3. FeTDCPP (SO₃Η⁺)₄]C|-: [meso-tetra- (2,6-dichloro-3-sulphonate-phenyl) porphyrinate of iron (III) chloride]

In a 100 cc twin-neck flask, 200 mg of the aforesaid porphyrin: [meso-tetra- (2,6- dichloro-sulphonatephenyl) porphyrin] are dissolved with 100 mg of FeSO₄, in 100 ml of H₂O. The solution is degassed for 10 minutes with Argon and then left to reflux still under Argon for 12 hours. The solvent is vacuum evaporated and the residue, collected with water, is subjected to ionic exchange chromatography according to the aforesaid methods for the free base. The moiety containing the desired product is evaporated and the residue is crystallized by CH₃OH/Acetone. Yield 80%. UVΛ is in methanol: 422, 521 . 2. H₂TPy: [meso-tetra-pyhdylporphyrin]

The meso-tetra-pyridylporphyrin has been obtained as reported in literature Pasternack R. F., Huber P. R., Boyd P., Engasser G., Francesconi L., Gibbs E., Fa-sella P., Cerio Ventura G., de C. Hinds L., J. Am. Chem. Soc. 1972, 94, 451 1- 4517.

2.1. MnTPy: [meso-tetra-pyridylporphyrinate of manganese (III) chloride] In a 100 cc twin-neck flask, 150 mg of the aforesaid porphyrin [meso-tetra- pyridylporphyrin] are dissolved with 100 mg of Mn (AcO)₂.4 H₂O, in 20 ml of DMF. 1.5 ml of 2,4,6 collidine are added and the solution obtained is degassed for 10 minutes with Argon and then left to reflux still under Argon for 12 hours. The solvent is vacuum evaporated and the residue washed on a glass filter with water. The raw product thus obtained is vacuum dried and chromatography is performed on an activated neutral alumina column, eluting with CH₃OH/CH₂CI₂ (3/97). The moiety containing metal-porphyrin is evaporated and the residue is directly used for the permethylation reaction. Yield 70%. 2.2. MnTMPy (AcO)₅: [meso-tetra-N-methylpyridylporphyrinate of manganese penta-acetate]

The above described product is methylated in DMF at 45°C with methyl iodide for three hours. After evaporation of the solvent, the residue is purified on a weak anionic ionic exchange column, conditioned with a solution of 2M acetic acid. The final product is recrystallized by CH₃OH/Acetone. Yield 95%. UV/Vis in methanol 462, 560, 588.

3. H₂TPHTA(SO₃Η⁺)₄: [tetra- (2,6,9, 12-tetrasulphonatephenyl) phthalocyanine] In a 25 cc single neck flask 200 mg of free base phthalocyanine are dissolved in 8 ml of fuming H₂SO₄ and is left under stirring at 180°C for 12 hours under Argon. Afterwards the mixture is very carefully poured into a flask containing H₂O and ice, rinsing the flask with the min. amount of H₂O. The solution, cooled with ice, is neutralized, taking great care, with a saturated NaOH solution. It is carefully dried by rotovapor at 60-70°C, at 20 mmHg. The residue is collected with CH₃OH and the salts remaining in the flask are filtered and washed with CH₃OH. The solvent is vacuum evaporated and the residue is redissolved with water. The solution is passed on a strong cationic exchange resin column, previously conditioned with a 10% HCI solution and then washed with H₂O up to pH 6-7 and eluted with H₂O; the moiety containing the mixture is vacuum evaporated and recrystallized by CH₃OH/Acetone. Yield 80%. UV/Vis in methanol: 345, 375, 570, 620,690 (sh)

3.1. [MnTPHTA(SO₃Η⁺)₄]C [tetra- (2,6, 9, 12-tetrasulphonatephenyl) phthalo- cyaninate manganese (III) chloride] In a 100 cc twin-neck flask, 200 mg of phthalocyanine and 100 mg of MnCI₂.2 H₂0, are dissolved in 100 ml of DMF. The solution is degassed for 10 minutes with Argon and then left to reflux still under Argon for 12 hours. The solvent is vacuum evaporated and the residue, collected with water, is subjected to ionic exchange chromatography with the aforesaid methods for the free base. The moiety containing the desired product is evaporated and the residue crystallized by CH₃OH /Acetone. Yield 65%. UVΛ/is in methanol: 390, 5570, 590, 650. Example 2 (examples of polymerization) The humic substances were extracted from oxidised carbon (HA1 ), a lignite of North Dakota (HA2), and a volcanic soil (lake Vico) (HA3), purified and treated according to usual and published processes [Conte, P. and Piccolo A. 1999. Conformational arrangement of dissolved humic substances. Influence of solution composition on the association of humic molecules Environmental Science and Technology 33. 1682 1690].

2 ml of aqueous solution containing 2 mg of each humic acid are added with 2.2 ml of a 0.15 mg ml^"1 solution of iron- or manganese- porphyrin (prepared in Examples 1-1 .2 and 1 -1 .3) and with 48 I of an 8.6M H₂O₂ solution. The volume is adjusted to 10.2 ml with a pH 7 phosphate buffer solution. This solution is left to incubate for a maximum of 100 hours. An aliquot of this solution is directly injected (100 I injection loop) into the HPSEC system while another aliquot is first added with acetic acid up to pH 3.5 and then injected into the HPSEC system. The resulting chromatograms are compared with those of the same humic solution without the addition of the polymerising mixture, before and after treatment with acetic acid at pH 3.5. Results and discussion

Figures 1 and 2 (HA1 ), 3 and 4 (HA2), 5 and 6 (HA3) contain the chromatograms of the three humic acid solutions, before and after polymerization, catalysed with iron- and manganese-porphyhn, (curves A and C respectively). In all the chromatograms of the humic solutions treated with polymerising mixtures it can be seen that, with respect to the humic control solution (curve A), a considerable increase (a doubling in the case of humic acids from oxidised carbon, Figures 1 and 2, and from lignite, Figures 3 and 4) of the intensity of the first chromatographic peak corresponding to the high molecular peaks. When just the humic solution is treated with acetic acid before the HPSEC injection (curve B), the resultant chromatogram shows a notable reduction in the peak intensity at high molecular weight and the presence of various new peaks at higher retention times showing that splitting up into particles with smaller molecular dimensions occurred. Instead, the acetic acid treatment on the solution polymerized with porphyrin catalysis (curve D) still shows a very high first peak intensity at high molecular weight (still considerably higher than the corresponding peak in the chromatogram of the humic control solution) and a shifting of the subsequent peaks to shorter elution times that highlights an increase in the molecular dimensions of the smaller humic moieties. Both the increase in the intensity of the peaks as well as their shifting to shorter elution times provide evidence that an increase in molecular dimension occurred in the humic matter subjected to polymerization catalysed by the two porphyhns. The differences in HPSEC chromatograms between the various humic acids can be attributed to the different chemical composition of their organic matter. The experimental results highlight the following points.

The polymerization produced by the porphyrin catalysis is more extensive than that which is possible with the enzymatic catalysis previously reported by Saccomandi, F., Conte, P., and Piccolo A. 1998 (Use of oxidase enzyme to increase polymerization of soil organic matter. Fresenius Environmental Bulletin 7, 537-543). In fact, the peroxidase catalysis produced a polymerization product from the various humic solutions that showed both a lower increase in peak intensity at the high molecular-weight peak and a smaller shift to lower elution volumes than for the the metal-porphyrins. Therefore polymerization catalysed by metal-porphyrins appears more effective and capable of being more efficient in ecological and environmental applications.

Claims

1. A process for oxidative polymerization of natural organic matter characterised in that the said organic matter is subjected to polymerization in an aqueous environment, in the presence of a catalyst consisting of a molecule that mimicks the activity of the heme group in enzymatic catalysts, and is functionalized to be soluble in the said environment.

2. A process according to claim 1 in which the catalyst is selected among metal- porphyrins, metaltetrasulphonate-phenyl phthalocyanines, and dimers and trimers of the said porphyrins, functionalized with hydrophilic groups.

3. A process according to claim 2 in which the hydrophilic groups are selected among chlorobenzensulphonic, ammoniummethyl-benzene, peptidic or carbohydratic groups.

4. A process according to claims 2 and 3 in which the metal is selected among iron, copper, manganese, cobalt.

5. A process according to claims 1-4 in which the organic matter, the catalyst and an oxidizing agent are mixed together and polymerization is carried out at a temperature ranging from room temperature up to 80°C, at a pH range between 3 and 14 and a time range of 2-100 hours.

6. A process according to claims 1 -5 in which polymerization is carried out in water at the following ratios:

- tens of mg/ml of natural organic matter,

- hundreds of μl of oxidizing agent for tens of nanomoles of catalyst.

7. A process according to claim 6 in which the amount of organic matter is 1-50 mg/ml and the amounts of oxidizing agent and catalyst vary between 100-300 μl and 60-100 nmoles, respectively.

8. A process according to claim 7 in which the oxidizing agent is selected among UV radiation, H₂O₂ at a concentration of 12M and potassium monopersulphate, at a concentration of 50 μmoles/ml.

9. A process according to claims 1 -8 in which the catalyst is immobilized on a carrier that is used in the said polymerization process.

10. A process according to claim 9 in which the carrier on which the catalyst is immobilized is selected among exogenous humic matter and synthetic resin of different chemical composition.

1 1. A process according to claim 9 in which the catalyst is immobilized on the carrier according to different types of bonds selected among weak dispersive bonds, covalent bonds.

12. A process according to claims 1 -1 1 in which the occurred polymerization is assessed by infrared spectrometry, recording the presence of bands at approx. 1250 cm^"1 and 1150 cm^"1 related to ethereal aryl and alkyl groups, respectively, which are characteristic of the polymerization product.

13. A process according to claims 1 -1 1 in which the occurred polymerization is assessed by solid-state NMR spectrometry (CPMAS-13C-NMR), measuring the proton relaxation time (T, H) that result to be from two to ten times larger than the T, H in the non polymerized material.

14. A process according to claims 1 -1 1 in which polymerization is carried out in situ, in the soil to be treated.

15. A process for physical stabilization of soil characterised in that the native organic matter present in the said soil or exogenous organic matter added to the said soil is subjected to polymerization according to claims 1-1 1.

16. A process to reduce microbial mineralization of the organic matter present in soil characterised in that the native organic matter present in the said soil or the exogenous organic matter added to the said soil is subjected to polymerization according to claims 1 -1 1 .

17. A process for stabilization of organic carbon present in soil, characterised in that the native organic matter present in the said soil or the exogenous organic matter added to the same soil is subjected to polymerization according to claims 1 -1 1 .

18. A process to inactivate contaminating organic compounds present in soil characterised in that the native organic matter present in the said soil or the exogenous organic matter added to the same soil is co-polymerized with the said contaminants by the polymerization reaction according to claims 1 -1 1 .

19. A process according to claim 18 in which the contaminating organic compounds are selected among PCB, dioxins, polyphenols, polychlorophenols, chlorinated aromatic hydrocarbons and corresponding mixtures.

20. A process according to claims 15-19 in which the exogenous organic matter is in concentrations ranging between 40 to 80 mg/g of soil.

21 . A process for an ex situ purification of water or soil polluted by contaminating organic compounds characterised in that aliquots of exogenous organic matter are added to aliquots of the said water or the said soil previously subjected to a non-exhaustive purification from the said contaminating organic compounds, the exogenous organic matter being used to promote the co-polymerization of the said contaminants by the polymerization reaction according to claims 1 -11.

22. A process according to claim 21 in which the contaminating substances are selected among PCB, dioxins, polyphenols, polychlorophenols, chlorinated aromatic hydrocarbons and corresponding mixtures.

23. The use of compounds which mimick the activity of the heme group in enzymatic catalysts as catalysts for polymerization according to claims 1-19.

24. A use according to claim 23 in which the catalyst is selected among metal- porphyrins, metal-tetrasulphonate-phenyl phthalocyanine, and dimers and trimers of the said porphyrins, functionalized with hydrophilic groups.

25. A use according to claim 24 in which the hydrophilic groups are selected among chlorobenzensulphonic, ammoniummethyl-benzene, peptide or carbohydrate groups.

26. A use according to claims 24 and 25 in which the metal is selected among iron, copper, manganese, cobalt.

27. A use of humic substance polymerized according to the polymerization process of claims 1-10 to physically stabilize the soil, inhibit its microbial respiration and reduce CO₂ emission into the atmosphere.

28. A use of humic matter co-polymerized by the polymerization reaction according to claims 1-1 1 with polluting organic compounds contained in polluted waters and/or soils to decontaminate and detoxify the said waters and the said soils.

29. A use according to claim 28 in which the polluting organic compounds are selected among PCB, dioxins, polyphenols, polychlorophenols, chlorinated aromatic hydrocarbons and corresponding mixtures.