EP0934000A1

EP0934000A1 - Chemical system generating reactive oxygen species continuously and methods of using same

Info

Publication number: EP0934000A1
Application number: EP97943087A
Authority: EP
Inventors: Neil Goldsmith; Troels Koch; Henrik Oerum; Paul Paradis
Original assignee: Gyre Ltd
Current assignee: Gyre Ltd
Priority date: 1996-10-15
Filing date: 1997-10-14
Publication date: 1999-08-11
Also published as: AU4469497A; GB2318349A; GB9621505D0; WO1998016109A1

Abstract

A method whereby reactive oxygen species (ROS) are generated continuously in situ in an environment in that a chemical system is provided in the environment which in the presence of an oxygen source and a reducing agent is capable of continuously generating reactive oxygen species. The system comprises a redox active transition metal ion and a chelating compound capable of binding to the metal ion to form a chelate. Such a system generates ROS in amounts which control fouling, degrade organic matter and control viability of microorganisms and macroorganisms. The chemical system is active in free form or when associated with a surface.

Description

CHEMICAL SYSTEM GENERATING REACTIVE OXYGEN SPECIES CONTINUOUSLY AND METHODS OF USING SAME

FIELD OF INVENTION

This invention relates to the fields of preventing the fouling of surfaces by organic molecules and biological organisms and to impairing the viability of organisms or degrading organic matter present in aqueous or humid environments by providing is such environments a chemical system which is capable of continuously generating or releasing reactive oxygen species.

TECHNICAL BACKGROUND AND PRIOR ART

The fouling of surfaces by settling and accumulation of organic matter, micro-organisms and macro-organisms is a well known problem. Such fouling occurs normally through a multistage process of which the most important initial stages are deposi- tion of a layer of organic molecules on the surface and settling and growth of micro-organisms on the organic layer. On surfaces which are immersed more or less permanently in an aqueous environment such as e.g. a ship's hull, these initial stages may be followed by settling of macro-organism larvae on the micro-organism layer and subsequent growth of larvae into sessile macro-organisms (e.g barnacles) .

Fouling can in principle be prevented by interfering with any one of these stages.

A wide variety of compounds have been proposed and used to prevent such fouling in particular on ships, most notably tin and copper containing paints which are applied to the ship's hull. To date all commercially viable compounds are based on long lasting poisonous or toxic compounds which have detrimental environmental effects. A wide variety of compounds and approaches have been proposed which, according to their proponents, do not have such detrimental effects. These include non-stick coatings of the Teflon type, antibiotics and other approaches. In WO 92/07037 is disclosed an anti- fouling composition based on the activity of copper ions which has a reduced toxicity. This composition comprises an ion exchange resin capable of selectively binding copper such as copper ions present in aquatic environments so as to provide a constant toxic level of copper.

Most compounds and approaches that can be applied to preventing the fouling of ships can, subject to safety concerns, also be applied to prevent the fouling of other surfaces, most obviously other submerged marine structures, but also other non-marine water transport or containment systems. One particularly important application is in the water inlet and outlet systems of power stations, which frequently require utilisation of water in a cooling or power generation capacity and which, in particular due to the heating of the water that occurs, can suffer major problems of fouling and hence blocking of intakes and outlets.

Surfaces do not need to be submerged constantly, or indeed to be submerged at all to face a problem of fouling. Given a sufficiently humid or wet environment, micro-organisms will colonise many forms of surfaces such as roofs, wooden structures, plastics, the walls of buildings, clothing or other fabrics, medical devices, packaging materials, bandages and dressings, skin and even industrial machinery such as oil drilling equipment. In many such cases the presence of micro- organisms in layers presents a significant aesthetic problem and it may also cause damages to the materials. In other cases the micro-organisms can, either directly or indirectly (by facilitating the settling of other organisms) cause a medical or structural problem to develop. Biocidally active compounds (biocides) are also presently used to reduce the presence, or prevent the build up, of undesired micro-organisms in water bodies or aquatic environments themselves. They can be either periodically applied directly to the body of water, or alternatively be introduced into the environment via diffusion from a surface which is in contact with the water body. The biocide is being used to partially or fully sterilise or disinfect the water body. Applications of such systems most obviously include the sterilisation or disin- fection of drinking water and the treatment of household effluent, sewage or industrial waste both in holding tanks and in the sewer system itself. They are also widely used in building air conditioning and ventilation systems (where the build up of such organisms as Legionella spp can be a health problem for the occupants) . They can further be used to prevent the contamination of aquatic environments such as processing tanks, filters and as a way of controlling residual or subsequent contamination of foodstuffs and drinks.

Active or reactive oxygen species (ROS) are highly reactive at normal temperatures. In particular they react with various component molecules of living organisms. This makes ROS potentially useful bactericidal and sterilising agents.

In addition, reactive oxygen species such as hydrogen peroxide are used to cause modification or degradation of organic matter molecules including bleaching of coloured or pigmented matters. However, currently used methods for this purpose are also based on feeding of the reactive oxygen species to the environment contaminated with organic matter.

The use of active oxygen species as antifouling or disinfecting agents in association with marine and other aquatic environments and wet surfaces in general is known e.g. from WO 93/02973 and JP 89-117227. JP 83-28345 discloses the use of a peroxide of a group Ila metal in a marine anti-foulant for application to a ship's hull to prevent adhesion of marine organisms .

Whereas known methods of controlling the association of submerged aqueous, wet (humid) or intermittently wet surfaces with accumulating organic matter, micro-organisms and/or macro- organisms, or the viability of such organisms or accumulation of organic matter in aqueous, humid or intermittently humid environments are either based on the use of toxic compounds or on the use of active oxygen species which has to be fed con- tinuously or intermittently to the surfaces or the environments, the present method is based on the use of an in- si tu chemical system comprising a redox active transition metal ion firmly associated with a chelating agent to form a chelate, the system being capable of continuously generating reactive oxygen species in amounts which are effective in preventing fouling and in controlling biological growth and degradation of organic matter.

SUMMARY OF THE INVENTION

Accordingly, the invention relates in a first aspect to a method of continuously generating reactive oxygen species (ROS) in an environment, the method comprising providing in said environment a chemical system which in the presence of an oxygen source and a reducing agent is capable of continuously generating reactive oxygen species, the system comprising (i) a redox active transition metal ion and (ii) a chelating compound capable of binding to the metal ion to form a chelate, the metal ion thereby being so firmly associated with the chelating compound that it essentially does not dissociate from the chelate into the environment.

An important aspect of the invention is to provide the means for controlling fouling of surfaces by organic matter including living micro- and macro-organisms . It is therefore an important objective of the invention that it provides a method of controlling in an aqueous, humid or intermittently humid environment the accumulation of organic matter on a surface delimiting or being in contact with the environment, the method comprising associating with said surface a chemical system as defined above which in said environment is capable of continuously generating reactive oxygen species in an amount which prevents or reduces accumulation of organic matter on said surface.

In a further aspect of the invention there is provided a method of impairing the viability of organisms or of degrading organic matter present in an aqueous, humid or intermittently humid environment, the method comprising providing in the environment a chemical system as defined above which is capable of continuously generating in said environment reactive oxygen species in an amount which impairs viability of the organisms and/or degrades the organic matter.

The invention provides in a still further aspect a composition comprising a chemical system which in the presence of an oxygen source and a reducing agent is capable of continuously generating reactive oxygen species in an aqueous, humid or intermittently humid environment, the system comprising (i) a redox active transition metal ion and (ii) a chelating compound capable of binding to the metal ion to form a chelating com- pound/metal ion complex (a chelate) , the metal ion thereby being so firmly associated with the chelating compound that it essentially does not dissociate from the chelate when it is present. Such a composition is capable of continuously generating reactive oxygen species in an amount which impairs the viability of organisms present in the environment or degrades organic matter in the environment . DETAILED DISCLOSURE OF THE INVENTION

The present invention relates, as it is mentioned above, in a first aspect to a method of continuously generating reactive oxygen species (ROS) in an environment containing water. In accordance with the invention, the in-situ reactive oxygen species generating system comprises at least two elements, i.e. (i) a redox active transition metal ion and (ii) a chelating compound which is capable of binding to the metal ion whereby a chelate is formed. Preferably, this binding is so firm that essentially no dissociation of free metal ions into the environment occurs.

In this context, the expression "transition metal" is used in its conventional meaning of elements in the ten columns near the centre of the Periodic Table of Elements which is generally referred to as the d-block. A general definition of transition metals would be: "those elements at least one of whose simple ions has an incomplete outer shell of d-electrons (that is contains between one and nine electrons) from Chemistry Facts, Patterns and Principles (Revised Edition), Kneen et al., Longman, London, 1987. Thus, the group of transition metals include the following atomic numbers: 21-30, 39-48, 57-80 and 89-107.

A range of different chelating compounds can be used in the present invention for forming chelates (also referred to as complexes) of transition metals. The term "chelating compound" may also be referred to as a "ligand" or a "chelator". Without being bound by examples these include the group of chelating compounds listed below:

EDTA: Ethylenediamine-tetraacetic acid or (ethylenedinitril) tetraacetic Acid,

ECTA: [N- (2-Ethylnitrilodiacetic Acid) -1, 2-Cyclohexylene dini- trilo] Triacetic Acid, EHPG: N,N' -Ethylene bis [2 - (2-hydroxy-5-chlorophenyl) -

Glycine] ,

HOEDTA: N- (Caroxymethyl) -N' - (2-hydroxyethyl) -N,N' Ethylene

Diglycine, CHCEDG: N- (Carboxymethyl) -N' -2-Hydroxycyclohexyl-N,N' -Ethylene-

Diglycine,

CH3EHPG: N,N' Ethylene bis [2- (2 -Hydroxy-5-methylphenyl)

Glycine] ,

CDTA: (1,2-Cyclohexylenedinitrilo) Tetraacetic Acid, EHCHG: N,N, Ethylene bis (N-2-Hydroxycyclohexylglycine) ,

NTA: Nitrilotriacetic Acid,

HEIDA: [ (2 -Hydroxyethyl ) Imino] Diacetic Acid,

DTPA: [(Carboxymethyl) Imino bis (ethylenenitrilo) ] Tetraacetic

Acid, EHPG: N,N' Ethylene bis [2- (o-Hydroxypheny1) Glycine],

DHEG: N,N'-bis (2 -hydroxyethyl) Glycine,

AP: Aminophosphonate,

IDA: Iminodiacetic acid,

EDAMC: Ethylenediamine-bis-methylenecarboxylate

The usefulness of the above-mentioned chelating compounds can be further increased by using either a combination of compounds or by derivatising the compounds. In the present context, derivatisation serves at least one of three important purposes : (i) to anchor the chelate to a surface, (ii) to enhance affin- ity/specificity of the ion chelation and (iii) to modulate surface polarity.

With respect to the above purpose (i) , the fixation of the chelate to the surface is essential to the in-situ/surface production of reactive oxygen species . One way to achieve this is by introducing substituents with long lipophilic chains in the chelator hence facilitating binding or anchoring of the chelator to a surface. One example of a suitable substituent is a substituent comprising an anthraquinone group. The substitution can be done via acid derivatives or by direct substi- tution on the carbon skeleton of the chelator. These chains may (or may not) have functional groups which e.g. can be functional groups participating in a hardening of a coating comprising the system according to the invention. Another way of fixating the chelating agent to a surface is by attaching the chelate to the surface either by chemical and/or physical activation of the surface followed by attachment of the chelate. Still another way is by direct attachment of the chelate to the unmodified surface.

In sea water the ion concentration of metals present herein is subject to considerable variations. The ions found in the highest concentration are Mg²⁺ and Ca²⁺. These metal ions are not active in the catalytic production of reactive oxygen species but they will bind to the anionic chelators described above although ions of transition metals such as Fe, Co, Ni, Cu or Zn which are redox active will bind preferentially. However, this "contamination" by Mg and Ca which is mainly caused by electrostatic interaction can be avoided by a derivatisation with a non-charged or neutral substituent such as e.g. hydroxylamine . It has been shown that non-charged hydroxylamine derivatives of e.g. EDTA and NTA chelate transition metal ions specifically and in fact co-ordinate e.g. Fe(II) even more strongly (Hirotsu et al . , 1985) (Scheme 1).

SCHEME 1

H₂NOR'

R COOH - FT ONHOR'

Another effect of having neutral chelates on the surface is that they enhance the lipophilicity of the surface. This feature has been shown to be an important property in anti- fouling (Wahl , 1989 ) .

In conclusion substituents on the basic chelator structures can be used for the following purposes:

1. Fixating the chelates to the surface,

2. Increasing the affinity of redox active transition metals to the chelator and the discrimination between redox inactive/redox active metals, or

3. Decreasing the polarity of the surface.

The terms R, R' as used in the above Scheme 1 and the below Scheme 2 designate different substituents. In the present context, the term "substituent" is used in its broadest sense and it encompasses most organic structures with or without functional groups. Most preferred are structures which comprise alkyl, alkenyl, alkynyl, cycloalkyl or aryl .

A number of different chelating compounds of variable complexity (and consequently varied activity) can be envisaged as it is illustrated in Scheme 2.

SCHEME 2

4. 5.

However, for the system to be effective in a given environment it is required that an oxygen source and a reducing agent is present in such environment. In accordance with the invention, suitable oxygen sources may be present in an aqueous or humid environment as free oxygen or as oxygen bound to organic or inorganic moieties. Alternatively, an organic or inorganic oxygen source may be added to the environment or incorporated into the chemical system according to the invention prior to its introduction into the environment.

Suitable reducing agents will normally be abundantly present in the aqueous or wet environment, but such agents may also, if required, be added to the system or the environment.

Chemical generation of reactive oxygen species is a stepwise electron transfer from the metal ion to molecular oxygen. Subsequent proton transfer forms hydrogen peroxide and/or hydroxyl radicals. The general reaction scheme is shown in Scheme 3 below: SCHEME 3

o- ₂- H₂0₂ OH H₂0

2H⁺ H⁺

Super- Hydrogen- Hydroxyl- vVater oxide peroxide radical

All three oxygen species are highly oxidative and have higher oxidation potentials than e.g. bromine and chlorine and as such they can be regarded as potent disinfectants. This electron transfer sequence can be catalyzed by different transition metal ions including as examples: Fe, Cu, Zn, Cd, Pb, Co and Ru ions.

The redox mechanism can be exemplified by the Fe (II) /Fe (III) system. Fe(II) is capable of transferring an electron to molecular oxygen giving rise to superoxide, with itself being oxidised to Fe(III). Fe(III) is easy to reduce and can be reduced by most reducing agents. Thus it has been shown that the reduction potential of EDTA-Fe (III) to EDTA-Fe (II) is very low (E°= 0.117 V) . This means that even very weak reducing agents such as the widely found agent hydrogen sulphide and 3- mercaptopropionic acid and DTE are capable of reducing EDTA- Fe(III) . Other reducing agents such as NADH and ascorbate are also able to carry out this reaction. Such compounds are found as components in heterogeneous biological aqueous environments. It has been shown that chelation of Fe(III) (e.g. by NTA, EDTA) makes the reduction of Fe(III) even easier. Superoxide and hydrogen peroxide may also be reduced and thus 3 electrons are required to form hydroxyl radicals which are the end product.

The reaction leading to the generation of reactive oxygen species is illustrated in the below Scheme 4 where M represents the metal ion and n its charge or oxidation state: SCHEME 4

M" ► M^{n+1 n} ► ⁿ⁺¹ Mⁿ >~ M^n+:

Red. Agent ^τ ' Red. Agent Red. Agent

In accordance with the invention, the key issue in the above chemical system is its continuous or cyclic nature whereby, in the presence of an oxygen source and a reducing agent, it will result in a steady flow of reactive oxygen species. As it is illustrated in Figure 1, the reaction is maintained by the continuous cycling of the metal ion complex, i.e. the chelate, between its reactive and its inert forms. The so-called inert complex is returned to its reactive state through reaction with a reducing agent. Each oxidation of the metal ion complex releases another electron to be transferred to oxygen, superoxide or peroxide, thus generating further ROS.

However, the generation of reactive oxygen species by the system is not absolutely dependent on the presence of an external reducing agent, since both superoxide and peroxide have an inherent capacity to act as reducing agents .

It has now been found that the above chemical system is capable of continuously generating reactive oxygen species in amounts which are sufficient to provide in an environment comprising an aqueous phase the means of preventing fouling of a surface by organic materials, micro-organisms and macro-organisms or of impairing the viability of organisms or degrading organic matter in such an environment . It is to be understood that when there is referred to "an environment comprising an aqueous phase" this implies any environment where there are sufficient amounts of water for the ROS generating system to become active. This will include moist, wet, humid or intermittently humid or wet environments and liquid containing only minor amounts or traces of an aqueous phase such as e.g. a cutting oil.

Thus, in an important aspect of the invention there is provided a method of preventing fouling. As used herein, the term "fouling" refers to the phenomenon of organic matter and living organisms being settled or accumulated on a solid surface. Such a surface may be the outer surface of constructions which are permanently submerged into a water body such as the outside of a ship's hull or a oil drilling platform construction. The surface may also be an outer surface of a construction or a construction element located in the open air or the inner surface delimiting a cavity such as e.g. the inner surfaces of constructions for containing or transport- ing a liquid including water or aqueous media.

In accordance with the invention, the method of controlling in an aqueous, humid or intermittently humid environment, the accumulation of organic matter including micro-organisms and macro-organisms on a surface, i.e. fouling of that surface, comprises that a chemical system as described above is associated with the surface.

In the present context, the expression "associated with a surface" indicates that the ROS generating system is either incorporated into a coating or surface layer covering the surface to be protected from fouling or that the system, i.e. the chelate is bound or linked to the surface itself.

The linking or binding of the metal ions to the surface is typically made by complexing the ions to metal chelating com- pounds to form chelates followed by binding or anchoring (fixating) these chelates to the surface. This binding or anchoring of chelates is advantageously carried out via substituents which are linked to the chelating compounds or chelators. One example of such a useful substituent is an anthraquinone group which can be firmly attached to a solid surface by a photochemical reaction.

Alternatively, the ROS generating system can be associated with the surface by incorporating it into an anti-fouling compo- sition or coating such as e.g. an anti-fouling paint comprising polymeric particles. In such an embodiment, the chemical system can be linked or anchored to the particles either during the polymerisation process leading to the paint or coating or the chemical system can attached to the polymeric particles inde- pendently of the polymerisation process.

It will be understood that the expression "associated with a surface" also refers to the situation where the object of the method according to the invention is not only to prevent fouling of a surface itself but also to impair the viability of organisms or to degrade organic matter in an aqueous or wet environment in the proximity to the surface. As used herein, the expression "in the proximity to the surface" is used to describe the distance from the surface over which the concentration of the generated reactive oxygen species is sufficient- ly high to impair the viability of undesired organisms present in the environment. It is contemplated that this distance, depending on the particular system applied, will be in the range of 0-100 cm such as in the range of 0-50 cm, including the range of 0-25 cm such as in the range of 0-10 cm.

The above method of controlling surface accumulation of organic matter and organisms is useful for controlling fouling of any submerged or wet surface, and the use of the method is contemplated for a variety of surfaces including as examples a ship's hull, a water storage reservoir, a waste water reservoir, the inner surface of a water pipe or tubing for transportation of liquid media, a liquor container, a fish net, an anchor, a rope, a roof, a fence, a boat, a submarine, a water inlet or outlet of a power station and other industrial plants, an aqueduct, a canalised waterway, a submerged marine structure, an industrial processing tank, an air conditioning system, a filter, a medical bandage or plaster, a packaging foil, a film, a plastic, paper for storage of foods and fluids, a medical device, a pharmaceutical, a household or industrial appliance and paper goods .

In another aspect, the present invention also relates to impairment of the viability of organisms or degradation of organic matter in an environment providing conditions under which the above chemical system is active including aqueous, humid or intermittently humid environments . In the present context, the expression "impairment of viability" refers to an effect whereby a substantial proportion of living organisms including micro-organisms such as bacteria that are present in the aqueous or humid environment are killed or inhibited with respect to growth or other manifestations of viability. Such an effect may also be referred to as a biostatic or biocidal effect.

During the experimentation leading to the invention it was found that a transition metal ion chelating compound in itself when added to an environment containing viable organisms may have a significant biocidal or biostatic effect. It is assumed that this effect is due to the metal scavenging effect of the chelator which results is a depletion of metal ions the presence of which are essential for the viability of the organisms. The effect of the chemical system as described herein may therefore be enhanced by adding a further amount of chelating compound to the environment . The present method of impairing the viability of organisms or of degrading organic matter is based on that a chemical system as defined herein is provided in the environment in an amount which is capable of continuously generating in said environment reactive oxygen species in an amount which impairs viability of the organisms and/or degrades the organic matter.

The types of organisms the growth of which can be controlled by the ROS-generating system include micro-organisms such as e.g. gram-positive and gram-negative bacteria, yeast, fungi, proto- zoa, algae, and macro-organisms including larval stages of animals having natural aqueous environments such as marine environments, rivers, lakes and ponds as their habitats. Depending on the type of organism and the concentration of ROS being formed in the particular environment, the effect the generation of reactive oxygen species on the organisms may a killing, i.e. a biocidal effect or the effect may be an inhibitory effect, i.e. a biostatic effect on the growth or other life manifestations of the organisms.

It is envisaged that in particular environment, the effect of the ROS may be biocidal for some organisms present and biostatic for other organisms.

A particularly interesting aspect of the invention is that reactive oxygen species can be generated in amounts which results in degradation of organic matter other than viable organisms . Natural water bodies and other aqueous environments such as e.g. water purification and water supply systems and sewage treatment facilities contain varying amounts of organic matter and in many instances is it desirable to obtain a reduc- tion of the organic matter content. It has been found that the chemical system when applied in accordance with the invention is capable of generating ROS which results in degradation of organic matter. One example of this effect is the bleaching of coloured organic matter. When the chemical system is provided in the form of particles carrying the system, such particles are typically of a size in the range of 0.1 to 1000 μm such as 0.5 to 500 μm including 1 to 100 μm.

In this context, the chemical system may be provided in the environment in the same manner as it is described above, i.e. it can be added directly to a water body either in the form of chelates (complexed transition metal ions) or in the form of particles to which the chelates are bound. Additionally, the chemical system may be provided by associating it with a surface delimiting or being in contact with the environment. Thus, the chemical system can e.g. be associated with a surface which is a surface delimiting a drinking water storage reservoir, a waste water reservoir or a swimming pool, the inner surface of a pipe or tubing for transportation of a liquid medium, a surface delimiting a liquor container, a water inlet or outlet of a power station and other industrial plants, an aqueduct, a canalised waterway, an industrial processing tank or the surface can be in an air conditioning system, a filter, a packaging foil, a film, a plastic, paper for storage of foods and fluids, a medical device, a pharmaceutical, a household or industrial appliance and paper goods.

The control of microbial growth and the accumulation of organic matter is of great concern in water supply systems and in the operation of private and public swimming pools. Presently, the control of the water quality is such systems is made by mechanical removal of organic matter e.g. by filtration and continuous feeding to the system of biocides such as chlorine. It is envisaged that the invention will provide alternative means of such control measures or that the system according to the invention can be used as a supplementary measure to control water quality. Thus, the chemical system can be associated with the filtering materials and/or it can be associated with surfaces delimiting water supply element or swimming pools. It is envisaged that the chemical system can be provided conveniently in a water environment in the form of particles coated with the system which are contained in a material permitting water to come into contact with the particles. Such contained particles can be inserted e.g. in a water supply system including a household water heating system to control microbial growth.

In interesting aspects of the invention there is also provided herein compositions which are useful in the above methods and which accordingly have the composition and features as described above. Although it is preferred that the composition does not include environmentally detrimental compounds, it is contemplated that the compositions may, for certain uses, advantageously include a biocidally active compound.

As it is mentioned above, such a composition may be in the form of particles carrying the chemical system. In accordance with the invention, the composition may also be a paint or coating composition such as an anti-fouling paint for building constructions or surfaces submerged in sea water, including a paint for ships or other vessels including an oil drilling platform.

The invention is further illustrated in the following non- limiting examples and the drawing wherein

Fig. 1 is a diagram illustrating the processes involved in the continuous generation of reactive oxygen species;

Fig. 2 illustrates the effect of Co (II) and Co(II)-NTA on Murexide concentration. (O) represents Murexide in the presence of Co (II) and (■) Murexide in the presence of Co(II)-NTA. The conditions were: [Co(II)] lxlO^"4 mol dm^"3, [NTA] l.SxlO^"4 mol dm^"3, [Murexide] 3xl0^"5 mol dm^"3. Reaction was carried out in distilled water at room temperature; Fig. 3 illustrates the effect of Fe(III) and Cu(II)-EDAMC coated chromatography beads on Murexide concentration. Hydrogen peroxide was added after 60 minutes (Cu (II) -EDAMC) and 75 minutes (FE (III) -EDAMC) , respectively. Conditions were: 0.5 g Fe(III) and Cu (II) -EDAMC were suspended in 50 ml distilled water. [Murexide] was 5.25 and 8.0xl0^"5 mol dm^"3, respectively, [H₂0₂] 7.0xl0^"7 mol dm^"3 was added to the reaction mixture. Reaction was followed at room temperature;

Fig. 4 illustrates multiple substrate bleaching in the presence of additional Cu(II) and Cu(II)-IDA coated chromatography beads. (•) represents Murexide, (O) amaranth and (A) methyl orange. Conditions were: [Murexide] 5.6xl0^"5 mol dm^"3, [Amaranth] 5, [Methyl orange] 1.5, additional [Cu(II)] 5xl0^"5 mol dm^"3 and 0.5 g of Cu(II)-IDA was suspended in distilled water. The reaction was followed at room temperature;

Fig 5 shows surface coupled metal ion-complexes. Structure (1) is a surface coupled metal ion-IDA complex with an associated water molecule and structure (2) is a surface coupled metal ion-EDAMC complex; and

Fig. 6 illustrates the effect of complexed and non-complexed aminophosphonate coated chromatography beads on E. coli growth, where (♦) represents a control sample of E. coli in nutrient broth solution, (D) E. coli in the presence of Fe (III) -aminophosphonate, (A) E. coli in the presence of Cu(II) -aminophos- phonate and (O) E. coli in the presence of aminophosphonate alone.

EXAMPLE 1

Charging of Murexide wi th Fe (III) and Cu (II)

1.1. Materials and Methods.

The presence of reactive oxygen species (ROS) was analyzed indirectly by following the bleaching over time of Murexide

(Mu) as described by Torreilles et al . (1989). Metal ions were immobilised on a Chelix resin (purchased from BIO-RAD^®) which serves as the surface in the experiments. Murexide was purchased from Aldrich. Ascorbate (As) was used as the reducing reagent and hydrogen peroxide was used as the co-oxidant.

1.2. Procedure

0.7 g of a dry Murexide product, Chelix™ (capable of binding 1.4 mmol of divalent cations) was mixed in a 15 ml tube with 10 ml of either a 0.1 M solution of Fe₂(S0₄)₃ (corresponding to 1.0 mmol of Fe₂(S0₄)₃) or 10ml of a 0.1M CuS0₄. After 10 min of incubation the Chelix is washed 3 times with 10 ml of water and 4 times with 10 ml of buffer (phosphate 25 mM, pH 7.0). Each round of washing included (i) a centrifugation step (5000 rpm for 3 min) to sediment the Chelix, (ii) decanting of the supernatant and (iii) addition of washing solution followed by resuspending of the beads by whirl mixing. After the final wash the charged beads were resuspended in 5 ml of phosphate buffer.

The charged resin was light brown in the case of iron charging and dark blue in the case of copper charging.

A control Chelix (uncharged resin) was prepared exactly as described above with the exception that no iron was added. This resin was colourless. EXAMPLE 2

Generation of ROS on a surface that has chelated iron

The following reactions were set up .

Absorbance at 520 nm 1 . 1 ml of Mu + 250 μl uncharged Chelix +

200 μl water . 0 .5878

2 . 1 ml of Mu + 250 μl uncharged Chelix + 100 μl As ( 0 .1M) +

100 μl H₂0₂. 0 .4062

3. 1 ml of Mu + 250 μl Fe charged Chelix + 200 μl water. 0.6109

4. 1 ml of Mu + 250 μl Fe charged Chelix +

100 μl As ( 0 .1M) + 100 μl H₂0₂ (0 .1M) . 0 .3316

After mixing the reactions were left at room temperature overnight after which the absorbance of the murexide was recorded at 520 nm. Some bleaching of Murexide was observed in the control reaction (reaction 2, absorbance = 0.4062) with uncharged resin/ascorbate and hydrogen peroxide (compare reaction 1 and 2) . This bleaching effect was attributed to the presence of trace amounts of divalent metal ions in the water used in the experiments. However, when the charged resin was used the bleaching effect increased significantly (reaction 4; absorbance = 0.3316) indicating that the bleaching effect was due to metal binding to the chelating surface and consequently production of ROS. The production of ROS was further supported by the observation that the light brown colour of the Fe-resin changed to dark brown during the reaction (oxidation of resin) .

EXAMPLE 3

Generation of ROS on a surface that has chelated copper

An experiment similar to that in Example 2 was set up, but using copper ions and less charged resin. The bleaching effect was recorded after incubation at room temperature after 3 hours .

Absorbance at 520nm

1 . 1 ml of Mu + 50 μl uncharged Chelix + 400 μl water . 0 .9004

2 . 1 ml of Mu + 50 μl uncharged Chelix + 100 μl As ( 0 .1M) + 100 μl H₂θ2 ( 0 .1M) + 200ml of H₂0 0 . 6574

3 . 1 ml of Mu + 50 μl Cu charged Chelix + 400 μl water . 0 .4752

4 . 1 ml of Mu + 50 μl Cu charged chelix + 100 μl As (0 .1M) + 100 μl H₂0₂ (0 . 1M) + 200ml of H₂0 0 . 0706

Again some bleaching was observed in the control reaction (reaction 2) with uncharged resin/ascorbate and hydrogen peroxide (compare reactions 1 and 2) . Also, the bleaching increased significantly when the ion charged resin was used. Compared to the Fe experiment in Example 2 , the bleaching effect observed with Cu was substantially larger.

EXAMPLE 4

The antimicrobial effect of complexed metal ions

2.5 g of Chelix was charged to the maximum capacity with Cu, Fe, Zn or Ni, respectively. Washing 5 times with water and 4 times with 25 mM phosphate buffer, pH 7.0. Each of the charged Chelix resins and an uncharged resin (control) were resuspended in culture tubes with 10 ml of LB growth media containing 10 μl of an overnight culture of E. coli strain JM103. The culture tubes were incubated in a gyroshaker at 37°C and inspected visually for growth at various time intervals. After 24 hours a substantial growth inhibition of the E. coli was observed in the culture tube containing the Cu charged Chelix compared to the culture tube containing uncharged resin. The clearest reaction was observed with Cu, and a lesser effect was found for Fe, Zn and Ni.

To ascertain that the growth inhibition observed with the Cu resin was not due to free copper in the solution (which may have a toxic effect on the bacteria) the amount of possible free Cu ions in the culture solution was analyzed by mixing 1 ml of Murexide (75 μM or 75 nmol) with 50 or 200 μl of cul- ture solution. No Cu induced UV- shift of Murexide was observed. Since a UV shift can be observed with 7.5 μM Cu in this assay this means that the 200 μl E. coli culture contained less than 37.5 μM of free Cu ions. A concentration of free Cu ions of 100 μM was subsequently shown to have no effect for E. coli growth. We conclude that the observed effect of Cu was due to Cu bound to the resin and therefore was the result of generation of ROS.

EXAMPLE 5

Degradation of organic materials (bleaching) in the presence of free or surface bound transition metal chelates

5.1. Materials

AnalaR grade metal nitrates, manganese dichloride, hydrogen peroxide nitrilotriacetic acid (NTA) , Murexide (purpuric acid, ammonium salt) methyl orange and amaranth were purchased from Sigma-Aldrich Ltd. Chromatography beads were manufactured by Supelco and purchased from Sigma-Aldrich Ltd. All reactants were made up in distilled water. 5.2. Investigation of substrate bleaching in the presence of transition metal ions

Reactant solutions of chromium (III) nitrate, manganese (II) dichloride, iron (III) nitrate, cobalt (II) nitrate, nickel (II) nitrate and copper (II) nitrate, concentrations ranging from 2xl0^"6 to lxlO^"3 mol dm^"3, were made up in distilled water. Nitrates were chosen because of their high solubility in water. The total volume of reactant solutions was 50 ml and consisted of, in the first instance, distilled water, 2xl0^"6 or lxlO^"4 mol dm^"3 metal nitrate and 5x10^"5 mol dm^"3 Murexide. Additionally other solutions were made up as above but with 1.7xl0^"4 mol dm^"3 hydrogen peroxide added. Reactions were initiated by addition of Murexide to the reactant solution. The concentration of Murexide was monitored over the course of the reaction using a UV visible spectrophotometer to measure changes in absorbance at 521 nm, which is the Xm-^ of Murexide. Aliquots were taken sequentially over regular periods of time.

Murexide formed a complex with Ni(II) and Cu(II) ions. This was seen by a rapid colour change and the ^-^ of Murexide was observed to shift 20-30 nm towards the UV region of the spectrum.

Non-complexed transition metal ions did not initiate Murexide bleaching. Hence no change in Murexide concentration was observed in the presence of Mn(II), Co(II) (Figure 2), Ni(II) or Cu(II) ions. Although decreases in Murexide concentration were seen in the presence of Cr(III) and Fe(III) this is due to other factors associated with the chemistry of these particular ions.

All of the metal ions studied had no apparent effect on the hydrogen peroxide/Murexide reaction, with the notable exception of Cu(II), which strongly catalyzed the reaction. When Cu(II) is present in conjunction with hydrogen peroxide Murexide concentration was observed to rapidly fall to zero. During the course of the reaction oxygen bubbles were seen to form on the sides of the reaction vessel.

5.3. Investigation of substrate bleaching in the presence of transition metal ion NTA complexes.

The procedure was as above with the exception that metal ion solutions (lxlO^"4 mol dm^"3) were incubated in 1.5xl0^"4 mol dm^"3 NTA for at least one hour prior to the addition of Murexide, which marks the initiation of the reaction.

The conditions were the following: [M(N0₃)_n] , 1x10^"4 mol dm^"3; [NTA], 1.5xl0^"4 mol dm^"3 and [Murexide], 5xl0^"5 mol dm^"3. All solutions were made up in distilled water.

Metal ion-NTA complexes generally initiated Murexide bleaching so that where Murexide concentrations remained stable in the presence of the non-complexed ion, in the presence of the NTA complex a decrease in concentration was observed, see Figure 2. It is assumed that this decrease was due to the formation of ROS.

5.3. Investigation of substrate bleaching in the presence of surface bound transition metal ion complexes.

5.3.1. Use and preparation of chromatography beads

Chromatography beads were used as convenient models for applications involving surface bound metal ion complexes. Three types of chelating compound coated chromatography beads were studied: iminodiacetic acid (IDA) , see Figure 5 (structure 1) ; ethylenediamine bis methylenecarboxylate (EDAMC) , see Figure 5 (structure 2) ; and aminophosphonate, coated beads. The required weight of beads were washed with distilled water and then added to an aqueous solution of excess metal nitrate where they were left to incubate for at least one hour.

After incubation the beads were washed and filtered using copious quantities of distilled water so that excess metal ions were removed. The washed beads were then re-suspended in the reactant solution.

Reactions were initiated by adding the substrate to distilled water which held the chromatography beads which usually formed a monolayer on the base of the reaction vessel. Additional Cu(II) ions, 5xl0^"5 mol dm^"3, were added to certain reaction solutions, otherwise the general procedure was as described in the above sections. Reactions were initiated by the addition of the substrate, amaranth, methyl orange or Murexide to the reaction solution. 7x10^"7 mol dm^"3 hydrogen peroxide was added to certain reaction solutions after approximately 60 minutes.

Murexide concentrations fell rapidly in the presence of Fe(III)- and Cu (II) -EDAMC complexes so that approximately 30% of the total added substrate was bleached, see Figure 3. After this initial burst of bleaching the concentration of substrate stabilised. Reaction was revived by the addition of hydrogen peroxide. Concentrations of Murexide fell much less steeply in the presence of Fe(III)- and Cu(II)-IDA and aminophosphonate coated chromatography beads, but the observed decrease was greater than that that can be attributed to hydrolysis.

The addition of additional Cu(II) ions induced a dramatic increase in the degree of substrate bleaching initiated by Cu(II) IDA and aminophosphonate complexes. Amaranth, methyl orange and Murexide were totally bleached within 150 minutes in the presence of the above Cu(II) complexes and additional Cu(II), see Figure 4. Additional Cu(II) had no effect on Cu(II) -EDAMC bleaching. EXAMPLE 6

The effect of Fe (III) - and Cu (II) -IDA and aminophosphonate chelate complexes on the viabili ty of bacteria

6 . 1 . Materials

Micro-organisms used in the study were Escherichia coli ATCC 8739 (grown at 37°C) , Bacillus cereus NCIMB 6349 (grown at 30°C) and Pseudomonas fluorescence NCIMB 8178 (grown at 25°C) all of which were obtained from South Bank University, London, culture library. The micro-organisms were cultivated in Oxoid nutrient broth purchased from Fisher Scientific UK.

The anti-microbial properties of Iminodiacetic acid (IDA) and aminophosphonate coated polystyrene chromatography beads, manufactured by Supelco and purchased from Sigma-Aldrich Ltd, and complexed with Cu(II) or Fe(III) were investigated. Other materials are as described above.

6.2. Procedure and results

Reactant solutions (total volume 10 ml) containing nutrient broth and lxlO^"4 mol dm^"3 Cu(II) or Fe(III), and nutrient broth and various weights of complexed or non-complexed IDA or amino- phosphonate coated chromatography beads were inoculated with

0.1 ml of well mixed bacterial solution that had been grown for 16 hours. Inoculated samples were incubated in pre-set incubators at their optimum growth temperatures e . g. 37°C for E. coli . 0.2 ml samples were taken from the inoculated solutions at regular intervals and pipetted into micro- titre plates. The level of growth in the sample was measured using an ELISA plate reader (Dynatech MR 4000) at 490 nm.

Fe(III)- and Cu(II)-IDA and -aminophosphonate complexes inhibited bacterial growth, see Tables 6.1 to 6.5 and Figure 6. Table 6 . 1 . Ef fect of IDA coated chromatography beads on E. coli growth . Readinqs are opt: ical density (OD) measurements made at

490 nm.

Incubation Time / Hrs

Sample 3 5 7 9 11

E. coli + broth 0.069 0.109 0.130 0.157 0.171

Control

Fe(III)-IDA lg 0.009 0.047 0.063 0.076 0.081

Fe(iπ)-IDA 2g 0.007 0.027 0.044 0.058 0.061

Cu(π)-IDA lg 0.008 0.033 0.46 0.055 0.063

Cu(π)-IDA 2g 0.005 0.017 0.028 0.043 0.051

IDA lg 0.008 0.006 0.007 0.009 0.009

IDA 2g 0.003 0.004 0.005 0.003 0.003

Fe(iπ) 0.08 0.122 0.144 0.168 0.186

Cu(E) 0.067 0.106 0.118 0.167 0.187

Table 6.2. Effect of aminophosphonate (AP) coated chromatography beads on E. coli growth. Readings are optical density (OD) measurements made at 490 nm.

Incubation Time / Hrs Sample 2 22 27 44 48

E. coli + broth 0.003 0.506 0.580 0.652 0.702

Control

Fe(III)-AP lg 0.002 0.284 0.299 0.352 0.376

Cu(II)-AP lg 0.002 0.260 0.279 0.321 0.340

AP l g 0 003 0 003 0 004 0 005 0 006 Table 6.3. Effect of IDA coated chromatography beads on B . cereus growth. Readings are optical density (OD) measurements made at 490 nm.

Incubation Time / Hrs Sample 3 5 7 9 10

B. cereus + broth 0.156 0.247 0 437 0 602 0.621

Control

Fe(III)-IDA lg 0.140 0.185 0.227 0.302 0.337

Fe(iπ)-IDA 2g 0.091 0.105 0 168 0 173 0 185

Cu(π)-IDA lg 0.080 0.120 0.156 0.206 0.216

Cu(π)-IDA 2g 0.024 0.041 0.102 0.116 0 121

IDA lg 0.005 0.005 0.015 0 016 0 015

IDA 2g 0.001 0.014 0.018 0.015 0.107

Table 6 .4 : Effect of AP coated chromatography beads on B . cereus growth . Readings are optical density (OD) measurements made at 490 nm.

Incubation Time / Hrs

Sample 4 5 7 9 10

B. cereus + broth 0 206 0 265 0 504 0 566 0 581

Control

Fe(III)-AP 2g 0 076 0 082 0 128 0 136 0 140

Cu(II)-AP 2g 0 018 0.021 0 042 0 054 0 063

AP 2g 0 020 0 020 0 023 0 024 0 024 Table 6.5. Effect of AP coated chromatography beads on P. fluorescence growth. Readings are optical density (OD) measurements made at 490 nm.

Incubation Time / Hrs Sample 22 28 47 50

P. fluorescens + broth 0.727 0.924 0.980 0.990

Control

Fe(III)-AP 0.5g 0.160 0.228 0.21 1 0.234

AP 0.5g 0.081 0.134 0.150 0.166

Fe(III), 100 μmol dm^"3 0.615 0.941 0.879 0.896

The non-complexed ions had no effect on bacterial growth at experimental concentrations (lxlO^"4 mol dm^"3) . Non-complexed beads had the greatest inhibitory effect on bacterial growth. It is likely that this is due to the non-complexed ligands sequestering nutrient metal ions from solution. This possibly has the dual effect of starving the bacteria and also poisoning them with ROS generated from the nutrient ion ligand complexes.

EXAMPLE 7

The effect of Cu (II) -chelates on biological growth in pond water

Aminophosphonate, IDA and EDAMC coated chromatography beads, non-complexed and complexed with Cu(II), respectively, were submerged in water taken from a pond having a varied and thriving ecology. Subsequent rates of algal and other forms of biological growth were compared in pond water only and pond water containing the various chromatography beads, non- comple - ed and Cu (II) -complexed. The results are summarized in the below Table 7.1. Cu(II)-IDA and Cu (II) -aminophosphonate and the corresponding non-complexed chelating compound (ligand) coated chromatography beads inhibited algal growth in the pond water samples . The effectiveness of the Cu(II) surface complexes was limited to approximately 2 months, after which time algal growth began. It should be noted that the experiments described in Table 7.1 represents a "worst case scenario" with regards to ROS generation. In a closed system, with no flow or current, the greatest concentration of ROS will be situated around the beads with much lower concentrations in the bulk solution, making it easier for micro-organisms to grow. Also resources, such as reducing agents, will be finite and thus will represent a limiting factor. But this experiment did give a very positive indication that self-sustaining systems generating ROS continu- ously can act as strong biocides and maintain clean aqueous solutions for relatively long periods even in difficult environments such as those manifested in this experiment.

Table 7.1. Results from environmental tests with pond water,

Environmental Sample Description after 7 Description after 42 Description after 75 days days days

Control pond water Solution is clear Solution is greenish, Solution is greenish, much algal growth much algal growth

Pond water with 10^"4 Small dots of Algae on Algal growth, film mol dm^"3 Cu(N0₃)₂ flask base (?) formed on solution surface

Cu(II) -IDA Solution is clear Solution is clear Algal growth beginning, solution has green tint

Cu (II) -aminophosphonate Solution is clear Solution is clear Algal growth beginning. Non-complexed IDA Solution is clear Solution is clear Algal growth beginning. Non-complexed aminoSolution is clear Algal growth around Algal growth beginning. phosphonate edges Cu(II) -EDAMC Solution clear Algal growth beginning. Non-complexed EDAMC Solution clear Algal (red) growth beginning.

REFERENCES

1. Torreilles, J. , M-C Guerin and M-L Carrie. 1989, Biochi - mie, 71, 1231-1234.

2. Hirotsu, T., S. Katoh, K. Sugasaka, M. Sakuragi, K. Ichi- mura, Y. Suda, M. Fujishima, Y. Abe and T. Misonoo. 1985,

Journal of Polymer Science : Part A : Polymer Chemistry, 24, 1953-1966.

3. Valentine, J.S., C.S. Foote et al . (eds.) Active Oxygen in Biochemistry, Chapman & Hall (London) 1995.

4. Wahl, M.. 1989, Marine Ecology Progress Series, 58, 175-189.

Claims

1. A method of continuously generating reactive oxygen species (ROS) in an environment, the method comprising providing in said environment a chemical system which in the presence of an oxygen source and a reducing agent is capable of continuously generating reactive oxygen species, the system comprising (i) a redox active transition metal ion and (ii) a chelating compound capable of binding to the metal ion to form a chelate, the metal ion thereby being so firmly associ- ated with the chelating compound that it essentially does not dissociate from the chelate into the environment.

2. A method according to claim 1 wherein the oxygen source and/or the reducing agent is added to the environment.

3. A method according to claim 1 wherein the chelating com- pound is selected from the group consisting of EDTA, ECTA,

EHPG, HOEDTA, CHCEDG, CH3EHPG, CDTA, EHCHG, NTA, HEIDA, DTPA, EHPG, IDA, EDAMC, aminophosphonate and DHEG.

4. A method according to claim 1 wherein the chelating compound is a neutral chelating compound

5. A method according to claim 4 wherein the chelating compound is in the form of a hydroxylamine derivative.

6. A method according to claim 1 wherein the chelating compound comprises a substituent moiety which is capable of associating with a surface delimiting or being in contact with the environment whereby the chelate becomes fixated thereto.

7. A method according to claim 6 wherein the substituent moiety comprises a lipophilic group.

8. A method according to claim 6 wherein the substituent moiety comprises an anthraquinone group.

9. A method according to any of claims 6-8 wherein the chelate is associated with a particulate matter present in the environment .

10. A method according to claim 1 wherein in the chemical system at least part of the redox active transition metal ions are such metal ions naturally present in the environment .

11. A method of controlling in an aqueous, humid or intermittently humid environment the accumulation of organic matter on a surface delimiting or being in contact with the environment, the method comprising associating with said surface a chemical system as defined in any of claims 1-10 which in said environment is capable of continuously generating reactive oxygen species in an amount which prevents or reduces accumulation of organic matter on said surface.

12. A method according to claim 11 wherein the organic matter accumulatable on the surface comprises viable organisms.

13. A method according to claim 11 wherein the chemical system is incorporated into a paint or a coating composition that has been applied to the surface.

14. A method according to claim 13 wherein the chelate is fixated to particulate matter in the paint or the coating composition.

15. A method according to claim 13 or 14 wherein said paint or coating composition comprises a biocide compound.

16. A method according to claim 11 wherein the surface is selected from the group consisting of a ship's hull, a water storage reservoir, a waste water reservoir, the inner surface of a water pipe or tubing for transportation of liquid media, a liquor container, a fish net, an anchor, a rope, a roof, a fence, a boat, a submarine, a water inlet or outlet of a power station and other industrial plants, an aqueduct, a canalised waterway, a submerged marine structure, an industrial processing tank, an air conditioning system, a filter, a medical bandage or plaster, a packaging foil, a film, a plastic, paper for storage of foods and fluids, a medical device, a pharmaceutical, a household or industrial appliance and paper goods .

17. A method of impairing the viability of organisms or of degrading organic matter present in an aqueous, humid or intermittently humid environment, the method comprising providing in the environment a chemical system as defined in any of claims 1-10 which is capable of continuously generating in said environment reactive oxygen species in an amount which impairs viability of the organisms and/or degrades the organic matter.

18. A method according to claim 17 wherein the amount of reactive oxygen species has a biocidal effect on at least some organisms present in the environment .

19. A method according to claim 17 wherein the chemical system is associated with a surface delimiting or being in contact with the environment.

20. A method according to claim 19 wherein the surface is selected from the group consisting of a drinking water storage reservoir, a waste water reservoir, the inner surface of a pipe or tubing for transportation of a liquid medium, a liquor container, a water inlet or outlet of a power station and other industrial plants, an aqueduct, a canalised waterway, an industrial processing tank, an air conditioning system, a filter, a packaging foil, a film, a plastic, paper for storage of foods and fluids, a medical device, a pharma- ceutical, a household or industrial appliance and paper goods .

21. A method according to claim 17 wherein the chemical system is provided as particles with which the chemical system is associated.

22. A composition comprising a chemical system which in the presence of an oxygen source and a reducing agent is capable of continuously generating reactive oxygen species in an aqueous, humid or intermittently humid environment, the system comprising (i) a redox active transition metal ion and (ii) a chelating compound capable of binding to the metal ion to form a chelating compound/metal ion complex (a chelate) , the metal ion thereby being so firmly associated with the chelating compound that it essentially does not dissociate from the chelate when it is present.

23. A composition according to claim 22 which when it is provided in an aqueous, humid or intermittently humid environment is capable of continuously generating reactive oxygen species in an amount which impairs the viability of organisms present in the environment or degrades organic matter in the environment .

24. A composition according to claim 22 wherein the metal ion-binding compound is a chelating compound selected from the group consisting of EDTA, ECTA, EHPG, HOEDTA, CHCEDG, CH3EHPG, CDTA, EHCHG, NTA, HEIDA, DTPA, EHPG, IDA, EDAMC, aminophosphonate and DHEG.

25. A composition according to claim 22 wherein the chelating compound is a neutral chelating compound

26. A composition according to claim 25 wherein the chelating compound is in the form of a hydroxylamine derivative.

27. A composition according to claim 22 wherein the chelating compound comprises a substituent moiety which is capable of binding to the surface or to a component of the composition whereby the chelating compound is fixated thereto.

28. A composition according to claim 27 wherein the substituent moiety comprises a lipophilic group.

29. A composition according to claim 27 wherein the substi- tuent moiety comprises an anthraquinone group.

30. A composition according to claim 22 which is a paint or a coating composition.

31. A composition according to claim 30 wherein said paint or coating composition comprises a biocide compound.

32. A composition according to claim 22 which comprises an oxygen source and/or a reducing agent .

33. A composition according to any of claims 22-32 wherein the chemical system is provided as particles with which the chemical system is associated.