AU766899B2 - Synergistic antimicrobial compositions containing methylene-bis(thiocyanate) and an organic acid - Google Patents

Description

P/00/Oil 128/5/91 Regulafion 3.2(2)

AUSTRALIA

Patents Act 1990 COMPLETE SPECIFICATION STANDARD PATENT Application Number: Lodged: Invention Title: SYNERGISTIC ANTIMICROBIAL COMPOSITIONS CONTAINING METHYLENE-BIS(THIOCYANATE) AND AN ORGANIC ACID The following statement is a full description of this invention, including the best method of performing it known to :-us 1 SYNERGISTIC ANTIMICROBIAL COMPOSITIONS CONTAINING METHYLENE-BIS (THIOCYANATE) AND AN ORGANIC ACID The present invention relates to certain compositions and fungi in various products, materials or media, particularly industrial products, materials or media.

These products, materials or media include paints, leather, wood, wood pulp, wood chips, starch, proteinaceous materials, lumber, animal hides, vegetable tanning liquors, cosmetics, toiletry formulations, acrylic latex paint emulsions, adhesives, coatings, metalworking fluids, cooling tower water, textiles, petrochemicals, pharmaceutical formulations, geological drilling lubricants, paper mill liquors, and agrochemical compositions.

The novel compositions and processes incorporating the composition of the present invention show unexpected, synergistic activity against microorganisms, including bacteria and fungi. Specifically, the invention is directed to the use of compositions and processes containing methylenebis(thiocyanate) and at least one carboxylic acid, its salt or ester.

Many of the products, materials or media referred to above, when wet or subjected to treatment in water, are susceptible to bacterial and/or fungal deterioration or degradation unless steps are taken to inhibit such degradation or 20 deterioration. To control deterioration or degradation caused by microorganisms, various industrial microbicides are used, but some of these biocides are of questionable utility because they have undesirable odours, are high in cost, show a low degree of effectiveness and/or create hazards with respect to storage, use and/or handling.

25 For instance, the use of such popular industrial microbicides as organomercury compounds, organotin compounds and chlorinated phenols has ooo• come under increasing regulatory pressure in recent times because of their high toxicity and concern about their adverse effects on the environment.

Consequently, the industry has continued to seek improved biocides that have low toxicity and are capable of exhibiting prolonged biocidal effect at normal use levels.

Organic acids can be used alone to control microorganisms, and while some of these compounds are generally regarded as safe, many of them have 2 low efficacy against bacteria and fungi unless extremely high concentrations are used. In excessive concentrations, these organic acids can be expensive and could even be corrosive to certain industrial materials. A method that can prevent excessive use of organic acids and hence decrease cost is therefore desirable.

Alternatively, methylene-bis(thiocyanate) can be used alone in low concentrations as low toxicity biocides. However, at low concentrations, methylene-bis(thiocyanate) tends to have a narrow antimicrobial spectrum and fails to completely prevent the growth of microorganisms.

Accordingly, the present invention is directed to microbicidal compositions and processes incorporating these compositions that substantially obviate one or more of the problems, limitations, and disadvantages of the prior art. In particular, the compositions of the present invention are capable of controlling the growth of at least one microorganism, especially fungi or bacteria, over prolonged periods of time, and are safe and economical to use. The present invention also is directed to methods or processes of controlling the growth of at least one microorganism.

The present invention provides a composition to control the growth of at least one microorganism comprising synergistically effective amounts of (A) methylene-bis(thiocyanate) and at least one of the following compounds selected from the group consisting of benzoic acid and derivatives thereof, sodium benzoate, p-hydroxybenzoic acid, methyl paraben, propyl paraben, :ii dehydroacetic acid, octanoic acid, nonanoic acid, formic acid, sorbic acid, potassium sorbate, acetic acid, oxalic acid, glycolic acid, citric acid, malic acid, propionic acid, lauric acid, undecylenic acid, their respective salts and esters, and S 25 mixtures thereof. The composition provides superior microbicidal activity at low concentrations against a wide range of microorganisms.

The present invention also provides a method for controlling the growth of at least one microorganism in or on a material or medium susceptible to attack by the microorganism which comprises the step of adding to the material or medium 30 a composition of the present invention, where the components of the composition are present in synergistically effective amounts.

Furthermore, the present invention provides a method of preventing spoilage of a product, material or medium caused by bacteria and/or fungi 3 comprising the step of adding to the product, material or medium, a composition of the present invention where the components of the composition are present in synergistically effective amounts.

The synergistically effective amounts vary in accordance with the material or medium to be treated and can, for a particular application, be routinely determined without undue experimentation by one skilled in the art.

The present invention also embodies the separate addition of methylenebis(thiocyanate) and at least one carboxylic acid, its salt or ester to the products, materials or media described above. According to this embodiment, the components are individually added to the system so that the final amount of methylene-bis(thiocyanate) and at least one carboxylic acid, its salt or ester present in the system at the time of use is that synergistically effective amount required to control the growth of at least one microorganism.

The compositions of the present invention are also useful in preserving various types of industrial products, media or materials that are susceptible to attack by microorganisms. Such products, media or materials include but are not limited to dyes, pastes, lumber, leather, textiles, pulp, wood chips, tanning liquors, paper mill liquors, polymer emulsions, paints, paper and other coating V6000 .:ee.i 0 ee 000.

0 0 000e eee 4 and sizing agents, metalworking fluids, geological drilling lubricants, petrochemicals, cooling water systems, pharmaceutical formulations, cosmetics and toiletry formulations.

The compositions can also be useful in agrochemical formulations for the purpose of protecting seeds or crops against microbial spoilage.

Additional advantages of the invention will be set forth in part in the description which follows, and in part will be obvious from the description, or may be learned by the practice of the invention. The advantages of the invention may be realized and obtained by means of the elements and combinations particularly pointed out in the claims.

It is to be understood that both the foregoing general description and the following detailed description are exemplary and explanatory only and are not restrictive of the present invention as claimed.

When two chemical microbicides are combined into one product or added separately three results are possible: 1) The chemicals in the product would produce an additive (neutral) effect.

2) The chemicals in the product would produce an antagonistic (negative) effect, or 3) The chemicals in the product would produce a synergistic (positive) effect.

An additive effect has no economic advantage over the individual components. The antagonistic effect would produce a negative impact. Only a synergistic effect, which is less likely than either an additive or antagonistic effect, would produce a positive effect and therefore possess economic S" advantage.

It is well-known in the microbicidal literature that there is no theoretical method to provide the likelihood of knowing, before actually testing, whether additive, go antagonistic or synergistic effects will be obtained when two biocides are mixed to yield a formulation.

The microbicidal compositions combining methylene-bis (thiocyanate) and at least one organic acid, its salt or ester demonstrate an unexpected, synergistic effect compared to the respective components alone. Thus these compositions achieve superior, i.e. greater than additive microbicidal activity at low concentrations against a wide variety of microorganisms. Examples of microorganisms include fungi and bacteria such as, but not limited to, Trichoderma hazianum and Pseudomonas aeruginosa. These two organisms are some of the most common organisms associated with spoilage of products, materials or media. Since these two are also some of the toughest organisms to control, the composition of the present invention is believed to be effective against most bacteria and fungi. Preferably, the compositions of the present invention have a low toxicity.

The preparation of methylene-bis(thiocyanate) (MTC) is described in U.S.

Pat. No. 3,524,871, which disclosure is fully incorporated by reference. MTC is commercially available and also easily synthesized from commercially available raw materials.

Effective carboxylic acids or esters thereof according to the invention include dehydroacetic acid, octanoic acid, nonanoic acid, formic acid, sorbic acid, acetic acid, oxalic acid, glycolic acid, citric acid, gluconic acid, malic acid, propionic acid, lauric acid, undecylenic acid, benzoic acid or derivatives of benzoic acid such as 2-hydroxybenzoic acid, 3-hydroxybenzoic acid 3- or 4hydroxybenzoic acid, methyl paraben or propyl paraben.

The salts of the carboxylic acids, preferably those containing calcium, zinc, potassium, or sodium, may be used, such as sodium benzoate and potassium sorbate. Preferred esters are parabens such as methyl paraben and propyl paraben.

a In accordance with the present invention, mixtures of these carboxylic acids, salts or esters can also be used. When such mixtures are used in combination with MTC, at least one of the carboxylic acids in the mixtures has a synergistic relationship with MTC. Carboxylic acids, salts and esters useful in the invention are commercially available or may be synthesized from commercially available raw materials.

The carboxylic acid may be chosen, for example, based on the compatibility of the acid with the products, materials or media. The compatibility is readily determined by adding the organic acid to the products, materials or media to be used.

Compatibility may be determined by criteria such as solubility in a fluid system and/or lack of reactivity with the fluid in question. When used in a fluid system, for example, it is preferable that the organic acid be freely soluble or dispersible in the particular fluid system, resulting in a uniform solution or dispersion. Examples of fluid systems are tanning liquor, paper mill liquor, cooling tower water, and paints.

In accordance with the present invention, the composition may be in the form of a solid, dispersion, emulsion or solution, depending on the particular application. Further, the components of the composition may be applied 20 separately or may be combined first and then applied to the product, material or medium.

The composition of the present invention may be prepared in liquid form by dissolving components and in an organic solvent.

In the following discussion of preferred embodiments, component is 2- S: 25 methylene-bis(thiocyanate) and component is at least one carboxylic acid, its salt or ester.

As described above, components and of the composition are used in synergistically effective amounts. The weight ratios of to vary depending on the type of microorganisms, as well as the products, materials or media to which the composition is applied. One skilled in the art can readily determine the appropriate weight ratios for a specific application.

In the present invention, the weight ratio of component to component preferably ranges from about 0.01:99 to about 99:0.01, more preferably from about 1:30 to about 30:1, and most preferably about 1:5 to about 5:1.

The following approximate range of weight ratios of MTC to the following carboxylic acids or esters thereof are also preferred according to the present invention: MTC: benzoic acid 1:5 to 0.1:100 MTC: sodium benzoate 1:100 to 0.1:400 MTC: methyl paraben 1:5 to 0.01:100 MTC: propyl paraben 1:5 to 0.01:100 MTC: p-hydroxybenzoic acid 1:5 to 0.1:250 MTC: dehydroacetic acid 1:5 to 1:400 MTC: octanoic acid 1:5 to 1:400 MTC: nonanoic acid 1:10 to 1:400 MTC: formic acid 1:50 to 1:800 MTC: sorbic acid 1:5 to 0.1:100 MTC: potassium sorbate 1:5 to 1:800 MTC: acetic acid 1:100 to 1:200 MTC: oxalic acid 1:50 to 1:200 20 MTC: glycolic acid 1:100 to 1:400 MTC: citric acid 1:100 to 1:400 MTC: malic acid 1:100 to 1:400 MTC: propionic acid 1:10 to 1:400 MTC: lauric acid 1:5 to 0.1:400 25 MTC: undecylenic acid 1:5 to 0.1:500 In general, however, an effective fungicidal and bactericidal response can be obtained when the synergistic combination is employed in concentrations ranging from about 0.01 to about 3000 ppm of methylene-bis(thiocyanate), preferably from about 0.1 to about 1000 ppm, and most preferably from about 0.1 to about 500 ppm, and from about 0.1 ppm to about 1% by weight of the carboxylic acid, preferably from about 0.1 to about 5000 ppm, and most preferably from about 0.1 to about 2000 ppm.

8 The present invention also provides a method for controlling the growth of at least one microorganism in or on a material or medium susceptible to attack by the microorganism which comprises the step of applying to the material or medium a composition of the present invention, where the components of the composition are present in synergistically effective amounts.

Furthermore, the present invention provides a method of preventing spoilage of a product, material or medium caused by bacteria and/or fungi comprising the step of applying to said product, material or medium, a composition of the present invention where the components of the composition are present in synergistically effective amounts. For example, the composition may be used to prevent the spoilage of seeds or crops, e.g. cotton, barley, rice, maize, tobacco, etc.

The mode and rate of application of the composition varies depending upon the intended use of the composition. For instance, the composition may be applied by spraying or brushing onto a material or product. The material or product could also be treated by dipping in a suitable formulation of the composition. For a liquid or liquid-like medium, the *ee0 e 0e0 9 composition could be added into the medium by pouring, or by metering with a suitable device, so that a solution or a dispersion containing the composition is produced. If used as a liquid preservative, for example, the composition may be prepared as an aqueous emulsion. If necessary, a surfactant may be added to the composition.

In accordance with the invention, additional components such as insecticides and the like may be added to the foregoing preparations without affecting the synergistic effects of the composition. Insecticides that may be used include but are not limited to pyrethrins, nicotine, chlordane, parathions, and methoxychlor.

The synergistic activity of the combinations described above has been confirmed using standard laboratory techniques, as illustrated below. The following examples are intended to illustrate, not limit, the scope of the present invention.

SMicrobioloaical evaluation A. Fungal evaluation A mineral salts-glucose medium was first prepared by adding to 1 liter of deionized water: 0.7 g of KH PO 4 0.7 g of MgSO 4 .7H 20 1.0g of NH 4

NO

3 0.005 g NaCL, 0.002 g FeSO4.7H20, 0.002 g ZnSO 47H20, 0.001 g MnSO4.7H20, and 10 g of Glucose. The pH of the medium was adjusted to 6 with 1N NaOH. The medium was then dispensed in 5 mL amounts into test tubes and autoclaved at 121 0 C for 20 minutes.

The fungus, Trichoderma harzianum, was grown on a potato dextrose agar slant for 7 to 10 days and a spore suspension prepared by washing down the spores from the slant into a sterile saline solution. After the addition of biocides in the desired concentrations to the sterile mineral saltsglucose medium, the fungal spore suspension was added. The final spore concentration was approximately 10@6 cfu/mL. The inoculated media was incubated at 28°C. for 14 days.

B. Bacterial Evaluation Nutrient broth (2.5 g/liter of deionized water) was prepared and the pH adjusted to 6 with 1N HCL. This was dispensed in 5 mL amounts into test tubes and autoclaved for 20 minutes at 121°C. After the addition of biocides in the.

desired concentrations to the nutrient broth, 100 microliters of a suspension of Pseudomonas aeruginosa cells of approximately 9.3.times.10@8 cfc/mL were added and incubated at 37°C. for 48 hours.

In Examples 1 through 19, a synergistic effect was demonstrated by testing the combination of methylene-bis(thiocyanate), designated as component A, and the corresponding carboxylic acid, its salt or ester, designated as component B, in a series of tests in varying ratios and a range of concentrations against the fungus Trichoderma harzianum and also against the bacterium Pseudomonas aeruginosa, using the methods described above.

For each component A and B in a mixture containing A and B and for each component A and B acting alone, the lowest concentration which completely prevented growth of the fungi for two weeks and the bacteria for 48 hours was 20 determined. These concentrations were used as end points for synergism calculations. End points for the components alone or in mixtures described above were then compared with the end points for the pure active ingredients alone in similarly prepared flasks or test tubes.

Synergism was demonstrated by the method described by Kull, E. C., 25 Eisman, P. Sylwestrwicz, H. and Mayer, R. L. 1961. Applied Microbiology.

9: 538-541 wherein: QA/Qa+QB/Qb is less than 1 Qa Concentration of compound A in parts per million, acting alone, which produced an end point.

Qb Concentration of compound B in parts per million, acting alone, which produced an end point.

QA Concentration of compound A in parts per million, in the mixture, which produced an end point.

11 QB Concentration of compound B in parts per million, in the mixture, which produced an end point.

When the sum of QA/Qa and QB/Qb is greater than one, antagonism is indicated and when the sum is equal to one, additivity is indicated. When the sum of this value is less than one, synergism exists.

This procedure for demonstrating synergism of the compositions of this invention is widely used and accepted. More detailed information is provided in the article by Kull et. al. Further information concerning this procedure is contained in U.S. Pat. No. 3,231,509, which disclosure is fully incorporated by reference.

The Examples demonstrate that in almost all of the samples, the combination of MTC with a corresponding carboxylic acid produced a synergistic result (indicated by a ratio value of less than one.) There were a few samples, such as Table 2, <1.25 or where synergistic results were inconclusive because endpoints for the acids used alone were not determined.

It will be apparent to those skilled in the art that various modifications and variations can be made in the compositions and methods of the present invention without departing from the spirit or scope of the invention. Thus, it is intended that the present invention cover the modifications and variations of this invention 20 provided they come within the scope of the appended claims and their ~equivalents.

00 0.00 e *l 12 Example 1 Component A MTC Component B BenzoiC 2cid Test organism Quantities producing end points (ppm) Q, QA Q. Q QA/Q. QA/Q.

Irichoerm 10 2.5 250 0.25 0.5 0.75 5.0 25 0.5 0.05 0.55 5.0 50 0.5 0.1- 0.6 5.0 100 0.5 0.2 0.7 5.0 250 0.5 0.5 <1I 500 Psuomns I 0.1 25 0.1 0.1 0.2 0.1 50 0.1 0.2 0.3 0.1 100 10.1 0.4 0.25 25 0.25 0.1 0.35 0.25 50 0.25 0.2 0.45 0.25 100 0.25 0.4 0.65 0.5 10 10.5 0.04 0.54 0.5 25 0.5 0.1 0.6 0.5 50 0.5 0.2 0.7 0.5 100 0.5 0.4 0.9 250 13 Example 2 Component A NTC Component B =Sodium benzoate Test organism Quantities producind end points (ppm) QA__I Q1 QJQA Q/.

Thechoderma j10 hazinm 500 0.5 -0.25 10.75 5 I.1000 0.5 0.5 F- 5s__ 2000 0.5 1 Pseudomonas 1->00 aeruprnosa I- 0.25 I-1000 0.25 1 1.25 J0.5 I-250 0.5 0.25 0.75 -0.5 I*500 0.5 0.5 1 I0.5 1000 0.5 V V >1000 14 Example 3 Component A =MTC Component B Sorbic acid Test organism Quantities producing end points (ppm) Q. QA Q Q. Q,11/. QAQ+ Tihdna 10 0.5 S00 0.05 0.5 0.55 -I 500 0.1 0.5 0.6 2.5 250 0.25 0.25 2.5 500 0.25 0.5 0.75 5 25 0.5 0.03 0.53 50 0.5 0.05 0.55 -100 0.5 0.1 0.6 250 0.5 10.25 0.75 5 7 Soo0 0.5 0.5 1 1000 Psedomnfl- acruginosa -0.1 100 0.1 0.4 0.25 100 0.25 0.4 10.65 0.5 10 0.5 0.04 0.54 0.5 25 0.5 0.1 0.6 50 0.5 0.2 0.7 0.5_ 100 0.5 0.4 0.9 250 1.5 Example 4 Component A MTC Component B Potassium sorbaxe Test organism Quantides producing end poinus (ppm) 0-QQ .I Q Q,/0 QaIQ QAIQO+ Ircoem 10 -250 0.25 0.13 0.38 2.5 500 0.25 0.25 2.5 1000 0.25 0.5 0.75 2.5 2000 0.25 1 1.25 5 25 0.5 0.01 0.51 5 s0 0.5 0.03 0.53 5 100 0.5 0.05 0.55 5 250 10.5 0.13 0.68 5 500 0.5 0.25 0.75 5 1000 0.5 0.5 I 5 2000 0.5 1 2000 Pseudomonas I 0.5 500 0.5 0.5 <1I 0.5 1000 0.5 1 1000 16 Example Component A MTC Component B p-bydroxybenzoic acid Test organism Quantities producing end points (ppm) Q. Q, Q Q. QAI/Q QSJQ OQQ,+ Iihdra 1 0 2.5 1000 0.25 0.5 0.75 2.5 2000 0.25 1 1.25 25 0.5 0.01 0.51 50 .0.5 0.03 0.53 5 100 0.5 0.05 0.55 250 0.5 0. 13 0.63 5 500 0.5 0.25 0.75 1000 0.5 0.5 1 2000 0.5 1 2000 Pseudons -0.1 100 0.1 0.2 0.3 -0.1 250 0.1 0.5 0.6 0.25 100 0.25 0.2 0.45 0.25 250 0.25 0.5 0.75 0.5 5 0.5 0.01 0.51 0.5 10 0.5 0.02 0.52 0.5 25 0.5 0.05 0.55 0.5 so 5 0.5 0.1 0.6 -0.5 -100 0.5 0.2 0.7 -0.5 -250 0.5 0.5 1

C

C

CC

.C

I 17- Exam~ple 6 Component A MTC Com~ponent B Debydroacetic acid Test organism Quantities producing end points (ppm) Q. Q Q. Q.SQ. QS/Qb Q~I+

QSIQS

Irechoderm 10 2.5 50 0.25 0.1 0.35 2.5 100 0.25 0.2 0.45 2.5 250 0.25 0.5 0.75 5 25 0.5 0.05 0.55 50 0.5 0.1 0.6 5 100 0.5 10.2 0.7 5 250 0.5 0.5 1 500 PeudomnaS aeru-o~i s 0.25 100 0.25 0.4 0.65 0.5 5o 0.5 0.2 0.7 -100 0.5 0.4 0.9 ~250 a.

a.

a a.

a a a a a a a p a.

18 Example 7 Comnponent A =MTC Component B Propionic acid T-n nS flu2ntiti~ nmducin9 end DOOLS (oow) r Q. Q. QA/Q *IQ~ QA'Q+ Trihoderma 10-- 5 50 0.5 0.03 0.53 5 100 0.5 0.05 0.55 5 250 0.5 0.13 0.63 5 500 0.5 0.25 0.75 5 1000 0.5 0.5 I 2000 Psedomona I -0.25 100 0.25 0.4 0.65 -0.5 50 0.5 0.2 0.7 -0.5 100 0.5 0.4 0.9 250

S.

S S

S

S.

S

S

S

S

I.

S

*5 S

SS

1.9 Example 8 Component A Component B Test organism

MTC

Metbyl parabcn Quantites producing end points T- ITichde

Q.

I QA/Q QI'Q -500 0.1 0.5 0.6

QA/Q-

255 10.25 0.05 0.3 100 0.25 0.1 0.35 2520 0.25 0.25 0.25 0.75

V

S

V

V

a.

5 25 0.5 0.03 0.53 5 so5 0.5 0.05 0.55 5 1 00 0.5 0.1 0.6 5 -250 0.5 0.25 0.75 5 -500 0.5 0.5 <1I 1000 1000 0.1 1 1.1 -0.25 500 0.25 0.5 0.75 -0.25 1000 0.25 1 1.25 50 0.5 0.05 0.55 -100 0.5 0.1 0.6 -250 0.5 0.25 0.75 -500 0.5 0.5 I -0.5 -1000 0.5 1 >1000 sea* a* e.

20 Example 9 Comnponent A NfTC Component B Propyl paraben Test organism Quantities producing eod points 0 00*000 0 0 0000 0 S S

S

Q, Q5 Q Q/ Trihoder 10 I 250 0.1 0.5 0.6 2.5 50 0.25 0.1 0.35 2.5 100 0.25. 0.2 0.45 2.5 250 10.25. 0.5 0.75 2.5 25 0.5 0.05 0.55 50 0.5 0.1 0.6 5 100 0.5 0.2 0.7 5 250 0.5 0.5 1 -L 500 om_ 0.1 500 0.1 0.5 0.6 0.1 1000 0.1 1 1.1 0.25 250 0.25 0.25 0.25 500 0.25 0.5 0.75 0.5 5 0.5 0.00 0.5 10 0.5 0.01 0.51 0.5 25 0.5 0.03 0.53 0.5 10 0.5 0.05 0.55 0.5 10 0.5 0.15 0.65 0.5 250 0.5 0.25 0.75 0.5 1500 10.5 0.5 I ~>00 Iwo- 21 Example Component A MTC Component B Nonanoic acid Test organism Quantities producing end points (ppm) Q. Q'I Q. Q& QAIQ. QsIQt QIQ- Ticderma 10 2.5 250 0.25 0.5 0.75 -100 0.5 0.2 0.7 5 250 0.5 0.5 <1 500 Psu dona 0.25 100 0.25 0.4 0.65 0.5 5 0.5 0.02 0.52 0.5 10 0.5 0.04 0.54 0.5 25 0.5 0.1 0.6 50 0.5 0.2 0.7 0.5 100 0.5 0.25 0.75 250 00 0 0 ii 22 Example I11 Component A MTC Component B Odtanic acid Test organism Quantities producing end points (ppm) Q. Q, Q Q, QIQ. Q./Q QI.Q Tichodrma 1 0- haaiamm_ 2.5 10 0.25 0.2 0.45 2.5 00 0.25 0. 2 0. 2.5 250 0.25 0.5 0.75 5 25 0.5 0.05 0.55 50 0.5 0.1 0.6 5 100 0.5 0.2 0.7 5 250 10.5 0.5 <1I 50o Pedomas I 1 0.25 100 0.25 0.4 0.65 0.5 50 0.5 0.2 0.7 0.5 100 0.5 0.4 0.9 250 0 23 Example 12 Compooent A =MTC Component B =Undecyleaic acid Test organism Quantities producing end points Q. Q Q, QA/QQ QO/Q" OA/Q-f+ 0.5 5w0 0.05 0.25 0.3 0.5 1000 0.05 0.5 0.55 I 500 0.1 0.25 0.35 I 1000 0.1 0.5 0.6 2.5 50 0.25 0.03 0.28 2.5 100 0.25 0.05 0.30 25 25 0.5 0.25 0.51 2 5 0 5 0 05 0 5 2 5 50 0.5 0.03 0.53 5 100 0.5 0.05 0.55 250 0.5 0.13 63 500 0.5 0.25 0.75 2000 0.2 2.50 0.1 0.25 0.35 0.1 500 0.1 0.5 0.6 0.25 250 0.25 0.25 0.25 500 0.25 0.5 0.75 0.5 100 0.5 0.2 0.6 0.5 250 0.5 0.25 0.75 24 Examnple 13 Component A MTC Component B Lairic acid Test orgaaism Quantiie pro~ducing end points (ppm) Q. Q Q, QJQ. QSIQ. 1QAQ Tih-ena 10- 1hazau,- 2.5 500 0.25 I0.25 1000 0.25 I0.5 0.75 2.5 2000 0.25 T0.1 1.25 5 25 0.5 10.01 0.51 5 50 0.5 0.03 0.53 5 100 0.5 0.05 0.55 5 250 0.5 0.13 0.68 5 500 0.5 0.25 0.75 5 1000 0.5 0.5 1 5 2000 0. 1 2000 0.25 500 0.25 0.5 0.75 0.25 1000 0.25 1 1.25 0.5 100 0.5 0.1 0.6 0.5 250 0.5 0.25 0.75 0.5 500 0.5 0.5 <1I 0.5 -1000 0.5 1 1000 25 Example 14 Component A NfTC Componenat B Formic acid Test organismf Quatities producing end points (ppm) Q. QA Q Q QJ/QA QaQ- QA/Q

QS'Q.

Trichoer-a 10 2.5 2000 0.25 1 1.25 5 250 0.5 0.13 0.63 5 500 0.5 0.25 0.75 5 1000 0.5 0.5 I 5 2000 0.5 1 20D0 PeudruinmSnaII- -0.25 -50 0.25 0.5 0.75 50 0.5 10.5 <1I 100 26 Example Component A TC Component B Acetic acid Test organism Quantities producing end points (ppm) Q. Q.1 Q. QJQA Q. Q,/Q 1 2000 0.1 1 1.1 -20010 0.25 1 1< 1.25 5 Soo0 0.5 0.25 0.75 -1000 0.5 0.5 <1I 2000 0.5 1 2000 1- 0.25 50 0.25 0.5 0.75 50 0.5 0.5 I 100

S

0 27 Example 16 Component A =MTC Component B =Oxalic acid Test organism Quantities producing end points Q. Q Q, Q/Q QI/Qs. QAQ.

10 S 5 100 0.5 0.5 <1I 2000 0.5 1 2000 Pseukrnonas I aeruoinosa 0.25 -100 025 0.1 0.65 -0.5 -25 0.5 0.1 0.6 -0.5 -50 0.5 0.2 0.7 100 0.5 0.4 0.9 250 11 t 28 Example 17 Comnponent A MTC Comtpoonn B Citric acid T~t nrganhi~m Ouantiti~ nroducin2 end DODLS (onm) 0- QA Q QJ/QA QJIQ. Q'1IQ TrichoderTna 10 -5 -1000 0.5 0.5 <1I 5 -2000 0.5 1 2000 PsuonnsI- 0.25 100 0.25 0.4 0.65 0.5 25 0.5 0.1 0.6 0.5 s0 0.5 0.2 0.7 1250 -29- Example 18 Component A =MTC Component B Maiic acid Test organism Quantities producing cad points (ppm) Q. Q, Q Q8 QJ/QA" Q4.Q QA/I Idt&;ML 10 -1000 0.5 0.5 I -2000 0.5 1 2000 25 -100 0.25 0.4 0.65 -50 0.5 0.2 10.7 -0.5 100 0.5 0.4 0.9 250 Example 19 Component A MTC Component B Glycolic acid Test organism Quantities producing end points (ppm)

S..

Qa QA Qb QB Qa/QA Qb/QB QA/Qa QB/Qb Trichoderma 10 barzianum 5 1000 0.5 0.5 1 5 2000 0.5 1 >2000 Pseudomonas 1 aeruqinosa 0.25 100 0.25 0.4 0.65 0.5 50 0.5 0.2 0.7 0.5 100 0.5 0.4 0.9 250 Comprises/comprising and grammatical variations thereof when used in this specification are to be taken to specify the presence of stated features, integers, steps or components or groups thereof, but do not preclude the presence or addition of one or more other features, integers, steps, components or groups thereof.

0 0 f o .00.

S

S.

00 0 :0

Claims (11)

1. A composition to control the growth of at least one microorganism comprising synergistically effective amounts of methylene-bis(thiocyanate) and at least one of the following compounds selected from the group consisting of benzoic acid and derivatives thereof, sodium benzoate, p- hydroxybenzoic acid, methyl paraben, propyl paraben, dehydroacetic acid, octanoic acid, nonanoic acid, formic acid, sorbic acid, potassium sorbate, acetic acid, oxalic acid, glycolic acid, citric acid, malic acid, propionic acid, lauric acid, undecylenic acid, their respective salts and esters, and mixtures thereof.

2. The composition of claim 1 wherein the microorganism is bacteria or fungi.

3. The composition of claim 2 wherein said bacteria or fungi is Trichoderma harzianum or Pseudomonas aeruginosa.

4. The composition of any one of claims 1 to 3 wherein the weight ratio of (A) to is from 1:30 to 30:1. The composition of claim 4 wherein the weight ratio of to is from to 5:1.

6. The composition of any one of 1 to 3 wherein the weight ratio of concentrations are from 0.1 to 500 ppm of component A, and from 0.1 to 2000 ppm of the component B.

7. A method of controlling the growth of at least one microorganism in or on a product, material or medium susceptible to attack by said microorganism comprising the step of applying to said product, material or medium a composition in any one of claims 1 to 6.

8. The method of claim 7 wherein said product, material or medium is wood S pulp, wood chips, lumber, paints, leather, adhesives coatings, animal hides, tanning liquors, paper mill liquors, metalworking fluids, petrochemicals, pharmaceutical formulations, cooling tower water, cosmetics, toiletry formulations, textiles, geological drilling lubricants or agrochemical compositions for crop or seed protection.

9. The method of claim 7 wherein said components and are applied separately to the product, material or medium, or are first combined and then added to the product, material or medium. A method for preventing spoilage of a product, material or medium caused by bacteria, fungi or both comprising the step of applying to said product, material or medium, a composition in any one of claims 1 to 6 in effective amounts to prevent said spoilage.

11. A composition according to claim 1 substantially as hereinbefore described with reference to Examples 1 to 19.

12. A method according to claim 7 substantially as hereinbefore described with reference to Examples 1 to 19.

13. A method according to claim 10 substantially as hereinbefore described with reference to Examples 1 to 19. 0 9 DATED this 15th day of August 2003 BUCKMAN LABORATORIES INTERNATIONAL, INC. WATERMARK PATENT TRADE MARK ATTORNEYS 290 BURWOOD ROAD HAWTHORN VICTORIA 3122 AUSTRALIA KJS/JPFNRH P3052AU00 9eo9e o* o 9