ES2402725B1 - LIQUID DETERGENT FOR THE ELIMINATION OF BIOFILMS IN SURFACES. - Google Patents

Abstract

The present invention relates to a liquid detergent for removing biofilms on surfaces characterized in that it comprises, in percentage by weight of the total composition: between 10% and 20% of at least one oxidizing agent and biocide; between 1.0% and 3.0% of at least one chelating agent; between 0.7% and 1.5% of at least one phosphonate; between 2.0% and 3.0% of at least one cationic surfactant; and between 69% and 85.45% of a dissolution medium, such as water. The various detergent formulations are applicable to all types of surfaces, preferably open surfaces and closed circuit surfaces (both external and internal). Another object of the present invention is a method for removing biofilms from surfaces by means of the detergent described, which comprises diluting said detergent in water, contacting the diluted detergent with the surface to be treated, leaving a time between 5 and 15 minutes to act. , and clarify the surface.

Description

Liquid detergent for biofilms removal on surfaces

TECHNICAL FIELD OF THE INVENTION

The present invention is framed in the field of the elimination of biofilms adhered to surfaces, also referred to as biofilms, biological films, bio-layers or microbial films, formed by microorganisms, technique linked in this particular case to the maintenance and sanitation of machinery and industrial facilities, surfaces, food processing equipment, and cosmetic and pharmaceutical products, among others.

BACKGROUND OF THE INVENTION

There are both compositions on the market that act as an inhibitor of the adhesion of biofilms to surfaces (ES 2,195,038), and compositions intended to remove said biofilm.

In general, cleaning products used for the elimination of biofilms in industrial facilities are based on products of biological origin, such as enzymes (ES 2162593), which degrade components of the biofilm matrix such as polysaccharides and proteins. Due to the heterogeneity of biofilms, it is necessary to incorporate in this type of products various types of enzymes (glycase, lipase, protease) to achieve a proper degradation of the matrix. These products are generally more expensive to produce than those based on ingredients of non-biological origin, which results in a higher selling price. Additionally, enzyme-based products require their application under relatively mild temperature and pH conditions, due to the nature of their ingredients and often require long application times. These factors make your performance limited.

Essentially chemical compositions can also be found, such as that described in US 6,762,160 B2 formed from detergents. However, although these types of compositions successfully manage to eliminate biofilms on surfaces, they require a prolonged period of action on them, which is usually at least one hour. In addition, these types of chemical compositions contain potentially corrosive substances, such as acids and sodium hydroxide, which can cause deterioration by corrosion in the materials of the surfaces to be treated (stainless steel, Teflon, aluminum), as well as irritating substances that require a special control in its application as sodium hypochlorite.

Faced with these drawbacks detected in the state of the art, the liquid composition or detergent proposed here is based exclusively on products of chemical origin that act synergistically when degrading and releasing the biofilm from the surface to be treated susceptible to biofilm growth, as well as eliminating the microorganisms present, without requiring substances capable of causing corrosion in the materials and using very short contact times.

DESCRIPTION OF THE INVENTION

A first object of the present invention constitutes a composition, referred to herein as "liquid detergent", for the removal of biofilms on surfaces characterized in that it comprises, in percentage by weight of the total composition:

-

 between 10% and 20%, including both limits, of at least one oxidizing agent and biocide;

-

between 1.0% and 3.0%, including both limits, of at least one chelating agent as a calcium and magnesium ion complexer;

-

between 0.7% and 1.5%, including both limits, of at least one phosphonate as a metal ion sequestering agent;

-

between 2.0% and 3.0%, including both limits, of at least one cationic surfactant; Y

-

 between 69% and 85.45%, including both limits, of a dissolution medium,

in which the sum of these components does not exceed 100%.

The biofilm removal product is composed of a series of essentially chemical components that act on different elements of said biofilm, so that the breakdown of the protective matrix, the shedding of the film and the elimination of bacteria inside it is achieved. The function of each component is as follows:

· The at least one oxidizing agent degrades components of the biofilm protective matrix, aiding its detachment from the surface. It also has a biocidal effect that helps eliminate the microorganisms that make up the biofilm.

· The chelating agent acts as a complexer of calcium and magnesium ions in solution, which helps to destabilize the biofilm protective matrix and inhibit the growth of biofilms on the treated surfaces.

· The phosphonate acts as a sequestering agent for metal ions, contributing to destabilize the biofilm structure.

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· The cationic surfactant helps to improve the wetting properties of the composition and to the detachment of the surface biofilm, as well as to eliminate biofilm microorganisms.

The mechanisms of action of these components complement each other synergistically, in such a way that their combination is highly effective when it comes to degrading and releasing the biofilm from the surface and eliminating the microorganisms present, thus being a very effective tool for sanitizing surfaces in all types of facilities where biofilms can grow.

Among the main advantages of this liquid detergent compared to other known ones, we can highlight:

· The present composition produces an effective elimination of the biofilm with a very short contact time of about 5 minutes, compared to other compositions (US 6,762,160 B2) that require at least one hour of action / contact.

· The detergent object of interest achieves the elimination of the biofilm already formed, compared to other products (ES 2,195,038) that act only as an inhibitor of the adhesion of biofilms to surfaces.

· The present composition does not contain substances capable of causing corrosion in the surface materials to be treated (stainless steel, aluminum, Teflon), compared to other products that contain potentially corrosive substances such as acids and sodium hydroxide.

Preferably, the at least one oxidizing agent is a peroxide, and more preferably the peroxide is selected from hydrogen peroxide, peracetic acid and its derivatives and organic peroxides, although the first one is the most preferred of all.

In a preferred embodiment, which includes any of the above variants, the at least one chelating agent is selected from sodium ethylenediaminetetraacetate and nitrilotriacetic acid.

In a preferred embodiment, which includes any previous variant, the at least one phosphonate is selected from the group consisting of 2-aminoethylphosphonic acid, sodium dimethyl methylphosphonate, 1-hydroxy ethylidene-1,1-diphosphonic acid, amino tris (methylene phosphonic acid ) and diethylenediamine penta acid (methylene phosphonic acid).

Preferably, the cationic surfactant is a quaternary ammonium salt, such as, for example, benzalkonium chloride, alkyldioctadecyldimethylammonium chloride and didecyldimethylammonium chloride, among others.

Preferably, the dissolution medium is water.

Optionally, the liquid detergent may further comprise at least one non-ionic surfactant. In a particular embodiment, it is a non-ionic foaming surfactant, such as amine oxides and their derivatives, in a percentage by weight of the total formulation of up to 3%, being more preferably between 1.5% and 2.5%, including both limits. This component improves the wetting power of the mixture and helps foam generation. Said amine oxide is preferably selected from the group consisting of C10-16-alkyldimethyl amine N-oxide, C12-18-alkyldimethyl amine oxide, and dimethyl laurylamine oxide.

Alternatively, the liquid detergent may further comprise at least one antifoam surfactant, in a percentage by weight of the total formulation of up to 2%, and more preferably between 0.5% and 1.5% including both limits, to reduce foam generated by the mixture. Any surfactant with these properties can be used, such as alkyl polyglycosides, ethoxylated alcohols or siliconized compounds, but the one preferably used is "Aldo LF", manufactured by LONZA.

Although the present composition is applicable to any surface susceptible to being affected by the growth of biofilms, it has been preferably designed for application on open surfaces such as floors, walls, work tables, transport belts, accessible equipment surfaces, etc. . or in closed circuits, through which fluids circulate, such as pasteurizers, bottling equipment, automatic washing machines, bottle cleaning equipment, etc.

In a preferred embodiment, when used on open surfaces, the liquid detergent has the following formulation in percentage by weight of the total:

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 water: 77.3%;

-

hydrogen peroxide: 15%;

-

sodium ethylenediaminetetraacetate: 2.0%;

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Sodium salt of diethylenetriamine penta acid (methylene phosphonic acid): 1.3%;

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benzalkonium chloride: 2.4%; Y

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-C10-16-alkyldimethyl amine oxide: 2.0%.

In another preferred embodiment, when used in closed circuits and installations, where foaming should be avoided, the liquid detergent has the following formulation as a percentage by weight of the total:

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 water: 78.3%;

-

hydrogen peroxide: 15%;

-

sodium ethylenediaminetetraacetate: 2.0%;

-

Sodium salt of diethylenetriamine penta acid (methylene phosphonic acid): 1.3%;

-

benzalkonium chloride: 2.4%; Y

-

Antifoam surfactant: 1.0%.

Preferably, the described liquid detergent can also be prepared diluted in water in doses of between 3% and 5%, including both limits.

A second object of the present invention is constituted by a method for removing biofilms on surfaces by means of the liquid detergent described above in any of its variants, characterized in that it comprises at least the steps of:

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dilute said detergent in water,

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contact the diluted detergent with the surface to be treated, and leave for a time between 5 and 15 minutes, including both limits, and

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clarify the surface.

Preferably, the detergent is diluted in water in doses of 3% and 5%, including both limits. Also preferably, the detergent is diluted in water at a temperature between 20 ° C and 70 ° C, including both limits; preferably, the temperature is 35 ° C.

The contact time is preferably 5 minutes.

Preferably, the contact between the liquid detergent and the surface can be carried out in one of the ways selected from: immersion of the surface in a bath of the detergent; application of the detergent on the surface in the form of foam, by suitable equipment; injection of the detergent into a circuit and recirculation thereof, etc.

The surface is preferably rinsed with pressurized water, net or higher.

It has been proven that the present method produces reductions in the bacterial population of up to one billionth of the initial population, and biomass detachments (adhered biofilm) greater than 90%.

It should be understood from this specification that the invention also encompasses the use of liquid detergent, in any of its variants, to remove biofilms on surfaces, said surface being an open surface or a closed circuit surface (internal or external). Particularly (that is, preferably), the surface to be treated is selected from the group consisting of surfaces and circuits found in the manufacturing and processing industry of food, cosmetic and pharmaceutical products or other chemical products.

EXAMPLES OF EMBODIMENT OF THE INVENTION

The following describes, by way of example and on a non-limiting basis, a study of the effectiveness of various formulations of the detergent to protect biofilms on surfaces. Specifically, the preferred embodiments defined for cleaning open surfaces and closed circuits were analyzed in the laboratory, where fluids pass through, in which biofilms of Pseudomonas fluorescens ATCC 948 were formed.

Example 1. Formation of biofilms

The formation of biofilms was carried out in a batch system called "carousel", in which a series of stainless steel coupons are simultaneously immersed in a container containing a culture medium. For this, the strain of Pseudomonas fluorescens ATCC 948 was revitalized for 24 hours at 20 ° C in TSB broth (Oxoid). The cells were recovered after two consecutive centrifugations at 7500 rpm / 10 minutes and resuspended in fresh medium. The absorbance at 600 nm was adjusted to a value corresponding to a cell density of between 103-104 cfu / mL suitable for inoculation in the carousel. The carousels were incubated at 20 ° C for a period of 24 hours with constant orbital agitation of 80 rpm. After incubation, the coupons with the biofilm were aseptically removed with tweezers, washed with 0.85% sterile saline (w / v) to remove weakly adhered cells and subjected to the different cleaning treatments.

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Example 2. Cleaning method: surface exposure to detergent

For the cleaning treatment, each coupon, according to the method of the present invention, was immersed for 15 min at 20 ° C in a Falcon tube containing 15 mL of each of the preparations cited as preferred embodiments, diluted to 5% (w / v). To test each treatment, the sample used was composed of four coupons from the same culture, two of which were used for plate count and the other two, for densitometric analysis. Each pair came from diametrically opposite positions on the carousel. Cultures were performed in triplicate, so that the results are expressed as the average of six coupons for each treatment, both for cell counts and for densitometric analysis. From the same carousel, the coupons used as controls were obtained, which were analyzed immediately.

Example 3. Evaluation of cleaning efficiency

The cleaning efficiency was evaluated by two parameters, reduction of the adhered population after the treatment and optical densitometry of the treated coupons with respect to the untreated ones, a technique that gives information of the residual biomass (cells + matrix) after the cleaning treatment .

For plate counts the coupons were scratched with a swab that was then placed in a tube with 1.5 mL of peptone water. This tube was stirred vigorously for 1 minute to undo the possible cell aggregates, before making decimal dilutions. All dilutions were seeded, from 100 to 10-5. From each one 1 ml was seeded in depth, using TSA as a medium for plate counting. The plates were incubated at 30 ° C for 48h.

For densitometric analysis, the coupons were allowed to dry at room temperature and stained with a solution of Coomassie blue, whose composition per liter is: 1 g of bright blue, 250 mL of methanol and 100 mL of glacial acetic acid. The following steps were followed: immersion of the coupons in the previous solution for 1.5 min, washing with distilled water and drying at room temperature. This process was repeated 2 times. The quantification of the stained biomass was performed by measuring the average optical density of each of the stained coupons, using the transmission mode, indicated for translucent samples. A resolution of 64 µm and a green filter, corresponding to wavelengths of 520-570 nm, were recommended for samples stained with Coomassie R-250. For the calculation of residual biomass, 100% of the adhered biomass was assigned to the control coupons.

Data were analyzed statistically using a multiple range test for comparison of multiple samples (Fisher´s LSD test). This procedure applies a multiple comparison that serves to determine which mean values have significant differences, with a 95% confidence level.

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Results The results obtained are indicated in the following table:

Sample

Log10 cfu · cm-2 ± SD Reduction of adhered population (log10 control-Log treated) Detached biomass (%) ± SD

Control Composition 1 (foaming) Composition 2 (not foaming)

7.47 ± 0.49 Absence Absence 7.47 7.47 0 54 ± 2 55 ± 10

Also, after submitting the coupons with biofilm to a rinse with water under network pressure (3 bar) for 8 5 seconds, an amount of residual biomass on the surface of less than 1.0% was determined

According to these results, it can be affirmed that the treatment with both formulations of a surface with biofilms achieves a detachment of adhered biomass greater than 50%, as well as a total elimination of the microorganisms present from an initial population of 108 cfu / cm2. The subsequent rinsing of the surface eliminates almost all of the biofilm biomass, so that levels of biomass detached from 10 99% are reached.

Thus, it is proved that the proposed formulations are effective both in the elimination of the microorganisms present in the biofilm and in the degradation of the protective structure and consequent detachment of the surface.

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Claims (21)

1. Liquid detergent for removing biofilms on surfaces characterized in that it comprises, in percentage by weight of the total composition:

-

 between 10% and 20%, including both limits, of at least one oxidizing agent and biocide;

-

between 1.0% and 3.0%, including both limits, of at least one chelating agent as a calcium and magnesium ion complexer;

-

between 0.7% and 1.5%, including both limits, of at least one phosphonate as a metal ion sequestering agent;

-

between 2.0% and 3.0%, including both limits, of at least one cationic surfactant; Y

-

 between 69% and 85.45%, including both limits, of a dissolution medium.

2.

 Liquid detergent according to claim 1, wherein the at least one oxidizing agent is a peroxide.

3.

Liquid detergent according to claim 2, wherein the peroxide is selected from hydrogen peroxide, peracetic acid and its derivatives and organic peroxides.

Four.

Liquid detergent according to any one of claims 1 to 3, wherein the at least one chelating agent is selected from sodium ethylenediaminetetraacetate and nitrilotriacetic acid.

5.

Liquid detergent according to any one of claims 1 to 4, wherein the at least one phosphonate is selected from the group consisting of 2-aminoethyl phosphonic acid, sodium dimethyl methylphosphonate, 1-hydroxy ethylidene-1,1-diphosphonic acid, amino tris (methylene phosphonic acid ) and diethylenediamine penta acid (methylene phosphonic acid).

6.

Liquid detergent according to any one of claims 1 to 5, wherein the cationic surfactant is a quaternary ammonium salt.

7.

 Liquid detergent according to claim 6, wherein the quaternary ammonium salt is selected from benzalkonium chloride, alkyldioctadecyldimethylammonium chloride and didecyldimethylammonium chloride.

8.

 Liquid detergent according to any one of claims 1 to 7, wherein the dissolution medium is water.

9.

Liquid detergent according to any one of claims 1 to 8, further comprising at least one foaming nonionic surfactant in a percentage by weight of the total formulation up to 3%.

10.

 Liquid detergent according to claim 9, wherein the nonionic surfactant is an amine oxide or its derivatives.

eleven.

 Liquid detergent according to claim 10, wherein the amine oxide is selected from the group consisting of C10-16-alkyldimethyl amine N-oxide, C12-18-alkyldimethyl amine N-oxide, and dimethyl laurylamine oxide.

12.

 Liquid detergent according to any one of claims 1 to 8, further comprising at least one antifoam surfactant, in a percentage by weight of the total formulation of up to 2%.

13.

Liquid detergent according to claim 12, wherein the surfactant is selected from the group consisting of alkyl polyglycosides, ethoxylated alcohols and silicone compounds.

14.

 Liquid detergent according to any one of claims 12 or 13, wherein the surfactant is Aldo LF.

fifteen.

Liquid detergent according to any one of claims 9 to 11, which has the following formulation in percentage by weight of the total:

- water: 77.3%; -hydrogen peroxide: 15%; - sodium ethylenediaminetetraacetate: 2.0%; -sodium salt of diethylenetriamine penta acid (methylenophosphonic acid): 1.3%; -benzalkonium chloride: 2.4%; Y -C10-16-alkyldimethyl amine oxide: 2.0%. 16. Liquid detergent according to any one of claims 12 to 14, which has the following formulation in percentage by weight of the total: ES 2 402 725 Al -water: 78.3%; -hydrogen peroxide: 15%; - sodium ethylenediaminetetraacetate: 2.0%; -sodium salt of diethylenetriamine penta acid (methylenophosphonic acid): 1.3%; 5-benzalkonium chloride: 2.4%; Y - Antifoam surfactant: 1.0%. 17. Method for removing biofilms on surfaces by means of the liquid detergent described in any one of the preceding claims, characterized in that it comprises at least the steps of: - diluting said detergent in water, - contacting the diluted detergent with the surface to try, and let act for a while between 5 and 15 minutes, including both limits, and -lighten the surface. 18. Method according to claim 17, wherein the detergent is diluted in water in doses of 3% and 5%, including both limits, at a temperature between 20 ° C and 70 ° C, including both limits. A method according to any one of claims 17 or 18, wherein the contact time is preferably 5 minutes.

twenty.

 Method according to any one of claims 17 to 19, wherein the surface is rinsed with pressurized water.

twenty-one.

 Method according to any one of claims 17 to 20, wherein the surface is selected from an open surface, an internal surface of a closed circuit and an external surface of a closed circuit.

SPANISH OFFICE OF THE PATENTS AND BRAND Application no .: 201131745 SPAIN Date of submission of the application: 28.10.2011 Priority Date: REPORT ON THE STATE OF THE TECHNIQUE 51 Int. Cl.: C11D1 / 62 (2006.01) C11D7 / 36 (2006.01) RELEVANT DOCUMENTS

Category

56 Documents cited Claims Affected

TO

US 4525291 A (SMITH et al.) 25.06.1985, the entire document. 1-3

TO

EN 8405839 A1 (INTEROX CHEMICALS LIMITED) 31.03.1981, the whole document. 1-3

TO

WO 2008034995 A1 (GERUSZ, ROMAN; VANLAER, ANTOINE) 27.03.2008, the whole document. one

Category of the documents cited X: of particular relevance Y: of particular relevance combined with other / s of the same category A: reflects the state of the art O: refers to unwritten disclosure P: published between the priority date and the date of priority submission of the application E: previous document, but published after the date of submission of the application

This report has been prepared • for all claims • for claims no:

Date of realization of the report 06.07.2012

Examiner A. Amaro Roldán Page 1/4

REPORT OF THE STATE OF THE TECHNIQUE Application number: 201131745 Minimum documentation sought (classification system followed by classification symbols) C11D Electronic databases consulted during the search (name of the database and, if possible, terms of search used) INVENES, EPODOC, WPI State of the Art Report Page 2/4  WRITTEN OPINION Application number: 201131745 Date of Completion of Written Opinion: 06.07.2012 Statement

Novelty (Art. 6.1 LP 11/1986)

Claims Claims 1-21 IF NOT

Inventive activity (Art. 8.1 LP11 / 1986)

Claims Claims 1-21 IF NOT

The application is considered to comply with the industrial application requirement. This requirement was evaluated during the formal and technical examination phase of the application (Article 31.2 Law 11/1986).  Opinion Base.- This opinion has been made on the basis of the patent application as published. State of the Art Report Page 3/4  WRITTEN OPINION Application number: 201131745  1. Documents considered.- The documents belonging to the state of the art taken into consideration for the realization of this opinion are listed below.

Document

Publication or Identification Number publication date

D01

US 4525291 A (SMITH et al.) 06.25.1985

D02

ES 8405839 A1 (INTEROX CHEMICALS LIMITED) 31.03.1981

D03

WO 2008034995 A1 (GERUSZ, ROMAN; VANLAER, ANTOINE) 03/27/2008

 2. Statement motivated according to articles 29.6 and 29.7 of the Regulations for the execution of Law 11/1986, of March 20, on Patents on novelty and inventive activity; quotes and explanations in support of this statement The present invention relates to a liquid detergent for removing biofilms on surfaces characterized in that it comprises, in percentage by total weight of the composition: a) 10-20% of at least one biocidal oxidizing agent (such as hydrogen peroxide) , or peracetic acid and its derivatives); b) 1.0-3.0% of at least one chelating agent as a calcium and magnesium ion complexer (such as sodium ethylenediaminetetraacetate and nitrilotriacetic acid; c) 0.7-1.5% of at least one phosphonate as a metal ion sequestering agent; d) 2.0-3.0% of at least one cationic surfactant (such as a quaternary ammonium salt); and e) 69-85.45% of a dissolution medium (claims 116). In addition, it includes the method for removing biofilms on surfaces by using the aforementioned detergent by diluting said detergent in water (3-5% at 20-70 ° C) and allowing it to act for 5 to 15 minutes before rinsing the surface with water (claims 17- twenty-one). D01 refers to an aqueous and alkaline detergent composition containing at least 4% of an anionic sulfate or sulphonate surfactant and an ethoxylated non-ionic surfactant in which the proportion of ethoxylate in the ethoxylated nonionic alcohol is 60 to 80% of the molecule; at least 5% of an adjuvant selected from alkali metal and carboxylic polyphosphates, at least 2% hydrogen peroxide and other stabilizing components of the composition (summary and claims 1-23 and 28). D02 refers to a detergent composition that contains at least 2% hydrogen peroxide, which presents some difficulties so that it does not decompose during storage; It also contains a surfactant such as an anionic sulfate or sulphonate surfactant and / or an ethoxylated ionic surfactant, at least 5% of an adjuvant selected from alkali metal polyphosphates and if desired a hydrotope and one or more detergency aids (summary). D03 refers to a solid form cleaning agent such as tablets, granules, tablets or tablets coated with sodium chloride which dissolves in contact with water, characterized in that it comprises sodium chloride, a dispersing detergent agent and / or a surfactant Non-ionic with detergent action and an oxidizing agent that is selected from hydrogen peroxide precursors in solid form, such as: alkaline perborates, alkaline percarbonates, alkaline persulfates, alkaline peroxides, potassium permanganate, potassium ferrate or mixtures thereof. The procedure for obtaining and its use in filters and water circuits for swimming pools and collective premises is also mentioned (claims 1-11).  NEW AND INVENTIVE ACTIVITY The documents cited D1-D3 only show the state of the art in general and are not considered of particular relevance, since it would not be obvious for a person skilled in the art to apply the characteristics of the cited documents and arrive at the invention as It is mentioned in claims 1-11. Therefore, the purpose of this application meets the requirements of novelty and inventive activity in accordance with Articles 6-8 of Patent Law 11/1986. State of the Art Report Page 4/4