EP3433349B1 - Enzymatic detergent compositions for the automatic cleaning of surgical and/or medical instruments and/or devices - Google Patents

Description

The present invention relates to the use of enzymatic detergent compositions for the automatic cleaning of surgical and / or medical instruments and / or apparatus.

In hospitals, surgical instruments are generally subjected to automatic cleaning in a washer-disinfector of instruments (LDI) using strongly alkaline detergents followed by thermal or chemical disinfection. These detergents generally have a pH higher than 12 which makes them very corrosive.

To avoid the damage caused by these alkaline detergents, enzymatic detergents have been proposed. We know for example the MEDICLEAN® forte from Dr. Weigert which is also the subject of the European patent EP 1 327 674 B1 . This detergent contains triethanolamine, nitrilotriacetic acid (NTA) as a chelating agent, phosphates and a protease. The simultaneous presence of an alkanolamine and an enzyme in the same composition poses stability problems for the enzyme, especially when the composition is very concentrated.

This is why Dr. Weigert has developed the neodisher® SYSTEM ALPHA which is composed of two distinct highly concentrated components, namely an enzymatic component, the neodisher® SystemAct, and an alkaline component, the neodisher® SystemClean. The enzyme component contains a subtilisin while the alkaline component contains triethanolamine and methylglycine diacetic acid (MGDA). This system is also the subject of the international patent application WO2015 / 043777 .

Even if this system makes it possible to solve the problems linked to the instability of the enzyme in the presence of high concentrations of alkaline agents such as alkanolamines, it has a major disadvantage in that it requires specific equipment to be able to feed the instrument washer-disinfector with the two components simultaneously. Instrument washer-disinfectors only have a supply of detergent composition.

The patent US 7,723,281 describes a detergent composition comprising a stabilized enzyme system. This enzymatic system comprises at least one enzyme but may include several, such as a protease, an amylase, a lipase, a gluconase, a cellulase, a peroxidase or a mixture thereof, as well as a chelating agent. According to the description, the stabilizing effect is obtained by incorporating an amine, in particular an antimicrobial, and an acid in a specific molar ratio.

There is therefore always a need for a means for effectively cleaning surgical and medical instruments in a machine without presenting the problems of stability of the solutions proposed in the prior art and without requiring specific apparatus in order to be able to supply the machines used for the cleaning.

It is to the credit of the Applicant to have found in the fruit of numerous researches that these drawbacks could be overcome by using a concentrated detergent composition which is based on the combination of three enzymes and a powerful chelating agent and which does not contain any alkanolamine.

• une subtilisine,
• une lipase,
• une amylase,
• un agent chélatant biodégradable choisi parmi l'acide méthylglycine diacétique (MGDA) et un iminodisuccinate (IDS) et
• de l'eau,

ladite composition détergente concentrée étant exempte d'alcanolamine.The invention therefore relates to the use of a concentrated aqueous detergent composition for cleaning surgical and / or medical instruments in a washer-disinfector, said concentrated aqueous detergent composition comprising:

• a subtilisin,
• lipase,
• an amylase,
• a biodegradable chelating agent chosen from acid diacetic methylglycine (MGDA) and an iminodisuccinate (IDS) and
• some water,

said concentrated detergent composition being free of alkanolamine.

Surprisingly, this specific combination of a subtilisin, a lipase and an amylase with MGDA or IDS eliminates the need for adding alkanolamine while maintaining effective cleaning of soiled surgical and / or medical instruments in a washer-disinfector. Thus, the concentrated aqueous detergent composition of the invention does not present the problems of enzymatic stability encountered in the enzymatic compositions of the prior art containing alkanolamines.

In addition, since the main components of biological tissues that constitute major stains in surgical and medical instruments are proteins, lipids (triglycerides) and carbohydrate polymers (glycogen), the tri-enzymatic cocktail of the concentrated aqueous detergent composition of the invention will simultaneously hydrolyze the three targets and thus facilitate detergency.

In addition, the absence of alkanolamine makes it possible to formulate the detergent composition used in accordance with the invention in a very concentrated manner. Thus, the water content of the concentrated aqueous detergent composition can be reduced up to 25% by weight relative to the total weight of the composition without having to add significant amounts of third-solvent, impact the stability of the enzymes and harm cleaning efficiency.

The composition used in accordance with the invention advantageously has a subtilisin content of 0.5 to 15% by weight relative to the total weight of the composition, preferably from 1 to 10% by weight relative to the total weight of the composition, preferably from 1 to 5% by weight relative to the total weight of the composition, more preferably around 3% by weight. relative to the total weight of the composition. The lipase content is advantageously from 0.05 to 2% by weight relative to the total weight of the composition, preferably from 0.5 to 1.5% by weight relative to the total weight of the composition, more preferably from about 1% by weight relative to the total weight of the composition. The amylase content is advantageously from 0.05 to 2% by weight relative to the total weight of the composition, preferably from 0.5 to 1.5% by weight relative to the total weight of the composition, more preferably from about 1% by weight relative to the total weight of the composition.

The biodegradable chelating agent is chosen from methylglycine diacetic acid (MGDA) and an iminodisuccinate (IDS), in particular the tetrasodium iminodisuccinate. Preferably, the biodegradable chelating agent is methylglycine diacetic acid (MGDA). Without wishing to be bound by any theory, the inventors believe that the chelating agent used in the present invention is not only used for softening water but also for complexing in a particularly effective way metal ions bi- and trivalent, in particular Ca ²⁺ , Fe ³⁺ , Fe ²⁺ and Cu ²⁺ , contained in the protein residues typically found on surgical and / or medical instruments. These chelating agents seem to sequester these metal ions, which destabilizes the tertiary structure of proteins and makes them more easily hydrolyzable.

The composition used in accordance with the invention advantageously has a content of biodegradable chelating agent of 10 to 50% by weight relative to the total weight of the composition, preferably from 20 to 40% by weight relative to the total weight of the composition, more preferably from 25 to 35% by weight relative to the total weight of the composition, more preferably still around 30% by weight relative to the total weight of the composition.

The concentrated aqueous detergent composition is advantageously free from any non-biodegradable chelating agent, in particular ethylene-diamine-tetraacetic acid (EDTA), diethylene-triamine-pentaacetic acid. (DTPA), phosphates and polyphosphates. Advantageously, the concentrated aqueous detergent composition is also free from any chelating agent classified as reprotoxic such as nitrilotriacetic acid and its sodium salt (NTA).

In one embodiment, the concentrated aqueous detergent composition is also free from alkali and alkaline earth hydroxides.

The water content of the concentrated detergent composition used in accordance with the invention is advantageously from 10 to 50% by weight relative to the total weight of the composition, preferably from 15 to 45% by weight relative to the total weight of the composition , more preferably from 20 to 40% by weight relative to the total weight of the composition and more preferably still from 25 to 40% by weight relative to the total weight of the composition. In addition to water, the concentrated detergent composition may also contain a third solvent such as a polyol. Examples of suitable polyols include alkylene glycols such as ethylene glycol, propylene glycol and glycerin (1,2,3-propanetriol). Preferably, the third solvent is glycerin. Glycerin has the advantage of not only serving as a third solvent but also of stabilizing the enzymes.

The content of third-solvent is advantageously from 5 to 50% by weight relative to the total weight of the composition, preferably from 10 to 40% by weight relative to the total weight of the composition, more preferably from 13 to 25% by weight. weight relative to the total weight of the composition.

The concentrated detergent composition used in accordance with the invention may also comprise one or more nonionic surfactants. The nonionic surfactant (s) may in particular be chosen from alkoxylated nonionic surfactants and alkylpolyglucosides. The alkoxylated nonionic surfactants are advantageously chosen from alkoxylated nonionic surfactants, in particular from alkoxylated fatty alcohols, in particular ethoxylated and / or propoxylated, and EO-PO block copolymers. In a preferred embodiment, the nonionic surfactant (s) are chosen from alkylpolyglucosides, preferably it is a C7 alkylpolyglucoside. This alkylpolyglucoside indeed brings the triple advantage of playing a stabilizing role, of bringing a wetting effect favorable to detergency and a non-foaming character compatible with an application in washer-disinfector.

The content of nonionic surfactant in the detergent composition used in accordance with the invention is advantageously from 1 to 10%, preferably from 2 to 5%, more preferably from approximately 3% by weight relative to the total weight of the composition.

The detergent composition used in accordance with the invention may further comprise an acid or a buffer system for adjusting the pH. Preferably, an organic acid is added, chosen from C 2 -C 6 mono-, di- and tricarboxylic acids, especially chosen from citric acid, tartaric acid, malic acid, lactic acid, glutaric acid, gluconic acid glycolic acid, succinic acid and adipic acid. Citric acid is particularly preferred. It has the advantage of serving at the same time as a complexing agent such as tartaric acid and gluconic acid.

Advantageously, the composition will not contain an antimicrobial agent, in particular no antimicrobial amine, and will therefore have no claim for antimicrobial activity at the dose of use. This is to be distinguished from a bacteriostatic effect which on the other hand can be sought. To this end, the detergent composition used in accordance with the invention may also comprise a preservative. The preservative can be chosen from the preservatives usually used, such as for example 5-chloro-5-methyl-4-isothiazoline-3-one, 2-methyl-4-isothiazolin3-one, p- acid hydroxybenzoic acid and its methyl esters, 5-bromo-5-nitro-1,3-dioxane and salicylic acid, benzyl alcohol and sodium benzoate. These preservatives prevent microbial and fungal contamination by bringing a bacteriostatic and fungistatic effect.

The detergent composition used in accordance with the invention may also comprise a hydrotropic agent. This hydrotropic agent can for example be chosen from xylenesulfonates, toluenesulfonates, cumenesulfonates, urea, methylacetamide, polyethylene glycol and polypropylene glycol.

The concentrated detergent composition used in accordance with the invention may also comprise other adjuvants such as, for example, dispersing agents and / or anti-foaming agents. The dispersing agents can for example be acids of homo-, co- or terpolymers based on acrylic acid or alkali metal salts thereof (polyacrylates).

In the use according to the invention, the detergent composition is used in a washer-disinfector (LD) such as a washer-disinfector of instruments (LDI) or a washer-disinfector of endoscope (LDE). The term "washer-disinfector" should be understood as defined in standard ISO 15883-1: 2006, namely a machine intended for cleaning and disinfecting medical devices and all other articles used in the medical, dental, pharmaceutical and veterinary. Also, the term "surgical and / or medical instruments" as used in the present application is to be understood broadly as designating medical devices and all other articles used in the medical, dental, pharmaceutical and veterinary environments requiring cleaning. with aqueous detergents before reuse.

In the present invention, this cleaning is advantageously carried out at a temperature of 20 to 60 ° C., preferably from 45 to 55 ° C., more preferably from 50 to 55 ° C. The cleaning time is advantageously between 20 s and 10 min, preferably between 30 s and 5 min.

The present invention also relates to the concentrated aqueous detergent composition used in the use according to the invention as described above. This composition is to be diluted with water, preferably softened, demineralized or osmosis, before use. The dilution is generally from 0.03% to 1% (v / v), preferably from 0.1% to 0.6% (v / v), more preferably from 0.2 to 0.4% (v / v) . The pH of the ready-to-use diluted composition is advantageously from 8 to 11, preferably from 8.5 to 9.5, more preferably from 8.8 to 9.2 and even more preferably from approximately 9. For the determination of the pH of the diluted composition, softened water is used (degree of hardness of approximately 5 ° f).

The invention will now be described with the aid of nonlimiting examples of embodiment of the invention.

FIGURES

• Figure 1 : Photos of on-board detergency tests for compositions C1, C2 and C3.
• Figure 2 : Evaluation of the activity of subtilisin by gel electrophoresis.
• Figure 3 : Evaluation of the activity of lipase by thin layer chromatography.
• Figure 4 : Evaluation of the activity of amylase by thin layer chromatography.

EXAMPLES

The following three concentrated detergent compositions (Table 1) were evaluated for their capacity to effectively clean residues typically found on surgical and medical instruments. <b> Table 1 </b> Composition (% by weight / total weight) C1 C2 C3 Demineralized Water 38.91 35.2 45.5 Citric acid 2.5 2.5 2.5 Technical glycerin 20 20 20 Ethoxylated fatty alcohol 3 3 3 Dioctyl dimethyl ammonium chloride 0.5 0.5 0.5 Methylisothiazolinone and methylchloroisothiazolinone 0.09 0.09 0.09 MGDA 30 - - IDS - 35 15 Polyacrylate - - 10 Savinase® Ultra 16 XL (subtilisin) 3 3 3 Lipolase 1 1 1 Amylase 1 1 1 pH (concentrated, 20 ° C) 9 9 9 pH (dilution to 0.5% (v / v) in softened water (5 ° f, 20 ° C) 8.9 8.5 8.2

EXAMPLE 1: On-board detergency tests in a MIELE G7836CD washer-disinfector 1) Cycle sequence

• Injection of detergent at 35 ° C, dilution of 0.3% (v / v) in softened water
• Wash with softened water for 3 minutes at 55 ° C
• Rinse with cold tap water
• Drying

2) The different on-board tests used

• Simicon (SIMICON PET-K-10 from SIMICON GmbH, Munich, Germany)
• Wash check H (Getinge, Sweden)
• Load Check Browne (Steris, USA)

The different tests were used according to the instructions for use provided by the respective manufacturer.

3) Test results

The performance tests have shown that the three compositions C1, C2 and C3 give completely satisfactory results under the test conditions. The best results are however obtained with composition C1. With this composition, we do not detect any trace on the three tests used ( Figure 1 ).

The tests were repeated at a cleaning temperature of 40 ° C and a dilution of the compositions to 0.5% (v / v). The results obtained are comparable to those obtained at 40 ° C and therefore confirm the effectiveness of the compositions according to the invention.

EXAMPLE 2: Comparative detergency tests in a MIELE PG 8528 washer-disinfector

The performance of composition C1 was compared with that of Anisoyme® DLM Maxi (Laboratoires Anios, Hellemmes, France), a trienzymatic detergent containing a protease, a lipase, an amylase and the chelating agent EDTA. The on-board test used was the TOSI AMCOR (reference 35002577).

• Prélavage : eau de réseau froide, 1 min,
• Lavage : eau adoucie (5°f), 5 min, température et dosage selon tableau 2 ci-dessous,
• Rinçage : eau de réseau froide, 1 min,
• Rinçage final : eau osmosée à 90°C, 1 min avec de l'Anios RDA à 0,05 % (v/v).

For each trial, the treatment cycle proceeds as follows:

• Prewash: cold mains water, 1 min,
• Washing: softened water (5 ° f), 5 min, temperature and dosage according to table 2 below,
• Rinsing: cold mains water, 1 min,
• Final rinse: osmosis water at 90 ° C, 1 min with 0.05% (v / v) Anios RDA.

Table 2 below summarizes the test conditions and results. <b> Table 2 </b> Detergent: C1 C1 Aniosyme DLM Maxi Aniosyme DLM Maxi Dosage (% v / v) 0.2 0.1 0.2 0.1 Wash temperature 45 ° C 50 ° C 45 ° C 50 ° C Result no trace no trace traces of fibrin traces of fibrin

These tests show that the composition according to the invention allows more effective cleaning at low temperatures and dosages. This is all the more surprising since the chelating activity of EDTA is at the base superior to that of MGDA which would allow waiting for a superior cleaning efficiency for Aniosyme® DLM Maxi.

EXAMPLE 3 Evaluation of the Activity of Enzymes in Composition C1

The respective stabilities of subtilisin, lipase and amylase in composition C1 were evaluated by the following tests.

1) Activity of subtilisin

The activity of the subtilisin contained in the detergent is followed by the technique of electrophoresis on polyacrylamide gel, the substrate used is a solution of human serum albumin at 0.3% (soiling representative of an application in health care establishment ).

After migration, the proteins and peptides are revealed by exposure in a staining solution with Coomassie blue followed by discoloration. A revelation with Coomassie blue reveals the degradation products resulting from the hydrolysis of human albumin serum.

The Figure 2 shows four tracks of electrophoresis: track 1 corresponds to the control (human albumin serum (0.3%) not degraded), tracks 2, 3 and 4 to composition C1 after manufacture, 3 weeks of storage in a hermetically closed container at 30 ° C and 7 weeks of storage in a tightly closed container at 30 ° C respectively. It is noted that the enzymatic activity of subtilisin does not degrade for at least 7 weeks.

2) Lipase activity

The activity of the lipase was followed by the technique of thin layer chromatography (TLC), the substrate used is a triglyceride and the eluent a mixture of 50 ml of heptane, 20 ml of diethyl ether, 0 , 5 mL of acetic acid and 1.5 mL of ethanol.

Lipase degrading triglycerides, the absence of triglycerides, and the presence of diglycerides and monglycerides highlights the enzymatic activity of lipase.

The Figure 3 shows six tracks of CCM: tracks 1, 2, 3 correspond to the controls (1: triglycerides, 2: diglycerides, 3: monoglycerides), tracks 4, 5 and 6 with composition C1 after manufacture, after 3 weeks of storage in a tightly closed container at 30 ° C and after 7 weeks of storage in a tightly closed container at 30 ° C respectively. It is noted that the enzymatic activity of the lipase does not degrade for at least 7 weeks.

3) Amylase activity

The activity of the amylase was followed by the technique of thin layer chromatography (TLC), the substrate used is glycogen and the eluent a mixture of 70 ml of ethanol, 1 ml of acetic acid and 29 mL of demineralized water.

Amylase degrading glycogen, the absence of glycogen and the presence of glucose demonstrate the enzymatic activity of amylase.

The Figure 4 shows five tracks of CCM: tracks 1 and 2 correspond to the controls (1: glycogen, 2: glucose), tracks 3, 4 and 5 to composition C1 after manufacture, after 3 weeks of storage in a container hermetically closed at 30 ° C and after 7 weeks of storage in a tightly closed container at 30 ° C respectively. It is noted that the enzymatic activity of amylase does not degrade for at least 7 weeks.

EXAMPLE 4 Influence of enzymes on the degreasing cleaning power

The influence of the presence of the three enzymes subtilisin, lipase and amylase in the detergent composition was evaluated by comparing composition C1 with compositions without enzymes (C4), with subtilisin (C5) and with lipase and amylase (C6). Composition (% by weight / total weight) C1 C4 C5 C6 Demineralized Water 38.91 43.91 40.91 41.91 Citric acid 2.5 2.5 2.5 2.5 Technical glycerin 20 20 20 20 Ethoxylated fatty alcohol 3 3 3 3 Dioctyl dimethyl ammonium chloride 0.5 0.5 0.5 0.5 Methylisothiazolinone and methylchloroisothiazolinone 0.09 0.09 0.09 0.09 MGDA 30 30 30 30 Savinase® Ultra 16 XL (subtilisin) 3 - 3 - Lipolase 1 - - 1 Amylase 1 - - 1

For these tests, the degreasing cleaning power of the various compositions was evaluated with respect to English soiling according to ISO standard 15883 which was applied with a brush on stainless steel blades (one blade per composition tested) and left to dry for one hour at ambient temperature. Then each blade was soaked with stirring (three-dimensional agitator, 50 rpm) for 15 min at room temperature in one of the four detergent compositions diluted to 0.3% (v / v) in softened water (5 ° f).

The best results have been obtained for composition C1. The slides treated with compositions C4, C5 and C6 indeed showed significant residues of English soiling visible to the naked eye. The determination of residual proteins by the MicroBCA technique (INTERCHIM UPTIMA MicroBC Assay kit, reference UP75860A) confirms these observations.

Claims (11)

1. Concentrated aqueous detergent composition comprising:

   - a subtilisin,

   - a lipase,

   - an amylase,

   - a biodegradable chelating agent selected from methyl glycine diacetic acid and an iminodisuccinate, and

   - water,

   said concentrated detergent composition being free of alkanolamine.
2. Detergent composition according to claim 1, characterized by the fact that it has a subtilisin content of between 0.5 and 15% by weight when compared to the total weight of the composition.
3. Detergent composition according to claim 1 or 2, characterized by the fact that it has a lipase content of between 0.05 and 2% by weight when compared to the total weight of the composition.
4. Detergent composition according to any one of the claims 1 through 3, characterized by the fact that it has an amylase content of between 0.05 and 2% by weight when compared to the total weight of the composition.
5. Detergent composition according to any one of the claims 1 through 4, characterized by the fact that the biodegradable chelating agent is methyl glycine diacetic acid.
6. Detergent composition according to any one of the claims 1 through 5, characterized by the fact that it has a biodegradable chelating agent content of between 10 and 50% by weight when compared to the total weight of the composition.
7. Detergent composition according to any one of the claims 1 through 6, characterized by the fact that it likewise comprises glycerin.
8. Detergent composition according to any one of the claims 1 through 7, characterized by the fact that it likewise comprises one or a plurality of non-ionic surfactants.
9. Use of a concentrated aqueous detergent composition according to any one of the preceding claims for the cleaning of surgical and/or medical instruments in a washer-disinfector.
10. Use according to claim 9, characterized by the fact that the cleaning is carried out at a temperature of between 20 and 60°C.
11. Use according to claim 9 or 10, characterized by the fact that the cleaning time period is between 20 seconds and 10 minutes.

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