WO1996029361A1

WO1996029361A1 - A process for preparing antimicrobial matrices

Info

Publication number: WO1996029361A1
Application number: PCT/GB1996/000605
Authority: WO
Inventors: Laurence George Evans; Ian Rorke Burr; Stephen John Cunningham
Original assignee: Bioshield (Uk) Limited
Priority date: 1995-03-17
Filing date: 1996-03-15
Publication date: 1996-09-26
Also published as: AU4840196A; GB9505162D0

Abstract

An antimicrobial matrix and a process for preparing such a matrix, which comprises a support matrix, an antimicrobial agent and a carrying agent, wherein the carrying agent and the antimicrobial agent are adapted to form at least one hydrogen bond or salt bridge therebetween. The antimicrobial agent migrates through the support matrix to the surface thereof. Such a matrix can be extruded and/or moulded into articles for pharmaceutical, medical, catering, forming or personal hygiene use.

Description

A PROCESS FOR PREPARING ANTIMICROBIAL MATRICES

The present invention relates to a process for preparing antimicrobial matrices. In particular, it relates to a process for preparing anti-bacterial and anti-fungal matrices. The present invention also relates to anti-bacterial and anti-fungal matrices and articles formed from the matrices.

At present, an effective and practical way of removing unwanted infectious agents, such as bacteria, from an article is to wipe the article with a disinfectant solution. A disinfectant solution applied in this way acts both as a bacteriostatic agent (to hinder the growth of bacteria) and as a bacteriocidal agent (to kill

bacteria) due to the antimicrobial agent contained

therein. However, disinfectants only have effect in a wet environment. This is because the disinfectant solution "wets" the cell wall of an organism so that the

antimicrobial agent can pass into the cell to affect either protein synthesis, cell reproduction or internal respiration/energy production or to damage the cell wall sufficiently to cause lysis. The effectiveness of an antimicrobial agent is dependent to some extent upon the "wetting" of a cell wall. Thus, when the disinfectant solution applied to an article dries, the disinfectant looses its bacteriostatic and bacteriocidal properties. As a result, it is necessary to regularly apply

disinfectant to an article in order to prevent a build up of infectious agents.

Earlier patents suggest methods of manufacturing antimicrobial articles such as shields for telephone handsets. These shields have been made out of materials such as rubber, plastics and paper, which have been impregnated or impressed with an anti-bacterial agent.

EP-A-0 262 921 discloses bactericidal shields for telephones, wherein the shields comprise a rigid polyvinylchloride or rigid polystyrene formulation

containing a bactericidal agent, such as a heterocyclic or halogenated cyclic bacterial agent. However, a later patent application PCT/GB 90/00404, which discloses an invention devised by the same inventor, states that the shields described in EP-A-0 262 921 do not meet the object of that invention because the bactericidal agent can not diffuse out of the matrix. Thus, the bactericidal agent is ineffective both on the surface of the shield and remote therefrom.

PCT/GB 90/00404 discloses a process for preparing bactericidal matrices comprising a plastics-type support matrix, an alkylated diaminoalkane-type antibacterial agent and an organoarsenic-type anti-bacterial agent. The anti-bacterial agents allegedly act

synergistically to enable the matrices to show

bactericidal activity. The document clearly states that a mixture comprising a plastics-type matrix and either one of the disclosed anti-bacterial agents alone showed substantially no bactericidal activity.

It is therefore an object of the present

invention to provide a process for preparing an improved antimicrobial matrix.

According to the present invention there is provided a process for preparing an antimicrobial matrix comprising mixing a support matrix with an antimicrobial agent and a carrying agent, wherein the carrying agent and the antimicrobial agent are adapted to form at least one hydrogen bond or salt bridge therebetween.

The resulting mixture may be extruded, and/or formed or moulded, to produce articles having

antimicrobial properties.

Preferably, a surfactant is also mixed with the support matrix.

According to the present invention there is further provided an antimicrobial matrix comprising a support matrix, an antimicrobial agent and a carrying agent, wherein the carrying agent and the antimicrobial agent are adapted to form at least one hydrogen bond or salt bridge therebetween.

The antimicrobial matrix may also comprise a surfactant.

According to the present invention in another aspect, there is provided an article incorporating or comprising an antimicrobial matrix as defined above.

In the antimicrobial matrix, the carrying agent and the antimicrobial agent are linked by hydrogen bonding or salt bridge and migrate over a period of time through the support matrix to the surface thereof. The

antimicrobial agent then acts to kill bacteria both on the surface of the matrix, and at a distance therefrom by virtue of further diffusion, thus forming a 'halo' area (inhibition zone).

The rate of migration and of diffusion of the carrying agent and the antimicrobial agent through the support matrix is dependent on a number of factors

including; the length of the polymer chain (molecular weight) of the carrying agent; the hydrophobicity or hydrophilicity of the carrying agent; and the molecular composition and/or density of the support matrix.

Thus, by selectively choosing the carrying agent and/or the support matrix, the rate of migration, the rate of diffusion and/or the surface concentration of the antimicrobial agent can be controlled. These factors will determine the degree and duration of activity of the antimicrobial agent.

Also, the carrying agent, by virtue of its wetting properties and viscosity, acts as a 'flypaper', adhering bacteria to it.

The addition of a surfactant improves the effectiveness of the antimicrobial agent by 'wetting' the cell wall of an organism to enable the antimicrobial agent to pass more easily therein. The surfactant is

particularly effective with respect to gram positive bacteria.

The antimicrobial agent is preferably active against both gram positive and gram negative bacteria. It is also preferably stable to light, heat and acid. In one example, the antimicrobial agent is chloramphenicol which has been environmentally and clinically approved as an antimicrobial agent world-wide in eye drops, for example. Chloramphenicol has the additional benefit of being effective against Escherichia Coli, Staphylococcus Aureus, Listeria, Salmonella enteritidis and Legionella.

Other antimicrobial agents include substituted isothiazolinones, bisguanadines eg chlorohexidine, and organic peracids.

Alternatively, the antimicrobial agent may be an anti-fungal agent such as undecenoic acid or griseofulvin.

Further, the antimicrobial agent may be an antiviral agent or an algaecidal agent.

The antimicrobial agent is preferable incorporated into the matrix at a concentration of from 0.5 to 5% weight by volume.

The carrying agent is a polymer and may be polyvinyl pyrolidone, coconut oil or polyethylene glycol (PEG). For example, PEG 300, PEG 400 or PEG 600 may be used, the numerals indicating the average molecular weight of each polymer. The molecular weight of the polymer used affects the rate of replenishment of the antimicrobial agent on the surface of the support matrix.

The support matrix may be of natural or synthetic plastics, natural or synthetic rubber, silicone rubber, nitriles, fabric, polymeric species, biopolymers or resin. In one example, the support matrix is poly vinyl chloride. In another example, it is a synthetic rubber such as Caraflex (trade mark). Other examples are low density polyethylene (LDPE), low density polypropylene (LDPD), polyurethane, ethylene vinyl acetate and

acrylonitrile-butadiene-styrene (ABS).

The choice of support matrix will affect the choice of carrying agent, since the carrying agent must not be incorporated within the molecular structure of the support matrix, else the carrying agent can not migrate and the matrix will show relatively little, or no,

antimicrobial activity.

The antimicrobial matrix is prepared by mixing the carrying agent with the antimicrobial agent and, optionally, the surfactant. The resulting mixture is added to the support matrix and kneaded until it becomes homogenous therewith.

The resultant homogenous antimicrobial matrix may be extruded and/or moulded into articles for

pharmaceutical, medical, catering, farming or personal hygiene use. For example, the antimicrobial matrix may be incorporated into containers or wrapping for medical eguipment or surgical gloves. Paint or cladding for the floors, wails and ceilings of hospitals, restaurants, breweries and abattoirs may also include the antimicrobial matrix. In addition, the antimicrobial matrix may be used in bathroom equipment such as toilet seats, in door seals and liners for fridges or freezers as well as a

disinfectant in washing-up liquids or washing powders.

The antimicrobial matrix can also be made into a shield for the mouthpiece or a telephone. In yet a further application the antimicrobial matrix may be incorporated in filters (eg as a foam) or baffles for air-conditioners.

The antimicrobial matrix prepared by the process of the present invention may therefore have any number of applications, including the non-limiting applications mentioned above.

Any screw slip occurring during the extrusion of the resultant homogenous antimicrobial matrix may be reduced by adding an inert material thereto.

The present invention is now described, by way of illustration only, in the following examples.

EXAMPLE 1 (Normal batching)

A batch of antimicrobial material was prepared by adding 40 millilitres of polyethylene glycol 400

(hereinafter referred to as PEG 400) to 2.5 grams of chloramphenicol and 0.5 grams of sodium dodecyl sulphate (hereinafter referred to as SDS). The resulting mixture was stirred until the chloramphenicol had dissolved. This mixture was then added to 1 kilogram of pellets of medical grade (tested for purity by microbiological assay)

polyvinylchloride (hereinafter referred to as PVC)

supplied by Hydropolymers, and the resulting mixture was kneaded until it became homogenous.

The homogenous mixture was added to the hopper of a compounding extruder and subsequently extruded. The temperature of extrusion did not exceed 200°C as

chloramphenicol starts to thermally decompose above this temperature. An extrusion temperature of from 160 °C to 130°C was found to be preferable for PVC. Any screw slip during extrusion was reduced by adding 1 to 5 grams of inert material per 1 kilogram of PVC to the hopper.

Preferred inert materials were calcium carbonate or talcum powder (Mg₃(OH)₂Si₄O₁₀).

The resulting extrudate was subsequently moulded. Again, the temperature of moulding did not exceed Z00°C because chloramphenicol is unstable above this temperature. A moulding temperature of from 160°C to 180 °C was found to be preferable for PVC.

A masterbatch of antimicrobial material may also be prepared, a masterbatch comprising an additive or additives mixed with a carrier that can be blended with other products such that the additive or additives can be dispersed in subsequent processing.

TESTING METHOD

The method or protocol for determining by antibiotic assay the bactericidal activity of the

antimicrobial matrices of this invention is described below.

Firstly, petri-dishes containing nutrient agar were seeded with, inter alia, one of the following test organisms:

Escherichia coli - NCTC 8196

Escherichia coli - NCIB 9484

Escherichia coli - NCTC 10418

Staphylococcus aureus - NCTC 4163

Staphylococcus aureus - NCTC 8532

Staphylococcus aureus - NCIB 8625

Escherichia coli NCTC 8196 and Staphylococcus aureus NCTC 4163 are standard disinfectant test strains.

Secondly, a 1 cm² sample of the antimicrobial matrix was placed aseptically onto the middle of the surface of each seeded plate. Each plate was then placed overnight in a fridge at 4°C to allow the antimicrobial agent to diffuse into the agar.

Thirdly, the plates were incubated at 30°C for 12 hours.

Finally, the plates were examined for the degree of growth inhibition. The inhibition zones were measured and photographed.

RESULTS

Table 1 shows the results of tests, read on 20 November 1993, conducted by Royal Hampshire County Hospital (District Pathology Laboratory) on samples of an antimicrobial matrix received there and inoculated on 18 November 1993.

Table 2 shows the results of tests, read on 28 November 1993, conducted by Royal Hampshire County

Hospital (Department of Microbiology) on samples of an antimicrobial matrix received there on 26 November 1993 and inoculated on 26 November 1993.

Table 3 shows the results of tests, read on 21 April 1994, conducted by Royal Hampshire County Hospital (Department of Microbiology) on the samples of the antimicrobial matrix received these on 26 November 1993 and inoculated on 19 April 1994.

Table 4 shows the results of cross-validation tests conducted by Southampton General Hospital (Public

Health Laboratory) on 9 May 1994 on the samples tested in relation to tables 2 and 3.

Table 5 shows the results of tests, read on 27 April 1994, conducted by Royal Hampshire County Hospital (Department of Microbiology) on samples of the

antimicrobial matrix received there on 26 November 1993 and inoculated on 25 April 1994.

Table 6 shows the results of tests performed on a flew moulded EVA pad conducted by Royal Hampshire County Hospital (Department of Microbiology) on samples received there on 19 December 1994, incubated on 21 December 1994 and read on 22 December 1994.

Table 7 shows the results of tests, read on 27 February 1995, conducted by Royal Hampshire County

Hospital (Department of Microbiology) on samples of the antimicrobial matrix received there on 26 November 1993, inoculated on 23 February 1995 and incubated on 24

February 1995. CONCLUSION

The antimicrobial matrices and articles according to the present invention exhibit excellent antimicrobial properties, even after a period of time.

Example 2

To prepare a batch of antimicrobial material from synthetic rubber, the process described in Example 1 was used but synthetic rubber was substituted for PVC. The synthetic rubber used was Caraflex (trade mark).

Example 3

A batch of antimicrobial material was prepared by adding 10 grams of polyethlene glycol 400 (hereinafter referred to as PEG400) to 20 grams of chlorhexidine mixture and this was then added to 1 kilo of polyurethane and the resultant compound was mixed until it became totally homogenous. The mixture was then poured into an aluminium mould and allowed to expand to form a foam block.

Testing Method

The method or protocol for determining by antibiotic assay the antibacterial activity of the antimicrobial matrices of this example is as described in Example 1.

Results

Table 8 shows the results of tests, read on 18 February 1996, conducted by Royal Hampshire County Hospital (department of Microbiology) on samples of an antimicrobial matrix received there on 16 February 1996 and innoculated on 17 February 1996.

Conditions: stored at 4 degrees C after 12 hours

incubation.

The expanded foam block formed was sliced into thin sheets 2 mm thick and from the sheets were cut a shape about 25 mm across suitable for covering the speech area of telephone mouthpiece, thus forming a telephone shield. To the back of the telephone shield an adhesive was applied so that it could be attached to the telephone mouthpiece. The adhesive was then covered with a peel-off covering.

Acoustic Test

Method

A circular pad 40 mm diameter was cut from a strip of 2 nm thick foam plastic material prepared as described above. The pad was fitted centrally over the mouthpiece of the telephone handset, so that all the holes in the mouthpiece were covered. The pad was fixed in place using a 'PrittStick' adhesive stick. The handset was an Eagle type TH7SP obtained from the NESCOT Media Services Department. It was held vertically using a retort stand 15 cm front of a Tannoy Loudspeaker (of cabinet dimensions 30 cm by 50 cm by 30 cm) with the mouthpiece facing towards the loudspeaker and opposite to its centre point.

The output signal from the microphone in the handset was measured using a Bruel and Kjaer sound level meter type 2203 fitted with octave band filters type 1613. The sound level meter was first checked to ensure that it was operating satisfactorily using a Bruel and Kjaer calibrator type 4232. The Bruel and Kjaer microphone was then removed and the signal from the microphone in the handset fed into the sound level meter using an adaptor.

The loudspeaker was fed with a white noise signal from a Bruel and Kjaer sine-random signal

generator, and the signal from the microphone in the handset measured in octave bands from 125 Hz to 8000 Hz. The foam plastic pad was removed from the mouthpiece of the handset and the measurements were repeated. The measurements were also repeated again, with the signal to the loudspeaker reduced to zero, to measure the background noise levels in the laboratory, to ensure that they were not affecting the results of the test.

Results

There was no measurable difference in any of the seven octave frequency bands from 125 Hz to 8000 Hz between the sound pressure level measurement with or without the foam plastic pad covering the mouthpiece of the handset.

Conclusion

When the tele:, .-.one handset was placed in front of a loudspeaker the presence of a 3 mm thick foam plastic pad fitted over the mouthpiece did not produce any

detectable difference to the level of sound received by the microphone in the handset. If any differences do exist they are much less than one decibel, which was the level of precision in these tests, and would be completely unnoticeable to the human ear.

Example 4

The mixture of PEG400, chlorhexidine and polyurethane was prepared as described in Example 3 and poured into a mould in the shape of a toilet seat and allowed to expand.

Claims

1. An antimicrobial matrix comprising a support matrix, an antimicrobial agent and a carrying agent, wherein the carrying agent and the antimicrobial agent are adapted to form at least one hydrogen bond or salt bridge therebetween.

2. The matrix according to Claim 1, comprising, in addition, a surfactant.

3. The matrix according to Claim 1 or Claim 2, comprising from 0.5% to 5% weight by volume antimicrobial agent.

4 . The matrix according to any one of Claim 1 to 3, wherein the support matrix is selected from the group comprising natural or synthetic plastics, polymeric species, natural or synthetic rubber, silicone rubber, nitriles, fabric, resin or biopolymer.

5. The matrix according to Claim 4, wherein the polymeric species is polyvinyl chloride, low density polyethylene, low density polypropylene, polyurethane, ethylene vinyl acetate or acrylonitrile-butadiene-styrene.

6. The matrix according to any one of the preceding claims, wherein the antimicrobial agent is active against gram positive and/or gram negative bacteria.

7. The matrix according to Claim 6, wherein the antimicrobial agent is selected from the group comprising chloramphenicol, substituted isothiazolinones,

bisguanadines, eg chlorhexidine and organic peracids.

8. The matrix according to any one of Claims 1 to 5, wherein the antimicrobial agent is an anti-fungal agent.

9. The matrix according to Claim 8, wherein the anti-fungal agent is undecenoic acid or griseofulvin.

10. The matrix according to any one of Claims 1 to 5, wherein the antimicrobial agent is an anti-viral or algaecidal agent.

11. The matrix according to any one of the preceding claims, wherein the carrying agent is a polymer.

12. The matrix according to Claim 11, wherein the polymer is selected from the group comprising polyvinyl pyrolidone, coconut oil or polyethylene glycol.

13. The matrix according to Claim 12, comprising polyvinyl chloride, chlorhexidine and polyethylene glycol.

14. A process for preparing an antimicrobial matrix comprising mixing a support matrix with an antimicrobial agent and a carrying agent, wherein the carrying agent and the antimicrobial agent are adapted to form at least one hydrogen bond or salt bridge therebetween.

15. The process according to Claim 14, wherein the carrying agent is mixed with the antimicrobial agent, and the resulting mixture is added to the support matrix and kneaded until homogenous.

16. The process according to Claim 15, wherein a surfactant is added to the carrying agent and

antimicrobial agent.

17. A process according to any one of Claims 14 to

16, wherein the mixture is extruded and/cr formed or moulded.

18. An article incorporating an antimicrobial matrix according to any one of Claims 1 to 13.

19. An article comprising an antimicrobial matrix according to any one of Claims 1 to 13.

20. The article according to Claim 19, which

comprises a toilet seat.

21. The article according to Claim 19, which

comprises a seal and liner for a fridge or freezer.

22. The article according to Claim 19, which

comprises a shield for a telephone mouthpiece.

23. An antimicrobial matrix substantially as

hereinbefore described and with reference to the example.