WO1997030718A1

WO1997030718A1 - Antimicrobial composition

Info

Publication number: WO1997030718A1
Application number: PCT/AU1997/000092
Authority: WO
Inventors: Thomas Keith Stacey
Original assignee: Thomas Keith Stacey
Priority date: 1996-02-20
Filing date: 1997-02-20
Publication date: 1997-08-28
Also published as: AUPN815696A0

Abstract

An antimicrobial composition including the aqueous fraction collected following the steam distillation of the foliage of melaleuca plants and a process for preparing the antimicrobial composition including: a) supplying foliage from melaleuca (linerafolia) and/or melaleuca (alternafolia) plants to a sealed vessel; b) passing steam through the foliage in the sealed vessel to extract oil therefrom and collecting the steam thereafter; c) cooling the steam to form an oil phase and an aqueous antimicrobial composition, and d) separating the oil phase from the antimicrobial composition.

Description

ANTIMICROBIAL COMPOSITION

The present invention relates to a process for preparing antimicrobial compositions, to the antimicrobial compositions so prepared and to methods of using the antimicrobial compositions.

BACKGROUND

An antimicrobial composition is a composition which includes natural or artificial agents which prevent the growth of or kill microorganisms.

There are many different types of antimicrobial compositions that are known. Some compositions may only be effective to prevent the growth of or kill a limited number or type of microorganisms. However, there are also other well known antimicrobial composition which can prevent the growth of or kill a wide range of types of microorganisms.

There are also a large range of different processes for preparing antimicrobial compositions. Some of these processes are very complicated and some are very simple.

Applicant has, however, developed a new process for the preparation of an antimicrobial composition.

Applicant has now surprisingly found that a by-product formed during the extraction of oil from the foliage of melaleuca plants is a very effective antimicrobial composition. It has previously been known that tea tree oil (ie oil extracted from melaleuca plants) is effective as an antibactericide. However, the present invention does not relate to this oil. The present invention relates to the composition remaining after the oil component produced from the distillation of melaleuca plants has been extracted. This composition has unexpectedly and surprisingly been found to have a very effective antimicrobial activity at levels far greater than tea tree oil.

SUMMARY

According to one aspect of the present invention there is provided a process for preparing an antimicrobial composition including:

a) supplying foliage from melaleuca (linerafolia) and/or melaleuca (altemafolia) plants to a sealed vessel,

b) passing steam through the foliage in the sealed vessel to extract oil therefrom and collecting the steam thereafter,

c) cooling the steam to form an oil phase and an aqueous antimicrobial composition, and

d) separating the oil phase from the antimicrobial composition.

According to another aspect of the invention there is provided an antimicrobial composition including the aqueous fraction collected following the steam distillation of the foliage of melaleuca plants.

Further according to the invention there is provided a method of treating a viral, fungal or bacterial infection which includes administering an effective amount of the antimicrobial composition of the invention to the site of the infection.

DETAILED DESCRIPTION

The process for preparing the antimicrobial composition may be conducted in any suitable apparatus. In a preferred embodiment the process is conducted in a standard steam distillation apparatus. The quantity of foliage supplied in step a) of the process will depend on the size of the sealed vessel. The sealed vessel may have a 1600 litre capacity. However, vessels up to and above 5 tonne (5000 litres) may be used. If a sealed vessel of 1600 litre capacity is used in the process, then there is usually about 0.2 to 0.5 tonne of foliage supplied to the vessel.

The sealed vessel used in the process of the invention may be of any suitable design and construction such that the foliage can be contained in a steam tight environment. Further, the sealed vessel must be designed such that steam can pass through the foliage.

The steam may be generated within the sealed vessel. Alternatively, the sealed vessel may include a steam inlet and the steam may be supplied to the inside of the vessel from a boiler or steam generator which is connected to the steam inlet. The steam inlet is preferably positioned such that the steam may enter the vessel and pass evenly through all of the foliage supplied thereto. In most cases the steam inlet will be positioned in the base or a lower side wall of the sealed vessel.

The sealed vessel may also include a grid adapted to hold the foliage above the base of the vessel to enable an even flow of steam to pass through the foliage. The optimum position of the grid above the base of the vessel will depend upon the amount of foliage to be steamed and the size of the vessel. For a 1,600 litre vessel the grid is preferably positioned about 200 - 500 mm (usually about 300 mm) above the base of the sealed vessel.

The mesh size of the grid will depend upon the size of the foliage. In a preferred embodiment the foliage is chipped prior to being placed in the vessel to maximise the surface exposure of the foliage to the steam. In one embodiment of the invention the foliage is chipped to sizes of about 50 mm² to 75mm² and the mesh size of the grid is 50 mm². The foliage in the sealed vessel is subjected to steam for any suitable time such that the optimum amount of oil is extracted from the foliage. The time will be dependent on the size of the vessel, the quantity of foliage and the temperature and pressure of the steam. For example, for a 1600 litre capacity sealed vessel holding 0.5 tonne of foliage the foliage is generally steamed for two to three hours at a temperature of 100°C.

After the steam has passed through the foliage and the oil has been extracted therefrom the steam is collected. Preferably the sealed vessel is provided with an outlet in the top or an upper side wall through which the steam may pass and be collected.

Following collection of the steam it is then necessary for the steam to be cooled to form an oil phase and an aqueous antimicrobial composition. This may be achieved by any suitable means, for example it may be achieved by passing the collected steam through a standard condenser.

On cooling of the steam it forms two immiscible phases, an oil phase which forms an upper layer and an aqueous antimicrobial composition which forms a lower layer. The oil phase may be separated from the antimicrobial composition by any suitable means. In a basic system the oil phase is simply skimmed or decanted from the top of the antimicrobial composition. Alternatively the oil phase may be drained from the top of the antimicrobial composition or the antimicrobial composition may be drained from the bottom of the container in which the two immiscible phases were collected.

The antimicrobial composition prepared by the above method has been found to be very effective with killing or preventing the growth of a wide range of microorganisms. For example, the antimicrobial composition has been used in the treatment of viral, bacterial and fungal infections in mammals including both humans and domestic pets. In particular the antimicrobial composition has been found to be effective in the treatment of viral infections such as parvovirus and heφes. It has also been found to be effective in the treatment of fungal infections such as ringworm or tinea. In addition, it has also been found to be effective in the treatment of bacterial infections such as Escherichia coli, Staphylococcus aureus. Leqionella pneumophila and acne.

Any suitable amount of the antimicrobial composition may be used to prevent the growth of or kill microorganisms. The preferred amount of antimicrobial composition to be used will depend on the method of application and the number and type of microorganisms.

In a preferred embodiment of the invention the antimicrobial composition as prepared by the process is directly applied to a microorganism. This may be achieved by directly dabbing the area of infection with enough composition to form a thin layer thereover.

In other embodiments of the invention the antimicrobial composition may be used as a preventive composition by directly applying the composition to a cut or abrasion.

In yet another embodiment of the invention the anitmicrobial composition may be used as a relief composition by directly applying the composition to a painful site, such as an insect bite or sting, sunburn or other burns and as an after body waxing lotion.

Those skilled in the art will appreciate that the antimicrobial composition may have many other uses and that the above examples should be read as only some examples of uses for the composition.

The effectiveness of the antimicrobial composition will now be described with reference to some particular examples. However, it should be understood that the use of the composition is not limited to the treatment of the microorganisms referred to therein.

Example 1

Test Procedure:

The relative efficacy of the antimicrobial composition was compared with a known effective disinfectant, 2.5% gluteraldehyde and a distilled water control.

Materials and Methods

Materials:

Product: Antimicrobial composition prepared by the method of the invention.

Virus: Canine Parvovirus, strain DV I333-4 obtained from Prof. Margaret Sabine, University of Sydney.

Virus Indicator

System: FK continuous cell line/pig red blood cells (PBRC)

Artificial soil: 10 per cent Foetal Bovine Serum (FBS)

Disinfectant Inactivator System: Amrad-Pharmacia Dextran 2000 Blue Chromatography

Method

Product and controls were mixed 50:50 with virus suspension in culture medium containing 10% FBS at room temperature. At 10 and 30 minutes, samples were taken and passed through a column of Dextran 2000 Blue to remove residual disinfectant. Ten-fold dilutions were made and inoculated into FK cells to titrate surviving viruses. After 5 days incubation, virus end-points were determined by the presence/absence of haemagglutination of PRBC. Virus titres were calculated by the method of Reed and Muench and the relative efficacy of the different test preparations determined. A 4 log₁₀ reduction or better is required for a disinfectant claim to be made. This may be reduced to a 3 log₁₀ reduction were the virus indicator/inactivation system does not enable a 4 log₁₀ reduction to be demonstrated.

Results

Virus % kill titre, log₁₀

Water control 6.00

2.5% Gluteraldehude (10 min) <2.40 >99.97

(30 min) <2.40 >99.97

Product (10 min) 5.40 75.00

(30 min) 4.84 93.00

< = less than > ^; = greater than

Comments

There was a reduction in the virus titre brought about by the antimicrobial composition. This was a 0.60 log₁₀ (75%) reduction at 10 minutes and a 1.16 log₁₀ (93%) at 30 minutes. To make a disinfectant claim, there should be a 3-4 log₁₀ (99.9 - 99.99%) reduction in 10 minutes. This was observed with the gluteraldehyde positive control.

Example 2

Test organisms: The organisms used in this example were:

Escherichia coli NCTC 8196

Staphylococcus aureus NCTC 4168

MRSA: hospital strain resistant to penicillin, ampicillin, cephalosporin, tetracycline, gentamicin and streptomycin.

Inoculum 8

All innocula were 18 hour cultures with a cell density of approximately 10⁸ cells/mL.

Test Procedure

1. 0.1 mL of each of the test organisms was inoculated into: (i) 9.9mL antimicrobial composition

(ii) 9.9mL 0.1 % peptone water (control).

2. Mixtures were incubated at 37°C throughout the test period.

3. Mixtures were sampled 0, 15, 30 and 60 minutes, in order to enumerate viable organisms. Serial dilutions were prepared in 0.1% peptone water, and 1mL aliquots used to prepare pour plates with Plate Count Agar. Colonies were counted after incubation at 37°C for 48 hours.

Results S. aureus

Cell concentration % kill log

Control 5.8 - Product 15 min 5.2 75 30 min 0.6 99.999 60 min ND* >99.999

MRSA

Cell concentration % kill logio Control 6.5

15 min <4 >99 30 min 1 99.999 60 min ND* >99.999 E. Coli

Cell concentration % logio

Control 6.6

15 min 5.1 97

30 min 3.7 99.9 60 min ND* >99.999

^*ND : Not detectable

Comments

Under the test conditions employed, the antimicrobial composition was responsible for a reduction in cell count of each of the three test organisms. At 15 minutes, this was a 75% reduction for S. aureus, a Gram-positive bacterium, >99% for the multi-resistant strain of S. aureus (MRSA) tested and 97% for E. coli, a Gram-positive bacterium. A 5-6 Iog10 reduction was achieved for each of the test organisms after a contact time of 60 minutes with the antimicrobial composition.

Those skilled in the art will appreciate that there may be many variations and modifications of the process and compositions described herein which are within the scope of the present invention.

Claims

CLAIMS:

1. A process for preparing an antimicrobial composition including:

d) separating the oil phase from the antimicrobial composition.

2. A process according to claim 1 wherein steam is passed through the foliage for two to three hours.

3. A process according to claim 1 or claim 2 wherein steam is passed through the foliage at a minimum temperature of about 100°C.

4. An antimicrobial composition when prepared by the proce<=^<- of any one of claims 1 to 3.

5. An antimicrobial composition including the aqueous fraction collected following the steam distillation of the forage of melaleuca plants.

6. A method of treating a microbial infection which includes administering an effective amount of the antimicrobial composition as defined by claim 4 or 5, to the site of the infection.

7. A method of treating a bacterial infection which includes administering an effective amount of the antimicrobial composition as defined by claim 4 or 5, to the site of the infection.

8. A method according to claim 7 wherein the antimicrobial composition is dabbed directly on to the site of infection to form a thin layer thereover.