WO2005087244A1

WO2005087244A1 - Antimicrobial composition

Info

Publication number: WO2005087244A1
Application number: PCT/GB2005/000914
Authority: WO
Inventors: Jacqueline Stringer; Peter Andrew Warn
Original assignee: The University Of Manchester
Priority date: 2004-03-12
Filing date: 2005-03-10
Publication date: 2005-09-22
Also published as: GB0405545D0

Abstract

The present invention relates to compositions comprising at least two essential oils derived from a plant genus independently selected from: Pelargonium; Cymbopogon; Mentha; Aniba; Lavandula; Origanum; Litsea; Citrus; Melissa; Pogostemon; Santalum; Valeriana; Styrax; Cinnamonum; and Rosa, for treating microbial infections and preventing microbial contamination of surfaces and devices. The compositions may be formulated as medicaments, consumer products or cleaning products.

Description

ANTIMICROBIAL COMPOSITION

The present invention relates to antimicrobial compositions, and particularly to antimicrobial compositions containing essential oils. In particular, the invention extends to antibacterial and antifungal compositions, and to methods of using the compositions to treat bacterial and fungal infections.

The use of plants and plant products for their healing properties has been recognized, and numerous plant products are currently used in medicine. "Unconventional" therapy, for example aromatherapy, is gaining increasing public acceptability, with many people now regularly purchasing complementary or homoeopathic remedies. A growth in expenditure on unconventional therapy of 50% between 1995 to 2000, was seen in the UK. Moreover, such increasing expenditure on herbal medicines is a worldwide phenomenon, with sales in the USA rising from US$1.6 billion in 1994 to US$4 billion in 1998. The reasons for the growing trend for people to look to complementary therapies instead of, or as an adjunct to, orthodox medicine over the last few years are complex, but include anxiety about ever more powerful drugs and dissatisfaction with the perceived limitations of orthodox medicine. The international move towards increasing use of a wide variety of therapies, including essential oils, has occurred despite the fact that up until recently there has been minimal scientific research demonstrating objective benefits for such remedies.

Aromatherapy is the therapeutic use of essential oils, with the assumption that the oils have multiple effects resulting from the complex nature of the constituents of the oils. Whilst more and more evidence of ways in which such essential oils can potentially be used within the clinical setting is coming to light through scientific study, the integration of such potentially useful therapies into traditional medical care requires a methodological approach. Specifically, a systematic strategy is required to qualify, quantify and justify how, why and when essential oils may be most effectively used.

In hospitals today, patients, who may or may not be immunosuppressed, are often subject to acute and chronic fungal and bacterial infections, some of which may be life-threatening. For example, multi-resistant organisms such as: methicillin resistant Staphylococcus aureus (MRS A) and vancomycin resistant Enterococcus (VRE) are real problems in hospitals, causing high rates of infection in patients. Immunosuppressed patients, (e.g. those undergoing cancer therapy) are especially susceptible to such microbial infections. As well as adversely affecting the clinical outcomes of infected patients, such bacterial and fungal infection impacts substantially on the health economy through increased health resource utilization for management of infection.

In addition to the development of medicaments for managing infection, the inventors also recognize that there is a need to develop antimicrobial agents for formulation in products that may be used in a broader clinical sense or as a personal healthcare product (e.g. soaps, shampoos, massage oils and the like). Furthermore such antimicrobial agents may be used in cleaning products (e.g. cleaning products for cleaning medical devices, hospital wards, kitchens etc).

The inventors have realized that there is a need to critically evaluate the anti-infective potential of essential oils (whether by aromatherapy or formulated in a composition) for use as medicaments, personal healthcare products or cleaning products. In the medical context that have particularly focused on the use of such oils as anti-infective medicaments (e.g. used as an adjunct to conventional anti-cancer therapy). It is, therefore, an aim of embodiments of the present invention to address the above problems and to provide a composition containing essential oils, which exhibits antimicrobial efficacy.

The inventors carried out an in depth study to investigate the antimicrobial activity of 37 essential oils, which are listed in Tables 1-4. In particular, the inventors studied the in vitro antimicrobial effects of these 37 essential oils against a range of fungi and bacteria that are commonly implicated in diseases in both immuno-compromised patients, and also patients that are not immuno-compromised. To their surprise they found that a subset of the oils tested had some efficacy as antimicrobial agents. However to their greater surprise they found that certain combinations, or blends, of oils had good efficacy and even synergistic effects on bacteria and fungi. Some of the essential oils that were tested have been used in aromatherapy for treating ailments, for example, aching backs etc.

According to a first aspect of the present invention, there is provided a composition comprising at least two essential oils selected from a group of oils derived from a plant genus independently selected from: Pelargonium; Cymbopogon; Mentha; Aniba; Lavandula; Cinnamonum; Origanum; Litsea; Citrus; Melissa; Pogostemon; Santalum; Valeriana; and Rosa, for use as a medicament. According to a second aspect of the present invention, there is provided use of a composition comprising at least two essential oils selected from a group of oils derived from a plant genus independently selected from: Pelargonium; Cymbopogon; Mentha; Aniba; Lavandula; Cinnamonum; Origanum; Litsea; Citrus; Melissa; Pogostemon; Santalum; Valeriana; and Rosa, for the manufacture of a medicament for the treatment of a microbial infection.

The inventors have also established that a composition comprising at least one viscous essential oil and at least one volatile essential oil are particularly useful for application to the skin. It is therefore preferred that the medicament comprising at least one viscous essential oil and at least one volatile essential oil wherein the oils are selected from a group of oils derived from a plant genus independently selected from: Pelargonium; Cymbopogon; Mentha; Aniba; Lavandula; Cinnamonum; Origanum; Litsea; Citrus; Melissa; Pogostemon; Santalum; Valeriana; and Rosa.

To the inventor's surprise, when individual essential oils in accordance with the first aspect of the invention were tested on bacteria and/or fungi, they exhibited exceptional antimicrobial efficacy, when compared to a range of other essential oils (see the Example). The oils exhibited such efficacy that the inventors realised that compositions comprising two or more of the essential oils according to the first aspect of the invention may be usefully formulated for antimicrobial use in a variety of clinical and non-clinical situations. Hence, it is the inventor's belief that they have demonstrated a new medical indication for essential oils according to the invention.

Essential oils are volatile, aromatic oils, which may be extracted from plants either by steam or hydrodistillation. Different parts of plants can be used to obtain essential oils, including the flowers, leaves, seeds, roots, stems, bark, wood, etc. Most essential oils contain terpenes, and their oxygenated derivatives (terpenoids). hi addition, the oils may also comprise variable mixtures of phenols, quinines, flavones, tannins, coumarins, alkaloids and sugars.

The inventors have found that essential oils may be used for the treatment of a variety of microbial infections, for example, a fungal infection and/or a bacterial infection. The inventors have demonstrated that the essential oils in accordance with the present invention, have a broad spectrum of antibacterial and antifungal activity against clinically relevant pathogens as described in the Example. For example, multi-resistant organisms such as the gram positive bacteria Nancomycin Resistant Enterococcus (NRE) and Methicillin Resistant Staphylococcus (MRS A) are especially problematic in hospitals. This is because they are resistant to many of the commonly used antibiotics, and so it is very difficult to prevent infection of these organisms using such conventional antibiotic treatment regimes. Because of this, hospitals are forced to spend large amounts of money by using very high concentrations of expensive antibiotics in an attempt to prevent bacterial infections. However, due to their resistance to conventional antibiotics, bacterial infections of MRS A and NRE are never totally erradicated. Accordingly, it is particularly advantageous that the essential oils in accordance with the present invention show effective antibacterial activity against not just one, but both of these bacteria, NRE and MRS A.

Furthermore, it is also particularly surprising that the essential oils with the greatest activity against bacteria were also highly active against fungi. This is surprising because antibacterials and antifungals tend to have different modes of action.

For example, the medicament may be used to treat a filamentous fungus infection, such as, an infection of an Ascomycete. Examples of preferred genera of Ascomycete, which the medicament may be used to treat, may be independently selected from a group of genera consisting of Aspergillus; Candida; Cryptococcus; Coccidiodes; Encephalitozoon; Fusarium; Pneumocystis; Histoplasma; Blastomycetes; Pythium; Trichophyton; Mucorales; Rhizopus; Mucor; Rhizomucor; Absidia; Apophysomyces; Cunninghamella; Saksenaea; Scytalidium; Malassezia; Madurella; Pseudallescheria; Sporothrix; Basidiobolus; Penicillium; Acremonium; Alternaria; Bipolaris; Aureobasidium; Exophilia; Scedosporium; Xylohypha; Paecilomyces; and Scopulariopsis.

In a preferred embodiment, the medicament is used for the treatment of an infection of Aspergillus and/or Candida. The medicament may be used to treat an infection of an organism independently selected from a group of species consisting of Aspergillus flavus; Aspergillus fumigatus; Aspergillus nidulans; Aspergillus niger; Aspergillus parasiticus; Aspergillus terreus; Candida albicans; Candida rusei; Candida glabrata; Candida parapsilosis; Candida tropicalis; Candida guilliermondii; Cryptococcus neoformans; Coccidiodes immitis, Encephalitozoon cuniculi; Fusarium solani; Pneumocystis cariniijiroveci; Histoplasma capsulatum; Blastomycetes dermatitidis; Pythium insidiosum; Trichophyton mentagrophytesinterdigitale; Trichophyton tonsurans; Trichophyton rubrurn; Epidermophyton floccosum; Microsporum canis; Mucorales; Rhizopus microsporus; Mucor circinelloides; Mucor racemosus; Rhizomucor pusillus; Absidia corymbifera; Apophysomyces elegans; Cunninghamella elegans; Cunninghamella bertholletiae; Saksenaea; Scytalidium dimidiatum; Malassezia furfur; Madurella mycetomatis; Madurella grisea; Pseudallescheria boydii; Sporoihrix schenckii; Basidiobolus haptosporus; Penicillium marneffii; Acremonium strictum; Alternaria alternata; Bipolaris spicifera; Aureobasidium pullulans; Exophilia jeanselmei; Exoseroiiilum; Scedosporium apiospermum; Xylohypha bantiana; Paecilomyces lilacinus; and Scopulariopsis brevicaulis.

It is preferred that the medicament is used to treat an infection caused by an Aspergillus sp. The infection may be caused by an organism independently selected from a group of species consisting of Aspergillus fumigatus; Aspergillus nidulans; Aspergillus niger; Aspergillus terreus; and Aspergillus flavus. A preferred Aspergillus fumigatus species includes A.fumigatus AF293 (NCPF 7367). A preferred Aspergillus terreus species includes A.terreus (AT7130). A preferred Aspergillus niger species includes A.niger (AN8). Furthermore, a preferred Aspergillus flavus species includes A.fl vus (AFL128).

It is also preferred that the medicament is be used to treat an infection caused by a Candida Sp. The infection may be caused by an organism independently selected from a group of species consisting of Candida albicans; Candida krusei; Candida guilliermondii; Candida glabrata; and Candida tropicalis. A preferred Candida albicans species includes Calbicans FA1936. A preferred Candida krusei species includes C.krusei FA3657. In addition, a preferred Candida glabrata species includes C.glabrata FA8829. Furthermore, a preferred Candida tropicalis species includes C.tropicalis FA3099.

The medicament may be used for the treatment of a variety of conditions caused by fungal infections, including: superficial fungal infections, ringworm, tinea pedis, tinea cruris, tinea corporis, tinea capitis, onychomycosis, pityriasis versicolor, superficial candidosis, oral candidosis, vaginal candidosis, paronychia, Candida intertrigo, tinea nigra, black peidra, white piedra, mycetoma (Madura foot), Sporotrichosis, subcutaneous zygomycosis, basidiobolomycosis, conidiobolomycosis, systemic mycosis, histoplasmosis, coccidioidmycosis, blastomycosis, paracoccidioidmycosis, cryptococcosis (pulmonary and disseminated), systemic candidosis, systemic aspergillosis, invasive zygomycosis, sporotrichosis, allergic bronchopulmonary aspergillosis, aspergilloma, invasive aspergillosis, mucormycosis, farmers lung and pneumocystis pneumonia. The medicament may also be used for the treatment of a bacterial infection, which may be either a gram positive bacterium, or a gram negative bacterium. Surprisingly, the medicament may be used to treat infections of both gram positive and gram negative bacteria.

Examples of gram positive bacteria, which the medicament according to the invention may be used to treat includes Firmicutes, such as Bacilli, for example, Bacillus anthracis, or Clostridia, for example, Clostridium botulinum.

In a preferred embodiment, the medicament may be used to treat Bacillales, preferably, Staphylococcus, for example, Staphylococcus aureus. An especially preferred gram positive bacterial strain, which the medicament may be used to treat, includes S. aureus ATCC33594, which is a methicillin resistant strain. In addition, the medicament may be used to treat an infection of S. aureus ATCC9144, which is a strain recommended for testing hand wash solutions. Additional gram positive Bacillales with which the composition according to the invention are effective include Streptococci, for example, Streptococcus pyogenes.

An example of a gram negative bacterium, which the medicament may be used to treat includes Pseudomonadales. An example of Pseudomonadales includes Pseudomonas aeruginosa, for example, P.aeruginosa ATCC25668. Further examples of gram negative bacteria against which the medicament is effective may include Gammaproteobacteria, which may be independently selected from a group consisting of Enter obacteriales, Proteus, Serratai, Pasteurellales, and Vibrionales.

Preferred Enterobacteriales include Escherichia, for example, E.coli, such as E.coli ATCC10536. Preferred Proteus include Proteus mirabϊlis. Preferred Serratai include Serratia marcescens. Preferred Pasteurellales include Haemophilus influenzae. Preferred Vibrionales include Vibrio cholerae.

Further examples of gram negative bacteria against which the composition in accordance with the invention is effective may include Betaproteobacteria, including Neisseriales, for example, Neisseria gonorrhoeae. Further examples of gram negative bacteria against which the composition in accordance with the invention is effective may include Delta/epsilon subdivided Proteobacteria, including Campylobacterales, for example Helicobacter pylori. Further examples of gram negative bacteria against which the composition in accordance with the invention is effective may include Actinobacteria, for example Mycobacterium tuberculosis and Nocardia asteroides.

The medicament may be used for the treatment of a variety of conditions caused by bacterial infections, including: streptococcal sore throat, scarlet fever, impetigo, erysipelas, cellulites, necrotizing fascitis, bacteraemia, pyoderma, neonatal streptococcal disease, puerperal sepsis, pneumonia (including hospital acquired pneumonia), bronchitis, asthma, acne vulgaris, folliculitis, boils, furuncles, paronychia, mastitis, sticky eye styes, otitis extema, otitis media, sinusitis, septicaemia, carbuncles, toxic shock syndrome, staphylococcal scalded skin syndrome, meningitis, urinary tract infections, diarrhoea, stomach ulcer, dacryocystitis, blepharitis, conjunctivitis, bacterial keratitis, and nasal carriage of pathogenic organisms (Staphylococcus aureus for example). However, it will be appreciated that this list is not exhaustive, and that there are many other bacterial infections which may be treated by the medicament.

In a preferred embodiment, the essential oils may be derived from plant species independently selected from: Pelargonium graveolens; Cymbopogon martinii; Mentha piperita; Aniba rosaeodora; Lavandula angustifolia; Cinnamonum camphora; Origanum marjorana; Litsea cubeba; Citrus aurantium; Melissa ojficinalis; Pogostemon cablin; Santalum album; Valeriana offϊcinalis; and Rosa damascena.

The composition may comprise a mixture of Palmarosa, Patchouli, Rosewood, Ho Leaf, Ho Wood, Sandalwood, Geranium, Lavender, Melissa true, Rose Otto, Peppermint Bigarade, May Chang, Melissa True, Rose Otto, Majoram, Benzoin Resinoid and Valerian oils in any combination.

IThe composition may compriseat least two specific oils selected from:

(1) An oil derived from Cymbopogon martinii var. This is a martinii species of the graminaceae family. The oil may comprise approx. 76-84% Geraniol; approx. 5-11% Nerol; approx. 2-5% Linalol; and under approx. 2% Gamma Terpinene. The oil is also known as Comoros Palmarosa. (2) A preferred oil is also derived from the Pelargonium graveolens species of the graminaceae family. The oil may comprise approx. 3.5-8% Geraniol; approx. 5% Iso menthone; approx. 2-5% Linalol; and approx. 40% Citronellol. The oil is also known as Yumian Geranium.

(3) Another preferred oil is derived from the Citrus aurantium var. amara species of the rutaceae family. The oil may comprise up to approx. 18% Beta Pinene; up to approx. 16% Limonene; approx. 30-40% Linalool; approx. 5% linalyl acetate; up to approx. 5% Alpha terpineol; and approx. 3% Geranyl acetate. The oil is also known as Neoli Bigarade.

(4) Another preferred oil is derived from the Pogostemon cablin species of the lamiaceae family. This oil may comprise up to approx. 60% Patchouli sesquiterpenes (including approx. 16%) alpha guaiene; approx. 7 % seychellene; approx. 6.4% alpha patchoulene; and approx. 16.4% Delta guaiene); and approx. 21-40% patchouliol. The oil is also known as Indonesian Patchouli.

(5) Another preferred oil is derived from the Mentha piperita species of the lamiaceae family. This oil may comprise approx. 11-31% menthone; approx. 30-45% menthol; approx. 3- 8% iso menthone: approx. 2-5% beta caryophyllene; and approx. 2% beta pinene. The oil is also known as Chinese Peppermint.

(6) Another preferred oil is derived from the Rosa damascena species of the rosaceae family. This oil may comprise up to approx. 50% citronellol; approx. 15% geraniol; approx. 3- 8% nerol; approx. 1-3% phenylethyl alcohol; approx. 5-10% nonadecane; and under approx. 5% farnesol. The oil is also known as Turkish Rose Otto.

(7) Another preferred oil is derived from the Aniba rosaeodora species of the lauraceae family. This oil may comprise less than approx. 1% alpha terpinene; up to approx. 2% beta pinene; approx. 78-90% linalool; and approx. 0.5-3% cineole-1.8. The oil is also known as Brazilian Rosewood.

(8) Another preferred oil is derived from the Litsea cubeba species of the lauraceae family. This oil may comprise approx. 66-76% citral; approx. 10-15% cineol 1.8; and up to approx. 3% limonene. The oil is also known as Chinese May Chang.

(9) Another preferred oil is derived from the Melissa officinalis species of the lamiaceae family. This oil may comprise approx. 50-60% citral; approx. 10% citronellal; under approx. 10% caryophyllene; under approx. 5% geraniol; and under approx. 5% geranyl acetate. The oil is also known as Irish Melissa True.

(10) Another preferred oil is derived from the Lavandula angustifolia species of the lamiaceae family. This oil may comprise over approx. 30% Linalyl acetate; up to approx. 5.5% Alpha terpineol; approx. 2-13% Terpinene - 4 - ol; approx. 34-45% Linalool; up to approx. 6% Camphor. The oil is also known as French Lavender.

(11) Another preferred oil is derived from the Origanum marjorana species of the lamiaceae family. This oil may comprise approx. 20-30% Terpinene-4-ol; approx. 5-15% Gamma terpinene; up to approx. 9% Sabinene; up to approx. 6% Alpha terpineol; approx. 10% Para cymene; up to approx. 20% Linalool; up to approx. 15% Alpha terpinene. The oil is also known as Sweet French Marjoram.

(12) Another preferred oil is derived from the Santalum album species of the santalaceae family. This oil may comprise approx. 47% Alpha santalol; approx. 20% Beta santalol. The oil is also known as East Indian Sandal Wood.

(13) Another preferred oil is derived from the Valeriana ofβcinalis species of the valerianaceae family. This oil may comprise approx. 30-38% Caryophyllene; approx. 20-25% Camphene; approx. 5-8% Alpha pinene; approx. under 5% Beta pinene; approx. under 2% Limonene. The oil is also known as Croatian Valerian.

(14) Another preferred oil is from the Linalool trees amd is known as Ho Wood Leaf or Ho Leaf oil. This oil may comprise linalool, 1,8 cineole, euginol, camphor, caryphyllene, 4- terpineol and alpha-terpineol

The composition may comprise at least two of the abovementioned oils. The composition may comprise at least three or at least four of the above-mentioned essential oils.

Surprisingly, the inventors noticed that combinations of such essential oils according to the invention may be combined to have a synergistic effect and improved antimicrobial efficacy.

For example, the composition may comprise a mixture of Rosewood, Palmarosa and Geranium oil, in any combination. Alternatively the composition may comprise a mixture of Palmarosa, Patchouli, Rosewood, Ho Leaf, Howood, Sandalwood, Geranium, Lavender, Melissa true, Rose Otto and Peppermint oils in any combination.

Preferred blends of two essential oils comprising blends of two oils selected from: Geranium, Patchouli, Rosewood, Peppermint, Lavender, Palmarosa or Ho leaf or Howood oils. Most preferred blends of two oils comprise Palmarosa and one of Geranium, Peppermint, Lavender and Rosewood oils. Such blends may comprise each of these oils in substantially equal proportions or the proportions may be varied in view of the viscosity required or the body site and condition to be treated.

Preferred blends of three oils comprise Geranium, Patchouli and one of Rosewood, Palmarosa or Ho leaf oils. A most preferred blend of three oils comprises Geranium, Patchouli and Rosewood oils. Such a blend may comprise each of these oils in substantially equal proportions or the proportions may be varied in view of the viscosity required or the body site and condition to be treated.

A skilled person will appreciate that the blend of oils used may be dictated by the body site to be treated and also the type of condition being treated. By way of example, Peppemiint oil, when blended with at least one further oil according to the present invention, is useful for gut orientated infections.

The inventors have demonstrated that such mixtures of oils surprisingly augment the activity of each other in respect of inhibition of microbial growth and particularly multi-drug resistant micro-organisms (see the Examples).

An important advantage of combinations of two or more essential oils according to the invention is that development of resistance to the agents by bacteria or fungi is less likely, as resistance would have to develop to all the components simultaneously.

Another important advantage of combinations of two or more essential oils according to the invention is that they are effective for inhibition of the growth of multi-drug resistant microorganisms that have already developed a resistance to most or all known conventional antimicrobial agents.

In an preferred embodiment, there is provided use of a composition comprising at least two essential oils derived from a group of plants consisting of Lavandula angustifolia; Cinnamonum camphora; Pelargonium graveolens; Origanum marjorana; Litsea cubeba; Cymbopogon martini; Mentha piperita; Aniba rosaeodora; Citrus aurantium; Melissa ofβcinalis; Styrax benzoin; Litsea cubeba; and Rosa damascena in the manufacture of a medicament for the treatment of a fungal infection (e.g. a Candida or Aspergillus infection). It is preferred that two or more of such oils are used to treat fungal infections. The oils may, for example, be the specific oils from these plants as disclosed above.

In another preferred embodiment, there is provided use of a composition comprising at least two essential oils derived from a group of plants consisting of Cymbopogon martini; Pogostemon cablin; Mentha piperita; Santalum album; Citrus aurantium; Melissa ofβcinalis; Cinnamonum camphora; and Rosa damascena for the manufacture of a medicament for the treatment of a gram positive bacterial infection. The oils are particularly effective against multidrug resistant infections. Accordingly, the gram positive bacteria may include Vancomycin Resistant Enterococcus; Methicillin Resistant Staphylococcus; Staphylococcus aureus; or Bacillus cereus. It is preferred that two or more of such oils are used to treat gram positive infections. The oils may, for example, be the specific oils from these plants as disclosed above. Advantageously, these essential oils (used singularly or preferably in combination) show antimicrobial activity against more than one gram positive strain (e.g. VRE and MRSA), which are resistant to conventional antibiotics, and which are a common cause of infection in hospitals.

In a still further preferred embodiment, there is provided use of a composition comprising at least two essential oils derived from a group of plants consisting of Eucalyptus globus; Cymbopogon martini; Mentha piperita; Aniba rosaeodora; Citrus aurantium; Cinnamonum camphora; and Melissa ofβcinalis for the manufacture of a medicament for the treatment of a gram negative bacterial infection. The gram negative bacteria may include Stenotrophomonas maltophilia; Pseudomonas aeroginosa; E.coli; and Klebsiella pneumoniae. It is preferred that two or more of such oils are used to treat gram negative infections. The oils may, for example, be the specific oils from these plants as disclosed above.

The inventors have found that essential oils derived from Cymbopogon martini; Melissa officinalis; Mentha piperita; and Citrus aurantium are active against fungi and/or gram positive bacteria and/or gram negative bacteria. Such oils are especially preferred for use as antimicrobial agents either on their own or in combination with each other. In addition, surprisingly, oil derived from Aniba rosaeodora is effective against fungal infections and gram negative bacteria, and oil derived from Rosa damascena is effective against fungal infections and gram positive bacteria.

The inventors have found that the composition defined by the first aspect of the invention may, as well as being suitable for use as an anti-infective medicament, may be used as consumer products. Therefore according to a third aspect of the invention there is provided a composition comprising at least two essential oils selected from a group of oils derived from a plant genus independently selected from: Pelargonium; Cymbopogon; Mentha; Aniba; Lavandula; Cinnamonum camphora; Origanum; Litsea; Citrus; Melissa; Pogostemon; Santalum; Valeriana; and Rosa for use as a consumer product.

By "consumer product" we mean products (such as body washes, shampoos, soaps, cosmetics, moisturizers, massage oils, sun-block, tooth pastes, mouth washes, chewing gums and the like) that are used by the public and are not necessarily regulated by the same authorities which control the marketing of medicaments. Such products are preferably healthcare consumer products (e.g. so called medicated shampoos and the like) and may be used by the consumer to prevent, or even treat, microbial infections.

It will be appreciated that consumer products according to the invention may comprise conventional formulations (e.g. of soaps, shampoos and the like) that are characterised by the inclusion of at least two of the essential oils as defined above.

According to prefened embodiments of the invention the consumer product or medicament may be applied to the skin of a subject to treat or prevent infections of the dermis. When this is the case the medicament or product is preferably a gel, ointment, cream, body wash, shampoo, soap or the like. Such medicaments or products may also be in the form of a neat blend or oils or may comprise such oils diluted in a suitable diluent for massage into the skin. Medicaments and products massaged into the skin may comprise between 0.5 - 100° o oils. It is preferred that the composition comprises less than 25% of the oils, preferably 0.5 — 10% and more preferably 1 - 3% (e.g. 2.5% oil)

It is most preferred that medicaments and products for application to the skin comprise at least one volatile oil and at least one viscous oil. The inventors have demonstrated that these blends have good efficacy and also act for a long time period following administration. In this respect Patchouli and Sandalwood are examples of a viscous oil whereas Rosewood, Ho Leaf and Palmarosa are considered to be volatile.

The inventors have realised that essential oils according to the invention may be put to a number of other antimicrobial uses (whether in a clinical context or otherwise). For instance, in addition to administering medicaments or consumer products to a subject, they may be used to coat surfaces and objects to prevent or treat bacterial or fungal contamination.

Therefore, in a fourth aspect there is provided a method of preventing and/or treating a microbial contamination comprising coating an object or a surface in need thereof with an amount of at least two essential oils as defined by the first or second aspects of the invention, that is effective for killing or preventing growth of micro-organisms

The micro-organisms are preferably bacteria or fungi as defined above.

According to a fifth aspect of the invention there is provided a composition comprising at least two essential oils selected from a group of oils derived from a plant genus independently selected from: Pelargonium; Cymbopogon; Mentha; Aniba; Lavandula; Cinnamonum camphora; Origanum; Litsea; Citrus; Melissa; Pogostemon; Santalum; Valeriana; and Rosa as a cleaning product.

By "cleaning product" we mean products used to clean surfaces in place such as the household, kitchens and particularly hospitals (e.g. wet wipes, tissue and cloths impregnated with oils, cleaning solutions, creams and gels etc). Such products may also be used to clean/sterilise objects that, if contaminated with micro-organisms, might cause infections in a humans or other animals of veterinary relevance (e.g. horses, cattle, pigs an sheep). Furthermore such products may be used to clean clothes and even the body (e.g. the hands) of people whose jobs, if they do not follow strict cleanliness guidelines, could put other people or animals at risk of being infected (e.g. doctors, nurses, butchers, farmers, kitchen workers, nursery workers etc)

It will be appreciated that the essential oils may be particularly useful for coating surfaces or objects that are required to be aseptic. As discussed above, many of the compositions have the advantage that they are both antifungal and antibacterial. Accordingly the polypeptide will have a broad anti-microbial effect.

The oils may be used to coat any object or device which is used in a biological or medical situation, such as a medical device, and for which it may be important to prevent bacterial or fungal contamination that may lead to any infection in a patient. Examples of medical devices that may be coated according to the invention include catheters, stents, wound healing dressings, contraceptive devices, surgical implants and replacement joints.

The oils may be used to coat surfaces in environments that are required to be aseptic. For instance the oils may be used in medical environments. The oils may be formulated in cleaning products used to keep hospital wards clean. They may be used to clean surfaces of equipment (e.g. operating tables) in operating theatres as well as theatre walls and floors. The inventors believe the oils will be useful to improve sterility in general and also to address the spread of VRE and MRSA in particular.

The oils may be formulated into solutions for cleaning objects and surfaces. For instance, they may be a routine constituent of cleaning solutions, soaps and handwashes.

It will be appreciated that the above list of objects and surfaces to which oils according to the invention may be applied is not exhaustive. Hence, the oils may be administered to any surface, which is prone to bacterial or fungal contamination, for example, kitchen and bathroom surfaces.

It will be appreciated that the composition according to the present invention may be used alone or in combination with other antimicrobial agents. For instance, medicaments according to the first or second aspect of the invention may be used in a monotherapy (i.e. use of the medicament according to the invention alone to prevent and/or treat a microbial infection). Alternatively, the medicament according to the invention may be used as an adjunct, or in combination with known therapies.

Suitable formulations for topical administration (as a medicament, consumer product or cleaning product) include solutions, suspensions, jellies, gels, creams, ointments, sprays, foams, powders, liposomes, pastilles, chewing gums, toothpastes and mouth washes. In the case of topical application to the scalp the medicaments may be formulated as sbampoos.

Most preferred medicaments are used in the treatment of skin conditions (e.g ringworm or athlete's foot).

In the case of microbial infections of the lungs (e.g. Aspergillvis) medicaments may be formulated as, for example, nasal drops, intranasal sprays or aerosols for inhalation.

In the case of topically applied compositions to be applied to the; skin the vehicle used to carry the oil may need to be in a vehicle capable of crossing the keratinous layer of the skin (e.g. if a medicament is used to treat an infection below the dermis).

Liquid preparations for oral application (e.g. as a mouthwash to prevent oral infections) represent a preferred form in which medicaments according to the invention may be formulated for oral administration. Suitable forms of liquid preparations include elixirs, syrups or suspensions. Such liquid preparations may comprise from about 0.05% to 25% or more by weight of the essential oil and may further comprise ingredients such as ethanol, water, glycerol, propylene glycol, sugar/sweeteners, and flavourings etc. Such mouth washes may be used to rinse the mouth for at least 10 seconds, preferably at least 20 seconds a_nd then either spat out or swallowed.

When used as a prophylactic (e.g. before there is a high risk of infection e.g. peri- operatively) the medicaments or consumer products of the invention sliould be administered as soon as the risk of infection has been recognised. For instance, it may be preferred to administer the medicament at the time of surgery, or in the hours or days preceding the treatment. Patients may be asked to wash in a soap or body-wash, comprising the oils, befoare being treated.

Medicaments manufactured according to the invention may be formulated such that they provide a daily dose of up to 500mg of the oil per kilogram bodyweight to a person receiving the medicament. Preferably the medicaments may be formulated such that they provide a daily dose of up to 250 mg per kilogram bodyweight, more preferably up to 125mg per kilogram bodyweight, even more preferably up to 62.5mg per kilogram bodyweight, more preferably still 30mg, 15mg or 5mg per kilogram, and most preferably lmg per kilogram. The preferred frequency of administration will depend upon the biological half-life of the selected oil. Typically a medicament in accordance with the invention should be administered to a target tissue such that the concentration of the oil is maintained at a level suitable to achieve a therapeutic effect. This may require administration daily or even several times daily.

Known procedures, such as those conventionally employed by the pharmaceutical and consumer product industries (e.g. in vivo experimentation, clinical trials etc), may be used to establish specific formulations of compositions and precise therapeutic regimes (such as daily doses of the oil and the frequency of administration).

In accordance with a sixth aspect of the invention, there is provided a method of preventing and/or treating a microbial infection, comprising administering to a subject in need of such treatment a therapeutically effective amount of a composition comprising at least two essential oils derived from a plant genus independently selected from: Pelargonium; Cymbopogon; Mentha; Aniba; Lavandula; Cinnamonum camphora; Origanum; Litsea; Citrus; Melissa; Pogostemon; Santalum; Valeriana; and Rosa.

The at least two essential oils may be derived from a plant species independently selected from: Pelargonium graveolens; Cymbopogon martinii; Mentha piperita; Aniba rosaeodora; Lavandula angustifolia; Cinnamonum camphora; Origanum marjorana; Litsea cubeba; Citrus aurantium; Melissa ofβcinalis; Pogostemon cablin; Santalum album; Valeriana ofβcinalis; and Rosa damascena.

It is preferred that cleaning products, as well as some medicament and consumer products, further comprises at least one surface-active agent. Such surface- active agents help the composition to spread over and/or coat a surface or object to which the composition is applied. The surface-active agent may be a surfactant or the like. The composition may be a shampoo and the vehicle and/or surface-active agent suitable for shampooing hair.

Alternatively the composition may be a cleaning composition, for example, a soap or handwash, and the vehicle and/or surface active agent suitable for cleaning the skin as an antimicrobial wash or scrub. In this case the oil may be formulated as a solid (e.g. a soap bar) or as a liquid (e.g. a liquid hand or body wash). Such compositions are particularly useful for use where sterility is important. For instance, doctors, surgeons, nurses, dentists, vets, cooks, people involved in the manufacture of food, nursery nurses and the like may advantageously use such compositions to reduce the risk of microbial contamination.

The composition may be a solution for cleaning surfaces. For instance the essential oil may be used to help clean/sterilize operating theatres, hospital wards, kitchen suαrfaces etc. However, it will be appreciated that the composition according to the invention! may be administered to any surface, which is prone to a microbial infection. Such compositioixs may also comprise detergents, bleaches, dyes and perfumes as is conventional for such cleaning solutions.

Solutions containing essential oils may also be used to help clean/sterilize lenses, catheters, stents, surgical instruments (such as scalpels), wound healing dressings, conixaceptives and the like.

The composition is also preferably used in coating objects such as medical devices and other biomaterials to prevent growth of broad categories of bacteria and fungi. For instance, an artificial joint used as a hip replacement may be coated in a gel comprising one or more essential oils according to the invention in order to prevent fungal or bacterial contamination of the device. The device may be stored in contact with the essential oil or the oil may be applied at a later date (e.g. prior to implantation).

All of the features described herein (including any accompanying claims, abstract and drawings), and/or all of the steps of any method or process so disclosed, may be combined with any of the above aspects in any combination, except combinations where at least some of such features and/or steps are mutually exclusive.

Embodiments of the invention will now be further described, by way of example only, with reference to the following Examples and Figures in which: -

Figure 1 is table showing antifungal activity of essential oils in accordance with an embodiment of the invention against Candida spp. and Aspergillus spp.;

Figure 2 is a table showing antibacterial activity of essential oils in accordance with the invention against gram positive and gram negative bacteria and fungi;

Figure 3 is a table showing antibacterial activity of essential oils in accordance with the invention against gram positive organisms; Figure 4 is a table showing antibacterial activity of essential oils in accordance with the invention against gram negative organisms;

Figures 5a & 5b are photographs of disc diffusion assays of essential oils in accordance with the invention.

Figure 6 illustrates survival of E. coli when exposed to 1% solutions of patchouli oil (O), rosewood oil(B), geranium oil(Δ); a 1% blend of all three oils (♦); or a control solution containing no oil (*);

Figure 7 illustrates survival of Staphylococcus aureus when exposed to 1% solutions of patchouli oil (O), rosewood oil(___), geranium oil(Δ); a 1% blend of all three oils (♦); or a control solution containing no oil (*);

Figure 8 illustrates survival of MRS A when exposed to 1% solutions of patchouli oil (O), rosewood oil(B), geranium oil(Δ); a 1% blend of all three oils (♦); or a control solution containing no oil (*);

Figure 9 illustrates survival of Vancomycin resistant enterococcus (VRΕ) when exposed to 1% solutions of patchouli oil (O), rosewood oil(H), geranium oil(Δ); a 1% blend of all three oils (♦); or a control solution containing no oil (*);

Figure 10 illustrates survival of Bacillus cereus when exposed to 1% solutions of patchouli oil (O), rosewood oil(B), geranium oil(Δ); a 1% blend of all three oils (♦); or a control solution containing no oil (*);

Figure 11 illustrates survival of Klebsiella pneumoniae when exposed to 1% solutions of patchouli oil (O), rosewood oil(B), geranium oil(Δ); a 1% blend of all three oils (♦); or a control solution containing no oil (#);

Figure 12 illustrates survival of Pseudomonas aeruginosa when exposed to 3% solutions of patchouli oil (O), rosewood oil(H), geranium oil(Δ); a 3% blend of all three oils (♦); or a control solution containing no oil (*);

Figure 13 illustrates survival of Candida albicans when exposed to 1% solutions of patchouli oil (O), rosewood oil(_B), geranium oil(Δ); a 1% blend of all three oils (♦); or a control solution containing no oil (*); Figure 14 is a grid illustrating an idealised score from a plate inhibition assay for combinations of two agents that act synergistically;

Figure 15 is a grid illustrating an idealised score from a plate inhibition assay for combinations of two agents that act antagonistically;

Figure 16 is a grid illustrating scores from a plate inhibition assay examining the effect of Geranium Oil and Palmarosa Oil on the growth of a strain of Candida albicans;

Figure 17 is a grid illustrating scores from a plate inhibition assay examining the effect of Rosewood Oil and Palmarosa Oil on the growth of a strain of Candida albicans;

Figure 18 is a grid illustrating scores from a plate inhibition assay examining the effect of Peppermint Oil and Palmarosa Oil on the growth of a strain of Candida albicans;

Figure 19 is a grid illustrating scores from a plate inhibition assay examining the effect of Lavender Oil and Palmarosa Oil on the growth of a strain of Candida albicans;

Figure 20 is a grid illustrating scores from a plate inhibition assay examining the effect of Geranium Oil and Palmarosa Oil on the growth of a strain of Aspergillus fumigatus;

Figure 21 is a grid illustrating scores from a plate inhibition assay examining the effect of Rosewood Oil and Palmarosa Oil on the growth of a strain of Aspergillus fumigatus;

Figure 22 is a grid illustrating scores from a plate inhibition assay examining the effect of Peppennint Oil and Palmarosa Oil on the growth of a strain of Aspergillus fumigatus; and

Figure 23 is a grid illustrating scores from a plate inhibition assay examining the effect of Lavender Oil and Palmarosa Oil on the growth of a strain of Aspergillus fumigatus.

Aspergillus fumigatus

blends of Palmarosa with one of Geranium, Peppermint, Rosewood and Lavender (compositions according to the invention) all had a synergistic effect on the survival of the fungi (see figures 16 - 23). EXAMPLE 1

Essential Oils

The essential oils used in this example were provided by Natural Touch Aromatherapy, Hampshire, UK. The same batches of oils were used in all experiments (additional batches were tested and had similar results, data not shown). All oils were stored in sealed bottles in the dark at 4°C. The quality and purity of all oils was confirmed by gas chromatography by the suppliers.

A number of oils were tested for antimicrobial activity. Details of the oils are as follows:-

Oil of Cymbopogon martinii (Comoros Palmarosa) - 78.46%) Geraniol; 6.8% Nerol; 5.86% Linalol; 1.63% Gamma Terpinene.

Oil of Pelargonium graveolens (Yunnan Geranium) - 6.3% Geraniol; 4.8% Iso menthone; 2.4% Linalool; 40.3% Citronellol.

Oil of Citrus aurantium (Nerolibigarade) - 16.0% Beta pinena; 14.0% Limonene; 34.0% Linalool; 5.5% Linalyl acetate; 4.1% Alpha terpineol; 2.6% Geranyl acetate.

Oil of Pogostemon cablin (Indonesian Patchouli) - 55.3% Patchouli sesqulterpenes including Alpha guaiene, Seycellene, Alpha patchoulene, Delta guaiene; 30.5% Patchouliol.

Oil of Mentha piperita (Chinese Peppermint) - 25.9% Menthone; 36.3%ι Menthol; 8.0% Iso menthone; 5.8% Beta caryophyllana; 1.5% Beta pinene.

Oil of Rosa damascena (Turkish Rose Otto) - 37.6% Citronellol; 17.2% Geraniol; 9.0% Nerol; 1.5% Phenylethyl alcohol; 9.2% Nonadecane; 1.4% Famesol.

Oil of Aniba rosaeodora (Brazilian Rosewood) - less than 1% Alpha terpinene; less than 1% Beta pinene; 81.7% Linalool; less than 1% Cineole -1.8.

Oil of Litsea cubeba (Chinese May Chang) - 69.9%> Citral; 11% Cineol 7.8; 1.4% Limonene.

Oil of Melissa ofβcinalis (Irish Melissa True) - 55.5% Citral; 8.2% Citronellal; 5.3% Caryophyllene; 4.5% Geraniol; 4.9% Geranyl acetate. Oil of Valeriana ofβcinalis (Croatian Nalerian) - 35% Caryophyllene; 24.1% Camphene; 7.7% Alpha pinene; 4.5% Beta pinene; 1.5% Limonene.

Oil of Styrax benzoin (Siam Benzoin Resinoid) - 73% Coniferyl benzoate; 15%> Coniferyl alcohol; 6% RT at c.49 (vanillin).

Oil of Citrus bergamia (Sicilian Bergamot) - 3.8% Beta pinene; 31.5% Limonene; 2.7% Gamma terpinene; 13.8% Linalool; 22.8% Linalyl acetate.

Oil of Piper nigrum (Indian Black Pepper) - 4.7% Alpha pinene; 8.3%) Beta pinene; 9.1% Sabinene; 16.6% Limonene; 22.9% Beta caryophyllene.

Oil of Elettaria cardamomum (Guatemalan Cardamom) - 37.4% Terpinyl acetate; 36.7% 1.8 Cineole; 4.3% Beta pinene; 4.2% Linalool; 3.8% Linalyl acetate.

Oil of Daucus carota (French Carrot Seed) - 33.09% Carotol; 16.47% Alpha pinene; 3.12% Beta pinene; 8.38%> Sabinene.

Oil of Cedrus atlantica (Atlas Cedarwood) - 17.4% Alpha Himachalene; 41.7% Beta Himachelene; 10.9% Gamma Himachelene; 1.5% Alpha Atlantone.

Oil of Salvia sclarea (French Clary Sage) - 21.2% Linalool; 1.8% Alpha terpineol; <1% Camphor; 58.7% Linalyl acetate; 1.4% Geranyl acetate.

Oil of Cupressus sempervirens (French Cypress) - 56.4% Alpha pinene; 19.4% RT c.ll (d-3- carene); 2.3%_> Myroene; 2.6% Limonene; 2.3% Alpha terpineol.

Oil of Eucalyptus globus (Spanish Eucalyptus) - 87.9% Cineol - 1.8; 3.6% Alpha pinene; 1.9% P-cymene.

Oil of Lavandula angustifolia (French Lavender) - 34.4% Linalyl acetate; 1.3% Alpha terpineol; 10.8% Terpinene - 4 - ol; 34.4% Linalool; <1% Camphor.

Oil of Boswellia carterii (Frankincense (Olibanum) - 49.1% Alpha - pinene; 4.9% Beta pinene; 3.4%o P-cymene; 4.0% Myrcene; 13.7% Limonene.

Oil of Zingiber officinale Chinese Ginger) - 41.8% Zingberene; 2.0% Alpha pinene; 6.4% Camphene; 7.5% Cineol - 1.8; 16.2% Farnesene; 6.3% Ar curcumene. Oil of Citrus paradisi (Brazilian Grapefruit) - 90.93% Limonene; 1.92% Myrcene.

Oil of Juniperus communis (French Juniper Berry) - 45.7% Alpha pinene; 2.1%o Beta pinene; 12.8%) Sabinene; 1.3% Terpinene - 4 - ol; 12.7% Myrcene.

Oil of Citrus limon (Spanish Lemon) - 63.04%> Limonene; 1.77% Alpha pinene; 10.86% Beta pinene; 1.19% Sabinene; 6Λ6% Gamma terpinene; 1.45%> Myrcene.

Oil of Citrus reticulata (Sicilian Mandarin) - 2.64% Alpha pinene; 1.75% Sabinene; 1.85% Myrcene; 1.51% P-cymene; 71.95% Limonene; 15.92%> Gamma - terpinene.

Oil of Origanum marjorana (Sweet French Marjoram) - 22.03% Terpinene-4-ol; 12.79% Gamma terpinene; 8.65%o Sabinene; 3.69% Alpha terpineol; 2.92% Para cymene; 15.24% Linalool; 8.21% Alpha terpinene.

Oil of Commiphora myrrha (African Myrrh) - 46% ISO Furano Germacrene; 28%> Furanodlene; 3.9%) Alpha cedrene; 3.00% Beta cedrene.

Oil of Myristica fragrans (Indonesian Nutmeg) - 21.9% Alpha pinene; 20.3% Sabinene; 15.3% Beta pinene; 5.0% Terpinene-4-ol; 8.7 Myristicin.

Oil of Rosmarinus ofβcinalis (Wild Spanish Rosemary) - 11.4% Alpha pinene; 40.5% Cineol- 1.8; 8.7%) Beta pinene; 12.6% Camphor; 4.6% Camphene; 4.9% Bomyl acetate.

Oil of Nardostachys jatamansi (Indian spikenard) - 11.73% Valeranone; 7.34%o Valerenal; 16.08% Calarene; 3.58% Valencene.

Oil of Melaleuca alternifolia (Australian Tea Tree) - 20.51% Gemma terpinene; 42.00% Terpinene-4-ol; 8.26% Alpha terpinene; 1.15% Cineol-1.8.

Oil of Jasminum officinale (Egyptian Jasmine Absolute) - 6.7% Linalool; 30.0% Benzyl acetate; 3.2% cis-Jasmene; 2.9% Eugenol; 6.6% iso-Phytol; 11.9% Phytol.

Oil of Matricaria recutica (German Chamomile) - 31.90% Alpha bisabolol; 5.38% Chamazulene; 3.04%> Famesene; 4.00% cis-en-in-dicyloether.

Oil of Achillea millefolium (Moroccan Yarrow) - 18.2% Sabinene; 16.1% Camphor; 7.9% Beta pinene; 6.8% Myrcene; 2.7% Chamazulene. Oil of Santalum album (East Indian Sandal Wood) - 49.36%> Alpha santalol; 24.96% Beta santalol.

Fungal and Bacterial strains

Fungi were isolated from recent clinical samples obtained from patients with a range of invasive diseases including septicaemia, wound infections, urinary tract infections and pulmonary aspergillosis. The fungal strains tested included Candida albicans FA1936, C. tropicalis FA3099, C. glabrata FA8829, C krusei FA3657, Aspergillus fumigatus AF293 (NCPF 7367), A. terreus AT7130, A. niger AN8 and A. flavus AFL128. The isolates were subcultured from frozen stock and were grown on Sabouraud dextrose agar (Difco, Surrey, UK) at 37°C in air for up to 10 days.

Bacteria were isolated from recent clinical samples obtained from patients with a range of invasive diseases or were obtained from reference strains used within the Microbiology Laboratory at Salford Royal Hospital, Manchester UK. The bacterial strains tested included methicillin resistant Staphylococcus aureus ATCC33591 (MRSA), which is a strain recommended for testing hand wash solutions, Oxford S. aureus ATCC9144, which is a strain recommended in susceptibility testing, Vancomycin resistant enterococcus (VRE) (clinical isolate), Bacillus cereus (clinical isolate), Pseudomonas aeruginosa ATCC25668 (clinical isolate, Klebsiella aerogenes (clinical strain), E. coli ATCC10536 and Stenotrophomonas maltophilia (clinical strain). The strains were sub-cultured from frozen stocks and grown on blood agar (Oxoid, Basingstoke UK) in air for up to 48 hours.

Media and h ocula

Aspergillus conidia were harvested by washing the surface of the culture with 25mL of phosphate buffered saline (PBST; Life Technologies, Paisley, UK) containing 0.05%> (v/v) Tween® 80 (Sigma, Poole, Dorset, UK). Candida blastoconidia were collected from 3 or 4 colonies and suspended in PBST. Colony-forming units (CFU) were counted using a haemocytometer and diluted as required in PBST. For determination of the minimal inhibitory concentration (MIC) using microdilution methods, a final inoculum of 0.5-2.5 x 10⁵ CFU/mL of Candida spp., or 0.5-5x10⁴ CFU/mL for Aspergillus spp., was prepared. For surface seeded plates used in disc diffusion assays, the inoculum was adjusted to provide confluent growth. Four to five morphologically identical colonies of bacterial strains were harvested and suspended in sterile distilled water. The inoculum was adjusted to lxlO⁶ CFU/mL for microdilution methods. For surface seeded plates in disc diffusion assays the inoculum was adjusted to provide confluent growth.

Minimal Inhibitory Concentration (MIC) determination using a microdilution assay

All MIC assays with fungi were determined in RPMI 1640 (Sigma) supplemented with 2% (w/v) glucose (Sigma) and buffered at pH 7.0 with MOPS (Sigma). MIC assays with all bacteria except NRE were determined in Isosensitest broth (Oxoid). VRE MIC assays were conducted in Brain Heart Infusion (BHI) broth (Oxoid).

The methods used for Aspergillus were based on ΝCCLS 38A (National Committee for Clinical Laboratory Standards, 2002, "Reference method for broth dilution antifungal susceptibility testing of conidium-forming filamentous fungi. Approved standard." Document M38-A 2002a. NCCLS, Wayne, PA, USA). The methods used for Candida were based on AFST-EUCAST (Gin. Microbiol Infect. 9, 467-474, 2003, "Multicenter evaluation of the reproducibility of the proposed antifungal susceptibility testing method for fermentative yeasts of the Antifungal Susceptibility Testing Subcommittee of the European Committee on Antimicrobial Susceptibility Testing". The methods used for bacteria were based on NCCLS M7-A5 (National Committee for Clinical Laboratory Standards. (2000). Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically: Approved standard M7-A5. 2000a NCCLS, Wayne, PA, USA.)

The essential oils were diluted 1:4 in either RPMI 1640 (fungi), or 1:20 in Isosensitest broth (bacteria other than VRE) or BHI broth (VRE) and mixed using a vortex mixer for at least 10 seconds to form an emulsion of the oil in culture medium. The oil suspension was then double diluted in flat-bottomed microdilution assay plates from 1:5-1:10240 with a final control row consisting of medium only. Plates were sealed with a microdilution plate-sealing strip (Sigma) and incubated at 37°C in air for 24-48 hours.

The minimal inhibitory concentration (MIC) of Aspergillus was determined visually and were defined as the concentration of the drug in the first well that showed no growth. Minimum fungicidal concentrations (MFC) were determined by culturing lOOμL from each well in the microdilution plate that had no visible growth. The MFC was taken as the first well with less than 5 CFU (>99.99%). The MICs of essential oils against Candida and bacteria were determined using a spectrophotometer (fitted with a 490nm filter) and a no growth endpoint. MICs were determined by culturing lOOμL from each well in the microdilution plate that had no determinable growth. The MFC was taken as the first well with less than 5 CFU (>99% kill for Candida or >99.99% kill for bacteria).

MIC determination using a disc diffusion assay

RPMI agar (RPMI 1640 medium containing 1.5% Bacto Agar (Becton Dickenson, Oxford, UK), Isosensitest agar (Oxoid) or BHI agar (Oxoid) were surface seeded with organisms and allowed to dry at room temperature in a laminar flow hood. A 6mm sterile paper disc (Mast Bootle, Merseyside, UK) was placed onto the dried surface in the centre of the plate and 10μL of the essential oil were pipetted onto the disc. Plates were incubated for 24 hours at 37°C in a moist atmosphere. Zones of inhibition were calculated by measuring the zone of inhibition from the disc to the perimeter of growth.

All MIC and MBC assays were performed in duplicate on at least two separate occasions.

Results

Y) Determination of Antifungal activity of Essential Oils

The minimal inhibitory concentrations (MICs) and minimal bactericidal concentrations (MBCs) of 37 essential oils against Candida and Aspergillus spp are shown in Figure 1. The MICs were determined in sealed microdilution plates in air at 37°C for 24 hours. All ratios shown in Figure 1 represent the dilution in culture medium.

Geranium, Palmarosa, Peppermint and Rosewood (essential oils according to the invention) demonstrated inhibitory activity at dilutions of >1: 1024 against both Candida and Aspergillus. Lavender, Marjoram sweet French, May Cheng, Neroli Bigarde, Melissa True and Rose Otto were also effective but demonstrated slightly lower activity with inhibitory concentrations of >1 :32. The other oils had lower activity (and therefore do not fall within the scope of the present invention), i.e. only effective at concentrations greater than >1:10, or were effective against two microbial strains.

2) Determination of Antibacterial Activity of Essential Oils against Gram Positive and Gram Negative Bacteria and Fungi

Results of the disc diffusion assay are shown in Figure 2, where NI - No zone of inhibition; and NA - not available. The overall pattern of inhibitory activity was similar to the microdilution assay with Palmarosa, Peppermint, Rosewood and Rose Otto (oils according to the invention) demonstrating the highest and most broad spectrum inhibitory activity.

P. aeruginosa and K. pneumoniae were most resistant to individual essential oils by disc diffusion, demonstrating resistance to 22 oils.

3) Determination of Antibacterial Activity of Essential Oils against Gram Positive Organisms

The MICs of the 37 essential oils against gram-positive bacteria are shown in Figure 3. MICs and MBCs were determined in sealed microdilution plates in air at 37°C for 24 hours. All ratios represent the dilution in culture medium.

Palmarosa and Melisa True demonstrated cidal activity against all gram-positive isolates at dilutions of >1:160. Peppermint, Neroli Bigarde and Rose Otto were cidal against all gram- positive isolates at dilutions of >1:40. Patchouli and Sandlewood were cidal at >1:160 against all gram-positive isolates other than B. cereus. Each of these oils represent oils which may be used according to the present invention.

Nineteen of the oils (not encompassed by the present invention) had lower activity, i.e. effective at concentrations greater than >1 :20 or were effective against 2 strains

4) Determination of Antibacterial Activity of Essential Oils against Gram Negative Organisms

The MICs of 37 essential oils against gram-negative bacteria are shown in Figure 4. MICs and MBCs were determined in sealed micro-dilution plates in air at 37°C for 24 hours. All ratios represent the dilution in culture medium. In general the essential oils were less effective against gram-negative bacteria. For instance, when used singularly only Neroli Bigarde (1 :320) and Eucalyptus (1:20) were effective against P. aeruginosa. However inhibitory and cidal effects against S. maltophilia, E. coli and K. pneumoniae were demonstrated with essential oils according to the invention (e.g. Palmarosa, Peppermint, Rosewood, Neroli Bigarde and Melisa True were effective at concentrations >1:160).

Twenty-three of the oils had lower activity against gram-negative bacteria (only effective at concentrations >1:20 or were effective against a single strain). All MIC and MBCs in replicate samples were within two dilution wells.

5) Synergistic Effects of Essential Oils According to the Invention Against Bacteria

An important feature of the present invention is that the inventors have established that the abovementioned preferred oils, which have individual anti-microbial activity, have great efficacy when used in combination (even under circumstances where the use of a single oil does not necessarily have significant efficacy against a specific micro-organism when used alone). This is demonstrated by the data discussed below.

Referring to Figures 5a and 5b, there is shown the antibacterial effect of a combination of three different essential oils 4, 6, 8 loaded on a plate 2. The essential oil 4 was Rosewood, the essential oil 6 was Palmarosa, and the essential oil 8 was Geranium oil.

To produce these plates Staphylococcus aureus is spread uniformly over the surface of the plate, small drops of oil (5-10μL impregnated onto a paper disc) are placed onto the growing area. The plates are then incubated for 24 hours to allow the bacteria to grow.

Bacterial growth 12 is clearly seen on the majority of the plate 2 in the Figures. The perimeter of the plate demonstrates the uninhibited growth pattern of Staphylococcus aureus with small colonies growing freely on the surface. As can be seen in Figure 5b, the expected antibacterial zones for each of the three essential oils 4, 6, 8 are shown as circles 14. The radius of these circles 14 are determined by the bacterial growth media on the plate 2, and the duration of the experiment. However, surprisingly, the three essential oils 4, 6, 8, are shown to have a synergistic effect in that the antibacterial zone or halo 10 around each oil is seen to extend beyond that of the circle 14. Within the plate there is clear inhibition of growth of the bacteria this is due to the growth suppressing effect of the oils. NorniaHy the inhibition pattern in this system is circular due to the radial diffusion of the agents (as indicated by the overlaid circles). If the oils had no interaction the growth inhibition would remain as distinct circles. If the combination of the oils reduced the bacterial killing effect the zones of inhibition would be similarly reduced; whereas if the combination supplemented the bacterial killing activity the zone of growth inhibition would increase in size. However, surprisingly, the three essential oils Patchouli, Rosewood and Geranium, are shown to have a dramatic synergistic effect (that is they demonstrate increased activity when in combination) in that the antibacterial zone 10 around each oil is seen to extend beyond that of the circle 14.

Interaction of the oils can also be measured in liquid culture. In these experiments dilutions of essential oils are mixed in wells in 96 different combinations. These plates test the killing ability of the mixtures over a 1000 fold range and are able to both measure inhibitory and killing ability of the combinations. In this system Patchouli, Rosewood, Geranium, Lavender and Peppermint oils were tested against fungi (Candida albicans and Aspergillus fumigatus). Mixing the oils, in any combination, resulted in a synergistic or enhanced activity of the oils. Mixing Patchouli, Rosewood and Geranium, in any combination, also had synergistic effects against Methicillin Resistant Staphylococcus aureus.

This data represents an important aspect of the invention because oils according to the invention, particularly in combination, have been demonstrated to be potent against a multi-drug resistant micro-organism (for which there are very few, if any, useful conventional microbicidal agents). The ability to blend oils into mixtures which have synergistic killing ability provides a huge advantage. There is a constant battle between antimicrobial agents and microbes, with the unfortunate result that microbes are sometimes able to develop resistance to the antimicrobial agent. If single agents are used to kill microbes there is a chance of development of resistance. If agents are used in combination the chance of resistance developing is vastly reduced. This is because the microbe would need multiple changes to occur in its genetic make-up (mutations) to occur simultaneously. This strategy is commonly used with conventional antimicrobial agents as seen with triple/quadruple therapy of tuberculosis in which three or four antibiotics are administered, or HAART therapy of HIV in which a cocktail of drugs is administered which is designed to affect multiple targets of the HIN virus. Conclusions

The results of the present study clearly demonstrate that specified essential oils, and in particular blends of at least two of such oils in accordance with the present invention, have a broad spectrum of antibacterial and antifungal activity against clinically relevant pathogens whereas as other essential oils (which fall outside of the scope of the present invention) had negligible activity. It is also of interest and very surprising that oils with the greatest activity against bacteria were also highly active against fungi. This is particularly surprising as antibacterials and antifungals often have different modes of action.

It is important to note that essential oils according to the present invention were particularly effective against microbial isolates with resistance to broad spectrum antibiotics. Specifically there was no obvious tendency to reduced activity of the oils between methicillin resistant S. aureus and a fully susceptible S. aureus with oils demonstrating bactericidal activity at dilutions of at least 1:1280 against both strains. The pattern of the antimicrobial effects of the essential oils determined by microdilution assays and disc diffusion assays are not identical but demonstrate similar spectrums of activity. While the inventors do not wish to be bound by any hypothesis, they believe that the differences are possibly due to variable solubility of the components in liquid medium and limited diffusion of large or highly charged molecules within agar. In both methods, Palmarosa, Peppermint and Rosewood oils demonstrate surprisingly impressive antimicrobial activity. Other data obtained within the study (not included) indicated that all the oils according to the present invention were sterile and therefore should not in themselves be a source of infection.

Essential oils have been used for many years by aromatherapists and herbalists. In general the oils are diluted in a drop-wise fashion into a careying medium (often oil), which is then used either directly onto the skin or is inhaled as a vapour. It is important to realize that sensitive balances and scales are not available to practitioners and the oils are diluted directly using droppers attached to the stock bottles. The inventors of the present invention have therefore attempted to present the data in a manner (using ratios) which represents the most common way in which the oils are used in medicine and is therefore directly transferable to the clinic where ratios of >1:50 are regularly used. A ratio of 1:1280 translates to approximately 780mg/L. In this Example, the oils have also been used directly as purchased from a wholesale supplier, Natural Touch Aromatherapy. The oils have been used in this form rather than as purified components because this is the form in which they are used by aromatherapists and herbalists in clinics and so again the findings of the study should be directly transferable to clinical use. Furthermore, essential oils are complex mixtures of compounds produced by plants and often have potential benefits unrelated to antimicrobial activity, such as enhanced wound healing and anti-inflammatory or analgesic effects, which might be degraded on purification or fractionation of the oils.

In the above described Example, both the range and activity for a number of essential oils have been established, enabling well-designed prospective clinical studies to be undertaken. The essential oils examined in the Example demonstrate a broad range of high level antimicrobial activity which appears unaffected by resistance to conventional antibiotics. In particular, these data have enabled the inventors to titrate doses of essential oils, which would be clinically effective for future in vivo studies. In particular these data illustrate that mixtures of two or more essential oils according to the first aspect of the invention have surprising synergistic effects on a broad spectrum of micro-organisms and particular multi-drug resistant micro-organisms.

EXAMPLE 2

The inventors conducted further experiments to evaluate the efficacy of essential oils to kill or prevent the growth of a number of bacteria or fungi.

Experiments were conducted to evaluate the efficacy of patchouli, rosewood and geranium oils when used singularly or as a most preferred composition according to the present invention (i.e. a blend of all three).

All oils in this data set were diluted 1:100 or 3:100 for use as specified. Accordingly the single oils were a 1% or 3% solution whereas the blend was used as a 1% or 3% solution.

For each experiment a dilution of oil was prepared in culture medium (Isosensitest broth for bacteria or RPMI 1640 for fungi) at double its final concentration i.e. either a 2% or 6% solution. Suspensions of bacteria or fungi were also prepared in culture medium (Isosensitest broth for bacteria or RPMI 1640 for fungi) at a concentration of 2 x 10⁵/mX. Cultures were prepared on the microbe suspension to determine the precise number of organisms. The suspensions were thoroughly vortexed to disperse the essential oil or microbes and then 2.5mL of the diluted essential oil was mixed with 2.5mL of the microbe suspension to produce a final solution of 1% or 3% essential oil containing 1 x 10⁵ microbes per mL. A timer was started and this solution was again thoroughly vortexed then placed in a shaking water bath, at 37°C. Additional samples were prepared without the addition of essential oils to act as culture controls.

Samples of 0.2mL of the suspension were removed immediately on mixing and at 2minutes, 10 minutes and 40 minutes. The removed samples were immediately placed onto the surface of a large Petri dish containing blood agar and spread widely over trie surface (to prevent further killing activity by the essential oils; additionally the removed samples were immediately diluted in saline (1:10, 1:100 and 1:1000) and these dilutions also spread over the surface of blood agar plates (to enable counting of discrete colonies).

The blood agar plates were incubated in air at 37°C for 48 hours. After incubation bacterial and fungal colonies were counted on all plates and compared to the control plate.

The effects of such oils are demonstrated on E. coli, Staphylococcus aureus, MRSA, Vancomycin resistant enterococcus (VRE), Bacillus cere s, Klebsiella pneumoniae, Pseudomonas aeruginosa and Candida albicans in Figures 6 - 13 respectively. Timings are minutes after contact with the oils. All data are means of experiments repeated on three occasions

In all experiments the inoculum was approximately 1 x 10⁵/ml. Results are expressed as percentage organisms surviving oil compared to the control sample.

The figures illustrate that control solution containing no oil (Jfc) had no effect on the survival of the bacteria and fungi used in these experiments. Patchouli oil ( ), rosewood oil(___l), geranium oil(Δ) when used alone had differing effects (depending on the micro-organism being treated. However a blend of oils according to the present invention (♦) consistently killed the cultured micro-organism.

EXAMPLE 3

The inventors carried out further experiments to demonstrate that a combination of essential oils according to the present invention results in a surprising synergistic effect for killing bacteria and fungi.

3.1 Methods

Plate inhibition assays were conducted by exposing two strains of fungi to a dilution matrix of essential oils.

A Microdilution 12 x 8 plate was constructed in which:

(a) Each well of every column of the plate contained a single dilution of a first oil and wells of subsequent columns filled with a increasing serial dilutions of the first oil. Accordingly the dilution of the first oil in each column was in the series 1:16; 1:32; 1:64; 1:128; 1:256; 1:512; 1:1,024 etc (as specified in the figures). The last column was a control only

(b) Each well of every row of the plate contained a single dilution of a second oil and wells of subsequent rows filled with a increasing serial dilutions of the second oil. Accordingly the dilution of the second oil in each row was in the series 1:16; 1 :32; 1:64; 1:128; 1:256; 1:512; 1:1,024 etc (as specified in the figures). The last row was a control only

The oils were either plated with CAL4112 (a strain of Candida albicans) or AF65 (a strain of Aspergillus fumigatus which is resistant to the major antifungal agent amphotericin B).

For each experiment either 8 or 12 dilutions of oil was prepared in culture medium (RPMI 1640) at four times their final concentration (normally dilutions of 1:16; 1:32; 1:64; 1:128; 1:256; 1:512; 1:1,024 etc). Suspensions of fungi were also prepared in RPMI 1640 culture medium at a concentration of 2 x 10⁵/mL (this is double its final concentration). Quantitative cultures were prepared on the microbe suspension to determine the precise number of organisms.

Entire vertical rows of the microdilution plate were partially filled with 50μL of the first oil. Then entire columns of the microdilution plate were partially filled with 50μL of the second oil. Finally lOOμL of the fungal suspension was added to all wells. The well contents in the plate were well dispersed on a plate mixer and the plate incubated overnight at 37°C in air. The following morning the plate was examined and the level of growth in every well recorded. After reading the plate lOOμL of the well contents was removed and inoculated onto the surface of a Sabouraud Dextrose agar plate. These plates were incubated for 48 hours at 37°C in air. The plates were then examined and the number of fungal colonies

Each well was scored to evaluate whether:

(a) there was no fungal growth (i.e. the oils killed the fungi) (blank grids in Figures 14 -23); the Minimum fungicidal concentration (MFC) was taken as the first well with less than 5 CFU (>99.99%) (the clear grids in Figures 14 -23).

(b) the Minimum inhibitory concentration (MIC) was taken as the first well with no visible growth (the lighter shaded grids in Figures 14 -23).

(c) there was no effect on fungal growth (i.e. the oils did not effect the fungi) (the darker shaded grids in Figures 14 -23)

3.2 Results

Mathematical formula may be applied to the scored microdilution plates to calculate whether or not the combination of two oils had a synergistic effect, an additive effect, no greater effect than a single oil or an antagonistic effect. Such calculations will be well known to one skilled in the art and we only propose to provide illustrative scored plates. However Figures 14 and 15 show idealized data for synergistic and antagonist combinations of agents.

Combinations of oils that fall outside the scope of the invention did not fit into the synergistic model (data not shown). However blends of Palmarosa with one of Geranium, Peppermint, Rosewood and Lavender (compositions according to the invention) all had a synergistic effect on the survival of the fungi (see figures 16 - 23). EXAMPLE 4

The antibacterial activity of essential oils by vapour contact against bacteria and fungi was assessed.

In these experiments undiluted oils were placed into the lids of petri dishes.The surface of the petri dishes were inoculated with >10⁵ of either bacteria or fungi (this is sufficient organisms to produce very heavy confluent growth on the plate surface). The halves of the plate were then closed (leaving a 10mm gap between the oil and the microbe and sealed using parafilm. The sealed plates were then incubated at 35°C for 24 hours.

After incubation the amount of growth was compared to control plates with no oil in the lid. plates contained 15mL of agar based culture medium and the vapour space was approximately 65cm in all cases.

Table 1

MRSA* Methicillin resistant Staphylococcus aureus. The Oil Blend ** consisted of Geranium, Patchouli and Rosewood oils (either 30μL or 5μL of each oil). The Oil Blend*** consisted of Geranium, Patchouli and Rosewood oils (either lOμL or 1.7μL of each oil).

These data illustrate that blends of oils (i.e compositions according to the invention) have significant efficacy for reducing growth of MRSA or Candida. EXAMPLE 5

The Experiments illustrated in Example 1 (see figures 1 and 2) were expanded upon and the data, generated using the same methods as Example 1, are presented in Ta^"ble 2 and 3. These tables illustrate the effectiviness of single oils that may be blended to form compositions according to the invention. Other single oils, that are not used to form compositions according to the invention, were found to be completely or relatively ineffective.

Table 2.illustrates antibacterial activity (MIC and MBC) of essential oils against gram positive organisms whereas Table 3.illustrates antibacterial activity (MIC and MBC) of essential oils against gram negative organisms.

MICs and MBCs determined in sealed microdilution plates in air at 37^C>C for 24 hours. All ratios represent the dilution in culture medium

^a>l:20 No inhibition

For gram positive organisms: Angelica oil, benzoin rezinoid, black pepper, cardomon oil, cedar wood, clary sage, cypress, eucalyptus, frankincense, ginger, grapefruiit, juniper berry, lemon, mandarin, nutmeg, rosemary and chamomile german were also tested but MIC and MFCs were <1:40

For gram negative organisms: Angelica oil, benzoin rezinoid, bergamot., Black pepper, cardomon oil, carrot seed, cedar wood, cypress, frankincense, ginger, grapefru.it, juniper berry, lemon, mandarin, nutmeg, sandalwood, spikenard, chamomile german and valerian oils were also tested but MIC and MFCs were <1 :40

Vancomycin Methicillin Resistant Resistant Staphylococcus Bacillus cereus enterococcus aureus Staphylococcus

MIC MBC MIC MBC MIC MBC MIC MBC Bergamot >1:20 >1:20 Citrus bergamia >1:20 >1:20 >1:20 >1:20 1:80 >1:20 Carrot seed Daucus carota >1:20 >1:20 1:40 >1:20 >1:20 >1:20 1:320 >1:20 Lavender Lavandula 1:20 >1:20 1:80 1:20 1:80 1:40 1:80 >1:20 angustifolia Geranium Pelargonium 1:640 1:40 1:1280 1:1280 1:640 1:80 1:1280 1:40 graveolens Howood Leaf Cinnamomum 1:320 1:40 1:640 1:320 1:3120 1:640 1:2560 1:640 camphora Marjoram sweet French Origanum 1:80 1:20 1:80 1:40 1:40 1:40 1:40 >1:20 marjorana May Chang 1:320 Litsea cubeba 1:320 1:160 1:160 1:640 1:160 >1:20 >1:20 Myrrh Commiphora 1:320 1:40 1:320 1:40 1:320 1:40 >1:20 >1:20 myrrha Palmarosa Cymbopogon 1:1280 1:160 1:1280 1:320 1:1280 1:320 1:2560 1:160 martinii Patchouli 1:320 Pogostemon cablin 1:160 1:12480 1 :2560 1:12480 1:12480 >1:20 >1:20 Peppermint 1:320 Mentha piperita 1:40 1:1280 1:1280 1:1560 1:1280 1:1280 1:160 Rosewood Aniba rosaeodora 1:640 1:40 1:1280 1:640 1:6240 1:1280 1:2560 1:640 Sandalwood Santalum album 1:12480 1:1280 1:12480 1:1280 1:12480 1:3120 >1:20 >1:20 Spikenard Nardostachys 1:40 1:20 1:40 1:20 1:40 1:20 >1:20 >1:20 jatamansi Tea Tree Melaleuca 1:20 1:20 1:320 1:80 1:320 1:80 1:80 >1:20 alternifolia Neroli Bigarde Citrus aurantium 1:320 1:40 1:320 1:320 1:1280 1:320 1:640 1:320 Melissa True 1:320 Melissa officinalis 1:320 1:1280 1:640 1:2560 1:1280 1:1280 1:320 Jasmine Absolute Jasminum officinale 1:40 1:40 1:80 >1:20 1:40 1:20 1:40 1:20 Rose Otto Rosa damascena 1:1280 1:80 1:3120 1:1280 1:2560 1:1280 1:320 1:160 Valerian Valeriana officinalis >1:20 >1:20 1:640 1:160 1:1560 1:320 1:160 1:40 Yarrow >1:20 Achillea millefolium >1:20 1:80 1:40 1:640 1:160 1:160 >1:20

Table 2 Stenotrophomonas Pseudomonas Klebsiella maltophilia aeruginosa Escherichia coli pneumoniae MIC MBC MIC MBC MIC MBC MIC MBC Clarey Sage 160 80 >1:20 >1:20 20 >1:20 >1:20 >1:20 Eucalyptus 160 80 1:20 1:20 40 1:40 1:40 1:20 Lavender 160 80 >1:20 >1:20 80 1:80 1:20 1:20 Geranium 640 160 >1:20 >1:20 :40 1:20 >1:20 >1:20 Not Not Howood Leaf available available >1:20 >1:20 1280 1:1280 1:1280 1:1280 Marjoram sweet French 1 160 1:80 >1:20 >1:20 1:80 1:40 1:80 1:40 May Cheng 1 160 >1:20 >1:20 1:40 1:40 1:20 1:20 >1:20 Myrrh 1 320 1:40 >1:20 >1:20 >1:20 >1:20 >1:20 >1:20 Palmarosa 1 1280 1:640 >1:20 >1:20 1:640 1:640 1:160 1:160 Patchouli 1 80 >1:20 >1:20 >1:20 >1:20 >1:20 >1:20 >1:20 Peppermint 1 2560 1:1280 >1:20 >1:20 1 :2560 1:1280 1:640 1:640 Rosemary 1 160 1:80 >1:20 >1:20 1:40 1:40 1:40 1:20 Rosewood 1 640 1:640 >1:20 >1:20 1:1280 1:1280 1:1280 1:1280 Tea Tree 1 320 1:80 >1:20 >1:20 1:160 1:80 1:320 1:40 Neroli Bigarde 1 320 1:160 1:120 1:320 1:320 1:320 1:320 1:320 Melissa True 1 2560 1:1280 >1:20 >1:20 1:2560 1:1280 1:1280 1:320 Jasmine Absolute 1 160 1:20 >1:20 >1:20 >1:20 >1:20 1:20 >1:20 Rose Otto 1 2560 1:1280 >1:20 >1:20 1:1280 1:640 1:20 1:20 Yarrow 1 320 1:80 >1:20 >1:20 1:40 >1:20 1:40 >1:20

Table 3

It will be appreciated from other parts of the specification that combinations of two or more of the effective oils produce a composition with all the benefits of the first aspect of the invention. However it will be appreciated from tables 2 and 3 (and figures 1 and 2) that some single oils have surprising efficacy. Therefore according to a final aspect of the invention there is provided use of an essential oil derived from a plant genus independently selected from: Pelargonium; Cymbopogon; Mentha; Aniba; Lavandula; Origanum; Litsea; Citrus; Melissa; Pogostemon; Santalum; Styrax; Cinnamonum; Valeriana; and Rosa, for the manufacture of a medicament for the treatment of a microbial infection. Such oils may be as defined above.

Claims

1. The use of a composition comprising at least two essential oil derived from a plant genus independently selected from: Pelargonium; Cymbopogon; Mentha; Aniba; Lavandula; Origanum; Litsea; Citrus; Melissa; Pogostemon; Santalum; Styrax; Cinnamonum; Valeriana; and Rosa, for the manufacture of a medicament for the treatment of a microbial infection.

2. The use according to claim 1, wherein the essential oils are derived from at least two plant species independently selected from: Pelargonium graveolens; Cymbopogon martinii; Mentha piperita; Aniba rosaeodora; Lavandula angustifolia; Cinnamonum camphora; Origanum marjorana; Litsea cubeba; Citrus aurantium; Melissa officinalis; Pogostemon cablin; Santalum album; Valeriana officmalis; Styrax benzoin; and Rosa damascena.

3. The use according to claim 1 wherein the essential oils are selected from the oils defined herein as Palmarosa, Geranium, Bigarade, Patchouli, Peppermint, Rose Otto, Rosewood, Nalarian, May Chang, Melissa True, Lavender, Marjoram Ho leaf or Sandalwood.

4. The use according to any one of claims 1 - 3 wherein the microbial infection is a filamentous fungus infection.

5. The use according to claim 4 wherein the filamentous fungus is an Ascomycete.

6. The use according to claim 5 wherein the Ascomycete is an Aspergillus sp.

1. The use according to claim 6 wherein the Aspergillus sp is an organism independently selected from a group of species consisting of Aspergillus fumigatus; Aspergillus nidulans; Aspergillus niger; Aspergillus terreus; and Aspergillus flavus.

8. The use according to claim 5 wherein the Ascomycete is a Candida sp.

9. The use according to claim 8 wherein the Candida sp is an organism independently selected from a group of species consisting of: Candida albicans; Candida guilliermondii; Candida krusei; Candida glabrata; and Candida tropicalis.

10. The use according to any one of claims 4 - 10 wherein the oil is derived from a plant species independently selected from: Lavandula angustifolia; Pelargonium graveolens; Origanum marjorana; Litsea cubeba; Cymbopogon martini; Mentha piperita; Aniba rosaeodora; Citrus aurantium; Melissa ofβcinalis; and Rosa damascena.

11. The use according to any one of claims 1 - 3 wherein the microbial infection is a bacterial infection.

12. The use according to claim 11 wherein the bacteria are gram positive.

13. The use according to claim 12 wherein the gram positive bacteria is a Bacillales.

14. The use according to claim 13 wherein the Bacillales is a Staphylococcus or a Streptococci.

15. The use according to any one of claims 12 - 14 wherein the oil is derived from a plant species independently selected from: Cymbopogon martini; Pogostemon cablin; Mentha piperita; Santalum album; Citrus aurantium; Melissa ofβcinalis; and Rosa damascena.

16. The use according to claim 11 wherein the bacteria are gram negative.

17. The use according to claim 16 wherein the gram negative bacteria is a Pseudomonadales, Gammaproteobacteria, Betaproteobacteria, Delta/epsilon subdivided Proteobacteria or an Actinobacteria.

18. The use according to any one of claims 16 or 17 wherein the oil is derived from a plant species independently selected from: Cymbopogon martini; Mentha piperita; Aniba rosaeodora; Citrus aurantium; and Melissa officinalis.

19. The use according to any preceding claim for the treatment of a multidrug resistant micro-organism.

20. The use according to any preceding claim wherein the medicament comprises at least one volatile oil and at least one viscous oil.

21. A composition comprising at least two essential oils selected from a group of oils derived from a plant genus independently selected from: Pelargonium; Cymbopogon; Mentha; Aniba; Lavandula; Origanum; Litsea; Citrus; Melissa; Pogostemon; Santalum; Valeriana; and Rosa, for use as a medicament.

22. A composition comprising at least two essential oils selected from a group of oils derived from a plant genus independently selected from: Pelargonium; Cymbopogon; Mentha; Aniba; Lavandula; Origanum; Litsea; Citrus; Melissa; Pogostemon; Santalum; Valeriana; and Rosa for use as a consumer product.

23. A composition comprising at least two essential oils selected from a group of oils derived from a plant genus independently selected from: Pelargonium; Cymbopogon; Mentha; Aniba; Lavandula; Origanum; Litsea; Citrus; Melissa; Pogostemon; Santalum; Valeriana; and Rosa for use as a cleaning product.

24. The composition according to any one of claims 21 - 23 wherein the essential oils are as defined in any one of claims 1- 3.

25. The composition according to claim 24 further comprising a surface-active agent.

26. The composition according to any one of claims 21 - 25 formulated as a shampoo, soap or handwash.

27. The composition according to any one of claims 21 - 25 formulated as a solution for cleaning surfaces, devices or utensils

28. A method of preventing and/or treating a microbial contamination comprising coating an object or a surface in need thereof with an amount of composition as defined by any one of claims 21 - 27, that is effective for killing or preventing growth of micro-organisms

29. The method according to claim 25 wherein it prevents or treats contamination with any of the micro-organisms defined by any one of claims 4-9, 11-14, 16, 17 or 20.