Substances having antiviral activity against one or more human disease-causing viruses are disclosed. These antiviral substances are obtained from the cuticular and extracuticular layers of plants. Pharmaceutical and nutraceutical formulations comprising the antiviral substances are also disclosed, as are methods of using such formulations to treat viral diseases.

SUMMARY OF THE INVENTION

In accordance with the present invention, substances from the cuticular or epicuticular layers of plants have been identified and are shown to possess potent antiviral activities. Also provided are methods for obtaining these antiviral substances from the cuticular and epicuticular material of plants. When prepared according to the method taught, these substances have particular therapeutic application for herpes simplex virus HIV, influenza virus and other viral infections. In addition, the invention also provides methods for the use of these substances in a therapeutic mode.

AS used herein, the term "cuticle" or "cuticular layer" refers to a substantially non-cellular multi-layered structure that lies over and merges into the outer walls of epidermal cells, as shown diagrammatically in FIG. 3 (taken from Martin & Juniper, The Cuticle of Plants, St. Martin's Press, New York 1970). The term "cuticular material" refers simply to material that is obtained from any part of the cuticle. Likewise, it is known that substances (such as waxes) are exuded from the cuticle onto the outermost surface of the plant, forming a layer referred to herein as the "epicuticular layer". The term "epicuticular material" refers to material or substances obtained from this layer.

It should also be understood that the term "substance" or "agent" when used herein in the singular is intended to encompass the plural as well. For example, a cuticular or extracuticular antiviral substance may contain a single substance, or it may contain a mixture of two or more substances.

According to one aspect of the present invention, an antiviral preparation obtained from the cuticular and/or epicuticular material of plants comprising substances soluble in an organic solvent, but not requiring disruption of the plant cells interior to the epidermal layer, is provided. In one embodiment, the active materials are localized externally to the epidermis of the plant, i.e., in the cuticle or deposited onto the surface of the plant in an "epicuticular" layer. The antiviral substances can be found in the cuticular and epicuticular layers of different plant parts, including but not limited to fruits, flowers, leaves, stems and roots. In a preferred embodiment, plant parts containing these antiviral agents are obtained from one or more readily available plant species, such as apple, avocado, bearberry, black cherry, blueberry, cabbage, cherry, cork oak, cucumber, eucalyptus, grape, holly, orange, pear, plum (including Japanese plum), rhododendron, tobacco, tomato, wax palm and willow, to name a few.

According to another aspect of the invention, a method for obtaining of an antiviral preparation from the cuticular and epicuticular layers of plants, is provided. The objective of the method is to obtain materials contained in the cuticle and epicuticular layers of the plant, while minimizing or eliminating extraction from internal plant tissues. The method comprises the following, steps: (a) plant tissues (preferably intact, but fruit peels also may be used) are exposed to a solvent which solubilizes the active material located external to the epidermis of the plant (i.e., the cuticle and epicuticular layer) under conditions that leave the internal plant tissues substantially unaffected; (b) the solvent is partly or completely removed, leaving behind the concentrated cuticular and epicuticular material.

The solvent is one that is capable of solubilizing compounds from a matrix of waxy or cuticular material, or compounds dispersed on the plant's surface that are not washed off by rain or dew; typically these would comprise substantially non-polar solvents. For instance, the solvent may be selected from the group consisting of hexane, chloroform, dichloromethane, heptane, ether, petrolether, t-butyl ether, DMSO, long-chain alcohols, petroleum derivatives, supercritical fluids and liquefied gases. The solvent may be removed by evaporation under vacuum, or by filtration, precipitation or lyophilization.

In one embodiment, the concentrated cuticular and epicuticular material is resolubilized or resuspended in a second solvent that, at the concentrations used, is minimally toxic or nontoxic to cells in vitro or organisms in vitro. Appropriate solvents for such purposes include combinations of one or more of ethanol, water, polyethylene glycol, DMSO, glycerol, phenol, and other such solvents. In another embodiment, the antiviral material is suspended in a combination of one or more of the above-mentioned solvents, or in a colloid, or in another medium.

According to another aspect of the present invention, a pharmaceutical formulation comprising an antiviral substance obtained from plant cuticular and epicuticular material is provided. The pharmaceutical formulation of this invention is effective in the treatment of a viral disease process, and especially useful for the treatment of Herpes Simplex Virus Type I, influenza and or HIV infections.

According to another aspect of the present invention, a method for the treatment of, or therapeutic intervention in, a viral disease process is provided. The method comprises administering to a patient the above described pharmaceutical formulation at a frequency and duration sufficient to result in reduction or alleviation of the symptoms or cause of the viral disease process. In a preferred embodiment, the viral disease is caused by HSV, HIV or influenza virus.

Also provided in accordance with the present invention is a nutraceutical formulation comprising the aforementioned antiviral substances obtained from plant cuticular and epicuticular layers. Since the antiviral substances of the present invention can readily be prepared front common food species, it is contemplated herein that such substances can readily be included with formulations of nutraceutical products which are now commonly available and used by millions of health conscious consumers interested in alternative approaches to health. Such nutraceutical products, comprising the antiviral substances include for example, health beverages, herbal drinks, elixirs, juices, or teas, powdered beverage mixes, nutritional supplements, vitamins, additives, natural remedy formulations, health bars, creams, balms, sprays, and the like.

DETAILED DESCRIPTION OF THE INVENTION

The development of antiviral substances that are safe and nontoxic to infected cells and yet inhibit the replication of the virus by one or more means is a area of great need and much research. Ethnobotanists, pharmacologists, and others have turned to the natural world, and especially the plant kingdom, as a vast resource of potentially therapeutic agents with such properties as high efficacy and low toxicity. Advances in this area will add greatly to the limited base of therapeutic options for treatment of viral disease processes.

In accordance with the present invention, plant-derived agents possessing antiviral properties are provided. It has been discovered in accordance with the present invention that agents with such desirable therapeutic properties can be removed from the cuticular and epicuticular material present in plants, with minimal disruption of the plant tissue internal to the epidermis of the plant. 3. The antiviral properties of these materials likely arises from one or more constituents which comprises one or more constituents of plant cuticular and epicuticular materials, such as waxes, plant wax components, cutins, terpenoids, triterpenoids, phenolics, primary alcohols, secondary alcohols, hydrocarbons, diketones, fatty acids and flavanoids, to name a few.

Whole plants or intact plant parts may used as a source material of the cuticular and epicuticular material, including fruits, flowers, leaves, roots, stems, and bark. Preferred for use in the present invention are fruits, leaves and stems. These plant parts preferably are intact; however, peels or rinds that are mechanically separated from fruits or other plant parts also may serve as an economical source of cuticular and epicuticular material, inasmuch as such peels or rinds are often discarded as refuse in the processing of fruits and vegetables.

In one embodiment, a plant species is selected which is a readily available agricultural or horticultural plant or crop. Examples include but are not limited to species of plants Such as apple (Malus spp.), pear (Pyrus spp.), grape (Vita spp), orange (Citrus spp.), tomato (Lycopersicon esculentum), cabbage (Brassica spp.), cucumber (Cucumis spp.), cherry, black cherry, plum, peach and apricot (all Prunus spp.), avocado (Persea spp.), blueberry (Vaccinium spp.) bearberry (Arctostaphylos spp.) olive (Olea) and wax palm (Copernicia spp. or Ceroxylon spp). Other examples include tobacco (Nicotianum spp.), cork oak (Quercus occidentialis), eucalyptus (Eucalyptus spp.), rhododendron (Rhodendendron spp.), holly (Ilex rotunda), Japanese plum (Eriobotrya Japonica) and willow (Salix spp.). Still other examples include plants of the following genera: Euphorbia, Pedilanthus, Syagrus Cocus, Attalea, Stipa, Glyceria, Saccharum, Myrica, Rhus, Sapium, Linum, Agave, Cannabis, Raphia, Coccus, Ligustrum, Fraxinus, Benincasa, Ricinus, Baxus, Mesembryanthemum, Rubus and Malaleuca. These genera, as well as certain of the genera listed before these genera, contain species known to produce an abundance of waxy or oily cuticular and epicuticular materials.

In one embodiment, tissues are selected which are separate from the main agronomic feature of the plant, for example: stem and leaf tissue from the tomato vine could be used for extraction after the tomato fruit is picked. Alternatively, in the case of fruits and vegetables, those items which due to damage or defect are not usable for the food or feed purpose intended may be used for extraction in lieu of diverting first quality produce. It is also contemplated that the plant tissues from which the antiviral extracts are prepared may consist of a mixture tissues from one or more of these plants in certain combinations as are useful. In addition, as discussed above, the peels or rinds of fruit provide a useful source of cuticular and epicuticular material. These examples are intended to illustrate, not to limit, the invention.

In accordance with the present invention, a method for preparing antiviral substances from cuticular and epicuticular plant materials is provided. The method comprises exposing the plant surfaces to a solvent that solubilizes the cuticular and epicuticular portions of the plant, while leaving, the remaining plant tissues substantially intact, followed by concentrating or otherwise preparing the antiviral substances for further use.

The step of solvent exposure typically involves dipping, spraying or otherwise briefly exposing the plant (or plant part) surface to a substantially nonpolar solvent of a type commonly used for dissolving waxes, lipids and similar substances found in the cuticle. An appropriate solvent typically comprises one or more solvents selected from the group consisting of hexane, chloroform, dichloromethane, heptane, ether, petrolether, t-butyl ether, DMSO. Other examples of suitable solvents include organic solvents, high molecular-weight alcohols, ethers, petroleum derivatives, supercritical fluids, liquefied gases, and other such solvents as are known to those skilled in the art.

The method includes an optional step of clarification. The clarification step comprises one or more separation steps to remove insoluble material and plant residue from the solubilized cuticular and epicuticular material. Examples of clarification processes include but are not limited to: gravity settling; screening;

Claim 1 of 12 Claims

1. A method for producing a preparation possessing antiviral activity comprising substances obtained from cuticular or epicuticular layers external to an epidermis of a plant or plant part, the method comprising:

a) exposing the plant or plant part to a solvent under conditions sufficient to solubilize materials in the cuticular and epicuticular layers of the plant, while leaving cells and tissues internal to the epidermis substantially unaffected;

b) obtaining a solution or suspension of plant cuticular and epicuticular materials, thereby producing the antiviral preparation; and,

c) formulating the antiviral preparation with a pharmaceutically acceptable carrier to form a pharmaceutical composition possessing antiviral activity in a form selected from a tablet and a capsule,

wherein the plant or plant part is selected from the group consisting of apple.

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