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  Pharmaceutical Patents  

 

Title:  Medicinal herbal composition for treating infection
United States Patent: 
7,556,830
Issued: 
July 7, 2009

Inventors:
 Chepkwony; Paul Kiprono (Eldoret, KE), Medina; Maria (Nairobi, KE), Medina; Mitchell (Nairobi, KE)
Assignee:
  International Patents Holdings, Ltd. (Hamilton, BM)
Appl. No.:
 11/352,931
Filed:
 February 13, 2006


 

Pharm Bus Intell & Healthcare Studies


Abstract

Herbal compositions derived from Kenyan plants are provided for the treatment of HIV and other infectious diseases. The herbal compositions can include the extracts of up to 14 plants, including the root of abyssinica and Clutia robusta. Also provided are methods for extracting alkaloids and other compounds from the plants. Also provided are methods of treating a subject having an infectious disease, particularly HIV.

Description of the Invention

SUMMARY OF THE INVENTION

The present invention is based upon the discovery of the unique antiviral properties of a herbal remedy composition prepared from a variety of plants native to Kenya. The herbal composition of the present invention can include plant material from between two and 14 different plants preferably including roots of abyssinica (representative seed of said line having been deposited under ATCC Accession No. PTA-6769) and Clutia robusta (representative seed of said line having been deposited under ATCC Accession No. PTA-6970). For treatment of infectious disease, the herbal composition of plant material may be extracted to produce a liquid herbal composition or further purified to obtain alkaloid compounds from the plant material. The liquid herbal composition prepared from aqueous extracts from the plants has demonstrated effectiveness in treating HIV-positive subjects, as subjects treated with the liquid herbal composition have experienced improvements in CD4+ cell counts, and in some cases, complete reversal of HIV positive status.

In one aspect, the invention provides a herbal composition for treating infectious diseases, such as for example, HIV. The composition containing plant material includes the roots of abyssinica and the roots of Clutia robusta. In other embodiments of the invention, the herbal pharmaceutical composition may also include plant material, as indicated, from one or more of the following: stem bark of Prunus africanastem bark of Croton macrostachyus, stem bark of Acacia nilotica (representative seed of said line having been deposited under ATCC Accession No. PTA-7378), roots of Rhamnus prinoides, roots of Adenia gummifera, roots of Asparagus africanus, stem bark of Anthocleista grandiflora, whole plant of Plantago palmata (representative seed of said line having been deposited under ATCC Accession No. PTA-7377), roots of Clematis hirsuta, stem bark of Ekebergia capensis, stem bark of Bersama abyssinica, and roots of Periploca linearifolia.

In another aspect, the invention provides a method for preparing a liquid extract of the solid herbal composition of the invention. The extraction of plant material can be done with hot water. In one embodiment, hot aqueous extraction is done under basic conditions, followed by hot aqueous extraction under acidic conditions. In further embodiments, desired alkaloid compounds purified from the liquid extracts are provided or produced from direct chemical synthesis.

The invention further provides aqueous extracts of the herbal compositions of the invention. Also provided are alkaloid compounds purified from aqueous extracts and the chemical synthesis of the herbal compositions of the invention.

In another aspect of the present invention a method for treating HIV-positive subjects is provided. Subjects are administered an effective amount of a herbal composition of the invention prepared from the aqueous extracts of Dovyalis abyssinica and Clutia robusta, alone or in combination with one or more of the following: Prunus africana, Croton macrostachyus, Acacia nilotica, Rhamnus prinoides, Adenia gummifera, Asparagus africanus, Anthocleista grandiflora, Plantago palmata, Clematis hirsuta, Ekebergia capensis, Bersama abyssinica, and Periploca linearifolia, in doses based on subjects' body weights. In other embodiments the herbal composition of the invention is prepared from purified alkaloid compounds obtained from the aqueous extracts. The herbal compositions are administered at least once a day. In other embodiments, the herbal composition is administered twice or three times daily, based upon the health of the subject. In other embodiments, the composition may be administered as a beverage, capsule, tablet, powder, candy, gel, nutritional product or pharmaceutical product.

In another aspect of the present invention provides an herbal composition for treating subjects having infection, such as for example, HIV or AIDS. The herbal composition consists essentially of extracts of abyssinica and Clutia robusta, and optionally one or more of the following: Prunus africana, Croton macrostachyus, Acacia nilotica, Rhamnus prinoides, Adenia gummifera, Asparagus africanus, Anthocleista grandiflora, Plantago palmata, Clematis hirsuta, Ekebergia capensis, Bersama abyssinica, and Periploca linearifolia. In one embodiment, the herbal composition of the invention is prepared from purified alkaloid compounds obtained from aqueous extracts.

In another aspect of the present invention a method is provided for treating subjects having infection, such as for example, HIV or AIDS. Subjects are administered an effective amount of a herbal composition consisting essentially of extracts of abyssinica and Clutia robusta, and optionally the extract of one or more of the following: Prunus africana, Croton macrostachyus, Acacia nilotica, Rhamnus prinoides, Adenia gummifera, Asparagus africanus, Anthocleista grandiflora, Plantago palmata, Clematis hirsuta, Ekebergia capensis, Bersama abyssinica, and Periploca linearifolia, in doses based on subjects' body weights. In other embodiments the herbal composition of the invention is prepared from purified alkaloid compounds obtained from the aqueous extracts. The herbal compositions can be administered at least once a day. In other embodiments, the herbal composition can be administered twice or three times daily, based upon the health of the subject. In other embodiments, the composition may be administered as a beverage, capsule, tablet, powder, candy, gel, nutritional product or pharmaceutical product.

DETAILED DESCRIPTION OF THE INVENTION

The present invention relates to the discovery that combinations of extracts from plants native to Kenya can be used in the treatment of infection, such as for example, HIV and AIDS. Herbal compositions prepared from combinations of the extracts of the following: the roots of abyssinica and Clutia robusta, Rhamnus prinoides, Adenia gummifera, Asparagus africanus, Clematis hirsuta, and Periploca linearifolia, the stem bark of Ekebergia capensis, Bersama abyssinica, Prunus africana, Croton macrostachyus, Acacia nilotica, and Anthocleista grandiflora, and the whole plant of Plantago palmata have been shown to be particularly effective in improving the health of infected subjects. Specifically, herbal compositions of the present invention are particularly well suited for the treatment of infectious diseases including HIV.

Compositions of the invention can be prepared from plant material collected from the Mau Forest Complex in Western Kenya. Herbal compositions prepared from aqueous extractions and purified extracts of plants from this region of Kenya exhibit increased potency in the treatment of infectious diseases. The Mau Forest Complex is located at 0.degree. 30' South, 35.degree. 20' East and in the Rift Valley Province, and spans four Kenyan administrative districts: Narok, Nakuru, Bomet and Kericho. Mean annual rainfall varies from 1000 to 1500 mm with peaks in April and August. The rainfall pattern at the western flanks is governed by the moist monsoon winds from the Indian Ocean and dry winds from the Great Rift Valley. The western flanks of the Mau Forest Complex are influenced by the Lake Victoria macroclimatic region and are generally wetter with annual rainfall greater than 2000 mm and more evenly distributed. Mean annual temperatures for the Mau Forest Complex range from 12-16.degree. C. The soil of the Mau Forest Complex is rich volcanic loam having a pH between 3.8-5.8.

The vegetational pattern follows an altitudinal gradient with local topographical ecolines. The closed canopy moist mountain forest at lower altitudes becomes increasingly intermixed with bamboo from 2200 m onwards. Between 2300 and 2500 m, pure bamboo (Arundinaria alpina) swards are found. Above 2500 m this gives way to mixed bamboo/tree stands, both associated with grass clearings that usually represent a sub-climax resulting from burning and cutting of bamboo. A marginal type of mountain sclerophyll forest, wherein the plants generally have hard leaves to prevent wilting during dry conditions, occupies the highest altitudes of the Mau complex.

Plants in the Western flank of the Mau Forest Complex have shown the highest potency for the herbal compositions. Plants growing in the Western flank, (which is generally a high rainfall, high altitude region), have fewer environmental stresses. It is therefore possible that plants of the Western flank have more biosynthetic pathways, which may in turn lead to the production of a greater number of diverse compounds, which may in turn explain the greater potency of plants from the Western flank (as compared to other regions of the Mau Forest Complex). Alternatively, the greater potency plant extracts from the Western flank plants may be a result of a greater variety and number of alkaloids and other compounds in the plant extracts, such that the combined effect is greater than the sum of their individual effects.

The East Mau Forest Complex has a drier vegetation of Cedar and Podo. Wherever these species have been extracted, colonizing species such as Neuboutonia marcrocalyx and Macaranga capensis can be found.

The compositions of the invention may be prepared using plants collected from three altitude ranges of the Mau Forest Complex: 2000 m (annual rainfall of 1000 mm), 2300 m (1500 mm), 2500 m (western Mau flank, annual rainfall greater than 2000 mm) above sea level. The Western flanks of the Mau Forest contain plants that are particularly preferred for preparing the herbal compositions of the invention. The plants grown in the drier. Eastern flank of the Mau Forest Complex also may be used.

Plant material for preparing compositions of the invention may also be obtained from plants grown in a greenhouse environment. The germination of the seeds of particular plants may be altitude or soil dependent. Seeds for greenhouse planting may require collection from the natural dispersal agents as they exist in the wild. Additionally, simulation of rainfall, sunlight (an average of 12 hours per day in the Mau Forest Complex), and soil conditions of the Mau Forest Complex (i.e., rich volcanic loam having a pH between 3.8-5.8) may be required to obtain plants of similar potency.

The seeds of abyssinica (representative seed of said line having been deposited under ATCC Accession No. PTA-6969) are contained in a fleshy fruit. There are about 4 seeds enclosed by the flesh. A ripe fleshy fruit can be soaked in water for about 4 days, to make it possible to squeeze with minimum force to release the small seeds, each being approximately the size of a tomato seed or slightly larger. The seeds are then washed, dried and stored, awaiting germination under Mau Forest-like environmental conditions. In the wild, the fruit flesh is soaked by rain water, which results in the release of the seeds. The seeds grow naturally under the environmental conditions of the Mau Forest Complex as described above.

The Clutia robusta (representative seed of said line having been deposited under ATCC Accession No. PTA-6970) seeds are much smaller and encased in berries having a nut-like outer covering which encases approximately 3 to 4 seeds the size of a grain of sand. When mature seeds are exposed direct sunlight, they disperse rapidly in a process called explosive dispersal. This is not a problem in the wild, but if one is interested in collecting the seeds, care and intelligence are required, or else all the seeds will fly away under the scattering effect of the hot sun.

To recover the clutia robusta seeds, the berries should be placed in a metallic container, and covered with a material that allows sunlight to enter, such as a transparent polyethylene film surrounding a container of appropriate wire mesh. Exposure to light will cause the shells to break open, releasing the seeds which can then be separated from the chaff.

The optimal time for planting the clutia and dovyallis seeds in their natural environment is during the long rains, typically around the month of April. However, in the wild, the plants will generally grow throughout the year, except during the dry season, as the plants require a considerable amount of water and light to grow.

Croton macrostachyus produces pale pea-sized capsules, on drooping spikes to 30 cm long, splitting open on the tree to release 3 shiny grey seeds, covered at one end by a soft, creamy aril, or envelope.

Prunus africana produces spherical fruit, about 10 mm in diameter and is pinkish brown in color.

The Acacia nilotica (representative seed of said line having been deposited under ATCC Accession No. PTA-7378) plant produces straight or curved pods measuring approximately 17.times.2 cm. When young, the pods are green and fleshy but get darker with age, and are usually velvety. Pods have a fruity odor and open on the ground to release seeds.

Ekebergia capensis produces rounded, thin skinned berries, up to 2.5 cm in diameter, on long stalks in heavy bunches, which are yellow to red in color when mature.

The berry-like fruits of Rhamnus prinoides are approximately the size of a pea (about 5 mm in diameter), roundish and clearly divided into three compartments. They are fleshy and green, turning red and then purple as they ripen.

The fruit of the Asparagus africanus is a round berry, approximately 0.5 cm in diameter, green aging to orange, found most of the year. It is spread mainly by birds carrying the seeds.

The Anthocleista grandiflora produces fruits that are oval in shape, measuring approximately 3 cm.times.2 cm, glossy, smooth and brown when mature. Multi-seeded, large fruits are found throughout the year.

The Bersama abyssinica produces a smooth, spherical capsule, measuring approximately 2.5 cm in diameter, golden velvety at first, losing most of the hair and becoming brown by maturity; splitting into four valves to reveal attractive bright red seeds, about 10 mm long, enveloped for about their half length by a yellow, cup-shaped aril.

Adenia gummifera produces a fruit which is a stalked 3-valved capsule, leathery or fleshy, often red; seeds compressed with bony testa in a fleshy aril.

Plantago palmata (representative seed of said line having been deposited under ATCC Accession No. PTA-7377) produces a capsule-like fruit with two seeds per capsule.

Periploca linearifolia (representative seed of said line having been deposited under ATCC Accession No. PTA-7375) produces black seeds measuring approximately 10 mm long and 2 mm wide with white wool measuring around 3 cm attached to the tips of the seeds. The seeds are enclosed in pods measuring about 12 cm long. Upon maturity, the pods break open upon exposure to sunlight. This releases the seeds, which are borne aloft by the wool as they are dispersed by wind. Alternatively, these plants may be cultivated from stem cuttings, which when laid on or planted in the ground, grow roots and propagate new plants.

Clematis hirsuta (representative seed of said line having been deposited under ATCC Accession No. PTA-7383) produces yellowish seeds measuring approximately 3mm in length and 1 mm in breadth. The seeds are surrounded by yellowish-white wool which measures about 5 mm long. The wool carries the seeds upon the wind, which is the dispersal agent.

HIV Testing

As noted previously, for purposes of this application, a person is considered HIV-negative if the subject tested negative on a two-part HIV screening tests, consisting of an initial screening test and a confirmatory test.

An infected individual usually goes for testing for one or more of the following reasons: 1) the individual feels ill, 2) the individual's sexual partner is ill and has tested positive, 3) the individual's sexual partner died of AIDS; or 4) the individual suspects his/her sexual partner is sexually promiscuous.

The initial screening test is ELISA (Enzyme-Linked Immunosorbent Assay), an enzyme immunoassay (EIA) to determine the presence of HIV antibodies. The ELISA test uses artificial HIV proteins that capture antibodies to the virus and is more than 99 percent accurate. If antibodies to HIV are present (positive result), the test is typically repeated. However, other antibodies can cause a false-positive result.

Generally, HIV-1 antibodies are detectable approximately 25 days after acute infection, with nearly all infected subjects testing HIV positive 12 weeks after infection. The process of developing antibodies to a virus is termed seroconversion, and individuals who become antibody-positive are often called seroconverters.

Two types of HIV have been identified: HIV-1 and HIV-2, of which, HIV-1 is more common. HIV-1 and HIV-2 are similar in the modes of transmission (sexual contact, sharing needles, etc.) and infected individuals are generally subject to the same opportunistic infections. However, HIV-2 appears to weaken the immune system more slowly than HIV-1.

In Kenya, individuals are generally tested for antibodies to both HIV-1 and HIV-2. HIV-1 is generally more common in the Western world and HIV-2 is more common in Africa. In Kenya however, most HIV-positive individuals have the HIV-1 infection. It is believed that 90% of the HIV-positive cases in Kenya are HIV-1, with the remaining 10% of HIV-positive cases being the HIV-2. While rare, subjects occasionally are HIV antibody-positive to both types of HIV (i.e. HIV-1 and HIV-2).

The second part of the HIV screening test is called the confirmatory test. In the U.S., the most often used confirmatory test is the Western blot, wherein an electrical field is used to separate the various components by their molecular weight prior to evaluating antibody binding. This allows identification of antibodies to specific viral antigens, which show up as identifiable "bands" on a strip of test paper. The Western blot test is more difficult to perform and accurately interpret than the ELISA test, but it is less likely to give a false-positive result because it can distinguish HIV antibodies from other antibodies that may react to the ELISA. Other confirmatory tests may be used, including the indirect fluorescent antibody assay (IFA) and the radioimmunoprecipitation assay (RIPA).

One major drawback of antibody tests is the "window" period (i.e. the time it takes the body to produce antibodies after infection has begun). The screening tests do not correlate to the presence or absence of symptoms. The standard HIV tests do not detect the virus itself, but instead detect the antibodies that the body produces in response to the virus. During the period before the antibodies are produced, a person may be infected with HIV and can infect others, and still test negative on the HIV antibody test. It is therefore important to tell subjects who test negative to avoid engaging in high-risk behavior and to return for retesting at a later date.

The p24 antigen test can be used in diagnosing HIV early in the course of infection. It is primarily used to screen the blood supply but in some places it is used for testing for HIV. The p24 antigen is a protein that is part of the HIV. Early in the infection, it is produced in excess and can be detected in the blood serum by a commercial test. The p24 test can detect HIV infection before the HIV antibody test can and it is recommended 2-3 weeks after a risk exposure.

Individuals that test positive for HIV are regularly administered two tests to monitor HIV levels in the blood and to determine how the virus is affecting the immune system. These tests are: (1) a viral load measurement, and (2) CD4+ cell counts.

Viral load measurement (also called the HIV plasma RNA test) determines how many HIV viral particles are present in a given amount of a person's blood. Test results help determine the best treatment for the HIV infection as the viral load test shows how fast the virus is multiplying in the body. Because HIV reproduces by making copies of itself, the results are given as copies per milliliter (mL). Viral load testing can also reveal the presence HIV infection before antibodies can be detected and can also accurately determine whether a baby born to an infected mother has HIV.

CD4+ cell counts (T-lymphocyte measurements) provide an estimate of the immunologic status of an individual and help determine the immediate risk of opportunistic infection. The CD4+ count measures the number of a certain type of white blood cell that is most affected by HIV, and are measured every 3 to 4 months in individuals infected with HIV. On average, an individual infected with HIV loses approximately, 30-60 CD4+ cells per year, although in some subjects, CD4+ T-lymphocyte counts may remain stable for years followed by rapid decline.

CD4(T4) or CD4+ cells are a type of T cell involved in protecting against infections, such as for example, viral, fungal, and protozoal infections. Destruction of these cells is the major cause of immunodeficiency observed in AIDS, and decreasing CD4+ lymphocyte counts appear to be the best indicator for the potential development of opportunistic infections. In judging the severity of HIV/AIDS cases, the CD4+ lymphocyte count is more indicative of the severity of the disease than gross symptomalogy, although it is also true that certain symptoms may be associated with particular CD4+ lymphocyte levels. See, for example, FIG. 1 (see Original Patent). Average normal adult CD4+ cell counts typically ranges from 500 to 1,500/2,000 cells per cubic milliliter of blood.

As CD4+ cell counts decrease below the normal adult levels during primary HIV infection, CD8+ or cytotoxic T-lymphocytes also increase. However, most studies indicate that an increase in CD8 count is not a prognostic indicator of disease progression. Some clinicians in the U.S. use the CD4/CD8 ratio as an indicator of disease progression, however, this ratio varies not only with the severity of the disease, but with the ethnicity of the subject.

There are several systems for classifying and staging HIV infection. The most commonly-used system is the CDC (Centers for Disease Control) Scheme. The CDC scheme has three classifications based upon CD4 counts. The definitions of the three CD4+ T-lymphocyte categories I as follow: Category 1: >500 cells/mm.sup.3 (or CD4%>28%); Category 2: 200-499 cells/mm.sup.3 (or CD4% 14% -28%); and Category 3: <200 cells/mm.sup.3 (or CD4%<14%).

In addition to the CDC classification scheme, there are also 3 possible categories of clinical conditions, which are designated by the letters A, B and C. Therefore, a given individual can have the following CDC classification and clinical categorization designation: 1-A, or 1-B, or I-C, 2-A, 2-B, 2-C, 3-A, 3-B or 3-C.

An individual in category A is identified as an adolescent or adult (>13 years) with documented HIV infection having one or more of the following conditions (and lacking any of the conditions associated with categories B and C): asymptomatic HIV infection; persistent generalized lymphadenopathy; and acute (primary) HIV infection with accompanying illness or history of acute HIV infection.

An individual in category B is identified as an adolescent or adult (>13 years) with documented HIV infection having one or more of the following conditions (and lacking any of the conditions associated with category C) and that meet at least one of the following criteria: (a) the conditions are attributed to HIV infection or are indicative of a defect in cell-mediated immunity; or (b) the conditions are considered by physicians to have a clinical course or to require management that is complicated by HIV infection. Examples of conditions in clinical category B include but are not limited to: bacillary angiomatosis; candidiasis (oropharyngeal, i.e. thrush); candidiasis (vulvovaginal, persistent, frequent, or poorly responsive to therapy); cervical dysplasia (moderate or severe/cervical carcinoma in situ); constitutional symptoms, such as fever (body temperature of 38.5.degree. C. or greater) or diarrhea lasting longer than 1 month; hairy leukoplakia (oral); herpes zoster (shingles), involving at least two distinct episodes or more than one dermatome; idiopathic thrombocytopenic purpura; listeriosis; pelvic inflammatory disease (particularly if complicated by tubo-ovarian abscess); and (11) peripheral neuropathy. For classification purposes, Category B conditions take precedence over Category A conditions. For example, an individual previously treated for oral or persistent vaginal candidiasis (but not exhibiting a Category C disease or condition) who is now asymptomatic, should be classified in Category B.

An individual in category C is identified as an adolescent or adult (>13 years) with documented HIV infection having one or more of the following conditions Category C conditions include the following: candidiasis of bronchi, trachea, or lungs; candidiasis (esophageal); invasive cervical cancer; coccidioidomycosis (disseminated or extrapulmonary); cryptococcosis (extrapulmonary); cryptosporidiosis (chronic intestinal, greater than 1 month's duration); cytomegalovirus disease (other than liver, spleen, or nodes); cytomegalovirus retinitis (with loss of vision); encephalopathy (HIV-related); herpes simplex: chronic ulcer(s) (greater than 1 month's duration), or bronchitis, pneumonitis, or esophagitis; histoplasmosis (disseminated or extrapulmonary); isosporiasis (chronic intestinal, greater than 1 month's duration); Kaposi's sarcoma; lymphoma (Burkitt's, or equivalent term), lymphoma, (immunoblastic, or equivalent term); Lymphoma (primary, of brain); mycobacterium avium complex or M. kansasii, disseminated or extrapulmonary; mycobacterium tuberculosis, (any site, pulmonary or extrapulmonary); mycobacterium, (other species or unidentified species, disseminated or extrapulmonary); pneumocystis carinii pneumonia; pneumonia (recurrent); progressive multifocal leukoencephalopathy; Salmonella septicemia (recurrent); toxoplasmosis of brain; and wasting syndrome due to HIV. For classification purposes, once a Category C condition has occurred, the individual will remain in Category C.

One method of treatment for HIV-positive individuals is the highly active antiretroviral therapy (HAART) regimen. HAART is a therapeutic treatment regime consisting of the combination of anti-HIV drugs, that is prescribed to HIV-positive individuals even before they develop symptoms of AIDS. The therapy usually includes one nucleoside analog, one protease inhibitor and either a second nucleoside analog or a non-nucleoside reverse transcription inhibitor (NNRTI). Frequently, the HAART regime is toxic to the individual, resulting in adverse side effects. For example, HAART can be toxic to blood because it almost always includes one or two nucleoside analogs, like AZT that are notorious for their toxicity to red and white blood cells and blood cell production. Various forms of anemia are very common and sometimes are irreversible. However, it is extremely rare for a subject on the HAART regimen reverse his/her HIV status in Kenya.

Examples of drugs administered for the HAART treatment regime include: azidovudine (AZT), didanosine (dideoxyinosine, ddI), zalcitabine (dideoxycytosine, ddC), lamivudine (epivir, 3TC), nevirapine (Viramune), abacavir (Ziagen), stavudine (Zerit, d4T), tenofovir (Viread), efavirenz (Sustiva), amprenavir (Agenerase), lopinavir (Kaletra), nefinavir (Viracept), saquinavir (Invirase), ritonavir (Norvir), indinavir (Crixivan), and delavirdine (Rescriptor).

Method for Extracting Alkaloid Compounds and Preparing Herbal Composition

The compositions of the invention are prepared using roots of abyssinica and Clutia robusta, and optionally one or more of the following: the stem bark of Prunus africana, stem bark of Croton macrostachyus, stem bark of Acacia nilotica, roots of Rhamnus prinoides, roots of Adenia gummifera, roots of Asparagus africanus, stem bark of Anthocleista grandiflora, whole plant of Plantago palmata, roots of Clematis hirsuta, stem bark of Ekebergia capensis, stem bark of Bersama abyssinica, and roots of Periploca linearifolia. Preferably, the ingredients collected are fresh, although dried samples may also be used. The ingredients are combined and chopped into small pieces and dried. Preferably, the dried ingredients are ground into a fine powder after drying. Alternatively, each ingredient may be processed individually and combined at a later stage. Preferably, if combined for the extraction process, the ingredients are combined in equal weight ratios. Optionally, Dovyalis abyssinica, Clutia robusta, Prunus africana, Croton macrostachyus, Acacia nilotica, Rhamnus prinoides, Adenia gummifera, Asparagus africanus, Anthocleista grandiflora, Plantago palmata, Clematis hirsuta, Ekebergia capensis, Bersama abyssinica and Periploca linearifolia can be present in a weight ratio of 2:2:2:2:2:2:1:2:2:1:2:2:2:2.

The herbal plant material mixture may be extracted with a non-polar solvent to remove fats from the chopped herbal ingredients. Preferably, approximately 20% by volume non-polar solvent is added to the herbal ingredient mixture. Non-polar solvents are generally organic solvents having a dielectric constant less than 20. Non-polar solvents that may be used include, but are not limited to: alkanes, 1,4-dioxane, carbon tetrachloride, chloroform, methylene chloride, benzene, ethers, ethyl acetate, tetrahydrofuran, acetic acid, butanol, chlorobenzene, cycloalkanes, xylene, and the like. Preferred non-polar solvents are xylene and ether.

The non-polar solvent is decanted and discarded. The defatted herbal solids, are then allowed to dry. Sufficient base is added to the defatted herbal material to achieve a pH of approximately 8. The concentration of the base added can be adjusted to provide sufficient liquid volume to cover the defatted herbal solid mixture. Any suitable base may be used, with preferred bases including NaOH, KOH, Ca(OH).sub.2, Mg(OH).sub.2, NH.sub.4OH, and the like. The base extract is then heated for 2-4 hours. Preferably, the ingredients are slowly simmered under reflux conditions, although the same effect can be achieved by simmering the mixture in a covered pot.

Acid is added to the base extract to achieve a pH of approximately 3. Preferably the acid is HCl, although other acids, including but not limited to, HBr, HNO.sub.3, H.sub.2SO.sub.4, H.sub.3PO.sub.4, or any other acid suitable for achieving a pH of approximately 3 may be used as well. The concentration of the acid can be adjusted as necessary to provide sufficient volume to the mixture. The acidified solution is then boiled for approximately 2-4 hours under the same conditions employed for the heating of the basic solution. After heating, the mixture is cooled, and the aqueous layer is separated from the mixture, such as for example, by decanting the liquid from the remaining solids. Acid is then added to the remaining residue sufficient to achieve a pH of approximately 3, and the mixture is then reheated for approximately 2-4 hours under the same conditions previously employed. The aqueous layer is separated from the ingredients and the two acidified layers are combined. If necessary, additional acid extractions may be performed.

The acidic filtrate is extracted several times with a non-polar solvent until little or no emulsion forms. Preferable non-polar solvents are ether and xylene. Base is added to the aqueous layer to precipitate the alkaloid compounds. Preferably, base is added to achieve a pH of approximately 9. The precipitate is separated from the aqueous solution, neutralized and dried.

The precipitate is preferably collected in either crystalline or powder form, and may administered to an subject as a beverage, capsule, tablet, powder, candy, gel, nutritional product or pharmaceutical product.

The precipitate can be further purified as desired to isolate individual alkaloid compounds by any known chromatographic means.

It is understood that at any point during the process of extracting the alkaloid compounds from the herbal ingredients that the aqueous solution can be concentrated and stored for later use without the need for precipitation of the compounds from solution.

Alternatively, the alkaloid compounds for use in the present invention can be synthesized by known methods once the chemical structure has been determined. Isolated compounds can be analyzed by chemical analysis, mass spectroscopy, infrared spectroscopy, X-ray diffraction, NMR (including .sup.1H NMR, .sup.13C NMR, COSY, NOSEY, and the like), and other known analytical techniques to obtain the chemical structures. For example, chemical structures for four extracts obtained from abyssinica have been previously determined. (See, for example, http://www.dfuni.dk/uploads/media/Naturstofgruppen_BonnieRasmussen.pdf).
 

Claim 1 of 14 Claims

1. A composition comprising: dried root of abyssinica; and dried root of Clutia robusta.
 

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