JP2002542286A - Cardiovascular and bone treatment with isoflavones - Google Patents

Abstract

  (57) [Summary] Optionally, with one or more carriers, excipients, adjuvants, and / or diluents, the therapeutically effective ratio of formononetin to the isoflavone is in the range of 15: 1 to 2: 1; And / or a composition comprising one or more isoflavones selected from biochanin, genistein and daidzein. Formononetin and one or more isoflavones selected from biochanin, genistein and daidzein, optionally, one or more carriers, excipients in a ratio of formononetin to the isoflavone of 15: 1 to 2: 1 with therapeutic effect Compositions that may be included with agents, adjuvants and / or diluents are described. It also contains such a composition and prevents and / or treats circulatory diseases, improves blood lipoprotein levels, or reduces the risk of vascular disease, or reduces the risk of ischemic heart disease, arteriosclerosis A method for improving or maintaining bone density, such as reducing the risk of osteoporosis and / or preventing and / or treating bone fractures, will be described.

Description

DETAILED DESCRIPTION OF THE INVENTION

The present invention relates to the treatment and / or prevention of cardiovascular diseases and osteoporosis using isoflavone compounds. More particularly, the present invention relates to compositions, uses and methods comprising certain plant isoflavones, and more particularly to the prevention and / or treatment of cardiovascular diseases, or the improvement of blood lipoprotein levels, or ischemic heart disease The present invention relates to a composition having a high formononetin content for reducing bone risk, reducing the risk of arteriosclerosis, preventing or treating osteoporosis, and / or improving or maintaining bone density in preventing / treating fractures.

BACKGROUND OF THE INVENTION Note: Citations are collected at the end of the specification. Cardiovascular disease and osteoporosis have become a major social health problem in the western world, where life expectancy is increasing, and in non-western areas, where lifestyle, especially diet, is increasing. Current treatment and prevention options for these two illnesses are unsatisfactory, such as those targeting specific symptoms and not addressing the underlying pathogenic mechanisms, or those whose doses are limited by undesirable side effects. Absent. Aimed at underlying biological events that cause cardiovascular disease and osteoporosis, can be used to treat already ill conditions, prevent the onset of illness, and can be used for long periods without side effects There is an urgent need to develop safer and more effective treatments.

[0003] A major cause of cardiovascular disease is the disease of the arterial wall known as atherosclerosis. Atherosclerosis is characterized by the deposition of fatty spots in the vessel wall and the inflammatory response induced by the spots. The consequence of this event is a hypertrophy of the vessel wall and a consequent decrease in the internal volume of the arterial lumen. This consequence leads to two main symptoms-(a) the end organs, usually the heart (causing ischemic heart disease), the kidney (causing renal failure), and the brain (causing senile dementia). Restriction of blood flow to
b) Acute cerebral ischemia by moving through the bloodstream until atherosclerotic flakes are detached and remain in narrow blood vessels as thrombogens, damaging certain tissues of the blood vessels.

An important factor that facilitates the progression of atherosclerosis is the level of cholesterol in the blood, or more specifically, the form in which cholesterol is present in the blood.
Cholesterol is an important structural component of cells and is needed by most cells every day. Cholesterol binds to proteins known as several different types of apoproteins and is transported to cells through the blood. Cholesterol and apoprotein combine to form particles known as lipoproteins. Cholesterol is a low-density lipoprotein that consists of a specific type of apoprotein combined with a small amount of cholesterol molecules (
(Abbreviated as LDL) are delivered to cells in the form of particles known as LDL. In that tissue, cholesterol is released from the carrier apoprotein and used in cells. Excess cholesterol exists in free form until collected by other types of apoproteins. This cholesterol returns to the liver for recycling in the form of particles known as high density lipoproteins (abbreviated HDL). In healthy people, the ratio of LDL to HDL ranges from about 2: 1 to 2.5: 1. It is generally believed that this ratio does not result in excessive cholesterol deposition in the tissue. As this ratio increases, the body's ability to recycle excess cholesterol decreases, leaving free cholesterol in tissues such as arterial walls. Free cholesterol is particularly susceptible to oxidation in the arterial wall. Oxidized cholesterol is very inflammatory and can cause inflammation in surrounding tissues.
Atherosclerotic plaques are a combination of accumulated oxidized cholesterol and inflammatory tissue.

[0005] Increasing the LDL: HDL ratio above 3: 1 is generally believed to be associated with an increased risk of atherosclerosis. The vast majority of Westerners are about 3: 1 to 7: 1. Apart from those with a familial predisposition to the disease,
The main causes of this imbalance are lifestyle factors and age. It is well known that this imbalance is due to abnormally elevated LDL levels, or abnormally low HDL levels, or both. L increased
Factors known to be associated with DL levels are mainly in foods, for example those containing high levels of animal fat and low levels of complex carbohydrates. Low level HD
Factors known to be associated with L are lack of exercise and aging. Age-related effects at the HDL level are primarily responsible for causing high LDL: HDL ratios in the elderly, especially women. This is because HDL production is related to estrogen levels in the body, and estrogen levels decrease with age in both women and men.

[0006] While total cholesterol levels in blood are considered to be a relevant risk factor for atherosclerosis, the range of normal values for total cholesterol in blood is now very wide, and the more relevant risk factor is LDL. : HDL ratio. That is, in cardiovascular risk terms, the absolute levels of both lipoprotein types are less important than the relative ratio of LDL and HDL.

[0007] It is believed that individuals with abnormally high LDL: HDL ratios may be able to restore LDL levels, increase HDL levels, or a treatment strategy with both to a normal ratio of about 2.5: 1. Currently, three general methods are used, with the primary treatment being aimed at lowering LDL levels. First, the first method uses a drug that inhibits cholesterol synthesis. So-called "statins", such as ethyl-2- (p-chlorophenoxy) -2-methylpropionate, inhibit cholesterol biosynthesis in the liver and reduce blood cholesterol levels. These drugs generally reduce blood LDL levels by between about 10-40%. The second method is to reduce the absorption of cholesterol from the intestinal tract, thereby reducing the storage of cholesterol in forms available to the body. Historically, binders have been used, such as insoluble high molecular weight polymers, which combine with bile acids to form complexes excreted from stool. More recently, plant sterols have been found to achieve similar results. Increasing bile acid excretion, including either substance, reduces LDL levels, typically in the range of 5-12%. The third method generally reduces total cholesterol and LDL levels to about 8-
It is a method of using soy protein which reduces 12%. The mechanism of action of this substance is unknown. This approach aimed at lowering LDL levels has a number of deficiencies. First, it has been hypothesized that there is a correlation between reduced LDL levels and reduced risk of atherosclerosis and ischemic heart disease, but no clear clinical evidence supports this assumption. Second, most of the current therapeutics have undesirable side effects. Statins are used for gastrointestinal symptoms such as nausea, vomiting, loose stools, indigestion, abdominal pain,
There are frequent side effects such as complications of cardiovascular diseases such as angina, arrhythmia, and skin diseases and various other common complications. Resin products have side effects such as gastrointestinal disorders, constipation, worse hemorrhoids and abdominal discomfort.

[0008] Another therapeutic option is to raise HDL levels. This choice has been considered by medical professionals to be the preferred choice for several reasons. First of all, the most critical evidence of clinical benefit arising from the re-adjustment of the LDL: HDL ratio is in strategies to increase HDL levels. Gord
on et al. (1989) (Circulation 79: 8-15) is 1mg / 100m of HDL cholesterol in blood
An increase in g (1%) was shown to reduce the risk of death from ischemic heart disease by 3%. Second, HDL appears to have a more effective effect on the arterial wall than on removing oxidized cholesterol. Third, abnormally high LD, especially in older women
The main reason for the L: HDL ratio is a decrease in HDL levels. The choice of treatment here is more limited when compared to those targeting LDL levels. The most effective treatments are steroidal estrogens such as estradiol. Estradiol and estrogen replacement therapy generally increase HDL levels between about 15-30% in postmenopausal women with little effect on LDL levels. However, estrogen replacement therapy is associated with a number of circulatory side effects, including increased propensity for thrombogenesis, leading to increased risk of blood clotting and heart attack. This makes estrogen replacement therapy less attractive for older women. The feminizing effect of estrogen does not make it a more attractive option for men. Another drug, known as clofibrate, raises HDL levels by about 10% in both men and women, but is rarely used due to its side effects. Given that the most conclusive clinical evidence of a beneficial effect on atherosclerosis by modulating cholesterol levels is from elevated HDL levels, drugs that provide a safe and effective means of achieving this result There is a big challenge, not the present.

[0009] Similar to cardiovascular disease, loss of bone density is becoming a major health problem in the prolonged life of the western world. As HDL levels decrease with age, a decrease in bone density appears to be primarily associated with a decrease in estrogen levels in the body. One of the biological effects of estrogen is the stimulation of osteoblasts, which are responsible for the production of new bone and also have an inhibitory effect on osteoclasts. Is responsible for bone resorption. Lower estrogen levels reduce osteoblast activity, maintain osteoclast activity, and reduce the production of new bone that replaces old bone that is removed. As a result, bone mass gradually decreases. The initial stage of this condition is known as osteopenia. The latter stage is known as osteoporosis. In osteoporosis, bone density falls into a state where fractures easily occur.

[0010] The body has two major bone tissues, trabecular bone and cortical bon.
e) there. Sponge bone, which makes up about 80% of the bones of the body, is a low density, reticulated bone.
The cancellous bone is rich in those bone parts that are not highly weight-bearing, such as the spine, ribs, skull, carpal and tarsal joints. Cortical bone makes up about 20% of the bones of the body and is very dense. Cortical bone is rich in load-bearing conditions such as the long bones of the extremities and the femoral neck of the hip. Bone density usually begins to decline from middle age in both men and women, affecting cancellous bone and cortical bone, and cancellous bone generally has a higher turnover speed in vivo compared to cortical bone, The cancellous bone generally decreases significantly in the early stages of this bone loss process. In women, menopause accelerates this process. As menopause approaches, trabecular bone has an approximately eight-fold turnover rate compared to cortical bone. Nearly menopausal women lose about 2-3% a year of cortical bone, while about 4% of cancellous bone
-8% reduction. By the age of about 60, both cancellous and cortical bones will decrease at a slower rate. This age-related effect is related to the phenomenon of menopausal women. In the age of 50-60, fractures are commonly found in the spine, carpal joints, and ribs (mainly trabecular bone). It is commonly found in hip joints and long bones (mainly cortical bone). Hip and femoral fractures are the most severe of the various fractures and require long hospital stays and usually major surgery. One-third of older women with hip fractures die from related complications within 12 months of the fracture.

There are various treatment options for treating and preventing osteoporosis. Steroidal estrogens such as estradiol and synthetic derivatives such as raloxifene are well known and widely used for this purpose. These compounds act in combination to promote bone deposition and suppress bone resorption. However, their effects are mainly on cancellous bone, with little or no effect on cortical bone. As a result of this choice, fractures of the carpal joints, ribs, spine, etc. may be prevented, but little or no severe hip and femoral fractures. A group of compounds known as bisphosphonates are also commonly used. These compounds reduce bone resorption and act on cancellous and cortical bone, but, like estrogen, act primarily on cancellous bone. Other treatments include calcitonin, which slows bone resorption, and ipriflavone, which suppresses bone resorption and increases osteoblast function. All of these drugs have undesirable side effects. If long-term treatment of 10-30 years is required for effective treatment and prevention of fractures, it is important that the patient is safe and tolerable, and all the above treatments are safe. The patient's consent is difficult to obtain due to the low level.
There is an urgent need to develop highly safe and tolerable treatments, especially for the protection of cortical bone and to promote long-term use.

In recent years, plant compounds known as isoflavones having an estrogenic function, such as genistein, daidzein and its methyl ester, biocanin and formononetin, are of particular interest. In part, this interest stems from epidemiological observations that people in areas where food is rich in isoflavones have less circulatory disease and osteoporosis. In part, it also stems from their estrogenic function and the possibility that they mimic the health benefits of estradiol, especially in the effects of estradiol in cardiovascular disease and in increasing bone density. Most scientific interest has focused on genistein and daidzein because of the four strongest estrogen agonists among the four isoflavones. Genistein and daidzein are said to have about 0.1% of the estrogenic function of estradiol, while formononetin and biochanin are about 10-100 times weaker.

[0013] There are few studies in the literature on osteoporosis and isoflavones. US Patent 5423331
The publication discloses the use of genistein and daidzein as components of an extensive mixture of specific compounds in the prevention and / or treatment of osteoporosis in humans. However, the patent does not mention the usefulness of isoflavones such as formononetin and biochanin, or the effect of specific isoflavone ratios, or the beneficial effect of isoflavones alone, or on cancellous or cortical bone alone or with other substances. It does not teach the relative effects of isoflavones in combination.

[0014] There is experimental evidence that genistein has beneficial effects on bone. Genistein promotes bone formation (Fanti, 1998) and suppresses osteoclast activity (US Pat. No. 5,506,621, Barne
s, S and Blair, HC: genistein for inhibiting osteoclasts). Low doses of genistein have been reported to increase bone mineral density in both cortical and trabecular bone in rats (Anderso, 1998). Although isoflavones such as genistein, daidzein, formononetin and biochanin are known to have certain common biological properties, it is well known that they differ significantly in biological potential. Thus, the effects of formonenetin, biochanin or daidzein on bone biology are still unknown.

The only clinical study using osteoporotic isoflavones involving 66 postmenopausal women treated for 6 months in a placebo randomized trial with soy products containing intermediate or high levels of isoflavones has been reported. (Potter SM et al., "Soy Proteins and Isoflavones: Effects on Blood Lipids and Bone Mineral Density in Postmenopausal Women."
n Journal of Clinical Nutrition 1998: 68 (supplement) 1375S-1379S). It is well known that soybeans contain daidzein and genistein at a ratio of about 1: 2, and contain little formononetin or biochanin. The results of this study showed that high isoflavone substances increased lumbar spine bone mineral content and density by 2%, but had no effect on femoral bone mineral content and density. These results suggest that daidzein and genistein have a modest effect on trabecular bone but no effect on cortical bone.

The literature is somewhat clear about isoflavones and lipoprotein levels. There are many studies on animals and humans that have administered foods themselves, such as soybeans and other legumes, and relatively crude extracts of soybeans and other legumes, and followed lipoprotein levels. At best, these data are very uncertain and variable. More importantly, however, the use of such crude products necessarily involves the use of many plant components that are rich in saponins and sterols, which are known to have a regulatory effect on cholesterol metabolism. This makes it impossible for those skilled in the art to derive a correlation between isoflavones and blood lipoprotein levels.

The most striking evidence came from studies in which the supplement was very rich in isoflavones and was unchanged for other foods. Three studies using soy extracts rich in isoflavones such as genistein and daidzein have been reported. Two studies found that supplemental diet failed to show a significant effect on LDL and HDL levels (Nsetel et al. (1997) "Soy isoflavones improved systemic arterial compliance in menopausal and near-menopausal women. Did not improve plasma lipids. "Arteriosclerosis, Thrombosis and Vascular Biology Biol 17:33
92-3398 ”and Hodgson et al. (1998)“ Isoflavonoid phytoestrogen supplementation did not alter serum lipid levels: a randomized controlled trial in humans ”,“ Journal of
Nutrition 128, 728-332). In a third study (such as Potter SM above), 6 months of treatment with soy flour in postmenopausal women with hypercholesterolemia increased HDL levels by an average of 4.3% and LDL levels by an average of 8%. Was seen. The first two studies show that isoflavones are added in a very concentrated form and are a more reliable indication that soy isoflavones are ineffective at lipoprotein levels where dietary adjustments are rarely needed. It is generally considered. A third study found that in addition to containing a wide variety of soy components, such as saponins and sterols, which have cholesterol-modulating effects, dietary habits can be altered by the weight of protein present in soy products. The isoflavones were administered in the form of soy flour.

US Pat. No. 5,585,892 (Potter) discloses a method of changing the concentration of a cholesterol component in human blood using isoflavone daidzein. Potter discloses the use of soy protein and the isoflavones genistein, daidzein, glycitein and their respective glycosides. An approximately 5.2% increase in HDL-cholesterol levels in patients receiving the soy protein / isoflavone composition was reported. It is believed that what is administered has soy protein components that can raise HDL levels very slowly, but it is not clear what the active substance of the composition is.

In a further study by Nestel et al. (1999 "Isoflavones from red clover improve systemic arterial compliance but not plasma lipids in menopausal women," Journal of Clinical Endocrinology and Metabolism 8
4: 895-898), discloses that administration of isoflavones from red clover containing biocanin, formonenetin, daidzein and genistein in a ratio of 1.8: 1.2: 0.2: 0.1 had no effect on plasma lipids. In other studies administered premenopausal women of normal cholesterol level using similar supplement diet isoflavones from red clover, but HDL ₃ subfractions increased slightly supplemented diet LDL Ya HDL levels isoflavones It was found that there was no statistically significant effect (S
Amman S et al. "Effects of isoflavone supplementation on plasma lipids and susceptibility to oxidation of low-density lipoproteins in premenopausal women" Atherosclerosis 147, 277-283 (1
999)).

A logical summary of the prior art would indicate that isoflavones from the group of genistein, daidzein, formononetin and biochanin, alone or in combination, have little or no effect on blood lipoprotein levels.

Summary of the Invention Formonenetin is present in a high ratio compared to one or more isoflavones selected from biocanin, genistein and daidzein, wherein the ratio of formonenetin to the isoflavones is from 15: 1 to 2 which is therapeutically effective. A composition which is optionally used together with one or more carriers, excipients, adjuvants and / or diluents, is useful for the prevention and / or treatment of cardiovascular diseases, Improve blood lipoprotein levels, reduce the risk of ischemic heart disease, or reduce the risk of atherosclerosis,
It is surprising that the inventors have found it useful for preventing or treating osteoporosis and / or improving or maintaining bone mineral density in preventing and / or treating bone fractures.

The unique health benefits of compositions containing such high levels of formonenetin are also surprising because they are completely unpredictable for two main reasons.
First, isoflavones' effects on the circulatory system and bone are linked to their effects on estrogen, and formonenetin is generally the weakest estrogen in the group of isoflavones, consisting of genistein, daidzein, formononetin and biochanin. That was thought. Second, formonenetin is efficiently demethylated into daidzein in the human body, and formonenetin was thought to have the same function as daidzein. According to a first aspect of the present invention, formononetin and one or more isoflavones selected from biochanin, genistein and daidzein are used as therapeutically effective ratios of formononetin and the isoflavones in a ratio of 15: 1 to 2 : 1 is provided, which can optionally be used with one or more carriers, excipients, adjuvants and / or diluents.

Preferably, the formononetin and one or more isoflavones selected from biochanin, genistein and daidzein are provided in the form of extracts from chickpea, clover or other plant sources having a high formononetin content. The extract can be prepared by extraction of the water / organic solvent system of legumes using isoflavone extraction procedures well known to those skilled in the art. Alternatively, isoflavones can be made by established synthetic techniques well known to those skilled in the art. Formonenetin can be used with one, two or three isoflavones selected from biochanin, genistein and daidzein. Preferably, formononetin is used with biocanin at the trace level or as low as 0.1-5% of genistein and daidzein or isoflavones.

According to another aspect of the present invention, treatment and / or prevention of circulatory disease, or improvement of blood lipoprotein level, or reduction of risk of ischemic heart disease, or reduction of risk of arteriosclerosis, Treatment or prevention of osteoporosis and / or prevention of fracture and / or
Or 15: 1 to 2 in the manufacture of medicines for improving or maintaining bone mineral density for treatment
The use of isoflavones selected from formononetin in a ratio of 1: 1 and one or more biocanins, genistein and daidzein is provided.

[0025] In another aspect of the invention, the treatment and / or prevention of circulatory disease, or the improvement of blood lipoprotein levels, or the reduction of the risk of ischemic heart disease, or the reduction of the risk of arteriosclerosis In order to improve or maintain bone density as in the treatment or prevention of osteoporosis and / or the prevention and / or treatment of fractures, formononetin and one or more isoflavones selected from biochanin, genistein and daidzein are used. A composition comprising a 2: 1 to 2: 1 ratio, optionally with one or more carriers;
There is provided a method comprising administering to a human patient with excipients, adjuvants and / or diluents.

DETAILED DESCRIPTION The composition of the present invention comprises formononetin and one or more isoflavones selected from biochanin, genistein and daidzein, wherein the ratio of formononetin to said isoflavones is a therapeutically effective ratio. : 1 to 2: 1, optionally 1
It contains the above carriers, excipients, adjuvants and / or diluents. Formononetin may be present with one or more biocanins, genistein and daidzein. When formononetin is present with a single isoflavone, the isoflavone is preferably biochanin, but biochanin can be replaced by genistein or daidzein. Two of biocanin and genistein, two of biochanin and daidzein,
Or when two of genistein and daidzein are present with formononetin, they may be in the same amount by weight, or the first isoflavone is 5-95% by weight and the second isoflavone is corresponding The amount may be.

If the composition consists of formononetin and biochanin, genistein and daidzein, and the ratio of formononetin to other isoflavones is from 15: 1 to 2: 1, biokanin, genistein and daidzein are in the same amount on a weight-to-weight basis. Alternatively, the amounts of these isoflavones are not critical to the invention. Thus, one unit of the combination of biochanin, genistein and daidzein is 0.1 to 0.99 units of biocanin, 0.1 to 0.99 units of daidzein, 0.1 to 0.9
Contains 9 units of daidzein, other isoflavones 1 (not formonenetin)
Unit. Of particular importance for the present invention is that the content of formonenetin is high relative to other isoflavones, especially biochanin, genistein and / or daidzein.

The effect of the high formononetin ratio is to increase HDL levels in the blood more unexpectedly, which is very beneficial in preventing atherosclerosis and ischemic heart disease It is known. As also shown below, the finding that the effects of this composition as described in detail in the present invention have a very dramatic effect on cortical bone density is also unexpected, and It suggests a clinical benefit that is important for the risk of fractures, such as the onset and progression of osteoporosis, and, in particular, the hip, humerus, femur, radius, and ulna. The magnitude of these biological effects and the clinical consequences that can be obtained with the compositions described in the present invention are much greater than the magnitude of the effects generally known for isoflavones, which This clearly shows that the function is at a specific isoflavone ratio.

[0029] Preferably, formononetin and one or more isoflavones selected from biochanin, genistein and daidzein are provided in the form of extracts from chickpea and clover having a high formononetin content. This extract is preferably an extract in a water / organic solvent system, and isoflavone extraction procedures are known to those skilled in the art.

Clover, for example, red clover, is a preferred source of formonenetin and other of the above isoflavones. The clover that can be used is red clover (T
. Pratense) or T. Subterranean. Many types of red clover and other varieties of clover are known and developed. These legumes can be used in the present invention. The above isoflavones are preferably prepared by extracting legumes with a water / organic solvent.

The collected plant material is ground or ground into small pieces, or is partially ground or cut into small pieces and chopped, or, generally, without any pre-treatment, in water and water miscible Contact with an organic solvent such as an organic solvent. The ratio of water to organic solvent is generally in the range of 1:10 to 10: 1, for example, in the same ratio of water and organic solvent, or in the range of 1% to 30% (vol / vol) of organic solvent. May be used. Any organic solvent or mixture of such solvents can be used. The organic solvent is preferably C2-10
, More preferably a C1-4 organic solvent (eg, methanol, chloroform,
Ethanol, propanol, propylene glycol, erythritol, butanol, butanediol, acetonitrile, ethylene glycol, ethyl acetate, glycidol, glycerol, dihydroxyacetone, or acetone) are preferred.
The extract is prepared in this regard by contacting the plant material with a water / solvent mixture. Optionally, the mixture may include an enzyme that breaks down the isoflavone glycosides to the aglycone form. The mixture may be stirred vigorously to form an emulsion. The mixing temperature may range, for example, from ambient temperature to the boiling point. The contact time can be from one hour to several weeks. One convenient extraction time is 24 hours at 90 ° C. The extract is separated from insoluble plant material and the organic solvent is removed by distillation, rotary evaporator or other standard procedures for removing organic solvents. The obtained extract containing the water-soluble substance and the water-insoluble component is dried to obtain an isoflavone-containing extract, which is formulated with one or more pharmacologically acceptable carriers, excipients and / or adjuvants. can do.

The extract after distillation contains a small amount of oil containing isoflavones in aglycone form (herein referred to as isoflavones). The isoflavone-enriched oil can be subjected to HPLC to adjust the isoflavone ratio, or if desired, e.g., drying the isoflavone in the presence of silica and extracting the isoflavone-containing extract with one or more carriers, excipients and And / or formulated with adjuvants. In another method, the isoflavones are concentrated by adding a non-aqueous, soluble organic solvent that is hexane, octane, acetone or a mixture of one or more such solvents to the oil. 1
As one example, 80% hexane, 20% acetone (weight / weight) is highly soluble in oil and does not dissolve isoflavones very well. The oil is easily partitioned into the organic solvent and an isoflavone-rich extract precipitates from the solvent. The recovered extract can be dried in an oven, for example, at 50 ° C. to about 120 ° C., and formulated with one or more pharmaceutically acceptable carriers, excipients, and / or adjuvants. The ratio of isoflavones in the extract from legumes is particularly high in the formofonenetin content relative to other isoflavones, the ratio of isoflavones is easily obtained, and, for example, the use of clovers with a high formononetin content By doing so, it can be adjusted by concentration, HPLC fractionation and the like using various solvents as described above.

As a preferred legume raw material, clover can be easily extracted with water / organic solvents.
A single source of clover may be used, or may be used in combination with one or more different clovers and / or chickpeas.

Formononetin and the other isoflavones referred to herein can be produced synthetically by methods well known in the art. For example, Kagal et al., Tetrahedron
Letters 1962,593; Mahal et al., J. Chem. Soc. 1934,1769; Wahala et al., Proc So
c See Biol Med 208, 18 (1995), 27-32. The compositions of the present invention can employ one or more pharmacologically acceptable carriers.
The carrier is selected to be acceptable in terms of the components of the composition and must not be harmful to the patient. The carrier may be solid or liquid, or both, and may be formulated with an extract containing the isoflavones as a unit dose in the desired ratio, for example, as a tablet, which may contain the carrier in an amount of 0.5% by weight of the extract. To 80%, or up to 100% by weight of the extract. The composition can be made using any of the known techniques for pharmacy. For example, the extracts can be prepared by techniques well known in the pharmaceutical arts, optionally mixing with excipients, diluents (eg, water) and adjuvants.

The composition of the present invention comprises vitamins (for example, vitamin A, vitamin B group, vitamin
Contains one or more drugs such as C, vitamin D, vitamin E or vitamin K) and minerals (e.g. magnesium, iron, zinc, calcium or manganese in the form of pharmaceutically acceptable salts) Is also good.

The compositions of the present invention include compositions suitable for oral, rectal, ocular, buccal (eg, sublingual), parenteral (eg, subcutaneous, intramuscular, intradermal, and intravenous) and transdermal administration. be able to. In any case, the most appropriate route will depend on the nature and severity of the condition being treated and the condition of the patient.

Compositions suitable for oral administration can be presented in discrete units such as capsules, cachets, troches or tablets. Each contains a predetermined amount of extract as a powder or granules; as a solution or suspension in a water-soluble or water-insoluble liquid; or as an oil-in-water or water-in-oil emulsion. Such compositions may be prepared in a suitable manner of preparing the preparation which comprises the step of bringing into association the active compound with one or more suitable carriers, which may contain one or more accessory ingredients as described above. In general, the compositions of the present invention are prepared by uniformly mixing the extract with a liquid or a finely divided solid carrier, or both, and, if necessary, shaping the resulting mixture. For example, a tablet may be a powder or granules containing the extract,
It is manufactured by incorporating or molding together with the above auxiliary components. Compressed tablets are made by compressing the extract, in powder or granule form, optionally with a binder, lubricant, inert diluent, and / or surface active agent / dispersant, in a suitable machine. Can be. Molded tablets may be made by molding in a suitable machine, the powdered compound moistened with an inert liquid binder.

Suitable carriers are sugars, for example, fillers such as lactose, sucrose, mannitol or sorbitol, cellulose preparations and / or calcium phosphates, for example tricalcium phosphate or hydrogen hydrogen phosphate, and also for example corn, wheat, rice or Starch paste using potato starch, gelatin, tragacanth, methylcellulose and / or polyvinylpyrrolidone, and, if desired, the above-mentioned starch, carboxymethyl starch, cross-linked polyvinylpyrrolidone, agar, alginic acid or a salt thereof such as sodium alginate, etc. Are used. For excipients, for example,
Glidants and lubricants such as silicic acid, talc, stearic acid or salts thereof, for example magnesium and calcium stearate, and / or polyethylene glycol can be used. Dragee cores are suitable, optionally enteric coatings, concentrated sugar solutions containing, inter alia, gum arabic, talc, polyvinylpyrrolidone, polyethylene glycol and / or titanium oxide, or suitable organic solvents or solvent mixtures. Or, to form an enteric coating, a solution of a suitable cellulose preparation such as, for example, acetylcellulose phthalate or hydroxypropylmethylcellulose phthalate is used. Dyestuffs or pigments may be added to the tablets or dragee coatings, for example, for identification purposes or to indicate different doses of active ingredient.

Other orally administrable pharmacological compositions are, for example, dry-filled capsules made of gelatin and soft, sealed capsules made of gelatin and a plasticizer, such as glycerol or sorbitol. Dry filled capsules may, for example, be prepared by mixing granules with a filler such as lactose, a binder such as starch, and / or a glicant such as talc or magnesium stearate, and, where appropriate, stabilizing agents. The extract may be contained in the form of

Formulations suitable for buccal tablet (sublingual) administration are lozenges containing isoflavone-based extracts, usually sucrose, and acacia or tragacanth, or inert bases such as gelatin and glycerin or sucrose and acacia. Includes incense tablets containing the compound.

Compositions suitable for parenteral administration of the present invention are easy to use, which include a sterile, water-soluble formulation of the extract, the formulation being similar to the osmotic pressure of the intended recipient's blood. Is preferred. These formulations are also effective for subcutaneous, intramuscular or intradermal administration, but intravenous administration is preferred. Suitable compositions include water-soluble extracts, suspensions of the active ingredients such as the corresponding oily injection suspensions, and fatty oils such as sesame oil, ethyl oleate, or synthetic fatty acid esters such as triglycerides. A suitable oily solvent, such as
Included are vehicles or aqueous injection suspensions containing, for example, sodium carboxymethylcellulose, a thickening agent such as sorbitol or dextran, and optionally, stabilizers and the like. By way of example, the composition can be prepared simply by mixing the extract with water or a glycine buffer and sterilizing the resulting solution to the same osmotic pressure as blood. The injectable preparations according to the invention contain the extract in a weight / volume of 0.1% to 60% and can be administered, for example, at a rate of 0.1 ml / min / kg.

Formulations suitable for rectal administration are preferably presented as unit dose suppositories. These can be made by mixing the extract with one or more solid carriers such as cocoa butter and molding the mixture.

Compositions suitable for transdermal administration are preferably provided as any adhesive suitable to be maintained in intimate contact with the epidermis of the recipient for an extended period of time. Such patches may optionally contain the extract in an aqueous buffer. Compositions suitable for transdermal administration can also be administered by iontophoresis (see, eg, Pharmaceutical Research 3 (6): 318 (1986)) and are generally in the form of any buffered aqueous solution of the extract. Is adopted. Such a composition may comprise, for example, citric acid or a bis / tris buffer (pH 6), or ethanol / water, eg, 0.05% of the extract.
To 30% w / w.

The compositions can be made in conventional manner, in dosage forms and in amounts. For example, Goodman & G
illman, The Pharmacological Basis of Therapeutics (7th edition, 1985) and Re
See mington's Pharmaceutical Science (Mack Publisher, 10th edition). Both are hereby incorporated by reference. The composition may be, for example, from 0.1 mg to 2 g.
For example, 0.1 mg to 200 mg, preferably 15 mg to 50 mg of isoflavones, and the ratio between isoflavones (weight / weight basis) is as described above.

The compositions of the present invention can be administered to a human subject in the form of a supplement. A supplement containing the active composition can be made by adding the composition to food in a food manufacturing process. Any food can be used, including but not limited to minced meat, emulsified meat, marinated meat; nutritional drinks, sports drinks, protein-fortified drinks, juices, milk, milk substitutes, diet drinks, etc. Beverages; cheeses such as hard and soft cheeses, cream cheeses, cottage cheeses; frozen desserts such as ice cream, ice milk, low fat frozen desserts, dairy-free desserts; yogurt; soups; puddings; Dressing; includes sources and spreads such as mayonnaise, margarine, butter, butter substitutes and other fat-containing spreads. The composition is added to the food in an amount selected to administer the desired amount of the composition to the consumer of the food.

The above-mentioned isoflavones may be in the form of powder, slurry, or aqueous solution (for example, containing a small amount of oil), particles, or dissolved in an organic solvent (for example, methanol, ethanol, ethyl acetate or dimethyl sulfoxide). What you have.

An effective amount of a composition of the present invention is administered to a subject. The actual dosage administered will depend on many factors, including the particular mode of administration, the condition being treated, the condition of the patient, and the judgment of the health care professional. An example of an isoflavone dosage volume is about 0.1 mg to 200 mg per day based on 1.5 mg / kg (body weight) / day. A convenient dosage comprises between 25 mg and 50 mg of isoflavones as described herein.

The composition of the present invention contains formonenetin as the main isoflavone. In the present invention, the preferred ratios of formononetin to the other three major isoflavones, genistein, daidzein and biochanin, also include the various naturally occurring forms of isoflavones, including their aglycone, glycoside, acetyl or malonyl forms. I found out.

According to another aspect of the present invention, formononetin and one or more isoflavones selected from biochanin, genistein and daidzein have a ratio of 15: 1 to 2: 1.
In the prevention and / or treatment of cardiovascular diseases, improvement of blood lipoprotein levels, reduction of the risk of ischemic heart disease, prevention and treatment of osteoporosis, and improvement and maintenance of bone density in the prevention and treatment of fractures Provided for use in the manufacture of a medicament.
Form ononetin and one or more of biochanin, genistein and daidzein,
The ratio of formononetin to other isoflavones alone or in combination is 15:
It is provided from 1 to 2: 1, preferably from 10: 1 to 5: 1. Compositions made therefrom can be administered to humans.

According to another aspect of the present invention, prevention and / or treatment of cardiovascular disease, improvement of blood lipoprotein level, reduction of risk of ischemic heart disease, reduction of risk of arteriosclerosis, osteoporosis The present invention provides a therapeutic method for improving and maintaining bone density in the prevention and / or treatment of bone fracture and / or the prevention and / or treatment of fracture. The method comprises formonenetin and one or more isoflavones selected from biochanin, genistein and daidzein, wherein the ratio of formonetin to the isoflavones is from 15: 1 to 2: 1 with a therapeutic effect, optionally Administering to a human patient a composition comprising one or more carriers, excipients, adjuvants and / or diluents.

According to a further aspect of the present invention, there is also a method for improving blood lipoprotein levels. In this aspect, HDL levels increase and / or LDL levels decrease. Accordingly, a method is provided for increasing a subject's HDL level. In another aspect, a method for reducing LDL levels in a subject is provided.

[0052] Aspects of the method of the invention can be extended to methods of reducing the likelihood of thrombogen formation in a human.

[0053] A further aspect of the method of the present invention provides a method for reducing the risk of cardiovascular disease, ischemic heart disease and / or arteriosclerosis.

A further aspect of the method of the present invention provides a method for improving or maintaining bone density in the treatment and / or prevention of osteoporosis.

A further method aspect is a method of preventing and / or treating a fracture involving mainly cortical bone tissue, including the femoral neck, femur, humerus, radius, ulna, tibia (including promoting healing). ) I will provide a.

Each of the above-described methods comprises a therapeutically effective formononetin, biocanin,
A therapeutically effective ratio of formononetin to said isoflavone of from 15: 1 to 2: 1, optionally comprising one or more carriers, excipients, supplements, comprising one or more isoflavones selected from genistein and daidzein Administering the composition comprising the agent together with the agent and / or diluent to a human subject. Subjects treated are postmenopausal women with normal cholesterol levels or hypercholesterolemia, women with atherosclerosis, postmenopausal women with low LDL, hypercholesterolemia or normal cholesterol and / or atherosclerosis. Men with sclerosis.

[0057] Oral administration in solid dosage forms such as tablets and capsules is preferred. Taking at least once a day is the standard method of administration. For example, administration can be continued until blood lipid levels are at an appropriate level. However, maximizing the lipid ratio, preventing circulatory disease, reducing the risk of ischemic heart disease, reducing the risk of arteriosclerosis, or preventing osteoporosis, and / or improving bone density to prevent fractures For a long period of time, such as one year or more.

As mentioned above, the ratio of formononetin to other isoflavones used in the compositions, methods, and uses of the present invention may increase HDL (albeit increasing total cholesterol) and increase cortical bone density. Produces a surprising and maximum beneficial effect on The present invention will be described with reference to the following examples, but is not limited to these examples.

Example 1 Tablets containing an extract enriched in isoflavones were prepared by methods known in the fields of pharmacology and phytochemistry. In particular, isoflavones are extracted from legumes such as red clover by a standard water / alcohol extraction procedure (patent PCT / AU980030).
No. 5 (disclosed in No. 5) and the extract was formed into tablets using standard methods. More specifically, the red clover used must contain formononetin, biochanin, daidzein and genistein.

In summary, red clover leaves are harvested and soaked in water to induce enzymatic degradation that degrades isoflavones from their glycosides to aglycone form. 2 at room temperature
After standing for a period of time, the plant material is flash frozen in liquid nitrogen. Raw materials can be stored in this form for several years. For extraction, crush the frozen raw material into fine powder, thaw and put it in a fine gauze bag,
Immerse in 60% ethanol aqueous solution. The extraction is performed at 60 ° C. for 24 hours. The supernatant is separated from the insoluble plant material and the solvent is distilled off. The aqueous layer containing the isoflavones is extracted again with an organic solvent (either petroleum ether, hexane or ethyl acetate) to remove oils and other polar compounds. The solvent is distilled off and the aqueous layer is made almost dry on a rotary evaporator. This gives a concentrated extract containing about 25% isoflavones on a dry weight basis.

In this process, isoflavones are essentially extracted on a non-selective basis and 4
The original ratio of the two isoflavones is retained in the final extract. In this example, red clover species containing the four isoflavones in an appropriate ratio of about 45% biocanin, 40% formononetin, 8% daidzein and 7% genistein (detectable by thin layer chromatography) were selected. Is done. The dried isoflavone extract is mixed with a standard excipient such as methylcellulose, and 40 mg isoflavone consisting of 160 mg of clover extract, more specifically, 18 mg of biochanin, 16 mg of formonenetin, 3 mg of daidzein and 3 mg of genistein. A tablet containing the compound is formed.

Example 2 Tablets are made by the method described in detail in Example 1, in which case a species of red clover with a high formonenetin content is selected. The selected species has a proportion of 82% formononetin, 12% biocanin, 3% daidzein and 3% genistein. After the solvent extraction detailed in Example 1, the dried isoflavone extract has about the same isoflavone ratio as the starting plant material. A 200 mg tablet is formulated with 100 mg dry plant extract containing 25 mg isoflavones, consisting of about 25 mg formonenetin, 3 mg biocanin, 1 mg daidzein and 1 mg genistein.

Example 3 Thirty-six postmenopausal women with normal cholesterol levels were recruited for a double-blind clinical trial and consisted of three treatment groups-(a) placebo tablets, (b) one daily isoflavone tablet, or (c) Randomly assigned to one of three isoflavone tablets. The tablet prepared in Example 1 was used. Treatment continued for three months. Blood was collected at the beginning and end of the study and analyzed for total cholesterol, LDL and HDL levels, and clotting factors. There were no significant changes in these parameters in any of the treatment groups throughout the study.

Example 4 Fifty postmenopausal women with normal cholesterol were recruited to a single-blind clinical trial and randomly assigned to three treatment groups. All three groups were taken daily for one month using placebo tablets. Then, a formofonetin-rich isoflavone supplement was taken in the form of the tablets prepared in Example 2. Three doses of 25 mg, 50 mg, or 75 mg isoflavones per day were used. The main results tracked are total cholesterol, HDL and non-HDL (mainly LDL) levels and bone density of the proximal forearm (mainly cortical) and distal forearm (mainly cancellous). The results are summarized below as the change (%) from the basal value after 6 months of treatment.

[0065]

[Table 1]

All three dose groups at this particular ratio of isoflavones showed various statistically significant and clinically significant changes. Total cholesterol levels were low in all three groups (7
%), Which was due to a dramatic increase in HDL levels. Although LDL levels did not change significantly, HDL levels increased by an average of 28% in the 50 mg isoflavone group, which is completely unexpected from the results that other studies of isoflavones lacked observation of effects on HDL. there were. A significant reduction in blood levels of apoprotein B is achieved,
This is also quite unexpected and shows significant clinical benefit to women in terms of cardiovascular disease risk factors.

In the treatment group, especially in the 50 mg treatment group, a very marked positive effect on cortical bone density (proximal brachial arm) (4.1% increase) was seen in the first 6 months. Trabecular bone (distal forearm)
Had no effect, indicating that this particular isoflavone ratio has a very specific effect on cortical bone. Also, this is a completely unexpected result, as no product that has no effect on cancellous bone and specifically increases cortical bone has not been described before.

Example 5 On the basis of the results of Example 4, 50 mg tablets prepared by the method according to Example 2 were treated, especially in the refractile treatment group, ie women with high LDL and low HDL, according to the invention. We tested to see if there was anything useful. It is believed that this particular group of subjects metabolizes cholesterol in a different manner than normal cholesterol subjects.

The study design consisted of 5 weeks with active composition or placebo, after which the treatment was switched (
The active group will be placebo and the placebo group will be active group) for another 5 weeks. All subjects are then given an additional 12 weeks of the active composition. In the results shown in the table below,
"Short-term activity" is data for women treated with an active composition for 5 weeks and "long-term activity" is 17
Data for women taking weekly (ie 5 weeks plus 12 weeks) active composition.

[0070]

[Table 2]

The study surprisingly showed that at the start of the study, 10 women had high HDL and low LDL.
% Showed significant LDL reduction. This finding is comparable or slightly better than the best drugs actually used, such as statins. There is no literature reference using isoflavones in women with this type of lipoprotein profile. Thus, the compounds of the present invention provide a treatment for this particularly refractile group of patients.

Example 6 25 mg, 50 mg and 75 mg tablets were molded and prepared so that the total tablet weight was 550 mg using the following excipients. The isoflavones were mixed with an acacia gum carrier and added to a tableting formulation containing mixed tocopherol, microcrystalline cellulose, calcium hydrogen phosphate, soy polysaccharide, magnesium stearate, and anhydrous silica colloid.

Throughout this specification and the claims, unless stated otherwise by context, “comprise”
Or variations such as "comprises" and "comprising" are meant to include the described complete step or steps, steps of complete steps or groups of complete steps, or other steps, but other complete forms It does not exclude bodies, steps, complete bodies, groups of steps, and the like.

Patents: PCT Patent No. PCT / AU98 / 00305 Kelly G. et al .: Preparation of isoflavones from legumes US Pat. No. 5,585,892, Potter SM, Henley EC, Waggle DH: Risk of atherosclerosis and vascular disease Method for lowering blood LDL-cholesterol level and increasing HDL-cholesterol level for reduction "US Pat. No. 5,506,621, Barnes S. and Blair HC: Genistein for inhibiting osteoclasts US Pat. No. 5,424,331, 6/1995 Shylankevich M., 514-456 Pharmacological composition and soy food product for prevention of osteoporosis

References Pharamceutical Research 3 (6): 318 (1986) Anderson, JJ, Ambrose, WW, Garaner, SC, (1998), "Biphasic Effect of Genistein on Bone Tissue in an Ovariectomized Lactating Rat Model" Proc .Soc. Exp
Biol. Med., March 217 (3) 345-350 Fanti, P., Monier-Faugere MC, Geng, Z., Schmidt, J., PE, Cohen, D. And Ma
lluche, HH, (1998), "Phytoestrogens genistein reduces bone mass in short-term ovariectomy." Osteoporosis Int., 8 (3): 274-81. Goodman Gilman A. (eds.) The Pharmacology of Goodman &Gilman's Therapy. Foundation, 7th Edition, McGraw Hill Publisher, 1985 Gordon DJ, Probsfield JL, Garrison RJ, Neaton JD, Castelli WP
, Knoke JD, Jacobs DRJr., Bangdiwala S., Tyroler HA, (1989) "High density lipoprotein-cholesterol and cardiovascular diseases. Four prospective American studies," Circulation 79: 8-15 Hodgson et al. 1998) "Isoflavonoid phytoestrogen replacement therapy did not alter serum lipid levels: a randomized controlled trial in humans." Journal of Nutrition
128: 728-332 Nestel et al., (1997) "Soy isoflavones improve systemic arterial compliance but not lipids in plasma in menopausal and perimenopausal women."
sis, Thrombosis and Vascular Biology 17: 3392-3398 Nestel et al., (1999) "Isoflavones from Red clover improve the systemic arterial compliance of menopausal women but not the lipids in serum."
Clinical Endocrinology and Metabolism 84: 895-898 Potter S., Baum, J. et al. (1998) "Soy Proteins and Isoflavones: Effects on Blood Lipids and Bone Density in Postmenopausal Women" American Journal of Clinical Nutr
ition 1998: 68 (supplement) 1375S-1379S Gennaro A.R. (ed.) Remington's Pharmaceutical Science, Mack Publishing Company, Easto
n PA, 17th edition, 1985 Samman S, Lyons Wall PM, Chan GSM, Smith SJ, Petocz. P, (1
999) "Effects of isoflavone supplementation on plasma lipids and susceptibility to oxidation of low-density lipoproteins in premenopausal women" Atherosclerosis 147,277-283

──────────────────────────────────────────────────続 き Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI Theme coat ゛ (Reference) A61P 9/10 A61P 9/10 101 101 19/08 19/08 19/10 19/10 (81) Designated country EP (AT, BE, CH, CY, DE, DK, ES, FI, FR, GB, GR, IE, IT, LU, MC, NL, PT, SE), OA (BF, BJ, CF, CG, CI, CM, GA, GN, GW, ML, MR, NE, SN, TD, TG), AP (GH, GM, KE, LS, MW, SD, SL, SZ, TZ, UG, ZW), EA (AM, AZ, BY, KG, KZ, MD, RU, TJ, TM), AE, AG, AL, AM, AT, AU, AZ, BA, BB, BG, BR, BY, CA, CH, CN , CR, CU, CZ, DE, DK, DM, DZ, EE, ES, FI, GB, GD, GE, GH, GM, HR, HU, ID, IL, IN, IS, JP, KE, KG, K P, KR, KZ, LC, LK, LR, LS, LT, LU, LV, MA, MD, MG, MK, MN, MW, MX, NO, NZ, PL, PT, RO, RU, SD, SE, SG, SI, SK, SL, TJ, TM, TR, TT, TZ, UA, UG, US, UZ, VN, YU, ZA, ZW (72) Inventor Husband Alan James Australia, New South Wales , 2063, Northbridge, Maleina Road 13F term (reference) 4B018 LB06 LB07 LB08 MD07 MD15 ME04 ME05 4C086 AA01 AA02 BA08 MA02 MA03 MA04 MA31 MA35 MA37 MA41 NA14 ZA36 ZA45 ZA96 ZA97 ZC33 ZC51 ZC52 CA08 BA08 MA07 MA08 MA 31 MA35 MA37 MA41 NA14 ZA36 ZA45 ZA96 ZA97 ZC33 ZC51 ZC52 ZC75

Claims (43)

[Claims] (1) Formononetin and one or more isoflavones selected from biocanin, genistein and daidzein, wherein formononetin and the isoflavone are in a ratio of 15: 1 to 2: 1 with a therapeutic effect, optionally 1: 1. A composition comprising the above carrier, excipient, adjuvant and / or diluent. 2. The ratio of formononetin to the isoflavones is from 10: 1 to 2: 1.
The composition according to claim 1, which is: 3. The composition according to claim 1, comprising formononetin and biochanin. 4. The composition according to claim 1, which is in the form of a solid dosage unit. 5. The composition according to claim 4, which is in the form of a tablet, capsule, granule, buccal or suppository. 6. The composition according to claim 1, wherein said formononetin and biochanin comprise an extract of chickpea or clover. 7. The method according to claim 6, wherein the clover is selected from red clover (T. pratense), subterranean clover (T. subterranean), or white clover (white clover: T. repens). Composition. 8. The composition according to claim 7, wherein said clover is clover. 9. A method for treating and / or preventing a circulatory disease, or improving blood lipoprotein levels, or reducing the risk of ischemic heart disease, or reducing the risk of arteriosclerosis, treating or preventing osteoporosis, A composition according to claim 1 for improving or maintaining bone density, such as for preventing and / or treating fractures. 10. The composition according to claim 1, which additionally contains one or more vitamins. 11. The method according to claim 11, wherein the vitamins are vitamin C, vitamin D, vitamin E, vitamin
The composition according to claim 10, wherein the composition is selected from one or more of K, vitamin A and vitamin B. 12. The composition according to claim 1, further comprising a calcium-containing compound (0.5 g to 2 g). 13. The composition according to claim 1, which is combined with a beverage. 14. The composition according to claim 13, wherein the beverage is selected from nutritional drinks, sports drinks, juices, milk and milk substitutes. 15. The composition according to claim 1, which is combined with a food. 16. The composition according to claim 15, wherein the food is yogurt, food bar, spread, hard cheese, soft cheese or cottage cheese. 17. The composition according to claim 1, which is in the form of a pharmaceutical composition. 18. The composition according to claim 17, wherein the composition is a pill, tablet, capsule, suppository, dragee or sublingual formulation. 19. A method for treating and / or preventing a circulatory disease, or improving the level of lipoprotein in blood, or reducing the tendency of humans to form trombogenic,
Or the manufacture of a medicament for improving or maintaining bone density, such as reducing the risk of ischemic heart disease, reducing the risk of arteriosclerosis, treating or preventing osteoporosis, and / or preventing and / or treating fractures. In addition, formononetin and one or more isoflavones selected from biochanin, genistein and daidzein from 15: 1 to 2
How to use in the ratio of 1: 1. 20. Formononetin and biochanin in a ratio of 15: 1 to 2: 1, mixed with one or more carriers, excipients, adjuvants or diluents to make a composition for human administration. 14. The use according to claim 13, comprising a step. 21. Formononetin and one or more isoflavones selected from biochanin, genistein and daidzein in a therapeutically effective ratio of 15: 1 to 2: 1, optionally one or more carriers and excipients. And / or prophylaxis of circulatory system diseases, or improvement of blood lipoprotein level, or ischemic heart disease, which comprises administering a composition containing the composition of the present invention to a human, together with an auxiliary, and / or a diluent. Risk reduction,
Or a method for improving or maintaining bone density, such as reducing the risk of arteriosclerosis, treating or preventing osteoporosis and / or preventing and / or treating fractures. 22. The method according to claim 21, which is a method for improving blood lipid protein levels. 23. The method according to claim 22, wherein the concentration of high-density lipoprotein in the blood of the individual is increased. 24. The method according to claim 23, wherein the concentration of low-density lipoprotein in the blood of the individual is reduced. 25. The method of claim 21, wherein the risk of vascular disease is reduced. 26. The method of claim 21, wherein the risk of ischemic heart disease is reduced. 27. The method of claim 21, wherein the risk of arteriosclerosis is reduced. 28. The method according to claim 21, for treating or preventing osteoporosis. 29. The ratio of formononetin to said isoflavones is from 10: 1 to 2:
22. The method according to claim 21, which is 1. 30. The method of claim 21, wherein said composition is a solid dosage form. 31. The method of claim 21, wherein said dosage form is a tablet, capsule, granule, buccal, or suppository. 32. The method of claim 21, wherein said formonenetin and biochanin comprise an extract of chickpea or clover. 33. The method of claim 32, wherein said clover is selected from red clover (T. pratense), subterranean clover (T. subterranean), or white clover (T. repens). 34. The method of claim 33, wherein said clover is a red clover. 35. The human subject is a pre-, post- or post-menopausal woman.
The described method. 36. The method of claim 35, wherein said female has normal cholesterol or hypercholesterolemia. 37. The method of claim 36, wherein said female has atherosclerosis. 38. The method of claim 21, wherein said human subject is a male with normal cholesterol or hypercholesterolemia. 39. The method of claim 38, wherein said male has hypercholesterolemia. 40. The method of claim 39, wherein said male has normal cholesterol. 41. The method according to claim 21, which is a method for preventing or treating a fracture. 42. The method of claim 41, wherein said method is a method of preventing or treating a fracture of a bone mainly enriched in cortical bone tissue. 43. The method of claim 42, wherein said bone is selected from a femoral neck, femur, humerus, radius, ulna or tibia.