TWI448250B - Muscadine composition with improved anti-oxidant activity - Google Patents

Description

Muscat composition with improved antioxidant activity

Cross-reference related application

The present application claims the benefit and priority of U.S. Provisional Application No. 61/085,369, filed on Jul. 31, 2008, and U.S. Provisional Application No. 61/085,371, filed on Jul. 31, 2008. Each is incorporated herein by reference in its entirety.

The present disclosure relates to a muscat grape composition comprising a muscadine pulp extract and resveratrol, which is used as an antioxidant, such as as an anti-aging supplement.

Reactive oxygen species (ROS) are obligate by-products of aerobic metabolism and thus continue to be produced in humans and other organisms. Humans also come into contact with ROS from sources/environmental sources such as pollution, sunlight and diet. Although there are different chemical forms of ROS, they all have harmful effects on the structure and function of cell components and macromolecules. The intensity of ROS production/exposure is called oxidative stress.

Oxidative stress and pathogenesis of chronic inflammatory diseases such as diabetes, cancer, atherosclerosis and other cardiovascular diseases, and degenerative effects such as Alzheimer's disease and Parkinson's disease Related to the disease. In addition, multiple lines of evidence support the view that oxidative stress is the primary mechanism of normal aging. Therefore, there is a need to develop a treatment for inhibiting oxidative stress.

Minimizing oxidative stress is considered a key to health and longevity. Epidemiological data show that diets rich in antioxidant nutrients confer significant benefits. In addition to the typical antioxidant vitamins (vitamins C and E), dietary plants also contain many polyphenols that appear to reduce oxidative stress in the short and long term after consumption. Several in-vitro assays have been developed to measure the antioxidant capacity of different foods and there is a strong correlation between polyphenol content and antioxidant capacity.

Although plants including grapes provide a promising source of antioxidants, there are significant barriers to the widespread use of phytochemicals in the human population. For example, the bioavailability of an active agent in a food product is generally sub-optimal for use as an antioxidant. The active agent may also be concentrated in certain parts of the plant that are neglected or discarded in the typical preparation of the food for consumption. Different types of related foods may also have biochemical differences that make them less than optimal for therapeutic use.

The present inventors have determined the antioxidant capacity of muscadine pulp extract and Japanese knotweed extract alone or in combination (in the form of a mixture or dietary supplement), such as by oxygen radical absorption capacity (Oxygen Radical Absorbance Capacity). , ORAC) verification measured. At the same time, the hydrophilic antioxidant capacity and lipophilic antioxidant capacity of the sample were measured. These studies have shown that muscadine pulp extracts and Japanese root extracts in the form of a mixture or dietary supplement have a strong synergistic effect in producing lipophilic antioxidant capacity. It was not expected to exhibit selective synergy under lipophilic conditions. These findings indicate that the Japanese cockroach extract (at least as a source of substantially pure resveratrol) and the muscadine pulp extract mixture and dietary supplement have improved antioxidant capacity compared to either individual extract. .

Compositions having improved antioxidant activity are disclosed herein. In some disclosed embodiments, the composition comprises a muscadine ( Vitis rotundifolia ) pulp extract having a polyphenol content of at least 2% and a source derived from a non-musk grape (such as Japanese radix root extract) a minimum purity of at least 5% of resveratrol, wherein the ratio of muscadine polyphenol to trans-resveratrol is in the range of 0.1/1 to 10/1 (weight/weight), thereby providing resistance A composition of oxidant activity. In a particular embodiment, the composition comprises a muscadine (musk) pulp extract having a polyphenol content of at least 2% and a minimum purity of at least 5 from a non-musk source (such as Japanese jaundice extract). % resveratrol, wherein the ratio of muscadine pulp extract to resveratrol is from 0.2/1 to 50/1 (weight/weight), such as from 5/1 to 50/1 (weight/weight) or 20 Within the range of /1 to 50/1 (weight/weight), a composition having antioxidant activity is thus provided. In some embodiments, the composition has a total ORAC of at least 24 micromoles of Trolox equivalent per microgram of polyphenol (micromolar TE per milligram of polyphenol).

In some embodiments, the muscadine (musk) pulp extract has a total polyphenol content of about 4% and the composition has about 18/1 (weight/weight) of muscadine pulp extract and trans white The ratio of resveratrol. In one embodiment, the composition has a ratio of muscadine polyphenol to resveratrol of about 0.75/1 (weight/weight). In certain embodiments, the muscadine pulp extract may comprise from 20% to 50% solids in the liquid. For example, the muscadine pulp extract contains about 40% solids in the liquid. In some embodiments, the resveratrol comprises at least 98% trans-resveratrol extracted from the roots of Japanese cockroach ( Polygonum cuspidatum ).

In some embodiments, the disclosed compositions can further include elderberry extract, purple carrot extract, excipients (such as glycerin, sorbitol, colloidal ceria or natural flavoring additives), or combinations thereof. In a particular embodiment, the composition comprises from 23% to 32% of the total composition of the muscadine pulp extract, 1.2% to 3.0% of the total composition of resveratrol, 0.12 of the total composition. % to 0.20% of purple carrot extract, 0.4% to 0.6% of total composition of elderberry extract, 50% to 61% of total composition of sorbitol, 8% to 12% of total composition % 99.7% glycerol composition and 1% to 2% of the total composition of colloidal cerium oxide.

In a more specific embodiment, the composition comprises about 28.9% of the total composition of the muscadine pulp extract (containing about 4.1% polyphenol), about 1.65% of the total composition of resveratrol, About 0.16% of the total composition of the purple carrot extract, about 0.48% of the total composition of the elderberry extract, about 54.8% of the total composition of sorbitol, about 10% of the total composition of 99.7 % glycerin composition, about 2% of the total composition of colloidal cerium oxide, about 1.27% of Concord grape extract, about 0.16% of Cabernet grape extract And about 0.48% of red grape powder.

In another specific embodiment, the composition comprises about 23.77% of the total composition of the muscadine pulp extract (containing about 5% polyphenol), about 1.65% of the total composition of resveratrol, About 0.16% of the total composition of the purple carrot extract, about 0.48% of the total composition of the elderberry extract, about 60% of the total composition of sorbitol, about 10% of the total composition of 99.7 % glycerin composition, about 2% of the total composition of colloidal cerium oxide, about 1.27% of Concord grape extract, about 0.16% of Cabernet grape extract and about 0.48% red Grape powder.

Any of the disclosed compositions can be provided in a non-beverage food, beverage or dietary supplement in liquid or solid form. In addition, any of the disclosed compositions can be used as an antioxidant, such as an anti-aging supplement, including a skin improving agent. For example, a method of preventing or inhibiting cellular senescence is disclosed comprising administering to a subject in need thereof a disclosed composition sufficient to prevent or inhibit one or more dose-related processes associated with cellular senescence, such as preventing or Free radical formation or activity in the individual ingesting the composition is inhibited.

Also provided herein are methods of preparing the disclosed antioxidant compositions. Such methods may include muscadine (musk) pulp extract having a polyphenol content of at least 2% and resveratrol having a minimum purity of at least 5% trans-resveratrol from a non-musk source. a combination wherein the ratio of muscadine polyphenol to trans-resveratrol is in the range of 0.1/1 to 10/1 (weight/weight), thereby producing an antioxidant active muscadine pulp extract and trans A mixture of resveratrol. In some embodiments, the methods can include the minimum purity of the muscadine (lof grape) pulp extract having a polyphenol content of at least 2% and the non-musk source (such as Japanese jaundice extract). a resveratrol combination of at least 5% trans-resveratrol, wherein the ratio of muscadine pulp extract to resveratrol is from 0.2/1 to 50/1 (weight/weight), such as from 5/1 to 50 Within the range of /1 (weight/weight) or 20/1 to 50/1 (weight/weight), including 18/1 (weight/weight), thereby providing a composition having antioxidant activity.

The disclosed method can further comprise preparing the muscadine pulp extract, followed by combining the muscadine pulp extract with trans-resveratrol. In a particular embodiment, preparing the muscadine pulp extract comprises combining the bronze muscadine pulp extract with the purple muscadine pulp extract. In other embodiments, preparing the muscadine pulp extract comprises preparing a extract from a mixture of bronze muscadine pulp and purple muscadine pulp.

In a particular embodiment, the method comprises preparing a muscadine pulp extract by combining a bronze muscadine pulp extract with a purple muscadine pulp extract, wherein the bronze muscadine pulp extract is purple The ratio of the muscadine pulp extract is in the range of 0.1 to 10 (weight/weight), such as 0.3 to 3 (weight/weight). In a more specific embodiment, the ratio of bronze muscadine pulp extract to purple muscadine pulp extract is about 2.25/1 (weight/weight).

The above and other features of the present disclosure will become more apparent from the following embodiments.

I. Abbreviations and terms:

(a) abbreviation

FRAP: Iron reduction ability of pulp

Mg: gram

Ml: liter

ORAC: free radical absorption capacity

ROS: Oxygen substances

TE: rolox equivalent

TEAC: rolox equivalent antioxidant capacity

Wt: amount

(b) terminology

The following explanation of terms and methods are provided to better describe the present disclosure and to guide the general practitioner in the practice of the present disclosure. As used herein and in the appended claims, the singular "a" or "the" The term "or" means a single element or a combination of two or more elements of the two alternative elements, unless the context clearly indicates otherwise. As used herein, "comprises" means "includes". Therefore, "comprising A or B" means "including A, B or A and B" without excluding other elements.

All technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art of the present disclosure. Although methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present disclosure, suitable methods and materials are described below. It should also be understood that any numerical value is an approximation, whether or not the word "about/approximately" or the like is stated. All percentages and ratios are by weight unless otherwise indicated.

Administration: An agent such as a composition comprising muscadine pulp extract and trans-resveratrol is provided or administered to an individual by any effective means. Exemplary routes of administration include, but are not limited to, oral, injection (such as subcutaneous, intramuscular, intradermal, intraperitoneal, intravenous, and intratumoral), sublingual, transdermal, intranasal, topical, and inhalation routes.

Antioxidant activity: includes the activity of scavenging and neutralizing oxidative free radicals. Antioxidant activity can be measured using the methods disclosed herein and methods known in the art, including oxygen free radical absorption capacity (ORAC) assay, and ferric reducing ability of plasma (FRAP). Verification and Trolox Equivalent Antioxidant Capacity (TEAC) assay. In one embodiment, if the composition has a total ORAC of at least 24 micromoles of Trolox equivalent per microgram of polyphenol (micromolar TE per milligram of polyphenol), it has antioxidant activity and can be used as an antioxidant.

Elderberry ( Sambucus nigra ): A plant belonging to the family Adoxaceae found in Europe and North America, having several regional varieties or subspecies. The flower is a flat-headed umbrella inflorescence. Berries are black to blue-green and contain anthocyanins and other polyphenolic compounds, of which the amount and type of anthocyanins and other polyphenolic compounds vary depending on the species.

"Elderberry extract" may be a substance obtained by extracting elderberry according to any extraction method known to those skilled in the art, as long as it has a desired activity (for example, color stabilizing activity, antioxidant activity, or The combination can be. For example, the elderberry extract may include juice obtained by pressing the elderberry fruit, or extracting the whole fruit or fruit of the elderberry by an optional extraction method and the like which are well known to those skilled in the art. Or an extract obtained from a suitable part of the seed. Similarly, crushed products of elderberry fruit or dried elderberry fruit concentrate can be used as "Sambucus extract".

Excipient : An inactive substance used as a carrier for the active ingredient of a composition. Excipients can include materials for increasing the volume of a formulation having a very effective active ingredient, allowing for conventional and precise dosages, stabilizing the active ingredients, and rendering the delivery system visually and/or organoleptically acceptable. Examples of pharmaceutical excipients include starch, glucose, lactose, sucrose, gelatin, malt, rice, flour, white peony, silicone, sodium stearate, glyceryl monostearate, talc, sodium chloride, skimmed milk powder, glycerin , propylene, ethylene glycol, water, ethanol and the like. In a particular embodiment, the disclosed anti-aging supplements include the following excipients: glycerin, sorbitol, colloidal ceria, and natural flavoring additives.

Inhibiting (including preventing) cellular aging : inhibiting (eg, preventing) one or more processes associated with cellular senescence, such as inhibiting free radical formation or activity in an individual ingesting a composition. Prevention of cellular aging refers to an intervention that improves the signs or symptoms of cellular aging. Prevention includes prevention to delay the onset of one or more processes associated with cellular aging. Blocking or inhibiting cellular senescence does not require complete absence of cellular senescence. In a particular embodiment, the disclosed compositions reduce or delay the process associated with cellular aging by at least 10%, at least 20%, at least 50%, or even at least 90%. Such reductions can be measured using the methods disclosed herein as well as methods known in the art.

Muscadine Grape (Clover) : native to southeastern United States, and from Delaware to the Gulf of Mexico and Missouri, Kansas Grapes found in the wilderness of the state (Kansas), Oklahoma, and Texas. Muscat is well adapted to the warm, humid conditions of the southeastern United States. The fruit grows into small loose ears of 3-40 grapes, which are completely different from the close string characteristics of European and American grapes. Round, 1 to 1-1/2 吋 fruit has a thick crust and contains at most 5 hard oval seeds. In terms of color, the fruit is in the range of bronze green to bronze, magenta, purple and near black.

Many different varieties of muscat grape are available, including female (only pistil) varieties, such as Black Beauty, Black Fry, Darlene, Fry, Higgins. Jumbo, Scuppernong, Sugargate, Summit, Supreme and Sweet Jenny; and self-cultivating species such as Carlos, Cowart, Dixieland, Dixie Red, Fry Seedless, Magnolia, Nesbitt, Noble, Redgate ), Regale and Sterling.

For example, the bronze colored varieties of muscat are identified by those skilled in the art as including Carlos, Chowan, Doreen, Higgins, Magnolia, Nevermiss, Pamlico, Roanoke, Scarperon, Sterling and peak cultivars. Purple to black colored varieties include Albermarle, Bountiful, Cowart, GA-1, Hunt, NC-1, Noble, Regal, North Carolina (Tarheel) and treasures.

The phytochemical component of the complete muscat grape is different from the Vitis vinifera . Musk grape has a higher total phenolic compound content characterized by high concentrations of citric acid glycosides, gallic acid glycosides and flavonoid glycosides. The presence of turkey ellagic acid in muscadine is unique and Turkish citric acid is present in the form of free Turkish citric acid, Turkish citrate glycosides, methoxylated derivatives and ellagitannin. Another unique feature is the anthocyanin chemistry observed in muscadine. 3,5-diglucoside (with 3-in a non-deuterated form of delphinidin, cyanidin, petunidin, peonidin, and malvidin) The presence of such compounds in the form of glycosides and the natural color of other anthocyanins present in the grapes render the juice and pulp dark purple. The purple pulp extract contains anthocyanins, while the bronze pulp extract does not contain anthocyanins.

Red and purple anthocyanins are polyphenolic compounds having antioxidant properties. Flavonol is the second most abundant flavonoid in intact muscat grape. The main phenolic compounds reported in the muscadine section (in descending order) are turmeric, myricetin, quercetin and kaempferol, as reported by the seed. The main phenolic compounds are epicatechin, catechin and gallic acid (Patrana-Bonilla et al . J. Agric. Food Chem . 51: 5497-5503, 2003).

Pharmaceutically Acceptable Vehicles : Pharmaceutically acceptable vehicles (carriers) suitable for use in the present disclosure are conventional. Remington's Pharmaceutical Sciences , EW Martin, Mack Publishing Company, Easton, PA, 19th edition (1995) describes compositions and formulations suitable for the delivery of one or more compositions (such as one or more muscadine compositions) and other pharmaceutical agents for pharmaceutical delivery. Things.

In general, the nature of the vehicle will depend on the particular mode of administration employed. For example, parenteral formulations typically comprise an injectable fluid as a vehicle, including pharmaceutically and physiologically acceptable fluids such as water, physiological saline, balanced salt solutions, aqueous dextrose, glycerol or Its analogues. For solid compositions (eg, powders, pills, troches, or capsules), conventional non-toxic solid vehicles may include, for example, pharmaceutical grade mannitol, lactose, starch, or magnesium stearate. In addition to the biologically neutral vehicle, the pharmaceutical composition to be administered may also contain small amounts of non-toxic auxiliary substances such as wetting or emulsifying agents, preservatives and pH buffers and the like, such as sodium acetate or sorbitan. Monolaurate.

Purified : The term purified does not require absolute purity; rather, it is a relative term. Thus, for example, a purified material is one that is more enriched with the substance in its natural environment (eg, in a fruit (eg, grape)). In one embodiment, the formulation is purified such that the material comprises at least about 5% of the total formulation (such as, but not limited to, at least 10%, 20%, 30%, 40%, 50%, 70%, 80%, 90%, 95%, 98% or 99%). In one embodiment, the disclosed composition having antioxidant activity comprises a minimum purity of at least 50%, 70%, 80%, 90%, 95%, 98% or 99% of the total resveratrol formulation (weight Trans-resveratrol.

Pomace: The skin, seeds and pulp remaining after juice extraction. In one embodiment, the pulp extract is a bronze muscadine pulp extract, a purple muscadine pulp extract, or a combination thereof. Many different varieties of muscadine pulp can be used as a starting material, and include female (only pistil) varieties, such as black beauty, black babies, Darin, small fish, Higgins, treasure, Scarperon, Sugar Gate, Peak, Crown and Sweet Jenny, and self-cultivating species such as Carlos, Cowart, Dixieland, Starred Red, Small Fish Seedless, Magnolia, Nesbitt, Noble, Red Gate, Regal and Sidley.

Muscat grape pulp contains phenolic compounds with antioxidant properties, including gallic acid and turmeric tannin.

Purple carrot ( Daucus carota ): A cultivar of carrots containing anthocyanin pigments. "Purple carrot extract" may be a substance obtained by extracting purple carrots according to any extraction method known to those skilled in the art as long as it has a desired activity (for example, color stabilizing activity, antioxidants) The activity or a combination thereof can be used. In one embodiment, the purple carrot extract has the property of stabilizing the muscat color pigment and thus can be subsequently used as a color stabilizing additive.

Resveratrol: A phytoalexin which is a stilbenoid (a derivative of stilbene) and is produced in plants by means of an enzyme stilbene synthase. Resveratrol exists in two structural isoforms: cis resveratrol and trans resveratrol. Trans-resveratrol can undergo isomerization to cis when heated or exposed to ultraviolet radiation.

Resveratrol is present in widely varying amounts in berries, raspberries, mulberries, plums, peanuts, berries of the Vaccinium species (including blueberries, bilberry and cranberries) and some pines ( Such as the European Scots pine and the eastern white pine, and the roots and stems of Polygonum cuspidatum and Japanese cockroach. In grapes, resveratrol is mainly found in the skin and seeds. The amount of resveratrol present in the grape skin varies with the grape cultivar, its geographic origin, and exposure to fungal infections.

As used herein, the term resveratrol may include natural trans-resveratrol or synthetic trans-resveratrol extracted from a plant such as grapes. As used herein, the term resveratrol can include a modified formulation of trans-resveratrol, such as a microencapsulated or water-dispersible form.

Subject: A living multi-cellular vertebrate organism, including the categories of human and veterinary individuals.

Therapeically Effective Amount: The amount of a composition that, alone or in combination with another (other) agent (eg, other antioxidants), induces a desired response (eg, prevents or inhibits cellular senescence). The formulations disclosed herein can be administered in a therapeutically effective amount.

In one embodiment, the desired response is to inhibit or reduce free radical production or activity associated with one or more cellular senescence processes in the subject to which the composition is administered. Regarding the effectiveness of the composition, free radical production or activity does not need to be completely eliminated. For example, the composition can reduce the production or activity by a desired amount, such as at least 10%, at least 20%, at least 50%, at least 60%, compared to free radical production or activity in the absence of the composition. At least 70%, at least 80%, at least 90%, at least 95% or at least 98%.

During treatment, a therapeutically effective amount of the disclosed muscadine pulp extract composition can be administered in a single dose or in several doses, such as daily administration. However, the therapeutically effective amount will depend on the individual being treated, the severity and type of condition being treated, and the mode of administration. For example, if administered orally, a therapeutically effective amount of such agents can vary from about 1 mg to about 500 mg per day of trans-resveratrol, muscadine polyphenols, or combinations thereof.

Unit dose: A physical discrete unit containing a predetermined amount of active substance calculated to produce a desired effect, such as a therapeutic effect, individually or collectively. A single unit dose or a plurality of unit doses can be used to provide the desired effect or activity, such as antioxidant activity. In one embodiment, the unit dose includes a desired amount of the agent that prevents or inhibits one or more processes associated with cellular aging.

II. Description of several specific examples

Plant agents have been well studied as potential sources of nutrients that can be used to reduce oxidative stress. Red grapes are one of many foods that have been the subject of extensive research. The antioxidant action of red grapes has been widely attributed to polyphenolic compounds such as resveratrol and procyanidin, the latter of which appear in grapes in substantial quality.

The inventors conducted a series of studies to develop a muscadine pulp extract sufficient to increase the maximum solubility of citric acid in Turkey. It has been found that the ratio of unconcentrated 65%/35% bronze muscadine pulp extract to purple muscadine pulp extract (vol/vol) increases the amount of turmeric tannin between the two pulp extracts. Maximum solubility.

The determination of the maximum solubility of citric acid in Turkey (which is unique to muscat) in muscadine pulp extracts allows the preparation of extracts with increased solubility of citric acid in Turkey (and thus the polyphenol distribution of the pulp), which in turn It is allowed to prepare an extract with improved antioxidant activity. Accordingly, the present invention discloses muscadine pulp extracts having improved Turkish citric acid solubility and methods of using such extracts. For example, the disclosed muscadine extract can be administered alone or in combination with other compounds in the form of a non-beverage food, beverage, dietary supplement or topical ointment. A method of preparing a muscadine pulp extract is also possible, the method comprising combining a bronze muscadine pulp extract with a purple muscadine pulp extract to produce a muscadine pulp extract, wherein the bronze muscat fruit The ratio of the pulp extract to the purple muscadine pulp extract is in the range of 0.1 to 10 (weight/weight), such as 0.3 to 3 (weight/weight).

In addition, the inventors determined the antioxidant capacity of the disclosed muscadine pulp extract and Japanese cockroach extract (standardized to 98% trans-resveratrol) alone or in combination (in the form of a mixture or a dietary supplement), such as Measured by ORAC test. At the same time, the hydrophilic antioxidant capacity and lipophilic antioxidant capacity of the sample were measured. These studies have shown that muscadine grape fruit extract in the form of a mixture or dietary supplement and Japanese root extract (standardized to 98% trans-resveratrol) have a strong synergistic effect in producing lipophilic antioxidant capacity. It was not expected to exhibit selective synergy under lipophilic conditions. Similarly, although the muscadine extract used in the composition may be a natural extract, it may vary depending on the species, the extraction method, etc., as long as the ratio of the disclosed muscadine polyphenol to resveratrol is maintained, but synergistically Lipophilic antioxidant activity is retained.

These findings indicate that the mixture containing Japanese cockroach extract (standardized as 98% trans-resveratrol) and muscadine pulp extract and dietary supplements has improved progeny compared to either individual extract. Lipid antioxidant capacity. Importantly, the oxidative process occurring in a lipophilic environment is thought to cause and/or trigger the pathogenesis of a variety of disease conditions, such as low density lipoprotein (LDL) oxidation in atherosclerosis and type II Obesity-induced insulin resistance in diabetes. In addition, oxidation of dietary lipids in the gastrointestinal tract results in absorption of cytotoxicity such as malondialdehyde (MDA) and genotoxic lipid peroxidation products. Therefore, the disclosed composition having high lipophilic antioxidant ability can impart a beneficial effect in counteracting the harmful effects of the above lipid oxidation.

This finding made it possible to make muscadine pulp extract compositions with improved antioxidant activity and methods of preparing such compositions. For example, disclosed herein is a muscadine pulp extract composition having antioxidant activity that can be administered in the form of a non-beverage food, beverage, liquid or solid dietary supplement or topical ointment. The method of preparing the disclosed composition may comprise adding a muscadine (circle grape) pulp extract having a polyphenol content of at least 2% to a trans-white cane having a minimum purity of at least 5% from a non-musk source. An alcohol combination in which the ratio of muscadine polyphenol to trans-resveratrol is in the range of 0.1/1 to 10/1 (weight/weight), thereby producing an antioxidant activity of muscadine pulp extract and anti-oxidant a mixture of resveratrol. The method of preparing the disclosed compositions may also comprise extracting a muscadine (lof grape) pulp extract having a polyphenol content of at least 2% with a trans-white peony having a minimum purity of at least 5% from a non-musk source. a rosin combination wherein the ratio of the muscadine pulp extract to trans-resveratrol is from 0.2/1 to 50/1 (weight/weight), such as from 5/1 to 50/1 (weight/weight), including 20 Within the range of /1 to 50/1 (weight/weight), such as 18/1 (weight/weight), thereby producing a mixture of muscadine pulp extract and trans-resveratrol having antioxidant activity.

Lipophilic antioxidants have been found to be effective in preventing various types of skin damage such as, but not limited to, products that inhibit lipid peroxidation and are produced by lipid peroxidation, such as crosslinkers. Oxidative stress is known to be the primary mechanism responsible for normal aging. The compositions disclosed herein are shown to have improved lipophilic antioxidant activity. Based on these observations, methods for inhibiting cellular senescence, such as preventing or inhibiting free radical production or activity, are disclosed. Such methods include the use of the disclosed compositions having improved antioxidant capacity (e.g., dietary supplements) as an anti-aging supplement. For example, a method of preventing or reducing one or more processes associated with cellular aging, such as reducing or inhibiting free radical production or activity, in an individual ingesting a mixture or dietary supplement composition is made possible.

A. Muscat grape puree extract

This article discloses extracts of muscadine pulp derived from bronze muscadine pulp and purple musk grape pulp. In some embodiments, the ratio of the bronze muscadine pulp extract to the purple muscadine pulp extract in the muscadine pulp extract is from 0.1 to 10 (weight/weight), such as from 0.3 to 3 (weight/weight). Within the scope. For example, the ratio of bronze muscadine pulp extract to purple muscadine pulp extract is about 2.75 / about 1 (weight / weight), 2.5 / about 1 (weight / weight), about 2.25 / about 1 ( Weight/weight), about 2/about 1 (weight/weight), about 1.5/about 1 (weight/weight) or about 1/about 1 (weight/weight). As used herein, the term "about" is defined as ±0.5. In a particular embodiment, the ratio of bronze muscadine pulp extract to purple muscadine pulp extract is about 2.25 / about 1 (weight / weight).

In certain embodiments, the disclosed muscadine (loft grape) pulp extract has a polyphenol content of at least 2%. For example, the polyphenol content is at least 3%, at least 3.5%, at least 4%, at least 4.5%, at least 5%, at least 6%, at least 8%, at least 10%, at least 12%, or at least 14%. In a particular embodiment, the muscadine (circle grape) pulp extract has a polyphenol content of about 4%.

In some embodiments, the disclosed muscadine pulp extract comprises 20% to 50% solids in a liquid, such as at least 25%, at least 30%, at least 35%, at least 37%, at least 40%, at least 42 %, at least 44%, at least 46% or at least 48%. In a particular embodiment, the extract comprises about 40% solids in the liquid.

B. Method for preparing muscadine pulp extract

Also disclosed is a method of preparing a muscadine pulp extract, wherein the ratio of the bronze muscadine pulp extract to the purple muscadine pulp extract is from 0.1 to 10 (weight/weight), such as from 0.3 to 3 (weight/weight). Within the scope. In a disclosed embodiment, the muscadine pulp extract is prepared by combining a bronze muscadine pulp extract with a purple muscadine pulp extract. In a particular embodiment, the muscadine pulp extract is prepared by combining a bronze muscadine pulp extract with a purple muscadine pulp extract, wherein the bronze muscadine pulp extract and the purple muscat fruit The ratio of the pulp extract is in the range of 0.1 to 10 (weight/weight), such as 0.3 to 3 (weight/weight), including about 2.75/about 1 (weight/weight), 2.5/about 1 (weight/weight), About 2.25 / about 1 (weight / weight), about 2 / about 1 (weight / weight), about 1.5 / about 1 (weight / weight) or about 1 / about 1 (weight / weight). In an even more specific embodiment, the muscadine pulp extract is prepared by combining a bronze muscadine pulp extract with a purple muscadine pulp extract, wherein the bronze muscat grape extract and the purple muscat fruit The ratio of the pulp extract was about 2.25/1 (weight/weight).

In some embodiments, a method of preparing a muscadine pulp extract can include separately preparing a bronze muscadine pulp extract and a purple muscadine pulp extract. For example, bronze muscadine pulp and purple musk grape pulp are extracted with water (preferably hot water). The preparation of the bronze muscadine pulp extract and the purple muscadine pulp extract may further comprise fermenting the bronze muscadine pulp extract and the purple muscadine pulp extract to remove the extracted sugar. In one embodiment, after extracting the bronze muscadine pulp and the purple muscadine pulp, but combining the bronze muscadine pulp extract with the purple muscadine pulp extract to produce the disclosed muscadine Fermentation is carried out before the pulp extract. In other embodiments, after combining the bronze muscadine pulp extract with the purple muscadine pulp extract at a desired post-extraction ratio (such as a ratio of bronze to purple of about 2:25/1), Fermented.

Fermentation can be carried out by any method known to those skilled in the art, including those described herein. For example, fermentation may include adding yeast and yeast nutrients to the pulp and fermenting the extract until the residual sugar content is converted to ethanol. Typically, 2 pounds of yeast is used per 1000 gallons of 1 x (unconcentrated) extract. In this embodiment, the fermentation is typically completed after 3 days. In other embodiments, the amount of yeast and/or the duration and temperature of the strain and fermentation may vary according to individual methods known to those skilled in the art. In some embodiments, the enzyme may be used to clarify and/or precipitate the residue or to improve the extraction yield of the pulp extract. For example, such enzymes may include pectinase or an enzyme blend from Aspergillus niger and may be purchased from sources such as Scott Laboratories. These enzymes can be added to the pulp extract before or during the fermentation.

In some embodiments, the preparation of the bronze muscadine pulp extract and the purple muscadine pulp extract comprises filtering the bronze muscadine pulp extract and the purple muscadine pulp extract. For example, bronze muscadine pulp extract and purple musk grape pulp extract can be filtered before and/or after fermentation. Filtration can be performed according to general methods known to those skilled in the art, including the filtration methods disclosed herein. In a particular embodiment, the extract is filtered through a screen of suitable mesh size, such as a USP mesh (typically 120 mesh) or a similar cloth filter (such as a filter available from Millipore Corporation).

In other embodiments, preparing the muscadine pulp extract may comprise combining the bronze muscadine pulp and the purple muscadine pulp in the desired ratio and simultaneously extracting the bronze muscadine pulp and the purple muscadine pulp. For example, the bronze muscadine pulp and the purple muscadine pulp are simultaneously extracted with water (preferably hot water) to produce the disclosed muscadine pulp extract. In some embodiments, the method can further comprise fermenting the muscadine pulp extract to remove the extracted sugar. Any of the methods of fermentation known to those skilled in the art, including those described herein, can be used as previously described. For example, fermentation may include adding yeast and yeast nutrients to the pulp and fermenting the extract until the residual sugar content is converted to ethanol. In one embodiment, the fermentation comprises adding 2 pounds of yeast per 1000 gallons of 1X extract and maintaining the extract at room temperature for about 3 days or until the residual sugar content is converted to ethanol. The disclosed method can also include filtering the muscadine pulp extract. For example, the muscadine pulp extract can be filtered before and/or after fermentation. Filtration can be performed according to general methods known to those skilled in the art, including the filtration methods disclosed herein.

In some embodiments, the method of preparing the muscadine pulp extract further comprises concentrating the bronze muscadine pulp extract and the purple muscadine pulp extract such that each extract is included in the liquid from 20% to 50% A solid such as at least 25%, at least 30%, at least 35%, at least 37%, at least 40%, at least 42%, at least 44%, at least 46% or at least 48% solids. In a particular embodiment, the extract is concentrated such that each extract comprises about 40% solids in the liquid. The bronze and purple extracts can typically be concentrated using known methods of concentrating the sample, including the methods of concentrating the samples disclosed herein.

In a particular embodiment, to prepare a 40% solids muscadine pulp extract, the muscadine pulp extract is dried to a powder form and reconstituted with 40% solids water. Alternatively, a more acceptable commercial method is to concentrate by removing the extraction solvent by evaporation under vacuum. This can be achieved via a batch or continuous process. The batch process involves placing the extract in a container and drawing a vacuum of 20-29 ''Hg) while heating the cannula to provide energy to increase the vapor pressure of the solvent. The solvent vapor is allowed to condense outside the vessel and the rate of condensation controls the temperature of the condensate. The same principle applies to the continuous evaporation process, but has the advantage that the time during which the condensate is exposed to high temperatures is shortened. Both methods are suitable for concentrating the muscadine pulp extract described herein.

In a particular embodiment, the method of preparing the muscadine pulp extract comprises separately extracting the bronze and purple pulp, followed by filtering the extracts, followed by combining the bronze and purple pulp extracts in the desired ratio. In some embodiments, the method can further comprise fermenting the combined muscadine pulp extract to remove the extracted sugar. Any of the methods of fermentation known to those skilled in the art, including those described herein, can be used as previously described. In one embodiment, the method includes more than one filtration step, such as filtering the extract before and after fermentation. In some embodiments, the method can further comprise concentrating the extract as described herein. For example, the method can include concentrating the extract by removing the extraction solvent by evaporation under vacuum.

C. Compositions having antioxidant activity

Compositions having improved antioxidant activity are disclosed herein. In some disclosed embodiments, the compositions include muscadine (musk) pulp extract having a polyphenol content of at least 2% and a minimum of non-musk source (such as Japanese radix extract) Trans resveratrol having a purity of at least 5%, wherein the ratio of muscadine polyphenol to trans-resveratrol is in the range of 0.1/1 to 10/1 (weight/weight), thereby providing an antioxidant Active composition.

In some embodiments, the composition comprises at least 3%, at least 3.5%, at least 4%, at least 4.5%, at least 5%, at least 6%, at least 8%, at least 10%, at least 12%, or at least 14% The total polyphenol content of the muscadine (musk) pulp extract and the minimum purity from the non-musk source is at least 5%, at least 10%, at least 25%, at least 50%, at least 60%, at least 70%, At least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 97% or at least 98% trans-resveratrol. In a particular embodiment, the composition comprises a muscadine (loft grape) pulp extract having a polyphenol content of about 4% and a trans-resveratrol having a minimum purity of at least 98% from a non-musk source. .

In a particular embodiment, the composition comprises from 20% to 50% solids in the liquid, such as at least 23%, at least 25%, at least 30%, at least 35%, at least 37%, at least 40%, at least 42%, Musk extract of muscadine at least 44%, at least 46% or at least 48% solids. In a particular embodiment, the extract comprises about 40% solids in the liquid.

In some embodiments, the resveratrol comprises at least 5% trans resveratrol, such as at least 10% trans resveratrol, at least 20% trans resveratrol, at least 30% trans white gourd Alcohol, at least 40% trans-resveratrol, at least 50% trans-resveratrol, at least 55%, at least 60%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, At least 95%, at least 97% or at least 98% trans-resveratrol. In a particular embodiment, the resveratrol comprises at least 50% trans-resveratrol extracted from the root of Japanese yam (polygonum cuspidatum), such as at least 55%, at least 60%, at least 70%, at least 75%, at least 80% At least 85%, at least 90%, at least 95%, at least 97% or at least 98% trans-resveratrol. Other trans-resveratrol sources, including synthetic trans-resveratrol, are contemplated for use in the disclosed compositions.

In certain embodiments, the disclosed antioxidant composition has a ratio of muscadine polyphenol to trans-resveratrol of at least 0.1/1 (weight/weight), such as 0.25/1, 0.5/1, 0.6/ 1, 0.65 / 1, 0.7 / 1, 0.75 / 1, 0.8 / 1, 0.9 / 1 or 1 / 1 (weight / weight). In other embodiments, the ratio of muscadine polyphenols to trans-resveratrol can be as high as 10/1 (weight/weight), such as 2/1, 3/1, 4/1, 5/1, 7.5/1 Or 9/1 (weight/weight). In a particular embodiment, the disclosed antioxidant composition has a ratio of 0.75/1 (weight/weight) of muscadine polyphenol to trans-resveratrol.

In certain embodiments, the disclosed antioxidant composition has a ratio of muscadine pulp extract to trans-resveratrol in the range of 0.2/1 to 50/1 (weight/weight), such as 0.5/1 1, 1/1, 5/1, 10/1, 15/1, 20/1, 22/1, 25/1, 30/1, 35/1, 40/1 or 45/1 (weight/weight).

In some embodiments, the disclosed composition having antioxidant activity has a total ORAC of at least 21 micromolar Trolox equivalent per microgram of polyphenol (micromolar TE per milligram of polyphenol), such as at least 22 micromoles TE/ Millions of polyphenols, at least 24 micromoles TE per milligram of polyphenols, at least 26 micromoles TE per milligram of polyphenols, at least 28 micromoles TE per milligram of polyphenols or at least 30 micromoles of TE per milligram of polyphenols. In one embodiment, the disclosed composition has a total ORAC of 24 micromoles TE per milligram of polyphenols.

In some embodiments, the disclosed compositions can further include elderberry extract, purple carrot extract, excipients (such as glycerin, sorbitol, colloidal ceria or natural flavoring additives), or combinations thereof. For example, elderberry extract and purple carrot extract can be included to provide composition color or to provide additional antioxidant activity.

In a particular embodiment, the composition comprises from 23% to 32% of the total composition (such as 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31% or 32%) muscadine pulp extract (containing about 4% to 5% polyphenol content), 1.2% to 3.0% of the total composition (such as 1.3%, 1.5%, 1.7%, 1.9%, 2.1%) , 2.3%, 2.5%, 2.7%, or 2.9%) of trans-resveratrol, from 0.12% to 0.20% of the total composition (such as 0.12%, 0.14%, 0.16%, 0.18%, or 0.20%) Purple carrot extract, from 0.4% to 0.6% of the total composition (such as 0.4%, 0.45%, 0.5%, 0.55% or 0.6%) of elderberry extract, accounting for 50% to 61% of the total composition ( Such as 51%, 52%, 53%, 54%, 55%, 56%, 57%, 58%, 59% or 60%) of sorbitol, accounting for 8% to 12% of the total composition (such as accounted for 99.7% glycerin composition of 8%, 9%, 10%, 11% or 12%) and colloidal state of 1% to 2% (such as 1.2%, 1.5%, 1.7% or 2%) of the total composition Ceria. In other specific embodiments, the composition comprises from 8% to 60% of the total composition of the muscadine pulp extract (containing 2% to 14% muscadine polyphenols by weight of the liquid) and 1.2% of the total composition. % to 3.0% trans-resveratrol.

In a more specific embodiment, the composition comprises from 28% to 30% of the total composition of the muscadine pulp extract, from 1% to 2% of the total composition of resveratrol, of the total composition. 0.1% to 0.2% purple carrot extract, 0.4% to 0.5% of the total composition of the elderberry extract, 53% to 56% of the total composition of sorbitol, 9% of the total composition to 11% of 99.7% glycerin composition, 1.5% to 2% of the total composition of colloidal cerium oxide, about 1% to 2% of Concord grape extract, accounting for 0.1% to 0.2% Nacca grape extract and 0.4% to 0.5% red grape powder.

In an even more specific embodiment, the composition comprises about 28.9% of the total composition of the muscadine pulp extract (containing 4.1% polyphenol content), and about 1.65% of the total composition of the trans-cigarette Alcohol, about 0.16% of the total composition of the purple carrot extract, about 0.48% of the total composition of the elderberry extract, about 54.8% of the total composition of sorbitol, about 10% of the total composition % 99.7% glycerol composition, about 2% of the total composition of colloidal cerium oxide, about 1.27% of Concord grape extract, about 0.16% of Cabernet grape extract and about 0.48 % red grape powder.

In another specific embodiment, the composition comprises from 23% to 25% of the total composition of the muscadine pulp extract, from 1% to 2% of the total composition of resveratrol, of the total composition. 0.1% to 0.2% purple carrot extract, 0.4% to 0.5% of the total composition of the elderberry extract, 58% to 61% of the total composition of sorbitol, 9% of the total composition to 11% of 99.7% glycerin composition, 1.5% to 2% of the total composition of colloidal cerium oxide, about 1% to 2% of Concord grape extract, accounting for 0.1% to 0.2% Nacca grape extract and 0.4% to 0.5% red grape powder.

In another even more specific embodiment, the composition comprises about 23.77% of the total composition of the muscadine pulp liquid extract (containing 5% polyphenol content), about 1.65% of the total composition of the trans white Resveratrol, about 0.16% of the total composition of the purple carrot extract, about 0.48% of the total composition of the elderberry extract, about 60.05% of the total composition of sorbitol, the total composition About 10% of 99.7% glycerin composition, about 2% of the total composition of colloidal cerium oxide, about 1.27% of Concord grape extract, about 0.16% of Cabernet grape extract and About 0.48% of red grape powder.

Any of the disclosed compositions can be provided in a non-beverage food, beverage, or liquid or solid dietary supplement. In some embodiments, the disclosed compositions are provided in the form of a beverage. The compositions herein (especially food, beverage, and dietary supplement compositions) may be supplemented with one or more nutrients, particularly one or more vitamins and/or minerals. U.S. Recommended Daily Intake (USRDI) is defined and clarified in the Recommended Daily Dietary Allowance-Food and Nutrition Board, National Academy of Sciences-National Research Council. Unless otherwise specified herein, in the case where a particular mineral is present in the product, the product comprises at least about 1% of USRDI (such as at least about 5%, at least about 10%, at least about 20%, at least about 50%, at least about 75). %, at least about 100%, at least about 150%) of the mineral. Unless otherwise specified herein, where a particular vitamin is present in the product, the product comprises at least about 1% of USRDI (such as at least about 5%, at least about 10%, at least about 20%, at least about 50%, at least about 75%). At least about 100%, at least about 150%) of the vitamin. The amount of vitamins and minerals to be added may depend on the concentration of the other ingredients in the processing and formulation.

Non-limiting examples of such vitamins and minerals include iron, zinc, copper, calcium, phosphorus, niacin, thiamine, folic acid, pantothenic acid, iodine, vitamin A, vitamin C, vitamin B2, vitamin B3, vitamin B6. , vitamin B12, vitamin D, vitamin E and vitamin K. Vitamins and minerals of commercial origin may also be included in the compositions of the present invention.

For example, any soluble salt suitable for such minerals contained in the edible product may be used, such as magnesium citrate, magnesium gluconate, magnesium sulfate, zinc chloride, zinc sulfate, potassium iodide, copper sulfate, copper gluconate. And copper citrate. Calcium sources may include, for example, amino acid chelated calcium, calcium carbonate, calcium oxide, calcium hydroxide, calcium sulfate, calcium chloride, calcium phosphate, calcium hydrogen phosphate, calcium dihydrogen phosphate, calcium citrate, calcium malate, titration Calcium titrate, calcium gluconate, calcium realate, calcium tartrate and calcium lactate, and especially citric acid-calcium malate. As used herein, "vitamin A" includes, but is not limited to, retinol, beta-carotene, retinyl palmitate, and retinol acetate. . Vitamin A can be in the form of, for example, an oil, a microcapsule or an encapsulate. It can also be used to encapsulate ascorbic acid and edible ascorbate. The source of iodine includes an iodide-containing salt such as sodium iodide, potassium iodide, potassium iodate, sodium iodate or a mixture thereof. These salts can be encapsulated. Acceptable forms of iron are well known in the art. The amount of iron compound incorporated into the composition will vary widely depending on the degree of replenishment desired in the final product and the target consumer. The iron-fortified compositions of the present disclosure typically comprise from about 5% to about 100% (such as about 15%, about 25%, about 40%, about 50%, about 70%, about 80%, or about 90%) of USRDI. Iron.

Ferrous iron is typically better utilized by the body than ferrous iron. Highly fertile ferrous salts that can be used in the disclosed ingestible compositions are ferrous sulfate, ferrous fumarate, ferrous succinate, ferrous gluconate, ferrous lactate, ferrous tartrate , ferrous citrate, ferrous amine acid chelate and mixtures of such ferrous salts. While ferrous iron typically has a higher bioavailability, certain iron salts can also provide a highly bioavailable source of iron. The highly bioavailable iron salts useful in the food or beverage compositions of the present disclosure are iron digluconate, ferric ammonium citrate, ferric citrate, ferric sulfate, and mixtures of such iron salts. Highly bioavailable combinations or mixtures of ferrous and iron salts can be used in such edible mixes and ready-to-eat beverages. In addition to the use of ferrous salts and iron salts in such high biomass, the food and beverage dietary compositions of the present disclosure may also include other sources of available iron. Other sources of iron that are particularly suitable for strengthening products include certain iron-sugar-carboxylate complexes.

The zinc compound which can be used in the disclosed composition can be in any conventional form such as zinc sulfate, zinc chloride, zinc acetate, zinc gluconate, zinc ascorbate, zinc citrate, zinc aspartate, zinc picolinate, Amino acid chelated zinc and zinc oxide.

In some embodiments, the food and beverage composition may also include one or more dietary fibers. "Dietary fiber" means a complex carbohydrate that is resistant to enzymatic digestion by mammals, such as those found in plant cell walls and algae, and those produced by microbial fermentation. Examples of such complex carbohydrates are bran, cellulose, hemicellulose, pectin, gums and viscous, seaweed extracts and biosynthetic gums. Sources of cellulosic fibers include plants, fruits, seeds, grains, and rayon (eg, by bacterial synthesis). Commercial fibers such as purified plant cellulose or cellulose powder can also be used. Naturally occurring fibers include fibers from whole citrus peel, citrus endocarp, sugar beet, citrus pulp and sac solids, apple, apricot and watermelon rind.

The acidity of the beverage can be adjusted and maintained within the necessary range by known and conventional methods (e.g., using a food grade acid buffer). Typically, the acidity of the beverage within the above range is the balance between the maximum acidity for microbial inhibition and the optimum acidity for the desired beverage flavor. In some embodiments, the beverage composition has a pH of from about 2 to about 8, such as from about 2 to about 4.5 or from about 2.7 to about 4.2.

Organic and inorganic edible acids can be used to adjust the pH of the beverage composition. The acid may be present in its undissociated form or as its individual salt (for example, dipotassium hydrogen phosphate or disodium hydrogen phosphate, potassium dihydrogen phosphate or sodium dihydrogen phosphate). In some embodiments, the acid comprises citric acid, malic acid, fumaric acid, adipic acid, phosphoric acid, gluconic acid, tartaric acid, ascorbic acid, acetic acid, phosphoric acid, pyruvic acid, or mixtures thereof. The acidulant can also be used as an antioxidant to stabilize the beverage component. Examples of commonly used antioxidants include, but are not limited to, ascorbic acid, ethylenediaminetetraacetic acid (EDTA), and salts thereof.

D. Method of preparing a muscat grape composition having antioxidant activity

Also disclosed herein is a method of preparing a muscadine pulp extract composition having antioxidant activity. The compositions of the present invention can be prepared according to methods well known to those skilled in the art. In general, all components may be dissolved, dispersed or otherwise mixed, separately or in a suitable combination, and dissolved, dispersed or otherwise mixed in water, and stirred with a mechanical stirrer until all ingredients have dissolved. Or suitably dispersed to prepare a composition. All individual solutions can be combined as appropriate. The final mixture may be pasteurized or aseptically filled under appropriate processing conditions, where storage of the stable composition is desired.

Such methods can include adding a muscadine (circle grape) pulp extract having a polyphenol content of at least 2% with a minimum purity of at least 5% (such as at least 10%, at least 20%, at least 30%, at least 40%) At least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 98% or at least 99%) of a combination of resveratrol derived from a non-musk source, wherein the muscadine is more The ratio of phenol to trans-resveratrol is in the range of 0.1/1 to 10/1 (weight/weight), thereby producing a mixture of muscadine pulp extract and trans-resveratrol having antioxidant activity. . The disclosed method can further comprise preparing a muscadine pulp extract, followed by combining the muscadine pulp extract with trans-resveratrol.

In a particular embodiment, preparing the muscadine pulp extract comprises combining the bronze muscadine pulp extract with the purple muscadine pulp extract. In other embodiments, preparing the muscadine pulp extract comprises simultaneously extracting a mixture of bronze muscadine pulp and purple muscadine pulp. In certain embodiments, the muscadine pulp extracts included in the disclosed compositions are prepared according to the methods disclosed herein. However, it is expected that the muscadine pulp extract can be prepared according to methods known in the art, including by crushing, pressing, extracting, filtering (several times) the extract, and concentrating by vacuum evaporation followed by freezing. . In one embodiment, the extraction process utilizes only water and does not add, for example, solvents, carriers, or other components of the preservative to the extract itself. This process is carried out under conditions that retain the polyphenolic compound while reducing the presence of other molecules, such as most sugars. In other embodiments, the extraction process may utilize ethanol or a mixture of ethanol and water. In a particular embodiment, the extraction process can further comprise using an enzyme to clarify or facilitate extraction. For example, an enzyme blend or a pectinase from Aspergillus niger can be used for such purposes. Commercial examples include Scottzyme KS and Scottzyme PEC5L from Scott Laboratories.

In other embodiments, the disclosed compositions are prepared by utilizing a muscadine pulp extract having a total phenol concentration of at least 2%. For example, commercially available muscadine extracts having a total phenol concentration of at least 2% can be used to prepare the disclosed compositions having antioxidant activity.

In a particular embodiment, the method of preparing a composition having antioxidant activity may further comprise extracting the muscadine pulp extract and resveratrol with elderberry extract, purple carrot extract, excipients (such as glycerin, Sorbitol, colloidal cerium oxide or natural flavoring additives) or combinations thereof. In a particular embodiment, the method comprises from 23% to 32% of the total composition of the muscadine pulp extract (containing from about 4% to about 5% polyphenol content) and from 1.2% to 3.0 of the total composition. % of resveratrol, 0.12% to 0.20% of total composition of purple carrot extract, 0.4% to 0.6% of total composition of elderberry extract, 50% to 61% of total composition Sorbitol, a composition of 8% to 12% of the total composition of 99.7% glycerin and a combination of 1% to 2% of the total composition of colloidal cerium oxide.

In a more specific embodiment, the composition comprises about 28.9% of the total composition of the muscadine pulp extract (containing about 4.1% polyphenol content) and about 1.65% of the total composition of resveratrol. About 0.16% of the total composition of the purple carrot extract, about 0.48% of the total composition of the elderberry extract, about 54.8% of the total composition of sorbitol, about 10% of the total composition 99.7% glycerin composition, about 2% of the total composition of colloidal cerium oxide, about 1.27% of Concord grape extract, about 0.16% of Cabernet grape extract and about 0.48% Red grape powder combination.

In another more specific embodiment, the composition comprises about 27.67% of the total composition of the muscadine pulp liquid extract (containing about 5% polyphenol content) and about 1.65% of the total composition of the trans Resveratrol, about 0.16% of the total composition of the purple carrot extract, about 0.48% of the total composition of the elderberry extract, about 60.05% of the total composition of sorbitol, the total composition About 10% of the 99.7% glycerin composition, about 2% of the total composition of colloidal cerium oxide, about 1.27% of Concord grape extract, about 0.16% of Cabernet grape extract and Approximately 0.48% of the red grape powder combination.

In the preparation of the ready-to-drink composition, the beverage concentrate may first be formed. One method of preparing a concentrate form of a beverage composition begins with less than the amount of water used to prepare the beverage composition. Another method is to partially dehydrate the final prepared beverage composition to remove only a portion of the water and any other volatile liquid present. Dehydration can be accomplished according to well known procedures such as evaporation under vacuum. The concentrate can be in the form of a relatively thick liquid. A syrup is typically formed by adding a suitable ingredient, such as an electrolyte or emulsion, to the beverage concentrate. The syrup is then mixed with water to form a final beverage or final beverage concentrate.

The substantially dry mixture of the disclosed compositions can be prepared by blending together all of the desired dry ingredients in the appropriate amounts and ratios. Alternatively, the final prepared beverage composition can be dewatered to produce a substantially dry mixture of the beverage composition. The substantially dry mixture, for example in the form of a powder, granule or lozenge, can be dissolved later in an appropriate amount of carbonated or non-carbonated water to produce the final drinkable beverage or combined with water.

As a food form, the disclosed compositions can be formulated in any optional form, such as particulate, crystalline, pasty, gel, solid or liquid. In such forms, various other materials may be included, such as binders, disintegrants, thickeners, dispersants, resorption enhancers, flavoring agents, buffers, surfactants, dissolution aids, preservatives, emulsifying agents. Agent, isotonicity agent, stabilizer or pH control agent. In certain embodiments, when the disclosed compositions are used as foods, functional foods, and the like that maintain health, including but not limited to improving skin quality, preferably, such that the intended effect of the present disclosure is sufficient to provide resistance The amount of oxidant activity comprises the active ingredients of the compositions and extracts of the invention (e.g., muscadine pulp extract, resveratrol, purple carrot extract, and elderberry extract).

E. Methods and kits for inhibiting (eg, preventing) cellular aging

The compositions disclosed herein are shown to have improved lipophilic antioxidant activity. Oxidative stress is known to be the primary mechanism responsible for normal aging. Lipophilic antioxidants are also known to inhibit various types of skin damage. Based on these observations, methods for inhibiting cellular senescence, such as inhibiting or reducing free radical production or activity, are disclosed. In one embodiment, the method comprises administering to a subject in need thereof a disclosed composition having antioxidant activity sufficient to inhibit or reduce one or more doses associated with cellular aging. For example, inhibiting one or more processes associated with cellular senescence includes reducing or inhibiting free radical formation or activity in the subject ingesting the composition.

In some disclosed embodiments, the method comprises administering an ingestible composition (such as described herein) comprising a muscadine (circle grape) pulp extract derived from bronze muscadine pulp and purple muscadine pulp. By). In some embodiments, the ratio of the bronze muscadine pulp extract to the purple muscadine pulp extract in the muscadine pulp extract is from 0.1 to 10 (weight/weight), such as from 0.3 to 3 (weight/weight). Within the scope. For example, the ratio of bronze muscadine pulp extract to purple muscadine pulp extract is about 2.75 / about 1 (weight / weight), 2.5 / about 1 (weight / weight), about 2.25 / about 1 ( Weight/weight), about 2/about 1 (weight/weight), about 1.5/about 1 (weight/weight) or about 1/about 1 (weight/weight). As used herein, the term "about" is defined as ±0.5. In a particular embodiment, the ratio of bronze muscadine pulp extract to purple muscadine pulp extract is about 2.25 / about 1 (weight / weight).

In some embodiments, the orally administered Muscat (Clover) pulp extract has a polyphenol content of at least 2%. For example, the polyphenol content is at least 3%, at least 3.5%, at least 4%, at least 4.5%, at least 5%, at least 6%, at least 8%, at least 10%, at least 12%, or at least 14%. In a particular embodiment, the muscadine (circle grape) pulp extract has a polyphenol content of about 4%.

In some embodiments, the method comprises administering an ingestible composition comprising a muscadine pulp extract and other additional ingredients including, but not limited to, resveratrol. In some embodiments, the method comprises administering an oral composition having a ratio of muscadine polyphenol to resveratrol of about 0.75/1 (weight/weight). In other embodiments, the method comprises administering at least 8% to 60% of the total composition of the muscadine pulp extract (containing 2% to 14% by weight of the liquid, such as 4%, 6%, 8%) , 10% or 12% muscadine polyphenols) and a composition of trans-resveratrol from 1.2% to 3.0% of the total composition.

In another embodiment, the method comprises administering a muscadine pulp extract comprising a polyphenol content of at least 2% and a white color having a minimum purity of at least 5% from a non-musk source (such as Japanese radix root extract) An oral composition of resveratrol wherein the ratio of muscadine polyphenol to trans-resveratrol is in the range of from 0.1/1 to 10/1 (weight/weight), thereby providing sufficient to inhibit or reduce one or more A composition of antioxidant activity associated with cellular aging, including but not limited to improving skin quality. In a particular embodiment, the method comprises administering a muscadine (musk) pulp extract comprising at least 2% polyphenol content and a minimum purity from a non-musk source (such as Japanese radix root extract). At least 5% of the composition of resveratrol, wherein the ratio of the muscadine pulp extract to resveratrol is from 0.2/1 to 50/1 (weight/weight), such as from 5/1 to 50/1 (weight) /weight) or in the range of 20/1 to 50/1 (weight/weight), thereby providing a composition having antioxidant activity. In some embodiments, the composition has a total ORAC of at least 24 micromoles of Trolox equivalent per microgram of polyphenol (micromolar TE per milligram of polyphenol).

In other embodiments, the methods are inhibited by coating a solution or topical ointment containing a separate muscadine pulp extract or a combination thereof with other active ingredients including, but not limited to, resveratrol. Reduces free radical formation or activity in skin cells to improve skin quality. In one embodiment, the solution or topical ointment comprises the disclosed muscadine pulp extract without resveratrol. In another embodiment, the solution or topical ointment comprises both the disclosed muscadine pulp extract and resveratrol, such as muscadine pulp extract comprising muscadine polyphenol concentrate to make muscadine polyphenols The ratio to resveratrol is about 0.75/1 (weight/weight). The method can be performed by a clinician or other healthcare provider. The method is also designed for home use. This method can reduce the occurrence of skin changes associated with aging, significantly reduce wrinkles in human skin, and improve the tissue quality of the skin. Compositions and kits for improving skin quality are also provided, which may include the disclosed compositions having antioxidant activity (including oral or topical) or muscadine extract and one or more other anti-aging compositions (such as a Or a variety of other antioxidants).

Any skin surface, such as the epidermis of the skin, can be treated by using the methods provided herein. The treatable skin surface includes, but is not limited to, the periosteum, the lips, the cheeks, the nasolabial folds, the forehead, the neck, the upper lip, the stomach, the neck, the back, the chest, the hands, the legs, the feet, or any combination thereof. In one embodiment, the skin of any facial surface can be treated using the methods provided herein. The method can be applied to any facial area and/or any body surface area, and other direct application areas are the chest and neck. More than one skin surface can be treated during the same treatment period. In a particular embodiment, the liquid or cream form of the disclosed composition or extract is applied substantially uniformly to the surface of the skin to form a layer of composition or extract on the skin.

The disclosed compositions or extracts may contain other substances commonly used in cosmetics, such as: perfumes; antimicrobial agents; antibacterial agents; fatliquors; complexing agents and spacers; pearlizing agents; plant extracts; Such as retinol or vitamin C; active agent; preservative; fungicide; surfactant, dye, colorant, pigment or any substance with coloring; emulsifier; thickener; softening substance, moisturizing substance and / Or a moisturizing substance; or other common components of a cosmetic or dermatological formulation, such as an alcohol, a polyol, a polymer, a foam stabilizer, an electrolyte, an organic solvent or a polyoxyl derivative. The composition may also contain functional additives such as exfoliants, oxidizing agents, sunscreens, and skin smoothing agents. The composition may also contain components believed to be beneficial for skin and subcutaneous tissue mesotherapy injection therapy, including antioxidants such as dimethylaminoethanol, alpha lipoic acid, and ascorbic acid. Agents may also be included in the composition to enhance the efficacy of the composition or extract, such as lipophilic and lipophobic penetration enhancers (e.g., triethyl citrate, propylene glycol, fatty acid esters, and other materials). Agents that enhance vascular perfusion of the skin, such as aminophyllin or pentoxifylline, may also be included in the composition. Agents may also be included in the composition which enhance the swell and tension of the skin and cause the underlying subcutaneous tissue to shrink or shrink, such as phosphatidyl choline and deoxycholate sulfate. Physiological substances that provide hormonal benefits, such as substances that are estrogen or testosterrogenic stimuli of the skin, including estriol and testosterone, may also be included in the composition. ).

The disclosed muscadine composition can include one or more preservatives. Such preservatives include, for example, Opthiphen ^TM (from International Specialty Products), Geogard Ultra (from Lonza), preservatives listed in the European Union Cosmetic Directive and other preservatives, such as formaldehyde donors (such as DMDM carbendazim, which can be traded under the trade name GLYDAN) Obtained from Lonza), iodopropyl butyl carbamate (for example, available from LONZATM under the trade name GLYCACIL-STM and/or from Jan Dekker under the trade name DEKABEN LMBTM), for example, p-hydroxybenzoate (for example, Alkyl hydroxybenzoate, such as methyl paraben, ethyl p-hydroxybenzoate, propyl p-hydroxybenzoate and/or butyl p-hydroxybenzoate, phenoxyethanol, ethanol, benzoic acid and water Salicylic acid. The preservative system may further comprise an anti-corrosion aid such as octoxyglycerin or glycine soya. Other preservatives or preservatives include dibromocyanobutane (2-bromo-2-bromomethylglutaronitrile), 3-iodo-2-propynyl butylcarbamate, 2-bromo-2- Nitropropane-1,3-diol, imidazolidinyl urea, 5-chloro-2-methyl-4-isothiazolin-3-one, 2-chloroacetamide, benzalkonium chloride and benzyl alcohol.

The composition may also include one or more conditioning agents, such as water soluble conditioning agents. Other regulators include, for example, compounds known as polytetra-ammonium (especially polytetra-ammonium-1 to poly-quaternary ammonium-56) according to the International Nomenclature for Cosmetic Ingredients (INCI).

The composition can be dispensed from a hose, jar, bottle, pump, can, spray can or spray bottle or from some other known container.

Also disclosed herein are muscadine pulp extract compositions and kits for use in improving skin quality for use by a family or clinician. In one embodiment, the kit is suitable for use by a clinician or beautician. In this embodiment, the kit may further include an applicator that assists in coating the composition. The kit usually also includes instructions for use. Such instructions may be handwritten or in digital format (such as videotape, DVD or CD) for use with electronic devices such as computers, CD players, mp3 players or DVD players and the like. In another embodiment, the kit is suitable for home use. This kit may include an applicator for coating the composition, such as a sponge or cloth. Alternatively, the composition can be applied using one or more fingers. The kit usually also includes instructions for use. Such instructions may be handwritten or in digital format (such as videotape, DVD or CD) for use with electronic devices such as computers, CD players, mp3 players or DVD players and the like.

In some embodiments, the kit includes the disclosed composition or extract and one or more other anti-aging compounds (such as another antioxidant) (eg, in two separate containers or in a single container in a single container) One of vitamin C, vitamin E, selenium and/or beta-carotene). In one embodiment, the composition or extract is applied to the target skin area. In a specific, non-limiting embodiment, the muscadine pulp extract anti-aging kit can be used by a household member as follows. First, to provide a uniform composition on the surface of the skin by using an applicator (such as a sponge or cloth) or by applying a measured composition or extract to the surface of the skin using one or more fingers. Extracts. The composition or extract is allowed to remain passively absorbed into the surface of the skin for a certain length of time, or the absorption of the composition/extract by the skin surface can be promoted by gently rubbing the composition into the surface of the skin with a fingertip. In general, normal skin care procedures such as cosmetic coating and application of other moisturizing agents may continue after the muscadine pulp composition/extract is applied.

This process can be performed twice daily as described, daily, every other day, biweekly, weekly, every other week or monthly, or for some other time interval, such as every 3 to 5 days. Improving skin quality includes reversing skin changes associated with natural or congenital aging, slowing the progression of such skin changes or preventing such skin changes. As used herein, preventing its change refers to the beginning of delaying skin changes to any degree. For example, improving skin quality includes reversing skin changes associated with free radical formation and activity, slowing the progression of such skin changes, or preventing such skin changes. In one embodiment, improving skin quality includes reversing skin changes associated with sunburn or light aging skin changes associated with exposure to sunlight or other forms of actinic radiation (eg, UV radiation and tanning chambers), slowing down such The progression of skin changes or prevents such skin changes. As another example, improving skin quality may also include reversing skin changes caused by extrinsic factors, slowing the progression of such skin changes, or preventing such skin changes, including but not limited to radiation, air. Pollution, wind, cold, humidity, heat, chemicals, smoke, cigarette smoke, and combinations thereof. Improving skin quality may also include reversing, preventing or reducing, for example, caused by certain skin conditions (eg, acne), infections (eg, leishmaniasis), or injuries (eg, abrasions, stab wounds, scratches, or surgical wounds). Scars. The improvement to the skin may also include at least one of the following: making the upper thread less visible, making the upper thread and/or wrinkles feel full, improving the appearance of the underline and/or the contour of the eye contour, improving the appearance of the crow's feet, and reducing / / to reduce wrinkles, especially on the cheeks, forehead (such as vertical wrinkles between the eyes, horizontal wrinkles above the eyes) and / or facial wrinkles around the mouth, and especially the appearance of deep wrinkles, wrinkles or wrinkles, improve skin softness Degree, reduces and/or eliminates fine lines and/or deep lines, wrinkles and wrinkles, and smoothes the skin. Methods for measuring improved skin quality are known in the art. See, for example, U.S. Patent Nos. 6,866,856 and 6,682,763.

Skin changes that can be treated by performing such methods and using the kits disclosed herein include, for example, wrinkles (including but not limited to human facial wrinkles), wrinkles, deep wrinkles, wrinkles, and fine lines, deepening of skin lines Thinning of the skin, preventing or reducing scarring, yellowing of the skin, pigmentation, hyperpigmentation, pigmentation and/or pigment-free age spots, leatheriness, loss of elasticity, loss of recoilability, loss of collagen fibers The elastic fiber changes abnormally, the small blood vessels of the dermis degenerate, and the surface of the skin forms a vascular structure with increased sunlight, and a combination thereof.

Improving skin quality includes reducing, reducing, and/or minimizing one or more of the above-described skin changes. Improving skin quality will give the skin a younger look. Improving skin quality will result in a smoother, moisturized (less dry) or less scaly appearance. For example, in certain embodiments, improving skin quality can include a reduction in roughness, dryness, or degree of scaling. Improving skin quality includes desalination and improvement of lines and wrinkles, and improvement of swell pressure and tension. The desired effect is lift and firmness.

The texture of the skin can be improved, including softness, softness and smoothness, resulting in enhanced gloss, clarity and brightness. Other and important qualities of the subject that can be subjectively and objectively measured include, but are not limited to, skin sagging or reverse skin firmness, and the presence and extent of fine lines and streaks in the skin.

These qualities are the same qualities that determine the external appearance of the appearance (eg skin aging). Improving the quality by the methods and kits disclosed herein produces benefits based on visual appearance judgments. Changing skin quality by the methods disclosed herein reduces the appearance of skin aging.

The desired benefit may include not only physiological benefits to the skin, but also therapeutic and pharmacological benefits, such as possible prevention and treatment of malignant diseases, whether by chemoprevention or enhanced photodynamic therapy. Benefits may also include treating and inhibiting acne by including a composition that inhibits sebaceous gland activity.

F. Pharmaceutical composition

The disclosed muscadine pulp extracts and compositions can be at least suitable for inhibiting (e.g., preventing) one or more oxidation processes, such as free radical formation associated with cellular events such as cellular senescence. Accordingly, a pharmaceutical composition comprising at least one of the disclosed muscadine pulp extracts alone or in combination with resveratrol is also described herein.

Formulations for use in pharmaceutical compositions are well known in the art. For example, Remington's Pharmaceutical Sciences , EW Martin, Mack Publishing Company, Easton, PA, 19th edition, 1995 describes suitable for the delivery of at least one of the disclosed muscadine pulp extracts alone or in combination with resveratrol. An exemplary formulation (and its components). In some embodiments, the compositions also include other agents, such as purple carrot root extract or elderberry extract. Pharmaceutical compositions comprising at least one such compound can be formulated for use in human medicine or veterinary medicine. Particular formulations of the disclosed pharmaceutical compositions may, for example, depend on the mode of administration (e.g., oral, topical or parenteral) and/or the condition to be treated (e.g., free radical production or activity). In some embodiments, the formulation also includes a pharmaceutically acceptable carrier in addition to at least one active ingredient such as a muscadine pulp extract.

Pharmaceutically acceptable carriers suitable for use in the disclosed methods and compositions are well known in the art. The nature of the pharmaceutical carrier will depend on the particular mode of administration employed. For example, parenteral formulations typically comprise an injectable fluid as a vehicle, including pharmaceutically and physiologically acceptable fluids such as water, physiological saline, balanced salt solutions, aqueous dextrose, glycerol or Its analogues. For solid compositions such as powders, pills, lozenges or capsules, conventional non-toxic solid carriers may include, for example, pharmaceutical grade mannitol, lactose, starch or magnesium stearate. In addition to the biological neutral carrier, the pharmaceutical composition to be administered may optionally contain minor amounts of non-toxic auxiliary substances or excipients such as wetting or emulsifying agents, preservatives and pH buffers and the like; for example, acetic acid Sodium or sorbitan monolaurate. Other non-limiting excipients include nonionic solubilizing agents such as cremote, or proteins such as human serum albumin.

The disclosed pharmaceutical compositions can be formulated as pharmaceutically acceptable salts. A pharmaceutically acceptable salt is a non-toxic salt of a compound in free base form having the desired pharmacological activity of the free base. These salts can be derived from inorganic or organic acids. Non-limiting examples of suitable inorganic acids are hydrochloric acid, nitric acid, hydrobromic acid, sulfuric acid, hydroiodic acid, and phosphoric acid. Non-limiting examples of suitable organic acids are acetic acid, propionic acid, glycolic acid, lactic acid, pyruvic acid, malonic acid, succinic acid, malic acid, maleic acid, fumaric acid, tartaric acid, citric acid, Benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, methanesulfonic acid, salicylic acid, formic acid, trichloroacetic acid, trifluoroacetic acid, gluconic acid, aspartic acid (asparagic) Acid/aspartic acid), benzenesulfonic acid, p-toluenesulfonic acid, naphthalenesulfonic acid and the like. A list of other suitable pharmaceutically acceptable salts is found in Remington's Pharmaceutical Sciences , 19th Edition, Mack Publishing Company, Easton, PA, 1995. Pharmaceutically acceptable salts can also be used to adjust the osmotic pressure of the composition.

The dosage form of the disclosed pharmaceutical composition will be determined by the mode of administration selected. For example, an oral dosage form can be used. The oral formulation can be a liquid, such as a syrup, solution or suspension; or a solid, such as a powder, pill, lozenge or capsule. Methods of preparing such dosage forms are disclosed herein or as will be apparent to those skilled in the art.

Certain specific examples of pharmaceutical compositions comprising the disclosed compounds may be formulated in unit dosage forms suitable for the precise administration of the precise dosage. The amount of active ingredient administered will depend on the individual being treated, the severity of the condition, and the mode of administration, and are known to those skilled in the art. Within such ranges, the formulation to be administered will contain an amount of the extract or compound disclosed herein in an amount effective to achieve the desired effect in the subject being treated. In a particular embodiment, for oral administration, the composition is provided as a liquid containing about 75 mg of muscadine pulp extract polyphenol and 100 mg of trans-resveratrol. In other embodiments, for oral administration, the composition contains from about 1.0 mg to about 500 mg of active ingredient, especially about 10.0 mg, about 50 mg, 100 mg, about 150 mg, about 175 mg, about 200 mg, about 210 mg, about 225 mg. A dosage form of about 250 mg, about 300 mg, about 400 mg or about 500 mg of the active ingredient is provided for symptomatic adjustment of the dosage administered to the individual being treated. In one exemplary oral administration regimen, a lozenge containing from about 1 mg to about 500 mg (such as from about 150 mg to about 225 mg) of the active ingredient is administered at least once a day. In other embodiments, a lozenge containing from about 1 mg to about 500 mg (such as from about 150 mg to about 225 mg) of active ingredient is administered at least once a week, such as at least twice, three times, at least four times, at least five per week or daily. Times or at least six times.

G. Administration of extracts and compositions disclosed

The method of administering any of the disclosed compositions and extracts is a conventional method. For example, the disclosed methods, such as methods comprising a composition having antioxidant activity for preventing or reducing one or more processes associated with aging, can be via injection, intratumoral, oral, topical, transdermal , administered parenterally or via inhalation or spray. In a particular embodiment, a mammalian individual, such as a human, is orally administered as a composition in the form of a non-beverage food, beverage or dietary supplement. In another embodiment, the composition is topically administered to the surface of the skin of a mammalian subject, such as a human.

The therapeutically effective amount of the agent administered can vary depending on the desired effect and the individual to be treated. In one embodiment, the method comprises administering to the individual (such as a human subject) at least 1 mg of the composition per day. For example, at least 1 g or at least 10 g of the composition can be administered daily to humans, such as 1 g to 5 g per day, 5 g to 10 g per day, for example 7 g per day. In one embodiment, at least 5 grams of the composition comprising the muscadine pulp extract and resveratrol are administered to the individual. In other embodiments, at least 6.3 g of the composition is administered orally to an individual. The dose may be administered in divided doses (such as 2, 3 or 4 doses per day) or in a single daily dose.

In a particular embodiment, the therapeutic composition is administered to the individual in a plurality of daily dosing schedules (such as at least 2 consecutive days, for 10 consecutive days, etc.), such as the resveratrol and muscat grape extracts disclosed. Anti-aging supplements, for example, for weeks, months or years. In one embodiment, the subject is administered a therapeutic composition (such as the disclosed anti-aging supplement comprising resveratrol and muscadine extract), including daily administration for at least 30 days, such as At least 2 months, at least 4 months, at least 6 months, at least 12 months, at least 24 months, or at least 36 months.

A therapeutic composition such as the disclosed anti-aging supplement comprising trans-resveratrol and muscadine pulp extract or muscadine pulp extract alone may further comprise one or more biologically active or inactive compounds (or Both), such as other antioxidants and conventional non-toxic pharmaceutically acceptable carriers. In a particular embodiment, the therapeutic composition further comprises one or more biologically inactive compounds. Examples of such biologically inactive compounds include, but are not limited to, carriers, thickeners, diluents, buffers, preservatives, and carriers. Pharmaceutically acceptable carriers suitable for use in such formulations are conventional (see Remington's Pharmaceutical Sciences, E. W. Martin, Mack Publishing Company, Easton, PA, 19th Edition (1995)). In general, the nature of the carrier will depend on the particular mode of administration employed. For example, a parenteral formulation can include an injectable fluid as a vehicle, including pharmaceutically and physiologically acceptable fluids such as water, physiological saline, balanced salt solutions, aqueous dextrose, glycerol or Its analogues. For solid compositions (eg, powders, pills, troches, or capsules), conventional non-toxic solid carriers can include, for example, pharmaceutical grade mannitol, lactose, starch, or magnesium stearate. In addition to the biologically neutral carrier, the pharmaceutical composition to be administered may also include minor amounts of non-toxic auxiliary substances such as wetting or emulsifying agents, preservatives and pH buffering agents and the like, such as sodium acetate or sorbitan. Monolaurate.

Although the present disclosure has provided a detailed description of the use of any of the disclosed compositions and extracts to prevent or inhibit one or more processes associated with cellular senescence, it is contemplated that the disclosed compositions and extracts can be used not only as a preventative Or an antioxidant that inhibits one or more processes associated with cellular aging, and can treat any condition associated with oxidative stress. For example, it is contemplated that the extracts and compositions of the present invention are useful for reducing, preventing or treating the pathogenesis of chronic inflammatory diseases such as diabetes, cancer, atherosclerosis, and other cardiovascular diseases, as well as with, for example, Alzheimer's Oxidative stress associated with degenerative diseases of Parkinson's disease.

The subject matter of the present disclosure is further illustrated by the following non-limiting examples.

Example

Example 1

Determination of Solubility of Turkish Tannin in Musk Grape Fruit Extract

This example illustrates the effect of various ratios of bronze and purple pulp extracts on the solubility of citric acid in Turkey.

The separately fermented bronze and purple pulp extracts were heated and then mixed in a total volume of 100 ml at the ratios provided in Table 1.

Since the individual bronze and purple pulp extracts were not concentrated, the resulting extract mixture was 1X concentration. The 1X concentration typically contains about 2% solids solution (100 grams of solution equals 2 grams of dry extract). Each of the approximately 20 ml ratios was transferred to individual glass tubes and covered with foil. The sample was heated at 85 ° C for 30 minutes, then removed from the heat source and allowed to reach room temperature. The sample was then filtered through a 0.45 μm PTFE filter w/GMP and analyzed by HPLC/MS to determine its Turkish tannic acid content. The individual bronze and purple muscadine pulp extracts were also analyzed by the same method to determine the baseline value of turmeric tannin in both the bronze and purple pulp extracts and to determine the anthocyanin content of the purple pulp extract. Then, UV-VIS detection was carried out by reverse phase HPLC at 254 nm, and the ion trap mass detector was used to detect the sample by using an extracted ion chromatogram (EIC) at [MH] ^- = 301 amu. The total range of citric acid in Turkey.

To prepare a bronze/purple pulp extract having various ratios of 40% solids, 1 x individual bronze and purple muscadine pulp extracts were dried to a powder form and reconstituted with water to produce a liquid containing 40% solids. The two separate solutions are then mixed in various ratios depending on the weight (wt) rather than the volume. The various ratios evaluated are provided in Tables 2A and 2B.

Each ratio of about 10 ml was transferred to a separate glass test tube and covered with a foil. The sample was heated at 85 ° C for 30 minutes, then removed from the heat source and allowed to reach room temperature. The sample was then filtered through a 0.45 μm PTFE filter w/GMP and analyzed by HPLC/MS to determine its Turkish tannic acid content. The individual bronze and purple musk grape pulp extracts were also analyzed by the same method to determine the baseline value of the Turkish tannic acid in both the bronze and purple musk grape pulp extracts and to determine the purple color in the purple musk grape pulp extract. The content of the prime. Combined with the ion trap mass detector, the extracted ion chromatogram (EIC) was used to detect at [MH] ^- = 301 amu, and the verification test method C2505 was used to determine the total range of citric acid in the sample.

These studies found that the ratio of 65%/35% bronze to purple pulp extract (volume by volume) is sufficient to increase the maximum solubility of the Turkish tannic acid in a mixture of the two pulp extracts at a concentration of 1x. In addition, at 40% solids, the solubility of citric acid in Turkey continued to increase with the increase in the content of the purple muscadine pulp extract in the mixture. These findings indicate that at 40% solids, a larger amount of turmeric acid is present in solid form, so a higher level of purple pulp extract is required to completely dissolve all of the turmeric acid present in the mixture. These studies may also indicate that the anthocyanins contained in the purple muscadine pulp extract affect the solubility of the Turkish tannic acid contained in the bronze musk grape pulp extract.

Example 2

Anti-aging dietary supplement and preparation thereof

This embodiment provides a dietary supplement that is edible to prevent or inhibit one or more processes associated with cellular aging.

The disclosed anti-aging dietary supplements are prepared according to the formulations provided in Table 3a or Table 3b.

Dietary supplements include instructions for dosage. These instructions indicate that 5 ml of supplement (about one tablespoon) can be taken daily before a meal and used as an anti-aging supplement to, for example, inhibit one or more processes associated with cellular aging.

Example 3

Antioxidant ability of muscadine grape fruit extract and Japanese root extract and anti-aging supplement

This example demonstrates that the mixture and supplement containing the Japanese cockroach extract and the muscadine pulp extract has improved antioxidant capacity, as measured by the Oxygen Free Radical Absorption Capacity (ORAC) assay.

The ORAC assay was used to assess the antioxidant capacity of the two plant extracts, either alone or in combination as a mixture or supplement. This assay has been used to quantify the antioxidant capacity of a variety of foods and beverages and this check is based on the information contained in the USDA ORAC database. Both hydrophilic antioxidant capacity and lipophilic antioxidant capacity can be measured by this test.

The following samples were analyzed: (1) normalized to a minimum of 98% trans-resveratrol dry/powdered Japanese root extract (actual content is 100% trans-resveratrol); (2) containing 14.4% total Phenol content dry/powdered muscadine pulp extract (2:1 ratio of bronze to purple pulp); (3) dried/powdered Japanese glutinous rice and dried/powdered muscadine pulp extract a mixture of 1:5.36 (wt:wt) ratio (total polyphenol content is 27.5% of the mixture and the ratio of muscadine polyphenol to trans-resveratrol is 0.75/1); and (4) Table 2a of Example 2 An anti-aging supplement provided in the product. The hydrophilicity, lipophilicity and total ORAC value were measured (the total ORAC value is the sum of the hydrophilic value and the lipophilic value) and the result is the number of micromolar Trolox equivalents per milligram of polyphenol (micromolar TE/mg polyphenol) Said. These results are shown in Table 4.

As shown in Table 4, the measured hydrophilic ORAC value of the mixture was similar to the predicted value based on the additive effect of the two extracts. However, the measured lipophilic ORAC value of the mixture was 5 times the predicted additive value, resulting in a 20% increase in the total ORAC value. A similar synergistic effect was observed for the composition, which is close to three times the predicted ORAC score based on additive. Table 5 shows the results expressed as ORAC values per gram of material (as opposed to each milligram of polyphenol shown in Table 4). As shown in Table 5, when the value is expressed in ORAC per gram of the substance, the synergistic effect of the mixture of the muscadine pulp extract and the Japanese jaundice extract and the anti-aging supplement in the production of lipophilic antioxidant capacity is maintained. :

These studies have shown a strong synergistic effect of the mixture of muscadine grape fruit extract and Japanese root extract and anti-aging supplements in producing lipophilic antioxidant capacity. It was not expected to exhibit selective synergy under lipophilic conditions. Thus, the findings of the present invention indicate that the mixture containing the Japanese cockroach extract and the muscadine pulp extract has improved antioxidant capacity as measured by the ORAC assay as compared to any of the individual extracts.

Example 4

Antioxidant capsule formulation and preparation

This embodiment provides an antioxidant capsule formulation and preparation thereof, which formulation can be used to inhibit one or more oxidation processes, including preventing or inhibiting free radical formation.

An aqueous extract of bronze and purple pulp with a ratio of solids content of 40% and a polyphenol content of 10% by solids (measured by Gallic Acid equivalent (GAE)) of 2:1 was treated as follows : 1.33 Kg of maltodextrin MD100 was added to 10 Kg of the pulp concentrate mentioned above. The mixture is blended to dissolve the maltodextrin. This liquid concentrate is then lyophilized to produce a dry matter. The chemical values of the dried muscadine/maltodextrin material are as follows: (1) total polyphenol (in % GAE), NLT 7.0% (dry basis weight); gallic acid (HPLC), NLT 0.3% (dry basis weight) And; free Turkish citric acid (HPLC), NLT 0.15% (dry basis weight). The characteristics of dried muscadine/maltodextrin are provided in Table 6.

Capsules are prepared by weighing out individual ingredients in divided batches. Passing these materials through Comil The grinder finally passes the maltodextrin through the grinder. The ground material was blended until uniform and then filled into a two-piece capsule of size "0" on the encapsulator. An example of the dosage form produced is provided in Table 7 and the results of the analysis of the muscadine/resveratrol capsules are provided in Table 8.

Example 5

Reduction of oxidative stress in human individuals

This example demonstrates the in vivo efficacy of the antioxidant capsules described in Example 4 as assessed in a placebo-controlled clinical trial.

The capsules contained a mixture of muscadine pulp extract and Japanese cockroach extract to provide 75 mg of muscadine polyphenol and 100 mg of trans-resveratrol per 3 capsules (Tables 7 and 8). Dietary consumption is known to increase oxidative stress in the short term and produce an inflammatory response. For example, consumption of 900kcal fast-food meals has been shown to cause a significant increase in reactive oxygen species (ROS) in circulating mononuclear cells (MNCs) and polymorphonuclear leukocytes (PMNs) in healthy normal-weight humans. (Aljada et al . Am. J. Clin. Nutr. 79:682-690, 2004). These changes are accompanied by an increase in the expression of MNC of the p47phox protein, which is a subunit of NADPH oxidase (mediator of ROS production).

To test the role of antioxidant capsules in this model of post-prandial oxidative stress, a group of 10 healthy individuals (age: 37 ± 4 years, BMI 22.6 ± 0.5 kg/m2) were provided, in separate two days, before meals. Eat 910kcal fast food meal and single dose (3 capsules) of antioxidant formulation or placebo (3 capsules) for 10 minutes. Blood samples were collected at baseline and at 1, 3, and 5 hours after dietary intake. The circulating concentrations of glucose, insulin and lipid were similar after each treatment, indicating that the disclosed antioxidant capsules did not alter the digestion/absorption of the diet. However, one hour after dietary consumption, the disclosed antioxidant capsule treatment significantly attenuated the increase in ROS production in MNC (15% vs. 62% in placebo) and PMN (8% vs. 64% in placebo). . In addition, dietary plus placebo intake resulted in a significant 148% increase in protein content of p47phox in MNC relative to baseline (P < 0.05), whereas dietary plus antioxidant capsule intake completely prevented any significant change in MNC p47phox content (P < 0.05, using double Factor RMANOVA). In addition, compared with the placebo-treated group in which the DNA-binding activity of the antioxidant stress transcription factor Nrf-2 was actually decreased, the DNA-binding activity of the antioxidant stress transcription factor Nrf-2 was relatively 3 hours after the intake of the dietary antioxidant capsule. Significantly increased by 150% at baseline (P < 0.05, according to RMANOVA and two-factor RMANOVA). Nrf-2 binds to an antioxidant response element (ARE) promoter sequence encoding multiple genes of an endogenous antioxidant enzyme (eg, glutathione-S-transferase) and, therefore, is ingested An increase in the DNA binding activity of Nrf-2 following the disclosed antioxidant formulation indicates the possibility of upregulating multiple cellular defenses.

Finally, at 3 hours, plasma endotoxin levels were significantly increased by 60% (P < 0.05) relative to baseline after dietary and placebo ingestion, while when the diet was combined with the disclosed antioxidant capsules, At 1 hour, the endotoxin concentration decreased significantly by 28% relative to baseline (P < 0.05, using RMANOVA and two-way RMANOVA). Endotoxin is an effective inflammatory signal that promotes the activation of a variety of cytokines that enhance oxidative stress.

These results indicate a novel in vivo antioxidant effect of the mixture of muscadine grape puree extract and Japanese glutinous rice (the ratio of muscadine polyphenols to trans-resveratrol is 0.75/1) and thus extend the synergistic resistance of this mixture. The oxidizing ability was found in a test tube (Example 3).

In view of the many possible specific examples to which the principles of the present disclosure may be applied, it is to be understood that the illustrated embodiments are only examples of the disclosed embodiments and should not be construed as limiting the scope of the disclosure. Rather, the scope of the disclosure is defined by the scope of the following claims. Accordingly, we have claimed that all of our inventions are within the scope and spirit of such patent application.

Claims (32)

A method of preparing an antioxidant oral dietary supplement composition comprising: a liquid solvent extract of muscadine ( Vitis rotundifolia ) pulp having at least 2% polyphenol content and a source other than muscat a resveratrol combination having a minimum purity of at least 5% trans-resveratrol, wherein the ratio of the polyphenol to trans-resveratrol in the muscadine pulp extract is 0.1/ From 1 to 10/1 (weight/weight), a mixture of muscadine pulp extract and trans-resveratrol having antioxidant activity as an oral dietary supplement composition is thereby produced. The method of claim 1, wherein the resveratrol comprises at least 98% trans-resveratrol extracted from the roots of Polygonum cuspidatum . The method of claim 1 or 2, wherein the muscadine (bougainvillea) pulp extract has a total polyphenol content of about 4% and the ratio of the muscadine pulp extract to the resveratrol It is about 18/1 (weight/weight). The method of claim 1, further comprising preparing the muscadine pulp extract, and subsequently combining the muscadine (musk) pulp extract with the resveratrol. The method of claim 4, wherein the extracting the muscadine pulp extract comprises combining a bronze muscadine pulp extract with a purple muscadine pulp extract, and wherein the bronze muscadine pulp extract is The mixture with the purple muscadine pulp extract promotes the solubility of turkey ellagic acid in the muscadine pulp extract. The method of claim 4, wherein the extracting the muscadine pulp extract comprises simultaneously extracting a mixture of purple muscadine pulp and bronze muscadine pulp to promote the solubility of citric acid in the muscadine pulp extract. . The method of claim 6, wherein the ratio of the bronze muscadine pulp extract to the purple muscadine pulp extract is in the range of 0.3 to 3 (weight/weight). The method of claim 6 or 7, wherein the muscadine pulp extract comprises 20% to 50% solids in the liquid. The method of claim 8, wherein the muscadine pulp extract comprises about 40% solids in the liquid. The method of claim 1 or 2, further comprising adding the elderberry extract, the purple carrot extract or a combination thereof to the mixture of the muscadine pulp extract and resveratrol. The method of claim 1 or 2, further comprising adding an excipient to the mixture of the muscadine pulp extract and resveratrol. The method of claim 11, wherein the excipient comprises glycerin, sorbitol, colloidal cerium oxide, a natural flavoring additive, or a combination thereof. The method of claim 5, wherein the bronze muscadine pulp extract and the purple musk grape pulp extract are filtered. And the fermented extract. An oral dietary supplement composition for use as an antioxidant comprising a muscadine (circle grape) pulp extract having at least 2% polyphenol content and a source other than muscadine having a minimum purity of at least 5 Resveratrol of trans-resveratrol, wherein the ratio of polyphenols to trans-resveratrol in the muscadine pulp extract is in the range of 0.1/1 to 10/1 (weight/weight), thereby An oral dietary supplement composition having antioxidant activity is provided. An oral dietary supplement composition according to claim 14, wherein the resveratrol comprises at least 98% trans-resveratrol extracted from the roots of Polygonum cuspidatum . An oral dietary supplement composition according to claim 14 or 15, wherein the muscadine (round leaf grape) pulp extract has a total polyphenol content of about 4% and the muscadine pulp extract and the white The ratio of resveratrol is about 18/1 (weight/weight). The oral dietary supplement composition of claim 14 or 15, wherein the muscadine pulp extract comprises a combination of muscadine pulp extract obtained from bronze muscadine pulp and purple muscadine pulp. Promotes the solubility of citric acid in muscadine grape fruit extract. The oral dietary supplement composition of claim 17, wherein the ratio of the bronze muscadine pulp extract to the purple muscadine pulp extract is in the range of 0.3 to 3 (weight/weight). The oral dietary supplement composition of claim 17, wherein the muscadine pulp extract is contained in the liquid in an amount of 20% to 50% solid. The oral dietary supplement composition of claim 19, wherein the muscadine pulp extract comprises about 40% solids in the liquid. The oral dietary supplement composition of claim 14 or 15, further comprising an elderberry extract, a purple carrot extract or a combination thereof. The oral dietary supplement composition of claim 14 or 15 further comprising an excipient. The oral dietary supplement composition of claim 22, wherein the excipient comprises glycerin, sorbitol, colloidal cerium oxide, a natural flavoring additive, or a combination thereof. An oral dietary supplement composition according to claim 14 or 15, wherein the composition has a total oxygen radical absorption capacity of at least 24 micromoles of Trolox equivalent per microgram of polyphenol (micromolar TE/mg polyphenol) (Oxygen Radical Absorbance Capacity, ORAC). The oral dietary supplement composition of claim 14 or 15, wherein the composition is provided in a non-beverage food, beverage, liquid or solid dietary supplement. An oral dietary supplement composition as claimed in claim 14 or 15 which is used as an anti-aging supplement. The oral dietary supplement composition of claim 14 or 15, wherein the composition has a ratio of muscadine polyphenol to resveratrol of about 0.75/1 (weight/weight). The oral dietary supplement composition of claim 17, wherein the bronze muscadine pulp extract and the purple musk grape pulp extract are filtered and fermented extracts. A composition for inhibiting cell senescence comprising an oral dietary supplement composition according to any one of claims 14 to 28, which comprises a dose sufficient to inhibit one or more processes associated with cell senescence. The composition of claim 29, wherein the inhibiting one or more of the processes associated with cellular senescence comprises preventing or inhibiting free radical formation or activity in an individual ingesting the composition. The use of an oral dietary supplement composition according to any one of claims 14 to 28, which is for the manufacture of a medicament for inhibiting cell senescence, wherein the dose of the composition in the medicament is sufficient to inhibit one Or multiple processes associated with cellular aging. The use of claim 31, wherein inhibiting one or more of the processes associated with cellular senescence comprises preventing or inhibiting free radical formation or activity in an individual ingesting the composition.