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WO2008108817A1 - Nutritional supplement with oleuropein - Google Patents

Nutritional supplement with oleuropein Download PDF

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Publication number
WO2008108817A1
WO2008108817A1 PCT/US2007/020953 US2007020953W WO2008108817A1 WO 2008108817 A1 WO2008108817 A1 WO 2008108817A1 US 2007020953 W US2007020953 W US 2007020953W WO 2008108817 A1 WO2008108817 A1 WO 2008108817A1
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WO
WIPO (PCT)
Prior art keywords
acid
oleuropein
juice
nutritional supplement
olive leaf
Prior art date
Application number
PCT/US2007/020953
Other languages
French (fr)
Inventor
Richard Hall
James Kwon
Original Assignee
Ameriden International, Inc.
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Ameriden International, Inc. filed Critical Ameriden International, Inc.
Publication of WO2008108817A1 publication Critical patent/WO2008108817A1/en

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    • AHUMAN NECESSITIES
    • A23FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23LFOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES, NOT OTHERWISE PROVIDED FOR; PREPARATION OR TREATMENT THEREOF
    • A23L2/00Non-alcoholic beverages; Dry compositions or concentrates therefor; Preparation or treatment thereof
    • A23L2/02Non-alcoholic beverages; Dry compositions or concentrates therefor; Preparation or treatment thereof containing fruit or vegetable juices
    • A23L2/04Extraction of juices
    • AHUMAN NECESSITIES
    • A01AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01NPRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N65/00Biocides, pest repellants or attractants, or plant growth regulators containing material from algae, lichens, bryophyta, multi-cellular fungi or plants, or extracts thereof
    • AHUMAN NECESSITIES
    • A01AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01NPRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N65/00Biocides, pest repellants or attractants, or plant growth regulators containing material from algae, lichens, bryophyta, multi-cellular fungi or plants, or extracts thereof
    • A01N65/08Magnoliopsida [dicotyledons]
    • A01N65/28Myrtaceae [Myrtle family], e.g. teatree or clove
    • AHUMAN NECESSITIES
    • A01AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01NPRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N65/00Biocides, pest repellants or attractants, or plant growth regulators containing material from algae, lichens, bryophyta, multi-cellular fungi or plants, or extracts thereof
    • A01N65/40Liliopsida [monocotyledons]
    • A01N65/44Poaceae or Gramineae [Grass family], e.g. bamboo, lemon grass or citronella grass
    • AHUMAN NECESSITIES
    • A23FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23LFOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES, NOT OTHERWISE PROVIDED FOR; PREPARATION OR TREATMENT THEREOF
    • A23L19/00Products from fruits or vegetables; Preparation or treatment thereof
    • AHUMAN NECESSITIES
    • A23FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23LFOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES, NOT OTHERWISE PROVIDED FOR; PREPARATION OR TREATMENT THEREOF
    • A23L2/00Non-alcoholic beverages; Dry compositions or concentrates therefor; Preparation or treatment thereof
    • AHUMAN NECESSITIES
    • A23FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23LFOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES, NOT OTHERWISE PROVIDED FOR; PREPARATION OR TREATMENT THEREOF
    • A23L2/00Non-alcoholic beverages; Dry compositions or concentrates therefor; Preparation or treatment thereof
    • A23L2/38Other non-alcoholic beverages
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18Magnoliophyta (angiosperms)
    • A61K36/185Magnoliopsida (dicotyledons)
    • A61K36/63Oleaceae (Olive family), e.g. jasmine, lilac or ash tree
    • YGENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02ATECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
    • Y02A50/00TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
    • Y02A50/30Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Definitions

  • the present invention relates, in general, to a nutritional supplement and, more particularly, to a nutritional supplement preferably comprised of a processed olive leaf extract in which oleuropein is captured within cyclodextrin with the aid of glycerin ester, thereby greatly suppressing the bitterness of oleuropein.
  • Oleuropein is a naturally occurring compound commonly found in the leaves, fruit, bark and roots of olive trees.
  • the leaves of an olive tree are richer in oleuropein than the fruit and/or bark of the tree.
  • Oleuropein can also be obtained from other plants belonging to the family Oleaceae; for example, ash and privet trees.
  • oleuropein is known to have a resistant activity against a wide spectrum of bacteria, viruses, and fungi, producing various therapeutic effects, such as alleviation of hypertension, antioxidation, control of glucose levels in blood, etc.
  • oleuropein hydrolyzes to d-elenolic acid and 1-elenolic acid that has powerful antimicrobial effects.
  • the d-elenolic acid acts as a strong antimicrobial.
  • oleuropein has the ability to interfere with critical amino acid processes that are essential for virus replication, thus inhibiting the spread of such viruses.
  • oleuropein can also directly stimulate the formation of immune system cells through a process called phagocytosis in order to combat various types of microbes. Therefore, oleuropein acts as a potent natural antibiotic against viruses and bacteria.
  • oleuropein has been the source of great interest concerning coronary dilation (the expansion/contraction ability of the arteries). Tests have shown that the injection of oleuropein in animals has improved blood circulation by up to 50%. In addition to the coronary artery dilating effect, oleuropein has been found to display other cardiovascular health-enhancing effects, such as inducing a long-lasting effect of lowering elevated blood pressure (as proven using animals when arrhythmia was induced through the injection of chlorides or calcium compounds). More specifically, a study, conducted by the Pharmacology University of Granada, Spain, disclosed that oleuropeoside, an ingredient isolated from oleuropein, dilates the blood vessels. That is, oleuropeoside acts to relax and smoothen contracted blood vessels to improve blood circulation and thus to decrease blood pressure.
  • Oleuropein is also believed to directly promote insulin release in the human body and maximize the use of blood sugar. At least one study has found that oleuropein may play a pivotal role in glycemic control. The recordings of this lab disclosed that when 16 mg of oleuropeoside was administered to animals when diabetes had been induced by alloxan injection, blood sugar levels were significantly reduced in these injected animals. Oleuropein was observed to have a proposed mechanism of action, of potentiation, of glucose-induced insulin release, and an increase in peripheral blood glucose uptake.
  • Pickling the extract with salt and acetic acid may break down oleuropein decreasing its nutritional value. Fermenting the extract is a complicated process and is of limited use in producing beverage and processed foods. The thermal method of heating oleuropein to reduce the bitterness also reduces the preventive and therapeutic effects oleuropein has on maladies.
  • the oleuropein used in this nutritional supplement will be in the form of an extract from olive leaves having a high concentration thereof; however, it is understood that the oleuropein to be included may be obtained from other sources as well.
  • the invention is directed toward a processed olive leaf extract, including oleuropein, cyclodextrin and glycerin ester as its essential ingredients.
  • the processed olive leaf extract contains oleuropein in an amount from 0.5 to 1,000 mg, cyclodextrin in an amount from 0.5 to 100 mg, and glycerin ester in an amount from 2 to 1 ,500 mg.
  • the processed olive leave extract is preferably embodied in the form of a powdered or candied nutritional supplement that further includes additional ingredients selected from a group consisting of xylitol, an acid, calcium, a sweetener, and/or any combination thereof.
  • the processed olive leaf extract is embodied in the form of a powdered or candied nutritional supplement that further includes a sweetener
  • the sweetener is preferably selected from the group consisting of, but not limited to, xylitol, rebaudiosides , erythritol, saccharin, sucralose cyclamate, sugar alcohol, aspartame, fructooligosaccharides, fructose, hydrogenated starch hydrolysates, isomalt, lactitol, maltitiol, mannitol, sorbitol, sugar, sugar cane extract, fruit juice, succanat, honey, rice juice, Yinnie juice, licorice extract, D-tagatose, tagatose, glucose, corn juice, Japanese maple juice, agave fruit juice, and/or any combinations thereof.
  • FIG.l represents a graphical depiction of alpha, beta and gamma cyclodextrin molecules utilized in a preferred embodiment of the present invention
  • FIG. 2 represents a graphical representation of how the cavity of a cyclodextrin molecule can be utilized to capture an oleuropein molecule
  • FIG. 3 is a graphical representation of how alpha, beta and gamma cyclodextrin molecules reduce the bitterness of oleuropein. DESCRIPTION OF THE PREFERRED EMBODIMENTS
  • Oleuropein has the following structural composition:
  • O-CeH -i 1 05 is O- ⁇ - D -g Euco py ranose
  • oleuropein molecules obtained from olive leaf extract are captured within dextrin molecules to alleviate the bitter taste thereof. While the oleuropein molecules come from olive leaf extract in a preferred embodiment, it is understood that the invention may also use oleuropein obtained from any suitable source, such as oleuropein derived from the olive fruit, bark of an olive tree, or from any other plant source containing suitable oleuropein.
  • Dextrin is a generic term for all intermediate products, consisting of monosaccharides, formed during the hydrolysis of starch into sugars. Generally, most monosaccharides form cyclic structures, that predominate in aqueous solution, by forming hemiacetals or hemiketals (depending on whether they are aldoses or ketoses) between an alcohol (-OH) and the carbonyl group (-CHO) of the same sugar molecule.
  • cyclodextrin is an intermediate monosaccharide cyclic structure form of dextrin and used as a capturer of oleuropein.
  • Cyclodextrins (sometimes called cycloamyloses) make up a family of cyclic oligosaccharides, composed of five or more ⁇ -D-glucopyranoside units linked together. More specifically, typical cyclodextrins contain a number of glucose monomers ranging from six to eight units in a ring, creating a cone shape. Cyclodextrins are produced from starch by means of enzymatic conversion. Over the last few years, they have found a wide range of applications in food, pharmaceutical and chemical industries as well as agriculture and environmental engineering.
  • cyclodextrins typically contain a number of glucose monomers - ranging from six to eight units - which are formed in a ring. Among these are included: alpha-cyclodextrin ( ⁇ -cyclodextrin) for a six sugar ring molecule, beta- cyclodextrin ( ⁇ -cyclodextrin) for a seven sugar ring molecule, and gamma-cyclodextrin ( ⁇ -cyclodextrin) for an eight sugar ring molecule. Cyclodextrins are able to form host- guest complexes with hydrophobic molecules given the unique nature imparted by their structure.
  • each cyclodextrin contains one or more water molecules that are produced as a result of the linkage between glucose monomers. These water molecules are easily displaced. Accordingly, when heated, cyclodextrins have a molecular formation that thereby forms a "cavity" suitable for capturing and bonding with the oleuropein molecule resident in an olive leaf extract or other sources.
  • cyclodextrins when heated, cyclodextrins form a hydrophobic cavity therein that allow cyclodextrins to form inclusion complexes with a variety of substrate molecules, that is, non-polar suitably-sized aliphatic and aromatic compounds, such as aroma compounds and lipophilic drugs.
  • cyclodextrin is heated to form this cavity and then combined with oleuropein in order to surround the oleuropein and mask or reduce its bitter taste.
  • alpha ( ⁇ ), beta ( ⁇ ) and gamma ( ⁇ ) cyclodextrin molecular formations are illustrated with detailed dimensions given thereto.
  • these cyclodextrins are represented by a doughnut-shaped (truncated cone) molecule having a central cavity. Because ⁇ -, ⁇ -, and ⁇ -cyclodextrin can form cavities of different sizes when heated, it is desirable to use a cyclodextrin that is capable of forming a cavity corresponding to the size of the oleuropein molecule to be captured therein. More specifically, it is desirable to use a cyclodextrin that when heated will form a large enough cavity to accommodate an oleuropein molecule (molecular size: 5.92 ⁇ 0.49* 10-6 cm/s, P ⁇ 0.001).
  • beta- cyclodextrin ( ⁇ -cyclodextrin) is the most desirable because when heated it will form a cavity suitable for accommodating an oleuropein molecule therein without leaving too much unused space.
  • the combination is desirable when developing a nutritional supplement that is to be orally administered. More specifically, the combination of ⁇ -cyclodextrin and oleuropein allows for formation of a cohesive compound wherein the oleuropein can remain captured within the cavity of the ⁇ -cyclodextrin until the combination passes over the gustatory buds, so that the bitter taste normally associated with oleuropein is reduced and/or masked.
  • FIG. 3 is a graphical representation of how alpha, beta and gamma cyclodextrin molecules reduce the bitterness of oleuropein.
  • ⁇ -cyclodextrin having a cavity suitably sized to accommodate oleuropein, can suppress the bitterness of oleuropein to the greatest extent. Therefore, the present invention features capturing oleuropein molecules within the cavities of cyclodextrins in order to reduce and/or mask the bitterness of oleuropein for easy ingestion.
  • cyclodextrins cannot completely surround them and envelope them. More specifically, as can be seen from the illustration shown in FIG.
  • oleuropein while contained within the cyclodextrin, remains at least partially exposed. Accordingly, it is desirable to further combine an additional agent for forming one complete and cyclical encapsulation of the oleuropein molecule. Therefore, there is a need for an additional, intermediary ingredient that can ensure the longevity of the bonding and joining of oleuropein molecules with cyclodextrin molecules and can complete the encapsulation of the oleuropein molecule so that it is not exposed, thereby more completely reducing and/or masking the bitter taste.
  • oleuropein is stably fixed to and contained within cyclodextrin by preferably using an additional ingredient such as glycerin.
  • Glycerin is a neutral, sweet-tasting, colorless liquid that freezes to a gummy paste and that has a high boiling point. Glycerin can be dissolved into water or alcohol, but not oils. On the other hand, many things will dissolve into glycerin easier than they do into water or alcohol. So glycerin is a good solvent. Glycerin is also highly “hygroscopic" which means that it absorbs water from the air. Glycerin has the following chemical formula.
  • Glycerin is joined with fatty acids by dehydration synthesis to form glycerin ester, represented by the following chemical formula.
  • glycerin ester is amphiphilic, having both a hydrophilic moiety and a lipophilic moiety. Because of its non-ionic surface active agent qualities glycerin ester is a preferred form of glycerin used to complete the encapsulation process. Because of the inhibitory activity of oleuropein against a broad spectrum of microbes, the present invention finds application in the medicinal and food industries, including, but not limited to, natural antibiotics and mouthwashes. It is further desirable to use the combination of oleuropein, cyclodextrin and glycerin ester in order to form a food product and/or nutritional supplement.
  • the above referenced combination can preferably be further implemented into an ingestible, consumable food product or nutritional supplement designed to inhibit harmful microbes in the mouth as well as in the body to provide the benefits enunciated hereinabove.
  • oleuropein, cyclodextrin and glycerin ester are further combined into a consumable nutritional supplement in the form of an ingestible food product that can be eaten/consumed.
  • this food product is formed as a crystallized candy-like or a powdered substance that can be consumed directly or mixed with a fluid product such as juice or water.
  • this food product may include, but not be limited to, the form of a gum, gel, a hard candy, a soft candy, or a liquid.
  • the oleuropein used in forming this product is preferably in the form of an olive leaf extract (that may include other minerals and nutrients derived from an olive leaf), although other forms and sources of oleuropein are intended to be included herewith.
  • Oleuropein 0.5 - 1,000
  • Milling Mill glycerin ester to a very fine powder and heat to 30°- 40° Celsius, 86°- 104° Fahrenheit and store. Mill olive leaf extract to a fine powder and mix together with the stored glycerin ester.
  • lactic acid esters of monoglyceride citric acid esters of monoglyceride, succinic acid esters of monoglyceride, diacetyltartaric acid esters of monoglyceride, polyglycerol esters of fatty acids, polyglycerol polyricinoleate, propylene glycol esters of fatty acids, sucrose esters of fatty acids, lecithin, or enzyme-treated lecithin may be used in place of the glycerin ester as an alternative thereto.
  • Blending/Sifting Combine heated glycerin with the milled olive leaf extract. Since the ratio between the glycerin ester and the olive leaf extract varies with the oleuropein content of the olive leaf extract, in the present invention, an olive leaf extract with a preferred oleuropein content of 20-25% by weight is used, with a weight ratio of 3:7 olive leaf extract to glycerin ester is preferred.
  • the combined mixture (the glycerin ester and olive leaf extract) is then blended - preferably using a commercial machine for 1 hour and 45 minutes - and then passed through a sieve until the mixture is homogeneous.
  • cyclodextrin preferably beta-cyclodextrin
  • Add cyclodextrin preferably beta-cyclodextrin
  • Heat the mixture to 40°-60° Celsius, 104°- 140° Fahrenheit for 2 hours and then allow the mixture to cool down to 20°Celsius/68° Fahrenheit. Then, pass the mixture through a narrow sieve until the total mixture is homogeneous and store.
  • Cyclodextrin which is used to mask or reduce the bitterness, may be exemplified by cellulosine, ⁇ -, ⁇ -, and ⁇ - cyclodextrin, ⁇ -hydroxypropyl cyclodextrin, and/or trapposal (trade name) cyclodextrin.
  • phosphoric acid boric acid, sodium dodecyl sulfate (SDS), octane, butan-1-ol, chromic acid, tetradecyltrimethylammonium bromide (TTAB), ethanesulfonic acid (MES), formic acid, imidazole, Methanol (MeOH), acetone (ACN), sodium acetate, acetic acid, or ammonium acetate may be used instead of cyclodextrin.
  • SDS sodium dodecyl sulfate
  • TTAB tetradecyltrimethylammonium bromide
  • MES ethanesulfonic acid
  • formic acid imidazole
  • Methanol Methanol
  • ACN acetone
  • sodium acetate acetic acid
  • ammonium acetate may be used instead of cyclodextrin.
  • Cyclodextrins are able to form host-guest complexes with hydrophobic molecules given the unique nature imparted by their structure. As a result these molecules have found a number of applications in a wide range of fields. In the food industry cyclodextrins are employed for the preparation of cholesterol free products. Other food applications further include the ability to stabilize volatile or unstable compounds and reduction of unwanted tastes. The ability of cyclodextrins to form complexes with hydrophobic molecules has led to their usage in supramolecular chemistryfg.h) and is part of this process.
  • Cyclodextrins can be represented as toroids and because of this arrangement cyclodextrins can host inclusion compounds/complexes that greatly modify the physical and chemical properties of the guest molecule, mostly in terms of water solubility. This is one reason why cyclodextrins have attracted much interest in many fields, especially pharmaceutical application.
  • the present embodiment presents a further development of potentiation in that when mixed with glycerin esters, encapsulates the potentially exposed end of the cyclodextrin thus disallowing the host complex to be realized at gustation.
  • the processed enclosure is referred to as cyclo-encapsulation.
  • xylitol used in this process has to be prepared prior to mixing with the primary homogeneous mixture by liquefying and adding natural or synthetic colors. Dry the prepared xylitol and coloring to less than 5% moisture. Add the mixture of the prepared xylitol as a sweetener to the stored primary homogeneous mixture, blend thoroughly and pass through a sieve several times.
  • a substitute sweetener may be selected and used from this set of sweeteners including, but not limited to, xylitol, rebaudiosides, erythritol, saccharin, sucralose cyclamate, sugar alcohol, aspartame, fructooligosaccharides, fructose, hydrogenated starch hydrolysates, isomalt, lactitol, maltitiol, mannitol, sorbitol, sugar, sugar cane extract, fruit juice, succanat, honey, rice juice, Yinnie juice, licorice extract, D-tagatose, tagatose, glucose, corn juice, Japanese maple juice, and agave fruit juice.
  • sweeteners including, but not limited to, xylitol, rebaudiosides, erythritol, saccharin, sucralose cyclamate, sugar alcohol, aspartame, fructooligosaccharides, fructose, hydrogenated starch hydrolysates,
  • citric acid (synthesized or naturally obtained from fruit, peanuts, vegetables or animals) may be used in place of the sweetener.
  • the following sour taste substitutes may include, but not be limited to, citric acid, tartaric acid, malic acid, lactic acid, acetic acid, and/or succinic acid, monoprotic acid, diprotic acid, triprotic acid, and polyprotic acid.
  • citric acid synthesized or naturally obtained from fruit, peanuts, vegetables or animals This citric acid may be in the form of monohydrate or anhydrate, liquid, powder or lyophilized.
  • the preceding method of manufacture should take place in a moisture controlled facility that has the ability to restrict moisture to less than 5%, store materials at less than 1%, and maintain a 20° Celsius (68° Fahrenheit) temperature. Moreover, the method of manufacture provided should follow Over The Counter (OTC) and or Certified Good Manufacturing Practice (cGMP) requirements intended for commercial distribution currently in the United States, Australia, Asia and Europe.
  • OTC Over The Counter
  • cGMP Certified Good Manufacturing Practice
  • the present invention provides a processed olive leaf extract that has oleuropein molecules captured within cyclodextrin, thus being greatly reduced in bitterness.
  • the processed olive leaf extract of the present invention is easy to consume. The reduced and/or masked bitterness allows the oleuropein to find various applications.

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Abstract

A nutritional supplement in which oleuropein molecules are captured within cyclodextrin with the aid of glycerin ester.

Description

NUTRITIONAL SUPPLEMENT WITH OLEUROPEIN Inventors: Richard Hall & James Kwon
FIELD OF THE INVENTION
The present invention relates, in general, to a nutritional supplement and, more particularly, to a nutritional supplement preferably comprised of a processed olive leaf extract in which oleuropein is captured within cyclodextrin with the aid of glycerin ester, thereby greatly suppressing the bitterness of oleuropein.
BACKGROUND OF THE INVENTION
Oleuropein is a naturally occurring compound commonly found in the leaves, fruit, bark and roots of olive trees. The leaves of an olive tree are richer in oleuropein than the fruit and/or bark of the tree. Oleuropein can also be obtained from other plants belonging to the family Oleaceae; for example, ash and privet trees.
The functional and therapeutic effects of oleuropein have been explored for years. More specifically, oleuropein is known to have a resistant activity against a wide spectrum of bacteria, viruses, and fungi, producing various therapeutic effects, such as alleviation of hypertension, antioxidation, control of glucose levels in blood, etc. When consumed, oleuropein hydrolyzes to d-elenolic acid and 1-elenolic acid that has powerful antimicrobial effects. In particular, the d-elenolic acid acts as a strong antimicrobial. Moreover, oleuropein has the ability to interfere with critical amino acid processes that are essential for virus replication, thus inhibiting the spread of such viruses. Additionally, in the case of retroviruses, it is able to neutralize the production of reverse transcriptase and protease, which are enzymes essential in order for a retrovirus to alter the RNA of a healthy cell. This action may lead to the prevention of various cancers. Further, oleuropein can also directly stimulate the formation of immune system cells through a process called phagocytosis in order to combat various types of microbes. Therefore, oleuropein acts as a potent natural antibiotic against viruses and bacteria.
Additionally, oleuropein has been the source of great interest concerning coronary dilation (the expansion/contraction ability of the arteries). Tests have shown that the injection of oleuropein in animals has improved blood circulation by up to 50%. In addition to the coronary artery dilating effect, oleuropein has been found to display other cardiovascular health-enhancing effects, such as inducing a long-lasting effect of lowering elevated blood pressure (as proven using animals when arrhythmia was induced through the injection of chlorides or calcium compounds). More specifically, a study, conducted by the Pharmacology University of Granada, Spain, disclosed that oleuropeoside, an ingredient isolated from oleuropein, dilates the blood vessels. That is, oleuropeoside acts to relax and smoothen contracted blood vessels to improve blood circulation and thus to decrease blood pressure.
Oleuropein is also believed to directly promote insulin release in the human body and maximize the use of blood sugar. At least one study has found that oleuropein may play a pivotal role in glycemic control. The recordings of this lab disclosed that when 16 mg of oleuropeoside was administered to animals when diabetes had been induced by alloxan injection, blood sugar levels were significantly reduced in these injected animals. Oleuropein was observed to have a proposed mechanism of action, of potentiation, of glucose-induced insulin release, and an increase in peripheral blood glucose uptake.
As explained earlier, olive leaves have a very high concentration of oleuropein. Recently, olive leaf extract containing oleuropein has been formulated into capsules and tablets for oral administration. Unfortunately, the bitter taste of oleuropein makes it difficult for people to ingest. Thus, it is desirable to be able to reduce and/or mask the bitter taste characteristic of oleuropein so that it can be orally administered and swallowed. Conventional methods for removing or alleviating the bitter taste of oleuropein have included 1) pickling the extract with salt and acetic acid, 2) fermentation, and 3) heating to change the properties of oleuropein. However, these efforts have proven unsuccessful for a myriad of reasons. Pickling the extract with salt and acetic acid may break down oleuropein decreasing its nutritional value. Fermenting the extract is a complicated process and is of limited use in producing beverage and processed foods. The thermal method of heating oleuropein to reduce the bitterness also reduces the preventive and therapeutic effects oleuropein has on maladies.
Accordingly, what is needed is a nutritional supplement having oleuropein, and a method for creating the same, that does not include the bitter taste commonly associated with oleuropein. Preferably, the oleuropein used in this nutritional supplement will be in the form of an extract from olive leaves having a high concentration thereof; however, it is understood that the oleuropein to be included may be obtained from other sources as well.
SUMMARY OF THE INVENTION
It is one object of the present invention to provide a nutritional supplement having oleuropein that is free of, or alleviated of, bitterness to the extent that it is suitable to consume. It is another object of the present invention to provide a processed olive leaf extract that contains intact oleuropein having its bitter taste greatly reduced, without destroying the nutritional benefits that are characteristic thereof. It is a further object of the present invention to provide a method of processing oleuropein to create a product that is suitable for oral administration. Finally, it is an object of the present invention to provide oleuropein in the form of a processed olive leaf extract as a food or a nutritional supplement (health aid). In accordance with each of these aspects, the invention is directed toward a processed olive leaf extract, including oleuropein, cyclodextrin and glycerin ester as its essential ingredients. In a preferred embodiment, the processed olive leaf extract contains oleuropein in an amount from 0.5 to 1,000 mg, cyclodextrin in an amount from 0.5 to 100 mg, and glycerin ester in an amount from 2 to 1 ,500 mg.
Furthermore, the processed olive leave extract is preferably embodied in the form of a powdered or candied nutritional supplement that further includes additional ingredients selected from a group consisting of xylitol, an acid, calcium, a sweetener, and/or any combination thereof.
In a preferred embodiment where the processed olive leaf extract is embodied in the form of a powdered or candied nutritional supplement that further includes a sweetener, the sweetener is preferably selected from the group consisting of, but not limited to, xylitol, rebaudiosides , erythritol, saccharin, sucralose cyclamate, sugar alcohol, aspartame, fructooligosaccharides, fructose, hydrogenated starch hydrolysates, isomalt, lactitol, maltitiol, mannitol, sorbitol, sugar, sugar cane extract, fruit juice, succanat, honey, rice juice, Yinnie juice, licorice extract, D-tagatose, tagatose, glucose, corn juice, Japanese maple juice, agave fruit juice, and/or any combinations thereof.
DETAILED DESCRIPTIONS OF THE DIAGRAMS
FIG.l represents a graphical depiction of alpha, beta and gamma cyclodextrin molecules utilized in a preferred embodiment of the present invention;
FIG. 2 represents a graphical representation of how the cavity of a cyclodextrin molecule can be utilized to capture an oleuropein molecule; and
FIG. 3 is a graphical representation of how alpha, beta and gamma cyclodextrin molecules reduce the bitterness of oleuropein. DESCRIPTION OF THE PREFERRED EMBODIMENTS
A key component to olive leaf extract is a bitter glucoside (phenolic) compound called oleuropein. Oleuropein has the following structural composition:
Figure imgf000006_0001
O-CeH -i 105 is O- β- D -g Euco py ranose
Structure of Oleuropein
As explained earlier herein, the health, medical and nutritional benefits of oleuropein continue to be expanded/discovered. Accordingly, ingestion of oleuropein as a nutritional supplemental is highly beneficial. Unfortunately, because of its very bitter taste, oleuropein is very difficult to consume directly. Accordingly, there exists a need for somehow reducing and/or masking the bitter taste associated with oleuropein.
In a preferred embodiment of the present invention, oleuropein molecules obtained from olive leaf extract are captured within dextrin molecules to alleviate the bitter taste thereof. While the oleuropein molecules come from olive leaf extract in a preferred embodiment, it is understood that the invention may also use oleuropein obtained from any suitable source, such as oleuropein derived from the olive fruit, bark of an olive tree, or from any other plant source containing suitable oleuropein.
Dextrin is a generic term for all intermediate products, consisting of monosaccharides, formed during the hydrolysis of starch into sugars. Generally, most monosaccharides form cyclic structures, that predominate in aqueous solution, by forming hemiacetals or hemiketals (depending on whether they are aldoses or ketoses) between an alcohol (-OH) and the carbonyl group (-CHO) of the same sugar molecule.
In accordance with a preferred embodiment of the present invention, cyclodextrin is an intermediate monosaccharide cyclic structure form of dextrin and used as a capturer of oleuropein. Cyclodextrins (sometimes called cycloamyloses) make up a family of cyclic oligosaccharides, composed of five or more α-D-glucopyranoside units linked together. More specifically, typical cyclodextrins contain a number of glucose monomers ranging from six to eight units in a ring, creating a cone shape. Cyclodextrins are produced from starch by means of enzymatic conversion. Over the last few years, they have found a wide range of applications in food, pharmaceutical and chemical industries as well as agriculture and environmental engineering.
As explained hereinabove, cyclodextrins typically contain a number of glucose monomers - ranging from six to eight units - which are formed in a ring. Among these are included: alpha-cyclodextrin (α-cyclodextrin) for a six sugar ring molecule, beta- cyclodextrin (β-cyclodextrin) for a seven sugar ring molecule, and gamma-cyclodextrin (γ-cyclodextrin) for an eight sugar ring molecule. Cyclodextrins are able to form host- guest complexes with hydrophobic molecules given the unique nature imparted by their structure. As a result, these molecules have found a number of applications in a wide range of fields. More specifically, each cyclodextrin contains one or more water molecules that are produced as a result of the linkage between glucose monomers. These water molecules are easily displaced. Accordingly, when heated, cyclodextrins have a molecular formation that thereby forms a "cavity" suitable for capturing and bonding with the oleuropein molecule resident in an olive leaf extract or other sources. More specifically, when heated, cyclodextrins form a hydrophobic cavity therein that allow cyclodextrins to form inclusion complexes with a variety of substrate molecules, that is, non-polar suitably-sized aliphatic and aromatic compounds, such as aroma compounds and lipophilic drugs. In a preferred embodiment, cyclodextrin is heated to form this cavity and then combined with oleuropein in order to surround the oleuropein and mask or reduce its bitter taste. Referring to FIG. 1, alpha (α), beta (β) and gamma (γ) cyclodextrin molecular formations are illustrated with detailed dimensions given thereto. As seen in the figures, when heated these cyclodextrins are represented by a doughnut-shaped (truncated cone) molecule having a central cavity. Because α-, β-, and γ-cyclodextrin can form cavities of different sizes when heated, it is desirable to use a cyclodextrin that is capable of forming a cavity corresponding to the size of the oleuropein molecule to be captured therein. More specifically, it is desirable to use a cyclodextrin that when heated will form a large enough cavity to accommodate an oleuropein molecule (molecular size: 5.92±0.49* 10-6 cm/s, P < 0.001). Of the three different cyclodextrins previously discussed, beta- cyclodextrin (β-cyclodextrin) is the most desirable because when heated it will form a cavity suitable for accommodating an oleuropein molecule therein without leaving too much unused space.
Referring to FlG. 2, illustrated here is the concept of capturing and retaining an oleuropein molecule within a β-cyclodextrin molecule. As shown in FIG. 2, because the cavity of a β-cyclodextrin molecule is of sufficient size to surround the oleuropein molecule, the combination is desirable when developing a nutritional supplement that is to be orally administered. More specifically, the combination of β-cyclodextrin and oleuropein allows for formation of a cohesive compound wherein the oleuropein can remain captured within the cavity of the β-cyclodextrin until the combination passes over the gustatory buds, so that the bitter taste normally associated with oleuropein is reduced and/or masked.
FIG. 3 is a graphical representation of how alpha, beta and gamma cyclodextrin molecules reduce the bitterness of oleuropein. As seen in the graph, β-cyclodextrin, having a cavity suitably sized to accommodate oleuropein, can suppress the bitterness of oleuropein to the greatest extent. Therefore, the present invention features capturing oleuropein molecules within the cavities of cyclodextrins in order to reduce and/or mask the bitterness of oleuropein for easy ingestion. Although able to capture oleuropein molecules, cyclodextrins cannot completely surround them and envelope them. More specifically, as can be seen from the illustration shown in FIG. 2, the oleuropein, while contained within the cyclodextrin, remains at least partially exposed. Accordingly, it is desirable to further combine an additional agent for forming one complete and cyclical encapsulation of the oleuropein molecule. Therefore, there is a need for an additional, intermediary ingredient that can ensure the longevity of the bonding and joining of oleuropein molecules with cyclodextrin molecules and can complete the encapsulation of the oleuropein molecule so that it is not exposed, thereby more completely reducing and/or masking the bitter taste. In a preferred embodiment, oleuropein is stably fixed to and contained within cyclodextrin by preferably using an additional ingredient such as glycerin.
Glycerin is a neutral, sweet-tasting, colorless liquid that freezes to a gummy paste and that has a high boiling point. Glycerin can be dissolved into water or alcohol, but not oils. On the other hand, many things will dissolve into glycerin easier than they do into water or alcohol. So glycerin is a good solvent. Glycerin is also highly "hygroscopic" which means that it absorbs water from the air. Glycerin has the following chemical formula.
OH-CH2-CH(OH)-CH2-OH
Glycerin is joined with fatty acids by dehydration synthesis to form glycerin ester, represented by the following chemical formula.
R-COO-CH2-CH(OH)-CH2-OH
As understood from the chemical formula, glycerin ester is amphiphilic, having both a hydrophilic moiety and a lipophilic moiety. Because of its non-ionic surface active agent qualities glycerin ester is a preferred form of glycerin used to complete the encapsulation process. Because of the inhibitory activity of oleuropein against a broad spectrum of microbes, the present invention finds application in the medicinal and food industries, including, but not limited to, natural antibiotics and mouthwashes. It is further desirable to use the combination of oleuropein, cyclodextrin and glycerin ester in order to form a food product and/or nutritional supplement. Having masked or reduced the bitterness of the oleuropein, the above referenced combination can preferably be further implemented into an ingestible, consumable food product or nutritional supplement designed to inhibit harmful microbes in the mouth as well as in the body to provide the benefits enunciated hereinabove.
In a further aspect of the present invention, oleuropein, cyclodextrin and glycerin ester are further combined into a consumable nutritional supplement in the form of an ingestible food product that can be eaten/consumed. In a preferred embodiment this food product is formed as a crystallized candy-like or a powdered substance that can be consumed directly or mixed with a fluid product such as juice or water. Alternatively, this food product may include, but not be limited to, the form of a gum, gel, a hard candy, a soft candy, or a liquid. Once again, it is understood that the oleuropein used in forming this product is preferably in the form of an olive leaf extract (that may include other minerals and nutrients derived from an olive leaf), although other forms and sources of oleuropein are intended to be included herewith.
The following table sets forth the different ingredients and the various amounts/weights thereof that are preferably used in forming a food product in accordance with a preferred embodiment of the present invention:
Composition Weight (mg)
Xylitol 0.5 - 1,500
Oleuropein 0.5 - 1,000
Citric Acid 0.5 - 1,500
Calcium 2 - 3,500
Cyclodextrin 0.5 - 100 Glycerin Esters 2 - 1 ,500
Sucralose 0.5 - 1,500
Colors 0.5 - 50
Flavors 0.2 - 3,000
The process of forming this food product/nutritional supplement is as follows:
(1) Milling: Mill glycerin ester to a very fine powder and heat to 30°- 40° Celsius, 86°- 104° Fahrenheit and store. Mill olive leaf extract to a fine powder and mix together with the stored glycerin ester.
Alternatively, lactic acid esters of monoglyceride, citric acid esters of monoglyceride, succinic acid esters of monoglyceride, diacetyltartaric acid esters of monoglyceride, polyglycerol esters of fatty acids, polyglycerol polyricinoleate, propylene glycol esters of fatty acids, sucrose esters of fatty acids, lecithin, or enzyme-treated lecithin may be used in place of the glycerin ester as an alternative thereto.
(2) Blending/Sifting: Combine heated glycerin with the milled olive leaf extract. Since the ratio between the glycerin ester and the olive leaf extract varies with the oleuropein content of the olive leaf extract, in the present invention, an olive leaf extract with a preferred oleuropein content of 20-25% by weight is used, with a weight ratio of 3:7 olive leaf extract to glycerin ester is preferred.
The combined mixture (the glycerin ester and olive leaf extract) is then blended - preferably using a commercial machine for 1 hour and 45 minutes - and then passed through a sieve until the mixture is homogeneous.
(3) Adding cyclodextrin: Add cyclodextrin (preferably beta-cyclodextrin) to the homogeneous mixture of olive leaf extract and glycerin ester with 15-20 weight % of cyclodextrin and blend continuously. Heat the mixture to 40°-60° Celsius, 104°- 140° Fahrenheit for 2 hours and then allow the mixture to cool down to 20°Celsius/68° Fahrenheit. Then, pass the mixture through a narrow sieve until the total mixture is homogeneous and store.
Cyclodextrin, which is used to mask or reduce the bitterness, may be exemplified by cellulosine, α-, β-, and γ- cyclodextrin, β-hydroxypropyl cyclodextrin, and/or trapposal (trade name) cyclodextrin. Alternatively, phosphoric acid, boric acid, sodium dodecyl sulfate (SDS), octane, butan-1-ol, chromic acid, tetradecyltrimethylammonium bromide (TTAB), ethanesulfonic acid (MES), formic acid, imidazole, Methanol (MeOH), acetone (ACN), sodium acetate, acetic acid, or ammonium acetate may be used instead of cyclodextrin.
Cyclodextrins are able to form host-guest complexes with hydrophobic molecules given the unique nature imparted by their structure. As a result these molecules have found a number of applications in a wide range of fields. In the food industry cyclodextrins are employed for the preparation of cholesterol free products. Other food applications further include the ability to stabilize volatile or unstable compounds and reduction of unwanted tastes. The ability of cyclodextrins to form complexes with hydrophobic molecules has led to their usage in supramolecular chemistryfg.h) and is part of this process.
Cyclodextrins can be represented as toroids and because of this arrangement cyclodextrins can host inclusion compounds/complexes that greatly modify the physical and chemical properties of the guest molecule, mostly in terms of water solubility. This is one reason why cyclodextrins have attracted much interest in many fields, especially pharmaceutical application. The present embodiment presents a further development of potentiation in that when mixed with glycerin esters, encapsulates the potentially exposed end of the cyclodextrin thus disallowing the host complex to be realized at gustation. The processed enclosure is referred to as cyclo-encapsulation.
(4) The inclusion of xylitol: The Xylitol used in this process has to be prepared prior to mixing with the primary homogeneous mixture by liquefying and adding natural or synthetic colors. Dry the prepared xylitol and coloring to less than 5% moisture. Add the mixture of the prepared xylitol as a sweetener to the stored primary homogeneous mixture, blend thoroughly and pass through a sieve several times.
Alternatively a substitute sweetener may be selected and used from this set of sweeteners including, but not limited to, xylitol, rebaudiosides, erythritol, saccharin, sucralose cyclamate, sugar alcohol, aspartame, fructooligosaccharides, fructose, hydrogenated starch hydrolysates, isomalt, lactitol, maltitiol, mannitol, sorbitol, sugar, sugar cane extract, fruit juice, succanat, honey, rice juice, Yinnie juice, licorice extract, D-tagatose, tagatose, glucose, corn juice, Japanese maple juice, and agave fruit juice.
As an alternative, if a sour tasting (instead of sweet tasting) nutritional supplement or food product is desired, then citric acid (synthesized or naturally obtained from fruit, peanuts, vegetables or animals) may be used in place of the sweetener. Likewise, the following sour taste substitutes may include, but not be limited to, citric acid, tartaric acid, malic acid, lactic acid, acetic acid, and/or succinic acid, monoprotic acid, diprotic acid, triprotic acid, and polyprotic acid.
(5) Addition of other support ingredients: Add the following ingredients together (as needed and/or desired) before mixing with the primary homogeneous mixture ;
(a) calcium obtained from fruit, peanuts, vegetables, shell fish, or animals;
(b) citric acid synthesized or naturally obtained from fruit, peanuts, vegetables or animals. This citric acid may be in the form of monohydrate or anhydrate, liquid, powder or lyophilized.
(c) any flavoring natural or synthetic that is used in the pharmaceutical or food industry;
(d) any coloring natural or synthetic that is used in the pharmaceutical or food industry. (6) Blend the entire mixture thoroughly and pass through a sieve several times. Once well blended, place in a container and preferably store at 20° Celsius (68° Fahrenheit).
The preceding method of manufacture should take place in a moisture controlled facility that has the ability to restrict moisture to less than 5%, store materials at less than 1%, and maintain a 20° Celsius (68° Fahrenheit) temperature. Moreover, the method of manufacture provided should follow Over The Counter (OTC) and or Certified Good Manufacturing Practice (cGMP) requirements intended for commercial distribution currently in the United States, Australia, Asia and Europe.
All of the above-mentioned ingredients may be used to prepare a nutritional supplement/food product having oleuropein in the form of an olive leaf extract in accordance with a preferred embodiment of the present invention. As described hitherto, the present invention provides a processed olive leaf extract that has oleuropein molecules captured within cyclodextrin, thus being greatly reduced in bitterness. Hence, the processed olive leaf extract of the present invention is easy to consume. The reduced and/or masked bitterness allows the oleuropein to find various applications.
Although a preferred embodiment of the present invention has been described for illustrative purposes, those skilled in the art will appreciate that various modifications, additions and substitutions are possible, without departing from the spirit and scope of the invention and the invention is not intended to be limited to any one particular embodiment except as claimed.

Claims

CLAIMSWhat is claimed is:
1. A nutritional supplement comprised of: oleuropein; and cyclodextrin.
2. The nutritional supplement of claim 1, further comprised of a glycerin.
3. The nutritional supplement of claim 1, wherein the oleuropein is in the form of a processed olive leaf extract.
4. The nutritional supplement of claim 3, wherein the olive leaf extract contains oleuropein in an amount from 0.5 to 1 ,000 mg.
5. The nutritional supplement of claim 4, wherein the nutritional supplement contains cyclodextrin in an amount from 0.5 to 100 mg and glycerin ester in an amount from 2 to 1,500 mg.
6. The nutritional supplement of claim 3, further comprising: an additional ingredient selected from the group consisting of xylitol, a sweetener, calcium, and/or an acid.
7. The nutritional supplement of claim 6, wherein the sweetener may be any sweetener selected from the group consisting of xylitol, rebaudiosides, erythritol, saccharin, sucralose cyclamate, sugar alcohol, aspartame, fructooligosaccharides, fructose, hydrogenated starch hydrolysates, isomalt, lactitol, maltitiol, mannitol, sorbitol, sugar, sugar cane extract, fruit juice, succanat, honey, rice juice, Yinnie juice, licorice extract, D-tagatose, tagatose, glucose, corn juice, Japanese maple juice, and agave fruit juice.
8. The nutritional supplement of claim 6, wherein the acid may be any acid selected from the group consisting of citric acid, tartaric acid, malic acid, lactic acid, acetic acid, succinic acid, monoprotic acid, diprotic acid, triprotic acid, and polyprotic acid.
9. The nutritional supplement of claim 6, wherein the supplement is in the form of a gum, a gel, a hard candy, a soft candy or a liquid.
10. A processed olive leaf extract comprised of
oleuropein; cyclodextrin; and glycerin.
1 1. The processed olive leaf extract of claim 10, wherein the processed olive leaf extract contains: oleuropein in an amount from 0.5 to 1 ,000 mg; cyclodextrin in an amount from 0.5 to 100 mg; and glycerin in an amount from 2 to 1,500 mg.
12. The processed olive leaf extract of claim 10, further comprising: an additional ingredient selected from the group consisting of a sweetener.
13. The processed olive leaf extract of claim 12, wherein the sweetener may be any sweetener selected from the group consisting of xylitol, rebaudiosides, erythritol, saccharin, sucralose cyclamate, sugar alcohol, aspartame, fructooligosaccharides, fructose, hydrogenated starch hydrolysates, isomalt, lactitol, maltitiol, mannitol, sorbitol, sugar, sugar cane extract, fruit juice, succanat, honey, rice juice, Yinnie juice, licorice extract, D-tagatose, tagatose, glucose, corn juice, Japanese maple juice, and agave fruit juice.
14. The processed olive leaf extract of claim 10, further comprising: an additional ingredient selected from the group consisting of an acid.
15. The processed olive leaf extract of claim 14, wherein the acid may be any acid selected from the group consisting of citric acid, tartaric acid, malic acid, lactic acid, acetic acid, succinic acid, monoprotic acid, diprotic acid, triprotic acid, and polyprotic acid.
16. A method for manufacturing a nutritional supplement comprised of:
combining glycerin with an olive leaf extract having oleuropein; and
adding cyclodextrin to the mixture.
17. The method of claim 16, wherein the glycerin is glycerin ester.
18. The method of claim 16 further comprising the additional step of adding a sweetener to the mixture in order to make the nutritional supplement taste sweet.
19. The method of claim 18, wherein the sweetener may be any sweetener selected from the group consisting of xylitol, rebaudiosides, erythritol, saccharin, sucralose cyclamate, sugar alcohol, aspartame, fructooligosaccharides, fructose, hydrogenated starch hydrolysates, isomalt, lactitol, maltitiol, mannitol, sorbitol, sugar, sugar cane extract, fruit juice, succanat, honey, rice juice, Yinnie juice, licorice extract, D-tagatose, tagatose, glucose, corn juice, Japanese maple juice, and agave fruit juice.
20. The method of claim 16 further comprising the additional step of adding an acid to the mixture in order to make the nutritional supplement taste sour.
21. The method of claim 20, wherein the acid may be any acid selected from the group consisting of citric acid, tartaric acid, malic acid, lactic acid, acetic acid, succinic acid, monoprotic acid, diprotic acid, triprotic acid, and polyprotic acid.
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JP2016132657A (en) * 2015-01-22 2016-07-25 国立研究開発法人産業技術総合研究所 Composition for reducing activity and / or anxiety
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WO2021053259A1 (en) * 2019-09-20 2021-03-25 Universidad De Sevilla Method for extracting, from olive leaf, oleuropein in a phenolic fraction; functional food composition; and use
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