Uses of curcuminoid compositions

Abstract

To provide a novel curcuminoid-containing composition for improving insufficient bioavailability of curcuminoids resulting from inadequate water solubility and absorption characteristics.SOLUTION: The present invention provides a composition comprising (i) curcuminoids, (ii) quillaja, and (iii) gum arabic, for use in improving bioavailability of curcumin for health conditions selected from joint health, inflammation, arthritis, atherosclerosis, liver steatosis, liver fibrosis, diabetes, cognition, mild cognitive impairment, and irritable bowel syndrome.SELECTED DRAWING: None

Description

[Technical field]

[0001] Technical Field The present invention relates to methods and uses of compositions comprising curcumin; in particular to methods and uses relating to improving the bioaccessibility, bioavailability, bioefficacy and / or bioactivity of curcuminoids in mammals. [Background technology]

[0002] Background technology The listing or discussion of an apparently prior-published document in this specification should not necessarily be taken as an acknowledgement that the document is part of the state of the art or is general knowledge.

[0003] Turmeric and compounds isolated from turmeric, such as curcuminoids, including curcumin, have long been used to treat a variety of diseases and conditions.

[0004] Curcuminoids are natural yellow-orange pigments and hydrophobic polyphenols derived from the rhizomes of the herb Curcuma longa. They are commonly isolated from the spice and food colorant turmeric.

[0005] Turmeric extract contains approximately 75-80% curcumin, 15-20% demethoxycurcumin (DMC), and 0-10% bisdemethoxycurcumin (BDMC). Curcuminoids have a unique conjugated structure, a bis-α,β-unsaturated β-diketone (commonly called diferuloylmethane), which exhibits keto-enol tautomerism with the keto form predominating in acidic and neutral solutions and the enol form stable in alkaline media (see Figure 1) (Hoehle SI, Pfeiffer E, Solyom AM, Metzler M. Metabolism of curcuminoids in tissue slices and subcellular fractions from rat liver. J Agric Food Chem. 2006 Feb 8;54(3):756-64).

[0006] Curcumin is a highly pleiotropic molecule that was first shown to exhibit antibacterial activity in 1949. Since then, this polyphenol has been shown to possess anti-inflammatory, hypoglycemic, antioxidant, wound healing, and antimicrobial activities. Extensive preclinical and clinical studies over the past three decades have demonstrated the therapeutic potential of curcumin for a wide range of human diseases (Gupta SC, Sung B, Kim JH, Prasad S, Li S, Aggarwal BB. Multitargeting by turmeric, the golden spice: From kitchen to clinic. Mol Nutr Food Res. 2013 Sep;57(9):1510-28).

[0007] Although curcumin has demonstrated efficacy for a number of human disorders, it is also known to have limited bioavailability due to poor absorption, rapid metabolism, and rapid systemic elimination.

[0008] Low serum levels have been reported when curcumin is administered orally, as well as limited tissue distribution and rapid metabolism. For example, it has been demonstrated that serum levels of curcumin did not exceed low micromolar levels even after oral administration of up to 12 g. In one clinical study with 4-8 g of curcumin, the maximum serum concentration observed was 1.3 μg / mL, while several other clinical and animal studies reported serum levels in the lower nanograms per milliliter range (Anand P, Kunnumakkara AB, Newman RA, Aggarwal BB. Bioavailability of curcumin: problems and promises. Mol Pharm. 2007 Nov-Dec;4(6):807-18).

[0009] Curcumin has been found to be poorly soluble in water: for example, the maximum solubility of curcumin in aqueous buffer (pH 5.0) is reported to be as low as 11 ng / ml (Tonessen et al., 2002).

[0010] Curcumin is relatively stable at acidic pH but decomposes rapidly above neutral pH to form ferulic acid and ferulomethane, the latter of which then gives rise to vanillin (FAO 2004. CURCUMIN Chemical and Technical Assessment (CTA) First draft prepared by Ivan Stankovic. Chemical and Technical Assessment 61st JECFA).

[0011] Curcumin also exhibits extremely low intestinal absorption: the apparent permeability coefficient of curcumin was found to be extremely low in an in vitro human intestinal cell model (Papp value: 0.1×10-6 cm / s), which could predict low (0-20%) absorption in humans according to a correlation between Papp values ​​measured in Caco-2 cells in vitro and in vivo human absorption (Dempe JS, Scheerle RK, Pfeiffer E, Metzler M. Metabolism and permeability of curcumin in cultured Caco-2 cells. Mol Nutr Food Res. 2013 Sep;57(9):1543-9).

[0012] Once absorbed, curcumin undergoes both phase I and phase II metabolism (see Figure 2). In phase I metabolism, curcumin and its two demethoxy congeners undergo sequential reduction to their dihydro-, tetrahydro-, hexahydro-, and octahydro-metabolites in the liver and in the intestinal mucosa. In phase II metabolism, both curcumin and its reduced metabolites are conjugated with glucuronic acid and sulfate to form phase II metabolites.

[0013] Reduction and conjugation appear to be common metabolic pathways of curcuminoids occurring in liver and intestinal tissues of rats and humans, and it is therefore believed by some authors that the biological effects induced by curcumin in tissues other than the gastrointestinal tract are likely due to curcumin metabolites.

[0014] Curcumin has also been shown to be metabolized by gut microbes (see Figure 3). The microbial metabolism of curcumin was found to involve a two-step reduction in which curcumin was sequentially converted to dihydrocurcumin and then to tetrahydrocurcumin by NADPH-dependent curcumin / dihydrocurcumin reductase (CurA) (Hassaninasab A, Hashimoto Y, Tomita-Yokotani K, Kobayashi M. Discovery of the curcumin metabolic pathway involving a unique enzyme in an intestinal microorganism. Proc Natl Acad Sci US A. 2011 Apr 19;108(16):6615-20).

[0015] A recent study reported that tetrahydroxycurcumin, but not curcumin, was able to accumulate in rat tissues, suggesting that the microbiota can also be considered as a potential actor in curcumin metabolism and bioavailability (Neyrinck AM, Alligier M, Memvanga PB, Nevraumont E, Larondelle Y, Preat V, Cani PD, Delzenne NM. Curcuma longa extract associated with white pepper lessens high fat diet-induced inflammation in subcutaneous adipose tissue. PLoS One. 2013 Nov 19;8(11):e81252).

[0016] Rapid metabolism, poor water solubility, instability at neutral pH and upon exposure to light and / or oxygen, and poor uptake by tissues significantly limit the potential utility of curcuminoids, including curcumin, in the treatment of conditions such as cancer.

[0017] Due to its poor water solubility and absorption properties, organic solvents, such as methyl sulfoxide (DMSO), are typically used to dissolve curcumin. However, although the use of such solvents helps to solubilize curcumin and improve its availability, the use of organic solvents as vehicles is controversial and undesirable, especially in an era of increasing consumer demand for natural ingredients.

[0018] Another possibility to aid in the water solubility of curcumin is through emulsification. However, natural organic grade emulsifiers are rare, and both synthetic emulsifiers such as polysorbate 80 or natural emulsifiers such as starch-based emulsifiers are typically only able to disperse low levels (<7%) of curcumin in water.

[0019] Other strategies that have been explored to address the above drawbacks and improve the therapeutic efficacy of curcumin include grafting curcumin into polymeric nanoparticles, polymeric micelles and hydrophilic polymers.

[0020] The present invention seeks to address the above-mentioned problems associated with curcuminoids, such as poor solubility of curcumin in water, and to improve the bioaccessibility, bioavailability, bioefficacy and / or bioactivity of curcuminoids in mammals by providing the following methods and uses: Summary of the Invention

[0021] Disclosure of the Invention The inventors have surprisingly found that the bioaccessibility, bioavailability, bioefficacy and / or bioactivity of curcuminoids in mammals can be improved by administration of said curcuminoids in the form of a composition comprising (i) a curcuminoid, (ii) quillaja, and (iii) modified starch and / or gum arabic.

[0022] Methods and Uses The present invention provides a method for improving the bioaccessibility, bioavailability, bioefficacy and / or bioactivity of curcuminoids in a mammal, the method comprising administration of said curcuminoids in the form of a composition comprising (i) a curcuminoid, (ii) an extract obtained or obtainable from Quillaja quilla, and (iii) modified starch and / or gum arabic. In particular, the present invention provides a method for improving the bioaccessibility, bioavailability, bioefficacy and / or bioactivity of curcuminoids in a mammal, the method comprising administration of said curcuminoids in the form of a composition comprising: (i) a curcuminoid; (ii) an extract obtained or obtainable from Quillaja quilla, and (iii) modified starch or gum arabic. Such a method may hereinafter be referred to as the "method of the invention".

[0023] The compositions used in the methods of the invention do not contain fenugreek, for example, some compositions of the invention do not contain fenugreek fiber (ie, fiber obtained or obtainable from fenugreek).

[0024] The compositions used in the methods of the invention may contain small amounts of polyols and / or low molecular weight sugars, preferably having one or two monosaccharide units, such as less than 5% by weight of the composition or less than 2.5% by weight of the composition. Alternatively, the compositions of the invention may be free of polyols and / or low molecular weight sugars, such as those having one or two monosaccharide units, i.e., some compositions do not contain any polyols and / or low molecular weight sugars, such as those having one or two monosaccharide units.

[0025] The present invention also provides use of a composition comprising (i) a curcuminoid, (ii) an extract obtained or obtainable from Quillaja quilla, and (iii) modified starch and / or gum arabic, for improving the bioaccessibility, bioavailability, bioefficacy and / or bioactivity of curcuminoid in a mammal.

[0026] The compositions used in the methods / uses of the invention described herein may be in the form of an emulsion or the compositions of the invention may be in the form of a solid, for example in the form of a powder.

[0027] As used herein, the term "emulsion" refers to a type of colloid formed by combining two mostly immiscible liquids, where one of the liquids typically contains a dispersion of the other liquid.

[0028] Sometimes the terms "colloid" and "emulsion" are used interchangeably, but as used herein, the term emulsion applies when both phases of the mixture are liquid. The particles in a colloid can be of any phase of matter. Thus, while emulsions are a type of colloid, not all colloids are emulsions. A colloidal solution, sometimes identified as a colloidal suspension, is a mixture in which a substance is periodically suspended in a fluid. Such a use may hereinafter be referred to as "the use of the invention".

[0029] In the method or use of the present invention, the curcuminoid in the composition may be obtained from any source. However, it is preferred that the curcuminoid is obtained from a natural source, i.e., the curcuminoid is not synthetic and is plant-based.

[0030] In the compositions used in the methods / uses described herein, the compositions may comprise particles having an average diameter of about 200 nm, 300 nm, 400 nm, 500 nm, 600 nm, 700 nm, 800 nm, 900 nm, 1000 nm, 1100 nm, 1200 nm, 1300 nm, 1400 nm or 1500 nm to about 9000 nm, 8000 nm, 7000 nm, 6000 nm, 5000 nm, 4000 nm, 3000 nm or 2000 nm, for example, about 1000 nm to about 6000 nm. The particles may also have an average diameter of about 200 nm to about 600 nm, or about 300 nm to about 500 nm or about 400 nm.

[0031] For example, in the compositions used in the methods / uses described herein, the composition may comprise particles having an average diameter of from about 100 nm to about 700 nm, such as from about 200 nm to about 600 nm or from about 300 nm to about 500 nm or about 400 nm.

[0032] For example, the composition may contain particles having an average diameter of about 550 nm to about 700 nm and particles having an average diameter of about 100 nm to about 250 nm, resulting in an average diameter of about 400 nm.

[0033] For example, when the composition is in the form of an emulsion, the composition may contain, for example, particles having an average diameter of about 550 nm to about 700 nm and particles having an average diameter of about 100 nm to about 250 nm, resulting in an average diameter of about 400 nm. When the composition is in the form of a solid, such as a powder, the composition may comprise particles having an average diameter of, for example, from about 1000 nm to about 6000 nm, such as from about 2000 nm to about 4000 nm.

[0034] The particles in the composition may be in the form of micelles. In compositions of the invention, for example, when the composition is in the form of a solid, particles may be formed using such techniques known in the art, such as spray drying. After the particles are formed (eg, after drying, such as by spray drying), the particles may be ground and / or milled (eg, ball milled) to provide a more uniform size.

[0035] The size and morphology of loaded curcumin micelles were analyzed by dynamic light scattering (DLS), and zeta potential (Z-potential), and scanning electron microscopy (SEM). For DLS and zeta potential analysis, a Zetasizer Nano ZS (NanoZS90, Malvern Instrument Ltd., UK) equipped with a He / Ne laser (λ=633 nm) at a fixed scattering angle of 90° at a temperature of (25±0.1° C.). For example, the size of the particles may be measured by the CQ-MO-304 method as defined in the examples below.

[0036] In the composition, the curcuminoid may be obtained from any source, however, it is preferred that the curcuminoid is obtained from a natural source, i.e., the curcuminoid is not synthetic and is plant-based. The compositions of the present invention may comprise at least about 2.5% curcuminoid, by weight of the composition. Typically, in the compositions used in the methods / uses described herein, the curcuminoids may be present in an amount of from about 2.5% to about 60% by weight, such as from about 5% to about 50% or from about 10% to about 45%, or from about 15% to about 40% by weight of the composition.

[0037] The curcuminoids may be provided by extraction and optional purification from turmeric (Curcuma longa) root (rhizome), oleoresin turmeric root, defatted oleoresin turmeric root and mixtures thereof, i.e. (i) the curcuminoids may be in the form of an extract or purified extract of turmeric containing from about 30% to about 100% curcuminoids, such as from about 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85% or 90% to about 95%, 90%, 85%, 80%, 75%, 70%, 65%, 60%, 55%, 50% or 45% curcuminoids, based on the percentage of total curcuminoids in the extract.

[0038] When the curcuminoids are provided as an extract of turmeric, the turmeric may be extracted using a water / alcohol mixture or an alcohol-based extraction solvent such as alcohol. For example, the alcohol-based extraction solvent may be water / methanol (i.e., a mixture of water and methanol) or water / ethanol (i.e., a mixture of water and ethanol) or methanol or ethanol.

[0039] When the extraction solvent comprises a water / alcohol mixture, the ratio of water to alcohol may be from about 25:75 to about 1:99, such as from about 20:80 to about 5:95 or about 10:90. For example, the extraction solvent may be water / ethanol in a ratio of from about 25:75 to about 1:99, such as from about 20:80 to about 5:95 or about 10:90.

[0040] The turmeric extract may then be further refined to provide an extract of curcuminoids containing from about 30% to about 100% curcuminoids, such as from about 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85% or 90% to about 95%, 90%, 85%, 80%, 75%, 70%, 65%, 60%, 55%, 50% or 45% curcuminoids based on the percentage of total curcuminoids in the extract.

[0041] Purification of the extract may be carried out using such techniques known in the art. Typically, the extract is purified using an alcohol-based solvent, such as 100% methanol or 100% ethanol. The turmeric extract may optionally be dried to remove excess solvent.

[0042] When the curcuminoids are provided in the form of a turmeric extract as defined above, the composition may comprise from about 2.5% to about 50% turmeric extract by weight, such as from about 5% to about 40 or about 35% by weight of the composition. For example, the composition may comprise from about 8% to about 40% turmeric extract by weight of the composition, where the turmeric extract comprises from about 65% to about 95% curcuminoids by weight of the turmeric extract, providing a composition comprising from about 6% to about 38% curcuminoids by weight of the composition, or the composition may comprise from about 8% to about 15% turmeric extract by weight, where the turmeric extract comprises from about 95% curcuminoids by weight of the turmeric extract, providing a composition comprising from about 6% to about 13.5% curcuminoids.

[0043] The curcuminoids may be provided as a liquid or powder, such as a powder, for example, a powdered turmeric extract. As used herein, the term "curcuminoid" includes curcumin, demethoxycurcumin (DMC), and bisdemethoxycurcumin (BDMC). For example, a turmeric extract may contain about 70% to about 85% curcumin (such as about 75% to about 80%), about 10% to about 25% DMC (such as about 15% to about 20%), and about 0% to about 10% BDMC.

[0044] When present, the modified starch in the composition may be present in an amount from about 30% to about 90% by weight of the composition, such as from about 40% to about 80% by weight of the composition. As used herein, the term "modified starch" includes starches that have been physically, enzymatically, or chemically treated, but does not include modified starches that have a cyclic structure, such as cyclodextrins. Modified starches that may be used in the compositions described herein include, but are not limited to, maltodextrins.

[0045] Gum arabic in the compositions of the present invention may be present in an amount of from about 40% to about 65% by weight of the composition, such as from about 50% to about 60% or about 58% by weight of the composition. The extract obtained or obtainable from Quillaja in the compositions of the present invention may be present in an amount of from about 0.1% to about 5% by weight, such as from about 0.5% to about 3% by weight of the composition.

[0046] The extract obtained or obtainable from Quillaja in the composition may contain at least 50% saponin by weight, such as at least 60% saponin by weight or at least 65% saponin by weight of the Quillaja extract. For example, the Quillaja used in the composition of the invention may contain about 50% to about 80% or about 60% to about 75% saponin by weight of the Quillaja extract.

[0047] The extract obtained or obtainable from Quillaja used in the process of the present invention may be in any form, such as a liquid or a solid. For example, the Quillaja extract may be used in the form of a solid, such as a powder.

[0048] The composition may optionally include a plant and / or vegetable oil. For example, the composition of the present invention may include a plant and / or vegetable oil selected from the group consisting of coconut oil, corn oil, cottonseed oil, olive oil, palm oil, peanut oil, rapeseed oil including canola oil, safflower oil, sesame oil, soybean oil, sunflower oil, and mixtures thereof. The plant and / or vegetable oil present in the compositions of the present invention may be present in an amount of from about 1% to about 20% by weight of the composition, such as from about 2.5% to about 10% or about 5% by weight. Unless otherwise stated herein, the weight percentages stated are based on the total weight of the resulting (dry) composition.

[0049] For the avoidance of doubt, preferences, options, specific features, etc. for a given aspect, feature or parameter of the invention should be considered as disclosed in combination with any and all other preferences, options, specific features, etc. as indicated for the same or other aspects, features and parameters of the invention, unless the context indicates otherwise.

[0050] The term "about" as used herein, by way of example, when referring to a measurable value (such as the amount or weight of a specific component in a reaction mixture), refers to a variation of ±20%, ±10%, ±5%, ±1%, ±0.5%, or specifically ±0.1% relative to the specified amount. For example, a variation of ±0.5% with respect to a percentage of a component in a composition of the invention means a variation of 0.5% relative to the given percentage, i.e., ±0.5% of 10% means a variation of 9.5% to 10.5%.

[0051] The compositions of the present invention may be provided in solid or liquid form, preferably in a solid form such as a powder. Solid form includes that the compound may be provided as an amorphous solid, or as a crystalline or partially crystalline solid.

[0052] The compositions of the present invention are typically highly water soluble and / or stable at a pH of 4 or above, such as a pH of about 4 to about 7. The term water-soluble means that at least about 50%, such as at least about 60%, 70%, 80%, 90% or 95% of the composition will dissolve in water at room temperature, i.e., a temperature of about 25°C.

[0053] In the methods or uses described herein, improved bioaccessibility, bioavailability, bioefficacy and / or bioactivity of curcuminoids in mammals may be due to improved gastrointestinal tolerance of the curcuminoids and / or improved absorption of the curcuminoids by intestinal cells and / or improved blood circulation. Therefore, the methods or uses described herein may be used to improve the gastrointestinal tolerance of curcuminoids and / or to improve the absorption of curcuminoids by intestinal cells and / or to improve blood circulation.

[0054] In the methods or uses described herein, improved bioaccessibility, bioavailability, bioefficacy and / or bioactivity of curcuminoids in mammals may be due to compositions providing improved water solubility and / or improved stability at a pH of about 4 to about 7.

[0055] Thus, the present invention provides a method for improving the water solubility and / or pH stability of a curcuminoid, said method comprising administration of said curcuminoid in the form of a composition of the present invention as defined above. The present invention also provides the use of a composition as defined above for improving the water solubility and / or pH stability of a curcuminoid.

[0056] In the methods or uses described herein, the curcuminoid may be selected from the group consisting of curcumin and its phase I or phase II metabolites, bisdemethoxycurcumin and its phase I or phase II metabolites, and mixtures thereof. For example, the phase I and / or phase II metabolites may be selected from the group consisting of curcumin glucuronide, curcumin sulfate, DMC glucuronide, DMC sulfate, BDMC glucuronide, BDMC sulfate, tetrahydrocurcumin (THC), THC glucuronide, THC sulfate, hexahydrocurcumin (HHC), HHC glucuronide, HHC sulfate, and mixtures thereof.

[0057] In the compositions, methods or uses described herein, the curcuminoids may be in their unmetabolized form (i.e., free form), such as forms without glucuronide or sulfate attachment or curcumin, DMC and BDMC. In the methods and uses described herein, the mammal may be a human.

[0058] As used herein, the term "bioavailability" can be defined as the fraction of an ingested component available at the site of action for utilization in normal physiological function, measured via in vivo assays (Guerra et al. 2012). Bioavailability is the result of three major steps: digestibility and solubility of the element in the gastrointestinal tract; absorption of the element by enterocytes and transport into circulation; and incorporation into a functional entity or target (Wienk KJH, Marx JJM, Beynen AC (1999) The concept of iron bioavailability and its assessment. Eur J Nutr 38:51-75; Etcheverry P, Grusak MA, Fleige LE (2012) Application of in vitro bioaccessibility and bioavailability methods for calcium, carotenoids, folate, iron, magnesium, polyphenols, zinc, and vitamins B6, B12, D, and E. Front Physiol 3:1-21).

[0059] As used herein, the term "bioaccessibility" can be defined as the fraction of a compound that is released from its food matrix in the gastrointestinal tract and thus becomes available for intestinal absorption (typically established from in vitro procedures). It encompasses the series of events that occur during food digestion for the conversion into potentially bioaccessible substances, but excludes absorption / assimilation via epithelial tissues and presystemic metabolism (both intestine and liver). (Alegria A., Garcia-Llatas G., Cilla A. (2015) Static Digestion Models: General Introduction. In: Verhoeckx K. et al. (eds) The Impact of Food Bioactives on Health. Springer, Cham)

[0060] As used herein, the term "bioactivity" can be defined as how a nutrient or bioactive compound is transported to reach a target tissue, how it interacts with biomolecules, the metabolic or biotransformations it may undergo, and the production of biomarkers and physiological responses elicited (Alegria A., Garcia-Llatas G., Cilla A. (2015) Static Digestion Models: General Introduction. In: Verhoeckx K. et al. (eds) The Impact of Food Bioactives on Health. Springer, Cham).

[0061] In the methods / uses described herein, the compositions may be administered or used to provide curcuminoids in an amount of about 100 mg / day to about 2000 mg / day, such as about 300 mg / day to about 1000 mg / day, or about 500 mg / day to about 1500 mg / day, or about 1000 mg / day. For example, the compositions may provide curcuminoids in an amount of about 1 to about 10 mg / kg body weight, such as about 2.5 to about 7.5 mg / kg body weight or about 5 mg / kg.

[0062] Compositions and Administration According to the invention, the compositions used in the methods and uses described herein may be provided in the form of, or form part of, a nutraceutical formulation, a human or animal dietary product or food (functional food formulation, i.e. food, beverage, feed or pet food or a food, beverage, feed or pet food supplement, etc.), a herbicide, a nutraceutical supplement, a fragrance or flavouring agent, a pharmaceutical or veterinary preparation, an oenological or cosmetic preparation.

[0063] For example, the invention provides a nutraceutical formulation, a human or animal dietary product or food (functional food formulation, i.e. food, beverage, feed or pet food or food, beverage, feed or pet food supplement), nutraceutical supplement, fragrance or flavoring agent, pharmaceutical or veterinary preparation, oenological or cosmetic preparation which consists, consists essentially of (i.e. at least 90% w / w of) the composition of the invention, for example at least 95% or 99% or 99.5%, of the composition of the invention) or which comprises a composition of the invention.

[0064] The present invention also provides the use of the compositions of the present invention in dietary supplement formulations, dietary products or foods for humans or animals (functional food formulations, i.e. foods, beverages, feeds or pet foods or food, beverage, feed or pet food supplements, etc.), dietary supplements, fragrances or flavoring agents, pharmaceutical or veterinary preparations, oenological or cosmetic preparations.

[0065] Where the composition of the invention is or may form part of a nutraceutical formulation, a dietary product or food for humans or animals (such as a functional food formulation, i.e. a food, beverage, feed or pet food or a food, beverage, feed or pet food supplement), a herbicide, a dietary supplement, a fragrance or flavoring agent, a pharmaceutical or veterinary preparation, an oenological or cosmetic preparation, it may optionally further comprise pharmaceutical / veterinary ingredients such as excipients or carriers or (functional) food-acceptable ingredients and mixtures thereof, as appropriate.

[0066] For the avoidance of doubt, when the term "comprising" or "comprises" is used herein, it means that the extract or composition described must contain the recited component(s), but may optionally contain additional components. When the term "consisting essentially of" or "consists essentially of" is used, it means that the extract or composition described must contain the recited component(s) and may contain small amounts (e.g., up to 5% by weight, or up to 1% by weight or 0.1% by weight) of other components, provided that any additional components do not affect the essential properties of the extract or composition. When the term "consisting of" or "consists of" is used, it means that the extract or composition described must only contain the recited component(s). It is also intended that, where appropriate, the terms "comprise" or "comprises" or "comprising" may be replaced with "consist" or "consisting" or "consisting essentially of" throughout the application.

[0067] As used herein, reference to a pharma- ceutically acceptable excipient may refer to a pharma- ceutically acceptable adjuvant, diluent and / or carrier as known to those skilled in the art. Food acceptable ingredients include those known in the art (including those also referred to herein as pharma- ceutically acceptable excipients) and can be natural or non-natural, i.e., their structure may or may not occur in nature. In some instances, they can be derived from natural compounds and modified prior to use (e.g., maltodextrin).

[0068] "Pharmaceutically / nutritional acceptable" means that the additional components of the composition are sterile and pyrogen-free. Such components must be "acceptable" in the sense of being compatible with the composition of the present invention and not harmful to the recipient thereof. Thus, "pharmaceutically acceptable" encompasses any compound used to form part of a formulation that is intended to act merely as an excipient, i.e., not to have biological activity itself. Thus, pharmaceutically acceptable excipients are generally safe, non-toxic, and not biologically or otherwise undesirable.

[0069] When the compositions of the invention form part of a nutraceutical formulation, a human or animal dietary product or food (such as a functional food formulation, i.e., a food, beverage, feed or pet food or a food, beverage, feed or pet food supplement), a dietary supplement, a fragrance or flavoring agent, a pharmaceutical or veterinary preparation, an oenological or cosmetic preparation, the compositions of the invention may be present in an amount of about 1 to about 99% by weight of the nutraceutical formulation, a human or animal dietary product or food (such as a functional food formulation, i.e., a food, beverage, feed or pet food or a food, beverage, feed or pet food supplement), a dietary supplement, a fragrance or flavoring agent, a pharmaceutical or veterinary preparation, an oenological or cosmetic preparation. , food, beverage, feed or pet food or food, beverage, feed or pet food supplements, etc.), dietary supplement, fragrance or flavoring agent, pharmaceutical or veterinary preparation, enology or cosmetic formulation, for example, from about 10% by weight, 20% by weight, 30% by weight, 40% by weight, 50% by weight, 60% by weight, 70% by weight, 80% by weight, 50% by weight, 40% by weight, 30% by weight, 50% by weight, 60% by weight, 70% by weight, 80% by weight, 90% by weight, 90% by weight, 80% by weight, 70% by weight, 60% by weight, 50% by weight, 40% by weight, 30% by weight, 50% by weight, 30% by weight, 50% by weight, 4 ...

[0070] Administration The skilled artisan will appreciate that in the methods and / or uses described herein, the compositions of the present invention, whether in the form of a nutraceutical formulation, a human or animal dietary product or food (such as a nutraceutical formulation, i.e., a food, beverage, feed or pet food or a food, beverage, feed or pet food supplement), a dietary supplement, a fragrance or flavoring agent, a pharmaceutical or veterinary preparation, an oenological or cosmetic preparation, or in the case where a nutraceutical formulation, a human or animal dietary product or food (such as a nutraceutical formulation, i.e., a food, beverage, feed or pet food or a food, beverage, feed or pet food supplement), a dietary supplement, a fragrance or flavoring agent, a pharmaceutical or veterinary preparation, an oenological or cosmetic preparation comprises the compositions of the present invention, may be administered to a patient or subject (for example a human or animal patient or subject) by any suitable route, such as oral, rectal, nasal, pulmonary, buccal, sublingual, transdermal, intracapsular, intraperitoneal, and parenteral (including subcutaneous, intramuscular, intrathecal, intravenous and intradermal) routes.

[0071] In particular, in the methods and / or uses described herein, the composition or compositions in the form of or forming part of a nutraceutical formulation, a human or animal dietary product or food product (such as a functional food formulation, i.e. a food, drink, feed or pet food or a food, drink, feed or pet food supplement), a nutraceutical supplement, a fragrance or flavoring agent, a pharmaceutical or veterinary preparation, an oenological or cosmetic preparation comprising the composition of the invention may be administered orally. In such cases, the pharmaceutical composition according to the invention may be specially formulated for administration by the oral route.

[0072] Pharmaceutical preparations for oral administration include solid dosage forms such as hard or soft capsules, tablets, troches, dragees, pills, lozenges, powders and granules. Where appropriate, they can be prepared with coatings, such as enteric coatings, or can be formulated so as to provide controlled release of the active ingredient, such as sustained or extended release, according to methods well known in the art. Liquid dosage forms for oral administration include solutions, emulsions, aqueous or oily / oil-based suspensions, syrups and elixirs.

[0073] The nutraceutical formulations, human or animal dietary products or foods (such as functional food formulations, i.e., foods, beverages, feeds or pet foods or food, beverage, feed or pet food supplements), nutraceutical supplements, fragrances or flavoring agents, pharmaceutical or veterinary preparations, oenological or cosmetic preparations, intended for oral administration, etc. described herein may be prepared according to methods known to those skilled in the art, such as by mixing the components of the composition.

[0074] Such nutraceutical formulations, human or animal dietary products or foods (such as functional food formulations, i.e., food, beverage, feed or pet food or food, beverage, feed or pet food supplements), nutraceutical supplements, fragrances or flavoring agents, pharmaceutical or veterinary preparations, oenological or cosmetic preparations described herein may contain one or more additional ingredients selected from the group consisting of food ingredients such as sweeteners, flavoring agents, colorants and preservatives. Tablets may contain the active ingredient(s) mixed with non-toxic pharmaceutically acceptable excipients (or ingredients) that are suitable for the manufacture of tablets. These excipients (or ingredients) may be, for example: inert diluents such as calcium carbonate, sodium carbonate, lactose, calcium phosphate or sodium phosphate; granulating and disintegrating agents, such as corn starch, maltodextrin or alginic acid; binding agents, such as starch, gelatin or acacia; and lubricants, such as magnesium stearate, stearic acid or talc. The tablets may be uncoated or may be coated by known techniques to delay disintegration and absorption in the gastrointestinal tract and thereby provide a sustained action over a longer period, for example a time delay material such as glyceryl monostearate or glyceryl distearate may be used.

[0075] Suitable pharmaceutical carriers include inert solid diluents or fillers, sterile aqueous solutions, and various organic solvents.The examples of solid carriers are lactose, terra alba, sucrose, cyclodextrin, maltodextrin, talc, gelatin, agar, pectin, acacia, magnesium stearate, stearic acid, gum arabic, modified starch, and lower alkyl ether of cellulose.The examples of liquid carriers are syrup, peanut oil, olive oil, phospholipids, fatty acids, fatty acid amines, polyoxyethylene, and water.In addition, carriers or diluents can include any sustained release material known in the art, such as glyceryl monostearate or glyceryl distearate, alone or mixed with wax.

[0076] Depending on the disorder and subject to be treated and the route of administration, the composition of the present invention can be administered in various doses (i.e., therapeutically effective doses administered to patients in need thereof).In this regard, those skilled in the art will understand that the dose administered to a mammal, particularly a human, in the context of the present invention should be sufficient to affect the therapeutic response of the mammal over a reasonable time frame.Those skilled in the art will recognize that the selection of the exact dose and composition and the most suitable delivery regimen will also be influenced by, among other things, the pharmacological properties of the formulation, the nature and severity of the condition to be treated, and the physical condition and mental acuity of the recipient, as well as the potency of the specific compound, the age, condition, weight, sex and response of the patient to be treated, and the stage / severity of the disease.

[0077] Typically, the compositions or nutraceutical formulations, human or animal dietary products or foods (such as nutraceutical formulations, i.e., foods, beverages, feeds or pet foods or food, beverage, feed or pet food supplements), nutraceutical supplements, fragrance or flavoring agents, pharmaceutical or veterinary preparations, oenological or cosmetic preparations of the present invention are administered to provide an amount of curcuminoids of about 100 mg / day to about 2000 mg / day, such as about 300 mg / day to about 1000 mg / day, or about 500 mg / day to about 1500 mg / day, or about 1000 mg / day. For example, a composition or nutraceutical formulation, a human or animal dietary product or food (such as a functional food formulation, i.e., a food, beverage, feed or pet food or a food, beverage, feed or pet food supplement), a nutraceutical supplement, a fragrance or flavoring agent, a pharmaceutical or veterinary preparation, an oenological or cosmetic preparation may provide curcuminoids in an amount of about 1 to about 10 mg / kg body weight, such as about 2.5 to about 7.5 mg / kg body weight or about 5 mg / kg body weight.

[0078] In any event, a physician or other skilled artisan may routinely determine the actual dosage amount most suitable for an individual patient. The dosage amounts above are exemplary of the average case; there can, of course, be individual instances where higher or lower dosage ranges are merited, and such are within the scope of this invention.

[0079] Process for preparing the composition of the present invention The present invention also provides a process for the preparation of the composition of the invention as defined above, said process comprising the following steps: (i) preparing an aqueous solution of curcuminoids; (ii) mixing the aqueous solution from (i) with an aqueous gum arabic solution, and an extract obtained or obtainable from Quillaja japonica, and optionally a vegetable and / or vegetable oil, to provide an emulsion; and optionally (iii) drying the product of (ii) to provide a composition comprising particles having an average diameter of from about 100 nm to about 10,000 nm, such as from about 100 nm to about 700 nm or from about 1,000 nm to about 6,000 nm; Includes.

[0080] For example, the present invention provides a process for the preparation of the composition of the invention as defined above, said process comprising the following steps: (i) preparing an aqueous solution of curcuminoids; (ii) mixing the aqueous solution from (i) with an aqueous gum arabic solution, and an extract obtained or obtainable from Quillaja japonica, and optionally a vegetable and / or vegetable oil, to provide an emulsion; and (iii) drying the product of (ii) to provide a composition comprising particles having an average diameter of from about 100 nm to about 700 nm, such as from about 100 nm to about 700 nm or from about 1000 nm to about 6000 nm; Includes. Such a process is hereinafter referred to as the process of the invention.

[0081] In the process of the present invention, the aqueous solution of gum arabic may be mixed with an aqueous solution of curcuminoids before being mixed with the plant and / or vegetable oil and the extract obtained or obtainable from Quillaja japonica, i.e. the process of the present invention comprises the steps of: (i) preparing an aqueous solution of curcuminoids; (ii) mixing the aqueous solution from (i) with an aqueous gum arabic solution; (iii) mixing an extract obtained or obtainable from Quillaja japonica, and optionally a plant and / or vegetable oil, with the product of (ii) to provide an emulsion; and optionally (iv) drying the product of (iii) (such as by spray drying) to provide a composition comprising particles having an average diameter of about 100 nm to about 700 nm; may also include After drying (such as spray drying), the particles may be ground and / or milled (such as ball milled) to provide a more uniform size.

[0082] In the process of the invention, the particles may have an average diameter of from about 200 nm, 300 nm, 400 nm, 500 nm, 600 nm, 700 nm, 800 nm, 900 nm, 1000 nm, 1100 nm, 1200 nm, 1300 nm, 1400 nm or 1500 nm to about 9000 nm, 8000 nm, 7000 nm, 6000 nm, 5000 nm, 4000 nm, 3000 nm or 2000 nm, such as from about 1000 nm to about 6000 nm. The particles may also have an average diameter of from about 200 nm to about 600 nm, or from about 300 nm to about 500 nm or about 400 nm.

[0083] For example, the composition may contain particles having an average diameter of about 550 nm to about 700 nm and particles having an average diameter of about 100 nm to about 250 nm, giving an average diameter of about 400 nm.

[0084] For example, when the composition of the present invention is in the form of an emulsion (i.e., before the drying step), the composition may contain particles having an average diameter of about 550 nm to about 700 nm and particles having an average diameter of about 100 nm to about 250 nm, resulting in an average diameter of about 400 nm. When the composition of the present invention is in solid form (ie, after the drying step), the composition may comprise particles having an average diameter of from about 1000 nm to about 6000 nm, such as from about 2000 nm to about 4000 nm. The particles in the composition may be in the form of micelles.

[0085] In the process of the present invention, the curcuminoids present in the aqueous solution of curcuminoids may be from any of the sources defined above for the composition of the present invention. Typically, in the process of the present invention, the curcuminoids may have a purity (based on total curcuminoids) of about 5% to about 100% by weight of the curcuminoid source, i.e., a turmeric or curcuminoid extract may contain about 30% to about 100% curcuminoids, such as about 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85% or 90% to about 95%, 90%, 85%, 80%, 75%, 70%, 65%, 60%, 55%, 50% or 45% curcuminoids based on the percentage of total curcuminoids in the extract.

[0086] In the process of the present invention, the curcuminoids may be mixed with water in a weight ratio of about 2:1 (curcuminoids:water) to about 4:1, such as about 3:1, to provide an aqueous solution of the curcuminoids. Typically, in the process of the present invention, the weight concentration of the curcuminoid in the aqueous solution may be from about 1% to about 95%, such as from about 5% to about 80% or from about 7% to about 40%.

[0087] In the process of the present invention, an aqueous gum arabic solution may be prepared by mixing gum arabic with water in a gum arabic:water ratio of about 2:1 to about 4:1, such as about 3:1. The aqueous gum arabic solution may have a weight concentration of gum arabic of about 30% to about 70%, such as about 40% to about 60%. Typically, the aqueous curcuminoid solution and the aqueous gum arabic solution are mixed using agitation.

[0088] In the process of the present invention, the plant and / or vegetable oil can be from any plant and / or plant source. For example, the plant and / or vegetable oil can be sunflower oil. Typically, in the process of the present invention, the vegetable and / or vegetable oil may be present in an amount of from about 1% to about 10%, such as from about 2.5% to about 7.5% or about 5%.

[0089] In the process of the invention, the extract obtained or obtainable from Quillaja may be as defined above for the composition of the invention. The extract obtained or obtainable from Quillaja used in the process of the present invention may be in any form, such as a liquid or a solid. For example, the Quillaja extract may be used in the form of a solid, such as a powder. Typically, in the process of the present invention, Quillaja may be present in an amount of from about 0.5% to about 5%, such as from about 1% to about 3%, or about 2%.

[0090] In the process of the present invention, the plant and / or vegetable oil and the Quillaja extract may be mixed using stirring. An emulsion is provided by mixing an aqueous solution of curcuminoids, an aqueous gum arabic solution, a plant and / or vegetable oil and quillaja as defined above.

[0091] The resulting emulsion may be dried using techniques as known in the art to provide a composition comprising micelles having an average diameter of about 200 nm, 300 nm, 400 nm, 500 nm, 600 nm, 700 nm, 800 nm, 900 nm, 1000 nm, 1100 nm, 1200 nm, 1300 nm, 1400 nm, or 1500 nm to about 9000 nm, 8000 nm, 7000 nm, 6000 nm, 5000 nm, 4000 nm, 3000 nm, or 2000 nm, such as about 1000 nm to about 500 nm, or about 400 nm. Typically, the emulsion is spray dried.

[0092] The process of the invention may optionally include the step of removing additional solvent as necessary to provide a substantially dry product, i.e., a product from which at least 90%, such as at least 95% or 99% of the water present has been removed. [Brief description of the drawings]

[0093] [Figure 1] Figure 1- Chemical structures of curcuminoids from turmeric [Diagram 2] Figure 2- Phase I and II metabolites of curcumin [Diagram 3] Figure 3- Bacterial metabolites of curcumin

[0094] [Figure 4] Figure 4 - LDH activity measured in apical supernatants collected from Caco-2 cells treated with samples from different gastrointestinal compartments after 4 h incubation; data from standard curcumin extract. Bars represent mean ± SEM. PP: pure product; SI: small intestine; ST: stomach; TB: transport buffer. (*), (**) and (***) correspond to significance at p<0.05, p<0.01 and p<0.001 compared to TB+colon 48 h, respectively. [Diagram 5] Figure 5 - LDH activity measured in apical supernatants collected from Caco-2 cells treated with samples from different gastrointestinal compartments after 4 h incubation, data from standard curcumin extract with data normalized to TB+Colon 48h sample (100%). Bars represent mean ± SEM. PP: pure product; SI: small intestine; ST: stomach; TB: transport buffer. (*), (**) and (***) correspond to significance at p<0.05, p<0.01 and p<0.001 compared to TB+Colon 48h, respectively.

[0095] [Figure 6] Figure 6 - LDH activity measured in apical supernatants collected from Caco-2 cells treated with samples from different gastrointestinal compartments after 4h incubation; data from turmeric phytosome formulations. Bars represent mean ± SEM. PP: pure product; SI: small intestine; ST: stomach; TB: transport buffer. (*), (**) and (***) correspond to significance at p<0.05, p<0.01 and p<0.001 compared to TB+colon 48h, respectively. [Figure 7]Figure 7 - LDH activity measured in apical supernatants collected from Caco-2 cells treated with samples from different gastrointestinal compartments after 4 h incubation, data from turmeric phytosome formulation with data normalized to TB+colon 48h samples (100%). Bars represent mean ± SEM. PP: pure product; SI: small intestine; ST: stomach; TB: transport buffer. (*), (**) and (***) correspond to significance at p<0.05, p<0.01 and p<0.001 compared to TB+colon 48h, respectively.

[0096] [Figure 8] Figure 8 - LDH activity measured in apical supernatants collected from Caco-2 cells treated with samples from different gastrointestinal compartments after 4 h incubation; data from Quillaja base formulations. Bars represent mean ± SEM. PP: pure product; SI: small intestine; ST: stomach; TB: transport buffer. (*), (**) and (***) correspond to significance at p<0.05, p<0.01 and p<0.001 compared to TB+colon 48 h, respectively. [Figure 9] Figure 9 - LDH activity measured in apical supernatants collected from Caco-2 cells treated with samples from different gastrointestinal compartments after 4 h incubation, data from Quillaja base formulations with data normalized to TB+Colon 48h samples (100%). Bars represent mean ± SEM. PP: pure product; SI: small intestine; ST: stomach; TB: transport buffer. (*), (**) and (***) correspond to significance at p<0.05, p<0.01 and p<0.001 compared to TB+Colon 48h, respectively.

[0097] [Figure 10] FIG. 10 - Evolution of food intake over the acclimation and habituation periods of groups of mice sacrificed at the same time point. Average food intake (g) per day of the different experimental groups during the acclimation (J0-J7) and habituation (J8-J14) periods. [Figure 11]FIG. 11 - Evolution of body weight over the adaptation and habituation period of groups of mice sacrificed at the same time point. Mean body weight (g) of the different experimental groups during the same period. Data are expressed as mean ± SEM.

[0098] [Figure 12] Figure 12 - Time course of total curcuminoid levels (sum of curcumin, DMC, BDMC and their associated metabolites curcumin glucuronide and sulfate, DMC glucuronide and sulfate, BDMC glucuronide and sulfate, THC, THC glucuronide and sulfate, HHC, HHC glucuronide and sulfate) in mouse plasma following a single oral dose of turmeric phytosome formulation versus standard turmeric extract containing 300 mg / kg curcuminoids. *, **, ***: turmeric phytosome formulation significantly different from standard turmeric extract at each time point (post-hoc t-test) (p<0.05, p<0.01, p<0.001, respectively). [Figure 13] Figure 13 - Time course of total curcuminoid levels (sum of curcumin, DMC, BDMC and their associated metabolites, curcumin glucuronide and sulfate, DMC glucuronide and sulfate, BDMC glucuronide and sulfate, THC, THC glucuronide and sulfate, HHC, HHC glucuronide and sulfate) in mouse plasma following a single oral dose of the composition used in the method / use of the invention versus a standard turmeric extract containing 300mg / kg of curcuminoids. *, **, ***: composition used in the method / use of the invention significantly different from the standard turmeric extract at each time point (post-hoc t-test) (p<0.05, p<0.01, p<0.001 respectively).

[0099] [Figure 14]Figure 14: Total curcuminoids and metabolites (sum of curcumin, DMC, BDMC and their associated metabolites, curcumin glucuronide and sulfate, DMC glucuronide and sulfate, BDMC glucuronide and sulfate, THC, THC glucuronide and sulfate, HHC, HHC glucuronide and sulfate) concentration (ppm) as a function of time (h) after ingestion of different formulations (n=72 per formulation). □, □□: Turmeric phytosome significantly different from standard turmeric extract at each time point (p<0.05, p<0.01 respectively). *, **, ***: Composition used in the method / use of the invention significantly different from standard turmeric extract (p<0.05, p<0.01, p<0.001 respectively).

[0100] [Figure 15] FIG. 15 - Corresponding area under the curve AUC(0-8h) of different formulations. [Figure 16] FIG. 16--Corresponding area under the curve AUC(0-∞) for different formulations. [Figure 17] Figure 17 - Cmax (concentration at Tmax) of different formulations. [Figure 18] FIG. 18-Effect of pH on the color of compositions of the invention dissolved in demineralized water (0.4%). [Figure 19] FIG. 19 - DLS profile of loaded composition of the present invention.

[0101] [Figure 20] FIG. 20 - Z potential of the composition of the present invention and individual components of the composition at different pH [Figure 21] FIG. 21-Scanning electron microscope (SEM) image of a composition of the present invention at 300×. [Figure 22] FIG. 22-Scanning electron microscope (SEM) image of the composition of the present invention at 1300×. [Diagram 23] FIG. 23-Scanning electron microscope (SEM) image of a composition of the present invention at 9200×. [Figure 24] FIG. 24—Graphical representation of dose-normalized AUC0-24h for the ITT population. [Diagram 25] FIG. 25—Graphical representation of dose-normalized AUC0-8h for the ITT population.

[0102] [Figure 26] FIG. 26—Graphical representation of dose-normalized AUC0-∞ for the ITT population. [Figure 27] Figure 27 - Graphical representation of AUC0-24h for the ITT population. [Figure 28] Figure 28 - Graphical representation of AUC0-8h for the ITT population. [Figure 29] Figure 29 - Graphical representation of AUC0-∞ for the ITT population. [Diagram 30] Figure 30 - Graphical representation of normalized Cmax for the ITT population.

[0103] [Diagram 31] FIG. 31 - Graphical representation of Cmax for the ITT population. [Diagram 32] FIG. 32—Graphical representation of relative bioavailability from 0 to 24 hours for the ITT population. [Diagram 33] FIG. 33—Graphical representation of relative bioavailability from 0 to 8 hours for the ITT population. [Diagram 34] Figure 34 - Graphical representation of relative bioavailability from 0 to infinity for the ITT population. [Diagram 35] FIG. 35—Graphical representation of half-life for the ITT population. [Diagram 36] FIG. 36—Graphical representation of the terminal elimination rate constants for the ITT population. [Figure 37] FIG. 37—Graphical representation of Tmax for the ITT population.

[0104] example The invention is further described by reference to the following non-limiting examples. Example 1 - Testing the Efficacy of the Compositions of the Present Invention in Enhancing the Bioavailability of Curcuminoids in an In-Vivo Model of the Human Gastrointestinal Tract and Intestinal Absorptive Cells The human gastrointestinal tract (GIT) is one of the main entries into the human body. Upon oral ingestion of food, beverages, or medicines, the intestine becomes the first site of contact between the ingested product and the host. To exert their biological activity, compounds must first pass through the stomach, where the acidic environment and the presence of digestive enzymes can result in chemical or enzymatic modifications. After leaving the stomach, ingested compounds reach the small intestine, where most of the host's metabolic enzymes are secreted, possibly causing further enzymatic modifications. Compounds, in their original or modified form, can subsequently be absorbed and enter the circulation or pass further through the intestine. Here, dietary compounds can have local biological activity by coming into contact with the complex microbial communities present in the terminal ileum (the last part of the small intestine) and colon (Alegria et al., 2015).

[0105] Human studies are certainly one of the most representative methods to study different intestinal processes, however, they are highly laborious and time-consuming, extremely costly and do not allow mechanistic studies. In humans, the gut can be considered as a black box that allows to quantify inputs and outputs, but due to sampling issues it is difficult to investigate the underlying gut processes in different compartments. Moreover, ethical constraints limit the general applicability of human studies.

[0106] Thus, well-designed in vitro simulation techniques provide an extremely useful alternative to human and animal studies. Such models, representative of specific processes, allow reproducible and thorough study of these processes without ethical constraints. Simpler set-up and sampling allow medium- to high-throughput studies at low cost. Nevertheless, the lack of a physiological host environment is the most important limitation of these models. However, the use of standardized in vitro cell cultures using human-derived cell lines provides a fast and reproducible way to study the net effect of compounds on the intestinal mucosa. Furthermore, the wealth of in vitro investigations allows the careful design of subsequent animal or human studies, thereby saving time and costs.

[0107] In vitro approaches simulating a carefully designed GIT provide an excellent high-throughput screening setup to evaluate the putative metabolic fate of selected dietary components at different concentrations. Such components may be modified or alter the bacterial community in the gut, thus reaching the intestinal mucosa intact or in the form of modified by-products. The oral bioavailability of a dietary compound is defined as the fraction of a dose that can be absorbed by the intestinal cells and is available for use or storage.

[0108] The bioavailability of a dietary compound depends on many factors, namely the nutritional and physiological state of the individual, the conjugation of the compound with other nutrients and / or bile salts, the enzymatic degradation of the compound by digestive enzymes, and the ability of gut-associated bacteria to metabolize it. In vitro gastrointestinal models offer the possibility to screen large sets of molecules in a rapid and cost-effective manner in short-term experiments.

[0109] The following approach allows rapid assessment of the intestinal fate of dietary compounds during digestion and colonic fermentation. It is associated with an in vitro cell model simulating the human intestinal epithelium and allows investigating the bioavailability of intact and modified compounds, thereby increasing both scientific output and commercial relevance.

[0110] First, a short-term screening assay was performed as a tool to evaluate the digestive fate of curcumin-based formulations with different solubility properties. The results of these experiments were then applied in vitro to Caco-2 cells to investigate the bioavailable fraction of the different formulations compared to the unmodified / undigested form. In addition, cytotoxicity was measured to compare the putative cytotoxic effect of the different digestion fractions.

[0111] The short-term screening assay consisted of sequential incubations (stomach, small intestine, colon) of representative dose(s) of selected lead compounds under simulated colonic conditions with representative bacterial inocula. Intestinal suspensions collected from the ascending colon compartment of the Simulator of Human Intestinal Microbial Ecosystem (SHIME) were used ( Van de Wiele et al., 2015 ). This inoculum consists of a stable microbial community that is adapted, both in structure and activity, to the environmental conditions present in the proximal colon.

[0112] The following curcumin-based formulations / compositions were evaluated: 1. Standardized curcumin extract (Curcuma longa) - contains a mixture of three curcuminoids (curcumin-75%, demethoxycurcumin (DMC)-15-20% and bidemethoxycurcumin (BDMC)-5-10%). 2. Control formulation (Meriva® Turmeric Phytosome Thorne product, this formulation contains 18-22% curcuminoids, the curcumin and soy lecithin present are blended in a 1:2 weight ratio (phytosome), then 2 parts microcrystalline cellulose are added to improve flowability, the overall content of curcumin in the final product is nearly 20%). 3. A composition used in the method / use of the present invention comprising 8.6% turmeric extract (having more than 6% curcuminoids), 15.9% sunflower oil, 2% quillaja extract, and 73.5% modified starch (also referred to as Type I).

[0113] The short-term screening assay consisted of sequential incubation of the three formulations under gastric, small intestinal, and colonic conditions. The formulations / compositions were tested to achieve a curcuminoid concentration of 0.5 g / L in the gastric compartment (actual amount in mg was calculated based on the % of curcuminoids in each product - as shown in Table 1). The different formulations / compositions were then incubated in the presence of pepsin at 37° C., pH 2.0 for 1 hour (h). The small intestine was then simulated by adding pancreatic enzymes and bile salts, and the samples were incubated at 37° C. for a total duration of 3 h.

[0114] Finally, in the third incubation stage, the colon was simulated by adding a representative fecal inoculum collected from SHIME and rich nutrient medium. Colon incubation was performed under anaerobic conditions with shaking at 37°C for a total of 48 h. Each formulation / composition was tested in triplicate to control for biological variability. It should be noted that these experiments were designed to respect the specific residence time of the dietary components in the gastrointestinal tract. Taking into account the volumes in each compartment, the concentrations of the tested curcuminoids were 0.5 g / L in the stomach, 0.35 g / L in the small intestine, and 0.1 g / L in the colon. For the cellular transport experiments, these samples were diluted 10-fold or more.

[0115] Table 1: Curcumin-based formulations / compositions tested and the respective percentage of curcuminoids in the formulations / compositions. The percentage of curcuminoids was taken into account to calculate the amount of mg added to the gastric compartment. The native form is shown in bold. [Table 1]

[0116] Samples of each formulation / composition were collected at the following time points: - Stomach: 30 and 60 minutes - Small intestine: 60, 120 and 180 minutes - Colon: 2, 4, 6, 24 and 48 hours The samples were then analyzed for curcuminoid content (curcumin, DMC, and BDMC) using high pressure liquid chromatography (HPLC) coupled with mass spectrometry.

[0117] Calibration curves were constructed for each of the three curcuminoids (Phytolab, Vestenbergsgreuth, Germany) in the range of 2–1000 ng / mL, and 54 ppb of curcumin-d6 (TLC pharmachem, Ontario, Canada) was added as an internal standard to ensure retention time stability and instrument correction variations. Acetonitrile was used as diluent for each solution. For free curcuminoid identification, exactly 450 μL of the internal standard solution (60 ng / mL) was loaded onto a Captiva 96-well plate (ND lipids from Agilent) via 50 μL of plasma sample. After mixing and filtration, the eluate is ready to be injected into the LC / MS system. The Captiva ND lipid plate is designed to effectively remove phospholipids from plasma. For the determination of total conjugated curcuminoid metabolites (glucuronide and sulfate metabolites), 100 μL of plasma sample was mixed with 100 μL of enzyme solution (either glucuronidase 1000 units / mL, Sigma#G7017; or sulfatase, Sigma#S9626, 100 units / mL) for 2 h at 37 °C. After this hydrolysis step, 50 μL of the solution was mixed with 450 μL of acetonitrile on a Captiva 96-well plate as well. The sample procedure was the same as for free curcuminoids, mixing and filtering before injection.

[0118] The LC / MS conditions were as follows: the autosampler (5 °C) and LC system used was an Agilent Infinity 1290 integrated system. An Agilent 6420 triple quadrupole mass spectrometer was used with electrospray ionization during the study. The metabolites were eluted from a BEH Shield RP 18 column (100x2.1, 1.7 μm; Waters) with a mobile phase consisting of 0.1% formic acid in HPLC grade water (solvent A) and 0.1% formic acid in acetonitrile (solvent B) at a flow rate of 0.5 mL / min. Elution was a gradient from 40–80% B in 0–6 min. The injection volume of standards and samples was 2 μL. For each reference compound, multiple reaction monitoring (MRM) mode was utilized to detect the associated transition from precursor to product ions. For each of the three analytes, they were identified in MS1 ​​full scan experiments and product ions were identified in MS / MS experiments. The MRM transitions of each analyte were optimized using direct injection and the Optimizer B.08.00 workstation software solution (Agilent technologies, Santa Clara, CA, USA). See Table 2 for optimal selection conditions. Mass spectrometer parameters were set as follows: ESI source in both negative and positive modes; drying gas (N2) flow rate, 10 L / min; gas temperature, 350 °C; nebulizer, 40 psi; and capillary, 4.0 kV. The MS system was fully calibrated before running according to the manufacturer's guidelines. Data analysis was performed with Agilent MassHunter Quantitative / Qualitative Analysis B.07.00 (Agilent technologies, Santa Clara, CA, USA).

[0119] Table 2: Retention times (Tr), multiple reaction monitoring (MRM) transitions, and optimized tandem mass spectrometry (MS / MS) detection parameters of the three curcuminoids and the internal standard. [Table 2]

[0120] Table 3 shows the concentrations of curcuminoids obtained between intestinal compartments during digestion. Surprisingly, the post-digestion concentrations in the stomach after 60 minutes and in the small intestine after 120 minutes and 180 minutes for the composition used in the method / use of the present invention were superior to those obtained for the standard turmeric powder extract, demonstrating a better resistance to digestion of the composition used in the method / use of the present invention compared to the standard extract. Also, the post-digestion concentrations in the stomach after 60 minutes and in the small intestine after 120 minutes and 180 minutes for the composition used in the method / use of the present invention were higher than those of the comparative turmeric phytosome formulation, demonstrating a better resistance to digestion of the composition used in the method / use of the present invention compared to the turmeric phytosome comparative.

[0121] Table 4 shows the percentage of curcuminoids remaining in the intestinal compartment after digestion with the initial concentration (0.5 g / l or 500 mg / l of curcuminoids in the stomach compartment at the beginning of the experiment).Surprisingly, the results clearly show that the curcuminoids from the composition used in the method / use of the present invention have much better resistance to digestion in the stomach and small intestine compared to the curcuminoids from standard turmeric extract, and that the composition used in the method / use of the present invention is more protected from degradation during digestion after oral intake than the curcuminoids from standard extract or turmeric phytosome preparation, and that the curcuminoids accessible for absorption in the small intestinal compartment after oral intake of the composition used in the method / use of the present invention are much more than those from standard extract or turmeric phytosome preparation.

[0122] [Table 3-1] [Table 3-2]

[0123] Table 4: Resistance to gastrointestinal (GIT) digestion: percentage of curcuminoids remaining in the intestinal compartment after digestion according to initial concentration [Table 4] Data show the mean ± SD of the percentage of curcuminoid concentrations in different compartments and at different times after digestion compared to the initial concentration.

[0124] For transport experiments in Caco-2 cells, the following samples were also collected: - Small intestine: 120 minutes and 180 minutes The pH of the samples applied to the cells was adjusted to 6.5 before use.

[0125] Caco-2 cells are widely used as a cellular model of intestinal function because they can spontaneously differentiate into enterocyte-like cells in culture. When cultured on semipermeable supports, these cells develop into a functional polarized monolayer resembling the intestinal epithelium in the presence of apical brush-border enzymes and microvilli. Thus, they are considered the "gold standard" model for transport experiments because they acquire in culture the morphological and functional characteristics of mature enterocytes (Sambuy et al., 2002).

[0126] Caco-2 cells (ATCC) were cultured at 1 × 10 5 0.9 x 10 cells / insert 5 cells / cm 2Cells were seeded in 12 transwell inserts (0.4 μm) at a density of 0.01 mg / mL. Cells were allowed to differentiate until a functional monolayer was reached (21 days); apical (600 μL) and basolateral (1500 μL) medium was replenished three times a week. On the day of the experiment, barrier function was assessed by measuring the transepithelial electrical resistance (TEER) of the monolayer. Cells were washed with HBSS to remove traces of medium, and 2 mL of transport buffer (TB) were added to the basolateral side. Samples collected from short-term experiments were diluted in a ratio of 1:10 (v / v) in transport buffer and given to the cells apically (600 μL). All products were also tested untreated, and powders were diluted in TB at a concentration of 0.025 mg / mL (final theoretical concentrations of curcuminoids tested for all formulations can be seen in Table 2). These dilutions were prepared from stock solutions (250 mg / mL) prepared in HBSS, except for the standard turmeric extract, which was dissolved in DMSO due to its poor solubility. As control wells, colon samples diluted 1:10 (v / v) in TB (48 h incubation) obtained by running a blank (without curcumin) during the short-term experiment were used. Transport buffer (TB) consisted of HBSS (pH 7.4) supplemented with 10 mM HEPES, 25 mM D-glucose, and 1× antibiotic-antimycotic. Cells were incubated at 37°C for a total duration of 4 h.

[0127] The following samples were collected: 1. Diluted samples (500 μL) used to stimulate the cells. These correspond to the 0 h time point, as they contain samples that were diluted before being given to the cells, and untreated formulations were also shipped diluted in TB at a concentration of 0.025 mg / mL. 2. Samples from the apical side (250 μL each) collected after 2 h and 4 h of incubation. 3. Samples from the basolateral side collected after 2 h (800 μL) and 4 h (1000 μL) of incubation. 4. Samples from cells after 4h incubation. These correspond to the fraction internalized in the cells. Briefly, ice-cold PBS1X was added to the cells to stop the transport. The cells were then washed once more with PBS1X to remove traces of non-internalized products, and the cells were permeabilized with a solution of PBS1X containing 20% ​​ethanol and 0.1% Tween-20 (600μL); after 20min in this solution, the cells were collected in a 1.5mL tube and disrupted and homogenized using a syringe and a 21G needle. 5. The tubes were centrifuged and the supernatant was transferred to a new tube (450 μL). All samples were stored at -20°C until HPLC analysis.

[0128] To evaluate the cytotoxicity of the different samples applied to Caco-2 cells, lactate dehydrogenase (LDH) released by apical Caco-2 cells (after 4 h of incubation) was assessed using an LDH-Activity Kit. LDH is released into the supernatant by cells upon membrane injury and is therefore a marker of cell death. Statistical analysis was performed using one-way ANOVA followed by Dunnett's post hoc multiple comparison test. * ), ( ** ) and ( *** ) correspond to significance at p<0.05, p<0.01 and p<0.001, respectively.

[0129] The control wells (TB+colon 48h) show an LDH activity of nearly 1.0, as shown in Figure 4. As can be seen, for all products, both the undigested form and the stomach samples collected after 60 minutes showed the highest LDH activity when compared to the control. In contrast, both the small intestine and colon samples showed comparable or lower levels than the control. These results demonstrated that, except for the stomach sample, all samples showed no toxicity to Caco-2 cells, and that the results of the assays testing the transport and bioavailability of curcuminoids from the Caco-2 cell samples can be utilized and judged as valid since they are obtained on viable cells.

[0130] Since Caco-2 cells are known to express UDP-glucuronosyltransferases and sulfotransferases ((Siissalo S, Zhang H, Stilgenbauer E, Kaukonen AM, Hirvonen J, Finel M), samples from the apical, basolateral and intracellular compartments of Caco-2 cells incubated with either undigested products or samples from the small intestine (120 or 180 min) were further analyzed for their curcuminoid content (curcumin, DMC and BDMC) and their associated metabolite content (curcumin sulfate, curcumin glucuronide, DMC sulfate and DMC glucuronide, BDMC sulfate and BDMC glucuronide). The expression of most UDP-glucuronosyltransferases (UGTs) is significantly increased during differentiation of Caco-2 cells, whereas UGT1A6 is highly expressed even in undifferentiated cells and has been shown to be highly expressed in undifferentiated cells (Drug Metab Dispos. 2008). Nov;36(11):2331-6), and thus can metabolize curcumin, DMC, and BDMC to their glucuronide or sulfate metabolites (Dempe JS, Scheerle RK, Pfeiffer E, Metzler M. Metabolism and permeability of curcumin in cultured Caco-2 cells. Mol Nutr Food Res. 2013 Sep;57(9):1543-9).

[0131] The apparent permeability coefficient of the apical to basolateral transition (P app) values ​​are calculated according to Artursson and Karlsson according to the following formula:

number

[0132] Table 5 shows the P values ​​at different time intervals after apical exposure of Caco-2 cells to a standard extract or to two different formulations (turmeric phytosome or the composition used in the method / use of the present invention). aap Indicates the value.

[0133] Table 5: Papp values ​​(10) of total curcuminoids and their metabolites calculated at different time intervals after apical exposure of Caco-2 cells to the standard extract or to the two different formulations. -7 (expressed as cm / s) [Table 5] Data are shown as mean ± SD. P compared to standard extract app The fold increase in P compared to the turmeric phytosome formulation is shown in parentheses. app The doubling is shown in square brackets [].

[0134] As shown in Table 5, the apparent permeability coefficient (P aap ) values ​​were surprisingly higher for the composition used in the method / use of the present invention than for the standard turmeric extract, with P values ​​at 2h and 4h respectively compared to the standard turmeric extract. appThere was a 19.3- and 20.3-fold increase in the values. The apparent permeability coefficient of the basolateral transition (P aap ) values ​​were also higher for the turmeric phytosome formulation used as a positive control for the enhanced bioavailability formulation, with P values ​​at 2h and 4h compared to the standard turmeric extract, respectively. app However, the results show that the absorption of curcuminoids by Caco-2 absorbent cells is greater for the composition used in the method / use of the present invention compared to the comparative turmeric phytosome formulation (4.1 and 4.2 times better Papp values ​​at 2h and 4h respectively).

[0135] Table 6 shows the P values ​​after apical exposure of Caco-2 cells to small intestinal digested samples (120 or 180 min) of a standard extract or two different formulations (turmeric phytosome or composition used in the method / use of the present invention) using the quantified concentration of curcuminoids at 120 or 180 min in the small intestinal compartment as Capi. app The values ​​shown are representative of the absorption capacity of curcuminoids from the digested preparation by the cells. When the standard extract or preparation is digested in the stomach and small intestine compartments, and thus samples from the small intestine (120 min or 180 min) are used to measure the absorption of curcuminoids by Caco-2 cells, the absorption of curcuminoids by Caco-2 absorptive cells was greater for the compositions used in the method / use of the present invention compared to the turmeric phytosome preparation (P 0.01 at 120 min or 180 min for the small intestine samples, respectively). app values ​​were demonstrated to be 1.8-fold and 16.4-fold higher.

[0136] Table 7 shows the P values ​​after apical exposure of Caco-2 cells to small intestinal digestion samples (120 or 180 min) of standard extract or two different formulations (turmeric phytosome or the composition used in the method / use), using the theoretical concentration of curcuminoids at 120 or 180 min in the small intestinal compartment as Capi to take into account not only the absorption capacity of the cells but also their resistance to the digestive process. app These data show that the levels of curcuminoids that can reach the gastrointestinal compartment (mimicking the blood circulation) after absorption and digestion by Caco-2 cells are higher for the composition used in the method / use of the present invention compared to standard turmeric extract turmeric (P app values ​​were 2.0-fold and 1.8-fold higher for small intestine samples at 120 or 180 min, respectively), and compared with the phytosome formulation ( P app values ​​were demonstrated to be much higher (3.4- and 7.1-fold higher for small intestine samples at 120 or 180 min, respectively).

[0137] Table 6: P of total curcuminoids and their metabolites after apical exposure (120 or 180 min) of Caco-2 cells to standard extracts or small intestinal digesta samples of two different formulations. app value [Table 6] Data are shown as mean ± SD. P compared to turmeric phytosome app The doubling is indicated in parentheses. app Values ​​have been calculated using the quantified concentrations of curcuminoids in the small intestinal compartment at 120 or 180 min as Capi.

[0138] Table 7: P of total curcuminoids and their metabolites after apical exposure (120 or 180 min) of Caco-2 cells to standard extracts or small intestinal digesta samples of two different formulations. app value [Table 7] Data are shown as mean ± SD. P compared to standard extract app The fold increase in P compared to the turmeric phytosome formulation is shown in parentheses. app The doubling is shown in square brackets []. app Values ​​are calculated using the theoretical concentration of curcuminoids in the small intestinal compartment at 120 or 180 min as Capi.

[0139] Example 2 - Efficacy of the Composition of the Present Invention in Enhancing the Bioavailability of Curcuminoids Through an In-Vivo Comparative Pharmacokinetic Study in Mice In view of the results obtained in the in vitro model which showed that the composition used in the method / use of the present invention exhibits better resistance to gastrointestinal digestion and better absorption through enterocytes, it is envisaged that the composition used in the method / use of the present invention (i.e. a composition comprising 8.6% turmeric extract (containing more than 6% curcuminoids), 15.9% sunflower oil, 2% quillaja extract and 73.5% modified starch) will improve the bioavailability of curcuminoids in mice compared to standard turmeric extract. Therefore, a comparative pharmacokinetic study was performed in mice.

[0140] Adult male C57Bl / 6J Rj mice from Janvier Labs (St-Berthevin, France) were 5 weeks old upon receipt and housed in groups in standard plastic cages (n=4 / cage). All animals had free access to water and standard pellet food (Pellet AO4; SAFE, Villemoisson-sur-Orge, France) and were maintained in a temperature-(24.0–26.0 °C) and humidity-(40.0–50.0%) controlled room with a 12-h light (07:00 AM–07:00 PM) / 12-h dark cycle.

[0141] All animals were acclimatized to their new environment for one week after receipt. Total food intake and animal weight were evaluated twice a week to ensure both correct acclimatization and standard growth curves. Following this acclimatization period, mice were acclimatized to receive daily oral administration of vehicle (Carboxymethylcellulose sodium salt 1% (w / v), dissolved in distilled water at room temperature, CMC; Ref# C4888, Batch number: SLBB5612V, SIGMA ALDRICH, St Quentin Fallavier, France) for 6 days before treatment. During this acclimatization period, total food intake and animal weight were measured daily. These measures made it possible to ensure optimal acclimatization of the animals to both the injection procedure and the manipulations by the experimenter.

[0142] During the acclimation and habituation period, mice were allowed free access to pre-weighed amounts of fresh food pellets (pellet AO4; SAFE, Villemoisson-sur-Orge, France). The remaining food was weighed the day after the measurement. Using a precision scale (THB-600G, PMC Millot; accuracy ±0.01 g), the total food intake per cage (08:40-09:20 AM) was determined by subtracting the remaining food from the amount of pre-weighed food. The average daily food intake was obtained by dividing this value by the number of days separating the two measures and the number of animals in each cage. At each weighing, mice were weighed in the morning (08:40-09:20 AM).

[0143] The day before treatment (last day of the habituation period), 120 mice were fasted in the evening (17:40-18:20 PM).The next day, mice (n=40 per preparation) were acutely treated in the morning (08:00-09:50 AM) with standard turmeric powder extract, comparative turmeric phytosome preparation or the composition used in the method / use of the present invention (i.e., a composition containing 8.6% turmeric extract (with more than 6% curcuminoids), 15.9% sunflower oil, 2% quillaja extract, and 73.5% modified starch) by oral gavage (30 ml / kg).

[0144] An appropriate volume of vehicle (1% CMC dissolved in distilled water) was added to a suitable recipient under stirring and the pH was adjusted to 5.5. An appropriate amount of pre-weighed formulation was gradually added to the vehicle under constant stirring. Once homogenous, the pH of the resulting suspension was measured and adjusted to 5.5 if necessary. To avoid degradation of the curcuminoids, the final suspension was administered systematically 1 h after preparation. The dose to animals was calculated (see Table 8) to receive 300 mg / kg total curcuminoids. This dose in mice was derived from the formula: (US Department of Health and Human Services, US Food and Drug Administration, Center for Drug Evaluation and Research (CDER). Guidance for Industry Estimating the Maximum Safe Starting Dose in Initial Clinical Trials for Therapeutics in Adult Healthy Volunteers. 2005); Human equivalent dose (mg / kg) = Animal dose mg / kg x (Animal weight kg / Human weight kg) 0.33 This corresponds to 21.37 mg / kg in humans and 1282 mg assuming a 60 kg human. The dose was adjusted to the body weight measured immediately before fasting. To avoid the impact of feeding and drinking behavior on intestinal absorption after oral gavage with the formulation, water and food were withheld for the first 12 h after treatment.

[0145] Table 8: Curcuminoid content of standard turmeric extract and two formulations and their respective concentrations in the suspension used to administer the products to mice at 300 mg / kg body weight of curcuminoids in the first in vivo study. [Table 8]

[0146] Blood was sampled by cardiac puncture in anesthetized mice at 0.5-, 1h-, 2h-, 4h-, 6h-, 8h-, 12h- or 24h after dosing (n=5 / time point / formulation). Anesthesia was achieved by intraperitoneal injection of a mixture of ketamine / xylazine (100mg / kg and 15mg / kg, respectively). For cardiac puncture, a 26G syringe was inserted at a 45° angle to the longitudinal axis formed by the animal's body between the 8th and 10th sternal ribs in order to directly penetrate the left ventricle. Blood was then gently drawn to obtain a final volume of 0.6-1ml. For the benefit of bioanalytical methods, blood was subsequently transferred to an Eppendorf tube, mixed with heparin sulfate (200U.I / ml blood) and gently stirred. All samples were centrifuged for 15 min at 3000g and 4°C within 30 min after blood collection to separate the plasma. The plasma (supernatant) was dispensed into a new 0.5 ml Eppendorf. Plasma aliquots were frozen at -80°C within 1 hour after centrifugation.

[0147] Plasma doses of parent curcuminoids (curcumin, DMC or BDMC) and their associated metabolites (curcumin glucuronide and sulfate, DMC glucuronide and sulfate, BDMC glucuronide and sulfate, THC, THC glucuronide and sulfate, HHC, HHC glucuronide and sulfate) were performed by LC-MS-MS method. Calibration curves were prepared for each of the five curcuminoids (Phytolab, Vestenbergsgreuth, Germany) in the range of 2-1000 ng / mL, and 54 ppb of curcumin-d6 (TLC pharmachem, Ontario, Canada) was added as an internal standard to ensure retention time stability and instrument compensation variations. Acetonitrile was used as diluent for each solution. For the identification of free curcuminoids, exactly 450 μL of the internal standard solution (60 ng / mL) was loaded into a Captiva 96-well plate (ND lipids from Agilent) via 50 μL of plasma sample. After mixing and filtration, the eluate is ready to be injected into the LC / MS system. Captiva ND lipid plates are designed to effectively remove phospholipids from plasma. For the determination of total conjugated curcuminoid metabolites (glucuronide and sulfate metabolites), 100 μL of plasma sample was mixed with 100 μL of enzyme solution (either glucuronidase 1000 units / mL, Sigma#G7017 or sulfatase, Sigma#S9626, 100 units / ml) for 2 hours at 37°C. After this hydrolysis step, 50 μL of the solution is also mixed with 450 μL of acetonitrile on a Captiva 96-well plate. The sample procedure is the same as for free curcuminoids, mixing and filtering before injection.

[0148] The LC / MS conditions were then as follows: The autosampler (5 °C) and LC system used was an Agilent Infinity 1290 integrated system. An Agilent 6420 triple quadrupole mass spectrometer was used with electrospray ionization during the study. The metabolites were eluted from a BEH Shield RP 18 column (100 × 2.1, 1.7 μm; Waters) with a mobile phase consisting of 0.1% formic acid in HPLC grade water (solvent A) and 0.1% formic acid in acetonitrile (solvent B) at a flow rate of 0.5 mL / min. Elution was a gradient from 40–80% B in 0–6 min. The injection volume of standards and samples was 2 μL. For each reference compound, multiple reaction monitoring (MRM) mode was utilized to detect the associated transition from precursor to product ions. For each of the five analytes, they were identified in an MS1 ​​full scan study and product ions were identified in an MS / MS experiment. The MRM transitions of each analyte were optimized using direct injection and the Optimizer B.08.00 workstation software solution (Agilent technologies, Santa Clara, CA, USA). See Table 9 for optimal selection conditions. Mass spectrometer parameters were set as follows: ESI source in both negative and positive modes; drying gas (N2) flow rate, 10 L / min; gas temperature, 350 °C; nebulizer, 40 psi; and capillary, 4.0 kV. The MS system was fully calibrated before running according to the manufacturer's guidelines. Data analysis was performed with Agilent MassHunter Quantitative / Qualitative Analysis B.07.00 (Agilent technologies, Santa Clara, CA, USA).

[0149] For the three formulations / compositions tested, the kinetics of plasma concentrations of each curcuminoid compound were determined 0.5-12 h after treatment by calculating the mean ± SEM plasma concentrations at each time point of blood sampling. The pharmacokinetic parameters T1 / 2 (half-life), Cmax, Tmax, AUC(0-12h) and AUC(0-∞) were determined from the 0-12 h kinetics by non-compartmental analysis using PKSolver. PKSolver is a menu-driven add-in program for Microsoft Excel written in Visual Basic for Applications (VBA) to solve pharmacokinetic problems (Zhang et al., 2010). The entire data are expressed as the mean ± SEM. Statistical analysis was performed using Statview 5.0.1 (Statview software, Cary, NC, USA) and Excel 2013 programs. Data were analyzed by Student's t-test at each time point. Risk α was fixed at 0.05.

[0150] Table 9: Retention times (Tr), multiple reaction monitoring (MRM) transitions, and optimized tandem mass spectrometry (MS / MS) detection parameters of curcuminoids, tetrahydrocurcumin and hexahydrocurcumin and internal standards. [Table 9]

[0151] During the two successive periods of acclimation (J1–J7) and habituation (J8–J14), 24 h-food intake and mouse weight were measured regularly to ensure both correct acclimation and standard growth curves before treatment. For the present study, mice were housed four per cage and each was used for blood collection at the same time point after treatment with one of the three formulations. As a consequence, food intake and weight data were first analyzed per group of 15 mice sacrificed at the same time point (8 groups; 5 mice / time point / formulation). Figures 10 and 11 show the food intake and weight of mice during the acclimation and habituation periods before treatment administration, respectively; the different groups showed classical weight curves and food intake before treatment. It should be noted that the significant decrease in body weight observed in all groups at J15 (Figure 11) occurred as a result of an overnight fasting performed the night before the treatment. These results confirmed that all mice used in this experiment behaved the same and could be compared as expected.

[0152] Figure 12 shows the pharmacokinetic profiles of total curcuminoids (sum of curcumin, DMC, BDMC and their associated metabolites, curcumin glucuronide sulfate, DMC glucuronide and sulfate, BDMC glucuronide and sulfate, THC, THC glucuronide and sulfate, HHC, HHC glucuronide and sulfate) obtained at each time point in mice after oral administration of turmeric phytosome and standard extract formulations (300 mg / kg bw of curcuminoids). The turmeric phytosome formulation allowed the plasma concentration of total curcuminoids to reach 41.5 ppm (μg / ml) after 30 min, significantly superior to that obtained with the standard at each time point except 24 h, whereas the plasma concentration of total curcuminoids of the standard turmeric extract only reached a maximum of 12.9 ppm at 1 h and was below 10 ppm at all other time points. Turmeric phytosome demonstrated a 3.2-fold increase in total curcuminoids Cmax and a 3.9-fold increase in AUC compared to standard turmeric extract. These results validate the use of the turmeric phytosome formulation as a positive control to enhance curcuminoid bioavailability and validate the in vivo model to test the ability of different formulations to enhance curcuminoid bioavailability versus standard turmeric extract.

[0153] Table containing the mean values ​​± SEM for each time point shown as Table 10. The numbers in brackets next to these values ​​indicate the number of samples that showed positive values ​​relative to the total number of samples. The results of statistical comparisons are also shown in the same table. Table 11 contains the PK parameters obtained from non-compartmental analysis using PKSolver software. The percentage of variation between groups is also shown (%Var°). Data are expressed as mean ± SEM.

[0154] [Table 10] Tables 10 and 11: Shows the mean ± SEM and PK parameters obtained from non-compartmental analysis using PKSolver software for each time point shown in FIG.

[0155] FIG. 13 shows the pharmacokinetic profile of the concentration of total curcuminoids (sum of curcumin, DMC, BDMC and their associated metabolites, curcumin glucuronide sulfate, DMC glucuronide and sulfate, BDMC glucuronide and sulfate, THC, THC glucuronide and sulfate, HHC, HHC glucuronide and sulfate) obtained in mice at each time point after oral administration (300 mg / kg bw of curcuminoids) of a mixture containing curcumin, quillaja oil and modified starch (Example 2 Type 1) and a standard extract formulation. The results showed that the composition used in the method / use of the present invention can significantly increase the total curcuminoid concentration from 0.5 h to 24 h compared to the standard turmeric extract. The composition used in the method / use of the present invention demonstrated a 1.8-fold increase in total curcuminoids Cmax and a 2.2-fold increase in AUC.

[0156] A table containing the mean values ​​± SEM for each time point is shown as Table 12. The numbers in brackets next to these values ​​indicate the number of samples that showed positive values ​​relative to the total number of samples. The results of statistical comparisons are also shown in the same table. A table containing the PK parameters obtained from non-compartmental analysis using PKSolver software is shown in Table 13. The percentage of variation between groups is also shown (%Var°). Data are expressed as mean ± SEM.

[0157] [Table 11] Tables 12 and 13: Shown are the mean values ​​± SEM and PK parameters obtained from non-compartmental analysis using PKSolver software for each time point shown in FIG.

[0158] As shown in Table 14, which gives the plasma concentration of parent curcuminoids at each time point after ingesting 300mg / kg of curcuminoids from standard turmeric extract, turmeric phytosome or the composition used in the method / use of the present invention, looking specifically at the parent compounds (natural, i.e. unmetabolized curcumin, DMC and BDMC), the composition used in the method / use of the present invention was the only one where detectable amounts of parent curcuminoids could be quantified during the first 4h after administration and therefore AUC(0-12h) and AUC(0-∞) could be calculated (Table 15). A 10.9-fold increase in Cmax of parent curcuminoids was obtained for the composition used in the method / use of the present invention compared to the standard turmeric extract.

[0159] As shown in Table 16, which gives the plasma concentration of parent curcumin at each time point after ingesting 300 mg / kg of curcuminoids from standard turmeric extract, turmeric phytosome or the composition used in the method / use of the present invention, looking specifically at the parent compound (natural, i.e. unmetabolized curcumin), the composition used in the method / use of the present invention was the only one that allowed quantification of detectable amounts of parent curcuminoids during the first 4 h after administration and therefore calculation of AUC(0-12h) and AUC(0-∞) (Table 16). A 521.8-fold increase in Cmax of parent curcumin was obtained for the composition used in the method / use of the present invention compared to the standard turmeric extract. Moreover, the composition used in the method / use of the present invention induced higher plasma levels of parent curcumin than the turmeric phytosome formulation (1.8-fold increase in Cmax).

[0160] From this first in vivo experiment it can be concluded that the composition used in the method / use of the present invention, having more than 6% curcuminoids and obtained with 8.6% turmeric extract, 15.9% sunflower oil, 2% quillaja extract and 73.5% modified starch and prepared according to type 1, is able to enhance the bioavailability of the total curcuminoids and their metabolites as well as the bioavailability of the parent compounds compared to standard turmeric extract. It can also be concluded that the composition used in the method / use of the present invention is able to improve the bioavailability of natural curcumin better than turmeric phytosome formulations.

[0161] Therefore, the compositions used in the methods / uses of the present invention represent an attractive way to enhance the bioavailability of parent curcumin without the use of soy-derived lecithin in the formulation as opposed to turmeric phytosome formulations. Also, curcumin is considered one of the most potent active substances of turmeric compared to DMC and BDMC and their associated reduced, glucuronide or sulfate metabolites (reson C, Orr S, Jones DJ, Verschoyle R, Lim CK, Luo JL, Howells L, Plummer S, Jukes R, Williams M, Steward WP, ​​Gescher A. Characterization of metabolites of the chemopreventive agent curcumin in human and rat hepatocytes and in the rat in vivo, and evaluation of their ability to inhibit phorbol ester-induced prostaglandin E2 production. Cancer Res. 2001 Feb 1;61(3):1058-64; Anand P, Thomas SG, Kunnumakkara AB, Sundaram C, Harikumar KB, Sung B, Tharakan ST, Misra K, Priyadarsini IK, Rajasekharan KN, Aggarwal BB. Biological activities of curcumin and its analogues (Congeners) made by man and Mother Nature. Biochem Pharmacol. 2008 Dec 1;76(11):1590-611; Pal A, Sung B, Bhanu Prasad BA, Schuber PT Jr, Prasad S, Aggarwal BB, Bornmann WG.Curcumin glucuronides: assessing the proliferative activity against human cell lines. Bioorg Med Chem. 2014 Jan 1;22(1):435-9), the composition used in the method / use of the present invention represents a good solution to improve the biological effectiveness of curcumin against different health conditions such as joint health, inflammation, arthritis, atherosclerosis, fatty liver, liver fibrosis, diabetes, cognition, mild cognitive impairment, irritable bowel syndrome, etc.

[0162] Table 14: Concentration of parent curcuminoids (sum of curcumin, DMC and BDMC) at each time point after ingestion of 300 mg / kg of curcuminoids from standard turmeric extract, turmeric phytosome or the composition used in the method / use of the present invention in the first in vivo study. [Table 12] The numbers in parentheses next to these values ​​indicate the number of samples that showed positive values ​​relative to the total number of samples.

[0163] Table 15: PK parameters obtained from non-compartmental analysis using PKSolver software for parent curcuminoids after ingestion of 300 mg / kg of curcuminoids from standard turmeric extract, turmeric phytosome or the composition used in the method / use of the present invention in the first in vivo study. [Table 13]

[0164] Table 16: Concentration of parent curcumin at various time points following ingestion of 300 mg / kg curcuminoids from standard turmeric extract, turmeric phytosome, or the composition used in the method / use of the present invention in the first in vivo study. [Table 14] The numbers in parentheses next to these values ​​indicate the number of samples that showed positive values ​​relative to the total number of samples.

[0165] Table 17: PK parameters obtained from non-compartmental analysis using PKSolver software for curcumin after ingesting 300 mg / kg of curcuminoids from standard turmeric extract, turmeric phytosome or the composition used in the method / use of the present invention in the first in vivo study. [Table 15]

[0166] Considering the results obtained in the first in vivo study in mice showing better bioavailability of total and parent curcuminoids and curcumin, it was decided to test in a second comparative pharmacokinetic study in mice an optimized formulation with a higher curcuminoids content (12% curcuminoids) prepared according to type 2 with 14.4% turmeric extract, 26.8% sunflower oil, 2% quillaja extract and 56.8% modified starch for its ability to improve the bioavailability of curcuminoids compared to the standard turmeric extract.

[0167] The same methodology (housing of mice, acclimation period, habituation period, quantification of curcuminoids and their metabolites using LC / MS methodology) was used as previously described in this example, but with a higher number of animals per group and time point (n=12 / time point / formulation) and with the addition of standard turmeric extract (79.5, 15.0, 3.0 g / 100 g curcumin, DMC and BDMC respectively and 97.5 g total curcuminoids / 100 g), turmeric phytosome formulation (18.6, 2.6 and 0.2 g / 100 g curcumin, DMC and BDMC respectively and 97.5 g total curcuminoids / 100 g), and turmeric phytosome formulation (18.6, 2.6 and 0.2 g / 100 g curcumin, DMC and BDMC respectively and 97.5 g total curcuminoids / 100 g). In order to clarify the kinetic profile in the earliest phase after oral intake of curcuminoids (300 mg / kg body weight) from the composition used in the method / use of the present invention prepared according to type 1 (9.8, 1.6 and 0.2 g / 100 g curcumin, DMC and BDMC and 11.6 ...

[0168] Figure 14 shows the total curcuminoid and metabolite concentrations as a function of time after ingesting different formulations (n=72 per formulation).The results clearly show a significant increase in the curcuminoid and metabolite concentrations of turmeric phytosome and the composition used in the method / use of the present invention compared to standard turmeric extract.The total curcuminoid concentration is higher at 0.5h and 0.75h after ingesting the composition used in the method / use of the present invention compared to the turmeric phytosome formulation, which surprisingly shows a better performance of the composition used in the method / use of the present invention in terms of improving the bioavailability of total curcuminoids and metabolites compared to the turmeric phytosome formulation.

[0169] This was confirmed while calculating the corresponding areas under the curves AUC(0-8h), AUC(0-∞) and Cmax (Figures 15, 16 and 17, respectively), which were 280%, 300%, 337% higher, respectively, for the composition used in the method / use of the present invention compared to the standard extract, and 6.5%, 30% and 63% higher, respectively, for the composition used in the method / use of the present invention compared to the turmeric phytosome formulation (Table 18).It also showed that the curcuminoids and metabolites from the composition used in the method / use of the present invention are absorbed more quickly, with a 1.5-fold reduction in Tmax compared to the standard extract (0.5h vs. 0.75h) and a 2-fold reduction in Tmax compared to the turmeric phytosome formulation (0.5h vs. 1h), respectively (Table 18). Surprisingly, the results also showed that curcuminoids and metabolites from the compositions used in the methods / uses of the present invention were not excreted more rapidly with a longer half-life (3.8h vs. 2.8h) compared to the turmeric phytosome formulation.

[0170] Looking specifically at the parent compounds (natural, i.e. unmetabolized curcumin, DMC and BDMC), as shown in Table 19, which gives the PK parameters of parent curcuminoids after ingestion of 300mg / kg from standard turmeric extract, turmeric phytosome or the composition used in the method / use of the present invention, surprisingly, the composition used in the method / use of the present invention was the only one for which the AUC(0-8h) could be calculated for parent curcumin. A 3.2-fold increase in Cmax of parent curcuminoids was obtained for the composition used in the method / use of the present invention compared to standard turmeric extract. The Cmax of curcumin could not be calculated because parent curcumin could not be found in the plasma samples after ingestion of the turmeric phytosome formulation.

[0171] From this second in vivo experiment, it is concluded that the composition used in the method / use of the present invention having higher curcuminoids content (12% curcuminoids) prepared according to type 2 with 14.4% turmeric extract, 26.8% sunflower oil, 2% quillaja extract and 56.8% modified starch is able to unexpectedly enhance the bioavailability of parent curcumin as well as total curcuminoids and their metabolites compared to standard turmeric extract and turmeric phytosome formulation. Therefore, the compositions used in the methods / uses of the present invention represent an attractive way to enhance the bioavailability of curcuminoids without the use of soy-derived lecithin in the formulation, as opposed to turmeric phytosome formulations.

[0172] Table 18: PK parameters obtained from non-compartmental analysis using PKSolver software for total curcuminoids and metabolites after ingesting 300 mg / kg of curcuminoids from standard turmeric extract, turmeric phytosome or the composition used in the method / use of the present invention in the second in vivo study. [Table 16] The fold increase in AUC or Cmax compared to the standard extract is shown in parentheses (). The fold increase in AUC or Cmax compared to the turmeric phytosome formulation is shown in brackets [ ].

[0173] Table 19: PK parameters obtained from non-compartmental analysis using PKSolver software for parent curcuminoids after ingestion of 300 mg / kg of curcuminoids from standard turmeric extract, turmeric phytosome or the composition used in the method / use of the present invention in the second in vivo study. [Table 17]

[0174] Example 3 - Comparative pharmacokinetic study in healthy volunteers to evaluate the ability of the compositions of the present invention to enhance the bioavailability of curcuminoids. This study had two objectives: 1. 1st purpose 300 mg of the composition of the present invention (Turmipure GOLD TM To evaluate the plasma concentration profile of total curcuminoids (curcumin, demethoxycurcumin (DMC), bisdemethoxycurcumin (DBMC) and their metabolites) 24 hours after a single dose of a 1500mg standardized turmeric powder extract 95% curcuminoids formulation compared to 1500mg standardized turmeric powder extract 95% curcuminoids.

[0175] 2. Secondary purpose 1425mg (standardized turmeric powder extract 95% curcuminoid, Curcumin C3 complex California Gold Nutrition), 200mg (Curcuma Platinum MannaVital), 90mg of the composition of the present invention (Turmipure GOLD TM To evaluate the plasma concentration profile of the following parameters after a single intake of the five study products containing either 30% curcuminoids) or 60 mg (Curcumin Cell'Innov) of the active substance: Total curcuminoids; Parent compounds (curcumin, DMC, BDMC) and their metabolites: curcumin glucuronide and sulfate; DMC glucuronide and sulfate; BDMC glucuronide and sulfate; tetrahydrocurcumin (THC) natural, glucuronide and sulfate; hexahydrocurcumin (HHC) natural, glucuronide and sulfate. This study was a monocentric, randomized, crossover, and open clinical trial.

[0176] The study began with a screening / inclusion visit (V0), followed by five experimental sessions (V1–V5), during which the studied products were ingested by subjects (one different product at each session for each randomized subject). The V1 visit took place up to 3 weeks after V0 and could also constitute the randomization visit. Each experimental session (V1-V5) was separated by a minimum of 1 week and a maximum of 2 weeks. During each experimental session, subjects underwent kinetic blood sampling over an 8-h period. The final kinetic blood sample was collected the day after each experimental session, 24 h after the start of the kinetic period. Urine collection was also performed during these visits for biobanking.

[0177] Subjects' first voided urine was collected on the morning of each study visit (the entirety of this first void), with additional collections from 0 to 8 hours during on-site dynamic blood collection, and from 8 to 24 hours upon discharge. The final urine collection was returned the day after the study visit (upon return for the final blood sample, T24H, of the dynamic blood collection). Standard meals were provided to the volunteers for diner before each experimental session and during all periods of each activity (breakfast, lunch, afternoon snack). The end of the study was the day after the last experimental session, V5 (V5-24H).

[0178] Thirty subjects were recruited for this study according to the following main inclusion and exclusion criteria: ■ I1: 18-45 years (encompassing limits); ■ I2: 19-25kg / m 2 BMI (including limits); ■ I3: stable weight, within ±3 kg in the past 3 months; ■ I4: routine blood chemistry values ​​within normal range;

[0179] ■ I5: For women: not menopausal, using the same reliable method of contraception for at least 3 cycles prior to the start of the study and agreeing to maintain it for the entire duration of the study (accepting condoms with spermicidal gel and estrogen / progestin combination contraceptives), or menopausal with or without hormone replacement therapy (excluding estrogen replacement therapy started less than 3 months ago); ■ I6: quit smoking or tobacco intake ≤5 cigarettes / day and agree not to smoke during all experimental sessions (V1–V5); ■ I7: Agree to not consume foods, beverages, and condiments containing curcumin or other curcuminoids (DMC, BDMC) for the entire duration of the study; ■ I8: Good general and mental health in the investigator's opinion: no clinically significant and relevant abnormalities of medical history or physical examination;

[0180] ■ E1: suffer from a metabolic or endocrine disorder such as diabetes, uncontrolled or controlled thyroid disorder, or other metabolic disorder; ■ E2: suffers from a severe chronic disease (e.g. cancer, HIV, renal failure, ongoing liver or biliary disorders, chronic inflammatory digestive diseases, arthritis or other chronic respiratory disorders) or a gastrointestinal disorder, or a gastrointestinal disorder found by the investigator to be incompatible with the conduct of the study (e.g. celiac disease); ■ E3: Having liver disease; ■ E4: Current medical conditions contraindicating dietary supplementation in subjects: chronic diarrhea, constipation or abdominal pain, inflammatory bowel disease (Crohn's disease or ulcerative colitis), liver cirrhosis, chronic laxative use...; ■ E5: suffers from irritable bowel syndrome (IBS) diagnosed by a physician and treated with chronic medication;

[0181] ■ E6: have a history of a current medical condition that, according to the investigator, may affect the study results or expose the subject to additional risk; ■ E7: Foodborne illness such as recent gastroenteritis or confirmed food poisoning (<1 month); ■ E8: Anyone who has donated blood within 3 months prior to the V0 visit or intends to donate within the next 3 months. ■ E9: In the opinion of the investigator, unable to perform blood sample dynamics, has low venous capital; ■ E10: have a known or suspected food allergy or intolerance or hypersensitivity (e.g., gluten intolerance, celiac disease) to any of the study product ingredients and / or standard diet; ■ E11: Women who are pregnant or breastfeeding, or intend to become pregnant within the next 3 months;

[0182] ■ E12: Presents with alcohol or drug dependence; ■ E13: Any chronic drug treatment, except oral and local contraceptives (e.g. anticoagulants, antihypertensive treatments, thyroid treatments, asthma treatments, anxiolytics, antidepressants, lipid-lowering treatments, corticosteroids, phlebotonics, veinotonics, drugs affecting blood circulation...). ■ E14: Currently (and in the past 3 months) taking any supplements from plants; ■ E15: consumed curcumin-containing food supplements (curcumin, turmeric, and curry) or foods (curcumin, turmeric, E100, and curry) at least three times a week for 2 weeks prior to the study; ■ E27: Control records with clinically significant abnormalities according to the investigator (blood glucose, GGT, ASAT, ALAT, urea, creatinine and complete blood count).

[0183] As part of the study, five dietary supplement products in capsule form were tested: 1. Standardized Turmeric Powder Extract 95% Curcuminoids 1500mg taken as capsules (4 capsules; 375mg powder per capsule) (STE), 2. C3 complex® 95% curcuminoids (1500 mg) + BioPerine® 95% piperine (15 mg) taken as the commercial product Curcumin C3 complex California Gold Nutrition (3 caps; 500 mg C3 complex powder + 5 mg BioPerine powder per capsule) (TEP); 3. Meriva® (1000 mg) Curcuma Platinum Mannavital 20% Curcuminoids (2 caps; 500 mg powder per capsule) (PHYT), taken as a commercial product; 4. Novasol® (1000 mg) Curcumin Cell innov 6% curcuminoids (2 caps; 500 mg liquid per capsule) (NOV), taken as a commercial product; 5. A composition as defined herein comprising turmeric extract, sunflower oil, quillaja extract, and gum arabic taken as a capsule (1 capsule; 300 mg of powder per capsule) (TURMIPURE GOLD).

[0184] Blood samples were collected during the V0 visit to ensure the subjects' health status and to confirm eligibility criteria, and control record analysis and pregnancy testing were performed in non-menopausal women (βhCG dose). Samples were taken after physical examination and verification of eligibility criteria. A maximum of 10 mL was collected. Blood pressure measurements were performed at each visit during the physical examination using an electronic blood pressure monitor (Carescape Dinamap® V100). Heart rate (HR, bpm), systolic blood pressure (SBP, mmHg), and diastolic blood pressure (DBP, mmHg) were also assessed. All subjects participated in a 12-h fasting state. In preparation for visits V1–V5, after clinical examination, subjects were fitted with an intravenous catheter in the elbow crease. This catheter allowed for blood sampling for kinetics without an additional puncture.

[0185] The dynamic samples lasted approximately 8 hours and all subjects stayed at the Clinical Trials Center. Ten blood samples were taken according to the following schedule: - T-10 (baseline), - T15>T30>T45>T60>T90>T120>T240>T360>T480, Margins of ±30 seconds were allowed for T15, ±1 minute for T30 and T45, ±2 minutes for T60 and T90, and ±5 minutes for T120 to T480. The TO time point corresponds to ingestion of the study product.

[0186] Volunteers were allowed to consume a standard lunch approximately 4 hours after study product intake (just after time T240) and a standard afternoon meal approximately 8 hours after study product intake. Lunch was consumed over a maximum of 30 minutes. No water was allowed 1 h before and 1 h after product administration. Catheters were removed after the final time point, T480.

[0187] Volunteers were then asked to return to the clinical trial site the day after the visit for the final dynamic blood draw, T24H, in a 12-hour fasting state. Classical venous blood sample material was used (single prick). The biological parameters assessed in these samples were analyzed in plasma, therefore only EDTA tubes were used (5 mL per sample).

[0188] Analysis population ■ ITT population: All subjects randomized in the study who received at least one dose of the product (n=30). ■ PP population: subjects included in the ITT population who completed the study without major protocol deviations (n=30). The following subjects were excluded from the PP population: ■ All parameters are subject to SN01-040-V5 ■ Safety population: All subjects randomized in the study who received at least one dose of the product (n=30) [Table 18] Table 20: Description of study population at baseline showing means and standard deviations.

[0189] Software Environment ■ Statistical analyses were performed by Biofortis using SAS® software version 9.3 (SAS Institute Inc., Cary, NC, USA). ■ Significance level ■ All statistical tests (two-tailed) used the 0.05 level of significance to justify claims of statistically significant effects.

[0190] ■ How to handle missing data on dynamics ■ If more than two values ​​or two consecutive values ​​were missing for kinetics, AUC calculations could not be performed and kinetics were considered missing in the statistical analysis (no replacement of missing data was performed); ■ In cases where data were missing at T-10, AUC calculations could not be performed and kinetics were considered missing in the statistical analysis (no replacement of missing data was performed); ■ If values ​​(except for baseline and last time point values) were missing for dynamics, they were replaced by values ​​obtained using the CopyMean method developed by Genolini (Genolini, 2013). ■ If values ​​were missing at the last dynamic time point (T1440 = T24h), no replacement of missing data was performed. ■ In cases where the dynamics are incomplete after missing data processing, the AUC cannot be calculated. This method was applied to the ITT and PP populations.

[0191] Derived variables ■ Total curcuminoids = curcumin + DMC + BDMC + THC + HHC + curcumin glucuronide + DMC glucuronide + BDMC glucuronide + THC glucuronide + HHC glucuronide + curcumin sulfate + DMC sulfate + BDMC sulfate + THC sulfate + HHC sulfate If all 15 of these components were missing, total curcuminoids could not be calculated. If at least one of these 15 components was quantified, total curcuminoids were calculated. ■ Total parent compounds = Curcumin + DMC + BDMC ■ Total parent compounds and their associated sulfate and glucuronide metabolites = curcumin + curcumin glucuronide + curcumin sulfate + DMC + DMC glucuronide + DMC sulfate + BDMC + BDMC glucuronide + BDMC sulfate

[0192] ■ Curcumin and its related sulfate and glucuronide metabolites = curcumin + curcumin glucuronide + curcumin sulfate ■ DMC and its related sulfate and glucuronide metabolites = DMC + DMC glucuronide + DMC sulfate ■ BDMC and its associated sulfate and glucuronide metabolites = BDMC + BDMC glucuronide + BDMC sulfate ■ Curcumin and all its related metabolites = Curcumin + Curcumin glucuronide + Curcumin sulfate + THC + THC glucuronide + THC sulfate + HHC + HHC glucuronide + HHC sulfate

[0193] ■ Relative bioavailability from 0 to 24 hours = the ratio of the dose-normalized AUC0-24h for the different test formulations to the dose-normalized AUC0-24h obtained for the reference product (turmeric extract 95% curcuminoids) ■ Relative bioavailability from 0 to infinity = the ratio of dose-normalized AUC0-∞ for the different test formulations to the dose-normalized AUC0-∞ obtained for the reference product (turmeric extract 95% curcuminoids)

[0194] Data processing for values ​​below limit of detection (LOD) ■ For curcumin (native, glucuronide and sulfate), expressed in the database as «0.62», several values ​​below the limit of detection (LOD) were identified. →The number and percentage of values ​​below the LOD were given for each parameter and visit.

[0195] Creating a graphical display ■ Boxplot of observed mean quantitative variables: AUC parameters (example shown in figure below) [Table 19]

[0196] Checking assumptions of statistical tests ■ Assumptions of normality and homogeneity of variance were investigated by graphical display of the residuals generated by the statistical model. In case of strong departures from normality and / or homogeneity of variance, log-transformation (log10) of the study endpoints was considered. Notes on producing results: ■ STE = Standardized Turmeric Extract Powder 95% Curcuminoids 1500mg ■ TEP=Curcumin C3 complex California Gold Nutrition(1500mgC3 complex(registered trademark)) ■ NOV=Curcumin Cell'Innov (1000 mg Novasol(registered trademark)) ■ PHYT = Curcuma Platinum MannaVital (1000mg Meriva (registered trademark)) ■ Turmipure GOLD TM=Turmipure Gold 30% curcuminoids300mg

[0197] statistical methodology ■ Primary endpoint: Dose-normalized AUC from 0 to 24 hours was analyzed using the following mixed model for repeated measures (SAS® PROCMIXED, statistical model n°1): Y = product + visit + baseline + target ランダム Along with: ■ Y: dose-normalized AUC of analyte plasma concentration from 0 to 24 h; ■ Product: Turmipure Gold TM , STE, TEP, NOV, PHYT; ■ Visit: Visit V1~V5; ■ Baseline: value of parameter at time point T-10 (T0 for AUC calculation); ■ Target ランダム : Random factor. ■ In case of a significant visit effect (p<0.05): a second analysis realised in the first period (visit) to assess the product effect. ■ Comparison between products of interest → Turmipure Gold compared to STE TM

[0198] Additional analysis: Examining gender effects ■ In this study, we investigated gender effects using the following mixed model (SAS® PROCMIXED, statistical model n°2) for repeated measures: Y = product + visit + gender + product * Gender + Baseline + Target ランダム Along with: ■ Y:Endpoint; ■ Product: Turmipure Gold TM , STE, TEP, NOV, PHYT; ■ Visit: Visit V1~V5; ■ Gender: female or male; ■ Products * Gender: The interaction between product and gender; ■ Baseline: value of parameter at time point T-10 (T0 for AUC calculation); ■ Target ランダム : Random factor. ■ Comparisons between products of interest; ■ Turmipure Gold compared to STE TM

[0199] ■ TEP compared with STE; ■ NOV compared with STE; ■ PHYT compared with STE; ■ Turmipure Gold TM TEP compared with; ■ Turmipure Gold TM NOV compared to; ■ Turmipure Gold TM Compared to PHYT. ■ In case of a significant visit effect (p<0.05): a second analysis realised in the first period (visit) to assess the product effect. ■ Significant products * For gender interaction effects (p<0.05): treatment effects examined separately for men and women (with the creation of descriptive statistics and graphical displays) ■ If there is a significant product*gender interaction effect (p<0.05): the treatment effect examined overall (women and men together)

[0200] Statistical Methodology for Secondary Endpoints (Excluding Relative Bioavailability) ■ The secondary endpoint was analyzed using the following mixed model (SAS® PROCMIXED, statistical model n°1) for repeated measures: Y = product + visit + baseline + target ランダム ■ In case of a significant visit effect (p<0.05): a second analysis realised in the first period (visit) to assess the product effect. ■ Comparisons between products of interest; ■ Turmipure Gold compared to STE TM ■ TEP compared with STE; ■ NOV compared with STE; ■ PHYT compared with STE; ■ Turmipure Gold TM TEP compared with; ■ Turmipure Gold TM NOV compared to; ■ Turmipure Gold TM Compared to PHYT.

[0201] ■ Statistical methodology for relative bioavailability ■ The secondary endpoint was analyzed using the following mixed model (SAS® PROCMIXED, statistical model n°1) for repeated measures: Y = product + visit + baseline + target ランダム ■ In case of a significant visit effect (p<0.05): a second analysis realised in the first period (visit) to assess the product effect. ■ Comparisons between products of interest; ■ Turmipure Gold TM TEP compared with; ■ Turmipure Gold TM NOV compared to; ■ Turmipure Gold TM Compared to PHYT.

[0202] Summary of primary endpoint results Tables 21 and 22: Initial Analysis of ITT and PP Populations [Table 20-1] [Table 20-2]

[0203] Tables 23 and 24: Additional analyses for the ITT and PP populations (examining gender effects) [Table 21]

[0204] [Table 22]

[0205] From the above analysis, the following was concluded for the ITT population: Between-group analyses (all genders combined) ■ No significant visits were identified → As a result, analysis was conducted on all visits. ■ Significant product effect (p<0.0001): ■ Primary endpoint: Turmipure GOLD TM and STE, there is a statistically significant difference (adjusted p<0.0001; diff[(adjusted CI95%]=1.32[1.18;1.46]). ■ Other comparisons: ■ TEP vs STE (adjusted p=0.6948) ■ NOV vs STE (adjusted p<0.0001; diff[adjusted CI95%]=1.62[1.48;1.76]) → NOV>STE ■ PHYT vs STE (adjusted p<0.0001; diff[adjusted CI95%]=0.48[0.34;0.62]) → PHYT>STE ■ TEP vs. Turmipure GOLD TM (adjusted p<0.0001; diff[adjusted CI95%]=-1.39[-1.53;-1.25]) → TEP <Turmipure GOLD TM ■ NOV vs Turmipure GOLD TM (adjusted p<0.0001; diff[adjusted CI95%]=0.29[0.15;0.43]) → NOV>Turmipure GOLD TM ■ PHYT vs Turmipure GOLD TM (adjusted p<0.0001; diff[adjusted CI95%]=-0.84[-0.99;-0.70]) → PHYT <Turmipure GOLD TM ■

[0206] ■ Additional analysis: Examining gender effects ■ Significant visits (p=0.2456) and products * No interaction effect of gender (p=0.3804): analyses were performed for all genders together and across all visits. ■ Significant product effect (p<0.0001): ■ TEP vs STE (adjusted p=0.7091) ■ NOV vs STE (adjusted p<0.0001; diff[adjusted CI95%]=1.61[1.47;1.75]) → NOV>STE ■ PHYT vs STE (adjusted p<0.0001; diff[adjusted CI95%]=0.48[0.34;0.62]) → PHYT>STE ■ Turmipure GOLD TM vs STE (adjusted p<0.0001; diff[adjusted CI95%]=1.32[1.18;1.46]) → Turmipure GOLD TM >STE ■ TEP vs. Turmipure GOLD TM (adjusted p<0.0001; diff[adjusted CI95%]=-1.39[-1.52;-1.25]) → TEP <Turmipure GOLD TM ■ NOV vs Turmipure GOLD TM (adjusted p<0.0001; diff[adjusted CI95%]=0.29[0.15;0.43]) → NOV>Turmipure GOLD TM ■ PHYT vs Turmipure GOLD TM(adjusted p<0.0001; diff[adjusted CI95%]=-0.84[-0.98;-0.70]) → PHYT <Turmipure GOLD TM

[0207] About PP Group: ■ Between-group analyses (all genders combined) ■ The results are similar to those observed in the ITT population. ■ Additional analysis: Examining gender effects ■ The results are similar to those observed in the ITT population.

[0208] Summary of secondary endpoint results The results of the secondary endpoints are shown in the tables below for both the ITT (Tables 26-42) and PP (Tables 43-59) populations, respectively. The results are displayed graphically in Figures 24-37. The following key applies to each table and indicates the statistical significance of each result. [Table 23]

[0209] [Table 24]

[0210] [Table 25]

[0211] [Table 26]

[0212] [Table 27]

[0213] [Table 28]

[0214]

Table 29

[0215]

Table 30

[0216]

Table 31

[0217]

Table 32

[0218]

Table 33

[0219]

Table 34

[0220]

Table 35

[0221]

Table 36

[0222]

Table 37

[0223]

Table 38

[0224]

Table 39

[0225]

Table 40

[0226]

Table 41

[0227]

Table 42

[0228]

Table 43

[0229]

Table 44

[0230]

Table 45

[0231]

Table 46

[0232]

Table 47

[0233]

Table 48

[0234]

Table 49

[0235] [Table 50]

[0236] [Table 51]

[0237] [Table 52]

[0238] [Table 53]

[0239] [Table 54]

[0240] [Table 55]

[0241] [Table 56]

[0242] [Table 57]

[0243] Serious Adverse Events : ■ Subject SN01-009: Neck pain / traffic accident (under no study product) at V0-V1 visit (mobile / rheumatoid body tissue, event unrelated to medical history, moderate intensity, no action on study product, event unrelated to investigation and study product, event unrelated to corrective treatment, recovery without sequelae).

[0244] ■ Treatment of emergent AEs of severe severity: ■ Target SN01-007: Lower back pain in V2~V3 visits (Turmipure GOLD TM under product) (Athletic / rheumatoid body tissue, events unrelated to medical history, severe intensity, no action on study product, events unrelated to investigation and study product, events unrelated to corrective treatment, recovery without sequelae).

[0245] ■ AEs related to research products: ■ Subject SN01-008: Headache on the day of V3 visit (Turmipure GOLD TM Product) (neurological / psychiatric somatic system, events unrelated to medical history, moderate intensity, no action on study product, events unrelated to investigation and study product, events unrelated to corrective treatment, recovery without sequelae). ■ Target SN01-030: Headache on the day of V3 visit (Turmipure GOLD TM under product) (neurological / psychiatric body system, events unrelated to medical history, moderate intensity, no action on investigational product, events possibly related to investigational and investigational products, events not related to corrective measures (paracetamol), recovery without sequelae). ■ Subject SN01-032: Headache on the day of V2 visit (under NOV product) (neurological / psychiatric system, event unrelated to medical history, moderate intensity, no action on study product, event possibly related to investigation and study product, event not related to corrective procedure, recovered without sequelae).

[0246] The results observed in the PP population were similar to those in the ITT population. The mean ± SD (ITT population) for total curcuminoids is shown below.

[0247] [Table 58]

[0248] conclusion The results demonstrate that there are few differences in the bioavailability of compounds found in TEP and STE (only 5 differences). Compositions within the scope of the present invention (Turmipure GOLD TM ) was found to provide better bioavailability of the compounds than STE, TEP and PHYT, was able to provide similar bioavailability to NOV despite being administered at a lower dose (300 mg compared to 1000 mg), and uses a natural source of curcuminoid compounds. More specifically, Novasol was used at 1000 mg, whereas the present invention (Turmipure GOLD TM ) composition was 300 mg. Turmipure gives an effect of 6520 at 300 mg, so when used in the same dosage as Novasol (1000 mg), it gives an effect of 21733, which is much higher than the effect of Novasol (8539) at the same dosage.

[0249] Example 4 - General preparation of compositions used in the methods / uses described herein. A mixture of curcuminoids in water was prepared using organic purified curcuminoid extract (at least 10% but preferably 95% purity (total curcuminoids)) in distilled water (3 volumes powder weight / water).

[0250] A 58% gum arabic mixture (substrate) was prepared using distilled water (3 volumes powder weight / water). To 500 ml of the aqueous gum arabic solution, 500 ml of the curcuminoid solution prepared in Example 1 was added under stirring (5000 rpm), to which was added 5% organic sunflower oil and 2% organic quillaja standardized with saponin. The resulting mixture was stirred at 5000 rpm for 10 minutes. The resulting emulsion was then spray dried.

[0251] Example 5 - Characterization of compositions used in the methods / uses described herein The size and morphology of the compositions of the present invention were analyzed by dynamic light scattering (DLS), zeta potential (Z-potential), and scanning electron microscopy (SEM). For DLS and zeta potential analysis, a Zetasizer Nano ZS (NanoZS90, Malvern Instrument Ltd., UK) equipped with a He / Ne laser (λ=633 nm) at a fixed scattering angle of 90° at a temperature of (25±0.1° C.) was used. The samples used were in the form of liquid emulsions (last step before drying). They were suspended in demineralized water at a volume concentration of 0.4% and subjected to 1 min of ultrasound. DLS analysis was carried out immediately on these samples (measurement time = 60 s). Zeta potential analysis was carried out over a wide pH range (2-11). Samples were prepared and analyzed at different pH by using 0.1 M HCl and 0.1 M NaOH solutions as follows: The resulting 10 samples (pH = 2, 3, 4, 5, 6, 7, 8, 9, 10, and 11) were stored at room temperature (23°C) (Figure 4).

[0252] As shown in Figure 18, the color of the composition of the present invention in water is triggered by pH. The keto form (yellow color) is the predominant form present in solution as the pH range changes from acidic to neutral (2-7). At pH 8 and 9, the color solution turns orange, and at pH 10 and 11, a translucent reddish color predominates. The color change is due to the successive deprotonation of the hydroxyl groups of the curcumin molecule caused by the increase in pH, which renders curcumin more soluble and unstable. The results of the DLS analysis are shown in Figure 19. There are two groups of particle size populations: one centered at 616 ± 160 nm (20.8% of the total population) and the most interesting one centered at 188 ± 42 nm (79.2% of the total population). The average hydrodynamic particle size of curcumin loaded in aqueous solution (pH 5.4) was found to be 476.5 nm with a PDI (polydispersity index) of 0.337.

[0253] FIG. 20 shows the Z-potential of the composition of the present invention at different pH (2-11). The higher the Z-potential, the more unstable the mixture. The composition of the present invention has a negative Z-potential between pH 2 and pH 11. The particles are negatively charged in the aqueous phase. At pH 2, the Z-potential is close to 0 (isoelectric point: pH at zero potential) and there is an instability zone of the emulsion. Between pH 2 and pH 4, the Z-potential is relatively low (<25 mV) and at pH 4 and above, the sample enters the stability zone. This stability is strongly confirmed from pH 5. A rapid shift in the isoelectric point was observed at pH 8.0 and the zeta potential of the loaded curcumin was surprisingly high in the pH range of 2-7. In the aqueous phase at pH < 4.0, the loaded curcumin is mostly in the lowest surface energy state. At pH 8.0, the loaded curcumin faces its low-charged side towards the gum arabic exposing its high-charged side and favoring interaction with water, which results in an elevated zeta potential.

[0254] Example 6 - Particle size distribution (PSD) of the composition of the present invention using CQ-MO-304 Materials and Reagents material -Mastersizer 3000 from Malvern Instrument, or equivalent -Hydro 2000SM sample dispersion unit, or equivalent (for liquid phase), -Malvern AERO S sample dispersion unit, or equivalent (for solid phase). reagent -water

[0255] procedure Analysis parameters -Background Time: 10 seconds - Measurement time @ 10 seconds -Refractive index of distilled water: 1.33 -Result calculation: General purpose -Pump / agitation speed: 1800RPM -Liquid dispersant: Water -Solid dispersion: ambient air Specific parameters -100705 (Refractive index: 1, Absorption: 1) -100019 (Refractive index: 1, Adsorption: 2) -3CAA0075 and 3CAA0076 (compositions of the invention) Samples of the compositions of the present invention were mixed with distilled water and the samples were tested using either a Hydro 2000SM unit or a Mastersizer 3000 (using a Scirocco 2000 unit).

[0256] result Several batches of the composition of the present invention obtained after drying and grinding were tested according to the method described above, and the results are shown in Table 61 below.

[0257] Table 61 Particle size distribution of compositions of the present invention, where (D90) corresponds to 90% of the particle size population and (D4:3) corresponds to the volume moment mean of the particle size population. [Table 59-1] [Table 59-2] [Table 59-3]

[0258] Example 7 - Morphology of the composition of the present invention (by scanning electron microscope, SEM) For SEM analysis, samples were prepared as follows: the composition of the invention in powder form was deposited on a sample holder by simple dusting, after which a platinum / palladium deposit was metallized and then observed and photographed under a scanning electron microscope equipped with an energy dispersive X-ray detector. The SEM images shown in Figures 21, 22 and 23 provide a visualization of the compositions of the present invention.

[0259] The compositions of the present invention show conjugates self-assembled into spherical micelles with a size of + / - 170 nm. The roughly spherical morphology in SEM analysis confirmed the size measurement analysis performed by dynamic light scattering. From the SEM, it can be seen that the particles in the composition of the present invention clearly exhibit a chitosan shell that was not present in the uncoated lecithin nanoparticles. The curcumin was found to be well dispersed in the lecithin core of the nanoparticles. The SEM measurements also support evidence of a roughly spherical shape, with surface roughness indicating surface adsorption. The driving force for this type of adsorption is suggested to be either direct electrostatic interactions or ion-ion interactions.

Claims

1. A method for improving the bioaccessibility, bioavailability, bioefficacy and / or bioactivity of a curcuminoid in a mammal, the method comprising administering the curcuminoid in the form of a composition comprising (i) a curcuminoid, (ii) quillaja, and (iii) modified starch and / or gum arabic.

2. Use of a composition comprising (i) a curcuminoid, (ii) Quillaja quinata, and (iii) modified starch and / or gum arabic to improve the bioaccessibility, bioavailability, bioefficacy and / or bioactivity of a curcuminoid in a mammal.

3. 3. The method or use according to claim 1 or 2, wherein the improved bioaccessibility, bioavailability, bioefficacy and / or bioactivity of curcuminoids in mammals is due to improved gastrointestinal tolerance of curcuminoids and / or improved absorption of curcuminoids by intestinal cells and / or improved blood circulation.

4. 4. The method or use according to any one of claims 1 to 3, wherein the curcuminoid is selected from the group consisting of curcumin and its phase I or II metabolites, demethoxycurcumin and its phase I or II metabolites, bisdemethoxycurcumin and its phase I or II metabolites and mixtures thereof.

5. The method or use according to any one of claims 1 to 4, wherein the mammal is a human.

6. The method or use according to any one of claims 1 to 4, wherein the curcuminoid is in free form.

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