EP3938071A1 - Novel solvent for extracting, solubilising and/or formulating volatile and non-volatile compounds of interest in animal nutrition and health, preparation method and uses thereof - Google Patents

Abstract

The subject matter of the invention is a method for preparing a solid and/or liquid totum or filtrate comprising - a first step of placing in contact via mixing a biological matrix or part of a biological matrix with a solvent; - at least a second step which is a phase of extracting, solubilising and/or formulating said mixture; optionally a filtration step; and - a final step of recovering the totum or the filtrate thus obtained. The invention is characterised in that the solvent is glycerol monolaurate.

Description

Description

Title of the invention: [New solvent for the extraction, solubilization and / or formulation of volatile and non-volatile compounds of interest in animal nutrition and health, preparation process and associated uses]

[0001] [The present invention relates to a process for preparing a totum or a solid and / or liquid filtrate from a biological matrix or part of a biological matrix and a solvent, as well as the totum or the filtrate which can be obtained according to said process, and its uses.

[0002] A totum can be defined as a mixture of a plant matrix, preferably in powder form with a solvent. The totum thus comprises active compounds or metabolites which may or may not be extracted from the plant matrix.

[0003] The active compounds or natural metabolites are molecules originating from a biological matrix or part of a biological matrix, generally plant, whose biological and technological activities have been demonstrated and described in the literature. These natural active compounds may be in pure form or contained in extracts. The benefit of these active compounds can be established in the context of food and / or human or animal welfare and / or health. Their use as additives may encompass different purposes, such as in particular:

improving health (anti-oxidant, anti-inflammatory, anti-microbial compounds in particular, alkaloids and polyphenols),

improving palatability (compounds that increase palatability such as aromatic compounds, terpenes or pigments such as carotenoids or chlorophylls),

contribution to nutrition (nutrients such as proteins, amino acids, vitamins, trace elements, etc.).

[0004] The natural metabolites of plant origin to which it is possible to attribute biological activities of interest in food and human or animal health can belong to different families of molecules. They are mainly secondary metabolites, which, unlike primary metabolites, are directly non-essential for plant nutrition, growth and development (Verpoorte, 2000, Secondary metabolism. In Metabotic engineering of plant secondary metabolism (pp. 1-29). Springer, Dordrecht). These are compounds whose biosynthetic pathways are quite specific to a taxonomic group and which generally participate in the interaction mechanisms between the plant and its environment (defense, resistance and responses to abiotic and biotic stresses, symbiosis, allelopathy. ..).

[0005] There are different families of secondary metabolites of interest in food and animal health.

[0006] First of all, the alkaloids (generally alkaline compounds which contain at least one nitrogen atom). These are compounds which generally have significant biological activity, in particular an action on the central and / or peripheral nervous system (stimulant or depressant), in particular as anesthetics, as hypertensives or anti-hypertensives, as antimalarials or even as anticancer drugs.

[0007] Alkaloids are generally grouped according to their nucleus (non-heterocyclic, derived from indole, pyrrole, pyridine, tropane, etc.). Among the alkaloids, we find well-known molecules such as caffeine, morphine, pi peri ne, nicotine, atropine, scopolamine or even quinine.

[0008] Capsaicinoids, including capsaicin and dihydrocapsaicin can represent up to 90% of total capsaicinoids. These are the active constituents from chili peppers that are part of the benzylamine alkaloids group. The consumption of capsaicin activates the TRPV1 receptors which activates a burning sensation. It also stimulates the production of two hormones, adrenaline and norepinephrine and therefore has a therapeutic interest given its anti-inflammatory, antioxidant and analgesic properties (Zimmer et al., 2012, Antioxidant and anti-inflammatory properties of Capsfcum baccatum . from traditional use to scientific approach. Journal of Ethnopharmacology, 139 (1), 228-233).

[0009] Then the carotenoid-type pigments (yellow, orange or red tetraterpenes), including carotenes, only composed of carbon and hydrogen, and xanthophylls, which additionally have oxygen atoms.

[0010] Chlorophylls (a, b, c1, c2 and d) are pigments present in all green plants (terrestrial and aquatic). Chlorophyll a (C ₅₅ H ₇₂ O ₅ MgN ₄ ) remains the form mainly found in the leaves of plants. [0011] Anthocyanins are water-soluble pigments (oxygenated heterosides) which range from red to blue.

[0012] Curcuminoids (orange pigments from the rhizome of Curcuma tonga) have been shown to significantly decrease the concentrations of C-reactive protein, an important factor in inflammation (Sahebkar, Are Curcuminoids Effective C-Reactive Protein- Lowering Agents in Clinical Practice? Evidence from a Meta-Analysis. Phytother Res. 2013 Aug 7. doi: 10.1002 / ptr.5045).

[0013] As for the flavonoids, they can range from a red to ultra-violet color depending on the pH and correspond to two aromatic rings linked by three carbons.

[0014] These different classes of pigments mainly have effects of regulating inflammation, protecting against light and also act as antioxidants (powerful anti-free radicals) (Stahl and Sies, Bioactivity and protective effects of natural carotenoids Biochimica et Biophysica Acta (BBA) -Molecular Bas / s of D / sease, 1740 (2), 101-107).

The advantage of their use in animal feed is in particular to couple their antioxidant activities with their participation in improving the visual quality (coloring and appearance) of the product formulation, as well as in coloring and conservation of animal products (meat, eggs).

[0016] Terpenes are also interesting secondary metabolites. These are volatile compounds whose structure corresponds to an aromatic ring and which have hydroxyl and terpenoids groups. They are at the origin of the aromatic properties of certain plants according to their taxonomy. According to the literature, there are around 25,000 different structures of terpenes.

[0017] To this are added the properties of another family which are essential components of essential oils: phenols.

[0018] Phenols are the metabolites which attribute to essential oils their very characteristic odor and their biological activities.

[0019] For example, the essential oil of oregano is predominantly composed of thymol (phenol monoterpenoids) and of its isomer, carvacrol and y-terpinene, the presence of which gives the essential oil its antioxidant and antimicrobial properties. [0020] Natural compounds of plant origin have very wide applications in the fields of cosmetics and perfumery, but also of health and human and animal food. Obtaining them first involves harvesting, drying, storing and conditioning the raw plant material. Nevertheless, it is important to note that the extraction processes (solid / liquid) are at the heart of various controversies.

[0021] Essential oils can be obtained by hydrodistillation or steam stripping. Other processes, making it possible to obtain extracts more complex than essential oils, are used such as cold maceration, hot digestion, decoction, leaching, percolation under pressure or cold, or even the infusion...

[0022] The most commonly used process remains the formation of oleoresin by extraction with volatile organic solvents such as petroleum ether, hexane, ethyl ether, acetone, carbon dioxide, benzene or toluene.

[0023] Although these solvents allow a much more efficient and less selective extraction than water (extraction mainly of hydrophilic compounds) and are easily removed at the end of the process by simple evaporation, most of them are:

- toxic, thus contributing to soil and water contamination,

- flammable and

- from non-renewable resources.

[0024] Furthermore, various studies have reported on the risks (carcinogens and mutagens in particular) incurred by prolonged exposure to these solvents.

[0025] In addition, the processes employed require a large amount of solvent and energy, necessary in order to achieve the desired yields.

Recently, European directives have been introduced in order to limit the use of this type of solvent in a global view of environmental protection (fight against the greenhouse effect, soil contamination and against exhaustion. petroleum resources).

In the list of solvents to be limited, hexane is placed at the very top of the scale.

It is indeed a saturated C6 hydrocarbon originating, for its marketing, distillation of petroleum or natural gas. It belongs to the category of type 3 CMR molecules (carcinogenic, mutagenic and reprotoxic; EU CMR list). It is also highly flammable (flash point 23.3 ° C), toxic to aquatic organisms, may damage fertility, causes skin irritation and possibly fatal if swallowed or enters the airways.

[0028] In view of the foregoing, there is today a need to develop or find new solvents or alternative solvents to the controversial solvents listed above.

[0029] Some alternative solvents have already been proposed, and may come from different sectors (wood, cereals, oilseeds). Terpene-type solvents are obtained from the distillation of oleoresins (usually pine) or by-products from other industries.

These are in particular alcohols (C ₆ H ₁₈ O) and hydrocarbons (C ₆ H ₁₀ ) which have numerous double bonds.

[0031] However, there is a brake on the use of these terpene derivatives, in particular because of their chemical reactivity due to their double bonds. For example, limonene exposed to air undergoes auto-oxidation reactions which induce the formation of oxygenated derivatives of the hydro-peroxide type, increasing the allergenic nature of this product and causing the formation of free radicals.

[0032] This is the case in particular with bio-ethanol, obtained by fermentation of sugars which come from the hydrolysis of field crops or by-products. On the same principle are produced butanol and 1, 3-propanediol. They are solvents commonly used in the pharmaceutical, cleaning and cosmetic industries.

Among the agro-solvents, mention may also be made of methyl-THF, a product derived from bio-based furfuraldehyde (from in particular a residue of corn and of sugar cane).

[0034] It appears that the physicochemical characteristics (lipophilic nature, less volatile), the cost and the technological limitations of the production of these alternative agro-solvents remain an obstacle to their use on an industrial scale.

In view of the above, a problem which the present invention proposes to solve consists in developing a new preparation process. a totum or a solid and / or liquid filtrate from a biological matrix or part of a biological matrix and a solvent which does not have the drawbacks listed above. In other words, by the use of a solvent which is in particular non-toxic and respectful of the environment.

The first object of the solution to this problem is a process for preparing a solid and / or liquid totum, which is a mixture of a plant matrix with a solvent, comprising:

a first step of bringing into contact by mixing a plant matrix or part of a plant matrix with a solvent;

at least a second step which is a phase of extraction, solubilization and / or formulation of said mixture; and

a final step of recovering the totum thus obtained;

characterized in that the solvent is glycerol monolaurate.

Preferably, the glycerol monolaurate is used at a temperature above 60 ° C, more preferably still above 63 ° C.

[0037] Its second object is a process for preparing a solid and / or liquid filtrate comprising:

a first step of bringing into contact by mixing a biological matrix or part of a biological matrix with a solvent;

at least a second step which is a phase of extraction, solubilization and / or formulation of said mixture;

a third step of filtering the mixture obtained in the second step; and

a final step of recovering the filtrate thus obtained;

characterized in that the solvent is glycerol monolaurate.

[0038] The third object of the invention is the use of Monolaurate glycerol taken alone or as a mixture with one or more vegetable oils such as sweet almond oil, peanut oil, argon oil, avocado oil, calophyllum oil, safflower oil, rapeseed oil, coconut oil, wheat germ oil, jojoba oil, corn oil , hazelnut oil, apricot kernel oil, virgin olive oil, palm oil, grape seed oil, castor oil, sesame oil, soybean oil, sunflower oil, oleic sunflower oil, non-oleic sunflower oil and hydrogenated sunflower oil, as extraction, solubilization and / or formulation solvent.

Its fourth object is a totum or filtrate capable of being obtained by the process according to the invention.

[0040] Its fifth object is the use of a totum or of a filtrate according to the invention, for the preparation of a food or cosmetic composition.

Finally, its last subject is a composition comprising a totum or a filtrate according to the invention, for its pharmaceutical use.

[0042] The subject of the invention is a process for preparing a totum or a solid and / or liquid filtrate comprising glycerol monolaurate as solvent.

Surprisingly, the Applicant has been able to demonstrate that glycerol monolaurate, uric acid monoglycerol, constitutes a particularly advantageous alternative solvent, the interests of which are multiple. Glycerol monolaurate or 2,3-dihydroxypropyl decanoate corresponds to the following general formula:

[0044] Glycerol monolaurate is a natural fatty acid mono-ester whose composition (more polar than certain vegetable oils) allows an extraction of amphiphilic compounds such as capsaicinoids. Its use does not pose any risk to humans or animals. It is known that its melting point is greater than 60 ° C, more particularly 63 ° C (CAS No. 142-18-7). It is therefore at this temperature that the wary uses it as a solvent.

[0045] In addition, it shows pronounced antibacterial, antifungal and anti-inflammatory activities. It can be used as an antimicrobial agent and in particular inhibits the growth, in vitro but also in vivo, of strains of Candida. It also acts against the growth of Gram + but also Gram- bacteria such as Staphy / ococcus, Streptococcus, Gardnereiia, Haemophi / us but also Listeria monocytogenes. It also acts as a bacteriostatic agent, against Baci / ius anthracis, that is to say it blocks its growth without nevertheless kill cells. In Staphy / ococcus aureus, glycerol monolaurate blocks the production of certain exo-enzymes and virulence factors such as protein A, α-hemolysin, b-lacfamase and toxin 1 responsible for toxic shock syndrome (TSST -1).

[0046] It would likewise reduce the risks of transmission and more precisely inhibit the transduction signal and the inflammatory response due to infection with HIV-1 and SIV.

Its action on the inflammatory cycle has also been demonstrated because a parallel and significant decrease in the amount of pro-inflammatory cytokines (IL-8 and TNF-a) was observed. Glycerol monolaurate can also act in synergy with other products, such as a minog lycosides, in particular in the destruction of biofilm of antibiotic resistant strains of Staphylococcus aureus. Indeed, prior treatment with glycerol monolaurate would improve the response of biofilms to antibiotics.

The Applicant has been able to demonstrate that glycerol is an interesting candidate in order to develop a process for extracting active compounds of natural origin.

For reasons of complexity of implementation, glycerol monolaurate has never been mentioned to replace any extraction solvent in an associated process.

[0050] Indeed, although its physicochemical properties make it an extraction solvent of choice, it is only for its antimicrobial and anti-inflammatory properties that it has been put forward so far.

Glycerol monolaurate is mentioned in particular in international application WO2017181784 as a food additive for laying hens.

[0052] International application WO2016169129 also describes the use of glycerol monolaurate in methods for manufacturing an antibacterial gel based on edible oil. Its antimicrobial properties are also a component of choice in the treatment of viral, fungal and bacterial infections, as described in particular in the international application.

WO2013159029. In food application, glycerol monolaurate is described in a process for treating raw fruits and vegetables. This is an application of 5 seconds to 30 minutes of a solution based on lactic acid and hydrogen peroxide or sodium benzoate or glycerol monolaurate. This process aims to reduce the presence of colonies of food-borne pathogens, such as certain known strains of Escherichia coli. An antimicrobial oil-in-water emulsion comprising glycerol monolaurate is also described in international application WO9531966. This emulsion has been developed for pharmaceutical purposes and shows an inhibitory action against a wide variety of infectious agents (bacteria, fungi, viruses).

As emerges from the above paragraphs, it appears that glycerol monolaurate has been mentioned as a component in mixtures for food decontamination applications, infection treatment processes (in human health) or as food additives. Its antimicrobial properties have been exploited in different fields of application. However, its use as a solvent, in a complete process of extraction, solubilization and / or formulation of plant material containing active natural metabolites of interest in animal supplementation has never been described. Its antimicrobial activity but also its physicochemical properties allowing the extraction of a wide range of metabolites make it a candidate as an intelligent solvent, or solvactive, combining significant efficiency as a solvent and providing the mixture with its antimicrobial properties. and anti inflammatory drugs described in a mont.

The invention and the advantages which result therefrom will be better understood on reading the description and the non-limiting embodiments which follow, illustrated with reference to the appended figures in which:

FIG. 1 represents a diagram of the process according to the invention using glycerol monolaurate (GML) as an innovative solvent in the extraction and / or solubilization and / or formulation from a plant matrix which is a plant or part of a plant.

FIG. 2 is a diagram showing the steps for obtaining active natural compounds according to the processes known from the prior art. FIG. 3 is a diagram showing the steps for obtaining the active natural compounds via the use of glycerol monolaurate (GML) according to the process of the invention.

[0059] FIG. 4 is an image showing a) micronized pepper powder before the process; b) glycerol monolaurate; and c) the cut totum (vegetable matrix of chilli and glycerol monolaurate loaded with metabolites of interest extracted from the chili during the process) obtained by the process according to the invention.

Figure 5 is an image showing a) micronized paprika powder before the process; (a), glycerol monolaurate (b), and c) the cut totum (chilli plant matrix and monolaurate glycerol loaded with metabolites of interest extracted from the chilli during the process) obtained by the process according to the invention.

The invention relates to a process for preparing a totum or a solid and / or liquid filtrate comprising a first step of bringing into contact by mixing a biological matrix or part of a biological matrix with a solvent. which is glycerol monolaurate as described above.

During the steps of solubilization and extraction of said process, the plant matrix, which may be in the form of powder, is brought into contact with the glycerol monolaurate and is heated. The duration and the heating temperature are optimized according to the plant matrix. Preferably, the monolaurate glycerol is used at a temperature above 60 ° C, more preferably still above 63 ° C, more preferably still above 80 ° C. The totum corresponds to the sum / total of

1) glycerol monolaurate having been loaded with active compounds extracted from the plant matrix (after extraction / solubilization)

2) the plant matrix depleted in certain active compounds that it contained and which were transferred to the solvent (glycerol monolaurate) during the extraction.

The totum forms a single mass containing as a mixture the products 1) and 2) resulting from said process. The therapeutic effect and the biological activities of the whole plant are superior to that of one of its constituents (a coherent set of active principles with synergistic actions). It must be considered that the therapeutic effect and the biological activities of the totum are even more important given that the active molecules of the plant matrix have been made more available by extraction, that the biological activities of the intelligent solvent, glycerol monolaurate, are added, or act in synergy with those of the active ingredients extracted from the plant. It should also be considered that the plant matrix, in which certain molecules of interest in terms of activities have remained trapped as in any extraction, is preserved in the totum.

The filtrate which is the subject of the invention is the term used to designate the liquid materials which have passed through a filter. As described above, therefore, it is 1) glycerol monolaurate loaded with active compounds extracted from the plant matrix. It is therefore the substance obtained after the totum has undergone a filtration step, allowing it to be freed from the powder of biological matrix or part of the biological matrix.

The first step of the method which is the subject of the invention consists in bringing the solvent which is glycerol monolaurate into contact, by mixing, with a biological matrix or part of a biological matrix.

[0066] The biological matrix or part of the biological matrix is preferably in powder form, that is to say in the form of a powder of dry matter.

Preferably, the biological matrix or the part of the biological matrix is, taken alone or as a mixture, chosen from wormwood, yarrow, garlic, artemisia, artichoke, basil, chamomile, cinnamon, lemon, cilantro, turmeric, cypress, eucalyptus, fenugreek, ash (leaves), juniper, cloves, bay leaf, lavender, lemon grass, alfalfa, peppermint, white mustard, walnut (leaves), orange, oregano, nettle, paprika, chili, pine, dandelion, pepper, rosemary, savory, sage, wild thyme, tansy, thyme, clover, goldenrod, or microorganisms or media derived from culture of microorganisms.

[0068] More preferably, the biological material is a plant material. Plant material is a plant or part of a plant such as a leaf, a fruit, a stem, a flower or even a root. By way of nonlimiting example of plant material or part of plant material that can be used, there may be mentioned, taken alone or as a mixture, wormwood, yarrow, garlic, artemisia, artichoke, basil, chamomile, cinnamon, lemon, cilantro, turmeric, cypress, eucalyptus, fenugreek, ash (leaves), juniper, cloves, bay leaf, lavender, lemon grass, alfalfa, peppermint, white mustard, walnut (leaves), orange, oregano, nettle, paprika, chili, pine, dandelion, pepper, rosemary, savory, sage, wild thyme, tansy, thyme, clover or even goldenrod.

Preferably, the biological matrix or part of the biological matrix is a plant material or a portion of plant material chosen from chilli, paprika, oregano and rosemary, taken alone or as a mixture.

As illustrated in Figure 1, preferably, the method according to the invention comprises a step, prior to the first step described above, of drying and / or grinding of the biological matrix or part of the biological matrix. , which is advantageously a plant matrix or part of a plant matrix as described above.

The method of the invention comprises a second step which is a phase of extraction, solubilization and / or formation of the mixture between the biological matrix or part of the biological matrix and the solvent.

As illustrated in FIG. 1, the Applicant has been able to demonstrate that the solvent which is glycerol monolaurate could be used in order to carry out three phases essential to obtaining active natural compounds from the matrix. biological or part of biological matrix. These are the extraction, solubilization and / or formulation phases. Glycerol monolaurate can be used to achieve these three phases in series, only two of these three phases or a single phase independently of the others.

[0073] According to a particular embodiment of the invention, the second step of the process according to the invention comprises at least two of the three phases chosen from the phases of extraction, solubilization and / or formulation of said mixture. Preferably, the two phases are extraction and solubilization or extraction and formulation.

[0074] The extraction step consists of separating elements (molecules) from a liquid (liquid / liquid extraction) or solid (solid / liquid extraction) matrix, most often using a solvent.

Conventional extraction methods vary depending on the desired composition of the extract: without solvent of press type for oils or juices, with solvents selected according to their chemical properties (polarity, etc.), their cost, their dangerousness. The classic example is extraction with hexane or ethanol with the intention of producing oleoresins. The use of toxic or harmful solvents makes the evaporation step mandatory. According to the invention, eco-extraction with glycerol monolaurate makes it possible to preserve the solvent, which is not harmful and with biological activities of interest, in the totum or the filtrate resulting from said process. Extractions by supercritical CO2 or by steam training (in the case of essential oils) are also commonly used. Extraction can be assisted by the use of ultrasound or microwaves.

[0076] The solubilization step makes it possible to make the active compounds of interest, previously extracted and taken out of the matrix, soluble in the extraction solvent, in other words in this case, glycerol monolaurate. This is facilitated by heat, stirring and is highly dependent on the chemical properties of the molecules extracted and the solvent used.

The formulation consists of homogeneously shaping the product resulting from the extraction and solubilization steps and containing the active ingredients, in order to target its field of use. Ultimately, it makes it possible to develop a product that can be marketed.

[0078] According to a preferred embodiment of the invention, the second step comprises the three phases of extraction, solubilization and formulation of said mixture.

As emerges from Figure 3, the second step of the process according to the invention corresponds to the steps of extraction, organic solubilization and formulation of the usual processes for the production of additives as illustrated in Figure 2. With the process according to the invention, no co-product, no waste, originating from the plant material itself or from the solvent is generated.

[0080] One of the advantages of using this solvent in this process lies in the direct 3-phase-in-1 extraction / solubilization / formulation procedure. As indicated above, each phase (extraction, solubilization or formulation) can be carried out independently of the other phases using monoglycerol laurate as solvent.

Likewise, only two of these steps out of three can be carried out using said solvent. The invention relates firstly to the use of glycerol monolaurate as a solvent. In addition, the Applicant has, despite the difficulty of implementation, thought of the process associated with the use of said solvent. The major obstacles unlocked by the Applicant were to implement the process, to destroy any degradation of the compounds extracted by the glycerol monolaurate and to make the extraction / solubilization and formulation efficient in terms of yield.

[0082] The method according to the invention optionally comprises a third step of filtering the mixture obtained in the second step. This filtration step makes it possible to recover, in the final step, a final product free of biological or vegetable powder and therefore only a loaded solvent which is a filtrate.

Alternatively, the method does not include this filtration step. The product thus recovered is called totum.

The entire product generated by the process, whether it is the totum or the filtrate, can be used in human or animal feed, preferably in animal feed.

It should be noted that the basic materials used to carry out said process are biological or plant material and the new proposed solvent: glycerol monolaurate. Before carrying out the process, the plant active agents (metabolites recognized for their biological activities) are contained and retained by the biological or plant matrix (not or partially bioavailable). Said process including the steps of extracting and solubilizing the plant active ingredients makes it possible to transfer these metabolites from the plant matrix to the glycerol monolaurate. The extraction and transfer into glycerol monolaurate increases the bioavailability of these active ingredients and their absorption during their transit in animals.

The different places of absorption can be modified thanks to the formulation included in said process, tn fine, the filtrate (glycerol monolaurate loaded with plant active ingredients) or the totum, corresponding to the remainder of plant material (which may have retained a low part of its plant active ingredients, whether available or trapped in the matrix) and with glycerol monolaurate highly loaded with active metabolites extracted from the plant matrix and made bioavailable, may be used.

Preferably, the method which is the subject of the invention comprises the following steps of: bringing the complex biological or plant matrix (dry matter powder) into contact with the solvent (100% solid glycerol monolaurate); vacuum (around 700 mbar to avoid oxidation of the metabolites of interest) or not, stirring, heating of the plant material and the solvent (depending on the compounds of interest, which can range from around 80 ° C) at approximately 180 ° C) and this for a period determined according to the plant matrix (approximately 20 minutes to approximately 60 minutes);

recovery of the solidified totum, or filtration in order to use only the filtrate which is the loaded solvent.

Advantageously, the method according to the invention is easy to implement because it is limited in production steps, which therefore limits the time constraints.

The method according to the invention also makes it possible to avoid logistics and the use of complex tools. Eliminating certain conventional steps in the extraction of plant active ingredients also limits the handling of equipment and the cleaning of machines, and is once again part of an ecological issue and respect for the environment. The equipment, solvent and materials necessary for the realization of the final product are inexpensive and non-hazardous for the handler, and adaptable in terms of volume.

A subject of the invention is also the use of an intelligent solvent, said to be solvactive, glycerol monolaurate, in a single, 3-step-in-l process of extraction, solubilization and formulation of natural compounds active in from plant matrices.

[0091] Glycerol monolaurate can advantageously be mixed with vegetable oils while not drastically modifying the extraction yields but by adding interesting properties to the totum obtained.

More particularly, the subject of the invention is the use of Monolaurate glycerol taken alone or as a mixture with one or more vegetable oils such as sweet almond oil, peanut oil, oil. argan oil, avocado oil, calophyllum oil, safflower oil, rapeseed oil, coconut oil, wheat germ oil, jojoba oil, l corn oil, hazelnut oil, apricot kernel oil, virgin olive oil, palm oil, grape seed oil, castor oil, sesame oil, soybean oil, sunflower oil, oil oleic sunflower oil, non-oleic sunflower oil and hydrogenated sunflower oil, as extraction, solubilization and / or formulation solvent.

The Applicant has been able to demonstrate that glycerol monolaurate could be used as a solvent with a broad extraction spectrum (polar compounds, polar α, amphiphilic compounds).

The properties of totum (solvent + plant matrix whose active compounds have been extracted and solubilized) can be multiple: antimicrobial, anti-inflammatory, antioxidant, nutritional, fragrance, flavoring or coloring additions.

Glycerol monolaurate can transmit to the product resulting from said process (in particular the filtrate) its emulsifying properties, a non-negligible benefit in animal nutrition and supplementation because it can in particular increase the growth performance or even the quality of the flesh in animals. 'breeding.

The filtrate (loaded solvent) or the totum can be, active material and energized by the solvactive which is glycerol monolaurate, used as additives in sectors such as food or human or animal health.

Said invention proposes the use of glycerol monolaurate as a new solvent which can, in addition to being used for the stages of extraction / solubilization / formulation of a plant matrix, be ingested by animals and humans. and bring its own properties of interest to the final product.

Advantageously, the method based on the use of glycerol monolaurate falls within the current problems of protection and respect for the environment. The ultimate advantage of these eco-extraction processes being to replace CMR risk solvents such as hexane.

Among the active compounds of interest and targeted, that is to say preferred, during the application of said method, there may be mentioned a family of alkaloids: capsaicinoids (capsaicins, dihydrocapsaicins, nordihydrocapsaicins).

By the method according to the invention, it is also possible to extract pigments of the carotenoid type (capsanthin, capsorubin, zeaxanthin, b-carotene, b-cryptoxanthin, b-cryptoxanthin, antheraxanthin) and chlorophylls. The temperature of the process can be adapted in order to extract and not to deteriorate molecules of aromatic and volatile types such as p-cymene, g-terpinene, a-pinene, 1, 8-cineole, cis-sabinene hydrate, linalool, camphor, bounded I, terpinen-4-ol, trans-p-mentha-l (7), 8-dien-2-ol, verbenone, bornylacetate, a-terpineol, carvone, thymol, carvacrol, piperitenone, eugenol, a-ylangene, carvacrol acetate, methyl-eugenol, caryophyllene, a-humulene, cis-calamenene, a-calacorene, caryophyllene oxide, 14-hydroxy- (Z) caryophyllene, abetatriene,

14-hydroxy-9-epi- (E) caryophyllene.

[0102] Furthermore, all of the extracted compounds can act in synergy with each other and with the solvactive (intelligent solvent) which is glycerol monolaurate, depending on their properties.

Given the physicochemical properties of glycerol monolaurate, a wide range of molecules (polar, polar, amphiphilic) can theoretically be extracted in the solvent and contribute to the biological activity of the totum. By way of nonlimiting example, mention may be made, among other compounds, of fatty acids, vitamins, oses or even amino acids.

[0104] The subject of the invention is also a totum or a filtrate, capable of being obtained by the process according to the invention.

[0105] The final product, filtrate or totum, as soon as it returns to a temperature equivalent to room temperature, generally solidifies and hardens.

The totum or the filtrate have, due to the presence of glycerol monolaurate, but also various pigments extracted during the process, a smooth, colored and shiny appearance.

[0107] In the event that the extracted plant material comes from an aromatic plant (rosemary or oregano to name just two), the totum could likewise have the characteristic odor of these plants. The Applicant has defined the firm supply of properties of glycerol monolaurate.

[0108] In this invention, the Applicant not only updates the use of a new intelligent solvent, known as a solvent, and its advantages for food and animal health applications, but likewise demonstrates the inference of the 3-to-3 process. 1 -steps of organic extraction / solubilization / formulation. The method is adaptable in terms of temperature, duration of contact with the vegetal matrix / solvent, and of volume. This makes it possible to extract a certain palette of active natural compounds from a wide variety of plant species. Heat-sensitive compounds such as aromatic compounds of terpene type can thus be extracted and integrated into the filtrate or the totum. The proposed new solvent is likewise thermostable and retains its properties and non-toxicity during the process steps.

[01 10] A subject of the invention is also the use of a totum or of a filtrate according to the invention, for the preparation of a food or cosmetic composition. Thus, the loaded filtrate or the totum can then be formulated according to the needs and the targeted animals and proposed for example for their different properties as feed additives for production animals. By way of nonlimiting examples, the charged filtrate or the totum is preferably in the form of powder, granule, pebble, ointment, paste, capsule, microcapsule or tablet.

[01 1 1] Finally, the last subject of the invention is a composition comprising a totum or a filtrate according to the invention, for its pharmaceutical use.

Examples

[0001] The present invention will now be illustrated by means of the following examples:

Example 1: Implementation of a process according to the invention:

[0002] The plant matrix in the form of dry material is first ground.

About 20g of plant material and about 20g of glycerol monolaurate (1/1) are inserted into a balloon, preferably a ground neck balloon.

The flask is installed on a rotary evaporator, and its contents are preferably protected from light.

[0005] A low rotation (less than 500 rpm, preferably less than 200 rpm) is then applied to the balloon.

[0006] The pressure inside the balloon is preferably fixed:

from 700 mbar, for compounds sensitive to oxidation, to 1000 mbar for aromatic compounds in particular.

The flask is then immersed in a heating bath, the temperature of which is between 80 ° C and 180 ° C, depending in particular: the solubility of the active metabolites to be extracted;

the thermal sensitivity of these metabolites;

of the duration defined to apply the process.

[0008] The heating and stirring time depends mainly on the plant material and the heating temperature. It can be set between 20 minutes and 60 minutes.

[0009] In order to increase the efficiency in terms of yield, time and energy required, an optimum temperature and duration can be defined specifically for each plant material depending in particular on the targeted active metabolites.

Example 2: Application of the method according to the invention to a plant matrix of chilli pepper (Capsicum) and quantitative analysis of the capsaicinoids transferred into the filtrate obtained.

As illustrated in Figure 4, the examination of the totum obtained makes it possible to determine that the product resulting from the process according to the invention has a shiny and smooth appearance, a deep red color and a non-grainy and malleable texture. hot.

[001 1] This totum can be formulated quite easily depending on the target animal and the end use, especially when making a premix.

[0012] As described above, the family preferentially targeted during the application of said method to a plant material such as chili pepper is a family of alkaloids: capsaicinoids.

[0013] The analyzes show that the micronized pepper powder used in said process contains 121 1 mg / 100 g of total capsaicinoids, divided into three main molecules: nordihydrocapsaicin, capsaicin, and dihydroca psaïci ne.

As emerges from Table 1 below, this is equivalent to approximately 1, 2% of capsaicinoids in the plant matrix.

On average, 1017.3 mg / 100 g of filtrate were extracted and transferred into the filtrate, that is to say 84% of the capsaicinoids of the plant matrix in terms of mass / mass proportion. [0016] It should be noted that the residue analyzed corresponds to the starting plant matrix solidified in a fraction of solidified glycerol monolaurate, loaded with extracted inferet compounds and which could not be filtered.

[0017] The advantage of the process is in particular that it can conserve:

the filtrate loaded with metabolites of interest; or

the totum containing the residual powder in which a portion of the active compounds of interest remains and said solvent having extracted most of the targeted active compounds and making them bioavailable upon ingestion.

[0018] The biological interest of capsaicinoids in animal supplementation is significant given that they have a panel of properties recognized in the literature. As can be seen from the table below, capsaicin (8-methyl-N-vanillyl-6-nonenamide) represents about 45% of total capsaicinoids, 43.6% in the present study.

[0019] It can in particular be used commercially in the preparation of cosmetic products or "hot" ointments for its antimicrobial and pungent properties.

[0020] Pepper extracts are used in food supplements in the context of specific diets and the anti-inflammatory activity of capsaicin is used in pharmaceutical creams.

[0021] In agronomy, resistance to certain fungal diseases seems to be correlated with the capsaicinoid content.

[0022] Used on other crops, capsaicinoids have antimicrobial and antifungal effects, in particular by inducing the stimulation of the synthesis of enzymes involved in the defense of plants such as chitinases.

Table 1: Capsaicinoid content (n ord i hyd roca psaïci ne, capsaicin, di hyd roca psaïci ne) of the micronized pepper powder, of the filtrates and of the filtration residues obtained by said process.

The results are expressed as the mean ± standard deviation (n = 3). The residue corresponds to the starting plant matrix solidified in the solidified solvent, which could not be filtered. It therefore constitutes a fraction of vegetable powder and a fraction of solvent loaded with extracted compounds.

The chemical analyzes in order to assay the capsaicinoids were carried out by UPLC (ACQUITY U P LC® (Waters) - C18 BEH column (2.1x50mm)). UV defection was made specifically at 280 nm and coupling with a mass spectrometer made it possible to identify and measure capsaicin, dihydrocapsaicin, and n ord i hyd roca psaïci ne (thanks to the passage of standards), respectively . Carotenoid assay was performed by UPLC and UV detection specifically at 470 nm. The coupling to mass spectrometry is carried out with the spectrometer described above equipped with an ICPA source (Chemical Ionization at Atmospheric Pressure). The aromatic compounds were assayed for its part by GC-MS (Agilent 7890A and 7000A, Santa Clara, USA) with a 60-meter DB-5 MS type column.

Example 3: COSMO-RS simulation and physical properties of the solvent

[0025] Solubilization studies of various compounds were carried out by simulation using the COSMO-RS software to predict the solubilization of the active substances in hexane or in glycerol monolaurate.

[0026] It appears that glycerol monolaurate extracts the various compounds better when it is heated to temperatures above 100 ° C.

[0027] For the study of aromatic compounds, a study at 80 ° C was carried out in an attempt to limit the degradation of the latter.

As emerges from Table 2 below, it can be observed that overall the glycerol monolaurate extracted as well or even better than hexane. Table 2:

Example 4: Application of said process to a paprika veal matrix

(Capsicum) and quantitative analysis of the carotenoids transferred to the filtrate

As emerges from Figure 5, the examination of the totum obtained allows to determine that the product resulting from said process has a dark orange color, a shiny and smooth appearance and a non-grainy and malleable texture when hot.

[0030] Just like the totum obtained during the application of the process with the pepper, the paprika totum can be transformed quite easily depending on the target animal and the demand when making a premix.

Table 3 below details the total content of free carotenoids after saponification of the samples (in mg / g). The results are expressed as mean ± standard deviation (n = 3).

Table 3:

[0032] It emerges from this example that the carotenoid profile of the paprika extracts shows the presence of carotenoid esters in the form of monoesters and diesters.

A small amount of carotenoids in free form is also identified in these extracts. Saponification is carried out in order to be able to quantify the carotenoids. The major neffenoid carotenoid in paprika is capsanfhin, which accounts for nearly 56% of the total carotenoids identified in paprika powder.

The other identified carotenoids are antheraxanthin (1 1, 8%), capsorubin (1 1, 1%), zeaxanthin (9.7%), b-cryptoxanthin (3.3%), and b-carotene (8.5%).

In the filtrate obtained by said method, 63.5% of the total carotenoids were extracted and transferred and 51% concerning the major carotenoid, capsanthin.

As in the example above for the pepper, it should be noted that the residue analyzed corresponds to the starting plant matrix taken up in a fraction of solidified glycerol monolaurate, loaded with compounds of interest extracted and n ' not having been filtered.

[0038] Thus, it appears that the conservation of the totum at the end of the process can be of significant interest.

[0039] Carotenoids can be used as a coloring agent for flesh and by-products (eggs) in animal nutrition, which in particular represents an advantage for consumers. Furthermore, carotenoids are powerful antioxidants that have the ability to absorb aggressive blue radiation and prevent the production of ROS (Reactive Oxygen Species) and therefore the associated cell damage.

This protective activity of plant cells is linked to their structure (double bonds, keto group and sometimes cycle with 5 centers).

[0041] The presence of these compounds is therefore advantageously targeted in said process.

Example 5: Application of said process to a plant matrix of oriaan and qualitative analysis of the aromatic compounds transferred to the filtrate

The method described in Example 1 is applied to a plant matrix of oregano.

Among all the metabolites derived from oregano extracted and identified in the hexane extract (= 100%), 63.5% metabolites were also identified in the essential oil of oregano. There is therefore a qualitative recovery of 63.5% between the essential oil of oregano and the extract of oregano with a hexane-type solvent. Among all the metabolites derived from oregano extracted and identified in the hexane extract (= 100%), 34.1% of the compounds present were likewise identified in the filtrate obtained during said process. 34.1% of the metabolites extracted in a hexane extract (transferred from the matrix to hexane) were transferred from the plant matrix of oregano to the glycerol monolaurate. Among them, the metabolites having a biological activity of interest. The rest of the metabolites are kept in the 'depleted' powder, which is present in the totum.

As emerges from Table 4 below, it is in particular p-cymene, y-terpinene, cis-sabinene hydrate, linalool, borneol, terpinen-4-ol, a-terpineol, carvone, thymol, carvacrol, eugenol, carvacrol acetate, caryophyllene, a-humulene, caryophyllene oxide, 14-hydroxy- (Z) caryophyllene and abetatriene.

[0046] Carvacrol is the major compound in the essential oil of this species of rosemary and is the compound most transferred to the filtrate.

[0047] It is, along with thymol, one of the compounds which participates mainly in the biological antioxidant and antimicrobial activities of essential oil of oregano.

[0048] Other compounds, including glycerol monolaurate, have been specifically annotated in the filtrate. They participate in the biological activity of the † o † ume \ tont the interest of said process in energizing the totum.

Regarding the compounds which have not been annotated in the filtrate and have been annotated in the hexane extract, two possibilities are to be considered:

or they are not transferred to the filtrate

thirsty than they are, but in too small a quantity, compounds that would therefore be below the defection limit of the mass spectrometer.

Table 4 below shows the qualitative aspect of the transfer of aromatic compounds from a plant matrix of oregano in the filtrate of said process (n = 3) in comparison with a hexanic extract of the plant matrix of rosemary and an essential oil of the same plant material.

In this table 4, the compounds denoted fhe, he, h and f respectively are compounds not annotated using the RI and the ADAMS, found in the filtrate, the essential oil and the hexane extract (fhel -1 1), in the essential oil and the hexane extract of rosemary (hel -4), specifically in the hexane extract (h 1-25) and specifically in the filtrate (f1 -33). The “xs” represent the qualitative presence of the compound in the extract and the “-” their absence.

Table 4:

Example 8 Application of said process to a rosemary plant matrix and qualitative analysis of the aromatic compounds transferred into the filtrate

Examination of the filtrate obtained is conclusive given that the filtrate retained the specific aromas of rosemary (olfactory examination). Likewise, it is smooth and shiny and characteristic green of the plant matrix of rosemary. It is strong enough to be formulated later (mix) but also fatty enough to be easily ingested by livestock.

Regarding the application of said process on rosemary, the study carried out on the aromatic compounds transferred into the filtrate was carried out qualitatively (the measurement corresponds to the number of molecules identified and not to their quantity). Among all the metabolites from rosemary extracted and identified in the hexane extract (= 100%), 25% of the compounds present were also identified in the filtrate obtained during said process. These are the compounds that are predominantly present (in terms of concentration) in the hexane extract and in the plant matrix and which are responsible for the biological activity of the plant. The rest of the metabolites are stored in the 'depleted' powder, which is present in the totum.

As emerges from Table 5 below, it was advantageously possible to find in the filtrate: α-pinene, 1, 8-cineole, linalool, camphor, borneol, terpinen-4-ol, a -terpineol, trans-p-mentha-l (7), 8-dien-2-ol, verbenone, bornyl acetate, piperitenone, eugenol, a-caryophyllene, a-humulene, cis-calamenene, a-calacorene, 14-hydroxy -9-epi- (E) -caryophyllene, hexadecanoic acid.

[0055] Regarding the compounds which have not been annotated, two possibilities are to be considered, as well as for oregano:

or they are not transferred to the filtrate

or in too small a quantity in the filtrate, compounds which would therefore be below the detection limit of the mass spectrum meter.

Table 5 below shows the qualitative aspect of the transfer of aromatic compounds from a plant matrix of rosemary into the filtrate of said process (n = 3) in comparison with a hexanic extract of the plant matrix of rosemary. The compounds denoted fh, h, and f respectively are compounds not annotated using RI and ADAMS, found in the filtrate and the hexane extract (fhl -16), specifically in the hexane extract ( h 1-75) and specifically in the filtrate (fl - 29). The “xs” represent the qualitative presence of the compound in the extract in question and the “-” their absence.

Table 5:

Example 9: Use of alvcerol monolaurate mixed with a vegetable oil and application of said process for the extraction, solubilization and formulation of a paprika powder

[0058] The efficiency in terms of β-carotene yield of mixtures of glycerol monolaurate and hydrogenated sunflower oil was tested by applying the scheme of said process. An experimental design involving different temperatures applied to said process and different glycerol monolaurate / hydrogenated sunflower oil ratios made it possible to evaluate the determining criteria in the efficiency of the process.

As can be seen from Table 6, the temperature is in fact a parameter that significantly modulates (p <0.005, ANOVA) the extraction yield of b-carotene, in particular from paprika powder (Table 6).

Conversely, modulating the percentage of glycerol monolaurate mixed with hydrogenated sunflower oil does not significantly modify the extraction yield of said process (p> 0.1, ANOVA).

It is therefore possible to use glycerol monolaurate, the new extraction / solubilization / formulation solvent proposed by the Applicant, as a mixture with a vegetable oil which can likewise provide its biological properties to the filtrate or to the totum obtained by said method.

Table 5 below is an ANOVA of the experimental design of said method set up in order to evaluate the possibility of using glycerol monolaurate (GML) mixed with a vegetable oil (hydrogenated sunflower oil ) for the extraction / solubilization / formulation of paprika.

Table 6:

Example 10: Use of alvercerol monolaurate and application of said method to microwave technology

A microwave-assisted heating and extraction experiment demonstrated that the monolaurate glycerol was made liquid by microwave heating and that the compounds of the vegetable raw material, that this thirst for chili, paprika, oregano or rosemary, could be extracted / solubilized and formulated using this eco-extraction technique and integrating it with said process.

[0064] The Applicant has observed that this technology saves time and energy for a similar quality of the products obtained.

Example 1 1: Use of alvercerol monolaurate and application of said method to ultrasound technology

Likewise, an ultrasound-assisted extraction experiment demonstrated that said process using glycerol monolaurate could integrate ultrasound-assisted extraction technology for the extraction / solubilization / formulation of a plant matrix of chilli. , paprika, oregano or rosemary.

[0066] The integration of this eco-extraction technology makes it possible to reduce the time required for said process and therefore to reduce the energy costs of said process at the same time.

Claims

Claims

[Claim 1] [A process for preparing a solid and / or liquid totum which is a mixture of a plant matrix with a solvent, comprising: a first step of bringing into contact by mixing a plant matrix or part of a matrix vegetable with a solvent;

at least a second step which is an extraction, solubilization and / or formulation phase of said mixture; and

a final step of recovering the totum thus obtained;

characterized in that the solvent is glycerol monolaurate.

[Claim 2] Process for preparing a solid and / or liquid filtrate comprising:

a first step of bringing into contact by mixing a biological matrix or part of a biological matrix with a solvent;

at least a second step which is a phase of extraction, solubilization and / or formulation of said mixture;

a third step of filtering the mixture obtained in the second step; and

a final step of recovering the filtrate thus obtained;

characterized in that the solvent is glycerol monolaurate.

[Claim 3] Method according to one of claims 1 or 2, characterized in that it comprises a prior step of drying and / or grinding the biological or plant matrix, part of the biological or plant matrix.

[Claim 4] Method according to one of the preceding claims, characterized in that the biological matrix, plant, part of biological or plant matrix is, taken alone or as a mixture, chosen from wormwood, yarrow, l ' garlic, artemisia, artichoke, basil, chamomile, cinnamon, lemon, coriander, turmeric, cypress, eucalyptus, fenugreek, ash (leaves), juniper, cloves, bay leaf, lavender, lemon grass, alfalfa, peppermint, white mustard, walnut (leaves), orange, oregano, nettle, paprika, chili, pine, dandelion, pepper, rosemary, savory, sage, wild thyme, tansy, thyme, clover, goldenrod, or also microorganisms or media derived from culture of microorganisms, preferably the biological matrix or part of biological matrix is chosen from chilli, paprika, oregano and rosemary.

[Claim 5] Method according to one of the preceding claims, characterized in that the second step comprises at least two of the phases chosen from the phases of extraction, solubilization and / or formulation of said mixture.

[Claim 6] A method according to claim 4, characterized in that the second step comprises three phases of extraction, solubilization and formulation of said mixture.

[Claim 7] Use of Monolaurate Glycerol taken alone or as a mixture with one or more vegetable oils such as sweet almond oil, peanut oil, argan oil, avocado oil , calophyllum oil, safflower oil, rapeseed oil, coconut oil, wheat germ oil, jojoba oil, corn oil, hazelnut oil , apricot kernel oil, virgin olive oil, palm oil, grape seed oil, castor oil, sesame oil, soybean oil, sunflower oil, oleic sunflower oil, non-oleic sunflower oil and hydrogenated sunflower oil, as an extraction, solubilization and / or formulation solvent.

[Claim 8] Totum or filtrate obtainable by the process according to one of claims 1 to 6.

[Claim 9] Use of a totum or a filtrate according to claim

8, for the preparation of a food or cosmetic composition.

[Claim 10] A composition comprising a totum or a filtrate according to claim 8, for its pharmaceutical use. |