Formulation of microparticipates based on polyphenolic compounds for the removal of free radicals in polluted air and smoke

Description

[0001] The invention relates to the field of free radical scavengers. More particularly, the invention relates to microparticles associated with polyphenolic compounds capable of capturing free radicals present in polluted air or fumes, in particular smoke from cigarettes containing tobacco, CBD and / or cannabis or mixtures thereof. Field of invention

[0002] The present invention relates to the use of polyphenolic compounds or their derivatives as free radical scavengers in devices for reducing the harmful effects of air pollution, smoke generated by the combustion of tobacco, smoke generated by the combustion of cannabis (hereinafter cannabis for medical use), or smoke generated by the combustion of CBD or a mixture of these fumes, or a mixture of these fumes and air pollution. Such use is described in patent EP1041899B1.

[0003] Many polyphenolic compounds are known for their beneficial properties in areas as diverse as hypertension, hypercholesterolemia, cardiovascular disease, viral infections, and inflammatory phenomena. The anti-lipoperoxidant and anti-carcinogenic activities of certain polyphenols have also been described.

[0004] In particular, epigallocatechin-3-gallate (EGCG), the main polyphenol present in green tea, has been shown to possess potent anti-inflammatory and antiproliferative activity capable of selectively inhibiting cell growth and inducing apoptosis in cancer cells without harming normal cells (DN Syed et al. Green tea polyphenol EGCG suppresses cigarette smoke condensate-induced NF-κB activation in normal human bronchial epithelial cells Oncogene Volume 26, pages673-682 (2007) doi.org / 10.1038 / sj.onc.1209829).

[0005] However, to date, the use of polyphenolic compounds as a means of capturing free radicals present in cigarette smoke and more generally in the air, and likely to be inhaled, has been very little studied and must be improved to be exploitable at the industrial level. CN 108 338 406 B describes antioxidant pearls consisting of liposoluble and soluble rosemary antioxidant and includes in particular essential oils and soluble oils. Statement of the invention

[0006] The authors of the present invention have now discovered that the incorporation of polyphenols or their derivatives into microparticles makes it possible to effectively eliminate free radicals from cytotoxic molecules in smoke from tobacco, cannabis (for medical purposes), CBD, air pollution or their mixtures. These microparticles are incorporated into a cigarette filter or onto a non-woven fabric (extruded blown, "meltblown"). The mixture of polyphenolic compounds formulated into microparticles makes it possible to selectively absorb free radicals from the gas phase and the semi-liquid phase (tars) as they pass through the filter or through the filter fabric.

[0007] The present invention therefore relates to the use of polyphenolic compounds formulated in microparticles as scavengers of free radicals. It relates in particular to a formulation of microparticles associated with polyphenolic compounds and their derivatives, in particular, in the form of a rosemary extract, in which the microparticles consist of a mixture of 2 to 4 saturated vegetable oils whose melting temperatures differ by at least 10°C.

[0008] This formulation can be used as free radical scavengers in tobacco cigarette filters, CBD cigarette filters, cannabis cigarette filters, or cigarette filters made from their blend and air filter fabric, especially in a mask.

[0009] The invention also relates to a method for preparing a cigarette filter and the filter obtained, as well as to a method for preparing a filter fabric and the fabric obtained. Advantages of the invention

[0010] The formulation of the polyphenolic compounds according to the invention has two main advantages.

[0011] On the one hand, this formulation based on at least 2 vegetable oils creates a surface effect that optimizes the anti-radical effectiveness of the polyphenolic compounds. After heating the oil / polyphenolic compound mixture and cooling it, the polyphenolic compounds are pushed towards the outside of the particle and are thus exposed, in contact with the air, so as to be able to interact with the free radicals. These phenolic compounds capture the free radicals, in particular the OH° radicals.

[0012] This composition has a neutralizing effect on the effect of tars due to the contacts established on the surface of the particles. In the case of cigarette smoke, tars compete with phenolic compounds to interact with free radicals. These free radicals in the semi-liquid phase are also trapped by polyphenolic compounds like those in the gaseous phase; this effect has been demonstrated in particular with a rosemary extract rich in polyphenols.

[0013] Furthermore, this approach provides particles whose size avoids the risk of ingestion, whether that of the particles present in the filter or on the filter fabrics. To this end, the particle size is adapted to the chosen application. The particles must be large enough to be retained on their support (filter or fabric), while taking into account the constraints linked to the preparation of the products and the temperature of use.

[0014] Cigarette filters can be used for the consumption of tobacco and / or cannabis, particularly for therapeutic use, or CBD to capture free radicals and thus counter their neurodegenerative effects.

[0015] Filter fabrics are particularly suitable for making anti-pollution masks that can be used in any environment polluted by fine particles such as tar, particularly for asthmatics, cyclists and athletes in urban environments. DETAILED DESCRIPTION OF THE INVENTION

[0016] A first subject of the invention concerns the formulation of microparticles associated with polyphenolic compounds and their derivatives, in particular in the form of a rosemary extract in which the microparticles consist of a mixture of 2 to 4 saturated vegetable oils whose melting temperatures differ by at least 10°C.

[0017] By "derivatives" is meant in particular compounds derived from polyphenolic compounds by substitution of the hydrogen atom of at least one of the hydroxy groups of the polyphenolic compounds by a C1-C6 alkyl group, or a (C1-C4 alkyl)carbonyl group. Acetates such as carnosic acid acetates and rosmarinic acid acetates are preferred derivatives of the polyphenolic compounds used in accordance with the invention.

[0018] On the other hand, the polyphenol derivatives used in accordance with the invention such as carnosol, rosmanol, rosmarinic acid, carnosic acid may correspond to the isomers of said polyphenols such as in particular epirosmanol and isorosmanol (Nakatani et al., Agric. Biol. Chem., 1984, vol. 48, n° 8, pp 2081-2085, https: / / doi.org / 10.1080 / 00021369.1984.10866436).

[0019] These compounds can be obtained by conventional chemical synthesis or by biotechnological means, according to methods known to those skilled in the art. They can also be isolated from plant extracts.

[0020] In a preferred embodiment of the invention, the polyphenols used are provided in the form of a rosemary extract ( Rosmarinus officinalis L .). Such a plant extract can be obtained by extraction with a polar solvent such as an alcoholic or hydro-alcoholic solvent. The alcohol used as solvent can be ethanol in particular. This extract can also be advantageously obtained using supercritical carbon dioxide and is then richer in polyphenolic compounds. Such a plant extract can be obtained by extraction with a polar solvent, an apolar solvent, a mixture of polar and apolar solvents, using supercritical carbon dioxide or the successive use of all these methods.

[0021] Preferably, the plant extract used according to the invention is obtained by extraction with a polar solvent followed by extraction with a non-polar solvent and followed by extraction with supercritical CO2.

[0022] Rosemary extraction is preferably carried out on dried plants, for example on rosemary branches, cut and dried in the sun for 4 to 5 days or distillation residues low in essential oil content.

[0023] Polyphenolic compounds or their derivatives, obtained by chemical synthesis, by biotechnological means, or by extraction from plants, can be used alone or in a mixture in accordance with the invention.

[0024] Polyphenolic compounds or their derivatives can be used in free form or can be conjugated or coupled to a carrier to promote and amplify the absorbent action of the polyphenol-carrier combination.

[0025] By "saturated vegetable oil" is meant any saturated vegetable oil known to those skilled in the art and in particular hydrogenated palm oil, hydrogenated castor oil and cutina, etc.

[0026] In a particular embodiment, the polyphenolic compounds are chosen from carnosol, rosmanol, rosmarinic acid, carnosic acid, and their derivatives.

[0027] In another particular embodiment, the polyphenolic compounds consist of a mixture of carmosol, carnosic acid and rosmarinic acid or their derivatives.

[0028] In another particular embodiment, the polyphenolic compounds consist of a mixture composed in whole or in part of carnosol.

[0029] In another particular embodiment, the polyphenolic compounds consist of a mixture composed in whole or in part of carnosic acid.

[0030] In a preferred embodiment of the invention, said polyphenolic compounds are provided by a rosemary extract purified to at least 60%.

[0031] The weight ratio of polyphenolic compounds to oil blend is between 25:75 and 65:35.

[0032] In the context of the present invention, the difference in melting temperature between the oils is at least 10°C, and may be at least 15°C.

[0033] A second subject of the invention relates to the use of a formulation as defined above as free radical scavengers in tobacco-based cigarette filters, CBD-based cigarette filters, cannabis-based cigarette filters, or cigarette filters based on their mixture and in an air filtering fabric, in particular in a mask.

[0034] The microparticles behave in an "ingenious" manner. Indeed, they have a multiplied contact surface for the redox reaction, which contributes to the elimination of free radicals. The increase in the free enthalpy of the reaction leads to an acceleration of the process of adduction of radicals in the gas phase to microparticles comprising polyphenolic compounds instead of their adduction to tar particles. This results in a 30% to 65% reduction of radicals in tars (semi-liquid phase), particularly semi-quinones, thus making the tars less oxidizing and less mutagenic. This can be referred to as a tar neutralization effect.

[0035] Thus, microparticles reduce the mutagenic and carcinogenic effects of fumes and slow down the stiffening effect that occurs in the arteries due to the presence of oxidizing compounds.

[0036] A third subject of the invention relates to a method for preparing a cigarette filter capable of capturing free radicals, in particular free radicals present in smoke, consisting of: mixing microparticles formed from a mixture of 2 to 4 saturated vegetable oils, the melting temperatures of which differ by at least 10°C, with polyphenolic compounds or their derivatives, in particular in the form of a purified rosemary extract, and incorporating said microparticles into a filter for tobacco cigarettes, filter for CBD cigarettes, filter for cannabis cigarettes or filter for cigarettes with a mixture density of 0.3 mg / mm 3< to 1 mg / mm 3<.

[0037] A fourth subject of the invention relates to a cigarette filter obtained by the method described above in which the microparticles have a size between 70-200 microns and the filter obtained has a draw resistance of less than 1200 mm of water.

[0038] In a particular embodiment, the microparticles are incorporated into the part of the filter located on the side of the substance to be consumed over a length of 3 mm to 10 mm. Preferably, the part of the filter containing the microparticles is located in a 6 mm long zone, located 3 mm from the internal end of the filter (tobacco side or other substance to be consumed). This arrangement makes it possible on the one hand to avoid direct contact of the microparticles with excessive heat which would cause them to melt and on the other hand to mechanically retain the microparticles so that they do not migrate towards the mouth, preventing their ingestion.

[0039] The use of such a filter allows a reduction of free radicals in the gas phase, a reduction of free radicals in the semi-liquid phase, with the consequence for the smoker of a reduction of oxidative stress, a reduction of the oxidative effect of smoke, a reduction of inflammatory effects, a reduction of accelerated aging, a reduction of the risk of exposure to diseases: stroke, neurodegenerative, heart attack, cancer, etc.

[0040] A fifth subject of the invention relates to a process for preparing a filter fabric capable of capturing free radicals, in particular free radicals present in fumes and polluted air, consisting of: mixing microparticles formed from mixtures of 2 to 4 saturated vegetable oils, the melting temperatures of which differ by at least 10°C, with polyphenolic compounds or their derivatives, in particular in the form of a purified rosemary extract and depositing these microparticles on the surface of a filter fabric (of the extruded blown type, "meltblown") with a density of 15 gsm (gram per square meter / gram per square meter) to 80 gsm.

[0041] A sixth subject of the invention relates to a filter fabric obtained by the method described above in which said microparticles have a size of between 1 and 20 microns and a pulling resistance of less than 150% of the FFP2, FFP3 or N95 standard.

[0042] This filter fabric is particularly suitable for the preparation of a mask to capture free radicals present in polluted air. In this case, the microparticles are applied to the external face of at least one of the meltblown layers (extruded blown fabric), preferably at least on the outermost meltblown layer of the mask. This type of mask is particularly suitable for asthmatics and athletes or users in polluted environments such as urban environments.

[0043] Free radical absorption and the induced reduction of the impact of cytotoxic molecules (see Alexandrov, K., Rojas, M., & Rolando, C. (2006). DNA damage by benzo(a)pyrene in human cells is increased by cigarette smoke and decreased by a filter containing rosemary extract, which lowers free radicals. Cancer research, 66(24), 11938-11945. DOI: 10.1158 / 0008-5472.CAN-06-3277) promote the reduction of oxidative stress, the reduction of cardiovascular diseases, the reduction of the rate of mutagenesis in cellular tissues, the reduction of lipid oxidation, the reduction of neurodegenerative diseases, the reduction of the risk of cardiovascular accidents, the reduction of the frequency of asthma attacks, particularly during pollution peaks, the reduction of the risk of cancers in smokers, particularly lung and lower respiratory tract cancer, the reduction of the risk of testicular cancers, the reduction of inflammations of the mouth, the reduction of inflammations of the tongue and upper respiratory tract and the reduction of inflammatory phenomena.

[0044] The present invention will be better understood from the following examples, provided for illustration purposes and in no way to be considered as limiting the scope of the present invention. DESCRIPTION OF FIGURES

[0045] Figure 1 : Schematic view of the process of preparing a filter containing rosemary extract EXAMPLES EXAMPLE 1: Preparation of a rosemary extract

[0046] Rosemary spikes ( Rosmarinus officinalis L .) are subjected to extraction with ethanol at 65°C. The volume of ethanol used (in liters) corresponds to five times the weight in kg of the rosemary spikes.

[0047] The extract is then purified and enriched with polyphenols by selective supercritical CO2 extraction. Depending on the temperature setting between 40°C and 100°C and the pressure between 1 and 170 bars, the extract is purified and selectively enriched in its various components to achieve a purity of carnosic acid and caronsol greater than 50%.

[0048] Washing with one or more solvents can significantly reduce the odor of the extract to 50% while increasing the concentration of carnosic acid and caronsol to 65% or more.

[0049] Such an extract contains the following compounds: carnosol, rosmanol, rosmadial, carnosic acid, genkwanin, rosmarinic acid...

[0050] The proportions of these different components vary depending on the rosemary plant used. Typically, the result is an extract comprising about 10% to 25% rosmarinic acid, about 40% to 50% carnosic acid, and about 5% to 10% carnosol. EXAMPLE 2: Incorporation of mixture into a cigarette filter or filter fabric.

[0051] A cigarette filter is prepared by incorporating microparticles onto the surface of cellulose acetate or into crepe paper during its rolling. It is preferred that the part of the filter containing the microparticles be located in a 6mm long area, located 3mm from the inner end of the filter (tobacco or other substance to be consumed side).

[0052] The filter fabric is prepared by spreading the particles on the fabric surface and using an electrostatic charge according to the process known to those skilled in the art. EXAMPLE 3: Method for evaluating the efficiency of the filter or fabric containing the mixture of microparticles and nanoparticles and polyphenols and their derivatives. 1. Computer-aided modeling

[0053] The effectiveness of the cigarette filter or filter fabric in capturing free radicals thus prepared is demonstrated initially by computer-assisted modeling, according to the Monte Carlo method, which makes it possible to calculate the number of encounters between a carcinogenic target molecule and a polyphenolic compound used in accordance with the invention.

[0054] The amount of cytotoxic free radical molecules present in cigarette smoke was calculated on both sides of the filter.

[0055] The quantity of cytotoxic molecules comprising free radicals present in the air passing through the filter fabric was calculated on both sides per cm2 of filter fabric surface.

[0056] The number of cytotoxic molecules after filtration is a function of the volume of smoke passing through the filter, the surface area of ​​the filter, the concentration of cytotoxic molecules in the smoke and the concentration of particles from a mixture of saturated vegetable oils and polyphenols in the filter.

[0057] The authors of the present invention have thus shown that 10 mg of particles in the filter of a cigarette makes it possible to reduce by more than 60% the level of cytotoxic free radical molecules in cigarette smoke based on tobacco, CBD or cannabis. 2. Quantitative mass spectrometry assay of free radical filtration from cigarette smoke

[0058] Quantitative mass spectrometry assay of free radical filtration is performed using liquid chromatography (LC) tandem mass spectrometry (LC / MS-MS) in Multiple Reaction Monitoring (MRM) mode to quantify the OH° free radical content in cigarette smoke. This mode allows for a more specific and sensitive analysis. Indeed, the spectrometer detects only a few transitions rather than an entire mass spectrum, resulting in a higher signal-to-noise ratio. The spin trap 4,5,5-trimethyl-pyrroline-N-oxide (TMPO) was used in benzene. The experiment was performed by specifying the mass of the parent ion and monitoring its transition. A commercial spin trap, 3-amino-2,2,5,5-tetramethyl-1-pyrrolidinyloxide, was used as an internal standard for free radical quantification.The absolute amount of free radicals measured here was in good agreement with the ESR experiments.

[0059] The results are presented in Table 1 below. Table 1: Comparison of quantitative mass spectrometry assays of cigarette smoke free radical filtration using dual-filter incorporation: the first reference is additive-free. Filter A contains 15 mg of rosemary extract distributed over a length of 6 mm. Filter B contains 15 mg of particles from mixtures of rosemary extract and saturated vegetable oils, as previously described, distributed over a length of 6 mm. All three filters are cellulose acetate-based. Reference Filter A Filter B Deposit length 6 mm and filter 0,27 6,27 6,27 Rosemary extract, or particles by weight (mg) deposited - 15 15 Pressure loss (mm H 2 O) 86 86 Puffs for 5 cigarettes 72 74 74 Number of free radicals OH°×10 -15< 9,2 6,8 1,5 Free radical scavenging efficiency (%) - 26 85 3. Quantitative mass spectrometry assay of free radical filtration through a filter fabric

[0060] To test the anti-radical effect associated with the incorporation of microparticles according to the invention in a filter fabric, 25 gsm of 5 micron microparticles were deposited on the outer surface of a meltblown.

[0061] The analysis methodology used is the same as that described previously for filters.

[0062] The results show that the presence of microparticles makes it possible to capture at least 70% of OH° free radicals.

Claims

1. Formulation of microparticles associated with polyphenolic compounds and / or their derivatives, especially in the form of rosemary extract, wherein the microparticles consist of a mixture of 2 to 4 saturated vegetable oils whose melting temperatures differ by at least 10° C.

2. Formulation according to claim 1 wherein said polyphenolic compounds are selected from carnosol, rosmanol, rosmarinic acid, carnosic acid, and derivatives thereof.

3. A formulation according to claim 1 wherein said polyphenolic compounds consist of a mixture of carnosol, carnosic acid and rosmarinic acid and / or derivatives thereof.

4. A formulation according to claim 1 wherein said polyphenolic compounds consist of a mixture composed wholly or partly of carnosol.

5. A formulation according to claim 1 wherein said polyphenolic compounds consist of a mixture composed wholly or partly of carnosic acid.

6. A formulation according to claim 1 in which said polyphenolic compounds are provided by a rosemary extract purified to at least 60%.

7. Use of a formulation as defined in claim 1 as a free radical scavenger in tobacco-based cigarette filters, CBD- based cigarette filters, cannabis-based cigarette filters, or cigarette filters based on a mixture thereof and in an air filter fabric, in particular in a mask.

8. A method of preparing a cigarette filter capable of capturing free radicals, in particular free radicals present in smoke, consisting in: mixing microparticles formed from a mixture of 2 to 4 saturated vegetable oils, whose melting temperatures differ by at least 10° C, with polyphenolic compounds or their derivatives, in particular in the form of a purified rosemary extract, and incorporate these microparticles in a filter for tobacco cigarettes, filter for CBD cigarettes, filter for cannabis cigarettes or filter for cigarettes with a mixture density of 0.3 mg / mm3 to 1 mg / mm3.

9. Cigarette filter obtained by the process according to claim 8 wherein said microparticles have a size between 70-200 microns and the filter has a pressure drop of less than 1200 mm of water.

10. Process for preparing a filter fabric capable of capturing free radicals, in particular free radicals present in smoke and polluted air, consisting of: mixing microparticles formed from a mixture of 2 to 4 saturated vegetable oils, whose melting temperatures differ by at least 10° C, with polyphenolic compounds or their derivatives, in particular in the form of a purified rosemary extract, and deposit these microparticles on the surface of a filter fabric with a density of 15 gsm to 80 gsm.

11. Filter fabric obtained by the process according to claim 10 wherein said microparticles have a size of between 1 and 20 microns and a draft resistance of less than 150% of the FFP2 or N95 or FFP3 standard.