ES2977206A1 - Antimicrobial hydrodistillates from olive pomace, their production process, and their encapsulation in microparticles for use in cosmetic and/or dermatological compositions - Google Patents

Abstract

The present invention comprises the production of antimicrobial hydrodistillates from olive pomace, a production method, and a process for encapsulating them in microparticles for use in cosmetic and/or dermatological compositions such as aqueous mist (aerosol) for topical use against pathogenic microorganisms that cause skin diseases and conditions such as dermatitis, acne, or psoriasis. The present invention includes: 1) A process for producing antimicrobial hydrodistillates from olive pomace that do not contain phenolic compounds or antioxidants, but do contain volatile compounds; 2) A process for encapsulating the antimicrobial hydrodistillates in microparticles using a water-soluble polymer by means of the spray-drying technique; 3) A cosmetic and/or dermatological composition, also known as aqueous mist (aerosol), in which the microparticles containing the antimicrobial hydrodistillates are dispersed in distilled, ultrapure, or thermal water.

Description

DESCRIPTION

Antimicrobial hydrodistillates from olive pomace, their production process, and their encapsulation in microparticles for use in cosmetic and/or dermatological compositions

Technical sector

The present invention consists in the production of antimicrobial hydrodistillates from olive pomace, encapsulation in microparticles and production of cosmetic and/or dermatological compositions, also called aqueous mist (aerosol), for topical use against microorganisms that cause skin diseases and conditions such as dermatitis, acne or psoriasis. Antimicrobial hydrodistillates, microparticles of antimicrobial hydrodistillates and cosmetic and/or dermatological compositions, or aqueous mist, are susceptible to being used in the field of cosmetics and dermatology.

Background of the invention

Olive pomace is a waste product generated in the olive oil sector after the extraction of olive oil. Depending on the extraction system, olive pomace will contain a higher or lower water content, so the compounds present will be the same, but in greater or lesser concentration [A. Paz et al., Waste Management; 2020, 118, 435-444].

Olive pomace is characterised by its acid pH, its intense odour and colour, as well as its high organic matter content (COD greater than 300 g/L), total solids and phenolic compounds (between 0.5 and 24 g/L) [S. Dermeche et al., Process Biochemistry; 2013, 48, 532-1552]. These characteristics make it a highly toxic waste for the environment, which requires specific treatments and procedures for its management and disposal. However, in recent years the scientific community has begun to show interest in this waste, as it is a source of high added value compounds. Specifically, phenolic compounds such as hydroxytyrosol, tyrosol, rutin, vanillic acid, hydroxylated from elenoic acid, p-coumaric acid, caffeic acid or oleuropein, among others [Contreras et al., Process Biochemistry; 2020, 97, 43 56]. In this sense, there is patent ES2395032 Al "Phenolic extract of heat-treated olive by-products" whose object of invention is the extraction of phenolic compounds present in fresh alperujo or any by-product derived from the extraction of olive oil and/or the olive dressing industry and subsequent use as an antioxidant for the preparation of preparations for food, cosmetic and/or pharmaceutical use, as an inhibitory agent of the action of the enzyme L-tyrosinase, or as an inhibitory agent of platelet aggregation in the prevention of coronary heart disease. More specifically, there are many procedures for the extraction of hydroxytyrosol, a phenolic compound with high antioxidant power and great industrial interest. In this regard, patent ES2451011 T3 "Process for producing an extract containing hydroxytyrosol from olives and solids containing olive oil extraction residues" develops a heat treatment and subsequent purification from products and residues from olive mills to obtain this polyphenol, or patent ES2395317 Al "Process for obtaining hydroxytyrosol extract, mixed extract of hydroxytyrosol and 3,4-dihydroxyphenylglycol, and hydroxytyrosyl acetate extract, from olive by-products and their purification."

In the state of the art of the present invention, no reference has been found to any patent or procedure related to obtaining antimicrobial hydrodistillates through green technologies such as hydrodistillation, to develop a natural antimicrobial product for topical use. In recent years, the search for new natural compounds with antimicrobial activity is a challenge contemplated within national plans [National Plan against Antibiotic Resistance] and international plans [World Health Organization, Global Action Plan on Antimicrobial Resistance] to deal with the new health emergency posed by infections by pathogenic microorganisms, and their resistance to antibiotics and drugs present. Therefore, the present invention is focused on obtaining antimicrobial hydrodistillates and their encapsulation in microparticles for use in cosmetic and/or dermatological compositions. As an embodiment, an aqueous mist (aerosol) for topical use is proposed in which the possibility of using distilled water, ultrapure water or thermal water is contemplated. The use of the latter is emphasized, since in a previous work carried out with synthetic phenolic compounds such as ferulic acid, vanillic acid, p-coumaric acid, tyrosol, hydroxytorol and hydroxymethylfurfural (trading company TCI Europe N.V.) in combination with thermal waters from the province of Ourense, the antimicrobial activity was significantly increased [Paz et al., IV International Symposium on Thermalism and Quality of Life, 2021].

Explanation of the invention

The present invention aims at obtaining an antimicrobial hydrodistillate from olive pomace with activity against microorganisms that cause skin conditions, which does not contain phenolic compounds or antioxidants in its composition. Antimicrobial is defined as a compound that has the capacity to kill a microorganism or inhibit its growth. These antimicrobial hydrodistillates will be encapsulated in microparticles to enhance their antimicrobial effect, and used in cosmetic and/or dermatological compositions that are administered under an aqueous mist (aerosol) for topical use.

Hydrodistillation is a variant of steam distillation in which the material is immersed in boiling water and the generated vapours are condensed in the cooling column and left to recirculate, or collected, in a clean container, obtaining the hydrodistillation. This variant is proposed because it is a green technology that allows the separation of volatile compounds from other non-volatile ones by evaporation. In contrast to other conventional solid:liquid extraction methods such as Soxhlet, the extraction is carried out with polar and/or non-polar solvents (ethanol, chloroform, petroleum ether, etc.) and all non-structural components of the biomass are extracted (phenols, pigments, sugars, lipids, waxes, etc.). The parameters to be taken into account to carry out the hydrodistillation technique efficiently are the solid:liquid ratio to be introduced into the flask, and the pH. Drastic drops in pH can cause hydrolysis or polymerization of certain compounds, modifying their volatile profile.

One aspect of the invention relates to a process for obtaining antimicrobial hydrodistillates present in olive pomace by means of hydrodistillation. The process comprises the following steps:

a) A value between 5-75 grams of olive pomace is introduced into an extraction flask to which between 50-250 mL of distilled water is added.

b) The pH of the mixture is adjusted to a value between 4-10.

c) The hydrodistillation system is set up with a temperature in the condenser between 5 and -1OC, and a temperature in the heating mantle that allows the boiling of the previous mixture. It is left in recirculation between 10 and 60 minutes, after which the antimicrobial hydrodistillation begins to be collected.

Another aspect of the invention is the antimicrobial hydrodistillates obtained, which are characterized by not having phenolic compounds or antioxidants, but rather volatile compounds comprising the following composition:

A value between 70 and 90% of medium chain aldehydes (6-12 carbons) A value between 2 and 4% of short chain aldehydes (3-5 carbons) A value between 3 and 4% of organosulfur compounds

A value between 1 and 3% of ketones

A value between 0 and 1% of organic acids

A value between 0 and 1% alcohol

A value between 0 and 0.1% of aromatic compounds

These antimicrobial hydrodistillates are characterized by having antimicrobial capacity against 3 pathogenic microorganisms: Candida albicans CECT 1002T, Staphylococcus epidermidis CECT 232T and Staphylococcus aureus CECT 59.

Another aspect of the invention relates to the process of encapsulating antimicrobial hydrodistillates in microparticles using a polymer by means of a spray drying process. Natural antimicrobial compounds, if not properly preserved, are biologically active for a limited time, in addition to being unstable when faced with sudden temperature variations. The spray drying technique allows the solvent matrix to be evaporated almost instantly, providing stability to the compounds of interest. For this reason, it is a technique widely used in different sectors. Through this procedure, spherical and homogeneous microparticles are obtained that contain a certain percentage of the antimicrobial hydrodistillate. The antimicrobial hydrodistillate microparticles are generated by incorporating a polymer and/or dispersing agent, preferably soluble in water, so that the solids content of the dispersion increases, thereby improving the production yield and the characteristics of the microparticles. This polymer will help to obtain homogeneous and dispersed microparticles, thus avoiding the agglomeration effect. Polymers can be of natural origin when they are obtained from plant or animal sources, or they can be synthetic when they are generated by synthesis. In this case, 3 natural polymers were selected, alginate and carrageenan of marine origin and mannitol of plant origin. The latter was selected because it is an inert agent widely used in the pharmaceutical industry due to its high compatibility with most active ingredients.

The process for obtaining micro particles comprises the following stages:

a) A suspension is prepared in a liquid medium, preferably water, with a proportion of water-soluble polymer between 0.5-20% (w/V)

b) A percentage of antimicrobial hydrodistilled water between 1-30% (V/V) is added to the soluble polymer suspension.

c) Spraying is carried out using the spray drying technique. The suspension is fed using a peristaltic pump through a spray nozzle. Air is used as the atomizing gas. The operating conditions in the spray dryer equipment are: drying chamber temperature between 103-115°C, the pump that will move the air flow will act between 20-40% with the total air flow being 400-500 L/m3.

Likewise, another aspect of the invention is the use of microparticles in cosmetic and/or dermatological compositions in the form of an aqueous mist (aerosol) for topical use. In this case, this cosmetic and/or dermatological composition is characterized by comprising the microparticles with the antimicrobial hydrodistillates produced resuspended in an aqueous matrix, either water or thermal water, using a concentration between 5 mg/mL and 15 mg/mL. The dispersion of these microparticles in an aqueous matrix allows their subsequent application to be more homogeneous and for the concentration to be adequate to achieve the desired antimicrobial activity. Additionally, the use of thermal water is proposed when it is intended for cosmetic and/or dermatological composition, also called mist (aerosol), due to the positive characteristics that these waters present at a dermatological level, from a moisturizing and nutritional point of view. For correct use of the aqueous mist, it is recommended to spray it on clean skin and allow it to penetrate naturally, without subsequent drying.

Brief description of the drawings

To complement the description being made, a drawing is attached as an integral part of said description, in which, for illustrative and non-limiting purposes, the following has been represented:

Figure 1.- Percentage of inhibition produced by antimicrobial hydrodistillates against the three pathogenic microorganisms.

Preferred embodiment of the invention

In order to describe the present invention, the following example of a process for obtaining antimicrobial hydrodistillates and subsequent encapsulation in microparticles for use in cosmetic and/or dermatological compositions is presented:

1. Procedure for obtaining antimicrobial hydrodistillates from olive pomace

The starting raw material was olive pomace with a moisture content of approximately 6% (w/w). Between 5 and 75 grams of olive pomace were introduced into the extraction flask and between 50 and 250 mL of distilled water was added. The resulting mixture was neutralized and placed in the hydrodistillation system, setting the temperature in the condenser between 5 and -1°C, and in the heating mantle a temperature that allows the boiling of the previous mixture. It is left in recirculation between 10 and 60 minutes, after which the antimicrobial hydrodistillate begins to be collected, once the siphon is full.

The antimicrobial hydrodistillates obtained were analyzed by gas chromatography using the headspace technique to identify the volatile compounds. The sample was injected into a chromatograph equipped with a HP-5MS column (60 m x 0.25 mm; 0.25 um). The injector temperature was 250°C, using helium as the carrier gas and a ramp method of 8°C/minute from 40°C to 280°C. The acquisition time was 42 minutes. The analysis of the hydrodistillates showed the presence of the compounds indicated in the following table:

The analysis of its antioxidant activity (Flórez-Femández et al. 2018, Food Chemistry 277, 353-361) was also carried out, as well as its phenol content (Paz et al.

2019, Bioresource Technology, 275 402-409), flavonoids and hydroxycinnamic acids by spectrophotometry techniques (Tao et al., 2019 Industrial Crops & Products 138, 111474).

The antimicrobial activity was determined by the agar diffusion method (Clinical and Laboratory Standards Institute, 2018) against 3 pathogenic microorganisms: Candida albicans (CECT 1002T), Staphylococcus epidermidis (CECT 232T) and Staphylococcus aureus (CECT 59). Once the activity was confirmed, the minimum inhibitory concentration and the IC50 were quantified using the official liquid medium dilution method (National Committee for Clinical Laboratory Standards, 2000) against the same 3 microorganisms. The IC50 is a parameter that indicates the concentration at which a compound is capable of inhibiting the growth of a microorganism by 50%.

When antimicrobial hydrodistillates with a percentage ranging from 20 to 100 were used, the composition of which did not contain any type of antioxidant compound, phenol, flavonoids or hydroxycinnamic acids, the inhibition percentages against Candida albicans CECT 1002T were between 75 and 100% (Figure 1). When antimicrobial hydrodistillates with a percentage ranging from 20 to 100 were used, the composition of which did not contain any type of antioxidant compound, phenol, flavonoids or hydroxycinnamic acids, the inhibition percentages against Staphylococcus epidermidis CECT 232T were between 39 and 100% (Figure 1). When percentages of antimicrobial hydrodistillates between 20 and 100 were used, whose composition did not contain any type of antioxidant compound, phenol, flavonoids or hydroxycinnamic acids, the inhibition percentages against Staphylococcus aureus CECT 59 were between 63 and 100% (Figure 1).

2. Encapsulation process of antimicrobial hydrodistillates in microparticles

The encapsulation of the antimicrobial hydrodistilled water in microparticles was carried out from a dispersion or suspension in an aqueous matrix, which can be water, distilled water, ultrapure water or thermal water. The suspension in liquid medium is formed by a polymer used as a support or carrier, and an antimicrobial hydrodistilled water. Three polymers of natural origin were tested, a polyalcohol such as mannitol, and two marine polymers, such as alginate and carrageenan. A solution was prepared in which the polymer had a percentage between 0.5-20% (w/V), while the hydrodistilled water was added in a percentage between 1-30% (V/V).

The production of microparticles was carried out by the atomization drying technique using a device known as a spray dryer, using a temperature in the drying chamber between 103-115°C, while the pump that will move the air flow will act between 20-40%, the total air flow being 400-500 L/m3.

Once the microparticles were produced with the antimicrobial hydrodistillation, their performance and behaviour were studied. In a preferred embodiment, the performance values obtained for mannitol were between 5 and 30% (w/w), for alginate between 5-30% (w/w) and for carrageenan between 5-30% (w/w).

3. Cosmetic and/or dermatological composition (aqueous mist) based on microparticles of antimicrobial hydrodistilled waters

The cosmetic and/or dermatological composition, also called aqueous mist (aerosol), was formulated by dispersing the microparticles in an aqueous matrix, in order to enhance the action of the hydrodistilled products against microorganisms that cause diseases and skin conditions. To do this, the amount necessary to obtain a concentration of between 5 mg/mL and 15 mg/mL of microparticles was weighed in bottles containing between 1-10 mL of water, distilled water, ultrapure water or previously sterilized thermal water. These bottles were hermetically sealed, introducing an aerosol nozzle, in order to ensure that the application of the cosmetic and/or dermatological product (aerosol) was homogeneous.

The use of microparticles containing the antimicrobial hydrodistillate increased the inhibition percentages against cutaneous pathogenic microorganisms in all cases. When the microorganism used was Candida albicans C<e>C<t>1002T, percentages between 20 and 50% of antimicrobial hydrodistillate encapsulated in microparticle format offered inhibition percentages between 75 and 100%. When the microorganism used was Staphylococcus epidermidis CECT 232T, percentages of antimicrobial hydrodistillate between 20 and 50% encapsulated in microparticle format offered inhibition percentages between 45 and 100%. When the microorganism used was Staphylococcus aureus CECT 59, percentages of antimicrobial hydrodistilled water between 20 and 50%, encapsulated in microparticle format, offered inhibition percentages between 65 and 100%. Additionally, when the water used for dispersion of the microparticles was thermal water, instead of distilled or ultrapure water, the inhibition percentages were increased in all cases between 10 and 20%.

Claims (8)

1. Antimicrobial hydrodistillates extracted from olive pomace that are characterized by not having phenolic compounds or antioxidants, but do contain volatile compounds with the following composition: - A value between 70 and 90% of medium chain aldehydes (6-12 carbons). - A value between 2 and 4% of short chain aldehydes (3-5 carbons). - A value between 3 and 4% of organosulfur compounds. - A value between 1 and 3% of ketones. - A value between 0 and 1% of organic acids. - A value between 0 and 1% alcohol. - A value between 0 and 0.1% of aromatic compounds. With antimicrobial capacity against 3 skin pathogenic microorganisms such as Candida albicans CECT 1002T, Staphylococcus epidermidis CECT 232T and Staphylococcus aureus CECT 59. 2. Process for obtaining antimicrobial hydrodistillates present in olive pomace by means of hydrodistillation according to claim 1, characterized by comprising the following stages: a) Introduce between 5-75 grams of olive pomace into an extraction flask, to which 50-250 ml of distilled water are added. b) Adjust the pH of the mixture to a value between 4-10 c) Configure the hydrodistillation system with a temperature in the condenser between 5 and -1°C, and a temperature in the heating mantle that allows the boiling of the previous mixture and the recirculation of the antimicrobial hydrodistillate between 10 and 60 minutes, and subsequent collection. 3. Process for encapsulating antimicrobial hydrodistillates in microparticles according to claim 1 to 2, characterized by comprising the following stages: a) Prepare a suspension in liquid medium, with a proportion of water-soluble polymer between 0.5-20% (w/V) b) Incorporate into the suspension of soluble polymer a percentage of antimicrobial hydrodistillation between 1-30% (V/V) c) Spray using the atomization drying technique, using a spray dryer, using a drying chamber temperature between 103-115°C, the pump that will move the air flow will act between 20-40%, the total air flow being 400-500 L/m3. 4. Process for encapsulating antimicrobial hydrodistillates in microparticles according to claim 3, wherein the liquid medium of step a) is water, distilled water or ultrapure water. 5. Process for encapsulating antimicrobial hydrodistillates in microparticles according to claim 3, wherein the soluble polymer of plant origin in step b) is of the inert type such as mannitol, or of the marine type such as alginate or carrageenan. 6. Use of the microparticles of olive pomace hydrodistillates according to any of claims 1 to 5 as a cutaneous antimicrobial compound in cosmetic and/or dermatological formulations. 7. Topical cosmetic and/or dermatological composition, according to claims 3 to 5, in the form of an aqueous mist (aerosol) characterized by comprising the microparticles containing the antimicrobial hydrodistilled water dispersed in distilled water, ultrapure water or thermal water, previously sterilized, in a final concentration between 5 mg/mL and 15 mg/mL. 8. Use of topical cosmetic and/or dermatological composition according to claim 7 in cosmetic and/or dermatological preparations for topical use against microorganisms that cause diseases and skin conditions such as dermatitis, acne or psoriasis.