RU2791329C1 - Silver water propolis extract - Google Patents

Abstract

FIELD: food industry.

SUBSTANCE: proposed aqueous extract of propolis contains propolis, royal jelly, cluster silver and water as initial components in the following ratio of initial components, by % of weight: propolis - 15.0 - 40.0; royal jelly - 1.0 - 6.0; cluster silver - 0.006 - 0.04; water is the rest. Moreover, the extract is obtained by placing propolis in an enamelled container, adding distilled water, heating to 60°C and boiling for 180 minutes. Then it is poured into a dark opaque glass container and allowed to stand for 48 hours, poured into an enamelled container, heated to 80°C, boiled for 180 minutes, poured into a dark opaque glass container and allowed to stand for 48 hours, in the third phase the broth is again placed in an enamelled container, boiled at 60°C for 180 minutes and allowed to stand for 48 hours. Then it is filtered through a fine sieve, cleaned of mechanical contamination - propolis particles, the purified extract is poured into a dark opaque glass container, royal jelly, cluster silver are added, stirred until evenly dissolved, left for 5 hours to settle and packaged. Preferably, the propolis extract additionally contains calcium alginate from kelp in the amount of 11% by weight.

EFFECT: invention is aimed at increasing the antiviral activity of propolis extract.

2 cl, 3 tbl, 6 ex

Description

Technical field

SUBSTANCE: invention relates to beekeeping derivatives, namely to propolis extract on silver water. The proposed invention can be used in the food industry, agriculture, medicine, pharmacology as a biologically active food supplement and / or as an external agent for the eyes, ears, nose.

State of the art

An alcoholic extract of propolis and a method for its production are known, which consists in the fact that it is purified from mechanical impurities and exposed to a solvent, which is ethyl alcohol or an aqueous solution of alcohol. The effect of alcohol on propolis is carried out for a long time in a container protected from light (Kivalkina V.P. and Barskova A.A. Dosage forms of propolis. journal "Beekeeping" N 11, p. 36, M. 1991).

The disadvantage of the alcohol extract of propolis is the limitation of the use of the obtained extract, due to the use of alcohol in one concentration or another, which has specific properties that limit their use, in particular in the treatment of stomach ulcers. Also, an alcohol solution is unacceptable for the treatment of children and persons sensitive to alcohol.

A known method for producing a propolis extract, involving the purification of propolis, grinding and extraction with water, and a propolis extract containing propolis and water (IN Solodukhin, Recommendations for the use of beekeeping products for preventive and therapeutic purposes. Volgograd, 1991, p. 35).

The disadvantage of the aqueous extract of propolis is the low content of biologically active substances due to the low extraction ability of water in relation to propolis.

A water-soluble pharmaceutical composition is known, disclosed in the RF patent for the invention No. 2715049 (publ. 26.02.2020). The composition is obtained by extracting a solution of L-arginine and crushed natural propolis in deionized water, and contains L-arginine and natural propolis in a ratio of 1:1.

Known biologically active composition based on chitosan gel, disclosed in the RF patent for invention No. 2209069 (publ. 27.07.2003). The known composition may contain propolis and silver ions.

A water-based propolis extract is known, disclosed in the RF patent for the invention No. 2751957 (publ. 21.07.2021). The known extract contains propolis and water, while obtaining the extract, it is homogenized in the presence of a stabilizer, which is used as silver ions.

However, known analogs do not have or have only a mild antiviral effect.

Known propolis extract on silver water Smart Bee (access mode : https://smartbee.expert/propolis , accessed: December 2021) - a prototype. Known product includes propolis in the amount of wt. 30%, water containing silver ions, royal jelly, royal jelly.

The disadvantage of the known extract of propolis is the low antiviral activity. This is due to the fact that in order to achieve a beneficial effect from the use of propolis, the accumulation of the substances contained in it in the body is required. At the same time, silver ions do not have a significant effect on the antiviral activity of the product; in addition, they have a high reactivity, which does not allow increasing the silver content in the product for reasons of safety for the human body. Therefore, the cumulative antiviral effect for humans from the use of this product is rather weakly expressed.

Disclosure of the essence of the invention

The technical problem underlying the present invention is to expand the range of environmentally friendly products with antiviral effect for internal/external use.

The technical result achieved in the implementation of the invention is to increase the antiviral activity of the propolis extract.

An aqueous propolis extract is claimed as an invention. The composition contains propolis, royal jelly and water. Unlike the prototype, cluster silver was introduced into the composition in the following ratio of components, wt. %:

Propolis 15.0 - 40.0 royal jelly 1.0 - 6.0 cluster silver 0.006 - 0.04 Water rest

Water propolis extract is obtained by placing propolis in an enameled container, adding distilled water, heating to 60 degrees and boiling for 180 minutes, then pouring into a dark opaque glass container and leaving to stand for 48 hours, pouring into an enameled container, heated to 80 degrees, boiled for 180 minutes, poured into a dark opaque glass container and left to stand for 48 hours, in the third phase the broth is again placed in an enameled container, boiled at 60 degrees for 180 minutes and left to infuse for 48 hours, filtered through a fine sieve, cleaned of mechanical contamination - propolis particles, pour the purified extract into a dark opaque glass container, add royal jelly, cluster silver, stir until evenly dissolved, leave for 5 hours to settle and pack.

In particular, the composition may additionally contain calcium alginate from kelp in the amount of 11 wt.%.

Beekeeping products, in particular, propolis, are known for their universal restorative effect on the human body, and are widely used both in traditional medicine and as concomitant drugs in the treatment of a wide range of diseases. Propolis has antibacterial properties, enhances the effect of antibiotics during treatment, activates the immune system, accelerates recovery after long illnesses and surgical intervention.

Another bee product, royal jelly, is an effective natural stimulant. Together with propolis, it activates the immune system and increases the energy potential of the body.

In turn, silver has antimicrobial, antiviral and anti-inflammatory effects. However, currently known drugs contain for the most part this element in the form of silver ions or colloidal silver. Their use is limited to the maximum allowable daily intake for a person. Therefore, it is not possible to enhance the beneficial effects of silver in the product by simply increasing its content, since an increase in the content also leads to an increase in the level of toxicity of the product. This makes it impossible to use the product inside, and when used externally, it can lead to undesirable side effects associated with inflammation, skin irritation.

Strengthening the beneficial effects of silver in the composition is achieved through the use of its other form - in the form of clustered silver. Cluster silver has a more pronounced antiviral effect by reducing the size of silver particles compared to silver ions or colloidal silver. At the same time, clustered silver is safe, belongs to the 4th hazard class (low hazard). In combination with bee products, cluster silver makes it possible to achieve a synergistic antiviral effect. In addition, cluster silver particles are not prone to long-term accumulation in the body, they are quickly excreted from the body.

The concentration of propolis in the composition is determined by the direction of application. For example, for children, the lowest concentration is chosen, while the maximum allowable, in order to maintain high efficiency. Royal jelly has stimulating properties. Therefore, the maximum allowable concentration was chosen for children so as not to cause the effect of hyperactivity. The silver content of the composition is limited to the range in which the product shows a significant increase in positive effects. At the same time, the threshold of the maximum permissible concentration for the body is not exceeded, which makes the product safe, non-toxic.

The proposed invention is illustrated by the following materials, which present:

Table 1 - Examples of samples of propolis extract on silver water;

Table 2 - The results of testing the cytotoxicity of the studied samples in relation to the Vero cell culture;

Table 3 - The results of the evaluation of antiviral efficacy and cytotoxicity of the studied samples.

Description of embodiments of the invention

Propolis has been used since ancient times in folk medicine as a preventive and therapeutic agent. It is now known that propolis is detrimental to some bacteria, in particular pyogenic (streptococci, staphylococci), is used in veterinary medicine and in the treatment of cracked nipples in nursing mothers; activates the functions of many protective mechanisms of the human body. Under its influence, phagocytosis increases, the content of a special protective protein (properdin) in the blood increases, and the production of specific antibodies against pathogens and toxins increases. Propolis helps to strengthen tooth enamel, most microorganisms do not become resistant to it; it is a nonspecific immunostimulant and increases the activity of antibiotics.

The composition of propolis includes resins (a mixture of organic acids), balms, tannins, essential oils, phenolic acids, aromatic aldehydes, flavonoids, ash elements; in addition, tocopherol, ascorbic and nicotinic acids, riboflavin, thiamine were found; from organic acids benzoic, cinnamic and others.

Propolis has antibacterial properties, enhances the effect of antibiotics during treatment, activates the immune system, accelerates recovery after long illnesses and surgical intervention. Propolis works at the cellular level, removes toxins from the gastrointestinal tract, while not disturbing the intestinal flora, cleanses the liver, lowers cholesterol, constricts blood vessels and strengthens their walls, and has a cumulative effect. Propolis extract is suitable for supporting the nervous system with high mental activity. During a cold, it supports the removal of inflammatory processes. Propolis has powerful anti-inflammatory properties, has analgesic properties and promotes healing and repair of cells and tissues. It can be used both externally and internally.

Royal jelly forms a high immune response, has so many substances that it can trigger regeneration and rejuvenation at the cellular level, activates the longevity regime, and is used to improve blood quality.

The claimed properties are due to the presence of such components as: amino acids, including essential ones, proteins (proteins) mainly albumins and globulins, enzymes, carbohydrates, trace elements and salts (sodium, potassium, magnesium, calcium, zinc, copper, manganese, selenium and etc.), vitamins (B1, B2, B3, B5, B6, BC, B12, C, E, A), acetylcholine and choline, organic acids (19 items), nucleic acids (DNA and RNA), lipids - mainly unsaturated fatty acids, hormones (progesterone, testosterone).

Royal jelly is the strongest natural stimulant. It activates the immune system and increases the energy potential of the body, slows down the aging process and is included in the diet of centenarians.

Cluster silver is an improved version of modernized colloidal silver, does not contain impurities, has a smaller size of silver nanoparticles. It does not harm the intestinal microflora, on the contrary, it contributes to its normalization, participates in biochemical and physiological processes, normalizes them, and helps to increase the protective and adaptive capabilities of the body, has a general strengthening effect on the body. Shows antibacterial action, strengthens action of antibiotics, not allowing harmful bacteria to adapt. Unlike antibiotics, which do not help with viral infections, cluster silver has antiviral activity.

Cluster silver is used in medicine, cosmetics, veterinary medicine, animal husbandry and poultry farming. It is added to medicines, food additives (BAA), feed additives, in the treatment of ENT diseases, SARS, tuberculosis, intestinal infections, etc. In therapeutic doses, cluster silver is safe, belongs to the 4th hazard class (low-dangerous).

The favorable effect of cluster silver on normal microflora makes it possible to use it as an auxiliary supplementary agent in the complex therapy of a number of diseases in order to correct microbiocenosis. One of the uses of propolis extract is aimed at improving metabolism and normalizing the functioning of the gastrointestinal tract. The addition of clustered silver, in symbiosis with propolis, fulfills the declared property.

Also, cluster silver enhances the antimicrobial, anti-inflammatory and immunostimulating effect. Due to the high stability of cluster silver, the shelf life of the product is increased. Additionally, the antifungal and antiviral effect of cluster silver expands the possibilities of using propolis extract.

As an additional component in the composition, calcium alginate from kelp can be used. The beneficial properties of the extract with marine calcium are aimed at replenishing the balance of calcium in the body, since its deficiency in the body can affect the condition of the teeth, the strength of the musculoskeletal system, and manifest itself in the form of convulsions. Also, calcium is important during pregnancy, with the rapid growth of children, and for the elderly. Calcium alginate is able to bind and remove metabolic by-products, heavy metal salts and radionuclides from the body. When taking it, the sorption of radionuclides (strontium and cesium) is 90%.

Calcium alginate helps to delay the absorption of water in the intestines, normalizing the stool. Effective for maintaining and restoring the tone of the immune system. It has antimicrobial, antifungal and antiviral activity. Anti-allergic effect: due to the content of calcium ions, it strengthens the membrane of specialized cells containing histamine, serotonin, etc., when they are released from the cells, a violent allergic reaction instantly develops. Has antacid properties: neutralizes hydrochloric acid of gastric juice. Thanks to this, calcium alginate is very effective in suppressing heartburn, a condition that, if repeated, can cause serious complications, up to cancer of the esophagus. Calcium alginate promotes the healing of ulcers and erosions of the stomach and duodenum, stops or reduces bleeding in these lesions of the gastrointestinal tract.

Propolis extract goes through 3 stages of infusion, during which useful substances are released into the infusion.

Propolis is placed in an enameled container, distilled water is added. Heat up to 60 degrees, cook for 180 minutes. Pour into a dark opaque container (glass) and leave to settle for 48 hours. Then it is poured into an enameled container, heated to 80 degrees, boiled for 180 minutes. Pour into a dark opaque container (glass) and leave to settle for 48 hours. In the third phase, the broth is again placed in an enameled container, boiled at 60 degrees for 180 minutes. The resulting broth is left to infuse for 48 hours. Then the resulting broth is filtered through a fine sieve, cleaned of mechanical contamination (propolis particles). The purified extract is poured into a dark opaque container (glass), royal jelly, cluster silver according to the recipe are added, stirred until evenly dissolved, left for 5 hours to settle. Further it is transferred to packaging in containers. In the case of propolis extract with marine calcium, calcium alginate from kelp is added along with the components: royal jelly, cluster silver. For all 6 examples shown in Table 1, the technological process is the same, the difference is in the content of the components.

In relation to experimental samples in which the proposed invention was used, laboratory studies were carried out in order to determine their antiviral activity. The following tasks were set before the study:

– study of in vitro cytotoxicity of experimental samples on Vero cell culture using the MTT test;

– evaluation of the antiviral activity of the samples using the plaque inhibition method;

– determination of the chemotherapeutic index (CTI) for the samples under study.

In the course of the study, it was found that moderate antiviral activity against one of the viruses that cause acute respiratory infection is observed for the samples of examples 1 and 2. The CTI value of these samples is 9.12. With regard to the samples of examples 3-6, it was found that their CTI value is 7.38, which indicates the presence of mild antiviral properties.

At the first stage of the study, the cytotoxicity of the samples against the Vero cell culture was determined using the MTT test method (Mossman test).

A cell culture in the form of a suspension with a cell content of 4.5*10 ⁵ cells/ml was added at 100 μl per well on the eve of the study. After 24 hours, the confluence of the monolayer was determined by visual inspection, selecting only those plates for which the confluence was 80-100%.

Solutions of the studied samples, starting from a concentration of 50%, were added in 100 µl per pre-washed cell culture and incubated for 24 hours at a temperature of 37°C and 5% CO ₂ . Next, the cells were washed with phosphate-buffered saline (PBS) and a solution of MTT (methyltetrazolium bromide) was applied on a maintenance medium at a concentration of 0.5 µg/ml, after which they were incubated for 1.5 hours at a temperature of 37°C and 5% CO ₂ . At the end of the incubation period, the MTT solution was removed, the precipitate was dissolved in 96% ethanol, and the optical density of the solution was determined at a wavelength of 535 nm. Based on the obtained data TsTD ₅₀ (the concentration of the drug (sample), causing the death of 50% of the cells).

The study of the antiviral activity of the samples was carried out according to the treatment and prophylactic scheme. For the study, 6-well plates with a daily monolayer of Vero cells were used. The sample at appropriate concentrations was added to the culture medium, after which it was incubated for 1 h at 37°C, 5% _CO2 . Next, the virus inoculum was added and incubated for 2 h at 37°C, 5% CO ₂ . Then the inoculum and the sample were removed, after which a solid cover medium containing 0.9% agar was added to the cells, to which the sample was added at appropriate concentrations and incubated for 4 days at a temperature of 37°C and 5% CO ₂ .

At the end of the incubation period, the cells were fixed and stained with a 0.2% solution of crystal violet in 10% formaldehyde, and the number of plaque forming units (PFU)/ml was counted. Based on the data obtained, ED ₅₀ was calculated - the concentration of the drug (sample), which reduces the reproduction of the virus by 50%.

Next, CTI (chemotherapeutic index) was calculated, that is, the ratio of CTD ₅₀ to ED ₅₀ . Samples with HTI more than 8 were recognized as having a pronounced activity.

At the first stage, 50% cytotoxic concentration (CTD ₅₀ ) was determined for each sample, the test results are presented in Table 2.

As can be seen from the data presented in the table, the values of CTD ₅₀ for all three studied samples were similar, ranging from 3.9 to 11.9%. Further, for each sample, a study of antiviral activity was carried out by the method of inhibition of plaque formation. The calculated values of antiviral activity, cytotoxicity and chemotherapeutic index, as well as the % reduction in the infectious activity of the virus at the maximum tolerated concentration are shown in Table 3.

From the obtained data it follows that for samples according to examples 3-6, the CTI value is very close to the threshold value (7.38 compared to 8), and the % reduction in the infectious activity of the virus is 88.1%. The data obtained indicate the presence of a small antiviral activity of the sample.

In samples according to examples 1-2, the CTI value exceeds the threshold value, while the percentage of reduction in infectious activity is 93.1%.

Various embodiments of the invention can be used in products having the following indications for use.

Example 1. Recovery of the body

Composition: propolis - 30 wt.%, royal jelly - 6 wt.%, cluster silver - 0.04 wt.%, water - the rest.

Strengthens the immune system and has a powerful bactericidal and antiviral effect, protects the body from infections, normalizes cholesterol levels. Contains oxidic acids. It helps to cleanse cell membranes, accelerate the removal of toxins, restore mucous membranes, reduce blood clotting, relieve vascular spasm, outflow of bile, normalize the functioning of the digestive tract, prolong the action of insulin, protect liver cells, improve metabolism, protect the body from free radicals, etc.

Application: adults 1 teaspoon (5 ml) 2-3 times a day 30 minutes before meals. The duration of admission is 1 month.

Example 2. To replenish calcium in the body

Composition: propolis - 35 wt.%, royal jelly - 4 wt.%, cluster silver - 0.04 wt.%, calcium alginate - 11 wt.%, water - the rest.

The beneficial properties of the extract are aimed at replenishing the balance of calcium in the body, since its deficiency in the body can affect the condition of the teeth, the strength of the musculoskeletal system, and manifest itself in the form of convulsions. Also, calcium is important during pregnancy, with the rapid growth of children, and for the elderly. The agent can be used externally in the treatment of atopic dermatitis, hard-healing ulcers, burns and other skin lesions.

Recommended for osteoporosis, bone fractures, rickets, diseases of the spine and joints, to strengthen the immune system and prevent colds and viral diseases, convulsions and anemia, dermatitis, dry skin, burns and other skin lesions.

Application: 1 teaspoon 2 times a day 30 minutes before meals.
Shake before use.

Example 3. Support and protection of the child's body

Composition: propolis - 15 wt.%, royal jelly - 1 wt.%, cluster silver - 0.006 wt.%, water - the rest.

Improving immunity, supporting a growing body, preventing viral and bacterial diseases. It has a beneficial effect on a growing organism - it supports the immune system and the musculoskeletal system of a child who experiences special loads during the growth period, improves sleep, strengthens the nervous system. The concentration of propolis allows the use of the product as drops in the ears and nose of children.

Inside: 30 minutes before meals, 5-10 drops dissolved in water or on a piece of sugar.

Nose: 1-2 drops 2-3 times a day.

Ears: warm the solution to 35 degrees Celsius, 2-4 drops 3-4 times a day

Example 4 Restoration of breathing

Composition: propolis - 40 wt.%, royal jelly - 5 wt.%, cluster silver - 0.006 wt.%, water - the rest.

The natural ingredients contained in the drops help in the complex treatment of inflammatory diseases of the nasopharynx, and also have an antiviral and immunomodulatory effect. The composition of propolis includes such components as: provitamin A; vitamins of group B, C, E; manganese; iron; silicon; zinc; calcium; phosphorus; magnesium; amino acids and organic acids, thanks to which, it is considered one of the best medicines for many ailments. The presence of royal jelly in the composition allows you to quickly alleviate the condition with acute respiratory infections and acute respiratory viral infections, because. it is a natural biostimulant and has clear healing properties.

Application: Recommended course use 2 drops 2-3 times a day for 14 days. If necessary, repeat the course.

Example 5 Anti-Inflammatory Agent for the Ears

Composition: propolis - 30 wt.%, royal jelly - 2 wt.%, cluster silver - 0.006 wt.%, water - the rest.

Contains propolis, which includes more than 50 components, it is they that have an immunomodulatory, antibacterial, anti-inflammatory and antiviral effect. The tool promotes cell renewal, moisturizes the mucous membrane, strengthens blood vessels, helps with wound healing and in the treatment of inflammatory diseases, is used for otitis media, colds and viral diseases, is used in combination with a runny nose to reduce pressure on the ears, relieves swelling.

Application: Recommended course use 2 drops 2-3 times a day for 14 days.

If necessary, repeat the course.

Example 6: Relieve tension and moisturise the eyes

Composition: propolis - 35 wt.%, royal jelly - 2 wt.%, cluster silver - 0.006 wt.%, water - the rest.

The therapeutic effect of propolis is observed at the initial stage of hyperopia, with retinopathy, conjunctivitis and myopia. The effectiveness of silver is also observed in conjunctivitis, blepharitis, keratitis, inflammation of the lacrimal sac and other inflammatory processes. Drops relieve inflammation of the eyes, moisturize and saturate the eye with nutrients and microelements, improve the conduction of the main nerve impulses, improve the sensitivity of the retina to various levels of light radiation and increase visual acuity, cleanse the fundus of the eye from old cells, strengthen the walls and improve the elasticity of the eye vessels, improve blood circulation of the eye vessels, retinal trophism and blood microcirculation in the eyes.

Recommended use:

- with increased eye loads, including prolonged work at the computer;

- with high sensitivity and susceptibility of the eyes to external factors;

- with tearing, dryness and redness of the eyes.

Application: Shake before use.

Adults and children over 12 years old: 1-2 drops 2-4 times a day

Claims (4)

1. An aqueous propolis extract containing propolis, royal jelly, cluster silver and water as initial components in the following ratio of initial components, wt.%: Propolis 15.0-40.0 royal jelly 1.0–6.0 cluster silver 0.006–0.04 Water rest, obtained by placing propolis in an enameled container, adding distilled water, heating to 60 degrees and boiling for 180 minutes, then poured into a dark opaque glass container and left to stand for 48 hours, poured into an enameled container, heated to 80 degrees, boiled for 180 minutes, poured into a dark opaque glass container and left to stand for 48 hours, in the third phase, the broth is again placed in an enameled container, boiled at 60 degrees for 180 minutes and left to infuse for 48 hours, filtered through a fine sieve, cleaned of mechanical contamination - particles propolis, pour the purified extract into a dark opaque glass container, add royal jelly, cluster silver, stir until evenly dissolved, leave for 5 hours to settle and pack. 2. Water propolis extract according to claim 1, characterized in that it additionally contains calcium alginate from kelp in the amount of 11 wt.%.

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