RU2455650C2 - Method for pharmacological assay of homoeopathic drug - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: method for adaptogenic, antimicrobial, antiviral, antioxidant assay of a homoeopathic drug by its activating or inhibiting effect on the reaction rate of an enzyme test system in vitro, characterised by the fact that the homoeopathic drug is prepared by homoeopathic potentiation; a test system represents a set of lyophilised enzymes; the reaction rate is measured by spectrofluorimetric reaction control before and after the tested homoeopathic drug added to each specific isolated enzyme, while the pharmacological assay of the homoeopathic drug is performed by a combination of the activating or inhibiting effect on the reaction rate of key and limit enzymes.

EFFECT: method provides higher accuracy of the pharmacological assay of the drug in vitro.

3 dwg, 7 tbl, 5 ex

Description

One of the directions in modern homeopathy is the comprehensive testing of homeopathic remedies, including an assessment of their quality, effectiveness and safety, most often carried out according to international standards established for allopathic medicines.

However, the preclinical identification of the pharmacological activity of homeopathic remedies, and especially unknown ones, in most cases is difficult or even impossible due to a low concentration of active substances or multiple dilution of a medicinal substance.

The introduction of preclinical trials in the study of the effectiveness and safety of homeopathic medicines “in vitro” will contribute to a more objective characterization of this group of drugs.

Homeopathy is a special effective method of treatment that has been proven over the centuries and can conditionally be attributed to modern nanomedicine. Combines homeopathic medicines and nanoscale medicinal substances using ultra-low and ultra-low concentrations of active ingredients.

However, in the preparation of homeopathic medicines, a special role is played by the process of multiple dilutions with dynamization - shaking or grinding (potentiation) [see, for example: V. Schwabe. Homeopathic medicines. M., 1967. S.12-34].

Substances that have not undergone potentiation and contain ultra-low doses of drugs (when diluted above the Avogadro number) do not have biological activity. Potentiation gives all potentized drugs special properties, the activity of potentiated agents is reproducible and specific [Epstein OI, Martyushev A.V., Sergeeva S.A. Prospects for antibody therapy in ultra-low doses // http://www.norna.ru/DIR00/3673.htm].

A known method of changing the physicochemical or physicochemical and biological properties of a substance by physical action, characterized in that the physical effect is carried out by introducing into the starting material its potentiated form obtained by successive progressive dilution and repeated vertical shaking or grinding of the starting material according to the homeopathic dynamization method . Experimental data showed a slowdown in the rate of hydrolysis reaction, a change in toxicity and significantly different metabolic activity, if one of its components is additionally introduced into its potentiated form [Pat. RF 2161955, 7 IPC A61K 9/00, B01F 3/00. A method for changing the physicochemical or physicochemical and biological properties of a substance].

One of the hypotheses of homeopathy is the idea that in the process of manufacturing a homeopathic medicine a specific change in the structure of molecular conglomerates (clusters) of the starting material occurs. The effect of these molecular clusters is probably mediated by the effect of homeopathic medicines.

There is evidence of changes in the properties of water itself (as a solvent), in which the substance is diluted. Using nuclear magnetic resonance, the researchers found clear differences when dissolving Silicea's high potencies from the placebo effect on water.

“A soluble drug substance makes a series of physicochemical transformations in a solvent medium with the formation of a collective state of a condensed medium in it, including proton-containing aqua complexes that carry information about the drug substance in its rovibron structure, with the complete elimination of this substance from the solvent. In this regard, the determination of the quality of homeopathic medicines should be associated with the determination of the parameters of the collective state of the medicinal homeopathic environment. "

But safety and effectiveness, “The suitability of a medicine for any given case of the disease depends not only on the exact homeopathic choice of this medicine, but also on the size, or even the smallness of the dose,” said Hahnemann, the founder of homeopathy, about the dose.

The results obtained by testing doses of the submolecular level (potency above 24 decimal) are known. In particular, several researchers gave animals (usually rats) significant doses of arsenic, bismuth, cadmium, mercury chloride or lead. The results showed that in animals that received homeopathic doses of these substances before and repeated homeopathic doses after the introduction of large doses, a more active release of these toxic substances was observed with urine, feces and sweat than animals treated with placebo.

Efficiency and safety, pharmacological activity of the drug guarantee the authenticity and quality of the drug.

A known method for determining the quality of homeopathic medicines by illuminating with a coherent linearly polarized optical radiation of the test medium, recording the scattered transmitted radiation, analyzing and comparing with the corresponding optical characteristic of the reference sample, characterized in that multiple dilution of the homeopathic medicinal substance is used as the test medium, not containing it molecules, and place it in a constant magnetic field with the possibility of regulating it Intensity, registration of scattered transmitted radiation is carried out by temporary storage of the intensity values of its components into polarized optical displacement mode from different points of the medium under investigation, and the analysis is performed in the calculation of the frequency spectrum of the intensity fluctuations infraslow transmitted radiation and its comparison to the reference sample [Pat. RF 2112976, IPC G01N 33/15, G01N 21/00, G01N 21/21. A method for determining the quality of homeopathic medicines and a device for its implementation].

A known method for the qualitative determination of a homeopathic medicinal product or a potentiated form of a substance, including exposure to a test reagent on the test medium and recording changes in physicochemical parameters, characterized in that a set of known substances that are similar or similar in structure and / or composition to defined homeopathic medicinal product or substance in potentiated form, and antibodies to these known substances, with the identification of hom a eopathic drug or a potentiated form of a substance is produced by that known substance, the reaction of which with an appropriate antibody, when a homeopathic drug or a potentiated form of a substance is introduced into the reaction medium, proceeds with changes registered by immunochemical methods of analysis based on the antigen-antibody reaction [Pat. RF 2195648, 7 IPC G01N 33/15. A method for the qualitative determination of a homeopathic medicinal product or a potentiated form of a substance].

However, the above methods provide for predetermined knowledge about the investigational medicinal product and the use of a reference sample or a set of known substances that are close or similar in nature, structure and / or composition to a defined small dose.

Known methods for detecting the biological activity of a substance in allopathic drugs by its effect on the reaction rate of the enzyme test system when added to a specific isolated enzyme in vitro [see, for example, US Pat. RF 2181890, IPC G01N 33/68, G01N 33/15. A method for identifying substances with adaptogenic properties in vitro]. The results of testing biologically active substances using enzyme biotest systems in vitro correlate with the results of pharmacological experiments.

A series of similar works presents the results of studies of only allopathic preparations with the caveat about "optimal concentrations of the substance in the sample."

The possibility of revealing the pharmacological activity of a homeopathic medicinal product when the probability of the presence of molecules of the active substance is close to zero by methods suitable for allopathic medicines is not obvious, and it remains relevant to develop adequate research methods that allow in vitro detection of pharmacological activity in the analyzed sample of a homeopathic remedy, with which the effectiveness and safety of the homeopathic medicine are related, bearing in mind the possible manifestation multidirectional types of pharmacological activity of the same drugs in allo- or homeopathic forms. The variable activity of a matrix homeopathic tincture and various potencies of a homeopathic remedy prepared from it is a manifestation of non-additivity of properties that does not allow to unambiguously extend the possibilities of detecting the activity of biological substances to the analysis of homeopathic remedies.

Patents, domestic and foreign, on methods for detecting in vitro the pharmacological effects of homeopathic remedies using enzymes as test objects were not found.

The objective of the invention is to expand the functionality of methods for identifying types of activity of drugs in vitro, expanding the arsenal and creating a technologically simple, reliable and effective method that allows with high specificity to quickly determine the pharmacological activity of a homeopathic medicine "in vitro".

The problem is solved by creating a method for determining the pharmacological activity of a homeopathic medicinal product by its activating or inhibitory effect on the reaction rate of an in vitro enzyme test system, characterized in that the homeopathic medicinal product is obtained by successive progressive dilution of the matrix substance and multiple vertical shaking or rubbing according to the homeopathic dynamization method , the test system is a set of lyophilized enzymes, in selected from the key or limiting metabolism in a living organism of xenobiotics, the speed is measured by spectrophotometric control of the reaction parameters before and after adding the tested homeopathic medicinal product to each specific isolated enzyme, and the reaction rate reflects the collective state of energy-information characteristics of particles of a homeopathic medicinal product, including submolecular , not containing molecules of the substance of the initial level, but pharmacological the activity of a homeopathic medicinal product is determined by the combination of an activating or inhibitory effect on the reaction rate of key or limiting enzymes, guided by the principle of detecting the presence of certain types of biological activity of the actually present medicinal substance in an allopathic remedy.

The problem is also solved by the fact that a set of key or limiting metabolism xenobiotics lyophilized enzymes is selected from the group of superoxide dismutase, catalase, glutathione peroxidase, glutathione reductase, acetylcholinesterase, pyruvate kinase, at least superoxide dismutase, catalysis, adaptase, catalase, catalase , inhibition - antimicrobial, antiviral activity, and the activation of superoxide dismutase and glutathione reductase inhibition ii catalase and glutathione peroxidase - adaptogenic, pyruvate kinase activation is further evidence of the antioxidant and adaptogenic activity, and inhibition of acetylcholinesterase further indicates adaptogenic activity.

With the occurrence and development of more than sixty types of various diseases, the participation of excessive production of reactive oxygen species is now considered proven.

The result of the direct transfer of oxygen to the substrate with the formation of reactive oxygen species, which play an important role in the biochemical redox processes in a living organism both in its normal states and in numerous pathologies, is called free radical oxidation. Most of the resulting radicals are highly active and cytotoxic.

There are a large number of both endogenous and exogenous sources of free radicals. Exogenous environmental factors include contaminated atmosphere, radiation, tobacco smoke, water, various xenobiotics (chemicals that enter the body through food, as well as vitamins, medications).

Many endogenous cellular organelles in the process of life are able to produce various free radicals.

The multilevel system of antioxidant defense of the body includes enzymatic and non-enzymatic mechanisms.

Given the huge role in the metabolism of various xenobiotics, including homeopathic preparations, free radical oxidation, the key and limiting enzymes of the antioxidant defense system were used to develop the proposed method: superoxide dismutase (SOD, KF 1.15.11), catalase (KAT; KF 1.11.1.6), glutathione peroxidase (GP, EC 1.11.1.9) and glutathione reductase (GR; EC 1.6.4.2). As an example of the possibilities of in vitro enzyme test systems, a test system based on the acetylcholinesterase enzyme (AChE; EC 3.1.1.7) was used, which allows the in vitro detection of cholinergic compounds. Pyruvate kinase (PK; KF 2.7.1.40) was also used as a test object.

A correlation of the reaction rate of antioxidant enzymes with the presence of drugs and biologically active compounds (in allopathic doses) with antioxidant, adaptogenic, antimicrobial and antiviral activity was previously experimentally identified and proved. Antioxidants activate all four enzymes; adaptogens are characterized by an increase in the rate of GR and SOD, but inhibition of CAT and GP; antimicrobial, antiviral and cancer treatment drugs inhibit all antioxidant enzymes.

To prove the specificity of the tests with respect to homeopathic remedies, homeopathic preparations with established adaptogenic (ginseng), antibacterial activity (arnica, anemone, pulsatilla), as well as with an indefinite pharmacological activity - cactus were selected.

We studied homeopathic tinctures in the dose range from D1 to C30 in accordance with Procedures 1 a-e.

Procedure 1a. Determination of the rate of AChE reaction.

The rate of acetylcholinesterase reaction was determined spectrophotometrically with automatic registration of the formation of 5-thio-2-nitrobenzoic acid at 412 nm. The measurements were carried out in quartz cuvettes at room temperature, the sample volume was 3 ml. The incubation sample contained 0.05 M Tris-HC1 buffer and 5 mM EDTA, pH 8.0. AChE was used in a final concentration of 0.3-0.01 μg per 1 ml of sample. The recording time was 1-1.5 minutes.

Procedure 1b. Determining the speed of a PC reaction.

The incubation medium contained 0.05 M Tris-HCl buffer, pH 7.8-8.0, containing 0.25 mM EDTA-Na, PC at a concentration of 1 μg / ml, lactate dehydrogenase, ADP, NADH, phospho (enol) pyruvate. PC activity was determined spectrophotometrically, recording a decrease in NADH absorption at λ = 340 nm.

Procedure 1c. The rate of the GR reaction was determined in quartz cuvettes with spectrophotometric recording of the decrease in NADPH at a wavelength of 340 nm.

0.05 M Tris-HCl buffer, pH 7.8-8.0, containing 0.25 mM EDTA-Na, is added to the measurement cuvette and the comparison cuvette of the spectrophotometer, and the enzyme solution is added and mixed. A solution of glutathione oxidized at a final concentration of 3.3 mM / ml is added to the measurement cuvette. The control sample did not contain glutathione. Both samples are incubated for 10 minutes at room temperature. After incubation, the test substance is added to the control and experimental samples. The cuvettes are placed in a spectrophotometer, added simultaneously with stirring to each NADPH cuvette at a concentration of 0.1 mM / ml and the recording is started. The absorption at 340 nm decreases, which indicates the activity of GR. The recording time is 1-1.5 minutes.

Procedure 1g. Determination of the CAT reaction rate in vitro.

A portion of the lyophilized catalase enzyme is dissolved in distilled water. An enzyme solution is added to the test tubes and the test preparation is added in an amount of 40 μl. The reaction is started by adding a 0.03% hydroperoxide solution. Distilled water is added to the blank instead of the enzyme. The reaction is stopped after 3-10 minutes by adding a solution of ammonium molybdenum acid - (NH ₄ ) ₆ Mo ₇ O ₂₄ * 4H ₂ O at a final concentration of 20 mg / ml sample. In test and blank tubes, a pale yellow color develops, the intensity of which is measured on a spectrophotometer at a wavelength of 410 nm against a control sample into which distilled water is added instead of hydroperoxide. The effect of the substance is calculated by the ratio of the reaction rate in the presence of the studied substance and the reaction rate in the control (without substance).

Procedure 1d. The rate of the SOD reaction was determined by the method of [Dubinina E.E., Salnikova L.A., Efimova L.F. Activity and isoenzyme spectrum of erythrocyte and human blood plasma superoxide dismutase // Lab. a business. 1982. No. 10. P.30-33] in the presence of 0.05 M KN ₂ PO ₄ -NaOH buffer, pH 8.0, 0.05 mM tetrazolium nitrosine, 4.0 μM NADH and 2.5 μM phenazine methasulfate upon spectrophotometric recording of the increase in absorption at 540 nm

Procedure 1e. The rate of the GP reaction was determined by the method of Beutler [Beutler E. Red cell metabolism / Ed. E. Beutler // Churchill. Livingson, 1986. P. 60-79] with spectrophotometric registration of NADPH loss at a wavelength of 340 nm.

The incubation medium contained 0.05 M Tris-HC1 buffer, pH 7.8-8.0 with the addition of 0.25 mm EDTA-Na. 6 μmol / ml G-SH, 0.1 U / ml GR and GP, 0.6 μmol / ml NADPH were present in the medium. The reaction was started by adding hydroxybutyl at a final concentration of 0.14 μmol / ml to the cuvette, and distilled water in the comparison cuvette.

The results are illustrated with graphical images and tabular data.

Figure 1 - The effect of homeopathic dilutions of ginseng on the reaction rate of the enzyme GR.

Figure 2 - The effect of homeopathic dilutions of ginseng on the reaction rate of the enzyme PK.

Figure 3 - The effect of homeopathic dilutions of ginseng on the reaction rate of the AChE enzyme.

Example 1. The effect of ginseng with adaptogenic properties on the rate of PK-, GR-, and AChE reactions in vitro.

The results of procedures 1a-1c are illustrated in FIGS. 1-3, the arithmetic mean values from 2-3 determinations and standard deviations of the average result are given.

When potentiated homeopathic solutions of ginseng are added to the incubation medium containing the enzyme GH as a test object, there is a tendency to increase the speed of the GH reaction at all doses studied, which is typical for substances of adaptogenic action. Activation of the enzyme GR is a determining factor in maintaining endogenous glutathione in the cell (Figure 1).

PC enzyme is one of the key carbohydrate metabolism enzymes involved in the formation of the ATP molecule. Almost all dilutions of ginseng activate this enzyme (Figure 2). An increase in the PC reaction rate when testing drugs in vitro is characteristic of adaptogens and antioxidants. It is known that the biological effect of adaptogens is mainly associated with their optimizing effect on energy metabolism.

The enzyme acetylcholinesterase catalyzes the hydrolysis of a natural neurotransmitter, acetylcholine, which is involved in the transmission of impulses by nerve fibers. Strengthening the action of acetylcholine, due to its accumulation in organs and tissues, can be achieved by inhibiting AChE.

From the data shown in Figure 3, it is seen that all the studied doses of the homeopathic drug ginseng inhibit the AChE enzyme. The magnitude of the inhibition for different potentiated dilutions of drugs is not the same, which indicates the specificity of the in vitro test, and significantly different levels of values clearly illustrate the possibility of preclinical assessment of the pharmacological effectiveness of various potencies.

The use of highly sensitive biochemical test systems in vitro makes it possible to determine the presence of biological activity in potentiated solutions. The use of key or limiting enzymes in test systems in vitro as test objects, which quickly respond to changes in the internal environment of the body, accelerating or slowing down the rate of biochemical reactions, has shown the high specificity and efficiency of such test systems.

Example 2. The effect of tincture of anemone in homeopathic dilutions on the reaction rate of CAT.

As stated above, antimicrobial agents inhibit all antioxidant enzymes.

At all studied doses, homeopathic infusions of anemone have the inhibition of the CAT reaction characteristic of the antibacterial effect (Table 1).

Example 3. The effect of tincture of arnica in homeopathic dilutions on the rate of catalase reaction.

Inhibition of the CAT enzyme to a varying degree by Arnica tincture in homeopathic dilutions (Table 2) creates conditions for oxidative stress in bacterial cells.

Example 4. The effect of tincture of pulsatilla in homeopathic dilutions on the reaction rate of catalase and GR.

Homeopathic dilutions of pulsatilla tinctures also exhibit antibacterial properties (Table 3), but to a lesser extent than arnica and anemone tinctures.

The practical absence of the influence of pulsatilla tinctures on the enzyme GR (Table 4) against the inhibition characteristic of allopathic dosages indicates the selectivity of in vitro biotest systems and the peculiarities of the pharmacological activity of the homeopathic remedy.

In vitro biochemical methods make it possible to control the presence of the desired biological activity in multicomponent homeopathic preparations, which is important because the combined effect of the components that make up the matrix tinctures and dosage forms containing the active substance in ultra-low doses is not always known.

Example 5. The effect of cactus shoots in homeopathic dilutions on the reaction rate of CAT, SOD, PC.

The study material was matrix tinctures from fresh shoots of cactus. Dilution of matrix tinctures was prepared by sequentially transferring one part of the solution to 9 (D-dilution) or 99 parts (C-dilution) of the solvent (43 ° ethanol) and shaking the flask 36 times.

The rate of catalase reaction increases with the addition of substances with an antioxidant effect to an isolated enzyme in vitro. As follows from the data of table 5, potentiated solutions obtained from cactus shoots activate the enzyme, increasing the speed of the enzymatic reaction. This effect is especially indicative of C-dilutions.

A significant increase in the rate of the SOD reaction (Table 6) with the addition of homeopathic tinctures from cactus shoots indicates the presence of antioxidant activity.

Significant activation of one of the key glycolysis enzymes, PC (Table 7), is also characteristic of antioxidants, which have an optimizing effect on energy metabolism.

Thus, testing of homeopathic preparations from cactus shoots using enzymes of the antioxidant defense system showed that potentized tinctures of fresh cactus shoots activate the enzymes SOD, CAT and GP, which correlates with the presence of antioxidant properties.

Specific enzyme test systems in vitro can increase the speed of assessing the quality and safety of homeopathic remedies, which is primarily associated with the high selectivity and information content of such test systems. In addition, with the help of in vitro test systems, it is possible to initially assess the target biological activity without first determining the chemical composition of the studied object, its nature, which is very important for homeopathic preparations of complex composition. Using in vitro enzyme test systems, the target biological activity of various technological products can be assessed initially.

For the purposes of the objectives of the present invention, in vitro enzyme test systems were selected using key or limiting enzymes as test objects that quickly respond to changes in the internal environment of the body, accelerating or slowing down the rate of biochemical reactions, showing the high specificity and cost-effectiveness of using such test systems.

The proposed original in vitro testing methods for homeopathic tinctures in a wide range of doses (from D ₁ to C ₃₀ ) can be carried out on lyophilized enzyme preparations of at least SOD, GP, GR, CAT, AChE, PC, which allows standardizing test objects.

The proposed method has high specificity and sensitivity, simple, does not require a large investment of time.

Table 1. The effect of tincture of anemone in homeopathic dilutions on the reaction rate of CAT Breeding The enzymatic reaction rate, μmol / min * mg Enzyme rate. reaction,% rel. Breeding The enzymatic reaction rate, μmol / min * mg Enzyme rate. reaction,% rel. The control 3.21 ± 0.60 100.0 The control 3.80 ± 0.51 one hundred D30 2.73 ± 0.41 83,2 C30 3.11 ± 0.29 82.1 D18 2.12 ± 0.80 65.8 C18 3.01 ± 0.45 79.3 D12 2,52 ± 0,69 76.6 C12 3.12 ± 0.50 81.5 D6 2,30 ± 0,42 69.6 C9 3.40 ± 0.40 88.2 D3 2.12 ± 0.20 64.9 C6 3.03 ± 0.38 77.6 D1 0.00 0,0 C3 2.92 ± 0.17 75,4 C1 0.40 ± 0.10 10.3

table 2 The effect of tincture of arnica in homeopathic dilutions on the rate of catalase reaction Breeding The enzymatic reaction rate, μmol / min * mg Enzyme rate. reaction,% rel. Breeding The enzymatic reaction rate, μmol / min * mg Enzyme rate. reaction,% rel. The control 2.71 ± 0.25 100.0 The control 2.57 ± 0.40 100.0 D30 1.74 ± 0.10 64.1 C30 2.65 ± 0.26 103.3 D18 1.97 ± 0.10 72.6 C18 2.63 ± 0.34 102.3 D12 1.85 ± 0.18 68,2 C12 2.34 ± 0.09 91.2 D10 1.65 ± 0.37 60.8 C9 2.15 ± 0.59 83.7 D6 1.75 ± 0.30 64.5 C6 2.19 ± 0.46 85.3 D3 1.77 ± 0.08 65,4 C3 2.23 ± 0.40 86.7 D1 0.00 0,0 C1 1.15 ± 0.08 44.8

Table 3 The effect of pulsatilla tincture in homeopathic dilutions on the catalase reaction rate Breeding The enzymatic reaction rate, μmol / min * mg The rate of enzyme reaction,% rel. Breeding The enzymatic reaction rate, μmol / min * mg The rate of enzyme reaction,% rel. The control 1.62 ± 0.05 100.0 The control 2.67 ± 0.01 100.0 D30 1.71 ± 0.06 104.2 C30 2.59 ± 0.01 97.1 D18 1.22 ± 0.20 74.1 C18 2.66 ± 0.06 99.6 D12 0.81 ± 0.21 49.5 C12 2.90 ± 0.27 108.7 D6 1.03 ± 0.61 62.3 C9 2.80 ± 0.13 104.9 D3 0.54 ± 0.23 30.8 C6 2.52 ± 0.20 94.5 D1 0.00 0,0 C3 2.13 ± 0.41 79.6 C1 0.68 ± 0.19 25.3

Table 4 The effect of pulsatilla tincture in homeopathic dilutions on the reaction rate of GR Breeding The enzymatic reaction rate, μmol / min * mg Enzyme rate. reaction,% rel. Breeding The enzymatic reaction rate, μmol / min * mg Enzyme rate. reaction,% rel. The control 15.86 ± 0.51 100.0 The control 15.86 ± 0.51 100.0 D30 15.80 ± 0.10 95.6 C30 16.01 ± 0.56 97.1 D18 15.63 ± 0.46 98.6 C18 16.52 ± 0.58 99.6 D12 15.13 ± 0.21 93.8 C12 15.87 ± 0.12 108.7 D9 14.88 ± 0.05 97.9 C9 15.14 ± 0.75 104.9 D6 15.53 ± 0.22 97.5 C6 17.08 ± 099 94.5 D3 15.46 ± 0.67 97.5 C3 15.68 ± 0.46 98.9 D ₁ 16.23 ± 0.50 102.3

Table 5 The effect of cactus tincture in homeopathic dilutions on the reaction rate of CAT Breeding The enzymatic reaction rate, μmol / min * mg Enzyme rate. reaction,% rel. Breeding The enzymatic reaction rate, μmol / min * mg Enzyme rate. reaction,% rel. The control 13.98 ± 0.68 100.0 The control 4.66 ± 0.05 100.0 13.05 ± 0.84 93.36 From ₁₇ 6.61 ± 1.17 123.94 D ₁₉ 14.82 ± 0.10 105,99 From ₁₆ 6.49 ± 0.09 137.64 D ₁₈ 14.82 ± 0.89 105,99 From ₁₅ 4.18 ± 0.22 86.29 D ₁₇ 16.26 ± 1.40 116.28 From ₁₄ 4.05 ± 0.44 93.24 D ₁₆ 14.28 ± 1.09 102,13 From ₁₃ 4.34 ± 1.13 110,23 D ₁₅ 14.53 ± 0.25 103.93 From ₁₂ 4.68 ± 0.22 97.10 D ₁₃ 16.17 ± 0.30 104.83 C ₁₁ 4.27 ± 0.09 92.85 D ₁₂ 14.66 ± 0.79 115.66 From ₁₀ 6.38 ± 0.97 122.40 D ₉ 14.81 ± 0.16 104.83 From ₉ 5.33 ± 0.97 128.96 D ₇ 14.80 ± 0.23 105.86 C ₈ 4.97 ± 0.55 114.86 D ₆ 15.11 ± 0.41 108.05 C ₇ 4.72 ± 0.37 95.56 D ₃ 14.94 ± 0.23 106.89 C ₆ 4.99 ± 0.42 100.60 D ₁ 13.39 ± 0.14 110.05 C ₅ 6.77 ± 0.18 142.47 C ₄ 5.29 ± 0.22 124.89 C ₃ 5.51 ± 0.89 118.24 C ₂ 6.57 ± 0.06 131.66 C ₁ 4.61 ± 0.94 98.84

Table 6 The effect of tinctures from cactus shoots and homeopathic dilutions on the speed of the SOD reaction Breeding The enzymatic reaction rate, μmol / min * mg The rate of enzymatic reaction,% Rel. The control 2.81 ± 1.68 100.00 D ₆ 3.24 ± 0.80 115.30 D ₅ 4.96 ± 0.52 176.51 D ₄ 5.78 ± 0.64 205.69 D ₃ 4.94 ± 0.23 175.80 D ₁ 3.39 ± 0.45 120.64

Table 7 The effect of tinctures from cactus shoots and homeopathic dilutions on the rate of PK reaction Breeding The enzymatic reaction rate, μmol / min * mg The rate of enzymatic reaction,% Rel. The control 18.94 ± 0.14 100.00 D ₆ 24.38 ± 1.80 128.71 D ₅ 24.24 ± 0.52 128.00 D ₄ 22.52 ± 1.64 118.92 D ₃ 22.63 ± 0.23 119.49 D ₁ 19.58 ± 1.45 103.37

Claims (1)

A method for determining the adaptogenic, antimicrobial, antiviral, antioxidant activity of a homeopathic medicinal product by its activating or inhibitory effect on the reaction rate of an in vitro enzyme test system, characterized in that the homeopathic medicinal product is obtained by sequential progressive dilution of the matrix substance and multiple vertical shaking or rubbing homeopathic method of dynamization, the test system is a set of lyophiliziro of enzymes selected from key or limiting metabolism in a living organism of xenobiotics, such as superoxide dismutase, catalase, glutathione peroxidase, glutathione reductase, acetylcholinesterase, pyruvate kinase, speed measurements are carried out by spectrophotometric control of reaction parameters before and after addition of a test drug to the isolated homeopath moreover, the reaction rate reflects the collective state of the characteristics of the particles of homeopathic medicines of a venous agent, including a submolecular one that does not contain molecules of the initial level, and the pharmacological activity of a homeopathic medicine is determined by the combination of the activating or inhibitory effect on the reaction rate of key or limiting enzymes, where the activation of superoxide dismutase and glutathione reductase inhibition of catalase and glutathione peroxidase corresponds to adapt activation of superoxide dismutase, catalase, glutathione peroxidase, glutathione reductase ie antioxidant activity, inhibition - antimicrobial, antiviral activity, the activation of pyruvate kinase indicates the antioxidant and adaptogenic activity, and inhibition of acetylcholinesterase evidence of adaptogenic activity.

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