1. Introduction

The development of medicines with anti-inflammatory activity is an actual task of pharmacology, since this typical pathological process forms basis of the pathogenesis of many human diseases. Despite the large number of medicines used in the treatment of acute and chronic inflammatory diseases, the effectiveness of their pharmacotherapeutic action is desired to make better, since the development of a number of side effects pose the lives of patients to the serious threat and reduce their value. For this reason, the modern range of anti-inflammatory medicines does not solve the problem of successful treatment of inflammatory diseases and their relapses, the frequency of which, after the abolition of this group of drugs can reach 100% [1]. There is no doubt that the prospect search of new anti-inflammatory drugs is promising, among compounds of natural origin - multicomponent extracts of medicinal plants and their collections [2].

The experience of developing new safe anti-inflammatory drugs convinces that great success can be achieved by searching for nonsteroidal anti-inflammatory medicines among the components of plant raw materials or by combining low toxic chemical compounds with the diverse mechanisms for restoring pathogenetic disorders during inflammatory processes [3-11]. According to the requirements of preclinical studies of new medicines, anti-inflammatory properties should be tested in experiments on models of inflammation induced by various compounds. Earlier, we established the efficacy of dry extract of medicinal plants (DEMP) in some models of aseptic arthritis [4,12-14]. However, the anti-inflammatory activity of DEMP in carrageenan-induced inflammation remained unexplored. This circumstance determined the purpose of this work.

2. Material and Methods

2.1. Plant Material and Preparation of Dried Extract

Dry extract of medicinal plants was obtained from plants: Hipericum scabrum L., Mediazia macrophylla, Glycyrrhiza glabra L. and Ziziphora pedicellata Pazij Vved. Aerial parts of Hypericum perforatum L., Ziziphora pedicellata Pazij et Vved. and Mediazia macrophylla as well as root and rhizome parts of Glycyrrhiza glabra L. were obtained in summer of 2017 from foothills to medium zones of mountains of Tashkent region, Fergana, Samarkand and Surkhandaryo regions of Uzbekistan. Plant material was dried under dark conditions at room temperature for 10 days. Taking into account that the soil is contained various bacterial spores, raw material of plants were treated with special methods. The dry material was milled, obtaining 4-6 mm particles and mixed in proportion 1,25:1,0:1,25:1,5 (productivity of dried extract was higher than other proportions) then extracted by water at 93-95°С temperature for 3 hours. The extract was then separated from the sample residue by filtration through filter paper. The resulting extracts were concentrated in vacuum until remaining a crude solid extract, which was then dried in a thermostat at temperature of 60°C.

2.2. Experiments

All experimental studies were carried out on adult male rats weighing 145-160 g, which were obtained from the vivarium of the Sanitary-Epidemiological Surveillance Department of the Chief Medical Administration under the Administration of the President of the Republic of Uzbekistan. Before the start of the experiment, all laboratory animals were carefully examined, weighed as well as their age, gender, and motor activity were taken into account. During the whole period of preparation for the experiment and its holding, all laboratory animals were kept in a vivarium at temperature 20-25°С, humidity not less than 50%, in a well-ventilated room and light mode day/night as well as by 6 animals in standard plastic cages with a standard diet. The daily requirement for food is compiled according to the age of the animals. Before the beginning of the experiment all laboratory animals had a healthy appearance and were active.

Inflammatory edema of the rat paws was modeled by the subplantational administration of the carrageenan, which are widely used to evaluate the anti-inflammatory activity of new potential drugs [15]. The experimental model of aseptic arthritis was reproduced by subplantary administration of a 1% carrageenin aqueous solution into the hind paw of rats in a volume of 0.1 ml. The preventative efficiency of various doses of DEMP (10, 25, 50 and 100 mg/kg) was studied in comparison with diclofenac sodium (10 mg/kg) and LIV-52 (100 mg/kg) [16]. The above drugs were administered intragastrically with a metal cannula 2 hours before the introduction of carrageenan. Measurement of the volume of paw of animals was performed by plethysmometer before and 1, 2, 3, 4 and 5 hours after the introduction of the carrageenan. The value of the anti-inflammatory activity (VAA) of the drugs was calculated according to the formula:

VAA = Vcon-Vexp/Vcon x 100 =%.

All experiments were performed in compliance with the requirements of the European Convention "On Protection of vertebrate animals used for experimental and other scientific purposes" (Strasbourg 1986).

2.3. Statistical Analysis

The received results were subjected to the statistic processing with the using of standard software package Biostat 2009 on well-known method of variation statistics with an estimation of the statistical significance of indicators (M±m) and differences between groups were analyzed using the Student’s t-test. P<0.05 was considered significant.

3. Results and Discussion

The obtained results (table 1) showed that in healthy animals under the influence of carrageenan, there was an expressed increase of the paw volume by 62.1% after one hour, and more than twice after three and four hours from the injection of carrageenan compared to the initial paw volume. At the same time, the volume of the paw even after 24 hours remained increased by 26.4%. It is considered that in the first hours of the inflammation process induced by carrageenan determines by the action of kinins, and in later periods (after three and four hours) - prostaglandins [17].

Table 1. Comparative study of the influence of dry extract of medicinal plants, LIV-52 and diclofenac sodium on course of aseptic arthritis induced by carrageenan

It is seen from the data given in table 1 that the increase of paw volume under the influence of carrageenan was noticeably low in animals, which were administered reference nonsteroidal anti-inflammatory drug, diclofenac sodium. After one hour, it increased to 41.1%, and after two hours - 46.6%, after three hours - 53.4% and by the end of the fourth hour - 37.0% compared to the initial volume of paw. At the same time, after one day from the start of the experiment, the paw volume of rats administered diclofenac sodium did not practically differ from the initial values.

The anti-inflammatory activity of diclofenac sodium in the first phase of carrageenan action was 33.3%, and in the second phase 50.6 - 60.3%. It is seen that diclofenac sodium has a more expressed influence on the prostaglandin than the kinin system. Similar in character of directionality of action, we found in rats preventively received LIV-52 - a preparation representing the sum of medicinal plants with an antioxidant effect [18,19]. However, its anti-inflammatory activity was noticeably low in carrageenan-induced inflammation compared with diclofenac sodium.

It is seen in table 1 that the suppression of the intensity of aseptic inflammation under the influence of DEMP was observed particularly in doses of 50 mg/kg and 100 mg/kg. At the same time, the paw volume decreased under the influence of the last doses of the preparation by the end of the 24 hours of the experiment and paw volume of animals did not practically differ from the initial values. It is characteristic that DEMP at a dose of 50 mg/kg was rather superior to diclofenac sodium by its influence on the kinin system, and in the second phase, the effect of the compared preparations is not significantly different.

Consequently, dry extract of medicinal plants is superior to LIV-52 in its anti-inflammatory activity in aseptic arthritis induced by carrageenan and is not inferior to diclofenac sodium. The modern concept of inflammation considers inflammation - a pathophysiological phenomenon from the standpoint of the decisive participation of oxidative stress in it [20].

It is notable that DEMP and diclofenac sodium have the same type of effect on both phases of carrageenan-induced inflammation. We have previously shown the low toxicity of DEMP [21], which has hepatoprotective [22], choleretic [23] activity and the mechanism of which is probably related to the inhibition of free radical processes [24]. Based on this, we assumed that using DEMP as a medicine for treatment a number of diseases, which the inflammation processes has significance in their pathogenesis, might be promising. This gives the right to raise the question of the possibility of including DEMP in complex traditional therapy of inflammatory diseases, after conducting adequate clinical studies.

As noted, obtained results by the authors, DEMP has both a high choleretic, hepatoprotective, anti-inflammatory activity and has an advantage over the known non-steroidal anti-inflammatory drug, since it is a compound with low toxicity and availability of plant raw materials in Uzbekistan.

Despite the evidence of the presented results, in our opinion, it is premature to declare about its clinical demand. Further, it is necessary to investigate the ability of DEMP to interfere to the course of various inflammatory processes in-depth studies not only in the experiment, but also in clinical studies.

4. Conclusions

1. The determined suppression of the exudative stage of inflammation by dry extract of medicinal plants indicates its effectiveness in aseptic inflammation.

2. The studied dry extract is not inferior to the reference non-steroidal anti-inflammatory drug- diclofenac sodium by its anti-exudative activity, especially at a dose of 50 mg/kg.

3. The studied dry extract of medicinal plants surpasses diclofenac sodium by the effect on the kinin system.

4. Dry extract of medicinal plants after further relevant in-depth studies can be recommended into practical medicine as an effective anti-inflammatory medicine.

ACKNOWLEDGEMENTS

The work was supported by national grant PZ -20170926458.