EXTRACTION OF SECONDARY METABOLITES FROM Agaricus bisporus FUNGUS

ABSTRACT

Considered one of the most widely consumed mushrooms worldwide, Agaricus bisporus, it combines properties with high biological activity. This species is suitable for breeding in most cases when growing in greenhouses without culture. But there is also a growing group in natural sharaoite, whose metabolites are different from the rest. As part of the study, its metabolites containing the fungus Agaricus bisporus, which grows naturally in sharaoite, were isolated. At the same time, several chromatography techniques and various solvents were used. Several metabolites were isolated using the chloroform: methanol (1:1) system as an elutant and were presented with several data on their biologic activity and scope of application.

АННОТАЦИЯ

Считающийся одним из наиболее широко употребляемых в пищу грибов в мире, Agaricus bisporus, он сочетает в себе свойства с высокой биологической активностью. Этот вид подходит для селекции в большинстве случаев при выращивании в теплицах без культивирования. Но существует также растущая группа в природном условим, метаболиты которого отличаются от остальных. В рамках исследования были выделены его метаболиты, содержащиеся в грибе Agaricus bisporus, который произрастает в природе в шараоите. При этом были использованы несколько хроматографических методов и различные растворители. Несколько метаболитов были выделены с использованием системы хлороформ:метанол (1:1) в качестве элюента и представлены с данными об их биологической активности и области применения.

Keywords: Agaricus bisporus, metabolite, fungus, chromatography, system, fraction.

Ключевые слова: Agaricus bisporus, метаболит, гриб, хроматография, система, фракция.

Introduction

In this review, the isolation of secondary metabolites from fungal species of the genus Agaricus and their activity are discussed. 2 novel and 3β,11,12-trihydroxydrimen driman sesquiterpenoids such as 11,12-dihydroxy-15-drimeneoic acid and 3α,11,15-trihydroxydrimen have been isolated from the cultural fluid of Agaricus arvensis basidiomycete [1, p. 305]. Another study of Agaricus bisporus fish to verify that it is suitable for its composition and qualitatively determined the presence of flavonoids, terpenoids, saponins, polyphenols and heart glycosides in an extract of A. bisporus made from methanol. In the petrolium essential extracts of A. bisporus biomass it has been known to be compounds belonging to the class such as terpenoids, alkaloids, polyphenols, tannins, heart glycosides, and quinins. 14 secondary metabolites in A. bisporushave been identified, of which 9 are: Tetradecanoic acid, Dodecanoic acid; Hexadecanoic acid, methyl ether; 2-Pentadecanoic acid; 6, 10, 14-trimethyl4-(6,6-Dimethyl-2-methylencyclohex-3-enylid)pentane-2-ol; Neophytadien; Tetradecanoic acid; Hexadecanoic acid; 9, 12, 15-Octadecactrinic acid, ethyl ether, (Z, Z, Z)- which have been found to be biolgic active compounds [2]. Agaricus fungi are distributed in large quantities in nature, containing mainly steroids, nitrogen compounds, sphingoids, terpenoids, aliphatic compounds and other structural species, some of which contain anti-tumor, anti-neurolysis, antibacterial, anti-angiotensin, anti-angiotensin conversion specific enzyme activity [3].  In another GC-MS study on butanol extracts, 44 compounds were identified for A. placomyces and 43 for A. pseudopratensis, namely: amino acids, fatty acids and their ethers, and sugar alcohols. In addition, the most common phenol and urea compounds, as well as the presence of ergosterol and 2 Delta7-sterols, were studied, and the main one was isolated from both fungi for the first time 5alpha,8alpha-Epidioxi-24(xi)-methylcholesta-6,22-diene-3beta-ol [4]. In Mongolia, 4 known substances such as urea, mannitol, ergosterol and ergosterol peroxide have been isolated for the first time in ethanol (95%) extracts of Agaricus silvaticus Schaeff ex. Secr [5, p. 92]. A methylated analogue of benzothiazole was shown in white-button mushrooms, which was subsequently isolated and identified as 2-amino-4-methyl-benzothiazole. Cell assay AHR transcription results revealed that 2-amino-4-methyl-benzothiazole has agonistic activity [6]. Agaritin (l-glutamic acid, 5-(2-(4-(hydroxymethyl) phenyl) hydrazide)) in spores of Agaricus bisporus was determined by mass spectrometric method using negative electron ionization using high-resolution liquid chromatography [7, p. 521]. Novel metabolites of agaridiol (1) Agaricus sp. and its structure was determined by mass spectrometric and NMR spectroscopic studies [8, p. 213].

Research methodology.

Together with the caps and fruits of the fungus Agaricus bisporus, grown in natural conditions, was dried at room temperature and crushed and biomass was obtained. The resulting mushroom mass was extracted by the tincture method 15 times in 96% ethanol solution. The resulting biomass was dried and placed for fractionation by column chromatography method. Fractionation was carried out in various solvents at different ratios. The obtained fractions were examined and analyzed by thin-layer chromatography and gas chromatography and mass spectrometric analysis.

Unknown volatile substances were detected in the mass spectrometric detector by gas chromatography of the YL 6900 GX/MS (30 m × 0.25 mm internal diameter, 0.25 μm film thickness) equipped with the DB-5MS column. Furnace was used as a carrier gas at an initial temperature of 80°C/3.0 min, heating rate from 15°C/min to 250°C, containment - 3.0 min, and Helium at a flow rate of 1.0 ml/min. Evaporator temperature was 280°C, flow section - 1/20, analysis time - 17 min. Liquid samples were injected into the spatula using a 1 μL micro syringe. The temperature of the transmission line was 300°C, the ionization voltage was –70 eV, and the temperature of the ion source was 230°C.  Scanning range – 30-350 a.m.u. The components were determined from the mass spectra of each component after comparing it with the spectral data available at MS Library NIST 2017.

Results and discussion

In the chloroform:methanol 1:1 system, GC-MS analysis of the formed fraction was obtained (Fig. 1), in which several metabolites were separated (Table 1).

Figure 1. Chromatogram of the fraction of the chloroform:methanol 1:1 system

Table 1.

Chloroform: methanol metabolites degraded in the 1:1 system

Most of the metabolites released in this fraction are considered high fatty acids and their ethers, while some are polyatomic alcohols (polyols) and lactones.

β-Sedogeptitol (D-glycero-D-gluco-heptitol) (Fig. 2) is a sweet-tasting, seven carb sugar alcohol (alditol), often found in combination with other sugar alcohols, such as volemitol, and exhibits diagnostic metabolite as well as antioxidant-like properties. 2(3H)-Furanone (Fig. 3), while dihyro-4-hydroxy is used as intermediate in organic synthesis, pharmaceutical research, and in several industries such as food and cosmetics.

The 1,4-benzenedicarboxylic acid, bis(2-ethylhexy) ester (Fig. 4) acts as an emollient and solvent in the cosmetic industry and acts as a plasticizer. And sorbitol (Fig. 5) Ais used in cosmetics, food industry and as medicine.

Conclusion

From the superior results, it can be seen that almost all of the obtained secondary metabolites contain unique biolgic activity, which is known to have function properties in various industries like this.

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