MD83Y - Fruit biomass of Aronia melanocarpa (Michx.) Elliot, obtained in vitro, for use as a product with antimicrobial properties - Google Patents

Abstract

The invention relates to a source of plant raw material with antimicrobial properties, which can be used in medicine. The essence of the invention is that the fruit biomass of Aronia melanocarpa (Michx.) Elliot, obtained in vitro, is proposed for use as a product with antimicrobial properties. The result of the invention consists in the fact that the aroma fruit biomass has a broader bacteriostatic and bactericidal spectrum than the aronia fruits.

Description

[0001] The invention relates to a source of plant raw material with antimicrobial properties, which can be used in medicine.

[0002] Recently, strains of microorganisms with increased resistance to the action of antibiotics have been increasingly encountered. The development of new antimicrobial remedies of plant origin, rich in phenolic content, known as natural substances with various therapeutic effects, including antibacterial and bacteriostatic, is one of the priority directions in research. In this context, the fruits of Aronia melanocarpa (Michx.) Elliot are of particular interest, the extract of which is characterized by different classes of phenolic compounds and shows sensitivity to microorganisms Staphylococcus aureus, Enterococcus faecalis, Escherichia coli [1], but not to other virulent microorganisms. Aronia fruits show antimicrobial action to a reduced spectrum of microorganisms, and the production of fruits in vivo is dependent on pedo-climatic conditions, natural cataclysms, seasonal rotation, the consequences of the ecological crisis - these representing essential disadvantages.

[0003] The problem solved by the invention consists in expanding the range of raw materials with antimicrobial properties, obtained under in vitro conditions.

[0004] The essence of the invention lies in the fact that the biomass of Aronia melanocarpa (Michx.) Elliot fruits, obtained in vitro, is proposed for use as a product with antimicrobial properties.

[0005] The result of the invention is that the biomass of chokeberry berries possesses a broader bacteriostatic and bactericidal spectrum compared to chokeberry berries.

[0006] Embodiment of the invention

[0007] Aronia melanocarpa (Michx.) Elliot fruit biomass was obtained from in vitro explants, taken from chokeberry fruits, subsequently sterilized with Diacid chemical agent for 7 min and washed 3 times with sterile double-distilled water. The explants were inoculated on Murashige-Skoog nutrient medium supplemented with growth regulators: naphthylacetic acid 3.0…3.5 mg/L; kinetin 0.5 mg/L; sucrose 60 g/L, having pH 5.7; cultivated at a temperature of 18…22°C, with a day-night photoperiod of 16 and 8 hours respectively and under aseptic conditions.

[0008] For the determination of antimicrobial activity, ethanolic extracts from chokeberry fruits and in vitro obtained fruit biomass of A. melanocarpa were prepared. The samples to be analyzed (chocolate berries and in vitro fruit biomass, respectively) in an amount of 20 mg, ground in a mortar and 80 mg of 20% ethyl alcohol (ratio 1:5) were left for extraction for 10 days, at a temperature of 4°C, in the dark. The 20% ethanolic extracts from the fruits and in vitro obtained fruit biomass served for the study of antimicrobial activity.

[0009] The antimicrobial action of the phenolic content of 20% ethanolic extracts of in vitro obtained fruit biomass and chokeberry fruits was determined by bacteriostatic activity (minimum inhibitory concentration - MIC), established in the absence of growth of microorganisms in the liquid nutrient medium, and bactericidal activity (minimum bactericidal concentration - MBC) based on the absence of growth of microorganisms after repeated seeding on peptone agar with subsequent thermostating for 24 hours. The following reference cultures served: Staphylococcus aureus (strain 209-P), Enterococcus faecalis, Escherichia coli (strain ATCC 25822), Pseudomonas aeruginosa (strain ATCC 27853), Proteus vulgaris (strain MX 19222).

[0010] Part of the reference cultures S. aureus (strain 209-P), E. faecalis, E. coli (strain ATCC 25822), P. aeruginosa (strain ATCC 27853), P. vulgaris (strain MX 19222), grown on agarose for 18 hours, washed with isotonic sodium chloride solution, was seeded at a dose of 500 thousand microbial bodies per 1 ml of nutrient medium (according to the optical turbidity standard) with the addition of 20% ethanolic extract of Aronia melanocarpa (Michx.) Elliot fruit biomass, obtained in vitro.

[0011] Another part of the same reference cultures was inoculated at a dose of 500 thousand microbial bodies per 1 ml of nutrient medium (according to the optical turbidity standard) with the addition of 20% ethanolic extract of Aronia melanocarpa (Michx.) Elliot fruit biomass. As a test-control, nutrient media with the same strains of reference bacteria without the content of the analyzed extracts served.

All experiments were performed in triplicate.

[0013] The phenolic content of the main groups of phenolic compounds, which determine the antimicrobial activity, was studied, and the results are presented in Tab. 1.

[0014] Table 1

[0015] Content of phenolic compounds in fruits and fruit biomasses obtained in vitro and from chokeberry fruits in vivo

The analyzed batch

Total polyphenol content (mg/kg-1)

Flavonoids (g % in rutoside)

Anthocyanins (mg/100g fruit)

C6-C3 phenylpropanic compounds (mg/ml)

Aronia berries

58 370

0.478

821

0.178

Aronia melanocarpa (Michx.) Elliot fruit biomass obtained in vitro

65 570

0.483

680

0.375

[0017] After studying the antimicrobial activity of the 20% ethanolic extracts of the analyzed batches, it was determined that the in vitro obtained aronia fruit biomasses possess bacteriostatic and bactericidal action against the microorganisms S. aureus (strain 209-P), E. faecalis, E. coli (strain ATCC 25822), P. aeruginosa (strain ATCC 27853), P. vulgaris (strain MX 19222), while aronia fruits only against S. aureus (strain 209-P), E. faecalis, E. coli (strain ATCC 25822). The intensity of the antimicrobial activity of the analyzed extracts correlates with the phenolic content (table 1).

[0018] The biomass of fruits obtained in vitro is characterized by a higher content of polyphenols, flavonoids and phenylpropanic compounds than chokeberry fruits and possesses a higher antimicrobial activity with a broader bacteriostatic and bactericidal spectrum (tab. 2).

[0019] Table 2

[0020] Bacteriostatic and bactericidal activity of ethanolic extracts of

[0021] fruit biomasses obtained in vitro and chokeberry fruits obtained in vivo

20% alcoholic extract analyzed

Reference cultures

S. aureus (t. 209-P)

E. faecalis

E. coli (strain ATCC 25822)

P. aeruginosa (strain ATCC 27853)

P. vulgaris (t. MX 19222)

CMI

WBC

CMI

WBC

CMI

WBC

CMI

WBC

CMI

WBC

In vivo obtained chokeberry fruits

1:2

1:2

1:2

1:1

1:2

1:1

-

-

-

-

In vitro fruit biomass

1:4

1:4

1:2

1:2

1:2

1:2

14

1:4

1:2

1:2

20% ethyl alcohol, control

1:1

1:1

1:1

1:1

1:2

1:1

1:4

1:1

1:1

1:1

[0023] Legend: MIC - minimum inhibitory concentration; MBC - minimum bactericidal concentration.

1. Valcheva-Kuzmanova S., Russev V., Bojkova K., Belcheva A. Study of natural Aronia Melanocarpa fruit juice for antibacterial and antiviral activity. Scr. Sci. Med. 2003, vol. 35, pp. 21-23

Claims (1)

1. Biomass of Aronia melanocarpa (Michx.) Elliot fruits, obtained in vitro, for use as a product with antimicrobial properties