STRUCTURAL AND METABOLIС DERIVATIVES OF BIFIDOBACTERIUM BIFIDUM AND LACTOBACILLUS REUTERI CELL-FREE EXTRACTS
About the author:
Knysh O. V., Pakhomov O. V., Pogorila M. S
CLINICAL AND EXPERIMENTAL MEDICINE
Type of article:
Probiotic strains of Bifidobacterium bifidum and Lactobacillus reuteri have beneficial properties that contribute to the promotion of human health. Most of the beneficial effects of probiotics are due to the action of their structural components and metabolic products. Bacterial metabolic products are usually obtained by cultivating the producer in a liquid nutrient medium. Structural components of bacteria can be obtained by applying various methods of disintegration: chemical, mechanical, enzymatic, physical and combined. Use of different in nature and intensity of destructive factors causes different in mechanism and depth of cell damage. Therefore, disintegrates obtained by various ways have different composition. This determines their various biological activity and purpose of application. It is known that bacteria are more resistant to freezing-thawing than eukaryotic cells. For their destruction, repeated freezing-thawing (thermal cycling) or combination with other disintegration methods is necessary. Preliminary studies have established the high biological activity of filtrates of B. bifidum and L. reuteri disintegrates, obtained by thermal cycling, and culture filtrates of these probiotics, cultured in disintegrates of their own cells. The aim of the study was to investigate the biochemical composition of cell-free extracts obtained from disintegrates and cultures of B. bifidum and L. reuteri cultured in their own disintegrates. Methods. Cell-free extracts were obtained by disintegration of L. reuteri and B. bifidum probiotic strains (L and B extracts) or by cultivation of these strains in their own disintegrates (ML and MB extracts). The pH values of the extracts, the content of protein, lactic, teichoic and amino acids, RNA and DNA were determined. The mass fraction of nitrogen by the Kjeldahl method was determined and the mass fraction of crude protein was calculated. Molecular weight distribution of peptide fractions of cell-free extracts was performed using high-performance gel permeation chromatography. Results. The pH values of the ML and MB extracts were significantly lower than the corresponding values of the L and B extracts. This indicated that during the cultivation of probiotic microorganisms in their own disintegrates, an accumulation of acidic metabolic products was occurred. The results of measuring the concentration of lactic acid confirmed this: the lactate content in ML extract was 2 times higher than in L extract, and in MB extract 1.5 times higher than in B extract. All investigated cell-free extracts contained a significant amount of teichoic acids and a small amount of nucleic acids. The content of protein, peptides, amino acids and nitrogen in ML and MB extracts was significantly higher than in L and B extracts. ML extract was superior to the rest of these extracts in the quantitative content of proteins, peptides, nitrogen and free amino acids. The highest amino acid content in the ML extract indicate d a higher productivity of lactobacilli compared to bifidobacteria in the selected study conditions. Conclusion. The protein components in the extracts are predominantly peptides with a molecular weight of less than 12 kDa. The content of protein components in extracts from cultures of probiotics is higher than in extracts from disintegrates. Teichoic, nucleic and lactic acids, low pH values (4,12 – 5,43) point to a potential antimicrobial and immunotropic activity of investigated extracts.
cell-free extracts, proteins, peptides, teichoic acids, lactic acid, DNA, RNA.
- Mu Q, Tavella V, Luo XM. Role of Lactobacillus reuteri in human health and diseases. Frontiers in microbiology. 2018;9:757. DOI: 10.3389/ fmicb.2018.00757
- Quigley EMM. Bifidobacterium bifidum. In Floch MН, Ringel Y, Walker WА, editors. The microbiota in gastrointestinal pathophysiology: implications for human health, prebiotics, probiotics, and dysbiosis. London: Academic Press; 2017. Part B, Chapter 14. P. 131-4. DOI: 10.1016/ B978-0-12-804024-9.00008-2
- Caselli M, Vaira G, Girolamo C, Papini F, Holton J, Vaira D. Structural bacterial molecules as potential candidates for an evolution of the classical concept of probiotics. Advances in Nutrition. 2011;2:372-6. DOI: 10.3945/an.111.000604
- Singh A, Vishwakarma V, Singhal B. Metabiotics: the functional metabolic signatures of probiotics: current state-of-art and future research priorities. Advances in Bioscience and Biotechnology. 2018;9:147-89. DOI: 10.4236/abb.2018.94012
- Bauer JA, Salvagni M, Vigroux JPL, Chalvet LLG, Chiavaroli C, inventors; Vifor Pharma (Switzerland), assignee. Immunomodulatory extracts from lactobacillus bacteria and methods of manufacturing and use thereof. European patent 2540818-A1. 2013 Jan 2.
- Kapustian A, Cherno N, Stankevich G, Kolomiіets I, Matsjuk O, Musiy L, Slyvka I. Determination of the enzyme destruction rational mode of biomass autolysate of lactic acid bacteria. Eastern-European Journal of Enterprise Technologies. 2018;1(1):1. DOI: 10.15587/1729- 4061.2018.120877
- Vora A. Bacterial lysates. RGUHS Journal of Medical Sciences. 2016;6(3):110-1.
- Lindquist JA, Mertens PR. Cold shock proteins: from cellular mechanisms to pathophysiology and disease. Cell Communication and Signaling. 2018;16(1):63. DOI: 10.1186/s12964-018-0274-6
- Knysh OV. Bifidogenic properties of cell-free extracts derived from probiotic strains of Bifidobacterium bifidum and Lactobacillus reuteri. Regulatory Mechanisms in Biosystems. 2019;10(1):124-8. DOI: 10.15421/021919
- Knysh OV, Isaenko OY, Babych EM, Kompaniets AM, Pakhomov OV, Polyanska VP, et al. Antimicrobial activity of bifidobacteria derivatives after storage in a frozen state. Problems of Cryobiology and Cryomedicine. 2018;28(3):237-48. DOI: 10.15407/сryo28.03.237
- Baddiley J, Davison AL. The occurrence and location of teichoic acids in lactobacilli. Microbiology. 1961;24(2):295-9.
- Lowry OH, Rosebrough NJ, Farr AL, Randall RJ. Protein measurement with the Folin phenol reagent. Journal of biological chemistry. 1951;193:265-75.
- Galchenko SE. Tissue and age specificity of aqueous-saline extracts from cryopreserved xenoorgans fragments. Problems of Cryobiology and Cryomedicine. 2005;15(2):153-8.
- International Organization for Standardization (ISO). (2005). ISO 13903: 2005. Animal feeding stuffs–Determination of amino acids content. Available from: https://www.iso.org/standard/37258.html
- ISO 5983-1:2005 Animal Feeding Stuffs – Determination of Nitrogen Content and Calculation of Crude Protein Content – Part 1: Kjeldahl Method. International Organization for Standardization: Geneva, Switzerland. Available from: https://www.iso.org/standard/39145.html
Publication of the article:
«Bulletin of problems biology and medicine» Issue 1 (155), 2020 year, 145-148 pages, index UDK 615.33:[579.873.13+579.864.1]:579.222