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Incapsulation of Pre- and Probiotics

#4053


Polyfunctional complexes of pre- and probiotics: the target delivery to the different regions of the human gastrointestinal tract with use of biocompatible polymers

Tech Area / Field

  • CHE-POL/Polymer Chemistry/Chemistry
  • BIO-MIB/Microbiology/Biotechnology

Status
3 Approved without Funding

Registration date
02.04.2010

Leading Institute
INEOS (Organo-Element Compounds), Russia, Moscow

Supporting institutes

  • Institute of Immunological Engineering, Russia, Moscow reg., Lyubuchany

Collaborators

  • University of the Pacific / School of Pharmacy and Health Sciences / Department of Chemistry, USA, CA, Stockton\nMcGill University / Department of Chemistry, Canada, QC, Montreal\nUniversité Catholique de Louvain / Institute of Condensed Matter and Nanosciences, Belgium, Louvain-la-Neuve

Project summary

The Project aim. The project aims are to develop a system of encapsulation and directed joint delivery of probiotics and prebiotics into the human gastrointestinal tract by creating a new macromolecular systems based on bio-compatible natural and synthetic polymers.

Current status. Probiotics are live microorganisms which, at prescribing in sufficient quantity has a positive impact on human health. The use of probiotics may have the following effects on the organism: anticontagious stimulation of protective mechanisms, improve the barrier function, metabolic effects, changes in bowel function and motility. Immunity of probiotic bacteria to antibiotics and other antibacterial drugs creates the possibility of using probiotics for the correction of dysbiosis arising during treatment with antibiotics.

Prebiotics (carbohydrates, which are nutrients for probiotics) may have great influence on intestinal microflora. By probiotic include Lactobacterium acidophilus, Streptococcus salivarius thermophilus, Lactobacillus delbrueckii subsp.bulgaricus, L. casei and Bifidobacterium, which contribute to the prevention of intestinal infections due to competitive inhibition of growth of pathogenic bacteria, including Staphylococcus, Salmonella and others, produced by the antimicrobial action of organic acids and antibacterial agents. For example, yogurt must contain around a million probiotic bacteria per gram, to provide the desired effect, and these bacteria-probiotics must survive at the the acidic environment if it enters the human stomach. One of the main directions in the design of new probiotic products is the encapsulation of probiotic microorganisms with the use of bio-compatible artificial and synthetic biopolymers.

Chitin, poly [β-2-acetamido-2-deoxy-D-glucopyranose], is the second most common organic material after cellulose. The main source of chitin is crustacean shells. Chitosan is a product of deacetylation of chitin.

Chitosan is a growing application in the food and medical industry as a thickener, foam and antibacterial and fungicidal tool fungicidal funds. Chitosan and its oligomers is practically non-toxic. United States Agency for food and medical products (US Food and Drug Administration) recognizes chitosan as a food additive since 1983. Chitosan oligomers recognized by the United States Agency for Environmental Protection (US Environmental Protection Agency) environmentally friendly c 1995, when they began to be used as pesticides in agriculture.

Oligohitozan is a mixture of chitooligosaccharides with a molecular weight of less than 16 kDa, obtained by partial depolymerization of chitosan. Oligohitozan as a prebiotic, helps the growth of the majority of probiotic bacteria, is a biocompatible non-toxic substance and immunomodulator. At the same time oligohitozan suppresses the growth of pathogens such as Candida sp., Bacillus spp, Aspergillus, Mucor and Penicillium species, at a concentration of 0.01-0.1%. In this project for the encapsulation of the probiotic microorganisms oligohitozan will be used.

The project’ influence on progress in this area. In case of successful completion of the planned experiment will be established probiotic products that effectively protect the passage of microorganisms in acidic conditions of the gastrointestinal tract, expelling pathogenic bacteria and allowing them to operate efficiently, providing a positive influence on the human body, including by strengthening its natural immunity.

The participants’ expertise. INEOS employees participating in the project have extensive experience in the synthesis of biocompatible polymers in the production and modification of oligochitosanes. IIE employees have extensive experience in providing various strains of microorganisms in the selection of antibodies and antigens of infectious diseases, have experience of working with living cells. This is supported by the available publications and presentations at international conferences.

Expected results and their application. As a result of the project will be the methods of chitosan hydrolysis development to the stage of the modified oligochitosanes with different molecular weights and high yields (70%). The hydrophobically-modified oligochitosanes also will be obtained. The synthesis of poly [(2-dimetilaminoetil) methacrylate will be carried out. There will be physical and chemical studies of polymers. Be Microporous materials based on oligohitozana using supercritical CO2 will be obtained and the porosity will be investigated. The synthesis of block copolymers based on poly [(2-dimetilaminoetil) methacrylate and polyethylene oxide with various units of methacrylic copolymer will be carried out. We will obtain new original strains of lactobacilli and bifidobacteria with promising probiotic properties. The new probiotic strains will be characterized by their ability to withstand the LCD-stress, resistance to oxygen, the presence of antioxidant and antimutagenic activity, stimulation of natural immunity, antagonistic activity and antibiotic resistance. Low molecular weight hydrophobically modified chitosan for use as prebiotics will be prepared. Microporous chitosan will be used for the probiotics encapsulation. The methods of producing microporous polymeric materials using supercritical carbon oxide to methacrylic block copolymers will be developed. The encapsulation of probiotics with amphiphilic block copolymers on the principle of "micelles" in polyelectrolyte microcapsules will be created based on the layered deposition of oppositely charged layers with the participation of the modified chitosan and a synthetic polymer. Graft-copolymers chitosan / synthetic polymer and porous materials obtained on their basis will be obtained. The probiotic strains, encapsulated in polymer matrices, in acid environment will be studyed. The increased accumulation titer of bacteria encapsulated in polymeric matrices that maintain their probiotic activity with increasing storage time and ability to withstand the LCD stress will be demonstrated. There will be an encapsulation of living cells in the polymer matrices and enzymatic and acid-resistance stability will be investigated. There will be physical and chemical studies. The results will be used to develop the food additives.

Meeting the ISTC goals and objectives. Running the project will contribute to solving important social problems, namely, the provision of engineering researchers at the Institute of Immunology and INEOS, who worked previously on government orders in anti biological and chemical protection, as well as possibilities to shift the terms of their scientific interests and accumulated experience for carrying out the basic and applied studies related with the decision of the international scientific and technological problems in biology, medicine and veterinary science for peaceful purposes, the establishment of long-term prospects for productive activities within the international scientific community. Also, to meet the goals of the ISTC is to involve the scientists from the participating organizations to participate in international conferences and seminars, publishing articles in domestic and international journals.

Scope of activities.

  1. Selection of probiotic strains with enhanced stability in acidic medium and with the ability to assimilate cholesterol.
  2. Development of technology for oligosaccharides of chitosan preparation.
  3. Creating of encapsulation and delivery system of probiotics. Encapsulation of probiotics, and then their directed delivery, and prolonged action can be realized through the creation of new macromolecular systems based on natural or synthetic biocompatible polymers. Several approaches for the encapsulation of bacteria and their directed delivery will be used:
    1. a) in the presence of prebiotics - oligosaccharides of chitosan;
    2. b) the use of supercritical CO2. Polymethacrylic biocompatible polymers, oligochitosane and their copolymers or blends will be used.
    3. c) based on diblock-copolymers, self-assembled on the principle of micelles, where the core is bacteria.
Role of Foreign Collaborators/Partners. As collaborators of this project have expressed a desire to serve members of the scientific group of Prof.. Prof. Jean-Francois Gohy of UCL Université catholique de Louvain. Institute of Condensed Matter and Nanosciences (IMCN), Department Bio-and Soft Matter (BSMA), Leuven, Belgium. Prof. Goya is an expert in the field of micelle formation of amphiphilic block copolymers. According to Professor. Goya on March 22, 2010, he took great interest in this field of research and would like to cooperate in terms of joint discussions on the results of the project at conferences and exchange visits.

As collaborators of the project have also expressed a desire to serve members of the scientific group Prof. Adi Eisenberg of McGill University, Otto Maass Chemistry Building, Department of Chemistry, Montreal, Canada. Prof. Eisenberg engaged biocompatible pH-dependent polymers, and their behavior in the presence of DNA. According to Professor. Eisenberg on March 22, 2010, their group is ready to cooperate fruitfully with INEOS and OAO IRS in the performance of the project.

Technical approach and methodology. The methods of technical chitosan to oligochitosan hydrolysis will be developed. The methods of radical polymerization, with the transition to the methods of controlled radical polymerization will be used. A method of supercritical CO2 for the creation of microporous polymeric structures, as well as study of the porosity of new materials by nitrogen adsorption (BET) will be developed. The search for finding and study of new strains of probiotic microorganisms using microbiological methods in aerobic and anaerobic conditions of cultivation will be made. Probiotic properties of strains of lactic acid bacteria will be evaluated on the ability to withstand exposure to low levels of acidity and resistance to bile (test in vitro, gastrointestinal stress). Bifidobacteria will be evaluated in the resistance test to oxygen. Microbiological method of determining the sensitivity of lactic acid bacteria to antibiotics will be used. The enzymatic method for estimating the antioxidant activity and anti mutagenic activity (multiorgan micronucleus test) of new strains of probiotics will be used. Microbiological method will determine the antagonistic activity of probiotics strains in relation to the test-strains of opportunistic microorganisms of gastrointestinal microflora. Using the microbiological methods the acid resistance of probiotic microorganisms encapsulated in polymer matrices will be determined.


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