Classifications

• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K8/00—Cosmetics or similar toiletry preparations
  • A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
  • A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
  • A61K8/60—Sugars; Derivatives thereof
  • A61K8/606—Nucleosides; Nucleotides; Nucleic acids
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
  • A61Q17/00—Barrier preparations; Preparations brought into direct contact with the skin for affording protection against external influences, e.g. sunlight, X-rays or other harmful rays, corrosive materials, bacteria or insect stings
  • A61Q17/04—Topical preparations for affording protection against sunlight or other radiation; Topical sun tanning preparations
• • C—CHEMISTRY; METALLURGY
  • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
  • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
  • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
  • C12N15/09—Recombinant DNA-technology
  • C12N15/10—Processes for the isolation, preparation or purification of DNA or RNA
  • C12N15/1003—Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses therefor

Definitions

• the present invention relates to a method for obtaining active products rich in deoxyribonucleic acid (DNA) from an animal source.
• DNA deoxyribonucleic acid
• Deoxyribonucleic acid is a molecule of capital importance in biology since it is the molecule generating life: it contains in coded form the genetic information whose programmed emission is the basis of the formation of any living organism.
• DNA has become a molecule of cosmetic interest, in particular for its. properties of absorption of ionizing radiation and, after biological depolymerization in contact with the human epidermis, by its capacity to supply in quality and quantity of the essential nutrients for the renewal and the regeneration of the epidermis.
• DNA is a product of current consumption in particular as a standard, precursor or initiator of biosynthesis.
• the sources' usual DNA are yeasts, certain plants and some animal products such as fish sperm or calf thymus. Extraction methods from these different sources are described in particular in the following documents:
• All of these known sources have the main drawback of requiring the elimination of contaminating macromolecules such as structural proteins, polysaccharides, ribonucleic acids (RNA), lipids, enzymes, as well as residues from cell walls.
• RNA ribonucleic acids
• lipids lipids
• enzymes as well as residues from cell walls.
• a major difficulty encountered during the production of DNA from plant cells relates to the inhibition of deoxyribonucease (DNAse) so that the DNA is not degraded before its extraction.
• DNAse deoxyribonucease
• One of the aims of the present invention is therefore, on the one hand, to select a DNA source comprising a priori few products capable of being considered as contaminants and, on the other hand, to provide a process extracting the DNA contained in this source in order to obtain active products rich in DNA
• Another object of the invention is to provide such a simple and economical method of obtaining which makes it possible to produce active products rich in DNA of good quality and at a low cost price.
• this process for obtaining comprises the following steps:
• the animal source used to obtain DNA is the blood of poultry or camelids.
• birds and camelids are the only homeothermic animals with nucleated erythrocytes, or red blood cells, whose DNA is metabolically inactive.
• the blood of birds such as poultry blood is currently a slaughter byproduct difficult to recover and having no market value due to its polluting nature.
• the blood is collected at the time of the bleeding of the poultry directly on the slaughter line, then conveyed to a storage tank in which the blood is mixed with a preservative or anticoagulant such as a solution of sodium citrate with 3%.
• the anti-coagulant can be introduced into the blood stream when it is transported to the storage tank.
• the temperature of the latter is maintained at a temperature of the order of 4 ° C. in order to ensure correct preservation of the blood.
• the separation of blood cells made up of approximately 98% erythrocytes, is carried out by centrifugation: the supernatant or plasma is eliminated and recovered with a view to recovering its components, some of which constitute interesting substitutes for the components of bovine plasma.
• a buffer solution is used at the rate of one liter of buffer per liter of blood to centrifuge.
• the buffer solution used is composed, for example, of: 150m M sodium chloride (NaCl), 1 mM Tris (hydoxy methyl) amino - methane - hydrochloric acid (Tris-Hcl), and its pH is 7.2.
• the supernatant essentially comprising the blood plasma is removed and a pellet containing the blood cells, including the erythrocytes, is recovered. If necessary, this pellet can be put back into solution in the above buffer and subjected to a new centrifugation.
• this pellet is mixed with a detergent in low concentration (for example Triton X 100; 1%; v / v) with stirring. .
• a detergent in low concentration for example Triton X 100; 1%; v / v
• the solution thus obtained is centrifuged at 250 g for 5 minutes at a temperature of + 4 ° C: the centrifugation pellet is resuspended in the above buffer solution and the suspension obtained is again centrifuged . This is repeated until a perfectly white product is obtained, therefore free of hemoglobin which is eliminated in the supernatant at each centrifugation: this product therefore only contains the nuclei
• the solution thus obtained is subjected to a tangential filtration step, thus making it possible to recover the nuclei.
• the membrane of these nuclei is burst by osmotic shock in distilled water and mixed at high speed.
• chromatin solution containing approximately 1 g / l of DNA and approximately 1 g / l of histones, ie a yield per liter of treated blood of approximately 4 g / l of DNA and approximately 4 g /! histones.
• the DNA fragments thus obtained can be classified according to their size into two categories: a highly polymerized part containing more than 2,500 base pairs, and a moderately polymerized part containing approximately 1,000 to 2,500 base pairs.
• This chromatin solution which is a mixed active ingredient of DNA and histones, constitutes a nutrient rich in nucleotides and their components: phospho ⁇ que acid, nitrogen bases, ribose and animated acids.
• This active ingredient can effectively enrich dietetic and cosmetic formulations.
• Two categories of DNA can be extracted from this chromatin solution, according to the process of the present invention: one with high molecular weight or type A, the other with low molecular weight or type B.
• Type A DNA is obtained by liquid chromatography in the presence of enzymes immobilized on a support from the chromatin solution.
• the product thus obtained contains an active ingredient 90% purified and constitutes a source of laboratory products, on the one hand, and of nucleotides, on the other hand.
• Type A DNA is obtained by acid precipitation and conditioned in this form: the DNA fragments are small (oligomers).

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• Health & Medical Sciences (AREA)
• Life Sciences & Earth Sciences (AREA)
• Chemical & Material Sciences (AREA)
• Engineering & Computer Science (AREA)
• General Health & Medical Sciences (AREA)
• Biomedical Technology (AREA)
• Genetics & Genomics (AREA)
• Bioinformatics & Cheminformatics (AREA)
• Public Health (AREA)
• Biotechnology (AREA)
• General Engineering & Computer Science (AREA)
• Veterinary Medicine (AREA)
• Molecular Biology (AREA)
• Animal Behavior & Ethology (AREA)
• Organic Chemistry (AREA)
• Biochemistry (AREA)
• Wood Science & Technology (AREA)
• Zoology (AREA)
• Microbiology (AREA)
• Analytical Chemistry (AREA)
• Crystallography & Structural Chemistry (AREA)
• Dermatology (AREA)
• Plant Pathology (AREA)
• Biophysics (AREA)
• Physics & Mathematics (AREA)
• Birds (AREA)
• Epidemiology (AREA)
• Cosmetics (AREA)

Applications Claiming Priority (3)

Publications (1)

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ID=9466648

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• 1994
  • 1994-09-02 FR FR9410542A patent/FR2724173B1/fr not_active Expired - Fee Related
• 1995
  • 1995-08-31 WO PCT/FR1995/001135 patent/WO1996007664A1/fr not_active Application Discontinuation
  • 1995-08-31 AU AU32615/95A patent/AU3261595A/en not_active Abandoned
  • 1995-08-31 EP EP95929149A patent/EP0778840A1/de not_active Ceased

Non-Patent Citations (1)

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