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  • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
  • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
  • C12P17/00—Preparation of heterocyclic carbon compounds with only O, N, S, Se or Te as ring hetero atoms
  • C12P17/16—Preparation of heterocyclic carbon compounds with only O, N, S, Se or Te as ring hetero atoms containing two or more hetero rings
  • C12P17/165—Heterorings having nitrogen atoms as the only ring heteroatoms
• • C—CHEMISTRY; METALLURGY
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  • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
  • C12N1/12—Unicellular algae; Culture media therefor
• • A—HUMAN NECESSITIES
  • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
  • A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
  • A01G33/00—Cultivation of seaweed or algae
• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
  • A23L5/00—Preparation or treatment of foods or foodstuffs, in general; Food or foodstuffs obtained thereby; Materials therefor
  • A23L5/40—Colouring or decolouring of foods
  • A23L5/42—Addition of dyes or pigments, e.g. in combination with optical brighteners
  • A23L5/46—Addition of dyes or pigments, e.g. in combination with optical brighteners using dyes or pigments of microbial or algal origin
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07K—PEPTIDES
  • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
  • C07K14/405—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from algae
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  • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
  • C12N1/32—Processes using, or culture media containing, lower alkanols, i.e. C1 to C6
• • C—CHEMISTRY; METALLURGY
  • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
  • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
  • C12P21/00—Preparation of peptides or proteins
  • C12P21/02—Preparation of peptides or proteins having a known sequence of two or more amino acids, e.g. glutathione
• • C—CHEMISTRY; METALLURGY
  • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
  • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
  • C12P23/00—Preparation of compounds containing a cyclohexene ring having an unsaturated side chain containing at least ten carbon atoms bound by conjugated double bonds, e.g. carotenes
• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
  • A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
• • 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
  • C12N2500/00—Specific components of cell culture medium
  • C12N2500/30—Organic components
  • C12N2500/34—Sugars
• • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
  • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
  • Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
  • Y02A40/00—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production
  • Y02A40/80—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production in fisheries management

Definitions

• the present invention relates to the cultivation of ARU for the production of biomass for the production of products of interest, such as dried biomass or compounds or mixtures of compounds of interest extracted from the biomass produced, in particular pigments or food coloring.
• the invention relates more particularly to the industrial production of this biomass, which must respond to an economic balance of profitability, with on the one hand an increase in the productivity (quantity of biomass and of compounds of interest in the biomass) and on the one hand other an economically acceptable production cost.
• Phycocyanins are pigments produced by unicellular microorganisms that can be used for food coloring. Today, they are mainly produced by the culture of cyanobacteria, basins and autotrophy, low yield of biomass produced. To increase the production of these phycocyanins, phycocyanin producing species have been sought and capable of being cultured under bioreactor fermentation conditions in order to increase the amount of biomass produced.
• Unicellular red algae (ARU) belonging to the class of cyanidiophyceae such as Galdieria sulphuraria are known to produce phycocyanins (Carfagna & al., 2018;) capable of developing under autotrophic, mixotrophic and heterotrophic conditions.
• the addition of one or more carbon sources in the culture medium makes it possible to significantly increase the growth rate.
• These microalgae are capable of consuming a large number of carbonaceous metabolites, in total more than one groin has been counted (glucose, glycerol, pentose, etc.) (Gross et al., 1995, Oesterhelt et al., 1999). - 2007, Sloth & al, 2006).
• the present invention relates to a new method for the cultivation of unicellular red algae (ARU) for the production of a biomass rich in phycocyanins, comprising the steps of (i) mixing in the mixotrophic or heterotrophic mode of said ARUs on a culture medium comprising a carbon source comprising glucose, and (ii) biomass recovery, characterized in that there is added to the culture medium an amount of glycerol sufficient to increase the production of phycocyanin relative to the culture without glycerol.
• ARU unicellular red algae
• the invention also relates to a process for preparing phycocyanins which comprises producing a biomass according to the invention and a step (iii) for extracting phycocyanins from the previously recovered biomass.
• Figure 1 shows the growth monitoring curves and production of phycocyanins under the culture conditions of Example 1.
• the present invention relates to a new method for the cultivation of unicellular red algae (ARU) for the production of a biomass rich in phycocyanins, comprising the steps of (i) mixing in the mixotrophic or heterotrophic mode of said ARUs on a culture medium comprising a carbon source comprising glucose, and (ii) biomass recovery, characterized in that there is added to the culture medium an amount of glycerol sufficient to increase the production of phycocyanin relative to the culture without glycerol.
• ARU unicellular red algae
• ARUs used to produce biomass by fermentation and fermentation methods, particularly for the production of phycocyanins are well known to those skilled in the art.
• patent applications WO 2017/050917, WO 2017/093345, WO 2017/050918 and FR 1752674 filed on March 30, 2017 are cited in particular.
• the ARUs are in particular chosen from the sub-division of Cyanidiophytina, in particular from the class of Cyanidiophyceae, more particularly from the order of cyanidiales, even more particularly chosen from the families of Cyanidiaceae or Galdieriaceae.
• the ARUs are chosen from the genera Cyanidioschyzon, Cyanidium or Galdieria.
• the ARUs employed in the process according to the invention are chosen from the species Cyanidioschyzon merolae 10D, Cyanidioschyzon merolae DBV201, Cyanidium caldarium, Cyanidium daedalum, Cyanidium maximum, Cyanidium partitum, Cyanidium rumpens, Galdieria daedala, Galdieria maxima, Galdieria partita or still Galdieria sulphuraria.
• These culture media comprise a carbon source comprising glucose. It may be glucose or glucose in a complex form such as lactose, fructose or polysaccharides comprising glucose.
• This source of carbon can come from the sugar industry, beet or cane, starch hydrolysates from starch plants such as corn, wheat, potatoes, or from the dairy industry such as milk permeate. (WO 2017/093345), high in lactose, used alone or in a mixture.
• the carbon source comprising glucose is selected from glucose and lactose.
• the culture medium generally comprises the carbon source comprising glucose in an amount of between 0.05 g / l and 200 g / l, advantageously between 1 g / l and 150 g / l, very advantageously between 10 g / l and 80 g / l. g / L.
• the culture medium may comprise other elements well known to those skilled in the field of fermentation microalgae culture, in particular a source of phosphorus and / or a source of nitrogen, and / or a source of sulfur.
• the sources of phosphorus can be chosen from among the following species: phosphoric acid, phosphorus salts, advantageously sodium hydrogenphosphorus (Na2HPC> 4), or sodium dihydrogenophosphorus (NahhPCU), or potassium dihydrogénophosphore (KH2PO4), or potassium hydrogénophosphore (K 2 HPO 4), or mixture, in any proportion of two or more of these sources.
• the medium may also include macroelements and microelements that promote the culture of ARU.
• the lighting conditions for the mixotrophic culture of the ARUs are also known to a person skilled in the art, in particular described in the patent applications WO 2017/050917, WO 2017/050918, WO 2017/093345 and FR 1752674 filed on 30 March 2017.
• Lighting may be continuous or discontinuous, in particular discontinuous in the form of flashes.
• the illumination is carried out with a blue light (WO 2017/050917), more particularly in the form of a radiation having a narrow spectrum of wavelength between 400 and 550 nm, advantageously a narrow spectrum wavelength between 420 nm and 500 nm, preferably between 430 and 480 nm, very preferably centered on 455 nm.
• a blue light WO 2017/050917
• the invention is characterized in that glycerol is added to the culture medium in an amount sufficient to increase the production of phycocyanins relative to the culture without glycerol.
• This addition can be made in the culture medium at the beginning of culture, or in culture media, once the biomass has reached a specific density, for example greater than 20 g / l of dry matter in the culture medium.
• the step (i) of culture can therefore be divided into two substeps, (ia) of growth to produce biomass with a carbon source comprising essentially glucose in a simple or complex form as defined above and (ib) ) of accumulation with the addition of glycerol to promote the production of phycocyanin.
• Industrial glycerol sources that can be integrated into production processes have no restriction on purity or refining (generally between 80% and 100%). Preferably they are food grade. Glycerol source suppliers are well known, particularly players in the biodiesel industry such as the Avril group with Oléon (Glycerine 4808 / 4808K, Glycerin 4827 / 4827K ...), or Cargill, which also offers a wide range of products. a product range based on glycerine refined between 86.5% and 99.7% possessing Kosher, Halal, RSPO, E442 or GMO-free certifications, among others.
• Glycerol source suppliers are well known, particularly players in the biodiesel industry such as the Avril group with Oléon (Glycerine 4808 / 4808K, Glycerin 4827 / 4827K ...), or Cargill, which also offers a wide range of products. a product range based on glycerine refined between 86.
• the carbon source comprising glucose is the main source of carbon in the culture medium and the amount of glycerol sufficient to increase the production of phycocyanin relative to the culture without glycerol will be readily determined by those skilled in the art by a simple experiment. fermenters for comparing the amount of phycocyanin produced with or without glycerol as shown in the examples.
• the desired increase in phycocyanin production will preferably be at least 0.15 mg / g / h, more preferably at least 0.30 mg / g / h, more preferably at least 1 mg / g / h; or at least 3.6 mg / g, more preferably at least 7.2 mg / g, more preferably at least 24 mg / gX.
• the amount of glycerol added to the culture medium is sufficient to have a glycerol / carbon source weight ratio comprising glucose (hereinafter referred to as Gly / Glu) of at least 1/15.
• Gly / Glu glycerol / carbon source weight ratio comprising glucose
• the Gly / Glu ratio is at least 1/14, more preferably at least 1/10, even more preferably at least
• the interest is to ensure optimum profitability between the cost of raw materials and the production of phycocyanin, it will be advantageous not to exceed a Gly / Glu ratio of 1/1.
• the glycerol is added to the culture medium for a Gly / Glu weight ratio of 1/10 to 1/1, preferably 1/7 to 1/2, more preferably about 1/5 to about 1/3.
• the method according to the invention may further comprise a step of recovering the biomass.
• Said recovery of the biomass can be carried out by any technique allowing the recovery of biomass, including filtration methods, gravimetric or under reduced pressure, decantation, or else precipitation methods followed by gravimetric filtration.
• the invention also relates to the biomass obtainable by any of the variants of the process according to the invention.
• biomass according to the invention is advantageously understood to mean a set of microorganism cells produced by the culture thereof, which cells may or may not have retained their physical integrity. It is thus understood that said biomass may comprise an amount of degraded microorganism cells ranging from 0% to 100%. By “degraded” it is meant that the physical integrity of said microorganism cells may have been altered such as, for example, lysed microorganisms, resulting for example from a homogenization or enzymatic lysis process. Once produced, this biomass can be raw, just separated from its culture medium, dried or not, degraded or not.
• Biomass depending on whether it is dried or not, in whole or in part, may comprise a moisture content of 1% to 90%.
• the biomass has a moisture content of 70% to 90%, preferably 80% to 85%. This is particularly the case when it consists essentially of industrial microorganisms, optimized and cultured, after filtration of the fermentation must to separate the cultured microorganisms from the culture medium, before drying.
• the biomass is dried in whole or in part and has a moisture content of 1% to 10%, preferably 2% to 7%.
• said biomass may have a density in ARUs of between 20 and 200 g / l of dry matter, preferably between 90 and 150 g / l of dry matter.
• said biomass may have a protein content of between 25% and 60%, even up to 70%, preferably between 30% and 55%, more preferably between 40% and 50% by weight of dry matter.
• the determination of nitrogen and the crude protein content is calculated using the bulk digestion method and steam distillation (NF EN ISO 5983-2).
• said biomass may have an intracellular content of phycobiliproteins (phycocyanin and allophycocyanin) of between 1 and 250 mg / g of dry matter, preferably between 20 and 150 mg / g of dry matter.
• phycobiliproteins phycocyanin and allophycocyanin
• said biomass may have an intracellular phycocyanin content of between 0.5 and 100 mg / g of dry matter, preferably between 10 and 40 mg / g of dry matter.
• Biomass may be conditioned for storage or for its use as such, for example as a food supplement or food for human or animal consumption.
• the cake that can be obtained after extraction of the phycocyanin from the biomass of ARUs that can be obtained by the method according to the invention can be used as a dietary supplement rich in proteins and carotenoids, in human food or animal.
• said biomass may have an intracellular content of carotenoids of between 0.1 and 10 mg / g of dry matter, advantageously between 0.250 and 1 mg / g of dry matter.
• ARUs have a significant potential for use in many fields, such as, for example, human or animal nutrition, cosmetics and medicine.
• said biomass of ARUs obtainable according to the invention can be used after harvest either directly, optionally dried or after transformation.
• said biomass may be used in the form of flours entering food compositions or in the form of food supplements.
• the biomass of ARUs that can be obtained according to the invention can be transformed into flour according to any method known to those skilled in the art. It can thus be envisaged, for example, that the ARUs can be separated from the culture medium, lysed and reduced to fine particles (average diameter of 10 microns), and then dried.
• the invention also relates to any use of the biomass of ARUs obtainable according to the invention in any known field of use of ARUs, particularly, food or feed, cosmetics, medicine. In the fields of human or animal nutrition and cosmetics, these are of course non-therapeutic uses for animals or healthy humans.
• the biomass obtained after culture of ARUs according to the method of the invention can make it possible to obtain in particular a flour rich in antioxidants, in particular carotenoids (particularly zeaxanthin and b-carotenes) in contents of between 0.1 and 10 mg / g of dry matter, advantageously between 0.25 and 1 mg / g of dry matter, in particular zeaxanthin in a content of between 0 and 10 mg / g.
• a flour rich in antioxidants in particular carotenoids (particularly zeaxanthin and b-carotenes) in contents of between 0.1 and 10 mg / g of dry matter, advantageously between 0.25 and 1 mg / g of dry matter, in particular zeaxanthin in a content of between 0 and 10 mg / g.
• 05 and 5 mg / g of dry matter advantageously between 0.1 and 1 mg / g of dry matter, and / or of b-carotene at a content of 0.05 and 5 mg / g of dry matter, advantageously between 0 and , 1 and 1 mg / g of dry matter, meeting a need particularly in the food industry, because more palatable, having better taste, providing antioxidants in large quantities and can be used in animal or human food.
• the invention therefore relates to a flour that can be obtained after transformation of the biomass into ARUs that can be obtained by the process according to the invention.
• said product can be used pure or mixed with other conventionally used ingredients, particularly in non-organic uses. therapeutics in food or cosmetics.
• the invention also relates to any product that may comprise at least the algal biomass obtainable according to the invention.
• the invention also relates to any product which may comprise at least flour resulting from the transformation of the algal biomass obtainable according to the invention.
• the phycocyanins produced by said biomass can be extracted for use, for example, in the diet or as a dye.
• the extraction of phycobiliproteins, and particularly phycocyanin, from said biomass may be carried out according to any extraction technique known to those skilled in the art such as that described by Moon et al. (2014) or by Jaouen & al. (1999) or in the applications FR 2 789 399, WO 2017/093345 and FR 1752674 filed March 30, 2017.
• the invention also relates to the use of phycocyanin obtainable according to the method of the invention, in food, animal or human, as a dietary supplement, or as a dye, particularly as food coloring.
• Example 1 Comparative growth of the G. sulphuraria strain on glycerol, glucose and a glucose-glycerol mixture.
• the growth of the strain is carried out by measuring the absorbance at 800 nm over time.
• the culture medium is the one known for this strain (medium Gross), the the only difference between the three media tested is the carbon substrate (s) used.
• Culture medium 30g / L of glucose or 30g / L of glycerol (glycerin 4808 Univar) or 25g / L of glucose + 5g / L of glycerol; 8 g / L of (NH4) 2SO4; 0.25 g / L of KH 2 PO 4; 716 mg / L MgSO4; 44mg / L CaCl2; 3 ml / L of Fe-EDTA stock solution (FeS04 at 6.9 g / L and Na2-EDTA at 9.3 g / L) and 4 ml of metal trace stock solution (3.09 g / L of Na2-EDTA; 08g / L CuS04.5H2O, 2.86 g / L H3B03, 0.04g / L NaV03.4H2O, 1.82g / L MnCl2, 0.04g / L CoCl2.6H2O, 0.22g / L ZnSO4.7H2O, 0.017g / L Na2
• Culture conditions The cultures are carried out on a stirring table (140 rpm) in a thermostatically controlled enclosure at 37 ° C. in the middle as described above in the presence of a light source.
• Crop monitoring The growth monitoring shows that we achieve similar OD regardless of the source (s) of carbon (s) used, dry masses and associated phycocyanin assays after 312h of culture are described In the picture.
• glycerol in the medium activates the production of phycocyanin despite the presence of glucose known as phycocyanin inhibitor.
• the growth of the strain is carried out by measuring the absorbance at 800 nm over time.
• the culture medium is that known for this strain (medium Gross), the only difference between the two media tested is the carbon substrate (s) used (s).
• Culture medium 30g / L of milk permeate or 25g / L of milk permeate + 5g / L of glycerol (glycerin 4808 Univar); 8 g / L of (NH4) 2SO4; 0.25 g / L of KH 2 PO 4; 716 mg / L MgSO4; 44mg / L CaCl2; 3 ml / L of Fe-EDTA stock solution (FeS04 at 6.9g / L and Na2-EDTA at 9.3 g / L) and 4 ml of metal trace stock solution (3.09 g / L Na2-EDTA; 08g / L CuS04.5H2O, 2.86g / L H3B03, 0.04g / L NaV03.4H2O, 1.82g / L MnCl2, 0.04g / L CoCl2.6H2O; 0.22g / L ZnSO4.7H2O; 0, 017g / L Na2SeO3; 0.03 g
• Culture conditions The cultures are carried out on a stirring table (140 rpm) in a thermostatically controlled enclosure at 37 ° C., in the medium as described above, in the presence or absence of a light source.
• Crop Monitoring The dry masses and associated phycocyanin assays after 168 h of culture are described in Table 2 (A) in the presence of light and in Table 2 (B) in the dark.
• Example 3 follow-up of a bioreactor culture method of the G.sulphuraria strain on a glucose-glycerol mixture.
• the growth of the strain is monitored by measuring the dry mass over time and the substrate consumption is identified by HPLC assay. An intracellular phycocyanin assay is also performed over time.
• the culture medium is that known for this strain (medium Gross) with 1/6 substrate of glycerol and 5/6 of glucose for the stock and a diet consisting of 1 ⁇ 4 glycerol and 3 ⁇ 4 glucose for the fed-batch .
• Foot of tank 25g / L glucose + 5g / L of glycerol (glycerin 4808 Univar); 8g / L of
• Culture conditions The cultures are carried out in a 2L bioreactor at a temperature controlled at 37 ° C., pH 3, in the presence of a light source.
• Example 4 follow-up of a bioreactor culture method of the G.sulphuraria strain on a permeate mixture of milk-glycerol.
• the growth of the strain is monitored by measuring the dry mass over time and the substrate consumption is identified by HPLC assay. An intracellular phycocyanin assay is also performed over time.
• the culture medium is the one known for this strain (medium Gross) with for 1/6 substrate of glycerol and 5/6 of lactose (brought by the permeate of milk) for the base of vat and a feed composed of 1 ⁇ 4 glycerol and 3 ⁇ 4 of lactose for fed-batch.
• Foot of tank 25g / L permeate of milk + 5g / L of glycerol (Glycerin 4808 Univar); 8 g / L of (NH4) 2SO4; 716 mg / L MgSO4; 3 ml / L of Fe-EDTA stock solution (FeS04 at 6.9 g / L and Na2-EDTA at 9.3 g / L) and 4 ml of metal trace stock solution (3.09 g / L of Na2-EDTA; 08g / L CuS04.5H2O, 2.86 g / L H3B03, 0.04g / L NaV03.4H2O, 1.82g / L MnCl2, 0.04g / L CoCl2.6H2O, 0.22g / L ZnSO4.7H2O, 0.017g / L Na2SeO3 0.03g / L
• Culture conditions The cultures are carried out in a 2L bioreactor at a temperature controlled at 37 ° C., pH 3, in the presence of a light source.
• Example 5 Variation of PC Production by Gly / GIc Proportion in the culture medium.
• a cross-over range of glucose and glycerol was performed to evaluate the effect of glycerol on the inhibition of inhibition of phycocyanin synthesis related to the presence of glucose in the culture medium.
• the total amount of organic carbon in each Erlenmeyer flask is 30 g / l.
• Glucose is symbolized by the contraction "glc” and glycerol by "gly”.
• the concentration of each element in the medium is shown in Table 3.
• Example 7 Variation of PC Production by Glycerol Proportion brought into a medium containing at the beginning of the culture 30g / l of milk permeate.
• glycerol Various amounts of glycerol are added to a medium composed of milk permeate in order to observe the effect of this on the phycocyanin synthesis and the removal of the lactose (glucose + sucrose) inhibition on the production of this pigment .
• the base medium contains 30g / l of milk permeate.
• the added glycerol is symbolized by the contraction "gly”. 6 glycerol concentrations are tested (0, 1, 2, 3, 4 and 5g / l).
• glucose has a limiting effect on the amount of phycocyanin produced by the Galdieria sulphuraria strain, with a concentration of less than 15 mg of phycocyanin per g of dry matter at the end of growth (15 mg PC / g of MS).
• the strain having grown in the presence of glycerol only accumulates about 40 mg PC / g of MS, for the same culture time (240h).
• the culture grown in glycerol may have accumulated up to 70 mg of PC / g of MS, while the one having grown in glucose will have a quantity of PC / g of MS which will remain more or less constant over time.
• the addition of small amounts of glycerol in the medium makes it possible to eliminate the inhibition of PC synthesis linked to the presence of glucose in the medium, whether it is present in the form of simple sugar (glucose), or complex (lactose, sucrose, or other ).
• This method makes it possible to grow the cells using glucose (or a carbon source containing glucose) as the main carbon source, and to use glycerol only as a PC synthesis inducer and not as a main substrate.
• Glucose or carbon sources containing glucose are usually cheaper than glycerol, and generally allow to obtain growth rates higher than glycerol, thus saving on fermentation costs.

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  • 2019-05-27 WO PCT/EP2019/063578 patent/WO2019228947A1/fr unknown

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