Classifications

• • 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
  • C12P7/00—Preparation of oxygen-containing organic compounds
  • C12P7/02—Preparation of oxygen-containing organic compounds containing a hydroxy group
  • C12P7/04—Preparation of oxygen-containing organic compounds containing a hydroxy group acyclic
  • C12P7/06—Ethanol, i.e. non-beverage
  • C12P7/14—Multiple stages of fermentation; Multiple types of microorganisms or re-use of microorganisms
• • C—CHEMISTRY; METALLURGY
  • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
  • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
  • C12M21/00—Bioreactors or fermenters specially adapted for specific uses
  • C12M21/12—Bioreactors or fermenters specially adapted for specific uses for producing fuels or solvents
• • C—CHEMISTRY; METALLURGY
  • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
  • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
  • C12M23/00—Constructional details, e.g. recesses, hinges
  • C12M23/58—Reaction vessels connected in series or in parallel
• • C—CHEMISTRY; METALLURGY
  • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
  • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
  • C12M29/00—Means for introduction, extraction or recirculation of materials, e.g. pumps
  • C12M29/24—Recirculation of gas
• • C—CHEMISTRY; METALLURGY
  • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
  • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
  • C12M33/00—Means for introduction, transport, positioning, extraction, harvesting, peeling or sampling of biological material in or from the apparatus
  • C12M33/22—Settling tanks; Sedimentation by gravity

Definitions

• the present invention relates to a process for producing carbon dioxide and ethanol by continuous multistage fermentation.
• Yeasts mainly of the genus Saccharomyces are well known and used for a very long time in brewing, winemaking ...
• productivity and yield are the two main parameters that allow us to judge the efficiency of a fermentation process. Since the speed of production is closely linked to the quantity of active microorganisms present in the fermenters, continuous fermentation processes must increase the active microbial population in order to increase their productivity. To combat the dilution phenomena, which would tend to decrease the microbial concentration, generated by high dilution rates, these methods use techniques for retaining efficient microorganisms.
• European patent application N ° EP. 0213005 offers a continuous fermentation process using flocculating microorganisms, the retention of which in the system is ensured by internal settling-recycling. This process achieves high concentrations of microorganisms. It is also insensitive to contaminants and avoids any mobile mechanical means essential in systems with external settling.
• flocculation is a biological phenomenon by which microorganisms combine to form particles of biomass or cellular aggregates of microscopic size, called flocs, the size of these flocs generally varying from 0.2 to 6 mm, with a average value of 2 mm.
• critical concentration In continuous fermentation, in a single mixed tank, it is not possible to reach ethanol concentrations higher than 50-70 grams / liter; from this concentration called critical concentration, the mortality rate of microorganisms exceeds their growth rate; and this all the more that the ethanol concentration is high.
• the value of the critical concentration depends on the strain of microorganism, the raw material used and the fermentation conditions (temperature, oxygenation, etc.).
• the alcoholic concentrations obtained largely exceed those obtained in a single-stage fermenter.
• the first fermenters are used to generate the microorganisms which ensure, before their destruction, the complementary production in the following stages, indeed the production of ethanol continues either until exhaustion of the sugar, or until the death of the last microorganism.
• FIG. 1 The evolution of the concentration of biomass and ethanol as a function of time, for a culture of Zymomonas mobilis is represented in FIG. 1 of the appended drawing, the concentrations of biomass (X) in DO unit at 510 nm are plotted on the ordinate , as well as those of ethanol (C 2 H 5 OH) in grams / liter, and the time in hours is plotted on the abscissa (t / h).
• Curve 1 corresponds to the change in the concentration of total biomass
• curve 2 to that of viable biomass
• curve 3 to the concentration of ethanol.
• the improvement in productivity generated by the direct recycling of the microorganisms from the effluent in the feed stream of a multi-stage system is only of marginal interest if the ethanol concentration of the effluent exceeds the critical concentration, the microorganisms having been, in fact, killed, at least partially, by ethanol.
• the first of these two methods proposes a control strategy allowing rational optimization of the operation of a multi-stage system with confinement of microorganisms; while recognizing the need to transfer biomass from upstream to downstream, as is the case in all traditional multi-stage continuous processes. Indeed, in the absence of transfer, as could be obtained with a system for absolute retention of microorganisms (ultrafiltration membrane for example), the system would evolve spontaneously either towards clogging or blocking of any flow by excessive accumulation of biomass (for concentrations between 120 and 250 grams / liter of dry mass of microorganisms; this concentration being a function of the microbial strain and of the stirring system used), i.e. towards growth arrest caused by an increase excessive concentration of ethanol and / or associated metabolic inhibitor products.
• the solution adopted in the British patent consists in slaving the operation of a microorganism transfer device (a pump for example), either to the ethanol concentration in the first stage of the system, or to the activity measured. by any means (the production speed of C0 for example).
• the control relates only to the first stage of fermentation.
• a process for the production of carbon dioxide and ethanol has been found, by continuous multi-stage fermentation of a culture medium inoculated with microorganisms in the form of flocs, of the bacteria or yeast type with retention and recycling of microorganisms, the operation is independent of that of any measurement system.
• a controlled transfer (regular, continuous or timed) of the biomass is carried out from one fermentation stage to the next stage.
• the average transfer rate reduced to the volume unit of the fermentation stage is equal to the growth rate of the microorganisms when equilibrium is reached.
• the transfer law is defined on the basis of prior knowledge of the relationship between the concentration of ethanol and the growth rate of the microorganism.
• This type of transfer can be carried out either using a pump, or by opening a valve, or by any other suitable device.
• the increase in the transfer rate q results in an increase in the growth rate / ⁇ and a decrease in the alcoholic concentration.
• ⁇ ⁇ MAX (1-P / P C ) in which ⁇ MAX is the maximum growth rate, without limitation and P is the critical ethanol concentration, from which microorganisms die.
• the volume of each fermentation stage, and of the first in particular is calculated in such a way that, for the nominal operating flow rate of the fermentation system and for a determined final ethanol concentration, the transfer of biomass from the upstream stages to the downstream stages is effected by overflow as soon as the biomass concentration exceeds the retention capacities of the various stages.
• Such behavior implies a continuous growth of microorganisms in the first stage; the volume V 1 will therefore be calculated from the following relation
• V 1 volume of the first stage
• X 1max limit concentration of microorganisms in the first stage.
• For a flocculating yeast X 1max is around 90 to 120 g / liter.
• Zymomonas flocculant X 1max is around 70 to 80 g / liter.
• P is defined in the same way as above. Furthermore, it is necessary that the microorganism retention system in each stage allows overflowing from one stage to another. This constraint therefore excludes the use of membrane type retention systems.
• the gradient is sufficient to cause the formation of bubbles at the center of the largest flocs.
• the fermenters used for the culture of flocculating microorganisms are of the fluidized bed reactors type: the flocs are kept in suspension in a tubular reactor of vertical axis by the upward flow of the fermentation medium.
• this operating principle is satisfactory: during the formation of the CO 2 bubble, there is a local break in the floc. The cell-cell links being irreversibly broken, the floc bursts. There is therefore self-regulation of the particle size.
• Figure 2 of the accompanying drawing illustrates this phenomenon through the volume occupied by the biomass, plotted on the ordinate V in liters, as a function of the sedimentation time in minutes t (min) plotted on the abscissa and the stirring speed ( rpm), curve 1 corresponds to a speed of 200 rpm, curve 2 to a speed of 300 rpm and curve 3 to a speed of 500 rpm.
• the volume occupied by the sedimented flocs after stirring has stopped is a function of the sedimentation time, on the one hand, and of the prior rotation speed of the turbine.
• the sediment compacts slowly, the duration of this phenomenon is much longer than that of the mixture in a fermenter and does not allow a flock broken by agitation to regain its maximum compactness.
• the agitation is optimal when it allows the breaking of the flocs weakened by the appearance of a CO 2 bubble in their center.
• a gas can be air, or recycled fermentation CO 2 or a mixture of the two;
• the stirring conditions are adjusted taking into account, on the one hand, the concentration of biomass to be kept in suspension, on the other hand the rate of fermentation which itself produces a gaseous release contributing to the stirring.
• This installation includes the fermenters 1a, 1b, provided with internal decanters 2a, 2b and overflows 3a, 3b the overflow circuit 3, the feed 4a, 4b in nutritive medium constituted by the feed rings 4.1 at the base of the fermenters , the nutrient medium metering pumps 4.2, the water metering pump 4.3 and the preheater 4.4, and the gas flows 5a, 5b, on the circuit of which there are the rotameters 5.1 (used to measure the production of CO 2 ) the condensers 5.2, the rotary masters 5.3 to measure the gas recirculation (CO 2 recycling), in 5.4 a short circuit, the compressors 5.5, the filters 5.6 and the gas injection crowns 5.7 at the base of the fermenters.
• the installation also includes the foam control devices 6, the pH control devices 7, the temperature control devices 8, a biomass transfer pump 9, a timer 10, and orifices for sampling 11.1 of the nutrient media, 11.2 for the sampling of fermenters and 11.3 for the sampling of the overflow of the 1st stage and 3b for that of the overflow of the 2nd stage.
• the fermentation was carried out in a first fermentation stage la, with a volume V 1 of 1 liter, and in a second fermentation stage lb, with a volume V 2 of 2 liters.
• 1st stage of fermentation Q 1 is 1.82 liters / hour and that of the second stage of fermentation Q2 is 0.32 liters / hour.
• the sugar concentration of the feed medium of the 1st fermentation stage S o 1 is
• S o 2 is 540 g / l
• the ethanol concentration at the outlet of the 1st stage is 60 g / liter and that at the outlet of the second stage is 90 g / liter.
• the transfer rate (q) of the biomass obtained by timed pumping from the first fermenter to the second is 0.02 liters / h; the second stage biomass is evacuated by overflow.
• the temperature is similar by 30 ° C.
• the pH is 4.9, and the pH of the second is 4.8.
• the fermentation media are agitated through the fermentation gases, the recirculation rate being 1.6 vvm in the first stage and 0.8 wm in the second stage.
• the overall ethanol productivity is equal to:
• the residual sugars are at a concentration of 20 grams / liter.
• the sugar use yield is 90.5% and the sugar conversion yield is 0.48 grams of ethanol per gram of sugar consumed.
• the experimental device is the same as above, but the feed rate of the first stage is brought to 3.3 1 / h and that of the second stage to 0.3 1 / h.
• the concentrations of the two sugar feeds are 150 and 650 g / l respectively.
• the concentration of biomass in the first stage reaches 70 g / 1.
• the transfer of biomass is then made by overflow into the second stage.
• the ethanol concentration reaches 92 g / l in the second stage, with a residual sugar concentration of 10 g / l.
• the overall productivity is equal to 110 g / l / h.
• the sugar use yield is 95% and the conversion yield is 0.505 grams of ethanol per gram of sugar consumed (ie 99% of the maximum theoretical yield).
• Example 3
• the two-stage system comprises the fermenters 1a and 1b, the decanters 2a and 2b, the overflows 3a and 3b and the overflow circuit 3 between the fermenters la and lb, (flow rate Q1), the substrate supplies 4a and 4b (flow rates Q1 and Q2), the circuits for circulating the CO 2 recycling gases 5a, 5b.
• the installation further comprises in 6a and 6b the instantaneous means of measuring the CO 2 produced in the first and the second fermenter, and by the circuit 7 where the flow rate at the outlet of the system is Q1 + Q2.
• the fermentation is carried out under the same general conditions as above; the substrate is formed by the P2 sewers from the sugar refinery.
• the feed rate Q1 of the first fermentation stage is 1.45 liters / hour and that of the second stage Q2 is 1.30 liters / hour; the volume of the first stage being 2 liters and that of the second stage 5 liters.
• Biomass transfers are ensured by overflow.
• the biomass concentration is 110 g / l in the first stage and 80 g / l in the second.
• the sugar concentration of the first stage feed medium is 110 g / liter, and that of the second stage is 250 g / liter.
• the amount of residual sugar leaving the 2nd stage is 10 g / liter.
• the ethanol concentration at the outlet of the 1st stage P1 is 51 g / liter and that at the outlet of the 2nd stage is 80 g / liter.

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• 1988
  • 1988-05-04 FR FR8805963A patent/FR2631040B1/fr not_active Expired - Lifetime
• 1989
  • 1989-05-02 JP JP1505543A patent/JPH02504107A/ja active Pending
  • 1989-05-02 EP EP89906371A patent/EP0365671A1/de not_active Withdrawn
  • 1989-05-02 BR BR898906945A patent/BR8906945A/pt not_active Application Discontinuation
  • 1989-05-02 AU AU36882/89A patent/AU3688289A/en not_active Abandoned
  • 1989-05-02 WO PCT/FR1989/000212 patent/WO1989010968A1/fr not_active Application Discontinuation
  • 1989-05-02 ZA ZA893238A patent/ZA893238B/xx unknown

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