WO2020020929A1 - Procédé et dispositif de préparation de suspensions de cellules - Google Patents

Procédé et dispositif de préparation de suspensions de cellules Download PDF

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Publication number
WO2020020929A1
WO2020020929A1 PCT/EP2019/069881 EP2019069881W WO2020020929A1 WO 2020020929 A1 WO2020020929 A1 WO 2020020929A1 EP 2019069881 W EP2019069881 W EP 2019069881W WO 2020020929 A1 WO2020020929 A1 WO 2020020929A1
Authority
WO
WIPO (PCT)
Prior art keywords
pressure
cell suspension
separation
process gas
cell
Prior art date
Application number
PCT/EP2019/069881
Other languages
German (de)
English (en)
Inventor
Katalin SOLYOM
Siegfried Egner
Ana Lucía VÁSQUEZ-CAICEDO
Ulrike Schmid-Staiger
Felix DERWENSKUS
Salima VARONA
Original Assignee
Fraunhofer-Gesellschaft Zur Foerderung Der Angewandten Forschung E.V.
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Fraunhofer-Gesellschaft Zur Foerderung Der Angewandten Forschung E.V. filed Critical Fraunhofer-Gesellschaft Zur Foerderung Der Angewandten Forschung E.V.
Publication of WO2020020929A1 publication Critical patent/WO2020020929A1/fr

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Classifications

    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01D—SEPARATION
    • B01D11/00—Solvent extraction
    • B01D11/02—Solvent extraction of solids
    • B01D11/028—Flow sheets
    • B01D11/0284—Multistage extraction
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01D—SEPARATION
    • B01D11/00—Solvent extraction
    • B01D11/02—Solvent extraction of solids
    • B01D11/0288—Applications, solvents
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01D—SEPARATION
    • B01D11/00—Solvent extraction
    • B01D11/04—Solvent extraction of solutions which are liquid
    • B01D11/0403—Solvent extraction of solutions which are liquid with a supercritical fluid
    • B01D11/0411—Solvent extraction of solutions which are liquid with a supercritical fluid the supercritical fluid acting as solvent for the solvent and as anti-solvent for the solute, e.g. formation of particles from solutions
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01D—SEPARATION
    • B01D11/00—Solvent extraction
    • B01D11/04—Solvent extraction of solutions which are liquid
    • B01D11/0488—Flow sheets
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01D—SEPARATION
    • B01D11/00—Solvent extraction
    • B01D11/04—Solvent extraction of solutions which are liquid
    • B01D11/0492—Applications, solvents used
    • 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
    • C12M47/00—Means for after-treatment of the produced biomass or of the fermentation or metabolic products, e.g. storage of biomass
    • C12M47/06—Hydrolysis; Cell lysis; Extraction of intracellular or cell wall material
    • 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
    • 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
    • C12N1/06—Lysis of microorganisms
    • C12N1/066—Lysis of microorganisms by physical methods
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01D—SEPARATION
    • B01D11/00—Solvent extraction
    • B01D11/02—Solvent extraction of solids
    • B01D11/0203—Solvent extraction of solids with a supercritical fluid
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01D—SEPARATION
    • B01D11/00—Solvent extraction
    • B01D11/04—Solvent extraction of solutions which are liquid
    • B01D11/0403—Solvent extraction of solutions which are liquid with a supercritical fluid

Definitions

  • the invention relates to a method and a device for processing cell suspensions comprising the following successive steps:
  • Process gas is enriched in the cells at an increased pressure and then the
  • a disadvantage of the previously known methods is that a comparatively large amount of liquid has to be carried along in the further process stages.
  • WO 2010/0153898 A1 discloses a method in which cell disruption and simultaneous extraction are carried out in countercurrent in a container. The object of the invention is therefore to make the method more economical.
  • Step of opening the cells and separating the mixture of process gas and cell suspension and separating at least one product, in particular by means of separation and / or extraction, a drainage is provided.
  • liquid obtained during dewatering can also be used to manufacture the liquid obtained during dewatering
  • Cell suspension can continue to be used.
  • Any known method can be used as drainage. It is particularly preferred if a PGSS method is used.
  • Intermediate dewatering can reduce the material flow by at least 50%, in particular by at least 60% and further in particular at least 70% and further in particular at least 80%, these being volume percentages. This has the advantage that subsequent process stages are dimensioned smaller and can therefore be operated more cost-effectively and the concentration of the products has already improved.
  • the dewatering is provided after the step of opening the cells without a further separation stage being provided.
  • the process is designed such that the pressure increase by means of the high-pressure pump takes place once for the entire process and the pressure drops over the process steps, in particular to atmospheric pressure (low pressure level).
  • atmospheric pressure low pressure level
  • this pressure energy which is applied before and for the step of cell disruption by decompression, is also available for the subsequent dewatering, so that methods which require pressure energy can also be used here.
  • the pressure energy is provided by a high pressure pump.
  • the closed printing system also includes all further process stages including any secondary branches.
  • the invention also encompasses an independent inventive method in which the separation of the mixture of process gas and cell suspension and separation of at least one product is designed in several stages, in particular by means of extraction and / or separation, and the method takes place in a closed pressure system.
  • material or energy flows can be added between the steps “cell disruption” and “dewatering” and / or “dewatering” and “separation” as well as between the separation steps or afterwards.
  • material or energy flows By means of these material or energy flows, the energy being added in the form of thermal energy in particular, the energy and / or pressure level can be adapted or raised and the necessary process parameters for the subsequent process steps can be set.
  • a process gas is added before the cell disruption stage, which is mixed with the suspension.
  • the process gas can in particular be argon, carbon dioxide or nitrogen.
  • a defined process temperature which can be, for example, 5 ° C., is set before cell disruption.
  • the separation process can be a mechanical separation, in particular a filtration. However, at least one "pressurized liquid extraction” is particularly preferred.
  • the separation in particular the extraction, is in several stages in series and / or in parallel.
  • a multi-stage process enables different products to be obtained and different separations to be made.
  • a solvent such as ethanol, ethyl acetate and / or water
  • a temperature of in particular at least 60 ° C., in particular at least 70 ° C. and further in particular at least 80 ° C. is preferred.
  • the solvents of the extraction steps can e.g. evaporated and recovered and returned to the cycle.
  • Subsequent process steps can be provided, such as, in particular, the separation of the liquid and solid phases, the liquid phase being able to be treated in further process and extraction stages to obtain further products.
  • a series of extractions and / or fractionations can be provided for this, which enables the cascaded extraction of different substances (products).
  • adiabatic temperature increase before dewatering. This can be done for example by means of conduction and / or induction and / or electromagnetic radiation.
  • the temperature increase can, for example, to at least 60 ° C, in particular to at least 70 ° C and further in particular to at least 80 ° C.
  • a gas stream for example carbon dioxide
  • the gas phase can be supercritical.
  • a separation of water-saturated gas and solid is then achieved by reducing the pressure below the critical point of the gas supplied.
  • the saturated gas can evaporate from the respective device unit and can be separated from the water at low pressure level.
  • the liquid is then also at a low pressure level, whereby the gas can be circulated again by conditioning to a defined pressure and a defined temperature and is available for renewed dewatering.
  • a second extraction with a polar solvent can take place after extraction from microalgae with a non-polar solvent.
  • the extract obtained by extraction and subsequent separation contains e.g. polar or nonpolar lipids, such as carotenoids, glyco- and / or phospholipids or other protein molecules.
  • a multi-strand separation can also be provided in which the steps do not run in series but in parallel, ie the separation stages are arranged in parallel. In any case, it remains that it is a closed printing system via the method or the device.
  • the invention further relates to a device for carrying out the method, as described above, comprising a feed for cell suspension and process gas, and a high-pressure pump for building up a required pressure level upstream of a digestion unit, a digestion unit for the cell suspension and a downstream dewatering and a downstream separation , in particular extraction and / or mechanical separation for an open cell mass.
  • the method and the device can be used to produce end products, intermediate products and / or by-products which result from the solvents used for the extraction or the separation steps. This enables targeted use of the fractions. Process gases and solvents from the individual stages can be recovered and reused in the respective process steps.
  • the invention is described below with reference to a drawing.
  • the single figure shows a flow chart of a preferred method example.
  • the presentation of a cell suspension is identified by the reference number 1.
  • Reference number 2 denotes a stage for the application of the maximum operating pressure and a first temperature level as well as the mixing of the material flows, the Cell suspension at least one process gas was added.
  • the reference number 17 denotes the mixing in of the process gas, which is recovered at reference number 14.
  • the material flows are kept at the pressure, characterized by the reference symbol 18, until an optimal diffusion of the gases into the cells is reached.
  • 3 denotes a stage for the disintegration of the microbial cells of the cell suspension (biomass) by lowering the pressure. The cells burst, so that the cell components that were previously enclosed in the cell membrane are now accessible.
  • the reference number 6 denotes any additional supplies of auxiliary materials as required.
  • Level 7 indicates a dewatering step that is carried out using supercritical gases.
  • the pressure below the critical point of the supplied gas, for example CO2 there is a separation of water-saturated gas and solid reached.
  • Reference number 8 now denotes a separation stage for the separation of gas saturated with water vapor, the gas being separated via a collecting vessel with a low pressure level for 9a gas or 9b liquid and the gas, namely the CO2, being returned to the circuit.
  • the dewatered solid is passed on to the extraction stage 11 as a dewatered mass flow for extraction at an elevated temperature and pressure level, 10 being a storage container for the solvent required for the extraction.
  • the extraction stage bears the reference symbol 11 and has a lower pressure stage, with the dashed line and the reference symbol 15 providing a plurality of extraction and fractionation stages in the bypass to the module 7.
  • the constituents dissolved during the extraction are separated from the solid stream, it being possible for any number of separation stages to be provided for the fractionation of the extract.
  • the extracted substances are collected together with the process gas (eg nitrogen) in the containers 14 at the ambient pressure level, as well as in the collecting container 13.
  • the process gas is returned to process steps 1 and 17 for re-mixing.
  • Reference number 16 denotes a stage for further processing, for example the drying and pelleting of the solid stream at a low pressure level.
  • the drainage in the stage designated by 7 is particularly characteristic, as a result of which a lower mass flow is passed on to the extraction stage and, in particular, that only once, namely in the area identified by reference numeral 2, a pressure energy into the System is introduced and, moreover, regardless of the number of subsequent process stages, including subordinate ones

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  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Biotechnology (AREA)
  • Organic Chemistry (AREA)
  • Zoology (AREA)
  • Wood Science & Technology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Genetics & Genomics (AREA)
  • Microbiology (AREA)
  • Biomedical Technology (AREA)
  • Biochemistry (AREA)
  • General Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Virology (AREA)
  • Tropical Medicine & Parasitology (AREA)
  • Medicinal Chemistry (AREA)
  • Cell Biology (AREA)
  • Molecular Biology (AREA)
  • Sustainable Development (AREA)
  • Mycology (AREA)
  • Extraction Or Liquid Replacement (AREA)

Abstract

L'invention concerne un procédé de préparation de suspensions de cellules, comprenant les étapes suivantes se suivant les unes les autres et consistant à : amener au moins une suspension de cellules ainsi qu'au moins un gaz de processus et mélanger une suspension de cellules et du gaz de processus ; amener le mélange composé d'une suspension de cellules et de gaz de processus dans un état surcritique ou sous-critique en augmentant la pression par une pompe haute pression sur une pression supérieure à 20 MPa et à une température définie ; relâcher brusquement la pression sous 20 MPa pour désagréger les cellules de la suspension de cellules. L'invention concerne en outre un dispositif pour réaliser le procédé.
PCT/EP2019/069881 2018-07-25 2019-07-24 Procédé et dispositif de préparation de suspensions de cellules WO2020020929A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
DE102018118021.7 2018-07-25
DE102018118021.7A DE102018118021A1 (de) 2018-07-25 2018-07-25 Verfahren sowie Vorrichtung zum Aufbereiten von Zellsuspensionen

Publications (1)

Publication Number Publication Date
WO2020020929A1 true WO2020020929A1 (fr) 2020-01-30

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Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/EP2019/069881 WO2020020929A1 (fr) 2018-07-25 2019-07-24 Procédé et dispositif de préparation de suspensions de cellules

Country Status (2)

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DE (1) DE102018118021A1 (fr)
WO (1) WO2020020929A1 (fr)

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20220240498A1 (en) * 2021-02-01 2022-08-04 GPS Innovations Tunable, simple, higher-yleld, higher-rate, lower-cost recovery method of biopharmaceutical products from cell factories

Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5380826A (en) * 1989-07-20 1995-01-10 Aphios Corporation Supercritical fluid disruption of and extraction from microbial cells
US5951875A (en) * 1996-12-20 1999-09-14 Eastman Chemical Company Adsorptive bubble separation methods and systems for dewatering suspensions of microalgae and extracting components therefrom
WO2010015398A1 (fr) * 2008-08-07 2010-02-11 Uhde High Pressure Technologies Gmbh Rupture cellulaire de matières de départ végétales ou animales, par combinaison d'un procédé par pulvérisation et d'une décompression, pour extraction sélective et séparation de matières réutilisables intracellulaires
WO2010053898A2 (fr) 2008-11-04 2010-05-14 3M Innovative Properties Company Elément de filtre et son joint d'étanchéité
EP2977439A1 (fr) * 2014-07-25 2016-01-27 VITO NV (Vlaamse Instelling voor Technologisch Onderzoek NV) Procédé et appareil pour la désagrégation de cellules de biomasse

Patent Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5380826A (en) * 1989-07-20 1995-01-10 Aphios Corporation Supercritical fluid disruption of and extraction from microbial cells
US5951875A (en) * 1996-12-20 1999-09-14 Eastman Chemical Company Adsorptive bubble separation methods and systems for dewatering suspensions of microalgae and extracting components therefrom
WO2010015398A1 (fr) * 2008-08-07 2010-02-11 Uhde High Pressure Technologies Gmbh Rupture cellulaire de matières de départ végétales ou animales, par combinaison d'un procédé par pulvérisation et d'une décompression, pour extraction sélective et séparation de matières réutilisables intracellulaires
DE102008036723A1 (de) 2008-08-07 2010-02-25 Uhde High Pressure Technologies Gmbh Zellaufschluss pflanzlicher oder tierischer Ausgangsmaterialien mittels Kombination von Sprühverfahren und Dekompression zur selektiven Extraktion und Abscheidung intrazellulärer Wertstoffe
WO2010053898A2 (fr) 2008-11-04 2010-05-14 3M Innovative Properties Company Elément de filtre et son joint d'étanchéité
EP2977439A1 (fr) * 2014-07-25 2016-01-27 VITO NV (Vlaamse Instelling voor Technologisch Onderzoek NV) Procédé et appareil pour la désagrégation de cellules de biomasse

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20220240498A1 (en) * 2021-02-01 2022-08-04 GPS Innovations Tunable, simple, higher-yleld, higher-rate, lower-cost recovery method of biopharmaceutical products from cell factories

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