DE102004022300A1 - Multicomponent solid phase-submerged culture system, useful for preparation and testing of high performance strains for e.g. enzyme production or composting - Google Patents

Abstract

Solid phase/submerged multicomponent culture system (A) and corresponding culture method. Solid phase/submerged multicomponent culture system (A) and corresponding culture method comprises: (i) one or more trickle bed/fluidized bed solid-state fermenters with, additionally in the trickle bed, stirrers that, before switching to fluidized bed mode, homogenize the mycelium formed (or the mycelial coating), either stepwise or, for fungi, continuosly, e.g to inhibit undesirable sporulation of strongly sporulating fungi; and (ii) one or more submerged fermenters in which, either with or after milling (using an Ultra-Turrax) of the mycelium transferred to them (after switching from trickle bed to to fluidized bed mode in the solid-substrate culture), the mycelium is cultured in batch or (semi) continuous mode. To generate stable, high capacity strains, the submerged fermenter comprises a quartz tube and closure plate (with standard fittings) and also includes a UV source and one or more solid-state cultures, particularly culture is on inert carriers.

Description

State of the art

In the own patent application PCT / EP03 / 01663 becomes a new bioreactor type and a new culture method using this new reactor type described in which the principle of the trickle bed bioreactor with that of the fluidized bed reactor is combined. Likewise are based on it Procedures such as a) the production of enzymes, b) the production of Secondary metabolites, c) the treatment of plant material (wood, annual plants), d) the use as a biocatalyst system in chemical synthesis, e) the use in pollutant removal from sewage and air and f) the degradation of xenobiotics associated with the simultaneous possibility Product analysis, product extraction and product pre-cleaning (a-d) claimed.

This novel cultivation principle can v.a. the main disadvantages of the falling film reactor (slight blockage of the reactor bed) by a synergistic Combination with the advantages of the fluidized bed reactor (no lumps and blockages with simultaneous high volumetric material turnover) cancel or reduce.

The main difficulty and the main disadvantage in the operation of trickle-film reactors and the corresponding processes are due to the fact that the microorganisms applied to, for example, support materials metabolize the substrates offered, for example to CO ₂ and H ₂ O, whereby biomass is produced.

The resulting biomass leads to congeal and eventually clog the reactor as the free spaces the packed bed or be added to the solid substrate. This increases the pressure loss above the Fixed bed and the mining capacity or production capacity is significantly reduced by channeling and death zones.

This is particularly in Rieselfilmreaktoren with high space-time yields the case, which accordingly can very quickly block.

It are several patented systems known from the literature, which show solutions for this problem by means of various process variants.

For example, VDI reports 1104 (1994) describe the delay in the growth of the biomass by, for example, N-limitation. The disadvantage is that the blocking is only delayed and not prevented in principle and also the corresponding mining services are greatly reduced. In DE 32 27 678 describes a method in which not only trickle water is supplied over the entire course of the reaction, but so much water that at the same time also excess growth is entrained by the water flows down through the interstices of the carrier body. However, this method is not suitable to successfully clean the packed bed, since the shearing action that results from the flooding of the filling body alone, is too low to sustainably prevent blocking. The same applies to the in DE 39 38 507 and DE 32 04 597 described methods that describe a loosening by means of stirrer installation, but this is also insufficient for longer life.

DE 196 08 805 describes a method which describes a loosening by means of screw stirring and flooding, wherein the stirring time is between 5 and a maximum of 60 minutes and the rotational speed is between 3 and a maximum of 30 rpm.

At the same time, a water flow of at least 0.05 m ³ per m ² and hour up to a maximum of 5 m ³ per m ² and hour is used for flooding the reactor space.

DE 196 09 053 describes a method in which the loosening is to be achieved by brief flooding of the reactor zone and simultaneous introduction of air with increasing flow velocity and shearing of the microorganisms mainly by pressure surges of a maximum of 5 sec. Duration. The excess biomass is discharged by subsequent rinsing.

Next the procedural unfavorable Relationship: Effort of such systems is also the duration of the action and the shear load too low to sustain blockages (blockages) to prevent.

Out Wat. Res. Vol 32 no. 5 pp 1593-1603, 1998 is a trickle-bed air biofilter (TBAB), but not as in the present Invention is operated alternately as a fluidized bed reactor.

Out DE 196 09 053 C1 a method is known in which a certain purification of trickle bed reactors is carried out by flooding the reactor, then flowing through air, applying short bursts or a pulsating air flow and by means of bottom-up flushing water. By this cleaning the cultivation process is interrupted, while in the present inventive process no interruption of the normal operation of the trickle bed operation is accompanied, but only the mode for the cultivation of microorganisms is changed.

In Patent Abstract of Japan, Vol 12, No. 165 8C-496), 1988 a fixed bed reactor for the purification of waste water is described in which the blocking of the upper layer of the fixed bed with suspended biomass is removed by introducing gas near the lower part of the fixed bed. In the present inventive method not only air but also additional water or nutrient solution is supplied. Furthermore, the ventilation device of the method according to the invention and the alternating driving style are missing. EP 0442110A and DE 32 27 678 relate essentially to processes for the microbial deposition of organic pollutants from an exhaust air stream, DE 43 08 832 C a process for the removal of organic impurities from raw waters, All mentioned processes do not work alternately in the trickle bed and fluidized bed process in contrast to the process of the invention. Likewise, other essential features such as aeration device are significantly different or absent in comparison to the method according to the invention.

The same applies to US 4,351,729 A . US 5 510 262 A , WO 01 02307 A, US 4746 615 A and Patent Abstracts of Japan, Vol. 016, no. 029 (C-0904) 1992.

Out DE 195 11 159 C1 For example, there is known a method of treating waste water, wherein there are cages in the treatment tank which are partially filled with spherical carriers floating in the waste water and microorganisms and operated as a trickle bed or fluidized bed reactor depending on the waste water load , whereby a reduction of the ventilation effort is achieved and the growth of the vegetation areas is prevented.

However, said method does not work in any way with solid substrates, as it is mainly provided in the own patent application PCT / EP03 / 01663, but on buoyant carriers. On these floatable carriers, wastewater is purified by feeding, alternating between falling film and fluidized bed. In the method according to the invention an alternating change is claimed for such a case with changing substrates, ie growth phase on the carriers, production phase in a fluidized bed or vice versa DE 195 11 159 C1 no pressure detection and control of the procedure with respect to the caused by the substrate growth pressure increase provided as in the application according to the invention. Likewise, the invention claimed ventilation device is missing.

• a) dieser die Möglichkeit des Rieselbettreaktors und des Wirbelreaktors dergestalt miteinander verbindet, dass eine Mikroorganismenkultur (Bakterien, Hefen oder andere Pilze) auf einem Festbett [unten begrenzt durch ein Sieb, welches die Bestandteile des Festbetts zurückhält, oben begrenzt durch ein Sieb, welches ein mögliches Ausschwemmen des Festbetts verhindert] unter Rieselreaktorbedingungen zum Anwachsen gebracht wird, wobei Wasser und Substrat durch bewegliche Rieseldüsen aufgebracht werden, die Belüftung von unten erfolgt und dass die durch die Festbettschicht ablaufende Lösung, welche die gebildeten Enzyme, Sekundär-Stoffwechselprodukte, umgewandelten Substrate, gereinigten Abwässer, Luftverunreinigungen oder umgewandelten Xenobiotika enthält, entweder bis die gewünschte Produktivität, das gewünschte Produkt bzw. das gewünscht Abbauprodukt oder der gewünschte Abbaugrad erreicht ist, zurückgeführt wird oder die Lösung, die durch das Sieb in den Bodentank abgelaufen ist zur weiteren Auftrennung (nach filtermäßiger Abtrennung von festen Partikeln und Mikroorganismen → Sterifiltration) weiterbehandelt bzw. mittels Ultrafiltration und/oder chromatographisch aufgetrennt wird und
• b) dadurch gekennzeichnet, dass bei z.B. kontinuierlicher Kulturführung bei Überschreitung eines bestimmten Druckverlustes über der Füllkörperschütthöhe oder Festbettfüllung von z.B. > 100 Pa/m Schütthöhe bzw. Festbettfüllung durch gleichzeitiges Einleiten von Wasser/Puffer (bzw. Nährlösung) und Luft die Kulturführung von einer Rieselbettreaktor-Verfahrensführung in eine Wirbelbettreaktor-Verfahrensführung übergeht, wobei ein Überlaufen von bewachsenem Füllkörper oder bewachsenen Festbettbestandteilen verhindert ist und die Kultur im Kreis gefahren wird und/oder durch Luft zugegeben wird, um das Bett im Wirbelbettverfahren zu lockern, solange bis bei der nachfolgenden Rieselfilmreaktor-Fahrweise die Überschreitung des Druckverlustes zurückgeführt ist und/oder die durch diese zusätzliche Fahrweise gewünschten Produktions-, Stoffumwandlungs-, Abbaueffekte erreicht worden sind.

The solution of the problems of the prior art has been achieved in our own application PCT / EP03 / 01663 by providing a trickling bed reactor (in principle), characterized in that

• a) this combines the possibility of the trickle bed reactor and the vortex reactor in such a way that a microorganism culture (bacteria, yeasts or other fungi) on a fixed bed [bounded below by a sieve, which retains the components of the fixed bed, bounded above by a sieve, which possible flocculation of the fixed bed prevented] under Rieselreaktorbedingungen is brought to growth, with water and substrate are applied by moving trickle nozzles, the ventilation is done from below and that running through the fixed bed layer solution, the enzymes formed, secondary metabolites, converted substrates, purified Waste water, air pollutants or converted xenobiotics, either to the desired productivity, the desired product or the desired degradation product or the desired degree of degradation is achieved, is returned or the solution, which drained through the sieve in the bottom tank is further treated for further separation (after filter-like separation of solid particles and microorganisms → Sterifiltration) or by ultrafiltration and / or chromatographically separated and
• b) characterized in that, for example, continuous culture control in excess of a certain pressure drop above the Füllkörperschütthöhe or fixed bed filling of eg> 100 Pa / m bed height or fixed bed filling by simultaneous introduction of water / buffer (or nutrient solution) and air, the culture guide of a trickle bed reactor Process procedure is followed in a fluidized bed reactor procedure, wherein overflow of overgrown packing or overgrown fixed bed components is prevented and the culture is circulated and / or added by air to loosen the bed in the fluidized bed process, as long as the subsequent Rieselfilmreaktor- Driving the excess pressure loss is attributed and / or the desired by this additional driving style production, Stoffumwandlungs-, degradation effects have been achieved.

General description the devices of the invention and procedures

A main problem of our own patent application PCT / EP03 / 01663 is that in cultivations of certain fungi, which show thick, leather-containing growths on the solid substrates, which may be the case especially with certain white rot fungi, a driving style change from falling film to fluidized bed without corresponding smashing the Mycelium mat only by blowing in air and water / medium difficult to perform.

Furthermore can e.g. corresponding enzyme optimizations usually only over volume time yields (im Comparison to submersed methods), i. through multiple harvestings and semicontinuous driving styles are achieved and no effective and real volume yield increases (compared to submitter) be achieved.

This but is an important requirement to be in upgescalten solid state procedures compete with submitter technology in terms of cost and acceptance to be able to.

This means that in terms of upscaliger enzyme activities or generally upscalable metabolic activities, whether anabolic or metabolically, or other metabolic factors (e.g., optimized Spore production, suppression sporulation, etc.) special equipment variants and procedures to generate the for required High-performance organisms provided could become.

This is possible in the submers range, because the transferability from bench scale shaking studs regarding almost all possible Process parameters with the exception of certain shearing loads etc., such as pH, temperature, oxygen transfer rates, Red / Ox potential on the Zig-ton scale possible today is. Likewise, this is a strain optimization with regard to C, N sources, Vitamin, trace element requirement, inducers etc. State of the art. This means that in addition to the genetic engineering procedures and more targeted Mutagenesis is another important pillar for stem optimization disposal stands.

This is in the solid state range in any case even approximately solved. Especially no method is known that a "classic" trunk optimization via SSF Piston cultures for next Scale up levels allowed, as can be adjusted in the piston experiments Parameters on pH regulation by buffer systems and relatively constant Restrict temperature control, but the reproducible attitude of other most important Parameters such as ventilation and humidity not possible are what continues to be a stem optimization with respect to C, N sources, Vitamin, trace element requirements, inducers, etc. prevented.

aim the present inventive method It is therefore to overcome the disadvantages of the prior art, that novel solid phase / submersed multicomponent growth systems and appropriate breeding methods are provided.

there could surprisingly be found that through appropriate combination and design of Falling film / fluidized bed reactors with solid state (solid carrier) Reactors or with stirred reactors (Submersreaktoren) and appropriate driving the disadvantages of Prior art no longer occur.

Thus, novel solid phase / submerged multicomponent growth systems and corresponding culture methods are provided, characterized in that as a component
one or more trickle bed / fluidized bed SSF fermenter and corresponding methods are provided (according to the own patent application PCT / EP03 / 01664), which in addition to the prior art in the trickle bed have a stirring device which also step before switching to the fluidized bed mode by step or continuously, the formed mycelia or mycelium can homogenize and, for example, can prevent the most unwanted sporulation of highly sporulating fungi by appropriate sporadic circulation and that as a further component
one or more Submersfermenter be made available in which either without crushing or smashed (by means of built-Ultra Turrax device) transferred into this submicron mycelia (after switching the Rieselbettfermenters on fluid bed mode) as a batch, semi-continuous or continuous cultures for cultivation which are made of a quartz tube and standard fitting-carrying end plates for the production of stable high-performance strains and are equipped with a corresponding UV irradiation device and that are provided as a further component solid state cultures in single or multiple are particularly preferred for growing on inert Carriers → Carrier cultures.

The Submersreaktorkomponente is inventively designed in such a way that the reactor is of the type: stirrer with guide tube, i. a Umwurfreaktor corresponds, but also the Types: of the airlift reactor or jet nozzle reactor or in general all Reactors that have a sufficient small gap depth to the quartz outer wall such that the UV source mounted around the cladding is capable of generating of stable mutants is sufficient (corresponding examples of suitable Reactor types are also in: W. Storhas; Bioreactors and peripheral facilities; Viehweg, 1994).

Detailed description the invention

The appropriately dressed microorga fungal culture, preferably fungal culture, which is preferred according to the task inductive or preferred in terms of optimal cell mass, is usually using Warring Blenders or similar. smashed or a spore suspension is prepared as preculture.

Prefers This preculture is also in one of the Submersfermenter invention (Inoculating fermenter) produced, wherein the appropriate culture medium either according to the criterion: formation of optimal cell yield or optimized enzyme production is selected by added inducers.

This Inoculating fermenter or the Impfpffermenter is (are) according to the invention as well Prefabricated fermenter provided and designed, wherein under Packaging fermenter is understood that in this vessel and media for the solid substrates / solid state cultures e.g. for their filling to disposal or media from solid state, solid substrates either unsubstituted or e.g. substituted with C / N sources, inducers, etc. continuously or step by step to running solid substrates, or solid state cultures or corresponding start cultures become.

As well the preculture is preferred (either from the seed fermenter originating or not dressed in the Anzuchtsubmersfermenter) on the corresponding preselected (in the case of enzyme production usually inductive) solid substrates growing substrate spent in the trickle bed / fluidized bed reactor, and up to one stable enzyme production or metabolite production (measured after multiple step by step harvestings or at the inflection point of the enzyme formation curve, Metabolite production curve (i.e., onset of decrease) in 1-step fermentations) fermented.

Also This preculture is also preferred for inoculation sodid state fermenter spent with inert carriers and also to stable enzyme production or metabolite production (measured after multiple step by step harvesting or at the inflection point of the enzyme formation curve, metabolite production curve (i.e., onset of decrease) in 1-step fermentations) After growth of the precultures according to the invention in the solid substrate fermenter (trickle bed / fluidized bed reactor), at the latest after blocking the SSF substrate bed is stirred after stirring Mushroom / Substrate bed flooded the bed by water, buffer or substrate and / or air and the culture in the submersed fermenter (mutation fermenter) transferred, wherein according to the invention possible the entire subtrate and the larger mushroom fragments in the SSF Fermener remain in which a new trickle cycle can start or the enzyme (product) is harvested.

Also preferred is also the transfer of the Pre-culture, dressed in the inoculum fermenter, not dressed in the growing submerged fermenter, grown in a solid substrate fermenter or solid state fermenter on inert carriers (Carriern) or as described above after appropriate enzyme production, Metabolite production (e.g., laccase production) into the submersed mutation reactor of the present invention spent.

In this submerged fermenter (submersed mutation fermenter) finds the actual mutation takes place. This submerged reactor component is according to the invention in the Designed so that the reactor type: stirrer with guide tube, i. one Discharge reactor corresponds, but also the types: the airlift reactor or jet nozzle reactor or in general all reactors, which have a sufficiently small gap depth for Quartz outer wall such that the UV source mounted around the cladding is capable of generating of stable mutants is sufficient. It can be the entire wall of the Transparent quartz glass fermenter, e.g. Suprasil) or only a part.

In This mutation fermenter is the mushroom mycelium Ultra Turrax smashed and started the submerged culture, either initially until attracted to an optimal cell density or Myceldichte stirred and then by UV irradiation the generation of stable Mutants begin or directly at the beginning of the fermentation the UV irradiation is started. The mutants generated are by appropriate choice the medium subjected to a selective pressure, so that only the adapted Organisms that carry appropriately directional mutation, grow through. This can be done by an additional continuous driving such as chemostat / turbidostat are supported. Such suitable selective substrates may e.g. Target substrates for that too be forming enzyme (as the sole C source or suplementiert with Minimal medium) or inducers, effectors or inhibitors only if enough high enzyme amount (metabolite amount) of the organisms to be attracted to enable growth.

Both Submersfermenter types (seed fermenter and mutation fermenter) are according to the invention with Ultra Turrax smashing equipment equipped to mushroom spawn or to break the mushroom pellets.

Both SSF fermenter (solid substrate or solid state fermenter) serve as reference fermenter for the performance comparison of mutation fermenter after mutation generated mutants in both SSF systems.

The so submerged high-performance strain cultures are then called solid state culture (= culture on solid supports) and on solid substrates Cultures simultaneously or sequentially) to further ferment the various Process variants in your performance to be able to test. This can be done directly after each mutation cycle or after appropriate Mutant selection (→ continuous culture) in the submerged procedure.

Both SSF fermenter (solid substrate or solid state fermenter) serve as a reference fermenter for the performance comparison of mutation fermenter after mutation generated mutants in both SSF systems.

The is called, that for the comparison of performance without mutation or after mutation and after cultivation of high-performance strains by the mutation according to the invention in the mutation remote a second set of solid substrates resp. solid state fermenters is used.

This represents a major advantage of the systems coupled according to the invention: solid substrate culture (trickle bed / fluid bed culture) and solid state Culture (cultivation on inert carriers) and submerged cultures a direct comparison of the individual mutation cycles becomes possible.

One Another advantage of the coupled systems: solid substrate culture (Trickle bed / fluidized bed culture) and solid state culture (breeding on inert carriers) and submerged culture is that the crops set up to the high-performance strain below Selection pressure remains and time-consuming pre-breeding and reprocessing can stay undone.

One Disrupting the culture after the mutation and mutant selection phase (Submerskultur) is definitely possible and can for certain Issues be preferred. If a substrate change in the SSF breeding necessary This can be through the above restart of the whole Systems or particularly preferably connected in parallel additional SSF reactor vessels take place.

Also preferred is the generation of mutants using state-of-the-art chemical mutagenic agents such as: alkylating agents such as N-ethyl-N-nitrosourea Etc..

With the new principle described: the combination of solid sustrat Trickle bed / fluidized bed reactors, solid state cultures as SSF variants and with Strain optimization in submerged culture makes it possible to optimize high performance strains optimized production of (preferred) enzymes for degradation or for Modification of all possible organic solid and liquid Substrates such as e.g. Wood, annual plants, other plant material, Humus, coal, organic waste all kinds of both plant and animal origin, yield-optimize Hydrolysates of these substances for subsequent fermentations and other improved metabolic activities such as degradation of pollutants or improved formation of spores for starter cultures etc. available and by the produced adapted high-performance strains these Applications also in large Upscale standards.

Detailed description a fermentation process:

Two submersed fermenters (mutation fermenter); two solid substrate fermenters, two solid state fermenters:
A disrupted culture, either from the seedling fermentor or not grown in the seed submerged fermenter, or a spore suspension is added to the appropriately preselected (usually inductive) enzyme production substrate in the trickle bed / fluidized bed reactor, or as inoculum into a sodid state fermentor with inert carriers and fermented until stable enzyme production or metabolite production (measured after multiple step-by-step harvestings or at the inflection point of the enzyme formation curve, metabolite production curve (ie onset of decrease) in 1-step fermentations).

To this phase in the solid according to the invention Substrate Fermenter Rieselfilm / fluidized bed reactor), at the latest after blocking the SSF substrate bed is stirred after stirring Mushroom / substrate bed by water, buffer or substrate and / or air flooded and transferred the culture in the submersed fermenter (mutation fermenter), where according to the invention possible the entire subtrate and the larger mushroom fragments in the SSF Fermener remain in which a new trickle cycle can start or the enzyme (product) is harvested.

In similar The same way, the cultivation in the solid state fermenter on inert carriers (Carriern) after appropriate enzyme production, metabolite production (e.g., laccase production) into another submerse mutant reactor of the invention.

In these submerged fermenters (Submersmutationsfermenter), the actual mutation for the production of high-performance strains takes place. This Submersreaktorkomponente is inventively designed in such a way that the reactor the Type: stirrer with guide tube, ie corresponds to a capping reactor, but also the types: the airlifter or jet nozzle reactor or in general all reactors, which have a sufficiently small gap depth to the quartz outer wall, so that the UV source mounted around the jacket sufficient to produce stable mutants. In this case, the entire wall of the fermenter from transparent quartz glass eg Suprasil) exist or only a part.

In These mutation fermenters is the mushroom mycelium usually means battered Ultra Turrax and launched the submerged culture, being either first attracted to an optimal cell density or Myceldichte stirred and then by means of UV irradiation the generation of stable mutants begins or directly at the beginning the fermentation the UV irradiation is started. The generated Mutants become selection pressure by appropriate choice of medium subjected to only the adapted organisms, the corresponding wear directional mutation, grow through. This can be done by an additional continuous Driving style as chemostat / turbidostat are supported. Such suitable selective substrates can e.g. Target substrates for be the enzyme to be formed (as sole C source or suplemented with minimal medium) or inducers, effectors or inhibitors, only if enough high amount of enzyme produced (metabolite amount) the organisms to be attracted to Enable growth.

Both SSF fermenter (solid substrate or solid state fermenter) serve as a reference fermenter for the performance comparison of mutation fermenter after mutation generated mutants in both SSF systems.

The is called, that for the comparison of performance without mutation or after mutation and after cultivation of high-performance strains by the mutation according to the invention in the mutation remote a second set of solid substrates resp. solid state fermenters is used.

More detailed description the fermenter function

The corresponding description of the falling film / fluidized bed reactor with additional stirring means, the corresponding description of the particular mutant production submersible reactor and the corresponding description of the growing reactor on solid inert carriers I refer to. The other combination options according to the invention can also be derived therefrom.

Of Further, the invention is explained by way of example, should but not limited to that.

Example 1: Increased Laccase yield of fungi in SSF cultures by UV mutation and under phytoalexin selection pressure in submerged culture

Certain Mushrooms among them also white rot fungi like Trametes versicolor or Pleurotus ostratus are in one Rieselfilm / fluidized bed reactor attracted to bagasse.

To 4-5 days Growth (highest Laccaseausscheidungsrate) is the reactor after stirring with Trace element solution / vitamin solution flooded → fluid bed mode and in a Submerskultur (Umwurfreaktor, airlifter or jet nozzle reactor etc.) (at extensive retention of the solid substrate medium).

The largely convicted culture is crushed using Ultra-Turrax and placed on a minimal medium, which in addition to C and N source (= glycerol / asparagine) contains attracted and then with UV according to the previously determined killing kinetics betrahlt.

The formed mutants are under increasing selection pressure under increasing addition of phytoalexins such as terpenoids, semiterpenoids, isoflavonoids, Furanoterpenoide, polyynes or Dihydroxyanthrene etc. in one of Mutation following continuous culture (chemostat / tubidostat) and the corresponding high performance mutants, i. the increased amounts of laccase against the increased amount of phytoalexins have enriched, enriched.

The stable mutants are then returned to the SSF cultures (either solid substrate trickle / fluid bed reactor or solid state carrier culture or in both) and the grown high-performance cultures investigated for increased Laccasausbeute out. There can be more Follow mutation cycles.

A

reactor

1

Rieseldrüsen

2

exhaust

3

air inlet (Print)

3a

air inlet (Print)

3b

air inlet (Print)

4

Reading recognition, Regulation, control

4a

pH control

4b

pO ₂ control

4c

temperature control

4d

humidity control

4e

print

5

Belüftungsfritte

6

storage tank

7

sterile filter

8th

prefilter (Particulate filter)

9

acid container

10

Base container

11

Water / buffer tank

12

nutrient reservoir

13

Screens / Filters

14

drain tank

15

circulating (Fluidized bed / fluidized bed)

16

circulating (Falling film)

17

valves

18

fixed bed

19

pressure port

20

procedure to FIA etc.

21

procedure for ultrafiltration (UF)

22

pump 22a . 22b → pumps)

23

Submersfermenter (e.g., mutation fermenter)

24

Solid state fermenter (on inert carriers / carriers

24a

Carrier

25

Ultra Turrax (smashing plant)

26

UV radiation source

27

Agitator: Rieselfimreaktor

Claims (7)

Novel solid phase / submerged multicomponent planting systems and corresponding cultivation methods, characterized in that one or more trickle bed / fluidized bed SSF fermenters and corresponding processes are provided as a component with additional stirring device present in the trickle bed, which, however, precedes switching to the fluidized bed mode also step by step or continuously (in the case of fungi) to homogenize the mycelia or mycelium covers formed and, for example, prevent the usually unwanted sporulation of highly sporulating fungi by appropriate sporadic circulation and that as a further component one or more submicron fermenters are provided in which either without Smashed or smashed (by means of built-in Ultra Turrax device) the mycelia transferred to this submerged fermenter (after switching the Rieselbettfermenters on fluidized bed mode of solid substrate culture) this as a batch, semikontinuier Rare or continuous cultures are grown and are made to produce stable high-performance strains from a quartz tube and standard fitting-carrying end plates and equipped with a corresponding UV irradiation device and that are provided as a further component solid state cultures in one or more especially preferred for cultivation on inert carriers → carrier cultures. Novel Solid Phase / Submers Multicomponent Growing Systems and corresponding cultivation method according to claim 1, characterized in that that as submersible fermenters in one or more all common types, preferably those of the type: Stirrer with guide tube (capping reactor), Airlift reactor or jet nozzle reactor be used in single or multiple. Novel Solid Phase / Submers Multicomponent Growing Systems and corresponding cultivation method according to claim 1 and 2, characterized characterized in that the Submersfermenter (fermenter tube) partially or entirely made of quartz glass and outside or inside with a UV irradiation device are equipped. Novel Solid Phase / Submers Multicomponent Growing Systems and corresponding cultivation method according to claim 1, characterized in that that the solid substrate fermenters such as drum reactors, Falling film reactors, falling film / fluidized bed reactors, screw reactors or all known in the art types. Novel Solid Phase / Submers Multicomponent Growing Systems and corresponding cultivation method according to claim 1, characterized in that that the solid state fermenters are equipped with inert carriers (carriers) may represent all known in the art types and that the corresponding used carrier types also all after May be prior art known types. Novel Solid Phase / Submers Multicomponent Growing Systems and corresponding cultivation method according to claim 1, characterized in that that these novel solid-phase / submerged multicomponent growth systems V. A. for manufacturing and in situ performance testing of generated High-performance strains serve and upscale appropriate solid state / solid substrate method. Novel solid-phase / submerged multicomponent growth systems and corresponding growth methods according to claims 1 and 6, characterized in that the high-performance strains and up-cooled processes produced by these novel solid phase / submerged multicomponent growth systems, in particular by the optimized production of (preferred) enzymes, Treatment (degradation or modification) of all possible organic solid and liquid substrates such as wood, annual plants, other plant material, humus, coal, organic waste of all kinds, as well as of plant and animal origin, for the production of yield-optimized hydrolysates from these materials for subsequent fermentations for the production of improved metabolic activities such as degradation of Pollutants or improved formation of spores for starter cultures, etc. are to be used.