Tech4Biowaste - User contributions [en]

Industrial fermentation

2023-03-30T13:28:36Z

Johan Thevelein: /* NovelYeast bv */

{{Infobox technology|Name=Industrial fermentation|Feedstock=[[Garden and park waste]], [[food waste]]|Product=Biomass, bioproducts (e.g., enzymes, biopolymers, organic acids, alcohols)|Category=[[Conversion]] ([[Conversion#Biochemical_processes_and_technologies|Biochemical processes and technologies]])}}
<onlyinclude>'''Industrial fermentation''' is a biotechnological process which uses microorganisms (genetically modified or not), in particular bacteria, yeasts, fungi or algae, to make useful products. The cells are real "cell factories" for the industrial conversion of a wide range of renewable feedstocks into bulk chemicals, fine chemicals, platform chemicals, pharmaceutical ingredients, bio-fuels, bio-plastics, etc. It is a multidisciplinary technology and includes the integrated application of disciplines such as biochemistry, microbiology, molecular genetics and process technology to develop useful processes and products.</onlyinclude>

== Feedstock ==

=== Composition and origin ===
Depending on the type of microorganisms and its genetic modifications, a various range of feedstocks can be used. The most commonly used feedstocks are listed below:

==== Lignocellulose and cellulose ====
Lignocellulose is present in [[garden and park waste]]. Cellulose is present in [[food waste]] such as fruit and vegetable waste. Via [[hydrolysis]], which is usually performed through enzymatic or thermal treatment, fermentable sugars can be obtained from lignocellulose and cellulose.

==== Starch ====
Starch is present in [[food waste]] such as potatoes, corn, wheat or cassava. Starch can directly be utilized by amylase-producing microorganisms, particularly filamentous fungi. However, to allow its use in a wider range of fermentations, starch is usually converted into glucose or dextrins by enzymatic [[hydrolysis]].

==== Oils and Fat ====
Oils and fats are present in [[food waste]] such as gravy, used cooking oil and grease. They can directly be used as fermentation substrate. As they are not water soluble, extensive mixing is required to allow a good contact between the liquid droplets and the fermentation water phase.

==== Dairy waste ====
Whey, the liquid by-product of cheese manufacturing, is used as a source of fermentable carbohydrate and nitrogen.

==== Sugars ====
Sugar-rich waste streams can be derived from food industry waste, e.g., from the candy industry.

=== Pre-treatment ===
Depending on the type of feedstock and its purity, specific pre-treatment technologies are required to provide fermentable substrates to the microorganisms. Generally, this involves a [[Sizing|size reduction]] step, after which the milled biomass can be processed to separate the desired substrate by e.g., [[centrifugation]], filtration, evaporation or [[Crystallisation and precipitation|crystallization]].

In addition, it should be taken into account that some of the above mentioned feedstocks only provide the carbon source (which compose about 50% of the weight of most microorganisms), in that case also other nutrients such as nitrogen, phosphate and potassium need to be added.

==Process and technologies==

=== Microorganisms ===
Microorganisms used in industrial fermentations include: bacteria, yeast, fungi or algae. In practice, these are well-known, productive and harmless (GRAS - Generally Regarded As Safe) production organisms, equipped with the new genetic information, that are used to produce the desired products in high yield and efficiency. A major advantage is that these often genetically modified microorganisms do their work under controlled conditions in a fermenter or bio-reactor, carefully contained and separated from the outside world (contained environment). They cannot escape from the factory so that ecological problems or concerns regarding the release of genetically modified organisms in the environment are avoided.
[[File:Bioreactor principle.svg|thumb|257x257px|Schematic representation of an industrial fermentation bioreactor]]

=== Equipment ===
A typical industrial fermenter consists of an CSTR equipped with:

* an aeration and agitation system: to provide good mixing and availability of oxygen for the cell culture
* a temperature and pH control system: to assure optimal conditions for growth or production
* a foam control system: to avoid excessive foam formation
* sampling ports
* addition ports
* a cleaning and sterilization system: to avoid contamination with other, undesired microorganism
=== Operating conditions ===
As it involves living organisms, a fermentation process is typically conducted under mild conditions (pH and temperature). As a result, the energy consumption is relatively low as well as the capital and operating costs. However, fermentation technologies are complex and sensitive requiring careful control of quality and safety of the raw materials, process parameters, contamination, etc.

Industrial fermentations may be carried out as batch, fed-batch, or continuous culture systems. Batch and fed-batch operations are quite common, continuous fermentations being relatively rare <ref>{{Cite book|author=Y. Chisti|year=2014|book_title=Encyclopedia of Food Microbiology (Second Edition)|publisher=Science Direct}}</ref>.

=== Scale-up of industrial fermentations ===
Typically, a pure starter culture (or seed), maintained under carefully controlled conditions, is used to inoculate sterile petri dishes or liquid medium in the shake flasks. After sufficient growth, the preculture is used to inoculate the seed fermenter. Because industrial fermentations tend to be large (typically 1–250 m3), the inoculum is built up through several successively larger stages, to 5–10% of the working volume of the production fermenter. However, scale-up of a fermentation process is not straightforward as an increase in fermenter size affects the various process parameters in different ways. Therefore, ample expertise is required to find a compromise between all process parameters.
==Product==
Depending on the type of microorganisms and its genetic modifications, a range of products can be synthesized. The most common products are listed and divided over two categories: (1) biomass, (2) bioproducts. In case of the latter, some products require complex genetic modifications.

=== Biomass ===

* Single Cell Protein
*Single Cell Oil
* Baker's yeast
* Lactic acid bacteria

=== Bioproducts ===

==== Enzymes ====

* Proteases
* Lipases
* Amylases
* Cellulases
* Peroxidases

==== Biopolymers ====

* Poly-hydroxyalkanoates (PHA)
* Polysaccharides: xanthan gum, dextran

==== Organic acids ====

* Acetic acid
*Lactic acid

* Citric acid
*Tartaric acid
*Fumaric acid

==== Alcohols ====

* Ethanol
*Butanol
*Glycerol
*Butanediol

==== Solvents ====

* Acetone

==== Pharmaceuticals ====

* Vitamins: vitamin C, B2, B12 ...
*Antibiotics: aminoglycosides, penicillins, cephalosporins, tetracyclines ...
*Hormones

==== Biocolorants ====

* cartenoids
*astaxanthins

==== Biosurfactants and bioemulsifiers ====

* glycolipids
*rhamnolipids

==== Amino-acids ====

* monosodium glutamate (MSG)
* Lysine
* Tryptophan
* Phenylalanine

=== Post-treatment ===
The first step in the post-treatment of fermentation broth cultures, also known as '''downstream processing (DSP)''', is to remove the cells from the medium. This is typically performed by a solid-liquid separation technology, such, as [[centrifugation]] or [[membrane filtration]]. Each fraction can then undergo further processing, depending on whether the product is the biomass itself or an intra- or extracellular product. While intracellular products require cell disruption to release the products, extracellular products are solubilized in the depleted fermentation medium. Cell disruption techniques can be divided into mechanical methods (f.e. [[homogenisation]], [[Sizing|grinding]], [[Ultrasonication|sonication]], [[microwave treatment]], [[steam explosion]]) and non-mechanical methods (f.e. osmotic or temperature shock, [[Enzymatic processes|enzymatic destruction]]). To further purify and concentrate the products several methods can be used including [[chromatography]], [[solvent extraction]], [[Crystallisation and precipitation|crystallization]], [[distillation]], [[drying]] etc. The choice of purification technology is depending on the characteristics of the desired products.

== Technology providers ==
{| class="wikitable sortable mw-collapsible"
|+'''Technology comparison'''
! class="cd-text-darkgreen" style="vertical-align:{{{va|bottom}}}"| Company name
! class="cd-text-darkgreen" style="vertical-align:{{{va|bottom}}}"| Country
! class="cd-text-darkgreen" style="vertical-align:{{{va|bottom}}}"| Technology subcategory
! class="cd-text-darkgreen" style="vertical-align:{{{va|bottom}}}"| Technology name
! class="cd-text-darkgreen" style="vertical-align:{{{va|bottom}}}"| TRL
! class="cd-text-darkgreen" style="vertical-align:{{{va|bottom}}}"| Capacity [kg/h]
! class="cd-text-darkgreen" style="{{writing-mode|s2}};vertical-align:{{{va|bottom}}}"| Feedstock: Food waste
! class="cd-text-darkgreen" style="{{writing-mode|s2}};vertical-align:{{{va|bottom}}}"| Feedstock: Garden & park waste
|-
! style="height:1.8em;"|
!
!
!
!
!
!
!
|-
| [[Industrial fermentation#Amphi-Star|Amphi-Star]]
| Belgium
| -
| BioSurf Biosurfactant Technology Platform
| 1-7
| -
| class="cd-background-lightgreen cd-text-darkgreen" style="text-align:center" |●
|
|-
| [[Industrial fermentation#Avecom|Avecom]]
| Belgium
| -
| PROMIC
| 4-7
| -
| class="cd-background-lightgreen cd-text-darkgreen" style="text-align:center" |●
|
|-
|[[Industrial fermentation#Blucon Biotech GmbH|Blucon Biotech GmbH]]
|Germany
| -
|Industrial Fermentation
|4-5
| -
| class="cd-background-lightgreen cd-text-darkgreen" style="text-align:center" |●
| class="cd-background-lightgreen cd-text-darkgreen" style="text-align:center" |●
|-
|[[Industrial fermentation#Cetaqua Galicia|Cetaqua Galicia]]
|Spain
| -
|TCP (The Carboxylic Platform)
|7
|0.43-0.63
| class="cd-background-lightgreen cd-text-darkgreen" style="text-align:center" |●
| class="cd-background-lightgreen cd-text-darkgreen" style="text-align:center" |●
|-
|[[Industrial fermentation#Holiferm|Holiferm]]
|United Kingdom
| -
|Fermentation intensification and in-line separation
|1-9
| -
| class="cd-background-lightgreen cd-text-darkgreen" style="text-align:center" |●
|
|-
|[[Industrial fermentation#Nature.27s Principles|Nature's Principles]]
|The Netherlands
| -
|Biorefinery
|5-6
| -
| class="cd-background-lightgreen cd-text-darkgreen" style="text-align:center" |●
| class="cd-background-lightgreen cd-text-darkgreen" style="text-align:center" |●
|-
|[[Industrial fermentation#Nosh.bio|Nosh.bio]]
|Germany
| -
|Sustainable functional ingredients for the food industry
|5
| -
| class="cd-background-lightgreen cd-text-darkgreen" style="text-align:center" |●
| class="cd-background-lightgreen cd-text-darkgreen" style="text-align:center" |●
|-
|[[Industrial fermentation#NovelYeast bv|NovelYeast bv]]
|Belgium
| -
|Yeast fermentation to biofuels and bio-based chemicals. Protein production
|3-5
| -
| class="cd-background-lightgreen cd-text-darkgreen" style="text-align:center" |●
| class="cd-background-lightgreen cd-text-darkgreen" style="text-align:center" |●
|-
|[[Industrial fermentation#PERSEO Biotechnology SL|PERSEO Biotechnology SL]]
|Spain
| -
|PERSEO Bioethanol ®
|7-8
|1000
| class="cd-background-lightgreen cd-text-darkgreen" style="text-align:center" |●
| class="cd-background-lightgreen cd-text-darkgreen" style="text-align:center" |●
|-
|[[Industrial fermentation#Sophie.27s BioNutrients|Sophie's BioNutrients]]
|The Netherlands
| -
|Fermentation of microalgae
|6
| -
| class="cd-background-lightgreen cd-text-darkgreen" style="text-align:center" |●
|
|}

=== Amphi-Star ===
{{Infobox provider-industrial fermentation|Company=AmphiStar|Webpage=https://www.amphistar.be|Country=Belgium|Contact=info@amphistar.be|Technology name=BioSurf Biosurfactant Technology Platform|Technology category=Microbial production of biosurfactants|TRL=1-7|Aeration=Yes|Agitator=Rushton|Biosafety lavel=1|Controlled parameters=Temperature, pH, Oxygen, Stirring speed, feed rates, etc.|Microorganism=Starmerella bombicola, Candida kuoi, Rhodotorula bogoriensis, etc. Open for collaboration on any BSL-1 biosurfactant producing strain|Reactor material=Glass or stainless steel|Feedstock=Vegetable oils and sugars from biomass|Product=Biosurfactants e.g. glycolipids such as sophorolipids}}

AmphiStar has developed a proprietary technology platform for the cost-efficient and ecological production of biosurfactants. We are a founders-led spin-off company established in July 2021 that is the result of 15 years joint development between Ghent University (Inbio.be) and the Bio Base Europe Pilot Plant. We derisk the early development stage for biosurfactant production, guide and support technology transfer to industrial manufacturers and collaborate intensely for further development and improvement of the licensed technology. Our technology platform is initially based on the fermentative production with the yeast ''Starmerella bombicola'', producing many different biosurfactants at a high volumetric productivity. Our biosurfactants are made from sustainable, renewable feedstocks and waste streams. Microbial fermentation is a clean production technology that is safe for people and the environment. Our biosurfactants are environmentally friendly, palm oil-free, sulfate-free, mild, non-toxic and non-irritant.

=== Avecom ===
{{Infobox provider-industrial fermentation|Company=Avecom|Image=avecomlogo.png|Country=Belgium|Contact=sales@avecom.be|Webpage=https://www.avecom.be|Technology name=PROMIC|TRL=4-7|Product=Single Cell Protein, PHB-rich biomass|Feedstock=Residual side streams and co-products from the food industry}}

Avecom has developed its PROMIC biomass fermentation platform for the efficient conversion of industrial and agricultural residual side streams and co-products towards high-value single cell proteins.

=== Blucon Biotech GmbH ===
{{Infobox provider-industrial fermentation|Company=BluCon Biotech GmbH|Country=Germany|Contact=Dr. Stefan Verseck; stefan.verseck@blucon-biotech.com|Webpage=https://blucon-biotech.com|Technology name=Industrial Fermentation|TRL=4-5|Product=Produkt L-lactic acid (polymer grade)|Feedstock=Variety of feedstock materials from agriculture, like grasses (straw, corn stover, bagasse, miscanthus, switchgrass, sorghum), wood type materials (poplar, spruce, cotton stalks) or from industrial processes ( cellulosic waste from paper recycling or pulp&paper production, Dried Distillers Grains with Solubles (DDGS), beet pulp)|Microorganism=thermophilic bacteria|Image=Blucon_Logo.jpg|Capacity=not relevant|Aeration=no (anaerobic process)|Biosafety lavel=Non-GMO|Agitator=not relevant|Controlled parameters=not relevant|Reactor material=not relevant|Other=not relevant}}

BluCon Biotech GmbH was founded in June 2017 and has its research laboratories and offices at the BioCampus, Cologne. The company develop a unique technology by which L-lactic acid can be produced at lowest possible production costs so that the bioplastic poly lactic acid (PLA) will become commercially competitive to fossil fuel-based plastics.

To reduce significantly the production costs low-cost raw materials are used, (e.g., cellulosic waste) and a sustainable process which most efficiently converts these feedstocks to the product, L-lactic acid (2nd generation L-lactic acid). Such technology development has been initiated and carried out by Direvo Industrial Biotech GmbH in Cologne. BluCon Biotech GmbH now drives the commercialization of this technology by modern strain and bioprocess engineering.

The unique technology of BluCon Biotech GmbH enables consolidated bioprocessing (CBP) of lignocellulosic feedstocks to L-lactic acid using thermophilic bacteria in a one step process. Our aim is to exclusively use cheap non-food cellulosic or lignocellulosic feedstock and to create a 2nd generation L-lactic acid process, thus to convert waste to value.

Besides developing BluCon technology to commercial stage, BluCon Biotech GmbH serves as a platform to acquire further technologies in the field of applied biotechnology. With its core team, the company shows outstanding expertise in microbiology of extremophilic bacteria, in strain and process development and in project management. It is very well connected both in Germany and internationally.

=== Cetaqua Galicia ===

{{Infobox provider-industrial fermentation|Company=Cetaqua Galicia|Country=Spain|Contact=anton.taboada@cetaqua.com|Technology name=TCP (The Carboxylic Platform)|Webpage=https://www.cetaqua.com/|TRL=7|Aeration=No (Anaerobic technology)|Capacity=0.43 - 0.63|Agitator=Pitched blade agitator|Controlled parameters=Temperature, pH, stirring speed, feed rates and moisture.|Microorganism=Anaerobic open mixed culture (uncontrolled)|Reactor material=Stainless steel|Biosafety lavel=1|Feedstock=Seawage sludge or urban biowaste|Product=Acetic acid, propionic acid, butyric acid and valeric acid.|Image=LogoCetGal.png}}

Cetaqua Galicia is a public-private research centre founded in 2011 by Viaqua, the University of Santiago de Compostela (USC) and the Superior Council of Scientific Investigations (CESIC). Through our three lines of research, we have positioned ourselves as a benchmark centre, at regional, national and European level, in the application of scientific knowledge to the water cycle, especially the fields of wastewater treatment and production and recovery of high value-added by-products in waste water.

Within the biofactory line, the Volatile Fatty Acids (VFAs) production technology from urban sewage sludge and urban biowaste has been developed. The technologies was validated on a laboratory scale in 2020, and two pilot prototypes were designed and built. The first one installed at Conservas Dardo to recover industrial wastewater from the canning industry, and the second at the Ourense wastewater treatment plant for the recovery of sludge from urban wastewater treatment plants. In addition, this project is currently preparing innovation proposals for the development of the corresponding patents for the VFA line.

=== Holiferm ===
{{Infobox provider-industrial fermentation|Company=Holiferm|Country=United Kingdom|Contact=Joana Pereira (info@holiferm.com)|Technology name=Fermentation intensification and in-line separation|Webpage=https://holiferm.com/|TRL=1-9|Capacity=Lab scale to pilot (600 L)|Biosafety lavel=Up to biosafety level 2|Aeration=Aerobic, semi-anaerobic, anaerobic|Agitator=Rushton|Controlled parameters=Temperature, pH, pressure, stirring rate, flows, dissolved oxygen, etc.|Microorganism=Bacteria and yeast|Feedstock=Vegetable oil and sugar streams|Product=Biosurfactants (other biomolecules in the future)|Reactor material=Glass (lab scale), stainless steel (pilot and commercial)|Other=https://www.linkedin.com/company/holiferm/|Image=Holiferm-Logo.jpg}}

Holiferm develops holistically improved fermentation technology and processes to deliver massive economic improvements, with biosurfactants being the initial focus. Holiferm’s patented integrated gravity separation and fermentation technology is a plug and play system that increases fermentation process productivity by three to four times, reducing production costs by more than 50% and enabling low cost production of biosurfactants: renewable, biodegradable and mild alternatives to the petrochemical derived surfactants used in household cleaning, laundry and personal care products. Holiferm is dedicated to the commercialisation of economic biosurfactant production processes, providing a complete platform technology for production, isolation and purification, enabling significant market disruption and growth.

=== Nature's Principles ===
{{Infobox provider-industrial fermentation|Company=Nature’s Principles B.V.|Country=The Netherlands|Contact=ir. Jerry Chen
Business Developer
info@naturesprinciples.eu
+31 6 3972 3401|Webpage=https://naturesprinciples.eu/|Feedstock=corn, sugar beet and sugar cane based feedstocks, to waste cellulose, lignocellulose, inulin or starch-based residual streams, pectin, any sugar-rich feedstock (juice from grass or tropical plants etc.)|Product=L-lactic acid and other organic acids|Image=Logo_NP_Whitebackground.jpg|Technology name=Biorefinery|TRL=5-6|Aeration=not disclosed|Biosafety lavel=not disclosed|Microorganism=not disclosed|Capacity=>100 m3 fermentor volume|Agitator=Not disclosed|Controlled parameters=Not to be given|Reactor material=Not disclosed|Other=Looking for feedstock providers}}
Nature’s Principles started as an innovative PhD project at Delft University of Technology, in The Netherlands. The lab experiments proved great results using mixed culture fermentation, which used Principles of Nature to make the microorganisms satisfied and productive, as opposed to genetically engineering them in a lab. We are now running the pilot scale production and will be ready soon for commercial volumes. Nature’s Principles is ready to face the challenge of making bio-based chemicals viable and circular at the same time.
=== Nosh.bio ===
{{Infobox provider-industrial fermentation|Company=Nosh.bio|Country=Germany|Contact=info@nosh.bio|Webpage=https://www.nosh.bio/|Technology name=Sustainable functional ingredients for the food industry|TRL=5|Aeration=Yes|Biosafety lavel=1|Microorganism=Filamentous fungi|Agitator=Pneumatically agitated (e.g., airlift)|Feedstock=Several|Product=High quality functional microbial biomass|Capacity=Not disclosed|Controlled parameters=Not disclosed|Reactor material=Not disclosed|Image=Nosh_biofoods_black_(002).png}}

Nosh Biofoods produce technical-functional ingredients and nutritional protein by exploring the natural microbial biodiversity, using proprietary fermentation and mild downstream processing technologies.
=== NovelYeast bv ===
{{Infobox provider-industrial fermentation|Company=NovelYeast bv|Agitator=Shake flasks, static tubes with magnetic stirring|Feedstock=1G and 2G feedstocks|Other=Construction of cell factories with recombinant DNA technology|Reactor material=Glass|Microorganism=Saccharomyces cerevisiae, other yeast species, Trichoderma|Controlled parameters=Standard parameters|Biosafety lavel=BSL-1|Aeration=Aerobic, semi-anaerobic|Webpage=https://www.linkedin.com/in/johan-thevelein-aab60a10/|Capacity=Lab-scale|TRL=3-5|Technology category=Industrial fermentation|Technology name=Yeast fermentation to biofuels and bio-based chemicals. Protein production|Contact=johan.thevelein@novelyeast.com|Country=Belgium|Product=Biofuels and bio-based chemicals, proteins, specialty sugars, specialty chemicals}}
NovelYeast bv was founded in 2019 by Prof. Johan Thevelein (KU Leuven and VIB) to continue his R&D activities after his retirement in 2020 as emeritus. The company focusses on the development and industrial implementation of yeast cell factories for the production of biofuels, bio-based chemicals as well as specialty sugars and ingredients with first- and second-generation feedstocks. It also develops cell factories for the production of specific proteins for food applications and enzymes for saccharification of lignocellulosic biomass. In addition, it uses yeast as a tool for biomedical and agroindustrial applications, including yeast probiotics and anti-cancer drugs selected by screening in yeast. NovelYeast has several R&D service collaborations with companies world-wide. Specific industrial yeast strains are available from NovelYeast for the production of 2G bioethanol with lignocellulosic biomass hydrolysates, the production of 1G 2,3-butanediol with first-generation sugars, the production of 1G isobutanol with first-generation sugars, the production of 2G isobutanol with lignocellulosic biomass hydrolysates, the production of 1G bioethanol with molasses, the production of lactic acid with first-generation substrates, the production of isomaltulose with sucrose as substrate. Other yeast strains are in development.

=== PERSEO Biotechnology SL ===
{{Infobox provider-industrial fermentation|Company=PERSEO Biotechnology SL|Country=Spain|Contact=informacion@perseobiotech.com|Webpage=https://www.perseobiotech.com/|Technology name=PERSEO Bioethanol ®|TRL=7 - 8|Capacity=1000|Aeration=If needed. Currently under anaerobic conditions.|Agitator=Vertical stirrers|Biosafety lavel=High, no dangerous biological material used.|Controlled parameters=Temperature, pH, pressure, stirring rate, flows, dissolved oxygen.|Reactor material=Stainless steel|Feedstock=Biodegradable waste (OFMSW, agro-industrial waste, cellulosic waste, etc.)|Product=Advanced Bioethanol + CO2+ valuable organic byproduct|Image=PERSEO_Biotechnology_logo.jpg|Microorganism=Yeasts and bacteria|Other=not applicable}}
PERSEO Biotechnology SL is a Spanish SME with track experience and know-how in the development of biotechnological processes, which range from the development phase at the laboratory level to the industrial upscaling of the process and its demonstration. Likewise, PERSEO Biotechnology offers complementary services to assess the feasibility and the scalability of the biotechnological processes.

PERSEO Biorefinery has its own laboratories and a versatile semi-industrial plant (L’Alcudia, Valencia, Spain) with a treatment capacity up to 25 tons / day of organic waste whose objective is to develop, test and validate biotechnological processes to generate bioproducts and bioenergy, integrating all R&D services for the global recovery of organic waste.

PERSEO Bioethanol® (<nowiki>http://www.perseobiotech.com</nowiki>) is a patented and innovative technology to convert organic waste, such as biodegradable municipal solid waste, horticultural waste, agro-industrial waste, HORECA channel or paper and cardboard, mainly into '''advanced bioethanol''', to be used as liquid biofuel or as raw material for the chemical industry, and in other '''bioproducts''' with high potential in the chemical industry (bioproducts from the fermentation of sugars, biosurfactants, biofertilizers or in biomethane by anaerobic digestion).

PERSEO Bioethanol® is a patented biotechnological process compatible with current existing waste treatment plants, under the concept of an '''integrated biorefinery'''. It is adaptable to each process and to the needs of each client. The process can be integrated as a previous recovery stage in existing plants, including incineration, anaerobic digestion or composting, increasing the value chain of waste treatment and significantly '''improving the economic and environmental results''' of waste management.

=== Sophie's BioNutrients (sophiesbionutrients) ===
{{Infobox provider-industrial fermentation|Company=Sophie's BioNutrients|Biosafety lavel=1|Reactor material=not relevant|Controlled parameters=not relevant|Agitator=not relevant|Aeration=not relevant|Capacity=not relevant|Microorganism=chlorella vulgaris|Feedstock=spent grains, okara, molasses|Image=SophiesBioNutrients_Logo.jpg|TRL=6|Technology name=Fermentation of microalgae|Product=Chlorella Protein Concentrate|Webpage=https://sophiesbionutrients.com|Contact=Eugene Wang; eugene@sophiesbionutrients.com|Country=Netherlands|Other=https://www.linkedin.com/company/sophie-s-bionutrients/}}

The next generation of future food with single cell protein such as micro-algae is a scalable and sustainable source of protein for human consumption. A Sophie’s Bionutrients, we grow micro-algae inside of bio-reactors. This process can be precisely controlled to produce protein in a matter of days. We use limited amount of water and local food waste to produce micro-algae protein within metropolitan areas in many parts of the globe. In addition we can generate whole-algae ingredients in a variety of food application in plant based meat and functional food.

== Open access pilot and demo facility providers ==
[https://biopilots4u.eu/database?field_technology_area_data_target_id=103&field_technology_area_target_id%5B87%5D=87&field_contact_address_value_country_code=All&field_scale_value=All&combine=&combine_1= Pilots4U Database]

== Patents ==
Currently no patents have been identified.

== References ==

[[Category:Conversion]]
[[Category:Technologies]]

Enzymatic processes

2023-03-30T13:25:19Z

Johan Thevelein: /* NovelYeast bv */

{{Infobox technology|Name=Enzymatic processes|Category=[[Conversion]] ([[Conversion#Biochemical_processes_and_technologies|Biochemical processes and technologies]])|Feedstock=[[Garden and park waste]]|Product=Cellulose, hemicellulose, lignin}}
<onlyinclude>'''Enzymatic processes''' utilise enzymes (/ˈɛnzaɪmz/) which are proteins that act as biological catalysts (biocatalysts).<ref>{{Cite web|year=2021|title=Enzyme|e-pub date=|date accessed=24-09-21|url=https://en.wikipedia.org/wiki/Enzyme}}</ref> In terms of lignocellulosic biomass valorisation, enzymes find two main applications: i) biomass pretreatment, and ii) polysaccharides hydrolysis. Biomass enzymatic pre-treatment falls under the category of "biological pre-treatment". Other pre-treatment methods for lignocellulosic biomass includes, physical (e.g., mechanical), [[Hydrolysis|chemical]] (e.g., acid and alkali), physico-chemical (e.g., [[steam explosion]] and AFEX), and a combination thereof<ref>{{Cite journal|author=E. Hosseini Koupaie, S. Dahadha, A.A. Bazyar Lakeh, A. Azizi, E. Elbeshbishy|year=2018|title=Enzymatic pretreatment of lignocellulosic biomass for enhanced biomethane production - A review|journal=Journal of Environmental Management|volume=233|page=774-784|doi=10.1016/j.jenvman.2018.09.106}}</ref>. polysaccharides hydrolysis is a concept mostly applied in biorefineries as part of the [[hydrolysis]] of plant cell wall constituents like cellulose, hemicellulose, and lignin. </onlyinclude>

== Feedstock ==

=== Origin and composition ===
Lignocellulosic biomass (LCB) can be collected as a waste material from forest residues, agricultural, and industrial activities. LCB is mainly characterized by the presence of two carbohydrate polymers, namely cellulose and hemicellulose, as well as an aromatic polymer called lignin. Other components in LCB, found in smaller amounts, are ash, pectin, and proteins. The percentage content of celluloce, hemicelllulose, and lignin are varied among different lignocellulosic materials. In general, the content of cellulose, hemicellulose, and lignin in LCB is in the range of 30-60%, 20-40%, and 15-25%, respectively.<ref>{{Cite journal|author=Sawatdeenarunat, C., Surendra, K., Takara, D., Oechsner, H., Khanal, S.K.|year=2015|title=Anaerobic digestion of lignocellulosic biomass: challenges and opportunities|journal=Bioresour. Technol.|volume=178|page=178-186|doi=10.1016/j.biortech.2014.09.103}}</ref> The physical appearance and strenght of the biomass depend on the varying concentration of these polymers and therefore greatly influences the type of pre-treatment strategy applied for its deconstruction.
[[File:Composition-of-the-plant-cell-wall.png|thumb|Composition of the plant cell wall]]

==== Structural features LCB: ====

===== Cellulose =====
Cellulose is a polysaccharide polymer of glucose disaccharide units, cellobiose, linked tightly by ß-1,4-glycoside bonds. Cellulose molecules are linked by hdyrogen bonds and have different orientations resulting in different levels of crystallinity. Its crystallinity plays a crucial role in the biodegradation of cellulose and, in general, the higher crystallinity level makes it harder to biodegrade the cellulose.

===== Hemicellulose =====
Hemicellulose is a random and branched heterogeneous polymer of different polysaccharides including pentoses (xylose and arabinose), hexoses (glucose, galactose, and mannose) and sugar acids. The branched nature of the hemicellulose allows it to form strong bonds with cellulose (through hydrogen bonds) and lignin (through covalent bonds).

===== Lignin =====
Lignin is a complex and large compound made out of phenylpropane units linked in a three-dimensional structure. The main monomers of lignin are p-hydroxyphenyl alcohol, coniferyl alcohol, and sinapyl alcohol. Lignin acts as a cementing material that links celluose and hemicellulose to form the rigid three-dimensional structure of the plant cell wall.

=== Pre-treatment ===
[[File:Schematic of lignocellulosic biomass pretreatment.PNG|thumb|Schematic of lignocellulosic biomass pretreatment]]
The following pre-treatments may be considered prior to enzymatic pre-treatment:

* [[Sizing]] (e.g., milling, grinding)
* [[Steam explosion]] (hybrid pre-treatment; e.g., combined with laccase pretreatment)<ref>{{Cite journal|author=Weihua Qiu, Hongzhang Chen|year=2012|title=Enhanced the ezymatic hydrolysis efficiency of wheat straw after combined steam explosion and laccase pretreatment|journal=Bioresource Technology|volume=118|page=8-12|doi=10.1016/j.biortech.2012.05.033}}</ref>

The following pre-treatments may be considered prior to enzymatic hydrolysis<ref>{{Cite journal|author=Rajeev Ravindran, Amit Kumar Jaiswal|year=2016|title=A comprehensive review on pre-treatment strategy for lignocellulosic food industry waste: Challenges and opportunities|journal=Bioresource Technology|volume=199|page=92-102|doi=10.1016/j.biortech.2015.07.106}}</ref>,<ref name=":0">{{Cite journal|author=Bikash Kumar, Nisha Bhardwaj, Komal Agrawal, Venkatesh Chaturvedi, Pradeep Verma|year=2019|title=Current perspective on pretreatment technologies using lignocellulosic biomass: An emerging biorefinery concept|journal=Fuel Processing Technology|volume=199|page=|doi=10.1016/j.fuproc.2019.106244}}</ref>:

* Physical (e.g., milling, grinding, [[ultrasonication]], extrusion)
* Chemical (e.g., acid, alkali, ionic liquid, organosolv)
* Physico-chemical (e.g., [[steam explosion]], hot water, [[Ammonia fibre expansion|AFEX]], wet oxidation)
* Biological (e.g., microbial and enzymatic)

== Process and technologies ==

=== Enzymatic pre-treatment (biological pre-treatment) ===

Biological pre-treatment systems rely on biological agents (e.g., enzymes) to delignify lignocellulose and make the process of enzymatic hydrolysis more convenient. The effect of enzymes on the lignocellulosic biomass depends on the type of enzymes as well as the composition of the biomass being treated. This is due to enzyme specificity in terms of the type of the reactions that they catalyze. Laccase (Lac), manganese peroxide (MnP) and versatile peroxide (VP) are enzymes that are used extensively to treat the lignocellulosic substrate<ref>{{Cite journal|author=Baruah J., Nath B.K., Sharma R., Kumar S., Deka R.C., Kalita E.|year=2018|title=Recent Trends in the Pretreatment of Lignocellulosic Biomass for Value-Added Products|journal=Front. Energy Res.|volume=141|page=|doi=10.3389/fenrg.2018.00141}}</ref>. Biological pre-treatment of LCB is often knows as a simple, inexpensive, selective, and environmentally-friendly technology. This is mainly due to the fact that biological pre-treatment does not require high energy inputs or chemicals addition. Furthermore, enzymatic treatment has found success in the removal of toxic inhibitory compounds (i.e., complete removal of phenolic compounds). The limitations asociated with enzymatic pretreatment is its production cost, stability, shelf life, and reusability<ref name=":0">{{Cite journal|author=Bikash Kumar, Nisha Bhardwaj, Komal Agrawal, Venkatesh Chaturvedi, Pradeep Verma|year=2019|title=Current perspective on pretreatment technologies using lignocellulosic biomass: An emerging biorefinery concept|journal=Fuel Processing Technology|volume=199|page=|doi=10.1016/j.fuproc.2019.106244}}</ref>.

=== Enzymatic hydrolysis ===
Enzymatic hydrolysis processes allow to produce monomeric sugars from (ligno)cellulosic biomass by using specific enzymes (e.g., cellulases and hemicellulases) able to break down the chemical bonds in cellulose and hemicellulose polymers. Several factors can affect the efficiency of this process: accessible surface area and crystallinity of the biomass, as well as pH, time and temperatures of the process<ref>{{Cite journal|title=Investigation of Enzymatic Hydrolysis of Coffee Silverskin Aimed at the Production of Butanol and Succinic Acid by Fermentative Processes|year=2019-06-01|author=Saverio Niglio, Alessandra Procentese, Maria Elena Russo, Giovanni Sannia, Antonio Marzocchella|journal=BioEnergy Research|volume=12|issue=2|page=312–324|doi=10.1007/s12155-019-09969-6}}</ref>. Enzymatic hydrolysis is gaining increased attention with respect to acid hydrolysis due to equipment corrosion, energy consumption, non-recyclability of reagents, and fermentation inhibitors production during acid hydrolysis <ref>{{Cite journal|title=Enzymatic hydrolysis of lignocellulosic biomass: converting food waste in valuable products|year=2015-02-01|author=Gabriela Piccolo Maitan-Alfenas, Evan Michael Visser, Valéria Monteze Guimarães|journal=Current Opinion in Food Science|volume=1|page=44–49|doi=10.1016/j.cofs.2014.10.001}}</ref>. Different enzymes play different roles in the hydrolysis of lignocellulosic biomass:

==== Cellulases ====
Cellulases are one of the key enzymes in biomass hydrolysis. Cellulases are a family of enzymes that synergistically act on cellulose to hydrolyze it to its monomers. Cellulases acts on the ß-1,4-glycosidic linkages in cellulose. The complete hydrolysis of cellulose is mediated by combination of three main cellulases, namely endoglucanases [https://enzyme.expasy.org/EC/3.2.1.14 (EC 3.2.14)], exoglucanases [https://enzyme.expasy.org/EC/3.2.1.91 (EC 3.2.1.91)], and glycosidase [https://enzyme.expasy.org/EC/3.2.1.21 (EC 3.2.1.21)]. ''T. reesei''-based cellulases have been the focus of research for the past several years and are widely used in laboratory- as well as pilot-scale studies for bioethanol application. Most commercially accessible enzymes for biomass hydrolysis are actually cocktails of cellulases from ''Trichoderma'' or ''Aspergillus'' supplemented with ß-glucosidases from other sources.

==== Hemicellulases ====
Hemicellulose consists of a mixture of glucose and sugar monomers. Xylan is the most abundant hemicellulose-containing pentose sugars, such as xylose. The enzyme xylanase helps to catalyze the hydrolysis of xylan. Hydrolysis of xylan is mediated by the action of multiple xylanases (i.e., endoxylanases and exoxylanases). Commercially, xylanases are produced from ''T. reesei'', ''A. niger'', ''humicola insolens'', and ''Bacillus'' sp.

==== Pectinases ====
Pectinase is a complex enzyme that degrades the pectin present in lignocellulosic biomass. Pectin is a polymer of α-1,4-linked D-galacturonic acid. This enzyme breaks down polygalacturonic acid (GalA) into a monomeric unit by opening glycosidic linkages. It helps the softening of biomass and therefore aids in the hydrolysis of biomass.

==== Other accessory enzymes ====
The accessory enzymes are also a crucial part in the hydrolysis of LCB and enhance the hydrolysis yield and reduce the enzyme cost and dosages. The breakdown of hemicelluloses is further accompanied by the addition of ß-xylosidases which produces a final product of oligomer with different length as intermediates. Another important accessory enzyme is α-arabinofuranidase for breaking down arabinose into monomers of furanose and pyranose.

== Product ==
Products enzymatic pre-treatment:

* Cellulose
* Hemicellulose
* Lignin

Potential products after fermentation:

* Bioethanol
* Biodiesel
* Biobutanol
* Methane
* Specialty chemicals

=== Post-treatment ===
Currently no post-treatment has been identified.

== Technology providers ==
{| class="wikitable sortable mw-collapsible"
|+'''Technology comparison'''
! class="cd-text-darkgreen" style="vertical-align:{{{va|bottom}}}"| Company name
! class="cd-text-darkgreen" style="vertical-align:{{{va|bottom}}}"| Country
! class="cd-text-darkgreen" style="vertical-align:{{{va|bottom}}}"| Technology subcategory
! class="cd-text-darkgreen" style="vertical-align:{{{va|bottom}}}"| Technology name
! class="cd-text-darkgreen" style="vertical-align:{{{va|bottom}}}"| TRL
! class="cd-text-darkgreen" style="vertical-align:{{{va|bottom}}}"| Capacity [kg/h]
! class="cd-text-darkgreen" style="vertical-align:{{{va|bottom}}}"| Processable volume [L]
! class="cd-text-darkgreen" style="vertical-align:{{{va|bottom}}}"| Temperature [°C]
! class="cd-text-darkgreen" style="{{writing-mode|s2}};vertical-align:{{{va|bottom}}}"| Feedstock: Food waste
! class="cd-text-darkgreen" style="{{writing-mode|s2}};vertical-align:{{{va|bottom}}}"| Feedstock: Garden & park waste
|-
! style="height:1.8em;"|
!
!
!
!
!
!
!
!
!
|-
| [[Enzymatic processes#MetGen Oy|MetGen Oy]]
| Finland
| -
| METNIN™
| 8
| 3
| -
|50
| class="cd-background-lightgreen cd-text-darkgreen" style="text-align:center" |●
| class="cd-background-lightgreen cd-text-darkgreen" style="text-align:center" |●
|-
|[[Enzymatic processes#NovelYeast bv|NovelYeast bv]]
|Belgium
| -
|Enzymatic saccharification, In-situ enzyme production
|3-5
| -
|5
|25-60
| class="cd-background-lightgreen cd-text-darkgreen" style="text-align:center" |●
| class="cd-background-lightgreen cd-text-darkgreen" style="text-align:center" |●
|-
|[[Enzymatic processes#University of Naples .28Naples.2C Italy.29|University of Naples (Naples, Italy)]]
|Italy
| -
|Enzymatic Hydrolysis
|2-4
| -
| -
| -
| class="cd-background-lightgreen cd-text-darkgreen" style="text-align:center" |●
| class="cd-background-lightgreen cd-text-darkgreen" style="text-align:center" |●
|}

=== MetGen Oy ===
{{Infobox provider-enzymatic processes|Company=MetGen Oy|Processable volume=3 tonnes|Feedstock=Lignin (waste stream in pulp and paper industries and biorefineries)|Other=|Temperature=50|Safety restrictions=|Reactor material=Acid durable steel quality S 316|Reactor=Steel Reactor|Controlled paramaters=Temperature, pressure, air flow, pH, DS|Webpage=www.metgen.com|Agitator=Rushton turbine|Capacity=50 kg/week (lignin)|TRL=8|Technology category=Conversion (Biochemical processes and technologies)|Technology name=METNIN™|Contact=alex@metgen.com|Country=Finland|Product=METNIN™ SHIELD - a sustainable BIO-BASED additive for fiber-based packaging boards.|Image=METGEN logo.jpg}}

MetGen has developed and commercially launched a novel lignin valorization technology, METNIN™ to valorize the underutilized lignin streams from modern biorefineries, and pulp and paper mills. METNIN™ is a unique market driven technology combines affordable engineering with advanced biotechnology and turns abundant industrial side stream into sustainable and recyclable alternatives for petrochemicals.

Potentially, the technology produces three main products such as METNIN™SHIELD for additives in packaging applications, METNIN™ lignopolyols in polyurethane application, METNIN™ resins for plywood adhesives. The technology is lignin agnostic and provides the missing link in the value-chain between crude lignin and high value lignin fractions for specific end user products. MetGen’s near-term goal is to accelerate the commercialization of METNIN™ technology and widen the bio-based products’ to market as fast as possible. This will open new opportunities for biorefinery lignin valorization thus paving the way for sustainable and more cost-efficient biorefinery business model. Currently, MetGen is in progress of bringing METNIN™ technology from pilot to demonstration scale with potential to reduce GHG emission by 85%.

METGEN is one of the leading innovators in creating and developing bio-based technologies. Our mission is to enable industries to enhance the value of lignocellulosic biomass through enzymatic solutions. MetGen addresses the demand for creating more sustainable production solutions, helping to meet the ever-growing desire to increase sustainability, reduce environmental pollution, minimize the carbon footprint and conserve biodiversity.

The switch to bio-based feedstocks allows the chemical industry to cut dependency on finite fossil feedstocks and increases the use of more resource-efficient biobased technologies. MetGen is at the core of this development. MetGen produces enzymes on an industrial scale, providing commercial enzymatic solutions to modern biorefineries and the pulp & paper and biogas industries. Our philosophy is to design enzymes for maximum impact in real-life process conditions.

After taking science from the labs to the brink of building factories, it is time to share a few experiences along the way. Innovation comes in many layers and MetGen’s presentation unravels the entangled enzymes, processes, chemistries, bio-products, financing, and business models to a more crisp view of the future of the bio-economy.

MetGen’s business model for METNIN™ technology and other enzymatic technologies is 2-fold: manufacturing and licensing of technologies. The market is too wide for any one company to capture alone and the societal and environmental impact can only be maximized through a parallel business model, licensing. MetGen shall produce the material on its own but also license the METNIN™ technology to be integrated into various types of biorefineries. This project will support and lay the foundation for both business avenues.

As the METNIN™ technology is lignin-agnostic, there are vast opportunities for future commercial replication. METNIN™ is a platform technology and can be operated independently to serve the entire industry. Production of large volumes of products maximizes the value of the technology. Even though the plant can be built next to an existing producer of lignin and take advantage of the synergies in utility sourcing as well as the chemicals recycling, the business model is best served if the operations are not dependent on any one operator of a source of raw material. The low energy demand for operations allows the facility to be located also as a stand-alone establishment. MetGen has received many indications of interest to co-locate the facility by the industry, showing the future expansion potential. The technology is modular, hence leaving flexibility for capacity scale-up to be realized after achieving the expected performance and up-time by first fractionation module.

METNIN™ technology can be used as a stand-alone operational unit but can also be integrated in existing P&P mills and Biorefineries.

LCA study has been carried out using third party services. METNIN™ technology demo plant with 3 selected products shows an emission reduction potential of 85% greenhouse gases (GHG) reduction compared to the fossil based manufacturing of the same products.

=== NovelYeast bv ===
{{Infobox provider-enzymatic processes|Company=NovelYeast bv|Processable volume=5|Feedstock=1G and 2G feedstocks|Other=Application of commercial enzyme cocktails, development of in situ enzyme production|Temperature=25-60|Safety restrictions=Standard microbiological practice|Reactor material=Glass|Reactor=Shake flasks, static tubes with magnetic stirring|Controlled paramaters=Standard parameters|Webpage=https://www.linkedin.com/in/johan-thevelein-aab60a10/|Agitator=Shake flasks, magnetic stirring|Capacity=Lab scale|TRL=3-5|Technology category=Enzymatic saccharification|Technology name=Enzymatic saccharification, In-situ enzyme production|Contact=johan.thevelein@novelyeast.com|Country=Belgium|Product=Fermentable sugars}}
NovelYeast bv was founded in 2019 by Prof. Johan Thevelein (KU Leuven and VIB) to continue his R&D activities after his retirement in 2020 as emeritus. The company focusses on the development and industrial implementation of yeast cell factories for the production of biofuels, bio-based chemicals as well as specialty sugars and ingredients with first- and second-generation feedstocks. It also develops cell factories for the production of specific proteins for food applications and enzymes for saccharification of lignocellulosic biomass. In addition, it uses yeast as a tool for biomedical and agroindustrial applications, including yeast probiotics and anti-cancer drugs selected by screening in yeast. NovelYeast has several R&D service collaborations with companies world-wide. Specific industrial yeast strains are available from NovelYeast for the production of 2G bioethanol with lignocellulosic biomass hydrolysates, the production of 1G 2,3-butanediol with first-generation sugars, the production of 1G isobutanol with first-generation sugars, the production of 2G isobutanol with lignocellulosic biomass hydrolysates, the production of 1G bioethanol with molasses, the production of lactic acid with first-generation substrates, the production of isomaltulose with sucrose as substrate. Other yeast strains are in development.

=== University of Naples (Naples, Italy) ===
{{Infobox provider-enzymatic processes|Company=University of Naples "Federico II"; Department of Chemical Sciences|Country=Italy|Contact=Professor Vincenza Faraco, PhD
Department of Chemical Sciences
Via Cintia, 4 IT-80126 Napoli|Webpage=https://www.docenti.unina.it/vincenza.faraco|Technology name=Enzymatic Hydrolysis|TRL=2-4|Feedstock=1G and 2G feedstocks|Product=Fermentable sugars|Image=Logo_istituzionale_federico_II.jpg|Reactor material=not relevant|Temperature=not relevant|Safety restrictions=not relevant|Capacity=not relevant|Reactor=not relevant|Processable volume=not relevant|Controlled paramaters=not relevant|Agitator=not relevant|Other=not relevant}}

The Department of Chemical Sciences of University of Napoli Federico II hosts about 100 researchers and 20 units of technicians and administrative personnel. The main activities of the Department are the Research, the Didactics and the so-called third mission activitities. The Research activities cover several areas of Chemistry, including the design and synthesis of new molecules, from low mass to macromolecules, the purification and the analytic characterization of natural and synthetic molecules, the structural characterization of new molecules through X-ray diffraction, nuclear magnetic resonance, optical and spin electron spectroscopy techniques, mass spectroscopy. The design, the synthesis and the characterization of new molecules are aimed, for example, to the production of innovative molecules with catalytic properties in important chemical and polymerization processes, or to the production of new functional materials, for several applications in a wide range of fields. Research activities also regard the study of biomolecules and biopolymers for applications in biotechnology, from the development of biosensors to biomedical applications, the study of organic functional molecules and organic polymers for special applications, such as the microelectronics, or the development of new materials with innovative mechanical properties, and the study of nanostructured materials for applications in several fields going from the biology to the medicine, from microelectronics to nanophotonics.

The Department of Chemical Sciences offer the following study programmes:

* Bachelor Degree in Chemistry
* Master Degree in Chemistry
* Bachelor Degree in Industrial Chemistry
* Master Degree in Science and Technology of the Industrial Chemistry
* Bachelor Degree in Molecular and Industrial Biotechnologies
* Master Degree in Molecular and Industrial Biotechnologies

The Department of Chemical Sciences offers the following PhD Courses:

* PhD Course in Chemical Sciences

== Open access pilot and demo facility providers ==
[https://biopilots4u.eu/database?field_technology_area_data_target_id=107&field_technology_area_target_id%5B72%5D=72&field_contact_address_value_country_code=All&field_scale_value=All&combine=&combine_1= Pilots4U Database]

== Patents ==
Currently no patents have been identified.

== References ==
<references />

[[Category:Conversion]]
[[Category:Technologies]]