Ultrasonic Bioreactors for Fermentation

Ultrasonics is an effective means to stimulate microorganisms by mechanical vibrations and cavitation. In a sonobioreactor / ultrasonic fermenter, the ultrasonic treatment of cells and tissue becomes highly controllable since the environmental factors can be exactly determined. With ultrasonic bioreactors, the fermentation output can be significantly enhanced.

Fermentation

The efficiency of fermentation depends on the process conditions: Nutrients, density of the medium, temperature, oxygen / gas content and pressure are important factors that influence the microbial activity. Microorganisms as well as mammalian cells thrive only under certain conditions. The right conditions combined with ultrasonic stimulation can maximize the yield of fermentation.

Ultrasonic Stimulation of Microorganisms

The UP400ST is an ultrasonic homogenizer used to intensify fermentation by improving mass transfer and stimulation microbial cell growth.Fermentation is a metabolic process that converts sugar to acids, gases, or alcohol. It occurs in yeast and bacteria, and also in oxygen-starved muscle cells, as in the case of lactic acid fermentation. Fermentation is also used more broadly to refer to the bulk growth of microorganisms on a growth medium, often with the goal of producing a specific chemical product.
Fermentation process are carried out on industrial scale using microorganisms such as bacteria and fungi for the fermentation. Fermented products are used in the food and general industry. Chemicals, such as acetic acid, citric acid, and ethanol are produced by fermentation. The fermentation rate is influenced by the concentration of microorganisms, cells, cellular components, and enzymes as well as temperature, and pH. For aerobic fermentation, oxygen is a key factor too. Nearly all commercially produced enzymes, such as lipase, invertase and rennet, are made by fermentation with genetically modified microbes.
 
In general, fermentations can be divided into four process types/stages:

  • Production of biomass (viable cellular material)
  • Production of extracellular metabolites (chemical compounds)
  • Production of intracellular components (enzymes and other proteins)
  • Transformation of substrate (in which the transformed substrate is itself the product)
Ultrasonic processor UIP2000hdT (2kW) with stirred batch reactor

Ultrasonic homogenizer UIP2000hdT (2kW) with batch reactor

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Sonication Before, During and After Fermentation

Sonication, the application of low-frequency ultrasound waves, can be used before, during, and after fermentation in various ways and at different stages of the fermentation process.

Ultrasonic Pre-Fermentation Treatment – Improvement of Biomass Availability

  1. Improved Mass Transfer: Sonication as pre-treatment is used to promote mass transfer and make the substrate more available to the microbes. Ultrasonic mixing promotes the mass transfer of substrates to, and products away from, the microbial cells. Ultrasonic intensification of mass transfer can be applied as pre-treatment as well as during fermentation.
  2. Cell Disruption: Sonication can be employed to disrupt cell walls and membranes, particularly in microbial or yeast cultures. This helps release intracellular components, such as enzymes or metabolites, which can improve fermentation performance or facilitate downstream processes.
  3. Extraction of Intracellular Compounds: Sonication can aid in the extraction of intracellular compounds from biological materials before fermentation. This includes extracting enzymes, proteins, nucleic acids, or other target compounds from cells, tissues, or plant materials for subsequent use in fermentation processes.

For instance, ultrasonic pretreatment of rice hull was used to enhance enzymatic hydrolysis for xylooligosaccharides production by Aspergillus japonicus (var. japonicus CY6-1). By sonication, the production of cellulolytic and xylanolytic enzymes from the rice hull was significantly enhanced. The hemicellulose yield was increased to 1.4-fold under sonication and production time was greatly shortened from 24 h to 1.5 h at 80ºC – with further potential of process optimization. The sonicated biomass is much easier convertible for the fungi so that the stability of enzyme activity is extended and the activity of CMCase, b-glucosidase, and xylanase is increased in comparison to the non-sonicated rice hull. The final fermentative products were xylotetraose, xylohexaose, and higher molecular weight xylooligosaccharides. The xylohexaose yield from the sonicated rice hull was 80% higher.

Ultrasonically-Assisted Fermentation – Stimulation of Microbes

  • Mixing and Homogenization: Sonication can be used as a mixing technique during fermentation. The application of ultrasound waves helps to create microstreaming and promotes homogeneity, ensuring uniform distribution of nutrients, gases, and microorganisms within the fermentation vessel.
  • Enhancement of Mass Transfer: Related to improved mixing and homogenization are the ultrasonically improved mass transfer rates during fermentation. Ultrasonic oscillation and cavitation create localized turbulence and enhance the diffusion of substrates, gases, and nutrients into the fermentation broth. This can improve the overall efficiency and productivity of the fermentation process.
  • Improving Cell Viability and Metabolic Activity: Sonication can be applied to microbial cultures during fermentation to enhance cell viability and metabolic activity. Mild sonication can stimulate certain microorganisms, promoting growth, biomass production, and the synthesis of desired metabolites or fermentation products.

Hielscher's ultrasonicators are used to intensify fermentation processes, e.g. the biosynthesis of proteinsPrecisely controllable and repeatable sonication helps to improve the productivity of various fermentation processes without damaging cells. The sonication intensity can be exactly adapted to the specific cell species and its requirements. By controlled sonication, the cell growth and metabolism is positively influenced and the conversions catalyzed by live cells is significantly improved, e.g. stimulating Bifidobacteria in milk.
For some fungal-driven fermentation processes, sonication is successfully used to modify growth morphology and broth rheology without affecting growth rate and yield of filamentous fungi.

Ultrasonic Post-Fermentation Treatment

  • Cell Harvesting and Separation: Sonication can assist in cell harvesting and separation after fermentation. It can aid in breaking cell aggregates, flocculants, or biofilms, facilitating the release of cells from the fermentation broth. This simplifies subsequent downstream processes, such as cell recovery or product purification.
  • Extraction of Intracellular Products: After fermentation, sonication can be used to extract intracellular products, such as enzymes, proteins, or secondary metabolites, from microbial or cellular biomass. This extraction process helps recover valuable compounds and improves the overall yield of the fermentation process.
  • Cell Disintegration for Analytical Purposes: Sonication can be applied to disrupt cells or microbial samples after fermentation, particularly for analytical purposes. It aids in cell lysis and the release of intracellular contents, facilitating the analysis of cellular components or performing downstream assays.

For the production of intracellular components such as microbial enzymes (e.g. catalase, amylase, protease, pectinase, glucose isomerase, cellulase, hemicellulase, lipase, lactase, streptokinase) and recombinant proteins (e.g. insulin, hepatitis B vaccine, interferon, granulocyte colony-stimulating factor, streptokinase), the cells have to be lysed / disrupted after the fermentation process to release the desired proteins. By sonication, the extraction of the intracellular and extracellular polysaccharide–protein complexes from the viscous mycelial fermentation broth is facilitated. Besides its outstanding extraction yield and efficiency, sonication is well-established and reliable for cell lysis and the extraction of intracellular matter.
Click here to read mor about ultrasonic lysis and extraction!

Ultrasonic Bioreactors for Improved Fermentation Processes

Hielscher Ultrasonics is long-time experienced with ultrasonically stimulated bio-processes such as cell stimulation, fermentation, cell disruption and extraction. We offer various standard ultrasonic reactors of different sizes and geometries for sonication in batch and flow-through mode. Alternatively, we offer customized solutions for the integration into your existing bioreactor. Since our ultrasonic processors are very versatile and require only small space, retrofitting into existing biotechnological plants can be realized without problems.
Read more about ultrasonic reactor types, designs and applications here!
The table below, indicates general device recommendations depending on the batch volume or flow rate to be processed. Click at the device type to get more information on each device.

Batch VolumeFlow RateRecommended Devices
0.5 to 1.5mLn.a.VialTweeter
1 to 500mL10 to 200mL/minUP100H
10 to 2000mL20 to 400mL/minUP200Ht, UP400S
0.1 to 20L0.2 to 4L/minUIP1000hdT, UIP2000hdT
10 to 100L2 to 10L/minUIP4000
n.a.10 to 100L/minUIP16000
n.a.largercluster of UIP16000
Ultrasonic irradiation promotes the extraction of active compounds from saffron

Sonicator UIP1000hdT for extraction

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Literature/References

Hielscher Ultrasonics manufactures high-performance ultrasonic homogenizers for dispersion, emulsification and cell extraction.

High-power ultrasonic homogenizers from lab to pilot and industrial scale.

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