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Microbial fermentation technology involves the use of microorganisms to mass produce substrates into final products on an industrial scale [1][2][3]. Microorganisms in fermentation, especially in its earliest stages, incorporate applications relating to food by enhancing; properties, aroma, shelf life, texture, and nutritional value [4]. Since the turn of the 21st century, two main fermentation techniques have been employed, Solid State Fermentation, and Submerged Fermentation. The commonly used fermentation bioreactors contain features such as agitators, aerators, and baffles. The models include Continuous Stirred Tanks, Bubble Columns, Packed Beds, Fluidized Bed, Photobioreactors, and airlift fermentors [3].

Because of the entrance into the genomic era, emerging applications in conjunction with technological drivers such as artificial intelligence (AI), have been able to expand the applicational uses of microbial fermentation, and biotechnology [5][6]. This has had direct results on the current market and trends of new research directions into health care, and the environment. Although this expansion is present, and rampant, it does come with its own restraints, and possible future improvements to tackle these inevitable challenges.

History

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Fermentation was historically used for food preservation during harsh weather conditions, or to enhance the sensory experience of food. The process has remained popular today mainly due to its health benefits, and ability to prolong shelf life[7].

In the past, fermentation processes made use of naturally occurring microbes found in the substrate [7]. However with the growth in demand for fermented products, approaches were regulated and safeguarded. Modern processes have standard production procedures, such as starter cultures or specific sanitization protocols[7]. 

Brief Timeline

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Pasteur substantiated fermentation in yeast cells in the mid 19th century, while disproving the longstanding theory of spontaneous generation [8].

Citric acid was produced via aerobic fermentation with novel technologies that could maintain sterile air in large quantities in 1919[9].

Large scale fermentors for yeast, penicillin and antibiotic production had been developed by the end of World War Il[8][9].

The production of amino acids and industrial enzymes was achieved via breakthroughs in genetic engineering from the 1960s to 70s [9].

Genome sequencing advances in the early 2000s allowed for beta analysis of the metabolic networks and physiology of microorganisms[9]

Main Fermentation Techniques:

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The two principle fermentation techniques widely used in modern manufacturing are Solid State Fermentation (SSF) and Submerged Fermentation (SmF).

SSF involves the use of a solid support, often a moist solid substrate, in the absence of spare liquid to produce microorganisms [8][10]. SSF is mainly used in the production of enzymes, secondary metabolites, organic acids and unsaturated fatty acids [8]. SSF has become widely applied due to its high yield, low energy requirement and lower risk of bacterial contamination [11].

SmF uses an enriched liquid broth to produce microorganisms [8]. The broth formulation is optimized to the target microbes, which have widespread applications in industry [10]. SmF is carried out in three modes: Batch mode, Fed-batch mode and continuous mode [8], which each have associated benefits and limitations.

Both techniques require appropriate conditions and proper maintenance to undergo optimal microbial fermentation.

Fermentation Models

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The relative success of modern fermentation correlates to the level of conditions achieved in a fermentor. If certain conditions such as pH and temperature are stabilized to an optimal level, this can contribute to the success of enzymatic function and microbial growth respectively [12].

Fermentation Device Requirements:

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Modern fermentors control microorganism activity by controlling the factors mentioned above [13]. Thus the following requirements should be met for devices.  

  • Containment: prevent viable cells from escaping during fermentation or downstream processing [3]
  • Continuous Monitoring: of pH, temperature and pressure via sensors [3]
  • Sustainability: minimal consumption of power, maintenance labor and construction cost while still providing a stable environment [3]

This will produce a specific fermentation process, and thus a specific fermentation product [13]. Modern fermentors require a varying degree of human labor, depending on the biochemical process and type of model being used [3].

Fermentation Device Features:

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The typical fermentation bioreactor has the following features:

Common bioreactor model shown with baffles and an agitation system.

An agitator, which mixes the media while keeping all cells in suspension [3].This allows for a homogenous medium which results in better distribution of ingredients, microbes, and enzymes [14]. This facilitates even air dispersion, the reduction of shear forces, acceleration of metabolism [15]. Stirrers or impellers can be used, in the form of disc turbines, vaned discs, propellers, etc [3].

An aeration system, which allows for a constant supply of O2 without inflicting damage [3]. A sparger is commonly used for delivery by bubbling air through the medium. The need for an agitator can be eliminated as the aeration system can simultaneously mix the medium when the broth is less viscous. Most fungal cultures require an aeration system [3].

Baffles, for further aeration and to prevent radial swirl formation [16]. They consist of vertically arranged metal plates [16]. Wider baffles have higher agitation efficiency. An opening between the baffles and the wall of the vessel is needed to prevent bacterial growth [3].

Commonly Used Fermentor Models:

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There are several bioreactors widely applied in industry, as follows.

Continuous Stirred Tank fermentors are distinguished by their continuous intake of raw materials [13]. Impellers are used for stirring, while spargers are used for aeration. A negative pressure system allows for aeration without culture damage. Steady state conditions are reached via chemostatic or turbidostatic operations [3]. Common applications include yogurt and ethanol [13].

Features of a bubble column reactor.

Bubble Column fermentors introduce air into the base of an elongated vessel via perforated pipes [3]. This creates a high pressure zone which promotes oxygen solubility. Gasses produced during the reaction can be expelled to the expanded head space. Common applications include citric acid and tetracycline[3].

Packed Bed fermentors pack biocatalysts such as microbial cells or immobilized enzymes into a hollow vessel [17]. The substrate is delivered via the bottom while the product is collected from the top of the vessel. The most common application is lactic acid fermentation [17].

Fluidised Bed fermentors suspend fine solids by a high speed upward flowing gas such that the medium behaves as a fluid [18]. This combines configurations of stirred tank and packed bed fermentors [19]. The suspension of the solids increases their metabolic activity, providing many benefits [19].

Photobioreactor uses an enclosed vessel with sunlight or artificial light exposure for biomass production via photosynthesis [20]. The vessels are typically constructed out of glass or plastic for light penetration. The culture is circulated within the solar receivers by centrifugal or airlift pumps. To provide aeration, CO2 is introduced to the vessel [21].

Photobioreactor in laboratory use.

Airlift fermentors utilize an air nozzle which injects high speed air into the liquid medium to form bubbles. The inserted gas achieves agitation[22]. A density difference is generated in the liquid as the average liquid density decreases on the ventilated side due to gas compression, and vice versa for the non-ventilated side [22]. Along with the air, solid substrates may also be delivered at a shallower depth of above the surface. Common subtypes include inner circulation, outer circulation and tension drum types [22]. The fermentor can be considered to be a subtype of fluidised bed bioreactors [23].

Limitations of Modern Fermentation

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Mechanical

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Besides expenses, modern fermentors may have limitations specifically relating to machinery. Most industrial level fermentors require thorough, and regular cleaning, which must be done in a certain way to prevent corrosion and damage of the fermentor’s surface. A similar protocol must be undertaken for regular painting of fermentors, to avoid corrosion overtime [24].

Manpower

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Technical limitations of the fermentor fall under manpower, as fermentors need to be calibrated in order to be as precise as possible. However, this can be eliminated with the integration of Machine Learning (ML). Oil level, filters, and electrical equipment required need to be monitored carefully to prevent damage, and contamination respectively [24].

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The Current Market

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In 2023, the industry generated 32 billion USD worldwide [25], and is predicted to surge to 45 billion USD by 2030 [25] , with Asia retaining dominance due to its historical applications with food, and North America established as the largest market [26]. This rapid growth can be attributed to various key players within the industry such as CRISPR-Cas9 gene editor, AI , and machine learning [27][28]. These players enable in depth research into various scientific fields such as genomics and proteomics [5][27], which are key in stages of development and research of microbial fermentation, with overall advancements in the aspects of production, analyses, and applications. AI and ML computer modeling can integrate real-time processing, automation to boost efficiency, and decrease production costs, and manpower [29].

Market Opportunities and Emerging Applications

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Increased technological advancements give rise to emerging applications of microbial fermentation away from food industries, to the direction of healthcare and the environment [29][30]. The rise in the prevalence of infectious diseases has resulted in demand for products such as antibiotics and vaccines. Both of these products are mass produced through microbial fermentation, specifically through the production of recombinants [26][29]. Recombinants manufacture materials with more than one genetic origin. Current established processes that utilize the mechanism of recombinants are the mass production of insulin, hormones, and erythropoietins . Through the mechanism of recombinants, mass production of antibiotics and vaccines can occur. The former is produced as a strategy to tackle antibiotic resistance present within circulating antibiotics [26]. The latter saw a spike after the rise of Covid-19 , specifically because of the mRNA vaccine which saw to become the most effective vaccine of the epidemic [31]. Subfields of microbial fermentation in health care are also in demand, these include production of more niche, and population specific medicine [26]

Because of these emerging applications compared to the traditional uses of microbial fermentation, there is projected to be a shift in the market and disciplines and their designated shares. The share that health care within microbial fermentation is projected to share is one third of the market [29]. These statistics are also accelerated through the increase in technological applications, and key players driving production. Global microbial fermentation technology has also been driven by demand for renewable energy in the form of biofuels as a fossil fuel alternative due to environmental concerns. Issues faced globally regarding the environment revolve around gas emissions, global warming, and air pollution, and microorganisms producing biofuels at a large scale, and efficiently, could reduce these emissions [9].

Market Restraints

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The increased market of microbial fermentation will be unable to take place on a global level due to market restraints. To effectively mass produce required products of these emerging applications, various expenses and its subsequent restraints are highlighted. Institutions will require specialized instruments, facilities, sterilization, materials, labor, utilities, and quality maintenance, all of which require a high amount of labor, control, and costs [32]. The production of biofuels has the potential to eliminate fossil fuels, and mitigate such environmental risks. Developing countries with inadequate funding from their government will retain more significant consequences in this technological realm [26].

Future Improvements

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Fermentors have not reached their mature stage relative to other established systems [33]. The use of single-use microbial fermentors offers solutions to challenges created by mass industrial fermentation, which also allows for down scale, and up scale production - increasing efficiency, whilst decreasing excess waste through specific outputs. Furthermore, with the integration of ML and AI, fermentors can undergo precise and real time adjustments, for the most effective process, to produce the most desirable product, in terms of both quantity and quality [5][6][9] . In modern fermentors, the optimal conditions are obtained through statistical tools such as AI and ML, to predict and improve these conditions [34].

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