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Industrial fermentation is the intentional use offermentation inmanufacturing processes. In addition to themass production offermented foods anddrinks, industrial fermentation has widespread applications inchemical industry.Commodity chemicals, such asacetic acid,citric acid, andethanol are made by fermentation.[1] Moreover, nearly all commercially producedindustrial enzymes, such aslipase,invertase andrennet, are made by fermentation withgenetically modified microbes. In some cases, production ofbiomass itself is the objective, as is the case forsingle-cell proteins,baker's yeast, andstarter cultures forlactic acid bacteria used incheesemaking.
In general, fermentations can be divided into four types:[2]
These types are not necessarily disjoined from each other, but provide a framework for understanding the differences in approach. The organisms used are typicallymicroorganisms, particularlybacteria,algae, andfungi, such asyeasts andmolds, but industrial fermentation may also involvecell cultures from plants and animals, such asCHO cells andinsect cells. Special considerations are required for the specific organisms used in the fermentation, such as the dissolvedoxygen level, nutrient levels, andtemperature. The rate of fermentation depends on the concentration of microorganisms, cells, cellular components, and enzymes as well as temperature,pH[3] and level of oxygen foraerobic fermentation.[4] Product recovery frequently involves theconcentration of thedilute solution.
In most industrial fermentations, the organisms oreukaryotic cells are submerged in a liquid medium; in others, such as the fermentation ofcocoa beans,coffee cherries, andmiso, fermentation takes place on the moist surface of the medium.[5][6]
There are also industrial considerations related to the fermentation process. For instance, to avoid biological process contamination, the fermentation medium, air, and equipment are sterilized. Foam control can be achieved by either mechanical foam destruction or chemicalanti-foaming agents. Several other factors must be measured and controlled such aspressure,temperature,agitator shaft power, andviscosity. An important element for industrial fermentations is scale up. This is the conversion of alaboratory procedure to anindustrial process. It is well established in the field ofindustrial microbiology that what works well at the laboratory scale may work poorly or not at all when first attempted at large scale. It is generally not possible to take fermentation conditions that have worked in the laboratory and blindly apply them toindustrial scale equipment. Although many parameters have been tested for use as scale up criteria, there is no general formula because of the variation in fermentation processes. The most important methods are the maintenance of constant power consumption per unit of broth and the maintenance of constant volumetric transfer rate.[3]
Fermentation begins once thegrowth medium is inoculated with the organism of interest. Growth of the inoculum does not occur immediately. This is the period of adaptation, called the lag phase.[7] Following the lag phase, the rate of growth of the organism steadily increases, for a certain period—this period is the log or exponential phase.[7]
After a phase of exponential growth, the rate of growth slows down, due to the continuously falling concentrations of nutrients and/or a continuously increasing (accumulating) concentrations of toxic substances. This phase, where the increase of the rate of growth is checked, is the deceleration phase. After the deceleration phase, growth ceases and the culture enters a stationary phase or a steady state. The biomass remains constant, except when certain accumulated chemicals in the culture chemically break down the cells in a process calledchemolysis. Unless other microorganisms contaminate the culture, the chemical constitution remains unchanged. If all of the nutrients in the medium are consumed, or if the concentration of toxins is too great, the cells may becomesenescent and begin to die off. The total amount of biomass may not decrease, but the number of viable organisms will decrease.[citation needed]
The microbes or eukaryotic cells used for fermentation grow in (or on) specially designedgrowth medium which supplies the nutrients required by the organisms or cells. A variety of media exist, but invariably contain a carbon source, a nitrogen source, water, salts, andmicronutrients. In the production of wine, the medium is grape must. In the production of bio-ethanol, the medium may consist mostly of whatever inexpensive carbon source is available.[citation needed]
Carbon sources are typically sugars or other carbohydrates, although in the case of substrate transformations (such as the production of vinegar) the carbon source may be an alcohol or something else altogether. For large scale fermentations, such as those used for the production of ethanol, inexpensive sources of carbohydrates, such asmolasses,corn steep liquor,[8] sugar cane juice, or sugar beet juice are used to minimize costs. More sensitive fermentations may instead use purifiedglucose,sucrose,glycerol or other sugars, which reduces variation and helps ensure the purity of the final product. Organisms meant to produce enzymes such asbeta galactosidase,invertase or other amylases may be fed starch to select for organisms that express the enzymes in large quantity.[citation needed]
Fixed nitrogen sources are required for most organisms to synthesizeproteins,nucleic acids and other cellular components. Depending on the enzyme capabilities of the organism, nitrogen may be provided as bulk protein, such as soy meal; as pre-digested polypeptides, such aspeptone ortryptone; or as ammonia or nitrate salts. Cost is also an important factor in the choice of a nitrogen source. Phosphorus is needed for production ofphospholipids in cellular membranes and for the production ofnucleic acids. The amount of phosphate which must be added depends upon the composition of the broth and the needs of the organism, as well as the objective of the fermentation. For instance, some cultures will not produce secondary metabolites in the presence of phosphate.[9]
Growth factors and trace nutrients are included in the fermentation broth for organisms incapable of producing all of the vitamins they require.Yeast extract is a common source of micronutrients and vitamins for fermentation media. Inorganic nutrients, includingtrace elements such as iron, zinc, copper, manganese, molybdenum, and cobalt are typically present in unrefined carbon and nitrogen sources, but may have to be added when purified carbon and nitrogen sources are used. Fermentations which produce large amounts of gas (or which require the addition of gas) will tend to form a layer of foam, since fermentation broth typically contains a variety of foam-reinforcing proteins, peptides or starches. To prevent this foam from occurring or accumulating,antifoaming agents may be added. Mineral buffering salts, such as carbonates and phosphates, may be used to stabilize pH near optimum. When metal ions are present in high concentrations, use of achelating agent may be necessary.[citation needed]
Developing an optimal medium for fermentation is a key concept to efficient optimization. One-factor-at-a-time (OFAT) is the preferential choice that researchers use for designing a medium composition. This method involves changing only one factor at a time while keeping the other concentrations constant. This method can be separated into some sub groups. One is Removal Experiments. In this experiment all the components of the medium are removed one at a time and their effects on the medium are observed. Supplementation experiments involve evaluating the effects of nitrogen and carbon supplements on production. The final experiment is a replacement experiment. This involves replacing the nitrogen and carbon sources that show an enhancement effect on the intended production. Overall OFAT is a major advantage over other optimization methods because of its simplicity.[10]
Microbialcells orbiomass is sometimes the intended product of fermentation. Examples includesingle cell protein,bakers yeast,lactobacillus,E. coli, and others. In the case of single-cell protein,algae is grown in large open ponds which allow photosynthesis to occur.[11] If the biomass is to be used for inoculation of other fermentations, care must be taken to preventmutations from occurring.
Metabolites can be divided into two groups: those produced during the growth phase of the organism, calledprimary metabolites and those produced during the stationary phase, calledsecondary metabolites. Some examples of primary metabolites areethanol,citric acid,glutamic acid,lysine,vitamins andpolysaccharides. Some examples of secondary metabolites arepenicillin,cyclosporin A,gibberellin, andlovastatin.[9]
Primary metabolites are compounds made during the ordinary metabolism of the organism during the growth phase. A common example is ethanol or lactic acid, produced duringglycolysis. Citric acid is produced by some strains ofAspergillus niger as part of thecitric acid cycle to acidify their environment and prevent competitors from taking over. Glutamate is produced by someMicrococcus species,[12] and someCorynebacterium species produce lysine, threonine, tryptophan and other amino acids. All of these compounds are produced during the normal "business" of the cell and released into the environment. There is therefore no need to rupture the cells for product recovery.
Secondary metabolites are compounds made in the stationary phase; penicillin, for instance, prevents the growth of bacteria which could compete withPenicillium molds for resources. Some bacteria, such asLactobacillus species, are able to producebacteriocins which prevent the growth of bacterial competitors as well. These compounds are of obvious value to humans wishing to prevent the growth of bacteria, either asantibiotics or asantiseptics (such asgramicidin S).Fungicides, such asgriseofulvin are also produced as secondary metabolites.[9] Typically secondary metabolites are not produced in the presence of glucose or other carbon sources which would encourage growth,[9] and like primary metabolites are released into the surrounding medium without rupture of the cell membrane.
In the early days of thebiotechnology industry, mostbiopharmaceutical products were made inE. coli; by 2004 more biopharmaceuticals were manufactured in eukaryotic cells, such asCHO cells, than in microbes, but used similarbioreactor systems.[6]Insect cell culture systems came into use in the 2000s as well.[13]
Of primary interest among the intracellular components are microbialenzymes:catalase,amylase,protease,pectinase,cellulase,hemicellulase,lipase,lactase,streptokinase and many others.[14]Recombinant proteins, such asinsulin,hepatitis B vaccine,interferon,granulocyte colony-stimulating factor,streptokinase and others are also made this way.[6] The largest difference between this process and the others is that the cells must be ruptured (lysed) at the end of fermentation, and the environment must be manipulated to maximize the amount of the product. Furthermore, the product (typically a protein) must be separated from all of the other cellular proteins in thelysate to be purified.
Substrate transformation involves the transformation of a specific compound into another, such as in the case ofphenylacetylcarbinol, andsteroidbiotransformation, or the transformation of a raw material into a finished product, in the case of food fermentations and sewage treatment.
In thehistory of food, ancient fermented food processes, such as makingbread,wine,cheese,curds,idli,dosa, among others can be dated to more thanseven thousand years ago.[15] They were developed long before humanity had any knowledge of the existence of themicroorganisms involved. Some foods such asMarmite are the byproduct of the fermentation process, in this case in the production ofbeer.
Fermentation is the main source[citation needed] of ethanol in the production ofethanol fuel. Common crops such assugar cane,potato,cassava, andmaize are fermented by yeast to produce ethanol which is further processed to become fuel.
In the process ofsewage treatment, sewage is digested by enzymes secreted by bacteria. Solid organic matters are broken down into harmless, soluble substances and carbon dioxide. Liquids that result are disinfected to remove pathogens before being discharged into rivers or the sea or can be used as liquid fertilizers. Digested solids, known also as sludge, is dried and used as fertilizer. Gaseous byproducts such as methane can be utilized asbiogas to fuelelectrical generators. One advantage of bacterial digestion is that it reduces the bulk and odor of sewage, thus reducing space needed for dumping. The main disadvantage of bacterial digestion in sewage disposal is that it is a very slow process.
A wide variety ofagroindustrial waste products can be fermented to use as food for animals, especially ruminants. Fungi have been employed to break downcellulosic wastes to increase protein content and improvein vitro digestibility.[16]
Precision fermentation is an approach to manufacturing specific functional products which intends to minimise the production of unwantedby-products through the application ofsynthetic biology, particularly by generating synthetic "cell factories" with engineeredgenomes andmetabolic pathways optimised to produce the desired compounds as efficiently as possible with the available resources.[17] Precision fermentation of genetically modified microorganisms may be used to manufacture proteins needed for cell culture media,[18] providing forserum-free cell culture media in the manufacturing process ofcultured meat.[19] A 2021 publication showed that photovoltaic-driven microbial protein production could use 10 times less land for an equivalent amount of protein compared to soybean cultivation.[20] Some Food Regulatory Agencies such as theFDA do not require the labeling of precision fermented foods as GMO since they are produced by, but do not contain the genetically engineered organisms.[21][self-published source?] It is unclear how regulation will be handled inEU markets, with some Startups such as Formo and Those Vegan Cowboys forming the Food Fermentation Europe (FFE) alliance together with other alt-protein startups to seek regulatory approval.[22]
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