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Presented BY Ashwini P. Kedar MSc. I Year (Sem II) Introduction Chemical process Fermentation Purpose Bioreacter design Downstream processing Application Uses References Lactic acid (2-hydroxypropanoic acid) was discovered and isolated in 1780 by the Swedish chemist Scheele in sour milk. It was first commercially produced in USA in 1881. Lactic acid fermentation is a biological process by which sugars such as glucose, fructose, and sucrose are converted into cellular energy and the metabolic byproduct lactate. The most commercially important genus of lactic acid-fermenting bacteria is Lactobacillus It is an anaerobic fermentation reaction that occurs in some bacteria and animal cells, such as muscle cells in the absence of oxygen. If oxygen is present in the cell, many organisms will bypass fermentation and undergo cellular respiration however, facultative anaerobic organisms will both ferment and undergo respiration in the presence of oxygen. In homofermentive fermentation, one molecule of glucose is ultimately converted to two molecules of lactic acid. Heterofermentative fermentation, in contrast, yields carbon dioxide and ethanol in addition to lactic acid, in a process called the phosphoketolase pathway. The process of lactic acid fermentation using glucose is summarized below. In homolactic fermentation, one molecule of glucose is converted to two molecules of lactic acid:C6H12O6 → 2 CH3CHOHCOOH In heterolactic fermentation, the reaction proceeds as follows, with one molecule of glucose converted to one molecule of lactic acid, one molecule of ethanol, and one molecule of carbon dioxide:C6H12O6 → CH3CHOHCOOH + C2H5OH + CO2 Before lactic acid fermentation can occur, the molecule of glucose must be split molecules of pyruvate. This process is called glycolysis. into two Lactic acid fermentation is the simplest type of fermentation. In essence, it is a redox reaction. In anaerobic conditions, the cell’s primary mechanism of ATP production is glycolysis. Glycolysis reduces – that is, transfers electrons to – NAD+, forming NADH. However, there is only a limited supply of NAD+ available in a cell. For glycolysis to continue, NADH must be oxidized – that is, have electrons taken away – to regenerate the NAD+. This is usually done through an electron transport chain in a process called oxidative phosphorylation however, this mechanism is not available without oxygen. Instead, the NADH donates its extra electrons to the pyruvate molecules formed during glycolysis. Since the NADH has lost electrons, NAD+ regenerates and is again available for glycolysis. Lactic acid, for which this process is named, is formed by the reduction of pyruvate In homolactic acid fermentation, both molecules of pyruvate are converted to lactate. In heterolactic acid fermentation, one molecule of pyruvate is converted to lactate; the other is converted to ethanol and carbon dioxide. Homolactic acid fermentation is unique in that it is one of the only respiration processes that do not produce a gas as a byproduct. Some bacteria and yeasts organisms are unable to cope with the presence of oxygen. These organisms use fermentation as a method of obtaining energy in the form of ATP .Because the production of lactic acid frees up NAD+ the process of glycolysis can continue. Lactic acid fermentation also occurs in animal muscle cells under conditions when oxygen is low. Extreme exercise would be an example of this. In this situation, the lactate is carried away by the circulatory system to the liver where it is converted back to pyruvate through the Cori cycle Fermentation, however, is far less effective than cellular respiration producing only two ATP molecules per glucose molecule consumed. The typical yield from cellular respiration is anywhere from 34-38 molecules of ATP. Thus, it is typically seen only in small organisms, such as bacteria and yeast, that can survive on this low energy yield. Based on the past success with membrane bioreacter They started work in1983 for contineous production of lactic acid . They design a continuous membrane bioreactor (CMB) as a continuous stirred tank reactor (CSTR) coupled in a semi closed loop configuration to a membrane module, as shown in the diagram. Synthetic semi-permeable membranes are used to separate and recycle the lactic acid bacteria, while simultaneously removing the lactate as it is formed. This has several advantages over batch fermenters. The continuous separation and recovery of the bacterial cells will reduce cycle time of the fermenters, since there will be little or no time lost due to start-up and shut down as in present batch fermenters. The recycle of the cells will allow us to obtain much higher cell densities than currently practiced. Laboratory studies have shown a 100-fold increase in cell numbers in the CMB during operation. The high concentration allows us to pump the feedstock through the fermenters much faster. "Cell wash-out" is eliminated, thereby allowing operation at dilution rates greater than the specific growth rate of the organism. The continuous removal of lactate allows us to maintain the fermenter at just below the lactate level which inactivates the cells. Thus the cells are always viable and producing lactate. The membrane bioreactors are very flexible, allowing a range of outputs that can be matched very easily to the demands of upstream and downstream operations. The product stream from the fermenter is clear, containing no suspended matter. This will improve the subsequent recovery and purification process, with a further reduction in cost. The membrane units are available in modular systems, making expansion easy. Due to the high productivity, floor space requirements for the membrane bioreactor system are much less than with present-day batch fermenters. Another configuration we investigated was the hollow fiber bioreactor, which was operated in a quasi plug flow mode. The CSTR-membrane configuration is preferred since it is a well-mixed system that allows us to efficiently neutralize the fermentation broth with the appropriate alkali (usually ammonium or sodium hydroxide). Lactic acid can be separated and substantially purified from fermentation broths by several membrane-based unit operations as shown in the diagram below. Microfiltration or ultrafiltration for cell separation and recycle Nanofiltration for separation of the lactic acid from other broth components using low rejection (LR) membranes Concentrating the lactate using reverse osmosis (RO) or a combination of high rejection (HR) and low rejection (LR) nanofiltration membranes Electrodialysis (ED) for simultaneous separation and concentration of lactate. A conventional anion-/cation-exchange membrane ED system will purify and concentrate the lactate, but the lactate product will still be in the salt form . On the other hand, a bipolar membrane ED system will result in the acid form of lactic acid and allow the recycle of the alkali used for neutralizing the fermentation broth. This minimizes alkali cost, as well as eliminating the waste product (e.g., calcium sulfate) generated in conventional downstream processes for organic acids Yogurt Production : The main method of producing yogurt is through the lactic acid fermentation of milk with harmless bacteria. The primary bacteria used are typically Lactobacillus bulgaricus and Streptococcus thermophilus and US law requires all yogurts to contain these two cultures These bacteria produce lactic acid in the milk culture, decreasing its pH and causing it to congeal. The bacteria also produce compounds that give yogurt its distinctive flavor. An additional effect of the lowered pH is the incompatibility of the acidic environment with many other types of harmful bacteria. For a probiotic yogurt, additional types of bacteria such as Lactobacillus acidophilus are also added to the culture. Sauerkraut : Lactic acid fermentation is also used in the production of sauerkraut. The main type of bacteria used in the production of sauerkraut is of the genus Leuconostoc As in yogurt, when the acidity rises due to lactic acid-fermenting organisms, many other pathogenic microorganisms are killed. The bacteria produce lactic acid, as well as simple alcohols and other hydrocarbons These may then combine to form esters contributing to the unique flavor of sauerkraut. Lactic acid is used as a humectant or moisturizer, in some cosmetics and as a mordant, a chemical that helps fabrics accept dyes, in textiles It is also used in making pickles and sauerkraut, foods for which a sour taste is desired. Lactic acid is used in the dairy industry not only in making yogurt but in making cheese as well. It is also used in tanning leather. Lactic acid is important in the pharmaceutical industry as a starting material for other substances and is involved in the manufacturing of lacquers and inks. A related compound that is made from lactic acid is calcium stearoyl-2-lactylate, which is used as a food preservative. Food Biotechnology; 2nd edition by Kalidas Shetty, Gopinathan Paliyath, Anthony Pometto, Robert E Levin ; pg no 144-156. Modern Industrial Microbiology & Biotechnology by Nduka Okafor; pg no.281-289. Thank You