Fermentation organic solvents

Apart from using an environmentally friendly solvent, it is also important to clean up the chemical reactions themselves by reducing the number and amount of side-products formed. For this purpose catalysts are a versatile tool. Catalysts have been used for thousands of years in processes such as fermentation and their importance has grown ever since. In synthetic oiganic chemistry, catalysts have found wide applications. In the majority of these catalytic processes, organic solvents are used, but also here the use of water is becoming increasingly popular . 

Isolation. Isolation procedures rely primarily on solubiHty, adsorption, and ionic characteristics of the P-lactam antibiotic to separate it from the large number of other components present in the fermentation mixture. The penicillins ate monobasic catboxyHc acids which lend themselves to solvent extraction techniques (154). Pencillin V, because of its improved acid stabiHty over other penicillins, can be precipitated dkecdy from broth filtrates by addition of dilute sulfuric acid (154,156). The separation process for cephalosporin C is more complex because the amphoteric nature of cephalosporin C precludes dkect extraction into organic solvents. This antibiotic is isolated through the use of a combination of ion-exchange and precipitation procedures (157). The use of neutral, macroporous resins such as XAD-2 or XAD-4, allows for a more rapid elimination of impurities in the initial steps of the isolation (158). The isolation procedure for cephamycin C also involves a series of ion exchange treatments (103). 

Polyethers are usually found in both the filtrate and the mycelial fraction, but in high yielding fermentations they are mosdy in the mycelium because of their low water-solubiUty (162). The high lipophilicity of both the free acid and the salt forms of the polyether antibiotics lends these compounds to efficient organic solvent extraction and chromatography (qv) on adsorbents such as siUca gel and alumina. Many of the production procedures utilize the separation of the mycelium followed by extraction using solvents such as methanol or acetone. A number of the polyethers can be readily crystallized, either as the free acid or as the sodium or potassium salt, after only minimal purification. 

Polar organic solvents readily precipitate exopolysaccharides from solution. The solvents commonly used are acetone, methanol, ethanol and propan-2-ol. Cation concentration of the fermentation liquor influences the amount of solvent required for efficient product recovery. In the case of propan-2-ol, increasing the cation concentration can lead to a four-fold reduction in die volume of solvent required to precipitate xanthan gum. Salts such as calcium nitrate and potassium chloride are added to fermentation broths for this purpose. 

Bioprocess plants are an essential part of food, fine chemical and pharmaceutical industries. Use of microorganisms to transform biological materials for production of fermented foods, cheese and chemicals has its antiquity. Bioprocesses have been developed for an enoimous range of commercial products, as listed in Table 1.1. Most of the products originate from relatively cheap raw materials. Production of industrial alcohols and organic solvents is mostly originated from cheap feed stocks. The more expensive and special bioprocesses are in the production of antibiotics, monoclonal antibodies and vaccines. Industrial enzymes and living cells such as baker s yeast and brewer s yeast are also commercial products obtained from bioprocess plants. 

The concentration of enzyme is veiy low, about several hundred milligrams per litre in the fermentation broth. Solvent extraction is a suitable process to recover a small amount of enzyme. The chance of some enzyme being intracellular is high, therefore cells are ruptured to liberate enzyme, which can then interact with organic solvents. Figure 7.1 shows a simple diagram for a jacketed fermentation vessel for operation at constant temperature. 

A typical penicillin broth contains 20-35 mg/1 of antibiotic. Filtration is used to remove mycelial biomass from fermentation broth. The filtration may be subjected to filter aided polymers. Neutralisation of penicillin at pH 2-3 is required. Amyl acetate or butyl acetate is used as an organic solvent to remove most of the product from the fermentation broth. Finally, penicillin is removed as sodium penicillin and precipitated by a butanol-water mixture. 

Product extraction from large volumes of fermentation broth can be complex, requiring large volumes of organic solvent or solid-phase extraction techniques, which can sometimes greatly reduce or even cancel out the benefits of the biotransformation itself, such as shorter route and environmentally benign conditions. 

Recently however, it has been recognized that liquid extraction is a potential method in the primary recovery of fermentation cell culture products, such as proteins and amino acids. The separation problem, however, is difficult because the product mixtures are often complex, including cell debris and enzymes. Proteins are not suitable for conventional solvent extraction because of incompatibility with organic solvents, but can be handled in aqueous two-phase systems or by extraction in reverse micellar systems (Chapter 15). 

The fermentation broth typically contains 20-30 mg/L of antibiotics, which is to say 30 parts per billion, and must be extracted into concentrated form using solvent extraction. The solvent extraction method was developed by Shell Oil and by Podbielniack and is based on the principle that penicillin is hydrolyzed in aqueous medium to H+ and RCOO ions. Thus, equilibrium in an acidic medium (i.e., one with low pH or high H+ concentration) is favored by the neutral RCOOH form, whereas equilibrium in an alkaline medium (i.e., one with high pH or low H+ concentration) is favored by the RCOO ionic form. The neutral form is more soluble in an organic medium, and the ionic form is more soluble in an aqueous medium. Thus, with amyl acetate as the organic solvent the partition coefficient of penicillin between solvent and water is about 100 at pH 3 and about 1 at pH 6. In the industrial process, the aqueous broth was acidified to pH 3 for the extraction into the organic solvent, and alkalized to a pH 6 for reverse extraction back into an aqueous medium. 

The crude paclitaxel is recovered from the rich aqueous fermentation broth by liquid/liquid extraction with a mixture of isobutyl acetate (IBA) and isopropanol (IPA), both class 3 solvents. The waste aqueous phase is stripped to remove residual organic solvents (IBA/IPA), treated with sodium hydroxide to deactivate any paclitaxel residues, and processed through a standard wastewater treatment facility. The amount of solid waste biomass generated in the process is negligible. 

CO2 can be easily removed from the purified product by decreasing the pressure of the collected fractions (the products are not soluble in the then gaseous CO2). This reduces or, in some cases, eliminates the problem of organic solvent removal encountered with liquid eluents. It must be stressed that the use of CO2 does not increase the greenhouse effect because it either comes from the chemical industry as a by-product or from natural processes like beverage fermentation. CO2, being... 

Special Organic Solvents (e.g., CH.CN) for post-fermentation modification or conjugation reactions such as PEGylation... 

This case represents a composite typical manufacturing process for a mediumsized protein produced by microbial cells. The key process steps are listed in Table 15.5. The overall yield from fermentation to API ranges from 15 to 30% with no recycle or recovery of used materials. Shown in Table 15.6, this composite process confirms the large usage of water in biologies manufacture 10000 to 20000 kg of water for every kg of protein produced. The amounts of organic solvents could be... 

Elfamycins are slightly acidic because of the 4-hydroxy-2-pyndone or the carboxylic acid moiety. They are soluble in most polar organic solvents and the alkali and ammonium salts are water-soluble. The extractability of the free acids from aqueous solution into solvents such as dichloromethane and ethyl acetate is utilized in their isolation from fermentation broths. 

Synonym marsh gas Formula CH4 MW 16.04 CAS [74-82-8] prime constituent of natural gas formed from petroleum cracking, decay of animal and plant remains, and anaerobic fermentation of municipality landfill contents occurs in marshy pools, landfill gas, and leachate from the landfill colorless and odorless gas lighter than air gas density 0.717 g/L liquefies at-161.4°C soluble in organic solvents, slightly soluble in water (25 mg/L) flammable gas. 

The use of extracellular lipases of microbial origin to catalyze the stereoselective hydrolysis of esters of 3-acylthio-2-methylpropionic acid in an aqueous system has been demonstrated to produce optically active 3-acylthio-2-methyl-propionic acid [41-43], The synthesis of the chiral side chain of captopril by the lipase-catalyzed enantioselective hydrolysis of the thioester bond of racemic 3-acetylthio-2-methylpropionic acid (15) to yield 5 -(-)-(15) has been demonstrated [44], Among various lipases evaluated, lipase from Rhizopus oryzae ATCC 24563 (heat-dried cells), BMS lipase (extracellular lipase derived from the fermentation of Pseudomonas sp. SC 13856), and lipase PS-30 from Pseudomonas cepacia in an organic solvent system (l,l,2-trichloro-l,2,2-tri-fluoroethane or toluene) catalyzed the hydrolysis of thioester bond of undesired enantiomer of racemic (15) to yield desired S-(-) (15), R-(+)-3-mercapto-2-methylpropionic acid (16) and acetic acid (17) (Fig. 8A). The reaction yield of... 

Using an enzymatic resolution process, chiral alcohol A-(+)-(43) was also prepared by the lipase-catalyzed stereoselective acetylation of racemic (43) in organic solvent [74], We evaluated various lipases, among which lipase PS-30 and BMS lipase (produced by fermentation of Pseudomonas strain SC 13856)... 

Triacylglycerol lipases (EC 3.1.1.3) are attracting renewed attention since they were demonstrated to be active in organic solvents and to be suitable catalysts in industrial important reactions, such as the synthesis of flavors, emulsifiers, and chiral compounds, and the transesterifi cation of tow-value fats to triacylglycerols of high commercial value. Fbngi are a particularly valuable source of lipases because the enzymes produced by the majority of them are extracellular and readily separable from the mycelia after fermentation. The recent availability of... 

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