Riboflavin fermentation

Late in the 1960s, concurrently with the development of the riboflavin fermentation process, a synthetic means of producing riboflavin was discovered. This synthetic process dominated the production of riboflavin until 1972 when some major fermentation strain and process improvements were made with the Ashbya gossypii strain. Since then yields have been significantly improved. The fermentation method now accounts for essentially all the riboflavin produced. 

For therapeutic use, riboflavin is produced by chemical synthesis, whereas concentrates for poultry and Hvestock feeds are manufactured by fermentation using microorganisms such as Jishbyagossypii and remothecium ashbyii which have the capacity to synthesi2e large quantities of riboflavin. 

Fermentative Manufacture. Throughout the years, riboflavin yields obtained by fermentation have been improved to the point of commercial feasibiUty. Most of the riboflavin thus produced is consumed in the form of cmde concentrates for the enrichment of animal feeds. Riboflavin was first produced by fermentation in 1940 from the residue of butanol—acetone fermentation. Several methods were developed for large-scale production (41). A suitable carbohydrate-containing mash is prepared and sterilised, and the pH adjusted to 6—7. The mash is buffered with calcium carbonate, inoculated with Clostridium acetohutylicum and incubated at 37—40°C for 2—3 d. The yield is ca 70 mg riboflavin/L (42) (see Fermentation). 

Manufacturing procedures of riboflavin have also appeared using Saccharomjces bacteria, eg, fermentation with a purine-independent S. reverse mutant (61) and with S. cerevisiae NH-268 (62) produced 2.79 g/L and 4.9 g/L, respectively. 

Further efficient fermentative methods for manufacture of riboflavin have been patented one is culturing C. famata by restricting the carbon source uptake rate, thereby restricting growth in a linear manner by restriction of a micronutrient. By this method, productivity was increased to >0.17 g riboflavin/L/h (63). The other method, using Bacillus subtilis AJ 12644 low in guanosine monophosphate hydrolase activity, yielded cmde riboflavin 0.9 g/ L/3 days, when cultured in a medium including soy protein, salts, and amino acids (64). 

For the industrial production of riboflavin as pharmaceuticals, the traditional methodology comprising the dkect condensation of (13) with (14) in an acidic medium with continuous optimisation of the reaction conditions is stiU used (28). A great part of riboflavin manufactured by fermentative methods is used for feeds in the form of concentrates. The present world demand of riboflavin may be about 2500 t per year. Of this amount, 60%, 25%, and 15% are used for feeds, pharmaceuticals, and foodstuffs, respectively. The main producers are Hoffmann-La Roche, BASF, Merck Co., and others. 

Biotransformations are carried out by either whole cells (microbial, plant, or animal) or by isolated enzymes. Both methods have advantages and disadvantages. In general, multistep transformations, such as hydroxylations of steroids, or the synthesis of amino acids, riboflavin, vitamins, and alkaloids that require the presence of several enzymes and cofactors are carried out by whole cells. Simple one- or two-step transformations, on the other hand, are usually carried out by isolated enzymes. Compared to fermentations, enzymatic reactions have a number of advantages including simple instmmentation reduced side reactions, easy control, and product isolation. 

Riboflavin (ii) riboflavin-5-phosphate E 101 Yes Semisynthetic, obtained from bacterial fermentation (Btakeslea O Q ... 

Niacin, riboflavin, pantothenic acid and vitamin B6 contents are greatly increased in tempeh during fermentation, whereas thiamin exhibits no significant change. H. oligosporus appears to have a great synthetic capacity for niacin, riboflavin, pantothenic acid, and vitamin B, but not for thiamin. 

Recovery of valuable products from penicillin, riboflavin, streptomycin, and vitamin B12 fermentation has been recommended as a viable waste control strategy when incorporated into animal feeds or supplements. Penicillin wastes, when recovered for animal feed, are reported to contain valuable growth factors, mycelium, and likewise evaporated spray-dried soluble matter [31,32,34]. 

Besides volatile solvents, fermentation with Clostridium acetobutyli-cum yields an appreciable amount of riboflavin. The medium should contain 1.5 and 2.0 ppm iron (Meade et al. 1945) and certain salts of organic acids or calcium carbonate (Yamasaki 1939, 1941 Yamasaki and Yositome 1938) for maximal synthesis of riboflavin and greatest fermentation rates. 

Yeast extract, liver extract, or cornmeal can be added to whey, thus achieving normal fermentation and avoiding addition of iron. The presence of these added solids (1%) in whey ensures normal fermentation and high yields of riboflavin. 

The physiological state of the organism influences the yield of riboflavin, as well as that of volatile solvents, and consequently requires control (Leviton 1946). Transfer of inocula when approximately 25% of the gaseous products of fermentation have envolved is conducive to high yields. Butanol-acetone fermentation was of considerable importance during and shortly after World War II. For years it was of little or no importance, but recently it has again become of commercial interest. 

Fermentation industry. Primary metabolites of importance in ihe fermentation field include amino acids, purine nucleotides, vitamins, and organic acids. Specific products include citric acid, riboflavin (vilamin B>). and cubalamin (vilamin B12). Check alphabetical index pertaining lo specific vitamins. Of the secondary metabolites, antibiotics are Ihe mosl... 

Commercial riboflavin dietary supplements are prepared (1) by the fermentation process (bacteria or veasti and (2) by chemical synthesis from alloxan, ribose, and o-xylene. 

The desugaring of cane juice concentrates the heat- and alkali-stable vitamins in the final molasses. Even after this accumulation, only myo-inositol may have reached the level of minimum dietary requirements.109 Niacin, pantothenic acid and riboflavin are also present in significant quantities109 the thiamine, pyridoxin, pantothenic acid, biotin and folic acid contents of molasses have been estimated by bioassay.110 111 The biotin content of Hawaiian and Cuban molasses was 2.1 and 1.7 gammas per gram, respectively.119 The antistiffness factor (closely related to stigmasterol) has been found in cane molasses.88 89 The distillery slop from the yeast fermentation of molasses is marketed as a vitamin concentrate this product also contains vitamins originating in the yeast. 

The many diverse components of milk have demonstrable effects on human health. Perhaps, the most commonly associated component of dairy food is that of dietary calcium. Dairy products provide the most significant contribution to dietary calcium intake in the modem Western diet. It has been estimated that dairy products contribute to >72% of dietary calcium in the United States (Huth et al., 2006). Calcium is an important mineral for maintenance of optimal bone health (Bonjour et al., 2009) and is an integral component of key metabolic pathways relating to, for example, muscle contraction both in skeletal and smooth muscle (Cheng and Lederer, 2008). Further, dairy products contribute other essential nutrients in the diet, such as proteins, phosphorus, potassium, zinc, magnesium, selenium, folate, riboflavin, vitamin B12, and vitamin A (Haug et al., 2007 Huth et al., 2006). Low-fat milk alternatives are fortified with vitamin A and vitamin D which is added to milk and fermented milk in many countries making it an important source for vitamin D (Huth et al., 2006). 

The first organism employed primarily for riboflavin production was Clostridium aceto-butylicum, the anaerobic bacterium used for the microbial production of acetone and butanol. Riboflavin was purely a byproduct and was found in the dried stillage residues in amounts ranging from 40 to 70 xg/g of dried fermentation solids. Later investigations disclosed that riboflavin could be produced by yeast such as Candida flareri or C. guillier-mondi, and the yield was as high as 200 mg/L. 

When a crystal line product is required, the fermented broth is heated for I hr at 121 °C to solubilize the riboflavin. Insoluble matter is removedby centrifugation, andriboflavin recovered by conversion to the less soluble form. The precipitated riboflavin is then dissolved in water, polar solvents, or an alkaline solution, oxidized by aeration, and recovered by recrystallization from the aqueous or polar solvent solution or by acidification of the alkaline solution. 

There are already several examples of chemicals being produced by microbial fermentation of engineered cell factories, whose production through metabolic engineering has been boosted by the use of genomics tools, e.g., 1,3-propanediol used for polymer production, riboflavin used as a vitamin, and 7-aminodeacetoxy-cephalosporanic acid (7-ADCA) used as a precursor for antibiotics production. Furthermore, in the quest to develop a more sustainable society, the chemical industry is currently developing novel processes for many other fuels and chemicals, e.g., butanol, to be used for fuels, organic acids to be used for polymer production, and amino acids to be used as feed. 

De novo fermentation has long been the method of choice for the manufacture of many natural L-amino acids, such as glutamic acid and lysine, and hydroxy acids such as lactic and citric acids. More recently, de novo fermentation is displacing existing multistep chemical syntheses, for example in the manufacture of vitamin B2 (riboflavin) and vitamin C. Other recent successes of white... 

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