Studies On Mannitol Production By Lactic Acid Bacteria

Mannitol is a naturally occurring sugar alcohol with six carbon atoms. Mannitol is usedextensive in several industrial areas, such as food industry,pharmaceutical industry,chemicalindustry,light industry. At present mannitol is produced commercially by natural extraction andcatalytic hydrogenation methods. Microbial (LAB, Lactic Acid Bacteria) transformation forproducing mannitol has many advantages, such as high reaction selectivity, mild reactionconditions, less sorbitol production and high yield.Firstly,determination of mannitol content in broth by TLC(Thin-Layer Chromatography )and spectrophotometry were studied. Thin-Layer Chromatography was performed with silica gel60 using a solvent system of butatone:acetic acid:water (40:104: by volume). While determinedby spectrophotometry, Influence of fructose in broth was eliminated by dehydration withhydrochloric acid.225 strains of gram-positive bacteria were isolated from traditional fermented foods,Sauerkraut. MRS fructose medium, Thin-Layer Chromatography and spectrophotometry wereused to analyze the abilities of production of mannitol. PM152 was selected as the optimal strainfor the bioconversion of mannitol. PM152was identified as Lactobacillus buchneri by APIidentification system.The growth characteristics of stain PM152 were studied. It grew at optimal temperature 32℃,and the minimum 13℃,maximum 43℃. As for initial pH for cell growth ,the optimal was 6.5,as well the upper and lower limitations was 9.4 and 3.6, respectively. The growth of PM152culture was tested in MRS broth. It was found that PM152 began logarithmic stage andmaximum growth at 2nd hour and 10th hour. By using screened bacteria,the effect of differentkinds of carbon sources,the ratio of fructose to glucose in carbon source, carbon sourceconcentration, nitrogen source concentration, the concentration of inorganic metal ion, theconcentration of buffer agent on bacterium species growth and mannitol's transformation wereoptimized. The results showed that the fermentate culture composition is: carbon sourceconcentration of 150g/L in which the fructose to glucose ratio is 2:1,the concentrations of thethree kinds nitro sources, peptone, meat extract and yeast extract, are 5 g/L,5 g/L,2.5 g/L,1g/Lrespectively, the concentrations of MgS04 .7H20 and MnS04 .4H20 are 0.55g/Land 0.28 g/L. Theinoculation volume, initial fermentative culture pH, fermentative time, and the shaker rotatespeed and fermentative temperature which effects on biotransformation were explored. Theoptimal process conditions are concluded: the rotate speed of 80 rpm, the inoculation volume of8%, initial pH of 6.8, fermentative time of 48h and the fermentative system temperature of 32℃,and the mannitol's production rate was 78.51%.