engleski

Development of new bioprocess for lactic acid production

Utilization of different substrates (glucose, sucrose, maltose, celobiose, starch, microcristalline cellulose ; combination of the two substrates), as well as complex raw material (corn grits) by three lactic acid bacteria (LAB), Lactobacillus rhamnosus DSM 20021T, L. amylovorus DSM 20531T (1) and L. coryniformis subsp. torquens 20004T, was investigated in order to develop new bioprocess for lactic acid (LA) production. The bioprocesses were carried out in lab-scale stirred tank bioreactors and horizontal rotating tubular bioreactor (HRTB) (V= 5, 6, 10 i 12 L) in batch, fed-batch, semicontinuous and continuous mode. Procedures for preparation of (heterogenous) samples and their analysis by array of the methods, especially for the samples whithdrawn during semi-solid substrate simultaneous saccharification and fermentation (S5F), was adapted and optimized. Concentration of different carbon sources (glucose, fructose, sucrose, celobiose, maltose, maltotriose, maltotetraose, maltopentaose, maltohexaose, maltoheptaose, starch) and products (LA, acetic acid, ethanol) was determined by HPLC method (1, 2), while concentration of stereoisomers of D- and L-LA was estimated by enzymatic method (1, 2). Derivatization of free aminoacids in water solution, in different media and in corn grits suspension, as well as extraction of the derivates to organic phase were optimized. Fourteen out of twenty amino acid esters were detected and their concentration was determined by using optimized GC method (3). Beside analysis of liquid phase (supernatant), content of (semi-)solid phase of heterogenous samples was estimated (portion of solid particles, concentration of reducing sugars, concentration of nitrogen, thickness of biofilm formed on the inner surface of HRTB). Amylolytic LAB L. amylovorus DSM 20531T catalyzes S5F and different ratios of D- and L-LA were produced, depending on carbon source and temperature of cultivation (1, 2, 4). Different substrates were fermented by L. rhamnosus DSM 20021T to L-LA while L. coryniformis subsp. torquens 20004T produces only D-LA. Amylolytic enzymes produced by L. amylovorus DSM 20531T were partially purified by affinity chromatography. Starch was hydrolyzed by purified amylases in similar manner as it was case with α- and β-amylase isolated from Bacillus sp. and barley, respectively. The tree LAB strains can be distinguished by spectrum of used substrates, consumption of metabolic energy in synthesis of bacterial biomass and/or in synthesys of amylolytic and/or proteolytic enzymes (5) and, thus, employed in different bioprocesses for LA production.

lactic acid bacteria; SSF; stirred tank bioreactor; HRCB; HPLC; GC

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