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Characterization of amylolytic activity of bacterium Lactobacillus amylovorus DSM 20531T by determining concentration of simpler carbohydrates by ion-exchange high-pressure liquid chromatography

Waste materials and the most abundant complex carbohydrates, such as starch and cellulose and their hydrolysates, are often used as substrates in the biotechnological production of alcohols, organic acids, enzymes and biopolymers. Amylolytic lactic acid bacterium (ALAB) Lactobacillus amylovorus DSM 20531T is capable of conducting simultaneous saccharification and fermentation (SSF) of starch containing substrates. A diversity of oligosaccharides and simple carbohydrates are produced during starch degradation in SSF and no single method is potent enough in separating and determining all of them. In this work amylolytic activity of the strain was characterized by determining concentration of a few simpler carbohydrates (glucose, G1 ; maltose, G2 ; maltotriose, G3 ; maltotetraose, G4 ; maltopentaose, G5 ; maltohexaose, G6 and maltoheptaose, G7) by ion-exchange high-pressure liquid chromatography. G1, G2, G3 and G4 have been eluted from the column and base-line separated in descending order of molecular size from their standard mixture and from the samples while G5 - G7 co-eluted from the column. Data obtained by HPLC analysis and with saccharogenic (Somogyi-Nelson or RS) and amyloclastic (iodometric) methods were compared and L. amylovorus DSM 20531T amylases have been found to form G2, G3, G4, G5 - G7 oligosaccharides, but also some with higher degree of polymerisation. G1 has not been detected in the samples. The ion-exchange HPLC method was proved to be the method of choice for efficient determination of carbohydrates from the samples that are derived from complex biologic matrices, such as corn grits.

Lactobacillus amylovorus; simultaneous saccharification and fermentation (SSF); amylolytic activity; simpler carbohydrates; HPLC analysis

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