ABSTRACT
A bacterial strain capable of producing extracellular alpha-galactosidase was isolated from sugar cane industrial waste soil sample. Microbiological, physiological, and biochemical studies revealed that isolate belonged to Bacillus sp,. Furthermore, 16S rDNA sequence analysis of new isolates was identified as Bacillus megaterium VHM1. The production of alpha-galactosidase was optimized by various physical culture conditions. Guar gum and yeast extract acted as the best carbon and nitrogen source, respectively for the production of alpha-galactosidase. The enzyme showed an optimum pH at 7.5 and was stable over a pH between 5 and 9. The enzyme was optimally active in 55degreesC and the enzyme was thermostable with half life of 120 minutes at 55 degrees C and lost their 90%, residual activity in 120 minutes at 60 degrees C. alpha-Galactosidase was strongly inhibited by Ag2, Cu2, and Hg2+ at 1mM concentration. The metal ions Fe2, Mn2+, and Mg2+ had no effect on alpha-galactosidase activity, Zn2+,Ni2+, and Ca2+ reduced the enzyme activity slightly. The B megaterium VHM1 enzyme treatment completely hydrolyzed flatulence-causing sugars of soymilk within one and half hour.
Subject(s)
Antifoaming Agents/metabolism , Bacillus megaterium/enzymology , Bacterial Proteins/metabolism , Soy Milk/chemistry , alpha-Galactosidase/metabolism , Antifoaming Agents/chemistry , Bacillus megaterium/classification , Bacillus megaterium/genetics , Bacillus megaterium/isolation & purification , Bacterial Proteins/chemistry , Bacterial Proteins/genetics , Carbohydrates/chemistry , Enzyme Stability , Flatulence , Molecular Sequence Data , Phylogeny , Saccharum/microbiology , alpha-Galactosidase/chemistry , alpha-Galactosidase/geneticsABSTRACT
Alpha-galactosidase was immobilized in a mixture of k-carrageenan and locust bean gum. The properties of the free and immobilized enzyme were then determined. The optimum pH for both the soluble and immobilized enzyme was 4.8. The optimum temperature for the soluble enzymes was 50 degrees C, whereas that for the immobilized enzyme was 55 degrees C. The immobilized enzyme was used in batch, repeated batch, and continuous modes to degrade the raffinose-family sugars present in soymilk. Two hours of incubation with the free and immobilized alpha-galactosidases resulted in an 80% and 68% reduction in the raffinose oligosaccharides in the soymilk, respectively. In the repeated batch, a 73% reduction was obtained in the fourth cycle. A fluidized bed reactor was also designed to treat soymilk continuously and the performance of the immobilized alpha-galactosidase tested at different flow rates, resulting in a 90% reduction of raffinose-family oligosaccharides in the soymilk at a flow rate 40 ml/h. Therefore, the present study demonstrated that immobilized alpha-galactosidase in a continuous mode is efficient for reducing the oligosaccharides present in soymilk, which may be of considerable interest for industrial application.
Subject(s)
Aspergillus oryzae/enzymology , Enzymes, Immobilized/metabolism , Raffinose/metabolism , Soy Milk/chemistry , alpha-Galactosidase/metabolism , Bioreactors , Carrageenan/metabolism , Galactans/metabolism , Hydrogen-Ion Concentration , Mannans/metabolism , Oligosaccharides/metabolism , Plant Gums/metabolism , TemperatureABSTRACT
Alpha-galactosidase from Aspergillus oryzae was immobilized on chitosan beads using glutaraldehyde as a cross-linking agent. The general properties of free and immobilized enzymes were determined. The optimum pH for the free and immobilized enzymes was 4.8 and 4.6 respectively. The optimum temperature for the free enzyme was 50 degrees C, whereas that of immobilized enzyme was increased to 56 degrees C. Kinetic parameters were determined with synthetic substrate (p-nitrophenyl alpha-D-galactopyranoside) and raffinose. Immobilized enzyme showed a higher Km and a lower Vmax than the free enzyme. The immobilized enzymes were used in batch, repeated and continuous mode. A level of 92% hydrolysis was observed at a flow rate of 60 ml/h. The immobilized enzyme was used repeatedly ten times without any change in the performance of the immobilized enzyme in fluidized-bed reactor. The results obtained are of considerable interest for industrial purposes.
Subject(s)
Aspergillus oryzae/enzymology , Flatulence/metabolism , Oligosaccharides/metabolism , Soy Milk/metabolism , alpha-Galactosidase/chemistry , Chitosan , Cross-Linking Reagents/chemistry , Enzyme Stability , Enzymes, Immobilized , Glutaral/chemistry , Hydrogen-Ion Concentration , Nitrophenylgalactosides/chemistry , Temperature , alpha-Galactosidase/metabolismABSTRACT
The hexasaccharide ajugose, alpha-D-galactopyranosyl-(1-->6)-alpha-D-galactopyranosyl-(1-->6)-O-alpha-D-galactopyranosyl-(1-->6)-alpha-D-galactopyranosyl-(1-->6)-alpha-D-glucopyranosyl-(1<-->2)-beta-D-fructofuranoside, generally uncommon in legumes, was detected in the seeds of Vigna mungo L. by TLC and paper chromatography. Ajugose was then isolated by silica gel chromatography and its structure was established by acid and enzymatic hydrolysis, fast atom bombardment mass spectrometry and both one- and two-dimensional 1H and 13C NMR techniques.
