R .A .B . PE SS O NI , R . C. L. F IG UE I RE DO -RI BE I RO AN D M. R. B RA GA . 1999. Extracellular inulinases from Penicillium janczewskii were obtained from the filtrate of 12 day-old cultures supplemented with inulin from Vernonia herbacea. Crude filtrates and partially-purified enzyme preparations (peaks I and II) were active on inulin, sucrose and raffinose. The apparent M r of the enzymes from peaks I and II were 48 and 66 kDa, respectively. The apparent K m (mmol l −1 ) values of peak I were 0·43 for inulin and 18·7 for sucrose; for peak II they were 0·87 and 18·5 for inulin and sucrose, respectively. Their temperature and pH optima were 55°C and 5·0, respectively. Both peaks catalysed the hydrolysis of b-(2,1) fructans more rapidly than b-(2,6) fructans. Free fructose was the predominant product released from inulin, indicating that these enzymes display exo-inulinase activity. In view of these characteristics, the yield and the high specific activity towards b-(2,1) fructans, inulinases from P. janczewskii can be utilized for the preparation of fructose syrup from inulin.

Penicillium janczewskii, a filamentous fungus isolated from the rhizosphere of Vernonia herbacea (Asteraceae), grows rapidly on media containing either sucrose or inulin as carbon sources. Maintenance of P. janczewskii on inulin medium induces secretion of proteins with high inulinase activity but results in a mycelium that easily collapses and breaks. We evaluated the influence of inulin on fungal growth and colony morphology and on cell-wall structure and composition in comparison with growth and wall characteristics on sucrose-containing medium. P. janczewskii grown on Czapek medium with agar containing 1% (w/v) sucrose or inulin showed differences in the color and morphology of the colonies, although growth rates were similar on both carbon sources. Scanning-electron microscopy revealed that the hyphae from fungus grown on inulin-containing medium are much thinner than those from fungus cultivated on sucrose. Ultrastructural analysis of 5 d old cultures using transmission-electron microscopy indicated significant differences in the cell-wall thickness between hyphae grown on inulin or sucrose media. No differences were detected in the overall carbohydrate and protein contents of cell walls isolated from cultures grown on the two carbon sources. Glycosyl composition analyses showed glucose and galactose as the predominant neutral monosaccharides in the walls but showed no differences attributable to the carbon source. Glycosyl linkage composition analyses indicated a predominance of 3-linked glucopyranosyl in the hyphal walls when P. janczewskii was grown on inulin-containing medium. Our results suggest that growth on inulin as the sole carbon source results in structural changes in the mycelia of P. janczewskii that lead to mycelial walls with altered physical and biological properties.

Penicillium janczewskii, isolated from the rhizosphere of Vernonia herbacea, grows rapidly on media containing either sucrose or inulin, although inulin more than sucrose induced the production of inulinases. Three different extracellular beta-fructofuranosidases (two inulinases and one invertase) were purified from fungal cultures grown on sucrose or inulin, through precipitation with ammonium sulfate, and anion-exchange, hydrophobic interaction and gel filtration chromatographies. The optimum temperature of the three enzymes was approximately 60 C, optimum pH 4-5.5 and apparent molecular mass of 80 kDa. K(m) and V(max) values determined for invertase on sucrose were respectively 3.7 10(-4) M and 7.9 10(-2) micromol/min/mL, and on inulin 6.3 10(-2) M and 2.09 10(-2) micromol/min/mL. The values of k(m) for the two inulinases were 8.11 10(-4) and 2.62 10(-3) M, being lower for inulin when compared to those obtained for sucrose. The inulinases did not produce oligofructans from inulin, indicating they are primarily exoinulinases. The differences found in inulinase induction patterns when inulin or sucrose was used seem to be related to modifications on the enzyme properties, mainly concerning substrate affinity.

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-(Sugarcane bagasse as a source of carbon for enzyme production by fi lamentous fungi). The aim of the present work was to assess the enzymatic activity of six strains of fi lamentous fungi grown in liquid media containing 1% sugarcane bagasse as the sole carbon source. All fungal strains were able to use this agro-industrial residue, producing various types of enzymes, such as cellulases, xylanases, amylases, pectinases, and laccases. However, Aspergillus japonicus Saito was the most effi cient producer, showing the highest enzymatic activity for laccase (395.73 U L -1 ), endo-β-1,4-xylanase (3.55 U mL -1 ) and β-xylosidase (9.74 U mL -1 ) at seven, fourteen and twenty-one days in culture, respectively. Furthermore, the endo-β-1,4-xylanases and β-xylosidases of A. japonicus showed maximum activity at 50°C, and pH 5.5 and pH 3.5-4.5, respectively. Thus, these results indicate that A. japonicus has a great biotechnological potential for the production of these enzymes using sugarcane bagasse as the sole source of carbon.

Penicillium janczewskii, a filamentous fungus isolated from the rhizosphere of Vernonia herbacea (Asteraceae), grows rapidly on media containing either sucrose or inulin as carbon sources. Maintenance of P. janczewskii on inulin medium induces secretion of proteins with high inulinase activity but results in a mycelium that easily collapses and breaks. We evaluated the influence of inulin on fungal growth and colony morphology and on cell-wall structure and composition in comparison with growth and wall characteristics on sucrose-containing medium. P. janczewskii grown on Czapek medium with agar containing 1% (w/v) sucrose or inulin showed differences in the color and morphology of the colonies, although growth rates were similar on both carbon sources. Scanning-electron microscopy revealed that the hyphae from fungus grown on inulin-containing medium are much thinner than those from fungus cultivated on sucrose. Ultrastructural analysis of 5 d old cultures using transmission-electron microscopy indicated significant differences in the cell-wall thickness between hyphae grown on inulin or sucrose media. No differences were detected in the overall carbohydrate and protein contents of cell walls isolated from cultures grown on the two carbon sources. Glycosyl composition analyses showed glucose and galactose as the predominant neutral monosaccharides in the walls but showed no differences attributable to the carbon source. Glycosyl linkage composition analyses indicated a predominance of 3-linked glucopyranosyl in the hyphal walls when P. janczewskii was grown on inulin-containing medium. Our results suggest that growth on inulin as the sole carbon source results in structural changes in the mycelia of P. janczewskii that lead to mycelial walls with altered physical and biological properties.

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Fructose, glucose, and an equimolar mixture of both sugars affected differently hyphae thickness, biomass production and secretion of β-fructofuranosidase in Penicillium janczewskii. Reduced growth, thinner hyphae and visible injuries were early observed during fungal cultivation in fructose-containing medium, reaching the maximum between 12 and 15 days of culture. Total sugar content from the cell wall was lower when fructose was supplied and polysaccharides lower than 10 kDa predominated, regardless the culture age. Maximal inulinase and invertase activities were detected in culture filtrates after 12 days, excepting in the glucose-containing medium. Structural changes in cell walls coincided with the increase of extracellular enzyme activity in the fructose-containing medium. The fragility of the hyphae might be related with both low carbohydrate content and predominance of low molecular weight glucans in the walls. Data presented here suggest changes in carbohydrate component of the cell walls are induced by the carbon source.