M. Gutiérrez-Rojas

Effects of different carbon sources on the synthesis of pectinase by Aspergillus niger in submerged and solid state fermentations

A study was made to compare the production of pectinase by Aspergillus niger CH4 in solid-state (SSF) and submerged (SmF) fermentations. Production of endo- (endo-p) and exo-pectinase (exo-p) by SSF was not reduced when glucose, sucrose or galacturonic acid (up to 10%) were added to a culture medium containing pectin. Moreover, both activities increased when concentrations of the carbon sources were also increased. In SmF, these activities were strongly decreased when glucose or sucrose (3%) was added to culture medium containing pectin. The addition of galacturonic acid affected endo-p activity production to a lesser extend than exo-p. Final endo-p and exo-p activities in SSF were three and 11 times higher, respectively, than those obtained in SmF. The overall productivities of SSF were 18.8 and 4.9 times higher for endo-p and exo-p, respectively, than those in SmF. These results indicate that regulatory phenomena, such as induction-repression or activation-inhibition, related to pectinase synthesis by A. niger CH4 are different in the two types of fermentation.

Production and properties of three pectinolytic activities produced by Aspergillus niger in submerged and solid-state fermentation.

Three extracellular pectinases were produced byAspergillus niger CH4 by submerged and solid-state fermentation, and their physicochemical and kinetic properties were studied. The highest productivities of endo- and exo-pectinase and pectin lyase were obtained with solid-state fermentation. The kinetic and physicochemical properties of these enzymes were influenced by the type of culture method used. All activities were very different in terms of pH and temperature optima, stability at different pH and temperature values and affinity for the substrate (Km values). In solid-state fermentation, all pectinase activities were more stable at extreme pH and temperature values but theKm values of endo-pectinase and pectin lyase were higher with respect to those activities obtained by the submerged-culture technique. The pectin lyase activity obtained by the submerged-culture technique showed substrate inhibition but the enzyme obtained by solid-state fermentation did not. Electrophoresis, using sodium dodecyl sulphate/polyacrylamide gel with enzymatic extracts obtained for both culture methods, showed the same number on protein bands but some differences were found in their electrophoretic position. The results obtained in this work suggest that the culture method (submerged or solid-state) may be responsible for inducing changes in some of the pectinolytic enzymes produced byA. niger.

Kinetics of growth of Aspergillus niger during submerged, agar surface and solid state fermentations

The effect of initial glucose concentration (30–450 g/litre) on the growth of Aspergillus niger 10 in submerged (SMF), agar surface (ASF) and solid state (SSF) fermentations was examined. Biomass production and specific growth rate were considerably less sensitive to changes in initial glucose concentration in SSF compared to either ASF or SMF. In all cases, maximum specific growth rates (0·323, 0·247 and 0·134 1h for SSF, ASF and SMF, respectively) were obtained with the lowest initial glucose concentration employed (50 g/litre for SSF and 30 g/litre for both ASF and SMF). The rates of glucose uptake in SSF were considerably less affected by initial glucose concentrations when compared with ASF and SMF. Complete consumption of glucose in SSF, ASF and SMF was observed when glucose was applied at initial concentrations of 200, 150 and 100 g/litre, respectively. These results indicate that SSF is more suitable than either ASF or SMF for the growth of filamentous fungi, especially under conditions where catabolite repression applies (e.g. due to high glucose concentrations).

In vitro evaluation of germination and growth of five plant species on medium supplemented with hydrocarbons associated with contaminated soils

The effect of a hydrocarbon mixture (HCM) of three polycyclic aromatic hydrocarbons (PAH) and Maya crude oil on germination, growth and survival of four grasses (Bouteloua curtipendula, Cenchrus ciliaris, Echinochloa crusgalli and Rhynchelytrum repens) was studied and compared to a control (Festuca arundinacea) under in vitro conditions. The species were cultured on MS medium with different HCM initial concentrations. Germination was not affected for any assayed concentration; however, the length of the stems and roots decreased when HCM increased and the survival of the four species also diminished. Except for F. arundinacea, a direct link between hydrocarbon concentration and plant survival was observed. In vitro studies are clean and easy to handle techniques allowing isolation of the plant activity from that derived from associations with microorganisms in non-sterile cultures. To our knowledge, this is the first work towards phytoremediation assisted by in vitro plant cultivation.

Effect of water activity on exo-pectinase production by Aspergillus niger CH4 on solid state fermentation

SummaryEthylene glycol, sorbitol and glycerol were used as water activity depressors to study the effect of water activity on pectinase production byAspergillus niger CH4. Ethylene glycol depressed aw without supporting growth nor strongly affecting pectinase production in petri dish cultures. This depressor was used to evaluate the influence of water activity on exo-pectinase production by SSF. It was found that although pectinase production decreased at low aw values, this activity was present at aw values as low as 0.90. The specific activity increased up to 4.5 fold by reducing aw from 0.98 to 0.90. The reducing groups accumulated extracellularly suggesting sugar transport limitation as a consequence of aw depression.

Citric acid and polyols production by Aspergillus niger at high glucose concentration in solid state fermentation on inert support

SummaryAspergillus niger cultures at high initial glucose concentration (up to 400 g/1) on Amberlite as inert support were carried out. Citric acid was accumulated in the support showing high concentration (94.54 g/l) and productivity (1.35 g/l h) without inhibition related to the presence of metals (Mn2+, Zn2+, Co2+, Cu2+, and Ca2+) at high concentrations. Citric acid accumulation was clearly associated with both, glycerol production and to the age of the culture. Glycerol and erythritol, the major osmoregulator metabolites, were also produced (8.16 and 24.57 g/l respectively) at 400 g/l of glucose.

HEAT TRANSFER IN CITRIC ACID PRODUCTION BY SOLID STATE FERMENTATION

Different mechanisms of heat removal (conductive, convective and evaporative) were studied in a SSF process using a packed bed reactor with an inert support (Amberlite IRA-900) previously inoculated. The cylindrical bioreactor was incubated in a temperature controlled chamber (30°C). Average temperature gradients obtained during the culture were 1·3 and 0·42°C cm−1 in the radial and axial directions, respectively. During maximal metabolic activity of Aspergillus niger the medium temperature rose from 32 to 48°C, and a maximum radial temperature gradient of 3·1°C cm−1 was achieved. At this stage, the unsteady-heat balances indicated that conductive heat transfer was the least efficient mechanism (8·65%) when compared with convective (26·65%) and evaporative (64·7%) mechanisms. Sugar consumption and glycerol and citric acid production were strongly affected by temperature.

Effect of nitrogen loading rate and carbon source on denitrification and sludge settleability in upflow anaerobic sludge blanket (UASB) Reactors

The combined effect of carbon source and nitrogen loading rate (NLR) on denitrification and sludge granular settleability in upflow anaerobic sludge blanket (UASB) bench-scale reactors was studied. Acetate, lactate and glucose were used as carbon sources for denitrification. Three NLR were evaluated: 500, 1000 and 2000 mg NO3−-N/ld. It was found that NO3−-N consumption efficiency was 99% for all the cases. Denitrification efficiencies were high (85-96%) for all substrates and NLR tested, but specific denitrifying rates were a better representation of the denitrifying process. The results showed that specific denitrifying rate was dependent on type of carbon source and NLR. It was also found that sludge settleability and reactor stability was influenced by the carbon source. The use of acetate did not influence the sludge settleability while, sludge flotation was always observed when lactate was used as carbon source. Foaming was present when glucose was used, causing reactor unstability. An increase in the exopolymeric protein and carbohydrate content was observed when lactate and glucose were fed, in comparison with acetate as carbon source. Results obtained in this work suggest that the exopolymeric composition here referred to as protein/carbohydrate ratio (P/C) could be a better stability indicator than the sludge volumetric index (SVI).

Biodegradation of hexadecane in liquid and solid-state fermentations by Aspergillus niger

The biodegradation and mineralisation of hexadecane (HXD) by Aspergillus niger were studied in SmF and Solid-state fermentation (SSF). HXD concentrations ranging from 45 to 180 g/l (SSF) and from 20 to 80 g/l (SmF) were tested. HXD consumption was three times higher and fungal growth was up to 30 times faster in SSF than in SmF. The maximum HXD consumption in SmF was 62% (18% mineralised) and in SSF 100% (52% mineralised) for initial HXD concentrations of 20 and 45 g/l, respectively. The respiratory quotient in SmF increased (from 0.85 to 1.08) with increase in HXD concentration, while it was independent (approximately 0.68) of the initial HXD concentration in SSF. These results showed that the consumption rate and biodegradation efficiency for HXD were higher in SSF than in SmF.

Criteria for the selection of strains of entomopathogenic fungi Verticillium lecanii for solid state cultivation

The objective of this study was the selection of strains of Verticillium lecanii for solid-state fermentation (SSF) containing cuticle of Sphenarium purpurascens as an inducer of proteases and chitinases. The selection criteria were: growth at low water activity (aW), enzymatic activities (proteases and chitinases) and CO2 production rate. Three strains of V. lecanii were studied ATCC 26854, ATCC 46578 and a wild strain (WS). The strains ATCC 26854 and WS presented the best biomass production at low aW (0.957). Highest rates of clearing zones of casein and chitin were obtained for strains ATCC 26854 and WS. Best results of CO2 production in SSF were obtained by using V. lecanii ATCC 26854 which showed a maximal value (2.3 mg CO2 g IDM−1 h−1) at 36 h of cultivation. Although clearing zones of casein and chitin were partial criteria for strain selection. It was concluded that growth a low water activities and CO2 production rate, were more reliable criteria for selecting strains of V. lecanii for solid state culture using cuticle of insect as the main C and N source.