5544
Eman Ibrahim Ibrahim
Production of mannanase enzyme by some local fungal strains and biomass utilization for the removal of some heavy metals from aqueous solutions
Fungal Mannanase, Aspergillus tammarii NRC3, Cell- Immobilization, Mannooligosaccharides, Prebiotic potential, Bioremediation, Biosorbents
In the present study forty locally fungal isolates were tested for their abilities to produce β-mannanase in both static and shaking cultures. The fungus Aspergillus tamarii NRC3 was found to be the most potent producer for βmannanase and therefore it was used throughout this study. The favorable cultural conditions for Aspergillus tamarii NRC3 β-mannanase production found to be; locust bean gum (1%), sodium nitrate (0.2%), inoculum age (6 days), inoculum size (one disk, 4 mm diameter, equal 2x107spores/ml), production medium volume (50ml), 7 days of static incubation, pH 5.0 , 25 ºC and no effect of surfactants as tween 20, 80 and SDS on β-mannanase production was detected. The agarose (3%) was the best matrix at entrapment cell immobilization technique, where gave a powerful effectiveness factor (1.01 Cimmo/Cfree). Acetone fractional precipitation of the crude enzyme at 25-50% concentration was the successful partial purification of Aspergillus tamarii NRC3 β-mannanase and exhibited the highest recovered activity (37.25u/mg). The enzyme was completely purified by ion-exchange chromatography (DEAEcellulose column) and gel filtration chromatography (Sephadex G-100 column) with 4.97 purification fold and 15.46% enzyme recovery. Molecular weight of the purified β-mannanase was estimated to be 80KDa. The pure enzyme was appeared to be rich with glutamic acid and aspartic acid in addition to traces of other amino acids. Optimal physicochemical properties of purified β-mannanase were at pH 6.0, 40°C and the enzyme was stable at pH 5.5 up to pH 6.5 and was thermostable up to 60°C for first 5 min; while the energy of activation was calculated as 3.052546 Kcal/ mol. The kinetic parameters, Michalis-Menten constant (Km) and maximum velocity (Vmax), of purified Aspergillus tamarii NRC3β-mannanase were about 0.10 mM, and 16.10 μmol/ml/min, respectively, with isoelectric point (pI) 4.2 based on pH– precipitation profile. Most of metal ions tested found to be had inhibition influence on β-mannanase activity at 1Mm or 10Mm by inhibition percentage of 1% - 22%, while, Hg+2 had a completely inhibition effect on β-mannanase activity. Glutathion and Mercaptoethanol had activation effect on purified β-mannanase activity. Chemical compounds as SDS, tween 20, tween 60, tween 80, EDTA had inhibitory effects on β-mannanase activity. The degree of polymerization (DP) of the hydrolysis process product value was ~3.5. Thin-layer chromatography (TLC) analysis of hydrolysis process product at various incubation times referred to produce a variety of mannooligosaccharides (MOS) including, mannobiose, mannotriose, mannotetrose, and mannopentose. Evaluation of possible prebiotic potential of the MOS indicated that MOS had ability for enhancement the growth of Lactobacillus spp., and had inhibitory effect on the growth of tested potentially pathogenic strains. Biomass produced as a byproduct from the mannanase production process was evaluated as a biosorbent for the removal and recovery of some heavy metal ions especially Cu2+from aqueous solutions. The maximum uptake of Cu+2 was achieved at the initial 5min (92%), 30°C, pH 5, and biomass concentration 5g w/w. In addition to Cu2+, the fungal biomass was able on removing considerable proportion of Pb2+, Co+2, Ni2+, Fe+3, and Cr3. The uptake of Cu+2 by pretreated biomass was studied. Recovery of the sorbed metal ions by desorbing agents and the potential reuse of the regenerated biomass in metal ions uptake (reloading) were evaluated. For the removal and recovery of metal ions from an industrial effluent (The Egyptian Company for leather tanning) by Aspergillus tamarii NRC3 biomass, it was observed that the efficiency of Cu2+ and Co2+ removal was high (90.94%, and 60.0%, respectively) followed by Ni2+, Fe3+, Pb2+, and Cr3+ (40.0, 34.47, 29.13 and 11.45%, respectively) also, unwanted color and odour were removed
2019
Ph.d
Ain Shams
Science