Raoudha Ghorbel

Extraction, characterization and antimicrobial activity of sulfated polysaccharides from fish skins

Sulfated polysaccharides were extracted from gray triggerfish (GTSP) and smooth hound (SHSP) skins. Their chemical and physical characteristics were determined using X-ray diffraction and Infrared spectroscopic analysis. The antibacterial activities of GTSP and SHSP against Listeria monocytogenes (ATCC 43251), Staphylococcus aureus (ATCC 25923), Enterococcus faecalis (ATCC 29212), Escherichia coli (ATCC 25922), Salmonella enterica (ATCC 43972) and Enterobacter sp were evaluated by determining clear growth inhibition zone diameters and the minimum inhibitory concentration (MIC) values and by essays in liquid media. GTSP and SHSP were fractionated by a Diethylaminoethyl-cellulose chromatography. Fraction FGII, from GTSP, and fraction FSII, from SHSP, showed the most important inhibitory effects against the tested bacterial species. The sulfated polysaccharides from fish skins did not show hemolytic activity towards bovine erythrocytes. Overall, the results suggested that those polysaccharides could offer promising sources of polysaccharides for future application as dietary ingredients in the nutraceutical industry.

Cloning and constitutive expression of His-tagged xylanase GH 11 from Penicillium occitanis Pol6 in Pichia pastoris X33: Purification and characterization

High-level constitutive expression of xylanase GH11 from Penicillium occitanis Pol6 termed PoXyn2 was achieved using the methylotrophic yeast Pichia pastoris. The PoXyn2 cDNA encoding for a mature xylanase of 320 amino acids was subcloned into the pGAPZαA vector, to construct recombinant xylanse with six histidine residues at the N-terminal and further integrated into the genome of P. pastoris X-33 under the control of the glyceraldehyde 3-phosphate dehydrogenase (GAP) constitutive promoter. Activity assay and SDS-PAGE demonstrate that the His-tagged xylanase was extracellularly expressed in P. pastoris and purified to homogeneity by a simple, one-step purification protocol using immobilized metal affinity chromatography (Ni-NTA resin). The purified PoXyn2 showed a single band on SDS-PAGE with an apparent molecular weight of 30 kDa. The xylanase activity was optimal at pH 3.0 and 50°C. The specific activity measured for Oat Spelt Xylan was 8549.85 U mg(-1). The apparent The K(M) and V(max) values were 8.33±0.7 mg ml(-1)and 58.82±0.9 μmol min(-1) ml(-1), respectively, as measured on Oat Spelt Xylan. This is the first report demonstrating the possibility of mass production of P. occitanis xylanase using P. pastoris.

Polysaccharide from garlic straw: extraction, structural data, biological properties and application to beef meat preservation

A novel polysaccharide (GSP) was isolated from garlic straw (Allium sativum L.) by hot water technique. The structural characterization, antioxidant and antimicrobial activities were investigated. The results showed that GSP was mainly composed of glucose, mannose, galactose and xylose and the major functional groups identified from FT-IR spectrum includes 1631.38 cm−1 (–COO–) and 3193 cm−1 (–OH). In addition, GSP had high DPPH radical scavenging activity, a strong reducing power and inhibited the peroxidation of linoleic acid. The antimicrobial activity of GSP was evaluated against a panel of 7 bacteria and 2 fungal strains using agar diffusion method. Results have shown that GSP exhibited moderate to strong antimicrobial activity against the tested species. These interesting results incite the experimental inoculation of GSP in minced beef meat preservation amended with different concentrations of the GSP and stored at 4 °C for 9 days. The obtained results showed significant inhibitions (p ≤ 0.05) of lipid oxidation over 9 days of aerobic storage and also improvement of meat colour stability while differences in total aerobic cell populations did not change noticeably over storage. Finally, sensory characteristics, e.g. colour, odour and texture, of treated meat with GSP, were higher than the control.

Improving the nutritive value of Olive Cake by solid state cultivation of the medicinal mushroom Fomes fomentarius

Olive Cake (OC) generated by the olive oil industries, well implanted in Tunisia, represents a major disposal and potentially severe pollution problem. This work presents the study of bioconversion of OC in solid state fermentation with the medicinal mushroom, Fomes fomentarius so as to upgrade its nutritional values and digestibility for its use as ruminants feed. The fungus was cultured on OC for 7-30 d, and subsequently the chemical composition, lignocellulolytic enzyme activities and in vitro digestibility of the resultant substrate were determined. The results obtained showed an increase in the crude protein ranging from 6% to 22% for the control and for treated OC, respectively. Significant (P<0.05) decreases in the values of neutral detergent fiber (hemicelluloses, cellulose and lignin), acid detergent fiber (lignin and cellulose) and acid detergent lignin were detected (23%, 13% and 10%, respectively). The estimated in vitro digestibility improved from 9% (control) to 25% (treated OC). The present findings revealed F. fomentarius to be an efficient organism for lignocellulolytic enzymes production and simultaneous enhancement in crude protein and in vitro digestibility of OC.

Functional characterization of Penicillium occitanis Pol6 and Penicillium funiculosum GH11 xylanases

Xylanases are hemicellulolytic enzymes, which are responsible for the degradation of heteroxylans constituting the lignocellulosic plant cell wall. Xylanases from the GH11 family are considered as true xylanases because of their high substrate specificity. In order to study in depth a crucial difference in the thumb region between two closely related xylanases from Penicillium in terms of kinetic parameters and inhibition sensitivity, the GH11 xylanases from Penicillium occitanis Pol6 (PoXyn3) and from Penicillium funiculosum (PfXynC) were heterologously expressed in Pichia pastoris. The PoXyn3 and PfXynC cDNAs encoding mature xylanases were cloned into pGAPZαA vectors and integrated into the genome of P. pastoris X-33 under the control of the glyceraldehyde 3-phosphate dehydrogenase constitutive promoter. PfXynC was expressed as a His-tagged recombinant protein and purified from the supernatant homogeneity by a one-step purification protocol using immobilized metal affinity chromatography. The recombinant PoXyn3 was purified using a single anion-exchange chromatography. The purified recombinant enzymes were optimally active at 45°C and pH 4.0 for PoXyn3 and 40°C and pH 3.0 for PfXynC. The measured kinetic parameters (k(cat) and Vmax) showed that PfXynC was five times more active than PoXyn3 irrespective of the substrate whereas the apparent affinity (K(m)) was similar. The recombinant enzymes showed distinct sensitivity to the Triticum aestivum xylanase inhibitor TAXI-I.

Antibacterial and antioxidant properties of mixed linkage beta-oligosaccharides from extracted β-glucan hydrolysed by Penicillium occitanis EGL lichenase

The aim of this study was first to ascertain the chemical composition and the physicochemical properties of cereal extracted β-glucan from barley flour. Secondly, to assess the antioxidant properties and the antibacterial properties of extracted β-glucan hydrolysates. The proximate composition, FT-IR and scanning electron microscopy of extracted β-Glucan were studied. Hydrolysates from extracted β-glucan, obtained by lichenase EGL from Penicillium occitanis, were a mixed linkage beta-oligosaccharides (MLBO) of trisaccharides and tetrasaccharides. MLBO showed a DPPH radical scavenger with IC50 about 1.8 ± 0.01 mg/mL whereas the IC50 of extracted β-glucan was about 5 ± 0.01 mg/mL. MLBO showed a high antioxidative capacity (175 μmol/mL α-tocopherol equivalents) at 5 mg/mL. The antimicrobial activity was confirmed against all tested bacteria especially at 20 mg/mL of MLBO while no inhibition was observed for all the strains used after the addition of either EGL or extracted β-glucan.

The protective potential of Nitraria retusa on penconazole-induced hepatic injury in adult rats

The protective effect of Nitraria retusa fruit extract against hepatotoxicity induced by penconazole at a dose of 67 mg/kg body weight given intraperitoneally every two days to male Wistar rats was investigated. Penconazole exposure increased malondialdehyde, hydrogen peroxide, and advanced oxidation protein product levels. Hepatic biomarkers as well as enzymatic (catalase, superoxide dismutase and glutathione peroxidase) and nonenzymatic (vitamin C, non-protein thiols and metallothioneins) antioxidant status were also altered. Treatment with N. retusa extract improved all parameters cited above. Liver histological studies confirmed the biochemical parameters. These results provide a scientific basis for the use of N. retusa fruit against hepatotoxicity.