Evaluation of different bacterial honey isolates as probiotics and their efficient roles in cholesterol reduction.

Continue to hypothesize that honey is a storehouse of beneficial bacteria, and the majority of these isolates are levansucrase producers. Accordingly, ten bacterial strains were isolated from different honey sources. Four honey isolates that had the highest levansucrase production and levan yield were identified by the partial sequencing of the 16S rRNA gene as Achromobacter sp. (10A), Bacillus paralicheniformis (2M), Bacillus subtilis (9A), and Bacillus paranthracis (13M). The cytotoxicity of the selected isolates showed negative blood hemolysis. Also, they are sensitive to the tested antibiotics (Amoxicillin + Flucloxacillin, Ampicillin, Gentamicin, Benzathine benzylpenicillin, Epicephin, Vancomycin, Amikacin, and Zinol). The isolates had strong alkaline stability (pHs 9, 11) and were resistant to severe acidic conditions (29-100 percent). The tested isolates recorded complete tolerance to both H2O2 and the bile salt (0.3% Oxgall powder) after 24 h incubation. The cell-free supernatant of the examined strains had antifungal activities against C. Albicans with varying degrees. Also, isolates 2M and 13M showed strong activities against S. aureus. The isolates showed strong adhesion and auto-aggregation capacity. Isolate 10A showed the highest antioxidant activity (91.45%) followed by 2M (47.37%). The isolates recorded different catalase and protease activity. All isolates produced cholesterol oxidase and lipase with different levels. Besides, the four isolates reduced LDL (low-density lipoprotein) to different significant values. The cholesterol-reducing ability varied not only for strains but also for the time of incubation. The previous results recommended these isolates be used safely in solving the LDL problem.

Optimization strategy of Bacillus subtilis MT453867 levansucrase and evaluation of levan role in pancreatic cancer treatment.

Pancreatic cancer is the fourth most lethal cancer type worldwide. Due to multiple levan applications including anticancer activities, studies related to levansucrase production are of interest. To our knowledge, levan effect on pancreatic cancer cells has not been tested previously. In this work, among eighteen bacterial honey isolates, Bacillus subtilis MT453867 showed the highest levan yield (33 g/L) and levansucrase production (8.31 U/mL). One-factor-at-a-time technique increased levansucrase activity by 60% when MgSO4 was eliminated. The addition of 60 g/L banana peels enhanced the enzyme activity (192 U/mL). Placket Burman design determined the media composition for maximum levan yield (54.8 g/L) and levansucrase production (505 U/mL). The identification of levan was confirmed by thin-layer chromatography, Fourier-Transform Infrared spectrometric analysis, 13C-nuclear-magnetic resonance, and 1H-nuclear-magnetic resonance. Both crude and dialyzed levan completely inhibited the pancreatic cancer cell line at 100 ppm with no cytotoxicity on the normal retinal cell line. The LD50 of crude levan was 4833 mg/kg body weight. Levan had strong antioxidant activity and significantly reduced the expression of CXCR4 and MCM7 genes in pancreatic cancer cells with significant DNA fragmentation. In conclusion, Bacillus subtilis MT453867 levan is a promising adjunct to pancreatic-anticancer agents with both anti-cancer and chemoprotective effects.

Biological applications study of bio-nanocomposites based on chitosan/TiO2 nanoparticles polymeric films modified by oleic acid.

The aim of the present study was to prepare and characterize nanocomposite films to improve the treatment of skin wounds by applying the film as a bandage. To modify chitosan (Cs) and to prepare nanocomposites, a mixture between titanium dioxide nanoparticles (TiO2 NPs) was performed at different concentrations (2, 5, 10 and 15 wt%) and oleic acid (OA). The thin nanocomposite films were prepared by using casting method. The prepared films (Cs, Cs/TiO2 NPs, Cs/OA and Cs/OA/TiO2 NPs) were described by water absorption (swelling study) and biological degradation. Physico-chemical characterizations of Cs, Cs/OA, Cs/TiO2 NPs and Cs/OA/TiO2 NPs (with only 15 wt% TiO2 NPs) films were determined by X-ray diffraction, transmission high-resolution electron microscopy, field emission scanning electron microscopy, thermal analysis and Fourier transform infrared spectroscopy as well as their mechanical properties. Antimicrobial activity against microorganisms has been studied to assess activity against bacteria. The prepared nanocomposite films showed good antimicrobial activity for both Gram-positive and Gram-negative bacteria. The therapeutic effects of Cs-TiO2 NPs-oleic acid nanocomposites on healing excision wounds were studied in rat animal model. The data obtained revealed that groups treated with nanocomposites showed enhancement wound closure and speed up wound healing time.

Microbiological and environmental assessment of human oral dental plaque isolates.

Plaque-related diseases are amongst the most common ailments of the oral cavity. Streptococcus mutans is the causal agent of dental caries in animals and humans and is responsible for the formation and accumulation of plaques. This study aimed to identify and evaluate the role of the dental plaque isolates and its surrounding environment in plaque formation or inhibition. The study started with the identification of human dental plaque isolates from high caries index patients based on 16S rRNA and Mitis salivarius bacitracin agar (MSB) was used for S. mutans growing. Unexpectedly, the Streptococcus mutans was completely absent. The disc diffusion assay recorded that all the isolates had antimicrobial activity against the S. mutans growth. Enzymes assay revealed that the isolates produced dextransucrase, levansucrase and levanase activity with wide variation degrees. Also, the lactic acid production assay was done based in pH shift assessment. The highest pH shift and dextran yield were detected by the isolates Bacillus subtilis_AG1 and Bacillus mojavensis_AG3. The adherence test revealed that Lysinibacillus cresolivorans_W2 (MK411028) recorded the highest adhesion property (60%). Oligo- and polysaccharides were synthesized by the action of dextransucrase enzyme and their cytotoxicity tests were negative. Dextran with a molecular weight (117521 Da) recorded the highest antimicrobial efficacy against Bacillus subtilis_AG1 and Bacillusmojavensis_AG3 (65%, 63.5%) respectively. The results concluded that the dextran was the most important factor causing the dental plaque pathogenicity. Also, oral oligo- and polysaccharides might play a role in dental plaque control.

Enzymatic synthesis using immobilized Enterococcus faecalis Esawy dextransucrase and some applied studies.

Dextrans enzymatic synthesis by immobilized Enterococcus faecalis Esawy dextransucrase was studied. Different parameters, such as: enzyme protein concentration (EPC), substrate concentration (SC), temperature and reaction time were evaluated. EPC played a fundamental role in controlling dextran molecular size with 0.1% dextran in reaction mixture. Dextran 38,397 and 125,471Da were yielded at EPC 4.78 and 5.78mg, respectively. Proper dextrans (73,378 and 117,521Da) demanded in pharmaceutical applications were achieved at 6% and 12% sucrose concentrations and at 4.78 and 5.78mg EPC, respectively. Optimum temperature for conversion of glucose to dextran was 30°C (73% and 80% at 5.78 and 4.78mg EPC, respectively). Varieties of maltooligosaccharides (MOS) were yielded by synergistic cooperation between sucrose and maltose. Six MOS and three dextrans samples in vitro have prebiotic effect on Lactobacillus casei with degree of variation. Two samples of MOS with different degree of polymerization (DP) and three samples of dextran with different molecular weight (MW) reported different fibrinolytic activity.

Covalent immobilization of Enterococcus faecalis Esawy dextransucrase and dextran synthesis.

Enterococcus faecalis Esawy dextransucrase was immobilized in Fe(3+)-cross-linked alginate/carboxymethyl cellulose (AC) beads. The gel beads were modified with polyethylenimine (PEI) followed by glutaraldehyde (GA) to form Fe(3+) (ACPG) beads. Fe(3+) (ACPG) was characterized using FTIR and DSC techniques. GA activated beads showed new two peaks. The first was at 1,717 cm(-1) which refers to (CO) group of a free aldehyde end of glutaraldehyde, and another peak was at 1,660 cm(-1) referring to (CN) group. The immobilization process improved the optimum temperature from 35 to 45°C. The immobilized enzyme showed its optimum activity in wide pH range (4.5-5.4) compared to pH 5.4 in case of free form. Also, the immobilization process improved the thermal and pH enzyme stability to great extent. Reusability test proved that the enzyme activity retained 60% after 15 batch reactions. Immobilized enzyme was applied successfully in the synthesis of oligosaccharides and different molecular weights of dextran.

Evaluation of free and immobilized Aspergillus niger NRC1ami pectinase applicable in industrial processes.

The Aspergillus niger NRC1ami pectinase was evaluated according to its hydrolysis efficiency of dry untreated orange peels (UOP), HCl-treated orange peels and NaOH-treated orange peels (HOP and NOP). Pectinase was entrapped in polyvinyl alcohol (PVA) sponge and the optimum pH and temperature of the free and immobilized enzymes were shifted from 4, 40 °C to 6, 50 °C respectively. The study of pH stability of free and immobilized pectinase showed that the immobilization process protected the enzyme strongly from severe alkaline pHs. The immobilization process improved the enzyme thermal stability to great instant. The unique feature of the immobilization process is its ability to solve the orange juice haze problem completely. Immobilized enzyme was reused 12 times in orange juice clarification with 9% activity loss from the original activity. Maximum reaction rate (V(max)) and Michaelis-Menten constant (K(m)) of the partially purified form were significantly changed after immobilization.