Mounir M. Salem-Bekhit

Sponge-Like: A New Protocol for Preparing Bacterial Ghosts

Bacterial Ghosts (BGs) received an increasing interest in the recent years for their promising medicinal and pharmaceutical applications. In this study, for the first time we introduce a new protocol for BGs production. E. coli BL21 (DE3) pLysS (Promega) was used as a model to establish a general protocol for BGs preparation. The protocol is based on using active chemical compounds in concentrations less than the Minimum Inhibition Concentration (MIC). Those chemical compounds are SDS, NaOH, and H2O2. Plackett-Burman experimental design was used to map the best conditions for BGs production. Normal and electronic microscopes were used to evaluate the BGs quality (BGQ). Spectrophotometer was used to evaluate the amount of the released protein and DNA. Agarose gel electrophoresis was used to determine the existence of any residue of DNA after each BGs preparation. Viable cells, which existed after running this protocol, were subjected to lysis by inducing the lysozyme gene carried on pLysS plasmid. This protocol is able to produce BGs that can be used in different biotechnological applications.

Plackett–Burman randomization method for Bacterial Ghosts preparation form E. coli JM109

Plackett-Burman randomization method is a conventional tool for variables randomization aiming at optimization. Bacterial Ghosts (BGs) preparation has been recently established using methods other than the E lysis gene. The protocol has been based mainly on using critical concentrations from chemical compounds able to convert viable cells to BGs. The Minimum Inhibition Concentration (MIC) and the Minimum Growth Concentration (MGC) were the main guide for the BGs preparation. In this study, Escherichia coli JM109 DEC has been used to produce the BGs following the original protocol. The study contained a detail protocol for BGs preparation that could be used as a guide.

A potential role of Lactobacillus acidophilus LA1 and its exopolysaccharides on cancer cells in male albino mice

The aim of the present study was to evaluate the potential role of Lactobacillus acidophilus (LA1), isolated from faecal samples of breast-fed Egyptian infants (15–90 days), and its exopolysaccharides (EPS), with regard to antitumour activity in vivo against Ehrlich ascites carcinoma (EAC) cells. L. acidophilus demonstrated antioxidant properties by suppression of malondialdehyde and nitric oxide serum levels. Also, the EPS showed a powerful antitumour effect based on the obtained results from liver function tests, such as aspartate aminotransferase (AST), alanine transaminase (ALT) and albumin concentration than positive control and toward the normal values, when compared with a positive control. The kidney function of the treated and non-treated groups was not affected and there were no significant differences between the negative control and the treated groups. There was a marked suppression of lactate dehydrogenase (LDH) and alkaline phosphatase (ALP) enzymes in groups treated with LA1 and its EPS.

Challenge of Moringa peregrina Forssk as an antimicrobial agent against multi-drug-resistant Salmonella sp.

ABSTRACT The emergence of multi-drug-resistant (MDR) pathogenic bacteria is considered as a global problem. The aim of this study was to evaluate the antimicrobial inhibitory effects of the oily aqueous extract of Moringa peregrina Forssk against MDR clinical Salmonella enterica isolates. Four MDR S. enterica isolates were proved to have a gene mutation in amino acids codon 83 and 87 of gyrA and 67, 76 and 80 of parC gene by polymerase chain reaction (PCR) amplification and sequencing. The active components of M. pregrina extract were purified using GLC and TLC techniques and by using IR, NMR and mass spectra. The M. peregrina Forssk extract effect on bacterial cells was determined using scanning and transmission electron microscopies. Results demonstrated that M. peregrina Forssk have an excellent inhibitory effect against 34 MDR S. enterica isolates with different minimum inhibitory concentration (MIC) (109.37–437.5 mg/mL). The active component was identified as oleic acid-3 hydroxy propyl ester. The main abnormalities of Salmonella cells were observed – destruction in the cell wall that led to a reduction of protoplast besides, disruption of cytoplasmic membranes and, consequently, loss in their metabolic functions and death. This is the first report that deeply highlights the antimicrobial activity of M. peregrina Forssk against MDR clinical S. enterica isolates.

Antioxidant lipoxygenase inhibitors from the leaf extracts of Simmondsia chinensis

OBJECTIVE To isolate and identify chemical constituents with antioxidant and lipoxygenase inhibitory effects of the ethanolic extract of Simmondsia chinensis (Jojoba) leaves. METHODS The alcoholic extract was subjected to successive solvent fractionation. The antioxidant active fractions (chloroform, ethyl acetate and aqueous fractions) were subjected to a combination of different chromatographic techniques guided by the antioxidant assay with DPPH. The structures of the isolated compounds were elucidated on the basis of spectroscopic evidences and correlated with known compounds. The antioxidant activity was assessed quantitively using DPPH and β-carotene methods. The inhibitory potential against enzyme lipoxygenase was assessed on soybean lipoxygenase enzyme. RESULTS Ten flavonoids and four lignans were isolated. Flavonoid aglycones showed stronger antioxidant and lipoxygenase inhibitory effects than their glycosides. Lignoid glycosides showed moderate to weak antioxidant and lipoxygenase inhibitory effects. CONCLUSIONS A total of 14 compounds were isolated and identified from Simmondsia chinensis; 12 of them were isolated for the first time. This is the first report that highlights deeply on the phenolic content of jojoba and their potential biological activities and shows the importance of this plant as a good source of phenolics in particular the flavonoid content.

Antimicrobial evaluation of novel buccoadhesive films containing Myrrh extract

This study describes the formulation of new buccoadhesive films containing Myrrh extract as a natural and safe antimicrobial agent to give a long local medication. Buccoadhesive films of Myrrh extract were developed using solvent evaporation technique. The fabricated Myrrh extract films were evaluated for their palatability, weight, surface pH, and their content uniformity. In addition, tensile strength and the percentage of elongation of the films as well as in vitro mucoadhesive time were determined. Also, the films were tested for their antimicrobial activities. Obtained results showed that the tested films are elegant in appearance and have neutral pH, palatable by volunteers and have good tensile strength and elasticity. They also exhibited antibacterial activities against different types of bacterial strains as well as antifungal activity against Candida albicans.

YIELD PRODUCTION OF RECOMBINANT PLASMID DNA WITH ESCHERICHIA COLI IN FED-BATCH CULTURE BY PSEUDO-EXPONENTIAL FEEDING

ABSTRACT For preventing and treating debilitating diseases, plasmid DNA [pDNA] is emerging as the vector of choice for gene therapy and DNA vaccination. In this study, the development of a new implementation of control strategies for high-cell-density (HCD) culture fermentation of Escherichia coli was described to produce high level of cell mass for the mass production of bacterial plasmid. A yield of 2 g of pDNA/L was achieved by application of the newly developed techniques for HCD culture. We were also able to reduce the fermentation time for the highest production to only 3 h rather than 72 h. Furthermore, application of the HCD culture technique permits to keep the organism at a low growth rate to ensure high quality of the produced plasmid. The kinetics of batch and fed-batch cultures of E. coli producing plasmid was investigated. The cell lag time, the maximum specific growth rate and (Y=D/U) were determined as 2 h., 1.7 h−1 and 1.61 g·g−1, respectively. In the fed-batch culture, different specific growth rates were set at 2.5 h−1, 3.2 h−1, and 4.5 h−1 by pseudo-exponential feeding, and the expressions for the specific rate of substrate consumption, the growth kinetics and the product formation kinetics of each phase were obtained. The result shows that the concentrations of cell and plasmid can reach 93.0 g·L−1 and 2.0 g·L−1 respectively, and the quality of the DNA vaccine met the requirements for medical use.

Novel chlorhexidine dermal patches, preparation characterization and antimicrobial evaluation

The purpose of this work is to develop chlorhexidine dermal patches using Eudragit® RL100 as the patch forming polymer. Solvent casting technique was employed in the preparation of the patches. The prepared patches, the corresponding physical mixtures, and the individual ingredients were physicochemically characterized by differential scanning calorimetry (DSC) and Fourier transform infrared spectroscopy (FTIR) to investigate drug-excipient interactions. The results of the incompatibility studies indicated that the drug was molecularly distributed in Eudragit RL100 matrix. The developed patches were evaluated in terms of tensile strength, percentage of elongation, patch weight, surface pH, content uniformity, and drug dissolution profile. The developed chlorhexidine-medicated patches were flexible, transparent, homogenous, elegant, and smooth. Evaluation of the antimicrobial activity of the developed patches showed effective antimicrobial action against varieties of microorganisms.

A novel protocol for bacterial ghosts’ preparation using tween 80

Bacterial ghosts (BGs) can be prepared by both genetic and chemical means. Genetic method include using lysis gene E. Chemical method include incubation with numerous agents for a short time at their minimum inhibitory or minimum growth concentrations (MIC or MGC). The aim of this study is to prepare the BGs with a new protocol via exposing the bacterial cells to tween 80 for an extended period of time followed by sudden reduction of the surrounding pH. Salmonella enterica serovar typhimurium ATCC 13311 was used for this purpose. The cells were incubated in 7% v/v tween 80 solution in Muller-Hinton broth for 24 h at 37 °C then pH was decreased to 3.6 by adding lactic acid for one hour. The bacterial pellets were separated by high speed centrifugation, and then washed three times by half normal saline solution. High quality BGs were visualized by scanning electron microscopy (SEM) revealing punctured cells with intact outer shells and at least one intramembranous tunnel. The absence of vital cells was confirmed by subculturing. The release of respective amounts of proteins and DNA is another evidence of ghost’s production. In addition, the integrity of cells was proved by visualization of Gram-stained cells using light microscopy. In conclusion, this new protocol is simple, economic and feasible for BGs preparation.