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We aimed to develop a simple yet novel method to prepare plasmid DNA-loaded nanoliposomes for cancer gene therapy. Murine interleukin-12 (mIL-12) pDNA-loaded nanoliposomes were prepared via novel freeze-drying of a monophase solution method. The physicochemical characteristics, cytotoxicity, and transfection efficiency of the prepared nanoliposomes in murine CT-26 colon carcinoma cells were evaluated. Furthermore, tumor progression and survival rate in CT-26 colon carcinoma-bearing BALB/c mice subsequent to direct intratumoral injections were investigated over a period of 40 days. Using this preparation method, nanoliposomes with particle size of around 300 nm and zeta potential of 96.5 mV were obtained. The transmission electron microscope results showed that the liposomes were nano-sized and almost spherical. The agarose gel retardation assay revealed the pDNA encapsulation in the nanoliposomes. The nanoliposomes with 72.4% encapsulation efficiency and low cell toxicity could significantly improve mIL-12 expression by approximately 25-fold relative to the naked mIL-12 pDNA. There was a significant tumor growth inhibition after repeated injections of mIL-12 pDNA-loaded nanoliposomes. This is the first study on the freeze-drying of a monophase solution method as a simple yet novel technique for the preparation of pDNA-loaded nanoliposomes. Given the ease of preparation method and promising in vitro and in vivo characteristics, this investigation demonstrates advances in pDNA lipid formulation for cancer gene therapy. [ABSTRACT FROM AUTHOR]

Protease is a kind of enzyme with different applications in medicine and industry. The limitation of the enzymes in industry is lack of tolerance for harsh conditions (high temperature, pH, and low water condition). This study intends to evaluate in silico study and screen the alkaline serine protease producing bacteria from honey and optimize the production of low water active alkaline serine protease. Plackett–Burman method was applied to improve effective factors on protease production, including pH, inoculum size, temperature, time, soybean protein, KH2PO4, MgSO4, CaCl2, NaCl, Glycerin and glucose concentration. Further, the Box–Behnken method was used to optimize the significant level of selected factors. Besides, in silico study was utilized to create the enzyme structure and investigate the stability in harsh conditions. The results showed that the highest protease production belongs to Bacillus sp. Khoz1 closed Bacillus safensis, Also temperature, glucose and soybean protein concentration were three significant factors for protease production and the optimized level for them were 35 °C, 0.5 g/l, and 38.32 g/l, respectively. The microorganism was able to produce protease until 98.36 U/ml and it was 98.68% similar to B. safensis. The stability of microbial alkaline serine protease was also determined in high pH and temperature conditions. The best stable condition for isolated protease was pH 9 and temperature at 50 °C. The in silico data showed that this protease has similar 3D structure to Bacillus subtilis Subtilisin E and highly charge amino acids on its surface were that caused this enzyme still activate and stable in low water condition. [ABSTRACT FROM AUTHOR]

This study aimed to screen alpha-amylase producing microorganisms from honey as a low water activity medium, a suitable source for selecting stable and cost-beneficial bacterial enzyme production systems. Plackett–Burman method was used to select twelve effective factors including pH, inoculum size, temperature, time, corn starch, KH2PO4, peptone, MgSO4, CaCl2, NaCl, glycerin, and yeast extract concentrations on bacterial alpha-amylases production yield. The Box–Behnken method was utilized to optimize the level of selected significant factors. The stability of bacterial alpha-amylases was also determined in low pH and high-temperature conditions. In addition, in silico study was used to create the alpha-amylase structure and study the stability in high-temperature and low water available condition. Among all isolated and characterized microorganisms, Bacillus megaterium produced the highest amount of alpha-amylases. The in silico data showed the enzyme 3D structure similarity to alpha-amylase from Halothermothrix orenii and highly negative charge amino acids on its surface caused the enzyme activity and stability in low water conditions. Based on Box–Behnken results, the temperature 35 °C, pH 6 and starch 40 g/l were determined as the optimum level of significant factors to achieve the highest alpha-amylases unit (101.44 U/ml). This bacterial alpha-amylases enzyme showed stability at pH 5 and a range of temperatures from 40 to 60 °C that indicates this enzyme may possess the potential for using in industrial processes. [ABSTRACT FROM AUTHOR]

Synthesis and application of layered nanomaterials are known as emerging field in nanotechnology. Inorganic layered nanomaterials with nanometer scale, high aspect ratios, and large-surface area generate various scientific and technological interests in several potential areas of application such as separation technology, medical sciences, chromatography drug delivery, and catalysis. In this research, naproxen was opted as a unique model of the drug to intercalate into three-dimensional layered double hydroxide (LDH) nanostructures by calcination–reconstruction method. The designed-naproxen nanostructures were synthesized and used as novel drug nanocarriers. The synthesized nanomaterial characteristics were confirmed by FTIR, XRD, and SEM. The results of antibacterial activity indicated that homemade nanostructure can inhibit bacterial growth. Furthermore, MTT assay analysis showed that synthesized-nanostructure in physiological concentration has not cytotoxicity on C2C12 myoblast cells in vitro. Subsequent to intercalation with naproxen according to calcination–reconstruction method, the basal spacing of LDH increased to 2.62 nm from primary 0.77 nm in diameter, which confirms successful intercalation of naproxen into inserted layer of LDH by bridging interaction. Also, in-vitro drug release tests in PBS (pH 7.4) showed constant release profile for naproxen. Consequently, due to the facile and low-cost fabrication of three-dimensional LDH nanostructure, it can be considered as a potential alternative for traditional and high cost drug delivery. [ABSTRACT FROM AUTHOR]

The goal of this study is to investigate the antibacterial activity and sustained release of Centella Asiatica (CA) extract loaded in gelatin-based hydrogels, reinforced with MIL-53(Fe) (Fe-MOF) for the potential use as wound dressing. To induce antibacterial activity to the designed wound dressing, CA-extract was incorporated within the nanocomposite hydrogels. FT-IR analysis showed the distinctive peaks related to the Fe-MOF and CA-extract, with little shift, for hydrogel samples, which confirm the successful synthesis of nanocomposite hydrogels containing Fe-MOF and CA-extract. GC-mass analysis showed the antibacterial potential for CA-extract because of the presence of phenol, caryophyllene, and propanoic acid. SEM micrograph demonstrates the presence of Fe-MOF enhancing the porous of hydrogels. The antibacterial activity of prepared nanocomposite hydrogels was investigated by disk diffusion method revealing the significant inhibition zone growth for Basillus, Staphylococcus, and Streptococcus, as gram-positive and E. coli, Klebsiella, and P. aeruginosa as gram-negative bacteria and the "Candida albicans as fungi. Also, the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were determined by the micro-broth dilution method. The results showed that the nanocomposite hydrogel has a MIC values of 3.125 mg/ml and 6.25 mg/ml against both groups of bacteria. Cytotoxicity results obtained from MTT tests showed that the prepared hydrogels are biocompatible. The immunohistochemistry-stained hydrogel images revealed that the growth of cells on the CA-extract containing hydrogels was significantly increased in comparison to neat hydrogels.Graphical abstract: The goal of this study is to investigate the antibacterial activity and sustained release of Centella Asiatica (CA) extract loaded in gelatin-based hydrogels, reinforced with MIL-53(Fe) (Fe-MOF) for the potential use as wound dressing. To induce antibacterial activity to the designed wound dressing, CA-extract was incorporated within the nanocomposite hydrogels. FT-IR analysis showed the distinctive peaks related to the Fe-MOF and CA-extract, with little shift, for hydrogel samples, which confirm the successful synthesis of nanocomposite hydrogels containing Fe-MOF and CA-extract. GC-mass analysis showed the antibacterial potential for CA-extract because of the presence of phenol, caryophyllene, and propanoic acid. SEM micrograph demonstrates the presence of Fe-MOF enhancing the porous of hydrogels. The antibacterial activity of prepared nanocomposite hydrogels was investigated by disk diffusion method revealing the significant inhibition zone growth for Basillus, Staphylococcus, and Streptococcus, as gram-positive and E. coli, Klebsiella, and P. aeruginosa as gram-negative bacteria and the "Candida albicans as fungi. Also, the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were determined by the micro-broth dilution method. The results showed that the nanocomposite hydrogel has a MIC values of 3.125 mg/ml and 6.25 mg/ml against both groups of bacteria. Cytotoxicity results obtained from MTT tests showed that the prepared hydrogels are biocompatible. The immunohistochemistry-stained hydrogel images revealed that the growth of cells on the CA-extract containing hydrogels was significantly increased in comparison to neat hydrogels. [ABSTRACT FROM AUTHOR]