Catalytic and anti-cancer properties of platinum, gold, silver, and bimetallic Au-Ag nanoparticles synthesized by Bacillus sp. bacteria.

Metallic nanoparticles play a significant role in the catalysis of chemical processes, besides, bimetallic nanoparticles with abundant active sites can reduce metallic nanoparticles toxicity in addition to increasing their catalytic performances. In this work, the platinum, gold, and silver nanoparticles are bio-synthesized using a native bacterium (GFCr-4). Also, the Au-Ag and Au@Ag bimetallic nanoparticles with alloy and core-shell structures, respectively, are biologically synthesized. To improve the synthesis, the effects of various factors like pH, temperature, electron donor, and ionic liquids were investigated. The as-synthesized nanoparticles were characterized with different techniques. The microscope images and dynamic light scattering (DLS) analysis confirm the uniform distribution of as-synthesized nanoparticles with average sizes of 25, 30, 47, 77, and 86 nm obtained for Ag, Au, Pt, Au-Ag alloy, and Au@Ag core-shell, respectively. The catalytic performances of as-synthesized nanoparticles were investigated. The Au-Ag alloy nanoparticles exhibit better catalytic performance than the as-synthesized metallic Au nanoparticles, according to the Gewald reaction. According to the photocatalytic study, the yield can be increased by up to 92% by using PtNPs in the presence of a green LED. Additionally, for the first time, PtNPs were utilized as an effective catalyst in a peroxyoxalate chemiluminescence (POCL) system in the presence of nuclear fast red (NFR) as a novel fluorophore. In addition, the results of the MTT (3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide) assay revealed that the synthesized eco-friendly nanoparticles have a low effect on the lethality of 3T3 normal cells whereas MCF-7 cancer cells were inhibited up to 77.3% after treatment by PtNPs nanoparticles.

Incorporation of white tea extract in nano-liposomes: optimization, characterization, and stability.

BACKGROUND: In the present study, an extraction method affected by sonication intensity (40%, 70% and 100%), sonication time (5, 10 and 15 min) and different solvents (ethanol, methanol and a combination of ethanol/methanol) was optimized to extract the white tea with the greatest polyphenolic compounds using a response surface methodology. To prepare the nano-liposomal vesicles, phospholipids and cholesterol in various proportions (60:0, 40:20, 30:30 and 20:40) were applied based on thin-film hydration and ultrasound method. The nano-capsules enriched in bioactive compounds were examined through particle characteristics, encapsulation efficiency, morphological analysis, thermal properties and Fourier transform infrared spectroscopy. RESULTS: The observations showed that the extraction yield highly depended on the type of solvent with varying permeability, sonication time and power. The highest total phenolic content (68.38 mg GA g-1 ) and free radical scavenging activity (77.65%) were observed for the following optimal conditions: 70% for sonication intensity, 15 min for sonication time and methanol as solvent. Characteristics of nanoliposomes within a compositional ratio of lecithin/cholesterol (40:20) and with a zeta potential of -56 ± 0.01 mV, as well as white tea extract (WTE) samples with an average particle diameter of 82.20 ± 0.08, microencapsulation efficiency of 76.5% ± 0.081, polydispersity index of 0.06 ± 0.02 and span value of 0.69 ± 0.03. are used as the optimal formulation for microencapsulation of antioxidant WTE. The results demonstrated an increment in thermal stability of liposomal WTE samples compared to other samples. CONCLUSION: The findings of the present study indicated that nano-liposomes comprise an effective technology for coating the WTE, as well as to increasing its stability and thermal properties. © 2021 Society of Chemical Industry.

CNT-based nanocarrier loaded with pyrimethamine for adipose mesenchymal stem cells differentiation and cancer treatment: The computational and experimental methods.

Pyrimethamine is an effective drug in the cancer cell treatment and is a dihydrofolate reductase inhibitor. In this work, the amount of drug loading up on CNT and its cytotoxicity effect upon MCF-7 cell lines was surveyed. The novel applications of some drugs and nanocarriers can induce the differentiation of adipose mesenchymal cells into nerve cells. Hence carbon nanotube-pyrimethamine was used to differentiate mesenchymal stem cells into the neural category, for the first time. The results of NSE and NFM gene expression level were evaluated using the real-time PCR. A detailed study on the interaction between pyrimethamine anticancer drug and (6, 0) zigzag single-walled carbon nanotube was performed by DFT/B3LYP and DFT/M06-2X with 6-31G* basis set calculations in gas phase and in solvent using the PCM. Different configurations of the adsorbed pyrimethamine onto the CNT surface were studied. Based on the results, the process of pyrimethamine adsorption on diff ;erent sites on the outer wall of the nanotube was exothermic and configurations were stable. The adsorption energy values indicated that the pyrimethamine molecule could be physically adsorbed on the external surface of the SWCNT. The QTAIM was used for characterizing the nature of the interactions between the pyrimethamine and the selected nanocarrier.

Extracellular biosynthesis of magnetic iron oxide nanoparticles by Bacillus cereus strain HMH1: Characterization and in vitro cytotoxicity analysis on MCF-7 and 3T3 cell lines.

Discovery of new properties and special functionalities at the nanoscale materials caused nanotechnology to become one of the leading parts in all sciences namely biology and medicine. Magnetic iron oxide nanoparticles (MIONPs) are among interesting nanomaterials in biomedical arena, which have attracted the attention of many researchers owing to their extensive capabilities. Due to the simple, cost-effective and environmentally-friendly production processes, biosynthesis is of paramount importance between different methods of nanoparticles production. In the current study, we succeeded to synthesize MIONPs using a newly extracted bacteria supernatant. Produced nanoparticles were characterized using FE-SEM, DLS, VSM, UV-vis, FT-IR and EDS spectroscopy. Analysis showed that the average particle size of very stable spherical MIONPs is about 29.3 nm. The bacteria protein profile obtained by SDS-PAGE analysis indicated induction of different proteins. In vitro cytotoxicity of nanoparticles on the viability of MCF7 and 3T3 cell lines was assessed by MTT assay. The results show that toxicity of the produced nanoparticles (IC50, MCF-7 > 5 mg/ml and IC50, 3T3 > 7.5 mg/ml) follows a concentration dependent manner.

Catalytic activation of Bacillus laccase after temperature treatment: Structural & biochemical characterization.

Laccases belong to a family of multicopper oxidases that have strong oxidation ability towards phenolic compounds. Here, more detailed investigations were carried out on a Bacillus laccase with remarkable behavior of activation after thermal treatment. The kcat of the enzyme was increased 2.5 fold after 50 min incubation at 70 °C. Copper content determination revealed a molar copper to protein ratio of 3.2 in the both sample. The present paper concerns the differences which are induced in enzyme structure after thermal treatment using common biochemical methods Intrinsic fluorescence of the enzyme was increased after incubation at 70 °C indicating higher compactness of the structure in comparison to untreated molecules. Quenching analysis did not show any significant changes in flexibility of the enzyme structure. The local changes in secondary structures were also obvious by far-UV circular dichroism when non-incubated and incubated laccase were compared. Oligomerization studies of the enzyme using gas-phase electrophoretic mobility macromolecule analysis (GEMMA) did not prove any oligomerization.

Comamonas sp. halotolerant bacterium from industrial zone of Jovein of Sabzevar introduced as good candidate to remove industrial pollution.

BACKGROUND AND OBJECTIVES: Heavy metals are considered as high risk biocides due to their harmful effects on human health, the environment and other living organisms. Bacterial strains showing resistance to heavy metals has been used for removing such toxic materials from the environment. In this study we isolated and characterized a heavy metals-resistance halophilic bacterial strains from Kal shoor Jovein of Sabzevar, one of the industrial zone of Khorasan-e-Razavi province in Iran and has naturally saline oils. MATERIALS AND METHODS: Strain JC-66 is heavy metals-resistance halophilic bacterial strains isolated from Kal shoor Jovein of Sabzevar. The 16S rDNA gene was sequenced to identify this bacterium. The appropriate conditions for its potency to remove the lead were tested in various temprature, pH and agitation speed. The resistance mechanism of JC-66 to lead were investigated. RESULTS: JC-66 is a Comamonas sp. according to 16S rDNA sequence analysis. Based on minimum inhibitory concentration (MIC) results, the isolated strain has high resistance to the lead metal. The optimal condition for lead removal was exhibited in neutral medium (pH 7) incubation temperature 37 °C, and shaking rate of 180 rpm for JC-66. X-Ray Diffraction results also are indicative of adsorption mechanism to lead metal uptake. Plasmid extraction was performed to confirm the role of plasmids in bacterial resistance to lead. CONCLUSION: It can be concluded that the mechanism of resistance to heavy metals in the studied strain, is the result of an expression plasmid, and adsorption. It was concluded that JC-66 is able to be one of the best candidates to remove industrial pollution because it showed high resistance to lead.

An efficient in vitro refolding of recombinant bacterial laccase in Escherichia coli.

Laccases (benzenediol oxygen oxidoreductases, EC 1.10.3.2) are important multicopper enzymes that are used in many biotechnological processes. A recombinant form of laccase from Bacillus sp. HR03 was overexpressed in Escherichia coli BL-21(DE3). Inclusion body (IB) formation happens quite often during recombinant protein production. Hence, developing a protocol for efficient refolding of proteins from inclusion bodies to provide large amounts of active protein could be advantageous for structural and functional studies. Here, we have tried to find an efficient method of refolding for this bacterial enzyme. Solubilization of inclusion bodies was carried out in phosphate buffer pH 7, containing 8M urea and 4mM ß-mercaptoethanol and refolding was performed using the dilution method. The effect of different additives was investigated on the refolding procedure of denaturated laccase. Mix buffer (phosphate buffer and citrate buffer, 100mM) containing 4mM ZnSO4 and 100mM sorbitol was selected as an optimized refolding buffer. Also Kinetic parameters of soluble and refolded laccase were analyzed.

Extra EF hand unit (DX) mediated stabilization and calcium independency of α-amylase.

It is the common feature of α-amylases that calcium ion is required for their structural integrity and thermal stability. All amylases have at least one Ca(2+) per molecule; therefore amino acids involved in calcium binding are specific and conserved. In this study, sequence analysis revealed the presence of EF-hand-like motif in calcium-binding loop of Bacillus megaterium WHO (BMW)-amylase that was previously isolated from BMW. The EF-hand motif and its variants (EF-hand-like motif) are the most common calcium-binding motifs found in a large number of protein families. To investigate the effect of calcium ion on the thermal stability and activity of BMW-amylase, we used site-directed mutagenesis to replace histidine 58 with Asp (D), Ile (I), Tyr (Y), Phe (F), and Arg (R) at the seventh position of EF-hand-like motif. Upon the addition of an extra DX unit to the calcium-binding loop in H58D variant, thermal stability, catalytic activity, and chelating power of the enzyme improved due to higher affinity toward calcium. H58D variant demonstrated calcium independency compared to the wild type and other created mutants. Conformational changes in the presence and absence of Ca(2+) were monitored using fluorescence technique.

Enhancement of thermal stability of chondroitinase ABC I by site-directed mutagenesis: an insight from Ramachandran plot.

The application of chondroitinase ABC I (cABC I) in damaged nervous tissue is believed to prune glycosaminoglycan chains of proteoglycans, thereby facilitates axon regeneration. However, the utilization of cABC I as therapeutics is notably restricted due to its thermal instability. In the present study, we have explored the possibility of thermostabilization of cABC I through release of its conformational strain using Ramachandran plot information. In this regard, Gln140 with non-optimal φ and ψ values were replaced with Gly, Ala and Asn. The results indicated that Q140G and Q140A mutants were able to improve both activity and thermal stability of the enzyme while Q140N variant reduced the enzyme activity and destabilized it. Moreover, the two former variants displayed a remarkable resistance to trypsin degradation. Structural analysis of all mutants showed an increase in intrinsic fluorescence intensity and secondary structure content of Q140G and Q140A compared to the wild type which indicated more compact structure upon mutation. This investigation demonstrated that relief of conformational tension can be considered as a possible approach to increase the stability of the protein.

A novel quantum dot-laccase hybrid nanobiosensor for low level determination of dopamine.

This work reports a novel nanobiosensor based on a thioglycolic acid (TGA)-capped CdTe quantum dot-laccase (Lac) enzyme system for sensitive detection of dopamine (DA). The enzyme used catalyzes the oxidation of DA to dopamine-o-quinone (DOQ), which can selectively quench the strong luminescence of CdTe nanocrystals at neutral pH. The relationship between luminescence intensity of CdTe nanocrystals and DA concentration is nicely described by the Stern-Volmer equation. At an optimum pH of 7.4, the proposed sensor gives a linear calibration over a DA concentration range of 0.3 to 100 µM, with a limit of detection of 0.16 µM and a response time of 2 min. The relative standard deviation for seven replicate determinations of 6.0 µM of DA was found to be 3.7%. The sensor was successfully applied to the determination of DA in a blood plasma sample and in a DA injection formulation.

Introduction of a New Diagnostic Method for Breast Cancer Based on Fine Needle Aspiration (FNA) Test Data and Combining Intelligent Systems.

BACKGROUND: Accurate Diagnosis of Breast Cancer is of prime importance. Fine Needle Aspiration test or "FNA", which has been used for several years in Europe, is a simple, inexpensive, noninvasive and accurate technique for detecting breast cancer. Expending the suitable features of the Fine Needle Aspiration results is the most important diagnostic problem in early stages of breast cancer. In this study, we introduced a new algorithm that can detect breast cancer based on combining artificial intelligent system and Fine Needle Aspiration (FNA). METHODS: We studied the Features of Wisconsin Data Base Cancer which contained about 569 FNA test samples (212 patient samples (malignant) and 357 healthy samples (benign)). In this research, we combined Artificial Intelligence Approaches, such as Evolutionary Algorithm (EA) with Genetic Algorithm (GA), and also used Exact Classifier Systems (here by Fuzzy C-Means (FCM)) to separate malignant from benign samples. Furthermore, we examined artificial Neural Networks (NN) to identify the model and structure. This research proposed a new algorithm for an accurate diagnosis of breast cancer. RESULTS: According to Wisconsin Data Base Cancer (WDBC) data base, 62.75% of samples were benign, and 37.25% were malignant. After applying the proposed algorithm, we achieved high detection accuracy of about "96.579%" on 205 patients who were diagnosed as having breast cancer. It was found that the method had 93% sensitivity, 73% specialty, 65% positive predictive value, and 95% negative predictive value, respectively. If done by experts, Fine Needle Aspiration (FNA) can be a reliable replacement for open biopsy in palpable breast masses. Evaluation of FNA samples during aspiration can decrease insufficient samples. FNA can be the first line of diagnosis in women with breast masses, at least in deprived regions, and may increase health standards and clinical supervision of patients. CONCLUSION: Such a smart, economical, non-invasive, rapid and accurate system can be introduced as a useful diagnostic system for comprehensive treatment of breast cancer. Another advantage of this method is the possibility of diagnosing breast abnormalities. If done by experts, FNA can be a reliable replacement for open biopsy in palpable breast masses. Evaluation of FNA samples during aspiration can decrease insufficient samples.

Enhancement of a bacterial laccase thermostability through directed mutagenesis of a surface loop.

Laccases (benzenediol oxygen oxidoreductases, EC 1.10.3.2) are used in many biotechnological processes, including removal of polyphenols in beverages, decolorizing and detoxifying effluents, drug analysis and bioremediation. In the present work, we have tried to increase thermal stability of laccase from Bacillus HR03 using site directed point mutations. Glu188 was substituted with 2 positive (Lys and Arg) and one hydrophobic (Ala) residues. All mutations showed improved thermal stability. Thermal activation of laccase was also increased after introducing the mutations. Remarkably, the Glu188Lys variant showed 3-fold higher thermal activation and higher T(50) (5 °C) with respect to the native enzyme. Furthermore steady-state k(cat) and K(m) values were influenced despite the distance between the mutated position and the catalytic site. In Glu188Arg mutation, the k(cat) was improved 3-fold and K(m) reduced by 25%. Interestingly, all three variants showed higher stability against urea as a chemical denaturant. Structural analyses of the native and mutated variants were carried out using fluorescence and far-UV circular dichroism.

Enhanced expression of a recombinant bacterial laccase at low temperature and microaerobic conditions: purification and biochemical characterization.

Laccases (benzenediol oxygen oxidoreductase; EC 1.10.3.2) have many biotechnological applications because of their oxidation ability towards a wide range of phenolic compounds. Within recent years, researchers have been highly interested in the identification and characterization of laccases from bacterial sources. In this study, we have isolated and cloned a gene encoding laccase (CotA) from Bacillus sp. HR03 and then expressed it under microaerobic conditions and decreased temperature in order to obtain high amounts of soluble protein. The laccase was purified and its biochemical properties were investigated using three common laccase substrates, 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) (ABTS), syringaldazine (SGZ) and 2,6-dimethoxyphenol (2,6-DMP). K(M) and k(cat) were calculated 535 microM and 127 s(-1) for ABTS, 53 microM and 3 s(-1) for 2, 6-DMP and 5 microM and 20 s(-1) for SGZ when the whole reactions were carried out at room temperature. Laccase activity was also studied when the enzyme was preincubated at 70 and 80 degrees C. With SGZ as the substrate, the activity was increased three-fold after 50 min preincubation at 70 degrees C and 2.4-fold after 10 min preincubation at 80 degrees C. Preincubation of the enzyme in 70 degrees C for 30 min raised the activity four-fold with ABTS as the substrate. Also, L-dopa was used as a substrate. The enzyme was able to oxidize L-dopa with the K(M) and k(cat) of 1,493 microM and 194 s(-1), respectively.

Purification and characterization of a thermostable phytate resistant alpha-amylase from Geobacillus sp. LH8.

A thermophilic and amylolytic bacterium (LH8) was isolated from the hot spring of Larijan in Iran at 65 degrees C. Identification of strain LH8 by 16S rDNA sequence analysis showed that LH8 strain belongs to the Geobacillus sp. with 99% sequence similarity with the 16S rDNA of Geobacillus thermodenitrificans. A new alpha-amylase (GA) was extracted from this strain and purified by ion-exchange chromatography. SDS-PAGE showed a single band with an apparent molecular mass of 52kDa. The optimum temperature and pH were 80 degrees C and 5-7, respectively. In the presence of Mn2+, Ca2+, K+, Cr3+ and Al3+, the enzyme activity was stimulated while Mg2+, Ba2+, Ni2+, Zn2+, Fe3+, Cu2+ and EDTA reduced the activity. The K(m) and V(max) values for starch were 3 mg ml(-1) and 6.5 micromol min(-1), respectively. The gene encoding alpha-amylase was isolated and the amino acid sequence was deduced. Comparison of GA and other alpha-amylase amino acid sequences suggested that GA has conserved regions that were previously identified in alpha-amylase family but GA exhibited some substitutions in the sequence. Its phytate resistant is an important property of this enzyme. 5 and 10 mM phytic acid did not inhibit this enzyme. Therefore, features of phytate resistant alpha-amylase from Geobacillus sp. LH8 are discussed.