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Pseudomonas aeruginosa is an opportunistic human pathogen that can cause life-threatening nosocomial infections. The alarming increase in antibiotic resistance has led to an urgent need for alternative therapeutic approaches, such as phage therapy, which has shown promising results in many studies. In this study, P121, a new lytic Pseudomonas phage, was isolated and characterized. Whole-genome sequencing showed that it has a genome of 73,001 bp that contains 91 predicted coding sequences. No genes involved in virulence or lysogeny were found in the genome, thus making it potentially safe for therapeutic applications. Genomic and phylogenetic analysis indicated that P121 is a member of the genus Litunavirus, family Schitoviridae. The present study provides some basic information for further research on treatment of P. aeruginosa infections.

Abstract Aims: Salmonellosis is a critical, common infectious disease afflicting people and animals alike, that is caused by Salmonellabacteria. With bacteria showing resistance to antibiotics, new methods should be introduced to prevent and treat this disease. To that end, bacteriophages are promising choices. Methods and Results: In this study, phage vB_SenS_TUMS_E19 (E19) was isolated from poultry wastewater against Salmonella enteritidis. Phage characteristics, including plaque formation, transmission electron microscopy, structural protein profile, host range, growth curve, pH and temperature stability were determined. The extracted genome was sequenced, and annotated using standard bioinformatics tools. Average Nucleotide Identity (ANI) and phylogeny were used to compare the phage genome with other Salmonella phages. Following morphological analysis, phage E19 turned out to belong to the Siphoviridae family. It worked on various strains of Salmonella but had no effect on bacteria of other genera. The burst size was approximately 156 plaque-forming units per cell (PFU per cell) and phage was stable at different temperatures as well as pH values. The genome of E19 is a linear dsDNA molecule of 42,813 bp whose G+C content stands at 49.8%. Out of 62 detected putative protein-coding genes, only 44 gene products with known functions were found in database searches. Neither tRNA genes nor genes associated with antibiotic resistance, virulence factor, and lysogeny formation were detected in the genome. Conclusion: The E19 showed lytic activity against some Salmonella enterica serovars and Salmonella pullorum. Genome-based studies exhibited the absence of antibiotic resistance and virulence genes, indicating the safety of this phage for environmental and clinical applications. Significance and Impact of the Study: The findings indicate that the lytic polyvalent E19 is an antibacterial agent that could control Salmonella in food samples and helps with Salmonellosis prevention and treatment.

The present investigation reports an in-vitro study using combination of laccase and an enhancer capable of inhibiting the growth of pathogenic microorganisms, preventing biofilm formation, and whitening teeth. Laccase-cinnamic acid system remarkably inhibited the growth of Aggregatibacter actinomycetemcomitans, Candida albicans, S. aureus, and Streptococcus mutans whilst showed no significant effects on Gram-negative bacteria. Data presented that cinnamic acid (10 mM) with laccase (0.125 U ml−1) led to a maximum decrease of about 90%, in S. mutans biofilm formation. The confocal laser scanning microscopy showed considerable detachment of S. mutans cells from glass substratum. The combined laccase-cinnamic acid system could remove teeth discoloration caused by coffee. SEM of the teeth surface exhibited no damages such as surface cracking or fracture. Liquid chromatography-tandem mass spectrometry (LC-MS) and cyclic voltammetry (CV) studies showed that laccase can catalyze the one-electron oxidation of cinnamic acid to the respective radical. This radical can then undergo several fates, including recombination with another radical to form a dimeric species, dismutation of the radical back to cinnamic acid or decarboxylation to give various reduced oxygen species. Therefore, the redox potential values of phenolic monomers/oligomers are related with their biological activities.

Maryam Zohri,1 Hamid Akbari Javar,2 Taraneh Gazori,3 Mohammad Reza Khoshayand,4 Seyed Hamid Aghaee-Bakhtiari,5,6 Mohammad Hossein Ghahremani1,7 1Nanotechnology Research Center, Faculty of Pharmacy, Tehran University of Medical Sciences, Tehran, Iran; 2Departments of Pharmaceutics, Faculty of Pharmacy, Tehran University of Medical Sciences, Tehran, Iran; 3Research and Development Department, Trita Nano Pharmaceutical Research Center, Tehran, Iran; 4Department of Drug and Food Control, Faculty of Pharmacy, Tehran University of Medical Sciences, Tehran, Iran; 5Bioinformatics Research Group, Mashhad University of Medical Sciences, Mashhad, Iran; 6Department of Medical Biotechnology, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran; 7Department of Pharmacology and Toxicology, Faculty of Pharmacy, Tehran University of Medical Sciences, Tehran, IranCorrespondence: Mohammad Hossein Ghahremani Tel/Fax +9866959102Email mhghahremani@tums.ac.irPurpose: In this study, chitosan/alginate nanoparticles are prospected as a carrier for controlled release of recombinant human bone morphogenetic protein-2 (rhBMP-2).Materials and Methods: The rhBMP-2-loaded chitosan/alginate nanoparticles (Cs/Alg/B NPs) were prepared using the ionic gelation (IG) method. The current research was conducted to optimize the effective factors for entrapping rhBMP-2 in Cs/Alg NPs using response surface methodology (RSM) and the Box–Behnken design (BBD). The variables were the Cs/Alg molecular weight (Mw) ratios (1– 3), pH (4.8– 5.5), stirring rates (900– 1300 rpm) and the responses included size, ζ-potential, polydispersity index (PDI), loading efficacy (LE), cumulative release (CR), and morphological degradation time (MDE). Then, the morphological properties of optimum formulation were studied for post-characterization. In the next step, the MTT assay for the optimized run was done for 24 and 48 hours.Results: The results revealed that the optimum conditions for the mentioned variables were stirring rate=1100 rpm, pH=5.15, and Cs/Alg Mw ratio=1.75 based on numerical optimization. It was shown that the average particle size and loading efficacy at optimum conditions were 253 nm and 67%, respectively. Other responses were as follows: CR=66%, ζ-potential=+35mV, PDI=0.5, and MDT=7 days.Conclusion: The results have suggested that the statistical optimization of rhBMP-2 offers the possibility of preparing Cs/Alg/B NPs with a favorable size, controlled release characteristics, and high loading efficiency. It is expected that the acquired optimum conditions will be useful for efficient rhBMP-2 delivery.Keywords: alginate, Box–Behnken design(BBD), chitosan, response surface methodology (RSM), recombinant human bone morphogenetic protein-2 (rhBMP-2), chitosan/alginate nanoparticles

Background Salmonellosis is a critical and common infectious malady between people and animals caused by Salmonella bacteria. With the advent of antibiotic resistance, it is essential for new methods to be replaced to prevent and treat infections. Bacteriophages are promising choices. Methods In this study, phage vB_SenS_TUMS_E4 against Salmonella enteritidis isolate has been separated from poultry wastewater. Determination of phage characteristics, including plaque formation, imaging with transmission electron microscopy, growth curve, structural proteins profile, host range, and pH and temperature parameters. Also, the Phage genome was extracted, sequenced, and annotated, and by utilizing Average Nucleotide Identity and phylogeny was compared with reference Salmonella phages. Results The burst size was large, nearly 287 plaque-forming units per cell (PFU/cell) and elevated stability to many temperatures and pH values. Phage vB_SenS_TUMS_E4 was effective on various clinical and environmental strains of Salmonella but did not affect bacteria of other genera. The morphological analysis indicated that phage vB_SenS_TUMS_E4 belongs to the Siphoviridae family. The genome of vB_SenS_TUMS_E4 is a linear dsDNA molecule of 43,018 bp with a G + C content of 49.7%. It includes 60 protein-coding genes, yet it contains no tRNA genes. Among the 60 detected putative protein-coding genes, just 43 gene products were contained in database searches. No genes associated with antibiotic resistance, virulence factor, and lysogenic were realized in the vB_SenS_TUMS_E4 genome. Conclusion The findings indicate that the high lytic potency vB_SenS_TUMS_E4 polyvalent phage is an antibacterial agent for controlling Salmonella in food production, prevention, and Salmonella treatment.

Nowadays, the trend to simplify and miniaturize sample preparation methods has resulted in the development of effective and low-cost microextraction techniques that utilize a very small volume of the extracting phase. Among them, the liquid-liquid microextraction (LLME) method is a simple and effective sample pre-treatment technique applicable to numerous analytical methods. A related miniaturized and environmentally friendly extraction technique, dispersive liquid-liquid microextraction (DLLME), has been developed within the last decade and shows a very high enrichment factor and very low solvent consumption compared to other liquid- or even solid-phase extraction methods. The inclusion of several effective parameters in DLLME and its variants has increased the need for optimization to obtain the best possible extraction results. In fact, experimental design and optimization of performance conditions are the most important applications of chemometrics in analytical chemistry. Thus, design of experiments (DoE) helps us to determine the best model of the relationship between variables, as well as the optimal experimental conditions. Here, a comprehensive review of recent advancements in the use of DoE methodologies including full factorial, fractional factorial, Plackett-Burman, orthogonal array, central composite, Box-Behnken, Doehlert, and D-optimal designs to optimize DLLME applications is provided. In addition, the preponderance and drawbacks of each optimal method are discussed. The overall purpose of this review is to present a general overview of the different DoEs that are currently used to optimize DLLME for various matrices and analytes.

Arise of multidrug resistance in cancer cells is a major obstacle for effective cancer chemotherapy. Multi-drug resistant (MDR) may be treated using combinations of encapsulated cytotoxic drugs and chemosensitizers. Dual drug delivery can control the drug-releasing pattern and improve the accumulation of chemotherapeutic agents at targeting sites. MDR reversal agent such as verapamil simultaneously with a traditional anticancer drug to cancer cells for enhanced chemotherapy efficacy have been limited due to the lack of efficient co-delivery methods. To optimize the effectiveness of this combinational, this study presents a system constructed of poly-lactid-co-glycolide (PLGA) carries SN38 and verapamil. SN38 is the active metabolite of irinotecan, a chemotherapeutic agent which acts as an inhibitor of topoisomerase 1 and verapamil which is a MDR1 reversal agent. Biological evaluation designed for determining cytotoxicity and cellular uptake evaluation on MDA-MB-231 breast cancer cells. Nanoparticles fabrication were optimized by small central composite design and characterized. Nanoparticles showed cytotoxic effects on MDA-MB-231 cells. Increasing of BAX expression and decreasing of Bcl-2 levels verified our hypothesis based on using verapamil to increase apoptosis following P-gp inhibition. PLGA nanoparticles also showed a selective distribution in animal body.

Choroidal neovascularization (CNV) is among the leading causes of blindness worldwide. Bevacizumab has demonstrated promising effects on CNV treatment; however, frequent intravitreal injection is its major drawback. Current study aimed to address this issue by developing a sustained release formulation through nanoparticles of bevacizumab imbedded in an ocular implant. Bevacizumab-loaded chitosan nanoparticles were prepared by ionic gelation method and inserted in the matrix of hyaluronic acid and zinc sulfate. Despite the common approaches in using ultraviolet (UV)-spectrophotometry, microprotein-Bradford, and bicinchoninic acid (BCA), assay for protein assessment, our results revealed a remarkable UV-Vis absorption overlap of protein and chitosan during these analysis and thus enzyme-linked immunosorbent assay was employed for the antibody concentration assay. The size of optimized nanoparticles obtained through statistical analysis based on design of experiments was 78.5 ± 1.9 nm with polydispersity index of 0.13 ± 0.05 and the entrapment-efficiency and loading-efficiency were 67.6 ± 6.7 and 15.7 ± 5.7%, respectively. The scanning electron microscopy and confocal microscopy images revealed a homogenous distribution of nanoparticles in the implant matrix and the release test results indicated an appropriate extended release of bevacizumab from the carrier over two months. In conclusion, the prepared system provided a sustained release bevacizumab delivery formulation which can introduce a promising ocular drug delivery system intended for posterior segment disease. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 2261-2271, 2018.

Backgrounds and Aims Selenium nanoparticles (SeNPs) have been reported to exhibit an inhibitory effect on cancer cells. In the present study, we aimed to compare the in vitro and in vivo effects of SeNPs and folic acid surface–coated selenium nanoparticles (FA@SeNPs) on breast cancer. Methods FA@SeNPs and SeNPs were chemically synthesized and characterized with different instrumental techniques. The cytotoxicity of both nanomaterials was evaluated against 4T1 cells. In addition, the intravenous administration effect of these nanomaterials (300 μg/week) on the lifespan and tumor size of cancer-bearing mice was investigated. Results Although the SeNPs showed an antiproliferative effect against the cell line, the cytotoxicity of the FA@SeNPs was higher than that of the SeNPs. A low concentration of FA@SeNPs (25 μg/mL corresponding to 8.75 μg/mL of elemental SeNPs) caused approximately 68% cell mortality. In the in vivo study, the nanomaterials decreased the tumor growth rate in cancerous mice in relation to the control group. FA@SeNPs were more effective than SeNPs. Conclusions The combination of SeNPs and FA has a potent antiproliferative effect against 4T1 cells, significantly increases the lifespan, and prevents tumor growth.

Chitosan/alginate (Chi/Alg) nanoparticles as a non-viral vector for the Smad4 encoding plasmid were optimized utilizing D-optimal design based on the nanoparticles/plasmid ratio, Chi/Alg MW, and preparation method type. Following the optimization and validation of the best formula, morphology studies and FTIR measurements were performed to evaluate the optimized Chi/Alg/S NPs. Toxicity (MTT assay) and transfection studies were performed for the best formula in comparison with Lipofectamine 2000, and Polyethyleneimine (PEI) and evaluated using Green Fluorescence Protein (GFP) assay, Flow cytometry, and RT-PCR. The model predicted a particle size of 111 nm, loading efficacy (LE) of 43%, cumulative release (CMR) of 39%, the ζ-potential of +50 mV, and PDI of 0.13. The predicted point condition was as follows: NP ratio = 13, Chi/Alg MW ratio = 2.35, and preparation method type = 1. Microscopic findings revealed that the shape of nanoparticles was spherical. The Chi/Alg/S nanoparticles showed no toxicity and transfection efficacy of 29.9% was observed in comparison with Lipofectamine (35.5%) and PEI (30.9%).

The principal purpose of the present study was to prepare and characterize a complex drug delivery system consisting of Nafarelin-poly (3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) nanoparticles (NPs) in combination with sodium alginate/poloxamer 407 in situ gel. Nafarelin-loaded PHBV NPs were prepared via double emulsion solvent evaporation technique. Box-Behnken Response Surface Methodology was utilized to optimize NPs. Mean particle size, polydispersity index (PDI), entrapment efficiency (EE), and drug loading (DL) of the optimized NPs were measured. Incorporation of Nafarelin within NPs was proven by differential scanning calorimetry (DSC). The combination delivery system (CDS) was prepared by adding Nafarelin-loaded PHBV NPs to sodium alginate/poloxamer 407 solution followed by physical mixing. Morphological properties of Nafarelin-loaded PHBV NPs and CDS were evaluated by SEM. Rheological properties were employed to investigate the effects of alginate concentration on sol-gel transition temperature. The release profile of Nafarelin from both PHBV NPs and CDS were individually assessed. The cumulative release percentage from CDS was significantly lower than Nafarelin released from PHBV NPs. Based on the favorable results in this study, the CDS consisting of sodium alginate/poloxamer 407 loaded with PHBV NPs could be a promising candidate for designing a long-lasting formulation of Nafarelin.

A polyaniline–multiwalled carbon nanotubes composite fiber was electrodeposited onto a platinum wire using cyclic voltammetry. This fiber was used for headspace solid-phase microextraction combined with gas chromatography and flame ionization detector of trace levels of benzaldehyde in some injectable pharmaceutical formulations. Three solid-phase microextraction parameters including temperature, time and stirring rate were investigated simultaneously using a three-level-three-factor Box–Behnken as experimental design on the extraction capability. The as-made fiber has a lifetime of over 300 extractions without any obvious decline in extraction efficiency. At the optimum conditions (extraction temperature 60°C, extraction time 15 min, stirring rate 700 rpm), the method displays excellent linearity over the concentration range of 25–1000 ng/mL of benzaldehyde with RSD values ranging from 1.0 to 6.8%. The limits of quantitation and detection were 25 and 10 ng/mL, respectively.

Gene therapy is a novel approach for cancer treatment and investigation for suitable gene delivery systems is remarkable. Here, preparation of a polyelectrolyte complex containing polysaccharides: trimethyl chitosan (TMC) as the positive and hyaluronate (HA), dextran sulfate and alginate as the negative part was studied. The optimized nanoparticles (TMC: between 0.2 and 0.47 mg/ml, HA: 0.35 mg/ml (≈131 nm, nearly full gene loading)) were obtained via primary screening followed by the D-optimal method. In vitro cellular study on the MCF7 cell line confirmed the non-toxicity and high cellular uptake (>90%) of prepared nanoparticles. Notably, in vivo study indicated noticeable tumor uptake of nanoparticles while low accumulation in vital organs such as heart, liver and lungs. Moreover, although a qualitative variable was considered, the applied method restricted the number of runs by selecting spots from the spherical atmosphere. The prepared nanoparticles could be suggested as an efficient and safe delivery system for cancer gene delivery.

This study describes the enzymatic elimination of imipramine and four other tricyclic antidepressants (TCAs) by laccase-mediated catalysis in aqueous solution. The results showed that TCAs demonstrated dissimilar enzymatic elimination behavior: up to 67% of clomipramine and 82% of imipramine were significantly removed during the first 6 h. The elimination percentages of amitriptyline, doxepin, and nortriptyline were 11%, 6%, and 23%, respectively, after 72 h laccase-based treatment. Due to the rapid and high percentage of removal, imipramine was selected for detailed toxicity evaluation. Optimal levels for the main factors in the enzymatic removal process of imipramine were obtained as pH (4.9), incubation time (5.7 h), imipramine concentration (0.12 μg mL −1 ), and enzyme activity (1.63 U mL −1 ). Laccase-catalyzed transformation of imipramine led to the formation of an unknown metabolite which was subsequently purified and spectroscopically characterized as 3-(2,7-dihydroxy-10,11-dihydro-5 H -dibenzo[ b,f ]azepin-5-yl)-1-hydroxy- N , N -dimethyl-3-oxopropan-1-amine oxide. The toxicity reduction of the bio-product was assessed by MTT cell and Caco-2 cell line.

The presence of recalcitrant synthetic dyes in environment is a major concern due to their possible threat on living organisms and aquatic ecosystems. To investigate the potential of combined thermal and enzymatic decolorization, betaine was applied to protect the secondary and tertiary structures as well as the enzymatic activity of laccase under thermal stress. At 60 °C, betaine 1 M significantly enhanced the stability of laccase in addition to keeping its catalytic efficiency (p T m of laccase in the presence of betaine. Once incubated at 60 °C, betaine exhibited superior protein-stabilizing properties than at 25 °C. Furthermore, fluorescence spectroscopy supported this concept that the overall folded structure of laccase became more rigid in the presence of betaine at 60 °C. Betaine-stabilized laccase was used for combined thermal and enzymatic decolorization of some recalcitrant organic dyes. This osmolyte could improve the enzymatic removal of most dyes at 60 °C. A detoxification study with respect to the inhibition of bacterial growth showed a decrease in microtoxicity of the laccase-treated dye solution in the presence of betaine at 60 °C.

In the present study, an electrically-assisted microextraction method termed electromembrane extraction (EME) followed by a simple high performance liquid chromatography-ultraviolet detection was developed and validated for the determination of phenytoin in human biological samples. Main parameters influencing the electromembrane extraction were evaluated and optimized using the Box-Behnken experimental design. The membrane consisted of 1-octanol immobilized in the pores of a hollow fiber. As a driving force, a 40 V electric field was applied to facilitate the migration of analytes from the sample solution to an acceptor solution through a supported liquid membrane. The optimum donor and acceptor solutions pHs were achieved 10 and 13, respectively. The enrichment factor was > 82 within 15 min led to 55% absolute extraction recovery. In optimum conditions, the method provided the linearity in the range of 10–1000 ng/mL (R2 >0.999). The repeatability of the method was indicated as relative standard deviations (%RSD) between 3.6% and 8.9% (n = 3). The limits of detection and quantitation were 3.0 and 10.0 ng/mL, respectively. The sensitivity of HPLC-UV for determining phenytoin was enhanced by electromembrane extraction. Also, in this study, the effects of some nano-sorbents like carbon nanotubes and molecularly imprinted polymer on membrane performance and EME efficiency were evaluated. The EME technique can be introduced as a new approach for screening of molecularly imprinted polymers in their primary steps of characterization. Finally, the proposed EME-HPLC-UV method was applied for phenytoin determination in human plasma and urine samples with relative recoveries ranged between 88–92% indicating the reliability of the method.

Some devices are used in the pharmaceutical industry to produce and control the quality of a variety of pharmaceutical products; therefore, it would be helpful to identify the factors affecting the performance of the devices and optimize them in terms of time and cost. In this regard, the use of experimental design approaches to optimization is important as it can help a device achieve its highest efficiency. One of the most popular devices in the pharmaceutical industry is high-performance liquid chromatography (HPLC). In view of its wide use in the industry and the number of factors affecting it, it is necessary to design and validate a new method that can simultaneously optimize its different parameters. In this study, the method of the simultaneous optimization of the parameters was implemented using the Box-Behnken design and multiple-criteria decision-making (MCDM) by Derringer's desirability function and technique for the Order of Preference by Similarity to Ideal Solution (TOPSIS) method. In this regard, mobile phase ratio, flow rate, and pH were simultaneously changed, and capacity factor, total analysis time, and resolution were considered as responses. To validate the method, the key parameters of the method were determined and used for all the validation steps. Therefore, this approach to optimizing HPLC factors is new and very cost-effective with diverse applications in the pharmaceutical industry.

Quantification and qualification of an analyte of interest in pharmaceutical tablets from different manufacturers/companies are a hard task because of the potential presence of various interfering molecules. Indeed, the composition of the tablets covers a wide range of interferents which can be even unknown. As a consequence, we propose to determine the concentration of an analyte of interest regardless of the interferents using the concept of universal calibration. Universal calibration paves the way to the quantification of a specific chemical entity in samples with various compositions and different interferents. This is possible by the trilinear structure of analyte's signal. In fact, the second-order advantage resulting from the second-order universal calibration models is exploited. However, a new second-order calibration strategy was conducted in this work using Trilinear Factor Extraction (TFE). A simulated data set was exemplified to highlight the ability of the proposed procedure in order to accurate extraction of the analyte's concentration profile. Additionally, two real data sets were also explored in order to test the TFE method. In the first case, Acetaminophen was quantified using fluorescence spectroscopy in tablets with different formulations from 6 companies. In the second experimental data, a peptide (Valine-Tyrosine-Valine) was successfully quantified in different samples using spectrofluorimetric data. Finally, these real data sets were analyzed by Multivariate Curve resolution – Alternating Least-Squares (MCR-ALS) under non-negativity and trilinearity constraints for the sake of comparison. The calculated Root Mean Square Error of Predictions (RMSEP) of Acetaminophen were 0.028 and 0.026 for the MCR-ALS and TFE models, respectively. On the other hand, for the second experimental data set, the RMSEP were 0.216 and 0.165, respectively. Finally, based on a paired t-test, the results of MCR-ALS and TFE were not significantly different.

The moisture content of many pharmaceutical products, like most vaccines, is an important control parameter in the product qualification. Freeze-drying is a widely applied technique for the preservation and storage of such products. Karl Fischer titration is the standard method for determination of residual moisture in freeze-dried samples; however, it is a time-consuming and destructive method that requires the handling of organic solvents. Instead, near-infrared (NIR) spectroscopy is a fast and non-invasive method that is well suited to the measurement of water due to the strong adsorption band of water in NIR. In this study, multivariate fitting regression with Gaussian function (MFRG) was applied to predict the water content of a freeze-dried sample drug using NIR spectra. Moreover, the prediction performance of the MFRG was compared to the results of the well-known PLS method. Several simulated datasets and experimental data were used to evaluate the performance of the MFRG method. In the case of experimental data, a number of mathematical pre-treatments were applied to evaluate whether they led to better models. In addition, several calibration and test sets were designed and applied to compare the quantification results of MFRG and PLS methods. It was found that the performance of the MFRG model was comparable or better compared to the PLS regression method in nearly all the examined cases, which is due to fact that MFRG models nonlinearity better than the PLS method.

Owing to their superior catalytic activity in the extreme conditions, extremozymes have found the potential biotechnological applications for industrial purposes. A robust extracellular protease activity was detected in the culture broth of

The fungal laccase isolated from Paraconiothyrium variabile was applied to optimize the enzymatic decolorization and detoxification of the azo dye reactive orange 7 using response surface methodology. Initial screening using fractional factorial design was carried out to select important independent variables that significantly affect the yield of decolorization including enzyme activity, pH, and incubation time. Optimum condition for maximal dye decolorization obtained by central composite design and surface plots were laccase activity 1.25 U/mL, pH 6.0, and incubation time 12.5 min. In addition, the kinetic and energetic parameters for enzymatic decolorization of reactive orange 7 were studied and the calculated values for Km, Vmax, Ea, ΔH, and ΔS were 1.14 mM, 283.28 mM/min.mg, 40 kJ/mol, 89 kJ/mol, and 285 J/mol.K, respectively. Decolorization is endothermic and spontaneous at temperatures higher than 37.8 °C. Microtoxicity evaluation demonstrated a decrease in toxicity of the laccase-treated dye in comparison with the parent dye.

Herein we reported the structure and several properties of a new biosurfactants produced by Corynebacterium xerosis strain NS5. This strain was capable of producing a novel lipopeptide biosurfactant that we have named coryxin. The biosurfactant structure was characterized by using Fourier transform infrared spectroscopy (FTIR), Nuclear magnetic resonance spectroscopy (NMR), and Liquid chromatography-mass spectrometry (LC-MS). It contained a hydrophobic moiety of 3-hydroxydecanoic acid and a peptide part predicted as a sequence of seven amino acids including Asn-Arg-Asn-Gln-Pro-Asn-Ser. Coryxin lowered the surface tension of water to 31.4 mN/m, with a critical micelle concentration of 25mg/l. It was a strong emulsifier with an emulsification index of 61% against n-hexane. Coryxin showed antibacterial activity against test organisms belonging to Gram-positive and Gram-negative bacteria and disrupted preformed biofilms of Staphylococcus aureus (82.5%), Streptococcus mutans (80%), Escherichia coli (66%) and Pseudomonas aeruginosa (30%). In conclusion, microbial surfactant from C. xerosis exhibited inhibitory and disruptive activities against biofilm formation that could be of use in biofilm-related menace.

Hypercholesterolemia is an important risk factor contributing to atherosclerosis and coronary heart disease. Lactic acid bacteria have attracted much attention regarding their promising effect on serum cholesterol levels. Tellurium (Te) is a rare element that has also gained considerable interest for its biological effects. There have been some recent in vivo reports on the reduction effect of Te on cholesterol content. In this study, Lactobacillus plantarum PTCC 1058 was employed for the intracellular biosynthesis of Te NPs. The UV-visible spectrum of purified NPs showed a peak at 214 nm related to the surface plasmon resonance of the Te NPs. Transmission electron microscopy showed that spherical nanoparticles without aggregation had the average size of 45.7 nm as determined by the laser scattering method. The energy dispersive X-ray pattern confirmed the presence of Te atoms without any impurities. A significant reduction was observed in group which received L. plantarum with or without Te NPs during propylthiouracil and cholesterol diet in compare with the control group which received just propylthiouracil and cholesterol. The levels of triglycerides also remarkably decrease (p

A new nanoparticle Cd(II) ion-imprinted polymer (IIP) has been prepared via precipitation polymerization using 2-vinylpyridine (the functional monomer), ethylene glycol dimethacrylate (the cross-linker), 2,2′-Azobisisobutyronitrile (the initiator), diphenylcarbazide (the ligand), and cadmium nitrate (the template ion). The prepared IIP was characterized by FT-IR spectroscopy, scanning electron microscopy, and transmission electron microscopy. The optimal pH value for the quantitative sorption was 6.0 and maximum sorbent capacity was 16.52 mg/g. Under the optimized conditions, the method had a linear analytical range from 1 to 50 μg/L. The detection limit, relative standard deviation, and recovery of the method were evaluated as 0.14 μg/L, 2.6% and 96–98%, respectively. The prepared IIP revealed good selectivity over a wide variety of other cations. The developed method was successfully applied for the selective extraction and determination of trace amount of cadmium ions in synthetic and real wate...

Controlled delivery of multiple therapeutic agents can be considered as an effective approach in skin tissue engineering. In this study, recombinant human epidermal growth factor (rhEGF) and recombinant human basic fibroblast growth factor (rhbFGF) encapsulated in PLGA microspheres were loaded in hybrid scaffolds of PLGA and PEO. The scaffolds with various formulations were fabricated through electrospinning in order to maintain dual, individual or different release rate of rhEGF and rhbFGF. Morphological, physical and mechanical properties of the scaffold were investigated. The scaffold possessed uniform morphology with an average diameter of 280 nm for PLGA and 760 nm for PEO nanofibers. Furthermore, the mechanical properties of the scaffolds were shown to be akin to those of human skin. Bioactivity of the scaffolds for human skin fibroblasts was evaluated. The HSF acquired significant proliferation and well-spread morphology on the scaffolds particularly in the case of different release rate of rhEGF and rhbFGF which implies the synergistic effect of the growth factors. Additionally, collagen and elastin gene expression was significantly up-regulated in the HSF seeded on the scaffolds in the case of individual delivery of rhEGF and dual delivery of rhEGF and rhbFGF. In conclusion, the prepared scaffolds as a suitable supportive substrate and multiple growth factor delivery system can find extensive utilization in skin tissue engineering.

Choroidal neovascularization (CNV) is among the leading causes of blindness worldwide. Bevacizumab has demonstrated promising effects on CNV treatment; however, frequent intravitreal injection is its major drawback. Current study aimed to address this issue by developing a sustained release formulation through nanoparticles of bevacizumab imbedded in an ocular implant. Bevacizumab-loaded chitosan nanoparticles were prepared by ionic gelation method and inserted in the matrix of hyaluronic acid and zinc sulfate. Despite the common approaches in using ultraviolet (UV)-spectrophotometry, microprotein-Bradford, and bicinchoninic acid (BCA), assay for protein assessment, our results revealed a remarkable UV-Vis absorption overlap of protein and chitosan during these analysis and thus enzyme-linked immunosorbent assay was employed for the antibody concentration assay. The size of optimized nanoparticles obtained through statistical analysis based on design of experiments was 78.5 ± 1.9 nm with polydispersity index of 0.13 ± 0.05 and the entrapment-efficiency and loading-efficiency were 67.6 ± 6.7 and 15.7 ± 5.7%, respectively. The scanning electron microscopy and confocal microscopy images revealed a homogenous distribution of nanoparticles in the implant matrix and the release test results indicated an appropriate extended release of bevacizumab from the carrier over two months. In conclusion, the prepared system provided a sustained release bevacizumab delivery formulation which can introduce a promising ocular drug delivery system intended for posterior segment disease. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 2261-2271, 2018.

The effects of vinegar and oxymel solutions (used in Iranian traditional medicine) and processing temperatures (75, 85 and 100°C) on the milk coagulation was studied to identify and determine the protein indicators of resulting whey. The HPLC (high performance liquid chromatography) analysis showed that the α-lactalbumin (ALA) and β-lactoglobulin (BLG) contents of resulting whey reduced significantly, when the coagulation temperature increased from 75 to 100°C. In both solutions and at 75°C processing temperature, the BLG of final whey protein was significantly more (∼ 60-70 %) than ALA content. The reverse process happened at 85°C, and the ALA content became much higher than BLG. A significantly higher content of ALA was recovered once the combination of oxymel and vinegar solutions with milk reached to respectively ∼ 10 and 0.3 % and milk protein was coagulated at 85°C. Overall, there is a good possibility to make a milk whey with high content of desirable ALA and negligible concentrations of BL...

Liquid protein formulations are prone to form aggregates. The effect of nonionic surfactants such as Polysorbate 20 (PS 20) and n-Dodecyl β-D-maltoside (DDM) on the prevention of aggregation and conformational changes of recombinant human IFNβ-1b (rhIFN β_1b) was explored. Polysorbate has been used in formulations of protein pharmaceuticals. There have been concerns about using PS 20 due to its residual peroxide content which may negatively affect protein efficacy. n-Dodecyl β-D-maltoside has been of interest and shown to be highly effective in prevention of aggregation. Fresh bulk of rhIFN β_1b was formulated using DDM or different concentrations of PS 20. Formulations were exposed to light stress condition according to the ICH guideline of Q1b. The overall conformational integrity of individual samples was characterized by a combination of Circular dichroism (CD), Fluorescence spectroscopy and RP_HPLC techniques. The CD spectrum depicting the conformational integrity of rhIFN β_1b showed 31.9% and 31.2% decreases in α-helix content of protein samples with 0.2% or 0.02% of PS20 compared to only18.2% of that containing 0.2% DDM. The RP-HPLC analysis also showed that the oxidized impurity in formulation containing DDM is less than those contain PS 20. Complementary analysis of the liquid formulations using IFR and UV methods also was in compliance with the data obtained by CD. Compared to PS 20, the sample of rhIFN β_1b formulation with DDM was more resistant to the destruction effect of light. Results were in accordance with previous studies and could suggest DDM as a reliable anti-aggregation surfactant in biopharmaceutical formulations.

Since industrial revolution heavy metals such as lead (Pb) and cadmium (Cd) have been extensively dispersed in environment which, unknown biological effects and prolong biological half-life make them as a major hazard to human health. In addition, the sharp increase in Multiple sclerosis incidence rateshas been recorded in Iran. The propose of this study was to measuring blood lead and cadmium concentration and their correlation with smoking habit in a group of 69 RRMS patients and 74 age/gender-matched healthy individuals resident in Tehran as most polluted city in Iran. All subjects were interviewed regarding age, medical history, possible chemical exposure, acute or chronic diseases, smoking and dietary habits. Blood Pb and Cd levels were measured by double beam GBC plus 932 atomic absorption spectrometer. Our result indicated a significant difference in Cd level (p = 0.006) in which, MS patients had higher blood concentration (1.82 ± 0.13 μg/L) in comparison with healthy individuals (1.47 ± 0.11 μg/L). A comparable blood Cd level to similar recent study (1.78 µg/L vs.1.82 µg/L) was observed. With respect to Pb there was no significant difference between cases and controls, however the geometric means of blood Pb concentration were considerably higher in males than in females in MS patients (57.1 ± 33.7 μg/L vs. 36.7 ± 21.9 μg/L. P = 0.02). Taking into consideration tobacco smoking, an elevated contents of each metal were observed in smoker subjects (p

Background Cancer stem cells (CSC) have been proposed as the reason of cancer relapse which are characterized mainly based on CD44+ phenotype with other supplementary markers. The aim of the present study is to fabricate cis-dichlorodiamminoplatinum (II) (CDDP) loaded glyconanoparticles using hyaluronic acid (HA) which is also known as the endogenous substrate for CD44 in vivo. Methods For this purpose, a drug-induced ionic gelation technique has been used to prepare CDDP-incorporated nanoparticles. To optimize the fabrication technique, stirring rate, stirring time, and HA/CDDP ratio have been selected as the main factors from other factors and subjected to face-centered central composite design for optimization purposes. The optimized nanoparticles were further characterized using different complementary methods including FTIR, SEM, AFM and DSC. To evaluate the biological effectiveness of CDDP nanoparticles release study, MTS assay, tumor cell clonogenicity and sphere formation assay have been performed as well. Results Spherical CDDP nanoparticles with Z-average approx. 150 nm with low PdI were prepared by adjusting the selected variables. FTIR results indicated the presence of inclusion complexes between CDDP and HA which lead to preparing nanoparticles with high entrapment efficiency and drug content of 87.4 and 43.74 percentage respectively. In vitro release study showed a sustained release of CDDP up to 4 days, and cellular studies confirmed that nanoparticles formation keeps the anticancer activity of formulated CDDP while moderate increase in cancer stem cell suppression. Conclusion It seems hyaluronic acid could be successfully exploited as carrier in cancer-targeted drug delivery with a look at targeting the CSCs.

In this work, polyaniline, polypyrrole, and polyaniline/polypyrrole composite fibers were synthesized in the absence and presence of oxidized multiwalled carbon nanotubes using electrochemical cyclic voltammetry with CF3COOH as dopant. Thermal stability of these fibers was studied by differential scanning calorimetry. Then, headspace solid-phase microextraction process coupled with gas chromatography and flame ionization detector was used for comparing extraction capability of benzaldehyde from aqueous solution. Since polyaniline fiber showed better extraction efficiency than the other fibers, its preparation conditions including acid concentration, aniline concentration, scan rate, and amount of multiwalled carbon nanotubes were studied by means of the “one-factor-at-a-time method”. The analytical performance of polyaniline fibers were investigated to determine benzaldehyde from the aqueous solution. The morphology and texture of polyaniline fibers were examined by field emission scanning electron microscopy and Fourier transform infrared spectroscopy analyses. The attained results revealed that the perfect conditions for acid concentration, aniline concentration, scan rate, and multiwalled carbon nanotubes content were 0.5 M, 0.2 M, 25 mV s−1, and 0.02 wt%, respectively. The limit of detection for the proposed polyaniline fiber was 15 ng ml−1.

Nanoparticles of ultra low molecular weight chitosan (ULMWCS)/hyaluronic acid (HA) were prepared by ion gelation. Three independent variables, namely, ratio of concentration of ULMWCS to HA (CS/HA), pH of solution and stirring time were studied to identify their effects on size, polydispersity and zeta potential of prepared nanoparticles using a Box-Behnken design. Results showed that pH and CS/HA have a direct effect on size, while increase of stirring time decreases the size of nanoparticles. Additionally, it was shown that all the independent parameters have direct effects on zeta potential. Also, the minimum polydispersity index was observed at lowest values of CS/HA. The model also predicted that the optimum values are 4.15, 4.14 and 180 (min) for the CS/HA, solution pH and stirring time, respectively. The obtained preparation had a size of 200 nm, polydispersity index of 0.37, and zeta potential of 13.0 mV.

Vincristine (VC) sulfate, a freely water-soluble cytotoxic agent was incorporated into cetyl palmitate solid lipid nanoparticles (SLNs) with the aid of dextran sodium sulfate (DS), a poly anion, using a microemulsion method. The manufacturing process was optimized using response surface methodology (Box–Behnken design). SLNs were characterized for size, zeta potential, morphology, crystallinity, and release behavior. The drug encapsulation efficiency reached up to 93% and release study revealed sustained drug release. SLN formulation showed comparable cytotoxic effect in comparison to VC sulfate solution against the MDA-MB-231 cells. The in vivo studies following injection to rat revealed higher plasma and tissue concentrations and longer drug mean residence times compared to VC solution. Using cumarin-6 as a model drug, it was shown that drug delivery to the brain was enhanced close to five times using SLNs prepared in this study compared to free cumarin-6. It can be concluded that complexes of cetyl palmitate SLNs with DS could produce high VC-loaded SLNs suitable for delivery of anticancer drugs to brain tumors. © 2013 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 102A: 2125–2136, 2014.

The objective of this study was to prepare poly lactic-co-glycolic acid (PLGA)-based microparticles as potential carriers for recombinant human epidermal growth factor (rhEGF). In order to optimize characteristic parameters of protein-loaded microspheres, bovine serum albumin (BSA) was selected as the model protein. To reduce burst release as a common problem of microspheres, a proper alteration in the particle composition was used, such as addition of poly vinyl alcohol and changes in initial drug loading. The effects of these parameters on particle size, encapsulation efficiency and in vitro release kinetics of BSA in PLGA microspheres were investigated using a Box-Behnken response surface methodology. The biological activity of the released rhEGF was assessed using human skin fibroblasts cell proliferation assay. The prepared rhEGF-loaded microspheres had an average size of 6.44 ± 2.45 µm, encapsulation efficiency of 97.04 ± 1.13%, burst release of 13.06 ± 1.35% and cumulative release of 22.56 ± 2.41%. The proliferation of human skin fibroblast cells cultivated with rhEGF releasate of microspheres was similar to that of pure rhEGF, indicating the biological activity of released protein confirming the stability of rhEGF during microsphere preparation. These results are in agreement with the purpose of our study to prepare rhEGF-entrapped PLGA microparticles with optimized characteristics.

Extremozymes have gained importance for their ability to efficiently develop the processes in rigorous industrial conditions with incidence in the recycling of especially robust natural wastes. The production of an extracellular laccase from the halophilic bacterium Chromohalobacter salexigens aided for the bio-delignification of almond shell was optimized using response surface methodology followed by one-factor-at-a-time, resulting in an 80-fold increase in the enzyme yield. Out of 10 different medium components, CuSO4, ZnSO4, glucose, and urea were shown to have the greatest effects on the laccase production. The crude laccase was surprisingly stable against the various solvents, salts, chemicals, pH ranges, and temperatures, and it exhibited a high catalytic efficiency to a wide range of phenolic and non-phenolic substrates. Laccase reduced the kappa number of the lignin of almond shell by approximately 27% without the aid of a mediator, and the delignification efficiency strengthened by up to 58% reduction in kappa number in the used harsh conditions. Due to the high potential of the enzyme in delignification, specifically under extreme conditions, laccase from C. salexigens can be considered as an ideal alternative for chemical treatment methods in cellulose fibres extraction of lignocellulosic bio-wastes or delignification of the lignin and lignin-derived industrial wastes.