Cloning and expression of recombinant arazyme with anti-inflammatory and anti-breast cancer potential.

Arazyme is an extracellular metalloprotease which is secreted by a Gram-negative symbiotic bacterium called Serratia proteomaculans. There are limited studies on various biological activities of arazyme. This preliminary study was designed to investigate the anti-cancer and anti-inflammatory capacities of recombinant arazyme (rAra) in vitro and in vivo. Arazyme gene, araA was cloned and expressed in E. coli BL21 (DE3) using pET-28a as a vector. Nickel column purification was used to obtain pure rAra. SDS-PAGE and protein assay were used to identify the product and to measure protein content, respectively. Skimmed milk test and casein assay were carried out to assess protease activity. MCF7 cells as a breast cancer cell model were exposed to different concentrations of rAra to study anti-breast cancer potentials using MTT assay. The anti-inflammatory property of rAra was investigated using a murine air-pouch model. PCR and SDS-PAGE data showed that cloning and expression of rAra was successful and the enzyme of interest was observed at 52 KDa. Protein assay indicated that 1 mg/ml of rAra was obtained through purification. A clear zone around the enzyme on skimmed milk agar confirmed the proteolytic activity of rAra and the enzymatic activity was 320 U/mg protein in the casein assay. Cytotoxic effects of rAra reported as IC50 were 16.2 µg/ml and 13.2 mg/ml after 24 h and 48 h, respectively. In the air-pouch model, both the neutrophil count and myeloperoxidase activity, which are measures of inflammation, were significantly reduced. The results showed that rAra can be used in future mechanistic studies and R&D activities in the pharmaceutical industry to investigate the safety and efficacy of the recombinant arazyme.

Modeling sunset yellow removal from fruit juice samples by a novel chitosan-nickel ferrite nano sorbent.

Analysis of food additives is highly significant in the food industry and directly related to human health. This investigation into the removal efficiency of sunset yellow as an azo dye in fruit juices using Chitosan-nickel ferrite nanoparticles (Cs@NiFe2O4 NPs). The nanoparticles were synthesized and characterized using various techniques. The effective parameters for removing sunset yellow were optimized using the response surface methodology (RSM) based on the central composite design (CCD). Under the optimum conditions, the highest removal efficiency (94.90%) was obtained for the initial dye concentration of 26.48 mg L-1 at a pH of 3.87, a reaction time of 67.62 min, and a nanoparticle dose of 0.038 g L-1. The pseudo-second-order kinetic model had a better fit for experimental data (R2 = 0.98) than the other kinetic models. The equilibrium adsorption process followed the Freundlich isotherm model with a maximum adsorption capacity of 212.766 mg g-1. The dye removal efficiency achieved for industrial and traditional fruit juice samples (91.75% and 93.24%), respectively, confirmed the method's performance, feasibility, and efficiency. The dye adsorption efficiency showed no significant decrease after five recycling, indicating that the sorbent has suitable stability in practical applications. variousThe synthesized nanoparticles can be suggested as an efficient sorbent to remove the sunset yellow dye from food products.

Synthesis and characterization of a novel magnetic chitosan-nickel ferrite nanocomposite for antibacterial and antioxidant properties.

A novel nanomagnet modified with nickel ferrite nanoparticles (NPs) coated with hybrid chitosan (Cs-NiFe2O4) was synthesized using the co-precipitation method. The resulting nanomagnets were characterized using various techniques. The size of the nanomagnetic particles was estimated to be about 40 nm based on the transmission electron microscopy (TEM) image and X-ray diffraction analysis (XRD) pattern (using the Debye-Scherrer equation). Scanning electron microscopy (SEM) images indicated that the surface of Cs-NiFe2O4 NPs is flatter and smoother than the uncoated NiFe2O4 NPs. According to value stream mapping (VSM) analysis, the magnetization value of Cs-NiFe2O4 NPs (17.34 emu/g) was significantly lower than NiFe2O4 NPs (40.67 emu/g). The Cs-NiFe2O4 NPs indicated higher antibacterial properties than NiFe2O4 NPs and Cs. The minimum inhibitory concentrations of Cs-NiFe2O4 NPs against S. aureus and E. coli were 128 and 256 mg/mL, respectively. Antioxidant activity (evaluated by 2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenging test) for NiFe2O4 NPs and Cs-NiFe2O4 NPs at the concentration of 100 µg/mL were 35% and 42%, respectively. Consequently, the synthesized Cs-NiFe2O4 NPs can be proposed as a viable material for biomedical applications.

Efficient biocatalytic removal and algal detoxification of Direct Blue-15 by the hierarchically structured, high-performance, and recyclable laccase@yttrium phosphate hybrid nanostructures.

From the environmental point of view, azo dye industrial effluent is a major public health concern due to its toxic, carcinogenic, and teratogenic characteristics. On the other hand, using enzyme-based technologies offers a promising systematic and controllable method for removing synthetic dyes from wastewater. In the present study, yttrium (Y3+) phosphate was applied for the synthesis of hybrid nanoparticles (HNPs) consisting of laccase as the green catalyst. When the association of HNPs was fixed by glutaraldehyde (GA), three-dimensional cubic structures with the regular arrangement were provided. GA increased the reusability of the fabricated hybrid nanostructures (HNSs) up to 32 successive cycles. About 85% of Direct Blue-15 was removed after a 4 h-treatment using laccase@YPO4•HNPs and laccase@GA@YPO4•HNSs. The azo dye removal data were well-fitted with a pseudo-second-order model for both types of the prepared HNSs. For the model freshwater green alga Raphidocelis subcapitata, the half maximal effective concentration (EC50) of the dye decreased 10- and 100-fold after the removal with laccase@YPO4•HNPs and laccase@GA@YPO4•HNSs, respectively. GA-treated HNSs (250 U L-1) inhibited the biofilm formation by approximately 78%, 82%, and 79% for Escherichia coli, Staphylococcus aureus, and Bacillus subtilis, respectively. Thus, the fabricated laccase@GA@YPO4•HNSs could be presented as a novel, efficient, and recyclable heterogeneous biocatalyst for wastewater treatment and clean-up.

Design, synthesis, in vitro, and in silico evaluations of benzo[d]imidazole-amide-1,2,3-triazole-N-arylacetamide hybrids as new antidiabetic agents targeting α-glucosidase.

α-Glucosidase as a carbohydrate-hydrolase enzyme is a crucial therapeutic target for type 2 diabetes. In this work, benzo[d]imidazole-amide containing 1,2,3-triazole-N-arylacetamide derivatives 8a-n were synthesized and evaluated for their inhibitory activity against α-glucosidase. In vitro α-glucosidase inhibition assay demonstrated that more than half of the title compounds with IC50 values in the range of 49.0-668.5 µM were more potent than standard inhibitor acarbose (IC50 = 750.0 µM). The most promising inhibitor was N-2-methylphenylacetamid derivative 8c. Kinetic study revealed that compound 8c (Ki = 40.0 µM) is a competitive inhibitor against α-glucosidase. Significantly, molecular docking and molecular dynamics studies on the most potent compound showed that this compound with a proper binding energy interacted with important amino acids of the α-glucosidase active site. Study on cytotoxicity of the most potent compounds 8c, 8e, and 8g demonstrated that these compounds did not show cytotoxic activity against the cancer and normal cell lines MCF-7 and HDF, respectively. Furthermore, the ADMET study predicted that compound 8c is likely to be orally active and non-cytotoxic.

Indole-carbohydrazide linked phenoxy-1,2,3-triazole-N-phenylacetamide derivatives as potent α-glucosidase inhibitors: design, synthesis, in vitro α-glucosidase inhibition, and computational studies.

BACKGROUND: A new series of indole-carbohydrazide-phenoxy-1,2,3-triazole-N-phenylacetamide hybrids 11a-o was designed based on molecular hybridization of the active pharmacophores of the potent α-glucosidase inhibitors. These compounds were synthesized and evaluated against α-glucosidase. METHODS: The 15 various derivatives of indole-carbohydrazide-phenoxy-1,2,3-triazole-N-phenylacetamide scaffold were synthesized, purified, and fully characterized. These derivatives were evaluated against yeast α-glucosidase in vitro and in silico. ADMET properties of the most potent compounds were also predicted. RESULTS: All new derivatives 11a-o (IC50 values = 6.31 ± 0.03-49.89 ± 0.09 µM) are excellent α-glucosidase inhibitors in comparison to acarbose (IC50 value = 750.0 ± 10.0 µM) that was used as a positive control. Representatively, (E)-2-(4-((4-((2-(1H-indole-2-carbonyl)hydrazono)methyl) phenoxy)methyl)-1H-1,2,3-triazol-1-yl)-N-(4-methoxyphenyl)acetamide 11d with IC50 = 6.31 µM against MCF-7 cells, was 118.8-times more potent than acarbose. This compound is an uncompetitive inhibitor against α-glucosidase and showed the lowest binding energy at the active site of this enzyme in comparison to other potent compounds. Furthermore, computational calculations predicted that compound 11d can be an orally active compound. CONCLUSION: According to obtained data, compound 11d can be a valuable lead compound for further structural development and assessments to obtain effective and potent new α-glucosidase inhibitors.

A survey on the stabilizing effect of osmolytes on the ultrasound-irradiated lipase for efficient enzymatic hydrolysis of coconut oil.

The stabilizing effect of some osmolytes including betaine, mannitol, proline, sorbitol, and trehalose (each 0.5 M) was investigated on the ultrasound-irradiated (60 kHz and 138 W, for 240 min) lipase by determination of the enzyme half-life time, evaluation of the enzymatic reaction velocity (Vmax), and hydrolysis of coconut oil for production of lauric acid (the main saturated fatty acid of the oil). The enzyme conformational stability was also assessed by circular dichroism (CD) and fluorescence spectroscopy. The average half-life time of mannitol- and sorbitol-treated lipase under the ultrasound irradiation was 511 ± 3 min and 531 ± 2 min, respectively; 3-fold higher than the unirradiated enzyme. The Vmax value of the ultrasound-treated lipase increased from 100 ± 3 nmol min-1 in the absence of osmolyte to 500 ± 7 nmol min-1 and 500 ± 9 nmol min-1 in the presence of mannitol and sorbitol, respectively. CD and fluorescence spectra indicated that mannitol and sorbitol enhanced the rigidity of the lipase molecular conformational structure, increasing the enzyme stability against the ultrasonic field. The ultrasound-irradiated lipase was then used to hydrolyze coconut oil in the absence or presence of the selected osmolytes, which led to liberate 310 ± 6 mg g-1, 413 ± 7 mg g-1, and 420 ± 4 mg g-1 of lauric acid in the absence or presence of sorbitol and mannitol, respectively. In the absence of an ultrasonic field, the non-osmotically-treated lipase was able to liberate only 211 ± 5 mg g-1 of lauric acid. These promising results indicate that sorbitol and mannitol stabilize the structural conformation of lipase under an ultrasonic field which in turn could improve the enzymatic hydrolysis of coconut oil.

Organic-inorganic hybrid nanoflowers: The known, the unknown, and the future.

Organic-inorganic hybrid nanoflowers (HNFs) are hierarchical flower-shaped microstructures that are assembled by nanoscale petal-like nanosheets composed of both organic and inorganic constituents. Generally, inorganic parts of HNFs are transition metal phosphates and organic components are mostly enzymes and proteins; however, non-protein molecules could be also used as organic phase in some types of newly described HNFs. Recent findings indicate that they are constructed through the coordination between organic and inorganic components. HNFs are mainly used for efficient biocatalysis and highly sensitive biosensing, while they have also some other noteworthy applications such as antimicrobial agents, antigen careers, and delivery platforms for anticancer drugs. It is believed that the high surface-to-volume ratio of HNFs could tackle mass transfer limitations leading to enhance the activity of their organic constituents. The environment-friendly route of synthesis and stabilization of biomolecules upon storage are the advantages of enzyme-based HNFs. In the present review, the focuses are on designs, preparations, formation mechanisms, and remarkable applications of the conventional forms and also magnetic, multi-component, and enzyme-free HNFs. Considering the fact that HNFs are in the early stages of development, the unknown aspects and future directions of research in this field are also discussed.

Production of fucoxanthin from the microalga Tisochrysis lutea in the bubble column photobioreactor applying mass transfer coefficient.

Fucoxanthin is one of the most vital pigments during photosynthesis and is extracted from golden-brown micro-algae such as Tisochrysis lutea. The present study investigates the constant volumetric mass transfer coefficient (kLa), for the first time as the scale-up strategy to change the scale from 500 mL to 2-L cultivation flasks, and 5-L bubble column photobioreactor for fucoxanthin production in T. lutea. The cell density and fucoxanthin content were improved because of through fine aeration, nutrients, and light availability by successful laboratory scale up. Fucoxanthin productivity obtained 21.20, 22.99, and 24.96 mg L-1day-1 for 500 mL, 2-L bottle, and 5-L bubble column photobioreactor, respectively. In addition, the biomass productivity enhanced from 267.5 to 275 and 284 mg L-1day-1 in three mentioned scales, respectively. Eventually, the scale up process for the production of fucoxanthin was succeeded from 500 mL bottle to 5-L photobioreactor using constant (kLa) under laboratory conditions.

Phytocatalytic and cytotoxic activity of the purified laccase from bled resin of Pistacia atlantica Desf.

This study reports an efficient and fast procedure for the purification of laccase (PaL) obtained from the resin of Pistacia atlantica Desf. It was purified by one-step affinity chromatography and showed the specific activity of 393 U/mg with 81.9-fold purification. The molecular weight of PaL was estimated to be approximately 60 kDa using gel electrophoresis SDS-PAGE. Moreover, it depicted diphenolase activity and high affinity towards 2,6-dimethoxy phenol (Km = 10.01 ± 0.5 mM) and syringaldazine (Km = 6.57 ± 0.2 mM) comparing with plant-origin polyphenol oxidases reported in the literature. It should be noted that PaL possessed optimal activity at pH 7.5 and 45 °C. It also remained stable under different conditions of pH (6.5-8.0), temperature (25-45 °C), and when it was exposed to several metal ions. The MTT and flow cytometry assays demonstrated that the enzyme treatment significantly affected growth of HeLa, HepG2, and MDA-MB-231 cells with LC50 values of 4.83 ± 0.02, 61 ± 0.31, and 26.83 ± 0.11 µM after 72 h, respectively. NOVELTY STATEMENT: This is the first attempt to isolate and characterize a new oxidoreductase from the resin of Pistacia atlantica Desf., native species of Iran, to recruit it in cytotoxicity researches. In the purification process by an efficient affinity column (SBA-NH2-GA), the enzyme was eluted promptly with a satisfied yield. The purified laccase exerted higher affinity to diphenolic compounds and pH-thermal stability compared to other plant-derived polyphenol oxidases. The purified enzyme was found to show anti-oxidant capacity and significantly inhibited the growth of cancerous cells in vitro. PaL showed more cytotoxic activity towards HeLa and MDA-MB-231 cells by induction of apoptosis. The cytotoxic activity of the laccase was measured by flow cytometry.

An organic solvent-tolerant lipase of Streptomyces pratensis MV1 with the potential application for enzymatic improvement of n6/n3 ratio in polyunsaturated fatty acids from fenugreek seed oil.

Lipase-catalyzed esterification is an efficient technique in the production of polyunsaturated fatty acid (PUFA) concentrates which are applied for nutrition and health purposes. In this project, a solvent-tolerant lipase from Streptomyces pratensis MV1 was immobilized and purified by a hydrophobic support. The purified lipase revealed enhanced activity and stability towards chemicals, organic solvents, and a broad range of pH values. The production of lipase was enhanced to 7.0 U/mL after optimization by a central composite design. Acylglycerols (AGs) rich in α-linolenic acid (45%, w/w) were produced and a favorable n-6/n-3 free fatty acid (FFA) ratio of 1.1 was achieved in fenugreek seed oil using the immobilized lipase. The ability of S. pratensis lipase in ester synthesis and the improvement of n6/n3 FFA ratio make it a suitable candidate in food production industries.

An efficient and targeted synthetic approach towards new highly substituted 6-amino-pyrazolo[1,5-a]pyrimidines with α-glucosidase inhibitory activity.

In an attempt to find novel α-glucosidase inhibitors, an efficient, straightforward reaction to synthesize a library of fully substituted 6-amino-pyrazolo[1,5-a]pyrimidines 3 has been investigated. Heating a mixture of α-azidochalcones 1 and 3-aminopyrazoles 2 under the mild condition afforded desired compounds with a large substrate scope in good to excellent yields. All obtained products were evaluated as α-glucosidase inhibitors and exhibited excellent potency with IC50 values ranging from 15.2 ± 0.4 µM to 201.3 ± 4.2 µM. Among them, compound 3d was around 50-fold more potent than acarbose (IC50 = 750.0 ± 1.5 µM) as standard inhibitor. Regarding product structures, kinetic study and molecular docking were carried out for two of the most potent ones.

Biomedical and Pharmaceutical-Related Applications of Laccases.

The oxidation of a vast range of phenolic and non-phenolic substrates has been catalyzed by laccases. Given a wide range of substrates, laccases can be applied in different biotechnological applications. The present review was conducted to provide a broad context in pharmaceutical- and biomedical- related applications of laccases for academic and industrial researchers. First, an overview of biological roles of laccases was presented. Furthermore, laccase-mediated strategies for imparting antimicrobial and antioxidant properties to different surfaces were discussed. In this review, laccase-mediated mechanisms for endowing antimicrobial properties were divided into laccase-mediated bio-grafting of phenolic compounds on lignocellulosic fiber, chitosan and catheters, and laccase-catalyzed iodination. Accordingly, a special emphasis was placed on laccase-mediated functionalization for creating antimicrobials, particularly chitosan-based wound dressings. Additionally, oxidative bio-grafting and oxidative polymerization were described as the two main laccase-catalyzed reactions for imparting antioxidant properties. Recent laccase-related studies were also summarized regarding the synthesis of antibacterial and antiproliferative agents and the degradation of pharmaceuticals and personal care products.

Purification and study of anti-cancer effects of Serratia marcescens serralysin.

BACKGROUND AND OBJECTIVES: Serralysin is an extracellular metalloprotease from Serratia marcescens which has been the subject of extensive biological investigations. The goal of this study was to extract and purify serralysin from S. marcescens and to investigate its cytotoxic activity on the colorectal cancer cell line. MATERIALS AND METHODS: The presence of the serralysin gene was confirmed using PCR. The supernatant of bacterial culture was collected and precipitated using ammonium sulfate. The precipitated protein was dialyzed and subjected to ion exchange chromatography for further purification. Casein assay and skim milk assay was used to confirm the enzymatic activity. SDS-PAGE was used to visualize the presence of serralysin. Metalloprotease inhibition activity was performed using 50 mM EDTA. Cytotoxic activity of serralysin was assessed on MTT assay. RESULTS: The PCR product corresponding to serralysin was estimated to be approximately 1500 bp. A transparent zone around the bacterial colonies on skim milk agar and casein digestion confirmed the proteolytic activity of serralysin. A 52 kDa band in SDS-PAGE corresponding to serralysin was observed before and after purification processes. MTT assay showed IC50 values 24.78 µg/ml and 19.16 µg/ml after 24 h and 48 h exposure of Caco-2 cells to serralysin, respectively. CONCLUSION: Our results showed that native serralysin has anticancer potential and may be a candidate for further pharmaceutical research and development. Further in vivo and in vitro mechanistic studies are suggested to confirm the biological activities.

PerioVax3, a key antigenic determinant with immunoprotective potential against periodontal pathogen.

Treponema (T.) denticola is one of the key etiological agents in the development of periodontitis. The major outer sheath protein (Msp) of T. denticola has been shown to mediate pathogenesis and to facilitate adhesion of T. denticola to mucosal surfaces. This study aimed to find short polypeptides in the amino acid sequence of Msp which may be immunogenic and might elicit protective antisera against T. denticola. The complete msp sequence was divided into six fragments and the corresponding genes were cloned and expressed. Antisera against the polypeptides were raised in rabbits and fragment 3 (F3), hereinafter called PerioVax3 was the most potent fragment of the Msp in terms of yielding high titer antiserum. An adhesion assay was done to examine the inhibitory effects of antisera on the attachment of T. denticola to human gingival fibroblasts (HGFs) and human fibronectin. Antiserum against PerioVax3 significantly inhibited attachment of T. denticola to the substratum. Also, antiserum against PerioVax3 inhibited detachment of HGFs upon T. denticola exposure. To begin examining the clinical relevance of this work, blood samples from 12 sever periodontitis patients were collected and the sera were used in western blotting against the recombinant polypeptides. Periodontitis patient antisera exclusively detected PerioVax3 in western blotting. The data suggest that PerioVax3 carries epitopes that may trigger humoral immunity against T. denticola, which may protect against its adhesion functions. The complexity of periodontitis suggests that PerioVax3 may be considered for testing as a component of an experimental multivalent periodontal vaccine in further preclinical and clinical studies.

2,4-Disubstituted Quinazoline Derivatives Act as Inducers of Tubulin Polymerization: Synthesis and Cytotoxicity.

BACKGROUND: During last recent years number of anti-tubulin agents were introduced for treatment of diverse kind of cancer. Despite of their potential in treatment of cancer, drug resistance and adverse toxicity such as peripheral neuropathy are some of the negative criteria of anti-tubulin agents. METHODS: Twenty seven quinazoline derivatives were synthesized using a multicomponent reaction. The cytotoxicity of compounds 1-27 was tested in SRB assays employing five different human tumor cell lines. Effect of two of active compounds on tubulin polymerization was also checked using a commercially available assay kit. Molecular modelling studies were also performed using autodock tools software. RESULTS: SRB assays showed that compounds 2, 9, 16 and 26, being highly cytotoxic with IC50 values ranging between 2.1 and 14.3µM. The possible mode of action of compounds, 2, 9, 16 and 26, and the taxol binding site of the protein tubulin, an important goal for antimitotic drugs, was also studied by molecular docking, which showed reasonable interactions with tubulin active site, followed by investigation of the effects of compounds 9 and 16 on the polymerization of tubulin. The results showed the tested compounds to be highly active as inducers of tubulin polymerization. CONCLUSION: Altogether, with respect to obtained results, it is attractive and beneficial to further investigation on quinazoline scaffold as antimitotic agents.

Novel trastuzumab-DM1 conjugate: Synthesis and bio-evaluation.

Antibody-drug conjugates are now of considerable interest and are recommended for the treatment of cancers. Linkers are having a crucial role in potency and efficacy of these drugs. Herein, for the first time, we have used a water-soluble poly-ethylene glycol based linker (succinimidyl-[(N-maleimido propionamido)-diethyleneglycol] [SM(PEG)2]) for lysine amide coupling of DM1 drug to trastuzumab considering evaluation of the effect of using a hydrophilic linker on physicochemical and biological properties of the resulting conjugate in comparison to the conjugate containing succinimidyl 4-(N-maleimidomethyl) cyclohexane-1-carboxylate (SMCC) linker, which has a relative hydrophobic nature. The physicochemical properties of synthesized conjugates were investigated in terms of drug to antibody ratio, size variants and free drug quantities. In vitro biological activity of trastuzumab-DM1 conjugates was assessed on breast cancer cell lines expressing different levels of HER2 using binding affinity, antiproliferative, apoptosis, and antibody-dependent cell-mediated cytotoxicity (ADCC) assays. Synthesized conjugate containing hydrophilic linker, showed higher drug to antibody ratio, no aggregated form and higher cellular toxicity in comparison to SMCC bearing conjugate. Binding affinity and ADCC potential of conjugates was not affected upon the usage of hydrophilic linker. In conclusion, application of SM(PEG)2 for coupling of DM1 to trastuzumab enhance desirable characteristics of the resulting conjugate.

Enhancing analgesic and anti-inflammatory effects of capsaicin when loaded into olive oil nanoemulsion: An in vivo study.

Topical preparations of capsaicin, the major pungent ingredient of hot pepper, are being used for management of pain and inflammatory disorders. Purpose of this study was to use nanoemulsion as an effective topical drug carrier for in vivo delivery of capsaicin. An oil-in-water nanoemulsion containing capsaicin was prepared by spontaneous emulsification method. Optimized formulation showed a median droplet diameter (d50) of 13-14 nm and was stable for more than 8 months in room and harsh temperature (i.e. 4 and 45 °C). The nanoemulsion was then formulated into topical cream and gel to compare its efficacy and safety profiles with conventional cream of capsaicin. Skin irritation study showed that topical application of capsaicin nanoemulsion was safe and no sign of edema and erythema was observed. The preparation significantly decreased inflammation of rats paw edema compared to the commercial cream and control group, especially in 2nd and 3rd hours of the test. Also, pretreated rats with capsaicin nanoemulsion gel showed very good resistance to the pain caused by heat stimulus. In total, the selected nanoemulsion showed great potential as carrier for topical delivery of capsaicin for improving its analgesic and anti-inflammatory effects. It was also found that the topical gel outperforms the topical cream as dosage form for the nanoemulsion.

Efficient Keratinolysis of Poultry Feather Waste by the Halotolerant Keratinase from Salicola Marasensis.

Sustainable development in the bio-treatment of large-scale biomass bulks requires high performance enzymes adapted to extreme conditions. An extracellular keratinolytic extract was obtained from the culture broth of a halotolerant strain of Salicola marasensis. Keratin hydrolyzing activity of the concentrated enzyme extract was observed on a 100 mg of pretreated feather waste. The concentrated enzyme was able to hydrolyze the poultry feathers by 25% after 12 h incubation. The bio-waste material was optimally hydrolyzed at pH 9 and temperature of 40 °C. Among reductants, 1,4-dithiothreitol, L-cysteine, 2-mercaptoethanol, glutathione, and sodium sulfate showed the most remarkable effect on the bio-waste keratinolysis, while the tested surfactants and urea had no significant effect on the keratinolytic activity. Hexane and hexadecane indicated strong effect on keratinase activity and bio-treatment in the presence of 1-butyl-3-methylimidazolium hexafluorophosphate ([BMIM][PF6]) as a hydrophobic ionic liquid resulted in a maximal of 80% extraction yield of soluble proteins from feathers. Considering the stability of the extracellular keratinolytic content in [BMIM][PF6], the observed keratinase activity was noteworthy suggesting that the secreted enzyme may contribute to the bioconversion of feather wastes.