Caffeic Acid Phenethyl Ester Loaded Electrospun Nanofibers for Wound Dressing Application.

Electrospinning is an advantageous method with a wide usage area, which enables the production of materials consisting of nano-thickness fibers. In this study, caffeic acid phenethyl ester (CAPE) molecule was loaded onto the poly(lactic-co-glycolic acid) (PLGA) nanofibers and obtained nanofibers were physicochemically and biologically investigated for the first time in the literature. The existence of CAPE molecules, loaded on PLGA membranes by dropping and spraying methods, was evaluated by a comparative investigation of Fourier-transform infrared (FTIR) spectra and X-Ray diffraction (XRD) patterns. Fiber morphology of the membranes was investigated by scanning electron microscope (SEM). CAPE release and swelling behaviors of the membranes were studied in vitro. The radical scavenging activity of CAPE-loaded wound dressing materials was determined by using an antioxidant assay. The antimicrobial properties of PLGA and CAPE-loaded PLGA membranes were evaluated against S. aureus, P. aeruginosa and C. albicans strains by the time-kill method. The biocompatibility study of the obtained CAPE-loaded fibers conducted on human fibroblast cell line and wound healing promoting effect of the fibers was investigated in vitro scratch assay. The results show that CAPE-loaded PLGA membranes are highly antimicrobial against all strains used in the experiment. Additionally, the results show that they are biocompatible and have wound healing properties on human fibroblasts.

Detection of bacteria using antimicrobial polymer derived via ring-opening metathesis (romp) pathway.

There is growing interest in the detection of bacteria in consumables, for example, in the food and water sectors. In this study, the aim was to produce a polymer-based bacteria biosensor via ROMP (ring opening metathesis polymerization). In the first part of the study, block and random copolymers were synthesized, and their biocidal activities were tested on the glass surface. Interdigitated electrode arrays coated with the polymers possessing the highest activity were used to screen the affinity towards different bacterial strains by monitoring impedance variations in real-time. The polymer-coated electrode could detect gram-positive and gram-negative bacteria strains at a concentration of 107 cfu/mL. The results show that ROMP-based polymer offers bacterial detection and can be used in developing biosensor devices for efficiently detecting pathogenic bacteria.

Assessment of the exclusion potential of suspects by using microbial signature in sexual assault cases: A scenario-based experimental study.

Sexual assault offences are one of the most serious crimes committed against a person, typically rank only second to homicide, and represent one of the major challenges in forensic sciences. In some cases of sexual assault, there may be more than one suspect and the analysis of the biological evidence with currently available methods such as human DNA analysis may not yield results. In this study using the designed experimental model (with different experimental scenarios that can be designed), it was aimed to investigate the effectiveness of the microbiome profile for the identification of the offender by comparing the microbiome structures of the suspects' saliva samples with the mixed samples on the victim (saliva transmitted on breast skin) within the first 48 h after a sexual assault. For this purpose, a total of 44 samples was collected from four healthy females and four healthy males aged 20-50 years. Microbiome profiles of 44 samples in four groups containing saliva, breast skin and mixed samples were determined with the IIlumina HiSeq platform. Differentiation between samples were calculated by beta-diversity analysis methods by using QIIME software (v1.80). To compare the differentiation among samples and groups, unweighted UniFrac distance values were applied. Eight dominant microbial genera accounted for 86.15% of the total bacterial population in male saliva samples and were composed of Fusobacterium, Haemophilus, Neisseria, Porphyromonas, Prevotella, Rothia, Streptococcus and Veillonella. These genera constituted 76.72% of the bacterial population in mixed samples, whereas they constituted 34.40% of the bacterial population in the breast skin samples. Results of this study show that bacterial DNA in saliva can be recovered from saliva transmitted breast skin within at least 48 h. In conclusion, it has been found that examination of the microbiota of the saliva transmitted to breast skin of a sexual assault victim as a forensic tool may have the potential to determine the offender of the incident among the suspects or to reduce the number of suspects. Supporting the results of this study with further studies using parameters such as different case models, different body regions, larger time periods and a higher number of participants will be beneficial to draw accurate conclusion of the judicial case.

In Vitro Evaluation of Immunogenicity of Recombinant OMP25 Protein Obtained from Endemic Brucella abortus Biovar 3 as Vaccine Candidate Molecule Against Animal Brucellosis.

BACKGROUND: Brucellosis is a zoonotic disease that causes serious economic losses due to factors, such as miscarriages and decreased milk yield in animals. Existing live vaccines have some disadvantages, so effective vaccines need to be developed with new technological approaches. OBJECTIVE: The primary objectives of this study were the expression and purification of recombinant Omp25 fusion protein from B. abortus, and the evaluation of the effect of the Omp25 protein on cell viability and inflammatory response. METHODS: The omp25 gene region was amplified by a polymerase chain reaction and cloned into a Pet102/D-TOPO expression vector. The protein expression was carried out using the prokaryotic expression system. The recombinant Omp25 protein was purified with affinity chromatography followed by GPC (Gel Permeation Chromatography). The MTS assay and cytokine-release measurements were carried out to evaluate cell viability and inflammatory response, respectively. RESULTS: It was determined that doses of the recombinant Omp25 protein greater than 0.1 µg/mL are toxic to RAW cells. Doses of 1 µg/mL and lower significantly increased inflammation due to Nitric Oxide (NO) levels. ELISA results showed that IFN-γ was produced in stimulated RAW 264.7 cells at a dose that did not affect the viability (0.05 µg/mL). However, IL-12, which is known to have a dual role in the activation of macrophages, did not show a statistically significant difference at the same dose. CONCLUSION: Studies on cell viability and Th1-related cytokine release suggest Omp25 protein to be a promising candidate molecule for vaccine development.

Antioxidant Activity and Hemocompatibility Study of Quercetin Loaded Plga Nanoparticles.

Quercetin (QU) is an important flavonoid compound presenting lots of biological activities, but its application has been limited due to its low aqueous solubility and instability. In this study, conducted to improve these properties of the quercetin, quercetin-encapsulated PLGA nanoparticles were prepared, characterized, and evaluated for antioxidant and hemolytic activity. Nanoparticles were produced by single emulsion solvent evaporation method. Four different process parameters initial QU amount, PVA concentration, PVA volume, and initial PLGA amount were investigated to obtain the nanoparticles which have minimum particle size and maximum entrapment efficiency. Synthesized nanoparticles were evaluated for particle size, entrapment efficiency, and reaction yield. Additionally, antioxidant properties and in-vitro hemolytic activity of quercetin loaded nanoparticles with different particle size were also evaluated for the first time in the literature. The antioxidant activity results showed that nanoparticles have different antioxidant activity, depending on the amount of quercetin release from nanoparticles at different particle sizes. The hemolytic activity results show that all nanoparticles exhibited favorable compatibility to red blood cells and no significant hemolytic effect was observed.

Cytotoxic and mutagenic potential of juglone: a comparison of free and nano-encapsulated form.

Despite its evidenced beneficial herbicidal, antibacterial, antiviral, antifungal, and antioxidant effects, the application of juglone (5-hydroxy-1,4,-naphthoquinone) is limited due to its low water solubility and allelopathic and toxic effects. In recent years, research has aimed to overcome these limitations by increasing its solubility and controlling its release through nanoparticular systems. This is the first study to have synthesised and characterised juglone-loaded polymeric nanoparticles and compared them with free juglone for cytotoxicity in mouse (L929 fibroblasts) and alfalfa cells and for mutagenic potential in Salmonella typhimurium TA98/100. Mouse and plant cells treated with free and nano-encapsulated juglone showed a decrease in cell viability in a dose and time-dependent manner, but this effect was significantly lower with the nano-encapsulated form at lower doses. In the TA98 strain with S9, nano-encapsulated juglone did not exhibit mutagenic effects, unlike the free form. Since all results show that juglone encapsulation with polymeric nanoparticles reduced the toxic and mutagenic effects, it has a promising potential to be applied in medicine, food safety, and agriculture.

An insight into the epitope-based peptide vaccine design strategy and studies against COVID-19.

SARS-CoV-2 is a new member of the coronavirus family and caused the pandemic of coronavirus disease 2019 (COVID-19) in 2020. It is crucial to design and produce an effective vaccine for the prevention of rapid transmission and possible deaths wcaused by the disease. Although intensive work and research are being carried out all over the world to develop a vaccine, an effective and approved formulation that can prevent the infection and limit the outbreak has not been announced yet. Among all types of vaccines, epitope-based peptide vaccines outshine with their low-cost production, easy modification in the structure, and safety. In this review, vaccine studies against COVID-19 have been summarized and detailed information about the epitope-based peptide vaccines against COVID-19 has been provided. We have not only compared the peptide vaccine with other types of vaccines but also presented comprehensive literature information about development steps for an effective and protective formulation to give an insight into on-going peptide vaccine studies against SARS-CoV-2.

The battle against biofilm infections: juglone loaded nanoparticles as an anticandidal agent.

In this study, juglone nanoparticles were prepared by single emulsion solvent evaporation method and their effect against Candida albicans biofilm was investigated in comparison with the free juglone and Fluconazole by performing XTT, crystal violet, standard plate count, confocal microscopy and membrane depolarization analyses. Juglone nanoparticles and free juglone were found to inhibit biofilm formation and pre-established biofilms (98-100%) at all doses tested, whereas Fluconazole did not cause a significant inhibition, even at the highest dose applied, especially against pre-established biofilms. Membrane depolarization analysis showed that free juglone and juglone loaded nanoparticles were effective on C. albicans membrane structure and have fluorescence quenching effect on DiSC3(5). It is extremely important that the antibiofilm activity of the juglone nanoparticles is similar to that of the juglone used at the same concentration, since similar effect is provided by using less active substance due to controlled release. Accordingly, it can easily be said that juglone loaded nanoparticles are much more effective in the formation and elimination of C. albicans biofilm than the free juglone and Fluconazole.

Quercetin-loaded nanoparticles enhance cytotoxicity and antioxidant activity on C6 glioma cells.

Quercetin (Qu) is a natural flavonoid present in many commonly consumed food items. The dietary phytochemical quercetin prevents tumor proliferation and is a potent therapeutic cancer agent. The purpose of this study was to synthesize and characterize quercetin-loaded poly(lactic-co-glycolic acid) nanoparticles (Qu1NP, Qu2NP, and Qu3NP) with different size and encapsulation properties and to evaluate their in vitro activity on C6 glioma cells. Nanoparticles were synthesized by single emulsion solvent evaporation method. Then, particle size, zeta potential, polydispersity index and encapsulation efficiency of nanoparticles were determined. Particle size of Qu1NP, Qu2NP, and Qu3NPs were determined as 215.2 ± 6.2, 282.3 ± 7.9, and 584.5 ± 15.2 nm respectively. Treating C6 glioma cells with all nanoparticle formulations effectively inhibited the cell proliferation. Qu1NPs were showed the lowest IC50 value in 48 h with 29.9 µg/ml and achieved higher cellular uptake among other nanoparticles and Qu. Additionally, 48-h treatment with Qu1NPs significantly decreased MDA level (14.90 nmol/µg protein) on C6 glioma cells which is related to reduced oxidative stress in cells. Findings of this study revealed that quercetin's cellular uptake and anti-oxidant activity is improved by small-sized Qu1NPs in C6 glioma cells.

Enhanced antibacterial and antiparasitic activity of multifunctional polymeric nanoparticles.

Due to the resistance to drugs, studies involving the combination and controlled release of different agents are gradually increasing. In this study, two different active ingredients, known to have antibacterial and antiparasitic activities, were encapsulated into single polymeric nanoparticles. After co-encapsulation their antibacterial and antileishmanial activity was enhanced approximately 5 and 250 times, respectively. Antibacterial and antileishmanial activities of caffeic acid phenethyl ester and juglone loaded, multifunctional nanoformulations (CJ4-CJ6-CJ8) were also evaluated for the first time in the literature comparatively with their combined free formulations. The antibacterial activity of the multifunctional nanoformulation (CJ8) were found to have a much higher activity (MIC values 6.25 and 12.5 µg ml-1 for S. aureus and E. coli, respectively) than all other formulations. Similar efficacy for CJ8 was obtained in the antiparasitic study against the Leishmania promastigotes and the IC50 was reduced to 0.1263 µg ml-1. The high activity of multifunctional nanoparticles is not only due to the synergistic effect of the active molecules but also by the encapsulation into polymeric nanoparticles. Therefore, it has been shown in the literature for the first time that the biological activity of molecules whose activity is increased by the synergistic effect can be improved with nanosystems.

Comparative evaluation of hesperetin loaded nanoparticles for anticancer activity against C6 glioma cancer cells.

The aim of this study was to evaluate anti-cancer properties of hesperetin (Hsp) and hesperetin-loaded poly(lactic-co-glycolic acid) nanoparticles (HspNPs) for glioblastoma treatment. Nanoparticles prepared by single emulsion method had a size of less than 300 nm with 70.7 ± 3.9% reaction yield and 26.4 ± 1.1% Hsp loading capacity. Treatment of C6 glioma cells with HspNPs for 24 and 48 h resulted in dose- and time-dependent decrease in cell viability, with approximate IC50 of 28 and 21 µg/mL, respectively (p = .036 for 24 h, p = .025 for 48 h). The percentage of PCNA positive cells decreased to 20% and 10%, respectively, for Hsp- and HspNP-treated cells at concentration of 100 µg/mL. Treatment with increasing concentrations of HspNPs (25, 50, 75 and 100 µg/mL) resulted in 9.1-, 7-, 12.5- and 12.7-fold in increase in apoptotic cell number. Optimum doses of Hsp and HspNPs were found to increase oxidative damage in C6 glioma cells. MDA levels, an indicator of lipid peroxidation, were found to be significantly elevated at 75 and 100 µg/mL exposure concentration of HspNPs with (p = .002) and (p = .018), respectively for 48-h treatment. The results obtained with this study showed biocompatible polymeric nanoparticle systems has great advantages to enhance anti-cancer activity and poor solubility of therapeutic agents. Overall our findings suggest that Hsp-loaded PLGA nanoparticle systems showed significant anti-cancer activity and HspNPs could be used as promising novel anti-cancer agent.

Chemical composition of the essential oils of Centaurea tomentella Hand.-Mazz. and C. haussknechtii Boiss. (Asteraceae) collected wild in Turkey and their activity on microorganisms affecting historical art craft.

In the present study the chemical composition of the essential oils from aerial parts of Centaurea tomentella Hand.-Mazz. and C. haussknechtii Boiss. collected in Turkey was evaluated by GC and GC-MS. The main components of C. tomentella L. were hexadecanoic acid (19.7%), caryophyllene oxide (6.6%) and spathulenol (4.8%) whereas C. haussknechtii was rich in hexadecanoic acid (26.2%), (Z,Z)-9,12-octadecadienoic acid (19.3%), heptacosane (5.3%) and nonacosane (5.1%). Antibacterial and antifungal activities against some microorganisms infesting historical art craft, were also determined.

Synthesis of hesperetin-loaded PLGA nanoparticles by two different experimental design methods and biological evaluation of optimized nanoparticles.

Hesperetin was effectively encapsulated into poly (d,l-lactic-co-glycolic acid) nanoparticles by using experimental design methods. A seven-factor Plackett-Burman design was used in order to determine the major process parameters. A significant linear equation, which shows the effect of each process parameter on encapsulation efficiency was developed, and then the most effective factors were determined. Further investigation and optimization was carried out by applying the three-factor three-level Box-Behnken design. Significant second-order mathematical models were developed by regression analysis of the experimental data for both responses: encapsulation efficiency and nanoparticle size. The two step experimental design allowed the synthesis of the desired nanoparticle formulations with maximum encapsulation efficiency (80.5 ± 4.9%) and minimum particle size (260.2 ± 16.5 nm) at optimum process conditions: 0.5% polyvinyl alcohol (PVA) concentration, 5.13 water:organic phase ratio, and 3.59 ml min-1 flow rate of the emulsified solution into 0.1% PVA. Furthermore, the biological activity of these optimized nanoparticles were determined with antimicrobial activity and cytotoxicity studies; results were then compared to the free hesperetin. The cytotoxicity result revealed that hesperetin and hesperetin-loaded nanoparticles were biocompatible with normal cell line L929 fibroblast cells up to 184.83 and 190.88 µg ml-1 for 24 h, and up to 133.24 and 134.80 µg ml-1 for 48 h, respectively. In the antimicrobial study, the optimized nanoparticle showed inhibition activity (minimal inhibitory concentration (MIC) values were 125 µg ml-1 for Escherichia coli, and 200 µg ml-1 for Staphylococcus aureus), while the free hesperetin did not demonstrate activity in both strains (MIC value >200 µg ml-1). These in vitro results may provide useful information for the investigation of hesperetin-loaded nanoparticles in diagnostic and therapeutic applications.

Assessment of the Antigenotoxic Activity of Poly(d,l-lactic- co-glycolic acid) Nanoparticles Loaded with Caffeic Acid Phenethyl Ester Using the Ames Salmonella/Microsome Assay.

In the present study, the antigenotoxic activity of poly(d,l-lactic- co-glycolic acid) (PLGA) nanoparticles (NPs) loaded with caffeic acid phenethyl ester (CAPE) was investigated in comparison to free CAPE using the Ames Salmonella/microsome assay. Additionally, to elucidate the impacts of the type of solvent effect on antigenotoxic activity, the following systems were tested: CAPE in water (poor solvent), ethyl alcohol (good solvent), and PLGA NPs (unknown). The effect of the NP system on solubility was investigated for the first time by assessing the antigenotoxic potential. In this study, the CAPE/PLGA NPs were synthesized using an oil-in-water (o/w) single-emulsion solvent evaporation method with an average size of 206.2 ± 1.2 nm, ζ potential of -19.8 ± 2.5 mV, encapsulation efficiency of 87.2 ± 2.5%, and drug loading of 53.3 ± 1.8%. According to the results of the antigenotoxic activity, the highest antimutagenic activity in both applied strains was found for CAPE in ethanol, and the lowest activity was detected for CAPE in water. Our study has shown that NP systems exhibit high antigenotoxic activity, which is similar to the results of CAPE dissolved in ethanol. These results have shown that NP systems increase biological activity of hydrophobic substances by increasing their solubility and that the use of PLGA instead of organic solvents in drug production may provide an increase in their medical utility.

Enhancement of Antifungal Activity of Juglone (5-Hydroxy-1,4-naphthoquinone) Using a Poly(d,l-lactic-co-glycolic acid) (PLGA) Nanoparticle System.

This study aimed to synthesize and characterize juglone-entrapped poly(d,l-lactic-co-glycolic acid) (PLGA) nanoparticles and compare the antifungal properties of free juglone with its PLGA nanoparticle formulation for the first time. The juglone-loaded nanoparticles prepared using the oil-in-water (o/w) single-emulsion solvent evaporation method were characterized by the reaction yield (RY), encapsulation efficiency (EE), polydispersity index (PDI), particle size, zeta potential (ZP), FT-IR, and in vitro release properties and evaluated for their morphological features using SEM. The nanoparticle formulation had size, RY, ZP, EE, and PDI values of 212 nm, 66.91 ± 2.4%, -16.3 ± 0.7 mV, 70.66 ± 3.1%, and 0.083 ± 0.024, respectively. In vitro release showed a triphasic pattern with initial burst followed by sustained release and dormant phase over the study period, releasing about 72.8% in total after 42 days. The antifungal studies against Aspergillus flavus, Candida albicans, and Fusarium spp. using agar dilution and top agar dilution methods indicated that the juglone-encapsulated nanoparticle was more effective than free juglone. This study showed that the top agar method, which was applied for the first time on antifungal activity, is more suitable for the nanoparticular system based on sustained release. Therefore, PLGA nanoparticle formulations may be an important tool for application in many areas for the effective and beneficial use of hydrophobic compounds such as juglone.

Comparative evaluation of antibacterial activity of caffeic acid phenethyl ester and PLGA nanoparticle formulation by different methods.

The aim of the present study was to evaluate the antimicrobial activity of nanoparticle and free formulations of the CAPE compound using different methods and comparing the results in the literature for the first time. In parallel with this purpose, encapsulation of CAPE with the PLGA nanoparticle system (CAPE-PLGA-NPs) and characterization of nanoparticles were carried out. Afterwards, antimicrobial activity of free CAPE and CAPE-PLGA-NPs was determined using agar well diffusion, disk diffusion, broth microdilution and reduction percentage methods. P. aeroginosa, E. coli, S. aureus and methicillin-resistant S. aureus (MRSA) were chosen as model bacteria since they have different cell wall structures. CAPE-PLGA-NPs within the range of 214.0 ± 8.80 nm particle size and with an encapsulation efficiency of 91.59 ± 4.97% were prepared using the oil-in-water (o-w) single-emulsion solvent evaporation method. The microbiological results indicated that free CAPE did not have any antimicrobial activity in any of the applied methods whereas CAPE-PLGA-NPs had significant antimicrobial activity in both broth dilution and reduction percentage methods. CAPE-PLGA-NPs showed moderate antimicrobial activity against S. aureus and MRSA strains particularly in hourly measurements at 30.63 and 61.25 µg ml(-1) concentrations (both p < 0.05), whereas they failed to show antimicrobial activity against Gram-negative bacteria (P. aeroginosa and E. coli, p > 0.05). In the reduction percentage method, in which the highest results of antimicrobial activity were obtained, it was observed that the antimicrobial effect on S. aureus was more long-standing (3 days) and higher in reduction percentage (over 90%). The appearance of antibacterial activity of CAPE-PLGA-NPs may be related to higher penetration into cells due to low solubility of free CAPE in the aqueous medium. Additionally, the biocompatible and biodegradable PLGA nanoparticles could be an alternative to solvents such as ethanol, methanol or DMSO. Consequently, obtained results show that the method of selection is extremely important and will influence the results. Thus, broth microdilution and reduction percentage methods can be recommended as reliable and useful screening methods for determination of antimicrobial activity of PLGA nanoparticle formulations used particularly in drug delivery systems compared to both agar well and disk diffusion methods.

Isolation of a flavonoid, apigenin 7-O-glucoside, from Mentha longifolia (L.) Hudson subspecies longifolia and its genotoxic potency.

Mentha is a medicinal and aromatic plant belonging to the Lamiaceae family, which is widely used in food, flavor, cosmetic and pharmaceutical industries. Recently, it has been found that the use of Mentha as a pharmaceutical source is based on its phytochemical constituents that have far been identified as tannins, saponins, phenolic acids and flavonoids. This study was designed to evaluate the mutagenic and antimutagenic activities of apigenin 7-O-glucoside (A7G), a flavonoid isolated from Mentha longifolia (L.) Hudson subspecies longifolia (ML). The possible antimutagenic potential of A7G was examined against mutagens ethyl methanesulfonate and acridine in an eukaryotic cell system Saccharomyces cerevisiae and sodium azide in Salmonella typhimurium TA1535 and 9-aminoacridine in S. typhimurium TA1537. According to our findings, any concentrations of the A7G used did not show mutagenic activity but exerted strong antimutagenic activities at tested concentrations. The inhibition rates for the Ames test ranged from 27.2% (S. typhimurium TA1535: 0.4 µM/plate) to 91.1% (S. typhimurium TA1537: 0.2 µM/plate) and for the yeast deletion assay from 4% to 57.7%. This genotoxicological study suggests that a flavonoid from ML owing to antimutagenic properties is of great pharmacological importance and might be beneficial to industries producing food additives, cosmetics and pharmaceuticals products.