Superparamagnetic iron oxide nanoparticles (spions) modified with sarcosine oxidase-enzymatic activity analysis by sds-page.

Sarcosine oxidase (SOX) is an enzyme that catalyzes the oxidative demethylation of sarcosine with the glycine as a product and is physiologically active in human body and other mammals. However prostate cancer cells have a high expression of sarcosine. In this study the superparamagnetic iron oxide nanoparticles (SPIONs) were prepared and their surface was modified with gold nanoparticles (AuNPs). These AuNPs were modified with chitosan (CS) and SOX. Obtained AuNPs were characterized by physicochemical methods, such as dynamic light scattering or spectrophotometry, where the pseudo-peroxidase activity of the AuNPs was used. Hydrogen peroxide decomposes because of the pseudo-peroxidase activity with the appearance of a blue coloration of 3,3',5,5'-tetramethylbenzidine (TMB). For the analysis of AuNPs enzymatic activity the SDSPAGE with silver staining has been used. Gels (7.5%) were prepared using acrylamide stock solution 30% (m/V) with bisacrylamide 1% (m/V). Separating gel contained: acrylamide 7.5% (m/V), bisacrylamide 0.5% (m/V), 0.4 M Tris/HCl, 0.1% (m/V) sodium dodecyl sulfate (SDS), pH 8.8. Stacking gel contained: 4.5% acrylamide (m/V), 0.15% bisacrylamide (m/V), 0.1% SDS (m/V), 0.1M Tris/HCl, pH 6.8. Nanoconstructs were diluted 2:1 with a loading buffer (PLB Max). Each well contained 15 μl of the diluted solutions. Electrophoretic measuring conditions were: 120 V, 1.5 hours in a running buffer (24mM Tris, 0.2M glycine and 3mM SDS). After measurement the gel was stained with silver, scanned and evaluated by Colortest in the laboratory system Qinslab. The SPIONs or AuNPs cannot be detected themselves alone using SDS-PAGE, therefore this method served as a confirmation, that all parts of the nanoconstruct are connected and we are able to analyze them and use them for other research or possible diagnostic purposes

Antioxidant activity of silver nanoparticles prepared by green synthesis.

At present, great attention is given to silver nanoparticles (AgNPs), which thanks to its unique properties, such as good electric conductivity, photoelectrochemical activity and antimicrobial activity are widely used. Green synthesis of nanoparticles uses biological molecules from living organisms. Biological extracts may contain molecules which exhibit significant antibacterial, antiviral and cytotoxic effects. The aim of this work was to study the diverse AgNPs synthesized from 10 different types of plant extracts. Extracts from dried plants (0.5g/25mL 18 MO water) were prepared at 70°C by heating for 20 minutes. After filtration, the leachates were mixed in the 1:1 ratio with 0.1 M AgNO3 and allowed to stir at room temperature for 18 hours. They were then mixed in the 1:1 ratio with methanol, shaken for 5 minutes on the rotary mixer, and centrifuged at 12,000g for 30 minutes. After removing the supernatant, the pellet was dried at 60°C for 24 hours. After weighing, the purified AgNPs were dissolved in 18 MO water. AgNPs were yellow, orange and brown. The extraction efficiency was monitored by organic solvents (methanol, ethanol, acetone, propanol). The yields of AgNPs ranged from 15 to 5%, and the most suitable solvent was methanol with an average AgNPs yield of about 10%.AgNPs were characterized spectrally (spectral maxima were in the range of 300-500 nm) by determining the zeta potential and the size of nanoparticles (30-80 nm). Furthermore, antioxidant activity was monitored using ABTS (670 nm), DPPH (517 nm), and FRAP (595 nm) methods. Two methods (ABTS and DPPH) based on the elimination of synthetic radicals and the FRAP method based on the reduction of iron complexes were used to monitor the antioxidant activity. Antioxidation assays were evaluated using calibration curve equations in which the standard was gallic acid. The results for the ABTS and DPPH methods were also expressed as percentage of inhibition of the radicals and in the FRAP method as percentage of reduction activity. The results were calculated using the ABTS method in the range (25.9-84.9%), in the DPPH method (19.2-86.6%) and in the FRAP method (8.5-93.2%). Most AgNPs prepared by green synthesis showed significant antioxidant activity

Melatonin Regulates Oxidative Stress Initiated by Freund’s Complete Adjuvant

Melatonin is a hormone with strong antioxidant properties. In this experiment, Freund’s complete adjuvant was used as a stressogenic substance given to laboratory outbred mice, whereas melatonin was investigated as a protectant against the stressogenic effect. Levels of low molecular weight antioxidants, thiobarbituric acid reactive substances, and tumor necrosis factor α and activity of glutathione reductase were determined in blood from the animals. Surprisingly, melatonin was not involved in direct regulation of antioxidants, thiobarbituric acid reactive substances and tumor necrosis factor α. On the other hand, melatonin regulated glutathione reductase activity. We can conclude on regulation of metabolism caused by melatonin in the model. The effect was more important than the expected regulation of immunity and basal oxidative homeostasis

G-Quadruplexes as Sensing Probes

Guanine-rich sequences of DNA are able to create tetrastranded structures known as G-quadruplexes; they are formed by the stacking of planar G-quartets composed of four guanines paired by Hoogsteen hydrogen bonding. G-quadruplexes act as ligands for metal ions and aptamers for various molecules. Interestingly, the G-quadruplexes form a complex with anionic porphyrin hemin and exhibit peroxidase-like activity. This review focuses on overview of sensing techniques based on G-quadruplex complexes with anionic porphyrins for detection of various analytes, including metal ions such as K+, Ca2+, Ag+, Hg2+, Cu2+, Pb2+, Sr2+, organic molecules, nucleic acids, and proteins. Principles of G-quadruplex-based detection methods involve DNA conformational change caused by the presence of analyte which leads to a decrease or an increase in peroxidase activity, fluorescence, or electrochemical signal of the used probe. The advantages of various detection techniques are also discussed