A fluorometric method for determination of the activity of T4 polynucleotide kinase by using a DNA-templated silver nanocluster probe.

The authors describe a turn-off fluorometric method for the determination of the activity of the T4 polynucleotide kinase (T4 PNK). It is based on the use of DNA-templated silver nanoclusters (AgNCs). DNA probes with terminal 5' hydroxy groups are used as substrates for DNA phosphatases. If subsequently treated with T4 PNK and Lambda exonuclease (λ exo), the AgNC DNA probes with a modified C-rich sequence and the G-rich sequence is separated. Upon their separation, the strong fluorescence (with excitation/emission maxima at 580/650 nm) that is caused by the proximity of the G-rich region and the C-rich region in the AgNCs decreases sharply. This enabled the fluorometric kinetic determination of the activity of T4 PNK. The assay is characterized by a wide linear range (from 0.01 to 12.5 U·mL-1), a low detection limit (0.01 U·mL-1) and short assay time (typically 60 min). This makes it a promising tool for use in studying processes related to DNA phosphorylation, in drug discovery and in diagnostics. Graphical abstract A turn-off fluorometric method for the determination of the activity of the T4 polynucleotide kinase (T4 PNK) has been developed. It is based on the use of DNA-templated silver nanoclusters (AgNCs). This makes it a promising tool for use in studying processes related to DNA phosphorylation, in drug discovery and in diagnostics.

Effect of disodium phosphate and homogenization on physico-chemical and rheological properties of buffalo skim milk based ultrafiltered retentate.

The concentration of pasteurized buffalo skim milk (PBSM) employing ultrafiltration (UF) alters the chemical composition of ultrafiltered retentate that adversely affect its proteins and salts equilibrium. Effect of stabilizing salts addition in concentrated milks or retentates was majorly dedicated to their thermal stability only. Therefore, this study was aimed to investigate the effect of disodium phosphate (DSP) addition and homogenization of 2.40 × UF retentate (0.60 protein to total solids ratio) on its ζ-potential, particle size, heat stability, turbidity, pH, viscosity and crossover temperature of storage (G') and loss (G″) modulus. Concentration of PBSM in UF process, significantly (P < 0.05) increased its percent TS, protein, fat and ash contents, but markedly decreased its lactose content. DSP addition significantly increased (P < 0.05) the ζ-potential, pH, viscosity and particle size in majority of the homogenized and non-homogenized retentates. Homogenized retentates containing 2.5 and 5% DSP exhibited Newtonian and Power law flow behaviour. However, rheological behaviour of non-homogenized retentates containing zero (control), 1 and 4% DSP was best explained by Bingham model. Further, non-homogenized retentates with 0.5, 2, 3, 5% DSP exhibited Newtonian flow, but retentates containing 6 and 7% DSP was best explained by Power law. The correlation among different attributes of DSP added non-homogenized and homogenized samples were also studied. Particle size and turbidity (r = + 0.999, P < 0.05) as well as ζ-potential and crossover temperature of G' and G″ (r = + 0.999, P < 0.05) showed positive correlation in 4% DSP added non-homogenized retentate.

Inhibitory effects of p-dodecylaminophenol on the invasiveness of human fibrosarcoma cell line HT1080.

Cancer is a major cause of death, and the development of new anticancer drugs is urgently needed. Invasion and metastasis are the primary causes of death due to cancer rather than growth of the primary tumor. In the current study, we examined the anti-invasive effects of p-dodecylaminophenol (1), which was developed based on N-(4-hydroxyphenyl)retinamide (2), a synthetic amide of all-trans-retinoic acid (3). In HT1080 cells 1 inhibited growth, induced apoptosis and arrested the cell cycle in S phase in a dose-dependent manner. In addition, 1 significantly suppressed cell invasion, and the activity and mRNA expression of matrix metalloproteinase-9 (MMP-9). Furthermore, the expression of the reversion-inducing cysteine-rich protein with Kazal motifs (RECK), which is a negative regulator of MMP-9, was increased by treatment with 1. These results suggest that 1 could be an effective anti-cancer agent that suppresses cell growth through apoptosis induction and cell cycle arrest, which also inhibits cell invasion by decreasing MMP-9 expression due to an increase in RECK. Compound 1 might be useful clinically as a new and potent anticancer agent that could overcome adverse side effects of the retinoids.

Polyphenol-driving assembly for constructing chitin-polyphenol-metal hydrogel as wound dressing.

The metal-polyphenol networks have attracted appealing attention for diverse biomedical applications due to their remarkable characteristics. Though various of metal-polyphenolic materials have been prepared, the homogeneous metal-polyphenol based hydrogel fabrication remains a challenge (e.g., quick aggregation). Herein, a facile and low-cost polyphenol-mediating non-covalent driven assembly strategy was developed for fabricating homogeneous chitin-polyphenol-metal hydrogels. Polyphenols not only noncovalently crosslinked chitin chains, but simultaneously captured metal nanomaterials from metal substrates and immobilized in chitin-polyphenol networks. A range of metal (Fe, Cu, Ti, Zn) and polyphenol (tannic acid, gallic acid, quercetin, pyrogallic acid) could be incorporated into this hydrogel framework. As a demonstration, the chitin-tannic acid-Cu hydrogel showed excellent antibacterial properties and significantly enhanced infected wound repair via promoting the cell proliferation and angiogenesis, showing the potential in wound dressing. The low cost, versatility and flexibility assembly process can be used to fabricate diverse polymer-polyphenol-metal hydrogel, thereby enabling their use in various applications.

Effect of molecular weight of chitosan on the formation and properties of zein-nisin-chitosan nanocomplexes.

This study evaluated the effect of molecular weight of chitosan (3 kDa,150 kDa, 400 kDa, and 600 kDa) on zein-nisin-chitosan nanocomplexes. The formation mechanism, physicochemical and antibacterial properties of the nanocomplexes (ZNC0.3, ZNC15, ZNC40, and ZNC60) were assessed. The nanocomplexes were characterized by DLS, ζ-potential, atomic force microscopy, scanning electron microscopy, circular dichroism, fourier transform infrared and UV-Vis spectroscopy. The results showed that the lowest molecular weight chitosan (LMWC, 3 kDa) formed nanocomplexes with nisin and zein structurally differed from the higher molecular weights chitosan (HMWC, >3 kDa). LMWC was doped on the surface of the nanocomplexes. HMWC linked and formed a network to adsorb zein and nisin. The antibacterial activity against Staphylococcus aureus showed that the minimum inhibitory concentration of ZNC0.3, ZNC15, ZNC40, and ZNC60 was 7.0625, 14.125, 14.125, and 28.25 µg/mL. ZNC0.3 could be a suitable nisin delivery system for its high encapsulation efficiency (85.38%) and antibacterial properties.

Phosphorylation of Maize Starch Enhanced with High-Voltage Electrical Discharge (HVED) Instead of Thermal Treatment.

The aim of this research was to explore the use of a high-voltage electrical treatment (HVED) as a substitute for heating during the phosphorylation of maize starch. Starch was treated with HVED, phosphorylated with Na2HPO4 or Na5P3O10 with and without thermal treatment and phosphorylated in combination with HVED prior to and after the chemical modification. When starch was phosphorylated with Na2HPO4, HVED was more efficient in catalyzing reaction (3.89 mg P/kg for 30 min HVED in relation to 0.43 mg P/kg for thermal treatment), whereas with Na5P3O10 similar P content was achieved as with thermal treatment (0.76 P/kg for 30 min HVED in relation to 0.86 mg P/kg). The order of HVED and chemical reactions did not have a marked effect on phosphorous content. In combination with Na2HPO4, HVED pre-treatment had a more pronounced effect on the solubility and water absorption, whereas post-treatment was favoured with Na5P3O10. Mean diameter was increased by all treatments, where HVED had a marked effect. Enthalpy of gelatinization ranged from 11.76 J/g for starch treated with Na5P3O10 and 10 min-HVED to 13.58 J/g for Na5P3O10 treated sample. G' and G″ increased after both thermally and HVED enhanced phosphorylations, with a slightly more pronounced effect of the HVED.

Naringin prevents cyclophosphamide-induced erythrocytotoxicity in rats by abrogating oxidative stress.

Earlier reports have shown that Cyclophosphamide (CYCP), an anti-malignant drug, elicited cytotoxicity; and that naringin has several beneficial potentials against oxidative stress and dyslipidaemias. We investigated the influence of naringin on free radical scavenging, cellular integrity, cellular ATP, antioxidants, oxidative stress, and lipid profiles in the CYCP-induced erythrocytotoxicity rat model. Rats were pretreated orally by gavage for fourteen consecutive days with three doses (50, 100, and 200 mg/kg) naringin before single CYCP (200 mg/kg, i.p.) administration. Afterwards, the rats were sacrificed. Naringin concentrations required for 50 % scavenging hydrogen peroxide and nitric oxide radical were 0.27 mg/mL and 0.28 mg/mL, respectively. Naringin pretreatment significantly (p < 0.05) protected erythrocytes plasma membrane architecture and integrity by abolishing CYCP-induced decrease in the activity of erythrocyte LDH (a marker of ATP). Pretreatment with naringin remarkably (p < 0.05) reversed CYCP-induced decreases in the erythrocytes glutathione levels, activities of glutathione-S-transferase, catalase, glutathione peroxidase, and glutathione reductase; attenuated CYCP-mediated increases in erythrocytes levels of malondialdehyde, nitric oxide, and major lipids (cholesterol, triacylglycerol, phospholipids, and non-esterified fatty acids). Taken together, different acute pretreatment doses of naringin might avert CYCP-mediated erythrocytes dysfunctions via its antioxidant, free-radical scavenging, and anti-dyslipidaemia properties.

Enhanced buffer capacity of fermentation broth and biohydrogen production from corn stalk with Na2HPO4/NaH2PO4.

The remarkable buffer capacity of buffer solution can significantly improve the biohydrogen production yield and energy conversion efficiency. In the present study, the effect of buffer solution Na2HPO4/NaH2PO4 on buffer capacity of fermentation broth and photo-fermentation biohydrogen production (PFHP) was studied. Gas characteristics, fermentation broth properties, and kinetic parameters in PFHP were investigated. With the increase in pH values (5-7) of buffer solution Na2HPO4/NaH2PO4, firstly hydrogen yield increased and then decreased. Maximum energy conversion efficiency 9.84%, hydrogen yield 132.69 mL/g corn stalk, and hydrogen content 53.88% were achieved at pH value of 6. The results of one-way ANOVA showed that pH values of fermentation broth and cumulative hydrogen production were strongly affected by pH values of buffer solution. Buffer solution Na2HPO4/NaH2PO4 retarded the decrease of pH value of photo-fermentation broth, and significantly improved the PFHP.

Monitoring zearalenone in corn flour utilizing novel self-enhanced electrochemiluminescence aptasensor based on NGQDs-NH2-Ru@SiO2 luminophore.

Accurate and early diagnosis of mycotoxin is particularly significant to the food and agricultural product safety. In the present work, a sensitive and effective monitoring method for zearalenone (ZEN) was exploited based on a novel self-enhanced electrochemiluminescence (ECL) aptasensor. The self-enhanced lumonophore was compounded by electrostatically combining amine-functionalized Ru(bpy)32+-doped silica nanoparticles (NH2-Ru@SiO2 NPs) and nitrogen doped graphene quantum dots (NGQDs) together. Since the emitter and co-reactant simultaneously existed in the same nanoparticle, shortened electron-transfer distance and decreased energy loss was obtained. Therefore, self-enhanced ECL aptasensor based on the novel complex expressed the widest linear range of 10 fg mL-1-10 ng mL-1 and the lowest detection limit of 1 fg mL-1 for ZEN detection. More importantly, ZEN produced during the mildew process of corn flour was monitored by the developed aptasensor, which exhibited superior determination and potential application in real samples.

A single metabolite production by Escherichia coli BW25113 and its pflA.cra mutant cultivated under microaerobic conditions using glycerol or glucose as a carbon source.

Abundant, low prices and a highly reduced nature make glycerol to be an ideal feedstock for the production of reduced biochemicals and biofuels. Escherichia coli has been paid much attention as the platform of microbial cell factories due to its high growth rate (giving higher metabolite production rate) and the capability of utilizing a wide range of carbon sources. However, one of the drawbacks of using E. coli as a platform is its mixed metabolite formation under anaerobic conditions. In the present study, it was shown that ethanol could be exclusively produced from glycerol by the wild type E. coli, while d-lactic acid could be exclusively produced from glucose by pflA.cra mutant, where the glucose uptake rate could be increased by this mutant as compared to the wild type strain. It was also shown that the growth rate is significantly reduced in pflA.cra mutant for the case of using glycerol as a carbon source due to redox imbalance. The metabolic regulation mechanisms behind the fermentation characteristic were clarified to some extent.

Prediction of the antiglycation activity of polysaccharides from Benincasa hispida using a response surface methodology.

Benincasa hispida is a popular vegetable in China. Our previous experiments suggested that polysaccharides isolated from B. hispida fruits (PBH) have antiglycation effect and DPPH free radical scavenging activity. Ultrasonic treatments can be used to extract polysaccharides from Benincasa hispida (PBH). The aim of this study was to investigate the correlation between the ultrasonic treatment conditions and the antiglycation activity of PBH. A mathematical model was generated with an artificial neural network (ANN) toolbox from MATLAB to analyze the effects of ultrasonic treatment conditions on antiglycation activity. The response surface plots showed relationships between ultrasonic extraction conditions and bioactivity. The R(2) value of the model was 0.9919, which suggested good fitness of the neural network. The application of genetic algorithms showed that the optimal ultrasonic extraction conditions resulted in the highest antiglycation activity for PBH. These were 150W, 46°C, and 35min. These conditions produced a predicted antiglycation activity of 41.2%; the actual activity was 40.9% under optimal conditions. This is very close to the predicted value. The experimental data indicated that the PBH possessed both antiglycation and antioxidant activities. The maximum actual value of antiglycation was 101.7% that of the positive control, and the PBH inhibited the DPPH free radicals with an EC50 value of 0.98mg/mL. This is 66.2% that of ascorbic acid. These results explained the observations that B. hispida can decrease glucose levels in diabetic patients. The experimental results also showed that the ANN could be used for optimization and prediction.

Forced degradation and impurity profiling: recent trends in analytical perspectives.

This review describes an epigrammatic impression of the recent trends in analytical perspectives of degradation and impurities profiling of pharmaceuticals including active pharmaceutical ingredient (API) as well as drug products during 2008-2012. These recent trends in forced degradation and impurity profiling were discussed on the head of year of publication; columns, matrix (API and dosage forms) and type of elution in chromatography (isocratic and gradient); therapeutic categories of the drug which were used for analysis. It focuses distinctly on comprehensive update of various analytical methods including hyphenated techniques for the identification and quantification of thresholds of impurities and degradants in different pharmaceutical matrices.