New genetic insights to consider coffee waste as feedstock for fuel,feed, and chemicals

Caffeine is a natural plant product found in many drinks, including coffee, tea, soft and energy drinks. Due to caffeine’s presence in the environment, microorganisms have evolved two different mechanisms to live on caffeine. The genetic maps of the caffeine N-demethylation pathway and C-8 oxidation pathway have been discovered in Pseudomonas putida CBB5 and Pseudomonas sp. CBB1, respectively. These genes are the only characterized bacterial caffeine-degrading genes, and may be of great value in producing fine chemicals, biofuels, and animal feed from coffee and tea waste. Here, we present preliminary results for production of theobromine and 7-methylxanthine from caffeine and theobromine, respectively, by two strains of metabolically engineered E. coli. We also demonstrate complete decaffeination of tea extract by an immobilized mixed culture of Klebsiella and Rhodococcus cells. These processes provide a first level demonstration of biotechnological utilization of coffee and tea waste.      

Application of UV detection in HPLC in the total antioxidant potential assay

HPLC has been already used for the TAP estimation. Phenylalanine, salicylic, p-hydroxybenzoic (pHBA) or terephthalic (TPA) acids have been used as sensor compounds. Products of their reaction with the hydroxyl radicals, generated in the Fenton-like reaction, were analyzed using electrochemical or fluorescence detection. This paper describes the TAP assay based on the hydroxyl radicals reaction with pHBA, reversed-phase-HPLC separation and UV photometric detection. The elaborated assay has been used to evaluate TAP values of some apiculture products.      

A study of the antioxidative behavior of phenolic acids, in aqueous herb extracts, using a dsDNA biosensor

Electrochemical DNA biosensors are promising tools for the fast, inexpensive and simple in vitro analysis for the determination of free radicals and antioxidants. High concentrations of antioxidants in such compounds as phenolic acids and plant extracts, act as free radical terminators which reduce the effect of the oxidative dam-age on DNA. The electrochemical behavior of three representative phenolic acids, caffeic acid, gallic acid and trolox were studied by cyclic voltammetry. Moreover, the determination of the above antioxidants under the optimized conditions (scan rate, deposition potential and time) using differential pulse voltammetry was also investigated. In vitro studies focused on their antioxidative effect were performed by adsorptive transfer stripping voltammetry and dsDNA biosensor. Using Fenton’s system, with FeSO₄ and H₂O₂ was chosen as a strong oxidative system. This biosensor was applied as a screening antioxidant test in order to estimate the antioxidant capacity of aqueous herb extracts.      

Surface plasmon resonance imaging biosensors for aromatase based on a potent inhibitor and a specific antibody: Sensor development and application for biological material

Aromatase (ARO) is an enzyme with potential diagnostic significance. Aberrant expression of aromatase in tissues is associated with a number of pathological conditions, including tumor of the breast, ovary, testes, liver, adrenal cortex and uterus, as well as endometriosis. Two methods for the highly selective determination of ARO concentration in human tissues by using of two different biosensors co-operating with the surface plasmon resonance imaging technique (SPRI) have been developed. One of the developed biosensors contains immobilised rabbit polyclonal antibody specific for aromatase (Y-ARO), while the other contains immobilized ARO inhibitor-exemestane (E-ARO). Both biosensors specifically bound ARO from analyzed samples. The analytically useful dynamic response range of both biosensors is between 0.3 and 5.0 ng mL^?1. The detection limit (3S.D.) of both biosensors is 90 pg mL^?1. Standard deviation of both biosensors is 1%. Recoveries of ARO spikes are between 97 and 108% for both biosensors under model conditions and for real samples. Albumin and alkaline phosphatase are tolerated for both biosensors up to 10,000 fold excess.      

A hydrogen peroxide biosensor based on direct electron transfer from hemoglobin to an electrode modified with Nafion and activated nanocarbon

A biosensor for hydrogen peroxide (HP) was developed by immobilizing hemoglobin on a glassy carbon electrode modified with activated carbon nanoparticles/Nafion. The characteristics of the sensor were studied by UV?Cvis spectroscopy and electrochemical methods. The immobilized Hb retained its native secondary structure, undergoes direct electron transfer (with a heterogeneous rate constant of 3.37?±?0.5?s^?1), and displays excellent bioelectrocatalytic activity to the reduction of HP. Under the optimal conditions, its amperometric response varies linearly with the concentration of HP in the range from 0.9???M to 17???M. The detection limit is 0.4???M (at S/N?=?3). Due to the commercial availability and low cost of activated carbon nanoparticles, it can be considered as a useful supporting material for construction of other third-generation biosensors.

Enantioselective inhibition of immobilized acetylcholinesterase in biosensor determination of pesticides

Chiral effects for the inhibition of acetylcholinesterase by organophosphorus pesticides were investigated for insecticide malathion and malaoxon, which is a metabolic product of malathion in living organisms. Studies were carried out using a bienzymatic biosensor with immobilized acetylcholinesterase, choline oxidase, and with Prussian Blue used as a mediator. In both cases the R enantiomers accelerate acetylocholinesterase inhibition. The chiral effect in inhibition was much more pronounced in fast flow measurements than in batch measurements.      

Antioxidative responses to seasonal changes and chemiluminescence assay of Astragalus onobrychis leaves extract

The aim of this study was to research the seasonal changes of antioxidant enzyme activity and total antioxidant capacity in leaves of Astragalus onobrychis L. subsp. chlorocarpus (Griseb.) S. Kozuharov et D.K. Pavlova. Leaves of A. onobrychis were collected during the different stages of growth and analyzed for antioxidant enzyme activity: superoxide dismutase, catalase, guaiacol peroxidase, glutathione peroxidase. Quantities of malonyldialdehyde, superoxide radicals, and hydroxyl radicals were measured as well as the content of soluble proteins. Furthermore, total antioxidant capacity was determined by the inhibition of chemiluminescence activity of blood phagocytes by leaf extracts. Stages of vegetation significantly affected the accumulation of superoxide radicals, but there were no significant differences in hydroxyl radical quantity and lipid peroxidation levels during vegetation. Soluble proteins vary greatly between different stages of growth. Seasonal changes were found to have an effect on enzymatic activities. During the spring season, guaiacol peroxidase showed the highest levels. Catalase and glutathione peroxidase increased their activities in summer, while, during the autumn season, superoxide dismutase showed maximum activity. On the basis of chemiluminescence assay, it can be concluded that leaf extract of A. onobrychis possesses a significant antioxidant capacity thus protecting plants during environmental stress.      

Organoruthenium(II) thiosemicarbazone complexes: synthesis, spectral characterization, DNA binding and DNA cleavage studies

A series of new ruthenium(II) complexes were synthesized with Schiff bases derived from salicylaldehyde / o-hydroxyacetophenone/ o-vanillin / 2-hydroxy-1-naphthaldehyde with thiosemicarbazide and acetyl furan. They are characterized by elemental analysis, IR, electronic, ¹H NMR, ¹³C NMR and mass spectral studies. The elemental analysis suggests the stoichiometry to be 1:1 (metal:ligand). Four of these complexes were tested for its binding with CT-DNA using absorption spectroscopic studies and two of these complexes exhibit efficient DNA cleavage activity.      

Electrochemical DNA biosensor for the detection of Trichoderma harzianum based on a gold electrode modified with a composite membrane made from an ionic liquid, ZnO nanoparticles and chitosan, and by using acridine orange as a redox indicator

An electrochemical DNA biosensor was developed that is based on a gold electrode modified with a nanocomposite membrane made from an ionic liquid, ZnO nanoparticles and chitosan. A single-stranded DNA probe was immobilized on this electrode. Acridine orange was used as the hybridization probe for monitoring the hybridization of the target DNA. The biosensor was capable of detecting target DNA in the concentration range from 1.0?×?10?C14 to 1.8?×?10?C4?mol?L-1, with a detection limit of 1.0?×?10?C15?mol?L-1. The approach towards constructing a DNA biosensor allows studies on the hybridization even with crude DNA fragments and also to analyze sample obtained from real samples. The results show that the DNA biosensor has the potential for sensitive detection of a specific sequence of the Trichoderma harzianum gene and provides a quick, sensitive and convenient method for the study of microorganisms.

Biosensor with Protective Membrane for the Detection of DNA Damage and Antioxidant Properties of Fruit Juices

With the purpose to prepare a DNA biosensor protected with an outer‐sphere membrane against high molecular weight interferences, a carbon film electrode was layer‐by‐layer modified with dsDNA and chitosan. Using cyclic and square‐wave voltammetry and impedance spectroscopy, the oxidative damage of DNA by the hydroxyl and superoxide anion radicals was detected which consists of opening of the helix structure followed by deep DNA chain degradation. The biosensor has been applied to the detection of the antioxidant effect of apple and orange juices. The investigation of the novel biosensor with a protective membrane represents a significant contribution to the field of DNA biosensors utilization.     

The first synthesis of 3-deoxyoripavine and its utilization in the preparation of 10-deoxyaporphines and cyprodime

The synthesis of 3-deoxyoripavine (7) was realized as a novel and promising intermediate towards the synthesis of the important class of dopaminergic and/or serotonergic 10-deoxyaporphines and the special pharmacological tool µ opioid antagonist cyprodime. Generally, the preparation of these valuable biologically active compounds was achieved in remarkable yields.      

Screen-printed electrodes modified with HRP-zirconium alcoxide film for the development of a biosensor for acetaminophen detection

The development of a biosensor based on the immobilization of horseradish peroxidase (HRP) within a zirconium alkoxide-polyetilenimine film onto screen-printed electrodes (SPE) for acetaminophen detection and acetaminophen quantification in pharmaceutical products is described. The biosensor operation mode is based on monitoring the amperometric signal produced by the electrochemical reduction of the enzymatically generated electroactive oxidized species of acetaminophen in the presence of hydrogen peroxide. The enzyme immobilization is performed by retention in a polyethylenimine-zirconium alcoxide porous gel film, a technique that offers good entrapping and a protective environment for the biocomponent due to the hydration properties of the immobilization layer. SPEs have the advantage of being easily mass-produced at low costs with superior characteristics in comparison with classical electrode materials. In this configuration, zirconium alkoxide demonstrates its electrocatalytic activity. The biosensor allows the quantification of acetaminophen with a limit of detection of 6.21×10^?8 M and a linear range between 4.35×10^?7 M and 4.98×10^?6 M. Finally, the biosensor is applied to the quantitative analysis of acetaminophen in Perdolan® tablets.

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Preparation of selectively protected protoescigenin derivatives for synthesis of escin analogs and neoglycoconjugates

Protoescigenin, the main aglycone of horse chestnut saponin mixture known as escin, was selected as substrate for exploratory chemistry towards selective protection, followed by propargyl ether formation and subsequent condensation with azido-monosaccharides, to obtain novel triazole linked conjugates of the triterpene.      

In vitro biological activity comparison of some hydroxyapatite-based composite materials using simulated body fluid

Hydroxyapatite composites are the main biomaterials used for metal implant coatings. Their in vitro study is very important. That is why their behavior was monitored in simulated body fluid (SBF), which is a solution with ion concentrations and pH value similar to those of human blood plasma. Silica, chitosan and gelatin-doped hydroxyapatite-based biomaterials were studied in SBF; the samples were characterized pre-, during and post-SBF immersion using infra-red, scanning and transmission electron spectroscopy and X-ray diffraction methods. The solubility of materials in SBF was determined, and the variation of Ca²⁺ and phosphorus concentration was also recorded during SBF experiments. The results were compared and their in vitro biological activity was determined.      

Reaction of barbituric acid with organic azides and phosphonium ylides

Amination by organic azides has been carried out to provide aminobarbiturates by fusion of a triazole ring to the 5,6-positions of barbituric acid followed by cleavage and thermal elimination of nitrogen, whereas aza-Wittig reaction gave phosphoranylidene barbituric acid salts.      

Self-assembly of acetylcholinesterase on gold nanoparticles electrodeposited on graphite

The immobilisation of AChE enzyme through chemisorption on Au-modified graphite was examined with view of its prospective application in the design of membraneless electrochemical biosensors for the assay of enzyme inhibitors. The developed immobilisation protocol has been based on a two-stage procedure, comprising i) electrodeposition of gold nanostructures on spectroscopic graphite; followed by ii) chemisorption of the enzyme onto gold nanoparticles. Both the coverage of the electrode surface with Au nanostructures and the conditions for enzyme immobilisation were optimised. The proposed electrode architecture together with the specific type of enzyme immobilisation allow for a long-term retaining of the enzyme catalytic activity. The extent of inhibition of the immobilised acetylcholinesterase enzyme by the organophosphorous compound monocrotophos has been found to depend linearly on its concentration over the range from 50 to 400 nmol mL^?1 with sensitivity 77.2% inhibition per 1 µmol mL^?1 of monocrotophos.      

Sensitive DNA-hybridization biosensors based on gold nanoparticles for testing DNA damage by Cd(II) ions

A DNA biosensor was constructed by immobilizing a 20-mer oligonucleotide probe and hybridizing it with its complementary oligomer on the surface of a glassy carbon electrode modified with gold nanoparticles. The properties of the biosensor and its capability of recognizing its complementary sequence were studied by electrochemical impedance spectroscopy. The oxidative stress caused by cadmium ions can be monitored by differential pulse voltammetry using the cobalt(III)tris(1,10-phenanthroline) complex and methylene blue as electrochemical indicators. The biosensor is capable of indicating damage caused by Cd(II) ions in pH 6.0 solution. The results showed that the biosensor can be used for rapid screening for DNA damage.

Metals content of soil,leaves and wild fruit from Serbia

The concentrations of Zn, Mn, Fe, Pb, Ni, Cu and Cd in soil, leaves and edible wild fruit (Crataegus laevigata L., Cornus mas L. and Prunus spinosa L.) from southeast Serbia were determined by atomic absorption spectroscopy. Metal translocations from soil to fruit were calculated as well as their oral intake and health risk indices. Positive correlations were found among metal concentrations in soil, leaves and fruit.      

Ultra-sensitive electrochemical DNA biosensor based on signal amplification using gold nanoparticles modified with molybdenum disulfide,graphene and horseradish peroxidase

We have developed an ultra-sensitive electrochemical DNA biosensor by assembling probe ssDNA on a glassy carbon electrode modified with a composite made from molybdenum disulfide, graphene, chitosan and gold nanoparticles. A thiol-tagged DNA strand coupled to horseradish peroxidase conjugated to AuNP served as a tracer. The nanocomposite on the surface acts as relatively good electrical conductor for accelerating the electron transfer, while the enzyme tagged gold nanoparticles provide signal amplification. Hybridization with the target DNA was studied by measuring the electrochemical signal response of horseradish peroxidase using differential pulse voltammetry. The calibration plot is linear in the 5.0?×?10^?14 and 5.0?×?10^?9 M concentration range, and the limit of detection is 2.2?×?10^?15 M. The biosensor displays high selectivity and can differentiate between single-base mismatched and three-base mismatched sequences of DNA. The approach is deemed to provide a sensitive and reliable tool for highly specific detection of DNA.