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In this study, tin oxide‑cobalt oxide nanocatalyst was prepared by a simple method, which grew in spherical particles with an average diameter of 30 nm. Tin oxide–cobalt oxide was further wrapped in alginate polymer hydrogel (Alg@tin oxide–cobalt oxide), and both materials were utilized as nanocatalysts for the catalytic transformation of different pollutants. Tin oxide–cobalt oxide and Alg@tin oxide–cobalt oxide nanocatalysts were tested for the catalytic reduction of 4-nitrophenol, congo red, methyl orange, methylene blue (MB) and potassium ferricyanide in which sodium borohydride was used as a reducing agent. Tin oxide–cobalt oxide and Alg@tin oxide–cobalt oxide nanocatalysts synergistically reduced MB in shorter time (2.0 and 4.0 min) compared to other dyes. The reduction conditions were optimized by changing different parameters. The rate constants for MB reduction were calculated and found to be 1.5714 min−1 and 0.6033 min−1 using tin oxide–cobalt oxide and Alg@tin oxide–cobalt oxide nanocatalysts, respectively. Implementing Alg@tin oxide–cobalt oxide nanocatalyst toward MB reduction in real samples proved its efficacy in sea and well water samples. The catalyst could be easily recovered, recycled and revealed a minimal loss of nanoparticles, which offering a competition and replacement with reputable commercial catalysts.

Photoanode sensing platforms with remarkable photoelectrochemical (PEC) response and satisfying visible-light absorption have become the most promising detection systems. Nevertheless, their inevitable electrophilic character limits their expansion in the bioassay because of reductive substances in serum or other body fluids that can severely interfere with the photocurrent to be read. To solve it, a PEC platform-assembled dual-active electrode is designed to realize the separation of biological monitoring from the photoanode. The ferricyanide ([Fe(CN)6]3-)-mediated redox cycle is first proposed to meet the gain and loss electron requirements of the PEC system. It can avoid the self-reaction in the electrolyte caused by the addition of a traditional electron donor and acceptor, for instance, ascorbic acid and hydrogen peroxide. As a consequence, the traditional counter electrode (Pt wire) is replaced by Fe2O3/AgInS2 heterojunction, which can amplify the PEC response of the cathode to meet the requirement of trace analysis. An aptasensor fabricated by the above strategies exhibits convincing data for 17β-estradiol (E2) detection from which a wide detection range is obtained in 10 fg/mL to 1 μg/mL with a detection limit of 2.74 fg/mL (S/N = 3). These advanced elements show a rosy prospect for environmental monitoring and point-of-care biomarker diagnosis.

Biological ion channels can realize delicate mass transport under complicated physiological conditions. Artificial nanochannels can achieve biomimetic ion current rectification (ICR), gating, and selectivity that are mostly performed in pure salt solutions. Synthetic nanochannels that can function under mixed ion systems are highly desirable, yet their performances are hard to be compared to those under pure systems. Seeking out the potential reasons by investigating the effect of mixed-system components on the ion-transport properties of the constructed nanochannels seems necessary and important. Herein, we report the effect of anions with different charges and sizes on the ICR properties of positively charged nanochannels. Among the investigated anions, the low-valent anions showed no impact on the ICR direction, while the high-valent component ferrocyanide [Fe(CN)64-] caused significant ICR inversion. The ICR inversion mechanism is evidenced to result from the adsorption of Fe(CN)64--induced surface charge reversal, which relates to solution concentration, pH conditions, and nanochannel sizes and applies to both aminated and quaternized nanochannels that are positively charged. Noticeably, Fe(CN)64- is found to interfere with the transport of protein molecules in the nanochannel. This work points out that the ion species from mixed systems would potentially impact the intrinsic ICR properties of the nanochannels. Replacing highly charged counterions with organic components would be promising in building up future nanochannel-based mass transport systems running under mixed systems.

Herein we present a general and turn-on strategy for enzymatic bioassays on the basis of redox state dependent emission of gold nanoclusters (AuNCs). The photoluminescence of AuNCs was quenched obviously by the oxidative ferricyanide while unaffected by its corresponding reduced state, i.e., ferrocyanide. The distinctive quenching abilities for AuNCs by the redox couple (ferricyanide/ferrocyanide) enabled their utility as new fluorescent sensing platforms to detect redox-related phenomena. The proposed protocols were conducted by using the model oxidoreductases of glucose oxidase (GOx) and the enzyme cascade of lactate dehydrogenase (LDH)/diaphorase to catalytically convert ferricyanide to ferrocyanide, which switched on fluorescence of the detection systems. The detection limit for glucose and lactate was found to be as low as 0.12 and 0.09 μM, respectively. This work features the first use of the redox couple of ferricyanide/ferrocyanide in fluorescent bioanalysis, which enables versatile, signal on and highly sensitive/selective detections as compared to the state of the art fluorescently enzymatic sensing platforms. Importantly, considering the significance of ferricyanide/ferrocyanide involves in numerous other oxidoreductases mediated biocatalysis, this protocol has wide versatility that enables combination with oxidoreductases related reactions for biosensing.

Carbon neutrality has been received more attention and emerged in wastewater treatment processes. Due to the development of treating technologies with the rising of new-emerging pollutants, the coupled chemical processes also should remain current for the goal of carbon-neutral operation. Among of those updated strategies, several advanced oxidation processes (AOPs) based on dithionite (DTN, S2O42−), a common water treatment agent, have been established for refractory organic contaminations removal. However, in terms of DTN detection, the traditional formol-titration method has several application limits including the low detection sensitivity and high consumption of formaldehyde. In this study, compared with traditional method, a low energy consumption technology has been developed based on the potassium ferricyanide with the carbon consumption decreasing by about 5 times. Moreover, detection limit of DTN (mmol/L level) also was lower than the titration method. The method was established based on the fact that every 1 mol of DTN can react with 2 mol [Fe(CN)6]3- under alkaline condition. According to that potassium ferricyanide (K3 [Fe(CN)6]) has the maximum absorption at 419 nm wavelength, a fitting equation based on the linear relationship between the absorbance variation of K3 [Fe(CN)6] and DTN amount in the ranges of 0–30 μmol with the detection limit of 0.6 μmol was established with the determination coefficient of 0.99935. It was found that there was no obvious influence of the ubiquitous foreign species with the amount lower than 6 mM, 4 mM, 6 mM, 4 mM and 1 mg/L for Cl−, HCO3−, NO3−, SO42− and NOM, respectively. Moreover, methanol and tert-butanol were employed to verify the influence of the presence of organic matters on the determination of DTN and no impact was observed in this study. The proposed method provides a new way for DTN detection with stable and countable performance in the related AOPs with the low electric energy and carbon source consumption and high detection efficiency.

Amide bond formation is one of the most important reactions in both chemistry and biology1–4, but there is currently no chemical method of achieving α-peptide ligation in water that tolerates all of the 20 proteinogenic amino acids at the peptide ligation site. The universal genetic code establishes that the biological role of peptides predates life’s last universal common ancestor and that peptides played an essential part in the origins of life5–9. The essential role of sulfur in the citric acid cycle, non-ribosomal peptide synthesis and polyketide biosynthesis point towards thioester-dependent peptide ligations preceding RNA-dependent protein synthesis during the evolution of life5,9–13. However, a robust mechanism for aminoacyl thioester formation has not been demonstrated13. Here we report a chemoselective, high-yielding α-aminonitrile ligation that exploits only prebiotically plausible molecules—hydrogen sulfide, thioacetate12,14 and ferricyanide12,14–17 or cyanoacetylene8,14—to yield α-peptides in water. The ligation is extremely selective for α-aminonitrile coupling and tolerates all of the 20 proteinogenic amino acid residues. Two essential features enable peptide ligation in water: the reactivity and pKaH of α-aminonitriles makes them compatible with ligation at neutral pH and N-acylation stabilizes the peptide product and activates the peptide precursor to (biomimetic) N-to-C peptide ligation. Our model unites prebiotic aminonitrile synthesis and biological α-peptides, suggesting that short N-acyl peptide nitriles were plausible substrates during early evolution.

Leishmania donovani (L. donovani) is one of the parasites that cause leishmaniasis. The mechanisms by which L. donovani fights against adverse environment and becomes resistant to drugs are not well understood yet. The present study was designed to evaluate the effects of different regulators on the modulation of Transplasma Membrane Electron Transport (transPMET) systems of susceptible and resistant L. donovani cells. Effects of UV, different buffers, and electron transport inhibitors and stimulators on the reduction of α-lipoic acid (ALA), 1,2-naphthoquinone-4-sulphonic acid (NQSA) and ferricyanide were determined. ALA reductions were inhibited in susceptible, sodium antimony gluconate (SAG)-resistant and paromomycin (PMM)-resistant AG83 amastigote cells, and stimulated in susceptible and SAG-resistant AG83 promastigote cells upon UV exposure. The results indicate that UV irradiation almost oppositely affect ALA reductions in amastigotes and promastigotes. ALA reductions were stimulated in sensitive and inhibited in resistant GE1 amastigotes upon UV exposure. Susceptible amastigotes and promastigotes inhibited, and resistant amastigotes and promastigotes stimulated NQSA reduction under UV irradiation. Thus, susceptible and drug-resistant amastigotes and promastigotes are different in the reduction of ALA. Susceptible and resistant AG83 amastigotes and promastigotes inhibited the ferricyanide reductions upon UV exposure, which indicates, there is no such difference in ferricyanide reductions among susceptible as well as resistant AG83 amastigotes and promastigotes. The reductions of extracellular electron excerptors in susceptible promastigotes requires the availability of Na+ and Cl− ions for maximal activity but susceptible amastigotes are mostly not dependent on the availability of Na+ and Cl− ions. Both in promastigotes and amastigotes, reductions of electron acceptors were strongly inhibited by carbonyl cyanide p-(trifluoromethoxy)phenylhydrazone. Furthermore, antimycin A, rotenone and capsaicin markedly inhibited the reductions of electron acceptors in promastigotes, but not in amastigotes. Results of this study suggest that the transPMET system is functionally different in wild and resistant strains of L. donovani.

The use of biosensors by using microorganisms such as bacteria have short life cycles and provide other advantages. One colorimetric biosensor technique that has been developed is the use of a biosensor utilizing the incorporation of Prussian blue formation reactions mediated by E. coli bioreactors with ferricyanide. Immobilization is a method that allows the bacteria can be used for long-term without reducing its ability as bioreceptor. This study aimed to develop a novel and rapid immobilized bacterial biosensor for the detection of toxic compound in water and to evaluate their analytical performances. Immobilization of E. coli performed by trapping method using alginate material support. The bacterial suspension was mixed with sodium alginate (1:1 v/v), and the mixture was continuously dropped in CaCl2 solution to be a form of beads. The beads were used as bioreceptor to detect toxicants regarding cadmium, arsenic, mercury, chromium and lead solutions with Prussian blue as a colorimetric indicator. The linearity and sensitivity of detection of beads to the toxicants were tested, the stability of repeated use and storage were evaluated as well. The results showed that E. coli could be immobilized using alginate with response value was correlated with toxic concentration. The developed biosensor was more stable when used repeatedly and could be stored in a long time. The immobilization of E. coli in calcium alginate bead was successfully performed as a biosensor system for monitoring acute toxicity in water.

Quantitation without relying on the calibration curve has long been an issue of overcoming analytical problems accompanied with the inherent limitations of the calibration curve fitting errors. Here, we report on a calibration curve-free method for electrochemical quantitation based on a multi-scale gap device (MGD). The MGD is an integrated device having a series of interdigitated electrodes (IDE) with micro-to-nano gap distances. The device shows a gap-dependent redox current of the analyte when subjected to the electrochemical cycling between the two facing electrodes of its componential IDEs. Based on the fact that the current increases as the gap distance decreases, the analyte concentration could be directly estimated: the rate of increase in the current was directly proportional to the analyte concentration. The calibration curve was not necessary for the quantitation. The accuracy of this MGD approach was better than that of an IDE collection of the same gap distance, which was deteriorated at the larger gap distances particularly. The MGD-based quantitation of dopamine, potassium ferricyanide, and aminophenol was demonstrated in a relatively broad range of concentrations (100 nM-5 mM).

A non-invasive aptamer-based electrochemical biosensor using disposable screen-printed graphene electrodes (SPGEs) was developed for simple, rapid, and sensitive determination of cortisol levels. Selective detection of cortisol based on a label-free electrochemical assay was achieved by specific recognition of the cortisol DNA aptamer (CApt). The CApt was modified with streptavidin magnetic beads (MBs) before simple immobilization onto the electrode surface using a neodymium magnet. The electrochemical behavior of the aptamer-based biosensor was assessed by using electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV) (vs Ag/AgCl). The specific binding between cortisol and CApt resulted in a decrease in charge transfer resistance (Rct) from EIS using [Fe(CN)6]3−/4− with increasing cortisol concentration. Under optimal conditions, a linear range from 0.10 to 100 ng/mL with a low detection limit (3SD/slope) of 2.1 pg/mL was obtained. Furthermore, the proposed biosensing system exhibited a satisfactory recovery in the range 97.4–109.2% with 5.7–6.6% RSD in spiked artificial human sweat. Regarding the applications of this tool, the aptamer-based biosensor has potential to be a versatile and point-of-care (POC) device for simple, sensitive, selective, disposable, and low-cost cortisol detection.

Deferoxamine (DFO), an iron chelator, is used therapeutically for the removal of excess iron in multiple clinical conditions such as beta thalassemia and intracerebral hemorrhage. DFO is also used as an iron chelator and hypoxia-mimetic agent in in vivo and in vitro basic research. Here we unexpectedly discover DFO to be a nitric oxide (NO) precursor in experiments where it was intended to act as an iron chelator. Production of NO from aqueous solutions of DFO was directly observed by ozone-based chemiluminescence using a ferricyanide-based assay and was confirmed by electron paramagnetic resonance (EPR). DFO also produced NO following exposure to ultraviolet light, and its incubation with sheep adult and fetal blood resulted in considerable formation of iron nitrosyl hemoglobin, as confirmed by both visible spectroscopy and EPR. These results suggest that experiments using DFO can be confounded by concomitant production of NO, and offer new insight into some of DFO's unexplained clinical side effects such as hypotension.

A novel ferricyanide/Prussian blue (PB) assay for total antioxidant capacity (TAC) determination was developed exploiting the formation of PB nanoparticles in the presence of polyvinylpyrrolidone (PVP) as stabilizer. This improved method, named as "nanoparticle-based ferricyanide/Prussian blue assay (PBNP)", was applied to the TAC measurement of Cynara Scolymus L. (globe artichoke). The calibration results of the novel (PBNP) method were compared with those of a similar nanoparticle PB method performed in the absence of PVP, and of a sodium dodecyl sulfate-modified and acid-optimized ferricyanide reference assay. Compared to similar common Fe(III)-based TAC assays, much higher molar absorptivities, pointing out higher response to different kinds of antioxidants, were obtained with PBNP for all tested antioxidants, and lower LOD and LOQ values were achieved for thiols. As an additional advantage, methionine, not responding to other electron-transfer based TAC reagents, could be measured. PBNP could detect various antioxidants with one-two orders-of-magnitude lower LOD values than those of widely used TAC assays like CUPRAC and Folin-Ciocalteau well correlating with the proposed assay.

8-Oxo-7,8-dihydro-2’-deoxyguanosine (8-oxodGuo) is a common primary product of cellular oxidative DNA damage. 8-OxodGuo is more readily oxidized than 2’-deoxyguanosine (dG); a two-electron oxidation generates a highly reactive intermediate (OG(ox)), which forms covalent adducts with nucleophiles, including OH(−), free amines, and the side chains of amino acids such as lysine. We determined here that K(3)Fe(CN)(6) oxidation of 8-oxodGuo in nucleosome core particles (NCPs) produces high yields, quantitative (i.e. 100%) in some cases, of DNA-protein cross-links (DPCs). The efficiency of DPC formation was closely related to 8-oxodGuo base pairing and location within the NCP and was only slightly decreased by adding the DNA-protective polyamine spermine to the system. Using NCPs that contained histone mutants, we determined that DPCs result predominantly from OG(ox) trapping by the N-terminal histone amine. The DPCs were stable under physiological conditions and therefore could have important biological consequences. For instance, the essentially quantitative yield of DPCs at some positions within NCPs would reduce the yield of the mutagenic DNA lesions spiroiminodihydantoin and guanidinohydantoin produced from the common intermediate OG(ox), which in turn would affect mutation signatures of oxidative stress in a position dependent manner. In summary, our findings indicate that site-specific incorporation of 8-oxodGuo into NCPs, followed by its oxidation, leads to DPCs with an efficiency depending on 8-oxodGuo location and orientation. Given that 8-oxodGuo formation is widespread in genomic DNA and that DPC formation is highly efficient, DPCs may occur in eukaryotic cells and may affect several important biological processes.

Electrochemical enzyme-linked sandwich assays on magnetic beads (MBs) refer to one of the most sensitive approaches for analysis of nonamplified nucleic acid samples, with redox enzymes being routinely used as labels. Here, we report a sensitive and inexpensive electrochemical nucleic acid sandwich assay on MBs that exploits a hydrolytic enzyme cellulase as a label, while MBs are used for preconcentration and bioseparation of analyzed samples. Binding of target DNA or RNA to capture DNA-modified MB triggers sandwich assembly and its labeling with cellulase. Application of the assembled sandwich to the electrodes covered with insulating nitrocellulose films induces film digestion by the cellulase label and pronounced changes in the electrical properties of the electrodes, the extent of the changes being proportional to the concentration of the analyzed nucleic acids. Down to 1 amol of Lactobacillus brevis specific synthetic DNA and rRNA isolated from L. brevis cells could be detected in 1 mL samples in the overall from 2 to 3 h assay. The assay is universal and can be adapted for point-of-care-testing and for in-field environmental and microbiomic analysis of unamplified samples of any DNA/RNA sequences.

The present report evaluates the effect of global perinatal asphyxia on several parameters of oxidative stress and cell viability in rat brain tissue sampled at an extended neonatal period up to 14 days, a period characterised by intensive neuritogenesis, synaptogenesis, synaptic consolidation, pruning and delayed cell death. Perinatal asphyxia was induced by immersing foetus-containing uterine horns removed by a caesarean section from on term rat dams into a water bath at 37 °C for 21 min. Asphyxia-exposed and sibling caesarean-delivered foetuses were manually resucitated and nurtured by surrogate dams for 1 to 14 postnatal (P) days. Brain samples (mesencephalon, telencephalon and hippocampus) were assayed for glutathione (reduced and oxidated levels; spectrophotometry), tissue reducing capacity (potassium ferricyanide reducing assay, FRAP), catalase (the key enzyme protecting against oxidative stress and reactive oxygen species, Western blots and ELISA) and cleaved caspase-3 (the key executioner of apoptosis, Western blots) levels. It was found that global PA produced a regionally specific and sustained increase in GSSG/GSH ratio, a regionally specific decrease in tissue reducing capacity and a regionally and time specific decrease of catalase activity and increase of cleaved caspase-3 levels. The present study provides evidence for regionally impaired redox homeostasis in the brain of rats subjected to global PA, an effect observed up to P14, mainly affecting mesencephalon and hippocampus, suggesting a sustained oxidative stress after the posthypoxia period. The oxidative stress observed postnatally can in part be associated to a respiratory apneic-like deficit, since there was a statistically significant decrease in respiration frequency in AS compared to CS neonates, also up to P14, together with the signs of a decreased peripheral blood perfusion (pink-blue skin colour in AS, compared to the pink colour observed in all CS neonates). It is proposed that PA implies a long-term metabolic insult, triggered by the length of hypoxia, the resuscitation/reoxigenation manoevres, but also by the developmental stage of the affected brain regions, and the integrity of cardiovascular and respiratory physiological functions, which are fundamental for warrantying a proper development.

Hypothesis Electrosynthesis of the MIP nano-film after binding of the separated domains or holo-cytochrome BM3 via an engineered anchor should result in domain-specific cavities in the polymer layer. Experiments Both the two domains and the holo P450 BM3 have been bound prior polymer deposition via a N-terminal engineered his6-anchor to the electrode surface. Each step of MIP preparation was characterized by cyclic voltammetry of the redox-marker ferricyanide. Rebinding after template removal was evaluated by quantifying the suppression of the diffusive permeability of the signal for ferricyanide and by the NADH-dependent reduction of cytochrome c by the reductase domain (BMR). Findings The working hypothesis is verified by the discrimination of the two domains by the respective MIPs: The holoenzyme P450 BM3 was ca. 5.5 times more effectively recognized by the film imprinted with the oxidase domain (BMO) as compared to the BMR-MIP or the non-imprinted polymer (NIP). Obviously, a cavity is formed during the imprinting process around the his6-tag-anchored BMR which cannot accommodate the broader BMO or the P450 BM3. The affinity of the MIP towards P450 BM3 is comparable with that to the monomer in solution. The his6-tagged P450 BM3 binds (30 percent) stronger which shows the additive effect of the interaction with the MIP and the binding to the electrode.

The study of trans-plasma membrane electron transport (tPMET) in oncogenic systems is paramount to the further understanding of cancer biology. The current literature provides methodology to study these systems that hinges upon mitochondrial knockout genotypes in conjunction with cell surface oxygen consumption, or the detection of an electron acceptor using colorimetric methods. However, when using an iron redox based system to probe tPMET, there is yet to be a method that allows for the simultaneous quantification of iron redox states while providing an exceptional level of sensitivity. Developing a method to simultaneously analyze the redox state of a reporter molecule would give advantages in probing the underlying biology. Herein, we present an electrochemical based method that allows for the quantification of both ferricyanide and ferrocyanide redox states to a highly sensitive degree. We have applied this system to a novel application of assessing oncogenic cell-driven iron reduction and have shown that it can effectively quantitate and identify differences in iron reduction capability of three lung epithelial cell lines.

The reductase domains of neuronal NOS, endothelial NOS and two constitutive nitric oxide synthase (cNOS) share higher sequence similarity (>60%). In order to evaluate the role of ferredoxin‑NADP+ reductase (FNR) module in adjusting NOS catalytic activities, chimeras were by interchanging the FNR‑like module between endothelial NOS and neuronal NOS in the present study. The assays of steady‑state enzymatic activities for cytochrome c and ferricyanide reduction, NO synthesis and NADPH oxidation were performed spectrophotometrically. The two NOS FNR modules transferred their ferricyanide reductase character to the chimera enzymes. Results showed that the FNR module was important in adjusting electrons flow through the reductase domain and out of the FMN module. Results indicated that the FNR module was critical in controlling the electron transfer capacities of the FMN module.

An electrochemical molecular sensor based on a new principle is reported. Nereistoxin (NRT, 4-N,N-dimethylamino-1,2-dithiolane), a naturally occurring neurotoxin (nicotinic acetylcholine receptor agonist), was adsorbed on an Au electrode via Au-S covalent bonding and accelerated the electron transfer between the electrode and the marker, ferricyanide anion. The contrast between the electrochemical responses obtained with the bare and NRT-modified Au electrodes was more pronounced at a low ionic strength of the supporting electrolyte, KCl. In the presence of 1 mM KCl, almost a 0/1 contrast between the signals was obtained through electrostatic interaction between the protonated tertiary amino group of NRT and the anionic ferricyanide ion. No current was observed with an electrode modified with mercaptopropionic acid. An unusually low ionic strength thickened the electric double layer to the degree where current was not observed with the bare electrode. The effect of the electrostatic concentration of the marker ion becomes obvious under such conditions. Commercially available NRT-related pesticides such as Cartap and Bensultap were also detected using the same format after pretreatments by hydrolysis/reduction. The present sensing method was successfully applied to human serum with satisfactory sensitivity.

In this research scanning electrochemical microscopy was applied for the investigation of immobilized yeast Saccharomyces cerevisiae cells. Two redox mediators based system was applied in order to increase the efficiency of charge transfer from yeast cells. 9,10-phenanthrenequinone (PQ) was applied as a lipophilic redox mediator, which has the ability to cross the cell’s membrane; another redox mediator was ferricyanide, which acted as a hydrophylic electron acceptor able to transfer electrons from the PQ to the working electrode of SECM. Hill’s function was applied to determine the optimal pH for this described SECM-based system. The influence of pH on cell viability could be well described by Hill’s function. It was determined that at pH 6.5 the PQ has a minimal toxic influence on yeast cells, and the kinetics of metabolic processes in cells as well as electron transfer rate achieved in consecutive action of both redox mediators were appropriate to achieve optimal current signals.

The autosomal recessive-hyper immunoglobulin E syndromes (AR-HIES) are inherited inborn primary immunodeficiency disorders caused mainly by mutations in the dedicator of cytokinesis 8 (DOCK8) gene. A method is described for the selection of DNA aptamers against DOCK8 protein. The selection was performed by using a gold electrode as the solid matrix for immobilization of DOCK8. This enables voltammetric monitoring of the bound DNA after each selection cycle. After eight rounds of selection, high affinity DNA aptamers for DOCK8 were identified with dissociation constants (Kds) ranging from 3.3 to 66 nM. The aptamer which a Kd of 8.8 nM was used in an aptasensor. A gold electrode was modified by self-assembly of the thiolated aptamer, and the response to the DOCK8 protein was detected by monitoring the change in the electron transfer resistance using the ferro/ferricyanide system as a redox probe. The aptasensor works in the 100 pg.mL−1 to 100 ng.mL−1 DOCK8 concentration range, has a detection limit of 81 pg.mL−1 and good selectivity over other proteins in the serum.

Respiration substrate (RS)-less isolation method was developed for enhancing the sensitivity of acute toxicity assessment of heavy metal ions. RS was removed from the first step of previous isolation method, which was an effective strategy for improving acute toxicity assessment. 50% inhibiting concentration (IC50) values of Cu2+, Cd2+, Zn2+, Hg2+ and Ni2+ were 0.39 mg L−1, 5.99 mg L−1, 3.99 mg L−1, 0.23 mg L−1 and 5.74 mg L−1, respectively. Beyond that, the complicacy of organic toxicants assessments was investigated by choosing 3,5-dichlorophenol (DCP) as model toxicant. Biofilm sensor, morphology method and suspended microbes-based methods including one-pot method, RS-isolation method, RS-less isolation method, RS-less isolation method with added potassium ferricyanide (+F), were compared. The sensitivity to DCP can be ranked as morphology method > suspended microbes-based methods > biofilm method. The difference of the present results implicated that the methodological interference, leading in different detection mechanisms of these methods. The relative investigations can provide theoretical guidance for developing comprehensive detection methods of pollutants.

Preparation and electrochemical interrogation of a novel redox active progesterone derivative progesterone thiosemicarbazone (PATC) is presented here together with an investigation into its suitability as conjugate in progesterone hormone immunosensing. PATC synthesis involved a condensation reaction between progesterone acetate and thiosemicarbazone hydrochloride. Voltammetric and pulse techniques confirmed the redox behaviour of the new compound with concentration and scan rate dependant irreversible behaviour evident at glassy carbon and gold transducers - ko (standard heterogeneous rate constant) was 2.56 � 10-3 cm2 /s (ν ¼ 100 mV/s in non-aqeuous media). Bioaffinity studies towards anti-progesterone antibodies involved a competitive ELISA format (optical) which confirmed recognition of the new progesterone derivative. Electrochemical impedance spectroscopy was employed as an interrogation technique in order to establish optimum binding and surface conditions for progesterone antigen-antibody interaction with the assistance of a redox probe (potassium hexacyanoferrate).

Polyhexamethylene biguanide (PHMB) is a cationic disinfectant widely used for personal-care products and for sanitizers in swimming pools. This paper describes a promotion effect of PHMB on a glucose oxidase (GOx) enzymatic reaction with ferricyanide ion and its analytical application. The promotion effect arose from a polyion complex formation between polycationic PHMB and polyanionic GOx. The promoted GOx reaction was analyzed by Michaelis-Menten equation, and the Michalis constant and catalytic constant were estimated to be 240 μM and 31 s-1, respectively. Utilizing the promotion effect, PHMB was successfully determined in the range of 0.05 to 0.40 ppm, and the detection limit was calculated to be 0.027 ppm. The visual detection and semi-determination of PHMB with the same concentration level were also possible. As an application, the method was applied to the determination of PHMB in a contact lens detergent and its model solution.

Herein, a simple and selective electrochemical method was developed for sulfadimethoxine detection based on the triggered cleavage activity of nuclease P1 by the formation of aptamer and sulfadimethoxine conjugate. After probe DNA was immobilized on gold electrode surface, aptamer DNA labeled with biotin at its 5′-terminal was then captured on electrode surface through the hybridization reaction between probe DNA and aptamer DNA. The formed double-stranded DNA (dsDNA) can block the digestion activity of Nuclease P1 towards the single-stranded probe DNA. Then, the anti-dsDNA antibody was further modified on electrode surface based on the specific interaction between dsDNA and antibody. Due to the electrostatic repulsion effect and steric-hindrance effect, a weak electrochemical signal was obtained at this electrode. However, in the presence of sulfadimethoxine, it can interact with aptamer DNA, and then the formation of dsDNA can be blocked. As a result, the probe DNA at its single-strand state can be digested by Nuclease P1, which leads to the failure of the immobilization of anti-dsDNA antibody. At this state, a strong electrochemical signal was obtained. Based on the change of the electrochemical signal, sulfadimethoxine can be detected with linear range of 0.1–500 nmol/L. The detection limit was 0.038 nmol/L. The developed method possesses high detection selectivity and sensitivity. The applicability of this method was also proved by detecting sulfadimethoxine in veterinary drug and milk with satisfactory results.

NAD(P)H:quinone oxidoreductase 1 (NQO1) is a key enzyme providing cytoprotection from quinone species. In addition, it is expressed at high levels in many human tumors, such as breast cancer. Therefore, it is considered to be a potential target in cancer treatment. In order to detect intracellular NQO1 activity in MCF-7 aggregates as a cancer model, we present, in this study, a double-mediator system combined with large-scale integration (LSI)-based amperometric devices. This LSI device contained 20 × 20 Pt working electrodes with a 250 μm pitch for electrochemical imaging. In the detection system, menadione (MD) and [Fe(CN)6]3- were used. Since MD can diffuse into cells due to its hydrophobicity, it is reduced into menadiol by intracellular NQO1. The menadiol diffuses out of the cells and reduces [Fe(CN)6]3- of a hydrophilic mediator into [Fe(CN)6]4-. The accumulated [Fe(CN)6]4- outside the cells is electrochemically detected at 0.5 V in the LSI device. Using this strategy, the intracellular NQO1 activity of MCF-7 aggregates was successfully detected. The effect of rotenone, which is an inhibitor for Complex I, on NQO1 activity was also investigated. In addition, NQO1 and respiration activities were simultaneously imaged using the detection system that was further combined with electrochemicolor imaging. Thus, the double-mediator system was proven to be useful for evaluating intracellular redox activity of cell aggregates.

Background: Sesamol is a widely used antioxidant for the food and pharmaceutical industries. The oxidation products of this compound may be accumulated in foods or ingested. Little is known about its effect on human health. Objective: It is of great interest to identify the oxidation products of sesamol that may be beneficial to humans. This study was undertaken to identify the oxidation products of sesamol and investigate their antioxidant and cytotoxic activities. Materials and Methods: Using the ferricyanide oxidation approach, four oxidation products of sesamol (2, 3, 20 & 21) have been identified. Structural elucidation of these compounds was established on the basis of their detailed NMR spectroscopic analysis, mass spectrometry and x-ray crystallography. Additionally, a formation mechanism of compound 20 was proposed based on high-resolution mass spectrometry-fragmentation method. The antioxidant activities of these compounds were determined by the DPPH, FRAP, and ABTS assays. The in vitro antiproliferative activity of these compounds was evaluated against a panel of human cancer cell lines as well as non-cancerous cells. Results: Two oxidation products of sesamol were found to contain an unusual methylenedioxy ring-opening skeleton, as evidenced by spectroscopic and x-ray crystallographic data. Among all compounds, 20 displayed impressive antiproliferative activities against a panel of human cancer cell lines yet remained non-toxic to noncancerous cells. The antioxidant activities of compound 20 are significantly weaker than sesamol as determined by the DPPH, FRAP, and ABTS assays. Conclusion:: The oxidation products of sesamol could be a valuable source of bioactive molecules. Compound 20 may be used as a potential lead molecule for cancer studies.

We characterize chemical reduction of a nitroxide biradical, TOTAPOL, used in dynamic nuclear polarization (DNP) experiments, specifically probing the stability in whole-cell pellets and lysates, and present a few strategies to stabilize the biradicals for DNP studies. DNP solid-state NMR experiments use paramagnetic species such as nitroxide biradicals to dramatically increase NMR signals. Although there is considerable excitement about using nitroxide-based DNP for detecting the NMR spectra of proteins in whole cells, nitroxide radicals are reduced in minutes in bacterial cell pellets, which we confirm and quantify here. We show that addition of the covalent cysteine blocker N-ethylmaleimide to whole cells significantly slows the rate of reduction, suggesting that cysteine thiol radicals are important toin vivoradical reduction. The use of cell lysates rather than whole cells also slows TOTAPOL reduction, which suggests a possible role for the periplasm and oxidative phosphorylation metabolites in radical degradation. Reduced TOTAPOL in lysates can also be efficiently reoxidized with potassium ferricyanide. These results point to a practical and robust set of strategies for DNP of cellular preparations.

Chicken feather-derived high-surface-area porous activated carbon (CFAC) material was prepared using chemical activation. A new composite composed of Ru-Pd nanoparticles supported on CFAC (Ru-Pd@CFAC) has been prepared by microwave-thermal reduction in the presence of the support. Characterization by XRD, Raman, BET, FE-SEM/TEM, FT-IR, TGA, XPS, HAADF-STEM-EDS, H2 -chemisorption, H2 -TPR, and ICP-AES was used to analyze the catalyst. This catalyst is found to be efficient for the reduction of hexavalent chromium (CrVI ), potassium ferricyanide (K3 [Fe(CN)6 ]), 4-nitrophenol (4-NP), and pendimethalin (PDM), at room temperature, and remains stable, even after several repeated runs. Moreover, it showed excellent catalytic activity compared with the monometallic counterparts.

Second-generation glucose biosensors are presently the mainstream commercial solution for blood glucose measurement of diabetic patients. Screen-printed carbon electrodes (SPCEs) are the most-used substrate for glucose testing strips. This study adopted hydrophilic and positively charged &alpha, poly-l-lysine (&alpha, PLL) as the entrapment matrix for the immobilization of negatively charged glucose oxidase (GOx) and ferricyanide (FIC) on SPCEs to construct a disposable second-generation glucose biosensor. The &alpha, PLL modification is shown to facilitate the redox kinetics of FIC and ferrocyanide on the SPCEs. The SPCEs coated with 0.5 mM GOx, 99.5 mM FIC, and 5 mM &alpha, PLL had better sensitivity for glucose detection due to the appreciable effect of protonated &alpha, PLL on the promotion of electron transfer between GOx and FIC. Moreover, the SPCEs coated with 0.5 mM GOx, 99.5 mM FIC, and 5 mM &alpha, PLL were packaged as blood glucose testing strips for the measurement of glucose-containing human serum samples. The glucose testing strips had good linearity from 2.8 mM to 27.5 mM and a detection limit of 2.3 mM. Moreover, the 5 mM &alpha, PLL-based glucose testing strips had good long-term stability to maintain GOx activity in aging tests at 50 &deg, C.

Nitric oxide (NO) has shown positive effects in tumor treatment. However, controlling NO release in specific targets is still a crucial challenge for antitumor therapy. Considering that sodium nitroprusside (SNP) and potassium ferricyanide have similar chemical structures, a near infrared (NIR) laser-controlled NO release nanoplatform has been fabricated by allowing SNP to participate in mesoporous Prussian blue (m-PB) nanoparticle formation. The resulting SNP-doped m-PB (m-PB-NO) exhibited a good NIR-controlled NO release behavior, and the amount of NO released can be controlled by adjusting the laser intensity and irradiation time. Given that m-PB-NO still has strong absorption in NIR region, it exhibited an excellent photothermal effect in vitro and in vivo. After carrying antitumor drug, docetaxel (DTX)-loaded m-PB-NO (DTX@m-PB-NO) can simultaneously achieve NIR-controlled NO release, good photothermotherapy, and chemotherapy. The combination therapy of DTX@m-PB-NO showed a significant synergistic effect compared with each monotherapy and can significantly improve the therapeutic effect. Combination therapy also significantly inhibited the lung metastasis of 4T1 breast cancer cells in tumor-bearing mice by ablating primary tumors.

In biosensors fabrication, entrapment in polymeric matrices allows efficient immobilization of the biorecognition elements without compromising their structure and activity. When considering living cells, the biocompatibility of both the matrix and the polymerization procedure are additional critical factors. Bio-polymeric gels (e.g. alginate) are biocompatible and polymerize under mild conditions, but they have poor stability. Most synthetic polymers (e.g. PVA), on the other hand, present improved stability at the expense of complex protocols involving chemical/physical treatments that decrease their biological compatibility. In an attempt to explore new solutions to this problem we have developed a procedure for the immobilization of bacterial cells in polyethersulfone (PES) using phase separation. The technology has been tested successfully in the construction of a bacterial biosensor for toxicity assessment. Biosensors were coated with a 300 μm bacteria-containing PES membrane, using non-solvent induced phase separation (membrane thickness ≈ 300 μm). With this method, up to 2.3 × 106 cells were immobilized in the electrode surface with an entrapment efficiency of 8.2%, without compromising cell integrity or viability. Biosensing was performed electrochemically through ferricyanide respirometry, with metabolically-active entrapped bacteria reducing ferricyanide in the presence of glucose. PES biosensors showed good stability and reusability during dry frozen storage for up to 1 month. The analytical performance of the sensors was assessed carrying out a toxicity assay in which 3,5-dichlorophenol (DCP) was used as a model toxic compound. The biosensor provided a concentration-dependent response to DCP with half-maximal effective concentration (EC50) of 9.2 ppm, well in agreement with reported values. This entrapment methodology is susceptible of mass production and allows easy and repetitive production of robust and sensitive bacterial biosensors.

Chemiluminescence (CL) assay is a promising point-of-care testing (POCT) technology due to the fast response, high sensitivity, and easy miniaturization. The application and performance of CL POCT method were highly dependent on the CL reaction. Herein, based on the CL reaction between luminol and in-situ generated K3Fe(CN)6, a low-cost, enzyme-free, and label-free CL POCT method was explored via a portable and handheld luminometer to detect telomerase activity. Telomerase elongated telomere substrate (TS) primer to form (TTAGGG)n repeats which hybridize with multiple short DNAs. The intercalation of SYBR Green I (SGI) into double-stranded DNA (dsDNA) generated singlet oxygen under the irradiation of LED light source. Singlet oxygen was then employed for in-situ oxidation of K4Fe(CN)6 to K3Fe(CN)6, which could react with luminol to generate a strong CL intensity. Thus, telomerase activity could be specifically, sensitively, and label-free detected. The detection limit was down to 98 HeLa cells. The detection process was very simple, and the cost was about $0.01 for each measurement. Furthermore, telomerase activity was detectable in human serum samples, with spike recoveries from 96% to 105%. According to our knowledge, it is the first effort to develop a low-cost, label-free and enzyme-free CL method with good repeatability for detecting biomarker based on the analyte-triggered and in-situ generated K3Fe(CN)6/luminol CL reaction.

We describe a new colorimetric assay method using glucose oxidase (GOx) to detect ε-poly-l-lysine (εPL). This method uses εPL's remarkable effect of promoting the enzymatic reaction of GOx with ferricyanide ion. This reaction reduces ferricyanide ion to ferrocyanide ion, accompanied by a color change from yellow to colorless. In this colorimetric assay, the detection limit of εPL was estimated to be approximately 0.5 mg/L when purified εPL samples were used. εPL has usually been produced by a fermentation process using Streptomyces albulus species. The components of the culture broth showed interference effects against the assay method. However, due to the high sensitivity of the assay method for εPL, εPL could be detected in the culture broth without any pretreatment. The detectable concentration of εPL in the culture broth, cPL,ac, was estimated to be approximately 20 mg/L. By combining the Berlin blue reaction with this method, the cPL,ac was reduced to 10 mg/L. In light of the proposed method's simplicity and sensitivity, it could be useful for screening εPL synthetic enzymes and microorganisms.

Acetaminophen (ACT) is one of the most frequently detected pharmaceuticals in aqueous environments, and treatment of ACT were generally carried out by photocatalytic degradations under high energy UV irradiation. In this study, potassium ferricyanide was utilized as a quadruple-elemental dopant in a TiO2 photocatalyst in order to enhance its visible-light activity. Two critical parameters (amounts of dopants and durations of calcination) of the synthesis of the photocatalyst by a sol–gel method were systematically evaluated. Crystal structure of the doping TiO2 was examined by X-ray diffraction while the effects of the two parameters on the photocatalytic activity were elucidated by various characterizations. Increasing the amount of dopant or the duration of calcination red-shifted the UV–vis DRS of the doped TiO2. The estimated band gap energy of the doped TiO2 decreased slightly as the amount of dopant increased, but it increased as the duration of calcination increased. The FT-IR yielded characteristic peaks that revealed the effects of the two parameters, whereas the SEM images revealed the morphological evolutions of each effect. The photocatalyst, synthesized at optimum conditions was able to remove 99.1 % acetaminophen with rate constant of 7.9 × 10−3 min−1, which was 4.88 times greater than virgin TiO2. In general, this study not only optimized synthetic conditions of the new visible-light active photocatalyst for ACT degradation but also presented characterizations conducted by SEM, XRD, UV–vis DRS, and FTIR to elucidate the relationship between modified structure and the photocatalytic activity.

Telomerase is closely related to cancers, which makes it one of the most widely known tumor marker. Recently, many methods have been reported for telomerase activity measurement in which complex label procedures were commonly used. In this paper, a label-free method for detection of telomerase activity in urine based on steric hindrance changes induced by confinement geometry in the porous anodic alumina (PAA) nanochannels was proposed. Telomerase substrate (TS) primer was first assembled on the inside wall of PAA nanochannels by Schiff reaction under mild conditions. Then, under the action of telomerase, TS primer was amplified and extended to repeating G-rich sequences (TTAGGG)x, which formed multiplex G-quadruplex in the presence of potassium ions (K(+)). This configurational change led to the increment of steric hindrance in the nanochannels, resulting in the decrement of anodic current of potassium ferricyanide (K3[Fe(CN)6]). Compared with previously reported methods based on PAA nanochannels (usually one G-quadruplex formed), multiplex repeating G-quadruplex formed on one TS primer in this work. As a result, large current drop (∼3.6 μA, 36%) was obtained, which gave facility to improve the detection sensitivity. The decreased ratio of anodic current has a linear correlation with the logarithm of HeLa cell number in the range of 10-5000 cells, with the detection limit of seven cells. The method is simple, reliable, and has been successfully applied in the detection of telomerase in urine with good accuracy, selectivity and reproducibility. In addition, the method is nondestructive test compared to blood analysis and pathology tests, which is significant for cancer discovery, development, and prognosis.

Fruits, vegetables and medicinal herbs rich in phenolics antioxidants contribute toward reduced risk of age-related diseases and cancer. In this study, Psidium guajava leaf extract was fractionated in various organic solvents viz. petroleum ether, benzene, ethyl acetate, ethanl and methanol and tested for their antioxidant and antimutagenic properties. Methanolic fraction showed maximum antioxidant activity comparable to ascorbic acid and butylated hydroxyl toluene (BHT) as tested by DPPH free radical scavenging, phosphomolybdenum, FRAP (Fe3 + reducing power) and CUPRAC (cupric ions (Cu2+) reducing ability) assays. The fraction was analyzed for antimutagenic activities against sodium azide (NaN3), methylmethane sulfonate (MMS), 2-aminofluorene (2AF) and benzo(a)pyrene (BP) in Ames Salmonella tester strains. The methanol extracted fraction at 80 μg/ml concentration inhibited above 70% mutagenicity. Further, phytochemical analysis of methanol fraction that was found to be most active revealed the pr...

Arbuscular mycorrhizal fungi (AMF) are integral functioning parts of plant root systems and are widely recognized for enhancing contaminants uptake and metabolism on severely disturbed sites. However, the patterns of their influence on the phytoremediation of iron-cyanide (Fe-CN) complexes are unknown. Fe-CN complexes are of great common interest, as iron is one of the most abundant element in soil and water. Effect of ryegrass (Lolium perenne L.) roots inoculation, using mycorrhizal fungi (Rhizophagus irregularis and a mixture of R. irregularis, Funneliformis mosseae, Rhizophagus aggregatus, and Claroideoglomus etunicatum), on iron-cyanide sorption was studied. Results indicated significantly higher colonization of R. irregularis than the mixture of AMF species on ryegrass roots. Series of batch experiments using potassium hexacyanoferrate (II) solutions, in varying concentrations revealed significantly higher reduction of total CN and free CN content in the mycorrhizal roots, indicating greater cyanide decrease in the treatment inoculated with R. irregularis. Our study is a first indication of the possible positive contribution of AM fungi on the phytoremediation of iron-cyanide complexes.

The synthesis of gold nanoparticles (AuNPs) having better dispersibility and catalytic ability than the conventional AuNPs is the challenging task. The fact that aldehydes and ketones results in the formation of catalytic hybrid material with amino functionalized silanes directed the use of carbonyl functional group (aldehydes and ketones) specifically formaldehyde, acetaldehyde, acetone and t-butyl methyl ketone alongwith 3-aminopropyltrimethoxysilane (3-APTMS) to meet such requirement. Accordingly, a comparative study on the synthesis of 3-APTMS and organic reducing agent mediated synthesis of AuNPs are reported herein. The findings reveal that 3-APTMS capped gold ions are converted into AuNPs with precise control of pH- and salt- sensitivity. The major findings reveal the following: (1) 3-APTMS being amphiphilic, dispersibility of as prepared AuNPs largely depends on the organic reducing agents. (2) An increase in the hydrocarbon content of the reducing agent facilitate the dispersibility of AuNPs in organic solvent whereas decrease of the same increases the dispersibility in water, (3) AuNPs made through aldehydic reducing agents (formaldehyde and acetaldehyde) have relatively better salt and pH tolerance as compared to ketonic reducing agents (acetone, t-butyl methyl ketone), and (4) an increase in 3-APTMS concentrations imparts better salt- and pH- resistant property to AuNPs irrespective of organic reducing agents. A typical example on the role of AuNPs in homogeneous catalysis during potassium ferricyanide mediated oxidation of ascorbic acid is also reported.

A proton-driven molecular pump is devised using a surface-confined dynamic 3D DNA scaffold. A dynamic DNA tetrahedral nanostructure is designed by incorporating a pH-sensitive i-motif sequence in one edge, which serves as the scaffold to ensure highly ordered orientation and spatial isolation of this nanomachine on the macroscopic gold surface. It is found that the switching ability of this dynamic tetrahedron is fully maintained on the surface. Importantly, this proton-driven nanomachine can reversibly pump water and ferricynide in response to pH variation in solution.

To find an efficient and cheap system for NAD+ regeneration A NADH-ferricyanide dehydrogenase was obtained from an isolate of Escherichia coli. Optimal activity of the NADH dehydrogenase was at 45 °C and pH 7.5, with a K m value for NADH of 10 μM. By combining the NADH dehydrogenase, potassium ferricyanide and laccase, a bi-enzyme system for NAD+ regeneration was established. The system is attractive in that the O2 consumed by laccase is from air and the sole byproduct of the reaction is water. During the reaction process, 10 mM NAD+ was transformed from NADH in less than 2 h under the condition of 0.5 U NADH dehydrogenase, 0.5 U laccase, 0.1 mM potassium ferricyanide at pH 5.6, 30 °C The bi-enzyme system employed the NADH-ferricyanide dehydrogenase and laccase as catalysts, and potassium ferricyanide as redox mediator, is a promising alternative for NAD+ regeneration.

Electrochemical or faradaic impedance spectroscopy (EIS) using the ferri/ferrocyanide couple as a redox probe at gold working electrodes was evaluated with respect to its ability to monitor consecutive surface modification steps. As a model reaction, the reversible hybridization and dehybridization of DNA was studied. Thiol-modified single stranded DNA (ssDNA, 20 bases, capture probe) was chemisorbed to a gold electrode and treated with a solution of short thiols to release nonspecifically adsorbed DNA before hybridization with complementary ssDNA (20 bases, target) was carried out. Reversible dehybridization was achieved by intense rinsing with pure water. The experimental procedures were optimized by kinetic surface plasmon resonance (SPR) and quartz crystal microbalance with dissipation (QCM-D) measurements to maximize the increase in reflectivity or decrease in frequency upon hybridization before hybridization/dehybridization was also monitored by EIS. In contrast to SPR and QCM-D, repeatable EIS measurements were not possible at first. Combined SPR/EIS and QCM-D/EIS measurements revealed that during EIS the gold surface is seriously damaged due to the presence of CN(-) ions, which are released from the ferri/ferrocyanide redox probe. Even at optimized experimental conditions, etching the gold electrodes could not be completely suppressed and the repeatability of the EIS measurements was limited. In three out of four experimental runs, only two hybridization/dehybridization steps could be monitored reversibly by EIS. Thereafter etching the gold electrode significantly contributed to the EIS spectra whereas the QCM-D response was still repeatable. Hence great care has to be taken when this technique is used to monitor surface modification at gold electrodes.

The development of an electrochemical immunosensor for the detection of human chorionic gonadotropin (hCG) is described with a limit of detection as low as 0.3 pg mL(-1) in phosphate buffer. In this immunosensor, cysteamine (Cys) and gold nanoparticles (AuNPs) were used to immobilize an anti-hCG monoclonal antibody onto a gold electrode (GE). The structure of AuNPs has been confirmed by EDS, SEM, and TEM analysis. Due to the large specific surface area and excellent electrical conductivity of AuNPs, electron transfer was promoted and the amount of hCG antibody was enhanced significantly. A systematic study on the effects of experimental parameters such as pH, incubation time in the hCG solution and urea solution used for experiments on the binding between the immobilized antibody and hCG has been carried out. Under optimal experimental parameters, differential pulse voltammetry (DPV) signal changes of the [Fe(CN)6](3-/4-) are used to detect hCG with two broad linear ranges: 0.001 to 0.2 and 0.2 to 60.7 ng mL(-1). The LOD value proves more sensitive in comparison with previously reported methods. The prepared immunosensor showed high sensitivity and stability. In addition, the immunosensor was successfully used for the determination of hCG in human serum.

Yeast cells are often used as a model system in various experiments. Moreover, due to their high metabolic activity, yeast cells have a potential to be applied as elements in the design of biofuel cells and biosensors. However a wider application of yeast cells in electrochemical systems is limited due to high electric resistance of their cell wall. In order to reduce this problem we have polymerized conducting polymer polypyrrole (Ppy) directly in the cell wall and/or within periplasmic membrane. In this research the formation of Ppy was induced by [Fe(CN)6](3-)ions, which were generated from K4[Fe(CN)6], which was initially added to polymerization solution. The redox process was catalyzed by oxido-reductases, which are present in the plasma membrane of yeast cells. The formation of Ppy was confirmed by spectrophotometry and atomic force microscopy. It was confirmed that the conducting polymer polypyrrole was formed within periplasmic space and/or within the cell wall of yeast cells, which were incubated in solution containing pyrrole, glucose and [Fe(CN)6](4-). After 24h drying at room temperature we have observed that Ppy-modified yeast cell walls retained their initial spherical form. In contrast to Ppy-modified cells, the walls of unmodified yeast have wrinkled after 24h drying. The viability of yeast cells in the presence of different pyrrole concentrations has been evaluated.

Objectives Many natural antioxidants have poor pharmacokinetic properties that impair their therapeutic use. For hydroxycinnamic acids (HCAs) and other phenolic antioxidants, their major drawback is their low lipophilicity and a rapid metabolism. The difluoromethyl group may be considered as a ‘lipophilic hydroxyl' due to its hydrogen bond donor and acceptor properties; this prompted us to assess it as a bioisosteric replacement of a phenolic hydroxyl for increasing the lipophilicity of HCAs. Methods Six difluoromethyl-substituted methyl cinnamates (4a-c, 5a-c) related to caffeic acid were synthesized and their antioxidant activity evaluated by chemical (FRAP, DPPH scavenging, inhibition of β-carotene bleaching, at 1–200 μm), electrochemical (differential pulse voltammetry, cyclic voltammetry) and cell-based (inhibition of lipid peroxidation in erythrocytes, at 1 and 50 μm) assays. Key fndings Analogues 4a-c and 5a-c were inactive in FRAP and DPPH assays and only those containing a free phenolic hydroxyl (4a and 5a) exhibited electrochemical activity although with high redox potentials. Compounds 4a,b and 5a,b were active in the inhibition of β-carotene bleaching assay and all analogues inhibited lipid peroxidation in the human erythrocytes assay. Conclusions Lipophilic difluoromethyl-substituted cinnamic esters retain radical scavenging capabilities that prove useful to confer antioxidant properties in a non-polar environment.

In the present research, a nanoaptasensor is proposed for electrochemical measurement of chloramphenicol (CAP). To this purpose, the nanocomposite prepared from graphene oxide and functionalized with (3‐Aminopropyl) triethoxysilane/silver nanoparticles to the abbreviated AgNPs/[NH2–Si]-f-GO, was utilized to modify the glassy carbon electrode (GCE). Furthermore, the modified electrode was also investigated using the electrochemical methods such as electrochemical impedance spectroscopy (EIS), cyclic voltammetry (CV) and differential pulse voltammetry (DPV). The AgNPs/[NH2–Si]-f-GO nanocomposite was investigated by UV–Vis spectrophotometry. Fourier transform infrared (FT-IR) spectrometry and transmission electron microscopy (TEM). Moreover, [Fe(CN)6]3-/4 solution in the role of an electrochemical probe was applied. The AgNPs/[NH2–Si]-f-GO nanocomposite was confirmed as a good layer to covalent immobilization of aptamer (Apt) onto the GCE surface. In this sense, the DPV was used as a sensitive electrochemical technique for the measurement of CAP with an appropriate linear concentration range which was found to be between 10 pM and 0.2 μM and, with a low limit of detection, it equaled 3.3 pM. CAP which was identified in the presence of other usual antibiotics existed in the real samples.

DNA 5-hydroxymethylcytosine (5-hmC) is an important epigenetic biomarker for tumorigenesis, and the loss of 5-hmC levels is associated with leukemia and melanoma cancers. However, it is a great challenge to discriminate 5-hmC from 5-methylcytosine (5-mC) using the conventional bisulfite conversion methods. Herein, we report a label-free and immobilization-free electrochemical magnetobiosensor for sensitive quantification of 5-hmC in genomic DNA based on a dual signal amplification strategy coupled with terminal deoxynucleotidyl transferase (TDT) enzymatic amplification and Ru(III) redox cycling. This screen-printed carbon electrode (SPCE)-based electrochemical magnetobiosensor shows distinct advantages of having low cost and simple fabrication and being label-free, immobilization-free, PCR-free, and radioactive-free. It exhibits high sensitivity with a detection limit of as low as 9.06 fM and a large dynamic range from 0.01 to 1000 pM. Importantly, this biosensor can discriminate 5-hmC from cytosine and 5-mC, and it can successfully detect 5-hmC in live cells.

There is a correlation between specific bases of DNA molecules and the chirality of single-walled carbon nanotubes (SWNTs), which contributes the recognition ability of DNA toward partner species of chiral SWNTs. A novel strategy of electrochemical characterization is reported here for the investigation on chirality selection of (7,6) and (6,5) SWNTs with various DNA sequences, and it is found that both DNA strand length and sequence composition significantly affected the interaction of chiral SWNTs with DNA. Then (7,6) and (6,5) SWNTs were distinguished from each other with DNA sequences chosen by electrochemical methods, which demonstrated an effective and excellent feasibility for the strategy and presented a new insight into DNA-SWNT applications. This strategy can also be applied to more chiral SWNTs and DNA sequence recognition and may serve as a prescreening method for the recognition and separation of single-chirality SWNTs, which would be a new contribution to the further development of DNA-SWNT hybrids.

Early screening of foodborne pathogenic bacteria is a key to prevent and control foodborne diseases. This study intended to develop a capillary biosensor for rapid and sensitive detection of Salmonella using the multi-column capillary for easy operation, the Fe-nanoclusters (FNCs) for signal amplification and the smart phone APP for image analysis. The multi-column capillary was successively preloaded the magnetic nanoparticle (MNP) column, the FNC column, two phosphate buffer solution with 0.05% Tween 20 (PBST) columns and the HCl column and the columns were separated by air gap. The iron spiral mixer was fabricated to accelerate the mixing, and the multi-ring magnets was developed to transfer the MNPs and their conjugates from column to column. The target bacteria were captured by the MNPs to form the magnetic bacteria and then conjugated with the FNCs to form the nanocluster bacteria. After washing with PBST, the nanocluster bacteria were transferred into the HCl column, and the iron ions were released and reacted with potassium hexacyanoferrate to form the Prussian Blue, which was finally measured and analyzed using the Hue-Saturation-Lightness color space based smartphone APP for the determination of the target bacteria. This proposed biosensor exhibited a wide linear range for detection of Salmonella typhimurium with the lower detection limit of 14 CFU/mL. The mean recovery of the target bacteria in spiked chicken samples was ~105.0%, indicating the applicability of this biosensor. The proposed biosensor had the potential for in-field detection of foodborne pathogens.

There is increasing evidence that the stability, structure, dynamics, and function of many proteins differ in cells versus in vitro. The determination of protein structure and dynamics within the native cellular environment may lead to better understanding of protein behavior. Electron spin resonance (ESR) has emerged as a technique that can report on protein structure and dynamics within cells. Nitroxide based spin labels are capable of reporting on protein dynamics, structure, and backbone flexibility but are limited due to nitroxide reduction occurring in cells. In order to overcome this limitation, we used the oxidizing agent potassium ferricyanide (K3Fe(CN)6) as well as the cleavage resistant spin label 3-malemido-PROXYL (5-MSL). Furthermore, we hypothesized that injection concentration is an important parameter regarding nitroxide reduction kinetics. By increasing the injection concentration of doubly 5-MSL labeled protein into Xenopus laevis oocytes, we found an increased nitroxide lifetime. Our work demonstrates unprecedented incubation times of 3-h in-cell and 5-h in-cytosol for double electron–electron resonance (DEER) experiments using nitroxide spin labels. This allows for more meaningful measurements of larger protein systems which may require longer incubation times for equilibration in the cellular milieu. Even longer incubation times are possible by combining our approach with more shielded nitroxides and Q-band.