Ultrasensitive DNAzyme‐Based Ca2+ Detection Boosted by Ethanol and a Solvent‐Compatible Scaffold for Aptazyme Design

Functional DNA includes aptamers and DNAzymes, and metal ions are often important for achieving the chemical functions of such DNA. Biosensors based on functional DNA have mainly been tested in aqueous buffers. By introducing organic solvents with much lower dielectric constants, the interaction between metal ions and DNA can be significantly enhanced, and this might affect the performance of DNA‐based biosensors. In this work, the effect of ethanol on the activity of the EtNa DNAzyme was studied for Ca²⁺ detection. With 30 % ethanol, the sensor has a detection limit of 1.4 μm Ca²⁺, which is a 16‐fold improvement relative to that in water. This EtNa DNAzyme is unique because other tested DNAzymes are all inhibited by 50 % ethanol. Finally, by using the EtNa DNAzyme as a scaffold, the adenosine monophosphate (AMP) aptamer was inserted to construct an aptazyme, which allowed the measurement of AMP in ethanol. In summary, this study has reported the most sensitive DNA‐based sensor for Ca²⁺, and its sensitivity and selectivity can approach those of proteins or small‐molecule ligands. This work also provides a way to measure aptamer binding in organic solvents.     

A New Na+‐Dependent RNA‐Cleaving DNAzyme with over 1000‐fold Rate Acceleration by Ethanol

Enzymes working in organic solvents are important for analytical chemistry, catalysis, and mechanistic studies. Although a few protein enzymes are highly active in organic solvents, little is known regarding nucleic acid‐based enzymes. Herein, we report the first RNA‐cleaving DNAzyme, named EtNa, that works optimally in concentrated organic solvents containing only monovalent Na⁺. The EtNa DNAzyme has a rate of 2.0 h^?1 in 54 % ethanol (with 120 mm NaCl and no divalent metal ions), and a K_d of 21 mm Na⁺. It retains activity even in 72 % ethanol as well as in DMSO. With 4 mm Na⁺, the rate in 54 % ethanol is >1000‐fold higher than that in water. We also demonstrated the use of EtNa to measuring the ethanol content in alcoholic drinks. In total, this DNAzyme has three unique features: divalent metal independent activity, Na⁺ selectivity among monovalent metals, and acceleration by organic solvents.     

An RNA‐Cleaving Catalytic DNA Accelerated by Freezing

The EtNa DNAzyme was isolated during the isopropanol precipitation step of an in vitro selection effort. Although inactive with the intended cofactor, its RNA cleavage activity was observed under a few conditions. With Na⁺, EtNa was highly active in ～50 % ethanol, whereas in water, it was highly active with Ca²⁺. In this work, we showed that the EtNa DNAzyme was accelerated by freezing in water in the presence of Na⁺. The apparent K_d value reached 6.2 mm Na⁺ under the frozen condition, over 20 times tighter than that in water at room temperature. With 10 mm Na⁺, EtNa had a cleavage rate of 0.12 h^?1 after freezing at ?20 °C. This effect was unique to EtNa, as all other tested DNAzymes were inhibited by freezing except for the Na⁺‐specific NaA43. Freezing also inhibited EtNa if Ca²⁺ was used. We attributed this to the concentrations of EtNa and Na⁺ in the micropockets between ice crystals, but divalent metals might misfold DNA. Overall, we have systematically studied the effect of freezing on the RNA‐cleavage activity of DNAzymes. The DNAzyme sequence and the metal ion species are both crucial to determine the effect of freezing.     

Competitive binding of divalent cations to poly(α-hydroxyacrylic acid)

The competitive binding of divalent cations (Mg²⁺, Ca²⁺, Mn²⁺, Co²⁺, Ni²⁺, Cu²⁺ and Zn²⁺) on poly(α-hydroxyacrylic acid) (PHA) and poly(acrylic acid) (PAA) was investigated by equilibrium dialysis. In the Mg/Ca mixed system, binding selectivity for Ca²⁺ over Mg²⁺ was significantly higher in PHA than in PAA; this was attributed to coordination of α-OH groups on PHA to Ca²⁺. The binding ability and selectivity for the transition metal cations were almost the same for PHA and PAA at neutral pH, while PHA maintained appreciably higher degrees of binding than PAA in acidic solutions (pH about 3). This cation binding ability of PHA was ascribed to the lower pK_a value in the relevant pH region. © 1998 SCI.     

Two Completely Different Mechanisms for Highly Specific Na+ Recognition by DNAzymes

Our view of the interaction between Na⁺ and nucleic acids was changed by a few recently discovered Na⁺‐specific RNA‐cleaving DNAzymes. In addition to nonspecific electrostatic interactions, highly specific recognition is also possible. Herein, two such DNAzymes, named EtNa and Ce13d, are compared to elucidate their mechanisms of Na⁺ binding. Mutation studies indicate that they have different sequence requirements. Phosphorothioate (PS) substitution at the scissile phosphate drops the activity of EtNa 140‐fold, and it cannot be rescued by thiophilic Cd²⁺ or Mn²⁺, whereas the activity of PS‐modified Ce13d can be rescued. Na⁺‐dependent activity assays indicate that two Na⁺ ions bind cooperatively in EtNa, and each Na⁺ likely interacts with a nonbridging oxygen atom in the scissile phosphate, whereas Ce13d binds only one Na⁺ ion in a well‐defined Na⁺ aptamer, and this Na⁺ ion does not directly interact with the scissile phosphate. Both DNAzymes display a normal pH–rate profile, with a single deprotonation reaction required for catalysis. For EtNa, Na⁺ fails to protect the conserved nucleotides from dimethyl sulfate attack, and no specific Na⁺ binding is detected by 2‐aminopurine fluorescence, both of which are different from those observed for Ce13d. This work suggests that EtNa binds Na⁺ mainly through its scissile phosphate without significant involvement of the nucleotides in the enzyme strand, whereas Ce13d has a well‐defined aptamer for Na⁺ binding. Therefore, DNA has at least two distinct ways to achieve highly selective Na⁺ binding.     

Synthesis of Starch-Stabilized Ag Nanoparticles and Hg2+ Recognition in Aqueous Media

The starch-stabilized Ag nanoparticles were successfully synthesized via a reduction approach and characterized with SPR UV/Vis spectroscopy, TEM, and HRTEM. By utilizing the redox reaction between Ag nanoparticles and Hg²⁺, and the resulted decrease in UV/Vis signal, we develop a colorimetric method for detection of Hg²⁺ ion. A linear relationship stands between the absorbance intensity of the Ag nanoparticles and the concentration of Hg²⁺ ion over the range from 10 ppb to 1 ppm at the absorption of 390 nm. The detection limit for Hg²⁺ ions in homogeneous aqueous solutions is estimated to be ~5 ppb. This system shows excellent selectivity for Hg²⁺ over other metal ions including Na⁺, K⁺, Ba²⁺, Mg²⁺, Ca²⁺, Fe³⁺, and Cd²⁺. The results shown herein have potential implications in the development of new colorimetric sensors for easy and selective detection and monitoring of mercuric ions in aqueous solutions.     

Binding of calcium and magnesium by modified onion skins

The use of modified onion skins for binding of Ca²⁺ and Mg²⁺ from solutions has been investigated. The effect of time of equilibration, temperature, and pH on the sorption of the metal ions have been studied. Batch and column experiments have been performed and the adsorption isotherms have been plotted. The capacities with respect to Ca²⁺ and Mg²⁺ were found to be 4 and 16 meq, respectively, per gram of the substrate when separate column experiments were conducted using 1 L of solution containing 1000 meq of the respective metal ions at pH6. With a solution containing 10 meq each of Ca²⁺ and Mg²⁺ together, however, the substrate seems to exhibit greater preference for Ca²⁺ than Mg²⁺. The sorbed metal ions from the substrate can be leached into solution with a decinormal solution of HCl and the washed bed can be reused. In view of the complex organic nature of the onion skin and its considerable capacity to bind Ca²⁺ and Mg²⁺, the possibility of its use for preventing scale formation in boilers is indicated.     

The Effect of Different Metal Cation Binding on the Proton Pumping in Bacteriorhodopsin

The first section of this paper is a detailed summary of studies made by us and others on metal cations binding to deionized bacteriorhodopsin (dIbR) and its variants. Our studies include the luminescence experiments of Eu³⁺ binding to dIbR and potentiometric studies of Ca²⁺ binding to dIbR, to deionized bR mutants, to bacterioopsin, and to dIbR with its C-terminus removed. The results suggest the presence of two classes of binding sites, one class has two high-affinity constants, and one has one low-affinity constant. For Ca²⁺ binding, there is one metal cation in each of the two high-affinity sites which are coupled to the charged aspartates 85 and 212 (known to be in the retinal cavity) but not coupled to each other. The low-affinity class can accommodate 0–6 Ca²⁺ ions and most of them are bound to the surface. Mg²⁺ has a slightly smaller value for its binding constant to the highest-affinity site. Thus, one expects more Ca²⁺ than Mg²⁺ bound to the two high-affinity sites. In the second section, we summarize our recent study on the effect of metal cation charge density (Ca²⁺, Mg²⁺, Eu³⁺, Tb³⁺, Ho³⁺, Dy³⁺) on the kinetics of both Schiff base deprotonation and proton transport to the extracellular surface. For all metal cations, the apparent rate constant of the slow components of the deprotonation process is the same as that for the transport process at 22 °C. The temperature studies, however, show this apparent equality to be fortuitous and to result from cancellation of the contribution of the energy and entropy of activation. Thus, while the entropy of activation is positive for the deprotonation process, it is negative for the proton transport process. These kinetic parameters depend weakly on the charge density, but in an opposite sense for the two processes. These results suggest that the deprotonation is not the rate-limiting step for the proton transport process. A possible mechanism is proposed in which a hydrated metal cation is used to induce the deprotonation of the protonated Schiff base and to dissociate one of its H₂O molecules to donate the proton in the L → M process.     

Transfer of Monovalent and Divalent Cations in Salt Solutions by Electrodialysis

Abstract

The selectivity mechanism of transport of Na⁺, Ca²⁺ and Mg²⁺ through commercial monovalent‐cation permselective membranes is investigated in batch electrodialysis experiments with synthetic salt solutions containing monovalent and divalent cations. The role of hydration energy, steric effect, kinetic effect as well as effects of permselectivity of cation exchange membrane has been elucidated with electrodialysis of single solutions (NaCl, CaCl₂, MgCl₂). The mechanism of interferences is investigated in (Na⁺/Ca²⁺, Na⁺/Mg²⁺, Ca²⁺/Mg²⁺ and Na⁺/Ca²⁺/Mg²⁺) mixtures.     

Lichen Substances Affect Metal Adsorption in <Emphasis Type="Italic">Hypogymnia physodes</Emphasis>

Lichen substances are known to function as chelators of cations. We tested the hypothesis that lichen substances can control the uptake of toxic metals by adsorbing metal ions at cation exchange sites on cell walls. If true, this hypothesis would help to provide a mechanistic explanation for results of a recent study showing increased production of physodalic acid by thalli of the lichen Hypogymnia physodes transplanted to sites with heavy metal pollution. We treated cellulose filters known to mimic the cation exchange abilities of lichen thalli with four lichen substances produced by H. physodes (physodic acid, physodalic acid, protocetraric acid, and atranorin). Treated filters were exposed to solutions containing seven cations (Ca²⁺, Cu²⁺, Fe²⁺, Fe³⁺, Mg²⁺, Mn²⁺, and Na⁺), and changes to the solution concentrations were measured. Physodalic acid was most effective at influencing metal adsorption, as it increased the adsorption of Fe³⁺, but reduced the adsorption of Cu²⁺, Mn²⁺, and Na⁺, and to a lesser extent, that of Ca²⁺ and Mg²⁺. Reduced Na⁺ adsorption matches with the known tolerance of this species to NaCl. The results may indicate a possible general role of lichen substances in metal homeostasis and pollution tolerance.     

Development of thin‐film composite membranes for carbon dioxide and methane separation using sulfonated poly(phenylene oxide)

Novel membranes based on sulfonated poly (phenylene oxide) (SPPO) was developed. SPPO membranes in the hydrogen form were converted to metal ion forms. The effect of exchange with metal ions including monovalent (Li⁺, Na⁺, K⁺), divalent (Mg²⁺, Ba²⁺, Ca²⁺) and trivalent (Al³⁺) ions was investigated in terms of permeation rate and permeation rate ratios for CO₂ and CH₄ gases. Both dense homogeneous membranes and thin‐film composite (TFC) membranes were studied for their gas separation characteristics. The effect of membrane preparation conditions and operating parameters on the membrane performance were also investigated. The selectivity of the TFC membrane increased as the cationic charge density increased as a result of electrostatic cross‐linking. TFC membrane of very high selectivity was achieved by coating a thin layer of SPPO‐Mg on a PES substrate. © 2000 John Wiley & Sons, Inc. J Appl Polym Sci 76: 735–742, 2000     

Binding of several heavy metal ions by polyaspartyl polymers and their application to some Chinese herbal medicines

Water‐insoluble polyaspartyl polymers were synthesized by using water as medium instead of organic medium. Taking Ca²⁺ as a reference, the binding of several heavy‐metal ions, including Pb²⁺, Cd²⁺, Hg²⁺, Cr³⁺, Cu²⁺, and Mn²⁺, by polyaspartyl polymers was studied. The experimental results revealed that polyaspartate is an excellent binding agent for the investigated heavy‐metal ions. These cation ions were bound to polyaspartate polymer by the same mechanism as Pb²⁺, which can be explained by ion exchange model. Since polyaspartate has a protein‐resembling structure that is sensitive to trace heavy metal, it was used to remove some trace heavy‐metal elements in Chinese herbal medicines. It was found that polyaspartate material was an effective agent for the removal of Pb²⁺, Cd²⁺, and Hg²⁺ ions from glycyrrhizin, angelica, and gynostemma pentaphyllum. © 2007 Wiley Periodicals, Inc. J Appl Polym Sci 2007     

Selection of RNA-Cleaving TNA Enzymes for Cellular Mg2+ Imaging

Catalytic DNA-based fluorescent sensors have enabled cellular imaging of metal ions such as Mg²⁺. However, natural DNA is prone to nuclease-mediated degradation. Here, we report the in vitro selection of threose nucleic acid enzymes (TNAzymes) with RNA endonuclease activities. One such TNAzyme, T17–22, catalyzes a site-specific RNA cleavage reaction with a k_cat of 0.017 min⁻¹ and K_M of 675 nM. A fluorescent sensor based on T17–22 responds to an increasing concentration of Mg²⁺ with a limit of detection at 0.35 mM. This TNAzyme-based sensor also allows cellular imaging of Mg²⁺. This work presents the first proof-of-concept demonstration of using a TNA catalyst in cellular metal ion imaging.     

Polythiophene based fluorescent probe for copper ions with high sensitivity

Two new conjugated polymers poly{3‐({4‐[(2‐hydrazino‐2‐oxoethyl)(methyl)amino]cyclohexylidene}methyl)thiophene}( P1 ) and poly{3‐({4‐[(3‐hydrazino‐3‐oxopropyl)(methyl)amino]cyclohexylidene}methyl)thiophene}( P2 ) were synthesized, and their optical properties were investigated. P1 exhibited excellent selectivity toward Cu²⁺ ions in 50% water solution, the fluorescence color of P1 changed distinctly from greenyellow to colorless in the presence of Cu²⁺ under UV‐light, while introduction of other metal ions could not induce such significant variation. Moreover, highly sensitive detection of Cu²⁺ ions was demonstrated in 90% water solution. Its high metal‐chelating capability allowed Cu²⁺ recognition with a detection limit of 3.2 × 10^?10 M. These results indicated that this kind of nonionic polymer containing multidentate ligand could be used as a highly selective and sensitive chemosensor for Cu²⁺ detection. The proposed binding mode of P1 with Cu²⁺ was supported by DFT calculation using Gaussian 03. Unlike P1 , P2 showed no obvious fluorescent change in the presence of various metal ions due to its space steric hindrance resulted from N/O distribution on the side chain of P2 . © 2015 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2015 , 132, 42440.     

Preparation and characterisation of a poly(acrylamidoglycolic acid-co-acrylamide) hydrogel for selective binding of Cu and application to diffusive gradients in thin films measurements

A poly(acrylamidoglycolic acid-co-acrylamide) [poly(AAGA-co-AAm)] hydrogel was prepared by copolymerising 2-acrylamidoglycolic acid (AAGA) with acrylamide (AAm). The copolymer hydrogel composition and structure was characterised by FTIR spectroscopy and elemental microanalysis and found to contain 3.5 AAGA monomer units for each AAm monomer unit. This was similar to the monomer ratios used in the synthesis. The metal ion binding properties of the hydrogel were characterised for a range of metal ions (Cu²⁺, Cd²⁺, K⁺, Na⁺, Mg²⁺ and Ca²⁺) under varying conditions of pH, ionic strength, metal concentration and time. The hydrogel was shown to bind Cu²⁺ and Cd²⁺ strongly under non-competitive binding conditions, with binding capacities of 5.3 and 5.1 μmol cm⁻², respectively. The binding capacity of each metal decreased, under competitive binding conditions (with a range of metal ions present at 17.8 μN), to 1.3 and 0.17 μmol cm⁻², respectively, indicating stronger selectivity for Cu²⁺. The metal ions were readily recovered (>94%) by eluting with 2 M nitric acid solution for 24 h. The binding capacities for Cu²⁺ and Cd²⁺ were also found to decrease with increasing ionic strength and at pH values <5. The copolymer was found to have an equilibrium swelling ratio (q_w) of over 500 at a maxima of pH 5.4 and at low ionic strengths. Finally, the copolymer hydrogel was tested as a binding phase with the diffusive gradients in thin films technique. A linear mass vs. time relationship was observed for Cu²⁺ in synthetic Windermere water with a recovery of approximately 100%.     

Synthesis,characterization, metal sorption,and biological activity of poly(N‐heterocylic acrylamide)

N‐heterocyclic acrylamide monomers were prepared and then transferred to the corresponding polymers to be used as an efficient chelating agent. Polymers reacted with metal nitrate salts (Cu²⁺, Pb²⁺_, Mg²⁺, Cd²⁺, Ni²⁺, Co²⁺_, Fe²⁺) at 150°C to give metal‐polymer complexes. The selectivity of the metal ions using prepared polymers from an aqueous mixture containing different metal ion sreflected that the polymer having thiazolyl moiety more selective than that containing imidazolyl or pyridinyl moieties. Ion selectivity of poly[N‐(benzo[d]thiazol‐2‐yl)acrylamide] showed higher selectivity to many ions e.g. Fe³⁺, Pb²⁺, Cd²⁺, Ni²⁺, and Cu²⁺. While, that of poly[N‐(pyridin‐4‐yl)acrylamide] is found to be high selective to Fe³⁺ and Cu²⁺ only. Energy dispersive spectroscopy measurements, morphology of the polymers and their metallopolymer complexes, thermal analysis and antimicrobial activity were studied. © 2015 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2015 , 132, 42712.     

Ultrafiltration recovery of alginate: Membrane fouling mitigation by multivalent metal ions and properties of recycled materials

Recovery of alginate extracted from aerobic granular sludge (AGS) has given rise to a novel research direction. However, these extracted alginate solutions have a water content of nearly 100%. Alternately, ultrafiltration (UF) is generally used for concentration of polymers. Furthermore, the introduction of multivalent metal ions into alginate may provide a promising method for the development of novel nanomaterials. In this study, membrane fouling mitigation by multivalent metal ions, both individually and in combination, and properties of recycled materials were investigated for UF recovery of sodium alginate (SA). The filtration resistance showed a significantly negative correlation with the concentration of metal ions, arranged in the order of Mg²⁺ < Ca²⁺ < Fe³⁺ < Al³⁺ (filtration resistance mitigation), and the moisture content of recycled filter cake showed a marked decrease. For Ca²⁺, Mg²⁺, Fe³⁺, and Ca²⁺+ Fe³⁺, the filtration resistances were almost the same when the total charge concentration was less than 5 mmol⋅L^–1. However, when the total charge concentration was greater than 5 mmol⋅L^–1, membrane fouling mitigation increased significantly in the presence of Ca²⁺ or Fe³⁺ and remained constant for Mg²⁺ with the increase of total charge concentration. The filtration resistance mitigation was arranged in the order of Fe³⁺ > Fe³⁺ + Ca²⁺ > Ca²⁺ > Mg²⁺. Three mechanisms were proposed in the presence of Fe³⁺, such as the decrease of SA concentration, change in pH, and production of hydroxide iron colloids from hydrolysis. The properties of recycled materials (filter cake) were investigated via optical microscope observation, dynamic light scattering, Fourier transform infrared, X-ray photoelectron spectroscopy (XPS), and scanning electron microscopy. The results provide further insight into UF recoveries of alginate extracted from AGS.     

Synthesis of a highly selective bis-rhodamine chemosensor for naked-eye detection of Cu ions and its application in bio-imaging

A bis-rhodamine based fluorescent chemosensor for naked-eye detection of Cu²⁺ with enhanced sensitivity as compared to mono-rhodamine derivative has been synthesized, and its selectivity for Cu²⁺ in the presence of other competitive metal ions (Li⁺, Na⁺, K⁺, Cs⁺, Mg²⁺, Ca²⁺, Sr²⁺, Cr³⁺, Mn²⁺, Fe²⁺, Fe³⁺, Co²⁺, Ni²⁺, Zn²⁺, Cd²⁺, Hg²⁺, and Pb²⁺), and application in bio-imaging are demonstrated.     

Selective removal of copper ions from aqueous solutions using modified silica beads impregnated with LIX 84

Silica beads immobilized with 2‐hydroxy‐5‐nonylacetophenoneoxime (LIX 84) were prepared after silica surface modification by γ‐aminopropyltriethoxysilane (SB‐L). Batch and packed‐column tests were conducted to evaluate the metal ion removal capabilities of the prepared adsorbent. Equilibrium isotherms of the SB‐L with aqueous solutions containing copper ions were obtained. In addition, the kinetic performances for copper ion removal from aqueous solutions were investigated. The results showed that the amount of extraction increases with solution pH in the range between 1.5 and 5. The selectivity was also experimentally investigated, these results showed that the SB‐L adsorbed copper ions selectively in the presence of other metal ions such as Ni²⁺, Co²⁺, Zn²⁺, Cd²⁺, Ca²⁺ and Mg²⁺. From the regeneration experiments, it was found that the copper ions adsorbed at the SB‐L surface were recovered by acidic solutions. The recovery ratios were between 78% and 90%, depending on the types of acidic solutions. The results showed that the SB‐L prepared may be used for the selective extraction of copper ions from aqueous solutions. © 1999 Society of Chemical Industry     

Synthesis of a highly selective bis-rhodamine chemosensor for naked-eye detection of Cu2+ ions and its application in bio-imaging

A bis-rhodamine based fluorescent chemosensor for naked-eye detection of Cu²⁺ with enhanced sensitivity as compared to mono-rhodamine derivative has been synthesized, and its selectivity for Cu²⁺ in the presence of other competitive metal ions (Li⁺, Na⁺, K⁺, Cs⁺, Mg²⁺, Ca²⁺, Sr²⁺, Cr³⁺, Mn²⁺, Fe²⁺, Fe³⁺, Co²⁺, Ni²⁺, Zn²⁺, Cd²⁺, Hg²⁺, and Pb²⁺), and application in bio-imaging are demonstrated.