Divalent Ions Research Articles

Small Molecule Inhibitors of Mycobacterium tuberculosis Topoisomerase I Identified by Machine Learning and In Vitro Assays

Tuberculosis (TB) caused by Mycobacterium tuberculosis is a leading infectious cause of death globally. The treatment of patients becomes much more difficult for the increasingly common multi-drug resistant TB. Topoisomerase I is essential for the viability of M. tuberculosis and has been validated as a new target for the discovery of novel treatment against TB resistant to the currently available drugs. Virtual high-throughput screening based on machine learning was used in this study to identify small molecules that target the binding site of divalent ion near the catalytic tyrosine of M. tuberculosis topoisomerase I. From the virtual screening of more than 2 million commercially available compounds, 96 compounds were selected for testing in topoisomerase I relaxation activity assay. The top hit that has IC50 of 7 µM was further investigated. Commercially available analogs of the top hit were purchased and tested with the in vitro enzyme assay to gain further insights into the molecular scaffold required for topoisomerase inhibition. Results from this project demonstrated that novel small molecule inhibitors of bacterial topoisomerase I can be identified starting with the machine-learning-based virtual screening approach.

Complexes of Iron(II), Cobalt(II), and Nickel(II) with DOTA-Tetraglycinate for pH and Temperature Imaging Using Hyperfine Shifts of an Amide Moiety.

Paramagnetic complexes of 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetate (DOTA4-) derivatives have shown potential for molecular imaging with magnetic resonance. DOTA-tetraglycinate (DOTA-4AmC4-) coordinated with lanthanide metal ions (Ln3+) demonstrates pH/temperature sensing with Biosensor Imaging of Redundant Deviation in Shifts (BIRDS) and Chemical Exchange Saturation Transfer (CEST), respectively, detecting nonexchangeable (e.g., -CHy, where 3 ≥ y ≥ 1) and exchangeable (e.g., -OH or -NHx, where 2 ≥ x ≥ 1) protons. Herein, we report paramagnetic complexes of divalent transition-metal ions (M2+ = Fe2+, Co2+, Ni2+) with DOTA-4AmC4- that endow a unique amide proton (-NH) moiety for pH/temperature sensing. Crystallographic data reveal that DOTA-4AmC4- coordinates with M2+ through oxygen and nitrogen donor atoms, ranging in coordination numbers from 8-coordinate in Fe(II)DOTA-4AmC2-, 7-coordinate in Co(II)DOTA-4AmC2-, and 6-coordinate in Ni(II)DOTA-4AmC2-. The -CHy protons in M(II)DOTA-4AmC2- displayed modest pH/temperature sensitivities, but -NH protons exhibited higher intensity, suggesting prominent BIRDS properties. The pH sensitivity was the highest for Ni(II)DOTA-4AmC2- (1.42 ppm/pH), followed by Co(II)DOTA-4AmC2- (0.21 ppm/pH) and Fe(II)DOTA-4AmC2- (0.16 ppm/pH), whereas temperature sensitivities were comparable (i.e., 0.22, 0.13, and 0.17 ppm/°C, respectively). The CEST image contrast for -NH in M(II)DOTA-4AmC2- was much weaker compared to that of Ln(III)DOTA-4AmC-. Given its high pH sensitivity and low cytotoxicity, Ni(II)DOTA-4AmC2- shows promise for use in preclinical BIRDS-based pH imaging.

A Hexanuclear Gallium(III) Complex with a {GaIII6(μ‐OH)10} Core Supported by Two Naphthalenediolate‐Based Ligands

AbstractWe have developed a family of 2,7‐disubstituted 1,8‐naphthalenediol ligands to pre‐organize two metal ions to the distance of two neighboring phosphate diesters of 6–7 Å in the DNA backbone by a rigid backbone. As complexes with divalent ions (CuII and NiII) bind to DNA, interferes DNA synthesis via PCR, and kills human cancer cells more efficiently than human stem cells of the same proliferation rate, we intended to increase the binding affinity by going from divalent to trivalent ions‐especially FeIII. Here, we apply GaIII as redox inactive model for FeIII and obtain for the first time a complex with trivalent ions. However, the structural characterization provides not a dinuclear GaIII complex, but a hexanuclear GaIII complex with a central {GaIII6(μ‐OH)10} core, that is yet unprecedented.

Monovalent Ion Effect on Liquid-Liquid Phase Separation of Aqueous Polyphosphate-Salt Mixtures.

Polyphosphate (polyP) is one of the most conserved biomacromolecules and can form aggregates, such as polyP granules in bacteria, which are generated through liquid-liquid phase separation (LLPS). Studies have examined the mechanism of polyP aggregation using LLPS systems containing artificial polyP molecules as aggregation system models, where LLPS is typically induced by multivalent salts and polyelectrolytes. Although the typical concentrations of monovalent ions in living cells are approximately 100 times higher than those of divalent ions, the effects of monovalent ions on the LLPS of polyP solutions are little known. This study demonstrated that submolar NaCl induces LLPS of polyP solutions, whereas other monovalent salts did not induce LLPS at the same concentrations. Small-angle X-ray scattering measurements revealed that NaCl significantly stabilizes the intermolecular association of polyP, inducing LLPS. These findings suggest that the modulation of monovalent ion concentrations is an underlying mechanism of polyP aggregate formation and deformation within living cells.

Thiol and amino co-grafting modification of sugarcane bagasse lignin nanospheres to enhance the adsorption capacity of cadmium ion

Divalent cadmium ions in the water environment are a serious threat to human health. The purpose of this study is to develop a biodegradable bio-adsorbent based on bagasse lignin and can effectively adsorb cadmium ions. In this study, it was found that the thiol amino co-grafted lignin nanospheres (TALNPs) had been successfully prepared by the Mannich reaction system, and the performance tests indicate that TALNPs have good adsorption potential. The evaluation experiment shows that factors such as pH value, dosage, time, initial concentration, and temperature significantly affect the ability of TALNPs to adsorb cadmium ions. The simulation calculation of isothermal adsorption experiment shows that the adsorption of cadmium ions follows the Langmuir model, and the adsorption kinetics is pseudo-second order. Compared with unmodified lignin, the ability of TALNPs has increased by about 3.5 times. Using XPS to analyze the structure of TALNPs in the adsorption and desorption process shows that only the bridge bond changes, indicating that it is a relatively stable adsorbent and can be reused. In general, the design strategy of thiol-amino co-grafting to modify lignin nanospheres can enhance the adsorption of cadmium ions, and also provide a new way for the development and utilization of bagasse lignin.

A DFT Study of Band-Gap Tuning in 2D Black Phosphorus via Li+, Na+, Mg2+, and Ca2+ Ions.

Black phosphorus (BP) and its two-dimensional derivative (2D-BP) have garnered significant attention as promising anode materials for electrochemical energy storage devices, including next-generation fast-charging batteries. However, the interactions between BP and light metal ions, as well as how these interactions influence BP's electronic properties, remain poorly understood. Here, we employed density functional theory (DFT) to investigate the effects of monovalent (Li+ and Na+) and divalent (Mg2+ and Ca2+) ions on the valence electronic structure of 2D-BP. Molecular orbital analysis revealed that the adsorption of divalent cations can significantly reduce the band gap, suggesting an enhancement in charge transfer rates. In contrast, the adsorption of monovalent cations had minimal impact on the band gap, suggesting the preservation of 2D-BP's intrinsic electrical properties. Energetic and charge analyses indicated that the extent of charge transfer primarily governs the ability of ions to modulate 2D-BP's electronic structure, especially under high-pressure conditions where ions are in close proximity to the 2D-BP surface. Moreover, charge polarization calculations revealed that, compared with monovalent cations, divalent cations induced greater polarization, disrupting the symmetry of the pristine 2D-BP and further influencing its electronic characteristics. These findings provide a molecular-level understanding of how ion interactions influence 2D-BP's electronic properties during ion-intercalation processes, where ions are in close proximity to the 2D-BP surface. Moreover, the calculated diffusion barrier results revealed the potential of 2D-BP as an effective anode material for lithium-ion, sodium-ion, and magnesium-ion batteries, though its performance may be limited for calcium-ion batteries. By extending our understanding of interactions between ions and 2D-BP, this work contributes to the design of efficient and reliable energy storage technologies, particularly for the next-generation fast-charging batteries.

Guanidinium‐Based Covalent Organic Framework Membranes for Superior Mono‐ and Divalent Cations Separation

Biological membranes exhibit extraordinary efficiency in processing ion permeability and selectivity. However, creating artificial membranes for an ideal ion separation is still challenging due to the subtle distinctions in valence and size among different ions. In the realm of biological recognition, the ultimate selectivity of ion channels is considered to stem from the specific binding interactions and appropriate charge density. Designing artificial membranes to achieve similar performance not only helps the understanding of complex ion transport in bioprocesses but also facilitates critical industrial separations. Inspiring by the remarkable performance in biological systems, a guanidinium‐based covalent organic framework membrane is designed, which exhibits an excellent capability to recognize mono‐/divalent cations, achieving K+/Mg2+ selectivity up to 202 in a mixed salt solution. Furthermore, the membrane displays rapid ion transport owing to the uniform sub‐2 nm channels. The experimental results and molecular dynamics simulations illustrate that the charge‐assisted hydrogen bonding sites and the Cl− counter ions within 1D channels play critical roles in cations sieving. Specifically, divalent ions passing through the positively charged channels need to overcome higher energy barriers than monovalent ions. These findings offer promising avenues for the development of advanced multifunctional membranes for efficient ion separation and sustainable water‐related separations.

Effects of mono- and divalent- ions and their strength on foaming properties of infant formula protein model system

Metal ions may affect the foaming properties of infant formula products that including milk protein during reconstitution. In this study, an infant formula protein model system (IFPMS) composed of both whey protein and casein was constructed, and the effect of mono (Na+, K+) and divalent (Mg2+, Ca2+) ions and their strength (0–100 mM) on foaming properties was investigated. The mono ions had no significantly impact on foaming capacity (FC), MgCl2 slightly increase FC and reaching a maximum of 116.09 ± 7.46% at 25 mM, whereas CaCl2 significantly decreased FC from 105.78 ± 7.42% to 83.14 ± 6.75% as 0-25 mM (p < 0.05). Foam stability (FS) was significantly improved as all salts above 10 mM (p < 0.05). The dramatically decreased net charge and larger protein aggregates (> 25 mM) of IFPMS with divalent ions indicating their stronger charge-shielding effect, which caused slow adsorption of protein and increased surface tension. Multiple spectral results confirmed that salts enhanced protein interactions via inter-molecular and form a viscoelastic layer to stabilize foam. Absorbed protein and SDS-PAGE analysis revealed that MgCl2 promoted protein adsorption, while CaCl2 reduced casein in the foam phase. Correlation analysis further implied that the FC showed highly significant positive with absorbed protein, solubility, and β-turn (p ≤ 0.01), and FS was significantly correlated with secondary structure and viscosity (p ≤ 0.05). This study may provide useful information for an in-depth understanding of the foaming properties of infant formula.

The role of multivalent cations in determining the cross-linking affinity of alginate hydrogels: A combined experimental and modeling study

This study explores the development of a comprehensive mathematical model to predict the cross-link density of alginate hydrogels using varying concentrations of divalent and trivalent ions. By systematically investigating the effects of Ca²⁺, Ba²⁺, Cu²⁺, Sr²⁺, Mg²⁺, Fe3⁺ and Al³⁺, the research describes the relationship between ion concentration and cross-link density. Experimental data reveals that the type and concentration of these ions critically influence the mechanical properties of the resulting hydrogels, with trivalent ions such as Fe³⁺ forming stronger, triple cross-links that significantly enhance the hydrogel's mechanical strength. Among the divalent ions, the trend in binding affinity is as follows: Ba²⁺ with the highest affinity followed by Sr²⁺, Ca²⁺ and Cu²⁺, while Mg²⁺ stands out with the lowest affinity, significantly differing from the others. The deviation of Cu²⁺ (transition metal ion) from the expected trend in ion interactions suggests that coordination chemistry, along with ionic radius, valence, and cation coordination abilities, plays a significant role in determining interaction strength with alginate. The proposed model, enhanced with fitting parameters k1 and k2 to account for ion-specific effects, leverages the unique binding affinities and coordination chemistry of each ion to tailor alginate hydrogels for specific applications. The parameter k1 reflects the affinity of the ions for the alginate chains, while k2 captures the coordination abilities and cross-linking efficiency. This work not only advances the understanding of ion-mediated cross-linking in alginate systems but also offers a valuable tool for the design and optimization of hydrogels with precise mechanical properties governed by various applications.

Role of divalent doping at A-sites (A=Ca2+, Sr2+, Ba2+ & Pb2+) on thermoelectric and magnetoresistive properties in La0.67(Bi0.0835Na0.0835) A0.165MnO3

The present work reports on the effect of divalent ion doping (Ca2+, Sr2+, Pb2+ & Ba2+) ions at A-site in the compound with compositional formula La0.67(Bi0.0835Na0.0835)A0.165MnO3 on structural, magnetic, magneto-transport and thermoelectric properties. X-ray diffraction reveals orthorhombic structure for calcium and barium doped with no peak splitting observed at ~ 32o whereas rhombohedral structure for strontium and lead dopants with a peak splitting in the main intensity. Doping with divalent ions at A-site in La0.67(Bi0.0835Na0.0835)A0.165MnO3 manganites has introduced Mn2+ ions, changing the ratio of Mn3+/Mn4+ in octahedral site and this has been confirmed from X-ray photospectroscopy studies. The field emission scanning electron microscope images confirm different grain sizes and morphologies for different doping elements and distinguished variation is noticed for strontium doped manganites with a grain size of 1.08µm. Room temperature ferromagnetic behavior was observed in the samples at room temperature except for calcium doped sample and magnetic behavior has been discussed on the basis of the various interactions between mixed valence manganese ions and Mn3+/Mn4+ ratio. Electrical resistivity (ρ) as a function of temperature (T) without magnetic field indicates the double peak behavior and with the application of external magnetic fields, the double peak disappears, in all the investigated samples. Highest MR% at their respective transition temperatures (TP1) has been obtained for lead doped samples when compared to barium, strontium and calcium samples. Activation energies (EP) estimated decreases initially for calcium and strontium whereas it increases for lead and barium dopants i.e. for further increases in ionic radii. The change of sign from positive to negative in Seebeck coefficient (S) was noticed with increasing ionic radii suggesting the variation in the nature of charge carriers. The electron-electron, electron-magnon, magnon-magnon scattering mechanism contributes to the temperature dependent resistivity and thermopower in the low temperature region while in the paramagnetic insulating region is governed by a small polaron hopping mechanism.

Functionalization of Magnetic Beads with Chelating Surfactants for Metal Ions Extraction

Divalent amino acid-based surfactants were utilized in the preparation and functionalization of polystyrene/superparamagnetic iron oxide nanoparticle (SPION) beads. These surfactants were derived from aminomalonic acid, aspartic acid, and glutamic acid, all containing two carboxylic groups and differing only in the spacer size between these groups, coupled with a fatty acid moiety, resulting in N-acyl amino acid surfactants. In the process of beads formation, the surfactant played a dual role as a stabilizer and a chelating agent towards the targeted metal. The beads were obtained using an emulsion solvent evaporation method. In this process, an oil-in-water emulsion was formed, containing polystyrene and iron nanoparticles. By concentrating the oil and evaporating it, solid nanoparticles were obtained. The morphology of these particles was characterized using atomic force microscopy (AFM), dynamic light scattering (DLS), while streaming potential measurements were used to determine their charge density. The surfactant's capability to extract divalent ions, specifically Zn2+ and Ni2+, was assessed both in its pure form and when they were attached to the particles. The aspartate-based surfactant showed the best chelating performance for divalent ions, and the combination of surfactant-functionalized particles with magnetic responsiveness led to highly efficient extraction (up to 90%) results. Additionally, the recyclability of the beads was also assessed, highlighting their practical utility.

Structural Studies on Mycobacterial NudC Reveal a Class of Zinc Independent NADH Pyrophosphatase

Non-tuberculous mycobacteria (NTM) have emerged as an increasing threat to public health, due to the extreme antibiotic resistance. NADH pyrophosphatase (NudC) was proposed involving in mycobacterial resistance to the first line anti-tubercular drug isoniazid (INH) or its analog ethionamide (ETH), by hydrolyzing their NAD modified active forms (NAD-INH and NAD-ETH). In this study, we performed enzymatic and structural studies on NudC from M. abscessus (NudCMab), which is highly resistant to isoniazid and emerging as the most worrisome NTM. We determined the crystal structures of NudCMab in apo form, substrate NAD-bound form and product AMP-bound form. We observed the mode for the Nudix motif of NudCMab capturing the pyrophosphate group of NAD mediated by three divalent cation ions, which provides details for understanding the mechanism on NudC hydrolyzing NAD(H) or NAD-capped substrate. Interestingly, our structures revealed a novel subclass NudC from mycobacteria characterized by a unique arginine residue on the conserved QPWPFPxS motif, as well as a unique tower domain that replaces a well-defined zinc-binding motif in E.coli NudC and catalytic domain of mammalian Nudt12. Thus, our structural studies on NudCMab not only present a class of zinc independent NADH pyrophosphatase in mycobacteria, but also may facilitate the design of NudC inhibitors for the treatment of mycobacteria infections in combination with INH or ETH.

Influence of high-molecular-weight glutenin subunits and salt types on dough rheology and gluten aggregation: A combined experimental and computational approach

This study investigates the impact of various salts (NaCl, KCl, CaCl₂, MgCl₂) on gluten aggregation, with an emphasis on the role of high-molecular-weight glutenin subunits (HMW-GS) in modulating these impacts. The results demonstrated that NaCl significantly improved dough springiness and adhesiveness, while divalent salts like CaCl₂ and MgCl₂ had a greater effect on increasing dough hardness, with KCl showing the least impact among all salts. Differential responses were observed in HMW-GS deletion lines, where Bx7 was more responsive to NaCl and CaCl₂, whereas By8 showed stronger interactions with KCl and CaCl₂. The observed reductions in zeta potential and decrements in ionic bond content during salt-induced gluten aggregation supported the 'electrostatic shielding' mechanism, with hydrophobic interactions and hydrogen bonding emerging as key contributors to aggregate stability. Molecular dynamics simulations further corroborated these findings, revealing stronger Coulomb-SR interactions and enhanced binding affinities for divalent ions, in agreement with experimental data. These insights provide a theoretical foundation for optimizing low-sodium formulations in wheat-based products.

Microstructural Evaluation and Linkage to the Engineering Properties of Metal-Ion-Contaminated Clay.

The rapid progress of urbanization and industrialization has led to the accumulation of large amounts of metal ions in the environment. These metal ions are adsorbed onto the negatively charged surfaces of clay particles, altering the total surface charge, double-layer thickness, and chemical bonds between the particles, which in turn affects the interactions between them. This causes changes in the microstructure, such as particle rearrangement and pore morphology adjustments, ultimately altering the mechanical behavior of the soil and reducing its stability. This study explores the effects of four common metal ions, including monovalent alkali metal ions (Na+, K+) and divalent heavy metal ions (Pb2+, Zn2+), with a focus on how ion valence and concentration impact the soil's microstructure and mechanical properties. Microstructural tests show that metal ion incorporation reduces particle size, increases clay content, and transforms the structure from layered to honeycomb-like. Small pores decrease while large pores dominate, reducing the specific surface area and pore volume, while the average pore size increases. Although cation exchange capacity decreases, cation adsorption density per unit surface area increases. Monovalent ions primarily disperse the soil structure, while divalent ions induce coagulation. Macro-mechanical tests reveal that metal ion contamination reduces porosity under loading, with compressibility rises as the ion concentration increases. Soils contaminated with alkali metal ions shows higher compression coefficients at all loads, while heavy metal ions cause higher compression under lower loads. Shear strength, the internal friction angle, and cohesion in metal-ion-contaminated clay decrease compared to uncontaminated field-state clay, with greater declines at higher ion concentrations. The Micropore Morphology Index and hydro-pore structural parameter effectively characterize both micro- and macrostructural properties, establishing a quantitative relationship between HPSP and the engineering properties of metal-ion-contaminated clay.

Alternative substrate kinetics of SARS-CoV-2 Nsp15 endonuclease reveals a specificity landscape dominated by RNA structure.

Coronavirus endoribonuclease Nsp15 contributes to the evasion of host innate immunity by suppressing levels of viral dsRNA. Nsp15 cleaves both ssRNA and dsRNA in vitro with a strong preference for unpaired or bulged U residues, and its activity is stimulated by divalent ions. Here, we systematically quantified effects of RNA sequence and structure context that define its specificity. The results show that sequence preference for U↓A/G, observed previously, contributes only ca. 2-fold to kcat/Km. In contrast, dsRNA structure flanking a bulged U residue increases kcat/Km by an order of magnitude compared to ssRNA while base pairing in dsRNA essentially blocks cleavage. Despite enormous differences in multiple turnover kinetics, the effect of RNA structure on the cleavage step is minimal. Surprisingly, although divalent ion activation of Nsp15 is widely considered to be important for its biological function, the effect on kcat/Km is only ∼2-fold and independent of RNA structure. These results reveal a specificity landscape dominated by RNA structure and provide a quantitative framework for identifying interactions that underlie specificity, determining mechanisms of inhibition and resistance and defining targets important for coronavirus biology.

Size-Dependent Nascent Sea Spray Aerosol Bounce Fractions and Estimated Viscosity: The Role of Divalent Cation Enrichment, Surface Tension, and the Kelvin Effect.

Viscosity, or the "thickness," of aerosols plays a key role in atmospheric processes like ice formation, water absorption, and heterogeneous kinetics. However, the viscosity of sea spray aerosols (SSA) has not been widely studied. This research explored the relationship between particle size and viscosity of authentic SSA particles through particle bounce, atomic force microscopy analysis, and predictive viscosity modeling from molecular composition. The study found that 40 nm SSA particles had estimated viscosities around 104 Pa·s and bounce fractions three times higher than 100 and 200 nm particles with less than 102 Pa·s at a relative humidity (RH) of 60%. Additional studies revealed the Kelvin effect and particle density, influenced by particle size, have a greater impact on size-dependent bounce fractions than changes in RH across impactor stages. While changes in the level of surfactants can impact particle bounce, the increased viscosity in smaller SSA is attributed to the formation of gel-like phase states caused by cation-organic cross-links between divalent calcium ions and organic anions enriched in the smaller particles. This work shows the smallest gel-like SSA particles observed in the field are highly viscous, which has implications for cloud formation, secondary aerosol growth, and pollutant transport in coastal environments.

Human epidermal growth factor receptor 2(Her2)-targeted pH-responsive MR/NIRF bimodal imaging-mediated nano-delivery system for the diagnosis and treatment of undifferentiated thyroid cancer.

Undifferentiated thyroid cancer (ATC) is highly malignant and does not respond well to sorafenib (SRF) treatment owing to the lack of specificity of SRF targeting. Drug delivery nanosystems can improve the efficiencies of drug in treating various cancer types. However, many conventional drug delivery nanosystems lack targeting and exhibit unresponsive drug release. Therefore, we developed a pH-responsive nano-targeted drug delivery systems using human serum albumin (HSA) as a carrier to generate manganese dioxide (MnO2)@HSA nanoparticles (NPs), then encapsulated SRF and the fluorescent dye indocyanine green (ICG) and finally modifyed the targeting antibody pertuzumab in the outer layer of the nano complexes, resulting in SRF/ICG/MnO2@HSA-pertuzumab (HISMP) NPs. This system targets human epidermal growth factor receptor 2 on the cell membrane surface of thyroid cancer cells and is designed to accumulate at tumor sites. Then, pH-responsive release of divalent manganese ions, ICG, and SRF enables magnetic resonance/fluorescence (MR/NIRF) dual-modality imaging and precise drug delivery for diagnostic and therapeutic integration. Various characterization analyses including transmission electron microscopy, Fourier infrared spectroscopy, and particle size analysis confirm that we successfully synthesized HISMP NPs with a diameter of 150.709nm. The results of CCK8 cytotoxicity and apoptosis assays show that HISMP NPs exhibited high cytotoxicity and induce apoptosis in thyroid cancer cells. In vivo MR/NIRF imaging experiments confirmed that the HISMP NPs specifically aggregated at tumor sites and have good in vivo MR/NIRF imaging ability and effective anti-tumor activity. The nano-delivery system is expected to provide a theoretical foundation for the efficient ATC diagnosis and therapy.

Probing the role of ligation and exonuclease digestion towards non-specific amplification in bioanalytical RCA assays.

Non-specific amplification (NSA, amplification in the absence of a target analyte) in bioanalytical rolling circle amplification (RCA) assays, especially those involving pre-synthesized circular DNA (cDNA), affects its analytical sensitivity. Despite extensive development of RCA-based bioanalytical methods, the NSA in RCA remains uncharacterized in terms of its magnitude or origin. NSA may originate from inefficient ligation or succeeding cDNA purification steps. This study comprehensively quantifies NSA across several ligation and digestion techniques for the first time since the innovation of RCA. To quantify the NSA in RCA, cDNAs were prepared using self-annealing, splint-padlock, or cohesive end ligations. The cDNAs were then subjected to nine different exonuclease digestion steps and quantified for NSA under linear as well as hyperbranched RCA conditions. We investigated buffer compositions, divalent ion concentrations, single or dual enzyme digestion, cohesive end lengths, and splint lengths. The optimized conditions successfully mitigated absolute NSA by 30-100-fold and relative NSA (normalized against primer-assisted RCA) to ∼5%. Besides understanding the mechanistic origin of NSA, novel aspects of enzyme-substrate selectivity, buffer composition, and the role of divalent ions were discovered. With increasing bioanalytical RCA applications, this study will help standardize NSA-free assays.

Charge‐Sign‐Independent Separation of Mono‐ and Divalent Ions With Nanofiltration Membranes

AbstractAchieving precise selective separation of monovalent and divalent cations, as well as anions, is vital yet challenging in practical applications involving complex component treatments. Current membranes are typically effective for separating either cation pairs or anion pairs. To address this issue, a straightforward strategy for fabricating a nanofiltration (NF) membrane is developed that selectively permeates monovalent ions. This study focused on neutralizing the surface charge and tuning the pore size distribution of the polyamide membranes through a secondary interfacial polymerization using a zwitterionic copolymer consisting of 2‐methacryloyloxyethyl phosphorylcholine and 2‐aminoethyl methacrylate hydrochloride. The optimized NF membrane prepared in this study, with a near‐neutrally charged membrane surface and appropriate pore size distribution, demonstrates favorable performance in precisely separating monovalent and divalent ions, irrespective of the ion charge sign. The optimum NF membrane features high selectivity for both Cl−/SO42− (93) and Li+/Mg2+ (67) ion pairs, along with high water permeance of 8.5 L m−2 h−1 bar−1, making it competitive with many reported membranes. This study offers new insights into the ion‐selective mechanisms of polyamide membranes for monovalent/divalent ions and may guide the development of advanced membranes with single‐solute selectivity.

Spectroscopic Properties of TmF3-Doped CaF2 Crystals.

In this study, we report the growth and comprehensive spectroscopic analysis of TmF3-doped CaF2 crystals, grown using the vertical Bridgman method. The optical absorption and photoluminescence properties of both trivalent (Tm3+) and divalent (Tm2+) thulium ions were investigated. Optical absorption spectra in the UV-VIS-NIR range reveal characteristic transitions of Tm3+ ions, as well as weaker absorption bands corresponding to Tm2+ ions. The Judd-Ofelt (JO) formalism was applied to determine the intensity parameters Ω2, Ω4, and Ω6, which were used to calculate radiative transition probabilities, branching ratios, and radiative lifetimes for the Tm3+ ions. The emission spectra showed concentration-dependent quenching effects, with significant emissions observed for the concentration of 0.1 mol% TmF3 under excitation at 260 nm and 353 nm for Tm3+ ions and at 305 nm for Tm2+ ions. A new UV emission associated with divalent Thulium is reported. The results indicate that higher TmF3 concentrations lead to increased non-radiative energy transfer, which reduces luminescence efficiency. These findings contribute to the understanding of the optical behavior of Tm-doped fluoride crystals, with implications for their application in laser technologies and radiation dosimetry.