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Coordination compounds of lanthanides are indispensable in biomedical applications as MRI contrast agents and radiotherapeutics. However, since the introduction of the chelator DOTA four decades ago, there has been only limited progress on improving their thermodynamic stability and kinetic inertness, which are essential for safe in vivo use. Here, we present ClickZip, an innovative synthetic strategy employing a coordination-templated formation of a 1,5-triazole bridge that improves kinetic inertness up to a million-fold relative to DOTA, expanding utility of lanthanide chelates beyond traditional uses. Acting as unique mass tags, the ClickZip chelates can be released from (biological) samples by acidic hydrolysis, chromatographically distinguished from interfering lanthanide species, and sensitively detected by mass spectrometry. Lanthanides enclosed in ClickZip chelates are chemically almost indistinguishable, providing a more versatile alternative to chemically identical isotopic labels for multiplexed analysis. The bioanalytical potential is demonstrated on tagged cell-penetrating peptides in vitro, and anti-obesity prolactin-releasing peptides in vivo., Competing Interests: Competing interests M.P., T.D., M.S. and A.J. are co-inventors on a patent application no. PCT/CZ2022/050087 filed in the name of applicant Ustav Organicke Chemie a Biochemie AV CR V.V.I. The application covers some of the compounds and their use discussed in this work. M.P., T.D., M.S. and A.J. declare no other competing interests. The other authors declare no competing interests., (© 2024. The Author(s).)

Traditional macromolecules or nanoscale Mn 2+ chelate-based magnetic resonance imaging (MRI) contrast agents (CAs) suffer from complicated and laborious synthesis processes, relatively low kinetic stability and T 1 relaxivity, limiting their clinical applications. Herein, we fabricated a series of kinetically inert Mn 2+ chelate-backboned polymers, P(MnL-PEG), through a facile and one-pot polymerization process. Particularly, P(MnL-PEG)-3 demonstrates a significantly higher T 1 relaxivity of 23.9 Mn mM -1 s -1 at 1.5 T than that of previously reported small molecules and macromolecules or nanoscale Mn 2+ chelate-based CAs. Due to its high T 1 relaxivity, extended blood circulation, hepatocyte-specific uptake, and kidneys metabolism, P(MnL-PEG)-3 presents significantly enhanced contrast in blood vessel, liver, and kidneys imaging compared to clinical Gd 3+ -based CAs (Gd-EOB-DTPA and Gd-DOTA) at a dosage of 0.05 mmol Mn/Gd kg -1 BW, and can accurately diagnose orthotopic H22 liver tumors in vivo in animal models. We anticipate that this work will promote the development of clinically relevant MRI CAs.

With increasing clinical applications and interest in targeted alpha therapy, there is growing interest in developing alternative chelating agents for [ 212 Pb]Pb 2+ and [ 212/213 Bi]Bi 3+ that exhibit rapid radiolabeling kinetics and kinetic inertness. Herein we report the synthesis and detailed investigation of diacetate and dipicolinate 18- and 21-membered macrocyclic chelators BADA-18, BADA-21, BADPA-18, and BADPA-21 for the complexation of Pb 2+ and Bi 3+ ions with potential use in the preparation of radiopharmaceuticals. The formation of mononuclear complexes was established by using ESI-mass spectrometry, and their stability constants were determined by potentiometric titration. A thorough study of the structure of the metal complexes was carried out by using X-ray diffraction and NMR spectroscopy. It was shown how the stability of the complex is influenced by an increase in the size of the macrocycle, the replacement of acetate arms with picolinate ones, the rigidity of the ligand, as well as the type of conformation ( syn - or anti -) of the metal complex. The new ligands were radiolabeled with [ 210 Pb]Pb 2+ and [ 207 Bi]Bi 3+ , and the in vitro stability of the resulting complexes in a competitive environment of serum and biologically significant metal ions was assessed. Rapid complex formation in 1-2 min at room temperature, as well as the high kinetic inertness of the complexes Pb(BADPA-18) and Bi(BADPA-18) in biological media, demonstrate its potential for use in targeted radionuclide therapy.

The effects of replacing nitrogen with sulfur atoms in the 18-membered macrocycle of the H 2 macropa chelator on the binding affinity and stability of "intermediate" (radio)metal [ 203 Pb]Pb 2+ and [ 213 Bi]Bi 3+ complexes are investigated. The 1,4,10,13-tetraoxo-7,16-diazacyclooctadecane backbone was replaced with derivatives containing sulfur in the 1,4- or the 1,4,10,13-positions to yield the novel chelators H 2 S 2 macropa (N 4 O 4 S 2 ) and H 2 S 4 macropa (N 4 O 2 S 4 ), respectively. Trends on the nat/203 Pb- and nat/213 Bi-complex stability constants, coordination chemistry, radiolabeling, and kinetic inertness were assessed via potentiometric titrations, UV-vis spectroscopy, NMR spectroscopy, X-ray crystallography and density functional theory (DFT) calculations. 1 H- 207 Pb NMR spectroscopy confirmed the involvement of backbone S and/or O donors in the metal coordination sphere. Overall, the trend demonstrated that increasing the softness of the donor atoms within the ligand backbone decreased the thermodynamic stability and kinetic inertness of both the Pb 2+ and Bi 3+ complexes. Conversely, DFT calculations with mock compounds dimethyl ether (DME) and dimethyl sulfide (DMS) demonstrated enhanced affinity of the S atom to both Pb 2+ and Bi 3+ with DMS compared to DME evinced by large Δ G ° values for both Pb 2+ and Bi 3+ complexes. The decreased stability of Pb/Bi-S x macropa ( x = 0, 2, 4) upon increased sulfur atom incorporation may be a result of the increased steric strain within the macrocyclic backbone upon sulfur atom introduction. Nonetheless, [ 203 Pb]Pb 2+ and [ 213 Bi]Bi 3+ labeling (pH = 7, 30 min reaction time; 10 -4 -10 -8 M chelator) resulted in both S 2 macropa 2- and macropa 2- attaining similarly high radiolabeling efficiency. Meanwhile, S 4 macropa 2- only possessed the ability to complex [ 213 Bi]Bi 3+ . Both [ 203 Pb][Pb(macropa)] and [ 203 Pb][Pb(S 2 macropa)] remained greater than 97% intact when challenged against human serum over 72 h. The results of this study reveal the effects of incorporating sulfur donor atoms into macrocyclic chelators for [ 203 Pb]Pb 2+ and [ 213 Bi]Bi 3+ radiopharmaceuticals.

Alzheimer's disease (AD) is one of the most common neurodegenerative diseases characterized by cognitive and memory impairment. Metal ion imbalance and Mitochondrial dysfunction, leading to abnormal aggregation of β-amyloid protein (Aβ), are key factors in the pathogenesis of AD. Therefore, we designed a composite nanometer system of red blood cell (RBC) membranes-encapsulated Prussian blue nanoparticles (PB/RBC). Prussian blue nanoparticles (PBNPs) can chelate Cu 2+ and reduce reactive oxygen species (ROS). The RBC membranes are a kind of natural long-lasting circulating carrier. At the same time, through NIR irradiation, the excellent photothermal ability of PBNPs can also temporarily open the blood-brain barrier (BBB), enhance the transmission efficiency of PB/RBC across the BBB, and depolymerize the formed Aβ deposits, thereby achieving the optimal therapeutic effect. In vitro and in vivo studies demonstrated that PB/RBC could inhibit Cu 2+ -induced Aβ monomers aggregation, eliminate the deposition of Aβ plaques, improve the quality of mitochondria, restore the phagocytic function of microglia, alleviate neuroinflammation in APP/PS1 mice, and repair memory damage. In conclusion, our biofilm-camouflaged nano-delivery system provides significant neuroprotection by inhibiting Cu 2+ -induced Aβ monomers aggregation, photothermally depolymerizing Aβ fibrils and reducing the level of ROS, thus effectively ameliorating and treating AD., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Pulmonary fibrosis, a disabling lung disease, results from the fibrotic transformation of lung tissue. This fibrotic transformation leads to a deterioration of lung capacity, resulting in significant respiratory distress and a reduction in overall quality of life. Currently, the frontline treatment of pulmonary fibrosis remains limited, focusing primarily on symptom relief and slowing disease progression. Bacterial infections with Pseudomonas aeruginosa are contributing to a severe progression of idiopathic pulmonary fibrosis. Phytic acid, a natural chelator of zinc, which is essential for the activation of metalloproteinase enzymes involved in pulmonary fibrosis, shows potential inhibition of LasB, a virulence factor in P. aeruginosa, and mammalian metalloproteases (MMPs). In addition, phytic acid has anti-inflammatory properties believed to result from its ability to capture free radicals, inhibit certain inflammatory enzymes and proteins, and reduce the production of inflammatory cytokines, key signaling molecules that promote inflammation. To achieve higher local concentrations in the deep lung, phytic acid was spray dried into an inhalable powder. Challenges due to its hygroscopic and low melting (25 °C) nature were mitigated by converting it to sodium phytate to improve crystallinity and powder characteristics. The addition of leucine improved aerodynamic properties and reduced agglomeration, while mannitol served as carrier matrix. Size variation was achieved by modifying process parameters and were evaluated by tools such as the Next Generation Impactor (NGI), light diffraction methods, and scanning electron microscopy (SEM). An inhibition assay for human MMP-1 (collagenase-1) and MMP-2 (gelatinase A) allowed estimation of the biological effect on tissue remodeling enzymes. The activity was also assessed with respect to inhibition of bacterial LasB. The formulated phytic acid demonstrated an IC50 of 109.7 µg/mL for LasB with viabilities > 80 % up to 188 µg/mL on A549 cells. Therefore, inhalation therapy with phytic acid-based powder shows promise as a treatment for early-stage Pseudomonas-induced pulmonary fibrosis., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

The chelation reaction of soluble soybean polysaccharide (SSPS) with zinc was investigated. Using response surface methodology, the optimum parameters for SSPS-Zn synthesis were obtained: pH 5.3, SSPS-ZnCl 2 mass ratio of 9.44:1, reaction temperature 50.44 °C, and reaction time 1.5 h, with the highest zinc content of 24.73 %. Compared with SSPS, SSPS-Zn increased in rhamnogalacturonan content and decreased in that of neutral monosaccharides (Fuc, Ara, Gal, Glu and Xyl). UV-vis spectra indicated that SSPS-Zn was lower than SSPS in protein content. FTIR spectra indicated that CO group of SSPS was bonded to Zn 2+ . X-ray diffraction spectra demonstrated that SSPS-Zn had higher crystallinity. Congo red reactions showed that SSPS possessed a triple-helix conformation while SSPS-Zn formed an irregular free-coiled conformation. EDX confirmed SSPS-Zn synthesis successfully. TGA curves exhibited that SSPS-Zn required higher temperature to undergo degradation. AFM revealed that SSPS-Zn was clustered while SSPS was filamentous. SEM micrographs showed the cracked fragments on the surface of SSPS-Zn. By in vitro simulation of gastrointestinal digestion, Zn 2+ release reached 68.87 % after 2 h digestion. Consequently, the chelation of SSPS with zinc could change structure and provide a basis for research and application of novel zinc supplements., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The design of novel chelators for therapeutic applications has been the subject of extensive research to address various diseases. Many chelators can manipulate the levels of metal ions within cells and effectively modulate the metal excess. In some cases, chelators show significant toxicity to cells. We investigated polyimidazole ligands by potentiometry and UV-Vis spectroscopy for their ability to form copper(II) complexes. We also compared the antiproliferative activity of the polyimidazole ligands and their copper(II) complexes with polypyridine ligands in CaCo-2 (colorectal adenocarcinoma), SH-SY5Y (neuroblastoma) and K562 (chronic myelogenous leukemia) cells and normal HaCaT (keratinocyte) cells. Polyimidazole ligands are less cytotoxic than their analogous polypyridine ligands. All polyimidazole ligands, except the tetraimidazole ligand for K562 cells, did not show any significant effect on the viability of cancer and normal cells. In contrast, the cytotoxic activity of polypiridine ligands was also observed in normal cells with IC 50 values similar to those of cancer cells. Tetraimidazole ligand, the only ligand active on the leukemic K562 cell line, induced caspase-dependent apoptosis and increased intracellular reactive oxygen species production with mitochondrial damage. The low cytotoxicity of the polyimidazole ligands, even if it limits their use as anticancer agents, could make them useful in other medical applications, such as in the treatment of metal overload, microbial infections, inflammation or neurodegenerative disorders., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. Graziella Vecchio reports financial support was provided by Italian Ministry of Research. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Inc.)

To fulfill the nutritional requirements of poultry, effective Zn supplementation is required due to Zn deficiency in basic feed. In this study, we investigated the effects of DMY-Zn (dihydromyricetin zinc chelate) on the growth performance, morphology, and biochemical indices; the expression of intestinal barrier-related genes; the intestinal microflora; and the cecum metabolome of Magang geese. A total of 300 14-day-old Magang geese (equal number of males and females) with an average body weight of 0.82 ± 0.08 kg were randomly divided into five groups and fed a basal diet; these groups were given DMY-Zn (low, medium, or high level of DMY-Zn with 30, 55, or 80 mg/kg Zn added to the basal diet) or ZnSO 4 (80 mg/kg Zn added) for 4 weeks. Our results revealed that DMY-Zn significantly impacts growth and biochemical indices and plays a significant role in regulating the intestinal barrier and microflora. DMY-Zn is involved in the upregulation of intestinal barrier gene (ZO1 and MUC2) expression, as well as upregulated Zn-related gene expression (ZIP5). On the other hand, a low concentration of DMY-Zn increased the ɑ diversity index and the abundance of Lactobacillus and Faecalibacterium. Additionally, a cecal metabolomics study showed that the main metabolic pathways affected by DMY-Zn were the pentose phosphate pathway, the biosynthesis of different alkaloids, and the metabolism of sphingolipids. In conclusion, DMY-Zn can reduce feed intake, increase the expression of intestinal barrier-related genes, help maintain the intestinal microflora balance, and increase the abundance of beneficial bacteria in the intestine to improve intestinal immunity., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Digital subtraction angiography (DSA) is considered the "gold standard" for the diagnosis of vascular diseases. However, the contrast agents used in DSA are limited to iodine (I)-based small molecules, which are unsuitable for patients with contraindications. Here, iodine-free DSA utilizing a bismuth (Bi) chelate, Bi-DTPA Dimeglumine, is proposed for vascular visualization for the first time. Bi-DTPA Dimeglumine possesses a simple synthesis process without the need for purification, large-scale production ability (over 200 g in the lab), superior X-ray imaging capability, renal clearance capacity, and good biocompatibility. Bi-DTPA-enhanced DSA can clearly display the arteries of the rabbit's head and lower limbs, with a minimum vascular resolution of 0.5 mm. The displayed integrity of terminal vessels by Bi-DTPA-enhanced DSA is superior to that of iopromide-enhanced DSA. In a rabbit model of thrombotic disease, Bi-DTPA Dimeglumine-enhanced DSA enables the detection of embolism and subsequent reevaluation of vascular conditions after recanalization therapy. This proposed iodine-free DSA provides a promising and universal approach for diagnosing vascular diseases., (© 2024 Wiley‐VCH GmbH.)

Rising concerns about global environmental degradation underscore the pressing need for effective solutions to combat heavy metal pollution. Industries such as semiconductor and steel production discharge vanadium into marine ecosystems, posing significant risks to both marine life and human health. The current study investigates efficacy of utilizing marine thraustochytrid for efficient vanadium removal outcompeting other microbial sources. By optimizing pH and temperature conditions during harvesting, achieved a remarkable 50.80 % enhancement in vanadium removal efficiency, from 19.31 to 29.12 mg/L. Furthermore, chelating agents EDTA and citric acid supplementation demonstrated promising enhancements, reaching up to 31.21 and 32.59 mg/L, respectively. Notably, vanadium-treated biomass supplemented with citric acid exhibited maximum enhancement in lipid content, from 58.47 to 75.34 %, indicating thraustochytrid's potential for biofuel production. This study presents a sustainable approach for industrial-scale vanadium bioremediation, aligning with Sustainable Development Goals focused on dual benefits of environmental protection and renewable energy., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Antifouling properties are crucial for enhancing the longevity and functionality of biomedical implants, drug delivery systems, and biosensors. Zwitterionic polymers are renowned for their exceptional surface hydration and charge neutrality, which effectively resist biomolecular adsorption and protein attachment. We propose an innovative approach to develop zwitterion-like antifouling surfaces by chelating divalent cations with anionic poly(3,4-ethylenedioxythiophene) (PEDOT) films, specifically PEDOT-PO 4 and PEDOT-COOH. The chelation behavior of these films was systematically evaluated using Na + , Mg 2+ , and Ca 2+ ions. Divalent ions, particularly Ca 2+ and Mg 2+ , exhibit a strong affinity for the anionic groups, leading to significant antifouling properties. These modified surfaces effectively repelled both negatively charged bovine serum albumin (BSA) and positively charged lysozyme (LYZ) proteins across various pH environments. This study offers valuable insights into the antifouling characteristics of charged surfaces, enhancing our understanding of how ion-mediated surface modifications influence protein adsorption and interactions.

We explore the potential of fluorine-containing small Mn 2+ chelates as alternatives to perfluorinated nanoparticles, widely used as 19 F MRI probes. In MnL1, the cyclohexanediamine skeleton and two piperidine rings, involving each a metal-coordinating amide group and an appended CF 3 moiety, provide high rigidity to the complex. This allows for good control of the Mn-F distance (r MnF =8.2±0.2 Å determined from 19 F relaxation data), as well as for high kinetic inertness (a dissociation half-life of 1285 h is estimated for physiological conditions). The paramagnetic Mn 2+ leads to a ~150-fold acceleration of the longitudinal 19 F relaxation, with moderate line-broadening effect, resulting in T 2 /T 1 ratios of 0.8 (9.4 T). Owing to its inner sphere water molecule, MnL1 is a good 1 H relaxation agent as well (r 1 =5.36 mM -1  s -1 at 298 K, 20 MHz). MnL1 could be readily visualized in 19 F MRI by using fast acquisition techniques, both in phantom images and living mice following intramuscular injection, with remarkable signal-to-noise ratios and short acquisition times. While applications in targeted imaging or cell therapy monitoring require further optimisation of the molecular structure, these results argue for the potential of such small, monohydrated and fluorinated Mn 2+ complexes for combined 19 F and 1 H MRI detection., (© 2024 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

The development of theranostic agents for radiopharmaceuticals based on therapeutic alpha emitters marks an important clinical need. We describe a strategy for the development of theranostic agents of this type via the functionalization of the ligand with the diagnostic radionuclide fluorine-18. An analogue of macropa, an 18-membered macrocyclic chelator with high affinity for alpha therapeutic radiometals, was synthesized and its complexation properties with metal ions were determined. The new macropa-F ligand was used for quantitative radiometal complexation with lead-203 and bismuth-207, as surrogates for their alpha-emitting radioisotopes. As a diagnostic partner, a radiofluorinated macropa ligand was used for quantitative bismuth(III) and lead(II) complexation. All fluorine-18 and radiometal complexes are highly stable in human serum over several days. This study presents a new proof-of-principle approach for developing theranostic agents based on alpha-emitting radionuclides and fluorine-18.

Mass cytometry is a bioanalytic tool based on atomic mass spectrometry for detecting biomarker expression on individual cells. Current reagents employ metal-chelating polymers binding isotopes of hard metal ions. Polymers bearing chelators for soft metal ions offer the promise for a large increase in multiplexing capabilities, but examples reported so far often have unacceptably high levels of nonspecific binding (NSB). We recently reported a new class of metal-chelating polymers with dipicolylamine (DPA) chelators that could bind Re and Pt. They also showed significant levels of NSB. Here, to reduce the NSB of the Pt-DPA polymer, we grafted water-soluble oligomers to the distal end of the dipicolylamine pendant group. Methoxy(polyethylene glycol) (DP = 24) was effective as was poly(sulfobetaine methacrylate) (DP = 29). Reacting the Pt-Cl bond of the metalated polymer with glutathione was remarkably effective at suppressing NSB. These results open the door to Pt-isotope-based metal-chelating polymers as new mass tags for mass cytometry.

Numerous studies have reported that mono-(2-ethylhexyl) phthalate (MEHP) (bioactive metabolite of Di(2-ethylhexyl) phthalate) has inhibitory effects on Leydig cells. This study aims to prepare an oyster peptide-zinc complex (PEP-Zn) to alleviate MEHP-induced damage in Leydig cells. Zinc-binding peptides were obtained through the following processes: zinc-immobilized affinity chromatography (IMAC-Zn 2+ ), liquid chromatography-mass spectrometry technology (LC-MS/MS) analysis, molecular docking, molecular dynamic simulation, and structural characterization. Then, the Zn-binding peptide (PEP) named Glu-His-Ala-Pro-Asn-His-Asp-Asn-Pro-Gly-Asp-Leu (EHAPNHDNPGDL) was identified. EHAPNHDNPGDL showed the highest zinc-chelating ability of 49.74 ± 1.44%, which was higher than that of the ethanol-soluble oyster peptides (27.50 ± 0.41%). In the EHAPNHDNPGDL-Zn complex, Asn-5, Asp-7, Asn-8, His-2, and Asp-11 played an important role in binding to the zinc ion. Additionally, EHAPNHDNPGDL-Zn was found to increase the cell viability, significantly increase the relative activity of antioxidant enzymes and testosterone content, and decrease malondialdehyde (MDA) content in MEHP-induced TM3 cells. The results also indicated that EHAPNHDNPGDL-Zn could alleviate MEHP-induced apoptosis by reducing the protein level of p53, p21, and Bax, and increasing the protein level of Bcl-2. These results indicate that the zinc-chelating peptides derived from oyster peptides could be used as a potential dietary zinc supplement.

Biodegradable radioactive microspheres labeled with positron emitters hold significant promise for diagnostic and therapeutic applications in cancers and other diseases, including arthritis. The alginate-based polymeric microspheres offer advantages such as biocompatibility, biodegradability, and improved stability, making them suitable for clinical applications. In this study, we developed novel positron emission tomography (PET) microspheres using alginate biopolymer radiolabeled with gallium-68 ( 68 Ga) through a straightforward conjugation reaction. Polyethylenimine (PEI)-decorated calcium alginate microspheres (PEI-CAMSs) were fabricated and further modified using azadibenzocyclooctyne- N -hydroxysuccinimide ester (ADIBO-NHS). Subsequently, azide-functionalized NOTA chelator (N 3 -NOTA) was labeled with [ 68 Ga]Ga to obtain [ 68 Ga]Ga-NOTA-N 3 , which was then reacted with the surface-modified PEI-CAMSs using strain-promoted alkyne-azide cycloaddition (SPAAC) reaction to develop [ 68 Ga]Ga-NOTA-PEI-CAMSs, a novel PET microsphere. The radiolabeling efficiency and radiochemical stability of [ 68 Ga]Ga-NOTA-PEI-CAMSs were determined using the radio-instant thin-layer chromatography-silica gel (radio-ITLC-SG) method. The in vivo PET images were also acquired to study the in vivo stability of the radiolabeled microspheres in normal mice. The radiolabeling efficiency of [ 68 Ga]Ga-NOTA-PEI-CAMSs was over 99%, and the microspheres exhibited high stability (92%) in human blood serum. PET images demonstrated the stability and biodistribution of the microspheres in mice for up to 2 h post injection. This study highlights the potential of biodegradable PET microspheres for preoperative imaging and targeted radionuclide therapy. Overall, the straightforward synthesis method and efficient radiolabeling technique provide a promising platform for the development of theranostic microspheres using other radionuclides such as 90 Y, 177 Lu, 188 Re, and 64 Cu.

Lead (Pb), a persistent and bio-accumulative contaminant, poses threats to the environment and human health. The effective removal of Pb from contaminated soil proves challenging due to its tendency to form stable complexes with soil components. Chelators have been extensively studied for their ability to extract metal contaminants, including Pb, from soil environment. However, the prolonged environmental persistence of traditional chelators and the high cost of biodegradable alternatives have hindered their practical application in remediation efforts. This study investigated a novel synergistic approach that combined a biodegradable chelator, [S,S]-ethylenediamine succinic acid (EDDS), with cationic and anionic surfactants to enhance Pb extraction efficiency. The study revealed that cationic surfactants, such as cetylpyridinium chloride (CPC) and cetyltrimethylammonium bromide (CTAB), significantly enhanced Pb extraction efficiency when combined with EDDS, whereas anionic surfactants, like sodium N-dodecanoyl-taurinate (SDT) and sodium dodecyl sulfate (SDS), inhibited the extraction process. Specifically, blending 5 mmol L -1 EDDS with 20 mmol L -1 CPC resulted in a 72.6% enhancement in Pb extraction efficiency. The proposed synergistic strategy offers a promising avenue for soil remediation, mitigating Pb contamination while preserving essential soil minerals. By addressing chelator limitations and improving efficiency, this approach presents a viable solution for enhancing soil remediation practices., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Fluoride (F) contamination in industrial waste is a significant challenge for sustainable materials recycling. Existing techniques for mitigating F contamination focus on immobilization, converting F compounds to insoluble forms while leaving the total F content untreated. Chelator-assisted washing is considered a promising alternative remediation strategy that can indirectly release F by entrapping and dissolving F-bearing minerals. This study evaluates the effectiveness of chelator-assisted washing in removing F from three real F-contaminated waste samples (TCS-49, TCS-51, and TCS-52) by treating with four different chelators, ethylenediaminetetraacetic acid (EDTA), ethylenediamine N,N'-disuccinic acid (EDDS), diethylenetriaminepentaacetic acid (DTPA), and 3-hydroxy-2,2'-imino disuccinic acid (HIDS). The influence of key washing variables, including chelator type, solution pH, chelator concentration, washing time, and liquid-to-solid (L/S) ratio toward F extraction was assessed and optimized for attaining the maximum extraction. All chelators exhibited the highest F extraction from TCS-49 and TCS-51 at pH 11, whereas in TCS-52 it showed a discrete extraction pattern. Under optimized conditions (concentration, 10 mmol L -1 ; pH, 11; washing duration, 3 h; and L/S ratio, 10:1), EDTA outperformed the other chelators, enhancing F extraction by 2.1 and 1.2 times for TCS-49 and TCS-51, respectively, compared with those of control treatments. However, for TCS-52, the efficiency of EDTA was analogous to that of the control under the same washing conditions. The linear correlation between the extracted F and F-containing minerals suggests that the chelator-induced F removal from contaminated waste involves the entrapment and dissolution of F-bearing minerals, especially Ca and Fe. The subsequent post-washing immobilization of chelator-washed waste residues using CaO or H 3 PO 4 significantly reduced the content of leachable F under the regulatory limit., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Phytoremediation, the use of plants to remove heavy metals from polluted environments, has been extensively studied. However, abiotic stresses such as drought, salt, and high temperatures can limit plant growth and metal uptake, reducing phytoremediation efficiency. High levels of HMs are also toxic to plants, further decreasing phytoremediation efficacy. This manuscript explores the potential of microbial-assisted and chelation-supported approaches to improve phytoremediation under abiotic stress conditions. Microbial assistance involves the use of specific microbes, including fungi that can produce siderophores. Siderophores bind essential metal ions, increasing their solubility and bioavailability for plant uptake. Chelation-supported methods employ organic acids and amino acids to enhance soil absorption and supply of essential metal ions. These chelating agents bind HMs ions, reducing their toxicity to plants and enabling plants to better withstand abiotic stresses like drought and salinity. Managed microbial-assisted and chelation-supported approaches offer more efficient and sustainable phytoremediation by promoting plant growth, metal uptake, and mitigating the effects of heavy metal and abiotic stresses. Managed microbial-assisted and chelation-supported approaches offer more efficient and sustainable phytoremediation by promoting plant growth, metal uptake, and mitigating the effects of HMs and abiotic stresses.These strategies represent a significant advancement in phytoremediation technology, potentially expanding its applicability to more challenging environmental conditions. In this review, we examined how microbial-assisted and chelation-supported techniques can enhance phytoremediation a method that uses plants to remove heavy metals from contaminated sites. These approaches not only boost plant growth and metal uptake but also alleviate the toxic effects of HMs and abiotic stresses like drought and salinity. By doing so, they make phytoremediation a more viable and effective solution for environmental remediation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

2-((1-(4-((2,4,6-trioxohexahydropyrimidin-5-yl)diazenyl) phenyl) ethylidene) amino) benzoic acid (H 3 L), and its V(IV), Co(II), Ni(II), Cu(II), Pd(II) and Ag(I) chelates were synthesized. They were defined using multiple spectral and analytical techniques. With the exception of Ag(I) chelate, all chelates possessed non-electrolytic character. Square pyramidal shape was proposed for V(IV) chelate and Square planar for the other chelates. The analysis of functional group bands of H 3 L and its coordination compounds alludes that H 3 L chelated as neutral tetradentate via nitrogen atoms of azo and azomethine groups, oxygen atom of carbonyl of barbituric acid and OH of the carboxylic group. TG/DTG predicted the thermal behaviors of all compounds. The antibacterial activity of H 3 L and its coordination compounds was conducted against Proteus mirabilis at concentrations of 250, 500, and 1000 µg/mL. Ag(I) at 1000 µg/mL, showed the most inhibiting potency against P. mirabilis and registered zone of inhibition of 28.33 ± 0.84 mm and highest biofilm inhibition of 70.31%. At 50 Gy of gamma irradiation, the reducing effect of Ag(I) chelate was improved. The protein interruption of P. mirabilis was greatly interrupted by increasing the concentration of the chaletes. Also, Ag(I) showed the highest cytotoxicity with IC 50 value of 11.5 µg/ mL. The novelty of this study is the synthesis of a new azo-Schiff base and this is almost the first publication of the effect of azo-Schiff ligands against that bacterial strain P. mirabilis., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Rapid monitoring of trace antibiotics in the field in real time is essential for environment forewarning and human health. High sensitivity and real-time on-site quantitative monitoring of antibiotic residues can be accomplished by integrating portable sensors alongside fluorescent optics to construct an intelligent sensing platform that smoothly eliminates the instability of conventional detection methods. In this study, a ratiometric fluorescence sensor for the ultrasensitive detection of pefloxacin was built employing the photoinduced electron transfer (PET) mechanism from red Eu-MOFs to Mn 2+ -PEF complex. A visual color change results from the photoinduced electron transfer process from manganese ions to pefloxacin weakening the ligand metal charge transfer (LMCT) process in Eu-MOFs. This enables the ultrafast visible detection of pefloxacin and produces a transient shift in visual color with a detection limit as low as 15.4 nM. For the detection of pefloxacin in water, tomato, and raw pork samples, various sensing devices based on the developed fluorescent probes exhibit good practicability and accuracy. With the development of the ratiometric fluorescence sensing probe, it is now possible to quickly and quantitatively identify pefloxacin residues in the environment, offering a new method for ensuring the safety of food and people's health., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Objective: Prostate-specific membrane antigen (PSMA) is a well-known biomarker of prostate cancer. Previously, our group reported that the succinimidyl-cystatin-urea-glutamate (SCUE) moiety has a high affinity for PSMA. In this study, we developed the novel technetium-99m-labeled PSMA-targeting probe "[ 99m Tc]Tc-(Ham-SCUE) 2 " based on a hydroxamamide chelate with a bivalent SCUE and evaluated its potential as a SPECT imaging probe for the diagnosis of PSMA-expressing prostate cancer., Methods: Ham-SCUE was synthesized by a one-step reaction with Ham-Mal and cysteine-urea-glutamine. Then, Ham-SCUE was reacted with [ 99m Tc]NaTcO 4 for 10 min at room temperature to obtain [ 99m Tc]Tc-(Ham-SCUE) 2 . [ 99m Tc]Tc-(Ham-SCUE) 2 was added to LNCaP (high PSMA expression) cells or PC3 (low PSMA expression) cells, and their radioactivity was measured 60 min after administration. The blocking study was performed by co-incubation of LNCaP cells with various concentrations of 2-PMPA (a PSMA inhibitor) for 15 min before adding [ 99m Tc]Tc-(Ham-SCUE) 2 . The biodistribution of [ 99m Tc]Tc-(Ham-SCUE) 2 in LNCaP/PC3 dual xenografted C.B.-17/Icr scid/scid Jcl mice was evaluated for 120 min after intravenous injection. The blocking study was performed by pretreatment of mice with 2-PMPA (10 mg/kg weight)., Results: [ 99m Tc]Tc-(Ham-SCUE) 2 was acquired at radiochemical yields of 56% with a radiochemical purity of over 95%. The cellular uptake level of [ 99m Tc]Tc-(Ham-SCUE) 2 by LNCaP cells was significantly higher than that by PC3 cells (LNCaP: 11.12 ± 0.71 vs. PC3: 1.40 ± 0.13%uptake/mg protein, p < 0.01), and the uptake was significantly suppressed by pretreatment with 2-PMPA (2.56 ± 0.37%uptake/mg protein, p < 0.05). IC 50 of 2-PMPA was 245 ± 47 nM. In the in vivo study, the radioactivity of LNCaP tumor tissue was significantly higher than that of PC3 tumor tissue at 120 min after the administration of [ 99m Tc]Tc-(Ham-SCUE) 2 (LNCaP: 9.97 ± 2.79, PC3: 1.16 ± 0.23%ID/g, p < 0.01), and was suppressed by pretreatment with 2-PMPA (2.50 ± 0.45%ID/g, p < 0.01)., Conclusion: [ 99m Tc]Tc-(Ham-SCUE) 2 has the potential to be a SPECT imaging agent for diagnosing high PSMA-expressing prostate cancer., (© 2024. The Author(s) under exclusive licence to The Japanese Society of Nuclear Medicine.)

Brachytherapy stands as an essential clinical approach for combating locally advanced tumors. Here, an injectable brachytherapy hydrogel is developed for the treatment of both local and metastatic tumor. Fe-tannins nanoparticles are efficiently and stably radiolabeled with clinical used therapeutic radionuclides (such as 131 I, 90 Y, 177 Lu, and 225 Ac) without a chelator, and then chemically cross-linked with 4-armPEG-SH to form brachytherapy hydrogel. Upon intratumoral administration, magnetic resonance imaging (MRI) signal from ferric ions embedded within the hydrogel directly correlates with the retention dosage of radionuclides, which can real-time monitor radionuclides emitting short-range rays in vivo without penetration limitation during brachytherapy. The hydrogel's design ensures the long-term tumor retention of therapeutic radionuclides, leading to the effective eradication of local tumor. Furthermore, the radiolabeled hydrogel is integrated with an adjuvant to synergize with immune checkpoint blocking therapy, thereby activating potent anti-tumor immune responses and inhibiting metastatic tumor growth. Therefore, this work presents an imageable brachytherapy hydrogel for real-time monitoring therapeutic process, and expands the indications of brachytherapy from treatment of localized tumors to metastatic tumors., (© 2024 Wiley‐VCH GmbH.)

The dyshomeostasis of metal ions in the brain leads to the accumulation of excess metals in extracellular and inter-neuronal locations and the Amyloid β peptide (Aβ) binds these transition metals, which ultimately cause the Aβ aggregation and severe oxidative stress in the brain. The aggregation of Aβ and oxidative stress are important factors to trigger Alzheimer's disease (AD). Metal chelation therapy is a promising approach to removing metals from Aβ-M species and relieve the oxidative stress. Therefore, 4 tetrahydrosalens containing benzothiazole moiety were designed and synthesized. Their biological activities for Alzheimer's disease therapy in vitro were determined by Turbidity assay, BCA protein assay, MTT assay and fluorescent probe of DCFH-DA. The results were comparing with that of non-specific chelator (cliquinol, CQ) and non-benzothiazole functionalized tetrahydrosalens, the results demonstrated that benzothiazole functionalized chelators had more efficient bio-activities in preventing Cu 2+ -induced Aβ aggregation, attenuating cytotoxicity mediated by Aβ-Cu 2+ species and decrease the level of reactive oxygen species (ROS) in Cu 2+ -Aβ treated PC12 cells than that of cliquinol and non-benzothiazole functionalized analogues., Competing Interests: Declaration of competing interest We solemnly promise that this manuscript is the original paper, and that all or part of contents of this manuscript has never published in any other publication in any form, and that there is no problem of multiple contributions. We also declare that we have no financial and personal relationships with other people or organizations that can inappropriately influence our works, there is no professional or other personal interest of any nature or kind in any product, service and/or company that could be construed as influencing the position presented in, or the review of, the manuscript entitled., (Copyright © 2024 Elsevier Inc. All rights reserved.)

D-penicillamine (PA) is the primary chelator of choice to treat Wilson disease (WD). There are limitations in obtaining comprehensive data on PA metabolites in biological specimens by conventional approaches. Hence, the aim of the present was to identify the major hepatic PA metabolites and draw clear conclusions of the drug's xenobiotic in WD. Urine samples were collected from children with hepatic WD (n = 63, aged 14.8 ± 4 years) 5 h after PA administration (16.3 ± 3.8 mg/kg/day) and age-matched healthy volunteers comprised as controls (n = 30). High-resolution 800 MHz nuclear magnetic resonance spectroscopy (NMR) and mass spectrometry was applied to reveal unambiguous appraisals of different excretory by-products of PA metabolism. Four new products comprising penicillamine disulphide (PD), penicillamine cysteine disulphide (PCD), S-methyl penicillamine (SMP), and N-acetyl penicillamine (NAP) of PA xenobiotic metabolites were identified using high-resolution NMR spectroscopy. Quantitative levels of PCD and SMP were approximately three-fold higher than those of PD and NAP, respectively. High-resolution NMR identifies the major PA metabolites with certainty. Reduction, sulfation, and methylation are the predominant pathways of PA metabolism. There is a potential application for assessing therapeutic monitoring of chelation in hepatic WD., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Abstract: During a nuclear/radiological incident or an accident involving internal intakes with radioactive cobalt or strontium, the recommended treatments, consisting of the administration of diethylenetriaminepentaacetic acid for 60 Co and calcium gluconate for 90 Sr, are of low specificity, and their effectiveness can be enhanced. In this manuscript, a liposomal formulation was developed to deliver potential chelating agents to the main retention organs of both radionuclides. A bisphosphonate, etidronate, has been selected as a possible candidate due to its satisfying decorporation activity for uranium, bone tropism, and potential affinity with cobalt. Pre-clinical studies have been carried out on rats using radionuclide contamination and treatment administration by the intravenous route. The effectiveness of free or liposomal etidronate was evaluated, with an administration at 30 min, 48 h post-contamination with 60 Co. Regarding 85 Sr, a more extended experiment with etidronate liposomes was performed over 6 d. The results were compared to those performed with reference treatments, diethylenetriaminepentaacetic acid for cobalt and calcium gluconate for strontium. Unexpected results were found for the reference treatments that were significantly less effective than previously reported or showed no effectiveness. Free etidronate revealed no significant efficacy after 48 h, but the liposomal form suggested an interaction with radionuclides, not sufficient to change the biokinetics. This study emphasizes the need for early treatment administration and further research to provide a more effective medical countermeasure., (Copyright © 2024 Health Physics Society.)

We report the synthesis, characterization, and in vitro biological activities of [Re(η 6 -arene) 2 ] + -terpyridine conjugates and their Cu II complexes. The terpyridine (terpy) chelators were attached to the [Re(η 6 -arene) 2 ] + scaffold via secondary amine linkers allowing for heteroleptic mono- and homoleptic bis-terpyridine-substituted chelators. Complexation with CuCl 2 afforded the respective square pyramidal [Cu(terpy)Cl 2 ] complexes hosted on the [Re(η 6 -arene) 2 ] + scaffold. The chelator conjugates and their respective complexes were found to be remarkably cytotoxic against malignant HT29 and A549 human cancer cell lines in vitro with IC 50 values in the low micromolar range. Mitochondrial respiration disruption was identified as a possible mode of action of these novel drug candidates. Crucially, the [Re(η 6 -arene) 2 ] + hosts delivered water solubility of the otherwise insoluble [Cu(terpy)Cl 2 ] motif. Importantly, the homoleptic [ 99m Tc(η 6 -arene) 2 ] + -terpyridine conjugate is available in a single step, which enables the presented system to be used as a theranostic approach to modern medicinal inorganic chemistry.

Lanthanum (La(III)) is recognized for its ability to mitigate heavy metal stress in plants. However, the inorganic La(III) salts and lanthanum oxide nanoparticles (La 2 O 3 NPs) extensively used in agriculture are prone to soil immobilization, thereby compromising their bioavailability and posing environmental risks. This study synthesized and characterized the lanthanum(III)-amino acid chelate (La(III)-AA) from soybean protein isolate (SPI) hydrolysates. Maximum chelating rate (94.95%) was achieved under the conditions of mole ratio 1:1.5, pH 8.0, 50 ℃ and 5 h. Glu, Asp and Pro represent the primary La(III)-binding ligands. UV-vis and FTIR demonstrated that amino nitrogen and carboxyl oxygen participate in metal-ligand recognition. Scanning and Transmission electron microscopy showed that La(III) chelates with amino acids in a core-shell structure of uniform size. Consequently, a proposed chemical structure for the La(III)-AA complex was presented. A concentration of 20 mg/L La(III)-AA outperforms inorganic La salts in growth promotion and Cu detoxification. La(III)-AA significantly reduces the content of Cu (II) in rice tissues and enhances seedling tolerance to Cu (II) stress. This study provides a novel La(III)-based candidate for crop protection and furthers our understanding of rare earth element-induced mitigation of heavy metal stress., (© 2024. The Author(s).)

Single-domain antibodies, or nanobodies (Nbs), are promising biomolecules for use in molecular imaging due to their excellent affinity, specificity, and fast clearance from the blood. Given their short blood half-life, pairing Nbs with short-lived imaging radioisotopes is desirable. Because fluorine-18 ( 18 F) is routinely used for clinical imaging, it is an attractive radioisotope for Nbs. We report a novel sortase-based, site-specific 18 F-labeling method applied to three nanobodies. Labeled nanobodies were synthesized either by a two-step indirect radiolabeling method in one pot or by a one-step direct labeling method using a sortase-mediated conjugation of either the radiolabeled chelator (H-GGGK((±)-Al[ 18 F]FH 3 RESCA)-NH 2 ) or the unlabeled chelator (H-GGGK((±)-H 3 RESCA)-NH 2 ) followed by labeling with Al[ 18 F]F, respectively. The overall radiochemical yields were 15-43% ( n = 22, decay-corrected) in 70 min (indirect labeling) and 23-58% ( n = 12, decay-corrected) in 50 min (direct labeling). The radiochemical purities of the labeled nanobodies prepared by both methods were >98% with a specific activity of 400-600 Ci/mmol ( n = 22) for each of the three Nbs tested and exhibited excellent stability profiles under physiological conditions. This simple, site-specific, reproducible, and generalizable 18 F-labeling method to prepare nanobodies (Nb-Al[ 18 F]F-RESCA) or other low molecular weight biomolecules can easily be adopted in various settings for preclinical and clinical studies.

Radiopharmaceuticals are currently a key tool in cancer diagnosis and therapy. Metal-based radiopharmaceuticals are characterized by a radiometal-chelator moiety linked to a bio-vector that binds the biological target (e.g., a protein overexpressed in a particular tumor). The right match between radiometal and chelator influences the stability of the complex and the drug's efficacy. Therefore, the coupling of the radioactive element to the correct chelator requires consideration of several features of the radiometal, such as its oxidation state, ionic radius, and coordination geometry. In this work, we systematically investigated about 120 radiometal-chelator complexes taken from the Cambridge Structural Database. We considered 25 radiometals and about 30 chelators, featuring both cyclic and acyclic geometries. We used quantum mechanics methods at the density functional theoretical level to generate the general AMBER force field parameters and to perform 1 µs-long all-atom molecular dynamics simulations in explicit water solution. From these calculations, we extracted several key molecular descriptors accounting for both electronic- and dynamical-based properties. The whole workflow was carefully validated, and selected test-cases were investigated in detail. Molecular descriptors and force field parameters for the complexes considered in this study are made freely available, thus enabling their use in predictive models, molecular modelling, and molecular dynamics investigations of the interaction of compounds with macromolecular targets. Our work provides new insights in understanding the properties of radiometal-chelator complexes, with a direct impact for rational drug design of this important class of drugs.

Thorium-232 (Th-232) is a promising fuel for advanced nuclear reactors. However, in case of internal human exposure to Th, there is currently no effective modality for its removal from liver and skeleton or for mitigating its effect. The FDA-approved agent, diethylenetriaminepentaacetate (DTPA), can remove Th and other actinides from blood circulation only. For the first time, a rationally-selected polyherbal hepatoprotective i.e. Liv52® (L52S), was evaluated in-combination with DTPA for its Th decorporation ability in Swiss mice. Inductively-coupled plasma mass spectroscopic analysis showed that oral administration of L52S in conjunction with DTPA significantly decreased Th burden from liver (20 %) and skeleton (33 %) as well as enhanced Th excretion (∼2.5 folds) through urine in comparison to DTPA or L52S alone. The combinatorial therapy was found to be complementary in-action, ameliorating Th-induced tissue damage in liver, spleen, and bone more effectively than monotherapy. Furthermore, markers of liver function (alanine transaminase) and liver inflammation and fibrosis (NF-κB & keratin) further validated the beneficial effect of L52S. The human consumption of L52S for various liver disorders further supports its clinical application for Th decorporation and mitigation of its health effects., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

This study employed a diverse approach to extract antioxidant peptides from red seaweed Palmaria palmata , recognized for its comparatively high protein content. Initially, an aqueous extraction of the entire seaweed was performed, followed by enzymatic hydrolysis of the solid residues prepared from the first step. The effects of three different pH levels (3, 6, and 9) during the aqueous extraction were also examined. Results indicated that the solid fraction from the sequential extraction process contained significantly higher levels of proteins and amino acids than other fractions ( p < 0.05). Furthermore, the solid fractions (IC 50 ranging from 2.29 to 8.15 mg.mL -1 ) demonstrated significantly greater free radical scavengers than the liquid fractions (IC 50 ranging from 9.03 to 10.41 mg.mL -1 or not obtained at the highest concentration tested) at both stages of extraction ( p < 0.05). Among the solid fractions, those produced fractions under alkaline conditions were less effective in radical scavenging than the produced fractions under acidic or neutral conditions. The fractions with most effective metal ion chelating activity were the solid fractions from the enzymatic stage, particularly at pH 3 (IC 50 = 0.63 ± 0.04 mg.mL -1 ) and pH 6 (IC 50 = 0.89 ± 0.07 mg.mL -1 ), which were significantly more effective than those from the initial extraction stage ( p < 0.05). Despite no significant difference in the total phenolic content between these solid fractions and their corresponding liquid fractions (3.79 ± 0.05 vs. 3.48 ± 0.02 mg.mL -1 at pH 3 and 2.43 ± 0.22 vs. 2.51 ± 0.00 mg.mL -1 at pH 6) ( p > 0.05), the observed antioxidant properties may be attributed to bioactive amino acids such as histidine, glutamic acid, aspartic acid, tyrosine, and methionine, either as free amino acids or within proteins and peptides.

Indium-111 ( 111 In) is a diagnostic radiometal that is important in nuclear medicine for single-photon emission computed tomography (SPECT). In order to apply this radiometal, it needs to be stably chelated and conjugated to a targeting vector that delivers it to diseased tissue. Identifying effective chelators that are capable of binding and retaining [ 111 In]In 3+ in vivo is an important research area. In this study, two 18-membered macrocyclic chelators, py-macrodipa and py 2 -macrodipa, were investigated for their ability to form stable coordination complexes with In 3+ and to be effectively radiolabeled with [ 111 In]In 3+ . The In 3+ complexes of these two chelators were characterized by NMR spectroscopy, X-ray crystallography, and density functional theory calculations. These studies show that both py-macrodipa and py 2 -macrodipa form 8-coordinate In 3+ complexes and attain an asymmetric conformation, consistent with prior studies on this ligand class with small rare earth metal ions. Spectrophotometric titrations were carried out to determine the thermodynamic stability constants (log  K ML ) of [In(py-macrodipa)] + and [In(py 2 -macrodipa)] + , which were found to be 18.96(6) and 19.53(5), respectively, where the values in parentheses are the errors of the last significant figures obtained from the standard deviation from three independent replicates. Radiolabeling studies showed that py-macrodipa and py 2 -macrodipa can quantitatively be radiolabeled with [ 111 In]In 3+ at 25 °C within 5 min, even at ligand concentrations as low as 1 μM. The in vitro stability of the radiolabeled complexes was investigated in human serum at 37 °C, revealing that ∼90% of [ 111 In][In(py-macrodipa)] + and [ 111 In][In(py 2 -macrodipa)] + remained intact after 7 days. The biodistribution of these radiolabeled complexes in mice was investigated, showing lower uptake in the kidneys, liver, and blood at the 24 h mark compared to [ 111 In]InCl 3 . These results demonstrate the potential of py-macrodipa and py 2 -macrodipa as chelators for [ 111 In]In 3+ , suggesting their value for SPECT radiopharmaceuticals.

Here we describe an anti-prostate-specific membrane antigen (PSMA) minibody (IAB2MA) conjugated to an octadentate, macrocyclic chelator based on four 1-hydroxypyridin-2-one coordinating units (Lumi804 [L804]) labeled with 89 Zr (PET imaging) and 177 Lu (radiopharmaceutical therapy), with the goal of developing safer and more efficacious treatment options for prostate cancer. Methods: L804 was compared with the current gold standard chelators, DOTA and deferoxamine (DFO), conjugated to IAB2MA for radiolabeling with 177 Lu and 89 Zr in cell binding, preclinical biodistribution, imaging, dosimetry, and efficacy studies in the PSMA-positive PC3-PIP tumor-bearing mouse model of prostate cancer. Results: Quantitative radiolabeling (>99% radiochemical yield) of L804-IAB2MA with 177 Lu or 89 Zr was achieved at ambient temperature in under 30 min, comparable to 89 Zr labeling of DFO-IAB2MA. In contrast, DOTA-IAB2MA was radiolabeled with 177 Lu for 30 min at 37°C in approximately 90% radiochemical yield, requiring further purification. Using europium(III) as a luminescent surrogate, high binding affinity of Eu-L804-IAB2MA to PSMA was demonstrated in PC3-PIP cells (dissociation constant, 4.6 ± 0.6 nM). All 4 radiolabeled constructs showed significantly higher levels of internalization after 30 min in the PC3-PIP cells than in PSMA-negative PC3-FLU cells. The accumulation of 177 Lu- and 89 Zr-L804-IAB2MA in PC3-PIP tumors and all organs examined (i.e., heart, liver, spleen, kidney, muscle, salivary glands, lacrimal glands, carcass, and bone) was significantly lower than that of 177 Lu-DOTA-IAB2MA and 89 Zr-DFO-IAB2MA at 96 and 72 h after injection, respectively. Generally, SPECT/CT and PET/CT imaging data showed no significant difference in the SUV mean of the tumors or muscle between the radiotracers. Dosimetry analysis via both organ-level and voxel-level dose calculation methods indicated significantly higher absorbed doses of 177 Lu-DOTA-IAB2MA in tumors, kidney, liver, muscle, and spleen than of 177 Lu-L804-IAB2MA. PC3-PIP tumor-bearing mice treated with single doses of 177 Lu-L804-IAB2MA (18.4 or 22.2 MBq) exhibited significantly prolonged survival and reduced tumor volume compared with unlabeled minibody control. No significant difference in survival was observed between groups of mice treated with 177 Lu-L804-IAB2MA or 177 Lu-DOTA-IAB2MA (18.4 or 22.2 MBq). Treatment with 177 Lu-L804-IAB2MA resulted in lower absorbed doses in tumors and less toxicity than that of 177 Lu-DOTA-IAB2MA. Conclusion: 89 Zr- and 177 Lu-L804-IAB2MA may be a promising theranostic pair for imaging and therapy of prostate cancer., (© 2024 by the Society of Nuclear Medicine and Molecular Imaging.)

Phyllanthus emblica L. fruit extract (PFE) was introduced to improve ZVI/H 2 O 2 technology, and the efficiency and mechanism of PFE promoting ZVI/H 2 O 2 technology were explored. With the introduction of PFE, the Norfloxacin (NOR) removal rate and k obs of the process were improved by 41.17% and 5.08 times, respectively. In the ZVI/H 2 O 2 /PFE process, the degradation of NOR by the attack of ROS is the main pathway for decontamination and is dominated by the heterogeneous reaction on the catalyst surface. PFE contains 13.92 g/L titratable acid and has good complexing ability and antioxidant ability. The mechanism of PFE promoting ZVI/H 2 O 2 technology was based on lowering the pH, complemented by chelation and antioxidant capacity. With the introduction of PFE, the utilization rate of the reagent was significantly increased (7.56 times for ZVI and 3.21 times for H 2 O 2 ), the applicable pH range was widened (6-9) and the iron sludge was reduced (32.80%). Meanwhile, the concept of UPR is proposed for the first time. The result is the key role to the selection of green promoters in the ZVI/H 2 O 2 process depends on the abundance of titratable acid, followed by a certain chelating ability and antioxidant capacity., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

In this research, a novel kind of walnut (Juglans regia L.) peptides-zinc (Zn-WPs) chelate was obtained using the mass ratio of the walnut peptides (WPs) to ZnSO 4 .7H 2 O of 3.5:1 at pH 8.5 and 50°C for 84 min, with the chelation rate of 84.5%. In comparison to walnut peptides (WPs), the contents of aspartic acid and glutamic acid in Zn-WPs chelate are approximately 27%, indicating that hydrophilic amino acids predominantly bind with walnut peptides. Following chelation with zinc ions, the ultraviolet-visible (UV) characteristic absorption peak shifted from 213 nm to 210 nm, while the average particle size of the chelate increased to 8.0 ± 0.14 µm, presenting a loose spherical structure under scanning electron microscopy. These findings suggest the formation of new substances. Fourier-transform infrared spectroscopy (FTIR) revealed carboxyl, amino, and peptide bonds as the chelation sites of WPs and zinc. The IC 50 of walnut peptides-zinc (Zn-WPs) chelate is 2.91 mg/mL, indicative of a favorable DPPH radical scavenging rate. Furthermore, Zn-WPs chelate microcapsules were produced via the spray drying method, achieving an encapsulation rate of 75.67 ± 0.83% under optimal conditions. These microcapsules demonstrate robust stability across diverse environmental conditions. This study underscores the potential of Zn-WPs and its chelate microcapsules to enhance stability and bioactivity under varying circumstances. PRACTICAL APPLICATION: In this study, a new walnut peptide-zinc (Zn-WPs) chelate was prepared. The presence of zinc ions changes the structure and properties of walnut peptides and improves its stability. The production of Zn-WPs chelate microcapsules enables Zn-WPs to have strong in vitro stability under different pH and simulated gastrointestinal digestion conditions. These results provide novel insights for developing the walnut peptides as bioactive ingredients in functional foods., (© 2024 Institute of Food Technologists.)

We analysed how citrate (CIT), a chelating agent potentially present in radioactive waste disposals, affects the mobility of four radionuclides (RN): 63 Ni, 233 U, 152 Eu, 238 Pu in portlandite, an important hydrated phase of cement, a commonly used material for waste isolation. Portlandite was synthetized in the laboratory and showed high purity and grain size of few μm. This solid, buffers the pH to 12.5 and shows high adsorption capability for the studied RNs: 152 Eu and 238 Pu exhibited the highest adsorption (K d ∼1·10 5  mL g -1 ) and 233 U the lowest (K d ∼8·10 2  mL g -1 ). CIT adsorption was also experimentally evaluated by batch sorption experiments and electrophoretic (ζ-potential) measurements: a non-lineal sorption behaviour was observed, with K d values decreasing (from ∼1·10 3  mL g -1 ) as CIT concentration increased up to 1·10 -2  M, according to portlandite sorption sites saturation. In the presence of CIT, a marginal decrease for 233 U adsorption in portlandite was observed, one order of magnitude reduction for 63 Ni, while 238 Pu and 152 Eu adsorption decreased significantly. The calculated sorption reduction factors (SRF) for the four RN in the presence of CIT at a concentration of 5·10 -3  M were: 2.4, 9.7, 37 and 50.9 for 233 U, 63 Ni, 238 Pu, and 152 Eu, respectively. According to the available thermodynamic databases, low complexation between CIT and RN is predicted at pH = 12.5, thus the RN adsorption decrease in the presence of CIT must be attributed to the organic adsorption on portlandite. However, current thermodynamic are still incomplete for this ligand and this pH range and this limits a precise interpretation of the experimental data., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Depletion and separation of histidine-rich proteins from complicated biosamples are crucial for various downstream applications in proteome research and clinical diagnosis. Herein, porous polymer microspheres coated with polyacrylic acid (SPSDVB-PAA) were fabricated through double emulsion interfacial polymerization technique and followed by immobilization of Cu 2+ ions on the surface of SPSDVB-PAA. The as-prepared SPSDVB-PAA-Cu with uniform size and nanoscale pore structure enabled coordination interaction of Cu 2+ with histidine residues in his-rich proteins, resulting in high-performance adsorption. As metal affinity adsorbent, the SPSDVB-PAA-Cu exhibited favorable selectivity for adsorbing hemoglobin (Hb) and human serum albumin (HSA) with the maximum adsorption capacities of 152.2 and 100.7 mg g -1 . Furthermore, the polymer microspheres were used to isolate histidine-rich proteins from human whole blood and plasma, underscoring their effectiveness. The liquid chromatography tandem mass spectrometry (LC-MS/MS) results indicated that the content of 14 most abundant proteins in human plasma was depleted from 81.6 % to 30.7 % and low-abundance proteins were enriched from 18.4 % to 69.3 % after treatment with SPSDVB-PAA-Cu, illustrating potential application of SPSDVB-PAA-Cu in proteomic research., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

This study focused on exploring the Zn 2+ chelating peptide GFLGSP: the characterization of structure/Zn 2+ chelating mode and the potential mechanisms for promoting Zn 2+ transport in Caco-2 cells. The findings revealed the bidentate chelating between Zn 2+ and carboxyl oxygen atom in Pro6 residue. Thereafter, the secondary structure of GFLGSP remained unchanged, but there was an increase in zeta potential and particle size. Notably, the GFLGSP-Zn 2+ complex enhanced the Zn 2+ transport rate and modulated ZIP4 and ZNT1 expression in a Caco-2 cells monolayer model. As revealed by molecular docking analysis, GFLGSP interacted with ZIP4 through intermolecular hydrogen bonds as well as Van der Waals forces. The Zn 2+ transport mechanisms of the GFLGSP-Zn 2+ complex encompassed ZIP4 (vital channel), endocytosis (primary pathway) and paracellular transport (supplementary pathway). Based on these results, the tilapia skin collagen-derived GFLGSP hold promise as the potential dietary Zn 2+ supplement., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Purpose: Accumulating evidence suggests that neurotensin (NTS) and neurotensin receptors (NTSRs) play key roles in lung cancer progression by triggering multiple oncogenic signaling pathways. This study aims to develop Cu-labeled neurotensin receptor 1 (NTSR1)-targeting agents with the potential for both imaging and therapeutic applications., Method: A series of neurotensin receptor antagonists (NRAs) with variable propylamine (PA) linker length and different chelators were synthesized, including [ 64 Cu]Cu-CB-TE2A-iPA-NRA ([ 64 Cu]Cu-4a-c, i = 1, 2, 3), [ 64 Cu]Cu-NOTA-2PA-NRA ([ 64 Cu]Cu-4d), [ 64 Cu]Cu-DOTA-2PA-NRA ([ 64 Cu]Cu-4e, also known as [ 64 Cu]Cu-3BP-227), and [ 64 Cu]Cu-DOTA-VS-2PA-NRA ([ 64 Cu]Cu-4f). The series of small animal PET/CT were conducted in H1299 lung cancer model. The expression profile of NTSR1 was also confirmed by IHC using patient tissue samples., Results: For most of the compounds studied, PET/CT showed prominent tumor uptake and high tumor-to-background contrast, but the tumor retention was strongly influenced by the chelators used. For previously reported 4e, [ 64 Cu]Cu-labeled derivative showed initial high tumor uptake accompanied by rapid tumor washout at 24 h. The newly developed [ 64 Cu]Cu-4d and [ 64 Cu]Cu-4f demonstrated good tumor uptake and tumor-to-background contrast at early time points, but were less promising in tumor retention. In contrast, our lead compound [ 64 Cu]Cu-4b demonstrated 9.57 ± 1.35, 9.44 ± 2.38 and 9.72 ± 4.89%ID/g tumor uptake at 4, 24, and 48 h p.i., respectively. Moderate liver uptake (11.97 ± 3.85, 9.80 ± 3.63, and 7.72 ± 4.68%ID/g at 4, 24, and 48 h p.i.) was observed with low uptake in most other organs. The PA linker was found to have a significant effect on drug distribution. Compared to [ 64 Cu]Cu-4b, [ 64 Cu]Cu-4a had a lower background, including a greatly reduced liver uptake, while the tumor uptake was only moderately reduced. Meanwhile, [ 64 Cu]Cu-4c showed increased uptake in both the tumor and the liver. The clinical relevance of NTSR1 was also demonstrated by the elevated tumor expression in patient tissue samples., Conclusions: Through the side-by-side comparison, [ 64 Cu]Cu-4b was identified as the lead agent for further evaluation based on its high and sustained tumor uptake and moderate liver uptake. It can not only be used to efficiently detect NTSR1 expression in lung cancer (for diagnosis, patient screening, and treatment monitoring), but also has the great potential to treat NTSR-positive lesions once chelating to the beta emitter 67 Cu., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Benzoic acids, which are commonly found in food, are also produced by human microbiota from other dietary phenolics. The aim was to investigate the interactions of 8 food-related benzoic acids with the physiological metals iron and copper under different (patho)physiologically relevant pH conditions in terms of chelation, reduction, impact on the metal-based Fenton chemistry, and copper-based hemolysis. Only 3,4-dihydroxybenzoic acid behaved as a protective substance under all conditions. It chelated iron, reduced both iron and copper, and protected against the iron and copper-based Fenton reaction. Conversely, 2,4,6-trihydroxybenzoic acid did not chelate iron and copper, reduced both metals, potentiated the Fenton reaction, and worsened copper-based hemolysis of rat red blood cells. The other tested compounds showed variable effects on the Fenton reaction. Interestingly, prooxidative benzoic acids mildly protected human erythrocytes against Cu-induced lysis. In conclusion, 3,4-dihydroxybenzoic acid seems to have a protective effect against copper and iron-based toxicity under different conditions., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

In general, Cu(II) is the critical factor in catalyzing reactive oxygen species (ROS) production and accelerating amyloid-β (Aβ) oligomer formation in Alzheimer's disease (AD). Natural chelating agents with good biocompatibility and appropriate binding affinity with Cu(II) have emerged as potential candidates for AD therapy. Herein, we tested the capability of guluronic acid disaccharide (Di-GA), a natural chelating agent with the moderate association affinity to Cu(II), in inhibiting ROS production and Aβ oligomer formation. The results showed that Di-GA was capable of chelating with Cu(II) and reducing ROS production, even in solutions containing Fe(II), Zn(II), and Aβ. In addition, Di-GA can also capture Cu(II) from Cu-Aβ complexes and decrease Aβ oligomer formation. The cellular results confirmed that Di-GA prevented SH-SY5Y cells from ROS and Aβ oligomer damage by reducing the injury of ROS and Aβ oligomers on cell membrane and decreasing their intracellular damage on mitochondria. Notably, the slightly higher efficiency of Di-GA in chelating with Cu(I) than Cu(II) can be benefit for its in vivo applications, as Cu(I) is not only the most active but also the special intermediate specie during ROS production process. All of these results proved that Di-GA could be a promising marine drug candidate in reducing copper-related ROS damage and Aβ oligomer toxicity associated with AD., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Inc.)

Anastomotic leaks are among the most dreaded complications following gastrointestinal (GI) surgery, and contrast-enhanced X-ray gastroenterography is considered the preferred initial diagnostic method for GI leaks. However, from fundamental research to clinical practice, the only oral iodinated contrast agents currently available for GI leaks detection are facing several challenges, including low sensitivity, iodine allergy, and contraindications in patients with thyroid diseases. Herein, we propose a cinematic contrast-enhanced X-ray gastroenterography for the real-time detection of GI leaks with an iodine-free bismuth chelate (Bi-DTPA) for the first time. The Bi-DTPA, synthesized through a straightforward one-pot method, offers distinct advantages such as no need for purification, a nearly 100 % yield, large-scale production capability, and good biocompatibility. The remarkable X-ray attenuation properties of Bi-DTPA enable real-time dynamic visualization of whole GI tract under both X-ray gastroenterography and computed tomography (CT) imaging. More importantly, the leaky site and severity can be both clearly displayed during Bi-DTPA-enhanced gastroenterography in a rat model with esophageal leakage. The proposed movie-like Bi-DTPA-enhanced X-ray imaging approach presents a promising alternative to traditional GI radiography based on iodinated molecules. It demonstrates significant potential in addressing concerns related to iodine-associated adverse effects and offers an alternative method for visually detecting gastrointestinal leaks., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Ltd.)

Background: Peanut peptides can chelate iron but their chelation mechanism remains unclear. The purpose of this study is to separate peanut ferrous-chelating peptides and explore the chelation mechanism of peanut peptides with iron., Results: Peanut peptide component F-122, which had a higher chelation rate, was separated using ultrafiltration, gel filtration chromatography, and ion exchange chromatography, achieving a ferrous chelation rate of 90.7%. Six peptide segments were screened and their amino acid sequences were identified by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Spectral analysis confirmed that the chelation between peanut peptides and ferrous ions occurred and a new substance was formed. Molecular docking simulation indicated that the amino acids in peanut peptides involved in the chelating reaction were glutamic acid, arginine, glycine, threonine, phenylalanine, and lysine. The binding sites included the main chain oxygen atom, side chain oxygen atom, and carboxyl oxygen atom of amino acid., Conclusion: The isolated peanut peptide had a higher ferrous-chelation rate. The chelating mechanism of peanut peptide with ferrous ion was elucidated. This study provides a theoretical basis for the development of new peptide-ferrous preparations. © 2024 Society of Chemical Industry., (© 2024 Society of Chemical Industry.)

Background: Peanut peptides have good chelating ability with metal ions. However, there are few studies on the chelation mechanism of peanut peptides with calcium and absorption properties of peptide-calcium complex., Results: Peptides with high calcium chelating rate were isolated and purified from peanut protein hydrolysate (PPH), and the chelation rate of component F21 was higher (81.4 ± 0.8%). Six peptides were identified from component F21 by liquid chromatography-tandem mass spectrometry, and the frequency of acidic amino acids and arginine in the amino acid sequence was higher in all six peptides. Peanut peptide-calcium complex (PPH21-Ca) was prepared by selecting component F21 (PPH21). Ultraviolet analysis indicated that the chelate reaction occurred between peanut peptide and calcium ions. Fourier transform infrared analysis showed that the chelating sites were carboxyl and amino groups on the amino acid residues of peptides. Scanning electron microscopy revealed that the surface of peanut peptide had a smooth block structure, but the surface of the complex had a granular morphology. Caco-2 cell model tests revealed that the bioavailability of PPH21-Ca was 58.4 ± 0.5%, which was significantly higher than that of inorganic calcium at 37.0 ± 0.4%., Conclusion: Peanut peptides can chelate calcium ions by carboxyl and amino groups, and the peptide-calcium complex had higher bioavailability. This study provides a theoretical basis for the development of new calcium supplement products that are absorbed easily. © 2024 Society of Chemical Industry., (© 2024 Society of Chemical Industry.)

The latest advancements in nuclear medicine indicate that radioactive isotopes and associated metal chelators play crucial roles in the diagnosis and treatment of diseases. The development of metal chelators mainly relies on traditional trial-and-error methods, lacking rational guidance and design. In this study, we propose the structure-aware transformer (SAT) combined with molecular fingerprint (SATCMF), a novel graph transformer network framework that incorporates prior chemical knowledge to construct coordination edges and learns the interactions between chelating agents and metal ions. SATCMF is trained on stability data collected from metal ion-ligand complexes, leveraging the SAT network to extract structural features relevant to the binding of ligands with metal ions. It further integrates molecular fingerprint features to refine the prediction of the stability constants of the chelating agents and metal ions. The experimental results on benchmark data set demonstrate that SATCMF achieves state-of-the-art performance based on four different graph neural network architectures. Additionally, visualizing the learned molecular attention distribution provides interpretable insights from the prediction results, offering valuable guidance for the development of novel metal chelators.

This study investigates the potential of using ionic liquids as cosolvents to enhance the solubility and activity of poorly soluble rhodium(III) complexes, particularly those with diene, pyridine derivatives, and camphor-derived bis-pyrazolylpyridine ligands, in relation to 5'-GMP, CT-DNA, and HSA as well as their biological activity. Findings indicate that ionic liquids significantly increase the substitution activity of these complexes toward 5'-GMP while only marginally affecting DNA/HSA binding affinities with molecular docking, further confirming the experimental results. Lipophilicity assessments indicated good lipophilicity. Notably, cytotoxicity studies show that Rh2 is selectively effective against HeLa cancer cells, with IL1 and IL10 modulating the cytotoxic effects. Redox evaluations indicate that rhodium complexes induce oxidative stress in cancerous cells while maintaining redox balance in noncancerous cells. By elucidating the role of ionic liquids in modulating these effects, the study proposes a promising avenue for augmenting the efficacy and selectivity of cancer treatments, thus opening new horizons in cancer therapeutics.

The chelation of protein-bound metal ions is typically thought to follow either a dissociative (D) or an associative (A) path. While the former mechanism involves the spontaneous dissociation of the metal from the protein prior to chelation, the latter route is characterized by the formation of an intermediate protein-metal-chelator ternary complex. Using the prototypical zinc protease carboxypeptidase A (CPA) and a variety of charged and neutral chelating agents, we demonstrate that inactivation of the enzyme (and likely other metalloproteins) proceeds through a split pathway with contributions from both D- and A-type mechanisms. In the case of charged chelators such as ethylenediaminetetraacetic acid (EDTA), the proportions of both paths could be tuned over a wide range through variation of the chelator concentration and the ionic strength, I (from ∼95% A type at low I values to ∼5% at high I values). By measuring the EDTA concentration and time dependence of CPA inactivation and fitting the obtained kinetic data to a modified time-dependent inhibition model, we obtained the rate constants for the A and D paths ( k inact and k off , respectively) and the inhibition constant ( K I ) for the formation of the CPA-Zn 2+ -EDTA ternary complex, indicating that the decreased contribution of the A-type mechanism at high ionic strengths originates from a large (40-fold; at I = 0.5 M) increase in K I . This observation might be related to a triarginine motif in CPA that electrostatically steers negatively charged substrates into the active site and may therefore also guide carboxylate-bearing chelators toward the Zn 2+ ion.

Tracer antibodies, which are labelled with fluorescent or other type of reporter molecules, are widely employed in diagnostic immunoassays. Time-resolved fluorescence immunoassay (TRFIA), recognized as one of the most sensitive immunoassay techniques, utilizes tracers labelled with lanthanide ion (Ln) chelates. The conventional approach for conjugating isothiocyanate (ITC) Ln-chelates to antibodies involves random chemical targeting of the primary amino group of Lys residues, requiring typically overnight exposure to an elevated pH of 9-9.3 and leading to heterogeneity. Moreover, efforts to enhance the sensitivity of the assays by introducing a higher number of Ln-chelates per tracer antibody are associated with an elevated risk of targeting critical amino acid residues in the binding site, compromising the binding properties of the antibody. Herein, we report a method to precisely label recombinant antibodies with a defined number of Ln-chelates in a well-controlled manner by employing the SpyTag/SpyCatcher protein ligation technology. We demonstrate the functionality of the method with a full-length recombinant antibody (IgG) as well as an antibody fragment by producing site-specifically labelled antibodies for TRFIA for cardiac troponin I (cTnI) detection with a significant improvement in assay sensitivity compared to that with conventionally labelled tracer antibodies. Overall, our data clearly illustrates the benefits of the site-specific labelling strategy for generating high-performing tracer antibodies for TRF immunoassays., (© 2024. The Author(s).)