Water Ligands Research Articles

Atomic View of Aqueous Cyclosporine A: Unpacking a Decades-Old Mystery.

An atomic view of a main aqueous conformation of cyclosporine A (CycA), an important 11-amino-acid macrocyclic immunosuppressant, is reported. For decades, it has been a grand challenge to determine the conformation of free CycA in an aqueous-like solution given its poor water solubility. Using a combination of X-ray and single-crystal neutron diffraction, we unambiguously resolve a unique conformer (A1) with a novel cis-amide between residues 11 and 1 and two water ligands that stabilize hydrogen bond networks. NMR spectroscopy and titration experiments indicate that the novel conformer is as abundant as the closed conformer in 90/10 (v/v) methanol/water and is the main conformer at 10/90 methanol/water. Five other conformers were also detected in 90/10 methanol/water, one in slow exchange with A1, another one in slow exchange with the closed form and three minor ones, one of which contains two cis amides Abu2-Sar3 and MeBmt1-MeVal11. These conformers help better understand the wide spectrum of membrane permeability observed for CycA analogues and, to some extent, the binding of CycA to protein targets.

Decomposition of dinitrosyl iron complex with thioformaldehyde ligands in water: reaction mechanisms and the role of chemical hardness of ligands

The mechanisms of hydrolysis of a model cationic dinitrosyl iron complex with a prototypic thioformaldehyde ligand have been studied using the density functional theory and polarizable continuum water model. The free-energy calculations have predicted that the associative mechanism of the thioformaldehyde ligand removal has a ∼34 kJ mol-1 lower activation barrier in water than the dissociative mechanism. The additional estimates of chemical hardness have provided useful qualitative characterization of the thio ligands binding.

Reversible Structural Isomerization of Nature's Water Oxidation Catalyst Prior to O-O Bond Formation.

Photosynthetic water oxidation is catalyzed by a manganese–calcium oxide cluster, which experiences five “S-states” during a light-driven reaction cycle. The unique “distorted chair”-like geometry of the Mn4CaO5(6) cluster shows structural flexibility that has been frequently proposed to involve “open” and “closed”-cubane forms from the S1 to S3 states. The isomers are interconvertible in the S1 and S2 states, while in the S3 state, the open-cubane structure is observed to dominate inThermosynechococcus elongatus (cyanobacteria) samples. In this work, using density functional theory calculations, we go beyond the S3+Yz state to the S3nYz• → S4+Yz step, and report for the first time that the reversible isomerism, which is suppressed in the S3+Yz state, is fully recovered in the ensuing S3nYz• state due to the proton release from a manganese-bound water ligand. The altered coordination strength of the manganese–ligand facilitates formation of the closed-cubane form, in a dynamic equilibrium with the open-cubane form. This tautomerism immediately preceding dioxygen formation may constitute the rate limiting step for O2 formation, and exert a significant influence on the water oxidation mechanism in photosystem II.

Analysis of Anion Binding Effects on the Sensitized Luminescence of Macrocyclic Europium(III) Complexes

AbstractFour triazamacrocyclic and two tetraazamacrocyclic ligands carrying two and three coordinating donor groups, respectively, and a carbostyril light‐harvesting antenna were synthesized. The antenna was linked to a macrocycle either via secondary or tertiary amide. Complexation with europium (Eu), gadolinium (Gd), terbium (Tb), and ytterbium (Yb) yielded overall +1 or +2 charged species. Paramagnetic 1H NMR, and steady‐state and time‐resolved luminescence spectroscopies showed that the complexes had 1–3 inner‐sphere water molecules and displayed a variety of coordination geometries in solution. The antennae sensitized Eu(III) and Tb(III) emission with quantum yields of 0.3–4.3 % and 9.9–24.5 %, respectively. The addition of excess fluoride or cyanide to buffered Eu(III) complex solutions resulted in anion‐dependent changes. Fluoride addition increased the Eu(III) luminescence intensity by displacing all inner‐sphere water molecules and stabilizing the +3 oxidation state of Eu. Eu(III) luminescence increased up to 25‐fold for one emitter. Cyanide quenched Eu(III) luminescence in all cases despite partial water ligand displacement.

Prediction of Stability Constants of metal-ligand Complexes Using Thermodynamic Radii of Metal Ions

ABSTRACT Linear Free Energy Relationships (LFER) logK j = a⋅logK i between the stabilityconstants (logK) of the Mn+L complexes of ith and jth metal ions Mn+ (Ca2+, Cd2+, Co2+, Cu2+, Fe2+, La3+, Mg2+, Mn2+, Ni2+, Pb2+, Th4+, VO2+, Y3+, Zn2+) with diverse organic ligands (L) in water were analyzed. From these 432 LFER it follows that each metal ion can bedescribed by one parameter - thermodynamic ionic radius r, and thestability constants of two metal ions M i and M j with a ligand L are related by the simple relationship logKj = (ri /rj )logKi . Predictive performance of this relationship was tested on 228 metal ion pairs which were not used forthe radii estimations, and the number of ligands varied from 20 to 221for these metal ion pairs. Rootmean squared error RMSE and squared determination coefficient of the logK predictions vary from 0.50 to 2.0 for 186 test sets and from 0.849 to 0.500 for 218 test sets, respectively.

Leveraging Nitrogen Linkages in the Formation of a Porous Thorium-Organic Nanotube Suitable for Iodine Capture.

We report the synthesis, characterization, and iodine capture application of a novel thorium-organic nanotube, TSN-626, [Th6O4(OH)4(C6H4NO2)7(CHO2)5(H2O)3]·3H2O. The classification as a metal-organic nanotube (MONT) distinguishes it as a rare and reduced dimensionality subset of metal-organic frameworks (MOFs); the structure is additionally hallmarked by low node connectivity. TSN-626 is composed of hexameric thorium secondary building units and mixed O/N-donor isonicotinate ligands that demonstrate selective ditopicity, yielding both terminating and bridging moieties. Because hard Lewis acid tetravalent metals have a propensity to bind with electron donors of rival hardness (e.g., carboxylate groups), such Th-N coordination in a MOF is uncommon. However, the formation of key structural Th-N bonds in TSN-626 cap some of the square antiprismatic metal centers, a position usually occupied by terminal water ligands. TSN-626 was characterized by using complementary analytical and computational techniques: X-ray diffraction, vibrational spectroscopy, N2 physisorption isotherms, and density functional theory. TSN-626 satisfies design aspects for the chemisorption of iodine. The synergy between accessibility through pores, vacancies at the metal-oxo nodes, and pendent N-donor sites allowed a saturated iodine loading of 955 mg g-1 by vapor methods. The crystallization of TSN-626 diversifies actinide-MOF linker selection to include soft electron donors, and these Th-N linkages can be leveraged for the investigation of metal-to-ligand bonding and unconventional topological expressions.

Structure Elucidation, Hirshfeld Surface Analysis, Molecular Docking and Computational Studies of a Jahn-Teller Distorted Octahedral Cobalt (II) Complex with Saccharin Ligand

A Jahn-Teller distorted octahedral cobalt(II) complex with saccharin and water ligands was synthesized and characterized using single-crystal X-ray diffraction, density functional theory calculation, surface analysis (MEP and Hirshfeld) and molecular docking studies. Co(II) center is six-coordinated in a distorted octahedral arrangement, with the two water O atoms located in the axial positions, and the two water O atoms and the saccharin N atoms located in the equatorial plane. Theoretical molecular modeling was carried out with Density Functional Theory (DFT) using UB3LYP with hybrid function LANL2DZ. To support intra-molecular and inter-molecular bondings in crystal structure, molecular electrostatic potential (MEP) and Hirshfeld surfaces are generated and visualized. The molecular docking analysis was investigated between cobalt complex ligand and the receptor of breast cancer mutant PDB: 3HB5-oxidoreductase.

Adsorption mechanism of hydrated Lu(OH)2+ and Al(OH)2+ ions on the surface of kaolinite

Density functional theory was used to study the adsorption mechanism of hydrated Lu(OH)2+ and Al(OH)2+ ions on (001) kaolinite surface. Calculation results showed that the respective most stable hydration configurations in the water system were Lu(OH)(H2O)72+ and Al(OH)(H2O)52+. Both hydrated ions were adsorbed on the outer layer of the (001) kaolinite aluminium octahedral and silica tetrahedral surfaces through hydrogen bonding. The two hydrated ions were adsorbed on the deprotonated (001) aluminium octahedral kaolinite surface via coordination bonds, with electrons transferred from the water ligand O 2p to the Al 2 s, Lu 5d, and the surface O 2p orbitals. Owing to its smaller radius and fewer water ligands, hydrated Al(OH)2+ was more easily adsorbed on the (001) kaolinite surface than hydrated Lu(OH)2+. The experimental adsorption and microcalorimetry results supported this conclusion, and it is hypothesised that hydrated ions mainly act on the kaolinite surface via inner-layer adsorption.

Negatively Charged MOF-Based Composite Anion Exchange Membrane with High Cation Selectivity and Permeability.

Every metal and metallurgical industry is associated with the generation of wastewater, influencing the living and non-living environment, which is alarming to environmentalists. The strict regulations about the dismissal of acid and metal into the environment and the increasing emphasis on the recycling/reuse of these effluents after proper remedy have focused the research community’s curiosity in developing distinctive approaches for the recovery of acid and metals from industrial wastewaters. This study reports the synthesis of UiO-66-(COOH)2 using dual ligand in water as a green solvent. Then, the prepared MOF nanoparticles were introduced into the DMAM quaternized QPPO matrix through a straightforward blending approach. Four defect-free UiO-66-(COOH)2/QPPO MMMs were prepared with four different MOF structures. The BET characterization of UiO-66-(COOH)2 nanoparticles with a highly crystalline structure and sub-nanometer pore size (~7 Å) was confirmed by XRD. Because of the introduction of MOF nanoparticles with an electrostatic interaction and pore size screening effect, a separation coefficient (SHCl/FeCl2) of 565 and UHCl of 0.0089 m·h−1 for U-C(60)/QPPO were perceived when the loading dosage of the MOF content was 10 wt%. The obtained results showed that the prepared defect-free MOF membrane has broad prospects in acid recovery applications.

Discrete, Cationic Palladium(II)-Oxo Clusters via f-Metal Ion Incorporation and their Macrocyclic Host-Guest Interactions with Sulfonatocalixarenes.

We report on the discovery of the first two examples of cationic palladium(II)‐oxo clusters (POCs) containing f‐metal ions, [PdII 6O12M8{(CH3)2AsO2}16(H2O)8]4+ (M=CeIV, ThIV), and their physicochemical characterization in the solid state, in solution and in the gas phase. The molecular structure of the two novel POCs comprises an octahedral {Pd6O12}12− core that is capped by eight MIV ions, resulting in a cationic, cubic assembly {Pd6O12MIV 8}20+, which is coordinated by a total of 16 terminal dimethylarsinate and eight water ligands, resulting in the mixed PdII‐CeIV/ThIV oxo‐clusters [PdII 6O12M8{(CH3)2AsO2}16(H2O)8]4+ (M=Ce, Pd6Ce8 ; Th, Pd6Th8 ). We have also studied the formation of host‐guest inclusion complexes of Pd6Ce8 and Pd6Th8 with anionic 4‐sulfocalix[n]arenes (n=4, 6, 8), resulting in the first examples of discrete, enthalpically‐driven supramolecular assemblies between large metal‐oxo clusters and calixarene‐based macrocycles. The POCs were also found to be useful as pre‐catalysts for electrocatalytic CO2‐reduction and HCOOH‐oxidation.

Discrete, Cationic Palladium(II)‐Oxo Clusters via f‐Metal Ion Incorporation and their Macrocyclic Host‐Guest Interactions with Sulfonatocalixarenes

AbstractWe report on the discovery of the first two examples of cationic palladium(II)‐oxo clusters (POCs) containing f‐metal ions, [PdII6O12M8{(CH3)2AsO2}16(H2O)8]4+ (M=CeIV, ThIV), and their physicochemical characterization in the solid state, in solution and in the gas phase. The molecular structure of the two novel POCs comprises an octahedral {Pd6O12}12− core that is capped by eight MIV ions, resulting in a cationic, cubic assembly {Pd6O12MIV8}20+, which is coordinated by a total of 16 terminal dimethylarsinate and eight water ligands, resulting in the mixed PdII‐CeIV/ThIV oxo‐clusters [PdII6O12M8{(CH3)2AsO2}16(H2O)8]4+ (M=Ce, Pd6Ce8; Th, Pd6Th8). We have also studied the formation of host‐guest inclusion complexes of Pd6Ce8 and Pd6Th8 with anionic 4‐sulfocalix[n]arenes (n=4, 6, 8), resulting in the first examples of discrete, enthalpically‐driven supramolecular assemblies between large metal‐oxo clusters and calixarene‐based macrocycles. The POCs were also found to be useful as pre‐catalysts for electrocatalytic CO2‐reduction and HCOOH‐oxidation.

Molecular Principles of Redox-Coupled Protonation Dynamics in Photosystem II.

Photosystem II (PSII) catalyzes light-driven water oxidization, releasing O2 into the atmosphere and transferring the electrons for the synthesis of biomass. However, despite decades of structural and functional studies, the water oxidation mechanism of PSII has remained puzzling and a major challenge for modern chemical research. Here, we show that PSII catalyzes redox-triggered proton transfer between its oxygen-evolving Mn4O5Ca cluster and a nearby cluster of conserved buried ion-pairs, which are connected to the bulk solvent via a proton pathway. By using multi-scale quantum and classical simulations, we find that oxidation of a redox-active Tyrz (Tyr161) lowers the reaction barrier for the water-mediated proton transfer from a Ca2+-bound water molecule (W3) to Asp61 via conformational changes in a nearby ion-pair (Asp61/Lys317). Deprotonation of this W3 substrate water triggers its migration toward Mn1 to a position identified in recent X-ray free-electron laser (XFEL) experiments [Ibrahim et al. Proc. Natl. Acad. Sci. USA 2020, 117, 12,624–12,635]. Further oxidation of the Mn4O5Ca cluster lowers the proton transfer barrier through the water ligand sphere of the Mn4O5Ca cluster to Asp61 via a similar ion-pair dissociation process, while the resulting Mn-bound oxo/oxyl species leads to O2 formation by a radical coupling mechanism. The proposed redox-coupled protonation mechanism shows a striking resemblance to functional motifs in other enzymes involved in biological energy conversion, with an interplay between hydration changes, ion-pair dynamics, and electric fields that modulate the catalytic barriers.

Crystallographic snapshots of ternary complexes of thermophilic secondary alcohol dehydrogenase from Thermoanaerobacter pseudoethanolicus reveal the dynamics of ligand exchange and the proton relay network.

Three-dimensional structures of I86A and C295A mutant secondary alcohol dehydrogenase (SADH) from Thermoanaerobacter pseudoethanolicus were determined by x-ray crystallography. The tetrameric structure of C295A-SADH soaked with NADP+ and dimethyl sulfoxide (DMSO) was determined to 1.85 Å with an Rfree of 0.225. DMSO is bound to the tetrahedral zinc in each subunit, with ligands from SG of Cys-37, NE2 of His-59, and OD2 of Asp-150. The nicotinamide ring of NADP is hydrogen-bonded to the N of Ala-295 and the O of Val-265 and Gly-293. The O of DMSO is connected to a network of hydrogen bonds with OG of Ser-39, the 3'-OH of NADP, and ND1 of His-42. The structure of I86A-SADH soaked with 2-pentanol and NADP+ contains (R)-2-pentanol bound in each subunit, ligated to the tetrahedral zinc, and connected to the proton relay network. The structure of I86A-SADH soaked with 3-methylcyclohexanol and NADP+ has alcohol bound in three subunits. Two of the sites have the alcohol ligated to the zinc in an axial position, with OE2 of Glu-60 in the other axial position of a trigonal bipyramidal complex. One site has 3-methylcyclohexanol bound noncovalently, with the zinc in an inverted tetrahedral geometry with Glu-60. The fourth site also has the zinc in a trigonal bipyramidal complex with axial Glu-60 and water ligands. These structures demonstrate that ligand exchange of SADH involves pentacoordinate and inverted zinc complexes with Glu-60. Furthermore, we see a network of hydrogen bonds connecting the substrate oxygen to the external solvent that is likely to play a role in the mechanism of SADH.

SYNTHESIS AND SPECTROSCOPIC CHARACTERIZATION OF SILVER (I) MEBENDAZOLE COMPLEXES

Objective: The aim of this research work is to synthesize silver-mebendazole complexes with imidazole (imi), 2,2-bipyridine (bpy) and 1,10-phenanthroline (phen) as secondary ligands that will be active and effective against microbes. Methods: Environmentally friendly techniques were employed to synthesize the new complexes by reacting silver nitrate and ligands in water and at ambient conditions. The formed complexes were isolated and dried at ambient conditions and characterized by UV and FTIR spectroscopies; the physicochemical and antimicrobial properties of the complexes against Proteus mirabilis and Salmonella typhi were also tested by agar well diffusion method. Results: Spectroscopic studies confirmed the formation of the complexes. In the newly synthesized complexes, silver is coordinated by N of the imidazole moiety of mebendazole as well as N atoms of imidazole, 2,2-bipyridine and 1,10-phenanthroline. The complexes are stable and possess low aqueous solubility. Antimicrobial analyses revealed that they were active against the tested organisms with antibacterial activities higher than the parent drug, mebendazole. Conclusion: The newly synthesized complexes were stable and the results show their antimicrobial potential. The resultant antimicrobial activity of the complexes is as a result of synergistic activities of silver and the ligands.

Role of water coordination at zinc binding site and its catalytic pathway of dizinc creatininase: insights from quantum cluster approach.

Creatininase is a key enzyme of creatinine-metabolizing pathway in mammals, and has a great potential for diagnostic application. It catalyzes the reversible conversion of creatinine to creatine. Here, we investigated its reaction mechanism with density functional theory in conjunction with the quantum cluster approach. Three reaction pathways in which several possible proton transfers assisted by either His178 or a water ligand to Zn1 (Wat2) or both were considered. DFT calculations reveal, depending on Wat2 coordination mode at Zn1, two competitive ring-opening pathways where His178 playing a central role as a proton shuttle or both His178 and Wat2 serving as a dual catalytic role as a base and an acid, respectively. Three elementary steps were proposed for the reaction: the first involves nucleophilic attack by a bridging hydroxide to the substrate and forms a gem-diolate intermediate, followed by a proton transfer from the gem-diolate to His178 (His178 protonation is a required step for efficient proton transfers). Finally, the second proton transfer from the protonated His178 or Wat2 to the amide of substrate leads to the ring opening. The first proton transfer is the rate-limiting step of the whole reaction, in consistent with previous experimental and computational studies. A detailed understanding of the reaction mechanism of the creatininase enzyme family will also be helpful for developing a biosensor for kidney function.

Oxygen-Atom Defect Formation in Polyoxovanadate Clusters via Proton-Coupled Electron Transfer.

The uptake of hydrogen atoms (H-atoms) into reducible metal oxides has implications in catalysis and energy storage. However, outside of computational modeling, it is difficult to obtain insight into the physicochemical factors that govern H-atom uptake at the atomic level. Here, we describe oxygen-atom vacancy formation in a series of hexavanadate assemblies via proton-coupled electron transfer, presenting a novel pathway for the formation of defect sites at the surface of redox-active metal oxides. Kinetic investigations reveal that H-atom transfer to the metal oxide surface occurs through concerted proton–electron transfer, resulting in the formation of a transient VIII–OH2 moiety that, upon displacement of the water ligand with an acetonitrile molecule, forms the oxygen-deficient polyoxovanadate-alkoxide cluster. Oxidation state distribution of the cluster core dictates the affinity of surface oxido ligands for H-atoms, mirroring the behavior of reducible metal oxide nanocrystals. Ultimately, atomistic insights from this work provide new design criteria for predictive proton-coupled electron-transfer reactivity of terminal M=O moieties at the surface of nanoscopic metal oxides.

Hydration Structures and Dynamics of Ga3+ Ion Based on Molecular Mechanics Molecular Dynamics Simulation (Classical DM)

The structure and hydration dynamics of Ga3+ ion have been studied using classical Molecular Dynamics (MD) simulations. The data collection procedure includes determining the best base set, constructing 2-body and 3-body potential equations, classical molecular dynamics simulations based on 2-body potentials, classical molecular dynamics simulations based on 2-body + 3 potential-body. The trajectory file data analysis was done to obtain structural properties parameters such as RDF, CND, ADF, and dynamic properties, namely the movement of H2O ligands between hydrations shells. The results of the research indicated that the hydration complex structure of Ga(H2O)83+ and Ga(H2O)63+ was observed in molecular dynamics simulations (MM-2 body) and (MM-2 body + 3-body), respectively. The movement of H2O ligands occurs between the first and second shell or vice versa in the MD simulation of MM-2 bodies but does not occur in MD simulations of (MM-2 bodies + MM-3 bodies). Therefore, the water ligands in the first hydrated shell are stable.

The Apparently Unreactive Substrate Facilitates the Electron Transfer for Dioxygen Activation in Rieske Dioxygenases.

Rieske dioxygenases belong to the non‐heme iron family of oxygenases and catalyze important cis‐dihydroxylation as well as O‐/N‐dealkylation and oxidative cyclization reactions for a wide range of substrates. The lack of substrate coordination at the non‐heme ferrous iron center, however, makes it particularly challenging to delineate the role of the substrate for productive O2 activation. Here, we studied the role of the substrate in the key elementary reaction leading to O2 activation from a theoretical perspective by systematically considering (i) the 6‐coordinate to 5‐coordinate conversion of the non‐heme FeII upon abstraction of a water ligand, (ii) binding of O2 , and (iii) transfer of an electron from the Rieske cluster. We systematically evaluated the spin‐state‐dependent reaction energies and structural effects at the active site for all combinations of the three elementary processes in the presence and absence of substrate using naphthalene dioxygenase as a prototypical Rieske dioxygenase. We find that reaction energies for the generation of a coordination vacancy at the non‐heme FeII center through thermoneutral H2O reorientation and exothermic O2 binding prior to Rieske cluster oxidation are largely insensitive to the presence of naphthalene and do not lead to formation of any of the known reactive Fe‐oxygen species. By contrast, the role of the substrate becomes evident after Rieske cluster oxidation and exclusively for the 6‐coordinate non‐heme FeII sites in that the additional electron is found at the substrate instead of at the iron and oxygen atoms. Our results imply an allosteric control of the substrate on Rieske dioxygenase reactivity to happen prior to changes at the non‐heme FeII in agreement with a strategy that avoids unproductive O2 activation.

Fragment-Based Drug Discovery for Trypanosoma brucei Glycosylphosphatidylinositol-Specific Phospholipase C through Biochemical and WaterLOGSY-NMR Methods.

Glycosylphosphatidylinositol-specific phospholipase C (GPI-PLC) of Trypanosoma brucei, the causative protozoan parasite of African trypanosomiasis, is a membrane-bound enzyme essential for antigenic variation, because it catalyses the release of the membrane-bound form of variable surface glycoproteins. Here, we performed a fragment-based drug discovery of TbGPI-PLC inhibitors using a combination of enzymatic inhibition assay and water ligand observed via gradient spectroscopy (WaterLOGSY) NMR experiment. The TbGPI-PLC was cloned and overexpressed using an Escherichia coli expression system followed by purification using three-phase partitioning and gel filtration. Subsequently, the inhibitory activity of 873 fragment compounds against the recombinant TbGPI-PLC led to the identification of 66 primary hits. These primary hits were subjected to the WaterLOGSY NMR experiment where 10 fragment hits were confirmed to directly bind to the TbGPI-PLC. These included benzothiazole, chlorobenzene, imidazole, indole, pyrazol and quinolinone derivatives. Molecular docking simulation indicated that six of them share a common binding site, which corresponds to the catalytic pocket. The present study identified chemically diverse fragment hits that could directly bind and inhibit the TbGPI-PLC activity, which constructed a framework for fragment optimization or linking towards the design of novel drugs for African trypanosomiasis.

Prospects and limitations of paramagnetic chemical exchange saturation transfer agents serving as biological reporters in vivo.

The concept of using paramagnetic metal ion complexes as chemical exchange saturation transfer agents (paraCEST) for molecular imaging of various biological processes first appeared in the literature about 20 years ago. The first paraCEST agent was based on a highly shifted, inner-sphere, slowly exchanging water molecule that could be activated at a frequency far away from bulk water, a substantial advantage for selective activation of the agent alone. Many other paraCEST agent designs followed that were based on activation of exchanging -NH or -OH proton on the chelate itself. Both types of paraCEST designs are attractive for molecular imaging because the rates of water molecule or ligand proton exchange can be designed to be sensitive to a biological or physiological property such as pH, enzyme activity, or redox. Hence, the intensity or frequency of the resulting CEST signal provides a direct readout of that property. Many molecular designs have appeared in the literature over the past 20 years, mostly reported as proof-of-concept designs but, unfortunately, only a few reports have explored the limitations of paraCEST agents for imaging a biological process in vivo. As a community, we now know that the sensitivity of paraCEST agents is lower than one might anticipate based upon simple chemical exchange principles and, in general, it appears the sensitivity of paraCEST agents is even lower in vivo than in vitro. In this short review, we address some of the factors that contribute to the limited sensitivity of paraCEST agents in vivo, offer some thoughts on approaches that could lead to dramatically improved paraCEST sensitivity, and challenge the scientific community to perform more in vivo experiments designed to test these ideas.