Monitoring Enzyme Activity Using Near-Infrared Fluorescent Single-Walled Carbon Nanotubes.

Enzymes serve as pivotal biological catalysts that accelerate essential chemical reactions, thereby influencing a variety of physiological processes. Consequently, the monitoring of enzyme activity and inhibition not only yields crucial insights into health and disease conditions but also forms the basis of research in drug discovery, toxicology, and the understanding of disease mechanisms. In this context, near-infrared (NIR) fluorescent single-walled carbon nanotubes (SWCNTs) have emerged as effective tools for tracking enzyme activity and inhibition through diverse strategies. This perspective explores the physicochemical attributes of SWCNTs that render them well-suited for such monitoring. Additionally, we delve into the various strategies developed so far for successfully monitoring enzyme activity and inhibition, emphasizing the distinctive features of each principle. Furthermore, we contrast the benefits of SWCNT-based NIR probes with conventional gold standards in monitoring enzyme activity. Lastly, we highlight the current challenges faced in this field and suggest potential solutions to propel it forward. This perspective aims to contribute to the ongoing progress in biodiagnostics and seeks to engage the wider community in developing and applying enzymatic assays using SWCNTs.

Rationally Designed Functionalization of Single-Walled Carbon Nanotubes for Real-Time Monitoring of Cholinesterase Activity and Inhibition in Plasma.

Enzymes play a pivotal role in regulating numerous bodily functions. Thus, there is a growing need for developing sensors enabling real-time monitoring of enzymatic activity and inhibition. The activity and inhibition of cholinesterase (CHE) enzymes in blood plasma are fluorometrically monitored using near-infrared (NIR) fluorescent single-walled carbon nanotubes (SWCNTs) as probes, strategically functionalized with myristoylcholine (MC)- the substrate of CHE. A significant decrease in the fluorescence intensity of MC-suspended SWCNTs upon interaction with CHE is observed, attributed to the hydrolysis of the MC corona phase of the SWCNTs by CHE. Complementary measurements for quantifying choline, the product of MC hydrolysis, reveal a correlation between the fluorescence intensity decrease and the amount of released choline, rendering the SWCNTs optical sensors with real-time feedback in the NIR biologically transparent spectral range. Moreover, when synthetic and naturally abundant inhibitors inhibit the CHE enzymes present in blood plasma, no significant modulations of the MC-SWCNT fluorescence are observed, allowing effective detection of CHE inhibition. The rationally designed SWCNT sensors platform for monitoring of enzymatic activity and inhibition in clinically relevant samples is envisioned to not only advance the field of clinical diagnostics but also deepen further understanding of enzyme-related processes in complex biological fluids.

Excitation-Dependent Fluorescence with Excitation-Selective Circularly Polarized Luminescence from Hierarchically Organized Atomic Nanoclusters.

Nanometer-scaled objects are known to have dimension-related properties, but sometimes the assembly of such objects can lead to the emergence of other properties. Here, we show the assembly of atomically precise gold nanoclusters into large fibrillar structures that are featuring excitation-dependent luminescence with an excitation-selective circularly polarized luminescence (CPL), even though all components are achiral. The origin of CPL in the assembly of atomic clusters has been attributed to the hierarchical organization of atomic clusters into fibrillar structures, mediated via a hydrogen bonding interaction with a surfactant. We follow the assembly process both experimentally and computationally showing the advance in the structural formation along with its chiroptical electronic properties, i.e., circular dichroism (CD) and CPL. Our study here can assist in the rational design of materials featuring chiroptical properties, thus leading to a controlled CPL activity.

The Role of Surface Groups in Dictating the Chiral-Solvent-Induced Assembly of Carbon Dots into Structures Exhibiting Circularly Polarized Luminescence.

Here, the use of achiral nanoparticles and solvent-induced chirality transfer is combined for the making of large structures exhibiting chiroptical properties in the form of circularly polarized luminescence (CPL). The nanoparticles that the authors use are carbon dots (C-Dots) that are known for their bright luminescence and the ability to tune their surface moieties by using different precursors in their synthesis. Here, the result of adding the chiral solvent limonene into an aqueous solution of various C-Dots is explored, differentiated by their surface group. It is shown that only nitrogen-containing C-Dots with amine functional groups see the emergence of a CPL signal and the formation of a large fibrillar assembled structure. The various forces happening in the interface between the C-Dots and the limonene phase and the role of the amine groups in both the chirality transfer interactions and the interactions between C-Dots in the assembly process are discussed, whereas these two processes intertwine with each other. The ability to form fluorescent chiral structures exhibiting CPL from achiral nanoparticles and the understanding of the various interactions in this process are both important to the rationale design of any supramolecular chiral assemblies.

Four orders-of-magnitude enhancement in the two-photon excited photoluminescence of homoleptic gold thiolate nanoclusters following zinc ion-induced aggregation.

Herein we report unprecedented determination of the molar mass of zinc mediated assemblies of homoleptic gold nanoclusters, based on charge detection mass spectrometry measurements. The accurate determination of the molar mass of zinc coordinated assemblies of gold clusters has further allowed unambiguous determination of the two-photon excited photoluminescence cross section of the same. Furthermore, in line with one-photon excited photoluminescence measurements, four orders-of-magnitude enhancement in two-photon excited photoluminescence of gold nanoclusters has been observed following complexation with zinc ions. The study reported herein is envisioned to not only deepen the fundamental understanding of the multiphoton excitation properties of atomic clusters but also widen their application potential as luminescence markers.

Controlling the Chemistry of Nanoclusters: From Atomic Precision to Controlled Assembly.

Metal nanoclusters have gained prominence in nanomaterials sciences, owing to their atomic precision, structural regularity, and unique chemical composition. Additionally, the ligands stabilizing the clusters provide great opportunities for linking the clusters in higher order dimensions, eventually leading to the formation of a repertoire of nanoarchitectures. This makes the chemistry of atomic clusters worth exploring. In this mini review, we aim to focus on the chemistry of nanoclusters. Firstly, we summarize the important strategies developed so far for the synthesis of atomic clusters. For each synthetic strategy, we highlight the chemistry governing the formation of nanoclusters. Next, we discuss the key techniques in the purification and separation of nanoclusters, as the chemical purity of clusters is deemed important for their further chemical processing. Thereafter which we provide an account of the chemical reactions of nanoclusters. Then, we summarize the chemical routes to the spatial organization of atomic clusters, highlighting the importance of assembly formation from an application point of view. Finally, we raise some fundamentally important questions with regard to the chemistry of atomic clusters, which, if addressed, may broaden the scope of research pertaining to atomic clusters.

Functionalized Au15 nanoclusters as luminescent probes for protein carbonylation detection.

Atomically precise, ligand-protected gold nanoclusters (AuNCs) attract considerable attention as contrast agents in the biosensing field. However, the control of their optical properties and functionalization of surface ligands remain challenging. Here we report a strategy to tailor AuNCs for the precise detection of protein carbonylation-a causal biomarker of ageing. We produce Au15SG13 (SG for glutathione) with atomic precision and functionalize it with a thiolated aminooxy moiety to impart protein carbonyl-binding properties. Mass spectrometry and molecular modelling reveal the key structural features of Au15SG12-Aminooxy and its reactivity towards carbonyls. Finally, we demonstrate that Au15SG12-Aminooxy detects protein carbonylation in gel-based 1D electrophoresis by one- and two-photon excited fluorescence. Importantly, to our knowledge, this is the first application of an AuNC that detects a post-translational modification as a nonlinear optical probe. The significance of post-translational modifications in life sciences may open avenues for the use of Au15SG13 and other nanoclusters as contrast agents with tailored surface functionalization and optical properties.

An ambient complexation reaction of zinc acetate and ascorbic acid leads to a new form of nanoscale particles with emergent optical properties.

We report the formation of nanoscale particles from the complexation reaction between zinc acetate and ascorbic acid under ambient conditions and in an aqueous medium. The reaction led to the formation of a molecular complex with the formula Zn x (AA) y (OAc) z (x, y, and z = possible smallest positive integer) with AA meaning ascorbate, based on the mass spectrometry results. Following this, the formation of luminescent nanoscale particles - the size of which increased with time - was observed. During 24 h of observation, the sizes increased to about 50 nm in the presence of different sizes at all times. Transmission electron microscopy results also indicated the formation of polycrystalline as well as amorphous nanoparticles in the medium. Further, the appearance of a UV absorption peak at 380 nm and photoluminescence peak at 473 nm marked the formation of the nanoparticles. The luminescence was also observed to be wavelength tuneable. FTIR and NMR spectroscopy results also supported the formation of a molecular complex with the above formula. The present work highlights the importance of emergent properties of nanoscale molecular materials for crystallization. Also, the present discovery is expected to contribute to the development of safe nanomaterials.

Second harmonic scattering from mass characterized 2D graphene oxide sheets.

In this communication, we report the second harmonic scattering from mass characterized 2D graphene oxide sheets dispersed in an aqueous suspension, in the femtosecond regime at 800 nm laser excitation. Charge-detection mass-spectrometry, performing at the single sheet level, allows for an exhaustive molar mass distribution and thus concentration for these 2D nanomaterials samples. The orientation-averaged hyperpolarizability value is (1.36 ± 0.15) × 10-25 esu as determined by the concentration-dependent harmonic scattering signal. In addition, the multi-photon excited fluorescence spectrum is characterized by a broad band in the visible range between 350 and 700 nm centered at about 500 nm.

Tailoring the luminescence of atomic clusters via ligand exchange reaction mediated post synthetic modification.

The growing prominence of atomic nanoclusters in fields of practical relevance has made modulation of their luminescent characteristics an important challenge for their future applications. Herein we report chemical reaction assisted modulation of luminescence of histidine stabilized gold nanoclusters via a ligand exchange reaction with cysteine. Upon addition of 3.8 mM cysteine, as evinced by X-ray photoelectron spectroscopy in conjunction with transmission electron microscopic analyses, histidine molecules were found to desorb from the surface of the Au NCs leading to the latter's aggregation into macroscopic units. Consequently, the luminescence of the His Au NCs underwent a large bathochromic shift from 475 nm to 500 nm with a concomitant decrease in the luminescence intensity. Thereafter, upon addition of 18 mM cysteine to a dispersion of His Au NCs, cysteine molecules by virtue of strong aurophilic interactions were found to adsorb on to the surface of the Au NCs, leading to the disaggregation of the macroscopic structures. This was accompanied by restoration of the luminescence features of the Au NCs to an emission maximum of 486 nm with partial recovery of the luminescence intensity. Thus, the work embodied herein demonstrates post-synthetic chemical reactions of nanoclusters as an effective and viable tool for tailoring the photoluminescence of atomic clusters to meet application demands.

Zinc-Ion-Induced Aggregation of Gold Clusters for Visible-Light-Excitation-Based Fluorimetric Discrimination of Geometrical Isomers.

Herein, we report discrimination of dicarboxylic acids - fumaric acid (FA) and maleic acid (MA) - exhibiting geometrical isomerism, using nanoclusters based luminescent probe having excitation under broad day light. The luminescent probe was designed via complexation reaction between zinc ions and ligands (mercaptopropioinc acid; MPA) stabilizing the gold nanoclusters. This resulted in formation of nanoaggregates exhibiting bright green luminescence upon excitation at 450 nm capable of discriminating between FA and MA upto nanomolar level. The basis of discrimination has been attributed to deprotonation of FA and MA following interaction with MPA moieties present on the surface of the nanoaggregates and being governed by the stability of the respective conjugate base of the geometrical isomers of the dicarboxylic acids. As a consequence of different extent of deprotonation of FA and MA upon interaction with the cluster aggregates, different effect on the luminescence of the aggregates was observed, thus enabling discernible fluorimetric discrimination between FA and MA under visible light excitation.

Complexation Reaction-Based Two-Dimensional Luminescent Crystalline Assembly of Atomic Clusters for Recyclable Storage of Oxygen.

In this work, we report storage of oxygen in two-dimensional (2D) crystalline nanosheets comprising luminescent gold nanoclusters (Au NCs). Complexation reaction between Au NCs (stabilized by l-phenylalanine and mercaptopropionic acid) and zinc ions led to the formation of crystalline assembly of Au NCs. The crystalline nature of the assembly of Au NCs was confirmed through transmission electron microscopy (TEM), high-resolution TEM, and selected area electron diffraction (SAED) analysis. Atomic force microscopy (AFM) analysis, in conjunction with field emission scanning electron microscopy (FESEM) analysis, confirmed the 2D nature of the assembly of the Au NCs. The 2D crystalline nanosheets formed out of reaction between Au NCs and Zn2+ were found to be of near-uniform thickness, with an average value of 3.8 ± 1.65 nm. These 2D nanosheets constituting of hierarchically organized Au NCs were further used for reversible storage of oxygen at ambient conditions of 20 °C and 20 bar pressure.

Photo induced chemical modification of surface ligands for aggregation and luminescence modulation of copper nanoclusters in the presence of oxygen.

Surface modification of nanoparticles has been a popular approach to tailor the properties of nanoparticles. Herein we report the unprecedented photo oxidation of cysteine moeties on the surface of copper nanoclusters (Cu NCs) leading to aggregation of Cu NCs, which further led to quenching of luminescence of the latter. Upon illumination of a dispersion of Cu NCs at 365 nm wavelength light, the luminescence of Cu NCs was completely quenched. Furthermore, the extent of luminescence quenching of Cu NCs upon photo illumination could be tuned by varying the area of exposure of light. Confirmation of photooxidation of cysteine molecules was made through Fourier transformed infrared (FTIR) studies, while the formation of submicron sized aggregates of Cu NCs as a result of photo oxidation of cysteine stabilizing the nanoclusters was evinced through transmission electron microscopy (TEM). The study embodied herein opens up new avenues for the tailoring of the chemical and optical properties of metal nanoclusters through chemical transformation of surface ligand moieties, which is envisioned to emerge as a powerful strategy for broadening the application potential of metal nanoclusters.

Visible Light Excitation-Induced Luminescence from Gold Nanoclusters Following Surface Ligand Complexation with Zn2+ for Daylight Sensing and Cellular Imaging.

Herein, we report a complexation reaction-mediated extended aggregation of gold nanoclusters exhibiting luminescence under visible light excitation. The complexation reaction between the carboxylate groups of mercaptopropionic acid and zinc ions induced the aggregation of gold nanoclusters, which featured bright green luminescence upon excitation with visible light of wavelength 450 nm and beyond. This luminescence of aggregated Au NCs, easily discernible with bare eyes (under broad daylight excitation), was used as a probe for luminescence-based detection of molecules based on the p Ka values of the latter. This aspect has been an unfilled dream of scientists pursuing research on the development of nanoscale sensors, as luminescence-based detection techniques offer a greater degree of accuracy and sensitivity compared to absorption-based methods, and was thus far an unexploited/untapped area by nanoscale materials. Moreover, facile imaging of mammalian cells was achieved using these aggregated clusters upon excitation with visible light. This study demonstrates the utility of luminescent nanoclusters, akin to organic dyes, as materials active under visible light excitation. Thus, the complexation reaction-based tailoring of the optical properties of nanoclusters served as an effective tool in pushing the absorption maxima of the nanoclusters from an ultraviolet to visible range, enabling the luminescence of nanoclusters under broad daylight excitation. Hence, the work embodied herein offers a unique route to widen the application potential of metal nanoclusters as sensors and bioimaging agents operating under visible light excitation.

Room-Temperature Delayed Fluorescence of Gold Nanoclusters in Zinc-Mediated Two-Dimensional Crystalline Assembly.

We report that complexation-reaction-mediated two-dimensional crystalline assembly of gold (Au14) nanoclusters (NCs) exhibits room-temperature delayed fluorescence at 605 nm, with an unprecedented long lifetime of 0.5 ms and an exceptionally high quantum yield of 19.1 ± 0.9%. Interestingly, the as-synthesized Au NCs had a very weak delayed fluorescence signal. The enhancement in delayed fluorescence of Au NCs upon formation of assembly has been attributed to the crystallization-induced structural rigidity, which restricted the nonradiative transitions and enhanced the excited-state lifetime. The attainment of crystalline organization was substantiated by electron diffraction analysis. A possible structure was established based on experimental results and computational optimizations. Atomic force microscopy revealed the formation of multilayered two-dimensional nanosheets with thickness of 2.44 ± 0.48 nm.

Crystallization-Induced Emission Enhancement of Nanoclusters and One-Step Conversion of "Nanoclusters to Nanoparticles" as the Basis for Intracellular Logic Operations.

Herein, we report the construction of intracellular logic operations using luminescent histidine stabilized gold nanoclusters (His Au NCs). The luminescence intensity of His Au NCs was found to be significantly enhanced following interaction with zinc ions, owing to "Crystallization induced emission enhancement". Further, the luminescence intensity of His Au NCs was found to be effectively quenched in presence of sulphide ions, owing to transformation of emissive His Au NCs to non-emissive gold nanoparticles. Thus, the collective and individual effects of zinc ions and sulphide ions causing significant variation in the luminescence intensity of His Au NCs, were used as input parameters for construction of intracellular logic operations such as Tri state buffer, "on-off" switch and INHIBIT gate within mammalian cells.

Crystalline assembly of gold nanoclusters for mitochondria targeted cancer theranostics.

Herein, we report the formation of a crystalline assembly of gold (Au) nanoclusters for cancer theranostics via active targeting of mitochondria. To the best of our knowledge, this is the first report of the target specific activity of an "assembly of gold nanoclusters". Au14 nanoclusters were stabilized with mercaptopropionic acid and l-tyrosine. The limited solubility of l-tyrosine in methanolic solution led to the formation of a polycrystalline assembly of Au nanoclusters via interactions between ligands (tyrosine) stabilizing the clusters. Further, complexation reaction between zinc ions and ligands stabilizing the Au nanoclusters led to the formation of a single crystalline assembly of gold nanoclusters. Transmission electron microscopic and selected area electron diffraction analyses revealed the formation of faceted crystals with a hexagonal arrangement of Au14 nanoclusters. A theoretical structure of the crystalline complex of Au nanoclusters and zinc ions has been proposed herein based on experimental observations and computational optimization. The crystalline assembly of Au nanoclusters, formed via ligand association as well as complexation reaction, exhibited mitochondria targeted anti-cancer activity - as verified by Mito Tracker staining experiments. However, the MTT assay, FACS analysis and JC-1 staining experiments revealed that the zinc mediated assembly of Au nanoclusters exhibited superior therapeutic action as compared to the methanol driven assembly of the clusters.

Zinc-Coordinated Hierarchical Organization of Ligand-Stabilized Gold Nanoclusters for Chiral Recognition and Separation.

Three-dimensional organization of d- or l-tryptophan and mercaptopropionic acid-stabilized gold nanoclusters has been achieved by complexation of ligands using zinc ions. Powder X-ray diffraction and transmission electron microscopy analyses substantiated the crystalline nature of the assembly of atomic nanoclusters. The hierarchical arrangement of the nanoclusters exhibited superior optical properties (namely, enhanced photoluminescence and excited-state lifetime) as compared to the non-assembled nanoclusters. Furthermore, photoluminescence of the crystalline assembly of nanoclusters served as a visual marker for chiral recognition of d and l enantiomers of tryptophan, with subsequent separation of the corresponding enantiomer. A theoretical structure based on various experimental observations has also been proposed herein. The mechanistic aspect of the chiral separation is proposed to have occurred through attachment of d or l-tryptophan to the coordinatively unsaturated zinc ions, thus forming super complexes. The degree of stabilization of the super complexes is dictated by a "three-point versus two-point" interaction between the enantiomers and the chiral selector.

Thumb Imprint Based Detection of Hyperbilirubinemia Using Luminescent Gold Nanoclusters.

Early and easy detection of diseases, using point-of-care and inexpensive devices, not only provides option for early treatment but also reduces the risk of propagation. Herein we report the fabrication of a robust film based luminescence indicator of bilirubin, which can indicate hyperbilirubinemia through the thumb imprint of the patient. The UV-light induced luminescence intensity of the film, made out of chitosan stabilised gold (Au) nanoclusters, which was effectively quenched in the presence of Cu2+ ions, recovered in the presence of bilirubin from skin or blood serum. Moreover, the sensitivity of detection of bilirubin was tuneable with the amount of Cu2+ added, thereby facilitating the detection of the desired concentration range of bilirubin.