Platinum Crosslinked Carbon Dot@TiO2-x p-n Junctions for Relapse-Free Sonodynamic Tumor Eradication via High-Yield ROS and GSH Depletion.

Sonodynamic therapy as a promising noninvasive modality is being developed for tumor therapy, but there is a lack of next-generation sonosensitizers that can generate full ROS at high yields and simultaneously deplete elevated levels of glutathione (GSH) in tumor cells. Semiconductor p-n junctions are engineered as high-efficacy sonosensitizers for sonodynamic tumor eradication using pyridine N-doped carbon dots (N-CDs) as a p-type semiconductor and oxygen-deficient TiO2-x nanosheets as a n-type semiconductor. The rate constants of 1 O2 and •OH generation by ultrasound-excited N-CD@TiO2-x p-n junctions are 4.3 and 4.5 times higher than those of TiO2 , respectively. A Z-scheme carrier migration mechanism in the p-n junction achieving the rapid spatial separation of the ultrasound-generated electron-hole pairs for enhanced full ROS production is proposed. GSH-cleavable, Pt-crosslinked, N-doped CD fluorescent probes to detect the presence of intracellular GSH are also constructed. A GSH-responsive, p-n junction platform (Pt/N-CD@TiO2-x ) with integrated GSH detection, GSH depletion, and enhanced sonodynamic performance is then assembled. Malignant tumors are completely eradicated without relapse via intravenous administration of low-dose Pt/N-CD@TiO2-x under ultrasound irradiation. This work substantiates the great potential of biocompatible, GSH-responsive p-n junctions as next-generation sonosensitizers via p-n junction-enhanced ROS generation and metal ion oxidation of intracellular GSH.

Hierarchical flower-like manganese oxide/polystyrene with enhanced oxidase-mimicking performance for sensitive colorimetric detection of glutathione.

Glutathione (GSH) is an important antioxidant and free radical scavenger that converts harmful toxins into harmless substances and excretes them out of the body. In this paper, 3D hierarchical flower-like nanozyme named MnO2/PS (polystyrene) was successfully prepared by template method for the first time. After the systematical studies, MnO2/PS nanozyme was evaluated to possess favorable oxidase activity and direct 3,3',5,5'-tetramethylbenzidine (TMB) catalytic ability in the near-neutral environment at room temperature. With the addition of different concentrations of GSH, oxidized TMB can be reduced to TMB with the whole process from blue to nearly colorless be observed by naked eyes. In addition, there is a good linear relationship in the range 1-50 µM and a detection limit of 0.08 µM. The method proposed can be successfully applied to the detection of reduced GSH in tablets and injections with good selectivity and high sensitivity. The analysis results exhibited good consistency with the results obtained by HPLC.

Facile synthesis of AgNPs@SNCDs nanocomposites as a fluorescent 'turn on' sensor for detection of glutathione.

The present study illustrates the facile synthesis of silver nanoparticles capped with sulfur and nitrogen co-doped carbon dots (AgNPs@SNCDs) nanocomposites and their application towards the sensitive and selective detection of glutathione (GSH) using a spectrofluorimetry method. SNCDs were synthesized using solvothermal treatment of cysteamine hydrochloride and p-phenylenediamine. The as-fabricated SNCDs were then utilized as capping and stabilizing agents for the preparation of AgNPs@SNCDs nanocomposites using wet chemistry. The size of AgNPs@SNCDs nanocomposites was characterized to be ~37.58 nm or even larger aggregates. Particularly, the quenched fluorescence of AgNPs@SNCDs nanocomposites could be significantly restored upon addition of GSH, and the colour of its solution changed to some extent. The fluorescence intensity ratio of AgNPs@SNCDs nanocomposites at ~450 nm and 550 nm was directly proportional to the GSH concentration within the ranges 8.35-66.83 µM and 66.83-200.5 µM, and the detection limit was 0.52 µM. Furthermore various common organic molecules had no obvious interference in the detection mode. The proposed nanosensor was successfully applied for GSH assay in actual water samples.

Bimetallic Oxide MnMoOX Nanorods for in Vivo Photoacoustic Imaging of GSH and Tumor-Specific Photothermal Therapy.

Accurate imaging of glutathione (GSH) in vivo is able to provide real-time visualization of physiological and pathological conditions. Herein, we successfully synthesize bimetallic oxide MnMoOX nanorods as an intelligent nanoprobe for in vivo GSH detection via photoacoustic (PA) imaging. The obtained MnMoOX nanoprobe with no near-infrared (NIR) absorption in the absence of GSH would exhibit strong GSH-responsive NIR absorbance, endowing PA imaging detection of GSH. Due to the up-regulated GSH concentration in the tumor microenvironment, our MnMoOX nanoprobe could be utilized for in vivo tumor-specific PA imaging. Moreover, MnMoOX nanorods with GSH-responsive NIR absorbance could also be employed to achieve tumor-specific photothermal therapy (PTT). Importantly, such MnMoOX nanorods show inherent biodegradability and could be rapidly cleared out from the body, minimizing their long-term body retention and potential toxicity. Our work presents a new type of GSH-responsive nanoprobe based on bimetallic oxide nanostructures, promising for tumor-specific imaging and therapy.

Enhanced fluorescence of gold nanoclusters composed of HAuCl4 and histidine by glutathione: glutathione detection and selective cancer cell imaging.

Glutathione (GSH) can significantly and selectively enhance the fluorescence intensity of Au nanoclusters (NCs) prepared by blending HAuCl4 and histidine in solution. The quantum yield of the Au NCs after adding GSH can reach above 10%. Besides, GSH capping shifts the excitation peak of Au NCs from ultraviolet (386 nm) to visible light (414 nm) and improves the stability of the Au NCs. The cytotoxicities of the Au NCs with and without GSH for normal lung cells (ATII) and cancerous lung cells (A549) are evaluated. The GSH-capped Au NCs have much less cytotoxicity to both normal and cancer cells, as compared to those without GSH. For Au NCs without GSH, less cytotoxicity is observed in cancer cells than in normal cells. In addtion, the Au NCs can selectively detect GSH over cysteine and homocysteine, the two biothiols which commonly exist in cells that can seriously affect GSH detection. Most importantly, Au NCs without GSH can selectively image the cancer cells, especially for the liver cancer cells whose GSH content is much higher than other cell types. This property makes the Au NCs a powerful probe to distinguish cancer cells from normal cells.

A Zn-modified PCN-224 fluorescent nanoprobe for selective and sensitive turn-on detection of glutathione.

Monitoring endogenous glutathione (GSH) levels in living cells is essential for cancer diagnose and treatment. In this work, GSH responsive fluorescent nanoprobe with turn-on property was constructed using Zn-modified porphyrinic metal-organic frameworks (PCN-224-Zn). The introduced Zn2+ could quench the fluorescence of PCN-224 by the metallization of organic ligand (TCPP) and serves as sensing site for GSH. When exposed to GSH, the strong binding affinity of GSH generates the formation of Zn-GSH complex, eliminating the fluorescence quenching effect of Zn2+. Based on the constructed PCN-224-Zn nanoprobe, selective determination of GSH was achieved in the range of 0.01-6 µM with a detection limit of 1.5 nM. Furthermore, the constructed nanoprobe can realize the fluorescence imaging of endogenous GSH in MCF-7 and HeLa cells. Meanwhile, PCN-224-Zn could also monitor GSH in cell lysate with recovery rates from 93.8 % to 102.3 %. The performance of PCN-224-Zn demonstrates its capacities in the application of fluorescence sensing and bio-imaging fields.

MXene/Carbon Dots Nanozyme Composites for Glutathione Detection and Tumor Therapy.

Co-N-CDs-based MXene nanocomposites (MXene@PDA/Co-N-CDs) were constructed by decorating Co-N-CDs on polydopamine-functionalized MXene nanosheets. Both Co-N-CDs and MXene nanosheets have peroxidase-like activity; when the two materials are combined to form MXene@PDA/Co-N-CDs nanocomposites, the peroxide-like activity can be further enhanced. MXene@PDA/Co-N-CDs could oxidize the substrate 3,3'5,5'-tetramethylbenziline (TMB) to form ox-TMB, as confirmed by detecting the absorption of the blue products. A highly selective colorimetric biosensor was developed for the determination of glutathione (GSH) in the concentration range of 0.3 to 20 µM with a lower detection limit (LOD) of 0.12 µM, which realized the accurate detection of GSH in human serum and urine samples. Moreover, in the tumor microenvironment, MXene@PDA/Co-N-CDs could catalyze hydrogen peroxide to produce hydroxyl free radicals and produce a photothermal effect under the exposure of NIR-I irradiation. The catalytic activity of MXene@PDA/Co-N-CD nanocomposites was fully achieved for the death of cancer cells through photothermal/photodynamic synergistic therapy. The MXene@PDA/Co-N-CDs nanozyme offers multiple applications in GSH detection and tumor therapy.

Multifunctional carbon dots for glutathione detection and Golgi imaging.

Glutathione (GSH) is present in almost every cell in the body and plays various integral roles in many biological processes. The Golgi apparatus is a eukaryotic organelle for the biosynthesis, intracellular distribution, and secretion of various macromolecules; however, the mechanism of GSH in the Golgi apparatus has not been fully elucidated. Here, specific and sensitive sulfur-nitrogen co-doped carbon dots (SNCDs) with orange-red fluorescence was synthesized for the detection of GSH in the Golgi apparatus. The SNCDs have a Stokes shift of 147 nm and excellent fluorescence stability, and they exhibited excellent selectivity and high sensitivity to GSH. The linear response of the SNCDs to GSH was in the range of 10-460 µM (LOD = 0.25 µΜ). More importantly, we used SNCDs with excellent optical properties and low cytotoxicity as probes, and successfully realized golgi imaging in HeLa cells and GSH detection at the same time.

The development of a novel copper-loaded mesoporous silica nanoparticle as a peroxidase mimetic for colorimetric biosensing and its application in H2O2 and GSH assay.

In recent years, the development of nanomaterials-based peroxidase mimics as enzyme sensors has been attracting considerable interest due to their outstanding features, including potent stability, and cost-effectiveness toward natural enzymes. In this work, mesoporous silica nanoparticles functionalized by copper (Cu-MSN) were prepared as a new artificial enzyme for the first time through the sol-gel procedure. A comprehensive investigation of the catalytic activity of Cu-MSN was done through the oxidation of chromogenic peroxidase substrates, 3,3',5,5'-tetramethylbenzidine (TMB), and (2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS), in the presence of H2O2. The results indicate that the peroxidase-like activity of the as-prepared sample is significantly higher than other nanoparticles. Additionally, for the study, a facile and rapid sensing method based on the enzyme-like activity of Cu-MSN to detect H2O2 and glutathione (GSH) was developed to examine the potency of the proposed biosensor. Preliminary analysis revealed that the limit of detection (LOD) of H2O2 and GSH is 0.2 and 0.0126 µM, in the range of 0.9-100 and 0.042-1 µM, respectively. These findings support the claims for the efficiency of the sensor in detection fields. Also, human serum was utilized as the real sample to obtain additional evidence.

In-situ formation of MnO2 nanoparticles on Ru@SiO2 nanospheres as a fluorescent probe for sensitive and rapid detection of glutathione.

Glutathione (GSH)-switched fluorescent assays have appealed much attention due to rapid signal changes of fluorescent probes. However, exposure to exterior environment of fluorescent probe causes photobleaching and premature leakage, leading to low sensitivity and poor photostability. Herein, luminescent SiO2 nanoparticles encapsulated with Ru(bpy)32+ (Ru@SiO2) were designed and synthesized as fluorescent probe to construct a GSH-switched fluorescent assay. The encapsulation of Ru(bpy)32+ in the SiO2 nanoparticles could effectively prevent the leakage of Ru(bpy)32+ molecules, improving the photostability of probe. The fluorescence of Ru@SiO2 nanoparticles was quenched by coating MnO2 nanoparticles on Ru@SiO2 surface (Ru@SiO2@MnO2 nanocomposites) through an in situ growth approach, which reduced background of the assay. The MnO2 nanoparticles not only further inhibited the leakage of Ru(bpy)32+ molecules, but also could serve as a recognition unit of GSH. In the presence of GSH, the MnO2 nanoparticles on the surface of Ru@SiO2 nanoparticles were reduced to Mn2+, resulting the fluorescence recovery of Ru@SiO2 nanoparticles. Thus, a signal-on fluorescent strategy was constructed for GSH detection. The assay displayed good analytical performance for GSH detection with a low detection limit of 16.2 nM due to excellent fluorescence quenching ability of MnO2 nanoparticles and special role of Ru@SiO2 nanoparticles to block probe leakage. The proposed assay was also applied to measure GSH levels in human serum samples. This work paves a new way to detect GSH with high sensitivity.

Construction of Bio-Nano Interfaces on Nanozymes for Bioanalysis.

Nanomaterials with enzyme-like activity (nanozymes) have been of great interest in broad applications ranging from biosensing to biomedical applications. Despite that much effort has been devoted to the development of the synthesis and applications of nanozymes, it is essential to understand the interactions between nanozymes and most commonly used biomolecules, i.e., avidin, streptavidin (SA), bovine serum albumin (BSA), immunoglobulin G (IgG), and glutathione (GSH), yet they have been rarely explored. Here, a series of bio-nano interfaces were constructed through direct immobilization of proteins on a variety of iron oxide and carbon-based nanozymes with different dimensions, including Fe3O4 nanoparticles (NPs, 0D), Fe3O4@C NPs (0D), Fe3O4@C nanowires (NWs, 1D), and graphene oxide nanosheets (GO NSs, 2D). Such interfaces enabled the modulation of the catalytic activities of the nanozymes with varying degrees, which allowed a good identification of multiplex proteins with high accuracy. Given the maximum inhibition on Fe3O4@C NP by BSA, we established molecular switches based on aptamer and toehold DNA, as well as Boolean logic gates (AND and NOR) in response to both DNA and proteins. Also importantly, we developed an on-particle reaction strategy for colorimetric detection of GSH with ultrahigh sensitivity and good specificity. The proposed sensor achieved a broad dynamic range spanning 7 orders of magnitude with a detection limit down to 200 pg mL-1, which was better than that of an in-solution reaction-based biosensor by 2 orders of magnitude. Furthermore, we explored the mechanisms of the interactions at bio-nano interfaces by studying the interfacial factors, including surface coverage, salt concentration, and the curvature of the nanozyme. This study offered new opportunities in the elaborate design and better utilization of nanozymes for bioanalysis in clinical diagnosis and in vivo detection.

Naturally Occurring Exopolysaccharide Nanoparticles: Formation Process and Their Application in Glutathione Detection.

Naturally occurring nanoscale exopolysaccharide (EPS) has attracted much attention in recent years. In this research, we obtained a new kind of naturally occurring spherical EPS nanoparticles (EPS-R503) from Lactobacillus plantarum R503. The secretion, self-assembly process, morphological structure, and surface characteristics of the as-prepared nanoparticles were comprehensively revealed with transmission electron microscopy (TEM) and atomic force microscope (AFM) for the first time. It was found that the EPS-R503 nanoparticles consist of negatively charged heteropolysaccharide composed of mannose, glucose, galactose, and glucuronide with several functional groups including -OH, -COOH, and -NH2. When different solvents were used to treat the EPS-R503 nanoparticles, the morphological structure and surface properties could be changed or manipulated. The forming mechanism of EPS-R503 was elucidated based on the aggregation processes from a fundamental point of view. Furthermore, EPS-R503 can serve as reducing and stabilizing agents for the biosynthesis of manganese dioxide nanosheets (MnO2 NSs), leading to EPS-MnO2 nanocomposite. The as-prepared nanocomposites can absorb fluorescein (FL) to form EPS-MnO2-FL, which can be used to detect glutathione (GSH) with a low limit of detection (0.16 µM) and a wide detection range from 0.05 to 4 mM. The excellent biocompatibility of EPS-MnO2-FL endows the feasibility of in vivo detection of GSH as well. Overall, the findings from this work not only benefit the exploitation of naturally occurring EPS nanomaterials but also provide a novel strategy for the green synthesis of metal-containing nanosheets for GSH detection.

Biocompatibility nanoprobe of MXene N-Ti3C2 quantum dot/Fe3+ for detection and fluorescence imaging of glutathione in living cells.

MXene quantum dots have attracted much attention due to their great optical performance and excellent water solubility. Glutathione (GSH) plays a key role in living cells. In this study, a biocompatibility nanoprobe was prepared for detecting intracellular GSH based on MXene N-Ti3C2 quantum dots (N-Ti3C2 QDs). The N-Ti3C2 QDs act as the fluorescence reporters and the ferric iron (Fe3+) as the quenchers based on nonradiative electron-hole annihilation. When Fe3+ encounters the amino group of N-Ti3C2 QDs, the electrons of N-Ti3C2 QDs in the excited state will transfer to the half-filled 3d orbitals of Fe3+, leading to the fluorescence quenching of N-Ti3C2 QDs. When the N-Ti3C2 QDs/Fe3+ nanoprobe acts on the cancer cell MCF-7, the abundant GSH in the cancer cells can reduce Fe3+ to Fe2+, which will restore the fluorescence of N-Ti3C2 QDs. The N-Ti3C2 QDs/Fe3+ nanoprobe displays a high sensitivity for GSH with a detection limit of 0.17 µM in range of 0.5-100 µM. It becomes a promising probe for detecting and showing cellular imaging of GSH in MCF-7 cells. The N-Ti3C2 QDs/Fe3+ nanoprobe might provide a new way for imaging-guided precision cancer diagnosis.

Bioinspired tailoring of fluorogenic thiol responsive antioxidant precursors to protect cells against H2O2-induced DNA damage.

Natural antioxidants, like phenolic acids, possess a unique chemical space that can protect cellular components from oxidative stress. However, their polar carboxylic acid chemotype reduces full intracellular antioxidant potential due to limited diffusion through biological membranes. Here, we have designed and developed a new generation of hydrophobic turn-on fluorescent antioxidant precursors that upon penetration of the cell membrane, reveal a more polar and more potent antioxidant core and simultaneously become fluorescent allowing their intracellular tracking. Their activation is stimulated by polarity alteration by sensing intracellular signals and specifically biothiols. In our design, the carboxylic group of phenolic acids that originally restricts cell entrance is derivatized and conjugated through Copper (I)-catalyzed azide-alkyne cycloaddition (CuAAC) to a coumarin derivative that its fluorescence properties are quenched with a biothiol activatable element. This more hydrophobic precursor readily penetrates cell membrane and once inside the cell the antioxidant core is revealed upon sensing glutathione, its fluorescence is restored in a turn-on manner and the generation of a more polar character traps the molecule inside the cell. This turn-on fluorescent antioxidant precursor that can be applied to phenolic acids, was developed for rosmarinic acid and the conjugate was named as RCG. The selectivity and responsiveness of RCG towards the most abundant biothiols was monitored through a variety of biophysical techniques including UV-Vis, fluorescence and NMR spectroscopy. The electrochemical behavior and free radical scavenging capacity of the precursor RCG and the active compound (RC) was evaluated and compared with the parent compound (rosmarinic acid) through cyclic voltammetry and EPR spectroscopy, respectively. The stability of the newly synthesized bioactive conjugate RC was found significantly higher than the parent rosmarinic acid when exposed to oxygen. Cell uptake experiments were conducted and revealed the internalization of RCG. The degree of intracellular DNA protection offered by RCG and its active drug (RC) on exposure to H2O2 was also evaluated in Jurkat cells.

Detection of glutathione within single erythrocyte of different ages and pathological state using microfluidic chips coupled with laser induced fluorescence.

As a major factor participating in the organism antioxidation and detoxification process, GSH is of vital importance to human beings. Detecting GSH content in single cells is significant to diagnosis and prevention of many diseases. In this work, the amount of GSH within single erythrocytes was detected and analyzed via statistical analysis. All erythrocytes tested were collected from people in different ages and people of different pathological states. The correlation between GSH level, age and pathological state were investigated. Results showed that the GSH level in erythrocytes decreased with the ages of patients increased. There was little difference between the GSH level in erythrocytes from people who had chronic diseases (hyperglycemia, hyperlipidemia and hypertension) and from healthy people. However, the GSH level in erythrocytes from people who had inflammation (myocarditis, nephritis and gastritis) was generally higher than that from the healthy people. This study provides basic data for researches of cell senescence and cytopathic effect and is helpful to diagnosis and prevention of diseases. In addition, it also provides a simple and effective method for rapid GSH detection within single cell.