Acid-Triggered Degradation of Three-In-One Ag2S Quantum Dots for In Situ Ratiometric NIR-II Fluorescence Imaging-Guided Ion/Gas Combination Therapy.

Smart theranostic nanoprobes with the integration of multiple therapeutic modalities are preferred for precise diagnosis and efficient therapy of tumors. However, it remains a big challenge to arrange the imaging and two or more kinds of therapeutic agents without weakening the intended performances. In addition, most existing fluorescence (FL) imaging agents suffer from low spatiotemporal resolution due to the short emission wavelength (<900 nm). Here, novel three-in-one Ag2S quantum dot (QD)-based smart theranostic nanoprobes were proposed for in situ ratiometric NIR-II FL imaging-guided ion/gas combination therapy of tumors. Under the acidic tumor microenvironment, three-in-one Ag2S QDs underwent destructive degradation, generating toxic Ag+ and H2S. Meanwhile, their FL emission at 1270 nm was weakened. Upon introduction of a downconversion nanoparticle (DCNP) as the delivery carrier and NIR-II FL reference signal unit, the formed Ag2S QD-based theranostic nanoprobes could achieve precise diagnosis of tumors through ratiometric NIR-II FL signals. Also, the generated Ag+ and H2S enabled specific ion/gas combination therapy toward tumors. By combining the imaging and therapeutic functions, three-in-one Ag2S QDs may open a simple yet reliable avenue to design theranostic nanoprobes.

Exceptional performance of Fe@carbon-rich nanoparticles prepared via hydrothermal carbonization of oil mill wastes for H2S removal.

Carbon-encapsulated iron oxide nanoparticles (CE-nFe) have been obtained from an industrial waste (oil mill wastewater-OMW, as a carbonaceous source), and using iron sulfate as metallic precursor. In an initial step, the hydrochar obtained has been thermally activated under an inert atmosphere at three different temperatures (600 °C, 800 °C and 1000 °C). The thermal treatment promotes the development of core-shell nanoparticles, with an inner core of α-Fe/Fe3O4, surrounded by a well-defined graphite shell. Temperatures above 800 °C are needed to promote the graphitization of the carbonaceous species, a process promoted by iron nanoparticles through the dissolution, diffusion and growth of the carbon nanostructures on the outer shell. Breakthrough column tests show that CE-nFe exhibit an exceptional performance for H2S removal with a breakthrough capacity larger than 0.5-0.6 g H2S/gcatalyst after 3 days experiment. Experimental results anticipate the crucial role of humidity and oxygen in the adsorption/catalytic performance. Compared to some commercial samples, these results constitute a three-fold increase in the catalytic performance under similar experimental conditions.

Depletion and Downregulation of Hydrogen Sulfide Using an Activatable Probe for Promoting Photothermal Therapy toward Colorectal Cancers.

Since hydrogen sulfide (H2S) is an important endogenous gaseous mediator, therapeutic manipulation of H2S is promising for anticancer treatment. In this work, we develop a novel theranostic nanoplatform with H2S-specific and photocontrolled synergistic activation for imaging-guided H2S depletion and downregulation along with promoted photothermal therapy. Such a nanoplatform is fabricated by integration of a H2S-responsive molecule probe that can generate a cystathionine-ß-synthase (CBS) inhibitor AOAA and a photothermal transducer into an NIR-light-responsive container. Our nanoplatform can turn on NIR fluorescence specifically in H2S-rich cancers, guiding further laser irradiation. Furthermore, prominent conversion of photoenergy into heat guarantees special container melting with controllable AOAA release for H2S-level downregulation. This smart regulation of the endogenous H2S level amplifies the PTT therapeutic effect, successfully suppressing colorectal tumor in living mice under NIR fluorescence imaging guidance. Thus, we believe that this nanoplatform may provide a powerful tool toward H2S-concerned cancer treatment with an optimized diagnostic and therapeutic effect.

Catalytic specificity and crystal structure of cystathionine γ-lyase from Pseudomonas aeruginosa.

The escalating drug resistance among microorganisms underscores the urgent need for innovative therapeutic strategies and a comprehensive understanding of bacteria's defense mechanisms against oxidative stress and antibiotics. Among the recently discovered barriers, the endogenous production of hydrogen sulfide (H2S) via the reverse transsulfuration pathway, emerges as a noteworthy factor. In this study, we have explored the catalytic capabilities and crystal structure of cystathionine γ-lyase from Pseudomonas aeruginosa (PaCGL), a multidrug-opportunistic pathogen chiefly responsible for nosocomial infections. In addition to a canonical L-cystathionine hydrolysis, PaCGL efficiently catalyzes the production of H2S using L-cysteine and/or L-homocysteine as alternative substrates. Comparative analysis with the human enzyme and counterparts from other pathogens revealed distinct structural features within the primary enzyme cavities. Specifically, a distinctly folded entrance loop could potentially modulate the access of substrates and/or inhibitors to the catalytic site. Our findings offer significant insights into the structural evolution of CGL enzymes across different pathogens and provide novel opportunities for developing specific inhibitors targeting PaCGL.

Endogenous H2S-activated Ag nanoparticles embedded in programmed DNA-cubes for specific visualization of colorectal cancer cells.

To avoid the unexpected aggregation and reduce the cytotoxicity of nanomaterials as optical probes in cell imaging applications, we propose a programmed DNA-cube as a carrier for silver nanoparticles (Ag NPs) to construct a specific hydrogen sulfide (H2S) responsive platform (Ag NP@DNA-cube) for diagnosing colorectal cancer (CRC) in this study. The DNA-cube maintains good dispersion of Ag NPs while providing excellent biocompatibility. Based on the characteristic overexpression of endogenous H2S in CRC cells, the Ag NPs are etched by H2S within target cells into silver sulfide quantum dots, thereby selectively illuminating the target cells. The Ag NP@DNA-cube exhibits a specific fluorescence response to CRC cells and achieves satisfactory imaging.

A multifunctional fluorescent probe for sequential detection of hydrogen sulfide and pH in foodstuffs, living cells and mice.

BACKGROUND: Cancer as a leading cause of premature death worldwide has become a major threat to human health due to the high incidence and mortality. Monitoring tumor markers are reliable and significantly important for early detection of cancers. In complex biological systems, it is of great urgency but still remains challenging to conceive a fluorescent probe with multiple tumor markers detection property. Hydrogen sulfide (H2S) and pH are two target biomarkers for diagnosis of early cancer. The preparation of a novel probe with H2S and pH dual detection functions is highly anticipated. RESULTS: Herein, a novel sequential detection probe HTPQ-HS for H2S and pH has been developed. In this system, HPQ (2-(2 -hydroxyphenyl)-4(3H)-quinazolinone) structure combined with triphenylamine is applied as the fluorophore, and 2, 4-dinitrophenylsulfonyl group is used as the recognition group. In the presence of H2S, HTPQ-HS is transformed into product HTPQ-OH which shows fluorescence enhancement (29-fold) at 525 nm in less than 4 min and further displays repeatable acid-base responsive ability. HTPQ-HS is able to sequentially response to H2S and pH in living cells and does not react directly with pH. Owing to the low cytotoxicity, HTPQ-HS is able to detect exogenous and endogenous H2S in colon cancer cells and mice, monitor H2S in inflammation model and in foodstuffs. As the environment changes from acidic to alkaline, the fluorescence intensity ratio (I470/I530) of product HTPQ-OH changes remarkably, illustrating the ratiometric fluorescent responsiveness to pH. SIGNIFICANCE AND NOVELTY: A multifunctional fluorescent probe HTPQ-HS for sequential detection of H2S and pH is synthesized. Probe HTPQ-OH realizes the monitoring of dynamic changes in intracellular pH and displays prospective application in security printing. We expect that our work could offer an important guidance on the development of multifunctional fluorescent probes for visualizing H2S and pH in biology and environment.

Cell-Membrane Coated Self-Immolative Poly(thiourethane) for Cysteine/Homocysteine-Triggered Intracellular H2S Delivery.

Hydrogen sulfide (H2S) is an important gaseous signaling molecule with unique pleiotropic pharmacological effects, but may be limited for clinical translation due to the lack of a reliable delivery form that delivers exogenous H2S to cells at action site with precisely controlled dosage. Herein, we report the design of a poly(thiourethane) (PTU) self-immolative polymer terminally caged with an acrylate moiety to trigger release of H2S in response to cysteine (Cys) and homocysteine (Hcy), the most used and independent indicators of neurodegenerative diseases. The synthesized PTU polymer was then coated with the red-blood-cell (RBC) membrane in the presence of solubilizing agent to self-assemble into nanoparticles with enhanced stability and cytocompatibility. The Hcy/Cys mediated addition/cyclization chemistry actuated the biomimetic polymeric nanoparticles to disintegrate into carbonyl sulfide (COS), and finally convert into H2S via the ubiquitous carbonic anhydrase (CA). H2S released in a controlled manner exhibited a strong antioxidant ability to resist Alzheimer's disease (AD)-related oxidative stress factors in BV-2 cells, a neurodegenerative disease model in vitro. Thus, this work may provide an effective strategy to construct H2S donors that can degrade in response to a specific pathological microenvironment for the treatment of neurodegenerative diseases.

Dual-Stimuli-Activatable Hybrid Prodrug for the Self-Immolative Delivery of an Anticancer Agent and Hydrogen Sulfide with Turn-On Fluorescence.

Stimuli-responsive fluorogenic prodrugs are advantageous for the targeted drug delivery enabling real-time non-invasive monitoring with turn-on fluorescence. We report herein the dual-stimuli (ROS and CA)-responsive thiocarbamate-based prodrug (AM-TCB) for the turn-on fluorogenic delivery of the naphthalimide-based anticancer agent amonafide along with the gasotransmitter hydrogen sulfide (H2 S). A carbamate-based prodrug AM-CB was also designed, capable of releasing the anticancer agent amonafide without any H2 S. The prodrugs were synthesized using multi-step organic synthesis. UV-Vis and fluorescence spectroscopic studies revealed selective reactivity of the boronate ester group of prodrugs towards ROS (primarily H2 O2 ) with the release of amonafide and COS/CO2 via self-immolative processes. Hydrolysis of the generated COS by carbonic anhydrase (CA) produces H2 S. While the prodrug AM-TCB retained the anticancer activity of free amonafide in cancer cells (MDA-MB-231 and HeLa), unlike amonafide, it enhanced the cellular viability of the non-malignant cells (HEK-293). Fluorescence imaging in HeLa cells revealed the simultaneous delivery of the anticancer agent and H2 S from AM-TCB with turn-on fluorescence. Western blot studies further revealed the cytoprotective effects of the released H2 S from AM-TCB. The present adjuvant strategy therefore would be helpful in future for ameliorating the anticancer drug-induced side-effects.

Redox and Nucleophilic Reactions of Naphthoquinones with Small Thiols and Their Effects on Oxidization of H2S to Inorganic and Organic Hydropolysulfides and Thiosulfate.

Naphthoquinone (1,4-NQ) and its derivatives (NQs, juglone, plumbagin, 2-methoxy-1,4-NQ, and menadione) have a variety of therapeutic applications, many of which are attributed to redox cycling and the production of reactive oxygen species (ROS). We previously demonstrated that NQs also oxidize hydrogen sulfide (H2S) to reactive sulfur species (RSS), potentially conveying identical benefits. Here we use RSS-specific fluorophores, mass spectroscopy, EPR and UV-Vis spectrometry, and oxygen-sensitive optodes to examine the effects of thiols and thiol-NQ adducts on H2S-NQ reactions. In the presence of glutathione (GSH) and cysteine (Cys), 1,4-NQ oxidizes H2S to both inorganic and organic hydroper-/hydropolysulfides (R2Sn, R=H, Cys, GSH; n = 2-4) and organic sulfoxides (GSnOH, n = 1, 2). These reactions reduce NQs and consume oxygen via a semiquinone intermediate. NQs are also reduced as they form adducts with GSH, Cys, protein thiols, and amines. Thiol, but not amine, adducts may increase or decrease H2S oxidation in reactions that are both NQ- and thiol-specific. Amine adducts also inhibit the formation of thiol adducts. These results suggest that NQs may react with endogenous thiols, including GSH, Cys, and protein Cys, and that these adducts may affect both thiol reactions as well as RSS production from H2S.

The biological functions of protein S-sulfhydration in eukaryotes and the ever-increasing understanding of its effects on bacteria.

As a critical endogenous signaling molecule, hydrogen sulfide may induce reversible post-translational modifications on cysteine residues of proteins, generating a persulfide bond known as S-sulfhydration. A systemic overview of the biofunctions of S-sulfhydration will equip us better to characterize its regulatory roles in antioxidant defense, inflammatory response, and cell fate, as well as its pathological mechanisms related to cardiovascular, neurological, and multiple organ diseases, etc. Nevertheless, the understanding of S-sulfhydration is mostly built on mammalian cells and animal models. We subsequently summarized the mediation effects of this specific post-transcriptional modification on physiological processes and virulence in bacteria. The high-sensitivity and high-throughput detection technologies are required for studying the signal transduction mechanism of H2S and protein S-sulfhydration modification. Herein, we reviewed the establishment and development of different approaches to assess S-sulfhydration, including the biotin-switch method, modified biotin-switch method, alkylation-based cysteine-labelled assay, and Tag-switch method. Finally, we discussed the limitations of the impacts of S-sulfhydration in pathogens-host interactions and envisaged the challenges to design drugs and antibiotics targeting the S-sulfhydrated proteins in the host or pathogens.

Adsorption of hydrogen sulfide by iron-based adsorbent derived from fly ash and iron slag.

In this article, an innovative sorbent (Fe-FA) is prepared from fly ash; ferrous sulfate-containing waste slag (FSS), which are industrial wastes; and NaOH by a hydrothermal method at 100 °C. As a result, in comparison to several conventional sorbents, such as ZnO, Fe2O3, 13X zeolite, and activated carbon, Fe-FA had the best adsorption performance for H2S adsorption. Fe-FA had not only a higher adsorption capacity (near 150 mg/g) but also a longer breakthrough time (near 400 min) when gas hourly space velocity was 8000 h-1. Then, characterizations of XRD, BET, NH3-TPD, FTIR, and XPS analyzed basic properties of Fe-FA and revealed reasons for the excellent adsorption performance. In general, the excellent adsorption performance of Fe-FA for H2S is mainly due to the high content of iron species (almost 50%) and suitable mesoporous structure in the Fe-FA.

Orally Administrable H2 S-Scavenging Metal-Organic Framework Prepared by Co-Flow Microfluidics for Comprehensive Restoration of Intestinal Milieu.

Intestinal milieu disorders are strongly related to the occurrence of inflammatory bowel diseases (IBDs), which results from mucosa destruction, epithelium disruption, and tight junction (TJ) proteins loss. Excess of H2 S in the intestinal milieu produced by the sulfate-reducing bacteria metabolism contributes to development of IBDs via epithelial barrier breakdown. Conventional interventions, such as surgery and anti-inflammatory medications, are considered not completely effective because of frequent recurrence and other complications. Herein, a novel oral delivery system, a hydroxypropyl methylcellulose acetate succinate (HPMCAS)-based polymer-coated Zr-based metal-organic framework (UiO-66) with a Cux -rhodamine B (CR) probe (hereinafter referred to as HUR), is produced via a co-flow microfluidic approach with the ability to reduce H2 S levels, thus restoring the intestinal lumen milieu. HPMCAS serves as an enteric coating that exposes UiO-66@CR at the pH of the intestine but not the acidic pH of the stomach. The synthesized HUR exhibits notable therapeutic efficacy, including mucosa recovery, epithelium integrity restoration, and TJ proteins upregulation via H2 S scavenging to protect against intestinal barrier damage and microbiome dysbiosis. Thus, HUR is verified to be a promising theranostic platform able to decrease the H2 S content for intestinal milieu disorder treatment. The presented study therefore opens the door for further exploitation for IBDs therapy.

Hydrogen Sulfide (H2S): As a Potent Modulator and Therapeutic Prodrug in Cancer.

Hydrogen sulfide (H2S) is an endogenous gaseous molecule present in all living organisms that has been traditionally studied for its toxicity. Interestingly, increased understanding of H2S effects in organ physiology has recently shown its relevance as a signalling molecule, with potentially important implications in variety of clinical disorders, including cancer. H2S is primarily produced in mammalian cells under various enzymatic pathways are target of intense research biological mechanisms, and therapeutic effects of H2S. Herein, we describe the physiological and biochemical properties of H2S, the enzymatic pathways leading to its endogenous production and its catabolic routes. In addition, we discuss the role of currently known H2S-releasing agents, or H2S donors, including their potential as therapeutic tools. Then we illustrate the mechanisms known to support the pleiotropic effects of H2S, with a particular focus on persulfhydration, which plays a key role in H2S-mediating signalling pathways. We then address the paradoxical role played by H2S in tumour biology and discuss the potential of exploiting H2S levels as novel cancer biomarkers and diagnostic tools. Finally, we describe the most recent preclinical applications focused on assessing the anti-cancer impact of most common H2S-releasing compounds. While the evidence in favour of H2S as an alternative cancer therapy in the field of translational medicine is yet to be clearly provided, application of H2S is emerging as a potent anticancer therapy in preclinical trails.

On-demand therapeutic delivery of hydrogen sulfide aided by biomolecules.

Hydrogen sulfide (H2S), known as the third gasotransmitter, exerts various physiological functions including cardiac protection, angiogenesis, anti-inflammatory, and anti-cancer capability. Given its promising therapeutic potential as well as severe perniciousness if improper use, the sustained and tunable H2S delivery systems are highly required for H2S-based gas therapy with enhanced bioactivity and reduced side effects. To this end, a series of stimuli-responsive compounds capable of releasing H2S (termed H2S donors) have been designed over the past two decades to mimic the endogenous generation of H2S and elucidate the biological functions. Further to improve the stability of H2S donors and achieve the targeted delivery, various delivery systems have been constructed. In this review, we focus on the recent advances of an emerging subset, biomolecular-based H2S delivery systems, which combine H2S donors with biomolecular vectors including polysaccharide, peptide, and protein. We demonstrated their basic structures, building strategies, and therapeutic applications respectively to unfold their inherent merits endued by biomolecules including biocompatibility, biodegradability as well as expansibility. The varied development potentials of biomolecular-based H2S delivery systems based on their specific properties are also discussed. At the end, brief future outlooks and upcoming challenges are presented as well.

Extract of Acanthopanax senticosus and Its Components Interacting with Sulfide, Cysteine and Glutathione Increase Their Antioxidant Potencies and Inhibit Polysulfide-Induced Cleavage of Plasmid DNA.

Aqueous root extract from Acanthopanax senticosus (ASRE) has a wide range of medicinal effects. The present work was aimed at studying the influence of sulfide, cysteine and glutathione on the antioxidant properties of ASRE and some of its selected phytochemical components. Reduction of the 2-(4-carboxyphenyl)-4,5-dihydro-4,4,5,5-tetramethyl-1H-imidazol-1-yloxy-3-oxide (●cPTIO) stable radical and plasmid DNA (pDNA) cleavage in vitro assays were used to evaluate antioxidant and DNA-damaging properties of ASRE and its individual components. We found that the interaction of ASRE and its two components, caffeic acid and chlorogenic acid (but not protocatechuic acid and eleutheroside B or E), with H2S/HS-, cysteine or glutathione significantly increased the reduction of the ●cPTIO radical. In contrast, the potency of ASRE and its selected components was not affected by Na2S4, oxidized glutathione, cystine or methionine, indicating that the thiol group is a prerequisite for the promotion of the antioxidant effects. ASRE interacting with H2S/HS- or cysteine displayed a bell-shaped effect in the pDNA cleavage assay. However, ASRE and its components inhibited pDNA cleavage induced by polysulfides. In conclusion, we suggest that cysteine, glutathione and H2S/HS- increase antioxidant properties of ASRE and that changes of their concentrations and the thiol/disulfide ratio can influence the resulting biological effects of ASRE.

Fluorescent detection of hydrogen sulfide (H2S) through the formation of pyrene excimers enhances H2S quantification in biochemical systems.

Hydrogen sulfide (H2S) is produced endogenously by several enzymatic pathways and modulates physiological functions in mammals. Quantification of H2S in biochemical systems remains challenging because of the presence of interferents with similar reactivity, particularly thiols. Herein, we present a new quantification method based on the formation of pyrene excimers in solution. We synthesized the probe 2-(maleimido)ethyl 4-pyrenylbutanoate (MEPB) and determined that MEPB reacted with H2S in a two-step reaction to yield the thioether-linked dimer (MEPB)2S, which formed excimers upon excitation, with a broad peak of fluorescence emission centered at 480 nm. In contrast, we found that the products formed with thiols showed peaks at 378 and 398 nm. The difference in emission between the products prevented the interference. Furthermore, we showed that the excimer fluorescence signal yielded a linear response to H2S, with a limit of detection of 54 nM in a fluorometer. Our quantification method with MEPB was successfully applied to follow the reaction of H2S with glutathione disulfide and to quantify the production of H2S from cysteine by Escherichia coli. In conclusion, this method represents an addition to the toolkit of biochemists to quantify H2S specifically and sensitively in biochemical systems.

Hydrogen sulfide alleviates chilling injury in peach fruit by maintaining cell structure integrity via regulating endogenous H2S, antioxidant and cell wall metabolisms.

Effects of hydrogen sulfide (H2S) on chilling injury (CI), H2S, antioxidant and cell-wall metabolisms of refrigerated peaches treated with H2S and hypotaurine (HT, H2S scavenger) were investigated in present study. Results revealed that H2S treatment enhanced endogenous H2S content, which was associated with increased related H2S synthase enzymes activities, while HT showed the opposite results. Moreover, H2S treatment induced the accumulation of ascorbic acid, glutathione and the enhancement of antioxidant enzymes activities compared to control and HT, contributing to lower hydrogen peroxide content and superoxide radical production. Furthermore, H2S suppressed the increase of cell-wall degradation enzymes accompanied by higher levels of water-insoluble pectin, 24% KOH-soluble hemicellulose and cellulose, while HT accelerated these components degradation. Therefore, results indicated that H2S mitigated CI of refrigerated peaches by regulating H2S, antioxidant and cell-wall metabolisms, maintaining higher H2S and antioxidants contents, suppressing cell-wall degradation, thereby contributing to redox homeostasis maintenance and cell structure integrity.

Hydrogen sulphide-triggered theranostic prodrugs based on the dynamic chemistry of tetrazines.

Dynamic nucleophilic aromatic substitution of tetrazines (SNTz) has been employed to build theranostic prodrugs that are activated by hydrogen sulfide. H2S is typically found in high concentrations in some kinds of cancer cells and it is able to trigger the disassembly of tetrazine prodrugs. In such a way, a dual release of drugs and/or fluorescent compounds can be selectively triggered.

Thioredoxin reductase-triggered fluorogenic donor of hydrogen sulfide: a model study with a symmetrical organopolysulfide probe with turn-on near-infrared fluorescent emission.

We describe herein the rational development of an organopolysulfide-based fluorogenic donor of hydrogen sulfide (H2S) DCI-PS, which can be activated by the antioxidant selenoenzyme thioredoxin reductase (TrxR) with concomitant release of the dicyanoisophorone-based near-infrared (NIR) fluorophore. Along with the polysulfide probe DCI-PS capable of releasing the NIR fluorophore and H2S, the corresponding disulfide-probe DCI-DS was also rationally designed and synthesized, which releases the fluorophore without donating H2S. Detailed spectroscopic and kinetic studies in an aqueous medium revealed significantly higher reactivity of the probes towards DTT (for TrxR activity) over the well-known cellular abundant biothiol GSH. Mechanistically, the nucleophilic attack at the disulfide/polysulfide linkage by the thiol/selenol group of the bio-analytes leads to the self-immolative cyclization process with the release of the turn-on fluorophore with/without H2S. Considering the overexpression of mammalian TrxR in cancer cells, the turn-on fluorogenic H2S donation process from the cellular non-toxic DCI-PS was validated in a representative breast cancer cell line (MDA-MB-231) for the sustained donation of H2S with concomitant release of the red-emitting NIR fluorophore. The TrxR-triggered fluorescence turn-on process in DCI-PS was further supported by the significant inhibition of the fluorogenic process in the presence of TrxR-selective small-molecule inhibitors and by the significant binding affinity predicted by the protein-ligand docking study. Results with the antioxidant enzyme-triggered intracellular sustained donation of H2S with concomitant fluorescence turn-on will certainly find wider biomedical applications in the near future, particularly in H2S-mediated therapeutics in disease states.

Cysteine-Activated Small-Molecule H2Se Donors Inspired by Synthetic H2S Donors.

The importance of selenium (Se) in biology and health has become increasingly clear. Hydrogen selenide (H2Se), the biologically available and active form of Se, is suggested to be an emerging nitric oxide (NO)-like signaling molecule. Nevertheless, the research on H2Se chemical biology has technique difficulties due to the lack of well-characterized and controllable H2Se donors under physiological conditions, as well as a robust assay for direct H2Se quantification. Motivated by these needs, here, we demonstrate that selenocyclopropenones and selenoamides are tunable donor motifs that release H2Se upon reaction with cysteine (Cys) at pH 7.4 and that structural modifications enable the rate of Cys-mediated H2Se release to be tuned. We monitored the reaction pathways for the H2Se release and confirmed H2Se generation qualitatively using different methods. We further developed a quantitative assay for direct H2Se trapping and quantitation in an aqueous solution, which should also be operative for investigating future H2Se donor motifs. In addition, we demonstrate that arylselenoamide has the capability of Cys-mediated H2Se release in cellular environments. Importantly, mechanistic investigations and density functional theory (DFT) calculations illustrate the plausible pathways of Cys-activated H2Se release from arylselenoamides in detail, which may help understand the mechanistic issues of the H2S release from pharmacologically important arylthioamides. We anticipate that the well-defined chemistries of Cys-activated H2Se donor motifs will be useful for studying Se biology and for development of new H2Se donors and bioconjugate techniques.