Immuno field-effect transistor (ImmunoFET) for detection of salivary cortisol using potentiometric and impedance spectroscopy for monitoring heart failure.

Cortisol, a steroid hormone mostly known as "the stress hormone," plays many essential functions in humans due its involvement in several metabolic pathways. It is well-known that cortisol dysregulation is implied in evolution and progression of several chronic pathologies, including cardiac diseases such as heart failure (HF). However, although several sensors have been proposed to date for the determination of cortisol, none of them has been designed for its determination in saliva in order to monitor HF progression. In this work, a silicon nitride based Immuno field-effect transistor (ImmunoFET) has been proposed to quantify salivary cortisol for HF monitoring. Sensitive biological element was represented by anti-cortisol antibody bound onto the ISFET gate via 11-triethoxysilyl undecanal (TESUD) by vapor-phase method. Potentiometric and electrochemical impedance spectroscopy (EIS) measurements were carried out for preliminary investigations on device responsiveness. Subsequently, a more sensitive detection was obtained using electrochemical EIS. The proposed device has proven to have a linear response (R2 always >0.99), to be sensitive (with a limit of detection, LoD, of 0.005 ± 0.002 ng/mL), selective in case of other HF biomarkers (e.g. N-terminal pro B-type natriuretic peptide (NT-proBNP), tumor necrosis factor-alpha (TNF-α), and interleukin 10 (IL-10)), and accurate in cortisol quantification in saliva sample by performing the standard addition method.

A novel electrochemical strategy for NT-proBNP detection using IMFET for monitoring heart failure by saliva analysis.

Heart failure (HF) is a chronic cardiovascular disease that represents main cause of mortality worldwide, particularly for elderly. N-terminal pro-brain natriuretic peptide (NT-proBNP) was identified as the gold standard biomarker for HF diagnosis and therapy monitoring. Presently, saliva analysis represents an emerging and powerful tool for clinical applications and electrochemical immunosensors have shown their potential in Healthcare applications as selective and reliable systems for detecting clinical biomarkers. This work presents the detection of NT-proBNP in saliva samples by an immunologically modified Field effect Transistor (IMFET). TESUD ((11-triethoxysilyl) undecanal) was used as cross-linker to immobilise anti-NT-proBNP antibody onto the device. Our IMFET that was then tested in different matrices (e.g. phosphate buffered saline (PBS), artificial saliva and human saliva) using electrochemical impedance spectroscopy (EIS), and it resulted selective to NT-proBNP with good sensitivity (detection limit of 0.02 pg/mL) and a wide linear range (0.02-1 pg/mL and 0.5-20 pg/mL). Finally, NT-proBNP concentration in ten saliva samples was determined by performing the standard addition method. An enzyme-linked immunosorbent assay was used for confirming IMFET results, highlighting both IMFET accuracy (analyte recovery of 99 ± 8%) and precision (coefficient of variation always <10%), and supporting the suitability of the device for determining salivary NT-proBNP.

Spatially hierarchical nano-architecture for real time detection of Interleukin-8 cancer biomarker.

In the present work we have developed two hierarchical nano-architectures based electrochemical immunosensors for the detection of interleukin-8 (IL-8) cytokine tumor biomarker. A comparative study has been performed for spatial nano-architectures and their relative sensing to establish the model for real time monitoring. With the first platform, the recognition layer consisted with immobilised IL-8 on aminothiol modified gold electrodes. In the second approach, the activated multi walled carbon nanotubes (MWCNT-COOH) were added in the functionalisation process by covalent attachment between the functionalities NH2 of aminothiol and the functionalities COOH of carbon nanotubes. The surface topology of the recognition layer has been characterised by atomic force spectroscopy (AFM) and contact angle (CA) measurements. The electrochemical response of the developed sensor was measured by electrochemical impedance spectroscopy (EIS). A side-by-side comparison showed that aminothiol/activated MWCNTs/anti-IL-8 based impedimetric immunosensor exhibits high reproducibility (The relative standard deviation (R.S.D) = 3.2%, n = 3) with high stability. The present sensor allows evaluating a lower detection limit of 0.1 pg mL-1 with a large dynamic sensitivity range from 1 pg mL-1to 1000 pg mL-1 covering the entire clinical therapeutic window. The developed MWCNTs based immunosensor has been calibrated by determining IL-8 in artificial plasma and showed a selective response to IL-8 even in the interfering environment of other cytokines such as Interleukin-1 (IL-1) and Interleukin-6 (IL-6).

Determination and stability of N-terminal pro-brain natriuretic peptide in saliva samples for monitoring heart failure.

Heart failure (HF) is the main cause of mortality worldwide, particularly in the elderly. N-terminal pro-brain natriuretic peptide (NT-proBNP) is the gold standard biomarker for HF diagnosis and therapy monitoring. It is determined in blood samples by the immunochemical methods generally adopted by most laboratories. Saliva analysis is a powerful tool for clinical applications, mainly due to its non-invasive and less risky sampling. This study describes a validated analytical procedure for NT-proBNP determination in saliva samples using a commercial Enzyme-Linked Immuno-Sorbent Assay. Linearity, matrix effect, sensitivity, recovery and assay-precision were evaluated. The analytical approach showed a linear behaviour of the signal throughout the concentrations tested, with a minimum detectable dose of 1 pg/mL, a satisfactory NT-proBNP recovery (95-110%), and acceptable precision (coefficient of variation ≤ 10%). Short-term (3 weeks) and long-term (5 months) stability of NT-proBNP in saliva samples under the storage conditions most frequently used in clinical laboratories (4, - 20, and - 80 °C) was also investigated and showed that the optimal storage conditions were at - 20 °C for up to 2.5 months. Finally, the method was tested for the determination of NT-proBNP in saliva samples collected from ten hospitalized acute HF patients. Preliminary results indicate a decrease in NT-proBNP in saliva from admission to discharge, thus suggesting that this procedure is an effective saliva-based point-of-care device for HF monitoring.

Salivary Biomarkers for Diagnosis and Therapy Monitoring in Patients with Heart Failure. A Systematic Review.

The aim of this study was to perform a systematic review on the potential value of saliva biomarkers in the diagnosis, management and prognosis of heart failure (HF). The correlation between saliva and plasma values of these biomarkers was also studied. PubMed was searched to collect relevant literature, i.e., case-control, cross-sectional studies that either compared the values of salivary biomarkers among healthy subjects and HF patients, or investigated their role in risk stratification and prognosis in HF patients. No randomized control trials were included. The search ended on 31st of December 2020. A total of 15 studies met the inclusion criteria. 18 salivary biomarkers were analyzed and the levels of all biomarkers studied were found to be higher in HF patients compared to controls, except for amylase, sodium, and chloride that had smaller saliva concentrations in HF patients. Natriuretic peptides are the most commonly used plasma biomarkers in the management of HF. Their saliva levels show promising results, although the correlation of saliva to plasma values is weakened in higher plasma values. In most of the publications, differences in biomarker levels between HF patients and controls were found to be statistically significant. Due to the small number of patients included, larger studies need to be conducted in order to facilitate the use of saliva biomarkers in clinical practice.

Sport in Town: The Smart Healthy ENV Project, a Pilot Study of Physical Activity with Multiparametric Monitoring.

BACKGROUND: Increasing evidence links meteorological characteristics and air pollution to physiological responses during sports activities in urban areas with different traffic levels. OBJECTIVE: The main objective of the Smart Healthy ENV (SHE, "Smart Monitoring Integrated System For A Healthy Urban Environment In Smart Cities") project was to identify the specific responses of a group of volunteers during physical activity, by monitoring their heart rates and collecting breath samples, combined with data on meteorological determinants and pollution substances obtained through fixed sensor nodes placed along city routes and remotely connected to a dedicated data acquisition server. METHODS: Monitoring stations were placed along two urban routes in Pisa, each two km long, with one located within the park beside the Arno river (green route) and the other in a crowded traffic zone (red route). Our sample participants were engaged in sports activities (N = 15, with different levels of ability) and were monitored through wearable sensors. They were first asked to walk back and forth (4 km) and then to run the same route. The experimental sessions were conducted over one day per route. A breath sample was also collected before each test. A questionnaire concerning temperature and fatigue perception was administered for all of the steps of the study over the two days. RESULTS: The heart rates of the participants were monitored in the baseline condition, during walking, and while running, and were correlated with meteorological and pollutant data and with breath composition. Changes in the heart rates and breath composition were detected during the experimental sessions. These variations were related to the physical activity and to the meteorological conditions and air pollution levels. CONCLUSIONS: The SHE project can be considered a proof-of-concept study aimed at monitoring physiological and environmental variables during physical activity in urban areas, and can be used in future studies to provide useful information to those involved in sports and the broader community.

A silicon nitride ISFET based immunosensor for tumor necrosis factor-alpha detection in saliva. A promising tool for heart failure monitoring.

According to the European statistics, approximately 26 million patients worldwide suffer from heart failure (HF), and this number seems to be steadily increasing. Inflammation plays a central role in the development of HF, and the pro-inflammatory cytokine Tumor necrosis factor-α (TNF-α) represents inflammation gold-standard biomarker. Early detection plays a crucial role for the prognosis and treatment of HF. An Ion Sensitive Field Effect Transistor (ISFET) based on silicon nitride transducer and biofunctionalized with anti-TNF-α antibody for label-free detection of salivary TNF-α is proposed. Electrochemical impedance spectroscopy (EIS) was used for TNF-α detection. Our ImmunoFET offered a detection limit of 1 pg mL-1, with an analytical reproducibility expressed by a coefficient of variance (CV) resulted < 10% for the analysis of saliva samples, and an analyte recovery of 94 ± 6%. In addition, it demonstrated high selectivity when compared to other HF biomarkers such as Inteleukin-10, N-terminal pro B-type natriuretic peptide, and Cortisol. Finally, ImmunoFET accuracy in determining the unknown concentration of TNF-α was successfully tested in saliva samples by performing standard addition method. The proposed ImmunoFET showed great promise as a complementary tool for biomedical application for HF monitoring by a non-invasive, rapid and accurate assessment of TNF-α.

Determination of peppermint compounds in breath by needle trap micro-extraction coupled with gas chromatography-tandem mass spectrometry.

Breath analysis is an alternative approach for disease diagnosis and for monitoring therapy. The lack of standardized procedures for collecting and analysing breath samples currently limits its use in clinical practice. In order to overcome this limitation, the 'Peppermint Consortium' was established within the breath community to carry out breath wash-out experiments and define reference values for a panel of compounds contained in the peppermint oil capsule. Here, we present a needle trap micro-extraction technique coupled with gas chromatography and tandem mass spectrometry for a rapid and accurate determination of alpha-pinene, beta-pinene, limonene, eucalyptol, menthofuran, menthone, menthol and menthyl acetate in mixed breath samples. Detection limits between 1 and 20 pptv were observed when 25 ml of a humidified standard gas mixture were loaded into a needle trap device at a flow rate of 10 ml min-1. Inter- and intra-day precisions were lower than 15%, thus confirming the reliability of the assay. Our procedure was used to analyse breath samples taken from a nominally healthy volunteer who was invited to swallow a 200 mg capsule of peppermint oil. Six samples were collected at various times within 6 h of ingestion. Analyte concentrations were not affected by the sampling mode (i.e. mixed vs. end-tidal fraction), whereas respiratory rate and exhalation flow rate values slightly influenced the concentration of the target compounds in breath samples.

Biosensors for Detecting Lymphocytes and Immunoglobulins.

Lymphocytes (B, T and natural killer cells) and immunoglobulins are essential for the adaptive immune response against external pathogens. Flow cytometry and enzyme-linked immunosorbent (ELISA) kits are the gold standards to detect immunoglobulins, B cells and T cells, whereas the impedance measurement is the most used technique for natural killer cells. For point-of-care, fast and low-cost devices, biosensors could be suitable for the reliable, stable and reproducible detection of immunoglobulins and lymphocytes. In the literature, such biosensors are commonly fabricated using antibodies, aptamers, proteins and nanomaterials, whereas electrochemical, optical and piezoelectric techniques are used for detection. This review describes how these measurement techniques and transducers can be used to fabricate biosensors for detecting lymphocytes and the total content of immunoglobulins. The various methods and configurations are reported, along with the advantages and current limitations.

Saliva as a non-invasive tool for monitoring oxidative stress in swimmers athletes performing a VO2max cycle ergometer test.

Biomarkers of oxidative stress are generally measured in blood and its derivatives. However, the invasiveness of blood collection makes the monitoring of such chemicals during exercise not feasible. Saliva analysis is an interesting approach in sport medicine because the collection procedure is easy-to-use and does not require specially-trained personnel. These features guarantee the collection of multiple samples from the same subject in a short span of time, thus allowing the monitoring of the subject before, during and after physical tests, training or competitions. The aim of this work was to evaluate the possibility of following the changes in the concentration of some oxidative stress markers in saliva samples taken over time by athletes under exercise. To this purpose, ketones (i.e. acetone, 2-butanone and 2-pentanone), aldehydes (i.e. propanal, butanal, and hexanal), α,ß-unsaturated aldehydes (i.e. acrolein and methacrolein) and di-carbonyls (i.e. glyoxal and methylglyoxal) were derivatized with 2,4-dinitrophenylhydrazine, and determined by ultra-high performance liquid chromatography coupled to diode array detector. Prostaglandin E2, F2/E2-isoprostanes, F2-dihomo-isoprostanes, F4-neuroprostanes, and F2-dihomo-isofuranes were also determined by a reliable analytical procedure that combines micro-extraction by packed sorbent and ultra-high performance liquid chromatography-electrospray ionization tandem mass spectrometry. Overall the validation process showed that the methods have limits of detection in the range of units of ppb for carbonyls and tens to hundreds of ppt for isoprostanes and prostanoids, very good quantitative recoveries (90-110%) and intra- and inter-day precision lower than 15%. The proof of applicability of the proposed analytical approach was investigated by monitoring the selected markers of oxidative stress in ten swimmers performing a VO2max cycle ergo meter test. The results highlighted a clear increase of salivary by-products of oxidative stress during exercise, whereas a sharp decrease, approaching baseline values, of these compounds was observed in the recovery phase. This study opens up a new approach in the evaluation of oxidative stress and its relation to aerobic activity.

Salivary lactate and 8-isoprostaglandin F2α as potential non-invasive biomarkers for monitoring heart failure: a pilot study.

Heart failure (HF) is a cardiovascular disease affecting about 26 million people worldwide costing about $100 billons per year. HF activates several compensatory mechanisms and neurohormonal systems, so we hypothesized that the concomitant monitoring of a panel of potential biomarkers related to such conditions might help predicting HF evolution. Saliva analysis by point-of-care devices is expected to become an innovative and powerful monitoring approach since the chemical composition of saliva mirrors that of blood. The aims of this study were (i) to develop an innovative procedure combining MEPS with UHPLC-MS/MS for the simultaneous determination of 8-isoprostaglandin F2α and cortisol in saliva and (ii) to monitor lactate, uric acid, TNF-α, cortisol, α-amylase and 8-isoprostaglandin F2α concentrations in stimulated saliva samples collected from 44 HF patients during their hospitalisation due to acute HF. Limit of detection of 10 pg/mL, satisfactory recovery (95-110%), and good intra- and inter-day precisions (RSD ≤ 10%) were obtained for 8-isoprostaglandin F2α and cortisol. Salivary lactate and 8-isoprostaglandin F2α were strongly correlated with NT-proBNP. Most patients (about 70%) showed a significant decrease (a factor of 3 at least) of both lactate and 8-isoprostaglandin F2α levels at discharge, suggesting a relationship between salivary levels and improved clinical conditions during hospitalization.

Micro-extraction by packed sorbent combined with UHPLC-ESI-MS/MS for the determination of prostanoids and isoprostanoids in dried blood spots.

This work presents a reliable analytical procedure combining micro-extraction by packed sorbent (MEPS) and ultra-high performance liquid chromatography-electrospray ionization tandem mass spectrometry to determine 8-iso prostaglandin F2α, 8-iso prostaglandin E2 and prostaglandin E2 in dried blood spots (DBSs). To reach this goal, we optimized a fast semi-automated MEPS procedure for the clean-up and pre-concentration of the analytes extracted from a single DBS (50 µL) by a 70:30 v/v methanol:water mixture. Limits of detection of about 20 pg mL-1, satisfactory recoveries (90-110%) and very good intra- and inter-day precisions (RSD ≤10%) were obtained for all the analytes. The innovative addition of internal standards on the filter paper before DBS sampling allowed to compensate changes in the amount of analyte during storage. Since prostanoids and isoprostanoids are biomarkers involved in the pathogenesis and progression of many diseases (e.g. ductal patency, diabetic nephropathy, and acute lung injury), our analytical method offers interesting diagnostic and prognostic opportunities in the medical field. The present method is currently used for the analysis of such biomarkers in DBSs from preterm newborns collected in the clinical setting.

Using labelled internal standards to improve needle trap micro-extraction technique prior to gas chromatography/mass spectrometry.

When working with humid gaseous samples, the amount of water vapour collected in a needle trap along with volatile analytes may vary from sample to sample and decrease during the storage. This has a major impact on desorption efficiency and recovery. We propose the addition of a labelled internal standards to nullify the effect of variable humidity on the analytical performance of needle trap micro-extraction combined with gas chromatography mass spectrometry. Triple-bed (Divinylbenzene/Carbopack X/Carboxen 1000) and single-bed (Tenax GR) needles were tested with standard gaseous mixtures prepared at different relative humidity levels (85%, 50% and 10%). The standard mixtures contained twenty-five analytes representative of breath and ambient air constituents, including hydrocarbons, ketones, aldehydes, aromatics, and sulphurs, in the concentration range 0.1-700 ppbv. The two needles showed different behaviours, as recovery was independent of humidity for single-beds, whereas a low recovery (10-20%) was observed when triple-beds trapped very volatile compounds at low humidity (e.g. pentane and ethanol, 10% relative humidity. Triple-beds showed an almost quantitative recovery (>90%) of all the analytes at 50% and 85% relative humidity. This big difference was probably due to the reduced action of water vapour pressure during the desorption step. The addition of 6D-acetone and 8D-toluene to the sorbent material before gas sampling and the normalization of raw data nullified this effect, thereby lowering the variations of analyte recovery at different humidity levels down to 20%. Internal standards were also exploited to limit within 10-20% alterations in peak areas of very volatile compounds during needle storage at room temperature. This variation may results from a loss of water vapour either retained from the sorbent material and/or condensed on triple-bed needle walls. After normalization, the inter- and intra-day precision were halved to 5% and 10% in the case of single-beds, respectively, and to 15% and 20% with three-beds. The addition of an internal standard to the sorbent helps to keep the overall analytical procedure under control and improves the reliability of needle trap micro-extraction for the analysis of volatile organic compounds at ultra-trace levels.

Ultrasensitive Immunosensor Array for TNF-α Detection in Artificial Saliva using Polymer-Coated Magnetic Microparticles onto Screen-Printed Gold Electrode.

Tumor necrosis factor-α (TNF-α) is a biomarker of inflammation that occurs in patients suffering from heart failure (HF). Saliva can be sampled in a non-invasive way, and it is currently gaining importance as matrix alternative to blood in diagnostic and therapy monitoring. This work presents the development of an immunosensor array based on eight screen-printed gold electrodes to detect TNF-α in saliva samples. Two different functionalization strategies of electrodes were compared. In the first, anti-TNF-α antibodies were chemically bonded onto the electrode by functionalization with 4-carboxymethylaniline. The other functionalization procedure involved the binding of antibodies onto polymer-coated magnetic microparticles, which were then deposited onto the electrode by pulsed chronoamperometry. Finally, the chronoamperometry technique was applied to characterize the modified SPEAu. The use of a secondary antibody anti-TNF-α (Ab-TNF-α-HRP) labelled with horseradish peroxidase (HRP, 2 µg·mL-1) was investigated using tetramethylbenzidine (TMB, pH = 3.75) as electrochemical substrate containing 0.2 mM of H2O2. A sandwich-type detection strategy with a secondary antibody anti-TNF-α provided chronoamperometric analyses in 10 s for each sample. Linearity, precision, limit of detection, and selectivity of devices were investigated. Interferences were evaluated by analyzing solutions containing other cytokine produced during the acute stage of inflammation. The immunosensor showed good performance within the clinically relevant concentration range, with a precision of 8%, and a limit of detection of 0.3 pg/mL. Therefore, it may represent a promising tool for monitoring HF in a non-invasive way.

Room temperature amine sensors enabled by sidewall functionalization of single-walled carbon nanotubes.

A new series of sidewall modified single-walled carbon nanotubes (SWCNTs) with perfluorophenyl molecules bearing carboxylic acid or methyl ester moieties are herein reported. Pristine and functionalized SWCNTs (p-SWCNTs and f-SWCNTs, respectively) were characterized by X-ray photoelectron spectroscopy (XPS), Raman spectroscopy and scanning electron microscopy (SEM). The nitrene-based functionalization provided intact SWCNTs with methyl 4-azido-2,3,5,6-tetrafluorobenzoate (SWCNT-N-C6F4CO2CH3) and 4-azido-2,3,5,6-tetrafluorobenzoic acid (SWCNT-N-C6F4CO2H) attached every 213 and 109 carbon atoms, respectively. Notably, SWCNT-N-C6F4CO2H was sensitive in terms of the percentage of conductance variation from 5 to 40 ppm of ammonia (NH3) and trimethylamine (TMA) with a two-fold higher variation of conductance compared to p-SWCNTs at 40 ppm. The sensors are highly sensitive to NH3 and TMA as they showed very low responses (0.1%) toward 200 ppm of volatile organic compounds (VOCs) containing various functional groups representative of different classes of analytes such as benzene, tetrahydrofurane (THF), hexane, ethyl acetate (AcOEt), ethanol, acetonitrile (CH3CN), acetone and chloroform (CHCl3). Our system is a promising candidate for the realization of single-use chemiresistive sensors for the detection of threshold crossing by low concentrations of gaseous NH3 and TMA at room temperature.

A computational approach for the estimation of heart failure patients status using saliva biomarkers.

The aim of this work is to present a computational approach for the estimation of the severity of heart failure (HF) in terms of New York Heart Association (NYHA) class and the characterization of the status of the HF patients, during hospitalization, as acute, progressive or stable. The proposed method employs feature selection and classification techniques. However, it is differentiated from the methods reported in the literature since it exploits information that biomarkers fetch. The method is evaluated on a dataset of 29 patients, through a 10-fold-cross-validation approach. The accuracy is 94 and 77% for the estimation of HF severity and the status of HF patients during hospitalization, respectively.

Determination of sevoflurane and isopropyl alcohol in exhaled breath by thermal desorption gas chromatography-mass spectrometry for exposure assessment of hospital staff.

Volatile anaesthetics and disinfection chemicals pose ubiquitous inhalation and dermal exposure risks in hospital and clinic environments. This work demonstrates specific non-invasive breath biomonitoring methodology for assessing staff exposures to sevoflurane (SEV) anaesthetic, documenting its metabolite hexafluoroisopropanol (HFIP) and measuring exposures to isopropanol (IPA) dermal disinfection fluid. Methods are based on breath sample collection in Nalophan bags, followed by an aliquot transfer to adsorption tube, and subsequent analysis by thermal desorption gas chromatography-mass spectrometry (TD-GC-MS). Ambient levels of IPA were also monitored. These methods could be generalized to other common volatile chemicals found in medical environments. Calibration curves were linear (r(2)=0.999) in the investigated ranges: 0.01-1000 ppbv for SEV, 0.02-1700 ppbv for IPA, and 0.001-0.1 ppbv for HFIP. The instrumental detection limit was 10 pptv for IPA and 5 pptv for SEV, both estimated by extracted ion-TIC chromatograms, whereas the HFIP minimum detectable concentration was 0.5 pptv as estimated in SIM acquisition mode. The methods were applied to hospital staff working in operating rooms and clinics for blood draws. SEV and HFIP were present in all subjects at concentrations in the range of 0.7-18, and 0.002-0.024 ppbv for SEV and HFIP respectively. Correlation between IPA ambient air and breath concentration confirmed the inhalation pathway of exposure (r=0.95, p<0.001) and breath-borne IPA was measured as high as 1500 ppbv. The methodology is easy to implement and valuable for screening exposures to common hospital chemicals. Although the overall exposures documented were generally below levels of health concern in this limited study, outliers were observed that indicate potential for acute exposures.

Pharmacological characterization of the vascular effects of aryl isothiocyanates: is hydrogen sulfide the real player?

Hydrogen sulfide (H2S) is an endogenous gasotransmitter, which mediates important physiological effects in the cardiovascular system. Accordingly, an impaired production of endogenous H2S contributes to the pathogenesis of important cardiovascular disorders, such as hypertension. Therefore, exogenous compounds, acting as H2S-releasing agents, are viewed as promising pharmacotherapeutic agents for cardiovascular diseases. Thus, this paper aimed at evaluating the H2S-releasing properties of some aryl isothiocyanate derivatives and their vascular effects. The release of H2S was determined by amperometry, spectrophotometry and gas/mass chromatography. Moreover, the vascular activity of selected isothiocyanates were tested in rat conductance (aorta) and coronary arteries. Since H2S has been recently reported to act as an activator of vascular Kv7 potassium channels, the possible membrane hyperpolarizing effects of isothiocyanates were tested on human vascular smooth muscle (VSM) cells by spectrofluorescent dyes. Among the tested compounds, phenyl isothiocyanate (PhNCS) and 4-carboxyphenyl isothiocyanate (PhNCS-COOH) exhibited slow-H2S-release, triggered by organic thiols such as L-cysteine. These compounds were endowed with vasorelaxing effects on conductance and coronary arteries. Moreover, these two isothiocyanates caused membrane hyperpolarization of VSM cells. The vascular effects of isothiocyanates were strongly abolished by the selective Kv7-blocker XE991. In conclusion, the isothiocyanate function can be viewed as a suitable slow H2S-releasing moiety, endowed with vasorelaxing and hypotensive effects, typical of this gasotransmitter. Thus, such a chemical moiety can be employed for the development of novel chemical tools for basic studies and promising cardiovascular drugs.