Active Sulfur, Oxygen, Carbon, And Species Analysis Based On Ratio-based Molecular Probes

Reactive sulfur species?RSS?,reactive oxygen species?ROS?,and reactive carbonyl species?RCS?participate extensively in a wide scope of physiological and pathological processes,including cell proliferation,cytoprotection,redox homeostasis regulation,signal conduction as well as growth of organism.However,many studies have demonstrated that these species also can induce many serious diseases,such as oxidative stress damage,neurodegenerative injury.As an important RSS,sulfur dioxide?SO₂?acts significant rloes in anti-oxidation,signal transduction and cell protection,and meanwhile it is also the metabolite of anti-oxidation in living system.As an representative ROS,hypochlorous acid?HClO?plays vital roles in immune defense and anti-inflammatory.Excessive hyprochlorous acid can consume antioxidant,break redox homeostasis,and thus cause serious mitochondrial permeabilization as well as cell apoptosis.As an important RCS,formaldehyde acts virtual role in memory storage and recognization.Unfortunately,formaldehyde also has potential harm to people healthy.Chronic exposure of formaldehyde can restrict the development of body.More importantly,once combined with free radical,formaldehyde can aggravate the oxidant damage to body.Recently,although some pathogenic mechanisms have discussed,many biological effects of these ROS,RSS and RCS still remain unknown.Therefore,developing powerful tools is of significance to the in-depth exploaration of cells and living system.Comparing with conventional detection technologies,fluorescent imaging features many advantages such as non-invasive,high resolution,real time visual imaging in living body.Recently,two-photon fluorescent probes based on near-infrared excitation have extensively applied owing to its deeper penetration depth,low photo toxicity and background.Furthermore,those probes that have high fluorescent quantum yield and ratiometric fluorescent signals were also beneficial to improving sensitivity and eliminating background interference.In this work,we designed and synthesized three fluorescent probes to detect SO₂,HCl O and FA,and further performed the pathogenic studies.Mainly content of this work was as follows:1)We designed a bright,dual-ratiometric fluorescenct probe 2-?4-?1-methyl-phenanthro-9,10-imidazole-2-yl?-benzylidene?malononitrile?MPIBA?for the detection of SO₂/HClO.MPIBA could report SO₂ and HClO with C=C reaction sites via the nucleophilic addition and oxidation respectively and induce the remarkable shift in fluorescent emission thereby leading to the dual-ratiometric toward SO₂/HClO,respectivly.This probe exhibited fast response?SO₂:<60s;HClO:<10 s?and high sensitivity?detection limits:SO₂:3.5 nM;HClO:12.5 nM?to analytes.The developed probe was successfully applied for monitoring SO₂ and endogenous HClO in respectively the HeLa cells and the RAW 264.7 cells,and also available to visualizing the dynamic of SO₂/HClO in zebrafish.Finally,this probe allowed investigation of oxidative and anti-oxidative effects of intracellular SO₂ under the NaCl O induced oxidative stress with the aid of fluorescent imaging studies and flow cytometry analysis.2)A bright ratiometric fluorescent probe 4-?1H-phenanthro[9,10-d]imidazol-2-yl?phenyl)but-3-en-1-amine?PIPBA?was designed to visually monitoring the endogenous formaldehyde.PIPBA has high quantum yield?f=0.62?and showed high selectivity toward formaldehyde.A80 nm red shift in emission wavelength and more than 90 fold rate of fluorescent intensity showed after probe reacted with formaldehyde.Probe have successfully applied to monitoring endogenous formaldehyde in cells,tissue and vivo.With the aid of our probe,we have demonstrated that exerted indirect oxydative damage by reacting with reactive free radicals,thus giving more oxidizing species,which eventually caused aggravated oxygen damage to organism.And The indirect oxidative damage due to FA could be alleviated by addition of exogenous or endogenous antioxidant.3)Based on the Naphthalene,a mitochondria-target two-photon ratiometric probe?4-??6-?2,2-dicyanovinyl?naphthalene-2-yl?oxy?butyl?triphenyl phosphonium?DNB?for the detection of SO₂/HClO in living cells and vivo have designed.The probe has large two-photon absorption cross sections,high sensitivity?limit of detection 8.0 nM for SO₂ and 15.2 nM for HClO?,and quick response?SO₂<50 s,HClO<20 s?in presence of formaldehyde.Probe was applied for monitoring SO₂/HClO in respectively the HeLa cells and the RAW 264.7 cells.Besides,during the process of oxidative stress and antioxidant repair,confocal imaging in zebrafish has verified that there exists a crosstalk between SO₂/HClO.SO₂ can endogenously produce as a metabolite of anti-oxidant through oxidation of intracellular Cys by HClO.