The Construction Of Novel CO And HNO Fluorescence/Photoacoustic Dual-Modality Ratiometric Molecular Probes And Their Biological Imaging Studies

Endogenous gas signaling molecules,including carbon monoxide(CO)and nitroxyl(HNO),play an important role in many physiological and pharmacological process and associate with various diseases in biological systems.To gain a deeper understanding of their role and mechanism of action,we need to develop a new technique for effective detection of gas signaling molecules.Optical imaging has been proven to be a powerful method for the detection and imaging of biological signals due to its non-invasive nature,high sensitivity and real-time in situ detection capability.However,the low spatial resolution of optical imaging limits the imaging of deeper tissues in living organisms.By contrast,photoacoustic imaging(PA),an emerging non-invasive and non-ionizing imaging method with the depth penetration capability of ultrasound imaging,can be used for imaging in deeper tissues,although the sensitivity of PA imaging are lower.With complementary advantages of fluorescence/photoacoustic imaging,the design of fluorescence/photoacoustic dual-mode imaging probe will be expected to improve the imaging efficacy.Since ratiometric measurement provides built-in self-calibration for signal correction,it enables more sensitive and reliable detection.Therefore,it is important to develop efficient dual-ratiometric fluorescent and photoacoustic probes to track signal molecules in biological samples.In this thesis,we have designed and synthesized two ratiometric fluorescence/photoacoustic dual-modality molecular probes for the detection of CO and HNO,respectively,and explore their imaging applications in cells and in mice.The research work was as follows:First,we reported the first dual ratiometric FL/PA probe DOP-CO for CO detection through palladium-mediated Tsuji-Trost chemistry.DOP-CO consisted of a dual ratiometric parent skeleton(thioxanthene-hemicyanine)and an allyl formate moiety,showing a 10.6-fold ratiometric fluorescence enhancement(F₇₈₅/F₇₂₀)and 2.14-fold ratiometric PA enhancement(PA₇₃₅/PA₆₇₀)upon activation by CO under physiological conditions.As expected,this unique probe DOP-CO was successfully employed for ratiometric imaging of exogenous and endogenous CO release behavior in Hep G2 cells.Notably,tail vein injection of probe DOP-CO resulted in liver-enrichment of mice and the endogenous CO enhancement and down regulation in the liver triggered by APAP-induced liver injury and liver repair,respectively,was confirmed by FL/PA dual-modality ratiometric imaging.More importantly,the dual ratiometric probe prepared in this study not only contributes to shed new light on the deep understanding of the relationship between the CO level and drug-induced liver injury and repair,but also provides a promising tool to study other CO-mediated biological processes.Second,we designed and synthesized the first Dual ratiometric probe DOP-HNO for the detection of endogenous HNO.DOP-HNO consisted of a dual ratiometric parent skeleton(thioxanthene-hemicyanine)and 2-(diphenylphosphine)benzoic acid,which can detect HNO changes in physiological conditions and biological systems with high sensitivity and specificity.After being activated by HNO under physiological conditions,a 4-fold ratio fluorescence enhancement(F₇₈₃/F₇₂₀)and a 2.6-fold ratio PA enhancement(PA₇₄₀/PA₆₇₀)could be observed.DOP-HNO has been successfully used to ratiometric imaging of exogenous and endogenous HNO release behavior in 4T1 cells.It can be used to visualize the crosstalk between NO and H₂S by the FL/PA imaging of HNO in cells and mice.Notably,by using DOP-HNO,we provided the direct visualization evidence of endogenous HNO in cancer models and further investigated its possible generation mechanisms.All of the results indicated that DOP-HNO could not only providing an effective tool for studying the complex relationship between HNO and cancer,but also serve as a new platform for exploring the pathological analysis of HNO-related diseases in complex biological systems.