Construction Of Nitrite Fluorescent Probes And Their Applications-detection And Imaging In Preserved Food And Biological Systems

Preserved foods were widely favored in people for their unique taste and texture.However,during the curing process,some anaerobic bacteria could reduce nitrate(NO3-)to nitrite(NO2-),which leaded to excessive production of NO2-in cured foods and made its content exceed the standard causing food safety problems.It was proven that NO2-had a strong toxicity could cause acute poisoning and potential carcinogenicity.Therefore,it was crucial to develop a highly sensitive,selective,and rapid detection method and tool for accurately assessing NO2content in pickled foods.Among the available molecular detection tools,fluorescent probes were widely acknowledged as an effective option due to their exceptional optical properties,high sensitivity,rapid response time,and ability to image in situ.Consequently,they have been widely employed in food safety testing and imaging analysis.This study aimed to design and develop three fluorescent probes with superior optical properties to investigate the rapid,accurate,and efficient detection and imaging analysis of NO2-in food and biological systems.The specific research contents of this study were as follows:1.A novel water-soluble nanoprobe(MTB)was constructed based on a photo-induced electron transfer(PET)strategy by primary-guest self-assembly of adamantane-labeled naphthalimide small molecule fluorophore NI-adH with polycyclodextrin(Poly-β-CD),which quenched the fluorescence signal of the fluorophore naphthalimide due to the strong PET effect of o-phenylenediamine(OPD).The spectral properties of MTB for NO2-were firstly investigated.When NO2-was present,OPD reacted rapidly with NO2-under weak acid conditions to form a triazole derivative,which inhibited the PET effect of OPD,a new gradually enhanced fluorescence emission peak appeared at 530 nm,and a bright green fluorescence was released from the detection solution.The probe showed good linearity with NO2-concentration in the range of 0-1 μM and its limit of detection(LOD)was as low as 21.8 nM.In moreover,MTB exhibited excellent selectivity,fast response time(60 s)and excellent biocompatibility.Noteworthy,MTB has been successfully used for the detection of NO2-in pickles and water samples as well as shrimp at low pH(～6.0)values with satisfactory results.2.Based on a host-guest self-assembly construction of a ratiometric fluorescent nanoprobe(RbNp-CD)with fluorescence resonance energy transfer(FRET),it consisted of two main parts:(1)an adamantane-labeled small molecule RN-adH as a guest containing an OPD with a chemically specific response to NO2-and a pH-sensitive rhodamine B fluorophore;(2)a Poly-β-CD with excellent water solubility and biocompatibility as a host.The spectral properties of RbNp-CD to NO2-were firstly investigated.After the response of the probe with NO2-,the fluorescence color of RbNp-CD solution changed from purplish red to pink,and the fluorescence intensity at both 530 nm and 576 nm was gradually enhanced with the increase of NO2-concentration In addition,the RbNp-CD showed a good linear relationship to NO2-within the NO2-concentration 0-1 μM with LOD as low as 22.3 nM.At the same time,RbNp-CD showed fast response time(60 s),high selectivity and sensitivity.Due to the above excellent properties of RbNp-CD,the final probe was successfully applied to the rapid assessment of NO2-in real food,water samples and aquatic animals with satisfactory results.3.Based on the FRET strategy,a dual-site responsive fluorescent probe CGT was constructed for the rapid detection of NO2-and bisulfite(HSO3-)in real samples and biological systems by linking a naphthalimide derivative as an energy donor and a xanthene derivative as an energy acceptor through a non-conjugated linker.The probes included an OPD structure and an oxonium ion as chemically specific response site for NO2-and HSO3-,respectively.First,the optical properties of the probe for NO2-and HSO3-were examined.When HSO3-was present,CGT reacted with HSO3-to form the adduct CGT-SO3-,and the fluorescence emission intensity at 628 nm gradually decreased and the red fluorescence signal disappeared.However,when CGT-SO3-responded with NO2-,the fluorescence emission intensity at 537 nm gradually increased and was accompanied by a bright cyan fluorescence emission.On the contrary,when CGT reacted with NO2-to form the triazole derivative CGT-N3,the fluorescence emission intensity at both 537 nm and 628 nm gradually increased,and a bright pink fluorescence signal appeared at this time.Upon the response of CGT-N3 with HSO3-the fluorescence emission intensity at 537 nm gradually increased and that at 628 nm gradually decreased.Moreover,CGT showed high sensitivity and selectivity for HSO3-and NO2-detection with LODs as low as 20.17 nM and 4.14 nM,respectively.More importantly,CGT showed good detection capability in complex water samples and was successfully used for the detection of HSO3-and NO2-in biological systems.