A Novel Technique For Bioluminescent Detection Of Cobalt,Ferrous And Sulfite

Cobalt and iron are essential trace elements in human body since they are very important for the maintenance of normal physiological activities of organism.Both loss and overload of these elements will lead to serious physiological diseases,thus causing tissue and organ damage.Sulphite is a common food additive used primarily as a preservative,including controlling microbial growth,preventing browning and decay,as well as bleaching some foodstuffs.However,it has been reported that long-term consumption of sulphite-containing foods can cause skin and respiratory allergies,and thus endangering huge,which requires us to control their content accurately.In addition to the traditional detection methods,small molecule fluorescent probe detection method also has a strong advantage,but these methods have common defects,can not be achieved in vivo level detection.Therefore,in this thesis,bioluminescence is used to explore the detection of the target to solve this problem,which enriches the detection methods for cobalt,ferrous and sulfite and strengthens the applications in practical life.Based on the reported design principles of small molecule fluorescent probes for cobalt,ferrous and sulfite,three types of bioluminescent probes were designed and evaluated for the detection of cobalt,ferrous and sulfite in vitro and in vivo.This thesis mainly includes four parts:1:Overview of cobalt,iron and sulfite,biological significance.Traditional detection methods;summary of the existing small-molecule cobalt,iron and sulfite fluorescence probes and design principles;introduced the basic situation of bioluminescence.2:This part introduces a ferrous ion biolu.inescence probe for Co2+ recognition.The recognition group of this probe is complexed with cobalt ion by the N30 structure.Under the participation of oxygen,the C-O bond cleaves to release the luciferase substrate,and the detection of cobalt ions is realized by the live imager.In this part,the design of probe,synthetic route,in vitro enzyme level detection,cell level detection and animal level of exogenous cobalt ions were tested.The results.show that the cobalt ion probe has good sensitivity and selectivity in aqueous solution;linearity detection of cobalt ions can also be achieved at the cellular level.In the animal level,cobalt ions are taken both by tail vein injection and gavage administration,which can both realize the dynamic visualization of cobalt ions and detect the metabolic accumulation of cobalt ions at different time in the body.3:This part introduces a ferrous ion bioluminescence probe for Fe2+ recognition.Iron is one of the essential trace elements in human body,and the iron element of human body ranks first among the trace elements,among which ferrous iron is very important for the maintenance of normal physiological function of organism.In view of this situation,we designed and synthesized a new ferrous bioluminescent probe for the detection of ferrous content in vitro and in vivo.The results show that the probe has good sensitivity and selectivity to ferrous ions in aqueous solution,and can quantitatively detect ferrous ions in vitro.It also enables visual detection of endogenous ferrous ions excreted by mice injected with ferrous ions and mouse sepsis models.Thus,it was proved that the ferrous ion bioluminescent probe can well detect ferrous ions in a multi-environment comprehensive manner.4:This section mainly introduces the detection of sulfite by bioliminescenct technique.Sulfites are currently used as preservatives,including controlling the growth of microorganisms,preventing browning and decay,and bleaching of some foodstuffs.However,excessive intake of sulfite also can have serious harm to human health.Therefore,this section designed and synthesized three sulfite probes.Levulinic acid and its derivatives as the main recognition group,and sulfite ring-release luciferase substrate to detect sulfite.In this part,the design of probe,synthetic route,enzyme level detection in vitro and exogenous sulfite detection at animal level were discussed.The results show that SOP-2 and SOP-3 are less sensitive to sulfites,so SOP-1,a highly sensitive and selective probe can be used for the detection of sulfite in vivo.