Study On The Application Of Evans Blue And Pontamine Sky Blue In The Detection Of Glycogen

Glycogen is a branched chain polysaccharide composed of glucose,which is a kind of energy storage biomolecules in vivo.In mammals,glycogen is primarily stored in the liver where it is used to maintain the balance of blood sugar,and glycogen in skeletal muscle provides energy for muscle stretching.When glycogen storage and breakdown function is impaired,people will suffer from a variety of related disease,such as glycogenosis and Pompe’s syndrome.Pompe’s syndrome is caused by the lack of enzyme/metabolic related enzymes like acid alpha-glucosidase,phosphorylase B kinase,glucose 6-phosphatase and glycogen synthetase kinase.Symptoms may include liver enlargement,hyperlipidemia,hypoglycemia,hyperuricemia,muscle atrophy.However,a small amount of glycogen is also stored in the kidney and brain.The glycogen in the kidney is very similar to the glycogen in the liver,which is mostly released into the blood to help maintain blood sugar balance.Brain glycogen is mainly involved in the regulation of the nervous system and provides energy for the nerves.The dysfunction of the system is related to a variety of neurological diseases including Alzheimer’s disease and Huntington’s disease.Therefore,based on various glycogen related diseases such as glycogen accumulation disease and neurological diseases,different glycogen detection methods began to emerge gradually.There are currently reported the methods of glycogen detection in the market,such as: PAS staining,anthrone method,antibody detection etc.Although they can identify glycogen,they all have some deficiencies.Anthrone require indirect conversion of glycogen into glucose to determine levels,which can lead to poor accuracy.However,the procedures involved in glycogen testing by kit method and PAS staining are complex and time-consuming.In addition,other detection methods can be expensive,which greatly limits its application.Therefore,it is very necessary to develop an economical,efficient,fast and convenient glycogen detection method.In this paper,two molecular dyes evans blue and pontamine sky blue were screened from our dye library as near-infrared fluorescence probes for the detection of glycogen.The results showed that the fluorescence of Ev B and PSB could be activated when the two dyes were mixed with glycogen respectively.Both PSB and Ev B showed linear response to glycogen concentration(0 μM～2 m M)with low detection limits.The detection method can identify glycogen stably even in complex environment simultaneously(such as different p H,temperature and various interferences or in the presence of cell extracts),and the supramolecular recognition between dyes(PSB/Ev B)and glycogen was demonstrated by fluorescence spectroscopy,isothermal calorimetry and other various methods.Most importantly,both were able to recognize glycogen specifically but not their assembly unit glucose.In addition,the mechanism of fluorescence luminescence between glycogen and Ev B is discussed by molecule docking.Finally,the present glycogen detection methods are compared and summarized.It is proved that PSB/Ev B-glycogen detection has some advantages: rapid efficient,good selectivity,strong stability.Therefore,this study shows that Ev B and PSB are convenient fluorescent probes with application prospect.