The Preparation Of Luminol Glow-type Chemiluminescent Hydrogels And Its Application In Cholesterol Detection

Chemiluminescence（CL）has been widely used in the fields of clinical diagnosis,food detection and environmental detection due to its advantages of no excitation light source,low background interference and high sensitivity.However,most CL systems are flash type with a fast decay rate,so it is depended on automatic and highly sensitive instruments for CL signal capture,which limits its application and development in point-of-care testing（POCT）.In recent years,the exploration of glow-type CL systems with high luminescence intensity and long-lasting emission has become a hot topic in this field,which is of great scientific significance in the development of new CL analysis and detection methods.In this paper,glow-type chemiluminescent hydrogels based on luminol CL system was prepared with alginate as the matrix,the CL and related properties of the glow-type chemiluminescent hydrogels were studied,and CL sensors for real-time detection of cholesterol was constructed.The main research contents are as follows:（1）Glow-type chemiluminescent hydrogels（HRP/COD/luminol/Alg hydrogels）containing horseradish peroxidase（HRP）,cholesterol oxidase（COD）and luminol was successfully prepared by a simple and mild chemical gelation method using alginate as substrate.Benefiting from the large amount of H₂O₂ produced by enzymatic cascade reaction,HRP/COD/luminol/Alg hydrogels reacted with H₂O₂ with high luminous intensity and obvious visualization effect.More importantly,due to the high viscosity and porous structure of alginate hydrogels,slow diffusion of H₂O₂ reacted with HRP/COD/luminol/Alg hydrogels to produce longer and more stable chemiluminescence.Therefore,the smartphone could be used as a detector instead of a special instrument to capture the CL signal.With the catalytic specificity of COD for cholesterol,the glow-type CL sensor was developed for the specific and sensitive detection of cholesterol.The linear range was from 0.01 to 0.35 m M（R~2=0.9913）,and the detection limit（LOD）was 5.3μM.The method was used for the evaluation of the level of cholesterol in serum.The results were approximate to the clinical reference value（RSD<10%）,and the standard recovery ranged from 92.8%to 107.5%.This work provides a new strategy for cholesterol detection in clinical diagnosis,which broadens the application of glow chemiluminescence sensor in POCT.（2）Glow-type chemiluminescence resonance energy transfer（CRET）hydrogels（HRP/COD/SF/luminol/Alg）was prepared and used for POCT of cholesterol.Luminol was used as the energy donor and sodium fluorescein（SF）was selected as the energy receptor for CRET.Based on CRET,we not only achieved green chemiluminescence with improved luminescence efficiency and visualization,but also reduced the amount of luminol and costs.In addition,profiting from the slow diffusion of molecules in high-viscosity alginate brine gels,CRET was sustained for up to 1 h in glow-type chemiluminescent hydrogels.On the basis of glow-type CL,a fast and convenient cholesterol detection was achieved by using a smartphone as the detector.The linear range was from 0.005 m M to 0.30 m M（R~2=0.9868）,and LOD was 0.29μM.This method was also used for the evaluation of the level of cholesterol in serum.The detection results were approximate to the clinical reference value（RSD<6%）,and the standard recovery ranged from 97.3%to 109.1%.Glow-type chemiluminescence based on CRET extends the spectrum range of chemiluminescence,improves the sensitivity of analysis method,and has great application potential in multivariate real-time detection,chemiluminescence imaging and color lamp source.