Study On Electrochemical Cholesterol Sensor Based On Graphene And Nano Signal Amplification

Cholesterol is closely related to cardiovascular disease.The simple and accurate method for detecting serum total cholesterol(TC)is of great significance for clinical medicine.Electrochemical sensors have the advantages of fast detection speed,high sensitivity and good selectivity,and cost-effective.Graphene-based nanomaterials have been widely used in nano-enzymes and signal amplification technologies due to their large specific surface area,many number of active sites and good composite design effect.In this paper,two different electrochemical cholesterol sensors were constructed based on the enzymatic method combined with the signal amplification method of nanomaterials by using graphene-chitosan-ferrocene nanocomposite(RGO-CS-Fc)and graphene-hemin nanocomposite(RGO-Hemin),which realized the high sensitivity and low cost detection of cholesterol.(1)Cholesterol biosensor based on RGO-CS-Fc/Pt NPs nanocomposites:A large number of cholesterol esterase(CHER)and cholesterol oxidase(CHOD)were modified on the surface of screen-printed electrodes by multi-point covalent binding between RGO-CS-Fc/Pt NPs nanocomposites and cholesterol enzymes.An electrochemical sensor for cholesterol detection was constructed by electrochemical i-t detection method.An enzyme electrode based on Fc medium was constructed and cholesterol was detected by the electrochemical i-t detection method.The morphology and properties of RGO-CS-Fc nanocomposites and enzyme electrodes were analyzed by scanning electron microscopy(SEM),infrared spectroscopy(FI-IR),X-ray photoelectron spectroscopy(XPS)and so on.Some key factors affecting the performance of the cholesterol biosensor were optimized.Under the optimal experimental conditions of pH 7,CHER&CHOD concentration of 0.5 mg/mL,RGO-CS-Fc/concentration of 0.5 mg/mL,incubation temperature of 15°C,and RGO-CS-Fc dosage of 6?L,the working curve of cholesterol(I(nA)=13.76396 c(mg/mL)+72.29672,R~2=0.99897)was obtained by detecting the cholesterol standard sample.The redox response current measured and cholesterol concentration had a good linearity in the range of 0.5-4 mg/mL with low detection limit(LOD)of5.7?g/mL(S/N=3)and good sensitivity of 1.654,?A/mM*cm~2.Moreover,the RGO-CS-Fc/Pt NPs cholesterol enzyme biosensor exhibited excellent specificity,acceptable reproducibility(RSD=1.58%)and higher recoveries(95.38-103.83%)in the detection of clinical serum samples,showing great potential for clinical diagnosis.(2)Cholesterase biosensor based on RGO-Hemin nanoenzyme and Fc-Tyr oxidative deposition signal amplification:The RGO-Hemin nanomaterials with high peroxidase catalytic activity and CHER&CHOD were co-immobilized on the screen-printed electrode surface to construct a cholesterol sensing interface.Through the combined catalysis of RGO-Hemin and cholesterol enzyme,cholesterol decomposition was efficiently catalyzed.Then,the signal of Fc-Tyr oxidative deposition triggered by hydrogen peroxide reduction was amplified,and the highly sensitive detection of cholesterol was realized by electrochemical DPV detection method.The morphology,structure and properties of RGO-Hemin nano-enzyme,Fc-Tyr and enzyme electrodes were analyzed by transmission electron microscopy(TEM),liquid-phase mass spectrometry(LC-MS),particle size and Zeta potential analysis.Some key experimental conditions that affect cholesterol sensor performance are optimized and the optimal experimental conditions were:incubation temperature of 37°C,pH of 7,ratio of RGO-Hemin to Tyr of 1:2,ratio of CHOD to CHER of 1:1and ratio of Fc-Tyr to Tyr of 2:1.The working curve of cholesterol(I(?A)=0.63209c(mg/mL)+5.07219,R~2=0.99712)was obtained by detecting the cholesterol standard sample,which showed good linearity in the range of 0.1-5mg/mL with low detection limit(LOD)of 24.43?g/mL(S/N=3)and good sensitivity of 97.22?A/mM*cm~2).Also,the cholesterol biosensor exhibited good stability and specificity,potential detection in clinical(Recovery:96.63-107.32%).(3)Two electrochemical biosensors constructed in this paper have realized the detection of cholesterol in serum.The current response signal and sensitivity of the cholesterol sensor based on RGO-Hemin nanoenzyme and Fc-Tyr oxidation deposition amplification are better than those of the cholesterol biosensor based on RGO-CS-Fc/PtNPs.The stability and specificity are good with the basically same.