The Design And Applications Of Dopamine Biosensor

Neurotransmitter is the important material in central nerve system of the mammal and mankind to transmit information. Dopamine participates in many physiological activities as an important neurotransmitter. A biosensor is designed to monitor the concentration of dopamine in rat caudate nucleus when drug is injected into rat. The research has considerable significance on the in vivo pharmaceutical analysis of dopamine and the diagnosis and treatment of diseases which are relevant to the abnormal metabolism of dopamine. In the thesis, the dopamine biosensor is made in two different ways which are the modification of maltodextrin and the technique of electrostatic self-assembly. The basic characteristics of the biosensor, the technique of development and in vivo monitoring have been investigated.The design and application of dopamine biosensor with the modification of maltodextrinFor the fabrication of the dopamine biosensor with the modification of maltodextrin, a pure platinum wire was settled in capillary, and then placed in stainless-steel pipe, with tyrosinase as sensitive substance, chitosan as carrier, maltodextrin as activating agent, Nafion as anti-interference material. The optimum conditions for modification of biosensor were determined through single factor test and orthogonal test, and the optimum conditions for immobilization were as follows: A 1.5% chitosan solution as the optimum concentration was coated 2 times on the surface of platinum, 0.15mg/ml tyrosinase and 10μl maltodextrin were immobilized on the support chitosan membrane for 5h in 4℃, then crosslinked with 0.25%glutaraldehyde for 4h in 4℃, too. Then the modified biosensor was immersed into 1% Nafion solution twice, 12 seconds once.The characteristical test of biosensor showed that the optimum pH value of the biosensor was 7.2~7.6, the optimum temperature is 37℃with the response time of which was 4min, and the linear range was 5×10-7mol/L~5×10-4mol/L with the detection limit of 5×10-7mol/L. The sensor could be continuously used for 6 days preserved in a wet state at 4℃, meanwhile, Na+, K+, Ca2+, NH4+ et al and antiscorbic acid almost had no effect on the biosensor. RSD of its reproducibility was less than 4.5%. The biosensor was employed to monitor the dynamic changes of dopamine in rat caudate nucleus. The results of test indicated that when madopar of different doses were injected into rat, the reponse time of sensor was about 10min and the response of sensor increased with the dose of madopar, in addition, when seligiline of different doses were injected into rat with the same dose of madopar administrated, the response of sensor also increased with the dose of seligiline, and the time of maximum concentration (Tmax) was about 40min.The design and application of dopamine biosensor with the technique of electrostatic self-assemblyThe technique of electrostatic self-assembly was applied to fabricate the dopamine biosensor with PO2 sensor as carrier, chitosan as polycation, tyrosinase as polyanion, Nafion as anti-interference material. The optimum conditions for immobilization of tyrosinase were determined through single factor test and orthogonal test, and the optimum conditions for immobilization were as follows: the pH value of the enzyme buffer was 7.0, the PO2 sensor was assembled in 1.0% chitosan solution and 0.15mg/ml tyrosinase alternately 10 times in 4℃, and then the modified biosensor was immersed into 1% Nafion solution twice, 10 seconds once.The characteristical test of biosensor showed that the optimum pH value of the biosensor was 7.2~7.8, the optimum temperature is 37℃with the response time of which was 3min, and the linear range was 5×10-7mol/L~5×10-4mol/L with the detection limit of 1×10-7mol/L. The sensor could be continuously used for 10 days preserved in a wet state at 4℃. The selectivity of the biosensor was good and RSD of its reproducibility was less than 5%. The results of in vivo test demonstrated that the trend of the dopamine concentration in rat caudate nucleus was relative to the dose of drug when selegiline hydrochlorides of low, medium, and high doses were injected into rat, while the more the drug was given, the shorter the response time of biosensor was, and the Tmax was about 45min.The dopamine biosensor fabricated by two different technologies had good characteristics, which could be applied to monitor the dynamic changes of dopamine in vivo, and the original construction of needle led to little injury of body and good biocompatibility. The research would provide the valuable informations on the in vivo or in vitro assay of dopamine.