标题: 建构修饰聚砒硌及奈米碳管之酪胺酸酶生物感测试片与其应用
Fabricating and application of the bioelectrochemical sensor strips of tyrosinase-immobolized screen printed carbon electrodes with polypyrrole and carbon nanotubes modification on the electrode surface
作者: 郭瀞韩
Kuo, Ching-Han
林志生
Lin, Chih-Sheng
生物科技学系
关键字: 电化学;酪胺酸酶;聚砒硌;奈米碳管;网版印刷碳电极;榖胱胺肽;electrochemistry;tyrosinase;polypyrrole;carbon nanotubes;screen printed carbon electrode;glutathione
公开日期: 2009
摘要: 此研究之目的在于架构一可携式应用于即时监控环境中酚类化合物、食物中农药残留及健康指标榖胱胺肽(glutathione; GSH)之酪胺酸酶网版印刷碳电极试片(screening printed carbon electrode; SPCE)。我们制备此可携带之抛弃型安培式生物感测试片并将其应用于快速检测酚类化合物、农药及榖胱胺肽。由于此生物感测系统所感测的为电子传递讯号,为提升此网版印刷碳电极试片之电子传递能力,我们于其上电聚合聚砒硌(polypyrrole; PPy)与奈米碳管(carbon nanotubes; CNTs)复合材料,再将酪胺酸酶以物理性吸附固定于其上做为生物辨识元件。由于聚砒硌具共轭π键之特性且与酸化之奈米碳管掺杂,此复合材料具良好之电子传递能力并可增加电极表面积,在利用循环伏安法探讨聚砒硌与奈米碳管复合材料于此系统之特性,结果显示修饰有聚砒硌或聚砒硌与奈米碳管复合材料之电极试片,在循环伏安法的还原波峰电流各别放大了3.9及4.8倍。
酪胺酸酶为一多酚氧化酶,可藉脱氢作用将酚类化合物氧化为醌 (o-quinone),生成的醌为电化学活性物质可在低电位下被还原,而此还原电流即为我们侦测的目标。于农药的检测上,因酪胺酸酶活性可被农药所抑制,所产生的醌也相对减少,导致还原电流降低,故在此侦测系统中农药的浓度,可由相对减少之反应电流数据量化得知。而在侦测榖胱胺肽这方面,则利用榖胱胺肽可和醌反应形成榖胱胺肽-醌(glutathione-quinone)复合物,导致还原电流降低,我们则可测定其电流抑制率估算榖胱胺肽之浓度。其中,影响此电极试片感测讯号之参数有pH值、缓冲液浓度及操作电压已最适化。此系统中最适之pH值为6.5及缓冲溶液浓度为50 mM之磷酸盐缓冲溶液,-400 mV的电压下进行反应,醌的最大反应电流可在一分钟内获得。
在此研究中,我们使用此酪胺酸酶生物感测试片侦测四种酚类化合物:酚(phenol)、邻苯二酚(catechol) 、对甲酚(p-cresol) 、邻苯三酚(pyrogallol),三种农药(含有机磷及氨基甲酸盐类):加保扶(carbofuran)、得灭莞(aldicarb)、马拉松(malathion) 及榖胱胺肽。所量测得之侦测极限为酚 50 □M、邻苯二酚 1 □M、对甲酚 2.5 □M、邻苯三酚 10 □M、加保扶 20 ppb、得灭莞 20 ppb、马拉松 20 ppb 及榖胱胺肽 6.25□□M。除此之外,此研究也针对各种化合物进行侦测范围的量测。研究证实,此修饰有砒硌及奈米碳管之酪胺酸酶网版印刷碳电极试片,具有相当潜力应用于环境中酚类化合物之监控、检测农药的残留,及健康指标榖胱胺肽的检测。
The purpose of this study is to fabricate the tyrosinase-SPCE strips possessing the potential to detect phenolic compounds for environment monitoring, pesticides for food safety and glutathione (GSH) for health care. Disposable amperometric biosensing strips were prepared for rapid determination of phenolic compounds, pesticides and GSH. Tyrosinase was immobilized on the surface of screening printed carbon electrode (SPCE) strips, which were modified by electrochemical polymerization of polypyrrole (PPy) and carbon nanotubes (CNTs). In the presence of PPy and PPy/CNTs modificaton, the cathodic peak current of the cyclic voltammogram of SPCE strips was significantly enhanced 3.9- and 4.8- fold, respectively.
Tyrosinase is a polyphenol oxidase that catalyzes the oxidation of phenolic compounds via dehydrogenation to o-quinones. The generated o-quinones are electoactive species that can be reduced at a low potential and the reduction current can be measured. In pesticides detection, tyrosinase activity inhibited by pesticides and the production of o-quinone was reduced. Thus, the concentration of the pesticides can be calculatedly obtained from the reduction of response current in the detection system. In GSH detection, GSH reacted with o-quinone and then produced glutathione-quinone complex. The complex reduced the quantity of o-quinone and made the reduction current decreasing. We obtained the inhibition percentage in the detection system to evaluate the concentration of GSH. The factors affecting the biosensing response of the SPCE strips such as pH, concentration of phosphate buffer and working potential were investigation. The optimum pH and concentration of phosphate buffer were 6.5 and 50mM, respectively. The tests were performed in presence of catechol substrate at -400 mV. The maximum response from the oxidation reaction of o-quinone was obtained within 1 minute.
Four phenolic compounds (phenol, catechol, p-cresol and pyrogallol), three pesticides (organophosphate: malathion and carbamate: aldicarb, carbofuran) and GSH were detected by the SPCE strips. The detection limit for phenol (50 □M), catechol (1□M), p-cresol (2.5 □M), pyrogallol (10 □M), aldicarb (20 ppb), malathion (20 ppb), carbofuran (20 ppb), and GSH (6.25 □M) was determined. Moreover, the detection range for each compound determined in this study was also evaluated. In conclusion, the SPCE strips modified with PPy and CNTs are potential for the forward applications in the detection of phenolic compounds to environment monitoring, pesticides in food to provide food safety and also GSH for body health.
URI: http://140.113.39.130/cdrfb3/record/nctu/#GT079728520
http://hdl.handle.net/11536/45293
显示于类别:Thesis