Department of Biochemistry, M D University, Rohtak 124001, Haryana, India.
Biosens Bioelectron. 2012 Jan 15;31(1):144-50. doi: 10.1016/j.bios.2011.10.007. Epub 2011 Oct 12.
A sulfite oxidase (SO(X)) (EC 1.8.3.1) purified from Syzygium cumini leaves was immobilized onto carboxylated gold coated magnetic nanoparticles (Fe(3)O(4)@GNPs) electrodeposited onto the surface of a gold (Au) electrode through N-ethyl-N'-(3-dimethylaminopropyl) carbodiimide (EDC)-N-hydroxy succinimide (NHS) chemistry. An amperometric sulfite biosensor was fabricated using SO(X)/Fe(3)O(4)@GNPs/Au electrode as working electrode, Ag/AgCl as standard and Pt wire as auxiliary electrode. The working electrode was characterized by Fourier Transform Infrared (FTIR) Spectroscopy, Cyclic Voltammetry (CV), Scanning Electron Microscopy (SEM) and Electrochemical Impedance Spectroscopy (EIS) before and after immobilization of SO(X). The biosensor showed optimum response within 2s when operated at 0.2V (vs. Ag/AgCl) in 0.1 M Tris-HCl buffer, pH 8.5 and at 35 °C. Linear range and detection limit were 0.50-1000 μM and 0.15 μM (S/N=3) respectively. Biosensor was evaluated with 96.46% recovery of added sulfite in red wine and 1.7% and 3.3% within and between batch coefficients of variation respectively. Biosensor measured sulfite level in red and white wines. There was good correlation (r=0.99) between red wines sulfite value by standard DTNB (5,5'-dithio-bis-(2-nitrobenzoic acid)) method and the present method. Enzyme electrode was used 300 times over a period of 4 months, when stored at 4 °C. Biosensor has advantages over earlier biosensors that it has excellent electrocatalysis towards sulfite, lower detection limit, higher storage stability and no interference by ascorbate, cysteine, fructose and ethanol.
从 Syzygium cumini 叶子中纯化的亚硫酸盐氧化酶（SO(X)）（EC 1.8.3.1）通过 N-乙基-N'-(3-二甲氨基丙基)碳二亚胺（EDC）-N-羟基琥珀酰亚胺（NHS）化学固定在通过电沉积在金（Au）电极表面上的羧基化金涂覆的磁性纳米颗粒（Fe(3)O(4)@GNPs）上。使用 SO(X)/Fe(3)O(4)@GNPs/Au 电极作为工作电极，Ag/AgCl 作为参比电极，Pt 丝作为辅助电极，制备了安培型亚硫酸盐生物传感器。在固定化 SO(X)前后，通过傅里叶变换红外（FTIR）光谱、循环伏安法（CV）、扫描电子显微镜（SEM）和电化学阻抗谱（EIS）对工作电极进行了表征。在 0.1 M Tris-HCl 缓冲液、pH 8.5 和 35°C 下，当在 0.2V（相对于 Ag/AgCl）下操作时，生物传感器在 2s 内显示出最佳响应。线性范围和检测限分别为 0.50-1000μM 和 0.15μM（S/N=3）。生物传感器在红葡萄酒中添加的亚硫酸盐的回收率为 96.46%，批内和批间变异系数分别为 1.7%和 3.3%。生物传感器测量了红葡萄酒和白葡萄酒中的亚硫酸盐水平。用标准 DTNB（5,5'-二硫代双-（2-硝基苯甲酸））法测定红葡萄酒中的亚硫酸盐值与本方法具有良好的相关性（r=0.99）。酶电极在 4 个月的时间内使用了 300 次，在 4°C 下储存。与早期的生物传感器相比，该生物传感器具有以下优点：对亚硫酸盐具有出色的电催化作用、更低的检测限、更高的存储稳定性以及不受抗坏血酸、半胱氨酸、果糖和乙醇的干扰。
