Plant tissue-based chemiluminescence flow biosensor for oxalate

W. Qin, Z. Zhang, Youyuan Peng, Baoxin Li
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引用次数: 6

Abstract

A novel plant tissue-based chemiluminescence (CL) biosensor for oxalate combined with flow injection analysis is proposed in this paper. The analytical reagents involved in the CL reaction, including luminol and cobalt(II), were both immobilized on an ion exchange resin column, while the biological material spinach tissue was packed in a mini-glass column. By the oxalate oxidase-catalyzed reaction in the plant tissue column, hydrogen peroxide was produced, which could react with luminol and cobalt(II) being released from the ion exchange column by hydrolysis to generate a CL signal. The CL emission intensity was linear with oxalate concentration in the range 0.6–100 µM and the detection limit was 0.2 µM. The biosensor was stable for 300 determinations and a complete analysis, including sampling and washing, could be performed in 2 min with a relative standard deviation of less than 5%.In recent years, traditional enzyme biosensors have been challenged by biosensors which use new biocatalytic materials, including animal and plant tissues1–3 and microorganisms.4–6 The catalytic function of these types of biosensors is due to the enzymes linked with metabolic pathways which exist either in the cytoplasmic membranes or directly inside the cells of these materials. Compared to biosensors with immobilized isolated and pure enzymes, such biosensors with immobilized tissues or whole cells show potential advantages of low cost, high stability, and a high level of enzyme activity. So far, most of them are bioselective membrane electrodes, in which the biocatalytic layer is usually retained physically at the detecting electrode surface with a support membrane and the analyte is sensed by diffusing through the test solution to the inner detector surface. However, these sensors often suffer from problems of long response time, low sensitivity and complex sensor assembly.Flow injection chemiluminescence (CL) analysis is becoming increasingly important in various fields for its high sensitivity, rapidity, simplicity and feasibility. Nowadays, CL flow sensing systems with immobilized reagents have received much attention and many applications have appeared in the literature.7–11 In these systems, analytes are detected by the CL reactions with the immobilized reagents directly or with the dissolved reagents which are released from the immobilized substrates by appropriate eluents. In this paper, a new type of biosensor, based on a plant tissue reactor, with flow injection CL detection for the determination of oxalate is proposed. It is prepared by using a spinach tissue column as the source of oxalate oxidase to catalyze the oxidation reaction of oxalate producing hydrogen peroxide, which is then detected by the CL reaction with luminol and cobalt(II) bleeding from an ion exchange column with immobilized reagents by hydrolysis.12
草酸盐植物组织化学发光流动生物传感器
提出了一种结合流动注射分析的草酸盐植物组织化学发光(CL)生物传感器。将参与CL反应的分析试剂鲁米诺和钴(II)固定在离子交换树脂柱上,将生物材料菠菜组织包装在微型玻璃柱上。草酸氧化酶在植物组织柱中催化反应生成过氧化氢,过氧化氢与离子交换柱水解释放的鲁米诺和钴(II)反应产生CL信号。在0.6 ~ 100µM范围内,氯离子发射强度与草酸浓度呈线性关系,检出限为0.2µM。该生物传感器在300次检测中稳定,可在2分钟内完成完整的分析,包括取样和洗涤,相对标准偏差小于5%。近年来,传统的酶生物传感器受到使用新型生物催化材料(包括动植物组织1 - 3和微生物)的生物传感器的挑战。这些类型的生物传感器的催化功能是由于与代谢途径相关的酶存在于细胞质膜中或直接存在于这些材料的细胞内。与固定化分离酶和纯酶的生物传感器相比,固定化组织或全细胞的生物传感器具有成本低、稳定性高、酶活性高的潜在优势。目前,大多数是生物选择性膜电极,其生物催化层通常物理保留在检测电极表面,并有支撑膜,分析物通过测试溶液扩散到检测内表面。然而,这些传感器往往存在响应时间长、灵敏度低和传感器组装复杂的问题。流动注射化学发光分析以其高灵敏度、快速、简单、可行等优点,在各个领域中发挥着越来越重要的作用。目前,固定化试剂的CL流量传感系统受到了广泛的关注,并在文献中出现了许多应用。7-11在这些系统中,通过直接与固定试剂或与溶解试剂的CL反应来检测分析物,这些试剂通过适当的洗脱剂从固定底物中释放出来。本文提出了一种基于植物组织反应器的流动注射CL检测草酸盐的新型生物传感器。以菠菜组织柱为草酸氧化酶源,催化草酸氧化反应生成过氧化氢,用固定试剂与离子交换柱流出的鲁米诺和钴(II)水解,用CL反应检测过氧化氢
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