Trace Analysis of Biochemical Substances by Near-Infrared Semiconductor Laser Fluorimetry

1. 激光荧光法灵敏度高，可用于痕量分析。然而，由于激光体积大且价格昂贵，因此它还没有被广泛用作光谱分析的常规工具。在这项研究中，我开发了一种新的光谱测量方法，使用小型和更便宜的半导体激光器。吲哚菁绿在近红外区有一个吸收带，但被OH自由基猝灭。用该反应方案灵敏地测定了过氧化物酶生成OH自由基的情况。该方法允许在10^<-4> U水平下检测过氧化物酶。该方法也适用于酶免疫测定。基于三明治法，胰岛素以U/ml水平测定。近红外半导体激光器的振荡波长限制在780 ~ 900 nm，其分析应用受到很大限制。当激光紧密聚焦成非线性晶体时，会产生390 ~ 450 nm的二次谐波发射。当使用KDP晶体时，转换效率为2.5 × 10^<-6>。当激光介质本身产生二次谐波发射时，它减小到1.7 × 10^<-11>。这种发射用作光纤传感器的光源，并用于中和滴定终点的检测。在近红外区域，只有少数种类是荧光的，而且大多数在水溶液中化学性质不稳定。因此，它很难应用到实际的分析应用中。最近出现的可见光半导体激光器（670纳米）可以克服这个问题。用罗丹明800离子对试剂溶剂萃取成间二甲苯，测定表面活性剂。在10^<-7> M的水平下可以检测到十二烷基硫酸钠。为了将可见半导体激光荧光法应用于生化分子，研究了用可见染料对蛋白质进行荧光标记的各种方法。发现恶嗪750最适合作为本研究的标记试剂。少

1. Laser fluorimetry is very sensitive and is useful for trace analysis. However, a laser is large and expensive, so that it has not been widely used as a conventional tool for spectrometric analysis. In this study I develop a new spectrometric method using a small and less-expensive semiconductor laser.2. Indocyanine green has an absorption band in a near-infrared region, ant it is found to be quenched by OH radical. By using this reaction scheme peroxidase generating OH radical is sensitively determined. This method allows detection of peroxidase at 10^<-4> U levels. This method is also applied to enzyme immunoassay. Based on sandwich assay, insulin is determined at U/ml levels.3. The oscillating wavelength of the near-infrared semiconductor laser is restricted to 780-900 nm, and then its analytical application is quite limited. When the laser is tightly focused into a non-linear crystal, it generates second harmonic emission (390-450 nm). When a KDP crystal is used, the conversion e … More fficiency was 2.5 x 10^<-6>. It decreases to 1.7 x 10^<-11> when second harmonic emission is generated by the laser medium itself. This emission is used as a light source for an optical fiber sensor and for detection of the end-point in neutralization titration.4. In the near-infrared region only minor species are fluorescent, and most of them are chemically unstable in an aqueous solution. Therefore, it is difficult to apply it to practical analytical applications. A visible semiconductor laser (670 nm), which becomes recently available, is used to overcome this problem. A surfactant is determined after solvent extraction into m-xylene with an ion-pair reagent of Rhodamine 800. It is possible to detect sodium lauryl sulfate at 10^<-7> M levels. For application of visible semiconductor laser fluorimetry to a biochemical molecule, a various procedures are investigated for fluorescence labeling of protein with visible dyes. It is found that oxazine 750 is most suitable as a labeling reagent in this study. Less