Two-Photon Circular Dichroism Spectrometer

双光子圆二色性（TPCD），即两个左旋圆偏振光子与两个右旋圆偏振光子的差异吸收，首先由Tinoco（1975）和Power（1975）在理论上进行了讨论。该技术在理论上类似于拉曼旋光，但尚未在实验上得到证实，尽管相关的TP-ORD效应已经得到证实（Gedanken Tamir，1987）。我们建议构建一个TPCD光谱仪的基础上，最近开发的商业可用的钛：蓝宝石飞秒激光技术，使用非常敏感的瞬态热光栅检测方法。我们主要感兴趣的领域是蛋白质的二级结构。鉴于这种蛋白质缺乏荧光，本申请规定了用于检测TPCD信号的直接吸收或基于热的方法。TPCD提供了几个潜在的优势，作为一种方法的补充，传统的圆二色性（CD）的蛋白质二级结构的测定和检测细微的构象变化。这些是（1）提供了一个新的、线性独立的数据集，潜在地允许二级结构的唯一测定，（2）300 ~ 600 nm的TPCD谱覆盖了对应于一个-从而将激发的范围扩展到更高的能量，并覆盖α-（3）有三个独立的TPCD光谱（对于相同频率的双光子和随机取向的样品）证明独立的信息。光谱仪是基于高重复率（1 kHz）飞秒激光技术。双光子CD信号的检测将基于差分瞬态热光栅方法。这种方法最近已被证明是一种非常灵敏的CD测量方法，以前已被证明是一个非常灵敏的方法进行双p光子吸收测量。我们的建议是将这两种能力联合收割机来提供新的双光子CD信息。详细的信号噪声计算的基础上，使用这种光栅的方法和预期的差分双光子截面的可靠估计的双光子测量表明，将实现良好的信号噪声。

Two-photon circular dichroism (TPCD), the differential absorption of two left circularly polarized photons vs. two right circularly polarized photons, was first discussed theoretically by Tinoco (1975) and by Power (1975). This technique, which is theoretically similar to Raman optical activity, has not yet been demonstrated experimentally although the related TP-ORD effect has been demonstrated (Gedanken & Tamir, 1987). We propose to construct a TPCD spectrometer based on recently developed commercially available Ti:sapphire femtosecond laser technology using the very sensitive transient thermal grating detection method. Our primary area of interest is the secondary structure of proteins. Given the lack of fluorescence of such proteins, this application dictates a direct absorption or thermally based methodology for detection of the TPCD signal. TPCD offers several potential advantages as a method complimentary to conventional circular dichroism (CD) for the determination of protein secondary structure and detection of subtle conformational changes. These are (1 ) a new, linearly independent data set is provided potentially permitting a unique determination of secondary structure, (2) the TPCD spectrum extending from 300 to 600 nm covers electronic excited states corresponding in one-photon energy to 150 to 300 nm thus extending the range of the excitations to higher energy and covering the expected higher exciton components of the a-helix, (3) there are three independent TPCD spectra (for two-photons of the same frequency and randomly oriented samples) proving independent information. The spectrometer to be constructed is based on high-repetition rate (1 kHz) femtosecond laser technology. The detection of the two-photon CD signal will be based on the differential transient thermal grating method. This method has been recently shown to provide a very sensitive method for CD measurements and has previously been shown to be a very sensitive method for performing two-p hoton absorption measurements. Our proposal is to combine these two capabilities to provide new two-photon CD information. A detailed signal to noise calculation based on prior two-photon measurements using this grating method and a reliable estimate of the expected differential two-photon cross section shows that excellent signal to noise will be achieved.