Fluorescence-Encoded Infrared Spectroscopy for Single-Molecule Chemical Dynamics

在这项由化学结构、动力学和机制-化学系计划支持的项目中，芝加哥大学的Andrei Tokmakoff教授和他的研究小组正在开发一种新型的光学光谱，可以检测单个分子，并提供关于其结构的信息(换句话说，分子中原子的几何排列)。在一个实验中同时实现单分子检测和结构信息是一个长期追求的目标，但非常具有挑战性。托克马科夫教授的策略涉及到同时使用分子的荧光和振动特性。在荧光中，暴露在可见光或紫外光下的分子重新发射特定波长范围(颜色)的光。这种发射可以非常强，甚至足以检测到单个分子。然而，发射的光并不能告诉我们太多关于分子结构的信息。另一方面，分子的振动频率相当于红外线(IR)，也就是我们感觉到的热量，也可以用在夜视镜上。观察分子如何吸收红外光可以告诉我们许多关于它们结构的信息，但需要高浓度的分子才能获得足够的信号。托克马科夫使用可见光和红外光的激光脉冲，以这种方式仔细排序强烈的荧光发射包含通常在红外实验中提供的信息，因此被称为荧光编码红外(FEIR)光谱。这项研究将Feir技术集中在简单的反应上，如酸解离，分子失去一个质子(带正电的氢原子)，以及双分子络合物形成，两个分子相互粘连并经历微妙的结构变化。FEIR代表了一种理解化学反应的全新方法，不仅对化学，而且对材料科学和生物物理学都有深远的影响。从事这个项目的研究生正在经历一个跨学科的环境，其中包括先进仪器的设计和建造、光学和光谱学方面的培训、分子生物学方法以及分子建模和计算机模拟。为了广泛分发课程材料，托克马科夫教授还向该领域的其他人提供了在线教学资源。这个项目中使用的方法，荧光编码红外(FEIR)光谱，通过将红外光的吸收编码到可见的荧光信号上，将结构特异性和SM敏感性结合在一起。托克马科夫小组利用超快傅里叶变换和多维光谱的整体测量，展示了FEIR的原理和能力，并发展了测量10-100纳摩尔浓度下的FEIR光谱和飞秒瞬变的实验能力，接近单个分子的敏感区域。该项目的目标是开发执行FEIR模拟荧光相关光谱(FCS)实验的能力，在该实验中，振动光谱被用于跟踪分子的时间演化状态，用于研究化学动力学和动力学。托克马科夫小组将把这项技术应用于质子转移和双分子缔合/解离平衡的动力学，并研究费尔光谱如何感知荧光探针的环境。这被认为是实现一种方法的长期目标的第一步，该方法通过将振动信息集成到SM荧光光谱分析的实时光子流中来跟踪溶液中单分子的随时间演变的结构。除了上述对研究生的培训外，该项目还支持在量子力学、光谱学和分子生物物理学领域以不断更新的在线教科书和相关资源的形式继续开发和分发教育材料。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

In this project supported by the Chemical Structure, Dynamics and Mechanisms-A Program of the Division of Chemistry, Professor Andrei Tokmakoff and his research group at the University of Chicago are developing a new type of optical spectroscopy that can detect single molecules, and provide information about their structure (in other words, the geometric arrangement of atoms in the molecule). Achieving both single molecule detection and structural information in a single experiment is a long-sought-after, but very challenging goal. Professor Tokmakoff's strategy involves the use of both the fluorescence and vibrational properties of molecules. In fluorescence, a molecule exposed to visible or ultraviolet light re-emits light of a specific wavelength range (color). This emission can be very strong, even enough for a single molecule to be detected. However, the emitted light does not tell us very much about the structure of the molecule. On the other hand, molecules vibrate with frequencies that correspond to infrared light ("IR"), what we feel as heat, and also exploit in night vision goggles. Observing how molecules absorb IR light can tell us a lot about their structure, but one needs a high concentration of molecules to obtain sufficient signal. Tokmakoff uses laser pulses of visible and infrared light, carefully sequenced in such a way the intense fluorescence emission contains the information normally provided in an IR experiment, hence the name Fluorescence-Encoded Infrared (FEIR) spectroscopy. The research is focusing the FEIR technique on simple reactions like acid-dissociation, where a molecule loses a proton (positively charged hydrogen atom), and bimolecular complex formation, where two molecules stick to each other and undergo subtle structural changes. FEIR represents an entirely new approach to understanding chemical reactions, and has far -reaching implications not only in chemistry, but materials science and biophysics. The graduate students engaged in this project are experiencing a cross-disciplinary setting that includes design and construction of advanced instrumentation, training in optics and spectroscopy, methods in molecular biology, and molecular modeling and computer simulation. For broad distribution of course materials, Professor Tokmakoff has also made online instructional resources accessible by others in the field. The method used in this project, fluorescence-encoded infrared (FEIR) spectroscopy, combines structural specificity and SM sensitivity by encoding the absorption of IR light onto a visible fluorescence signal. The Tokmakoff group has demonstrated the principles and capabilities of FEIR, using ensemble measurements of ultrafast Fourier-transform and multidimensional spectroscopy, and developed the experimental capability for measuring FEIR spectra and femtosecond transients at 10-100 nanomolar concentrations, approaching the sensitivity regime of individual molecules. The objective of this project is to develop the capabilities for performing an FEIR analog of fluorescence correlation spectroscopy (FCS) experiments, in which the vibrational spectrum is used to track the time-evolving state of molecules for studies of chemical kinetics and dynamics. The Tokmakoff group will apply this technique to the kinetics of proton transfer and bimolecular association/dissociation equilibria, and study how FEIR spectra sense the environment of the fluorescent probe. This is imagined as a first step in the longer-term goal of realizing a method to track the time-evolving structures of single molecules in solution by integrating vibrational information into the real-time photon streams analyzed in SM fluorescence spectroscopy. In addition to the aforementioned training of graduate students, this project supports the continued development and distribution of educational material in the form of continually updated online textbooks and related resources, in the areas of quantum mechanics, spectroscopy, and molecular biophysics.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.