Katrin F. Domke

Quantitative coherent anti-Stokes Raman scattering (CARS) microscopy.

The ability to observe samples qualitatively at the microscopic scale has greatly enhanced our understanding of the physical and biological world throughout the 400 year history of microscopic imaging, but there are relatively few techniques that can truly claim the ability to quantify the local concentration and composition of a sample. We review coherent anti-Stokes Raman scattering (CARS) as a quantitative, chemically specific, and label-free microscopy. We discuss the complicating influence of the nonresonant response on the CARS signal and the various experimental and mathematical approaches that can be adopted to extract quantitative information from CARS. We also review the uses to which CARS has been employed as a quantitative microscopy to solve challenges in material and biological science.

Studying Surface Chemistry beyond the Diffraction Limit: 10 Years of TERS†

The use of an illuminated scanning probe tip to greatly enhance Raman scattering from the sample underneath the tip is one of the most intriguing developments in optical spectroscopy, and the steeply increasing number of publications per year shows that chemists, physicists and biologists alike recognize the importance and great potential of this technique. With tip-enhanced Raman spectroscopy (TERS), one of the main goals in surface science has been achieved, namely the combination of scanning probe microscopy and optical spectroscopy such as Raman spectroscopy. Important here is the use of the tip as an optical antenna to substantially increase the emitted radiation and to simultaneously improve the optical resolution much beyond the Abbe diffraction limit. This permits the correlation of topographic and chemical information of the same surface region. The synergy of detailed insight in morphology and the chemical nature of the target species facilitates data interpretation significantly and enables characterization of interfaces at the nanometer scale. A wide variety of substrates and sample molecules have been studied with TERS since the first publication of tip-enhanced Raman spectra, and the technique has reached a first level of maturity on its 10th birthday, with TERS applications extending into various research fields from surface chemistry over biology to nanoscale physics.

Label-Free Imaging of Lipophilic Bioactive Molecules during Lipid Digestion by Multiplex Coherent Anti-Stokes Raman Scattering Microspectroscopy

The digestion and absorption of lipophilic, bioactive molecules such as lipids, physiologically active nutrients (nutraceuticals), and drugs play a crucial role in human development and health. These molecules are often delivered in lipid droplets. Currently, the kinetics of digestion of these lipid droplets is followed by in vitro models that simulate gastrointestinal conditions, while phase changes within the lipid droplets are observed by light or electron microscopy. However real-time, spatially resolved information about the local chemical composition and phase behavior inside the oil droplet is not accessible from these approaches. This information is essential as the surface and phase behavior determine the local distribution of molecules in the oil droplets and thus may influence the rate of uptake, for example, by impairing the effective transfer of bioactive molecules to intestinal cells. We demonstrate the capability of multiplex coherent anti-Stokes Raman scattering (CARS) microspectroscopy to image the digestion process non-invasively, with submicrometer resolution, millimolar sensitivity, and without the need for labeling. The lipolysis of glyceryl trioleate emulsion droplets by porcine pancreatic lipase is imaged, and the undigested oil and the crystalline lipolytic products are distinguished by their different vibrational signatures. The digestion of droplets containing the phytosterol analogue ergosterol is also probed, and the crystals are observed to dissolve into the lipolytic products. The lipophilic drug progesterone and Vitamin D(3) are dissolved in glyceryl trioctanoate emulsion droplets, and the local concentration is mapped with millimolar sensitivity. The bioactive molecules are observed to concentrate within the droplets as the oil is hydrolyzed. This observation is ascribed to the low solubility of these molecules in the lipolytic products for this system. Neither the type of bioactive molecule nor the initial radius of the emulsion droplet had a large effect upon the rate of digestion under these conditions; lipolysis of the triglyceride by pancreatic lipase appears insensitive to the type of bioactive molecule in solution. These findings shed important new light on lipid digestion and open new possibilities for the chemical visualization of lipid digestion and phase changes in lipid droplets containing bioactive molecules, which in combination with other existing techniques will provide a full picture of this complex physicochemical process.

Label-Free Chemical Imaging of Catalytic Solids by Coherent Anti-Stokes Raman Scattering and Synchrotron-Based Infrared Microscopy†

Take a look inside: The combination of coherent anti-Stokes Raman scattering and synchrotron-based IR microscopy during the catalytic conversion of thiophene derivatives on zeolite crystals yields space- and time-resolved chemically specific information without the need for labeling (see picture). The thiophene reactant is mostly present in the center of the crystal, and the product is aligned within the straight pores of the zeolites.

Tip-Enhanced Raman Spectroscopic Studies of the Hydrogen Bonding between Adenine and Thymine Adsorbed on Au (111)

Although the hydrogen bond is one of the weakest chemical interactions, it plays a key role in forming Watson–Crick base pairs (adenine–thymine and guanine–cytosine) that hold together the two helical chains of nucleotides in DNA molecules and form the basis of the genetic code. [1–3] The bonding process is easily influenced by molecular structures, steric hindrance, [4] the base-pair geometry, [5] and the surrounding environment. [5–7] X-ray crystal structure analysis and NMR spectroscopy, [8] infrared spectroscopy, [9–11] Raman spectroscopy, [12] electrochemistry [13] and the quartz crystal microbalance technique [14] have been employed to explain the basic biological, chemical and physical mechanisms of hydrogen bonding. Most of the methods are based on the data collected from large amounts of molecules. Due to its extremely weak chemical interaction, hydrogen bonding gives rather subtle signals that are easily covered by signals from other stronger interactions between the molecules. Highly sensitive and selective methods are needed to get more accurate insight into hydrogen bonding. Tip-enhanced Raman spectroscopy (TERS) is a recently developed near-field spectroscopic method, which can simultaneously achieve high spatial resolution from the scanning probe microscopy (SPM) images and rich chemical information from the Raman spectra. [15, 16] Benefitting from the greatly enhanced electromagnefic field generated in the cavity of the SPM tip and the substrate by illuminating the SPM tip, TERS has been successfully applied to Raman studies on atomically smooth surfaces with high reproducibility. [16] The sample volume in surface-enhanced Raman spectroscopy (SERS) is defined by the laser focal spot projected on a rough coinage metal surface, that is, about 1 to 2 mm 2 ( 10 12 m 2 ) from where large amounts of molecules are excited. By using an STM tip to focus the enhanced electromagnetic field into an area of ~ nm range (~ 10 15 m 2 for a tip of 40 nm in diameter) [17] at an atomically smooth surface, the number of the excited molecules is dramatically reduced, allowing one to study molecular events at approximately the single-molecule level for resonant molecules [17] and picomole level for non-resonant molecules, such as DNA bases. [18] Very recently, a single brilliant cresyl blue molecule has been spectroscopically detected and imaged by using the UHV–TERS setup in our group. [19] A sensitive study of hydrogen bonding can therefore be achieved at single crystalline surfaces without information-averaging between large amounts of molecules and interference from ill-defined substrates.

In Situ Discrimination between Axially Complexed and Ligand-Free Co Porphyrin on Au(111) with Tip-Enhanced Raman Spectroscopy

Simultaneous chemical and topographic information about cobalt tetraphenyl-porphyrin (CoTPP) adlayers formed on a Au(111) single crystal is obtained with tip-enhanced Raman (TER) spectroscopy. We distinguish in situ between sample areas covered with an ordered adlayer of CoTPP and areas covered with a spontaneously formed disordered phase. The Raman vibrational fingerprints collected from the nanometer-sized near-field region just below a scanning tunnelling microscope (STM) tip are correlated with the adsorbate structures seen in the STM images. We assign the TER spectral features of the disordered phase to CoTPP complexes with CO and/or NO axial ligands, whereas the TER spectrum obtained from the ordered phase does not show any indication of additional axial complexation of CoTPP.

Versatile Side-Illumination Geometry for Tip-Enhanced Raman Spectroscopy at Solid/Liquid Interfaces

In situ characterization of surfaces with tip-enhanced Raman spectroscopy (TERS) provides chemical and topographic information with high spatial resolution and submonolayer chemical sensitivity. To further the versatility of the TERS approach toward more complex systems such as biological membranes or energy conversion devices, adaptation of the technique to solid/liquid working conditions is essential. Here, we present a home-built side-illumination TERS setup design based on a commercial scanning tunneling microscope (STM) as a versatile, cost-efficient solution for TERS at solid/liquid interfaces. Interestingly, the results obtained from showcase resonant dye and nonresonant thiophenol monolayers adsorbed on Au single crystals suggest that excitation beam aberrations due to the presence of the aqueous phase are small enough not to limit TER signal detection. The STM parameters are found to play a crucial role for solid/liquid TERS sensitivity. Raman enhancement factors of 10(5) at μW laser power demonstrate the great potential the presented experimental configuration holds for solid/liquid interfacial spectroscopic studies.

Host–Guest Geometry in Pores of Zeolite ZSM‐5 Spatially Resolved with Multiplex CARS Spectromicroscopy

Pore relations: Reagent molecules form head-to-tail chains in the pores of ZSM-5 zeolite particles in the presence or absence of acidic sites, according to multiplex coherent anti-Stokes Raman scattering spectromicroscopy (mCARS). The molecular ordering in the pores makes it possible to characterize the crystallographic subunits of individual ZSM-5 particles with (sub)micrometer spatial resolution in three dimensions.

Tracing Catalytic Conversion on Single Zeolite Crystals in 3D with Nonlinear Spectromicroscopy

The cost- and material-efficient development of next-generation catalysts would benefit greatly from a molecular-level understanding of the interaction between reagents and catalysts in chemical conversion processes. Here, we trace the conversion of alkene and glycol in single zeolite catalyst particles with unprecedented chemical and spatial resolution. Combined nonlinear Raman and two-photon fluorescence spectromicroscopies reveal that alkene activation constitutes the first reaction step toward glycol etherification and allow us to determine the activation enthalpy of the resulting carbocation formation. Considerable inhomogeneities in local reactivity are observed for micrometer-sized catalyst particles. Product ether yields observed on the catalyst are ca. 5 times higher than those determined off-line. Our findings are relevant for other heterogeneous catalytic processes and demonstrate the immense potential of novel nonlinear spectromicroscopies for catalysis research.

Nanoscale Distribution of Sulfonic Acid Groups Determines Structure and Binding of Water in Nafion Membranes

Abstract The connection between the nanoscale structure of two chemically equivalent, yet morphologically distinct Nafion fuel‐cell membranes and their macroscopic chemical properties is demonstrated. Quantification of the chemical interactions between water and Nafion reveals that extruded membranes have smaller water channels with a reduced sulfonic acid head group density compared to dispersion‐cast membranes. As a result, a disproportionally large amount of non‐bulk water molecules exists in extruded membranes, which also exhibit larger proton conductivity and larger water mobility compared to cast membranes. The differences in the physicochemical properties of the membranes, that is, the chemical constitution of the water channels and the local water structure, and the accompanying differences in macroscopic water and proton transport suggest that the chemistry of nanoscale channels is an important, yet largely overlooked parameter that influences the functionality of fuel‐cell membranes.