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We developed a state-of-the-art, high-sensitivity, low-stray-light standoff deep-ultraviolet (DUV) Raman spectrometer for the trace detection of resonance Raman-enhanced chemical species. As an excitation source for Raman measurements, we utilized our recently developed, second-generation, miniaturized, diode-pumped, solid-state neodymium-doped gadolinium orthovanadate (Nd:GdVO4) laser that generates quasi-continuous wave 228 nm light. This 228 nm excitation enhances the Raman intensities of vibrations of NOx groups in explosive molecules, aromatic groups in biological molecules, and various aromatic hydrocarbons. Our DUV Raman spectrograph utilizes a custom DUV f /8 Cassegrain telescope with an ∼200 mm diameter primary mirror, high-efficiency DUV transmission gratings, custom DUV mirrors, and a custom 228 nm Rayleigh rejection filter. We utilized our new standoff DUV Raman spectrometer to measure high signal-to-noise ratio spectra of ∼50 μg/cm2 drop-cast explosives: ammonium nitrate (AN), trinitrotoluene, pentaerythritol tetranitrate as well as aromatic biological molecules: lysozyme, tryptophan, tyrosine, deoxycytidine monophosphate, deoxyadenosine monophosphate at an ∼3 m distance within 10–30 s accumulation times. We roughly estimate the average ultraviolet resonance Raman (UVRR) detection limits for the relatively homogeneous drop-cast films of explosives and biological molecules to be ∼1 μg/cm2 when utilizing a continuous raster scanning that averages Raman signal over ∼1 cm2 sample area to avoid quick analyte depletion due to ultraviolet (UV) photolysis. We determined 3 m standoff UVRR detection limits for drop-cast AN films and identified factors impacting UVRR detection limits such as analyte photochemistry and analyte morphology. We found a detection limit of ∼0.5 μg/cm2 for drop-cast AN films on glass substrates when the Raman signal is averaged over ∼0.5 cm2 of sample surface using a continuous raster scan. For a step raster scan, when the probed sample area is limited to the laser spot size, the detection limit is approximately tenfold higher (∼5 μg/cm2) due to the impact of UV photochemistry. Graphical abstract This is a visual representation of the abstract. [ABSTRACT FROM AUTHOR]

The ultraviolet resonance Raman (UVRR) spectra of the two proteins bovine serum albumin (BSA) and human serum albumin (HSA) in an aqueous solution are compared with the aim to distinguish between them based on their very similar amino acid composition and structure and to obtain signals from tryptophan that has only very few residues. Comparison of the protein spectra with solutions of tryptophan, tyrosine, and phenylalanine in comparative ratios as in the two proteins shows that at an excitation wavelength of 220 nm, the spectra are dominated by the strong resonant contribution from these three amino acids. While the strong enhancement of two and one single tryptophan residue in BSA and HSA, respectively, results in pronounced bands assigned to fundamental vibrations of tryptophan, its weaker overtones and combination bands do not play a major role in the spectral range above 1800 cm–1. There, the protein spectra clearly reveal the signals of overtones and combination bands of phenylalanine and tyrosine. Assignments of spectral features in the range of Raman shifts from 3800 to 5100 cm–1 to combinations comprising fundamentals and overtones of tyrosine were supported by spectra of amino acid mixtures that contain deuterated tyrosine. The information in the high-frequency region of the UVRR spectra could provide information that is complementary to near-infrared absorption spectroscopy of the proteins. Graphical Abstract This is a visual representation of the abstract. [ABSTRACT FROM AUTHOR]

Raman spectra obtained by UV laser excitation above 260 nm are contaminated by spectrally overlapping UV‐excited fluorescence. Here, we demonstrate a picosecond UV Kerr‐gated Raman spectroscopy setup to suppress the fluorescence in Raman spectra excited by 280 nm laser pulses. Specifically, we use a model sample that consists of acetonitrile contaminated by a small amount of toluene for generating UV‐excited fluorescence. We were able to acquire a Raman spectrum of acetonitrile by suppressing the UV‐excited fluorescence of toluene by the action of a 1‐mm thick fused silica‐based UV Kerr gate operated by 1030 nm, ~1 ps gating pulses. These results also indicate the challenges that must be overcome in the future for realizing UV‐fluorescence‐free UV resonance Raman (UVRR) spectroscopy studies of molecules exhibiting electronic resonances around 280 nm using laser excitation wavelengths right at or close to this resonance. [ABSTRACT FROM AUTHOR]

Ultraviolet resonance Raman (UVRR) scattering is a highly sensitive and selective vibrational spectroscopic technique with a broad range of applications from polyaromatic hydrocarbons (PAHs) to biomolecular systems (peptides/proteins and nucleic acids) and catalysts. The interpretation of experimental UVRR spectra is not as straightforward as in purely vibrational Raman scattering (Placzek approximation) due to the involvement of higher lying electronic states and vibronic coupling. This necessitates the comparison with theoretical UVRR spectra computed by electronic structure calculations. Anthracene is an ideal model system for such a comparison between experiment and theory because it is rigid, symmetric, and of moderate size. By taking into account Herzberg–Teller contributions including Duschinsky effects, bulk solvent effects, and anharmonic contributions, a good qualitative agreement close to the resonance condition is achieved. The present study shows that within the framework of time‐dependent density functional theory (TD‐DFT), a general and robust approach for the analysis and interpretation of resonance Raman spectra of medium‐ to large‐size molecules is available. [ABSTRACT FROM AUTHOR]

Ultraviolet resonance Raman (UVRR) spectroscopy is a powerful vibrational spectroscopic technique for the label-free monitoring of molecular recognition of peptides or proteins with supramolecular ligands such as guanidiniocarbonyl pyrroles (GCPs). The use of UV laser excitation enables Raman binding studies of this class of supramolecular ligands at submillimolar concentrations in aqueous solution and provides a selective signal enhancement of the carboxylate binding site (CBS). A current limitation for the extension of this promising UVRR approach from peptides to proteins as binding partners for GCPs is the UV-excited autofluorescence from aromatic amino acids observed for laser excitation wavelengths >260 nm. These excitation wavelengths are in the electronic resonance with the GCP for achieving both a signal enhancement and the selectivity for monitoring the CBS, but the resulting UVRR spectrum overlaps with the UV-excited autofluorescence from the aromatic binding partners. This necessitates the use of a laser excitation

The ultraviolet resonance Raman spectra of the adenine-containing enzymatic redox cofactors nicotinamide adenine dinucleotide and flavin adenine dinucleotide in aqueous solution of physiological concentration are compared with the aim of distinguishing between them and their building block adenine in potential co-occurrence in biological materials. At an excitation wavelength of 266 nm, the spectra are dominated by the strong resonant contribution from adenine; nevertheless, bands assigned to vibrational modes of the nicotinamide and the flavin unit are found to appear at similar signal strength. Comparison of spectra measured at pH 7 with data obtained pH 10 and pH 3 shows characteristic changes when pH is increased or lowered, mainly due to deprotonation of the flavin and nicotinamide moieties, and protonation of the adenine, respectively. Graphical Abstract [ABSTRACT FROM AUTHOR]

We describe a new compact diode‐pumped solid‐state frequency quadrupled quasi‐three‐level neodymium‐doped gadolinium vanadate (Nd:GdVO4) laser that generates ~50 mW of 228‐nm quasi‐continuous wave light as ns pulses at a tunable kilohertz repetition rate. We developed two generations of this laser. The first generation has a high duty cycle and a tunable repetition rate. The second generation is optimized for maximum output power. We utilize these new lasers to measure ultraviolet resonance Raman (UVRR) spectra of many important chromophores that absorb in deep ultraviolet (UV). We demonstrate the utility of this excitation by measuring the 228‐nm absolute differential Raman cross sections of explosives, peptides, aromatic amino acids, and DNA/RNA nucleotides. Deep UV excitation at 228 nm occurs within the π → π* electronic transitions of these molecules. The 228‐nm resonance excitation enhances the Raman intensities of vibrations of NOx groups, peptide bonds, aromatic amino acid side chains, and DNA/RNA nucleotides. The measured 228‐nm UVRR cross sections of these molecules are 103–104 fold greater than those excited in the visible spectral region. These new lasers should be of great interest for UVRR spectroscopy and for other applications that benefit from compact, high average power deep UV laser light sources with low peak powers. [ABSTRACT FROM AUTHOR]

The effects of two pulp pretreatments, impregnation with ascorbic acid (AA) or purified kraft lignin (KL), on bleached pulp refining were investigated by examining and testing handsheets made from these pulps. The AA pretreatment of the pulp amplified the depolymerization of the cellulose and notably impaired the strength properties of the pulp handsheets. The effects were enhanced upon the combination of the AA pretreatment and intensive refining. Furthermore, heat treatments (at 225 °C, 30 min, in water vapor atmospheres of 1 and 75% (v/v)) promoted the depolymerization of cellulose and the total color difference in the AA impregnated handsheets more than for the KL impregnated and reference handsheets. In contrast to AA, the KL pretreatment of the pulp improved the burst index stability of the refined pulp handsheets after the humid thermal treatment (75% (v/v)). In addition, the total color difference of the KL impregnated handsheets was lower than the AA impregnated and reference handsheets.

Molecular insights on the β-lactoglobulin thermal unfolding and aggregation are derived from FTIR and UV Resonance Raman (UVRR) investigations. We propose an in situ and in real-time approach that thanks to the identification of specific spectroscopic markers can distinguish the two different unfolding pathways pursued by β-lactoglobulin during the conformational transition from the folded to the molten globule state, as triggered by the pH conditions. For both the investigated pH values (1.4 and 7.5) the greatest conformational variation of β-lactoglobulin occurs at 80 °C and a high degree of structural reversibility after cooling is observed. In acidic condition β-lactoglobulin exposes to the solvent its hydrophobic moieties in a much higher extent than in neutral solution, resulting on a highly open conformation. Moving from the diluted to the self-crowded regime, the solution pH and consequently the different molten globule conformation select the amyloid or non-amyloid aggregation pathway. At acidic condition the amyloid aggregates form during the heating cycle leading to the formation of transparent hydrogel. On the contrary, in neutral condition the amyloid aggregates never form. Information on the secondary structure conformational change of β-lactoglobulin and the formation of amyloid aggregates are obtained by FTIR spectroscopy and are related to the information of the structural changes localized around the aromatic amino acid sites by UVRR technique. Our results highlight a strong involvement of the chain portions where tryptophan is located on the formation of amyloid aggregates. • Vibrational spectroscopies to characterize the molten globule state of β-lactoglobulin • Amyloidogenic and non-amyloidogenic aggregation pathways determined by the molten globule conformation • Tryptophan involvement on the amyloid aggregates formation • Responsibility of the amyloid aggregates for the hydrogel network existence [ABSTRACT FROM AUTHOR]

The effects of two pulp pretreatments, impregnation with ascorbic acid (AA) or purified kraft lignin (KL), on bleached pulp refining were investigated by examining and testing handsheets made from these pulps. The AA pretreatment of the pulp amplified the depolymerization of the cellulose and notably impaired the strength properties of the pulp handsheets. The effects were enhanced upon the combination of the AA pretreatment and intensive refining. Furthermore, heat treatments (at 225 °C, 30 min, in water vapor atmospheres of 1 and 75% (v/v)) promoted the depolymerization of cellulose and the total color difference in the AA impregnated handsheets more than for the KL impregnated and reference handsheets. In contrast to AA, the KL pretreatment of the pulp improved the burst index stability of the refined pulp handsheets after the humid thermal treatment (75% (v/v)). In addition, the total color difference of the KL impregnated handsheets was lower than the AA impregnated and reference handsheets. [ABSTRACT FROM AUTHOR]

Recent advances in optical and spectroscopic technologies have enabled a plethora of Raman spectrometers that are suitable for studies of protein pharmaceuticals. Highly sensitive Raman spectrometers have overcome the handicap of the fundamentally weak Raman effect that hampered their applications to protein pharmaceuticals in the past. These Raman spectrometers can now routinely measure protein therapeutics at the low concentration of 1 mg/mL, which is on par with other spectroscopic methods such as CD, fluorescence and FTIR spectroscopies. In this article, various Raman techniques that can be used for protein pharmaceutical studies are reviewed. Novel Raman marker of proteins discovered from fundamental studies of protein complexes are examined along with established Raman spectra and structure correlations. Examples of Raman spectroscopic studies of protein pharmaceuticals are demonstrated. Future applications of Raman spectroscopy to protein pharmaceuticals are discussed. © 2007 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 96: 2861–2878, 2007 [ABSTRACT FROM AUTHOR]

Abstract: A facile synthetic scheme for the preparation of methyl 4-deoxy-β-l-threo-hex-4-enopyranosiduronic acid utilizing the commercially available methyl α-d-galactopyranoside as starting material has been developed. The synthesis sequence comprises six high yielding reaction steps: TEMPO oxidation, acetylation, methanolysis of the lactone, acetylation, β-elimination, and final removal of the protecting groups. Only one column chromatographic purification is needed throughout the whole sequence. The overall yield is 60%. The final product has been characterized by NMR, Raman, UVRR, FTIR, and HRMS. [Copyright &y& Elsevier]

Ultraviolet resonance Raman (UVRR) spectroscopy is a powerful vibrational spectroscopic technique for the label-free monitoring of molecular recognition of peptides or proteins with supramolecular ligands such as guanidiniocarbonyl pyrroles (GCPs). The use of UV laser excitation enables Raman binding studies of this class of supramolecular ligands at submillimolar concentrations in aqueous solution and provides a selective signal enhancement of the carboxylate binding site (CBS). A current limitation for the extension of this promising UVRR approach from peptides to proteins as binding partners for GCPs is the UV-excited autofluorescence from aromatic amino acids observed for laser excitation wavelengths >260 nm. These excitation wavelengths are in the electronic resonance with the GCP for achieving both a signal enhancement and the selectivity for monitoring the CBS, but the resulting UVRR spectrum overlaps with the UV-excited autofluorescence from the aromatic binding partners. This necessitates the use of a laser excitation <260 nm for spectrally separating the UVRR spectrum of the supramolecular ligand from the UV-excited autofluorescence of the peptide or protein. Here, we demonstrate the use of UVRR spectroscopy with 244 nm laser excitation for the characterization of GCP as well as guanidiniocarbonyl indole (GCI), a next generation supramolecular ligand for the recognition of carboxylates. For demonstrating the feasibility of the UVRR binding studies without an interference from the disturbing UV-excited autofluorescence, benzoic acid (BA) was chosen as an aromatic binding partner for GCI. We also present the UVRR results from the binding of GCI to the ubiquitous RGD sequence (arginylglycylaspartic acid) as a biologically relevant peptide. In the case of RGD, the more pronounced differences between the UVRR spectra of the free and complexed GCI (1:1 mixture) clearly indicate a stronger binding of GCI to RGD compared with BA. A tentative assignment of the experimentally observed changes upon molecular recognition is based on the results from density functional theory (DFT) calculations., (Copyright © 2020, Holtum et al.; licensee Beilstein-Institut.)

UV resonance Raman (UVRR) spectroscopy is a powerful tool for investigating the structure of biological molecules, such as proteins. Numerous UVRR spectroscopic markers that provide information on the structure and environment of the protein backbone and of amino acid side chains have recently been discovered. Combining these UVRR markers with hydrogen-deuterium exchange and advanced statistics is a powerful tool for studying protein systems, including the structure and formation mechanism of protein aggregates and amyloid fibrils. These techniques allow crucial new insights into the structure and dynamics of proteins, such as polyglutamine peptides, which are associated with 10 different neurodegenerative diseases. Here we summarize the spectroscopic structural markers recently developed and the important insights they provide.