Comprehensive multi-gas study by means of fiber-enhanced Raman spectroscopy for the investigation of nitrogen cycle processes.

The extensive use of synthetic fertilizers has led to a considerable increase in reactive nitrogen input into agricultural and natural systems, resulting in negative effects in multiple ecosystems, the so-called nitrogen cascade. Since the global population relies on fertilization for food production, synthetic fertilizer use needs to be optimized by balancing crop yield and reactive nitrogen losses. Fiber-enhanced Raman spectroscopy (FERS) is introduced as a unique method for the simultaneous quantification of multiple gases to the study processes related to the nitrogen cycle. By monitoring changes in the headspace gas concentrations, processes such as denitrification, nitrification, respiration, and nitrogen fixation, as well as fertilizer addition were studied. The differences in concentration between the ambient and prepared process samples were evident in the Raman spectra, allowing for differentiation of process-specific spectra. Gas mixture concentrations were quantified within a range of low ppm to 100% for the gases N2, O2, CO2, N2O, and NH3. Compositional changes were attributed to processes of the nitrogen cycle. With help of multivariate curve resolution, it was possible to quantify N2O and CO2 simultaneously. The impact of fertilizers on N-cycle processes in soil was simulated and analyzed for identifying active processes. Thus, FERS was proven to be a suitable technique to optimize fertilizer composition and to quantify N2O and NH3 emissions, all with a single device and without further sample preparation.

Isotopomeric Peak Assignment for N2O in Cross-Labeling Experiments by Fiber-Enhanced Raman Multigas Spectroscopy.

Human intervention in nature, especially fertilization, greatly increased the amount of N2O emission. While nitrogen fertilizer is used to improve nitrogen availability and thus plant growth, one negative side effect is the increased emission of N2O. Successful regulation and optimization strategies require detailed knowledge of the processes producing N2O in soil. Nitrification and denitrification, the main processes responsible for N2O emissions, can be differentiated using isotopic analysis of N2O. The interplay between these processes is complex, and studies to unravel the different contributions require isotopic cross-labeling and analytical techniques that enable tracking of the labeled compounds. Fiber-enhanced Raman spectroscopy (FERS) was exploited for sensitive quantification of N2O isotopomers alongside N2, O2, and CO2 in multigas compositions and in cross-labeling experiments. FERS enabled the selective and sensitive detection of specific molecular vibrations that could be assigned to various isotopomer peaks. The isotopomers 14N15N16O (2177 cm-1) and 15N14N16O (2202 cm-1) could be clearly distinguished, allowing site-specific measurements. Also, isotopomers containing different oxygen isotopes, such as 14N14N17O, 14N14N18O, 15N15N16O, and 15N14N18O could be identified. A cross-labeling showed the capability of FERS to disentangle the contributions of nitrification and denitrification to the total N2O fluxes while quantifying the total sample headspace composition. Overall, the presented results indicate the potential of FERS for isotopic studies of N2O, which could provide a deeper understanding of the different pathways of the nitrogen cycle.

Spectrally Resolved and Highly Parallelized Raman Difference Spectroscopy for the Analysis of Drug-Target Interactions between the Antimalarial Drug Chloroquine and Hematin.

Malaria is a severe disease caused by cytozoic parasites of the genus Plasmodium, which infiltrate and infect red blood cells. Several drugs have been developed to combat the devastating effects of malaria. Antimalarials based on quinolines inhibit the crystallization of hematin into hemozoin within the parasite, ultimately leading to its demise. Despite the frequent use of these agents, there are unanswered questions about their mechanisms of action. In the present study, the quinoline chloroquine and its interaction with the target structure hematin was investigated using an advanced, highly parallelized Raman difference spectroscopy (RDS) setup. Simultaneous recording of the spectra of hematin and chloroquine mixtures with varying compositions enabled the observation of changes in peak heights and positions based on the altered molecular structure resulting from their interaction. A shift of (-1.12 ± 0.05) cm-1 was observed in the core-size marker band ν(CαCm)asym peak position of the 1:1 chloroquine-hematin mixture compared to pure hematin. The oxidation-state marker band ν(pyrrole half-ring)sym exhibited a shift by (+0.93 ± 0.13) cm-1. These results were supported by density functional theory (DFT) calculations, indicating a hydrogen bond between the quinolinyl moiety of chloroquine and the oxygen atom of ferric protoporphyrin IX hydroxide (Fe(III)PPIX-OH). The consequence is a reduced electron density within the porphyrin moiety and an increase in its core size. This hypothesis provided further insights into the mechanism of hemozoin inhibition, suggesting chloroquine binding to the monomeric form of hematin, thereby preventing its further crystallization to hemozoin.

Towards therapeutic drug monitoring of antibiotic levels - analyzing the pharmacokinetics of levofloxacin using DUV-resonance Raman spectroscopy.

Therapeutic drug monitoring (TDM) plays an important role in clinical practice. Here, pharmacokinetics has a decisive influence on the effective antibiotic concentration during treatment. Moreover, different kinetics exist for different administration forms. Accordingly, adjusting the correct concentration depends, in addition to gender, age, weight, clinical picture, etc., on the dosage form of the antibiotic. This study investigates the capability of deep UV resonance Raman spectroscopy (DUV-RRS) to simulate the pharmacokinetics of fluoroquinolone levofloxacin after two different administration forms (intravenous and oral). Three different pre-processing methods were applied, and the best agreement with the simulation was achieved using the extended multiplicative scatter correction. The resulting spectra were used for partial least squares (PLS) regression and ordinary least squares (OLS) regression. The kinetic parameters were compared with the simulated data, with PLS showing the best performance for intravenous administration and a comparable result to OLS for oral administration, while the errors are smaller. The acquired results show the potential of DUV-RRS in combination with PLS regression as a promising supportive method for TDM.

Parallelized Raman Difference Spectroscopy for the Investigation of Chemical Interactions.

Raman spectroscopy provides an extremely high chemical selectivity. Raman difference spectroscopy is a technique to reveal even the smallest differences that occur due to weak interactions between substances and changes in the molecular structure. To enable parallelized and highly sensitive Raman difference spectroscopy in a microtiter-array, a diffractive optical element, a lens array, and a fiber bundle were integrated into a Raman spectroscopy setup in a unique fashion. The setup was evaluated with a microtiter-array containing pyridine-water complexes, and subwavenumber changes below the spectrometer's resolution could be resolved. The spectral changes were emphasized with two-dimensional correlation analysis. Density functional theory calculation and "atoms in molecule" analysis were performed to simulate the intermolecular long-range interactions between water and pyridine molecules and to get insight into the involved noncovalent interactions, respectively. It was found that by the addition of pyridine, the energy portion of hydrogen bonds to the total complexation energy between pyridine and water reduces. These results demonstrate the unique abilities of the new setup to investigate subtle changes due to biochemically important molecular interactions and opens new avenues to perform drug binding assays and to monitor highly parallelized chemical reactions.

Hydrogen and C2-C6 Alkane Sensing in Complex Fuel Gas Mixtures with Fiber-Enhanced Raman Spectroscopy.

Power-to-gas is a heavily discussed option to store surplus electricity from renewable sources. Part of the generated hydrogen could be fed into the gas grid and lead to fluctuations in the composition of the fuel gas. Consequently, both operators of transmission networks and end users would need to frequently monitor the gas to ensure safety as well as optimal and stable operation. Currently, gas chromatography-based analysis methods are the state of the art. However, these methods have several downsides for time-resolved and distributed application and Raman gas spectroscopy is favorable for future point-of-use monitoring. Here, we demonstrate that fiber-enhanced Raman gas spectroscopy (FERS) enables the simultaneous detection of all relevant gases, from major (methane, CH4; hydrogen, H2) to minor (C2-C6 alkanes) fuel gas components. The characteristic peaks of H2 (585 cm-1), CH4 (2917 cm-1), isopentane (765 cm-1), i-butane (798 cm-1), n-butane (830 cm-1), n-pentane (840 cm-1), propane (869 cm-1), ethane (993 cm-1), and n-hexane (1038 cm-1) are well resolved in the broadband spectra acquired with a compact spectrometer. The fiber enhancement achieved in a hollow-core antiresonant fiber enables highly sensitive measurements with limits of detection between 90 and 180 ppm for different hydrocarbons. Both methane and hydrogen were quantified with high accuracy with average relative errors of 1.1% for CH4 and 1.5% for H2 over a wide concentration range. These results show that FERS is ideally suited for comprehensive fuel gas analysis in a future, where regenerative sources lead to fluctuations in the composition of gas.

Fiber-Enhanced Raman Gas Spectroscopy for the Study of Microbial Methanogenesis.

Microbial methanogenesis is a key biogeochemical process in the carbon cycle that is responsible for 70% of global emissions of the potent greenhouse gas methane (CH4). Further knowledge about microbial methanogenesis is crucial to mitigate emissions, increase climate model accuracy, or advance methanogenic biogas production. The current understanding of the substrate use of methanogenic microbes is limited, especially regarding the methylotrophic pathway. Here, we present fiber-enhanced Raman spectroscopy (FERS) of headspace gases as an alternate tool to study methanogenesis and substrate use in particular. The optical technique is nondestructive and sensitive to CH4, hydrogen (H2), and carbon dioxide with a large dynamic range from trace levels (demonstrated LoDs: CH4, 3 ppm; H2, 49 ppm) to pure gases. In addition, the portable FERS system can provide quantitative information about methanol concentration in the liquid phase of microbial cultures through headspace gas sampling (LoD 25 ppm). We demonstrate how FERS gas sensing could enable us to track substrate and product levels of microbial methanogenesis with just one instrument. The versatility of Raman gas spectroscopy could moreover help us to elucidate links between nitrogen and carbon cycle in microbial communities in the near future.

Counterfeit and Substandard Test of the Antimalarial Tablet Riamet® by Means of Raman Hyperspectral Multicomponent Analysis.

The fight against counterfeit pharmaceuticals is a global issue of utmost importance, as failed medication results in millions of deaths every year. Particularly affected are antimalarial tablets. A very important issue is the identification of substandard tablets that do not contain the nominal amounts of the active pharmaceutical ingredient (API), and the differentiation between genuine products and products without any active ingredient or with a false active ingredient. This work presents a novel approach based on fiber-array based Raman hyperspectral imaging to qualify and quantify the antimalarial APIs lumefantrine and artemether directly and non-invasively in a tablet in a time-efficient way. The investigations were carried out with the antimalarial tablet Riamet® and self-made model tablets, which were used as examples of counterfeits and substandard. Partial least-squares regression modeling and density functional theory calculations were carried out for quantification of lumefantrine and artemether and for spectral band assignment. The most prominent differentiating vibrational signatures of the APIs were presented.

Vibrational spectroscopic characterization of arylisoquinolines by means of Raman spectroscopy and density functional theory calculations.

Promising new antimalarial agents were investigated using FT-NIR and deep-UV resonance Raman spectroscopy. The Raman spectra of the seven arylisoquinolines (AIQ) were calculated with the help of density functional theory (DFT). Very good agreement with the experimental data was achieved and a convincing mode assignment was performed with the help of the calculated potential energy distribution (PED). For the non-resonant Raman spectra the most prominent bands were assigned to ν(C[double bond, length as m-dash]C) stretching modes of the isoquinoline system. To differentiate between substances with similar structures, deep-UV resonance Raman spectra were recorded. Raman bands in the range between 1250 and 1210 cm-1 were assigned to ν(C[double bond, length as m-dash]C) stretching vibrations in combination with δ(HCC) deformation vibrations of the aryl rests. These vibrations of the aryl part of the molecules were selectively enhanced, which, thus, enabled the differentiation of similar active agents from each other. For λexc = 257 nm excitation, strong ν(C[double bond, length as m-dash]C) vibrations of the isoquinoline (benzo-) part dominate the Raman spectrum in the range between 1685 and 1585 cm-1 and for λexc = 244 nm the Raman signals between 1430 and 1350 cm-1 were enhanced and assigned to ν(C[double bond, length as m-dash]C) of the isoquinoline (pyridino-) system.

Ultrasensitive Detection of Antiseptic Antibiotics in Aqueous Media and Human Urine Using Deep UV Resonance Raman Spectroscopy.

Deep UV resonance Raman spectroscopy is introduced as an analytical tool for ultrasensitive analysis of antibiotics used for empirical treatment of patients with sepsis and septic shock, that is, moxifloxacin, meropenem, and piperacillin in aqueous solution and human urine. By employing the resonant excitation wavelengths λexc = 244 nm and λexc = 257 nm, only a small sample volume and short acquisition times are needed. For a better characterization of the matrix urine, the main ingredients were investigated. The capability of detecting the antibiotics in clinically relevant concentrations in aqueous media (LODs: 13.0 ± 1.4 µM for moxifloxacin, 43.6 ± 10.7 µM for meropenem, and 7.1 ± 0.6 µM for piperacillin) and in urine (LODs: 36.6 ± 11.0 µM for moxifloxacin, and 114.8 ± 3.1 µM for piperacillin) points toward the potential of UV Raman spectroscopy as point-of-care method for therapeutic drug monitoring (TDM). This procedure enables physicians to achieve fast adequate dosing of antibiotics to improve the outcome of patients with sepsis.

Fiber enhanced Raman spectroscopic analysis as a novel method for diagnosis and monitoring of diseases related to hyperbilirubinemia and hyperbiliverdinemia.

Fiber enhanced resonance Raman spectroscopy (FERS) is introduced for chemically selective and ultrasensitive analysis of the biomolecules hematin, hemoglobin, biliverdin, and bilirubin. The abilities for analyzing whole intact, oxygenated erythrocytes are proven, demonstrating the potential for the diagnosis of red blood cell related diseases, such as different types of anemia and hemolytic disorders. The optical fiber enables an efficient light-guiding within a miniaturized sample volume of only a few micro-liters and provides a tremendously improved analytical sensitivity (LODs of 0.5 µM for bilirubin and 0.13 µM for biliverdin with proposed improvements down to the pico-molar range). FERS is a less invasive method than the standard ones and could be a new analytical method for monitoring neonatal jaundice, allowing a precise control of the unconjugated serum bilirubin levels, and therefore, providing a better prognosis for newborns. The potential for sensing very low concentrations of the bile pigments may also open up new opportunities for cancer research. The abilities of FERS as a diagnostic tool are explored for the elucidation of jaundice with different etiologies including the rare, not yet well understood diseases manifested in green jaundice. This is demonstrated by quantifying clinically relevant concentrations of bilirubin and biliverdin simultaneously in the micro-molar range: for the case of hyperbilirubinemia due to malignancy, infectious hepatitis, cirrhosis or stenosis of the common bile duct (1 µM biliverdin together with 50 µM bilirubin) and for hyperbiliverdinemia (25 µM biliverdin and 75 µM bilirubin). FERS has high potential as an ultrasensitive analytical technique for a wide range of biomolecules and in various life-science applications.