Chiral Monitoring Across Both Enantiomeric Excess and Concentration Space: Leveraging Quantum Cascade Lasers for Sensitive Vibrational Circular Dichroism Spectroscopy.

Recently, high-throughput quantum cascade laser-based vibrational circular dichroism (QCL-VCD) technology has reduced the measurement time for high-quality vibrational circular dichroism spectra from hours to a few minutes. This study evaluates QCL-VCD for chiral monitoring using flow-through measurement of a changing sample in a circulating loop. A balanced detection QCL-VCD system was applied to the enantiomeric pair R/S-1,1'-bi-2-naphthol in solution. Different mixtures of the two components were used to simulate a racemization process, collecting spectral data at a time resolution of 6 min, and over three concentration levels. The goal of this experimental setup was to evaluate QCL-VCD in terms of both molar and enantiomeric excess (EE) sensitivity at a time resolution relevant to chiral monitoring in chemical processes. Subsequent chemometric evaluation by partial least squares regression revealed a cross-validated prediction accuracy of 2.8% EE with a robust prediction also for the test data set (error = 3.5% EE). In addition, the data set was also treated with the least absolute shrinkage and selection operator (LASSO), which also achieved a robust prediction. Due to the operating principle of LASSO, the obtained coefficients constituted a few discrete spectral frequencies, which represent the most variance. This information can be used in the future for dedicated QCL-based instrument design, gaining a higher time resolution without sacrificing predictive capabilities.

In situ based surface-enhanced Raman spectroscopy (SERS) for the fast and reproducible identification of PHB producers in cyanobacterial cultures.

The production of polyhydroxybutyrate (PHB) by autotrophic fermentation of cyanobacteria has received increasing interest in the light of carbon emission reducing process strategies. Biotechnological approaches are in development to optimize the yield of PHB, including adapted cultivation media, characterized by a limitation of key nutrients: cyanobacteria accumulate PHB as energy storage molecules under limited growth conditions. Since there is an increasing demand for fast, simple and reliable analytics, we report the establishment of surface enhanced Raman spectroscopy (SERS) as a suitable monitoring tool for up scaled PHB production processes. Both, pure Ag-colloids mixed with bacterial culture, and in situ prepared colloids (Ag-Synechocystis), generated on the cell surface directly, were successfully applied and evaluated for this purpose. SERS measurements with in situ prepared Ag-colloids improved the reproducibility of Raman signals from 54.8% to 93.9%. The measurement time could be reduced significantly, completing our secondary goal. The quality of classically and in situ prepared Ag-colloids was monitored by zeta potential measurements and scanning electron microscopy (SEM) respectively. For data interpretation and statistical model-building an in house written code in the open source software RStudio was implemented. It was applied for the differentiation of PHB producers at the cellular level, revealing heterogeneities within sample groups regarding the PHB amount accumulated. The results obtained using the statistical model were validated as well and were complementary to the reference HPLC analysis. Therefore, a fast and reliable identification in situ SERS tool for the selection of the most promising cyanobacterial PHB production was established.