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Double emulsions show great potential for encapsulating active substances and protecting them against external influences. However, they tend to become unstable during storage. Research on double emulsions, therefore, focuses on maintaining their microstructure during their shelf life. Optical measurement methods, such as Raman spectroscopy, have hardly been used to date to analyze the microstructure of double emulsions, mainly due to multiple scattering effects. This study investigates the influence of refractive index matching of double emulsion phases by measuring the Raman signal strength of the inner water phase for different refractive index combinations. Ammonium nitrate and glycerol are added to the inner and outer water phase, respectively, to change the refractive indices of both phases. Additionally, polyvinyl alcohol serves as an emulsifier in the outer water phase. The oil phase consists of silicone oil and Dowsil Resin XR 0497 as the emulsifier. The refractive index of the oil phase is kept constant. For individual phase boundaries of single droplets, the refractive index matching plays a minor role. However, if there are many droplets with correspondingly numerous phase boundaries, which leads to multiple scattering during the measurement, the matching has a significant influence on the signal strength of the inner phase. When measuring double emulsions, the phases should always be matched, as this results in higher signals and improves the sensitivity of the measurement.

Multimodal imaging gains increasing popularity for biomedical applications. This article presents the design of a novel multimodal imaging system. The centerpiece is a light microscope operating in the incident and transmitted light mode. Additionally, Raman spectroscopy and VIS/NIR reflectance spectroscopy are adapted. The proof-of-concept is realized to distinguish between grey matter (GM) and white matter (WM) of normal mouse brain tissue. Besides Raman and VIS/NIR spectroscopy, the following optical microscopy techniques are applied in the incident light mode: brightfield, darkfield, and polarization microscopy. To complement the study, brightfield images of a hematoxylin and eosin (H&E) stained cryosection in the transmitted light mode are recorded using the same imaging system. Data acquisition based on polarization microscopy and Raman spectroscopy gives the best results regarding the tissue differentiation of the unstained section. In addition to the discrimination of GM and WM, both modalities are suited to highlight differences in the density of myelinated axons. For Raman spectroscopy, this is achieved by calculating the sum of two intensity peak ratios (I2857 + I2888)/I2930 in the high-wavenumber region. For an optimum combination of the modalities, it is recommended to apply the molecule-specific but time-consuming Raman spectroscopy to smaller regions of interest, which have previously been identified by the microscopic modes.

Raman and mid-infrared (MIR) spectroscopy are useful tools for the specific detection of molecules, since both methods are based on the excitation of fundamental vibration modes. In this study, Raman and MIR spectroscopy were applied simultaneously during aerobic yeast fermentations of Saccharomyces cerevisiae. Based on the recorded Raman intensities and MIR absorption spectra, respectively, temporal concentration courses of glucose, ethanol, and biomass were determined. The chemometric methods used to evaluate the analyte concentrations were partial least squares (PLS) regression and multiple linear regression (MLR). In view of potential photometric sensors, MLR models based on two (2D) and four (4D) analyte-specific optical channels were developed. All chemometric models were tested to predict glucose concentrations between 0 and 30 g L−1, ethanol concentrations between 0 and 10 g L−1, and biomass concentrations up to 15 g L−1 in real time during diauxic growth. Root-mean-squared errors of prediction (RMSEP) of 0.68 g L−1, 0.48 g L−1, and 0.37 g L−1 for glucose, ethanol, and biomass were achieved using the MIR setup combined with a PLS model. In the case of Raman spectroscopy, the corresponding RMSEP values were 0.92 g L−1, 0.39 g L−1, and 0.29 g L−1. Nevertheless, the simple 4D MLR models could reach the performance of the more complex PLS evaluation. Consequently, the replacement of spectrometer setups by four-channel sensors were discussed. Moreover, the advantages and disadvantages of Raman and MIR setups are demonstrated with regard to process implementation.