Diamond atr crystal

Spectra from all standard mixture solutions and harvested media were collected using a Nicolet iS5 FT-IR spectrometer with a diamond ATR crystal (Thermo Scientific). The spectra were collected at 8 cm⁻¹ resolution and 32 averaged scans. Deionized (DI) water was used to collect the background signal for the experimental data. In addition, an air background was used for some specific comparisons. The spectra were processed using the Unscrambler X 10.4 software (Camo). To analyze the spectra in detail, we obtained 2^nd derivatives by applying the Savitzky-Golay algorithm. This is a mathematically objective approach used to enhance resolution of underlying and overlapping peaks and allow analysis of spectral features with greater specificity(25 (link)). The 2^nd derivative spectra were inverted (multiplication by −1) to transform to positive peaks. The spectral region from 1200–950 cm⁻¹ was selected for further analysis as it comprises the main glucose and lactate peaks and it has less confounding absorption bands from proteins in the medium. Multivariate analysis was performed by principal component analysis (PCA) of the spectral region of interest and the first principal component (PC-1) scores and loadings plot were assessed. The 2^nd derivative 1122/1035 peak intensity ratio was also quantified as a more direct approach to evaluate the lactate-to-glucose content of the samples.