Determination of glucose and ethanol after enzymatic hydrolysis and fermentation of biomass using Raman spectroscopy

Raman spectroscopy has been used for the quantitative determination of the conversion efficiency at each step in the production of ethanol from biomass. The method requires little sample preparation; therefore, it is suitable for screening large numbers of biomass samples and reaction conditions in a complex sample matrix. Dilute acid or ammonia-pretreated corn stover was used as a model biomass for these studies. Ammonia pretreatment was suitable for subsequent measurements with Raman spectroscopy, but dilute acid-pretreated corn stover generated a large background signal that surpassed the Raman signal. The background signal is attributed to lignin, which remains in the plant tissue after dilute acid pretreatment. A commercial enzyme mixture was used for the enzymatic hydrolysis of corn stover, and glucose levels were measured with a dispersive 785 nm Raman spectrometer. The glucose detection limit in hydrolysis liquor by Raman spectroscopy was 8 g L−1. The mean hydrolysis efficiency for three replicate measurements obtained with Raman spectroscopy (86 ± 4%) was compared to the result obtained using an enzymatic reaction with UV–vis spectrophotometry detection (78 ± 8%). The results indicate good accuracy, as determined using a Student's t-test, and better precision for the Raman spectroscopy measurement relative to the enzymatic detection assay. The detection of glucose in hydrolysis broth by Raman spectroscopy showed no spectral interference, provided the sample was filtered to remove insoluble cellulose prior to analysis. The hydrolysate was further subjected to fermentation to yield ethanol. The detection limit for ethanol in fermentation broth by Raman spectroscopy was found to be 6 g L−1. Comparison of the fermentation efficiencies measured by Raman spectroscopy (80 ± 10%) and gas chromatrography–mass spectrometry (87 ± 9%) were statistically the same. The work demonstrates the utility of Raman spectroscopy for screening the entire conversion process to generate lignocellulosic ethanol.