Abstract
BackgroundThe availability and low cost of lignocellulosic biomass has caused tremendous interest in the bioconversion of this feedstock into liquid fuels. One measure of the economic viability of the bioconversion process is the ease with which a particular feedstock is hydrolyzed and fermented. Because monitoring the analytes in hydrolysis and fermentation experiments is time consuming, the objective of this study was to develop a rapid fluorescence-based method to monitor sugar production during biomass hydrolysis, and to demonstrate its application in monitoring corn stover hydrolysis.ResultsHydrolytic enzymes were used in conjunction with Escherichia coli strain CA8404 (a hexose and pentose-consuming strain), modified to produce green fluorescent protein (GFP). The combination of hydrolytic enzymes and a sugar-consuming organism minimizes feedback inhibition of the hydrolytic enzymes. We observed that culture growth rate as measured by change in culture turbidity is proportional to GFP fluorescence and total growth and growth rate depends upon how much sugar is present at inoculation. Furthermore, it was possible to monitor the course of enzymatic hydrolysis in near real-time, though there are instrumentation challenges in doing this.ConclusionWe found that instantaneous fluorescence is proportional to the bacterial growth rate. As growth rate is limited by the availability of sugar, the integral of fluorescence is proportional to the amount of sugar consumed by the microbe. We demonstrate that corn stover varieties can be differentiated based on sugar yields in enzymatic hydrolysis reactions using post-hydrolysis fluorescence measurements. Also, it may be possible to monitor fluorescence in real-time during hydrolysis to compare different hydrolysis protocols.
Highlights
The availability and low cost of lignocellulosic biomass has caused tremendous interest in the bioconversion of this feedstock into liquid fuels
We showed that a higher amount of sugar in solution will cause the cells to stay in the log growth phase, and have higher fluorescence, for a longer period of time than when sugar levels are lower. (This is assuming these sugar solutions are within the dynamic range of the microbial assay.) These data support our revised hypothesis that instantaneous fluorescence is proportional to the sugar catabolism rate
Because the microbe expresses green fluorescent protein (GFP) constitutively, it allows the use of fluorescence as a measure of sugar concentration when the solution is too turbid to measure absorbance accurately or when other compounds interfere with this culture density measurement
Summary
The availability and low cost of lignocellulosic biomass has caused tremendous interest in the bioconversion of this feedstock into liquid fuels. One approach is to complete a simultaneous saccharification and fermentation (SSF) process [3] and to assay residual sugars and inhibitory products such as glucose, cellobiose, and acetic acid by high performance liquid chromatography (HPLC), and ethanol concentration by gas chromatograph (GC) or HPLC. Weimer et al have developed a higher throughput method to predict the fermentability of cellulosic biomass to ethanol through in vitro gas production [5]. In this procedure, fermentations are carried out in sealed serum bottles, and the gas produced is measured as an indicator of the digestibility of the cellulosic biomass
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
