A national laboratory of the U.S. Department of Energy Office of Energy Efficiency & Renewable Energy National Renewable Energy Laboratory Innovation for Our Energy Future Determination of Sugars, Byproducts, and Degradation Products in Liquid Fraction Process Samples Laboratory Analytical Procedure (LAP) Issue Date: 12/08/2006 A. Sluiter, B. Hames, R. Ruiz, C. Scarlata, J. Sluiter, and D. Templeton Technical Report NREL/TP-510-42623 January 2008 NREL is operated by Midwest Research Institute Battelle Contract No. DE-AC36-99-GO10337 National Renewable Energy Laboratory 1617 Cole Boulevard, Golden, Colorado 80401-3393 303-275-3000 www.nrel.gov Operated for the U.S. Department of Energy Office of Energy Efficiency and Renewable Energy by Midwest Research Institute Battelle Contract No. DE-AC36-99-GO10337 Technical Report NREL/TP-510-42623 January 2008 Determination of Sugars, Byproducts, and Degradation Products in Liquid Fraction Process Samples Laboratory Analytical Procedure (LAP) Issue Date: 12/08/2006 A. Sluiter, B. Hames, R. Ruiz, C. Scarlata, J. Sluiter, and D. Templeton DISCLAIMER These Standard Biomass Analytical Methods (“Methods”) are provided by the National Renewable Energy Laboratory (“NREL”), which is operated by the Midwest Research Institute (“MRI”) for the Department Of Energy. Access to and use of these Methods shall impose the following obligations on the user. The user is granted the right, without any fee or cost, to use, copy, modify, alter, enhance and distribute these Methods for any purpose whatsoever, except commercial sales, provided that this entire notice appears in all copies of the Methods. Further, the user agrees to credit NREL/MRI in any publications that result from the use of these Methods. The names NREL/MRI, however, may not be used in any advertising or publicity to endorse or promote any products or commercial entity unless specific written permission is obtained from NREL/MRI. The user also understands that NREL/MRI is not obligated to provide the user with any support, consulting, training or assistance of any kind with regard to the use of these Methods or to provide the user with any updates, revisions or new versions. THESE METHODS ARE PROVIDED BY NREL/MRI “AS IS“ AND ANY EXPRESS OR IMPLIED WARRANTIES, INCLUDING BUT NOT LIMITED TO, THE IMPLIED WARRANTIES OF MERCHANTABILITY AND FITNESS FOR A PARTICULAR PURPOSE ARE DISCLAIMED. IN NO EVENT SHALL NREL/MRI BE LIABLE FOR ANY SPECIAL, INDIRECT OR CONSEQUENTIAL DAMAGES OR ANY DAMAGES WHATSOEVER, INCLUDING BUT NOT LIMITED TO CLAIMS ASSOCIATED WITH THE LOSS OF DATA OR PROFITS, WHICH MAY RESULT FROM AN ACTION IN CONTRACT, NEGLIGENCE OR OTHER TORTIOUS CLAIM THAT ARISES OUT OF OR IN CONNECTION WITH THE ACCESS, USE OR PERFORMANCE OF THESE METHODS. Procedure Title: Determination of Sugars, Byproducts, and Degradation Products in Liquid Fraction Process Samples Laboratory Analytical Procedure 1. Introduction 1.1 Carbohydrates make up a major portion of biomass samples. These carbohydrates are polysaccharides constructed primarily of glucose, xylose, arabinose, galactose, and mannose monomeric subunits. During certain pretreatments of biomass, a portion of these polysaccharides are hydrolyzed and soluble sugars are released into the liquid stream. This method is used to quantify the total amount of soluble carbohydrates released into solution as well as the amount of monomeric sugars released into solution. The soluble sugars in the liquid fraction of process samples can be quantified by HPLC with refractive index detection. If the sugars are present in oligomeric form further processing into their monomeric units is required prior to HPLC analysis. 1.2 The liquid portion may also contain carbohydrate degradation products, such as HMF and furfural, as well as other components of interest, such as organic acids and sugar alcohols. This method is used to measure the level of these degradation products and byproducts. These components are analyzed by HPLC with refractive index detection to determine optimal production process parameters or to monitor ongoing processes. 1.3 The concentrations of monomeric sugars (soluble monosaccharides) and cellobiose, total sugars (monosaccharides and oligosaccharides), as well as carbohydrate degradation products and sugar alcohols can be determined using this procedure. Monomeric sugars are quantified by HPLC with refractive index detection. Oligomeric sugars are converted into the monomeric form using acid hydrolysis and quantified by HPLC with refractive index detection. Byproducts and degradation products are quantified by HPLC with refractive index detection. 2. Scope 2.1 This procedure is used to characterize liquid process samples, including pretreatment liquors, liquid fermentation samples, and liquid fractions of process solids. 2.2 This procedure is appropriate for biomass containing the components