Enzymatic Activities and Composition Analysis

FPase, pectinase, α-amylase, and α-glucosidase activities were measured as described elsewhere [50 (link)–52 (link)]. The protein concentration was assayed using the Lowry method [53 (link)]. For composition analysis of SPRs, glucose, xylose and galactose were produced according to the National Renewable Energy Laboratory protocols [54 ]. The content of glucose was determined as described elsewhere [6 (link)]. The contents of xylose and galactose were determined using a Dionex ICS 2500 system (Thermo Scientific, Waltham, MA, USA) with a CarboPac™ PA1 analytical column (2 × 250 mm). The elution solution was a mixture of water and 400 mM NaOH at a volume ratio of 95:5. The production and measurement of galacturonic acid followed the protocols as previously described [55 (link)]. The pectin content was based on the released galacturonic acid. Ash and lignin were measured as described elsewhere [56 (link)]. Starch content was determined with the method described previously [48 (link)]. The cellulose content was determined by subtracting the starch content from the glucan content. Three independent replicates were performed.