Water insoluble solids were prepared by washing solids with
25 ml DI water, vortexing 60 s, and centrifuging at 3500 rpm
(2000 g) for 3 min, which was adapted from the NREL protocol,
‘‘Determination of insoluble solids of pretreated biomass material
(NREL/TP-510-42627).” The washing step was repeated until the
pH of the samples reached 5–7. Compositions of water insoluble
solids were determined by the two step acid hydrolysis method
as described in the raw corn stover composition analysis.