Heat and Mass Transport in Processing of Lignocellulosic Biomass for Fuels and Chemicals - Sustainable Biotechnology: Sources of Renewable Energy - page 14

cellulases from binding to cellulose [42]. Other indirect mechanisms that impede
complete cellulose hydrolysis are also possible such as non-productive binding
of cellulases to lignin [34-36], however reports that contradict this theory also
exist [57].
Enzymatic hydrolysis of biomass pretreated under alkaline conditions, which
hydrolyzes less xylan than acidic pretreatments, supports the steric hindrance
concept. Elevated cellulolytic activity is observed on alkaline pretreated biomass
when cellulases are supplemented with xylanases and other hemicellulose degrading
enzymes, likely a function of removing additional barriers to cellulose accessibility
[58, 59]. A study in pretreatment variability by Selig and co-workers suggested that
cellulose digestibility is improved directly by xylan removal, but only indirectly
by lignin removal [47]. Removal of lignin by pretreatment appeared to increase
enzymatic removal of xylan, which in turn increased cellulose digestibility. Lignin
removal alone had little impact on cellulose digestion. Lignin modifying enzymes,
however, have been shown to synergistically work with cellulases during digestion
of steam-pretreated biomass, improving sugar yields through at least partial removal
of the lignin barrier [60]. In spite of a general consensus in the scientific community
about the significance of the lignin barrier to cellulose digestibility, only limited
attention has been given to the fate of lignin during widely used high tempera-
ture dilute acid, hot water, and steam pretreatments which only partially remove
lignin [1, 8].
A recent study investigated the fate of lignin during high temperature acid and
neutral pretreatments using electron microscopy and spectroscopy techniques [40].
This study revealed that lignin could be mobilized within the cell wall matrix at
temperatures as low at 120 ◦ C during both neutral and low pH pretreatments, and
appears to be, at least in part, dependent on pretreatment severity. On a relatively
macro scale, part of the mobilized lignin deposits back on to biomass surfaces as
spherical bodies, suggesting that lignin undergoes the following sequence of events
during these pretreatments - phase-transition or melting, mobilization into bulk
solution, coalescence, and deposition onto solid surfaces. Scanning- and transmis-
sion electron microscopy (SEM and TEM) of pretreated cell walls shows that the
lignin droplets (stained with KMnO 4 ) take a wide range of sizes (<50 nm to 2

μ

m)
and shapes (Fig. 4a, b and Fig. 5), though the “free” shapes are uniformly spheri-
cal. Other shapes observed appear to be dictated by the physical constraints of the
structures surrounding them. In addition to redeposition, there also appears to be
a reorganization of lignin structure within the cell walls. A fraction of the lignin
remains within the walls during pretreatment. This fraction apparently melts, but is
unable to escape into the bulk liquid phase before coalescing back into droplets, as
evidenced by the KMnO 4 stained lignin droplets that appear between layers in the
cell wall (Fig. 4b-d).
Aside from the obvious implications of lignin mobility, coalescence, and rede-
position observed during high temperature pretreatments, chemical modification of
the lignin should also be considered. These may range from covalent bond break-
age and formation to changes in inter- and intramolecular interactions. Although
FTIR and NMR studies did not distinctly show chemical changes in the mobilized