Rapid flow-through fractionation of biomass to preserve labile aryl ether bonds in native lignin

Article abstract of DOI:10.1039/c9gc02315a

Lignin is the second largest component of vascular plants and is the most abundant renewable aromatic polymer on our planet. The attractiveness of lignin valorization lies in its conversion into high value aromatic chemicals and biofuels through fractionation and upgrading. The literature has demonstrated that the presence of aryl ether bonds in native lignin was a key factor for the conversion, while the conventional technical lignins from carbohydrate-first processes, e.g. pulp and cellulose ethanol production, are intensively condensed and lack these linkages due to the intense delignification conditions. Here, by using the β-O-4 lignin model dimer GG, we reveal the dramatic degradation of GG and the synchronous formation of relatively stable intermediate β-O-4 dimers, C₆C₃ enol ether and the formylated enol ether, within the first 5 min under the conditions of 72 wt% aqueous formic acid and 130 °C, conditions suitable for biomass fractionation. Based on these findings, we propose a simple but effective strategy of rapid flow-through fractionation (RFF), which separates the dissolved lignin from the reactor in time and space, thereby preserving these labile aryl ether bonds in native lignin. The application of RFF of poplar wood with a short residence time of 2.6 min attained 75% delignification with an equivalent of the β-O-4 motif in native lignin. Structure-preserved lignins (β-O-4 retention, 75.0%-85.4%) were also harvested from wheat straw with good lignin yields (61.7%-78.5%). Contrarily, batch fractionation acted as a protracted war and resulted in extensive cleavage of aryl ether bonds as suggested by 92%-100% loss of the β-O-4 motif under the same conditions. Because of the well-preserved structure, RFF lignin can be used as a good feedstock to boost its downstream valorization, especially for hydrogenolysis into monophenolic chemicals and fuels. It is noteworthy that the carbohydrate fraction from RFF retained structural integrity and almost reached theoretical yields for glucan and xylan.

Full text of DOI:10.1039/c9gc02315a

View Article Online
View Journal
Green
Chemistry
Cutting-edge research for a greener sustainable future
Accepted Manuscript
This article can be cited before page numbers have been issued, to do this please use: H. Zhou, J. Xu, Y.
Fu, H. Zhang, Z. Yuan, M. Qin and Z. Wang, Green Chem., 2019, DOI: 10.1039/C9GC02315A.
Volume 20
This is an Accepted Manuscript, which has been through the
Number
May 2018
Pages 1919-2160
9
7
Royal Society of Chemistry peer review process and has been
accepted for publication.
Green
Chemistry
Cutting-edge research for a greener sustainable future
rsc.li/greenchem
Accepted Manuscripts are published online shortly after acceptance,
before technical editing, formatting and proof reading. Using this free
service, authors can make their results available to the community, in
citable form, before we publish the edited article. We will replace this
Accepted Manuscript with the edited and formatted Advance Article as
soon as it is available.
You can find more information about Accepted Manuscripts in the
Information for Authors.
Please note that technical editing may introduce minor changes to the
text and/or graphics, which may alter content. The journal’s standard
Terms & Conditions and the Ethical guidelines still apply. In no event
shall the Royal Society of Chemistry be held responsible for any errors
or omissions in this Accepted Manuscript or any consequences arising
from the use of any information it contains.
ISSN 1463-9262
PAPER
Paul T. Anastas et al.
The Green ChemisTREE: 20 years after taking root with the 12
principles
rsc.li/greenchem

Page 1 of 8
Green Chemistry
View Article Online
DOI: 10.1039/C9GC02315A
COMMUNICATION
Rapid Flow-Through Fractionation of Biomass to Preserve Labile
Aryl Ether Bonds in Native Lignin
Hao Zhou,[a] Jia Yun Xu,[a] Yingjuan Fu,[a] Haiguang Zhang,[a] Zaiwu Yuan,[a] Menghua Qin,*[b]
Zhaojiang Wang*[a]
Abstract :Lignin is the second largest component of vascular plants chemicals and fuels. It is noteworthy that carbohydrate fraction
and is the most abundant renewable aromatic polymer on our from RFF retained structural integrity and almost reached
theoretical yields for glucan and xylan.
planet. The attractiveness of lignin valorization lies in the
conversion into high value aromatic chemicals and biofuels
through fractionation and upgrading. Literatures demonstrated
the presence of aryl ether bonds in native lignin was a key factor
for the conversion, while the conventional technical lignins from
carbohydrate-first processes, e.g. pulp and cellulose ethanol
production, are intensively condensed and lack these linkages due
to the intense delignification conditions. Here, by using β-O-4
lignin model dimmer GG, we reveal the dramatic degradation of
GG and the synchronous formation of ralatively stable
intermediate β-O-4 dimers, C6C3 enol ether and the formylated
enol ether, within the first 5 min under the condition of 72wt%
aqueous formic acid and 130 °C, a condition suitable for biomass
fractionation. Based on these findings, we propose a simple but
effective strategy of rapid flow-through fractionation (RFF), which
separates the dissolved lignin from reactor in time and space,
thereby preserving these labile aryl ether bonds in native lignin.
Application of RFF of poplar wood with a short residence time of
2.6 min attained 75% delignification with an equivalent of β-O-4
motif in native lignin. Structure-preserved lignins (75.0%-85.4% β-
O-4 retention) were also harvested from wheat straw with good
lignin yields (61.7%-78.5%). Contrarily, batch fractionation acted
as a protracted war and resulted in extensive cleavage of aryl
ether bonds as suggested by 92%-100% loss of β-O-4 motif under
the same conditions. Because of the well-preserved structure, RFF
lignin can be used as good feedstock to boosting its downstream
valorization, especially for hydrogenolysis into monophenolic
Lignocellulosic biomass is a renewable organic resource from
plants photosynthesis, and is chemically composed of
cellulose, hemicellulose and lignin. Lignin is an aromatic
polymer formed in nature by the radical polymerisation of p-
coumaryl alcohol, coniferyl alcohol and sinapyl alcohol.1, 2 The
radical polymerisation process leads to a variety of inter-unit
linkages including the β-O-4, β–β, β-5, 5–5 and 4-O-5 structural
motifs. Their relative abundance in native lignin varies from
plant to plant but the β-O-4 aryl ether bond is typically
predominant among these inter-unit linkages.3, 4
Although a highly abundant aromatic feedstock, lignin is still
treated as a waste product and commonly burned to supply
heat and energy in carbohydrate-oriented processes, e.g. pulp,
cellulose ethanol, xylose, and furfural production due to its
chemical recalcitrance and structural complexity. Valorization
of lignin is increasingly recognized as being crucial to the
economic viability of integrated biorefinery. Transformation of
lignin into aromatic chemicals and drop-in fuels through
depolymerization and upgrading is now attracting much
attention.5-7 Literatures demonstrated the presence of aryl
ether bonds in native lignin was a key factor for the conversion
of lignin to aromatic monomers.6, 8 Unfortunately, chemical
structure of these technical lignins from the existing
carbohydrate-oriented processes is highly modified.
Specifically, under the intense delignification conditions, lignin
aryl ether bonds are cleaved, and stable carbon-carbon bonds
are formed as result of condensation.2
In response, biomass fractionation strategies that are capable
of preventing structural lignin degradation are being
developed with the expectation of efficient and selective
a. H.Zhou, J. Xu, Prof. Y. Fu, Dr. H. Zhang, Prof. Z. Yuan, Prof. Z. Wang*
State Key Laboratory of Biobased Material and Green Papermaking
Qilu University of Technology, Shandong Academy of Sciences
3501 Daxue Rd, Changqing District, Jinan, 250353(China)
E-mail: wzj@qlu.edu.cn
lignin-to-aromatic conversion.
A
new liquid ammonia
b. Prof. M. Qin
pretreatment methodology called extractive ammonia (EA)
9
Laboratory of Organic Chemistry
Taishan University
was developed by Costa Sousa et al.,
which can produce
lignin with well preserved functionalities but limited lignin
yield lower than 50%. Study from Huy Quang Lê et al. showed
the delignification is more than 90% from a γ-valerolactone
(GVL)/water binary mixture treatment. However, the isolated
525 Dongyue Street, daiyue District, Taian, 271021(China)
E-mail: qmh@qlu.edu.cn
Electronic Supplementary Information (ESI) available: [details of any
supplementary information available should be included here]. See
DOI: 10.1039/x0xx00000x

Green Chemistry
Page 2 of 8
COMMUNICATION
Journal Name
lignin only contain 12% β-O-4 per C9 motif.10 These passive HMV (Fig. 1A, Fig. S5, S8). C6C3 enol-ether (EE), 3-(4-hydroxy-3-
View Article Online
DOI: 10.1039/C9GC02315A
strategies demonstrate trade off between lignin yield and methoxyphenyl)-2-(2-methoxyphenoxy)-2-propenol,
was
structure integrity, which means that high aryl-ether bond identified by GC-MS as the primary β-O-4 intermediate dimer
contents go hand in hand with low isolated lignin yields, and of GG acidolysis according its mass spectrum (Fig. S9). EE was
vice versa.11 An alternative concept of particular interest, reported by Satoshi Kubo and Shuji Hosoya as a stable product
called active stabilisation, is emerging, that relies on active from GG degradation in ionic liquid 30. Formylation of EE
lignin stabilisation and thereby prohibits the problem of lignin occurred at γ-OH as verified by the formation of 3-(4-hydroxy-
condensation during biomass fractionation. Shuai et al. 3-methoxyphenyl)-2-(2-methoxyphenoxy)-2-propenol methyl
reported a novel lignin stabilization strategy by applying ester (fγ-EE) from analysis of mass spectrum in Fig. S10. This
formaldehyde during an acid-catalysed delignification process finding is consistent with the study of Stahl et al 15. The time
to form relatively stable acetals (i.e., 1,3-dioxane structure) course study demonstrated a quick degradation of GG with a
with lignin’s α- and γ-OH side-chain groups.12 Deuss et al. reduction of 58 % in 5 min, and a complete consumption in 60
reported an elegant solution to suppress lignin condensation min (Fig. 1C), as determined by GC-MS (Fig. S5). The
by capturing lignin-derived unstable aldehyde products by intermediate dimer EE and fγ-EE were stable in aqueous formic
reaction with diols to form acetal structures.13, 14 Two step acid as indicated by the yield of 38% at 5 min and 50% at 60
strategies
consisting
of
oxidation
and min (Fig. 1C). Degradation of GG consists of synchronous
depolymerization/hydrogenation also succeed in attaining formation of β-O-4 intermediate dimmers, depolymerization
remarkable aromatic monomers yields due to effective into monomers, and repolymerization into condensed
suppressing the undesired condensation reactions.15-17 products. Yields of monomers, dimmers, and condensed
However, these active lignin stabilization strategies require products were quantified using peak area of GPC (Fig. 1 C, Fig.
extra chemicals or catalysts. Downsides also include underside S4). Time course study showed that approximately 80%
side reactions, i.e. grafting of formaldehyde on carbohydrate bonds (sum of GG and EE) were preserved at first 5 min. The
in the case of formaldehyde stabilization and formation of quantity of -O-4 decreased further to 44% at 60 min, as
alkyl sugars. suggested by the yields of GG and EE dimmers (Fig. 1C). The
Different from the previous defending strategies, i.e. lignin gradually decrease of -O-4 dimers is ascribed to
β-O-4
β
β
stabilization using capping agent, here we contemplate a depolymerization and condensation reactions. Condensation
simple but effective strategy to preserve lignin structures by reaction was aggressive and accounted for 8% and 35% of GG
employing rapid flow-through formic acid on the basis of the at 5min and 60 min, respectively. Condensation involves
plethora of information about formic acid fractionation over repolymerization of intermediate dimmers, monomeric
the last few decades.18-22 Rapid flow-through fractionation products from acidolysis of GG via the α-benzyl carbocation by
(RFF) offers a large surface-to-volume ratio, good mixing and its electrophilic aromatic substitution on the electron-rich
heat transfer properties, and therefore can accelerate the aromatic rings.31, 32 The condensation mechanism is verified by
speed of delignification and reduce the lignin condensation.23- the formation of CD-I and CD-II as identified by 2D HSQC NMR
27
The most important is the ability of RFF in separating the (Fig. 2), and consistent with other studies.33-35 Theoretically,
formic-acid-dissolved lignin fraction in time and space, thereby the above-discussed formylation at
quenching lignin condensation reaction and protecting liable possibility of carbon-carbon bonds formation at C
C
α
eliminates the
. However,
α
aryl ether bonds. Expanding the arsenal of biomass formylation is reversible, and the formyl group is not stable
fractionation methods could play an important role in enough in acid condition. Besides acting as acidic catalyst in
developing viable lignin valorization method within future the depolymerization reaction, formic acid plays the role of
biorefinery. Here, we describe the significance of RFF for lignin reductant as indicated by the hydrodeoxygenation of PHHM
valorization via revisiting the cleavage of aryl ether bonds in into PMP (Fig. 1B). The mechanism of the reduction reaction of
aqueous formic acid, evaluating time effect on lignin PHHM to PMP under treatment with formic acid can be
degradation, and our successful application of RFF in extracting interpreted by Eschweiler-Clarke reaction (Fig. S7a) or two
high quality lignin with good yields.
steps reaction involving transfering hydride from formic acid to
Oxidation
Guaiacylglycerol-β-guaiacyl ether (GG), a phenolic model lignin ketone and further reduciton of diols to PMP 36
.
dimer with a methoxy substituent on each of the phenyl rings, reaction was observed that HMV was converted in to DMP
was used to investigate the cleavage of β-O-4 aryl ether bond with formaldehyde as by-product (Fig. 1B, Fig. S7b). The
and the formation of intermediate dimmers and lignin oxidation reaction could interpret the absence of HMV in
monomers in 72wt % aqueous formic acid at 130 °C for products mixture from GG acidolysis using formic acid, while
different time. The conditions were chosen because it is HMV was identified as the main product of GG acidolysis using
37
suitable and widely applied for biomass fractionation and sulphuric acid
and hydrochloric acid.35 Oxidative product
Organosolv pulping.18, 28, 29 Reaction pathways were proposed DMP is then converted into BDME as result of adding a
based on analysis of resultant products from GC/MS formaldehyde molecule (Fig. S7c), or converted into VAN as
determination (Fig. 1, Fig. S5). The acknowledged acid catalytic result of eliminating a formaldehyde molecule (Fig. 1B). Totally,
mechanism was confirmed that the aryl ether bond cleaves the sum yield of all identified monomeric phenolics is
along with the formation of a new-formed phenolic compound approximately 21% at 60 min (Fig. 1C).
GOH and Hibbert’s ketone-type substructures, PHHM and

Page 3 of 8
Green Chemistry
Journal Name
COMMUNICATION
for GG degradation at 5 min, respectively. Totally,VGiewGA,rtfi-cGleGO,nlfinγe-
DOI: 10.1039/C9GC02315A
EE and CD-II kept 83% of β-O-4 in untreated GG at 5 min,
consistent with the result in Fig. 1C from GPC analysis. 2D NMR
confirmed the formation f-GG, fγ-EE, CD-I, CD-II. Most
importantly, the dependence of
again verified.
β-O-4 on treatment time was
Figure 1. Possible Reaction pathways of GG in 72w% aqueous formic acid at 130°C, (A)
Acid catalyzed cleavage of aryl ether bond and the formylation of GG at γ-OH, (B)
Reduction and oxidation reactions of resultant monomers from acidolysis, (C) Yield of
monomers, dimmers (GG + EE), and condensed compounds from GG degradation in
time course.
2D HSQC NMR was used to study the cleavage of aryl ether
bonds and synchronous condensation reactions (Fig. 2, Table
S5). Cleavage product PHHM was identified by 2D NMR. The
assignment of the Hibbert’s ketone PHHM was made by the
cross peaks at δC/δH 44.5/3.67 ppm (α-position) and 67.1/4.19
ppm (γ-position) according to the study of Miles-Barrett et al.38
It is worth noting that esterification reaction between GG and
formic acid occurred to both aliphatic-OH at Cα and Cγ, and
phenolic-OH at the first 5 min of reaction, leading to formation
39
of totally formylated GG (f-GG), as revealed by 2D NMR.
However, formyl groups at Cα and phenol-C4 are not stable as
indicated by the diminishing peaks at 2D NMR spectra (not
shown). Consistent with GC-MS analysis, fγ-EE was identified
again by 2D NMR with the assignment of δC/δH 120.6/6.33
ppm at α position. Condensation products CD-I and CD-II were
also unveiled by 2D NMR (Fig. 2) and LC-MS (Fig. S6). The
structures of CD-I and CD-II are consistent with the well-
acknowledged mechanism that condensation mostly occurred
via the α-benzyl carbocation by its electrophilic aromatic
substitution on the electron-rich aromatic rings, forming a new
carbon-carbon linkage. The participation of GOH in formation
of condensed products interprets the descending trend of GOH
along with reaction (Fig. S11). The identification of
condensation products brings new insight into the competition
between the depolymerization and condensation via the α-
benzyl carbocation intermediate as catalyzed by aqueous
formic acid. Carbon-carbon linkages in CD-I and CD-II show
characteristic correlation signals from the α-position at δC/δH
50.8/4.18 and 48.9/4.40 ppm, respectively. Signal intensity at
α-position was used to quantify GG, f-GG, fγ-EE and CD-II
during reaction with the normalization of untreated GG to 100
(Fig. 2). Quantity of GG decreases to 41.9 at 5 min of reaction.
Products f-GG, fγ-EE and CD-II account 10.2%, 24.5% and 6.9%
Figure 2. HSQC NMR spectra of untreated GG, and reaction products at 5 min, 130 °C,
72wt % aqueous formic acid. Contours are colour coded according to the structure they
are assigned to. Gray cross peaks correspond to currently unassigned signals.
Figure 3. Comparison of batch and flow-through reaction system in lignin chemistry,
Top: homogeneous batch reaction of lignin model dimmer (guaiacylglycerol-β-guaiacyl
ether) in 72wt% formic acid at 130 °C results in 20-57% cleavage of aryl ether bonds in
5-60 min; Bottom: heterogeneous rapid flow-through fractionation of lignocellulose in

Green Chemistry
Page 4 of 8
COMMUNICATION
Journal Name
72wt% formic acid at 130 °C attains 50-94% lignin yield and 78-99% retention of aryl
ether bonds due to the significantly reduced residence time of 2.6 min.
reactor is fixed and equal to the reaction timVeie.wTAhrtiicrlde ,Onflioner
heterogeneous reaction, mass transfer oDf OflIo: w10.1r0e3a9c/tCo9rGiCs0m23u1c5hA
more efficient than batch reactor due to the elevated
concentration gradient in the surface of solid reactant.27 The
advantage of RFF in preserving lignin structures was confirmed
Lignocellulosic biomass is chemically composed by cellulose,
hemicellulose and lignin. Components fractionation is the
inevitable and crucial step for subsequent biorefinery
upgrading. The conventional alkaline cooking for pulp
production and dilute acid pretreatment for cellulosic ethanol
production focus only on carbohydrate and therefore employ
forcing conditions for a maximized delignification, which
results in invertible degradation of lignin in terms of
depolymerization and condensation. To facilitate the
production of lignin monomer through reductive catalytic
by a comparison with lignins from batch treatments using
41, 42
alkali 41, ethanol 2, and formic acid
(Fig. 4). Last but not
least, esterification reaction between lignin and formic acid
leads to formation of -O-formyl group at Cα, eliminating the
possibility of the acidolysis reaction and the subsequent
cleavage of β-O-4. Formation of -O-formyl group at Cα also
suppresses condensation reactions at Cα. Clearly, if the goal of
lignin valorizaiton is production of aromatic hydrocarbon,43, 44
lignin from RFF is a good alternative for an easier catalytic
upgrading.
processes, extracting
a stream of lignin with native
uncondensed structure is very important. The above
discussions suggest that the cleavage of aryl ether bonds is
dependent on treatment time in homogeneous aqueous
formic acid reaction system, with 20% loss at 5 min, 57% loss
at 60 min (Fig. 1C). Furthermore, amount of new-formed
condensed structures with carbon-carbon linkages increase
with treatment time, with yields of 9% at 5 min, 35% at 60 min
(Fig. 1C). For true biomass fractionation in heterogeneous
system, our strategy is to attempt to shorten the residence
time of lignin in reactor with an elaborate flow-through
reactor, in which the flowing solvent takes the dissolved lignin
from lignocellulose out of reactor in a really short time
depending on flow rate (Fig. 3). The application of rapid flow-
through fractionation (RFF) using 72wt % aqueous formic acid
as solvent with a residence time of 2.6 min and total running
reaction time of 10 min demonstrates the ability of extract
structure-preserved lignin from wheat straw with good yields
from 61.7% to 78.5% (Table 1, Fig. S1). The lignin dissolution in
flow-through reactor showed a rapid decline trend and almost
finishes within 8 min (Fig. 3 bottom right). Noteworthy is the
ability of preserving lignin structure as suggested by the good
retention rates of β-O-4 aryl ether bonds from 75.0% to 85.4%
compared with milled wood lignin (i.e. WS/MWL) depending
on treatment severities (Table 1). In contrast, only 9.2% β-O-4
linkages retains for lignin from batch fractionation at the same
conditions (WS/F72T130t10/Batch). RFF also showed excellent
performance for poplar wood. Lignin yield attained
50.55~94.2% with remarkable retention rates of β-O-4 aryl
ether bonds from 77.9~99.6% under various conditions (Table
1). By contrast, no β-O-4 aryl ether bonds survive for lignin
from batch fractionation under the same conditions
(Poplar/F72T130t30/Batch). Take delignification ability into
Fig. 5 shows the comparison of HSQC spectra of the milled
wood lignin (MWL), and lignins from RFF and batch
fractionation under the same conditions. Main inter-unit lignin
linkages (A: β-O-4, B: β-β, C: β- 5) and substructures (I, FA, PCA,
G, H, S, PB, T) are assigned to correlation peaks accordingly.17,
45, 46
MWL is obtained by solvent extraction (dioxane:H2O,
96v:4v) and commonly used as a representative of native
lignin.47 There are no obvious difference between MWL and
RFF lignin (WS/F72T130t10/RFF, Poplar/F72T130t10/RFF) in
aryl ether linkages (A, A’). However, crosspeaks of aryl ether
linkages (A, A’) decreased significantly or dispeared for lingin
from
batch
treatments
(WS/F72T130t10/Batch,
Poplar/F72T130t30/Batch). Signals from guaiacyl (G), syringyl
(S), and p-hydroxyphenyl (H) units are observed almost
equivalently in the spectra of the MWL and RFF lignin. In
contrast, strong correlation peaks of condensed S were
observed
for
lignin
from
batch
treatment
of
Poplar/F72T130t30/Batch, which means formation of carbon-
carbon bonds. Formyl groups were identified both for lignin
from batch and flow-through reaction system using formic acid,
but not for native MWL lignin (Fig. 5). Formylation makes lignin
soluble in formic acid and accelerates the delignificaiton
process.
Table 1. Lignin yield and amount of β-O-4 aryl ether bonds of MWL, RFF lignin and
batch lignin from wheat straw and poplar wood under various conditions
Lignin yield
Samples
β-O-4 (A/A')c
(%)b
WS/MWL
44.4
WS/F72T120t10/RFFa
WS/F72T130t10/RFF
WS/F72T140t10/RFF
WS/F72T130t10/Batch
Poplar/MWL
61.7
72.4
78.5
73.8
37.9 (85.4)d
36.5 (82.2)
33.3 (75.0)
4.1 (9.2)
account, formic-acid-RFF showed comparative performance to
40
acid-hydrotrope-RFF
in protecting lignin structures from
degradation (Fig. 4). This ability can be explained from multiple
points of view. First, lignin dissolution is a fast diminishing
process as indicated by the plot of reaction time against
concentration of solubilized lignin in Fig. 3. This means a
majority of lignin flows out of reactor and therefore suffers
less degradation than lignin in batch reactor. Second,
residence time of the solubilized lignin in flow reactor can be
infinitely shortened to reduce lignin degradation in reactor by
increasing the flow rate, while the residence time for batch
46.1
Poplar/F72T130t10/RFF
Poplar/F72T120t30/RFF
Poplar/F72T130t30/RFF
Poplar/F72T140t30/RFF
Poplar/F72T130t30/Batch
50.5
76.0
90.3
94.2
91.7
45.9 (99.6)
42.2 (91.5)
38.4 (83.3)
35.8 (77.7)
NDe

Page 5 of 8
Green Chemistry
Journal Name
COMMUNICATION
[a] WS is wheat straw; numbers after F, T, t are concentration of formic acid, treatment
temperature, and total running time, respectively. [b] lignin yield is calculated using the
equation ‘100 - 100 × Lsr × Ysr /Lub ’, where Lsr is lignin content in solid residue, Ysr is
yield of solid residue, and Lub is lignin content in untreated biomass. [c] Expressed per
100 Aromatic units based on the relative volume integrals of characteristic peaks in 2D-
HSQC spectra using computational formula in Table S1 and Table S2. [d] Numbers in
parentheses are retention rate of β-O-4 on basis of MWL. [e] Not detected.
observed due to the short residence time of soluVbielwe AsruticgleaOrsnliinne
reactor. The majority of xylan is dissolveDdOoI:u1t0.1o0f3b9/ioCm9GaCs0s2i3n1t5oA
liquor as indicated by the xylose yield of 88.3% and 79.4% for
poplar wood (Poplar/F72T130t30/RFF) and wheat straw
(WS/F72T130t10/RFF), while more than 95% glucan remains in
solid. The delignification ability of RFF is reflected by the
soluble lignin yield of 90.3% and 72.4% for poplar wood and
wheat straw, respectively (Fig. 6). The mass balance data for
other treatment conditions were provided in Table S3 and
Table S4. Due to the good selectivity toward non-cellulose
components, the cellulose-rich solid residue from RFF
treatment can be utilized for paper and paperboard
production. The soluble substances, e.g. xylose and lignin, can
be separated from solvent by spray dry. The condensed
solvent can be reused for RFF. Powders from spray dry process
mainly consist of sugars and lignin. These sugars can be
converted selectively to biofuels through biotransformation or
furfural through dehydration, or purified for sugars production
through redissolution-crystallization in water since lignin is
water insoluble. The structure integrity of sugars and lignin
from RFF ensures good compatibility to current biorefinery
schemes. The obtained lignin can then be converted to high
value dispersant or surfactant through sulfonation, or low
molecular phenol compouds by catalytic hydrogenolysis. These
flexible lignin valorization appoaches will enhance the process
economic competitiveness of RFF.
55
50
Flow-through fractioantion
45
Formic acid, poplar wood
40
35
30
25
20
15
10
5
Formic acid, wheat straw
p-TsOH, poplar wood
Batch fractioantion
Alkali, poplar wood
Ethanol, wheat straw
Ethanol, poplar wood
Ethanol, Spruce
Formic acid, bamboo
Formic acid, wheat straw
Formic acid, poplar wood
0
20
30
40
50
60
70
80
90
100
Delignification (%)
Figure 4. Correlation between delignification and β-O-4 linkages.
Continuous RFF showed nearly theoretical recovery of
carbohydrate from the mass balance analysis (Fig. 6). No
dehydration products of carbohydrate, e.g. furfural from
pentose and 5-hydroxymethylfurfural from hexose, were

Green Chemistry
Page 6 of 8
FeiCtghuOerreMs 5w.MitHhSUQacNCylsaIpCteeAcdtrTaanIoOdffNtohremmylaillteeddwγ-oOoHd;li(gBn)inph(MenWylcLo),uamnadrlaignns;in(sC)frroemsinRoFlFs;a(nFd) sbpaitrcohdiferancotnioensa; t(iIo)nc.inMnaaminysltarulccotuhroelsepnrde-sgernotuparse;:(F(AA)) βfe-Oru-l4a′taelsk;y(lP-aCrAy)l ept-hceorusm; J(aAor′a)utβer-snO; -(a4Gl′)aNglkuyaal-imaacryyell
units; (H) p-hydroxyphenyl units; (S) syringyl units. (S′) oxidized syringyl units with a Cα ketone; (T) tricin. (PB) p-hydroxybenzoate substructures. See Table S1 for details of signal
assignments.
View Article Online
DOI: 10.1039/C9GC02315A
Figure 6. Mass balance of polysaccharide (glucan and xylan) and lignin fractions of biomass. (A) Poplar wood (B) wheat straw, the standard deviation of determination is
±2%.
Conflicts of interest
There are no conflicts to declare
Experimental Section
Degradation of GG in aqueous formic acid
Stock solution of GG was prepared by dissolving GG (10 mg) in acetone (10 ml).
Stock solution (1.0 ml) was introduced into a glass vial. Solvent acetone was
evaporated using stream of N2. Aqueous formic acid (2 ml, 72wt %) was added to
the vial to dissolve GG, which gives a GG concentration of 0.5 mg /ml. The vial
was sealed, and incubated at 130 °C for 60 min in an oven. Reaction was stopped
at pre-set time by taking the vial out from the oven and placing it in icy water.
The resultant products were dried through evaporation of aqueous formic acid
using stream of N2. Acetone (1.0 ml) was added to the vial to dissolve the
resultant products for GC/MS analysis.
Acknowledgements
This work was supported by the Natural Science Foundation of
China (31570571) and Key Research and Development
Program of Shandong Province (2016GGX108002).
Notes and references
Flow-through and batch fractionation of biomass
1.
2.
R. Vanholme, B. Demedts, K. Morreel, J. Ralph and W.
Boerjan, Plant physiology, 2010, 153, 895-905.
S. Constant, H. L. Wienk, A. E. Frissen, P. de Peinder, R.
Boelens, D. S. Van Es, R. J. Grisel, B. M. Weckhuysen, W. J.
Huijgen and R. J. Gosselink, Green Chemistry, 2016, 18,
2651-2665.
The RFF system consists of a solvent bottle, an HPLC pump (LC-10AT, Shimadzu,
Corp. Japan), an extraction column (300 mm length × 8 mm diameter, 15 ml
volume), a temperature adjustable silicone oil bath, a bottle for liquid sample
collection, and stainless steel tubes connecting them, as illustrated by Fig. S1.
Aqueous formic acid (72wt%) was used as solvent. Biomass (3.0 g) was loaded in
the extraction column. The remaining space of the extraction column for flowing-
solvent after biomass loading is 13 ml since biomass takes up approximately 2.0
ml (solid density of biomass 1.5 g/ml). After end-fittings installation and tubes
connection, pump was started at 10 ml/min for a fast filling-in of the extraction
column with solvent. Pump was then stopped, and the extraction column was
placed in an oil bath at pre-determined temperature for 5 min to allow heat
transfer and liquid diffusion. RFF of biomass was conducted at a flow rate of 5
ml/min (residence time 2.6 min) for 10 min, or at a flow rate of 1.67 ml/min
(residence time 7.8) for 30 min. RFF consumes totally 63 ml (50 ml +13 ml)
solvent, which corresponds to solid/solvent of 1:21. Extract was collect from the
outlet of RFF reactor at room temperature. Solid residue of biomass in the
column was washed by pumping 100 ml of water through the column, and then
taken out of column, stored at 4 ºC for further analysis.
3.
4.
J. S. M. A. Luterbacher, D.; Dumesic, J. A., Green
Chemistry, 2014, 16, 4816-4838.
J. Ralph, K. Lundquist, G. Brunow, F. Lu, H. Kim, P. F.
Schatz, J. M. Marita, R. D. Hatfield, S. A. Ralph and J. H.
Christensen, Phytochemistry Reviews, 2004, 3, 29-60.
W. Schutyser, T. Renders, S. Van den Bosch, S.-F.
Koelewijn, G. T. Beckham and B. F. Sels, Chem. Soc. Rev.,
2018, 47, 852-908.
R. Rinaldi, R. Jastrzebski, M. T. Clough, J. Ralph, M.
Kennema, P. C. Bruijnincx and B. M. Weckhuysen,
Angewandte Chemie International Edition, 2016, 55,
8164-8215.
5.
6.
7.
8.
Z. Sun, B. l. Fridrich, A. de Santi, S. Elangovan and K. Barta,
Chem. Rev., 2018, 118, 614-678.
Batch fractionation was performed in a 120 ml pressure flask in an oil bath at
temperatures of 130 °C. Biomass (3 g) and 72wt % aqueous formic acid (63 ml)
were introduced into the flask with a biomass/liquid of 1/21 (w/v). The flask was
shaken every 5 min during fractionation. At the end of batch fractionation, flask
was taken out from the bath and placed in icy water to quench the reaction.
Extract was then separated from solid residue of biomass by vacuum filtration
through a filtering crucible. Solid residue and extract were collected for further
analysis.
F. P. Bouxin, A. McVeigh, F. Tran, N. J. Westwood, M. C.
Jarvis and S. D. Jackson, Green Chem., 2015, 17, 1235-
1242.
L. da Costa Sousa, M. Jin, S. P. S. Chundawat, V. Bokade, X.
Tang, A. Azarpira, F. Lu, U. Avci, J. Humpula, N.
Uppugundla, C. Gunawan, S. Pattathil, A. M. Cheh, N.
Kothari, R. Kumar, J. Ralph, M. G. Hahn, C. E. Wyman, S.
Singh, B. A. Simmons, B. E. Dale and V. Balan, Energy &
Environmental Science, 2016, 9, 1215-1223.
9.

Page 7 of 8
Green Chemistry
Journal Name
COMMUNICATION
10.
11.
H. Q. Lê, Y. Ma, M. Borrega and H. Sixta, Green Chemistry,
2016, 18, 5466-5476.
36.
37.
D. Cao, C. Yan, P. Zhou, H. Zeng and C. J. Li, Chemical
View Article Online
communications, 2019, 55, 767-77D0O. I: 10.1039/C9GC02315A
T. Renders, S. Van den Bosch, S.-F. Koelewijn, W.
Schutyser and B. Sels, Energy & Environmental Science,
2017, 10, 1551-1557.
L. Shuai, M. T. Amiri, Y. M. Questell-Santiago, F. Héroguel,
Y. Li, H. Kim, R. Meilan, C. Chapple, J. Ralph and J. S.
Luterbacher, Science, 2016, 354, 329-333.
P. J. Deuss, M. Scott, F. Tran, N. J. Westwood, J. G. d. Vries
and K. Barta, Journal of the American Chemical Society
2015, 137, 7456-7467.
P. J. Deuss, C. S. Lancefield, A. Narani, J. G. de Vries, N. J.
Westwood and K. Barta, Green Chem., 2017, 19, 2774-
2782.
A. Rahimi, A. Ulbrich, J. J. Coon and S. S. Stahl, Nature,
2014, 515, 249.
M. R. Sturgeon, S. Kim, K. Lawrence, R. S. Paton, S. C.
Chmely, M. Nimlos, T. D. Foust and G. T. Beckham, ACS
Sustainable Chemistry & Engineering, 2014, 2, 472-485.
D. M. Miles-Barrett, A. R. Neal, C. Hand, J. R.
Montgomery, I. Panovic, O. S. Ojo, C. S. Lancefield, D. B.
Cordes, A. M. Slawin and T. Lebl, Organic & Biomolecular
Chemistry, 2016, 14, 10023-10030.
S. A. Ralph, J. Ralph and L. Landucci, eds., NMR database
of lignin and cell wall model compounds, 2004.
Z. Wang, S. Qiu, K. C. Hirth, J. Cheng, J. Wen, N. Li, Y. Fang,
X. Pan and J. Zhu, ACS Sustain. Chem. Eng., 2019, 7.
H. Zhou, L. Tan, Y. Fu, H. Zhang, N. Liu, M. Qin and Z.
Wang, ChemSusChem, 2019, 12, 1213-1221.
Y. Zhang, Q. Hou, W. Xu, M. Qin, Y. Fu, Z. Wang, S. Willför
and C. Xu, Industrial Crops and Products, 2017, 108, 864-
871.
H. Guo, B. Zhang, Z. Qi, C. Li, J. Ji, T. Dai, A. Wang and T.
Zhang, Chemsuschem, 2017, 10, 523-532.
L. D. C. Sousa, M. J. Jin, S. P. S. Chundawat, V. Bokade, X.
Y. Tang, A. Azarpira, F. C. Lu, U. Avci, J. Humpula and N.
Uppugundla, Energy & Environmental Science, 2016, 9,
1215-1223.
T.-Q. Yuan, S.-N. Sun, F. Xu and R.-C. Sun, J. Agric. Food.
Chem., 2011, 59, 10604-10614.
J. C. D. Río, J. Rencoret, P. Prinsen, Á. T. Martínez, J. Ralph
and A. Gutiérrez, Journal of Agricultural & Food
Chemistry, 2012, 60, 5922.
12.
13.
14.
38.
39.
40.
41.
42.
15.
16.
17.
C. Zhang, H. Li, J. Lu, X. Zhang and W. Feng, Acs Catalysis,
2017, 7, 3419-3429.
C. S. Lancefield, O. S. Ojo, F. Tran and N. J. Westwood,
Angewandte Chemie International Edition, 2015, 54, 258-
262.
U. Baeza, D. M. Erismann and S. Freer, Bioresour.
Technol., 1991, 37, 1-6.
B. M and J. RK, Pulp & Paper, 1983, 57, 102-104.
S. ap a, . Santos and . Para , Biomass Bioenergy,
2002, 22, 213-221.
J. Snelders, E. Dornez, B. Benjelloun-Mlayah, W. J. J.
Huijgen, P. J. D. Wild, R. J. A. Gosselink, J. Gerritsma and C.
M. Courtin, Bioresource technology, 2014, 156, 275-282.
F. Abdelkafi, H. Ammar, B. Rousseau, M. Tessier, R. El
Gharbi and A. Fradet, Biomacromolecules, 2011, 12, 3895-
3902.
43.
44.
18.
19.
20.
45.
46.
21.
22.
47.
A. Björkman, Nature, 1954, 174, 1057.
23.
24.
E. M. Anderson, M. L. Stone, R. Katahira, M. Reed, G. T.
Beckham and Y. Román-Leshkov, Joule, 2017, 1, 613-622.
I. Kumaniaev, E. Subbotina, J. Sävmarker, M. Larhed, M. V.
Galkin and J. S. M. Samec, Green Chemistry, 2017, 19,
5767-5771.
25.
26.
27.
V. Jafari, H. Sixta and A. van Heiningen, Industrial &
Engineering Chemistry Research, 2014, 53, 8385-8394.
V. Jafari, K. Nieminen, H. Sixta and A. van Heiningen,
Cellulose, 2015, 22, 2055-2066.
E. M. Anderson, M. L. Stone, M. J. Hülsey, G. T. Beckham
and Y. Román-Leshkov, ACS Sustainable Chemistry &
Engineering, 2018, 6, 7951-7959.
28.
29.
30.
J. Xu, Y. Fu, G. Tian, Q. Li, N. Liu, M. Qin and Z. Wang,
Bioresour. Technol., 2018, 254, 353-356.
M. Zhang, Q. Wei, L. Rui, R. Su, S. Wu and Z. He, Biomass
& Bioenergy, 2010, 34, 525-532.
S. Kubo, K. Hashida, T. Yamada, S. Hishiyama, K. Magara,
M. Kishino, H. Ohno and S. Hosoya1, Journal of wood
chemistry and technology, 2008, 28, 84-96.
L. Ning, Y. Li, G. Y. Chang, X. Yang, X. Lin, J. Ralph and X.
Pan, Green Chem., 2018, 20, 4224-4235.
L. Shuai, M. T. Amiri and J. S. Luterbacher, Current Opinion
in Green and Sustainable Chemistry, 2016, 2, 59-63.
T. Yokoyama and Y. Matsumoto, Holzforschung, 2008, 62,
164-168.
31.
32.
33.
34.
R. M. Ede and G. Brunow, Holzforschung-International
Journal of the Biology, Chemistry, Physics and Technology
of Wood, 1989, 43, 317-322.
35.
K. Lundquist and R. Lundgren, ACTA Chem. Scand., 1972,
26, 2005-2023.

Green Chemistry
Page 8 of 8
Graphical Abstract
View Article Online
Hao Zhou,[a] Jia Yun DXOuI,:[a1]0Y.1i0n3g9ju/Ca9nGFCu0,2[a3]15A
Haiguang Zhang,[a] Zaiwu Yuan,[a]
Menghua Qin,*[b] Zhaojiang Wang*[a]
Rapid Flow-Through Fractionation of
Biomass to Preserve Labile Aryl Ether
Bonds in Native Lignin
Effective isolation of Lignin Containing Native Aryl Ether Linkages