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Article

Towards the Potential of Using Downstream-Separated Solvents as the Pulping Liquor of Upstream Lignocellulose Fractionation for Enhanced Acetone–Butanol–Ethanol Production

by
Changsheng Su
1,2,3,†,
Yunxing Gao
1,2,†,
Gege Zhang
1,2,
Hao Wen
1,2,
Rui Chen
1,2,
Jiajing Wang
1,2,
Yujie Li
1,2,
Mingyuan Sun
3,
Jikang Cao
1,2 and
Di Cai
1,2,3,*
1
State Key Laboratory of Green Biomanufacturing, Beijing University of Chemical Technology, Beijing 100029, China
2
National Energy R & D Centre for Biorefinery, Beijing University of Chemical Technology, Beijing 100029, China
3
College of Life Science and Technology, Beijing University of Chemical Technology, Beijing 100029, China
*
Author to whom correspondence should be addressed.
†
These authors contributed equally to this work.
Fermentation 2025, 11(9), 514; https://doi.org/10.3390/fermentation11090514
Submission received: 30 June 2025 / Revised: 29 August 2025 / Accepted: 30 August 2025 / Published: 1 September 2025
(This article belongs to the Special Issue Bioprocesses for Biomass Valorization in Biorefineries)
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Abstract

Developing efficient, clean, and sustainable lignocellulose pretreatment technologies is essential for second-generation biofuel production. In this study, we attempted to use downstream-separated binary acetone-water, n-butanol-water, and ethanol-water solutions as the initial liquor for upstream organosolv pulping, in order to achieve the efficient and economic closed-circuit clean fractionation of the lignocelluloses for biological acetone–butanol–ethanol (ABE) production. Parameters, including concentration and temperature of the organosolv pulping, were optimized systematically. Results indicated that the 50 wt% ethanol and 30 wt% acetone aqueous solutions and pulping at 200 °C for 1 h exhibited better corn stover fractionation performances with higher fermentable sugar production. The total monosaccharide recovery (including glucose and xylose) was 50.92% and 50.89%, respectively, in subsequent enzymatic saccharification. While pulping corn stover using n-butanol solution as initial liquor showed higher delignification 86.16% (50 wt% of n-butanol and 200 °C for 1 h), the hydrolysate obtained by the organosolv pulps always exhibited good fermentability. A maximized 15.0 g/L of ABE with 0.36 g/g of yield was obtained in Ethanol-200 °C-50% group, corresponding to 112 g of ABE production from 1 kg of raw corn stover. As expected, the lignin specimens fractionated by closed-circuit organosolv pulping exhibited narrow molecule weight distribution, high purity, and high preservation of active groups, which supports further valorization. This novel strategy tightly bridges the upstream and downstream processes of second-generation ABE production, providing a new route for ‘energy-matter intensive’ and environmentally friendly lignocelluloses biorefineries.

1. Introduction

Biobutanol (n-butanol) is an advanced renewable biofuel and an important bulk chemical [1]. The primary strategy to produce biobutanol is the biotransformation of the carbohydrates via acetone–butanol–ethanol (ABE) fermentation, using the solventogenic Clostridium sp. [2]. In recent years, the production of biobutanol from the abundant lignocellulose biomass, especially agriculture and forest residues, is increasing, as it showed promise in response to the increasingly concerning issues of environmental pollution, climate change, and energy security [3]. From a technical viewpoint, in the whole production chain of biobutanol from lignocellulose, one of the crucial steps is the biomass fractionation to properly depolymerize the complex recalcitrance matrix structure of cell wall, in order to improve the accessibility of cellulase for fermentable sugar production [4]. In fact, lignocellulose fractionation has long been recognized as a high-pollution and economically infeasible process, as it has always required severe acid or alkaline conditions, an energy-intensive heating process, and a low-value lignin-rich side stream [5].
Among the candidate techniques for lignocellulose fractionation, organosolv pulping is recognized as a more economic and cleaner strategy, as it co-generates high purity technical lignin that is capable of valorizing into valuable materials and chemicals [6,7], while the solvents can be easily recycled by vacuum evaporation, thereby eliminating the contamination [8]. Basically, organosolv pulping will dissolve lignin and hemicellulose under relatively mild conditions, leaving insoluble cellulose-rich pulp that is suitable to enzymatic hydrolysis. This process also leads to the extensive degradation of the polymerized native lignin in lignocellulose matrixes, mainly manifested in the formation of benzyl cations under hydrogen ion catalysis [9].
Generally, one of the primary principles for the solvent’s selection for organosolv pulping is volatility, as it is closely related to energy requirement in solvent recycling post-fractionation [10]. Another principle is the solubility of solvent to lignin fractions [7]. Indeed, solvents that possessed similar polarity to lignin molecule always exhibit better fractionation performances [11]. Meanwhile, in a systematic concern, it is important to reduce the complexity of the process and the supply chain of solvents [12]. Therefore, bulk and cheap solvents, especially the biological solvent productions from biorefinery processes, are more favorable in a growing number of reports, which allows additional economic and environment benefits from the closed-circuit streams [13,14].
In this context, for the lignocellulosic biobutanol production, the previous researches have attempted to use binary acetone and ethanol aqueous solutions as the pulping liquor, and realized high biobutanol yield using the corresponding lignocellulose hydrolysate [15,16]. To further improve the hemicellulose and lignin removal rates, researches also suggested to add acid and alkaline catalysts in organosolv pulping process, which further facilitated the hemicellulose and lignin hydrolysis [17]. Meanwhile, when using binary n-butanol solution as pulping liquor, the literature also pointed out that the naturally separated bi-phasic system guaranteed the lignocellulose decomposition while partially preserving lignin recondensation in pulping [18,19]. To shine light on the benefits of bi-phasic n-butanol pulping, Jang et al. combined the reductive catalysis fractionation techniques with n-butanol-water pulping, and obtained high yield of mono-lignols, though noble metal catalyst and hydrogen were required [20]. Moreover, in whole process concerns, Yang et al. used the quaternary ABE-water solution as the pulping liquor for upstream biomass fractionation, and revealed the synergistic effect of different components in initial liquor on lignocellulose depolymerization [21]. Nonetheless, from a technical viewpoint, the existing literature was not comprehensively compared to the effect of A-B-E and water binary solutions that are easily obtained in downstream processes on the efficiency of lignocellulose fractionation and subsequential fermentative ABE production.
This study aims to demonstrate the feasibility of recycling the downstream-separated ABE solvents as pulping liquors for upstream lignocellulose fractionation, thereby establishing a closed-loop biorefinery process for cost-effective and eco-friendly second-generation biobuthanol production. The possibility of using downstream-separated solvents as the initial liquor in upstream processes was highlighted (see Figure 1). After optimization of the type of solvent, the pulping temperature, and the solvent concentration in organosolv pulping process, the enzymatic hydrolysate of the corn stover pulp was directly used as the substrate for ABE fermentation without detoxification. Then, the fractionated lignin specimens in side stream were isolated and characterized to unlock the valorization potentials. Finally, the appropriate upstream pulping strategy was selected by comprehensively comparing the mass balance information.

2. Materials and Methods

2.1. Materials

Corn stover was obtained from a local farm in Qinhuangdao, Hebei province, China. After harvesting, the corn stover was dried out and milled into ~60 meshes. The main chemical components of corn stover were 40.2 ± 1.1% of glucan, 15.1 ± 0.8% of xylan, 18.3 ± 0.6% of acid insoluble lignin (AIL), and 6.1 ± 0.8% of acid soluble lignin (ASL). The organic solvents used in the experiments were under chromatographic grade, and were obtained from the Beijing Chemical Work (Beijing, China). Other chemicals were obtained from Sinopharm Chemical Reagent Co., Ltd. (Shanghai, China).

2.2. Organosolv Pulping

Corn stover bagasse was mixed with different concentrations of binary A-B-E and water solutions under a loading rate of 10% (w/v). The organosolv pulping was conducted in a 300 mL stirred high-temperature reactor (Yanzheng Technology Co., Ltd., Beijing, China) at 120–220 °C for 1 h. After termination of the reaction, the pulp was vacuum filtered and was washed by the initial liquor for 3 times. Then, the pulp was washed with deionized water and dried at 105 °C for 24 h.
For the isolation of the fractionated lignin after organosolv pulping, the pulping liquor was mixed with the post-washing solution, and the pH of mixture was adjusted to 5.5 by 6M HCl. Then, the liquid was concentrated by rotary evaporation, and 3 volumes of 95% ethanol were added. The supernatant of suspension was centrifuged, and was further concentrated. The precipitate lignin can be obtained after adjusting pH to 2.0 by 6M HCl. Finally, the lignin specimens can be obtained after centrifugation, washing, and freeze-drying. Calculations for solid recovery, delignification, and pulp recovery were referred to previous studies [22].

2.3. Enzymatic Hydrolysis

Cellulase was obtained from Shandong KDN Biotechnology Co., Ltd. (Jinan, China). The filter paper enzyme activity of the cellulase was 65 ± 5 FPU/mL. Briefly, the fractionated pulp was mixed with 0.05M citric acid/sodium citrate buffer at 6% (w/v) solid loading, and the pH was adjusted to 4.7. The cellulase loading was 20 FPU/g of glucan. The enzymatic hydrolysis of pulps was conducted in 250 mL of conical flask (with 100 mL working volume), and the temperature and rotation rate were 50 °C and 200 rpm. After 48 h of hydrolysis, the supernatant liquid was obtained by centrifugation. The enzymatic hydrolysis efficiency and monosaccharide yield can be calculated according to the method explained in detail in our previous studies [23,24].

2.4. ABE Fermentation

The strain used for ABE fermentation was the laboratory-preserved Clostridium acetobutylicum ABE1401 (derived from ATCC 824 by adoptive laboratory evolution) [25]. The strains were stored in anaerobic bottles and activated in 3% corn flour medium before inoculation. The seed medium contained glucose 40 g/L, ammonium acetate 2.2 g/L, KH2PO3 1 g/L, K2HPO3 1 g/L, MgSO4 0.2 g/L, MnSO4 0.01 g/L, FeSO4 0.01 g/L, p-aminobenzoic acid 0.001 g/L, biotin 0.0001 g/L.
For the fermentation using the corn stover hydrolysate, nutrients (similar components from seed medium components except glucose) were added into the initial hydrolysate, and pH was adjusted to ~7.0 by ammonium hydroxide. Before inoculation with a size of 10 vol%, the hydrolysates were autoclaved at 116 °C for 30 min. High-purity nitrogen (99.9%) was purged into the sterilized hydrolysate to create a strict anaerobic environment. After inoculation, fermentation was carried out at 37 °Cand 50 rpm.

2.5. Analytical Methods

The chemical composition of raw corn stover and fractionated pulps were determined according to the standard of NREL [26]. Monosaccharides (glucose and xylose) and organic acids (acetic acid and butyric acid) were detected by high-performance liquid chromatography (HPLC, Thermo Fisher U3000, Waltham, MA, USA) equipped with a refractive index detector (RID) and Aminex HPX-87P column (300 × 7.8 mm). The conditions were as follows: column temperature of 65 °C, 5 mM sulfuric acid solution as mobile phase, flow rate of 0.6 mL/min, injection volume of 10 μL [27]. Organic solvents including acetone, n-butanol, and ethanol were detected by gas chromatography (GC, Thermo Fisher Trace 1300, Waltham, MA, USA) equipped with a flame ionization detector (FID) and a packed column (Porapack Q, 80/100 mesh, 2 m ×3 mm) [28].
For the pulps’ characterization, the surface morphology was observed by a scanning electron microscope (SEM, Hitachi S-300N, Minato, Japan). The crystallinity of the pulp was detected by X-ray diffraction (XRD, Rigaku 2500VB2, Akishima, Japan). The scanning range was 5–40°, and the scanning speed was 2 degrees/min. The calculation formula of crystallinity is as follows:
CrI = (I002 − Iam)/I002 × 100%
where I002 is the intensity of the strongest absorption peak around 2θ = 22.5°, and Iam is the intensity of the weaker absorption peak around 2θ = 15.5°.
Fourier transform infrared spectroscopy (FTIR, Thermo Fisher Nicolet 6700, Waltham, MA, USA) was conducted to characterize the chemical bonds. The scanning range was 400–4000 cm−1 and the resolution was 2 cm−1. Thermal stability of pulps was characterized by a thermogravimetric analyzer (TGA-Q500, TA Company, New Castle, DE, USA). Specifically, the specimens were placed in an alumina crucible, and the heating rate was 10 °C /min. The nitrogen flow rate was 30 mL/min, and the temperature range was from room temperature (~25 °C) to 600 °C.
For the lignin characterization, gel permeation chromatography (GPC, Agilent 1200 workstation, Santa Clara, CA, USA) was used to characterize the molecular weight of lignin specimens, equipped with an HT5 column (7.8 mm × 300 mm, Waters Styragel, Milford, MA, USA) and a refractive index detector. The injection volume is 10 μL, and the flow rate of tetrahydrofuran, the eluent, was 1 mL/min. Heteronuclear single quantum coherence (HSQC) was measured using Magnet system 400 MHz (Bruker, Karlsruhe, Germany). Specifically, lignin specimens were dissolved in deuterated dimethyl sulfoxide (DMSO-d6) and scanned from 13C and 1H dimensions, with scanning intervals of 18,000 Hz and 5000 Hz, respectively. In addition, the FTIR characterization and thermal stability characterization of lignin specimens were generally consistent with the methods for pulp characterization.

3. Results and Discussions

3.1. Organosolv Pulping Using Binary A-B-E and Water Solution

Firstly, the possibility of using binary A-B-E aqueous solution as pulping liquor for lignocellulose fractionation is investigated. Regardless of the initial liquor constitutions, the mechanism of organosolv pulping has been sufficiently clarified in previous researches [29]. In typical organosolv pulping process, the bonds between hemicellulose and lignin in lignocellulose were broken. At the same time, they were degraded into small molecular weight fragments that dissolved in the liquidous phase, and the insoluble cellulose can be easily separated. Lignin and hemicellulose dissolved in pulping liquor can be recovered by concentration, precipitation, and drying. Furthermore, the addition of catalysts such as acids and alkalis promoted the removal of lignin and hemicellulose, and reduced the optimal pulping temperature [30].
Therefore, the depolymerization efficiency of lignocellulose matrixes are highly dependent on the temperature of organosolv pulping [31]. In higher temperature conditions, lignocellulose undergoes self-hydrolysis to produce organic acids such as acetic acid, formic acid, and levulinic acid, which can promote the depolymerization of lignocellulose matrix. Under acidic conditions, the α-OH in the β-O-4 unit of lignin may be protonated and dehydrated to form a reactive benzyl cation intermediate [32]. This will lead to the self-condensation of subsequent lignin units to form stable C-C bonds or cleavage to form ketone/aldehyde compounds, leading to subsequent repolymerization of lignin fragment [33,34]. As expected, results in the current work were in accordance with the phenomenon in related references [35,36]. Specifically, with the increase of pulping temperature, the delignification and hydrolysis of hemicellulose in corn stover were increased rapidly (Figure S1).
As shown in Figure 2, the highly branched structure rich in acetyl and hydroxyl groups and the non-crystalline structure lead to relatively low thermal and chemical stability of hemicelluloses in organosolv pulping [37]. Because of the enhanced delignification and hydrolysis of hemicellulose in higher temperature conditions, the crystallinity of the pulp was also increased [29]. However, the crystallinities of the pretreated pulps were similar to or even lower than that of the raw corn stover (Table S1). This was due to the destruction of cellulose crystals by the alkaline solvent and high temperature [38]. Previous studies have shown that the amorphous cellulose was more susceptible to enzymatic hydrolysis [39], which will be clarified in Figure 3. Consequently, the glucan content was always higher than the xylose and lignin retention in the fractionated pulp, no matter the type and concentration of solvents in initial liquor, as shown in Figure 2.
The effectiveness of organosolv pulping on decomposition of lignocellulose matrixes is also visualized in SEM images (Figure S4). The surface feature of corn stover, which was originally dense and smooth before organosolv pulping, became rough and porous with wrinkles and fragments no matter the pulping conditions and constitution of the initial liquor [40]. It was well-reorganized that the rough surface of organosolv pulps provided much more enzyme adsorption sites compared to raw material, improving the accessibility of cellulase [41].
As mentioned earlier, according to the principle of organosolv pulping, the solubility coefficient of the initial liquor was a decisive factor to the delignification performances and thereby influenced the lignocellulose fractionation performances [42]. Previous researches indicated that the initial liquor with closer Hanssen solubility coefficient to the basic lignin units (24.47–24.15 MPa1/2) was more favorable to the depolymerization of lignocelluloses [43]. The Hansen coefficient of n-butanol is 23.2 MPa1/2 [44], which is closer to lignin. Thus, under the conditions of Butanol-200 °C-50 wt% group, a maximized delignification of corn stover of 86.16% was obtained.

3.2. Enzymatic Hydrolysis of Pretreated Pulps

Monosaccharide production from the organosolv pulps was further evaluated. Generally, the increased pulping temperature led to the increase, while the increased solvent concentration in initial liquor resulted in the decreases of overall monosaccharide recovery. Correspondingly, compared with other groups, the n-butanol containing liquors always exhibited higher sugar recovery rates from pulp (Figure S2). In particular, in Butanol-200 °C-50 wt% group, 49.8 g/L total monosaccharides (47.5 g/L glucose and 2.26 g/L xylose) were obtained after 48 h of enzymatic hydrolysis. The enzymatic hydrolysis efficiency of glucan and total monosaccharides reached 89.41% and 89.39%, respectively, in the same group (Figure S3). For the yield of mono-sugars, the Ethanol-200 °C-50 wt%, Acetone-200 °C-30 wt%, and the Butanol-180 °C-50 wt% groups recovered 50.92%, 50.89%, and 45.98% of monosaccharides from raw corn stover, respectively (see Figure 4), which were selected for further analysis.

3.3. Characterization of the Fractionated Lignin

Previous studies have reported that low molecular weight and high uniformity are the prerequisites for high solubility, homogeneity, and rich functional groups in lignin molecules [45]. The organosolv pulping could co-generate high-purity value technical lignin with narrow molecule weight distribution and high functional group preservation [46]. This motivates us to further characterize and compare the lignin specimens isolated from the binary A-B-E and water solutions. GPC results (see Table 1) indicated that the Mw and Mn of lignin specimens from acetone pulping groups were 1084 g/mol and 1032 g/mol, respectively, and the polydispersity index (PDI, Mw/Mn) was 1.05, confirming the good uniformity and dispersibility. The TGA/DTG curves suggested the lignin specimens also possessed good thermos stability, which further extends the potential of valuable applications of the lignin specimens (Figure S6). In contrast, relatively higher Mw, Mn, and PDI of lignin specimens were observed in the ethanol and n-butanol pulping groups.
Figure 5 illustrates that the lignin specimens exhibited similar FTIR spectrums. The absorption peak at 3340 cm−1 is the stretching vibration peak of aromatic and aliphatic hydroxyl groups [47], and 2900 cm−1 and 2850 cm−1 are the stretching vibration absorption peaks of methylene and methyl C-H [48]. It can be seen that the absorption peak intensity of n-butanol fractionated lignin is the largest, which indicates that the molecules with methyl and methylene side chains are more abundant [49]. The absorption peaks at 1610 cm−1, 1510 cm−1, and 1420 cm−1 clearly indicate the vibration of the benzene ring skeleton, which is a typical lignin structure [50]. Corn stover is a typical grass that is rich in guaiacylphenylpropane (G-type), syringylphenylpropane (S-type), and p-hydroxyphenylpropane (H-type) lignin units, so the absorption peaks at 1150 cm−1, 1110 cm−1, and 830 cm−1 are obvious [51,52]. The peak at 1320 cm−1 is the infrared absorption peak of S-type lignin and condensed G-type lignin, 1260 cm−1 is the absorption peak of G-type lignin C=O, and 1220 cm−1 is the stretching vibration peak of S-type lignin C-C, C-O, and C=O [53,54]. Comparatively, the S type unit of lignin accounts for a larger proportion in lignin specimens, indicating less demethylation effect [55]. The appearance of the 920 cm−1 absorption peak is attributed to the impurity residual structure of glucan or xylan [56].
The HSQC spectrums indicated the organosolv pulping promoted the breakage of the α- and β-aryl ether bonds (Figure S7) [57]. Relatively strong methoxyl and xylan signals are also apparent in HSQC spectrums, which also confirmed the polysaccharide residues in lignin [58]. In fact, due to the relatively stable C-C structure of β-β and β-5, the content in lignin changed little after fractionation [59]. Meanwhile, it can be found that the β-O-4 bond signal is weaker after acetone pulping, indicating that the inter-molecule lignin breakage is more efficient than that in ethanol and n-butanol groups. Additionally, there are still large number of β-O-4 interlinkages preserved in the lignin specimens after organosolv pulping [60,61]. The aromatic ring region signals of lignins mainly include basic lignin units (G, S, H) and hydroxycinnamic acid units (ferulic acid and coumaric acid). Meanwhile, ferulate and p-coumarate structures are also apparent in the aromatic ring region (for details, please see the Supplementary Information).

3.4. ABE Fermentation Using the Enzymatic Hydrolysate of Pulps

The hydrolysates with the highest fermentable monosaccharide yield from organosolv pulps for each solvent were used as the substrate for ABE fermentation using the hyper- and high-inhibitor tolerant Clostridium acetobutylicum ABE1401. The total monosaccharide concentrations in the hydrolysates of Ethanol-200 °C-50 wt%, Acetone-200 °C-30 wt%, and the Butanol-180 °C-50 wt% groups were 41.5 g/L (38.6 g/L glucose and 2.93 g/L xylose), 36.5 g/L (35.0 g/L glucose and 1.5 g/L xylose), and 37.7 g/L (32.6 g/L glucose and 5.1 g/L xylose), respectively. Fermentations using the synthetic mediums containing similar sugars were set as the control groups.
Results in Figure 6 demonstrated that the organosolv pulps’ hydrolysates exhibited excellent ABE fermentation performances no matter the conditions. The ABE yield and concentration in experimental groups were very similar to those in control, indicating negligible inhibition of the hydrolysates to the metabolism of microorganisms. After 96 h inoculation in batch mode, a highest ABE concentration of 15.0 g/L was obtained in the Ethanol-200 °C-50 wt% group (9.35 g/L n-butanol, 4.62 g/L acetone, and 1.06 g/L ethanol), with total solvent yield of 0.36 g/g sugar (n-butanol yield 0.20 g/g sugar), while the results obtained in Acetone-200 °C-30 wt% group were a little behind the other two groups. We speculate the higher AIL and ASL content in Acetone-200 °C-30 wt% pulp might be the reason for the lower solvent yield, as the phenolic compounds always possessed inhibitions to Clostridia cells [62].

3.5. Mass Balance

As shown in Figure 1, in the whole diagram of the proposed biorefinery process, the fermentation broth can be efficiently separated and purified in the distillation system [63]. In turn, the outlet streams of the distillation sequence can be diluted and used as the initial liquor for upstream corn stover fractionation. In side stream, the solvents in lignin-rich post-pulping liquor are easily separated by vacuum evaporation and recycled into the beer column. Thus, technical lignin can be easily obtained after simple suspension or extraction–evaporation processes.
In this context, as shown in Figure 7, mass balance indicated 1 kg of corn stover produced 312 g of total monosaccharides and 112 g of ABE as final products in Ethanol-200 °C-50 wt% group, while 96.0 g of total ABE production was received in Butanol-180 °C-50 wt% group. As shown in Table 2, the results of this study are still competitive in terms of biobutanol production compared with previous studies. This confirmed the closed-circuit clean fractionation for second-generation biobutanol production was of high efficiency. At the same time, it was obvious that the closed-circuit organosolv pulping strategy under ‘energy-matter intensification’ concept yielded green and economic benefits.

4. Conclusions

The acetone–n-butanol–ethanol-enriched streams from downstream separation unit of second-generation biobutanol production processes can be recycled as the pulping liquor in upstream organosolv fractionation unit. This closed-circuit clean fractionation strategy could efficiently depolymerize the complex recalcitrance structure of lignocellulose, supporting the subsequent fermentable monosaccharide production and fermentative biobutanol production. In side stream, the fractionated lignin can be easily separated, which exhibited narrow molecule weight distribution and high preservation of functional groups, and showing prospects for further valorization. Under the optimized conditions (200 °C, 50 wt% ethanol, and 1 h), mass balance indicated 112 g of total ABE can be produced from 1 kg of dry corn stover. The closed-circuit organosolv pulping strategy based on the concept of ‘energy-matter intensification’ showed promise extending to other biorefinery processes.

Supplementary Materials

The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/fermentation11090514/s1, Figure S1: Solid recovery after organosolv pulping using mono-A-B-E aqueous solution under different conditions; Figure S2: Monosaccharide concentrations in enzymatic hydrolysate of the organosolv pulps; Figure S3: Enzymatic hydrolysis of the organosolv pulps; Figure S4: SEM morphologies of the organosolv pulps; Figure S5: XRD patterns of the organosolv pulps; Figure S6: (a) TGA and (b) DTG curves of the isolated lignin after organosolv pulping; Figure S7: 2D-HSQC NMR spectrums of the isolated lignin specimens after organosolv pulping; Table S1: Crystallinity of pulps after organosolv pulping. References [54,67,68,69] are cited in the Supplementary Materials.

Author Contributions

Conceptualization, C.S. and Y.G.; Methodology, C.S., Y.G., G.Z., R.C., Y.L. and J.C.; Formal analysis, R.C., J.W. and M.S.; Data curation, C.S. and H.W.; Writing—original draft, C.S. and H.W.; Writing—review & editing, D.C.; Funding acquisition, D.C. All authors have read and agreed to the published version of the manuscript.

Funding

This work is funded by the National Key R&D Program of China (Grant No. 2023YFB4203500), the National Natural Science Foundation of China (Grant No. 22478021) and the Guangxi Science and Technology Program (Grant No. AA24206052).

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

The original contributions presented in the study are included in the article, further inquiries can be directed to the corresponding authors.

Conflicts of Interest

The authors declare no conflict of interest.

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Fermentation 11 00514 g001
Figure 1. The organic solvent fractionation of lignocellulosic based on self-circulation of ABE fermentation solvent proposed in this study.
Figure 1. The organic solvent fractionation of lignocellulosic based on self-circulation of ABE fermentation solvent proposed in this study.
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Figure 2. Chemical composition of the pulps after organosolv pulping using binary A-B-E and water solutions under different conditions.
Figure 2. Chemical composition of the pulps after organosolv pulping using binary A-B-E and water solutions under different conditions.
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Figure 3. Carbohydrate recovery and delignification rates after organosolv pulping using binary A-B-E and water solutions under different conditions. (a) Glucan recovery; (b) xylan recovery; (c) total polysaccharides recovery; (d) delignification.
Figure 3. Carbohydrate recovery and delignification rates after organosolv pulping using binary A-B-E and water solutions under different conditions. (a) Glucan recovery; (b) xylan recovery; (c) total polysaccharides recovery; (d) delignification.
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Figure 4. Monosaccharide yield from pulps after enzymatic saccharification. (a) Glucose yield; (b) xylose yield; (c) total monosaccharide yield.
Figure 4. Monosaccharide yield from pulps after enzymatic saccharification. (a) Glucose yield; (b) xylose yield; (c) total monosaccharide yield.
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Figure 5. FT-IR spectrums of the isolated lignin after organosolv pulping.
Figure 5. FT-IR spectrums of the isolated lignin after organosolv pulping.
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Figure 6. ABE fermentation performances using hydrolysates of corn stover pulps. The control 1, 2, 3 represent the fermentations using synthetic mediums containing similar sugars of the Ethanol-200 °C-50%, Acetone-200 °C-30%, and Butanol-180 °C-50% groups, respectively.
Figure 6. ABE fermentation performances using hydrolysates of corn stover pulps. The control 1, 2, 3 represent the fermentations using synthetic mediums containing similar sugars of the Ethanol-200 °C-50%, Acetone-200 °C-30%, and Butanol-180 °C-50% groups, respectively.
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Figure 7. Mass balance of the lignocellulosic ABE production from corn stover based on organosolv pulping using downstream-separated streams.
Figure 7. Mass balance of the lignocellulosic ABE production from corn stover based on organosolv pulping using downstream-separated streams.
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Table 1. Molecular weight and polydispersity of the fractionated lignin specimens after organosolv pulping.
Table 1. Molecular weight and polydispersity of the fractionated lignin specimens after organosolv pulping.
SampleMw (g/mol)Mn (g/mol)PDI (Mw/Mn)
Ethanol-200 °C-50%127011501.10
Acetone-200 °C-30%108010301.05
Butanol-180 °C-50%128011601.10
Table 2. Summary of studies on n-butanol production from lignocellulose biorefining based on organosolv pretreatment.
Table 2. Summary of studies on n-butanol production from lignocellulose biorefining based on organosolv pretreatment.
Solvent ConditionRaw MaterialsPretreatment ConditionsEnzymatic Hydrolysis Efficiency (%)StrainABE Concentration (g/L)ABE Yield from Monosaccharide (g/g)Solvent Yield per kg of Straw (g/kg)Ref.
75% (v/v)Ethanol + 1% (w/w) H2SO4Elm wood180 °C, 60 min—Clostridium acetobutylicumnrrl B-59111.6 —121.1 [40]
75% (v/v) Ethanol + 1% (w/w) H2SO4Rice straw180 °C, 30 min—Clostridium acetobutylicumnrrl B-59110.5 —123.9 [16]
75% (v/v) Ethanol + 1% (w/w) H2SO4Elm woodAutohydrolysis (180 ℃, 60 min) + Organosolv (180 ℃, 60 min)—Clostridium acetobutylicumnrrl B-59112.7 —133 [64]
60% Ethanol + 4% NaOHCornstalks110 °C, 90 min Cellulose 85%, hemicellulose 82%Clostridium beijerinckii NCIMB 411012.8 0.43—[65]
50% Acetone + 0.1% H2SO4Sweet sorghum bagasse180 °C, 60 min94.2%Clostridium acetobutylicumnrrl B-59111.4 0314125 [15]
65% (v/v) Ethanol + 1% (w/w) H2SO4Loblolly pine170 °C, 60 min92.0%Clostridium acetobutylicum ATCC 82411.11 0.21 —[66]
168.24 g/l ABE solvent a +1% NaOHCorn stover160 °C, 1 h 92.6%Clostridium acetobutylicum ABE140113.5 0.331106.6[24]
50%wt EthanolCorn stover200 ℃, 1 h89.01%Clostridium acetobutylicum ABE140115.030.36112.4This study
30%wt AcetoneCorn stover200 ℃, 1 h90.26%Clostridium acetobutylicum ABE140111.680.3299.8This study
50%wt ButanolCorn stover180 ℃, 1 h76.05%Clostridium acetobutylicum ABE140112.820.3496.0This study
a: the ABE solvent contained 11.39 g/L ethanol, 44.06 g/L acetone, and 112.79 g/L n-butanol. —: Not applicable.
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Su, C.; Gao, Y.; Zhang, G.; Wen, H.; Chen, R.; Wang, J.; Li, Y.; Sun, M.; Cao, J.; Cai, D. Towards the Potential of Using Downstream-Separated Solvents as the Pulping Liquor of Upstream Lignocellulose Fractionation for Enhanced Acetone–Butanol–Ethanol Production. Fermentation 2025, 11, 514. https://doi.org/10.3390/fermentation11090514

AMA Style

Su C, Gao Y, Zhang G, Wen H, Chen R, Wang J, Li Y, Sun M, Cao J, Cai D. Towards the Potential of Using Downstream-Separated Solvents as the Pulping Liquor of Upstream Lignocellulose Fractionation for Enhanced Acetone–Butanol–Ethanol Production. Fermentation. 2025; 11(9):514. https://doi.org/10.3390/fermentation11090514

Chicago/Turabian Style

Su, Changsheng, Yunxing Gao, Gege Zhang, Hao Wen, Rui Chen, Jiajing Wang, Yujie Li, Mingyuan Sun, Jikang Cao, and Di Cai. 2025. "Towards the Potential of Using Downstream-Separated Solvents as the Pulping Liquor of Upstream Lignocellulose Fractionation for Enhanced Acetone–Butanol–Ethanol Production" Fermentation 11, no. 9: 514. https://doi.org/10.3390/fermentation11090514

APA Style

Su, C., Gao, Y., Zhang, G., Wen, H., Chen, R., Wang, J., Li, Y., Sun, M., Cao, J., & Cai, D. (2025). Towards the Potential of Using Downstream-Separated Solvents as the Pulping Liquor of Upstream Lignocellulose Fractionation for Enhanced Acetone–Butanol–Ethanol Production. Fermentation, 11(9), 514. https://doi.org/10.3390/fermentation11090514

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