DOI: 10.3303/CET25117038 Paper Received: 5 December 2024; Revised: 25 March 2025; Accepted: 1 May 2025 Please cite this article as: Ferreira B.X., De Q. Nobrega R., Maciel Filho R., Vieira C.F., Mariano A.P., 2025, A Short Bibliometric Analysis of Detoxification Processes in Butanol Production from Biomass Hydrolysates via Fermentation, Chemical Engineering Transactions, 117, 223- 228 DOI:10.3303/CET25117038 CHEMICAL ENGINEERING TRANSACTIONS VOL. 117, 2025 A publication of The Italian Association of Chemical Engineering Online at www.cetjournal.it Guest Editors: Fabrizio Bezzo, Flavio Manenti, Gabriele Pannocchia, Almerinda di Benedetto Copyright © 2025, AIDIC Servizi S.r.l. ISBN 979-12-81206-17-5; ISSN 2283-9216 A Short Bibliometric Analysis of Detoxification Processes in Butanol Production from Biomass Hydrolysates Via Fermentation Bruno X. Ferreira, Riann Q. Nóbrega, Rubens Maciel Filho, Carla F. S. Vieira, Adriano P. Mariano* Universidade Estadual de Campinas (UNICAMP), Faculdade de Engenharia Química (FEQ), Laboratory of Optimization, Design, and Advanced Control – Fermentation Division (LOPCA-Ferm), Campinas, SP, Brazil. adpm@unicamp.br The fermentative production of products from biomass is hindered by microbial inhibitory compounds generated during thermochemical pretreatment. To address this challenge, various detoxification strategies have been implemented either before or during fermentation. An overview of the most explored strategies can help identify promising directions for further investigation. This study thus examines research on detoxification techniques used in fermentation processes, specifically focusing on butanol production from biomass hydrolysates. A bibliometric analysis was conducted using the Web of Science (WoS) database alongside the Bibliometrix tool (RStudio) and the Biblioshiny app for statistical and network analyses, as well as bibliometric mapping. A search using the terms "fermentation" and "detoxification" resulted in 2,307 documents (1960 to 2024). The search was refined by adding the terms "hydrolysate" or "hemicellulosic hydrolysate," and finally "butanol." Brazil was the most cited country when "detoxification" was associated with "hemicellulosic hydrolysate." Butanol production was predominantly associated with Acetone-Butanol-Ethanol (ABE) fermentation by Clostridium species. Highly cited articles utilized hydrolysates derived from various biomass sources, including sugarcane bagasse, corn stover, and wood. Among the most cited articles (excluding reviews), the predominant detoxification techniques are overliming with Ca(OH)2, activated charcoal, and resins (or membranes). All these techniques were applied before fermentation, suggesting a potential gap in research on in-situ detoxification strategies. 1. Introduction Using residual biomass as a renewable source to produce different bio-based compounds usually obtained by the petrochemical route is an important aspect of biorefineries. Biomass can have distinct origins, such as forestry and agriculture residuals and food waste (Raj et al., 2022). Agriculture residuals are lignocellulosic materials, which are mainly composed of three main structures: cellulose [17 – 50 %], hemicellulose [13 – 33 %], and lignin [9 – 35 %]. From the cellulosic (C6) and hemicellulosic (C5) fractions, fermentable sugars can be obtained using different hydrolysis strategies (Candido et al., 2020; Gyan et al., 2024). Biomass hydrolysates containing sugar can be used to produce second-generation alcohols such as ethanol and butanol. However, the traditional yeast Saccharomyces cerevisiae can only metabolize hexoses. An alternative is to use other microorganisms that can also consume C5 sugars as substrate, such as Clostridium species. These bacteria can convert both C6 and C5 sugars into butanol by the Acetone-Butanol-Ethanol (ABE) fermentation (Nogueira et al., 2021). C5 sugars, such as xylose, are the main substrate in hemicellulosic hydrolysates, often obtained by dilute acid pretreatment (DA), and the second most relevant sugar in hydrolysates of cellulose and hemicellulose. However, DA also generates microbial inhibitory compounds, such as furan derivatives (furfural), organic acids (acetic acid), and phenolic derivatives (p-coumaric acid), which can reduce or inhibit microbial growth (Candido et al., 2020; Xu et al., 2023). Due to the challenges posed by lignocellulose-derived microbial inhibitory compounds in the production of second-generation alcohols, significant research has focused on understanding their impact on fermentation 223 processes and developing strategies to mitigate these effects (Gao et al., 2016; Raj et al., 2022; Gyan et al., 2024). This study provides a concise bibliometric analysis and an overview of advances in detoxification strategies for butanol production from hydrolysates derived from various biomass sources. It highlights key journals, examines country-specific scientific output, and identifies emerging trends in the field. 2. Methodology Data for the bibliometric analysis was collected from the Web of ScienceTM (WoS) database (ClarivateTM), applying different combinations of topic words and logic operation terms (Table 1) to refine the search window. The search was also limited by the type of documents (only research, review, and conference proceeding articles were considered). The bibliometric analysis was conducted using the Bibliometrix tool (RStudio v.4.3.2) and the Biblioshiny app (Aria and Cuccurullo, 2017) for statistical and network analyses and bibliometric mapping. Table 1: Combinations of terms explored in the WoS database Scenario Topics Total of documents 1 “fermentation” AND “detoxification” 2302 2 “fermentation” AND “detoxification” AND “hydrolysate” 818 3 “fermentation” AND “detoxification” AND “hemicellulosic hydrolysate” 108 4 “fermentation” AND “detoxification” AND “hydrolysate” AND “butanol”a 101 5 “fermentation” AND “detoxification” AND “hydrolysate” AND “ABE”b 73 6 “fermentation” AND “detoxification” AND “hemicellulosic hydrolysate” AND “butanol”a 5 a – “butanol” OR “biobutanol” OR “bio-butanol” OR “bio butanol” OR “n-butanol” b – “ABE” OR “acetone-butanol-ethanol” OR “acetone-butanol-ethanol (ABE)” 3. Results and Discussions 3.1 General information Among the groups generated by the combinations, the majority of the documents are research articles (90 – 95 %), and the review articles are the minority group (1.4 – 7.1 %). The exception was scenario 6, in which all five documents were research articles. Interestingly, the analysis of the publications through the years (Figure 1) shows time marks for the association between the terms and research fields that are emerging. Figure 1: Annual growth of publications for each combination of terms used in the bibliometric analysis. 3.2 Main journals Considering the combinations of terms explored in the WoS database, the journal “Bioresource Technology” emerged as the most relevant source, publishing at least twice as many articles as the second-ranked journal. In scenarios 1 to 3, the top five sources are mostly the same journals. However, when the terms “butanol” or “ABE” were added to the search (scenarios 4 to 6), the journals “Renewable Energy” and “Biotechnology for Fuels” emerged as second and third in the ranking, respectively. 224 3.3 Country-specific scientific output Countries were compared for productivity and citation impact of publications. China and the United States are the leading countries in terms of productivity across all scenarios, except in those related to hemicellulosic hydrolysate (scenario 3), where Brazil and India are the most prolific producers of articles. Brazil ranks among the top five in scenarios 1, 2, and 4, but falls to sixth place in scenario 5. When comparing countries based on citation levels, Sweden ranks first in scenario 1 and third in scenario 2, while China and the United States are the most prominent in the majority of cases. Brazil and India keep their leading positions in scenario 3. Brazil also ranked in the top four for scenarios 1 and 2 but placed sixth in scenario 4 and ninth in scenario 5. Other countries that appear in the rankings for productivity and citation level are Spain, Canada, and Finland, mainly when the terms “butanol” or “ABE” are added. These comparisons highlight Brazil's significant role in research on the fermentation of hydrolysates. Furthermore, as its publications primarily focus on ethanol production, this expertise could be applied to butanol production in the future. 3.4 Trending topics Trending topics were first identified by assessing authors’ keywords that appeared in the publications found in the search scenarios (Table 2). In scenarios 1 to 3, the keywords indicate that detoxification and fermentation processes are primarily associated with ethanol and xylitol production, suggesting that these products are trends in research on the fermentation of biomass hydrolysates. When the terms “butanol” or “ABE” were included in the search (scenarios 4 and 5), the resulting top five keywords were the same. The term “ABE fermentation” notably ranked second in both scenarios, suggesting that most studies focus on n-butanol rather than isobutanol, which is primarily produced at the commercial level by genetically modified yeast (Lakshmi et al., 2021). This differentiation is presumably reinforced by adding the genus and species name of the microorganism (note that solventogenic Clostridia are xylose-fermenting microorganisms) to the keywords list. Table 2: Most frequently used terms as authors’ keywords in scenarios of the bibliometric analysis. Word (occurrence) # Scenario 1 Scenario 2 Scenario 3 Scenario 4 Scenario 5 1 detoxification (406) detoxification (188) xylitol (43) butanol (37) butanol (32) 2 fermentation (333) ethanol (143) detoxification (29) ABE fermentation (30) ABE fermentation (30) 3 ethanol (296) fermentation (133) hemicellulosic hydrolysate (28) Clostridium (15) Clostridium (12) 4 lignocellulose (106) xylitol (82) fermentation (16) detoxification (15) detoxification (12) 5 pretreatment (105) inhibitors (63) ethanol (15) Clostridium acetobutylicum (13) Clostridium acetobutylicum (10) To further confirm the trending topics, a co-occurrence network analysis was performed considering scenarios 2 and 4 (Figure 2). In scenario 2 (Figure 2A), butanol production is a secondary topic (represented by green nodes), while the primary emphasis is on the production of xylitol and ethanol. Additionally, detoxification is the most prominent node, highlighting significant efforts made to tackle the challenge of converting biomass hydrolysates into these products. In scenario 4 (Figure 2B), on the other hand, detoxification appears as a secondary topic, with a node size similar to terms not used in the search such as gas stripping and liquid-liquid extraction — both in-situ product recovery techniques. This suggests that research on the fermentative production of butanol from biomass hydrolysates may have focused more on minimizing butanol inhibition than on advancing existing methods to detoxify microbial inhibitors derived from lignocellulose. 225 Figure 2: Authors’ keywords co-occurrence ([A] – scenario 2; [B] – scenario 4). The size of each node is proportional to the number of occurrences. 3.5 Trends in detoxification strategies Articles found in scenario 4 were examined to find major trends in detoxification strategies applied to butanol production from biomass hydrolysates. Among the top 25 most cited articles (with the top 10 presented in Table 3), several general observations can be made. First, the most common types of biomass are derivatives from crops, featured in 17 articles, and wood, found in 5 articles. For the preparation of hydrolysates, 16 studies employed pretreatment in combination with enzymatic hydrolysis while the remaining studies used pretreatment alone. Among the pretreatment techniques, dilute acid (DA) and liquid hot water (LHW), whether used individually or in combination, are the most prevalent (Table 3). The microorganisms used in the fermentation were Clostridium species, with C. beijerinckii and C. acetobutylicum being the most commonly utilized. Each of these species is found in 11 articles. Regarding the detoxification methods, eight studies do not employ any detoxification strategies. In the remaining 17 studies, the prevalent methods for removing microbial inhibitory compounds from hydrolysates are overliming (typically using Ca(OH)2), followed by activated charcoal (at concentrations ranging from 2 to 7 % w/v), synthetic resins, and membranes. These are methods extensively used in the fermentative production of ethanol from biomass hydrolysates (Nogueira et al., 2021). In almost all 25 studies, detoxification was performed prior to fermentation, adding an extra step in the production of second- generation butanol. The only exception was Sun et al. (2020) (ranked 19th), who added 1 g of biochar during fermentation to remove inhibitors from switchgrass hydrolysate. Since most of the detoxification techniques have been applied before fermentation, this implies a potential gap in research regarding in-situ detoxification strategies oriented to solventogenic Clostridium fermentation. Such strategies could enhance the fermentative production of butanol from biomass while eliminating one process step for the preparation of fermentation broths containing hemicellulosic hydrolysates. [A] [B] 226 Table 3: List of most cited publications on biomass hydrolysate production, detoxification, and fermentation strategies identified in scenario 4. # Document TC TC/year Biomass Hydrolysate production Microorganism Detoxification method 1 Ezeji et al. (2007) 446 24.78 Corn fiber DA C. beijerinckii BA10 Overliming (Ca(OH)2) 2 Ezeji et al. (2008) 218 12.82 Distillers dried grains and solubles (DDGS) LHW and DA + Enzymatic C. beijerinckii BA101, C. acetobutylicum (260, 824), C. saccharobutylicum 262, and C. butylicum 592 Overliming (Ca(OH)2) 3 Lu et al. (2012) 182 14.00 Cassava bagasse LHW + Enzymatic C. acetobutylicum JB200 - 4 Wang et al. (2011) 129 9.21 Chipped corn stover SE and AP + Enzymatic C. acetobutylicum ATCC 824 Activated charcoal (7.5 %) 5 Lu et al. (2013) 107 8.92 wood pulp (maple, beech, and birch trees) DA C. beijerinckii CC101 (adaptive mutant) Ion exchange resins, overliming (Ca(OH)2), Activated charcoal (2 %) 6 Cai et al. (2013) 92 7.67 Sweet sorghum bagasse (SSB) DA C. acetobutylicum ABE1201 PDMS 7 Zhang et al. (2018) 85 12.14 Hybrid poplar (Populus) DA C. saccharobutylicum BAA-117 Overliming (Ca(OH)2), Activated charcoal (5 %) 8 Sun et al. (2012) 76 5.85 Sugar maple (Acer saccharum) wood LHW and DA C. acetobutylicum ATCC824 Membrane (nano)filtration + overliming (Ca(OH)2) 9 Lopez-Linares et al. (2019) 76 12.67 Brewer’s spent grain (BSG) MAH + Enzymatic C. beijerinckii DSM 6422 Activated charcoal (2 %) 10 Gottumukkala et al. (2013) 73 6.08 Rice straw DA+ Enzymatic C. sporogenes BE01 Filtration with Amberlite and anionic resin TC - Total of citation; LHW - Liquid Hot Water pretreatment; DA - Dilute Acid pretreatment; SE - Steam Explosion pretreatment; AP - Alkaline Peroxide pretreatment; MAH - Microwave-assisted hydrothermal pretreatment; PDMS - Pervaporation membrane composed of polydimethylsiloxane. 4. Conclusions The most commonly employed detoxification strategies among the top-cited articles for butanol production from biomass hydrolysate were overliming (using Ca(OH)2) and activated charcoal (at concentrations of 2 – 7 %w/v). The hydrolysates are primarily derived from crop or wood biomass, employing pretreatments such as dilute acid or liquid hot water, with or without enzymatic hydrolysis. In the fermentation, various species of Clostridium were utilized, with C. beijerinckii and C. acetobutylicum being the most commonly used. The country-specific bibliometric analysis of productivity and citation impact revealed that China and the United States play a significant role in scientific production. Brazil is emerging as a significant player and has the potential to leverage 227 its expertise in cellulosic ethanol production for ABE fermentation. Butanol production accounts for approximately 15% of scientific production related to the detoxification of biomass hydrolysates, with most detoxification processes occurring prior to fermentation. Therefore, future studies should also investigate the further development of in-situ detoxification methods for lignocellulose-derived microbial inhibitors during fermentation. Acknowledgments This study was financed, in part, by the São Paulo Research Foundation (FAPESP), Brazil (process numbers 2023/01072-7 and 2023/01219-8). We also thank the financial support from Coordenação de Aperfeiçoamento de Pessoal de Nível Superior – Brasil (CAPES/PROEX). 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