Hydrothermal pretreatment is a widely used, cost-effective, and environmentally friendly method for processing lignocellulosic biomass (Singh et al., 2023). It separates biomass into solid and liquid fractions, with the solid phase primarily containing cellulose and lignin for saccharification and fermentation and the liquid phase containing dissolved hemicellulose. This process enhances the accessibility of cellulose for enzymatic hydrolysis and increases the profitability of biorefining by efficiently utilizing both fractions. Operating at temperatures between 160°C and 270°C and pressures over 5 MPa, hydrothermal pretreatment leverages auto-hydrolysis, where water ionizes to produce hydronium ions that facilitate selective hydrolysis of ether bonds in lignin and acetyl group in hemicellulose decomposition (Shen et al., 2016), lead to polysaccharide and lignin depolymerization. This method significantly improves enzymatic hydrolysis efficiency, and supports the overall economic viability of biorefinery processes (Yang et al., 2018).

Yuan et al. (2019) conducted a short-term two-step hydrothermal pretreatment on tobacco stalks, effectively removing non-structural carbohydrates (mainly hemicellulose) and dissolving them in water. Initially, at a lower temperature of 144°C for 21 min, 11.8 wt% of sugars (based on dry biomass weight) were dissolved. The temperature was then increased to 200°C and maintained for 0 min, resulting in the extraction of 1.56 wt% of high-molecular-weight hemicellulose and 3.95 wt% of low-molecular-weight hemicellulose oligosaccharides, removing 86.98% of non-structural polysaccharides. The residue was primarily composed of cellulose and lignin. Additionally, the study found that after hydrothermal treatment, the residue surface changed from highly ordered to rough, with lignin forming spherical deposits on the surface.

Furthermore, Santana et al. (2023) increased the solid content to 17% and performed hydrothermal pretreatment at 190°C for 10 min, selectively converting hemicellulose in tobacco stalks into prebiotic oligosaccharides, achieving a maximum yield of 132 g of oligosaccharides per kilogram of tobacco stalks (Figure 4). These prebiotics can be used in food formulations, dietary supplements, and even pharmaceuticals to help restore gut microbiota. This study highlights the potential of tobacco-derived biomolecules, reinforcing the concept that “tobacco goes far beyond cigarettes,” a concept known for its significant health impacts on millions. Building on this, hydrothermal pretreatment at a higher solid content (20%) and at 190°C for 8 min was found to be the most effective, yielding 49.54% of xylooligosaccharides with a concentration of 11.11 g/L (Santana et al., 2024). Additionally, the residue could be further bioconverted to produce succinic acid with a concentration of 16.88 g/L and a yield of 42%. The entire process can produce 56 kg of oligosaccharides and 78 kg of succinic acid per 1,000 kg of tobacco. This demonstrates the feasibility of using tobacco stalks as valuable raw materials for producing oligosaccharides and succinic acid, laying the foundation for future optimization in clean and sustainable industrial applications. Although lignin removal at the macroscopic level was minimal during hydrothermal treatment, lignin degradation occurred due to extensive cleavage of ether bonds, with its molecular weight drastically reduced from 5,160 to 2,910 g/mol (Sun et al., 2019). This degradation and subsequent condensation caused the lignin to form spherical deposits on the biomass surface upon cooling, which hinders subsequent enzymatic hydrolysis of cellulose. Recently, Li et al. (2024) introduced rhamnolipid surfactants into the hydrothermal pretreatment process, achieving efficient conversion of tobacco stalks into reducing sugars with a concentration of 11.86 g/L, and the residue produced 611.3 mL of hydrogen through enzymatic hydrolysis and fermentation.

Hydrothermal pretreatment is a green and efficient method, but relying solely on temperature increase to enhance the concentration of hydrogen ions and activate biomass components may not achieve optimal results. Utilizing hydrothermal pretreatment as a solvent and incorporating acid, alkali, or metal catalysts to lower the pretreatment temperature and enhance conversion efficiency could be the future direction of hydrothermal pretreatment development.

Steam explosion pretreatment is a widely adopted method for lignocellulosic biomass processing (Hoang et al., 2023). This technique employs high-temperature and high-pressure steam to alter the biomass structure, enhancing enzymatic hydrolysis efficiency. Conducted at temperatures between 160°C and 240°C and pressures of 0.7–4.8 MPa for durations from seconds to min, the process involves rapid depressurization post-steam treatment, causing explosive decompression (Wang et al., 2015). This primarily hydrolyzes hemicellulose, improving cellulose accessibility for enzymatic hydrolysis and increasing the enzymatic hydrolysis rate. The acetyl group hydrolysis in hemicellulose releases acetic acid, which further catalyzes hemicellulose hydrolysis, converting it into fermentable glucose and xylose monomers. Key advantages of steam explosion include high sugar recovery efficiency, cost-effectiveness, and environmental benefits. However, challenges such as potential fermentation inhibitors, stringent pretreatment conditions, and high energy consumption exist (Yu et al., 2022). Despite these, steam explosion remains prevalent in industrial applications requiring high sugar recovery.

Han et al. (2015) conducted steam explosion pretreatment of tobacco stems at 0.5 MPa for 20 S. This treatment rendered the tobacco stems loose, facilitating polysaccharide enzymatic hydrolysis. Compared to untreated tobacco stems, the enzymatic hydrolysis efficiency of steam-exploded tobacco stems increased by 231%. Martin et al. (2008), Martin et al. (2002) further investigated steam explosion pretreatment at different durations and found that longer pretreatment times resulted in higher sugar concentrations in the hydrolysate, with relatively low inhibitor concentrations, minimizing the impact on fermentation. The ethanol yield reached 0.39 g/g, outperforming the hydrolysate from sugarcane bagasse. However, nicotine in tobacco stems significantly inhibited enzyme activity and microbial fermentation. Song and Zhang et al. (2021) treated tobacco stems with dilute sulfuric acid followed by steam explosion pretreatment. This method removed up to 85.54% of the nicotine from the tobacco stems and improved the efficiency of carbohydrate enzymatic hydrolysis and ethanol fermentation by 103.36% compared to steam explosion pretreatment alone. In a 5 L fermentation setup, the ethanol concentration reached a maximum of 23.53 g/L, with a glucose-to-ethanol conversion yield of 71.40%. Besides enzymatic hydrolysis for glucose and ethanol fermentation, the steam-exploded tobacco stem residue, due to its larger surface area and higher dissociation degree, can be used for pulping and nanocellulose production. Moreover, Tuzzin et al. (2016) introduced an alkaline activation step in the steam explosion pretreatment process. By pretreating at 175°C for 7 min, followed by bleaching, they found that the nanocellulose content in the cellulose from tobacco stems was higher than that in eucalyptus sulfate pulp, with no significant chemical differences between the two. Incorporating alkali-treated steam-exploded tobacco stem pulp as “backbone” fibers into low-quality secondary fiber paper pulp upgraded the paper from substandard to premium grade (Grade A), significantly increasing the breaking length (64.79%–90.14%) and ring crush index (34.36%–162.96%) (Figure 5) (Wang et al., 2024).

Overall, steam explosion pretreatment is a green and environmentally friendly technology that can significantly enhance the structural looseness of tobacco stem biomass in a short period, facilitating subsequent utilization. However, the process involves high pressure and high temperature, necessitating pressure-resistant equipment and high energy consumption. Future research should focus on optimizing steam explosion pretreatment conditions to reduce energy consumption, improve processing efficiency, and enhance the feasibility of industrial applications.

Acidic reagents have demonstrated the ability to break glycosidic bonds in lignocellulose, effectively dissolving cellulose, hemicellulose, and small amounts of lignin, thereby promoting the release of fermentable sugars (Lv et al., 2024). Compared to hydrothermal pretreatment, acid pretreatment is more commonly used and effective for degrading hemicellulose (and partially lignin) and disrupting amorphous cellulose (Jiang et al., 2023). The primary mechanism involves disrupting interactions between lignocellulosic components, such as van der Waals forces, hydrogen bonds, and covalent bonds, resulting in the dissolution of hemicellulose and the disruption of cellulose and lignin structures. This makes acid pretreatment more effective in lowering pretreatment temperatures and reducing energy consumption in biomass refining compared to hydrothermal methods. Due to the corrosive nature of acids, diluted acids are typically used to reduce equipment corrosion and minimize environmental harm. Although acid pretreatment enhances biomass degradation more effectively than hydrothermal pretreatment, it also generates inhibitors (such as furfural, 5-hydroxymethylfurfural, levulinic acid, and lactic acid), which can impede subsequent enzymatic hydrolysis and fermentation (Lv et al., 2024). Therefore, optimizing acid pretreatment involves balancing its high degradation efficiency with the generation of inhibitors to maximize overall biomass conversion efficiency.

The acid pretreatment of tobacco stems generally employs dilute acids, primarily to remove hemicellulose and enhance the accessibility of cellulose to enzymes. For example, after pretreatment with dilute sulfuric acid at 121°C for 90 min, the total reducing sugar in the hydrolysate reached 20.3 g/L, mainly consisting of hemicellulose-based monosaccharides (Schneider et al., 2017). The residue’s enzymatic hydrolysis yielded 38.1% glucose, and subsequent fermentation produced 0.19 g ethanol per gram of raw material. Besides sulfuric acid, other inorganic acids, such as HCl and HNO₃, have also been studied (Temitayo et al., 2019). However, their efficiency is lower than that of sulfuric acid, possibly because H₂SO₄ being a diprotic acid, can continuously provide hydrogen ions, whereas HCl and HNO₃ are monoprotic and volatile acids, making them less effective for tobacco stem pretreatment. Using a weaker organic polyacid like citric acid for the pretreatment of tobacco stems has shown higher efficiency than sulfuric acid. This may be because citric acid, being a weak polyacid, can continuously supply hydrogen ions to promote acid pretreatment, while its mild acidity can reduce the condensation of lignin structures in the biomass, thus decreasing inhibition of cellulose accessibility to enzymes. The significant advantage of acid pretreatment of tobacco stems is the removal of toxic substances such as nicotine, ensuring safe and high-value utilization in subsequent processes (Zhang et al., 2021). Studies have shown that dilute acid pretreatment can remove up to 85.54% of nicotine while enhancing enzymatic hydrolysis. Although dilute acid pretreatment can promote the acid hydrolysis of hemicellulose to produce oligosaccharides and even monosaccharides like xylose, it can also lead to side reactions that produce furfural and fatty acids and promote partial degradation of lignin to phenolic compounds, which inhibit subsequent enzymatic hydrolysis and fermentation (Li et al., 2023). To improve ethanol fermentation yield, tobacco stems subjected to dilute acid pretreatment can undergo further mild alkaline pretreatment, achieving the removal of 47% hemicellulose and 65% lignin (Gong et al., 2023). After enzymatic hydrolysis, the residues can then be co-fermented, producing 0.19 L ethanol per kilogram of tobacco stems. In summary, acid pretreatment is an efficient method for processing tobacco stems. By optimizing the types of acids and pretreatment conditions, it is possible to further enhance biomass conversion efficiency and reduce the generation of inhibitors, laying the foundation for industrial application.

Different from hydrothermal and acid pretreatment, alkaline pretreatment can effectively dissolve lignin (Xu et al., 2016; Ntakirutimana et al., 2022). Through saponification and solvent reactions, the bonds in lignin-carbohydrate complexes (LCC), such as ester bonds, aryl ether bonds, alkyl-aryl bonds, and glycosidic bonds, are broken (Dong et al., 2019). Subsequently, acetyl and uronic acid groups in hemicellulose are also removed (Xu et al., 2016). These reactions lead to structural changes and degradation of lignocellulosic biomass, increasing the exposure of reactive sites and promoting cellulose swelling and partial decrystallization, thereby facilitating further enzymatic hydrolysis (Kim et al., 2016). Additionally, tobacco stalk biomass contains ash components like silica, which exist as silica deposits and protect the cell wall from digestibility (Khaleghian et al., 2017). Alkaline pretreatment can dissolve these silica compounds into the solution as silicates, promoting the breakdown and utilization of cell wall components. Generally, unless extensive delignification is implemented, alkaline pretreatment is less effective in removing the recalcitrance of plant biomass. However, it has the advantage of eliminating fermentation inhibitors (Jönsson and Martín, 2016). Nevertheless, the recovery of alkaline substances requires expensive capital investment and remains a significant barrier to alkaline pretreatment.

Alkali pretreatment of tobacco stalks not only mitigates the formation of fermentation inhibitors but also promotes the solubilization of lignin, thus enhancing the enzymatic hydrolysis efficiency of carbohydrates (Figure 6) (Yuan et al., 2019; Guo et al., 2021). The pretreatment conditions for tobacco stalks are relatively mild compared to other biomass types. For instance, sodium hydroxide pretreatment at 90°C has been shown to increase enzymatic hydrolysis efficiency by 165.7%, with subsequent simultaneous saccharification and fermentation yielding a 138.0% increase in ethanol production (Guo et al., 2021). Moreover, CaO has also been used effectively for the alkali pretreatment of tobacco stalks (Sophanodorn et al., 2020a; Sophanodorn et al., 2020b). This method resulted in a 79.35% monosaccharide yield upon enzymatic hydrolysis (Sophanodorn et al., 2020b). During alkali pretreatment, components such as lignin, hemicellulose, extractives, and a portion of amorphous cellulose are partially removed, enhancing the thermal stability of the residue but lowering the activation energy (Dallé et al., 2021). Additionally, the crystalline cellulose in the residue transforms from cellulose I to cellulose II, with the residue surface becoming rougher due to the removal of lignin and hemicellulose, making it suitable for applications as reinforcing fillers in composite materials. Despite the effectiveness of alkali pretreatment in lignin removal, hemicellulose remains in the residue. To address this, a sequential hydrothermal-alkali pretreatment can be employed to separate cellulose, hemicellulose, and lignin from tobacco stalks (Sophanodorn et al., 2020c; Wang et al., 2022). This combined pretreatment method significantly enhances enzymatic hydrolysis and fermentation efficiency, achieving ethanol yields of up to 75.74 g/L. During the hydrothermal-alkali process, the bonds between lignin and carbohydrates are cleaved, resulting in lignin with low carbon content and slightly increased molecular weight due to the cleavage of aryl-ether bonds and partial condensation during hydrothermal treatment (Liu et al., 2021). Interestingly, the regenerated lignin exhibits regular nanoparticle morphology and excellent UV absorption properties, making it suitable for lignin-based UV shielding materials. Furthermore, incorporating oxidants during alkali pretreatment improves the efficiency of tobacco stalk processing, yielding residues with 92.41% cellulose purity (Martin et al., 2008; Zhan et al., 2024). The cellulose crystallinity changes and its thermal stability increases from 261°C to 369°C. This high-purity cellulose can be used to produce food preservation films using plasticizers such as glycerol and sorbitol through solution casting, achieving high-value utilization of cellulose (Qian et al., 2023).

Overall, alkali pretreatment operates at relatively mild temperatures and effectively removes lignin, enhancing the accessibility of carbohydrates to enzymes. The transformation of cellulose crystalline structure is advantageous for producing cellulose-based materials. Compared to acid pretreatment, alkali pretreatment effectively eliminates inhibitors, facilitating subsequent enzymatic hydrolysis and fermentation. Therefore, alkali pretreatment is a prevalent method for processing tobacco stalks, demonstrating significant potential for industrial applications.

Compared to hydrothermal, acid, and alkaline pretreatment methods, organosolv pretreatment is more effective at dissolving lignin, efficiently removing it from the biomass, and dissolving it in the solvent (Ferreira and Taherzadeh, 2020). This process also penetrates and breaks down the internal chemical bonds of cellulose and hemicellulose. During this process, the dissolution of hemicellulose and delignification enhance the pore volume and surface area of cellulose, improving its availability for saccharification and enzymatic hydrolysis (Xu et al., 2023). Since the biomass components are highly polar substances rich in hydroxyl groups, low-boiling alcohols (such as ethanol and methanol) and high-boiling alcohols (such as glycerol, ethylene glycol, and tetrahydrofurfuryl alcohol) are commonly used in organosolv pretreatment (Sun et al., 2022; Sun et al., 2023). However, due to the large molecular weight of biomass, its solubility is relatively poor. To achieve more effective pretreatment, acid or alkaline catalysts are added to accelerate the hydrolysis of aryl ether bonds in lignin and hemicellulose, allowing them to dissolve in the organic solvent. Despite the effectiveness of organosolv in dissociating biomass, most organic solvents are costly, increasing the overall cost of the pretreatment process (Ferreira and Taherzadeh, 2020). Additionally, the flammability and volatility of organic solvents are significant drawbacks that must be considered in organosolv pretreatment.

Currently, research on the organic solvent pretreatment of tobacco stems is relatively limited. This is primarily due to the higher cost of organic solvents compared to hydrothermal treatments, which hinders industrial promotion. Additionally, the presence of nicotine and solanesol in tobacco stems affects the efficiency of the pretreatment, requiring additional extraction steps and further increasing costs. Studies have shown that using ethylene glycol as an organic solvent to pretreat tobacco stems results in a lignin removal rate of only 13.9%, significantly lower than the delignification rates for wheat straw and corncobs. This is mainly because solanesol in tobacco stems weakens the strong hydrogen bond interactions between free hydroxyl groups in lignin and ethylene glycol. To mitigate the influence of solanesol, Chen et al. (2022) used n-hexane extraction to remove solanesol before ethylene glycol pretreatment, successfully increasing the lignin removal rate to 40.5%, an improvement of 191%. This method provides an effective solution for the organic solvent pretreatment of tobacco stems.

Despite the potential of organic solvent pretreatment in addressing lignin removal in tobacco stems, further research and optimization are needed for large-scale industrial applications. This includes finding more cost-effective solvents and improving pretreatment processes to reduce costs and enhance efficiency.

According to the principles of green chemistry, environmentally friendly solvents are crucial for sustainable processes (Roy et al., 2020). Industrial solvents often contain toxic substances, driving the demand for green alternatives like ionic liquids (ILs) and deep eutectic solvents (DES) (Roy et al., 2020). ILs, particularly imidazole-based ones, excel in dissolving cellulose, hemicellulose, and lignin, aiding enzymatic hydrolysis and biomass conversion into fermentable sugars and biofuels (Achinivu et al., 2024; Hernandez et al., 2024). They can be recycled due to their lack of vapor pressure, but their high cost and toxicity to cellulases limit large-scale application. Residues of lignin and hemicellulose in ILs can increase viscosity, complicating further processing. DES, derived from natural, non-toxic, and inexpensive resources, offers a promising alternative to ILs (Hong et al., 2020). They are compatible with enzymes, cost-effective, and simple to synthesize without complex purification. DES forms competitive hydrogen bonds with carbohydrates and hydroxyl groups in lignin, hydrolyzing LCC and breaking ether bonds between lignin and hemicellulose (Shen et al., 2019a; Wang et al., 2019). DES selectively dissolves lignin, enhancing biomass deconstruction and high-value utilization (Shen et al., 2019b; Wang et al., 2020). Despite these benefits, both ILs and DES face challenges with increased viscosity due to residual lignin and hemicellulose, complicating subsequent processing and applications.

IL pretreatment and DES pretreatment, as relatively green and advanced technologies, have been applied to tobacco stalks to a lesser extent, mainly focusing on lignin separation and characterization, interpreting structural changes of lignin during pretreatment, and identifying its unique properties. Similar to other biomass, imidazolium-based ionic liquids exhibit high solubility for tobacco stalk lignin, especially 1-ethyl-3-methylimidazolium diethylphosphate ([Emim][DEP]), which has the highest solubility due to its strong acidity and hydrogen bonding capability (Fan et al., 2018). After [Emim][DEP] pretreatment, the removal of lignin renders the surface of tobacco stalks rough, facilitating subsequent enzymatic hydrolysis and fermentation. The purity of separated lignin exceeds 85%, with extraction rates reaching up to 90.21%. As reaction conditions become more severe, the regeneration rate of lignin decreases, likely due to lignin degradation during IL pretreatment, resulting in lignin fragments that cannot precipitate and regenerate. At higher temperatures, lignin further undergoes condensation.

DES also exhibits strong solubility for lignin in tobacco biomass (Wang et al., 2022; Wang et al., 2022). Studies on the pretreatment of tobacco stalk lignin using acidic, neutral, and basic DES have shown that acidic DES (e.g., lactic acid/choline chloride) can efficiently separate and extract tobacco stalk lignin. This is likely because acidic DES not only has high solubility for lignin but also efficiently cleaves hemicellulose, promoting lignin separation. Conversely, basic DES (monoethanolamine/choline chloride) and neutral DES (ethylene glycol/choline chloride) show less significant effects on hemicellulose degradation, and the dispersion of lignin decreases, resulting in lignin with a uniform molecular weight. However, lignin in acidic DES undergoes significant degradation and condensation, whereas basic DES pretreatment maintains the lignin aryl ether bond content (Figure 7). Therefore, for DES pretreatment of tobacco stalks, if the goal is to improve lignin separation efficiency without focusing on its structural changes, an acidic DES system should be used. If the objective is to obtain structurally intact lignin for subsequent use, basic DES solvents should be employed.

Although the application of ionic liquid pretreatment and DES pretreatment in tobacco stalks is currently limited, their green solvents, zero vapor pressure, high safety, and recyclability significantly promote the green and efficient pretreatment of tobacco stalks.