Efficient Bioethanol Production from Spent Coffee Grounds Using Liquid Hot Water Pretreatment without Detoxification

Abstract

Coffee beans, a popular commodity in the world, are processed into coffee, which generates a considerable quantity of spent coffee grounds (SCGs). However, SCGs, a byproduct rich in hemicellulose, poses a challenge due to fermentable sugar loss during conventional pretreatment. This study investigates the efficient production of bioethanol from SCG using an optimized liquid hot water (LHW) pretreatment combined with separate hydrolysis and fermentation (SHF) process. LHW pretreatment at 180 °C for 20 min with a high solid-to-liquid ratio (SLR) of 1:6 (w/v) was optimized to disrupt the lignocellulosic structure and retain high levels of fermentable sugars, which included mannose and glucose. This approach achieved a bioethanol concentration of 15.02 ± 0.05 g/L and a productivity rate of 1.252 g/(L·h), demonstrating the efficiency of this integrated process. Interestingly, the high SLR LHW pretreatment significantly reduces water usage and enhances product concentration, offering a promising, environmentally friendly, and economically viable method for industrial bioethanol production from SCGs without the necessity of detoxification.

1. Introduction

Tightening resource constraints, serious environmental pollution, and ecosystem degradation pose significant challenges to the sustainable development of human society. One of the most promising approaches for promoting comprehensive resource conservation and recycling, and for establishing a clean, low-carbon, safe, and efficient energy system, is through research on advanced and efficient biomass conversion and biorefinery technologies. These technologies hold the potential to convert renewable lignocellulosic biomass resources into high-value-added platform compounds or biofuels [1]. Coffee is among the most popular beverages globally. According to statistics [2], the global consumption of coffee beans in 2023 was 177 million bags (each bag weighing 60 kg), totaling approximately 10 million tons. Moreover, each ton of coffee beans produces 650 kg of spent coffee grounds (SCGs) [3], resulting in an estimated 6.5 million tons of SCGs in 2023. SCGs are solid waste rich in caffeine, coffee melanin, and phenolic compounds [4]. Once these compounds are introduced into the soil in large quantities, they become ecologically destructive, making large-scale composting of SCGs infeasible [5]. Most SCGs are either landfilled or incinerated, leading to significant losses of reusable resources, waste of land resources, and exacerbation of climate issues [6]. In summary, discovering a cost-effective way to utilize SCGs would address the challenge of high-value utilization of SCGs and contribute to achieving global carbon neutrality.

Second-generation bioethanol is a green fuel with significant potential that is produced from lignocellulosic feedstock [7]. A variety of lignocellulosic resources, such as vine pruning residue, sugarcane bagasse, bamboo, poplar, and wheat straw, have been increasingly used for bioethanol production, with glucose and xylose being the primary components [8,9,10,11,12]. On one hand, both C5 and C6 sugars are utilized simultaneously in the fermentation process to maximize the efficiency of lignocellulose utilization [13]. However, common industrial yeast strains ferment C5 inefficiently, resulting in low yields [14,15]. Compared to most lignocellulosic sources, SCGs predominantly contain mannose, glucose, and galactose, with minimal C5 sugars [16]. Furthermore, it has been demonstrated that glucose and mannose derived from the hydrolyzed products of SCG can be fermented to produce ethanol using yeast [17,18]. Theoretically, the hydrolyzed products of SCGs can facilitate rapid fermentation, thereby increasing the efficiency of cellulosic ethanol production.

Lignocellulose, a major component of plant cell walls, has a complex three-dimensional structure that is recalcitrant to deconstruction. In this structure, the self-association and crystallinity of cellulose, in conjunction with the compaction and aggregation of lignin, effectively preclude the enzyme from accessing its internal cleavable bonds [19]. Pretreatment is an effective method for destructing the dense structure of lignocellulose, thereby enhancing the conversion of lignocellulosic resources [20]. The main pretreatment methods include physical, chemical, and physicochemical techniques. The pretreatment of SCGs is typically accomplished through various methods, including acid and alkali pretreatment, gas explosion pretreatment, potassium permanganate pretreatment, ferric chloride pretreatment, and H₃PO₄-acetone combined ammonia fiber explosion pretreatment [21,22,23,24,25,26]. Kwon et al. [27] developed a method for the co-production of bioethanol and biodiesel from SCGs, yielding 22 g/L of ethanol and 98% biodiesel, thereby demonstrating the hierarchical utilization of SCG. López-Linares et al. [28] employed microwave-assisted dilute sulfuric acid (1.5%) to hydrolyze SCGs, recovering 79% hemicellulosic sugar and 100% glucose at 160.47 °C from the pretreatment liquid. Although this method optimizes the recovery of fermentable sugars from SCGs, the development of industrial-scale microwave equipment remains a significant challenge. Titiri et al. [22] pretreated the SCG with 0.06% (w/v) NaOH at 99.5 °C for 1 h, resulting in the enzymatic hydrolysis of the remaining alkaline pretreated solids, which yielded 79.4% glucan, 74.5% mannan, and 54.8% hemicellulose hydrolysis. Recently, the application of deep eutectic solvent (DES) pretreatment has been regarded as an effective methodology for enhancing the utilization efficiency of lignocellulose [12,29]. The fundamental principle involves dissolving the lignin and separating the cellulose from the lignocellulose, thus increasing the accessibility of cellulose to enzymes. Liu et al. [7] pretreated sugarcane bagasse with triethylbenzyl ammonium chloride/lactic acid (TEBAC/LA) DES at 120 °C for 4 h, removing 80% lignin and 78% xylan. However, along with the removal of lignin, some hemicellulose is inevitably lost, as with alkali pretreatment. It has been demonstrated that the pretreatment of SCGs with DES and alkali does not fully retain the sugars present in SCGs, which are rich in hemicellulose. Consequently, the key issue in the pretreatment of SCGs is effectively disrupting the structure while maximizing the retention of cellulose and hemicellulose.

Liquid hot water (LHW) pretreatment is a process that disrupts the structure of lignocellulose by hydrogen ions generated by water ionization at high temperatures, using only water as the reagent [30]. After pretreatment, the hydrolysate was subjected to whole-slurry saccharification and fermentation in the same reactor [31]. The advantage of this method is that it reduces equipment usage while eliminating sugar loss during transfer [32]. Additionally, hemicellulose can be dissolved in LHW, effectively separating it from lignocellulose and disrupting the overall structure [31]. For the pretreatment of SCGs, LHW is a superior choice due to its high retention efficiency of lignocellulose, low cost, low energy consumption, and minimal environmental impact. The process involves two key steps: LHW pretreatment, separate hydrolysis, and fermentation (SHF). After LHW pretreatment, bioethanol is produced using the SHF process. This study aims to utilize high-temperature liquid water for the pretreatment of SCGs and the SHF process for the efficient preparation of bioethanol. The goal is to develop a high-efficiency, low-energy, water-saving, environmentally friendly ethanol preparation process. By comprehensively utilizing cellulose and hemicellulose from SCGs through enzyme fermentation, the work seeks to improve the utilization rate of lignocellulose and achieve the high-value utilization of SCGs.

2. Materials and Methods

2.1. Materials

SCGs were generously provided by a Starbucks coffee shop in Guangzhou, China. The SCGs were dried at 105 °C for 24 h to achieve a moisture content of less than 3% and then stored at −20 °C until use. The primary components of untreated and treated SCGs (cellulose, hemicellulose, and lignin) were quantified using standard procedures for biomass analysis at the U.S. National Renewable Energy Laboratory (NREL) [33]. The main composition was as follows (% w/w): cellulose, 14.10 ± 0.54%; hemicellulose, 36.15 ± 0.49% (mannose, 0.30 ± 0.00 g/g SCG; galactose, 0.10 ± 0.00 g/g SCG; xylose and arabinose monomers were not detected); acid-insoluble lignin (AIL), 39.48 ± 0.01%; and acid-soluble lignin (ASL), 4.71 ± 0.02%. Celluclast 2.0 L (75 FPU/mL) was provided by Novozymes (Beijing, China) Investment Co., Ltd. (Beijing, China). β-mannanase (50,000 U/g) was provided by Macklin Biochemical Technology Co., Ltd. (Shanghai, China). These enzymes were used for saccharification. The ethanol-fermentation active dry yeast (Saccharomyces cerevisiae CCTCC M94055(AQ)) [34] used in this study was kindly provided by Angel Yeast Co., Ltd., located in Yichang, China.

2.2. Washing of Spent Coffee Grounds

To exclude the effect of coffee melanin on the experiment, the extractives from SCGs were removed by a Soxhlet extraction system using ultrapure water and absolute ethanol in two sequential stages [35]. The extractive-free SCG sample was dried at 105 °C (moisture content < 3%), then stored at −20 °C until use.

2.3. Liquid Hot Water (LHW) Pretreatment

LHW pretreatment was carried out in a 100 mL high-pressure autoclave (Anhui Kemi Instrument Co., Ltd., Hefei, China). The experiments were performed at two different temperatures (170 and 180 °C) with varying pretreatment times (20, 40, and 60 min) at five different solid-to-liquid ratios (SLR) (1:6, 1:9, 1:12, 1:15, and 1:18 w/v). After the pretreatment, the reactor was cooled to room temperature naturally. In this study, the pretreatment was initially conducted at different temperatures and times with an SLR of 1:15 (w/v). Subsequently, the optimal temperature and time conditions were determined for different SLRs.

2.4. Separate Enzymatic Hydrolysis and Fermentation

Following pretreatment, the whole slurry of SCG hydrolysate was directly retained in a reactor for subsequent saccharification and fermentation. Enzymatic hydrolysis experiments were carried out in a shaking incubator (Shanghai Zhichu Instrument Co., Ltd., Shanghai, China) at 50 °C and 150 rpm for 48 h, utilizing various β-mannase loadings (1000, 2000, 3000, 4000, and 5000 U/g SCG) along with a cellulase loading of 24 FPU/g SCG. Furthermore, the effects of different pretreatment conditions on cellulose and hemicellulose conversion, as well as the yields of glucose, mannose, and galactose, were investigated at enzyme loadings of 24 FPU/g SCG and 5000 U/g SCG.

Yeast was inoculated into a fermentation medium and incubated at 30 °C for 18 h to prepare a seed medium, which contained 10 g/L yeast extract, 20 g/L peptone, 20 g/L glucose, 1.5 g/L KH₂PO₄, 4 g/L (NH₄)₂SO₄, and 0.5 g/L MgSO₄ [31]. Prior to the commencement of fermentation, nutrient supplements and the seed medium (10% v/v) were added to the slurry. Fermentation was conducted following the completion of enzymatic hydrolysis under conditions of 30 °C and 100 rpm for 12 h.

2.5. Analytical Methods

2.5.1. High-Performance Liquid Chromatography (HPLC)

The concentrations of ethanol, formic acid, acetic acid, furfural, 5-hydroxymethylfurfural (5-HMF), and levulinic acid were determined by HPLC (Shimadzu LC-10ATvp, Tokyo, Japan) equipped with a refractive index detector (RID) and an Aminex@HPX-87H Column (300 mm × 7.8 mm, Bio-Rad Laboratories, Hercules, CA, USA) at 55 °C. The mobile phase used was 5 mM sulfuric acid at a flow rate of 0.5 mL/min. For the determination of glucose, galactose, and mannose, an Aminex@HPX-87P Column (300 mm × 7.8 mm, Bio-Rad Laboratories, Hercules, CA, USA) at 80 °C was utilized with ultrapure water as the eluent at a flow rate of 0.4 mL/min. In addition, all samples were filtered using 0.2 μm nylon filters prior to analysis.

The equations used to calculate the conversion of glucose, mannose, and total fermentable sugars via enzymatic hydrolysis, ethanol yield, productivity, mass yield, and conversion are as follows:

$$\mathrm{Glucose} \mathrm{conversion} (\%)={\displaystyle \frac{{\mathrm{c}}_1\times {\mathrm{V}}_1}{{\mathrm{m}}_1\times 1.111}}\times 100\%$$

(1)

$$\mathrm{Mannose} \mathrm{conversion} (\%)={\displaystyle \frac{{\mathrm{c}}_2\times {\mathrm{V}}_1}{{\mathrm{m}}_2\times 1.136}}\times 100\%$$

(2)

where c₁ and c₂ (g/L) are the concentrations of glucose and mannose in the enzymatic hydrolysate, respectively; V₁ (L) is the volume of the hydrolysate; and m₁ and m₂ (g) are the masses of cellulose and mannan in the substrate, respectively. Fermentation experiments of the pretreated SCB were implemented using separate hydrolysis and fermentation (SHF).

$$\mathrm{Ethanol} \mathrm{yield} (\%)={(\mathrm{c}}_3+{\mathrm{c}}_4)\times 0.51\times 100\%$$

(3)

$$\mathrm{Ethanol} \mathrm{productivity} (\mathrm{g}/(\mathrm{L}·\mathrm{h})={\displaystyle \frac{{\mathrm{c}}_5}{\mathrm{t}}}$$

(4)

$$\mathrm{Ethanol} \mathrm{massyield} (\mathrm{g}/({\mathrm{g}}_{\mathrm{SCG}})={\displaystyle \frac{{\mathrm{c}}_5\times {\mathrm{V}}_2}{\mathrm{t}}}$$

(5)

$$\mathrm{Ethanol} \mathrm{conversion} (\%)={\displaystyle \frac{{(\mathrm{c}}_5\times {\mathrm{V}}_2)\times {\mathrm{m}}_4}{{\mathrm{m}}_3}}\times 100\%$$

(6)

where c₃ is the initial concentration of glucose in the fermentation broth (g/L); c₄ is the initial concentration of mannose in the fermentation broth (g/L); c₄ is the initial concentration of mannose in the fermentation broth (g/L); c₅ is the final concentration of ethanol in the fermentation broth (g/L); t is the time of the fermentation (h); V₂ is the volume of the fermentation broth (L); m₃ is the mass of the raw material (g); and m₄ is theoretical mass of ethanol (g).

2.5.2. X-ray Analysis

The crystallographic behavior of samples before and after LHW pretreatment and enzymatic hydrolysis was determined using an X-ray diffractometer (Rigaku Miniflex 600, Tokyo, Japan) operating at 40 kV and 45 mA utilizing a Cu Kα radiation source. The samples were scanned over a range of 3–90 degrees (2θ), with a step size of 0.01 degrees and a scanning speed of 10 degrees per minute.

2.5.3. Fourier Transform Infrared (FT-IR) Analysis

The presence of functional groups in untreated, hydrothermally pretreated, and enzymatic hydrolysis SCG samples was analyzed using Fourier Transform Infrared (FT-IR) spectroscopy (Thermo Fisher Scientific, Karlsruhe, Germany). The analysis was performed using the KBr pellet method, covering a frequency range of 4000 to 500 cm⁻¹ with a resolution of 4 cm⁻¹.

2.5.4. Scanning Electron Microscope (SEM) Analysis

The surface morphology of untreated, hydrothermally pretreated, and enzymatic hydrolysis SCG samples were analyzed by scanning electron microscopy (SEM, ZEISS Gemini 300, Jena, Germany). The SCG samples were sputter-coated with a thin layer of gold and mounted on carbon tape. The SEM analysis was conducted under an accelerating voltage of 5 kV and a working distance of 4.0 mm.

2.6. Statistical Analysis

The results of the experiment were statistically analyzed using analysis of variance (ANOVA), and p < 0.05 was considered significant.

3. Results and Discussion

3.1. Effects of LHW Pretreatment Conditions on Enzymatic Hydrolysis

3.1.1. Pretreatment Temperature and Retention Time

In LHW pretreatment, it is essential to investigate the optimal temperature and retention time conditions, as these parameters significantly influence the solubilization effect of hemicellulose, the disruption of the structure of SCG, and the production of inhibitory compounds. Therefore, identifying the appropriate reaction temperature and time is beneficial for enhancing the subsequent enzymatic conversion of SCG and increasing ethanol yield during fermentation.

The effects of LHW pretreatment temperature and retention time on glucose and mannose conversion during the enzymatic hydrolysis process were significant (p < 0.05, Table S1). As shown in

Table 1. Effects of liquid hot water pretreatment temperature and time on enzymatic hydrolysis and fermentation of SCG.

Pretreatment	Conversion (%)		Ethanol
	Glucose	Mannose	Concentration (g/L)	Yield b (%)	Mass Yield c (mg/g SCG)
170 °C—20 min	73.52 ± 1.45 a	42.64 ± 2.11	6.95 ± 0.15	93.80 ± 0.17	116.09 ± 0.10
170 °C—40 min	80.78 ± 1.75	42.21 ± 0.28	7.00 ± 0.10	90.61 ± 2.19	117.53 ± 0.07
170 °C—60 min	81.85 ± 2.20	41.04 ± 0.51	7.05 ± 0.36	91.31 ± 3.10	124.02 ± 0.08
180 °C—20 min	81.05 ± 1.86	44.66 ± 1.34	7.25 ± 0.08	89.67 ± 4.13	124.81 ± 0.03
180 °C—40 min	81.86 ± 1.27	38.98 ± 3.06	6.94 ± 0.07	91.49 ± 4.25	119.96 ± 0.03
180 °C—60 min	83.15 ± 0.24	30.53 ± 0.50	6.32 ± 0.17	94.69 ± 3.20	112.80 ± 0.04

^a Data: means ± standard deviations. ^b Yield: Percentage of glucose and mannose converted to ethanol in the fermentation broth. ^c Mass yield: Amount of ethanol produced per gram of SCG.

, glucose conversion gradually increased with prolonged retention time and higher pretreatment temperature, reaching a maximum conversion of 83.15% at 180 °C for 60 min. This is likely due to the high crystallinity of cellulose, which presents a barrier to structural disruption by hot water [36]. At 170 °C, mannose conversion did not change significantly with increasing retention time, suggesting that an extended retention period at 170 °C may be required to achieve a significant pretreatment effect on hemicellulose. Conversely, at 180 °C, the conversion of mannose decreased from 44.66% to 30.53% as the retention time increased from 20 to 60 min, with maximum mannose conversion occurring at 180 °C for 20 min. The substantial decline in mannose conversion at 180 °C indicates a more destructive effect of hot water on hemicellulose, highlighting the importance of regulating the retention time to ensure optimal results.

Furthermore, the pronounced decline in mannose conversion with the increasing retention time suggests hemicellulose degradation, as evidenced by the observed changes in inhibitor levels, which are shown in Figure 1. Zheng et al. [31] reported a similar decline in hemicellulose conversion for sugarcane bagasse subjected to LHW at 180 °C, with a marked decrease in conversion when the pretreatment duration exceeded 20 min. These data indicate that a pretreatment temperature of LHW at 180 °C and a retention time of approximately 20 min are beneficial for reducing the sugar loss of hemicellulose. Najafi et al. [37] found that LHW pretreatment of olive pomace at 180 °C removed 13% of xylan. Comparatively, Romero-García et al. [38] pretreated olive tree pruning at 180 °C, 200 °C, and 220 °C for 5 min using LHW, resulting in xylose concentrations in the liquid phase of 22.95 ± 0.25 g/L, 6.72 ± 0.17 g/L, 0.09 ± 0.0 g/L, respectively. These findings demonstrate that hemicellulose is poorly retained due to rapid degradation at elevated temperatures. In conclusion, hot water at 180 °C effectively solubilizes hemicellulose, but it is necessary to control the retention time to prevent sugar degradation.

The effects of different temperatures and retention times on the inhibitor concentration are demonstrated in Figure 1. All pretreatment conditions exhibited low inhibitor concentrations. At each pretreatment temperature, the inhibitor content increased with longer retention time, indicating that inhibitor production can be minimized at shorter retention times. Furthermore, furfural was only produced at retention times of 40 and 60 min, as its formation in water requires a longer reaction time [39]. Furfural is known to have a significant inhibitory effect on the yeast fermentation process [40]. Therefore, shorter pretreatment durations offer advantages for subsequent fermentation, such as reduced inhibitor content, shorter pretreatment time, and minimized sugar loss.

To further examine the effect of pretreatment, the fermentation outcomes were compared. The experimental results of whole slurry fermentation are shown in Table 1. The highest ethanol concentration of 7.25 ± 0.08 g/L was achieved under pretreatment conditions of 180 °C for 20 min, along with the highest mass balance effect of 124.81 ± 0.03 mg/g SCG. Similar results were obtained under the condition of 170 °C for 60 min, where the highest mannose conversion, ethanol concentration, and mass accounting were observed. In addition, the inhibitor concentration remained relatively low, with a total inhibitor concentration below 1 g/L, indicating the potential for enhanced performance. In conclusion, it can be posited that pretreatment at 180 °C for 20 min is more efficacious than at 170 °C for 60 min in terms of SCG pretreatment. Furthermore, the hydrolysate produced exhibited enhanced biocompatibility due to the low inhibitor concentrations. As evidenced by the preceding analysis, the application of LHW at 180 °C for a 20 min period was selected for subsequent experiments.

3.1.2. Solid-to-Liquid Ratio

During pretreatment, the SLR directly influences the fermentable substrate and inhibitor content, which in turn indirectly affects the fermentation result. Moreover, the cost of subsequent ethanol separation is dependent on the ethanol concentration and can be reduced as the ethanol concentration increases. A significant proportion of the expense associated with ethanol production is attributable to the separation process, with the cost of distillation columns being closely correlated with the ethanol concentration. To enhance product concentration, productivity, and recovery efficiency, as well as to reduce consumption of freshwater resources and electricity, higher substrate concentrations are often required in industrial fermentation applications [41].

Hence, the effects of different SLRs on enzymatic digestion and fermentation were investigated. As shown in Table S2, it’s noteworthy to mention that the effects of SLRs on glucose conversion, mannose conversion, and ethanol yield were significant (p < 0.05). The enzymatic conversion of glucose and mannose showed a decreasing trend with increasing solids loading (see Figure 2A). As expected, the highest enzymatic conversions of glucose and mannose, 85.37 ± 1.04% and 43 ± 1.76%, respectively, were achieved at an SLR of 1:18. This is attributed to the lower viscosity of the system at low solids loading, which facilitates contact and mass transfer between the solid and liquid phases, resulting in improved enzyme interaction with cellulose and hemicellulose. More effects of different SLRs on fermentation are shown in Table S3. As the solids content increased, the ethanol mass yield and conversion tended to decrease. Comparing with the maximum theoretical ethanol mass yield (235.88 mg/g SCG), the maximum mass yield of ethanol prepared from SCG was 121.86 ± 4.45 mg/g SCG, with a maximum conversion of 58.13 ± 0.17%. Comparing the ethanol mass yield and conversion of the 1:6 and 1:18 groups, the differences were 14.31 mg/g SCG and 10.45%, respectively. However, when the SLR was set at 1:6, the maximum ethanol concentration achievable was 15.87 ± 0.17 g/L, while the maximum productivity was 1.322 ± 0.014 g/L. These values are 164.5% higher than those observed in the 1:18 group. Conversely, the pretreatment of the same mass of SCG at a high SLR of 1:6 demonstrated a notable reduction in water usage, at a rate of 66.7% compared to the 1:18 condition. The combination of a higher product concentration, a higher productivity, and lower water consumption was achieved through fermentation with a high SLR, which suggests the potential for enhanced production efficiency. Additionally, there was no discernible change in ethanol yield at SLRs ranging from 1:18 to 1:9, as shown in Figure 2B. There was a positive correlation between ethanol concentration and solid loading. Notably, the study revealed that the maximum yield of 89.63 ± 1.39% was observed at the SLR of 1:6, which was significantly higher than the yield observed under other conditions. This may be due to the higher substrate concentration at increased solids loading, allowing the yeast to better adapt to the hydrolysate environment.

The study examined the impact of various SLRs on SCG hydrolysates, revealing that whole-slurry fermentation was successfully achieved, as shown in Figure 3A–E. Hydrolysates with SLRs ranging from 1:18 to 1:9 completed fermentation within 6 h, while those with an SLR of 1:6 required approximately 9–12 h. The variation in fermentation time can be attributed to the concentration of substrates and inhibitors present in the hydrolysate. During fermentation, yeast metabolized glucose prior to mannose, with a faster metabolism rate for both sugars, aligning with the metabolic curve reported by Burniol-Figols et al. [42]. It is generally believed that direct whole-slurry fermentation at high SLRs is challenging due to the presence of high levels of inhibitory substances [43]. However, the highest ethanol concentration (15.87 ± 0.24 g/L) was observed at an SLR of 1:6. Although inhibitor levels increased with higher solids contents, the total inhibitor content remained below 2.0 g/L at an SLR of 1:6. Notably, as depicted in Figure 4, the hydrolysate at an SLR of 1:6 completed fermentation without water washing or detoxification in approximately 12 h. Cao et al. [44] reported that LHW pretreatment of corn stover at an SLR of 1:10, 190 °C, and 15 min yielded 2.2 g/L acetic acid, 0.1 g/L 5-HMF, and 0.7 g/L furfural. Zheng et al. [31] pretreated sugarcane bagasse at an SLR of 1:6, but did not achieve the desired fermentation outcome. Conversely, Xian et al. [34] successfully achieved fermentation following detoxification of high-SLR hydrolysate of sugarcane bagasse using chromatographic separation with resins such as SY-01. Compared to other lignocellulosic resources, the high SLR hydrolysate obtained from pretreating SCG with LHW exhibited lower inhibitor levels. Unlike many previous studies, the hydrolysate of SCG prepared at an SLR of 1:6 could be directly fermented without solid–liquid separation, water washing, or detoxification. The results demonstrate that the hydrolysate of SCG prepared under high SLR conditions retained biocompatibility, a quality highly desirable in numerous applications.

3.1.3. Optimization of Enzymatic Hydrolysis Process

Although SCG did not yield the most favorable enzymatic conversion results at an SLR of 1:6, it exhibited higher ethanol concentrations and conversion rates compared to an SLR of 1:18. Thus, high SLR pretreatment combined with whole-slurry saccharification and fermentation in one pot can reduce water consumption in ethanol production and avoid the generation of pretreated wastewater. This advancement presents a novel avenue for commercialization, addressing the long-standing challenge of elevated water consumption inherent to hydrothermal processes [45]. Moreover, from an economic standpoint, the production costs may be more favorable, making this method a potentially advantageous choice for ethanol production. Therefore, an SLR of 1:6 was selected for the subsequent experiments.

One of the primary limitations in cellulosic ethanol production is the high cost of enzymes. To improve the economics of cellulosic ethanol production, experimental optimization of β-mannase loadings (1000, 2000, 3000, 4000, and 5000 U/g SCG) was performed under the previously established optimal conditions: pretreatment temperature of 180 °C, pretreatment time of 20 min, and an SLR of 1:6. The outcomes of mannose conversion at varying β-mannase loadings were significant (p < 0.05, Table S2). As illustrated in Figure 5A, the concentration of mannose exhibited a rapid increase from 0 to 12 h, stabilizing after 36 h. Comparing enzymatic digestion results at 48 h, the mannose concentration obtained from 1000–2000 U/g SCG was significantly lower than that from 3000–5000 U/g SCG. Further analysis of the conditions with 3000–5000 U/g SCG showed that the mannose concentrations for these three conditions were relatively close, with values of 19.67 ± 0.09 g/L, 20.35 ± 0.43 g/L, and 20.03 ± 0.08 g/L and conversion rates of 39.21 ± 0.19%, 40.58 ± 0.87%, and 37.2 ± 4.05%, respectively, as shown in Figure 5B. Finally, considering both the overall economic benefits and the mannose conversion rate, a β-mannase dosage of 3000 U/g SCG was selected for subsequent study.

3.2. Mass Balance

To assess the conversion process of the preparation technology in this work, a mass balance experiment was conducted under the optimal conditions for the SHF process: pretreatment at 180 °C for 20 min, an SLR of 1:6, saccharification, and β-mannanase loading of 3000 U/g SCG. As shown in Figure 6, both glucan and mannan were retained in the liquid and solid fractions throughout the preparation process, demonstrating the advantages of whole-slurry preparation. Furthermore, the ethanol yield from LHW-pretreated SCGs was 71.92 ± 1.3 mg/g SCG higher than that from untreated SCGs. The total fermentable sugar and ethanol yields of SCGs pretreated with LHW were enhanced by 221.21% and 185.78%, respectively, compared to raw SCG material. This demonstrates that LHW pretreatment facilitated the utilization of cellulose and hemicellulose from SCGs, thereby enhancing the efficiency of SCGs for cellulosic ethanol production.

3.3. Bioethanol Generartion Analysis

The bioethanol mass yield (101.79 ± 0.23 mg/g SCG) based on the optimal conditions (180 °C, 20 min, SLR at 1:6) of this process, in conjunction with the global amount of SCGs, was estimated as detailed in Table S4 of the Supplementary Materials. Based on the global coffee consumption (177 million bags (each bag weighing 60 kg)) outlined in the introduction section, assuming that all these beans were used for coffee production, 6.9 million tons of SCGs would be produced. The estimated yield of bioethanol is 702,351 tons (converted to 890 million liters by volume). Furthermore, global bioethanol production and consumption are relatively close to each other, and yield a more accurate comparison. Compared to global bioethanol production in 2023 [46], if all SCGs were used to produce bioethanol in 2023, it would account for 1.52% of global production.

3.4. Structural Characterization Studies

3.4.1. FT-IR Analysis

The chemical bond structure and functional groups of the untreated, LHW-pretreated, and enzymatically hydrolyzed SCG were investigated by FT–IR analysis, as shown in Figure 7. The broad peak near 3000–3600 cm⁻¹ is attributed to the stretching of large amounts of –OH in cellulose and hemicellulose. The bands at 2926 and 2854 cm⁻¹ correspond to the asymmetric stretching of C–H bonds. The new peak at 1743 cm⁻¹ is associated with C=O stretching vibrations [47]. The absorption bands between 1500 and 1700 cm⁻¹ are attributed to C=O stretching, C–N stretching, and N–H bending in protein molecules [48]. The decrease in peaks near 1743 cm⁻¹ and 1500–1700 cm⁻¹ after enzymatic hydrolysis of SCG indicates the removal of hemicellulose. Additionally, two absorption phenomena were observed in the SCG spectra, with ion peaks at 1633 and 1603 cm⁻¹, which are associated with caffeine residues in the substance [49]. The peak at 779 cm⁻¹ is attributed to stretching of β–D–mannopyranose unit bonds [50,51]. The presence of this peak in the spectrograms of all three samples indicates that neither pretreatment nor enzymatic hydrolysis completely separated the mannan from the SCGs, consistent with the mass balance data presented in Section 3.2.

3.4.2. SEM Analysis

To facilitate a more comprehensive comparison of the alterations in the surface structure of coffee grounds following pretreatment and enzymatic hydrolysis, untreated SCGs, LHW-pretreated coffee grounds, and enzymatic hydrolysis SCGs were observed and analyzed via SEM. As shown in Figure 8A–C, the untreated SCG particles have flatter surfaces with fewer gaps. Further magnification in Figure 8C reveals that the surfaces of SCGs have many uneven basin-like structures, yet can still be considered relatively flat and dense. After LHW pretreatment, the SCG particles exhibited flakes of broken components and numerous folds (see Figure 8D–F). These folds enhanced the surface area of the SCG, creating a conducive environment for enzymes to more readily access the cellulose and hemicellulose. Upon further comparison, the enzymatically hydrolyzed SCGs (see Figure 8G–I), displayed an increasing number of fragmented and fractured particles. This phenomenon is attributed to the continued degradation of hemicellulose and cellulose facilitated by the enzyme.

3.4.3. X-ray Analysis

To evaluate the changes in crystallinity, the XRD spectra of the raw material, LHW-pretreated SCGs, and enzymatically digested SCGs were compared. The XRD spectra of SCGs align with the findings of Batista et al. [52] and Ravindran et al. [26]. To ascertain the distinctions between the treatments, the intensities of the peaks at 2θ ≈ 20° in the samples’ XRD spectra were compared, as peak intensity is regarded as an indicator of the degree of crystallinity [52,53]. Generally, the crystallinity of lignocellulose is primarily contributed by the cellulose fraction, while hemicellulose and lignin are predominantly amorphous fractions. As shown in Figure 9, the peak at 2θ ≈ 20.5° in the LHW-pretreated SCG exhibited a notable enhancement compared to the raw material. This suggests that the hydrothermal pretreatment effectively removed a portion of the amorphous hemicellulose and lignin from the material, which is consistent with the experimental results presented in Section 3.2. The peak at 2θ ≈ 16° is considered an amorphous region in lignocellulose, mainly composed of hemicellulose [24]. After enzymatic digestion, there was a significant decrease in the peak values of both 2θ ≈ 16.5° and 2θ ≈ 20.5°, which correlated with the removal of cellulose and hemicellulose. The remaining residue contained components including 2.78 ± 0.15% cellulose and 8.35 ± 0.49% mannan.

4. Conclusions

SCGs have shown significant promise for high-SLR cellulosic ethanol production due to the low levels of inhibitors in the hydrolysate following LHW pretreatment. Optimal conditions of 180 °C for 20 min at an SLR of 1:6 yield 17.95 ± 0.23 g/L glucose and 19.67 ± 0.13 g/L mannose after saccharification. In addition, the hydrolysate was successfully fermented to produce 15.02 ± 0.05 g/L ethanol with a high productivity of 1.252 g/(L·h). Moreover, the bioethanol mass yield was 101.79 ± 0.23 mg/g SCG. Comparing the maximum theoretical mass yield, the ethanol produced via this process from SCG exhibited a conversion of 43.2%. Based on the ethanol mass yield of this process and the 2023 coffee consumption data, it is estimated that 554 million liters of bioethanol can be produced. This study highlights that LHW pretreatment of SCGs at a high SLR of 1:6, followed by separate hydrolysis and fermentation, not only achieves rapid and efficient pretreatment and fermentation, but also reduces water usage by 60% (compared with an SLR of 1:15) and increases product concentration. These advancements significantly enhance the potential for the industrialization and commercialization of bioethanol production processes using SCGs, contributing to the sustainable and high-value utilization of this abundant waste resource.