Owing to a significant increase in biodiesel production, the search for sustainable and non-edible feedstocks for biodiesel production, such as microalgae [1,2], sewage sludge [3,4], waste coffee grounds (WCGs) [5,6], and waste cooking oil (WCO) [7,8], has gained importance. Among these feedstocks, WCO does not require a lipid separation process, which requires a large amount of extraction solvent; biomass such as microalgae, sewage sludge, and WCGs requiring lipid separation generally results in the discharge of a large amount of wastewater and biomass debris for disposal [7-10]. According to Korean government statistics, the number of franchised fried chicken stores, the main source of WCO, soared from approximately 9,000 in 2002 to 22,529 in 2013, 25,687 in 2019, and 28,627 in 2021 [11].

Catalysts are essential for accelerating the transesterification reaction in biodiesel production [8-10]. Alkaline catalysts facilitate high reaction rates and require much lower amounts of methanol and shorter reaction times than acidic catalysts [7,12]. However, alkaline catalysts cannot be used in feedstock with a high content of free fatty acids (FFAs) because they cause saponification of the FFAs. According to previous studies, lipids with FFA levels exceeding 0.5% [13,14], 1.0% [15-17], or 2.0% [18] undergo distinct saponification in the presence of alkaline ions. Another study reported that alkaline catalysts could be used when the FFA content was < 3.0% [19,20]. We previously reported that biodiesel is successfully produced from WCG lipids containing 1.9% FFAs using alkaline catalysts [10], while WCO containing 4.2% FFAs resulted in a very low biodiesel conversion (4.0%) with alkaline catalysts. Thus, we proposed a novel two-step process in which the first step aims to lower the FFA content to an acceptable level (e.g., 1.5%) for the use of alkaline catalysts in the second step [7]. As the FFA content threshold for the possible use of alkaline catalysts may depend on the lipid source, the use of alkaline catalysts should be preliminarily investigated for each individual feedstock [7]. Another issue with catalysts in the biodiesel process, from economic and environmental viewpoints, is their reuse. Homogeneous chemical catalysts, such as sodium hydroxide (NaOH) and sulfuric acid (H₂SO₄), have been used extensively in industry- and laboratory-scale studies because they are relatively inexpensive and can produce biodiesel with high reaction rates [21,22]. Despite their extensive use, these chemical catalysts cannot be reused because they are homogeneously dispersed in the reaction mixture and become nonseparable [23]. Accordingly, processes using these chemical catalysts require the repeated addition of catalysts in every batch of the operation. Additionally, they necessitate large amounts of water for the neutralization and washing of biodiesel, inevitably leading to the discharge of a significant volume of wastewater [24,25]. In contrast to homogenous chemical catalysts, heterogeneous solid catalysts have the significant advantages of possible reuse and easy separation from the reaction mixture, which would substantially decrease the catalyst requirements and wastewater discharge [23-25]. We previously prepared solid alkaline catalysts for biodiesel production by calcining waste eggshells and waste scallop shells (WSS) for biodiesel production from the WCGs [5,22]. In particular, a large amount of WSS have been discharged in the city of Gangneung located on the East seashore of Korea, and its treatment has become a perplexing problem—most of these shells are currently being landfilled [5]. Therefore, the recycle of WSS as solid catalysts holds significant importance for waste management in this city.

To easily separate the solid catalysts from the WCG debris after each round of biodiesel production, we devised a cartridge containing the solid catalysts. Biodiesel can be easily produced from WCGs by removing the cartridge from a reactor after biodiesel production and transferring it to another reactor containing fresh WCGs, methanol, and an extraction solvent [5,22]. Despite the distinct advantages of adopting a cartridge, the mass-transfer limitation, long reaction time, and requirement of large amounts of methanol in this process are difficult to overcome. Unlike WCGs, WCO does not require a cartridge because the solid catalysts and reaction mixtures can be readily separated from each other by settling the solid catalysts.

In this study, WCO and WSS discharged from local restaurants were recycled as feedstock for biodiesel production and sources of solid catalysts, respectively. The calcination conditions of the WSS for the preparation of the solid catalysts were optimized, and various treatment methods for the reuse of the solid catalysts were evaluated. The reaction conditions for biodiesel production using the solid catalysts and WCO were determined, and the solid catalysts were reused for the repeated production of biodiesel from WCO.

2.1. Materials and chemicals

WCO was collected from three fried chicken restaurants near Gangneung-Wonju National University (GWNU). After simple filtration to remove the solid substances, the WCO was used for biodiesel production. WSS were collected at a local seafood restaurant near GWNU. After carefully removing the impurities attached to the WSS, they were washed several times with distilled water. Following this, they were dried in an oven at 105°C for 12 h and ground using a MicroHammer mill (MFC CZ 13; Culatti AG, Switzerland) [5]. To determine the optimal calcination conditions, 20 g of WSS was calcined at various temperatures and times to prepare the WSS-based alkaline solid catalysts (WASCs) in an electronic furnace (FX-03; DAIHAN Scientific, Korea) [22]. Analytical grade methanol (99% purity, Fisher Scientific, UK), n-hexane (96% purity, Showa, Japan), sulfuric acid (99% purity, Showa, Japan), and sodium hydroxide (98% purity, Daejung, Japan) were used. Standard materials for gas chromatography (GC) (methyl palmitate, methyl oleate, methyl linoleate, methyl stearate, and methyl heptadecanoate) were purchased from Sigma-Aldrich (USA).

2.2. Catalyst characterization and biodiesel analysis

The composition of the WASCs were analyzed using an Xray fluorescence spectrometer (XRF) (ZSX100e, Rigaku, Tokyo, Japan) [5]. The size distribution of the WASCs used in this study was determined using a particle size analyzer (MasterSizer 2000, Malvern Panalytical Ltd., UK). The FFA content of the WCO was measured as previously described [7]. The biodiesel conversion was determined using a GC (7850 A, Agilent, USA) equipped with an autoinjector (G4513 A, Agilent, China), an HP-5 column (30m × 0.32mm × 0.25 μm film thickness), and a flame ionization detector (FID) [5,22]. The temperatures of the injector and the detector were 250°C, and that of the column was increased from 150°C to 250°C at a rate of 5°C/min after the oven temperature was initially maintained at 150°C for 2 min. Helium was used as the carrier gas. Methyl heptadecanoate was used as the internal standard.

2.3. Biodiesel production and reuse of the WASCs

The biodiesel was produced in a 30 mL glass bottle with a Teflon-coated cap. This glass bottle reactor containing 1 g of WCO, methanol, and WASC was placed in a shaking incubator (SI-600R, JEIO Tech, Seoul, Korea) and rotated in orbital mode for biodiesel production. The catalyst concentration (wt%) was defined as the weight of the WASC relative to that of the WCO. In the experiments using homogeneous catalysts, after the reaction was completed, the reaction mixture was cooled to room temperature, and 2 mL of distilled water was added to the bottle. Biodiesel was then extracted twice with 10 mL of n-hexane by vigorously agitating the mixture for 1 min [5,22]. After centrifuging the mixture at 23 relative centrifugal force (RCF) for 10 min (Gyro 1730MG, Gyrozen, Daejeon, Korea), the organic layer was carefully separated with a syringe (Kovax-Syringe, Korea Vaccine, Korea) to measure the biodiesel conversion by GC analysis.

In the experiments on the reuse of the WASCs, the shaking incubator was stopped after the reaction and maintained for 5 min to allow the solid catalysts to settle, and the liquid reaction mixture was separated from the reactor using a syringe. The solid catalysts were then reused in situ or ex situ. For the in-situ mode, fresh WCO and methanol were introduced into the reactor, where the WASC used in the previous round remained for the next round of biodiesel production. For the ex-situ mode, the WASC was also recovered and treated using one of the following methods: simple drying, washing with methanol and/or n-hexane, or washing with methanol and/or n-hexane followed by drying, and then placed into a reactor containing fresh WCO and methanol for the next round of biodiesel production. All the experiments were conducted in triplite, and the data are presented as the mean ± standard deviation.

3.1. Effect of catalyst type on biodiesel production

The FFA content of the WCO from the three restaurants was in the range of 1.5-1.9%. Compared to the FFA content of WCO reported in a previous study (4.2%), this value was significantly low [7]. The owners of the restaurants said that they were voluntarily reducing the reuse of cooking oil for frying their chickens to comply with the strict demands of Korean consumers, which would substantially reduce the FFA content of the WCO. To investigate the possible use of alkaline catalysts which have many advantages over acid catalysts, biodiesel was produced from the WCO under favorable reaction conditions (0.5 wt% catalysts, 15 mLmethanol/ g-WCO, 70ºC, 12 h reaction time, and 200 rpm) using acid (H₂SO₄) and alkaline (NaOH) catalysts, which are the most extensively used chemical catalysts [7]. As shown in Fig. 1, the biodiesel conversion using the acidic and alkaline catalysts were 20.0% and 86.2%, respectively. In a previous study using WCO with a 4.2% FFA content, the biodiesel conversions were only 16.7% for the acid catalysts and 4.0% for the alkaline catalysts under the same reaction conditions, indicating that the FFA content of the feedstock of biodiesel is a critical factor not only for the possible use of an alkaline catalyst but also for biodiesel conversion. Because the FFA content of the WCO was relatively low and sodium hydroxide is readily used for biodiesel production, WASCs was expected to be successfully used for biodiesel production from WCO in this study.

Figure 1. Effect of the catalyst type on biodiesel production from waste cooking oil (0.5wt% catalyst, 1.5 mL-methanol/g-WCO, 70ºC, 12h, 200 rpm). ▨: acid catalyst, ■: alkaline catalyst

3.2. Optimal calcination conditions for the manufacturing of the WASCs

The WASCs were prepared by the calcination of WSS, as described above, and the effects of calcination temperature and time on the calcination efficiency were investigated. The major component of WSS is calcium carbonate, which results in 98.3% of calcium oxide (CaO) in the WASC, as previously reported [5]. CaO, which acts as an alkaline catalyst in biodiesel production, is obtained by calcination at high temperatures, as described by the following chemical reaction:

According to the stoichiometric analysis, the weight loss of calcium carbonate after the complete calcination of calcium oxide is 44.0%. Based on this calculation, the calcination efficiency was defined as the percentage of weight loss due to calcination divided by 44.0%, as shown in Equation (2).

Table 1 presents the calcination efficiencies at different calcination temperatures and times. The calcination efficiency was slightly greater than 100% when components other than calcium carbonate were present in the WSS. The results show that the calcination efficiency increases as temperature and/or time increases, and reaches a value of over 95% at 700°C for 5 h, and 800°C for 1, 3, and 5 h. To investigate the performance of the WASCs as alkaline solid catalysts, biodiesel conversions from WCO using WASCs manufactured under various calcination conditions were also investigated as presented in Table 1. The reaction conditions were determined based on previous studies [23,26,27]; 3.0 wt% catalysts, 0.5 mL-methanol/g-WCO, 65°C, 12 h and 200 rpm. The intact WSS without calcination did not produce biodiesel, resulting in zero biodiesel conversion. WASCs manufactured at 600°C for 1 h resulted in a very low biodiesel conversion (2.5%) but much higher biodiesel conversions were obtained using WASCs manufactured at 600°C for 3 h (69.6%) and 600°C for 5 h (72.8%). When the calcination temperature was increased to 700°C, biodiesel conversion was significantly increased to 73.4%, 78.1%, and 79.7% for 1, 3, and 5 h of calcination time, respectively. When the calcination temperature was further increased to 800°C, biodiesel conversions were rather decreased to 68.9%, 68.4%, and 58.8% for 1, 3, and 5 h of calcination, respectively. Based on these observations, calcination at 750°C for 1, 3, and 5 h, and that at 800°C for 0.5 were additionally investigated. As shown in Fig. 2, the biodiesel conversions were 78.5%, 81.3%, and 80.3% when using WASCs manufactured at 750°C for 1, 3 and 5 h, respectively. The biodiesel conversion was 77.8% using a WASC manufactured at 800°C for 0.5 h. From these experimental data, the optimal calcination conditions were determined to be 750°C for 3 h. The size distribution of the WASCs were relatively unimodal, and the volume-weighted mean diameter of the WASC particles was 536 μm as shown in Fig. 3.

Figure 2. Effect of calcination temperature and time on biodiesel conversion. (3.0 wt% catalysts, 0.5 mL-methanol/g-WCO, 65 ℃, 12 h and 200 rpm)

Figure 3. Size distribution of the alkaline solid catalysts prepared by the calcination of waste scallop shells.

Table 1 Effect of calcination temperature and time on calcination efficiency and biodiesel conversion

Temperature (oC)	Time (h)	Calcination efficiency (%)	Biodiesel conversion (%)a
25	0	0	0.0
600	1	57.6 ± 1.8	2.5 ± 0.7
	3	59.3 ± 2.3	69.6 ± 1.3
	5	60.7 ± 2.1	72.8 ± 0.1
700	1	66.8 ± 3.5	73.4 ± 3.1
	3	80.8 ± 2.7	78.1 ± 0.5
	5	94.8 ± 0.9	79.7 ± 1.4
800	1	99.4 ± 1.2	68.9 ± 0.6
	3	99.8 ± 3.1	68.4 ± 2.3
	5	100.3 ± 1.1	58.8 ± 8.3

a: reaction conditions. 3.0 wt% catalysts, 0.5 mL-methanol/g-WCO, 65°C, 12 h and 200 rpm

3.3. Optimization of reaction conditions for biodiesel production

In the process of biodiesel production from WCO using solid catalysts, five variables should be considered: catalyst concentration, methanol loading, reaction temperature, reaction time, and agitation speed. When a cartridge containing solid catalysts is employed in the biodiesel production process, the agitation speed is a critical factor because the mass transfer rates of the reactants (oil and methanol) and products (biodiesel and glycerol) across the cartridge are very important [5]. When solid catalysts are directly added to a reaction mixture, complete mixing of the reaction mixture, complete suspension or no settlement of the solid catalyst, is important [28]. By the naked eye’s observation, the agitation speed, the number of shaking cycles in the orbital mode of the shaking incubator, should be at least 150 rpm to ensure complete mixing; thus the agitation speed was fixed at 150 rpm throughout the experiments. The other variables under the reaction conditions were optimized using the one-factor-at-a-time technique. The initial reaction conditions were set as stated above; 3.0 wt% catalysts, 0.5 mL-methanol/g-WCO, 65°C, 12 h, and 150 rpm.

3.3.1. Effect of catalyst concentration

In this study, the WASC concentration was set in the range of 0-10 wt% relative to the WCO. As presented in Fig. 4(a), the biodiesel conversion was 10.6% at 1.0 wt% of solid catalysts and dramatically increased to 53.4% at 2.0 wt% of solid catalysts, 82.5% at 3.0 wt% of solid catalysts, and further increased until 3.5 wt% of WASCs (86.7%); only minor increases in biodiesel conversion was observed thereafter. Therefore, the optimal WASC concentration was determined to be 3.5 wt%. Compared to the homogeneous catalysts (NaOH), much higher amounts of WASCs (0.5% vs. 3.5%) are required to obtain similar biodiesel conversion owing to poor mass and heat transfer. Another critical reason for the larger requirement of solid catalysts compared to homogeneous ones could be that transesterification occurs only on the surface of the solid catalysts, not the entire volume of the reaction mixture. That is unlike heterogeneous catalysts, homogeneous catalysts are completely dissolved and dispersed in the reaction mixture, thereby facilitating high reaction rates. Therefore, despite the advantages of solid catalysts stated above, solid catalysts need to be reused to compensate for the larger requirements of solid catalysts in biodiesel production.

Figure 4. Effect of reaction conditions on biodiesel conversion. (a) Effect of catalyst concentration (0.5 mL-methanol/g-WCO, 65ºC, 12h, 150 rpm). (b) Effect of methanol loading (3.5wt% catalyst, 65ºC, 12h, 150 rpm). (c) Effect of reaction temperature (3.5wt% catalyst, 0.7 mL-methanol/g-WCO, 12h, 150 rpm) (d) Effect of reaction time (3.5wt% catalyst, 0.7 mL-methanol/g-WCO, 60ºC, 150 rpm)

3.3.2. Effect of methanol loading

Next, the effect of methanol loading on the biodiesel conversion was investigated. Methanol is an acyl donor that produces biodiesel via transesterification with WCO. Fig. 4(b) shows that biodiesel conversion dramatically increased from 9.0% to 69.9% as the methanol loading increased from 0.1 to 0.3 mL/g-WCO. When the methanol loading was further increased to 0.7 mL/g-WCO, the biodiesel conversion reached a maximum value (90.4%); methanol loadings exceeding this value resulted in a slight decrease in biodiesel conversion, which may be due to the dilution of the solid catalysts [3]. Therefore, the optimal methanol loading was determined to be 0.7 mL/g-WCO. Considering the molecular weights of methanol and WCO (32.0 g/mol and 870 g/mol, respectively) [29], 0.7 mL/g-WCO corresponds to an approximately 15.1 molar ratio of methanol to WCO. Compared to our previous study of biodiesel production from soybean oil with low FFA contents (0.15%) under alkaline solid catalysts manufactured from fly ash supplemented with NaOH, a much smaller amount of methanol is required (0.7 mL vs. 5 mL methanol/ g-oil) in this study [23].

3.3.3. Effect of reaction temperature and time

High reaction temperatures generally accelerate the reaction rate, resulting in a high biodiesel conversion. As shown in Fig. 4(c), biodiesel conversion increased from 50.5% to 64.9% as the temperature went from 40°C to 50°C. The biodiesel conversion reached a maximum value of 89.4% at 60°C and a slight decrease was observed at temperatures higher than this. Based on the experimental data, we concluded that 60°C is enough to obtain high biodiesel conversion in this study. Finally, the effect of the reaction time on biodiesel conversion was investigated. As shown in Fig. 4(d), biodiesel conversion was as low as 20.0% at 3 h but dramatically increased to 78.6% after 4 h of reaction. When the reaction time was increased to 5 h, the biodiesel conversion further increased to 90.2% but slightly decreased when the reaction time was further increased to 6 h. From these investigations, we concluded that 5 h was the optimal reaction time. Fig. 5 presents a GC chromatogram of the biodiesel produced from WCO, which exhibits five distinct peaks, including the internal standard, indicating that the biodiesel from the WCO was composed of methyl linoleate (52.8%), methyl oleate (30.9%), methyl palmitate (11.7%), and methyl stearate (4.48%).

Figure 5. GC-chart of biodiesel from WCO.

3.4. Reuse of solid catalysts for repeated biodiesel production

In this study, various WASC treatment methods used in previous batch reactions were evaluated in terms of biodiesel conversion, as described in the Materials and Methods section. As presented in Table 2, the simple separating out and refilling method showed the highest performance in terms of biodiesel conversion in the second round of the batch reaction, that is, a marginal decrease in biodiesel conversion was observed. Washing the solid catalysts with methanol and/or n-hexane did not yield satisfactory results. Drying the solid catalyst alone or drying the solid catalyst after washing it with methanol and/or n-hexane resulted in lower biodiesel conversion. In addition to high biodiesel conversion, the simple separation and refiling method has the advantages of much less time and fewer chemical requirements compared to the other treatment methods. In addition, the loss of solid catalysts observed after washing with or without drying treatment could be avoided. Using the separating-out and refilling methods, a negligible decrease in biodiesel conversion was observed until seven rounds of batch operation, maintaining over 85% of biodiesel conversion, as shown in Fig. 6.

Figure 6. Biodiesel conversion with the number of reuse of alkaline solid catalysts. (3.5wt% catalyst, 0.7 mL-methanol/g-WCO, 60ºC, 5h, 150 rpm)

Table 2 Evaluation of various solid alkaline solid catalysts used in a previous round for the reuse of alkaline solid catalysts for the repeated biodiesel production

Treatment method		Biodiesel conversion (%)
In-situ mode	Separating out and refilling	89.7 ± 1.3
Ex-situ mode	Simple drying	73.5 ± 0.9
	Washing with methanol	81.6 ± 1.3
	Washing with n-hexane	82.8 ± 0.1
	Washing with methanol and n-hexane	84.1 ± 1.8
	Washing with methanol + drying	71.4 ± 3.1
	Washing with n-hexane + drying	76.1 ± 0.5
	Washing with methanol and n-hexane + drying	77.7 ± 1.4

Owing to the relatively low FFA content of WCO (1.9%) from local restaurants, alkaline catalysts can be used in biodiesel production. The solid alkaline catalysts were prepared from WSS via calcination for reuse. The optimal calcination conditions were 750°C for 3 h. The optimal reaction conditions for biodiesel production from WCO were 3.5 wt% solid catalysts, 0.7 mL-methanol/g-WCO, 60°C, 5 h of reaction time, and 150 rpm. Under these optimal conditions, a biodiesel conversion of 90.2% was achieved. Among the various treatment methods of the solid catalysts used for the next round of the reaction, a simple separating-out and refilling method, separating the reaction mixture using a syringe from a previous reaction, and introducing fresh WCO and methanol into the reactor containing the solid catalysts used, was the most effective for repeated biodiesel production. The alkaline solid catalysts were successfully reused for biodiesel production in seven rounds of repeated batch operations, maintaining more than 85% biodiesel conversion.

This work was supported by the Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education (NRF-2018R1 D1A1B07051113 and 2022R1F1A1072189). The authors greatly appreciate these supports. The authors declare no conflict of interest. Neither ethical approval nor informed consent was required for this study