Biomass Valorization of Walnut Shell for Liquefaction Efficiency

: Globally, lignocellulosic biomass has great potential for industrial production of materials and products, but this resource must be used in an environmentally friendly, socially acceptable and sustainable manner. Wood and agricultural residues such as walnut shells as lignocellulosic biomass are one of the most affordable and important renewable resources in the world, which can partially replace fossil resources. The overall objective of the research is to provide background information that supports new applications of walnut shells in a bioreﬁnery context and to increase the economic value of these non-wood forest products. This paper presents the properties characterization of liqueﬁed biomass according to their chemical composition. All results were compared to liqueﬁed wood. In this study, the liquefaction properties of ﬁve different walnut shell particle sizes were determined using glycerol as the liquefaction reagent under deﬁned reaction conditions. The liqueﬁed biomass was characterized for properties such as percentage residue, degree of liquefaction, and hydroxyl OH numbers. The chemical composition of the same biomass was investigated for its inﬂuence on the liquefaction properties. Accordingly, the main objective of this study was to determine the liquefaction properties of different particle sizes as a function of their chemical composition, also in comparison with the chemical composition of wood. The study revealed that walnut shell biomass can be effectively liqueﬁed into glycerol using H 2 SO 4 as the catalyst, with liquefaction efﬁciency ranging from 89.21 to 90.98%.


Introduction
Agriculture is very important for the overall development of individual countries and regions, but at the same time it can have a negative impact on the state of the environment. Agricultural production also produces waste and agricultural waste is available in large quantities in both the EU and the Republic of Croatia and therefore has the potential to become a source of renewable raw materials. Agricultural biomass is an important source of renewable raw materials [1] with great production potential, and the use of biomass as a raw material is continuously increasing throughout Europe [2,3]. A large part of agricultural waste consists of lignocellulosic material, and the main properties of lignocellulosic biomass are very good strength, flammability, biodegradability and reactivity [4]. Agricultural biomass can be used as a raw material for the production of natural fibers and is particularly interesting from the point of view of environmental protection, since it is available, renewable and acceptable as a raw material source independent of petroleum products [5][6][7].
In the 20th and 21st centuries, there has been intense technological and social progress in all spheres of human life. Industrialization and globalization, as well as the consumer Although there have been many studies on wood biomass liquefication, there are gaps in the literature on agricultural lignocellulose biomass liquefication, which led to the main objective of this study: to determine the liquefaction properties of walnut shell, as well as compare them to the chemical composition of wood. In this research the percentage residues, liquefaction degree and hydroxyl OH-numbers were determined as liquefaction properties of walnut shell biomass.

Materials and Methods
For this research we used biomass of walnut shell from the domestic cultivar Šejnovo (Sisak-Moslavina County, Dvor municipality, geographical coordinates of the experimental orchard: 45 04 23.5 N 16 • 22 35.0 E). Walnuts were collected by random selection of only healthy fruits from eight different trees in the orchard. An amount of 20 kg was collected from each tree. From the total amount (160 kg) of walnuts, a final sample (40 kg) was prepared by the method of quartering for further chemical analyzes. After separation of the shell and the kernel of the walnut, the shell air-dried naturally.
For screening, laboratory electromagnetic sieves shaker Cisa RP-08 (shaking time t = 15 ± 1 min) was used (TAPPI T 264 cm-97). After sample grinding and sieving, three smaller samples were taken on which all the chemical analysis and liquefaction were performed, and the results are presented as the mean values of these three samples.
The data were analyzed by means of the statistical software package SAS v 9.3 (Cary, NC, USA). The graphs are generated based upon the results of experiments using SageMath version 9.2 NoteBook package. The functions used are list_plot3D and list_plot2D.

Chemical Composition Analysis
In our research, non-combustible and combustible properties, heating value, lignocellulose composition and macro-element content were investigated. The chemical characterization of walnut shell was made on one granulometric fraction-0.63-1.25 mm. The analyses of walnut shell included determination of the moisture, ash, coke, fixed carbon and volatile matter content, C, H, N, S the content and the higher (HHV) and lower (LHV) heating values.
Moisture Isolation methods for determining the chemical composition (wet chemistry), namely ash, accessory materials (extractives), cellulose, hemicellulose (polyoses) and lignin were conducted in compliance with previous studies [32][33][34][35][36][37]. Sample chemical compositional analysis consisted of a series of isolation methods of the main components, which can be schematically presented as shown in Figure 1. Isolation methods for determining the chemical composition (wet chemistry), namely ash, accessory materials (extractives), cellulose, hemicellulose (polyoses) and lignin were conducted in compliance with previous studies [32][33][34][35][36][37]. Sample chemical compositional analysis consisted of a series of isolation methods of the main components, which can be schematically presented as shown in Figure 1. Isolation methods for determining the content of chemical composition (wet chemistry), namely ash, accessory materials (extractives), cellulose, hemicellulose (polyoses) and lignin were conducted in compliance with previous studies [33,34]. A small portion of the prepared sample was first used to determine the ash content, and the other major part for prior sample extraction (treatment with a solvent mixture of methanol, CH3OH and benzene, C6H6 in the volume ratio 1:1) to remove the accessory materials from the sample which could interfere during further chemical analysis. Thus, additional residual solid content was determined as a content of accessory materials or extractives [34]. Furthermore, sulfonic acid lignin or Klason's lignin (treatment with 72% sulfuric acid, H2SO4) and the polysaccharides cellulose (by treatment with a solvent mixture of nitric acid, HNO3 and ethanol, C2H5OH in a volume ratio of 1:4) was isolated from the extracted sample. The content of hemicellulose (polyose) was determined by calculation according to share of other mentioned components in the samples. The hemicellulose content was calculated according to next expression: WP = 100 − (% A + % AM + % C + % L) in % [34]. All used chemicals were high purity (p.a.) and were obtained from commercial sources.

Liquefied Biomass Preparation
Liquefied biomass was prepared based on a previous study according to Antonović et   Isolation methods for determining the content of chemical composition (wet chemistry), namely ash, accessory materials (extractives), cellulose, hemicellulose (polyoses) and lignin were conducted in compliance with previous studies [33,34]. A small portion of the prepared sample was first used to determine the ash content, and the other major part for prior sample extraction (treatment with a solvent mixture of methanol, CH 3 OH and benzene, C 6 H 6 in the volume ratio 1:1) to remove the accessory materials from the sample which could interfere during further chemical analysis. Thus, additional residual solid content was determined as a content of accessory materials or extractives [34]. Furthermore, sulfonic acid lignin or Klason's lignin (treatment with 72% sulfuric acid, H 2 SO 4 ) and the polysaccharides cellulose (by treatment with a solvent mixture of nitric acid, HNO 3 and ethanol, C 2 H 5 OH in a volume ratio of 1:4) was isolated from the extracted sample. The content of hemicellulose (polyose) was determined by calculation according to share of other mentioned components in the samples. The hemicellulose content was calculated according to next expression: WP = 100 − (% A + % AM + % C + % L) in % [34]. All used chemicals were high purity (p.a.) and were obtained from commercial sources.

Liquefied Biomass Preparation
Liquefied biomass was prepared based on a previous study according to Antonović et  Smaller obtained samples (without any prior chemical treatment) of different particle size were liquefied with mixture of glycerol and sulfuric acid (H 2 SO 4 ) by acid catalyst (biomass/glycerol = 1:5, sulfuric acid as catalyst (3 wt%)) method for 120 min at 150 • C. Undissolved residue (UR) percentage and wood liquefaction degree (LD), as well as hydroxyl number (OH-number) were determined as values that describe polymer properties of liquefied biomass for the purpose of selecting optimal liquefaction parameters. Smaller obtained samples (without any prior chemical treatment) of different particle size were liquefied with mixture of glycerol and sulfuric acid (H2SO4) by acid catalyst (biomass/glycerol = 1:5, sulfuric acid as catalyst (3 wt%)) method for 120 min at 150 °C. Undissolved residue (UR) percentage and wood liquefaction degree (LD), as well as hydroxyl number (OH-number) were determined as values that describe polymer properties of liquefied biomass for the purpose of selecting optimal liquefaction parameters.

Undissolved Residue and Liquefaction Degree
After the liquefaction reaction, undissolved residue of liquefied biomass was determined by the dioxane/water mixture, which is recommended as a universal diluent for liquefied biomasses. An amount of 1 g of liquefied biomass was diluted with dioxane/water mixture in a ratio 8/2, and then stirred in a magnetic stirrer for 60 min. After that mixture was filtrated through a glass-fiber filter B2 in a vacuum. The residue was rinsed by the same diluent repeatedly until a colorless filtrate was obtained, and then the undissolved residue was dried in oven at 105 ± 2 °C to a constant weight. Undissolved residue percentage was calculated by the following equation (according to Antonović et al., [34]): Liquefaction degree (LD) percentage was calculated according to the next equation:

Undissolved Residue and Liquefaction Degree
After the liquefaction reaction, undissolved residue of liquefied biomass was determined by the dioxane/water mixture, which is recommended as a universal diluent for liquefied biomasses. An amount of 1 g of liquefied biomass was diluted with dioxane/water mixture in a ratio 8/2, and then stirred in a magnetic stirrer for 60 min. After that mixture was filtrated through a glass-fiber filter B2 in a vacuum. The residue was rinsed by the same diluent repeatedly until a colorless filtrate was obtained, and then the undissolved residue was dried in oven at 105 ± 2 • C to a constant weight. Undissolved residue percentage was calculated by the following equation (according to Antonović et al. [34]): Liquefaction degree (LD) percentage was calculated according to the next equation:

Hydroxyl Number (OH-Number)
An amount of 1.5-2.5 g of liquefied biomass was weighed into two 250 mL volume Erlenmeyer's flasks and then 10 mL of reagent was added. The reagent was a mixture of pyridine and phthalic acid anhydride. In the third flask we added only reagent, for determination of a blank solution. Each flask was equipped with a condenser and magnetic stirrer with heater, and each magnetic stirrer had an oil bath, which was used for keeping a constant temperature at 115 • C, and with the help of a condenser we condensated the reagent. The mixture in the flask was heated for exactly one hour measured from the moment when the first drop condensed. After, 50 mL of pyridine was added to the cooled mixture and titrated with the 0.5 M sodium hydroxide solution with the presence of phenolphthalein until an equivalent point was reached (the bright red staining should not appear for at least 30 s). The hydroxyl number for liquefied biomass sample in mg KOH/g was calculated from the following equation (according to Antonović et al. [34]): where: A-volume of the NaOH solution used for sample titration (ml); B-volume of the NaOH solution used for blank solution titration; C NaOH -normality of the NaOH solution (M); m-weight of liquefied biomass sample. The dry matter content of different particle sizes was determined by drying a sample in an oven, at two temperatures (102 • C and 150 • C), until a constant weight was obtained.

Chemical Composition of Walnut Shell
The analysis of non-combustible properties determined: nitrogen (N), water (H 2 O), ash, coke, fixed carbon (C fix ), and the obtained values are shown in Table 1.  Biomass chemical composition namely accessory materials (extractives) content, ash content, cellulose content, polyoses (hemicellulose) content and lignin content of the different species are given in Table 3.

Liquefaction Properties of Liquefied Biomass
The liquefaction properties of liquefied different particle biomass size namely undissolved residue, liquefaction degree and hydroxyl number are given in Table 4. Among the liquefied different biomass particle sizes of walnut shell, the lowest undissolved residue content was achieved with granulation 0.3-0.6 mm, so the highest liquefaction degree was achieved with the same particle size (Figure 3). A-ash; AM-accessory material; H-holocellulose; C-cellulose; P-polyoses; L-lignin.

Liquefaction Properties of Liquefied Biomass
The liquefaction properties of liquefied different particle biomass size namely undissolved residue, liquefaction degree and hydroxyl number are given in Table 4. Among the liquefied different biomass particle sizes of walnut shell, the lowest undissolved residue content was achieved with granulation 0.3-0.6 mm, so the highest liquefaction degree was achieved with the same particle size (Figure 3). Obtained results have shown that liquefaction degree depends on particle size and with a decrease in granulation there is an increase in liquefaction degree. Among the liquefied different biomass particle sizes, the lowest OH-number was achieved with granulation 0.3-0.6 mm (Figure 4). Error bars with trendline -Liquefaction degree Obtained results have shown that liquefaction degree depends on particle size and with a decrease in granulation there is an increase in liquefaction degree. Among the liquefied different biomass particle sizes, the lowest OH-number was achieved with granulation 0.3-0.6 mm (Figure 4).  Obtained results have shown that dry matter depends on temperature and particle size. The lowest content of dry matter was at the smallest and at the biggest particle size of walnut shell (Table 5 and Figure 5). Table 5. Relationship between dry matter (%) and particle size (the same small letter (a) indicates that there is no significantly difference at p < 0.05). Results have also shown that the relationship between dry matter and particle size remains the same regardless of temperature, i.e., given the particle size interval, the change of dry matter is the same no matter the temperature ( Figure 5). Error bars with trendline -OH number Obtained results have shown that dry matter depends on temperature and particle size. The lowest content of dry matter was at the smallest and at the biggest particle size of walnut shell (Table 5 and Figure 5). Table 5. Relationship between dry matter (%) and particle size (the same small letter (a) indicates that there is no significantly difference at p < 0.05). Results have also shown that the relationship between dry matter and particle size remains the same regardless of temperature, i.e., given the particle size interval, the change of dry matter is the same no matter the temperature ( Figure 5). . Dry matter dependence on temperature and particle size (x-particle size; y-temperature; z-dry matter)

Discussion
Moisture content depends on environmental air temperature during the winter season, and it is one of the most important parameters when it comes to fuel properties. The moisture content was 12.23%. Ash content, which is also one of the main factors of biomass quality, since higher amounts of ash diminishes the quality of fuels, especially solid ones, was 1.26%. Ash originates simultaneously from natural and technogenic inorganic, organic and fluid matter during biomass combustion [35]. This result is in the range reported for nut shells, although a large variation of values may be found for a specific nut shell. Queirós [36] also reported a low content of ash for walnut shell (0.7%). Coke content and fixed carbon content are considered positive properties of biomass because they represent the quantity of energy released by the combustion of a specific amount of biomass [38]. In our study, the obtained fixed carbon content in walnut shell was 15.89% and coke content was 17.15%. From an environmental point of view, nitrogen contributes to the increase in greenhouse gases, and is considered to be an unfavorable element in biomass. Content of nitrogen in walnut shell was 0.56%. The obtained results are slightly lower than those obtained by Demirbas [39] who determined the values of non-combustible substances in walnut shell as 2.80% ash, 37.90% fixed carbon and 1.50% nitrogen (N).
In the chemical composition of fuel, carbon (C) is one of the basic elements and makes up to 95%. The amount of carbon also determines the quality of fuel; i.e. by increasing the carbon content the quality of fuel improves. The carbon content in the walnut shell was 52.11% while the hydrogen content was 5.86%. The obtained results are slightly higher than those obtained by Matin et al., [40] who found the carbon content in walnut shell to be from 57.55 to 58.01%, depending on the variety, while Demirbaş [39] found that walnut shell contains 53.50% carbon (C). Hydrogen is the second most important ingredient in fuel, which with its high energy increases the thermal value of the fuel, and by burning it creates a visible flame.
By its share in fuel, oxygen reduces the calorific value of fuel. Oxygen content was 42.20%. Figure 5. Dry matter dependence on temperature and particle size (x-particle size; y-temperature; z-dry matter).

Discussion
Moisture content depends on environmental air temperature during the winter season, and it is one of the most important parameters when it comes to fuel properties. The moisture content was 12.23%. Ash content, which is also one of the main factors of biomass quality, since higher amounts of ash diminishes the quality of fuels, especially solid ones, was 1.26%. Ash originates simultaneously from natural and technogenic inorganic, organic and fluid matter during biomass combustion [35]. This result is in the range reported for nut shells, although a large variation of values may be found for a specific nut shell. Queirós [36] also reported a low content of ash for walnut shell (0.7%). Coke content and fixed carbon content are considered positive properties of biomass because they represent the quantity of energy released by the combustion of a specific amount of biomass [38]. In our study, the obtained fixed carbon content in walnut shell was 15.89% and coke content was 17.15%. From an environmental point of view, nitrogen contributes to the increase in greenhouse gases, and is considered to be an unfavorable element in biomass. Content of nitrogen in walnut shell was 0.56%. The obtained results are slightly lower than those obtained by Demirbas [39] who determined the values of non-combustible substances in walnut shell as 2.80% ash, 37.90% fixed carbon and 1.50% nitrogen (N).
In the chemical composition of fuel, carbon (C) is one of the basic elements and makes up to 95%. The amount of carbon also determines the quality of fuel; i.e. by increasing the carbon content the quality of fuel improves. The carbon content in the walnut shell was 52.11% while the hydrogen content was 5.86%. The obtained results are slightly higher than those obtained by Matin et al., [40] who found the carbon content in walnut shell to be from 57.55 to 58.01%, depending on the variety, while Demirbaş [39] found that walnut shell contains 53.50% carbon (C). Hydrogen is the second most important ingredient in fuel, which with its high energy increases the thermal value of the fuel, and by burning it creates a visible flame.
By its share in fuel, oxygen reduces the calorific value of fuel. Oxygen content was 42.20%.
Sulfur (S) is the least represented element and it is usually found in traces in biomass. The obtained values (0.23%) are slightly higher than stated by Demirbaş [39] who found that walnut shell contains 0.10% sulfur (S) while Matin et al. [40] found even lower values (0.04-0.05%).
Obtained results have shown that walnut shell biomass has different chemical composition compared to wood chemical composition investigated by Antonović et al. [34].
The biggest difference between walnut shell biomass and wood is in the content of cellulose and lignin (cellulose content from 39.18-48.38%, polyoses content from 22.69-32.41% and lignin content from 21.82-27.96%).
Liquefaction comprises a complex set of reactions taking place on the polymeric components of biomass. They include derivatization such as esterification or etherification of free hydroxyl groups in cellulose or lignin as well as reactions that break the polymer chain of cellulose. In addition, liquefaction is affected by physical constraints on biomass reactivity such as the high crystallinity of cellulose. The tight packing of cellulose in the crystalline domains makes the reaction kinetics of otherwise reactive functional groups dependent on the diffusion of reagents into the tightly packed system. To overcome this limitation and speed up the liquefaction, increasingly harsh catalysts and reaction conditions, mainly mineral acids and high temperatures, have been employed. In short, macromolecule compounds in biomass are degraded into micro molecules and the obtained small molecules are unstable, reactive and can re-polymerize into oily products with a wide range of molecular weight distribution [33,34].
Undissolved residue content among the liquefied walnut shell varied from 9.02-10.79%, liquefaction degree content from 89.21-90.98% and hydroxyl (OH)-number from 359-450 mg KOH/g. Obtained results have shown that walnut shell biomass has a different chemical composition compared to wood chemical composition investigated by Antonović et al., [34]. The hardwood species exhibited higher undissolved residue (11.53% for beech to 8.02% for common oak) and lower liquefaction degree content (88.47% for beech to 91.98% for common oak) as compared to softwood species (undissolved residue was 5.06% for spruce and 6.20% for common oak). Within hardwood species differences were also observed in the same liquefaction properties. Although softwoods show the smallest undissolved residue content and the highest liquefaction degree content compared with hardwood, the hardwoods show a much higher OH-number. Hydroxyl number is very important in further liquefied wood application for various bioproducts, therefore should be given priority over undissolved residue and liquefaction degree [34]. According to that all hardwoods (692-798 mg KOH/g), and especially beech (798 mg KOH/g), show better properties in further application compared to walnut shell (359 to 450 mg KOH/g). So, the biggest difference between walnut shell biomass and wood is OH-number.
Liquefaction degree content in walnut shell varied from 89.21-90.98%, which is similar to the results obtained for liquified wood (88.47-94.94%).
Results have shown that OH-number depends on particle size and with an increase in granulation there is an increase in OH-number. Results also shown that the relationship between dry matter and particle size remains the same regardless of temperature, i.e., given the particle size interval, the change of dry matter is the same no matter the temperature.

Conclusions
The study revealed that walnut shell biomass can effectively be liquefied in glycerol using H 2 SO 4 as a catalyst with liquefaction efficiency ranging from 89.21-90.98% under the same liquefaction conditions, which is similar to the results obtained for liquified wood (88.47-94.94%).
Hydroxyl number as the most important liquefaction property in further application for various bioproducts have priority over undissolved residue and liquefaction degree, and according to that bigger particle size of walnut shell biomass show better properties in further application compared to smaller. Results have shown that OH-number depends on particle size and with the increase in granulation there is an increase in OH-number, so the biggest particle size (>1.4) had the biggest OH number (450). The lowest content of dry matter was at the smallest and at the biggest particle size of walnut shell so the obtained results have shown that the relationship between dry matter and particle size remains the same regardless of temperature, i.e., given the particle size interval, the change of dry matter is the same no matter the temperature.
The use of a catalytic liquefaction method with polyhydric alcohol glycerol was found to be suitable for liquefying walnut shells. Finally, the present study revealed unimaginable opportunities for scientific research and development aimed at novel bioproducts from liquefied biomass and opened new challenges in exploring natural, ecologically sound products with unlimited raw materials. Data Availability Statement: Upon a reasonable request, all data in three replicates will be provided by authors.

Conflicts of Interest:
The authors declare no conflict of interest.