Optimization and characterization of cellulose extraction from olive pomace

Olive pomace (OP) was used as raw material to extract cellulose by multi-step chemical method, and the extracted cellulose was characterized. The two steps of alkali treatment (alkali concentration, reaction time and reaction temperature) and bleaching (H2O2 concentration, NaOH concentration, bleaching time and bleaching temperature) were optimized. The results showed that the optimum conditions for alkali treatment were as follows: alkali concentration 6%, reaction time 2 h, reaction temperature 95 °C; the optimal bleaching conditions were as follows: 7.5% H2O2 solution, 5% NaOH, bleaching time 2 h, bleaching temperature 75 °C. After multi-step chemical treatment, the cellulose content of the final OP cellulose was (86.41 ± 0.27) %, the lignin content was (6.77 ± 0.55) %, the extraction process yield was 20.36%. By scanning electron microscopy, the surface of OP cellulose changes from dense smooth structure to rough irregular structure with different treatment degrees. Fourier transform infrared spectroscopy (FTIR) proved the removal of hemicellulose and lignin in OP fibers after chemical treatment. The crystallinity of OP fiber increased from 34.80 to 68.69% after treatment. Thermogravimetric analysis (TGA) showed that OP cellulose had high thermal stability. The study provides a new idea for the conversion of industrial processing by-products into high-quality cellulose, and proves that it is feasible to extract cellulose from olive pomace.


Introduction
With increasing public attention to materials that are renewable resources, cellulose is the most abundant natural resource in the world and can be obtained from plants, algae, bursal animals, and bacteria (Shi and Liu 2021). Cellulose is a linear polymer composed of D-glucose linked by β (1 → 4) glycosidic bonds. Cellulose fibrils are mainly composed of α-cellulose, hemicellulose, lignin, pectin and wax, and the content of these components varies from plant to plant. At present, there are three main sources of cellulose: the first is cellulose fiber as the main product of plants: wood, cotton, hemp and flax; the second is agricultural residues, such as the coconut shell of straw of wheat, corn and rice crops; the third is waste from industrially processed biomass, including by-products from the food and beverage industry, such as bagasse (Pennells et al. 2020). Wood has been the research object of cellulose in the past. In recent years, more and more studies have turned the research object of cellulose to agricultural waste and industrial waste.
Olive pomace (OP) is a solid waste produced during the production of olive oil and is mainly composed of pulp and kernel. Each processing of 1 t of olive will produce 0.5-0.6 t of olive pomace (Foti et al. 2022). The OP produced by processing olive oil is mostly discarded directly by olive enterprises, or a small part is used as animal feed or plant fertilizer. OP is mainly composed of cellulose, hemicellulose, and lignin. Considering the cellulose content in OP, OP may be a good source of cellulose. Therefore, how to efficiently extract pure cellulose from OP and improve the economic value of OP has become an urgent problem to be solved.
The key to obtaining pure cellulose is to effectively remove hemicellulose, lignin and other impurities. At present, the extraction of cellulose from lignocellulosic materials generally first crushes the raw material to reduce the fiber particle size and increase the reaction area, and then uses toluene and ethanol for dewaxing treatment to remove wax, pigment, and soluble sugar. After dewaxing treatment, the alkaline solution of sodium hydroxide (NaOH) or potassium hydroxide (KOH) will be used, and hemicellulose and remaining pectin, wax, and fat will be eliminated at this stage. Due to the removal of some hemicellulose after alkali treatment, the fiber structure was destroyed, the fiber size decreased and the aspect ratio increased, the fiber surface became rough, and the fiber matrix interface bonding performance was increased (Borchani et al. 2015). The effect of alkali treatment on fiber composition, structure and morphology depends on alkali concentration, reaction time and reaction temperature. Therefore, some studies have been carried out to obtain high-performance cellulose. In Chen's study (2017), when the concentration of NaOH increased to 10%, the hemicellulose in bamboo fiber was completely removed, and the microfiber aggregates began to change from random interlaced structure to granular structure. When the concentration increased to 15%, the structure of bamboo fiber changed from cellulose I to cellulose II. Sarak et al. (2022) compared Khlum fibers treated with alkali for 1 h and 2 h, the fibers treated with alkali for 1 h had higher thermal stability and high crystallinity, and the cellulose structure was seriously damaged after alkali treatment for 2 h. In the alkali treatment optimization experiment, Gomaa et al. (2021) observed that the cellulose yield decreased with the increase of extraction temperature and time, and the effect of NaOH concentration on cellulose yield was linear. Finally, fibers were bleached with oxidizing compounds such as sodium hypochlorite (NaClO), sodium chlorite (NaClO 2 ) or hydrogen peroxide (H 2 O 2 ). Oxidizing compounds can destroy the selected bonds (aryl ether bonds, C−C bonds) in lignin, generate small molecule compounds, and remove the remaining hemicellulose and lignin (Ventura-Cruz and Tecante 2021). Lignin has physical rigidity due to its strong polycyclic bonds of C−O−C, C−C and hydrophobic bonds, and lignin is located deep in the cell wall and tends to recombine (Ciftci et al. 2018), making complete removal of lignin difficult. As the effective extraction of cellulose depends largely on the removal of lignin, bleaching can be considered as a key step in the separation of cellulose in chemical treatment. The bleaching conditions for extracting cellulose from lignocellulosic materials have been studied comprehensively, including bleaching concentration, bleaching pH, bleaching time and bleaching temperature. Rizwan et al. (2021) compared the bleaching effects of hydrogen peroxide, sodium hypochlorite and sodium chlorite, and found that the fiber treated with hydrogen peroxide had higher thermal stability and crystallinity, and higher cellulose extraction efficiency. Suriyatem et al. (2020) found that with the increase of H 2 O 2 concentration, the cellulose yield and lignin content of palm bundles and bagasse decreased, and the fiber brightness and whiteness increased. Wang and Zhao (2021) optimized the bleaching conditions of microcrystalline cellulose extracted from apple pomace and kale pomace. It was found that 2-3% NaClO was the critical condition for bleaching, and higher than this concentration would lead to a decrease in the quality of extracted cellulose.
At present, there is no complete research on the optimization of cellulose extraction process from olive pomace. In this study, the extraction process of cellulose from OP was optimized with the aim of removing hemicellulose and lignin from OP fiber as much as possible to obtain OP cellulose with high cellulose content. In order to reduce the pollution to the environment, the more environmentally friendly alkaline H 2 O 2 solution was selected for bleaching. The two-step process of alkali treatment (alkali concentration, reaction time, reaction temperature) and bleaching (H 2 O 2 concentration, NaOH concentration, bleaching time, bleaching temperature) was optimized by orthogonal test. The extracted OP cellulose was characterized by various techniques, and the surface morphology of cellulose treated by different steps was analyzed by scanning electron microscopy (SEM). The functional group structure of cellulose was analyzed by Fourier transform infrared spectroscopy (FTIR). The crystallinity of cellulose was analyzed by X-ray diffraction (XRD). The thermal stability of the extracted cellulose was analyzed by a thermogravimetric analyzer (TGA). All measured properties were compared with microcrystalline cellulose (MCC).

Chemicals and raw material
The raw material is represented by olive pomace (OP) resulting from the three-phase centrifugal separation process of olive, which is provided by Longnan Longjinyuan Olive Development Co., Ltd. The fresh OP was dried at 60 °C in a blast oven until a constant weight was reached. The OP was crushed by a pulverizer and passed through a 60-mesh sieve. Hydrogen peroxide (H 2 O 2 , 30%) was purchased from Rongman Biotechnology Co., Ltd.; sodium hydroxide (NaOH, > 96%) was purchased from Guangshi Technology Co., Ltd.

Pretreatment
Cellulose was extracted from pomace by continuous chemical treatment to remove lipids, hemicellulose and lignin. The raw olive pomace (R-OP) was dewaxed in a Soxhlet apparatus for 3 h at 75 ℃ using 95% ethanol solution at a solid-liquid ratio of 1:10 (w/v). After dewaxing treatment, it was rinsed with ethanol solution to neutral and dried in an oven at 60 ℃ to constant weight to obtain dewaxed olive pomace (D-OP).

Optimization of alkali treatment conditions
The reaction of D-OP was carried out according to the solid-liquid ratio of 1:10 (w/v), NaOH solution (3%, 4%, 5%, 6% and 7%(w/w)), reaction time (1.0, 1.5, 2.0, 2.5 and 3.0 h) and reaction temperature (75, 80, 85, 90 and 95 °C). After the reaction, the fiber was repeatedly washed with distilled water until the pH was neutral, and dried to constant weight in an oven at 60 °C to obtain alkali-treated olive pomace (A-OP).

Orthogonal experimental design
According to the results of previous single factor experiments (see supplementary data), the two steps of alkali treatment and bleaching in cellulose extraction process were optimized by orthogonal design. The effects of alkali concentration (5%, 6% and 7%), reaction time (1.5, 2.0 and 2.5 h) and reaction temperature (85, 90 and 95 °C) on cellulose content, hemicellulose content, and lignin content were studied. The effects of H 2 O 2 concentration (5.0%, 7.5% and 10.0%), NaOH concentration (4%, 5% and 6%), bleaching time (3, 4 and 5 h) and bleaching temperature (70, 75 and 80 °C) on cellulose content, hemicellulose content, and lignin content were studied. These variables and levels are selected based on our preliminary study. With cellulose content as the main basis, hemicellulose content and lignin content as the auxiliary, the optimal reaction conditions of the twostep process of alkali treatment and bleaching were determined, and the optimal cellulose extraction process conditions of olive pomace were obtained.

Cellulose based material composition
Neutral detergent fiber (NDF) and acid detergent fiber (ADF) were determined by Van Soest method (Van Soest 1967). The 0.5 g raw material (dry basis) was put into 50 mL neutral detergent solution or acidic detergent solution, heated to maintain a slightly boiling state, and reacted for 1 h. The material was filtered, washed with water many times to no foam and neutral, and finally washed with acetone. The filter residue was dried and weighed. The samples after ADF analysis were treated with 72% sulfuric acid for 3 h to remove cellulose, residual lignin and ash. The remaining residue was calcined to remove ash to obtain lignin content. The hemicellulose content is calculated by: NDF minus ADF content (%), and the cellulose content is calculated by: ADF minus lignin content (%).

Cellulose extraction yield
After each step of OP treatment, it is dried to constant weight in an oven at 60 °C. The yield (%) of the resulting cellulose is calculated by Eq. (1):

Characterization
The morphology of the samples was observed with a transfield emission scanning electron microscope (Merlin, Zeiss). An appropriate amount of powder was fixed on the sample table with conductive glue, and then gold spraying was carried out. High-resolution images were taken at a high voltage of 10 kV at various magnifications.
In order to confirm the existence and purity of cellulose, the infrared spectrum of OP obtained in different steps of cellulose extraction was analyzed, and the changes of OP functional groups after different steps were analyzed. The samples were scanned by FTIR spectrometer (Nicolet IS50-Nicolet Continuum, Thermo Fisher Scientific) with a scanning range of (1) Yield (%) = W Dried cellulose W Dried pomaces × 100% 500-4000 cm −1 , a resolution of 4 cm −1 , and a scanning number of 32 times. The crystalline structure of the sample was tested by a multi-position X-ray diffractometer X 'pert (X 'pert Powder, PANalytical). The diffraction target CuKα, wavelength (λ) is 1.54059 Å, working at 44 mA and 40 kV, scanning angle is 5-50°, scanning speed is 4°/min, the crystallinity of the sample is calculated by Segal method Eq. (2): I 200 is the maximum intensity of the (200)-lattice peak at 2θ = 22.5°, and I am is the intensity from the amorphous phase at approximately 2θ = 18°.
The sample was subjected to thermogravimetric analysis using a TG thermogravimetric analyzer (TG209F1, Natch, Germany) to protect the odor N2, the gas flow rate was 20 mL/min, the heating rate was 10 °C/min, and the heating range was 30-700 °C.
Alkali treatment can reduce the hemicellulose, lignin, pectin, wax and oil contained in the raw fiber. The mechanism is that OH − weakens the hydrogen bond between cellulose and hemicellulose, saponifies the ester bond between hemicellulose and lignin, increases the pores between fibers, breaks the chemical bond, and leads to the dissolution of hemicellulose (Ng et al. 2015). Cellulose reacts with NaOH, as shown below (Jayamani et al. 2020).
The increase of cellulose content is mainly due to the decrease of hemicellulose and lignin. Reaction temperature is the biggest factor affecting the content of extracted cellulose. The second is the solution Cellulose-OH+ NaOH → Cellulose-ONa + H 2 O + Impurities concentration, and the least effect is the reaction time. Similar to Wu 's studies (Wu et al. 2022), the increase of reaction temperature is more effective in removing hemicellulose and lignin from bamboo fibers than the extension of reaction time. This may be because the hydroxyl group (OH − ) in the cellulose is temporarily converted to O−Na during the alkali treatment process, and the Na ion is washed away by water, which destroys the hydrogen bond of the cellulose and weakens the hydrogen binding of the cellulose and hemicellulose. The increase of reaction temperature caused strong molecular movement, and the combination of Na ions and cellulose hydroxyl groups was more active, which weakened the hydrogen binding between cellulose and hemicellulose, and more hemicellulose was removed, thus increasing the cellulose content in OP fibers (Jayamani et al. 2020;Lamo et al. 2022). Alkali treatment destroys biomass cells by dissolving amorphous hemicellulose, lignin and other impurities wrapped on the surface of fiber cell wall, and destroys ether bonds and ester bonds between lignin and hemicellulose. However, in high concentration of alkali solution, cellulose is degraded, so the cellulose content decreases. Therefore, when the concentration of alkali solution increases to 7%, the cellulose content decreases (Kathirselvam et al. 2019; Van et al. 2022). Table 1 shows that the hemicellulose content of the sample treated with 5% NaOH is 8.31-9.90%, while that treated with 7% NaOH is 6.2-7.69%. It can be seen that the increase of NaOH concentration helps to better remove hemicellulose. After alkali treatment, the hemicellulose in OP fiber was effectively removed and the lignin content increased, which was similar to that of Parsley Stalks (Cakmak and Dekker 2022), Thespesia Lampas plant (Reddy et al. 2014) and Ensete ventricosum plant fiber (Teli and Terega 2019). The reasons for the increase of lignin content may be: First, the lignin content increases with the proportion of hemicellulose removal; Second, alkali treatment cannot remove all non-cellulose components, the effect and reaction of alkali treatment on lignin are complex, due to the different stability of different types of lignin bonds and structural elements, studies (Keikhosor et al. 2013) have shown that high concentration NaOH (6-20%) process on lignin removal effect is not obvious, and cellulose degradation will occur. However, lignin can be removed at a lower concentration of NaOH (less than 4%) and a higher reaction temperature. Therefore, the lignin content of OP fiber increased. Therefore, according to the results of orthogonal test of alkali treatment, the optimum conditions for alkali treatment were: alkali concentration 6%, reaction time 2 h, reaction temperature 95 °C. At this time, the cellulose content of alkali treated fiber was (55.17 ± 0.42)%, hemicellulose content was (7.78 ± 0.30)%, lignin content was (35.44 ± 0.29)%.
After alkali treatment, cellulose fibers still retained a certain amount of hemicellulose and lignin, which needed to be further removed by bleaching. In this study, cellulose was further purified by alkaline H 2 O 2 solution. When the H 2 O 2 solution was alkaline, a large amount of hydrogen peroxide anion (HOO·) was released to produce more hydroxyl radicals  HO·). The reaction between lignin and free radicals produced by H 2 O 2 mainly acted on the benzene ring and side chain of lignin. H 2 O 2 will destroy the quinine structure of lignin, make the colored structure colorless, and finally decompose into low molecular fat compounds. It also destroys the conjugated double bonds on the lignin side chain, turns the colored conjugated double bonds into colorless, and further decomposes the side chain, eventually leading to the oxidative decomposition of the benzene ring and quinone structure of lignin, and the lignin is removed (Qu et al. 2005).
In the bleaching process, the influence degree of each factor on cellulose content is: bleaching temperature > bleaching time > NaOH concentration > H 2 O 2 concentration. The influence degree of each factor on hemicellulose content is: H 2 O 2 concentration > bleaching time > NaOH concentration > bleaching temperature. The influence degree of each factor on lignin content is: H 2 O 2 concentration > NaOH concentration > bleaching temperature > bleaching time. According to the results of orthogonal test, the four factors of bleaching reaction had different effects on cellulose, hemicellulose and lignin. H 2 O 2 concentration is the most important factor affecting hemicellulose and lignin removal. It is well known that lignin surrounds cellulose and hemicellulose, and lignin interaction hinders the decomposition of hemicellulose. With the increase of H 2 O 2 concentration, more HOO· and HO· were produced in the reaction system, and more free radicals will act on the benzene ring and conjugate double bond on the side chain of lignin, resulting in the oxidation and decomposition of lignin, and the removal of more lignin. The Mittal study (Mittal et al. 2017) also confirmed the importance of H 2 O 2 concentration on lignin removal. At lower H 2 O 2 concentration, the lignin side chain structure was only oxidized, while the aromatic ring did not change significantly. At high H 2 O 2 concentration (> 250 mg H 2 O 2 /g dry matter), most of the side chain structure and aryl ether bonds of lignin were cracked. With the decomposition of lignin surrounding hemicellulose, hemicellulose was exposed to the H 2 O 2 system for decomposition, and the residual hemicellulose after alkali treatment was further removed. NaOH concentration is also an important factor affecting lignin removal during bleaching. Because H 2 O 2 can only generate HOO· under alkaline conditions with pH close to 11.5, and then produce more active HO· and superoxide anion radicals (O 2-·), prompting lignin oxidation and depolymerization into small molecule compounds (Ho et al. 2019). Too low or too high concentration of NaOH solution will deviate the pH of the reaction conditions from 11.5 to 11.6, resulting in poor lignin removal. At the same time, too high NaOH concentration will lead to unexpected dissolution of cellulose, resulting in a decrease in cellulose content in the obtained OP fiber. Bleaching time is equally important for the removal of lignin and hemicellulose. Because alkaline H 2 O 2 produces HOO·, HO· and O 2 -·, it takes a certain time to decompose lignin and hemicellulose into small molecular compounds. The longer the action time, the better the removal effect of both, but it also needs to be noted that too long time will cause free radicals to attack cellulose, resulting in cellulose decomposition. Wu et al. (2019) found that the bleaching time was prolonged from 30 to 150 min, the lignin content decreased from 19.5 to 14.9%, and the hemicellulose content decreased from 16.8% to 15.7%. The degree of alkaline hydrogen peroxide delignification is also affected by the reaction temperature. The reason is: H 2 O 2 decomposition is very dependent on temperature. At low temperature, H 2 O 2 is relatively stable, and the redox potential is not high. With the increase of temperature, H 2 O 2 is easy to decompose spontaneously, generating active free radicals such as hydroxyl radicals, which participate in the degradation reaction of lignin and hemicellulose, so that the relative content of cellulose in the fiber is increased. Sun 's study (2000) confirmed this point. When the H 2 O 2 bleaching temperature increased from 20 to 70 °C, the solubility of raw lignin increased from 52.7 to 87.8%, and the yield increased from 7.8 to 13.0%. It is worth noting that there are two forms of hydrogen peroxide reaction: one is self-oxidative decomposition into water and oxygen, and the other is decomposition to produce free radicals. Therefore, if the temperature is too high, it will aggravate the decomposition of H 2 O 2 itself, reduce the free radicals that actually act on hemicellulose and lignin, and affect the bleaching effect.

Variation of cellulose components and yield
The cellulose content of R-OP was (40.85 ± 0.32)%, the hemicellulose content was (14.27 ± 0.39)%, and the lignin content was (25.05 ± 0.32)% (Fig. 1 a). After dewaxing treatment, the soluble extracts (impurities) and waxy substances in the fiber were removed to obtain D-OP cellulose. After dewaxing treatment, the overall content of OP cellulose, hemicellulose and lignin in OP increased, and the yield of dewaxing treatment was about 86.96% (Fig. 1b). After D-OP was treated with 6% NaOH solution, some hemicellulose was removed. In addition, wax, grease and pectin materials that form the outer non-cellulose layer of cellulose can also be removed by alkali treatment (Ng et al. 2015). The semi-fiber content of A-OP was (7.78 ± 0.30)%, the cellulose content increased to (55.17 ± 0.42)%, and the yield of alkali treatment was lower, only 61.50%. B-OP was obtained by bleaching A-OP with alkaline hydrogen peroxide. After three times of bleaching, the resulting B-OP3 has not detected hemicellulose, proving that hemicellulose has been completely removed. After five times of bleaching, the cellulose content of B-OP5 increased from (40.85 ± 0.32)% to (86.41 ± 0.27)%, the lignin content decreased from (25.05 ± 0.32)% to (6.77 ± 0.55)%, and the yield of each bleaching was about 82%. After dewaxing, alkali treatment and bleaching treatment, the final yield of R-OP cellulose was about 20.36%.
The comparison between the extracted OP cellulose and other studies is shown in Table 3. Compared with other lignocellulosic materials, hemicellulose was completely removed from the cellulose extracted by the optimal alkali treatment and bleaching conditions. After alkali treatment and bleaching treatment, most of the cellulose extracted from lignocellulosic materials still remained a certain amount of hemicellulose. This may also be related to the relatively low hemicellulose content of OP fiber itself. The extracted OP cellulose is also slightly higher than other fibers in cellulose content, and the lignin content and yield are also superior to some lignocellulosic materials. Therefore, it can be seen that the extraction of OP cellulose under optimized conditions is completely desirable.

Morphology analysis
Hemicellulose, lignin, pectin and wax are the outer layers of cellulose fibers, which play a role in protecting cellulose. Therefore, untreated lignocellulose usually presents a dense structure. As can be seen from Fig. 2a, R-OP without any chemical treatment, due to the presence of wax and lipid substances, the fiber surface is smooth and the structure is dense (Rizwan et al. 2021). It can be seen from Fig. 2b that after dewaxing treatment of OP, some wax and lipid substances were removed, the smooth and dense structure   Lamo et al. (2022) of cellulose surface was destroyed, and the surface was uneven. The biomass shell composed of hemicellulose, lignin and pectin began to be destroyed. It can be seen from Fig. 2c that after alkali treatment, some hemicellulose was removed, and the surface of the fiber became rough and porous. Highly interwoven fiber bundles can be observed. Fiber bundles indicate the presence of lignin. There are hydrogen bonds, covalent O-H bonds and van der Waals interactions between cellulose and lignin. Lignin acts as a binder for cellulose fibers, which is similar to the Asif study (Asif et al. 2022). Because cellulose is covered by lignin and hemicellulose, both of them prevent chemicals from passing through lignocellulose (Keikhosor et al. 2013). After alkali treatment, the OP biomass shell is completely destroyed, hemicellulose is removed, and holes are formed. Alkaline hydrogen peroxide can enter the interior of lignocellulose and act on lignin. After alkaline hydrogen peroxide bleaching treatment, as shown in Fig. 2 d, the lignin as a physical barrier was removed, the fiber surface became more irregular and rough, the porous structure was more obvious, and the fiber structure inside was completely exposed.
FTIR spectra analysis Figure 3 shows the FTIR spectra of OP treated by different steps. FTIR spectroscopy was used to determine the effect of different chemical treatments on the chemical structure of OP, which was of great significance to verify the removal of hemicellulose and lignin of OP in this study. It can be seen from Fig. 3 that all the spectra show roughly similar peaks, indicating that the same functional groups exist, and it is proved that the multistep treatment does not change the structure of OP cellulose, and the cellulose is successfully extracted from OP after multi-step treatment. The peak near 3440 cm −1 is the −OH stretching vibration peak of cellulose. With the deepening of treatment, the peak value of this peak increases, which means that the content of −OH group increases and the content of cellulose increases. The −CH stretching vibration peak at 2900 cm −1 indicated the asymmetric stretching vibration of aliphatic saturated C=H in cellulose, hemicellulose and lignin, which were the characteristic peaks of cellulose (Rohadi et al. 2020 Gomaa et al. (2021) are related to the C−O stretching vibration, C−O−C asymmetric stretching vibration and C−H vibration of cellulose, respectively, which correspond to the FTIR spectra of cellulose reported by Pongchaiphol et al. (2021), Ventura-Cruz and Tecante (2019). 1645 cm −1 is the bending vibration peak of H−O−H in adsorbed water. Although the sample was dried before FTIR analysis, the interaction between cellulose and water in the sample still existed. 896 cm −1 is the bending vibration peak of −C 1 −O−C 4 of cellulose β-glycosidic bond. The peaks near 1735 and 1250 cm −1 , representing carboxyl groups, may be caused by the acetyl C−O stretching vibration of hemicellulose or the ester bond of lignin. As the process progresses, the peak at 1735 cm −1 disappears, and the peak intensity at 1250 cm −1 decreases, indicating that hemicellulose and lignin are better removed after alkali treatment and bleaching (Nagarajan et al. 2021). 1515 cm −1 is the C=O stretching vibration peak of ketone and carbonyl on lignin, and the peak intensity decreases with the process, indicating that the lignin is partially removed.

XRD analysis
Cellulose is orderly arranged through intramolecular and intermolecular hydrogen bonding interactions, and cellulose has a crystal structure. In the process of cellulose extraction, chemical treatment will affect the crystal structure of cellulose. The XRD patterns of OP treated by different steps are shown in Fig. 4. The crystallinity (CrI) of each OP fiber calculated by the Segal method is shown in Table 4.
It can be seen from Fig. 4 that the OP fibers obtained by different treatment steps have four main characteristic peaks:2θ = 14.7°, 16.3°, 22.5° and 34.6°, corresponding to the (1-10), (110), (200) and (004) crystal planes of the cellulose I structure (French 2014). There are three diffraction peaks at 2θ = 14.7°,16.3°,22.5° and the characteristic diffraction peak of cellulose I. Two peaks at 2θ = 18.5°(I am ) and 22.5°(I 200 ) represent the amorphous and crystalline regions of cellulose, respectively (Lim et al. 2021). The small peak at 2θ = 34.6° proved that the fiber had the natural cellulose Iβ structure (Hachaichi et al. 2021). The peak shape of the treated OP fiber remains the same, indicating that the treatment does not change the crystal structure of the OP fiber. As the sample was treated, the peak intensity at 2θ = 22.5° increased, meaning that hemicellulose and lignin in the amorphous region of the fiber were removed, and cellulose content and crystallinity increased. After dewaxing, alkali treatment and bleaching, the amorphous non-cellulose compounds in the fiber were removed, and the crystallinity of OP increased from 34.80 to 68.60%. Compared with the crystallinity of microcrystalline cellulose 81.23%, there is still a gap. This is due to the extracted cellulose still has a small amount of lignin is not completely removed, cellulose content is lower than MCC.

TG/DTG analysis
It can be seen from Fig. 5a that the thermal decomposition process of OP in different steps is divided into three stages. The first stage is 30-130 °C. At this stage, the weight of OP fiber decreased slightly (< 6%), mainly due to the evaporation of some volatile compounds and water. The second stage is 195-403 °C, which is mainly caused by dehydration, decarboxylation and depolymerization of glycosidic bond. At this stage, cellulose, hemicellulose and lignin in lignocellulose will be degraded. Hemicellulose is the easiest to degrade, and its thermal degradation zone is mainly at 220-315 °C. Because hemicellulose is composed of a variety of sugars, it is randomly amorphous structure, so it is easy to degrade into volatile substances such as CO and CO 2 . Compared with hemicellulose, cellulose is composed of long chains of unbranched glucose, which is arranged in order. Its thermal stability is higher than that of hemicellulose, and the thermal degradation zone is 315-400 °C (Yang et al. 2007). The third stage is 403-700 °C, which mainly occurs oxidation and decomposition of carbonized residue. Lignin also decomposes at this stage. Because lignin contains a large number of aromatic rings, its thermal degradation is slow and can decompose from 150 to  900 °C. The char residue is the result of the conversion of biomass into a solid residue with an aromatic ring structure, so lignin produces more char residue than cellulose (Dada et al. 2021). As shown in Fig. 5 a, the residual contents of R-OP, D-OP, A-OP and B-OP were 22.51%, 24.29%, 27.96% and 25.72% respectively. The residue content of A-OP is the highest, which may be the highest lignin content of A-OP, and lignin is related to the formation of carbon residue. As can be seen from Fig. 5b, R-OP has a smaller shoulder peak at 283 °C due to hemicellulose decomposition (Lin et al. 2021). Figure 5b shows that the weight loss peaks of R-OP, D-OP and B-OP are 335.0 °C, 339.9 °C and 331.9 °C respectively. The maximum weight loss peaks of the three fibers are all around 335 °C, which indicates that the thermal stability of cellulose treated by different methods has not changed much, but the maximum weight loss peak of A-OP is 318.6 °C, which is lower than the other four fibers, which may be that A-OP contains the highest lignin content.

Conclusion
In this paper, olive pomace was subjected to multistep chemical treatment to extract cellulose. The two steps of alkali treatment and bleaching were optimized respectively. It was found that the most influential factor on the change of cellulose content of extracted fibers in alkali treatment was reaction temperature., followed by alkali concentration. Bleaching temperature has the greatest influence on cellulose content, followed by bleaching time.
After alkali treatment and bleaching treatment, OP cellulose content increased from (40.85 ± 0.32)% to (86.41 ± 0.27)%, lignin content decreased from (25.05 ± 0.32)% to (6.77 ± 0.55)%, and hemicellulose was completely removed. The yield of each step of dewaxing and bleaching was above 82%, the yield of alkali treatment was about 61%, and the final yield of OP cellulose extraction process was 20.36%. SEM analysis of the extracted OP fibers showed that with the process treatment, the dense surface structure of cellulose was destroyed, becoming porous and irregular, and the internal cellulose was displayed. It was proved by FTIR that chemical treatment did not change the chemical structure of the fiber, OP fiber was cellulose I, and the hemicellulose and lignin in the fiber were almost completely removed after alkali treatment and bleaching. After treatment, the crystallinity of cellulose increased from 34.80% to 68.69%, indicating that hemicellulose and lignin in the amorphous region were removed and the cellulose content in OP increased. In this paper, the alkali treatment and bleaching treatment of cellulose extraction process were optimized to provide reference for the extraction of high purity and high crystalline cellulose from waste produced by industrial processing of biomass.