Properties of dietary fiber from citrus obtained through alkaline hydrogen peroxide treatment and homogenization treatment

Journal Pre-proofs Properties of dietary fiber from citrus obtained through alkaline hydrogen peroxide treatment and homogenization treatment Yue Zhang, Junru Qi, Weiqi Zeng, Yingxing Huang, Xiaoquan Yang PII: DOI: Reference:

S0308-8146(19)32011-4 https://doi.org/10.1016/j.foodchem.2019.125873 FOCH 125873

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Food Chemistry

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10 July 2019 6 November 2019 6 November 2019

Please cite this article as: Zhang, Y., Qi, J., Zeng, W., Huang, Y., Yang, X., Properties of dietary fiber from citrus obtained through alkaline hydrogen peroxide treatment and homogenization treatment, Food Chemistry (2019), doi: https://doi.org/10.1016/j.foodchem.2019.125873

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Properties of dietary fiber from citrus obtained through alkaline hydrogen peroxide treatment and homogenization treatment.

Yue Zhang, Junru Qi*, Weiqi Zeng, Yingxing Huang, Xiaoquan Yang

School of Food Science and Engineering, South China University of Technology, Guangzhou 510640, China

*Corresponding author: [email protected] Research and Development Center of Food Proteins, School of Food Science and Engineering, South China University of Technology, Guangzhou 510640, Guangdong, China Tel.: +86 20 87114262 Fax: +86 20 87114263

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Abstract In this research, citrus dietary fiber (CF) was modified by two methods, homogenization or alkaline hydrogen peroxide (chemical) treatment, to improve its physicochemical properties. The homogenization and chemical treatment highly increased the CF’s water swelling capacity by 433 % and 276 %, respectively. The water holding capacity of CF significantly increased by 253 % after homogenization and 197 % after chemical treatment. Both treatments increased CF’s total dietary fiber content and thermal stability. Moreover, the chemically treated CF thermal stability was higher than the homogenized one. Scanning electron microscopy (SEM) results showed the modified CFs exhibited porous structure. XRD and NMR results showed that the CF’s crystalline region could be disrupted by both treatments. Overall results suggest that the two treatments could effectively improve CF physicochemical properties. The modified fibers might be potentially used as functional food ingredients.

Abbreviations CF, citrus dietary fiber; DF, dietary fiber; AHP, alkaline hydrogen peroxide; WHC, water holding capacity; WSC, water swelling capacity; OHC, oil holding capacity

Keywords Citrus dietary fiber; Alkaline hydrogen peroxide treatment; Homogenization; NMR; Physicochemical properties; Water holding capacity.

1. Introduction Pectins, an acid polysaccharide, have been widely used as food ingredients, such as thickening and gelling agents. Citrus is one of the main sources of pectin (Wang, Jing, Yin, & Xie, 2014). The pectin industry is growing rapidly in the world (Marića, Grassino, Zhu, Barba, Brnčić, & Rimac Brnčić, 2018), a large amount of citrus by-products (citrus peel) are produced and discarded during pectin production. Citrus peel is rich in dietary fiber (DF) and other high-added-value compounds (i. e. 2

fats, vitamins and pigments) (Wicker, Hart, & Parish, 1989). Studies have shown that the citrus fiber is more valuable than the cereal fiber since the former produces higher total dietary fiber（TDF） content than the latter, with better functional properties (i.e. water holding and water swelling capacities) (Larrauri, 1999). It has been widely accepted that DF used as food ingredients can not only be of benefit to people’s health, but also improve foods’ functional properties, such as increasing a beverage’s emulsibility and prolonging shelf-life (Dervisoglu & Yazici, 2006). Incorporation of dietary fiber with better physicochemical properties, especially high water holding and water swelling capacities, in some food products is beneficial to food properties because it could act as an emulsifier and provide food viscosity as well as an ability to form gels (Mudgil & Barak, 2013). To maximize the utilization of resources and minimize the pollution of the environment, citrus fiber can be recovered from the orange peel after the pectin extraction process. However, there is a low recovery rate of the citrus byproducts from the pectin industry at present, because the physicochemical properties of the untreated citrus fiber are not good enough to improve the food quality. To improve the citrus fiber’s commercial value and hence its utilization rate, the physicochemical properties of citrus fiber were modified and analyzed in this research. Research has shown that the modification treatments could change the dietary fiber’s composition and micro-structure, resulting in both good and bad effects on its physicochemical properties (Peerajit, Chiewchan & Devahastin, 2012). The strong alkaline conditions could break the glycosidic linkages in DF, influencing the ratio between IDF and SDF. The alkali modified soluble fiber exhibited higher thermal stability (Margareta & Nyman, 2002; Zhang et al., 2017). The insoluble dietary fibers extracted by the ultrasound-assisted alkali method had higher water retention capacity than those of untreated soybean residues (Sun, Zhang, Xiao, Wei, & Jing, 2018). Although several studies have simply evaluated the chemical composition of alkaline hydrogen peroxide treated dietary fibers, the physicochemical properties (i. e. water swelling and oil adsorption capacities) of the 3

modified fiber have been rarely characterized and compared with those of untreated ones (Azzam, 1989; Maes, & Delcour, 2001; Aslanzadeh, Mizani, Gerami, & Alimi, 2012). In this study, we modified the citrus fiber by chemical treatment and alkaline hydrogen peroxide (AHP) methods, to improve its physicochemical properties, so that it can be applied to the food producing process to improve food properties. Homogenization is also an effective method to modify dietary fiber. Bengtsson & Tornberg (2011) stated that carrot and potato pulp suspensions consisted of large cell clusters or aggregates, which were degraded to smaller cell clusters when homogenized (9 MPa). Wang, Yuan, Xiang, & Gao (2015) found that it was feasible to use homogenization (37 MPa) to achieve the extraction of the SDF from ponkan pomace. Xie et al. (2017) reported that dietary fiber obtained from purple-fleshed potato modified by high pressure homogenization (200 MPa) showed a higher ratio of the soluble fraction. There is little research on the functional properties of homogenized fibers obtained from citrus peel. Homogenization under 30 MPa is easy to operate in the food industry and may have better effects on the citrus fiber’s functional properties. Therefore, we also treated the citrus fiber with homogenization under 30 MPa for two times to improve its properties in this research. The study on the modification of dietary fiber extracted from the citrus (orange) peel is rare. Moreover, few studies have focused on the analysis of citrus fiber functional properties (especially, water holding and water swelling capacities) improvement. The citrus fiber with better functional properties could be added to beverages or jams to improve its properties and reduce its production costs. In this research, the citrus fiber was modified by two methods, the chemical (alkaline hydrogen peroxide) or physical (homogenization) method, to improve its functional properties. We simulated two different industrial pectin extraction processes and got two natural citrus fiber which were different from the pectin content, to preliminarily study whether the pectin content had any influence on fiber’s physicochemical properties during the modification. The composition, 4

structure and physicochemical properties of the natural and modified citrus fibers were evaluated and compared.

2. Materials and methods 2.1. Materials and reagents The orange peel used in this project was acquired from Guangzhou Laimeng Biotechnology Co., LTD., China. The dried samples were kept in a desiccator at room temperature (ca. 25 ℃) until use. The reagents used in this study were all analytical grade, and purchased from Guangzhou chemical reagent factory.

2.2. Citrus dietary fiber extraction procedure We simulated two different industrial pectin extraction processes and got two natural citrus fibers with different pectin content, which were named S1-N and S2-N, respectively. To prepare S1-N, orange peel (50 g) was initially suspended in deionized water (5 % w/v), the pH was adjusted to 1.7 with saturated oxalic acid solution, and the suspension was transferred to a water bath (70 ℃, 2 h). The mixture was then centrifuged at 6000 g for 15 min, and then the residue was collected and dried in a drying oven at 60 ± 1 °C for 4 h. To prepare S2-N, S1-N (50 g) was firstly suspended in deionized water (5% w/v), pH was adjusted to 1.7 with saturated oxalic acid solution, and the suspension was transferred to a water bath (80 ℃, 1 h). The rest of the treatment was the same as the S1-N producing process. S1-N and S2-N were crushed into powder (80 mesh) using a grinder (HX-YM01, Foshan Haixun Electric Appliance Co., LTD., China), and then kept in a desiccator at room temperature (ca. 25 ℃) until used.

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2.3. Modification of citrus dietary fiber 2.3.1. Alkaline hydrogen peroxide modification To modify CF by AHP treatment, we dispersed the natural citrus fiber (S1-N or S2-N) in 1.0 % hydrogen peroxide solution (1:20, w/v), and the pH of the suspension was adjusted to 11.50 with 0.5 M NaOH solution. The suspension was then stirred in a water bath at 60 ℃ for 4 h., followed by adjusting the pH to 6.0 with HCl (1.0 M) at room temperature (ca. 25 ℃). The mixture was centrifuged at 6000 g for 15 min, the residue was then collected and washed in pure ethanol (the alcohol-fiber ratio was 1:1) and dried in a drying oven at 60 ± 1 °C for 7 h. The modified CFs were named as S1-AHP and S2-AHP referring to the natural samples S1-N and S2-N, respectively. S1-AHP and S2-AHP were crushed into powder (80 mesh) using the grinder，and then kept in a desiccator at room temperature (ca. 25 ℃) until use.

2.3.2. Homogenization modification To modify CF by homogenization, we dispersed the natural citrus fiber (S1-N or S2-N) in water (1:100, w/v) first, and then the suspension was stirred, in a magnetic stirrer, for 30 min at room temperature (ca. 25 ℃). The suspension was then treated twice by homogenization under a pressure of 30 MPa at 60 ℃. The mixture was centrifuged at 6000 g for 15 min, and the residue was then collected and washed in pure ethanol (the alcohol-fiber ratio was 1:1) and dried in a drying oven at 60 ± 1 °C for 7 h. The modified citrus fibers were named as S1-H and S2-H referring to the natural samples S1-N and S2-N, respectively. S1-H and S2-H were crushed into powder (80 mesh) by using the grinder, and then kept in a desiccator at room temperature (ca. 25 ℃) until use.

2.4. Chemical composition TDF, IDF and SDF contents were determined by AACCI method 32-07 (2000). Cellulose, hemicellulose and lignin were measured by following the method of Habibi, Mahrouz, & Vignon, 6

2002. Protein was determined by the Kjeldahl method and was calculated by the timing factor of 6.25. Fat was measured by AOAC method 996.06. The pectin was determined by using the method described by Donaghy & Mckay (1994).

2.5. Physicochemical properties 2.5.1. Water holding capacity (WHC) We calculated the samples’ WHC according to the method described by Luo, Wang, & Zheng (2017) with some modifications. Each sample (0.100 g) were well mixed with distilled water (10 ml) at room temperature (ca. 25 ℃) for 24 h, and the slurry was then centrifuged at 3000g for 15 min. Finally, the supernatant was completely removed by slowly tilting the centrifuge tube. WHC was calculated by the following equation: WHC (g/g) = (m2-m0)/(m1-m0) where m0 is the weight of centrifuge tube, m1 is the weight of centrifuge tube and sample prior to hydration, and m2 is the weight of centrifuge tube and hydrated CF sample.

2.5.2. Water swelling capacity (WSC) and oil adsorption capacity (OAC) Water swelling capacity was measured by the method of Sowbhagya, Suma, Mahadevamma & Tharanathan (2007). Each Sample (0.200 g) was well mixed with distilled water (15 ml) in the graduated cylinder at room temperature (ca. 25 ℃) for 18 h, and WSC was calculated by the following equation: WSC (ml/g) = (v1-v0) / m0 where v0 is the volume of the dried CF sample, v1 is the volume of the hydrated DF sample, and m0 is the weight of dried CF sample.

Oil adsorption capacity of CF sample was determined by the method of Li, S. (2014) with some modifications. Each samples (0.100 g) was well mixed with 10 ml of Arawana soybean oil (Yihai 7

Kerry Grain and Oil Industry Co. Ltd, Guangzhou) at room temperature (ca. 25 ℃) for 24 h, and the slurry was then centrifuged at 3000g for 15 min. OAC was calculated by the following equation : OAC (g/g) = (m2-m0)/(m1-m0) where m0 is the weight of centrifuge tube, m1 is the weight of centrifuge tube and dried sample, and m2 is the weight of centrifuge tube and DF sample, which contained the oil.

2.6. Structural analysis 2.6.1. Scanning electron microscopy (SEM) The microstructure of natural and modified samples were observed by SEM (EVO 18, ZEISS, Germany) at 15 KV. Before observation, the dehydrated samples were coated with gold and placed in a support. Each micrograph of sample was taken at 100-1000x magnification.

2.6.2. Fourier-transformed infrared spectroscopy (FT-IR) The FT-IR spectrum of samples was performed in a total reflection Fourier Transform Infrared (ATR-FTIR) instrument (VERTEX 33, Bruker Co. Ltd., Germany). The sample was mixed with KBr (1:100, w/w), and FT-IR spectra were obtained between 500 and 4000 cm−1 with a resolution of 4 cm-1.

2.6.3. X-ray diffraction (XRD) X-ray diffractometer (D8 Advance, Bruker, Germany) was used to analyze the crystalline structures of citrus fibers samples at the operating voltage of 40 kV and an incident current of 40 mA. The angular region was scanned from 3° to 55° with a step width of 0.02° and a speed of 0.2°/min. The crystallinity index (CI) of the samples was calculated by following Mohamad Haafiz, M. K. & S.J. Eichhorn’s method (2013).

2.6.4. 13C solid-state NMR The cross polarization/magic angle spinning (CP/MAS) 13C NMR spectra were performed on a Bruker DSX-400 spectrometer (Bruker Biospin Gmbh, Rheinstetten, Germany) at 100 MHz. The 8

samples were packed in a 4 mm ZrO2 rotor. All spectra were run with contact times of 1 ms. The delay time after the acquisition of the FID signal was 2 s.

2.6.5. Thermogravimetric analysis (TGA) Thermogravimetric analysis of fiber samples was carried out using Netzsch STA 449 F3 Jupiter. Each citrus sample (15 mg) was heated from 50 to 800 °C at a speed of 10 °C/min.

2.7. Statistical analyses All data has been given as means with standard deviation. SPSS package (SPSS for Windows, 16.0, 2007, SPSS Inc., USA) software programs were used for the analyses. Least significant differences (LSD) were determined by Turkey test at P < 0.05.

3. Results and discussion 3.1. Chemical composition The composition of citrus fibers were presented in Table 1. The TDF mass fractions of the six citrus fibers were all over 65 %. It showed that the TDF content of citrus fibers highly increased after the AHP treatment or the homogenization. These results may be attributed to the loss of small molecular weight oligosaccharides and some non-fiber components during the modified process, such as flavonoids, essential oils and carotenoids. Compared to the natural fibers, the chemically modified CFs showed higher IDF content (S1-AHP: 74.50 g/100 g & S2-AHP: 78.15 g/100 g). High pH-values of the AHP treatment could promote a beta-elimination of the pectin backbone, which might result in the disruption of pectin (Renard & Thibault, 1996), thus reducing the CF’s pectin and SDF content, and increasing its IDF content. The TDF (IDF) content of homogenized fiber was higher than that of the natural one, but was lower than that of the chemically treated one. These results indicated that the effects of the homogenization under 30 MPa on TDF (IDF) was slighter compared with that of AHP treatment. The 9

homogenization treatment had less impact on the fiber and non-fiber compositions of each natural fiber, and resulted in small changes in citrus fiber composition. The SDF content of S1-H (15.89 g/100 g) and S2-H (12.83 g/100 g) was higher than that of their natural ones. The homogenization treatment has the effects of turbulence, shear and cavitation, which may convert some IDF into SDF. The content of citrus fiber’s cellulose, hemicellulose and lignin was similar to that of IDF, indicating that cellulose, hemicellulose, and lignin might be the main components of citrus fiber’s IDF. Cellulose and hemicellulose content of the two raw fibers increased after the AHP or homogenization treatment. However, this change may not be caused by the increase of cellulose and hemicellulose, but by a dissolution or reduction of other small molecule polysaccharide and non-fiber components during the treatments. Besides, we could easily calculate that each fiber’s cellulose-TDF ratio and hemicellulose-TDF ratio had decreased after the AHP or homogenization treatment from Table 1. This result was consistent with our hypothesis. The pectin content of S2-N (9.58 g/100 g) was lower than that of S1-N (13.23 g/100 g), which were caused by the difference in the CF extraction processes. As described above, to obtain S2-N, we soaked S1-N in the saturated oxalic acid solution (80 ℃, 1 h). This was consistent with the previous description that suitable acid solution extraction treatment could help to release pectin (Grohmann et al., 1995). The pectin content of each chemically modified citrus fiber was lower than that of the raw sample. The loss of pectin may be due to the breaking of glycosidic linkages in the strong alkaline conditions (Margareta & Nyman, 2002). The pectin content of S1-H and S2-H was similar to that of the two natural ones, indicating that the homogenization under 30 MPa may not be sufficient to destroy the pectin structure. Pectin content was similar to SDF content in each sample, indicating that pectin might be the main component of SDF in citrus fiber. Results showed that there were low lignin, fat, and protein contents in CF. In addition, the lignin fraction in the six samples was similar to each other. 10

3.2. Physicochemical properties 3.2.1. Water holding capacity (WHC) Fig. 1 (A) demonstrated WHC of citrus fibers. The WHC value of S1-N, S2-N, S1-AHP and S2-AHP was 8.64 g/g, 6.24 g/g, 13.25 g/g and 12.28 g/g, respectively. After the AHP treatment, the WHC value of S2-N (S1-N) increased by 197 % (153 %), indicating that the chemical treatment could effectively increase the citrus fiber’s water holding capacities. Compared to AHP treatment, the CF modified by homogenization had higher WHC values (S1-H: 14.97 g/g & S2-H: 15.75 g/g), indicating that the homogenization is also an effective method to improve the functional properties of citrus fiber. Besides, the WHC of S1-N was higher than that of S2-N. As shown in Table 1, S1-N had higher pectin content than S2-N, this result indicated that pectin content may have positive effects on citrus fiber’s water holding capacity. The AHP or homogenization treatment may break the hydrogen bonding in CF, which could help CF lose its structure and expose more hydrophilic hydroxyls to water. The AHP treatment may cause the loss of some non-hydrophilic components (lignin, Hemicellulose and cellulose, etc.). The homogenization under 30 MPa may have a greater effect on CF surface and inner structure than chemical treatment. These changes in citrus fiber’s compositions, inner structure and surface may be the main reasons for the increase of its water holding capacity. In the food industry, dietary fiber with high water holding capacity could be added into jam or fruit juice to improve its properties, prolong its shelf-life and reduce its production costs. These results showed that the AHP or homogenization treatment could effectively improve citrus fiber’s water holding capacities, which could further promote CF’s potential use as a food additive in the food industry.

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3.2.2. Water swelling capacity (WSC) and oil adsorption capacity (OAC) The WSC and OAC results are shown in Fig. 1 (B) and (C). The WSC property is related to the micro-structure (surface, honey-comb appearance and cavities) and chemical composition (the SDF and IDF content) of CF (Yalegama, Nedra Karunaratne, Sivakanesan & Jayasekara, 2013). The WSC of S1-N, S2-N, S1-AHP, S2-AHP, S1-H and S2-H was 7.32 ml/g, 3.64 ml/g, 11.63 ml/g, 10.04 ml/g, 15.15 ml/g and 15.76 ml/g, respectively. The data showed that both chemical treatment and the homogenization under 30 MPa could significantly improve the water swelling capacity of citrus fibers. The growth of citrus fiber’s water swelling capacity may be due to its wrinkled surface, porous structure and disruption of hydrogen bonds improved by the chemical and homogenized methods. The data also revealed that the water swelling capacity of homogenized CFs was higher than that of AHP treated ones. The homogenization could effectively loosen the citrus fiber’s inner structure and damage its crystal structure, which lead to a greater increase in citrus fiber’s water swelling capacity than the chemical treatment. Besides, the WSC of S1-N was higher than that of S2-N, which could be explained by the higher pectin content in S1-N. There was no significant difference in WSC values of citrus fibers modified by the same method, and the previous citrus fiber’s WHC analysis results had similar results. In addition, the citrus fibers’ WHC values (Fig. 1-A) and WSCs had similar increasing trends after the two modification treatments, suggesting that the citrus fiber’s water holding capacity and water swelling capacity properties had similar mechanisms. The oil adsorption capacity of dietary fiber is commonly considered as an important parameter to evaluate its adsorption capacity for lipophilic components. Citrus fiber with higher oil adsorption capacity has a better ability to prevent fat loss during food processing (Navarro-González, García-Valverde, García-Alonso & Periago, 2011). The OAC of S1-N, S2-N, S1-AHP, S2-AHP, S1-H and S2-H was 2.69 g/g, 2.69 g/g, 2.62 g/g, 2.68 g/g, 3.50 g/g and 3.52 g/g, respectively. The results 12

showed that the the homogenized citrus fibers had higher oil adsorption capacity than the raw samples and chemically modified ones. This may be because homogenized fibers had a rougher surface and more porous structure than the chemically modified ones (Fig. 2.), which could help citrus fiber absorb and retain more oil. Besides, the OAC values of raw samples and chemically modified ones were similar to each other, indicating that AHP modification had little effect on citrus fiber oil adsorption capacity.

3.3. Structural analysis 3.3.1. Scanning electron microscopy The SEM images of the citrus fibers are shown in Fig. 2. It showed that no significant difference was observed in the surface structure of chemically modified fibers (S1-AHP and S2-AHP), this was because they were made by the same chemical method, and their natural fibers’ structure were similar to each other as presented in Fig. 2. The chemically modified CFs seemed significantly different from their natural ones. A wrinkled surface and tiny pores clearly appeared in S1-AHP (Fig. 2-b2) and S2-AHP (Fig. 2-b4). The strong alkaline conditions could break the glycosidic linkages in DF (Margareta et al. 2002), destroy the micro-structure and result in the degradation of some cellulose, hemicellulose and pectin which may cause many holes in the citrus fiber, thus each AHP modified citrus fiber had a clearer wrinkled surface and looser inner structure than the raw one. Compared to AHP treatment, the CF modified by the homogenization under 30 MPa demonstrated a more wrinkled surface, looser microstructure and more porous (Fig. 2-b6 and b7), this may be because the inner structure and surface area of citrus fiber were largely damaged or loosened by physical factors, such as collision and shear. These results indicated that the homogenization might be a more effective method than the AHP treatment to improve CF microstructure.

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Research showed that the porous structure could help fiber to absorb and retain water (Biswas, Kumar, Bhosle, Sahoo & Chatli, 2011). Citrus fiber with a loose structure and wrinkled surface might have high water holding and water swelling capacities. The results of SEM suggested that both the alkaline hydrogen peroxide and homogenization treatments could improve the citrus fiber microstructure, which could further help to improve its physicochemical properties, such as water holding capacity. In fact, this was consistent with the results of physicochemical properties showed in Fig. 1.

3.3.2. FT-IR analysis Fig. 3. shows the infrared spectrums of citrus fibers. As for the IR spectra of the six CF samples, they showed similar profiles but various absorption intensity. The broad absorption at about 3411 cm−1 ascribed to the vibrations of the hydrogen bound of the hydroxyl groups (Cui, Phillips, Blackwell & Nikiforuk, 2007), which was mainly from cellulose, hemicellulose, lignin. Blue shifts were observed in both the AHP treated citrus fibers (S1-AHP & S2-AHP ) and the homogenized ones (S1-H & S2-H) as compared to the natural groups (S1-N & S2-N), which could be due to the destruction of hydrogen bonds formed in the hydroxyl groups of polysaccharides during AHP treatment and homogenization. The breaking of hydrogen bonding could help citrus fiber form a coarse and porous structure, and increase the hydrogen bonds between the citrus fiber and water, which may lead to the increase of citrus fiber’s water holding capacity (Fig. 1). The vibration signal in the range 2800-3000 cm−1 ascribed to C-H strething (Cui et al., 2007). The absorbance peak at about 1730 cm−1 was responsible for uronic acid from pectin (Coates, 2006). This peak intensity of the chemically modified CFs was weaker than that of their natural ones. These results indicated that the AHP treatment had removed some pectin, this might be because the strong alkaline treatment could promote a beta-elimination of the pectin’s backbone and result in the degradation of some pectin (Table 1). The band at 1640 cm-1 of homogenized CFs was responsible for the carboxyl groups that were interconnected with cellulose chains by forming inter-molecular hydrogen bonds, and this band

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intensity decreased after homogenization. This indicated that the effects of turbulence, shear and cavitation of the homogenization might result in the destruction of some hydrogen bonds between CF cellulose chains. This hydrogen bond changes in CF could help the citrus fiber loosen its inner structure and form more fiber-water hydrogen bonds when it is exposed to water. The peak at about 1245 cm-1 ascribed to the vibration of C-O methoxyl groups in lignin and hemicellulose (Sain, 2006), the peak at 898cm-1 and 811 cm-1 were related to the b-glycosidic linkages of polysaccharides (Reddy, Reddy, Zhang, Zhang & Varada Rajulu, 2013). However, these peaks of the chemically modified fibers decreased or even disappeared, indicating the degradation of CF’s pectin, lignin and hemicellulose. The removal of hemicellulose and lignin caused by chemical treatment could help to increase the CF surface roughness and improve its water holding capacity. The peaks at 1245 cm-1 and 818 cm-1 of homogenized modified fiber disappeared, suggesting that the homogenization under 30 MPa could cause the reduction of some hydrogen bonds in citrus fiber (Mealer, Jones, Newman, Mcfann, Rothbaum & Moss, 2012), and loose its inner and surface structure.

3.3.3. X-ray diffraction Fig. 4 (A) shows the X-ray diffraction of citrus fibers. The prominent intensity peak was clearly observed at about 22 °, indicating that the natural citrus fibers were cellulose I type (Segal, Creely, Martin & Conrad, 1959). The peak position of the natural samples was similar to the chemically modified and homogenized fibers, indicating that the crystalline type of citrus fibers did not change after modification. The crystallinity indices of S1-N, S2-N, S1-AHP, S2-AHP, S1-H and S2-H were 40.42 %, 42.00 %, 43.39 %, 44.17 %, 39.96 % and 37.94 %, respectively. The crystallinity indice (CI) of the two natural fibers was reduced by 0.5 %-4.1 % after processing by homogenization. The results suggested that the crystal structure of citrus fiber was damaged by physical factors, such as collision 15

and shear, leading to the citrus fiber’s crystalline structure shifted from ordered to less ordered. On the contrary, the CI of citrus fiber was slightly increased after AHP treatment. This was due to the disruption of some cellulose chains and the removal of amorphous constituents during the chemical process, such as lignin, amorphous cellulose, hemicellulose and pectin. The result also revealed that the amorphous constituents were more reactive than crystalline cellulose. Besides, the CI of S1-N was lower than that of S2-N, which may be because S1-H had higher pectin content (Table 1). 13

3.3.4.

C solid-state NMR

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C CPMAS NMR spectra of all studied citrus fibers were recorded in Fig.4 (B). It showed large

signals in the carbohydrate region between 60 and 110 ppm. The resonance at δ60-70 ppm is attributed to C6; The cluster of resonances at δ70-81 ppm is assigned to the ring carbons C2, C3 and C5; The signal at 79-86 ppm is assigned to the C4 of amorphous cellulose; The signal at 86-93 ppm corresponds to C4 of crystalline cellulose; The resonance at δ98-108 ppm is attributed to C1 (Wawer, Wolniak & Paradowska, 2006; Liitiä, Maunu, Hortling, Tamminen, Pekkala & Varhimo, 2003). The signal intensity at 79-86 ppm (C4 of amorphous cellulose) of the AHP modified fibers (S1-AHP & S2-AHP) and homogenized fibers (S1-H & S2-H) was higher than that of their natural ones. This result indicated that some of the crystalline region in cellulose was disrupted and changed into the amorphous region by AHP or homogenization treatment. Besides, The peak ratio of C4(79–86 ppm)/C4(86–92 ppm)，which

is often used to estimate the cellulose crystallinity (Liitiä et al., 2003),

decreased after the AHP treatment. This revealed that the crystallinity indice of the citrus fibers increased after this treatment, and amorphous cellulose was more reactive during the chemical treatment process. These results were consistent with the previous X-ray diffraction findings. The pectin characteristic signals appear at 170-174 (C=O), 98-104 (C1), 60–84, 53 (OCH3) and 18 ppm (CH3). Compared to the natural fibers, these signal peaks of chemically modified samples (S1-AHP & S2-AHP) decreased or even disappeared, indicating that the AHP treatment could cause 16

the degradation of pectin, similar results were found in the previous FTIR analysis. Besides, the peak intensity of the natural fibers was similar to that of homogenized fibers, indicating that the pectin content was not significantly changed by the homogenization under 30 MPa. 3.3.5. Thermogravimetric analysis The thermal decomposition of the raw citrus fibers (S1-N & S2-N), and their modified samples (S1-AHP, S2-AHP, S1-H and S2-H) has been carried out through thermogravimetric analysis. TGA curves are shown in Fig. 5. The weight loss of the samples mainly occurred at the temperature range of 50–600 °C, which can be roughly divided into three stages: 50-150 ℃,150-400 ℃, 400-600 ℃ (Zhang et al., 2017). The weight loss at the first stage (50-150 ℃) was mainly caused by absorbed water evaporation and the decomposition of low molecular weight polysaccharides (Juan, Alvarez, Cyras & Analia, 2008). In this stage, the curves showed similar weight loss among the samples. The second stage (150-400 ℃) was mainly due to the degradation of hemicellulose, lignin, pectin, etc. At this stage, the weight loss was rapid, and the initial degradation temperature for S1-N, S2-N, S1-AHP, S2-AHP, S1-H and S2-H was at about 150 ℃, 160 ℃, 180 ℃, 195 ℃, 204 ℃ and 205 ℃, respectively. After the AHP or homogenization treatment, the initial degradation temperature of CF highly increased, indicating that the AHP or homogenization treatment could result in the improvement of CF thermal stability. This might be because the AHP or the homogenization under 30 MPa increased the CF’s crystallinity indice and caused the disruption and even removal of its active components, such as amorphous cellulose or pectin. The weight loss of CFs at the third stage (400-600 ℃) was slow, probably due to decomposition of some lignin and complex polymer compounds. The residue weight of S1-N, S2-N, S1-AHP, S2-AHP, S1-H and S2-H was 24.96%, 24.02%, 33.31%, 32.06%, 30.14% and 26.74%, respectively. It showed that the residue weight of citrus fiber increased after the two treatments respectively. Residue 17

weight of chemically modified fibers was higher than that of homogenized ones, indicating that the thermal stability of CFs was improved after both treatments, and chemically treated fibers had a better thermal stability than the homogenized fibers. The crystalline region of dietary fiber has higher stability than the amorphous region. The result showed that the chemical treatment had a greater influence on the CF’s amorphous region than the the homogenization under 30 MPa, the former could remove more amorphous areas in citrus fiber and increased its thermal stability.

4. Conclusion The results of this study demonstrate that the citrus fiber physicochemical properties could be significantly improved by alkaline hydrogen peroxide treatment or homogenization under 30 MPa. The modified citrus fibers present significantly higher water holding and water swelling capacities than the natural ones. Moreover, the homogenized citrus fibers water holding and water swelling capacities, as well as oil adsorption capacity values, are all higher than the chemically treated ones. Besides, the citrus fibers’ thermal stability increases after both treatment. The alkaline hydrogen peroxide treatment increases citrus fiber’s total dietary fiber content, degrades some hemicellulose, lignin and pectin, and loosens the microstructure, and the homogenization effectively damages citrus fiber’s crystal structure and also loosens its inner structure. These modified citrus fibers with good functional properties, such as high water holding and water swelling capacities, could be used as emulsifiers in some food, such as juice or jam, and help to increase its viscosity and other properties. Overall this comprehensive study shows that the homogenization and alkaline hydrogen peroxide treatment may be of great significance to the improvement of citrus fiber’s application in the food industry. Our research will be continued to improve citrus fiber functional properties to a greater extent.

Acknowledgements This work was supported by the National Natural Science Foundation of 351 China (31370036). 18

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figure captions

Fig. 1. The WHC (A), WSC (B) and OAC (C) of natural citrus fibers (S1-N and S2-N), chemically modified and homogenized citrus fibers (S1-AHP, S2-AHP, S1-H and S2-H).

Fig. 2. Scanning electron micrographs of natural citrus fibers (S1-N and S2-N), chemically modified and homogenized citrus fibers (S1-AHP, S2-AHP, S1-H and S2-H). (a1 : S1-N × 100; a2 : S1-AHP × 22

100; a3 : S1-H × 100; a4 : S2-N × 100; a5 : S2-AHP × 100; a6 : S2-H × 100; b1: S1-N × 1000; b2 : S1-AHP × 1000; b3 : S1-H × 1000; b4 : S2-N × 1000; b5 : S2-AHP × 1000; b6 : S2-H × 1000 ).

Fig. 3. FTIR spectra of natural citrus fibers (S1-N and S2-N), chemically modified and homogenized citrus fibers (S1-AHP, S2-AHP, S1-H and S2-H).

Fig. 4. The X-ray diffraction (A) and 13C CPMAS NMR spectra (B) of natural citrus fibers (S1-N and S2-N), chemically modified and homogenized citrus fibers (S1-AHP, S2-AHP, S1-H and S2-H).

Fig. 5. TGA analysis of natural citrus fibers (S1-N and S2-N), chemically modified and homogenized citrus fibers (S1-AHP, S2-AHP, S1-H and S2-H).

(A)

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(B)

（C） Fig. 1.

24

Fig. 2.

Fig. 3.

25

(A)

(B)

Fig. 4.

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Fig. 5.

Table 1 Chemical composition of natural citrus fibers (S1-N, S2-N), chemically treated citrus fibers (S1-AHP, S2-AHP ), and homogenized citrus fibers (S1-H, S2-H). Chemical composition

S1-N

S1-AHP

S1-H

S2-N

S2-AHP

S2-H

Protein

5.24±0.16a

5.47±0.12a

5.30±0.10a

5.34±0.13a

5.42±0.18a

5.27±0.16a

Fat

2.62±0.057a

1.21±0.061b

2.54±0.076a

2.30±0.045a

0.95±0.023b

2.38±0.065a

TDF

65.60±0.72d

81.81±1.26b

75.33±1.00d

78.10±0.83c

85.02±1.05a

84.14±0.56a

IDF

52.26±0.73f

74.50±0.55b

60.33±0.43e

68.23±0.61d

78.15±0.57a

72.23±0.66c

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SDF

13.32±0.53b

7.24±0.67d

15.89±0.56a

9.80±0.45c

6.25±0.53d

12.83±0.55b

Cellulose

32.68±0.45e

40.44±0.97a

36.57±0.56c

35.03±0.75d

41.51±0.84a

39.14±0.42b

Hemicellulose

12.96±0.75d

28.46±069b

16.99±0.77e

26.95±0.35c

30.61±0.71a

27.35±0.75c

Lignin

6.09±0.31a

5.45±0.81a

6.01±0.61a

6.01±0.71a

5.75±0.57a

5.99±0.51a

Pectin

13.23±039b

7.07±0.75e

14.50±0.46a

9.58±0.65d

6.07±0.35e

11.75±0.76c

Values are means of three determinations ± standard deviation. Data were expressed as “g/100 g dry basis”. TDF (total dietary fiber); SDF (soluble dietary fiber); IDF (insoluble dietary fiber). Values of different letters are significantly different in the same line, P < 0.05.

►The citrus fiber was recovered from the orange peel after pectin extraction process. ►The properties of citrus fiber improve during the chemical treatment or

homogenization. ►The water holding capacity of citrus fiber could increase by 253% after

homogenization. ►The modified citrus dietary fibers present obvious cavity and a wrinkled surface. ►The chemically treated citrus fiber had higher thermal stability than the homogenized

treated one.

Declaration of interests

☒ The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. ☐The authors declare the following financial interests/personal relationships which may be considered as potential competing interests:

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