- Email: [email protected]
Impact of binding interaction characteristics on physicochemical, structural, and rheological properties of waxy rice flour
Accepted Manuscript Impact of binding interaction characteristics on physicochemical, structural, and rheological properties of waxy rice flour Zhenni Li, Li Wang, Zhengxing Chen, Qiusheng Yu, Wei Feng PII: DOI: Reference:
S0308-8146(18)30800-8 https://doi.org/10.1016/j.foodchem.2018.05.010 FOCH 22843
To appear in:
Food Chemistry
Received Date: Revised Date: Accepted Date:
2 January 2018 26 March 2018 2 May 2018
Please cite this article as: Li, Z., Wang, L., Chen, Z., Yu, Q., Feng, W., Impact of binding interaction characteristics on physicochemical, structural, and rheological properties of waxy rice flour, Food Chemistry (2018), doi: https:// doi.org/10.1016/j.foodchem.2018.05.010
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Impact of binding interaction characteristics on physicochemical, structural, and rheological properties of waxy rice flour Zhenni Lib, Li Wangab, Zhengxing Chenabc*, Qiusheng Yub, Wei Fengb a
State Key Laboratory of Food Science and Technology , Jiangnan University, Lihu Road 1800, Wuxi 214122, China
b
School of food science and Technology, National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, Lihu Road 1800, Wuxi 214122, China
c
Jiangsu Provincial Research Center for Bioactive Product Processing Technology, Jiangnan University, Lihu Road 1800, Wuxi 214122, China
*Corresponding author
Tel: +86-510-8519 7110; Fax: +86-510-8519 7110; E-mail: [email protected]
Abstract
Impact of covalent and non-covalent binding interactions on physicochemical, structural, and rheological properties of waxy rice flour were investigated using several typical types of chemicals. Dynamic rheometer was used to investigate the viscoelastic properties and resistance to deformation or shearing. Both hydrogen bonds and hydrophobic interactions were necessary for waxy rice flour to form a material with stable structure matrix. Besides this, X-ray diffraction pattern and pasting property measurements showed that the relative crystallinity and pasting viscosity values of waxy rice flour exposed to SDS and L-cysteine treatments increased whereas those of urea decreased compared with the control. Addition of acid, alkali, and NaCl might result in electrostatic forces, ion interactions, and indirectly change hydrogen bonds and hydrophobic interactions between starch and protein, which played a synthetic role on property changes of waxy rice flour.
Keywords: Waxy rice; Starch; Protein; Binding interaction characteristics;
1
1. Introduction
Rice (Oryza sativa L.) is the staple food for about 3.5 billion people worldwide. Starch is the main storage carbohydrate in plant organisms. Normal starch in granular form is generally composed of two types of molecules, amylose and amylopectin. Waxy starches consist almost exclusively of amylopectin, a high molecular weight molecule. Protein is the most abundant component in rice grain next to starch, and the interaction between protein and starch are important as they can have potential effects on processing and cooking properties of waxy rice and its products. In food systems, there are covalent and non-covalent interactions among compounds involving disulfide bonds, hydrogen bonds, hydrophobic interactions, electrostatic forces, ionic interactions, and so on. Recent advances in understanding these interaction modes further illustrate diverse aspects of their nature and importance in food systems. For starch gels, non-covalent chain interactions were generally believed to involve intermolecular double helix formation stabilized by hydrogen bonds (Šárka & Dvořáček, 2017). Decreasing in hydrogen bonds within starch led to a significant decreasing in relative crystallinity and retrogradation enthalpies of corn starch (Niu, Zhang, Xia, Liu, & Kong, 2018). Schober, Bean, Tilley, Smith, and Ioerger (2011) claimed that hydrophobic interactions rather than disulfide bonds were the key to gluten-like functionality of zein and kafirin. According to Renzetti et al. (2012), the formation of large protein complexes induced by transglutaminase treatment was ascribed to the new and stronger hydrophobic interactions among proteins. These complexes could promote a more viscoelastic 2
matrix and modify physicochemical properties of rice flour (Marco & Rosell, 2008). The interactions between starch and protein were mediated by a number of non-covalent interactions included site recognition with the aromatic side chain of Trp, Tyr or, less frequently, Phe of the protein. Binding of carbohydrates to protein also involved electrostatic interactions and H-bonding. Charged residues (Arg, Lys, Asp and Glu) were generally found in the binding sites in carbohydrate–protein (Fadda & Woods, 2010). Recognition and binding of carbohydrates to proteins always take place through the formation of complex H-bonding networks and involve charge–charge interactions. The objective of this paper was to explore the effects of binding interaction characteristics on physicochemical, structural and rheological properties of waxy rice flour system. The results reported were expected to provide significant clues for separating pure starch from protein in rice system and lay a foundation for illuminating property changes of other similar starch-protein enriched systems.
2. Materials and methods
2.1. Materials
One Oryza sativa Thai native cultivar of waxy rice was kindly provided by Ingredion Incorporated (Shanghai, China). Moisture, total starch and protein content of waxy rice were 12.5%, 77.4% and 6.3%, respectively, determined by AACC Approved Methods (AACC International, 2000). All other chemicals utilized were of analytical grade or higher. 3
2.2. Methods
2.2.1. Samples preparation Waxy rice was soaked in a 45 ◦C water bath for 1 hr. After soaking, waxy rice was milled under continuous addition of water and sieved through an 80-mesh sifter to obtain uniformity of particle size. The rice slurry and different chemicals with concentration of 2% (w/w) were mixed to achieve the required final concentration of 25% (w/w). A series of chemicals were added, including HCl and NaOH at pH of 5 and 9, NaCl, SDS, urea, and L-cysteine (L-Cys). The mixtures were reacted with agitation using a controlled-rotational speed mixing device (IKA® RW 20; Wilmington, NC, USA) at 25 °C for 4 hr. Afterwards, the mixtures were centrifuged at 4000 × g for 5 min, the supernatant was removed, and the precipitate (solid residue) was retained and rinsed with water three times for removal of redundant chemical solution. The remaining precipitate was oven dried at 40 ◦C, grounded and sieved through a 100-mesh sifter for further determinations. 2.2.2. Chemical composition and solubility determination Approved AACC Methods were used to determine starch content using a total starch assay kit (Megazyme International Ireland, Wicklow, Ireland) and protein content (N × 5.95) according to the Kjeldahl method of waxy rice flour samples (AACC International, 2000). Solubility properties were determined by the method of Derycke, Veraverbeke, Vandeputte, Man, Hoseney, and Delcour (2005) with some modifications. About 1.5 g flour and 25 mL deionized water were weighted in tube and shocked at room 4
temperature for 1 hr. This was then incubated at constant temperature (30 ◦C, 50 ◦C, 70 ◦C and 90 ◦C) at 160 rpm for 30 min in a water bath. Sample was cooled down to room temperature and centrifuged (4000 × g, 15 min). Water solubility (WS, g/100g) was calculated as follow: WS = dried supernatant weight / dry flour weight × 100. 2.2.3. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) SDS-PAGE (12% separating gel and 4% stacking gel) was performed on a discontinuous buffered system according to the method of Xia, Wang, Gong, Yang, Yin, and Qi (2012). About 1.5 mg waxy rice flour sample was dissolved in 100 μL of sample buffer (0.125 mol/L Tris-HCl buffer, 1% SDS (w/v), 20% glycerol (v/v), 2% 2-mercaptoethanol (2-ME, v/v), pH 6.8), and heated at 95 °C for 5 min. Gels were stained with Coomassie Brilliant Blue R-250 (0.1%) in methanol/acetic acid/water (45:10:45, v/v/v) and de-stained in methanol-water solution containing 10% acetic acid (methanol/acetic acid/water =1:1:8, v/v/v). 2.2.4. X-ray diffractometry X-ray diffraction patterns of samples were obtained using an X-ray diffractometer (D2 Phaser, Bruker-AXS, Germany). The scanning region of the diffraction angle (2θ) ranged from 3 ◦ to 40 ◦ at a step size of 0.02 ◦ with a count time of 0.6 s. Relative crystallinity (%) was calculated as the percentage ratio of the sum of total crystalline peak areas to that of the total diffractogram (sum of total crystalline and amorphous peak areas) by software (MDI Jade 6). 2.2.5. Raman spectroscopy All spectra of the samples were collected using the LabRAM HR Evolution 5
system (HORIBA Jobin Yvon S.A.S., Longjumeau Cedex, France) with a 532 nm laser source, motorized microscope and a cooled charge-coupled device (CCD) detector. The spectral data were collected in the range of 400-1800 cm-1 at a resolution of 0.80 cm-1 with constant measurement parameters (acquisition time: 10 s; laser power: 100 mW). As the spectrophotometer was sensitive to the change of outer environment condition, the spectra were collected at controlled temperature (25 °C) and humidity (60%). The software package LabSpec 6 (HORIBA Jobin Yvon S.A.S., Longjumeau Cedex, France) was used for spectral data acquisition. 2.2.6. Pasting properties Pasting properties were determined using a Rapid Visco-Analyzer (Perten RVA 4500, Australia), as reported previously (Jeong, Kim, & Lee, 2017). Each sample (3 g of flour at 14% moisture basis) was directly weighed into an aluminum RVA canister. Then, 25 mL of distilled water was added to achieve a total weight of 28 g. The formed slurry sample was heated to 50 °C and stirred at 960 rpm for 10 s and fixed at 160 rpm throughout rest of the run. Then, sample was held at 50 °C for 0.5 min, and then heated up to 95 °C for 3.5 min and held for 2.5 min, and cooled down to 50 °C for 3.5 min and held for 2 min. Parameters recorded were peak, trough, final viscosity, breakdown, setback, pasting temperature and pasting time. 2.2.7. Rheological measurements Rheological measurements were conducted using a DHR-3 rheometer (TA Instrument, USA) as reported previously (Matos, Sanz, & Rosell, 2014). Waxy rice flour dispersions (10% w/w) were moderately stirred and heated in a boiling water 6
bath for 10 min by a magnetic stirrer until fully gelatinized. The hot pastes obtained were immediately cooled down at 30 °C for 10 min and measurements were measured at 30 °C using 40 mm diameter plate. Oscillatory frequency test from 0.1 to 10 Hz was performed in the linear viscoelastic region at a target strain of 1%, and storage modulus (G'), loss modulus (G''), and tan δ (G''/ G') versus frequency values were recorded. Steady shear flow measurement was carried out at shear rate lineally from 0 s-1 to 300 s-1. Shear stress (τ) was recorded as a function of shear rate. Creep and recovery measurements were carried out as follows: The creep phase was recorded at a shear stress of 10 Pa for 60 s, followed by a recovery phase of 120 s at a stress of 0 Pa. Creep and recovery curves were recorded and analyzed by RHEOPLUS/32 version 3.21 software to obtain the parameters, including the maximum creep compliance (Jmax), zero shear viscosity (η0), relative elastic part of the maximum creep compliance (Je/Jmax), and the relative viscous part of the maximum creep compliance (Jv/Jmax). 2.2.8. Statistical analysis All tests were performed in triplicate unless specified. Data were evaluated by analysis of variance (ANOVA), and a comparison of means were carried out with Tukey’s HSD test. Differences were considered to be significant at p < 0.05. Statistical computations and analyses were conducted using SPSS software (version 19.0).
7
3. Results and Discussion
3.1. Physicochemical properties
3.1.1. Chemical composition The starch and protein content of waxy rice flour with different treatments were reported in Fig. 1a. Treatments with urea and NaCl had pronounced effect on starch and protein content of waxy rice flour. Addition of urea led to a weakening in hydrogen bonds, thus promoting starch and protein leaching into solution and decreasing by 2% and 9% compared with that of the control. The addition of NaCl had positive effect on the starch content (91%), which might be because NaCl weakened the electrostatic attraction between the protein and polysaccharide molecules and thereby decreasing the protein content in systems. Besides this, it was reported that increasing the salt level enhanced the mechanical strength of starch through direct electrostatic interactions with the hydroxyl groups of starch molecules (Chuang, Panyoyai, Shanks, & Kasapis, 2017). The control sample exhibited the highest protein content (7.0%) and the content in decreasing order of other treatments was pH 5, pH 9, NaCl, L-Cys, urea and SDS. The content decreased by values of 2-23% compared with protein content in control. The protein extractabilities of all treated samples increased even more, indicating that covalent disulfide bonds and non-covalent interactions were involved in the formation of the protein network of waxy rice. SDS-PAGE of waxy rice flour samples were measured to investigate changes of polypeptide profiles, as shown in Fig. 2. The major polypeptide glutelin (19–25 and 8
30–39 kDa for basic and acidic subunits respectively) and 49–62 kDa for pro-glutelin were identified. The 10–16 kDa fraction corresponded to prolamin with the 13 kDa being predominant, which was known to be an integral part of rice protein. Two rice endosperm globulin polypeptides, having MW of 16 kDa and 25 kDa, had been identified as reported previously (Amagliani, O'Regan, Kelly, & O'Mahony, 2017).The profiles of waxy rice flour exposed to pH 5 and pH 9 were similar compared with the control. Upon NaCl and SDS treatments, the band at 23 kDa disappeared. Waxy rice flour treated with SDS and urea exhibited a decrease of higher molecular weight marks (> 75 kDa), indicating the disruption of bigger peptides and release of smaller peptides into solution. Apart from other treatments, high molecular weight (>75 kDa) aggregates of sample with addition of L-Cys were not observed in separated bands, which suggested that L-Cys could interact with waxy rice flour proteins by splitting the disulfide bonds and decrease the content of large macromolecular complexes (Xu et al., 2016). 3.1.2. Solubility The water solubility of waxy rice flour with different treatments at 30-90 °C were shown in Fig. 1b. Values of all samples increased dramatically with increasing temperature and the solubility of control was increased from 2.2% to 41.7% with temperature increased from 30 °C to 90 °C. Addition of urea showed the highest effect on solubility of waxy rice flour at any test temperature. Addition of SDS, which disrupted protein structure mainly affecting hydrophobic interactions, showed the most significant changes in water solubility of waxy rice flour next to that of treated 9
with urea. This suggested that non-covalent, particularly hydrogen bonds type and hydrophobic interactions seemed to be the most important in stabilizing the waxy rice flour matrix. This was consistent with previous studies which had investigated the interaction between maize starch and amaranth protein (Condés, Añón, Mauri, & Dufresne, 2015). Solubility of samples with pH, NaCl and L-Cys treatments changed more markedly and more rapidly under higher temperatures.
3.2. Structural properties
3.2.1. X-ray diffraction pattern and crystallinity The X-ray diffraction patterns of waxy rice flour with different treatments were presented in Fig. 3a. All samples displayed the typical A-type diffraction patterns with strong diffraction peaks at about 15 ° and 23 ° (2θ), and an unresolved doublet at around 17 ° and 18 ° (2θ). The relative crystallinity of waxy rice flour exposed to pH 5, pH 9, SDS, and L-Cys treatments increased whereas those of urea, NaCl decreased compared with that of the control. According to Sangpring, Fukuoka, and Ratanasumawong (2015), the formation of hydrogen bonds and starch retrogradation were limited with NaCl addition, because of co-action of electrostatic repulsion and steric hindrance effect. They explained that the H+ ions of the starch hydroxyl group migrate from starch molecules into water, leading to the Na+ ions being pushed and entrapped by the starch molecules. Since the radius of Na+ ions is greater than for H+ ions, the formation of hydrogen bonds and starch retro-gradation are limited. Renzetti et al. (2012) suggested that polymerization through disulfide bonds promotes 10
hydrophobic interactions among proteins. The reducing agent L-Cys broke disulfide bonds and more hydrophobic groups were separated. After the protein matrix was disrupted, starch granules were exposed and had more opportunity to interact with each other, thus increasing the relative crystallinity of waxy rice flour. 3.2.2. Raman spectra analysis Raman spectra of waxy rice flour samples with different treatments in the 400-1800 cm-1 region was presented in Fig. 3b. In general, the characteristics of Raman bands were observed with some changes of the bands’ positions and intensities after treatments. Each Raman curve of sample was classified and its various spectral components were associated with vibrations, rotations, etc., of the chemical bonds in the waxy rice flour’s constituent components. Changes in the relative intensity, position, and width of these bands were related to the composition (such as starch, protein, and lipids) of the sample. The main Raman bands and their preliminary assignments of waxy rice flour were listed in Table 1. Observed from Raman scans, the most obvious changes of positions and intensities happed to the bands at 874, 951, 1061, 1118, 1270, and 1349 cm-1. According to Fan et al. (2016), it might be caused by the changes in νs(C-O-C) ring mode, νs(C-O-C) α-1, 4-glycosidic linkage, C1-H bending α-configuration and changes in hydrogen bond network involving CH2OH side-chain and C-OH groups. The peak 1014 cm-1 was attributed to ring-breathing vibration of phenylalanine molecules and was observed diminishing distinguishingly with treatments except pH 5. The phenomenon indicated the treatments except pH 5 had a serious impact on certain 11
vibration of side chain of proteins (Feng, Zhang, Cong, & Zhu, 2013). Regarding characteristic bands of the starch, the bands around 781 cm-1 and 487 cm-1 were assigned to ν(C-C) ring mode and skeletal mode of pyranoid ring, respectively. For all treatments with waxy rice flour, a marked decrease and increase in the area of the band at 781 cm-1 and 487 cm-1 were observed respectively, indicating a change in the molecular order or crystallinity of the starch (Cruz, Soto, Barajas, Rodríguez, Morales, & Nicanor, 2017).
3.3. Pasting properties
Determining the pasting properties of waxy rice flour is important not only for actual applications during food processing, but also for understanding the structural changes with different treatments (Baxter, Blanchard, & Zhao, 2014). The pasting behaviors of waxy rice flour slurry were presented in Fig. 4a and different treatments showed various significant influences on pasting properties of waxy rice flour. Some treatments (NaCl and SDS) had positive effect on viscosity values and the viscosity values of waxy rice flour treated with SDS were significantly higher than that of the control (ranging from 8% to 12%). This could be due to the effective removal of protein layers that help to increase the swelling power of waxy rice flour (Chan, Bhat, & Karim, 2010). It was consistent with the result of Debet and Gidley (2006), wherein a big increase in peak viscosity was observed after SDS treatment. On the contrary, some other treatments (urea, L-Cys, and pH) had negative effects and the addition of urea significantly decreased the peak, trough, and final viscosity values of curve by 12
15%, 9%, and 16%, respectively, compared with that of the control. This result indicated that urea might block the formation of hydrogen bonds to impact intra- and inter-particle interactions of systems and resulting in lower viscosities (Niu, Wu, & Xiao, 2017). Setback viscosity values represented the differences between trough viscosity and final viscosity, and it was mainly resulted from the rearrangements between amylose and amylopectin in the system when temperature decreased from 95 to 50 °C. The setback values were decreased among all treatments by from 5% to 39%, which suggested that the transformation of the nuclei might change the environment of the starch molecules and influence the short-term rearrangements of the sample microstructure under higher water content.
3.4. Rheometer measurements
3.4.1. Oscillation All tested waxy rice flour samples behaved like a soft gel with G' > G'' and with tan δ (ratio of G''/G') < 1 (Fig. 4b) at all studied frequencies. In general, samples with addition of urea and L-Cys produced higher tanδ values than control, showing that their viscous properties were more pronounced than the elastic properties. The result agreed with the water solubility of waxy rice flour treated with urea reported previously (Fig. 1b). Blocking in hydrogen bonds promoted starch and protein leaching into solution, which might indicate a weak gel matrix of waxy rice flour with addition of urea. L-Cys and SDS disrupted protein network and decreased the viscoelasticity of waxy rice flour, indicating that disulfide bonds and hydrophobic 13
regions of internally linked protein complexes were necessary for waxy rice flour to form a material with better viscoelastic properties (Xie, Chen, Duan, Zhu, & Liao, 2008). 3.4.2. Steady flow All of the examined samples exhibited a non-Newtonian flow and were shear thinning with a tendency towards shear stress under the current experimental conditions, as observed in Fig. 4c. Higher value of the shear stress meant that the tested sample was more resistant to deformation and a more stable shear structure in terms of speed (Rożnowska & Fortuna, 2017). Among samples, a lowest value of shear stress was determined in the sample reacted with urea, followed successively by L-Cys, SDS, pH 9, Ctrl, NaCl, and pH 5. The results indicated that covalent (disulfide bonds) and non-covalent (hydrogen bonds and hydrophobic interaction) binding interactions intensified the intermolecular entanglement of waxy rice flour system, which had a significant impact on shearing sensitivity of flour paste (Smith, Bean, Selling, Sessa, & Aramouni, 2014). 3.4.3. Creep-recovery The results of creep and recovery phase curves and their analysis of all treatments were given in Fig. 4d and Table 2. Wang and Sun (2002) suggested that the maximum creep compliance (Jmax) could be used to characterize the rigidity (firmness) of dough samples. They reported that stronger dough samples which had greater resistance to deformation had smaller creep compliance than softer dough samples. The sample treated with urea exhibited the lowest deformation resistance, and the 14
decrease in Jmax values followed the order: L-Cys < NaCl < SDS < Ctrl. As seen in Table 2, the zero shear viscosity (η0) gave the flowability of samples at the end of applied stress (10 Pa) and η0 values of SDS and urea were higher than the others. This suggested that it might be difficult for particles to attract each other and to maintain their shape of waxy rice flour pastes, which resulted in higher flowability. This trend was consistent with the result of the solubility on sample treated with SDS and urea. The creep-recovery curves also exhibited a typical viscoelastic behavior combining both viscous fluid and elastic components. The higher the relative elastic part of maximum creep compliance (Je/Jmax), the higher the recovery capacity of sample (Sivaramakrishnan, Senge, & Chattopadhyay, 2004). In the creep recovery phase, approximately 48.4% elastic recovery could be seen for the NaCl treated sample. But for urea treated sample, the viscous part constituted 60.7%. Based on all the information in Figs. 4c and 4d, the trends were consistent with the results of previous work on oscillation properties (Fig. 4b) treated with urea and SDS. These results again indicated that waxy rice flour treated with urea and SDS might cause a weaker gel structure under current tested conditions and was more susceptible to food operations. Addition of L-Cys had minor effects on starch-protein leaching and water solubility of waxy rice flour on lower temperature (Fig. 1). However, it had notable effect on rheological properties. These might be explained that disulfide bond sensitivity toward reduction depends on their accessibility and it has been suggested that some disulfide bonds are inaccessible for reduction in raw rice (Morel, Redl, & Guilbert, 2002; Borght, Vandeputte, Derycke, Brijs, Daene, & 15
Delcour, 2006). Higher temperature and steady shear or flake might promote their accessibility on micro-environments. Addition of acid, alkali, and NaCl might act to give electrostatic forces, ion interactions, and indirectly change hydrogen bonds and hydrophobic interactions (Fig. 3), which played a synthetic role on property changes of waxy rice flour (Chuang et al., 2017; Sangpring et al., 2015; Sun, Liang, Yu, Tan, & Cui, 2016).
4. Conclusion
In summary, the main interaction characteristics in waxy rice flour involved hydrogen bonds, disulfide bonds, and hydrophobic interactions. Addition of urea led to a weakening in hydrogen bonds, thus dramatically influencing the intermolecular entanglement between starch and protein and remaining an unstable structure matrix. It suggested that the complex network structure was maintained mostly through hydrogen bonding and the binding interaction characteristics could be fully reflected by rheological property parameters. Hydrophobic regions internally linked protein complexes and the starch-protein aggregates were necessary for waxy rice flour to form a material with better viscoelastic properties. Unlike wheat gluten, viscoelastic properties of rice products were determined both by starch and protein. Thus, property changes of waxy rice flour must be influenced by covalent and non-covalent binding interactions between starch and protein, possibly relating to changes in composition type, contents and structures. This research would extend the way of producing pure rice starch (protein content < 0.5%) from a technological point of view and aid in the 16
future utilization of rice flour in food and other industrial applications.
Acknowledgments
This research was supported by National Key R&D Program of China [No. 2017YDF0401100]; National Natural Science Foundation of China [No. 31471616]; National
Top
Youth
Talent
for
Grain
Industry
[No.
LQ2016301];
Industry-Academia-Research Joint Innovation Fund of Jiangsu Province [No. BY2016022-30]; and Ingredion Incorporated (Shanghai, China) for its financial support to this research project.
Conflict interest
The authors declare no conflict of interest.
17
References
AACC International (2000). Approved methods of analysis (10th ed.). St. Paul, Minnesota: AACC International. Amagliani, L., O'Regan, J., Kelly, A.L., & O'Mahony, J.A. (2017). The composition, extraction, functionality and applications of rice proteins: a review. Trends in Food Science and Technology, 64, 1–12. Baxter, G., Blanchard, C., & Zhao, J. (2014). Effects of glutelin and globulin on the physicochemical properties of rice starch and flour. Journal of Cereal Science, 60, 414–420. Borght, A.V.D., Vandeputte, G.E., Derycke, V., Brijs, K., Daenen, G., & Delcour, J.A. (2006). Extractability and chromatographic separation of rice endosperm proteins. Journal of Cereal Science, 44, 68–74. Chan, H.T., Bhat, R., & Karim, A.A. (2010). Effects of sodium dodecyl sulphate and sonication treatment on physicochemical properties of starch. Food Chemistry, 120, 703–709. Chuang, L., Panyoyai, N., Shanks, R.A., & Kasapis, S. (2017). Effect of salt on the glass transition of condensed tapioca starch systems. Food Chemistry, 229, 120–126. Condés, M.C., Añón, M.C., Mauri, A.N., & Dufresne, A. (2015). Amaranth protein films reinforced with maize starch nanocrystals. Food Hydrocolloids, 47, 146–157. Cruz, L.G., Soto, J.L.M., Barajas, E.C., Rodríguez, M.X.N., Morales, A.F., & Nicanor, 18
A.B. (2018). Spectroscopic, calorimetric and structural analyses of the effects of hydrothermal treatment of rice beans and the extraction solvent on starch characteristics. International Journal of Biological Macromolecules, 107, 965-972. Debet, M.R., & Gidley, M.J. (2006). Three classes of starch granule swelling: influence of surface proteins and lipids. Carbohydrate Polymers, 64, 452–465. Derycke, V., Veraverbeke, W.S., Vandeputte, G.E., Man, D.W., Hoseney, R.C., & Delcour, J.A. (2005). Impact of protein on pasting and cooking properties of nonparbioled and parboiled rice. Cereal Chemistry, 82, 468–474. Fadda, E., & Woods, R.J. (2010). Molecular simulations of carbohydrates and protein-carbohydrate interactions: motivation, issues and prospects. Drug Discovery Today, 15, 596–609. Fan, D.M., Liu, Y.X., Hu, B., Lin, L.F., Huang, L.L., Wang, L.Y., Zhao, J.X., Zhang, H., & Chen, W. (2016). Influence of microwave parameters and water activity on radical generation in rice starch. Food Chemistry, 196, 34–41. Feng, X.W., Zhang, Q.H., Cong, P.S., & Zhu, Z.L. (2013). Preliminary study on classification of rice and detection of paraffin in the adulterated samples by Raman spectroscopy combined with multivariate analysis. Talanta, 115, 548–555. Jeong, S., Kim, H.W., & Lee, S. (2017). Rheological and secondary structural characterization of rice flour-zein composites for noodles slit from gluten-free sheeted dough. Food Chemistry, 221, 1539–1545. 19
Marco, C., & Rosell, C.M. (2008). Effect of different protein isolates and transglutaminase on rice flour properties. Journal of Food Engineering, 84, 132–139. Matos, M.E., Sanz, T., & Rosell, C.M. (2014). Establishing the function of proteins on the rheological and quality properties of rice based gluten free muffins. Food Hydrocolloids, 35, 150–158. Morel, M.H., Redl, A., & Guilbert, S. (2002). Mechanism of heat and shear mediated aggregation of wheat gluten protein upon mixing. Biomacromolecules, 3, 488–497. Niu, H.L., Zhang, M.C., Xia, X.F., Liu, Q., & Kong, B.H. (2018). Effect of porcine plasma protein hydrolysates on long-term retrogradation of corn starch. Food Chemistry, 239, 172–179. Niu, L.Y., Wu, L.Y., & Xiao, J.H. (2017). Inhibition of gelatinized rice starch retrogradation by rice bran protein hydrolysates. Carbohydrate Polymers, 175, 311–319. Renzetti, S., Behr, J., Vogel, R.F., Barbiroli, A., Iametti, S., Bonomi, F., & Arendt, E.K. (2012). Transglutaminase treatment of brown rice flour: a chromatographic, electrophoretic and spectroscopic study of protein modifications. Food Chemistry, 131, 1076–1085. Rożnowska, I.P., & Fortuna, T. (2017). Effect of conditions of modification on thermal and rheological properties of phosphorylated pumpkin starch. International Journal of Biological Macromolecules, 104, 339–344. 20
Sangpring, Y., Fukuoka, M., & Ratanasumawong, S. (2015). The effect of sodium chloride on microstructure, water migration, and texture of rice noodle. LWT-Food Science and Technology, 64, 1107–1113. Šárka, E., & Dvořáček, V. (2017). New processing and applications of waxy starch (a review). Journal of Food Engineering, 206, 77–87. Schober, T.J., Bean, S.R., Tilley, M., Smith, B.M., & Ioerger, B.P. (2011). Impact of different isolation procedures on the functionality of zein and kafirin. Journal of Cereal Science, 54, 241–249. Sivaramakrishnan, H.P., Senge, B., & Chattopadhyay, P.K. (2004). Rheological properties of rice dough for making rice bread. Journal of Food Engineering, 62, 37–45. Smith, B.M., Bean, S.R., Selling, G., Sessa, D., & Aramouni, F.M. (2014). Role of non-covalent interactions in the production of visco-elastic material from zein. Food Chemistry, 147, 230–238. Sun, N.X., Liang, Y., Yu, B., Tan, C.P., & Cui, B. (2016). Interaction of starch and casein. Food Hydrocolloids, 60, 572–579. Wang, F.C., & Sun, X.S. (2002). Creep-recovery of wheat flour doughs and relationship to other physical dough tests and breadmaking performance. Cereal Chemistry, 79, 567-571. Xia, N., Wang, J.M., Gong, Q., Yang, X.Q., Yin, S.W., & Qi, J.R. (2012). Characterization and in vitro digestibility of rice protein prepared by enzyme-assisted microfluidization: comparison to alkaline extraction. Journal of 21
Cereal Science, 56, 482–489. Xie, L., Chen, N., Duan, B., Zhu, Z., & Liao, X. (2008). Impact of proteins on pasting and cooking properties of waxy and non-waxy rice. Journal of Cereal Science, 47, 372–379. Xu, X.F., Liu, W., Liu, C.M., Luo, L.P., Chen, J., Luo, S.J., McClements, D.J., & Wu, L.X. (2016). Effect of limited enzymatic hydrolysis on structure and emulsifying properties of rice glutelin. Food Hydrocolloids, 61, 251–260.
22
Figure captions Fig.1. Effect of different treatments on chemical compositions (a) and water solubility (b) of waxy rice flour. Data are expressed as the means ±SD (n=3). The error bar represents the standard deviation. Values marked with different letters are significantly different at the same temperature (p < 0.05).
Fig.2. SDS-PAGE profiles of waxy rice flour with different treatments.
Fig.3. X-ray diffraction patterns (a) and Raman (b) with different treatments of waxy rice flour.
Fig.4. Pasting curves (a) and rheological [(b) tan δ (ratio of G''/G'); (c) steady flow; (d) creep-recovery] properties of different treatments of waxy rice flour.
23
Figure 1
24
Figure 2
25
Figure 3
26
Figure 4
27
Table 1 The approximate peak positions and their tentative assignments of waxy rice flour with different treatments. Approximate peak positions (cm−1)
Assignments
Ctrl
pH 5
pH 9
NaCl
SDS
Urea
L-Cys
1470 1388 1349 1270 1118 1061 1014 951 874 781 487
δ(CH2) twisting and C-H bending δ(C-H), C-H bending and C-H scissoring vibrations CH2–,δ(CH2) and C-OH bending a complex mode involving CH2OH side-chain. ν(C-O) and C-O-H deformation δ(C-OH) and γ(C-OH) ring-breathing vibration of phenylalanine molecules νs (C-O-C) α-1,4-glycosidic linkage νs(C-O-C) ring mode and C1-H bending α-configuration. ν(C-C) ring mode and C-C stretching (C-O-C) ring mode and δ(C-C-O)
1470 1388 1249 1270 1118 1061 1014 951 874 781 487
1470 1391 1350 1275 1120 1061 1014 951 879 781 487
1472 1391 1349 1275 1121 1062 -* 951 879 780 487
1469 1391 1349 1272 1120 1064 -* 951 879 780 487
1470 1391 1350 1275 -* 1064 -* 953 881 780 487
1473 1391 1350 1275 1120 1064 -* 951 881 780 487
1472 1391 1349 1272 1121 1062 -* 951 879 780 487
* designate not detectable.
28
Table 2 Parameters obtained from the creep-recovery curves of waxy rice flour with different treatments. Samples Ctrl pH 5 pH 9 NaCl SDS Urea L-Cys
Creep phase η0×10 (Pa·s) Jmax ×101 (1/Pa) 0.92 0.66 0.84 0.64 0.82 0.48 0.90 0.74 0.94 0.72 1.31 1.90 0.92 1.69 3
Recovery phase Je /Jmax (%) Jv /Jmax (%) 42.2 57.8 44.6 55.4 46.1 53.9 48.4 51.6 42.2 57.8 39.5 60.5 41.7 58.4
Jmax, the maximum creep compliance; η0, the zero shear viscosity; Je /Jmax, the relative elastic part of maximum creep compliance; Jv /Jmax, the relative viscous part of maximum creep compliance.
29
Highlights:
The main interactions involved hydrogen, disulfide, and hydrophobic bonds.
Both hydrogen and hydrophobic bonds were necessary for a viscoelastic structure.
Urea and SDS showed different effects on crystallinity and pasting properties.
This research would extend the way of producing pure rice starch in technological.
30
S0308-8146(18)30800-8 https://doi.org/10.1016/j.foodchem.2018.05.010 FOCH 22843
To appear in:
Food Chemistry
Received Date: Revised Date: Accepted Date:
2 January 2018 26 March 2018 2 May 2018
Please cite this article as: Li, Z., Wang, L., Chen, Z., Yu, Q., Feng, W., Impact of binding interaction characteristics on physicochemical, structural, and rheological properties of waxy rice flour, Food Chemistry (2018), doi: https:// doi.org/10.1016/j.foodchem.2018.05.010
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Impact of binding interaction characteristics on physicochemical, structural, and rheological properties of waxy rice flour Zhenni Lib, Li Wangab, Zhengxing Chenabc*, Qiusheng Yub, Wei Fengb a
State Key Laboratory of Food Science and Technology , Jiangnan University, Lihu Road 1800, Wuxi 214122, China
b
School of food science and Technology, National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, Lihu Road 1800, Wuxi 214122, China
c
Jiangsu Provincial Research Center for Bioactive Product Processing Technology, Jiangnan University, Lihu Road 1800, Wuxi 214122, China
*Corresponding author
Tel: +86-510-8519 7110; Fax: +86-510-8519 7110; E-mail: [email protected]
Abstract
Impact of covalent and non-covalent binding interactions on physicochemical, structural, and rheological properties of waxy rice flour were investigated using several typical types of chemicals. Dynamic rheometer was used to investigate the viscoelastic properties and resistance to deformation or shearing. Both hydrogen bonds and hydrophobic interactions were necessary for waxy rice flour to form a material with stable structure matrix. Besides this, X-ray diffraction pattern and pasting property measurements showed that the relative crystallinity and pasting viscosity values of waxy rice flour exposed to SDS and L-cysteine treatments increased whereas those of urea decreased compared with the control. Addition of acid, alkali, and NaCl might result in electrostatic forces, ion interactions, and indirectly change hydrogen bonds and hydrophobic interactions between starch and protein, which played a synthetic role on property changes of waxy rice flour.
Keywords: Waxy rice; Starch; Protein; Binding interaction characteristics;
1
1. Introduction
Rice (Oryza sativa L.) is the staple food for about 3.5 billion people worldwide. Starch is the main storage carbohydrate in plant organisms. Normal starch in granular form is generally composed of two types of molecules, amylose and amylopectin. Waxy starches consist almost exclusively of amylopectin, a high molecular weight molecule. Protein is the most abundant component in rice grain next to starch, and the interaction between protein and starch are important as they can have potential effects on processing and cooking properties of waxy rice and its products. In food systems, there are covalent and non-covalent interactions among compounds involving disulfide bonds, hydrogen bonds, hydrophobic interactions, electrostatic forces, ionic interactions, and so on. Recent advances in understanding these interaction modes further illustrate diverse aspects of their nature and importance in food systems. For starch gels, non-covalent chain interactions were generally believed to involve intermolecular double helix formation stabilized by hydrogen bonds (Šárka & Dvořáček, 2017). Decreasing in hydrogen bonds within starch led to a significant decreasing in relative crystallinity and retrogradation enthalpies of corn starch (Niu, Zhang, Xia, Liu, & Kong, 2018). Schober, Bean, Tilley, Smith, and Ioerger (2011) claimed that hydrophobic interactions rather than disulfide bonds were the key to gluten-like functionality of zein and kafirin. According to Renzetti et al. (2012), the formation of large protein complexes induced by transglutaminase treatment was ascribed to the new and stronger hydrophobic interactions among proteins. These complexes could promote a more viscoelastic 2
matrix and modify physicochemical properties of rice flour (Marco & Rosell, 2008). The interactions between starch and protein were mediated by a number of non-covalent interactions included site recognition with the aromatic side chain of Trp, Tyr or, less frequently, Phe of the protein. Binding of carbohydrates to protein also involved electrostatic interactions and H-bonding. Charged residues (Arg, Lys, Asp and Glu) were generally found in the binding sites in carbohydrate–protein (Fadda & Woods, 2010). Recognition and binding of carbohydrates to proteins always take place through the formation of complex H-bonding networks and involve charge–charge interactions. The objective of this paper was to explore the effects of binding interaction characteristics on physicochemical, structural and rheological properties of waxy rice flour system. The results reported were expected to provide significant clues for separating pure starch from protein in rice system and lay a foundation for illuminating property changes of other similar starch-protein enriched systems.
2. Materials and methods
2.1. Materials
One Oryza sativa Thai native cultivar of waxy rice was kindly provided by Ingredion Incorporated (Shanghai, China). Moisture, total starch and protein content of waxy rice were 12.5%, 77.4% and 6.3%, respectively, determined by AACC Approved Methods (AACC International, 2000). All other chemicals utilized were of analytical grade or higher. 3
2.2. Methods
2.2.1. Samples preparation Waxy rice was soaked in a 45 ◦C water bath for 1 hr. After soaking, waxy rice was milled under continuous addition of water and sieved through an 80-mesh sifter to obtain uniformity of particle size. The rice slurry and different chemicals with concentration of 2% (w/w) were mixed to achieve the required final concentration of 25% (w/w). A series of chemicals were added, including HCl and NaOH at pH of 5 and 9, NaCl, SDS, urea, and L-cysteine (L-Cys). The mixtures were reacted with agitation using a controlled-rotational speed mixing device (IKA® RW 20; Wilmington, NC, USA) at 25 °C for 4 hr. Afterwards, the mixtures were centrifuged at 4000 × g for 5 min, the supernatant was removed, and the precipitate (solid residue) was retained and rinsed with water three times for removal of redundant chemical solution. The remaining precipitate was oven dried at 40 ◦C, grounded and sieved through a 100-mesh sifter for further determinations. 2.2.2. Chemical composition and solubility determination Approved AACC Methods were used to determine starch content using a total starch assay kit (Megazyme International Ireland, Wicklow, Ireland) and protein content (N × 5.95) according to the Kjeldahl method of waxy rice flour samples (AACC International, 2000). Solubility properties were determined by the method of Derycke, Veraverbeke, Vandeputte, Man, Hoseney, and Delcour (2005) with some modifications. About 1.5 g flour and 25 mL deionized water were weighted in tube and shocked at room 4
temperature for 1 hr. This was then incubated at constant temperature (30 ◦C, 50 ◦C, 70 ◦C and 90 ◦C) at 160 rpm for 30 min in a water bath. Sample was cooled down to room temperature and centrifuged (4000 × g, 15 min). Water solubility (WS, g/100g) was calculated as follow: WS = dried supernatant weight / dry flour weight × 100. 2.2.3. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) SDS-PAGE (12% separating gel and 4% stacking gel) was performed on a discontinuous buffered system according to the method of Xia, Wang, Gong, Yang, Yin, and Qi (2012). About 1.5 mg waxy rice flour sample was dissolved in 100 μL of sample buffer (0.125 mol/L Tris-HCl buffer, 1% SDS (w/v), 20% glycerol (v/v), 2% 2-mercaptoethanol (2-ME, v/v), pH 6.8), and heated at 95 °C for 5 min. Gels were stained with Coomassie Brilliant Blue R-250 (0.1%) in methanol/acetic acid/water (45:10:45, v/v/v) and de-stained in methanol-water solution containing 10% acetic acid (methanol/acetic acid/water =1:1:8, v/v/v). 2.2.4. X-ray diffractometry X-ray diffraction patterns of samples were obtained using an X-ray diffractometer (D2 Phaser, Bruker-AXS, Germany). The scanning region of the diffraction angle (2θ) ranged from 3 ◦ to 40 ◦ at a step size of 0.02 ◦ with a count time of 0.6 s. Relative crystallinity (%) was calculated as the percentage ratio of the sum of total crystalline peak areas to that of the total diffractogram (sum of total crystalline and amorphous peak areas) by software (MDI Jade 6). 2.2.5. Raman spectroscopy All spectra of the samples were collected using the LabRAM HR Evolution 5
system (HORIBA Jobin Yvon S.A.S., Longjumeau Cedex, France) with a 532 nm laser source, motorized microscope and a cooled charge-coupled device (CCD) detector. The spectral data were collected in the range of 400-1800 cm-1 at a resolution of 0.80 cm-1 with constant measurement parameters (acquisition time: 10 s; laser power: 100 mW). As the spectrophotometer was sensitive to the change of outer environment condition, the spectra were collected at controlled temperature (25 °C) and humidity (60%). The software package LabSpec 6 (HORIBA Jobin Yvon S.A.S., Longjumeau Cedex, France) was used for spectral data acquisition. 2.2.6. Pasting properties Pasting properties were determined using a Rapid Visco-Analyzer (Perten RVA 4500, Australia), as reported previously (Jeong, Kim, & Lee, 2017). Each sample (3 g of flour at 14% moisture basis) was directly weighed into an aluminum RVA canister. Then, 25 mL of distilled water was added to achieve a total weight of 28 g. The formed slurry sample was heated to 50 °C and stirred at 960 rpm for 10 s and fixed at 160 rpm throughout rest of the run. Then, sample was held at 50 °C for 0.5 min, and then heated up to 95 °C for 3.5 min and held for 2.5 min, and cooled down to 50 °C for 3.5 min and held for 2 min. Parameters recorded were peak, trough, final viscosity, breakdown, setback, pasting temperature and pasting time. 2.2.7. Rheological measurements Rheological measurements were conducted using a DHR-3 rheometer (TA Instrument, USA) as reported previously (Matos, Sanz, & Rosell, 2014). Waxy rice flour dispersions (10% w/w) were moderately stirred and heated in a boiling water 6
bath for 10 min by a magnetic stirrer until fully gelatinized. The hot pastes obtained were immediately cooled down at 30 °C for 10 min and measurements were measured at 30 °C using 40 mm diameter plate. Oscillatory frequency test from 0.1 to 10 Hz was performed in the linear viscoelastic region at a target strain of 1%, and storage modulus (G'), loss modulus (G''), and tan δ (G''/ G') versus frequency values were recorded. Steady shear flow measurement was carried out at shear rate lineally from 0 s-1 to 300 s-1. Shear stress (τ) was recorded as a function of shear rate. Creep and recovery measurements were carried out as follows: The creep phase was recorded at a shear stress of 10 Pa for 60 s, followed by a recovery phase of 120 s at a stress of 0 Pa. Creep and recovery curves were recorded and analyzed by RHEOPLUS/32 version 3.21 software to obtain the parameters, including the maximum creep compliance (Jmax), zero shear viscosity (η0), relative elastic part of the maximum creep compliance (Je/Jmax), and the relative viscous part of the maximum creep compliance (Jv/Jmax). 2.2.8. Statistical analysis All tests were performed in triplicate unless specified. Data were evaluated by analysis of variance (ANOVA), and a comparison of means were carried out with Tukey’s HSD test. Differences were considered to be significant at p < 0.05. Statistical computations and analyses were conducted using SPSS software (version 19.0).
7
3. Results and Discussion
3.1. Physicochemical properties
3.1.1. Chemical composition The starch and protein content of waxy rice flour with different treatments were reported in Fig. 1a. Treatments with urea and NaCl had pronounced effect on starch and protein content of waxy rice flour. Addition of urea led to a weakening in hydrogen bonds, thus promoting starch and protein leaching into solution and decreasing by 2% and 9% compared with that of the control. The addition of NaCl had positive effect on the starch content (91%), which might be because NaCl weakened the electrostatic attraction between the protein and polysaccharide molecules and thereby decreasing the protein content in systems. Besides this, it was reported that increasing the salt level enhanced the mechanical strength of starch through direct electrostatic interactions with the hydroxyl groups of starch molecules (Chuang, Panyoyai, Shanks, & Kasapis, 2017). The control sample exhibited the highest protein content (7.0%) and the content in decreasing order of other treatments was pH 5, pH 9, NaCl, L-Cys, urea and SDS. The content decreased by values of 2-23% compared with protein content in control. The protein extractabilities of all treated samples increased even more, indicating that covalent disulfide bonds and non-covalent interactions were involved in the formation of the protein network of waxy rice. SDS-PAGE of waxy rice flour samples were measured to investigate changes of polypeptide profiles, as shown in Fig. 2. The major polypeptide glutelin (19–25 and 8
30–39 kDa for basic and acidic subunits respectively) and 49–62 kDa for pro-glutelin were identified. The 10–16 kDa fraction corresponded to prolamin with the 13 kDa being predominant, which was known to be an integral part of rice protein. Two rice endosperm globulin polypeptides, having MW of 16 kDa and 25 kDa, had been identified as reported previously (Amagliani, O'Regan, Kelly, & O'Mahony, 2017).The profiles of waxy rice flour exposed to pH 5 and pH 9 were similar compared with the control. Upon NaCl and SDS treatments, the band at 23 kDa disappeared. Waxy rice flour treated with SDS and urea exhibited a decrease of higher molecular weight marks (> 75 kDa), indicating the disruption of bigger peptides and release of smaller peptides into solution. Apart from other treatments, high molecular weight (>75 kDa) aggregates of sample with addition of L-Cys were not observed in separated bands, which suggested that L-Cys could interact with waxy rice flour proteins by splitting the disulfide bonds and decrease the content of large macromolecular complexes (Xu et al., 2016). 3.1.2. Solubility The water solubility of waxy rice flour with different treatments at 30-90 °C were shown in Fig. 1b. Values of all samples increased dramatically with increasing temperature and the solubility of control was increased from 2.2% to 41.7% with temperature increased from 30 °C to 90 °C. Addition of urea showed the highest effect on solubility of waxy rice flour at any test temperature. Addition of SDS, which disrupted protein structure mainly affecting hydrophobic interactions, showed the most significant changes in water solubility of waxy rice flour next to that of treated 9
with urea. This suggested that non-covalent, particularly hydrogen bonds type and hydrophobic interactions seemed to be the most important in stabilizing the waxy rice flour matrix. This was consistent with previous studies which had investigated the interaction between maize starch and amaranth protein (Condés, Añón, Mauri, & Dufresne, 2015). Solubility of samples with pH, NaCl and L-Cys treatments changed more markedly and more rapidly under higher temperatures.
3.2. Structural properties
3.2.1. X-ray diffraction pattern and crystallinity The X-ray diffraction patterns of waxy rice flour with different treatments were presented in Fig. 3a. All samples displayed the typical A-type diffraction patterns with strong diffraction peaks at about 15 ° and 23 ° (2θ), and an unresolved doublet at around 17 ° and 18 ° (2θ). The relative crystallinity of waxy rice flour exposed to pH 5, pH 9, SDS, and L-Cys treatments increased whereas those of urea, NaCl decreased compared with that of the control. According to Sangpring, Fukuoka, and Ratanasumawong (2015), the formation of hydrogen bonds and starch retrogradation were limited with NaCl addition, because of co-action of electrostatic repulsion and steric hindrance effect. They explained that the H+ ions of the starch hydroxyl group migrate from starch molecules into water, leading to the Na+ ions being pushed and entrapped by the starch molecules. Since the radius of Na+ ions is greater than for H+ ions, the formation of hydrogen bonds and starch retro-gradation are limited. Renzetti et al. (2012) suggested that polymerization through disulfide bonds promotes 10
hydrophobic interactions among proteins. The reducing agent L-Cys broke disulfide bonds and more hydrophobic groups were separated. After the protein matrix was disrupted, starch granules were exposed and had more opportunity to interact with each other, thus increasing the relative crystallinity of waxy rice flour. 3.2.2. Raman spectra analysis Raman spectra of waxy rice flour samples with different treatments in the 400-1800 cm-1 region was presented in Fig. 3b. In general, the characteristics of Raman bands were observed with some changes of the bands’ positions and intensities after treatments. Each Raman curve of sample was classified and its various spectral components were associated with vibrations, rotations, etc., of the chemical bonds in the waxy rice flour’s constituent components. Changes in the relative intensity, position, and width of these bands were related to the composition (such as starch, protein, and lipids) of the sample. The main Raman bands and their preliminary assignments of waxy rice flour were listed in Table 1. Observed from Raman scans, the most obvious changes of positions and intensities happed to the bands at 874, 951, 1061, 1118, 1270, and 1349 cm-1. According to Fan et al. (2016), it might be caused by the changes in νs(C-O-C) ring mode, νs(C-O-C) α-1, 4-glycosidic linkage, C1-H bending α-configuration and changes in hydrogen bond network involving CH2OH side-chain and C-OH groups. The peak 1014 cm-1 was attributed to ring-breathing vibration of phenylalanine molecules and was observed diminishing distinguishingly with treatments except pH 5. The phenomenon indicated the treatments except pH 5 had a serious impact on certain 11
vibration of side chain of proteins (Feng, Zhang, Cong, & Zhu, 2013). Regarding characteristic bands of the starch, the bands around 781 cm-1 and 487 cm-1 were assigned to ν(C-C) ring mode and skeletal mode of pyranoid ring, respectively. For all treatments with waxy rice flour, a marked decrease and increase in the area of the band at 781 cm-1 and 487 cm-1 were observed respectively, indicating a change in the molecular order or crystallinity of the starch (Cruz, Soto, Barajas, Rodríguez, Morales, & Nicanor, 2017).
3.3. Pasting properties
Determining the pasting properties of waxy rice flour is important not only for actual applications during food processing, but also for understanding the structural changes with different treatments (Baxter, Blanchard, & Zhao, 2014). The pasting behaviors of waxy rice flour slurry were presented in Fig. 4a and different treatments showed various significant influences on pasting properties of waxy rice flour. Some treatments (NaCl and SDS) had positive effect on viscosity values and the viscosity values of waxy rice flour treated with SDS were significantly higher than that of the control (ranging from 8% to 12%). This could be due to the effective removal of protein layers that help to increase the swelling power of waxy rice flour (Chan, Bhat, & Karim, 2010). It was consistent with the result of Debet and Gidley (2006), wherein a big increase in peak viscosity was observed after SDS treatment. On the contrary, some other treatments (urea, L-Cys, and pH) had negative effects and the addition of urea significantly decreased the peak, trough, and final viscosity values of curve by 12
15%, 9%, and 16%, respectively, compared with that of the control. This result indicated that urea might block the formation of hydrogen bonds to impact intra- and inter-particle interactions of systems and resulting in lower viscosities (Niu, Wu, & Xiao, 2017). Setback viscosity values represented the differences between trough viscosity and final viscosity, and it was mainly resulted from the rearrangements between amylose and amylopectin in the system when temperature decreased from 95 to 50 °C. The setback values were decreased among all treatments by from 5% to 39%, which suggested that the transformation of the nuclei might change the environment of the starch molecules and influence the short-term rearrangements of the sample microstructure under higher water content.
3.4. Rheometer measurements
3.4.1. Oscillation All tested waxy rice flour samples behaved like a soft gel with G' > G'' and with tan δ (ratio of G''/G') < 1 (Fig. 4b) at all studied frequencies. In general, samples with addition of urea and L-Cys produced higher tanδ values than control, showing that their viscous properties were more pronounced than the elastic properties. The result agreed with the water solubility of waxy rice flour treated with urea reported previously (Fig. 1b). Blocking in hydrogen bonds promoted starch and protein leaching into solution, which might indicate a weak gel matrix of waxy rice flour with addition of urea. L-Cys and SDS disrupted protein network and decreased the viscoelasticity of waxy rice flour, indicating that disulfide bonds and hydrophobic 13
regions of internally linked protein complexes were necessary for waxy rice flour to form a material with better viscoelastic properties (Xie, Chen, Duan, Zhu, & Liao, 2008). 3.4.2. Steady flow All of the examined samples exhibited a non-Newtonian flow and were shear thinning with a tendency towards shear stress under the current experimental conditions, as observed in Fig. 4c. Higher value of the shear stress meant that the tested sample was more resistant to deformation and a more stable shear structure in terms of speed (Rożnowska & Fortuna, 2017). Among samples, a lowest value of shear stress was determined in the sample reacted with urea, followed successively by L-Cys, SDS, pH 9, Ctrl, NaCl, and pH 5. The results indicated that covalent (disulfide bonds) and non-covalent (hydrogen bonds and hydrophobic interaction) binding interactions intensified the intermolecular entanglement of waxy rice flour system, which had a significant impact on shearing sensitivity of flour paste (Smith, Bean, Selling, Sessa, & Aramouni, 2014). 3.4.3. Creep-recovery The results of creep and recovery phase curves and their analysis of all treatments were given in Fig. 4d and Table 2. Wang and Sun (2002) suggested that the maximum creep compliance (Jmax) could be used to characterize the rigidity (firmness) of dough samples. They reported that stronger dough samples which had greater resistance to deformation had smaller creep compliance than softer dough samples. The sample treated with urea exhibited the lowest deformation resistance, and the 14
decrease in Jmax values followed the order: L-Cys < NaCl < SDS < Ctrl. As seen in Table 2, the zero shear viscosity (η0) gave the flowability of samples at the end of applied stress (10 Pa) and η0 values of SDS and urea were higher than the others. This suggested that it might be difficult for particles to attract each other and to maintain their shape of waxy rice flour pastes, which resulted in higher flowability. This trend was consistent with the result of the solubility on sample treated with SDS and urea. The creep-recovery curves also exhibited a typical viscoelastic behavior combining both viscous fluid and elastic components. The higher the relative elastic part of maximum creep compliance (Je/Jmax), the higher the recovery capacity of sample (Sivaramakrishnan, Senge, & Chattopadhyay, 2004). In the creep recovery phase, approximately 48.4% elastic recovery could be seen for the NaCl treated sample. But for urea treated sample, the viscous part constituted 60.7%. Based on all the information in Figs. 4c and 4d, the trends were consistent with the results of previous work on oscillation properties (Fig. 4b) treated with urea and SDS. These results again indicated that waxy rice flour treated with urea and SDS might cause a weaker gel structure under current tested conditions and was more susceptible to food operations. Addition of L-Cys had minor effects on starch-protein leaching and water solubility of waxy rice flour on lower temperature (Fig. 1). However, it had notable effect on rheological properties. These might be explained that disulfide bond sensitivity toward reduction depends on their accessibility and it has been suggested that some disulfide bonds are inaccessible for reduction in raw rice (Morel, Redl, & Guilbert, 2002; Borght, Vandeputte, Derycke, Brijs, Daene, & 15
Delcour, 2006). Higher temperature and steady shear or flake might promote their accessibility on micro-environments. Addition of acid, alkali, and NaCl might act to give electrostatic forces, ion interactions, and indirectly change hydrogen bonds and hydrophobic interactions (Fig. 3), which played a synthetic role on property changes of waxy rice flour (Chuang et al., 2017; Sangpring et al., 2015; Sun, Liang, Yu, Tan, & Cui, 2016).
4. Conclusion
In summary, the main interaction characteristics in waxy rice flour involved hydrogen bonds, disulfide bonds, and hydrophobic interactions. Addition of urea led to a weakening in hydrogen bonds, thus dramatically influencing the intermolecular entanglement between starch and protein and remaining an unstable structure matrix. It suggested that the complex network structure was maintained mostly through hydrogen bonding and the binding interaction characteristics could be fully reflected by rheological property parameters. Hydrophobic regions internally linked protein complexes and the starch-protein aggregates were necessary for waxy rice flour to form a material with better viscoelastic properties. Unlike wheat gluten, viscoelastic properties of rice products were determined both by starch and protein. Thus, property changes of waxy rice flour must be influenced by covalent and non-covalent binding interactions between starch and protein, possibly relating to changes in composition type, contents and structures. This research would extend the way of producing pure rice starch (protein content < 0.5%) from a technological point of view and aid in the 16
future utilization of rice flour in food and other industrial applications.
Acknowledgments
This research was supported by National Key R&D Program of China [No. 2017YDF0401100]; National Natural Science Foundation of China [No. 31471616]; National
Top
Youth
Talent
for
Grain
Industry
[No.
LQ2016301];
Industry-Academia-Research Joint Innovation Fund of Jiangsu Province [No. BY2016022-30]; and Ingredion Incorporated (Shanghai, China) for its financial support to this research project.
Conflict interest
The authors declare no conflict of interest.
17
References
AACC International (2000). Approved methods of analysis (10th ed.). St. Paul, Minnesota: AACC International. Amagliani, L., O'Regan, J., Kelly, A.L., & O'Mahony, J.A. (2017). The composition, extraction, functionality and applications of rice proteins: a review. Trends in Food Science and Technology, 64, 1–12. Baxter, G., Blanchard, C., & Zhao, J. (2014). Effects of glutelin and globulin on the physicochemical properties of rice starch and flour. Journal of Cereal Science, 60, 414–420. Borght, A.V.D., Vandeputte, G.E., Derycke, V., Brijs, K., Daenen, G., & Delcour, J.A. (2006). Extractability and chromatographic separation of rice endosperm proteins. Journal of Cereal Science, 44, 68–74. Chan, H.T., Bhat, R., & Karim, A.A. (2010). Effects of sodium dodecyl sulphate and sonication treatment on physicochemical properties of starch. Food Chemistry, 120, 703–709. Chuang, L., Panyoyai, N., Shanks, R.A., & Kasapis, S. (2017). Effect of salt on the glass transition of condensed tapioca starch systems. Food Chemistry, 229, 120–126. Condés, M.C., Añón, M.C., Mauri, A.N., & Dufresne, A. (2015). Amaranth protein films reinforced with maize starch nanocrystals. Food Hydrocolloids, 47, 146–157. Cruz, L.G., Soto, J.L.M., Barajas, E.C., Rodríguez, M.X.N., Morales, A.F., & Nicanor, 18
A.B. (2018). Spectroscopic, calorimetric and structural analyses of the effects of hydrothermal treatment of rice beans and the extraction solvent on starch characteristics. International Journal of Biological Macromolecules, 107, 965-972. Debet, M.R., & Gidley, M.J. (2006). Three classes of starch granule swelling: influence of surface proteins and lipids. Carbohydrate Polymers, 64, 452–465. Derycke, V., Veraverbeke, W.S., Vandeputte, G.E., Man, D.W., Hoseney, R.C., & Delcour, J.A. (2005). Impact of protein on pasting and cooking properties of nonparbioled and parboiled rice. Cereal Chemistry, 82, 468–474. Fadda, E., & Woods, R.J. (2010). Molecular simulations of carbohydrates and protein-carbohydrate interactions: motivation, issues and prospects. Drug Discovery Today, 15, 596–609. Fan, D.M., Liu, Y.X., Hu, B., Lin, L.F., Huang, L.L., Wang, L.Y., Zhao, J.X., Zhang, H., & Chen, W. (2016). Influence of microwave parameters and water activity on radical generation in rice starch. Food Chemistry, 196, 34–41. Feng, X.W., Zhang, Q.H., Cong, P.S., & Zhu, Z.L. (2013). Preliminary study on classification of rice and detection of paraffin in the adulterated samples by Raman spectroscopy combined with multivariate analysis. Talanta, 115, 548–555. Jeong, S., Kim, H.W., & Lee, S. (2017). Rheological and secondary structural characterization of rice flour-zein composites for noodles slit from gluten-free sheeted dough. Food Chemistry, 221, 1539–1545. 19
Marco, C., & Rosell, C.M. (2008). Effect of different protein isolates and transglutaminase on rice flour properties. Journal of Food Engineering, 84, 132–139. Matos, M.E., Sanz, T., & Rosell, C.M. (2014). Establishing the function of proteins on the rheological and quality properties of rice based gluten free muffins. Food Hydrocolloids, 35, 150–158. Morel, M.H., Redl, A., & Guilbert, S. (2002). Mechanism of heat and shear mediated aggregation of wheat gluten protein upon mixing. Biomacromolecules, 3, 488–497. Niu, H.L., Zhang, M.C., Xia, X.F., Liu, Q., & Kong, B.H. (2018). Effect of porcine plasma protein hydrolysates on long-term retrogradation of corn starch. Food Chemistry, 239, 172–179. Niu, L.Y., Wu, L.Y., & Xiao, J.H. (2017). Inhibition of gelatinized rice starch retrogradation by rice bran protein hydrolysates. Carbohydrate Polymers, 175, 311–319. Renzetti, S., Behr, J., Vogel, R.F., Barbiroli, A., Iametti, S., Bonomi, F., & Arendt, E.K. (2012). Transglutaminase treatment of brown rice flour: a chromatographic, electrophoretic and spectroscopic study of protein modifications. Food Chemistry, 131, 1076–1085. Rożnowska, I.P., & Fortuna, T. (2017). Effect of conditions of modification on thermal and rheological properties of phosphorylated pumpkin starch. International Journal of Biological Macromolecules, 104, 339–344. 20
Sangpring, Y., Fukuoka, M., & Ratanasumawong, S. (2015). The effect of sodium chloride on microstructure, water migration, and texture of rice noodle. LWT-Food Science and Technology, 64, 1107–1113. Šárka, E., & Dvořáček, V. (2017). New processing and applications of waxy starch (a review). Journal of Food Engineering, 206, 77–87. Schober, T.J., Bean, S.R., Tilley, M., Smith, B.M., & Ioerger, B.P. (2011). Impact of different isolation procedures on the functionality of zein and kafirin. Journal of Cereal Science, 54, 241–249. Sivaramakrishnan, H.P., Senge, B., & Chattopadhyay, P.K. (2004). Rheological properties of rice dough for making rice bread. Journal of Food Engineering, 62, 37–45. Smith, B.M., Bean, S.R., Selling, G., Sessa, D., & Aramouni, F.M. (2014). Role of non-covalent interactions in the production of visco-elastic material from zein. Food Chemistry, 147, 230–238. Sun, N.X., Liang, Y., Yu, B., Tan, C.P., & Cui, B. (2016). Interaction of starch and casein. Food Hydrocolloids, 60, 572–579. Wang, F.C., & Sun, X.S. (2002). Creep-recovery of wheat flour doughs and relationship to other physical dough tests and breadmaking performance. Cereal Chemistry, 79, 567-571. Xia, N., Wang, J.M., Gong, Q., Yang, X.Q., Yin, S.W., & Qi, J.R. (2012). Characterization and in vitro digestibility of rice protein prepared by enzyme-assisted microfluidization: comparison to alkaline extraction. Journal of 21
Cereal Science, 56, 482–489. Xie, L., Chen, N., Duan, B., Zhu, Z., & Liao, X. (2008). Impact of proteins on pasting and cooking properties of waxy and non-waxy rice. Journal of Cereal Science, 47, 372–379. Xu, X.F., Liu, W., Liu, C.M., Luo, L.P., Chen, J., Luo, S.J., McClements, D.J., & Wu, L.X. (2016). Effect of limited enzymatic hydrolysis on structure and emulsifying properties of rice glutelin. Food Hydrocolloids, 61, 251–260.
22
Figure captions Fig.1. Effect of different treatments on chemical compositions (a) and water solubility (b) of waxy rice flour. Data are expressed as the means ±SD (n=3). The error bar represents the standard deviation. Values marked with different letters are significantly different at the same temperature (p < 0.05).
Fig.2. SDS-PAGE profiles of waxy rice flour with different treatments.
Fig.3. X-ray diffraction patterns (a) and Raman (b) with different treatments of waxy rice flour.
Fig.4. Pasting curves (a) and rheological [(b) tan δ (ratio of G''/G'); (c) steady flow; (d) creep-recovery] properties of different treatments of waxy rice flour.
23
Figure 1
24
Figure 2
25
Figure 3
26
Figure 4
27
Table 1 The approximate peak positions and their tentative assignments of waxy rice flour with different treatments. Approximate peak positions (cm−1)
Assignments
Ctrl
pH 5
pH 9
NaCl
SDS
Urea
L-Cys
1470 1388 1349 1270 1118 1061 1014 951 874 781 487
δ(CH2) twisting and C-H bending δ(C-H), C-H bending and C-H scissoring vibrations CH2–,δ(CH2) and C-OH bending a complex mode involving CH2OH side-chain. ν(C-O) and C-O-H deformation δ(C-OH) and γ(C-OH) ring-breathing vibration of phenylalanine molecules νs (C-O-C) α-1,4-glycosidic linkage νs(C-O-C) ring mode and C1-H bending α-configuration. ν(C-C) ring mode and C-C stretching (C-O-C) ring mode and δ(C-C-O)
1470 1388 1249 1270 1118 1061 1014 951 874 781 487
1470 1391 1350 1275 1120 1061 1014 951 879 781 487
1472 1391 1349 1275 1121 1062 -* 951 879 780 487
1469 1391 1349 1272 1120 1064 -* 951 879 780 487
1470 1391 1350 1275 -* 1064 -* 953 881 780 487
1473 1391 1350 1275 1120 1064 -* 951 881 780 487
1472 1391 1349 1272 1121 1062 -* 951 879 780 487
* designate not detectable.
28
Table 2 Parameters obtained from the creep-recovery curves of waxy rice flour with different treatments. Samples Ctrl pH 5 pH 9 NaCl SDS Urea L-Cys
Creep phase η0×10 (Pa·s) Jmax ×101 (1/Pa) 0.92 0.66 0.84 0.64 0.82 0.48 0.90 0.74 0.94 0.72 1.31 1.90 0.92 1.69 3
Recovery phase Je /Jmax (%) Jv /Jmax (%) 42.2 57.8 44.6 55.4 46.1 53.9 48.4 51.6 42.2 57.8 39.5 60.5 41.7 58.4
Jmax, the maximum creep compliance; η0, the zero shear viscosity; Je /Jmax, the relative elastic part of maximum creep compliance; Jv /Jmax, the relative viscous part of maximum creep compliance.
29
Highlights:
The main interactions involved hydrogen, disulfide, and hydrophobic bonds.
Both hydrogen and hydrophobic bonds were necessary for a viscoelastic structure.
Urea and SDS showed different effects on crystallinity and pasting properties.
This research would extend the way of producing pure rice starch in technological.
30