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Effects of wheat tempering with slightly acidic electrolyzed water on the microbial, biological, and chemical characteristics of different flour streams
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Effects of wheat tempering with slightly acidic electrolyzed water on the microbial, biological, and chemical characteristics of different flour streams Yun-Xia Chena,b, Xiao-Na Guoa,b, Jun-Jie Xinga,b, Xiao-Hong Sunc, Ke-Xue Zhua,b,∗ a
State Key Laboratory of Food Science and Technology, Jiangnan University, 1800 Lihu Avenue, Wuxi, 214122, Jiangsu Province, PR China School of Food Science and Technology, Jiangnan University, 1800 Lihu Avenue, Wuxi, 214122, Jiangsu Province, PR China c College of Food and Biological Engineering, Qiqihar University, Qiqihar, 161006, Heilongjiang Province, PR China b
ARTICLE INFO
ABSTRACT
Keywords: Tempering with SAEW Clear flour Bran Microbial control Quality properties
Wheat tempering was conducted with slightly acidic electrolyzed water (SAEW) at different active chlorine contents. Microbial, biological, and chemical properties of different flour streams milled from SAEW-tempered wheat grains were investigated. Results showed that the total plate count (TPC) and yeast/mould count (YMC) in flour products were significantly (p < 0.05) reduced as SAEW concentration increased. The total polyphenols, lipase (LA) activity, polyphenol oxidase (PPO) activity of clear flour and bran, along with starch viscosity and protein extractability under reduced conditions of clear flour were all decreased significantly (p < 0.05) when SAEW concentration raised. Meanwhile, farinograph properties of clear flour were slightly improved. However, there was no significant (p > 0.05) difference in biological and chemical characteristics of straight flour. Based on our findings, SAEW has potential to function as a novel tempering solution for producing clean flour.
1. Introduction Wheat grains are easily contaminated by microbes during growth, harvest, transportation, and storage (Li, Li, Luo, & Yoshizawa, 2002). The contaminated microorganisms in wheat grains can easily grow during the followed processing procedures, especially in tempering process due to the factors of long time, appropriate temperature and proper humidity (Berghofer, Hocking, Miskelly, & Jansson, 2003). The living microorganisms in flour can remain in a dormant state for a long period since flour generally has a low water activity (Eglezos, 2010; Rose, Bianchini, Martinez, & Flores, 2012). However, when flour is made into food and water is added, the microorganisms will proliferate immediately and lead to the degeneration of end products (Li et al., 2013). Thus, flour with high microbial load is considered as one of the major reasons for shortening the shelf-life of flour-based products, especially for fresh-wet noodle products. Meanwhile, some of heat-resistant spores, such as Bacillus spp., cannot be killed during production, and moulds can form allergenic spores and even produce mycotoxins (Schnürer, Olsson, & Börjesson, 1999). Therefore, microbial contamination within wheat grains is one of the biggest food safety issues of flour and flour-derived products, which urgently needs attention. To reduce the microbial load in wheat flour, some researchers replaced tempering water with ozonated water (İbanoǧlu, 2001), chlorinated water (Dhillon, Sandhu, Wiesenborn, Manthey, & Wolf-Hall, ∗
2007), organic acids or saline solutions (Sabillón, Stratton, Rose, Flores, & Bianchini, 2016). However, many problems limited the applications of these methods, such as instability, low efficacy, chemical residuals, and pipeline corrosion. SAEW as a novel antimicrobial agent has recently attracted lots of attention for its advantages. SAEW has been certified as a food additive by Japan Ministry of Health, Labour and Welfare (MHLW) and United States Food and Drug Administration (FDA). It can be generated by electrolyzing dilute NaCl and/or HCl solution in a non-membrane electrolytic chamber, resulting in a high concentration of hypochlorous acid with pH of 5.0–6.5 (Jadeja, Hung, & Bosilevac, 2013). The strong germicidal effect of SAEW is due to the available chlorine, including hypochlorite ion, hypochloric acid, and chlorine (Zheng et al., 2012). Compared with other bactericides, SAEW is more environment-friendly, non-toxic, non-residual, non-corrosive to organic matters, more stable and less expensive. In recent years, SAEW has been widely applied to extend shelf life of chicken meat (Rahman, Park, Song, Al-Harbi, & Oh, 2012), wash vegetables (Zhang, Cao, Hung, & Li, 2016) and clean shell eggs (Cao, Zhu, Shi, Wang, & Li, 2009). However, few research has been conducted on the applications of SAEW on wheat processing. The present study was designed to evaluate the disinfection efficacy of SAEW at different concentrations and its impacts on the microbial, biological, and chemical characteristics of different flour streams. At the same time, this paper also aimed to introduce a new concept of
Corresponding author. State Key Laboratory of Food Science and Technology, Jiangnan University, 1800 Lihu Avenue, Wuxi, 214122, Jiangsu Province, PR China. E-mail address: [email protected] (K.-X. Zhu).
https://doi.org/10.1016/j.lwt.2019.108790 Received 23 April 2019; Received in revised form 20 August 2019; Accepted 27 October 2019 0023-6438/ © 2019 Elsevier Ltd. All rights reserved.
Please cite this article as: Yun-Xia Chen, et al., LWT - Food Science and Technology, https://doi.org/10.1016/j.lwt.2019.108790
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production of “low-bacteria flour” that is mainly used to make freshwet noodle products.
2.7. Determination of activities of polyphenol oxidase (PPO), lipase (LA) and lipoxidase (LOX) in bran and clear flour The PPO activity was determined according to the method of Li et al. (2012) with some modifications. 1.0 g of sample was mixed into 10 mL extraction buffer (0.1 M phosphate buffer, pH 6.0) and then shaken under 4 °C for 24 h. The mixture was centrifuged at 10,000 rpm under 4 °C for 20 min. The test mixture was composed of 2 mL of the extract and 200 μL of catechol solution. One PPO activity unit was calculated as an increase in absorbance of 0.001 at 420 nm within 1 min. The LA activity was evaluated as described by Cai et al. (2011). 50 mM Tris-HCl buffer (10 mL, pH 8.0) was for the extraction of LA from 2.0 g of sample. 200 μL of the extract was reacted with 20 μL of 10 mM p-nitrophenyl octanoate in 1.78 mL of Tris-HCl buffer at 37 °C for 1 min. One LA activity unit was defined as an increase in absorbance of 0.1 at 405 nm within 1 min. The LOX activity was determined following the method of Cato, Halmos, and Small (2006). 2.0 g of sample was incubated in 10 mL of 0.1 M phosphate buffer (pH 7.5) and stirred on ice for 30 min. The mixture was then centrifuged at 8000 rpm at 4 °C for 10 min. The reaction mixture consisted of 50 mM sodium acetate buffer (2.89 mL, pH 5.5), linoleic acid substrate (90 μL), and LOX extract (20 μL). One LOX activity unit was defined as an increase in absorbance of 0.01 at 234 nm within 1 min.
2. Materials and methods 2.1. Wheat samples Wheat grains (Xinong-979, a kind of Henan hard white wheat grains) were obtained from China Oil & Foodstuffs Corporation in 2017 and were cleaned by sieves and hand. 2.2. Slightly acidic electrolyzed water (SAEW) SAEW was prepared with a hyper optimized chlorinated liquid device (HOCL 0.2 t pro, Meiloon Industrial Company Ltd., Taiwan, China). The active chlorine content (ACC) of prepared SAEW was 70 mg/L. SAEM with different ACC was obtained by mixing original SAEW with distilled water in various proportions and was used immediately after preparation. 2.3. Wheat tempering 500 g of cleaned wheat grains were placed in a sterile plastic bag and were tempered to 16% moisture with distilled water (control) and SAEW with different ACC (10, 30, 50, and 70 mg/L). Samples were shaken vigorously every 15 min during the first 2 h and were equilibrated for 24 h under 25 °C.
2.8. Analysis of rheological characteristics of straight flour and clear flour The rheological characteristics were evaluated by Mixolab 2 (Chopin Technologies, Paris, France) according to the ICC standard method No. 173 and Chopin+ protocol (ICC, 2006). The settings of tests were followed: 8 min at 30 °C, heating to 90 °C (at a rate of 4 °C/min), holding at 90 °C for 8 min, cooling to 55 °C (at a rate of 4 °C/min) and holding for 6 min.
2.4. Wheat milling Tempered wheat grains were ground using a Bühler MLU-202 Laboratory Flour Mill (Bühler, Switzerland). Flour was divided into six different fraction streams, including the first, the second, the third break and reduction. The six streams were mixed well to get straight flour, and the third break and the third reduction were mixed to obtain clear flour.
2.9. Analysis of pasting properties of straight flour and clear flour The pasting properties were studied using a Rapid Visco Analyzer (RVA, Model Super-3, Newport Scientific, Warriewood, Australia) following the AACC method 76–21 (AACC, 2000).
2.5. Analysis of microbial concentrations in tempered wheat grains, straight flour, clear flour, and bran
2.10. Analysis of protein extractability in sodium dodecyl sulfate (SDS) of straight flour and clear flour
25 g of samples was added into 225 mL 0.85% sterilized physiological saline solution and the mixture was homogenized for 90 s. Serial ten-fold dilutions were made with sterilized physiological saline solution. The total plate count (TPC) was performed according to GB 4789.2–2016 (Code of National Standard of China, 2016). 1 mL of the dilutions was poured onto sterile agar plates and incubated at 36 °C for 48 h. The yeast/mould count (YMC) was tested following GB 4789.15–2016 (Code of National Standard of China, 2016). Dilutions were placed onto bengal rose agar plates and incubated aerobically at 28 °C for 120 h. For mesophilic aerobic spores (MAS), dilutions were heat-treated under 80 °C for 15 min, cooled and poured onto nutrient agar plates, and incubated aerobically at 36 °C for 48 h (Berghofer et al., 2003).
Protein extractability in SDS was analyzed by size-exclusion high performance liquid chromatography (SE-HPLC) (Li, Sun, Han, Chen, & Tang, 2018). Samples containing 1.0 mg of proteins were extracted with 1.0 ml of 0.05 M sodium phosphate buffer (pH 7.0) including 2.0% (w/ v) SDS, hereafter referred to as SDS buffer, and were shaken for 60 min. Under reducing conditions, samples were extracted with 1.0 mL of SDS buffer containing 1.0% (w/v) dithiothreitol (DTT). The mixture was centrifuged at 8000 rpm for 10 min, and the supernatant was filtered through a 0.45 μm membrane. The extract was loaded onto a TSKgel G4000SWXL column (8 μm, 300 mm × 7.8 mm, TOSOH Corporation, Tokyo, Japan), and determined by HPLC (Shimadzu, Kyoto, Japan). The elution solvent was SDS buffer with a flow rate of 0.7 mL/min and a column temperature of 30 °C. Protein elution was monitored at 214 nm and samples were loaded every 30 min.
2.6. Determination of total polyphenols in bran and clear flour Samples (0.5 g) were put into 5 mL of methanol (70%, v/v) that was preheated at 70 °C for 30 min. The mixture was extracted at 70 °C for 10 min and then centrifuged at 3500 rpm for 10 min. The residue was repeatedly extracted with 5 mL of 70% methanol in the same conditions. All supernatants were combined and diluted with 70% methanol to a final volume of 10 mL. The total polyphenols were quantified by Folin-Ciocalteu method (ISO, 2005).
2.11. Statistical analysis SPSS 23.0 (SPSS Inc., Chicago, IL, USA) was used for data analysis, and all data were expressed as the means ± standard deviation. The variance analysis was conducted to determine the significance between the data at p < 0.05 level. 2
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Fig. 1. Effects of wheat tempering with SAEW under different ACC on TPC (a), YMC (b) and concentration of MAS (c) in tempered wheat grains, bran, clear flour and straight flour.
which surrounds the endosperm and the germ. However, compared to previous studies (Zhang et al., 2016), the decreases in TPC and YMC at similar ACC were relatively smaller. It may be due to the facts that: 1) the lower liquid-solid ratio (about 0.05) cannot make wheat grains be soaked in SAEW and is unable to provide enough SAEW for sterilization; 2) the rough surface of wheat grains may protect microorganisms to some extent; 3) organic matters in layers of wheat grains can compete with microbes to react with HClO.
3. Results and discussion 3.1. Microbial loads in tempered wheat grains, straight flour, clear flour, and bran The bactericidal effect of SAEW has been mainly attributed to the combination of high oxidation-reduction potential (ORP-reactions) and high concentration of hypochlorous acid (HClO) (Issa-Zacharia, Kamitani, Miwa, Muhimbula, & Iwasaki, 2011) that accounts for nearly 97% of the available chlorine in SAEW (Wang et al., 2018). In our study, the microbial load except MAS of wheat grains was increased significantly (p < 0.05) after tempering with distilled water (TPC: 5.58 lgCFU/g, YMC: 4.26 lgCFU/g, MAS: 2.41 lgCFU/g) compared to wheat grains before tempering (TPC: 4.52 lgCFU/g, YMC: 3.97 lgCFU/g, MAS: 2.31 lgCFU/g). This may be due to the fact that the long time, appropriate temperature and proper humidity during tempering process are beneficial to microbial growth. With the increase of SAEW concentration, TPC (Fig. 1a) and YMC (Fig. 1b) in all samples were decreased significantly (p < 0.05) while MAS only slightly changed (Fig. 1c). This is mainly because spores are mostly in the dormant state and difficult to be killed (Iurlina, Saiz, Fuselli, & Fritz, 2006). As was shown in Fig. 1a, after tempering with 70 mg/L SAEW, TPC of tempered wheat grains, straight flour, clear flour, and bran were reduced by 0.65, 0.72, 0.91, and 0.93 lgCFU/g, respectively. At the same time, nearly 0.49, 0.62, 0.73, and 0.78 lgCFU/g of YMC were inactivated in tempered wheat grains, straight flour, clear flour, and bran, respectively (Fig. 1b). The results also suggested that TPC and YMC reductions after SAEW treatment were the most in bran followed by those in clear flour. A possible explanation for this phenomenon may be that most of the microorganisms present in wheat grains are located in the pericarp
3.2. Changes of total polyphenols in bran and clear flour Because of strong oxidizing abilities of HClO in SAEW, tempering with SAEW may exert effects on bioactive compounds. It could be seen from Fig. 2 that the total polyphenols content was reduced significantly (p < 0.05) with the rise of SAEW concentration. The results were in line with the findings of Jia, Shi, Song, and Li (2015) who reported this was possibly due to the large number of hydroxyl radicals generated by HClO in SAEW. However, the reductions of total polyphenols in bran and clear flour (11.52% and 6.49%, respectively) were both at a relatively low level despite the SAEW concentration was 70 mg/L. It might be because: 1) The amount of SAEW was not enough; 2) SAEW expressed an obvious inhibition of PPO activity which consequently decreased the degradation of polyphenols (Bai, Guo, Zhu, & Zhou, 2017). 3.3. Enzyme activities in bran and clear flour As presented in Fig. 3a, the activity of PPO in bran was much higher than that in clear flour, and it showed a decline tendency when SAEW concentration increased. When SAEW concentration changed from 0 mg/L to 70 mg/L, PPO activities in bran and clear flour were 3
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radicals produced by HClO; and this oxidization might destroy the structure of PPO. However, the reduction of PPO activity in clear flour was smaller. It might be owing to that the outside bran could contact and react with SAEW firstly. On the other hand, SAEW could also inhibit the LA activity in bran and clear flour (Fig. 3b). The LA activity was decreased significantly (p < 0.05) after tempering with SAEW. Interestingly, the changes in LOX activity (Fig. 3c) were not significant (p > 0.05). 3.4. Rheological characteristics of straight flour and clear flour Rheological characteristics including dough mixing behavior, protein and starch behavior of flour were shown in Fig. 4, with corresponding parameters listed in Table 1A. Rheological properties characterized the protein quality include the following parameters: water absorption (W.A.), dough development time (D.T.T.), dough stability, and C2, and the latter part of the curve mainly indicates starch properties characterized by C3, C4, and C5 (Torbica, Hadnađev, & Dapčevic', 2010). As seen in Fig. 4a, there was little or no difference among straight flour samples. It is known that water diffusion begins from the germ and bran to the endosperm layer by layer gradually during tempering (Delcour & Hoseney, 2010). So HClO in SAEW is possibly consumed by organic matters in outer layers of wheat grains and attached microbes before water penetrating into the endosperm. Therefore, the influences of SAEW on straight flour might be negligible or difficult to detect. However, as shown in Fig. 4b and Table 1A, the rheological characteristics of clear flour were affected after SAEW treatment. It could be concluded that dough development time and dough stability had an increasing tendency which could be attributed to the stronger dough
Fig. 2. The total polyphenols in clear flour or bran as influenced by SAEW (mg/ g dry basis).
decreased by 37.82% and 15.85%, respectively. As such, SAEW treatment could significantly (p < 0.05) inhibit the PPO activity, which was in accordance with previous reports (Jia et al., 2015; Li, Ren, Hao, & Liu, 2017). Considering the conditions of nearly neutral pH and the high HClO concentration in SAEW, it was speculated that the inhibitory effect on PPO activity was resulted from strong oxidizing abilities of OH
Fig. 3. The activities of PPO (a), LA (b) and LOX (c) in clear flour and bran as affected by SAEW. 4
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Fig. 4. Mixolab complete test curves of straight flour (a) and clear flour (b), as well as RVA pasting curves of straight flour (c) and clear flour (d) as affected by SAEW. Table 1A Mixolab parameters of clear flour.a. concentration (mg/L) W.A. (%) D.T.T. (min) Stab. (min) C2 (Nm) C3 (Nm) C3–C4 (Nm) C5 (Nm) C5–C4 (Nm)
0 68.3 6.52 9.88 0.58 1.40 0.16 1.88 0.64
10 ± ± ± ± ± ± ± ±
a
0.1 0.01a 0.12a 0.00ab 0.00c 0.02a 0.01a 0.02b
68.9 6.73 9.98 0.58 1.38 0.14 1.85 0.61
30 ± ± ± ± ± ± ± ±
b
0.0 0.53a 0.11a 0.00ab 0.00b 0.00a 0.01b 0.01b
71.9 6.78 9.98 0.60 1.35 0.16 1.72 0.57
50 ± ± ± ± ± ± ± ±
d
0.3 0.15a 0.04a 0.01c 0.01a 0.01a 0.01a 0.01a
70.7 6.68 9.92 0.59 1.36 0.16 1.82 0.62
70 ± ± ± ± ± ± ± ±
c
0.2 0.32a 0.15a 0.00bc 0.01a 0.01a 0.01c 0.00b
70.9 ± 0.1c 6.74 ± 0.25a 10.04 ± 0.05a 0.57 ± 0.01a 1.34 ± 0.01a 0.17 ± 0.00a 1.78 ± 0.01b 0.60 ± 0.01b
Data was presented as mean ± standard deviation. Values with different superscript letters in rows were significantly different, p < 0.05. a W.A., water absorption; D.T.T., dough development time; Stab., stability.
structure. The oxidizability of SAEW can result in crosslinks between proteins, which improve the dough strength, whereas the C2 value indicating the protein weakening under the mechanical and thermal stress had no significant (p > 0.05) change. For starch features, it could be observed that system viscosity was decreased as SAEW concentration increased from 0 mg/L to 30 mg/L, but it was increased with the concentration increasing from 30 mg/L to 70 mg/L. SAEW treated groups had lower peak viscosity (smaller C3 value), unchanged cooking stability (constant C3–C4 value), lower final viscosity (smaller C5 value), and better anti-retrogradation ability (lower C5–C4 value) compared to the control. The decrease in starch viscosity may be due to the depolymerization of starch molecules caused by the oxidation process (Lawal, 2004; Zhou, Liu, Zhang, Chen, & Kong, 2016), while the protein aggregation caused by SAEW can inhibit the degressive tendency of starch viscosity. Moreover, the water absorption was increased
significantly (p < 0.05) as the concentration increased. This may be attributed to the structural weakening of starch granules. 3.5. Pasting properties of straight flour and clear flour As shown in Fig. 4c, there was no difference between straight flour samples, which was coincident with the results in Mixolab tests. However, pasting properties of clear flour were strongly affected by SAEW (Fig. 4d, Table 1B). As listed in Table 1B, compared with the control, the starch viscosities of all treated groups were decreased significantly (p < 0.05). Flour with lower breakdown and setback has better thermal paste stability, which can prompt clear flour to resist shear thinning and retrogradation. The peak viscosity, trough viscosity, breakdown, final viscosity, and setback of clear flour were decreased when the concentration increased from 0 mg/L to 30 mg/L. These 5
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Table 1B RVA parameters of clear flour. concentration (mg/L) 0 10 30 50 70
peak viscosity (mPa·s) e
1652 ± 13 1577 ± 3d 1271 ± 4a 1373 ± 8b 1425 ± 4c
trough viscosity (mPa·s) d
breakdown (mPa·s) c
1145 ± 8 1105 ± 24c 856 ± 18a 939 ± 12b 967 ± 15b
507 ± 6 472 ± 26bc 414 ± 17a 434 ± 17ab 458 ± 15b
final viscosity (mPa·s) e
2080 ± 13 2007 ± 23d 1659 ± 25a 1770 ± 11b 1822 ± 8c
setback (mPa·s) 935 ± 6c 902 ± 25c 803 ± 7a 831 ± 23ab 855 ± 22b
Data was presented as mean ± standard deviation. Values with different superscript letters in columns were significantly different, p < 0.05.
As was shown in Fig. 5a, compared to control group, neither the peak shape nor the peak area of SAEW treated samples changed, which indicated that SAEW did not catalyze the crossing-links between proteins in straight flour. However, it exited obvious differences in SEHPLC curves of clear flour. Generally, the total peak area was decreased as SAEW concentration increased, which indicated that the protein extractability in SDS declined with ACC increasing. Compared with the control, the area of F3, F4, F5, and F6 was decreased obviously when SAEW concentration increased from 30 mg/L to 70 mg/L. The protein extractability of clear flour declined significantly (p < 0.05) with the concentration increasing from 30 mg/L to 70 mg/L. These changes demonstrated the protein aggregation induced by crosslinking was caused by HClO in SAEW. This could also account for the increase of starch viscosity of clear flour mentioned above. In order to explore the nature of crosslinking happened in proteins of clear flour after SAEW treatment, reduced SE-HPLC profiles were carried out. Compared to non-reduced SE-HPLC chromatograms (Fig. 5b), the total peak area in reduced SE-HPLC curves (Fig. 5c) was increased largely. This is mainly due to the DTT effect that changed some unextractable proteins into SDS-extractable forms. Moreover, the total area of peaks had no significant (p > 0.05) change with the increase of SAEW concentration, and this indicated that the above polymerizations occurred mainly through disulphide bonds.
parameters, however, were increased as the concentration was up to 70 mg/L. The results were generally consistent with those obtained by Mixolab. In Mixolab tests, 70 mg/L samples showed lower viscosity than 50 mg/L groups while the results in RVA measurements presented the opposite trend. Different examination matrix used in RVA tests and Mixolab tests may account for the slight difference. The matrix measured in Mixolab tests is a dough and limited water is available for starch gelatinization, while RVA experiments are carried out with a suspension that provides enough water for starch gelatinization (Bucsella, Ágnes Takács, Vizer, Schwendener, & Tömösközi, 2016). The underlying mechanism of SAEW treatment on the pasting properties is still not elucidated. In previous reports, HClO was the main form of chlorine compounds in SAEW with pH value ranging from 5.8 to 6.3 (Issa-Zacharia et al., 2011), so the oxidizing activities of HClO may cause the decrease of pasting viscosity. The final and peak viscosity of SAEW treated groups were significantly (p < 0.05) lower than the control, which was mainly owing to the depolymerization of amylose and amylopectin molecules of starches (Lawal, 2004; Zhou et al., 2016). The breakdown was also reduced and this might be due to the carboxyl groups produced by the oxidation of hydroxyl groups, which could inhibit the association of starch molecules, promote the uniform distribution of starch molecules in the paste, and eventually improve the thermal stability of starch paste (Zhou et al., 2016). The retrogradation tendency of starches is mainly affected by the association of hydroxyl groups (Sandhu, Kaur, Singh, & Lim, 2008). It was speculated that the hydroxyl groups of starches were replaced with carbonyl groups generated during the oxidation, which might hinder the interaction between hydroxyl groups and consequently lead the reduction in setbacks of treated groups (Tavares, Zanatta, Zavareze, Helbig, & Dias, 2010; Zhou et al., 2016). Lyu, Gao, Zhou, Zhang, and Ding (2018) also found that the carbonyl and carboxyl content of starch granules isolated from wheat grains that were treated with acid electrolyzed water were increased. On the other hand, the changes in proteins might account for the increase of viscosity when ACC raised from 30 mg/L to 70 mg/L. The oxidation caused by HClO in SAEW can lead to the aggregation and conformational changes of wheat proteins. According to Neill, AlMuhtaseb, and Magee (2012), cross-linkage reactions might improve gluten viscosity. These changes in proteins might promote the starch granules to absorb more water (Hu, Wang, Zhu, & Li, 2017; Neill et al., 2012), which could result in higher viscosity and inhibit the downtrend of RVA viscosity.
4. Conclusions Tempering with SAEW altered microbial, biological, and chemical properties of different flour milling streams. TPC and YMC in straight flour, clear flour, and bran were all decreased significantly (p < 0.05) compared to the control after SAEW treatment. This improved the safety of end products and flour-based food. The total polyphenols, PPO activity, LA activity in clear flour and bran were significantly (p < 0.05) reduced as affected by SAEW. Moreover, clear flour presented a stronger dough structure, a better thermal paste stability and anti-retrogradation ability because of the oxidizing ability of HClO in SAEW, which indicated SAEW treatment could slightly improve qualities of clear flour. Results also showed that it had no significant (p > 0.05) difference on quality properties of straight flour. Since SAEW is environment-friendly, non-toxic, non-residual, cheap, and can effectively control the microbes of straight flour with no negative influence on its qualities, it has an excellent potential as a tempering solution to be widely used during the production of flour.
3.6. Protein extractability in SDS of straight flour and clear flour
Declaration of competing interest
The total extracted proteins were separated into six major peaks in the SE-HPLC profile referred to as F1 to F6 under non-reduced conditions. According to previous studies (Dewaest et al., 2017), the major peaks can be divided into three fractions. Fraction F1 corresponds to large polymeric proteins that are highly aggregated. Fraction F2 is likely to consist of smaller aggregated peaks with a continuous range of molecular size. F1 and F2 mainly indicate glutenin macropolymers. Peaks F3 and F4 correspond to large monomeric proteins. Fraction F5 and F6 represent small molecular weight proteins.
We declare that we have no financial and personal relationships with other people or organizations that can influence our work, there is no professional or other personal interest of any nature or kind in any product, service and/or company that could be construed as influencing the position presented in, or the review of, the manuscript entitled, “Effects of Wheat Tempering with Slightly Acidic Electrolyzed Water on the Microbial, Biological, and Chemical Characteristics of Different Flour Streams”.
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Fig. 5. Un-reduced SE-HPLC profiles of SDS extractable proteins in straight flour (a) and clear flour (b) and reduced SE-HPLC profiles of SDS extractable proteins in clear flour (c) as affected by SAEW.
Acknowledgments
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Effects of wheat tempering with slightly acidic electrolyzed water on the microbial, biological, and chemical characteristics of different flour streams Yun-Xia Chena,b, Xiao-Na Guoa,b, Jun-Jie Xinga,b, Xiao-Hong Sunc, Ke-Xue Zhua,b,∗ a
State Key Laboratory of Food Science and Technology, Jiangnan University, 1800 Lihu Avenue, Wuxi, 214122, Jiangsu Province, PR China School of Food Science and Technology, Jiangnan University, 1800 Lihu Avenue, Wuxi, 214122, Jiangsu Province, PR China c College of Food and Biological Engineering, Qiqihar University, Qiqihar, 161006, Heilongjiang Province, PR China b
ARTICLE INFO
ABSTRACT
Keywords: Tempering with SAEW Clear flour Bran Microbial control Quality properties
Wheat tempering was conducted with slightly acidic electrolyzed water (SAEW) at different active chlorine contents. Microbial, biological, and chemical properties of different flour streams milled from SAEW-tempered wheat grains were investigated. Results showed that the total plate count (TPC) and yeast/mould count (YMC) in flour products were significantly (p < 0.05) reduced as SAEW concentration increased. The total polyphenols, lipase (LA) activity, polyphenol oxidase (PPO) activity of clear flour and bran, along with starch viscosity and protein extractability under reduced conditions of clear flour were all decreased significantly (p < 0.05) when SAEW concentration raised. Meanwhile, farinograph properties of clear flour were slightly improved. However, there was no significant (p > 0.05) difference in biological and chemical characteristics of straight flour. Based on our findings, SAEW has potential to function as a novel tempering solution for producing clean flour.
1. Introduction Wheat grains are easily contaminated by microbes during growth, harvest, transportation, and storage (Li, Li, Luo, & Yoshizawa, 2002). The contaminated microorganisms in wheat grains can easily grow during the followed processing procedures, especially in tempering process due to the factors of long time, appropriate temperature and proper humidity (Berghofer, Hocking, Miskelly, & Jansson, 2003). The living microorganisms in flour can remain in a dormant state for a long period since flour generally has a low water activity (Eglezos, 2010; Rose, Bianchini, Martinez, & Flores, 2012). However, when flour is made into food and water is added, the microorganisms will proliferate immediately and lead to the degeneration of end products (Li et al., 2013). Thus, flour with high microbial load is considered as one of the major reasons for shortening the shelf-life of flour-based products, especially for fresh-wet noodle products. Meanwhile, some of heat-resistant spores, such as Bacillus spp., cannot be killed during production, and moulds can form allergenic spores and even produce mycotoxins (Schnürer, Olsson, & Börjesson, 1999). Therefore, microbial contamination within wheat grains is one of the biggest food safety issues of flour and flour-derived products, which urgently needs attention. To reduce the microbial load in wheat flour, some researchers replaced tempering water with ozonated water (İbanoǧlu, 2001), chlorinated water (Dhillon, Sandhu, Wiesenborn, Manthey, & Wolf-Hall, ∗
2007), organic acids or saline solutions (Sabillón, Stratton, Rose, Flores, & Bianchini, 2016). However, many problems limited the applications of these methods, such as instability, low efficacy, chemical residuals, and pipeline corrosion. SAEW as a novel antimicrobial agent has recently attracted lots of attention for its advantages. SAEW has been certified as a food additive by Japan Ministry of Health, Labour and Welfare (MHLW) and United States Food and Drug Administration (FDA). It can be generated by electrolyzing dilute NaCl and/or HCl solution in a non-membrane electrolytic chamber, resulting in a high concentration of hypochlorous acid with pH of 5.0–6.5 (Jadeja, Hung, & Bosilevac, 2013). The strong germicidal effect of SAEW is due to the available chlorine, including hypochlorite ion, hypochloric acid, and chlorine (Zheng et al., 2012). Compared with other bactericides, SAEW is more environment-friendly, non-toxic, non-residual, non-corrosive to organic matters, more stable and less expensive. In recent years, SAEW has been widely applied to extend shelf life of chicken meat (Rahman, Park, Song, Al-Harbi, & Oh, 2012), wash vegetables (Zhang, Cao, Hung, & Li, 2016) and clean shell eggs (Cao, Zhu, Shi, Wang, & Li, 2009). However, few research has been conducted on the applications of SAEW on wheat processing. The present study was designed to evaluate the disinfection efficacy of SAEW at different concentrations and its impacts on the microbial, biological, and chemical characteristics of different flour streams. At the same time, this paper also aimed to introduce a new concept of
Corresponding author. State Key Laboratory of Food Science and Technology, Jiangnan University, 1800 Lihu Avenue, Wuxi, 214122, Jiangsu Province, PR China. E-mail address: [email protected] (K.-X. Zhu).
https://doi.org/10.1016/j.lwt.2019.108790 Received 23 April 2019; Received in revised form 20 August 2019; Accepted 27 October 2019 0023-6438/ © 2019 Elsevier Ltd. All rights reserved.
Please cite this article as: Yun-Xia Chen, et al., LWT - Food Science and Technology, https://doi.org/10.1016/j.lwt.2019.108790
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production of “low-bacteria flour” that is mainly used to make freshwet noodle products.
2.7. Determination of activities of polyphenol oxidase (PPO), lipase (LA) and lipoxidase (LOX) in bran and clear flour The PPO activity was determined according to the method of Li et al. (2012) with some modifications. 1.0 g of sample was mixed into 10 mL extraction buffer (0.1 M phosphate buffer, pH 6.0) and then shaken under 4 °C for 24 h. The mixture was centrifuged at 10,000 rpm under 4 °C for 20 min. The test mixture was composed of 2 mL of the extract and 200 μL of catechol solution. One PPO activity unit was calculated as an increase in absorbance of 0.001 at 420 nm within 1 min. The LA activity was evaluated as described by Cai et al. (2011). 50 mM Tris-HCl buffer (10 mL, pH 8.0) was for the extraction of LA from 2.0 g of sample. 200 μL of the extract was reacted with 20 μL of 10 mM p-nitrophenyl octanoate in 1.78 mL of Tris-HCl buffer at 37 °C for 1 min. One LA activity unit was defined as an increase in absorbance of 0.1 at 405 nm within 1 min. The LOX activity was determined following the method of Cato, Halmos, and Small (2006). 2.0 g of sample was incubated in 10 mL of 0.1 M phosphate buffer (pH 7.5) and stirred on ice for 30 min. The mixture was then centrifuged at 8000 rpm at 4 °C for 10 min. The reaction mixture consisted of 50 mM sodium acetate buffer (2.89 mL, pH 5.5), linoleic acid substrate (90 μL), and LOX extract (20 μL). One LOX activity unit was defined as an increase in absorbance of 0.01 at 234 nm within 1 min.
2. Materials and methods 2.1. Wheat samples Wheat grains (Xinong-979, a kind of Henan hard white wheat grains) were obtained from China Oil & Foodstuffs Corporation in 2017 and were cleaned by sieves and hand. 2.2. Slightly acidic electrolyzed water (SAEW) SAEW was prepared with a hyper optimized chlorinated liquid device (HOCL 0.2 t pro, Meiloon Industrial Company Ltd., Taiwan, China). The active chlorine content (ACC) of prepared SAEW was 70 mg/L. SAEM with different ACC was obtained by mixing original SAEW with distilled water in various proportions and was used immediately after preparation. 2.3. Wheat tempering 500 g of cleaned wheat grains were placed in a sterile plastic bag and were tempered to 16% moisture with distilled water (control) and SAEW with different ACC (10, 30, 50, and 70 mg/L). Samples were shaken vigorously every 15 min during the first 2 h and were equilibrated for 24 h under 25 °C.
2.8. Analysis of rheological characteristics of straight flour and clear flour The rheological characteristics were evaluated by Mixolab 2 (Chopin Technologies, Paris, France) according to the ICC standard method No. 173 and Chopin+ protocol (ICC, 2006). The settings of tests were followed: 8 min at 30 °C, heating to 90 °C (at a rate of 4 °C/min), holding at 90 °C for 8 min, cooling to 55 °C (at a rate of 4 °C/min) and holding for 6 min.
2.4. Wheat milling Tempered wheat grains were ground using a Bühler MLU-202 Laboratory Flour Mill (Bühler, Switzerland). Flour was divided into six different fraction streams, including the first, the second, the third break and reduction. The six streams were mixed well to get straight flour, and the third break and the third reduction were mixed to obtain clear flour.
2.9. Analysis of pasting properties of straight flour and clear flour The pasting properties were studied using a Rapid Visco Analyzer (RVA, Model Super-3, Newport Scientific, Warriewood, Australia) following the AACC method 76–21 (AACC, 2000).
2.5. Analysis of microbial concentrations in tempered wheat grains, straight flour, clear flour, and bran
2.10. Analysis of protein extractability in sodium dodecyl sulfate (SDS) of straight flour and clear flour
25 g of samples was added into 225 mL 0.85% sterilized physiological saline solution and the mixture was homogenized for 90 s. Serial ten-fold dilutions were made with sterilized physiological saline solution. The total plate count (TPC) was performed according to GB 4789.2–2016 (Code of National Standard of China, 2016). 1 mL of the dilutions was poured onto sterile agar plates and incubated at 36 °C for 48 h. The yeast/mould count (YMC) was tested following GB 4789.15–2016 (Code of National Standard of China, 2016). Dilutions were placed onto bengal rose agar plates and incubated aerobically at 28 °C for 120 h. For mesophilic aerobic spores (MAS), dilutions were heat-treated under 80 °C for 15 min, cooled and poured onto nutrient agar plates, and incubated aerobically at 36 °C for 48 h (Berghofer et al., 2003).
Protein extractability in SDS was analyzed by size-exclusion high performance liquid chromatography (SE-HPLC) (Li, Sun, Han, Chen, & Tang, 2018). Samples containing 1.0 mg of proteins were extracted with 1.0 ml of 0.05 M sodium phosphate buffer (pH 7.0) including 2.0% (w/ v) SDS, hereafter referred to as SDS buffer, and were shaken for 60 min. Under reducing conditions, samples were extracted with 1.0 mL of SDS buffer containing 1.0% (w/v) dithiothreitol (DTT). The mixture was centrifuged at 8000 rpm for 10 min, and the supernatant was filtered through a 0.45 μm membrane. The extract was loaded onto a TSKgel G4000SWXL column (8 μm, 300 mm × 7.8 mm, TOSOH Corporation, Tokyo, Japan), and determined by HPLC (Shimadzu, Kyoto, Japan). The elution solvent was SDS buffer with a flow rate of 0.7 mL/min and a column temperature of 30 °C. Protein elution was monitored at 214 nm and samples were loaded every 30 min.
2.6. Determination of total polyphenols in bran and clear flour Samples (0.5 g) were put into 5 mL of methanol (70%, v/v) that was preheated at 70 °C for 30 min. The mixture was extracted at 70 °C for 10 min and then centrifuged at 3500 rpm for 10 min. The residue was repeatedly extracted with 5 mL of 70% methanol in the same conditions. All supernatants were combined and diluted with 70% methanol to a final volume of 10 mL. The total polyphenols were quantified by Folin-Ciocalteu method (ISO, 2005).
2.11. Statistical analysis SPSS 23.0 (SPSS Inc., Chicago, IL, USA) was used for data analysis, and all data were expressed as the means ± standard deviation. The variance analysis was conducted to determine the significance between the data at p < 0.05 level. 2
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Fig. 1. Effects of wheat tempering with SAEW under different ACC on TPC (a), YMC (b) and concentration of MAS (c) in tempered wheat grains, bran, clear flour and straight flour.
which surrounds the endosperm and the germ. However, compared to previous studies (Zhang et al., 2016), the decreases in TPC and YMC at similar ACC were relatively smaller. It may be due to the facts that: 1) the lower liquid-solid ratio (about 0.05) cannot make wheat grains be soaked in SAEW and is unable to provide enough SAEW for sterilization; 2) the rough surface of wheat grains may protect microorganisms to some extent; 3) organic matters in layers of wheat grains can compete with microbes to react with HClO.
3. Results and discussion 3.1. Microbial loads in tempered wheat grains, straight flour, clear flour, and bran The bactericidal effect of SAEW has been mainly attributed to the combination of high oxidation-reduction potential (ORP-reactions) and high concentration of hypochlorous acid (HClO) (Issa-Zacharia, Kamitani, Miwa, Muhimbula, & Iwasaki, 2011) that accounts for nearly 97% of the available chlorine in SAEW (Wang et al., 2018). In our study, the microbial load except MAS of wheat grains was increased significantly (p < 0.05) after tempering with distilled water (TPC: 5.58 lgCFU/g, YMC: 4.26 lgCFU/g, MAS: 2.41 lgCFU/g) compared to wheat grains before tempering (TPC: 4.52 lgCFU/g, YMC: 3.97 lgCFU/g, MAS: 2.31 lgCFU/g). This may be due to the fact that the long time, appropriate temperature and proper humidity during tempering process are beneficial to microbial growth. With the increase of SAEW concentration, TPC (Fig. 1a) and YMC (Fig. 1b) in all samples were decreased significantly (p < 0.05) while MAS only slightly changed (Fig. 1c). This is mainly because spores are mostly in the dormant state and difficult to be killed (Iurlina, Saiz, Fuselli, & Fritz, 2006). As was shown in Fig. 1a, after tempering with 70 mg/L SAEW, TPC of tempered wheat grains, straight flour, clear flour, and bran were reduced by 0.65, 0.72, 0.91, and 0.93 lgCFU/g, respectively. At the same time, nearly 0.49, 0.62, 0.73, and 0.78 lgCFU/g of YMC were inactivated in tempered wheat grains, straight flour, clear flour, and bran, respectively (Fig. 1b). The results also suggested that TPC and YMC reductions after SAEW treatment were the most in bran followed by those in clear flour. A possible explanation for this phenomenon may be that most of the microorganisms present in wheat grains are located in the pericarp
3.2. Changes of total polyphenols in bran and clear flour Because of strong oxidizing abilities of HClO in SAEW, tempering with SAEW may exert effects on bioactive compounds. It could be seen from Fig. 2 that the total polyphenols content was reduced significantly (p < 0.05) with the rise of SAEW concentration. The results were in line with the findings of Jia, Shi, Song, and Li (2015) who reported this was possibly due to the large number of hydroxyl radicals generated by HClO in SAEW. However, the reductions of total polyphenols in bran and clear flour (11.52% and 6.49%, respectively) were both at a relatively low level despite the SAEW concentration was 70 mg/L. It might be because: 1) The amount of SAEW was not enough; 2) SAEW expressed an obvious inhibition of PPO activity which consequently decreased the degradation of polyphenols (Bai, Guo, Zhu, & Zhou, 2017). 3.3. Enzyme activities in bran and clear flour As presented in Fig. 3a, the activity of PPO in bran was much higher than that in clear flour, and it showed a decline tendency when SAEW concentration increased. When SAEW concentration changed from 0 mg/L to 70 mg/L, PPO activities in bran and clear flour were 3
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radicals produced by HClO; and this oxidization might destroy the structure of PPO. However, the reduction of PPO activity in clear flour was smaller. It might be owing to that the outside bran could contact and react with SAEW firstly. On the other hand, SAEW could also inhibit the LA activity in bran and clear flour (Fig. 3b). The LA activity was decreased significantly (p < 0.05) after tempering with SAEW. Interestingly, the changes in LOX activity (Fig. 3c) were not significant (p > 0.05). 3.4. Rheological characteristics of straight flour and clear flour Rheological characteristics including dough mixing behavior, protein and starch behavior of flour were shown in Fig. 4, with corresponding parameters listed in Table 1A. Rheological properties characterized the protein quality include the following parameters: water absorption (W.A.), dough development time (D.T.T.), dough stability, and C2, and the latter part of the curve mainly indicates starch properties characterized by C3, C4, and C5 (Torbica, Hadnađev, & Dapčevic', 2010). As seen in Fig. 4a, there was little or no difference among straight flour samples. It is known that water diffusion begins from the germ and bran to the endosperm layer by layer gradually during tempering (Delcour & Hoseney, 2010). So HClO in SAEW is possibly consumed by organic matters in outer layers of wheat grains and attached microbes before water penetrating into the endosperm. Therefore, the influences of SAEW on straight flour might be negligible or difficult to detect. However, as shown in Fig. 4b and Table 1A, the rheological characteristics of clear flour were affected after SAEW treatment. It could be concluded that dough development time and dough stability had an increasing tendency which could be attributed to the stronger dough
Fig. 2. The total polyphenols in clear flour or bran as influenced by SAEW (mg/ g dry basis).
decreased by 37.82% and 15.85%, respectively. As such, SAEW treatment could significantly (p < 0.05) inhibit the PPO activity, which was in accordance with previous reports (Jia et al., 2015; Li, Ren, Hao, & Liu, 2017). Considering the conditions of nearly neutral pH and the high HClO concentration in SAEW, it was speculated that the inhibitory effect on PPO activity was resulted from strong oxidizing abilities of OH
Fig. 3. The activities of PPO (a), LA (b) and LOX (c) in clear flour and bran as affected by SAEW. 4
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Fig. 4. Mixolab complete test curves of straight flour (a) and clear flour (b), as well as RVA pasting curves of straight flour (c) and clear flour (d) as affected by SAEW. Table 1A Mixolab parameters of clear flour.a. concentration (mg/L) W.A. (%) D.T.T. (min) Stab. (min) C2 (Nm) C3 (Nm) C3–C4 (Nm) C5 (Nm) C5–C4 (Nm)
0 68.3 6.52 9.88 0.58 1.40 0.16 1.88 0.64
10 ± ± ± ± ± ± ± ±
a
0.1 0.01a 0.12a 0.00ab 0.00c 0.02a 0.01a 0.02b
68.9 6.73 9.98 0.58 1.38 0.14 1.85 0.61
30 ± ± ± ± ± ± ± ±
b
0.0 0.53a 0.11a 0.00ab 0.00b 0.00a 0.01b 0.01b
71.9 6.78 9.98 0.60 1.35 0.16 1.72 0.57
50 ± ± ± ± ± ± ± ±
d
0.3 0.15a 0.04a 0.01c 0.01a 0.01a 0.01a 0.01a
70.7 6.68 9.92 0.59 1.36 0.16 1.82 0.62
70 ± ± ± ± ± ± ± ±
c
0.2 0.32a 0.15a 0.00bc 0.01a 0.01a 0.01c 0.00b
70.9 ± 0.1c 6.74 ± 0.25a 10.04 ± 0.05a 0.57 ± 0.01a 1.34 ± 0.01a 0.17 ± 0.00a 1.78 ± 0.01b 0.60 ± 0.01b
Data was presented as mean ± standard deviation. Values with different superscript letters in rows were significantly different, p < 0.05. a W.A., water absorption; D.T.T., dough development time; Stab., stability.
structure. The oxidizability of SAEW can result in crosslinks between proteins, which improve the dough strength, whereas the C2 value indicating the protein weakening under the mechanical and thermal stress had no significant (p > 0.05) change. For starch features, it could be observed that system viscosity was decreased as SAEW concentration increased from 0 mg/L to 30 mg/L, but it was increased with the concentration increasing from 30 mg/L to 70 mg/L. SAEW treated groups had lower peak viscosity (smaller C3 value), unchanged cooking stability (constant C3–C4 value), lower final viscosity (smaller C5 value), and better anti-retrogradation ability (lower C5–C4 value) compared to the control. The decrease in starch viscosity may be due to the depolymerization of starch molecules caused by the oxidation process (Lawal, 2004; Zhou, Liu, Zhang, Chen, & Kong, 2016), while the protein aggregation caused by SAEW can inhibit the degressive tendency of starch viscosity. Moreover, the water absorption was increased
significantly (p < 0.05) as the concentration increased. This may be attributed to the structural weakening of starch granules. 3.5. Pasting properties of straight flour and clear flour As shown in Fig. 4c, there was no difference between straight flour samples, which was coincident with the results in Mixolab tests. However, pasting properties of clear flour were strongly affected by SAEW (Fig. 4d, Table 1B). As listed in Table 1B, compared with the control, the starch viscosities of all treated groups were decreased significantly (p < 0.05). Flour with lower breakdown and setback has better thermal paste stability, which can prompt clear flour to resist shear thinning and retrogradation. The peak viscosity, trough viscosity, breakdown, final viscosity, and setback of clear flour were decreased when the concentration increased from 0 mg/L to 30 mg/L. These 5
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Table 1B RVA parameters of clear flour. concentration (mg/L) 0 10 30 50 70
peak viscosity (mPa·s) e
1652 ± 13 1577 ± 3d 1271 ± 4a 1373 ± 8b 1425 ± 4c
trough viscosity (mPa·s) d
breakdown (mPa·s) c
1145 ± 8 1105 ± 24c 856 ± 18a 939 ± 12b 967 ± 15b
507 ± 6 472 ± 26bc 414 ± 17a 434 ± 17ab 458 ± 15b
final viscosity (mPa·s) e
2080 ± 13 2007 ± 23d 1659 ± 25a 1770 ± 11b 1822 ± 8c
setback (mPa·s) 935 ± 6c 902 ± 25c 803 ± 7a 831 ± 23ab 855 ± 22b
Data was presented as mean ± standard deviation. Values with different superscript letters in columns were significantly different, p < 0.05.
As was shown in Fig. 5a, compared to control group, neither the peak shape nor the peak area of SAEW treated samples changed, which indicated that SAEW did not catalyze the crossing-links between proteins in straight flour. However, it exited obvious differences in SEHPLC curves of clear flour. Generally, the total peak area was decreased as SAEW concentration increased, which indicated that the protein extractability in SDS declined with ACC increasing. Compared with the control, the area of F3, F4, F5, and F6 was decreased obviously when SAEW concentration increased from 30 mg/L to 70 mg/L. The protein extractability of clear flour declined significantly (p < 0.05) with the concentration increasing from 30 mg/L to 70 mg/L. These changes demonstrated the protein aggregation induced by crosslinking was caused by HClO in SAEW. This could also account for the increase of starch viscosity of clear flour mentioned above. In order to explore the nature of crosslinking happened in proteins of clear flour after SAEW treatment, reduced SE-HPLC profiles were carried out. Compared to non-reduced SE-HPLC chromatograms (Fig. 5b), the total peak area in reduced SE-HPLC curves (Fig. 5c) was increased largely. This is mainly due to the DTT effect that changed some unextractable proteins into SDS-extractable forms. Moreover, the total area of peaks had no significant (p > 0.05) change with the increase of SAEW concentration, and this indicated that the above polymerizations occurred mainly through disulphide bonds.
parameters, however, were increased as the concentration was up to 70 mg/L. The results were generally consistent with those obtained by Mixolab. In Mixolab tests, 70 mg/L samples showed lower viscosity than 50 mg/L groups while the results in RVA measurements presented the opposite trend. Different examination matrix used in RVA tests and Mixolab tests may account for the slight difference. The matrix measured in Mixolab tests is a dough and limited water is available for starch gelatinization, while RVA experiments are carried out with a suspension that provides enough water for starch gelatinization (Bucsella, Ágnes Takács, Vizer, Schwendener, & Tömösközi, 2016). The underlying mechanism of SAEW treatment on the pasting properties is still not elucidated. In previous reports, HClO was the main form of chlorine compounds in SAEW with pH value ranging from 5.8 to 6.3 (Issa-Zacharia et al., 2011), so the oxidizing activities of HClO may cause the decrease of pasting viscosity. The final and peak viscosity of SAEW treated groups were significantly (p < 0.05) lower than the control, which was mainly owing to the depolymerization of amylose and amylopectin molecules of starches (Lawal, 2004; Zhou et al., 2016). The breakdown was also reduced and this might be due to the carboxyl groups produced by the oxidation of hydroxyl groups, which could inhibit the association of starch molecules, promote the uniform distribution of starch molecules in the paste, and eventually improve the thermal stability of starch paste (Zhou et al., 2016). The retrogradation tendency of starches is mainly affected by the association of hydroxyl groups (Sandhu, Kaur, Singh, & Lim, 2008). It was speculated that the hydroxyl groups of starches were replaced with carbonyl groups generated during the oxidation, which might hinder the interaction between hydroxyl groups and consequently lead the reduction in setbacks of treated groups (Tavares, Zanatta, Zavareze, Helbig, & Dias, 2010; Zhou et al., 2016). Lyu, Gao, Zhou, Zhang, and Ding (2018) also found that the carbonyl and carboxyl content of starch granules isolated from wheat grains that were treated with acid electrolyzed water were increased. On the other hand, the changes in proteins might account for the increase of viscosity when ACC raised from 30 mg/L to 70 mg/L. The oxidation caused by HClO in SAEW can lead to the aggregation and conformational changes of wheat proteins. According to Neill, AlMuhtaseb, and Magee (2012), cross-linkage reactions might improve gluten viscosity. These changes in proteins might promote the starch granules to absorb more water (Hu, Wang, Zhu, & Li, 2017; Neill et al., 2012), which could result in higher viscosity and inhibit the downtrend of RVA viscosity.
4. Conclusions Tempering with SAEW altered microbial, biological, and chemical properties of different flour milling streams. TPC and YMC in straight flour, clear flour, and bran were all decreased significantly (p < 0.05) compared to the control after SAEW treatment. This improved the safety of end products and flour-based food. The total polyphenols, PPO activity, LA activity in clear flour and bran were significantly (p < 0.05) reduced as affected by SAEW. Moreover, clear flour presented a stronger dough structure, a better thermal paste stability and anti-retrogradation ability because of the oxidizing ability of HClO in SAEW, which indicated SAEW treatment could slightly improve qualities of clear flour. Results also showed that it had no significant (p > 0.05) difference on quality properties of straight flour. Since SAEW is environment-friendly, non-toxic, non-residual, cheap, and can effectively control the microbes of straight flour with no negative influence on its qualities, it has an excellent potential as a tempering solution to be widely used during the production of flour.
3.6. Protein extractability in SDS of straight flour and clear flour
Declaration of competing interest
The total extracted proteins were separated into six major peaks in the SE-HPLC profile referred to as F1 to F6 under non-reduced conditions. According to previous studies (Dewaest et al., 2017), the major peaks can be divided into three fractions. Fraction F1 corresponds to large polymeric proteins that are highly aggregated. Fraction F2 is likely to consist of smaller aggregated peaks with a continuous range of molecular size. F1 and F2 mainly indicate glutenin macropolymers. Peaks F3 and F4 correspond to large monomeric proteins. Fraction F5 and F6 represent small molecular weight proteins.
We declare that we have no financial and personal relationships with other people or organizations that can influence our work, there is no professional or other personal interest of any nature or kind in any product, service and/or company that could be construed as influencing the position presented in, or the review of, the manuscript entitled, “Effects of Wheat Tempering with Slightly Acidic Electrolyzed Water on the Microbial, Biological, and Chemical Characteristics of Different Flour Streams”.
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Fig. 5. Un-reduced SE-HPLC profiles of SDS extractable proteins in straight flour (a) and clear flour (b) and reduced SE-HPLC profiles of SDS extractable proteins in clear flour (c) as affected by SAEW.
Acknowledgments
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