Effects of electron beam irradiation on storability of brown and milled rice

Journal of Stored Products Research 81 (2019) 22e30

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Effects of electron beam irradiation on storability of brown and milled rice Xiaohu Luo a, b, c, 1, Yulin Li d, 1, Dan Yang c, Jiali Xing e, f, Ke Li c, Ming Yang c, Ren Wang c, Li Wang c, Yuwei Zhang g, Zhengxing Chen c, * a Key Laboratory of Agro-products Quality and Safety Control in Storage and Transport Process, Ministry of Agriculture and Rural Affairs/Institute of Food Science and Technology, Chinese Academy of Agricultural Sciences, Beijing, 100193, China b Beijing Advanced Innovation Center for Food Nutrition and Human Health, Beijing Technology and Business University (BTBU), Beijing, 100048, China c National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, Wuxi, 214122, China d Hubei Key Laboratory of Edible Wild Plants Conservation and Utilization (Hubei Normal University), Huangshi, 435002, China e Ningbo Institute for Food Control, Ningbo, 315048, China f School of Marine Science, Ningbo University, Ningbo, 315211, China g Wuxi EL PONT Radiation Technology Co., Ltd, Wuxi 214151, China

a r t i c l e i n f o

a b s t r a c t

Article history: Received 20 March 2018 Received in revised form 28 November 2018 Accepted 18 December 2018

The effects of electron beam irradiation (EBI) treatment on the freshness and quality of brown and milled rice at different irradiation doses were investigated. The colors of brown rice (P > 0.05) and milled rice (P < 0.05) slightly changed after 5 kGy irradiation. After irradiation, the viscosities and amylose contents of the samples decreased (P < 0.05), crystal forms remained unchanged, but crystallinities decreased (P < 0.05). The microstructures of the samples did not change according to the results of the scanning electron microscopy. During the 120-day storage process, the free fatty acids of the irradiated samples increased at a slower rate than the non-irradiated samples did. Lipase activity was inhibited effectively, and the total viable bacterial count was significantly reduced (P < 0.05). Results indicated the potential of EBI to improve the quality and extend the shelf lives of brown and milled rice during storage. © 2018 Elsevier Ltd. All rights reserved.

Keywords: Brown rice Milled rice Electron beam irradiation Storage Quality

1. Introduction Rice (Oryza sativa) is a staple food for more than half of the world's population, especially Asian populations. This crop is mainly consumed as cooked rice and is partly used as seed or as an ingredient for processed food. This pattern of use implies the need to store rice over varying periods (Zhou et al., 2016). However, rice easily spoils during storage due to the growth of insects, bacteria, and molds (Gregory et al., 2008). Chemical reactions such as lipid oxidation and decomposition are also common causes of rice spoilage during storage. Thus, when rice is exposed to an unfavorable external environment (such as high temperature conditions), enzymatic activity may increase and accelerate lipid degradation during the storage period, thereby reducing the

* Corresponding author. E-mail addresses: [email protected], [email protected] (Z. Chen). 1 These authors contributed equally to this work and should be considered cofirst authors. https://doi.org/10.1016/j.jspr.2018.12.003 0022-474X/© 2018 Elsevier Ltd. All rights reserved.

sensory quality of rice (Zhou et al., 2015). Brown rice deteriorates more easily than milled rice does due to the lack of protection from the rice hull, and it carries greater nutritional value than white rice ~ o et al., 2017; Arthur, 2016). During storage, the does (Castan nutritional quality and storability of rice can deteriorate because of several mechanisms caused by changes in the rice's internal chemistry. Approximately 10% of post-harvest losses in stored rice throughout the world are caused by insects, mites, rodents, and microbes that are attracted to the abundant nutrient content of rice. Such losses are also attributed to the difficulty in controlling the storage environment. Approximately 1% and 10%e30% of these losses have been reported in developed and developing nations, respectively (Yadav et al., 2014). The overall damage caused by insects and microbiological factors affects approximately 27% of the annual total rice yield during storage (Alfonso-Rubí et al., 2003). Effective storage technologies are required to protect rice from undesirable changes. Traditional technologies include cryogenic storage, controlledatmosphere storage, and chemical storage (Li, 2013; Wijayaratne

X. Luo et al. / Journal of Stored Products Research 81 (2019) 22e30

et al., 2018). However, these methods have several drawbacks. Although traditional low-temperature or atmospheric storage can inhibit the growth of eggs by reducing the temperature or changing the air composition, insect pests in larvae and adult stages cannot be killed with such methods. Moreover, these methods perform poorly in mildew and pest control, as well as in preservation. Chemical storage methods require the long-term use of chemicals. However, these chemicals promote pest resistance, thereby increasing the difficulty of prevention while polluting the environment and threatening human health due to their large-scale use. New approaches to grain storage are continuously being explored (Li and Farid, 2006). For example, Ding et al. (2016) used infrared radiation heating to improve the shelf life of brown rice, and Subramanyam et al. (2017) investigated the effects of different ozone concentrations on pests to prolong the storage period of wheat. Food irradiation has been recognized as a preservation technique to extend the shelf life of food products for several decades (Li et al., 2016; Sirisoontaralak and Noomhorm, 2007; Watters and MacQueen, 1967). Sources of ionizing radiation originate from electron beams or X-rays generated in electrically driven machines or gamma rays from radioactive 60Co or 137Cs (Farkas, 2006). Radionuclides 60Co and 137Cs have been widely used in food irradiation (Nayak et al., 2007; Wang and Chao, 2003). However, these radionuclides are made disadvantageous by the continuous emission of gamma rays. Strict safety measures are needed in facilities to protect personnel and the environment even when these substances are not being used. High-energy electrons are mechanically produced by electric energy in electron accelerators without the use of any radioactive source. Accelerators are switched off when not in use. This practice reduces the risk of fatal failures. Furthermore, electron beams do not continuously emit gamma rays. In 1980, the Joint FAO/IAEA/ WHO Expert Committee on Food Irradiation recommended an increased dose of 10 kGy (1 Gy is a dose equal to 1 J/kg of absorbing material) for food irradiation processes. This method is considered safe and does not induce radiation (Yang et al., 2014). Despite this safety, the effects of electron beam irradiation (EBI) on the physicochemical and storage properties of food, especially rice, have received limited attention. Thus, in the present study, the effects of EBI on the physicochemical properties of brown and milled rice, including their microstructures, pasting properties, relative crystallinity, apparent amylose contents, and color differences, were investigated. Storability, free fatty acids, lipase activities, and total viable bacterial counts were also determined.

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thickness of
0.02 mm. Then, the samples were subjected to electron beam treatment with doses of 1, 3, and 5 kGy at a dose rate of 2 kGy/h at room temperature. The dosage was selected on the basis of previous studies (Chen et al., 2016a,b), and long shelf life was sufficiently ensured by insect disinfestation. The samples did not show changes in temperature and moisture content during the process. The irradiation treatments were performed at Wuxi EL PONT Radiation Technology Co. Non-irradiated samples (0 kGy dose) were used for comparison. 2.3. Preparation of rice flour Brown and milled rice were polished into powder by using a high-speed crusher (JP-150A type, Yongkang Jiu Pin Industry and Trade Co). One portion was passed through a sieve with a pore size of 0.180 mm to detect relative crystallinity. The other portion was sieved using a pore size of 0.250 mm to determine color and viscosity differences. 2.4. Starch isolation The extraction method for rice starch was modified in accordance with the method described in the literature (Zhong et al., 2009). A certain amount of rice was soaked in deionized water for 12 h. The solideliquid mixture was beaten using a beater for 2 min, followed by a colloid mill for 5 min. The mixture was subsequently centrifuged to remove water and then soaked in 0.2% sodium hydroxide solution for 24 h. The mixture was stirred thoroughly and passed through a sieve with a pore size of 0.150 mm. The filtrate was repassed through a 300-mesh sieve (pore size of 0.05 mm) twice and centrifuged at 4500 r/min for 10 min. Then, the upper gray matter (protein) was scraped off. This gray matter was redissolved and centrifuged, and the upper layer was scraped. The pH was adjusted to 7.0 using dilute hydrochloric acid. Three centrifugations were then performed to remove salt. The precipitate was lyophilized to obtain the rice starch for detecting apparent amylose content. 2.5. Color measurement

2. Materials and methods

The color of the rice was determined using a Konica Minolta Chroma CR400 (Konica Minolta Inc., Osaka, Japan). The rice was crushed into powder and mixed evenly beforehand to avoid measurement errors caused by the uneven distribution of rice grain color. The parameters of lightness (L*), redegreenness (a*), and yelloweblueness (b*) values were measured.

2.1. Rice

2.6. Starch viscosity determination

Rice (Zhengnong 1, late-cropping season japonica Oryza sativa) was cultivated in Yancheng City, Jiangsu Province and was harvested in August 2016 with approximately 22% moisture content. The grains were transported to the National Engineering Laboratory for Cereal Fermentation Technology of Jiangnan University and cleaned and dried to achieve 14% moisture content. Thereafter, a portion of the grains was dehusked to produce brown rice, and another portion was hulled and ground in hulling (model JLGJ2.5, Yiujiang Machinery Corporation in Zhejiang) and milling machines (model RSKM5B, Tohoku Satake Corporation) to produce milled rice.

Pasting properties were determined using a rapid viscosity €gersten, Sweden), analyzer (RVA 4500, Perten Instruments Ha which was controlled by computer software. The temperature was first maintained at 50  C for 1 min to obtain a stable temperature and then heated to 95  C at a rate of 10  C/min. The sample was kept at 95  C for 1.5 min and cooled to 50  C at a rate of 10  C/min. The temperature was maintained at 50  C for 1.5 min, and the detection of pasting behavior was obtained. The experiments were carried out in triplicate to inspect the reproducibility of the sample, and the average values were noted. The pasting properties were recorded to describe the divergence of the RVA profile caused by EBI. These pasting properties included peak viscosity (PKV), cool pasting viscosity (CPV), hot pasting viscosity (HPV), breakdown (PKV minus HPV), setback (CPV C minus PKV), breakdown (PKV minus HPV), and peak time (PKT).

2.2. Irradiation of samples Samples of 500 g rice were packed in polyethylene bags with a

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X. Luo et al. / Journal of Stored Products Research 81 (2019) 22e30

2.7. X-ray diffraction and relative crystallinity The moisture of the samples was equilibrated in a desiccator containing water for 18 h in room temperature. The X-ray diffraction of the native and irradiated samples was conducted using an Xray diffractometer (Shimadzu XRD 7000) at the scanning speed of 2 /min at 40 kV and 30 mA. The 2q ranged from 5 to 45 . Relative crystallinity was quantitatively determined by the ratio of the peak to total areas, as described by Sullivan et al. (2017), using the MDI Jade 6.0 software. 2.8. Apparent amylose content The apparent amylose contents of the samples were determined by the method set by the Chinese National Standard (GAQS, 2008b) and ISO methodology (1987) using a spectrophotometer (UV-1100). The wavelength was set to 720 nm. 2.9. Microstructural observation The microstructures of the samples irradiated at different doses were observed to investigate the effects of EBI on the structural properties of brown and milled rice using a scanning electron microscope (S-4800, Hitachi, Tokyo, Japan). The rice grains used in this experiment were broken off along the short axis. On each broken surface, a location at 0.1e0.2 mm from the lateral surface was selected for observation. The accelerator voltage was 50 kV, and the picture was obtained in multiples of 40 and 2400 times. Similarly, the starch extracted from the brown rice and milled rice was also scanned by a scanning electron microscope. The accelerator voltage was 50 kV, and the pictures were obtained in multiples of 2400 times. 2.10. Storability determination 2.10.1. Storage Samples of 200 g each of brown and milled rice were packed in polyethylene bags separately. The samples were stored in an incubator at 35  C for four months, and the related indexes, including free fatty acids, lipase activities, and total viable bacterial counts, were determined every month. 2.10.2. Free fatty acids The free fatty acids of the samples were detected in accordance with the Chinese National Standard (GAQS, 2015) and previous studies (Tan et al., 2017) with some modification. The modifications were as follows. Samples (10.0 g) were weighed in a 250 mL stoppered conical flask. Absolute ethanol (50 mL) was added, oscillated for 30 min, and allowed to stand for 5 min after filtration. The 25 mL filtrate was pipetted into a 150 mL Erlenmeyer flask, and 50 mL of distilled water free of carbon dioxide was added. A magnetic stirrer was turned on after placing the composite electrode and stir bar. The mixture was then titrated with a standard potassium hydroxide (0.1 mol/mL) ethanol solution to adjust the pH to 8.0. The final results were expressed as milligrams of potassium hydroxide needed to neutralize free fatty acids in 100 g of rice flour (dry basis). 2.10.3. Lipase activity The lipase activity values of the samples were measured according to the Chinese National Standard (GAQS, 2008a) and previous studies (Hernandez-Garcia et al., 2017). Samples (2.00 g) were weighed into a mortar, to which a small amount of quartz sand and 1 mL of pure oil were added. The mixture was combined evenly after adding 5 mL of buffer (11.846 g of disodium hydrogen phosphate dissolved in 11 mL of water and 4.539 g of potassium

dihydrogen phosphate dissolved in 11 mL of water to form a 4:1 mixture). The mixture was then ground into a thin paste, transferred into a stoppered conical flask, and incubated at 30  C for 24 h. A volume of 50 mL was removed from the mixture of ethanol and ether and allowed to stand for 1e2 min after filtration. A 50 mL filtrate was collected. The 25 mL filtrate was pipetted into a 150 mL Erlenmeyer flask and titrated with potassium hydroxide (0.05 mol/ mL) ethanol solution to adjust the pH to 8.0. The results were expressed in the form of milligrams of potassium hydroxide. The blank test operation followed nearly the same procedure, except that the mixture was kept warm for 24 h. 2.10.4. Total viable bacterial count The total viable bacterial count was determined in accordance with the method described in the national standard method (MOHC, 2010) and previous studies (Lu et al., 2011). We used a tenfold dilution method. Three dilution gradients (i.e.,  10,  100, and  1000) were selected. 2.11. Experimental design and data analysis The design of the experiments can be divided into two major parts. In the first part, the samples irradiated with different doses of electron beam (0, 1, 3, and 5 kGy) were prepared immediately to determine their properties, including color differences, starch viscosity, relative crystallinity, apparent amylose content, and microstructure. The aim for this part was to determine whether EBI had favorable or adverse effects on the quality of milled rice and brown rice. In the second part, the irradiated milled rice and brown rice were stored in an incubator at 35  C for four months (accelerated storage) to investigate the effects of the EBI treatment on the storability of the samples. The samples used for index determination in the two parts were randomly selected from a large number of experimental samples. For experiment 2.5 on color measurement, six samples were randomly selected from the experimental samples, crushed into powder and then mixed evenly beforehand separately to avoid measurement errors caused by the uneven distribution of rice grain color. Each sample was measured once, and the procedure was repeated six times. Therefore, each sample was related to six sets of parallel experimental data. In determining starch viscosity, relative crystallinity, and apparent amylose content, three parallel samples were selected from each section randomly, and the parallel samples were processed to obtain three sets of parallel data. In experiment 2.9 on microstructural observations, image analysis was carried out by randomly selecting three samples from each batch and capturing images of four different regions from each sample. All images presented in this paper were randomly selected. In experiment 2.10 on storability, three samples were randomly selected from each batch every other month (30 days) in the span of four months of accelerated storage. All the data obtained from the experiments were based on a completely randomized design. The results were expressed as standard deviations of the means and were analyzed with one-way analysis of variance (ANOVA) procedure using SPSS17.0 software package (SPSS Inc., Chicago, IL, USA). The means of the data were later separated by Tukey's test, and the results were considered significant only when P
0.05. 3. Results 3.1. Effects of irradiation on rice color The color values, including L*, a*, and b* of native and irradiated rice, are summarized in Table 1. L* represents the brightness value,

X. Luo et al. / Journal of Stored Products Research 81 (2019) 22e30 Table 1 Effect of EBI on the color of rice. Type

Dose/kGy

L*

a*

b*

Brown rice

0 1 3 5 0 1 3 5

87.78 ± 0.39a 87.59 ± 0.51a 86.25 ± 0.25b 86.18 ± 0.43b 96.48 ± 0.13a 96.01 ± 0.16b 96.05 ± 0.01b 95.92 ± 0.09c

1.32 ± 0.09a 1.32 ± 0.10a 1.33 ± 0.06a 1.21 ± 0.09a 0.34 ± 0.02a 0.37 ± 0.02a 0.36 ± 0.01a 0.50 ± 0.02b

11.34 ± 0.31a 11.66 ± 0.36a 12.58 ± 0.08b 12.33 ± 0.20b 5.70 ± 0.11a 6.03 ± 0.09b 6.03 ± 0.03b 6.47 ± 0.07c

Milled rice

Values expressed are means of 3 standard deviations. Means in the row with different superscripts are significantly different at p
0.05.

a* represents the redegreen value, and b* represents the blueeyellow value (Hayashi and Yanase, 2016; Oli et al., 2016). The results of the color difference measurement in Table 1, that is, the lightness, redegreen values, and blueeyellow values of brown rice, were not greatly affected by irradiation (P > 0.05). Moreover, the color values of milled rice did not change significantly after irradiation with 1 and 3 kGy doses (P > 0.05). However, after irradiation with an electron beam at 5 kGy, the brightness and redegreen values of the milled rice showed a significant decrease (P < 0.05) from 96.48 to 95.92 and from 0.34 to 0.50, respectively. Furthermore, the blueeyellow values increased significantly (P < 0.05) from 5.70 to 6.47. 3.2. Effects of irradiation on starch viscosity The pasting properties of rice flour are listed in Table 2. The starch viscosities of brown and milled rice were significantly altered by the effect of EBI. Six major parameters of starch pasting curve, namely, peak viscosity (PV), trough viscosity (TV), breakdown viscosity (BV), final viscosity (FV), setback viscosity (SV), and peak time (PT), were considerably decreased with the increase of dose at different velocities. With the increased irradiation dose from 0 kGy to 5 kGy, the values of PV, TV, BV, FV, and SV decreased by 1032.0, 862.0, 173.0, 1575.0, and 713.0 cP for brown rice, respectively; and 1823.0, 835.0, 399.0, 2630.0, and 1206.0 cP for milled rice, respectively. 3.3. Effects of irradiation on relative crystallinity Starch consists of ordered crystalline and disordered amorphous regions (Amagliani et al., 2016). In a diffraction pattern, the crystalline region corresponds to a peak diffraction characteristic, whereas the disordered amorphous region corresponds to a diffractive diffraction characteristic (Jin and Tian, 2016). Given the Xray diffraction pattern of starch, the crystalline forms of natural

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starch can be divided into three types: A, B, and C. Under normal circumstances, most cereal starches (e.g., corn starch and wheat starch) show A-type crystalline structures. The characteristic diffraction angles are 15 , 17, 18 , and 23 . Root starch (e.g., potato starch) exhibit a B-type crystal structure with characteristic diffraction angles of 5 , 17, 22 , and 24 . The C-type crystal structure (e.g., seed starches) involves types A and B. The characteristic diffraction angles are 5.73 , 15.3 , 17.3 , and 23.5 . The X-ray diffraction patterns of native and irradiated rice starches are illustrated in Fig. 1. The angular position and intensity of these different peaks correspond to the A-type morphology of cereal starches. After EBI, the relative crystallinity of brown rice starches gradually decreased from 31.98% to 27.28%, whereas that of milled rice starches decreased from 20.47% to 15.09% as the dose increased from 0 kGy to 5 kGy. 3.4. Effects of irradiation on amylose content The amylose content of native and irradiated brown and milled rice is presented in Table 3. The amylose content of brown rice ranged from 19% to 15%, and that of milled rice ranged from 15% to 13%. Moreover, the amylose content of milled rice was higher than that of brown rice. The amylose content of non-irradiated brown rice and that of milled rice were 15.96 ± 0.03% and 18.52 ± 0.11%, respectively. After EBI at 1, 3, and 5 kGy, a gradual decrease in amylose content was observed as the radiation dose increased. The amylose content of brown rice decreased by 6.39%, 13.10%, and 16.23%; and that of milled rice decreased by 4.86%, 11.34%, and 16.31% relative to the original percentages. 3.5. Effects of irradiation on microstructure The scanning electron microscopy (SEM) microphotographs presented in Fig. 2 show that the rice grain microstructure was not affected by EBI. From the images of the cross sections of brown rice and milled rice, we did not find significant damage in the form of broken particles and holes. Although a number of small cracks were noted in the cross section, they were already observed before the analysis possibly as a result of the drying process. Similarly, the SEM images of the starches extracted from the milled and brown rice grains showed that the shape, size, and uniformity of the starch granules did not change significantly. 3.6. Effects of irradiation on free fatty acids and lipase activity After EBI, the free fatty acids of the samples slightly increased (P < 0.5) (Table 4). The free fatty acid content of brown rice increased from 8.32 ± 1.02 mg KOH/100 g to 9.03 ± 0.02, 9.23 ± 0.12, and 8.98 ± 0.09 mg KOH/100 g, representing increases of 8.53%,

Table 2 Pasting properties of native and irradiated rice flour (n ¼ 3). type Brown rice

Dose (kGy)

0 1 3 5 Milled rice 0 1 3 5

Peak viscosity (CP)

Trough viscosity (CP)

Breakdown viscosity (CP)

Final viscosity (CP)

Setback (CP)

Peak time (min) Pasting temperature ( C)

1717.00,±,12.13a 1321.00 ± 21.11b 1207.00 ± 32.77c 682.00 ± 12.72d 2946.00 ± 54.33a 2234.00 ± 12.33b 1784.00 ± 32.33c 1123.00 ± 10.11d

1045.00 ± 10.01a 650.00 ± 9.08b 561.00 ± 12.88c 183.00 ± 12.11d 1765.00 ± 42.77a 1143.00 ± 11.02b 774.00 ± 45.65c 341.00 ± 12.34d

672.00 ± 6.09a 671.00 ± 7.77a 646.00 ± 14.21b 499.00 ± 13.42c 1181.00 ± 13.56a 1091.00 ± 9.83b 1010.00 ± 10.82c 782.00 ± 7.87d

2104.00 ± 12.11a 1488.00 ± 17.87b 1352.00 ± 34.11c 529.00 ± 22.33d 3462.00 ± 32.77a 2434.00 ± 21.32b 1775.00 ± 17.89c 832.00 ± 3.21d

1059.00 ± 10.09a 838.00 ± 7.92b 791.00 ± 1.23c 346.00 ± 12.32d 1697.00 ± 18.92a 1291.00 ± 12.56b 1001.00 ± 19.94c 491.00 ± 0.07d

6.07 ± 0.02a 5.87 ± 0.01b 5.67 ± 0.01c 5.27 ± 0.02d 6.00 ± 0.01a 5.87 ± 0.01b 5.64 ± 0.03c 5.33 ± 0.04d

Values expressed are means of 3 standard deviations. Means in the row with different superscripts are significantly different at p
0.05.

88.80 ± 0.07 88.75 ± 0.02 88.00 ± 0.04 88.05 ± 0.02 84.70 ± 0.01 84.85 ± 0.02 84.75 ± 0.00 84.70 ± 0.02

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X. Luo et al. / Journal of Stored Products Research 81 (2019) 22e30

Fig. 1. XRD patterns of rice flour at different EBI doses; (A) XRD patterns of brown rice; (B) XRD patterns of milled rice.

Table 3 Amylose contents of the native and irradiated rice flours (n ¼ 3). Type

Dose (kGy)

Amylose content (%)

Brown rice

0 1 3 5 0 1 3 5

15.96 ± 0.03a 14.94 ± 0.04b 13.87 ± 0.03c 13.37 ± 0.02d 18.52 ± 0.11a 17.62 ± 0.07b 16.42 ± 0.01c 15.50 ± 0.01d

Milled rice

Values expressed are means of 3 standard deviations. Means in the row with different superscripts are significantly different at p
0.05.

10.94%, and 7.93%, respectively. That of milled rice increased from 6.32 ± 0.02 mg KOH/100 g to 7.03 ± 0.06, 7.33 ± 0.72, and 8.08 ± 0.18 mg KOH/100 g, denoting increases of 11.23%, 15.98%, and 27.06%, respectively. However, during the 120-day storage process, the amount of free fatty acids of the irradiated sample was lower than that of the non-irradiated samples (Fig. 3 A and B). The lipase activities of the irradiated brown and milled rice were lower than those of the non-irradiated samples (Table 4). The lipase activities also decreased with prolonged storage time (Fig. 3 C and D). As the storage time increased from 0 day to 120 days, the slope of the enzyme activity curve became gentle.

Fig. 2. Scanning electron microscopy (SEM) microphotographs of rice at different EBI doses; (A) SEM of brown rice at different EBI doses. The leftmost and middle columns show the images of the cross section of brown rice (40  ) and the images of a location at 0.1e0.2 mm from the lateral surface of brown (2400  ). The images in the right column are magnified views of brown rice starch (2400  ); (B) SEM of milled rice at different EBI doses. The leftmost and middle columns show the images of the cross section of milled rice (40  ) and the images of a location at 0.1e0.2 mm from the lateral surface of the milled rice (2400  ). The images in the right column are magnified views of milled rice starch (2400  ). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

X. Luo et al. / Journal of Stored Products Research 81 (2019) 22e30

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Table 4 Free fatty acids contents and lipase activity of the native and irradiated rice (n ¼ 3). Type

Dose (kGy)

Free fatty acids (mg KOH/100g)

Lipase activity (mg KOH/100g)

Brown rice

0 1 3 5 0 1 3 5

8.32 ± 1.02a 9.03 ± 0.02b 9.23 ± 0.12c 8.98 ± 0.09b 6.32 ± 0.02a 7.03 ± 0.06b 7.33 ± 0.72c 8.08 ± 0.18d

6.72 ± 0.06a 6.01 ± 0.06b 5.96 ± 0.08b 5.49 ± 0.04c 6.21 ± 0.06a 5.98 ± 0.08b 5.78 ± 0.07c 5.39 ± 0.06d

Milled rice

Values expressed are means of 3 standard deviations. Means in the row with different superscripts are significantly different at p
0.05.

3.7. Effects of irradiation on total viable bacterial count Fig. 3 shows the effects of different EBI doses on the total viable bacterial count in brown and milled rice during storage. The total viable bacterial counts in brown and milled rice before the EBI treatment were 3  103 and 2.8  103 cfu/g, respectively; these values generally increased with prolonged storage time (P < 0.5). After irradiation at doses of 1, 3, and 5 kGy, the total viable bacterial counts of brown rice and milled rice decreased to 800, 120, and 10 cfu/g; and to 720, 100, and 0 cfu/g, respectively. The inactivation rates reached 73.33%, 96.00%, and 99.67% for the brown rice; and 74.29%, 96.43%, and 100% for the milled rice. Moreover, the increase rate of the total viable bacterial count in the irradiated rice was slower than that in the control during storage time. 4. Discussion We observed the effects of EBI on quality properties and storability of brown rice and milled rice. Color difference, pasting properties, relative crystallinity, apparent amylose contents, and microstructures were determined, along with the free fatty acids, lipase activities, and total viable bacterial count during storage. In choosing a rice product, consumers consider color as an important factor in their decision because color provides them with a first impression of the product. The color difference results in this study indicate that the EBI in the test dosage range did not affect the visual appearance of brown rice. For milled rice, the color values did not change significantly after irradiation at low doses (P > 0.05). However, at a relatively high irradiation dose, i.e., 5 kGy, the brightness and redegreen values declined. A certain degree of nonenzymatic browning reaction in rice leads to the deepening of color. As for consumers’ preference for milled rice color, the dose should be controlled below 5 kGy when milled rice is irradiated. The changes in pasting properties were mainly related to the characteristics of starch, such as the swelling property, ratio of straight chain starch, degree of gelatinization, water absorption capacity, and polymerization degree of starch granules after rupture (Amagliani et al., 2016). The increased viscosity during the heating of starch suspension is mainly caused by the swelling of starch granules, whereas the breakdown of viscosity is due to the rupture of swollen granules (Rani and Bhattacharya, 1995). The effects of various doses of irradiation on starch have been shown to include depolymerization and degradation, which often result in decreased viscosity (Liu et al., 2012b). The decrease of pasting viscosity after EBI could be attributed to the damage of starch granules and the scission of starch chains caused by free radicals generated by EBI (Hallman, 2013). Reduced pasting viscosity could yield positive effects, such as delayed retrogradation of starch, ease of cooking, and increased water absorption during cooking (Sirisoontaralak and Noomhorm, 2006). Furthermore, detrimental effects on end-use quality, such as increased total solids in cooking

water and decreased hardness of cooked rice, may be observed. However, in the current work, the effects of irradiation on longterm storage was minimal and showed a decreasing trend with storage time (Sirisoontaralak and Noomhorm, 2007). The taste value of irradiated rice was discussed in our previous paper (Yang and Luo, 2017). The results showed that although the taste value of the samples after EBI decreased slightly, the decline rate of their taste value was slower than that of the non-irradiated samples during storage. Therefore, given the comprehensive factors, the effects of EBI on the end-use quality of rice during storage have more benefits than defects. The results obtained by X-ray diffraction are consistent with those of Ocloo et al. (2016) and Liu et al. (2012a). After EBI, the relative crystallinity of brown rice starches and milled rice starch gradually decreased with the increasing dose. The degree of crystallinity in the starch granules is due to the long chains of amylopectin; the amylose chains are responsible for amorphous regions (Pandey et al., 2016). Thus, the structure of branched amylopectin chains is affected by EBI doses. This relation can explain the changes in the amylose content of rice after irradiation. The effects of EBI on the amylose content of the samples in the current work are the same as the tendencies found by Atrous et al. (2015) and Yu and Wang (2007a). These results may be due to the severe degradation of amylose fractions. This degradation reduces the iodine binding ability of amylose under low amylose contents (Gani et al., 2012). This result may be explained by the free radicals that are produced by irradiation, destroy part of the starch macromolecules, and cause the rupture of some starch particles. The texture of rice is obviously affected by amylose content. Amylose content affects the viscosity change during rice gelatinization (Zhou et al., 2015). Rice with high amylose content has high reduction value, recovery value, hot paste viscosity, and final viscosity. Specifically, rice flour is difficult to gelatinize, and rice texture is hard, features a slag feel, and lacks elasticity when chewed. On the contrary, rice with low amylose content has low reduction value, recovery value, hot paste viscosity, and final viscosity. Specifically, rice has a low retrogradation value, and the rice texture is sticky, features no slag feel, and is elastic when chewed. The relationship between amylose content and the highest viscosity value and disintegration value is not a simple linear one. When the amylose content is low, the highest viscosity and disintegration value increase with increasing amylose content. When the amylose content is higher than a certain amount, the increase in amylose decreases the highest viscosity and disintegration value (Tao et al., 2019). The resulting SEM images in the current work differ from those of Yu and Wang (2007b), who observed that the starch granules were destroyed apparently by gamma irradiation at 5, 8, and 10 kGy. The irradiation dose (
5 kGy) in this experiment may not be sufficiently high to visibly destroy starch. EBI penetration may also be weaker than that of gamma ray irradiation (Calado et al., 2014). However, most studies showed that irradiation does not

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Fig. 3. (A) Free fatty acids of brown rice during storage; (B) free fatty acids of milled rice during storage; (C) lipase activity of brown rice during storage; (D) lipase activity of milled rice during storage; (E) total viable bacterial count of brown rice during storage; (F) total viable bacterial count of milled rice.

cause evident damage to starch. Abu et al. (2006) found that up to 50 kGy irradiation did not present any visible physical effect on cowpea starch granules under SEM (2500 microphotographs). Similar results have been found in the irradiation of Indian horse chestnut starch (Wani et al., 2014). This result indicates the positive effects of irradiation on rice preservation.

Free fatty acid quantities are commonly used as an index of rice quality deterioration during storage because lipid dissolution progresses faster than protein and starch dissolution does (Genkawa et al., 2008). Changes in fatty acid profiles and increased free fatty acids of rice are generally observed during storage. Free fatty acid quantities increase during storage and bind with amylose in

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starch, and this increase in fatty acideamylose complexes may have detrimental effects on the end-use quality of rice (Park et al., 2012). China's national standard stipulates the range of free fatty acid quantities for stored rice. Fatty acid quantities should preferably be lower than 25 mg KOH/100 g to ensure the quality of rice (GAQS, 2006). Rice in the storage process is known to generate free radicals. These free radicals are responsible for lipid degradation and oxidation and produce unpleasant odors from volatile compounds. Such changes limit the shelf life of products (Park et al., 2012). In general, the safe level of free fatty acid in rice is 25 mg KOH/100 g. Beyond this value, rice begins to degenerate (Yasumatsu et al., 2014). Lipases are enzymes that are commonly known to catalyze the release of free fatty acids from acyl fats by cleavage of acyl ester bonds (Rose and Pike, 2006). Lipase is the first enzyme involved in the reaction of lipolytic metabolism. Lipases are generally believed to play a role in regulating the transformation rate of fat and is a main cause of free fatty acid rancidity in rice storage (Goffman and Bergman, 2003). In rice, lipases continuously hydrolyze lipids to produce free fatty acids. This result increases the amount of free fatty acids in rice (Li et al., 2016). In the present study, the lipase activities of the irradiated brown and milled rice were lower than those of the nonirradiated samples. This result confirms the inference that EBI inhibits lipase activities (Fig. 3). The lipase activity decreased with prolonged storage time. This observation may explain why the rate of increase in free fatty acids diminished during the storage process. The lipase activity in irradiated rice was inhibited from our experiments. As a result, the reaction rate was reduced, and the deterioration of rice quality could be delayed. Fig. 3 shows that EBI significantly inhibited the increase in the total number of stored rice colonies. High irradiation doses resulted in low total viable bacterial counts. This finding is consistent with previous research results (Chen et al., 2016a,b). Therefore, EBI can effectively extend the storage period of brown and milled rice. We also noticed that EBI exerted a minimal effect on the total viable bacterial count of brown rice at 1 kGy dose. We deduced that this phenomenon is due to the different sensitivities of various molds, bacteria, and spores toward irradiation. In similar studies conducted 52 years ago, some specific pest adults were found to require an irradiation dose of >1 kGy to prevent reproduction (Cogburn et al., 1966); a greater dose may be necessary for specific microorganisms. Unlike milled rice, brown rice has an external layer of aleurone protection, thereby complicating the penetration of irradiation. Therefore, brown rice may need a higher irradiation dose (greater than 1 kGy) than milled rice does to inhibit the growth of bacteria. In conclusion, storage quality was greatly improved during storage, and the physicochemical properties of rice slightly changed after EBI treatment. Although irradiation caused color deepening in milled rice, controlling the radiation dose below 5 kGy can prevent this effect. The results obtained in this study indicate that EBI can effectively delay the deterioration of rice quality during the storage process. The proposed method can be recommended as a new means for rice storage to improve the quality of brown and milled rice and extend their shelf life. However, further study is necessary to determine the mechanisms behind the observed changes in the physical and chemical properties of rice after EBI treatment. Acknowledgements National Key R&D Program of China (2017YFD0401200), Jiangsu Agriculture Science and Technology Innovation Fund CX(17)1003, National first-class discipline program of Food Science and Technology (JUFSTR20180203), Open Foundation of Beijing Advanced Innovation Center for Food Nutrition and Human Health

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