Method for preparing instant rice by far infrared combined solid state fermentation

Abstract

The application provides a far infrared combined solid-state fermentation method for preparing instant rice, and relates to the technical field of food processing. The method comprises the following steps: preparing an aspergillus oryzae spore suspension; irradiating a colored rice sample with far infrared rays for 470-490 s and then cooling the colored rice sample to room temperature to obtain a colored rice treatment product; the irradiation temperature of the far infrared rays is 150-170 DEG C; mixing the colored rice treatment product with nutritional ingredients, adding the aspergillus oryzae spore suspension, and then performing solid-state fermentation to obtain nutritional rice by drying the fermentation product at low temperature. The method can significantly shorten the ripening time of the colored rice, improve the total phenol content and total flavonoid content of the colored rice, and delay the increase of the fatty acid value and peroxide value of the black rice during storage.

Description

Technical Field

[0001] This invention relates to the technical field of food processing, and in particular to a method for preparing quick-cooking rice using far-infrared combined solid-state fermentation. Background Technology

[0002] Hulled colored rice, such as black rice, has a high fat content. During storage, the oil inside the black rice is prone to rancidity, producing an off-odor and reducing its shelf life. This rancidity is caused by lipase hydrolysis, producing free fatty acids that lead to hydrolytic rancidity. These fatty acids act as substrates for lipoxygenase, producing primary oxidation products called lipid hydroperoxides. These hydroperoxides are then acted upon by cellular enzymes, including peroxidase and polyphenol oxidase, producing reactive oxygen species (ROS). These ROS attack the free fatty acids, generating secondary oxidation products, which cause the black rice to become rancid during storage.

[0003] Meanwhile, colored rice is difficult to cook thoroughly, mainly because the gaps between starch granules are small, allowing water to penetrate the rice grains more slowly, resulting in incomplete starch gelatinization and longer cooking times. Furthermore, the outer layer of colored rice is hard and contains more hemicellulose, cellulose, and lignin, making it difficult to soften during cooking and resulting in a hard texture that affects taste. Therefore, it is necessary to provide a solution to improve these problems. Summary of the Invention

[0004] The purpose of this invention is to provide a method for preparing quick-cooking rice using far-infrared combined solid-state fermentation, which can significantly shorten the cooking time of colored rice, increase the total phenolic and total flavonoid content of colored rice, and delay the increase in fatty acid value and peroxide value of colored rice during storage.

[0005] This invention provides a method for preparing quick-cooking rice using far-infrared combined solid-state fermentation, which adopts the following technical solution and includes the following steps:

[0006] Preparation of Aspergillus oryzae spore suspension;

[0007] The colored rice sample was irradiated with far-infrared rays for 470-490 seconds and then cooled to room temperature to obtain the colored rice processed product; the irradiation temperature of the far-infrared rays was 150-170℃.

[0008] After mixing colored rice products with nutrients, Aspergillus oryzae spore suspension is added for solid-state fermentation. The fermentation product is then dried at low temperature to obtain nutritious rice.

[0009] The beneficial effects of the far-infrared combined solid-state fermentation method for preparing quick-cooking rice provided by this invention are as follows: Irradiating colored rice samples with far-infrared rays causes some starch granules to gelatinize and degrade into small molecules, providing a basis for the growth of Aspergillus oryzae. Simultaneously, far-infrared treatment enhances the fermentation effect of Aspergillus oryzae, promotes starch hydration, increases water absorption, improves cooking performance, and significantly reduces cooking time. Furthermore, the activities of α-amylase, protease, cellulase, carboxymethyl cellulase, and pectinase in the prepared quick-cooking rice are increased by 7.03 times, 9.10 times, 3.91 times, 22.82 times, and 2.45 times, respectively, compared to untreated black rice. It also improves the utilization rate of nutrients in colored rice, mitigates its poor storage resistance and difficulty in cooking, and delays the increase in fatty acid value and peroxide value during storage.

[0010] Optionally, the step of preparing the Aspergillus oryzae spore suspension includes: inoculating Aspergillus oryzae into a culture medium and culturing it to obtain the Aspergillus oryzae spore suspension.

[0011] Optionally, the Aspergillus oryzae spore suspension contains 2-3 × 10⁻⁶ Aspergillus oryzae spores. 6 cfu / mL.

[0012] Optionally, during the solid-state fermentation process of mixing colored rice with nutrients and adding Aspergillus oryzae spore suspension, the ratio of the volume of Aspergillus oryzae spore suspension to the weight of colored rice is controlled to be 0.05-0.15 L / kg.

[0013] Optionally, when inoculating the culture component with Aspergillus oryzae, the selected Aspergillus oryzae is Aspergillus oryzae M6, which was deposited at the China Center for Type Culture Collection on April 12, 2023, with accession number CCTCC NO: M2023533, and the deposit address is Wuhan University, Wuhan, China.

[0014] Optionally, during the solid-state fermentation process of mixing colored rice with nutrients and adding Aspergillus oryzae spore suspension, the Aspergillus oryzae can simultaneously metabolize α-amylase, protease, cellulase, carboxymethyl cellulase and pectinase during the solid-state fermentation process.

[0015] Optionally, the colored rice sample is at least one of black rice, purple rice, and red rice; when the colored rice sample is black rice, during the process of mixing the black rice treatment product with nutrients and adding Aspergillus oryzae spore suspension for solid-state fermentation, the activities of α-amylase, protease, cellulase, carboxymethyl cellulase, and pectinase in Aspergillus oryzae during solid-state fermentation reach 334.5 U / g, 350.9 U / g, 7.5 U / g, 22.8 U / g, and 36.1 U / g, respectively.

[0016] Optionally, during the step of mixing the colored rice product with nutrients and then adding a suspension of Aspergillus oryzae spores for solid-state fermentation, the ratio of Aspergillus oryzae colony-forming units in the Aspergillus oryzae spore suspension to the weight of the colored rice product is controlled to be 2-3 × 10⁻⁶. 8 cfu / kg.

[0017] Optionally, the process of mixing the colored rice product with nutrients and then adding Aspergillus oryzae spore suspension for solid-state fermentation also includes: controlling the ambient temperature of the solid-state fermentation environment to be 36-38℃, the ambient humidity to be 88%-92%, and the solid-state fermentation time to be 70-74h.

[0018] Optionally, when performing the process of mixing colored rice products with nutrients and then adding Aspergillus oryzae spore suspension for solid-state fermentation, the solid-state fermentation product is turned over every 24 hours, and the total number of times it is turned over is controlled to be 2-3 times. Attached Figure Description

[0019] Figure 1 This is a flowchart illustrating the steps of a method for preparing quick-cooking rice using far-infrared combined solid-state fermentation, as described in an embodiment of the present invention.

[0020] Figure 2 A is the X-ray diffraction pattern of black rice in Comparative Example 1 and quick-cooked black rice in Comparative Example 1 and Example 1 of the present invention;

[0021] Figure 2 B is the infrared spectrum of black rice in Comparative Example 1 and quick-cooked black rice in Comparative Example 1 and Example 1 of the present invention;

[0022] Figure 3 A1-A3 are appearance diagrams of black rice in Comparative Example 1 and quick-cooked black rice in Comparative Example 1 and Example 1, respectively.

[0023] Figure 3 B1-B3 are scanning electron microscope images of black rice in Comparative Example 1 and quick-cooked black rice in Comparative Example 1 and Example 1, respectively.

[0024] Figure 3 C1-C3 are microscopic images of black rice in Comparative Example 1 and quick-cooked black rice in Comparative Example 1 and Example 1 after staining, taken using a laser confocal microscope.

[0025] Figure 4 A is a graph showing the change in fatty acid value of black rice in Comparative Example 1 and black rice in Comparative Example 1 and Example 1 during accelerated storage;

[0026] Figure 4 B is a graph showing the change in peroxide value of black rice in Comparative Example 1 and black rice in Comparative Example 1 and Example 1 during accelerated storage. Detailed Implementation

[0027] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below. Obviously, the described embodiments are only some embodiments of the present invention, not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention. Unless otherwise defined, the technical or scientific terms used herein should have the ordinary meaning understood by those skilled in the art. The terms "comprising" and similar expressions used herein mean that the element or object preceding the word covers the element or object listed after the word and its equivalents, but does not exclude other elements or objects.

[0028] See Figure 1 This invention provides a method for preparing high-quality rice using far-infrared combined solid-state fermentation, comprising the following steps:

[0029] S1. Prepare a suspension of Aspergillus oryzae spores;

[0030] S2. After treating the colored rice sample with far-infrared irradiation for 470-490s, cool it to room temperature to obtain the colored rice treated product; wherein the far-infrared irradiation temperature is 150-170℃.

[0031] S3. After mixing the colored rice product with nutrients, add Aspergillus oryzae spore suspension for solid-state fermentation, and dry the fermentation product at low temperature to obtain quick-cooking rice.

[0032] In some embodiments, when performing the method steps for preparing quick-cooking rice using far-infrared combined solid-state fermentation, step S2 can be performed first, followed by step S1, or steps S1 and S2 can be performed simultaneously.

[0033] In some embodiments, the process of performing step S1 includes step S11: inoculating Aspergillus oryzae into the culture component and culturing to obtain an Aspergillus oryzae spore suspension.

[0034] In some further embodiments, before performing step S11, step S10 is performed. S10: *Aspergillus oryzae* M6, which was deposited on April 12, 2023, at the China Center for Type Culture Collection (CCTCC), Wuhan University, Wuhan, China, with accession number CCTCC NO: 2023533, is selected. This *Aspergillus oryzae* M6 (CCTCC NO: 2023533) can simultaneously metabolize α-amylase, protease, cellulase, carboxymethyl cellulase, and pectinase during solid-state fermentation.

[0035] In some further embodiments, the culture medium used in step S11 is potato dextrose agar. In fact, in other embodiments, the culture medium can be any other culture medium suitable for the growth and reproduction of *Aspergillus oryzae*.

[0036] In some further embodiments, when performing step S11, Aspergillus oryzae is inoculated into the culture medium and cultured at 36-38°C for 4-5 days.

[0037] In some further embodiments, during step S11, after inoculating Aspergillus oryzae into the culture medium, the Aspergillus oryzae spore concentration is adjusted so that the Aspergillus oryzae spore suspension obtained contains 2-3 × 10⁻⁶ Aspergillus oryzae. 6 cfu / mL.

[0038] In some embodiments, when performing step S2, the colored rice sample can be at least one of black rice, purple rice, and red rice, and after performing step S2, the corresponding colored rice processed product, such as black rice processed product, purple rice processed product, and red rice processed product, is obtained.

[0039] In some embodiments, the nutrient medium used in step S3 is potato dextrose broth. In other embodiments, the nutrient medium can be any other liquid culture medium suitable for the growth and reproduction of Aspergillus oryzae.

[0040] In some further embodiments, when performing step S3, the colored rice treatment product is mixed with potato glucose broth medium and left for 18 hours to ensure that the potato glucose broth medium and the colored rice treatment product are fully mixed and achieve nutrient balance.

[0041] In some further embodiments, during step S11, after inoculating Aspergillus oryzae into the culture medium, the Aspergillus oryzae spore concentration is adjusted so that the Aspergillus oryzae spore suspension obtained contains 2-3 × 10⁻⁶ Aspergillus oryzae. 6 cfu / mL; In step S3, after balancing the colored rice treatment with potato glucose broth medium, Aspergillus oryzae spore suspension is added for solid-state fermentation, and the ratio of the volume of Aspergillus oryzae spore suspension to the weight of colored rice treatment is 0.05-0.15 L / kg.

[0042] In some further embodiments, during step S3, after nutrient balancing of the colored rice treatment product and potato glucose broth culture medium, Aspergillus oryzae spore suspension is added for solid-state fermentation, such that the ratio of Aspergillus oryzae colony-forming units in the Aspergillus oryzae spore suspension to the weight of the colored rice treatment product is 2-3 × 10⁻⁶. 8 cfu / kg.

[0043] In some embodiments, when performing step S3, the ambient temperature of the solid-state fermentation environment is controlled at 36-38°C, the ambient humidity is controlled at 88%-92%, and the solid-state fermentation time is controlled at 70-74 hours.

[0044] In some further embodiments, when performing step S3, the solid fermentation material is turned over every 24 hours, and the total number of times it is turned over is controlled to be 2-3 times.

[0045] Isolation of Aspergillus oryzae M6 (CCTCC NO: 2023533): Using a sterile spatula, 1g of naturally fermented soybean paste was mixed with 9mL of 0.9% physiological saline solution and stirred to dilute. After shaking, 1mL of the diluted soybean paste was mixed with 9mL of physiological saline solution and diluted to a dilution of 10. -2 The soybean paste dilution was prepared simultaneously with a dilution of 10. -3 10 -4 10 -5 10 -6 Diluent, and take 200 μL of 10 -3 10 -4 10 -5 10 -6 The diluted solution was evenly spread on Bengal Red agar medium containing chloramphenicol using a sterile spreader. After incubation at 28°C for 96 hours, typical mold single colonies were selected and streaked onto potato dextrose agar solid medium. After purification 2-3 times until single colonies were obtained, the purified mold strains were streaked onto potato dextrose agar solid slant medium to obtain Aspergillus oryzae strains for low-temperature preservation.

[0046] Mutagenesis: After the isolated Aspergillus oryzae strain was cultured to maturity, the Aspergillus oryzae plates were washed with sterile water to prepare a homogeneous spore suspension, and the concentration of the Aspergillus oryzae spore suspension was adjusted to 10. 8 CFU / mL, then 10 mL of spore suspension was evenly spread on the slide. The mutagenesis conditions were set as follows: aeration rate of 10 SLM, power of 100 W, and time of 0 s, 15 s, 30 s, 45 s, 60 s, 75 s, 90 s, and 120 s, respectively. After completing the atmospheric pressure and room temperature plasma mutagenesis treatment, the slide was washed in PBS and then resuspended in sterile plasma water for later use.

[0047] Screening: The mutagenized Aspergillus oryzae suspension was spread on casein agar and incubated at 30°C for 72 hours in the dark. Single colonies exhibiting vigorous mycelial growth and large clear zones compared to the unmutated Aspergillus oryzae strain were streaked onto soybean protein agar and incubated at 30°C for 48 hours. Single colonies with large diameters and vigorous growth were then selected and transferred to soybean protein agar for maturation and preservation. The strain was then activated and cultured to produce 1×10⁻⁶ cells / years. 6A spore suspension of CFU / mL was prepared and placed on a solid-state fermentation medium. Solid-state fermentation was carried out at 30℃ for 72 hours, and the activities of amylase, protease, cellulase, carboxymethyl cellulase, and pectinase were measured. Strains with normal growth and high enzyme activity were selected for preservation. The casein medium consisted of 10 g / L casein, 10 g / L beef extract, 2 g / L disodium hydrogen phosphate, 5 g / L sodium chloride, 15 g / L agar powder, and 1 L deionized water. The raw materials were autoclaved at 121℃ for 20 min. The soybean protein medium consisted of 10 g soybean protein isolate, 0.5 g MgSO4·7H2O, 30 g xylose, 1 g KH2PO4, and 15 g agar powder, dissolved in 1000 mL distilled water. After adjusting the pH to 6.4, the solution was sterilized at 121℃ for 15 min. The solid-state fermentation medium consisted of 97% cooked black rice and (NH4)2SO4. 3%, adjust the solid-to-water ratio to 1:0.8, and sterilize at 121 degrees Celsius for 15 minutes;

[0048] Identification: The bacterial strain was sent to Beijing Qingke Biotechnology Co., Ltd. for gene identification, and the gene sequence of Aspergillus oryzae M6 was obtained as follows:

[0049] .

[0050] Example 1

[0051] This embodiment 1 provides a method for preparing quick-cooking rice using far-infrared combined solid-state fermentation, including the following steps:

[0052] S1. Inoculate Aspergillus oryzae M6 (CCTCC NO: 2023533) into potato dextrose agar medium and incubate at 37°C for 4-5 days. Adjust the Aspergillus oryzae spore concentration to obtain an Aspergillus oryzae content of 2-3 × 10⁻⁶. 6 CFU / mL Aspergillus oryzae M6 spore suspension;

[0053] S2. After irradiating the colored rice with far-infrared rays for 470-490 seconds, cool it to room temperature to obtain black rice processed product; wherein the far-infrared irradiation temperature is 160℃.

[0054] S3. Take 10 kg of black rice treatment product and mix it with 2.5 L of potato glucose broth culture medium. After 18 h of nutrient balance, add 1 L of Aspergillus oryzae M6 spore suspension and stir evenly. Place it in a fermentation environment with a temperature of 37℃ and a humidity of 90% for solid fermentation for 72 h. During the fermentation process, turn the solid fermentation material once every 24 h. After the fermentation is completed, dry it at low temperature to obtain quick-cooking black rice.

[0055] Comparative Example 1

[0056] Comparative Example 1 provides a method for preparing nutrient-rich rice through solid-state fermentation, comprising the following steps:

[0057] S1. Inoculate Aspergillus oryzae M6 (CCTCC NO: 2023533) into potato dextrose agar medium and incubate at 37°C for 4-5 days. Adjust the Aspergillus oryzae spore concentration to obtain an Aspergillus oryzae content of 2-3 × 10⁻⁶. 6 CFU / mL Aspergillus oryzae M6 spore suspension;

[0058] S2. Take 10 kg of black rice and mix it with 2.5 L of potato glucose broth culture medium and balance the nutrients for 18 h. Then add 1 L of Aspergillus oryzae M6 spore suspension and stir evenly. Place it in a fermentation environment with a temperature of 37℃ and a humidity of 90% for solid fermentation for 72 h. During the fermentation process, turn the solid fermentation material once every 24 h. After the fermentation is completed, dry it at low temperature to obtain quick-cooking black rice.

[0059] Compare with Example 1

[0060] Comparative Example 1 is untreated black rice, and the type and batch of black rice are the same as those in Example 1 and Comparative Example 1.

[0061] Performance testing and analysis

[0062] (1) Determination of hydrolytic enzyme activity, soluble protein and polysaccharide

[0063] Following the methods of Chen G, Liu Y, Zeng J, et al. Enhancing three phenolic fractions of oats (Avena sativa L.) and their antioxidant activities by solid-state fermentation with Monascus anka and Bacillus subtilis[J]. Journal of Cereal Science, 2020, 93: 102-940. and Li WY, Zhao LL, He XL. Degradation potential of different lignocellulosic residues by Trichoderma longibrachiatum and Trichoderma afroharzianum under solid state fermentation[J]. Process Biochemistry, 2022, 112: 6-17., α-amylase, protease, cellulase, and pectinase in the samples were extracted and determined.

[0064] The sample was crushed and passed through a 60-mesh sieve. 1.0 g of black rice flour was weighed and added to 20 mL of sodium citrate buffer (50 mM, pH 5.5). The mixture was extracted at 30 °C for 30 min and then centrifuged at 4 °C for 10 min. The supernatant was collected as the crude enzyme solution of the fermentation substrate, and its enzyme activity was immediately measured.

[0065] α-Amylase activity: Take 250 μL of diluted crude enzyme solution, add 1.75 mL of 1% soluble starch solution (w / v), react in a 50℃ water bath for 10 min, then immediately add 4 mL of DNS reagent and boil in a water bath for 5 min, and finally bring the volume to 10 mL. After the sample cools to room temperature, measure the absorbance at 540 nm, and simultaneously plot a standard curve using glucose as a standard via the DNS method. Enzyme activity unit (U) is defined as the amount of enzyme required to hydrolyze starch to produce 1 μmol of glucose per minute under experimental conditions, and is expressed as U / g.

[0066] Protease activity: Mix 2 mL of crude enzyme solution with 2 mL of 1% casein solution, react in a 40℃ water bath for 15 min, stop the reaction by adding 4 mL of 0.4 mol / L trichloroacetic acid solution, let stand for 30 min, and then centrifuge at 12000 r / min for 5 min. Collect the supernatant and measure the absorbance at 280 nm, and plot a standard curve using tyrosine standards. Enzyme activity unit (U) is defined as the amount of enzyme required to hydrolyze casein to produce 1 μg of tyrosine per minute under experimental conditions, and is expressed as U / g.

[0067] Cellulase activity: Take 1 mL of diluted crude enzyme solution, add 2 mL of sodium citrate buffer (0.1 mol / L, pH 4.8), and add 0.1 g of filter paper strip (1.0 × 6.0 cm) and 1 mL of 0.5% sodium carboxymethyl cellulose solution as reaction substrates. Measure the activities of filter paper cellulase (FPase) and carboxymethyl cellulase (CMCase). After reacting in a 50℃ water bath for 1 h, immediately add 4 mL of DNS reagent and react in a boiling water bath for 10 min, then bring the volume to 10 mL. After the sample cools to room temperature, measure the absorbance at 540 nm. Simultaneously, use glucose as a standard to plot a standard curve using the DNS method. Enzyme activity unit (U) is defined as the amount of enzyme required per minute to hydrolyze filter paper to produce 1 μmol of glucose under experimental conditions, expressed as U / g.

[0068] Pectinase activity: Take 1 mL of diluted crude enzyme solution, add 2 mL of sodium acetate buffer (0.04 mol / L, pH 4.6) and 1 mL of 1% pectin solution, react in a 50℃ water bath for 30 min, then immediately add 4 mL of DNS reagent and react in a boiling water bath for 10 min, and then bring the volume to 10 mL. After the sample cools to room temperature, measure the absorbance at 540 nm, and simultaneously plot a standard curve using D-galacturonic acid as a standard using the DNS method. Enzyme activity unit (U) is defined as the amount of enzyme required to hydrolyze pectin to produce 1 μmol of galacturonic acid per minute under experimental conditions, expressed as U / g.

[0069] The soluble proteins and polysaccharides in the samples were determined according to the method described in Shen D, Labreche F, Wu C, et al. Preparation and aroma analysis of flavonoid-rich ginkgo seeds fermented using rice wine starter[J]. FoodBioscience, 2021, 44: 101-459.

[0070] Soluble protein content: 1.0 g of rice flour was mixed with 0.05 mol / L NaOH solution (1:30 w / v), stirred in a water bath at 35 ℃ for 1.5 h, centrifuged, and the supernatant was brought to a final volume of 50 mL. Bovine serum albumin solutions of different concentrations were prepared, and the soluble protein content was determined using the Coomassie brilliant blue method.

[0071] Soluble polysaccharide content: 1.0g of rice flour was mixed with water (1:30w / v), stirred in a water bath at 35℃ for 1.5h, centrifuged and the supernatant was collected. Different concentrations of glucose solutions were prepared and the soluble polysaccharide content was determined by the phenol-sulfuric acid method.

[0072] All the above measurements were repeated three times. The data were analyzed by one-way ANOVA (Duncan's test) using SPSS 25.0. The significance level was p<0.05. The results are expressed as mean ± standard deviation.

[0073] The enzyme activity and soluble components of the quick-cooked black rice in Example 1 and Comparative Example 1 are shown in Table 1.

[0074] Table 1. Enzyme activity and soluble components in quick-cooked black rice and black rice.

[0075]

[0076] Note: Data are expressed as mean ± standard deviation. Different letters in the same row indicate significant differences (p < 0.05).

[0077] As shown in Table 1, compared with the black rice in Control Example 1, the quick-cooked black rice in Comparative Example 1, after solid-state fermentation, showed a significant increase in the activities of α-amylase, protease, cellulase, carboxymethyl cellulase, and pectinase, increasing by 3.54 times, 5.12 times, 2.86 times, 15.55 times, and 0.70 times, respectively. In contrast, the quick-cooked black rice in Example 1, after combined far-infrared irradiation and solid-state fermentation treatment, showed an increase in the activities of α-amylase, protease, cellulase, carboxymethyl cellulase, and pectinase, increasing by 7.03 times, 9.10 times, 3.91 times, 22.82 times, and 2.45 times, respectively, compared with the black rice in Control Example 1.

[0078] (2) Gelatinization and cooking properties determination

[0079] The gelatinization characteristics of the samples were determined using a rapid viscosity analyzer (RVA) in accordance with the method described in Zhong Y, Xiang X, Zhao J, et al. Microwave pretreatment promotes the annealing modification of rice starch[J]. Food Chemistry, 2020, 304: 125432.

[0080] Weigh 3g of sample powder and add it to an aluminum cylinder containing 25mL of deionized water. Place the cylinder into the RVA groove, then select the standard program to stir, heat (50~95℃) and cool. The single run time is 12.5min. Measure the gelatinization characteristics of the sample.

[0081] The cooking quality and textural properties of the samples were determined according to the method described in Wu J, Chen J, Liu W, et al. Effects of aleurone layer on rice cooking: ahistorical investigation[J]. Food Chemistry, 2016, 191: 28-35.

[0082] Take 5g of sample rice grains and place them in 50mL of boiling water (98±1℃) to steam and start timing. After 15 minutes, randomly take 10 grains of rice, press them between two clean glass plates and squeeze to observe whether there is any ungelatinized starch core inside the rice grains. When 90% of the rice grains do not have a white core (ungelatinized starch), record the steaming time. At the same time, the water absorption rate should also be measured after the rice grains have reached the cooking time.

[0083]

[0084] Where: W0 represents the weight of the sample before cooking, g; W1 represents the weight of the sample after cooking, g; W i The weight is on a dry basis, in grams.

[0085] Take cooked rice (not exceeding the probe area) that has reached the required cooking time and place it on the texture analyzer testing platform while still hot. The measurement is performed using a two-cycle compression (TPA) mode. The probe speeds before, during, and after measurement are 1.0, 0.5, and 1.0 mm / s, respectively. The rice is compressed to 75% deformation using a 26 mm diameter cylindrical probe (P / 36). Each sample is measured 10 times. The instrument's built-in Texture Expert Exceed software automatically analyzes the texture curves to obtain the sample's hardness, adhesiveness, elasticity, cohesiveness, chewiness, and resilience.

[0086] All the above measurements were repeated three times. The data were analyzed by one-way ANOVA (Duncan's test) using SPSS 25.0. The significance level was p<0.05. The results are expressed as mean ± standard deviation.

[0087] The cooking quality, textural properties, and gelatinization properties of the quick-cooked black rice in Example 1 and Comparative Example 1 are shown in Table 2.

[0088] Table 2. Cooking quality, textural properties, and gelatinization properties of quick-cooking black rice and regular black rice.

[0089]

[0090] Note: Data are expressed as mean ± standard deviation. Different letters in the same row indicate significant differences (p < 0.05).

[0091] As shown in Table 2, the cooking time of black rice in Comparative Example 1 was 31.58 min, while the cooking time of quick-cooking black rice that underwent solid-state fermentation in Comparative Example 1 was reduced to 20.68 min. Furthermore, the cooking time of quick-cooking black rice treated with a combination of far-infrared irradiation and solid-state fermentation in Example 1 was reduced to 15.38 min. This demonstrates that solid-state fermentation can reduce the cooking time of black rice, and in particular, the combined treatment of solid-state fermentation and far-infrared irradiation can significantly reduce the cooking time.

[0092] Meanwhile, the water absorption rate of black rice in Comparative Example 1 was 125.21%, while the water absorption rates of quick-cooked black rice in Comparative Example 1 and Example 1 increased to 160.90% and 205.09%, respectively. This indicates that solid-state fermentation of black rice can increase its water absorption rate during cooking. This may be because water can rapidly migrate into the interior of the cooked sample during cooking, and the hydrogen bond interaction between starch and water is enhanced during the cooking process. Furthermore, as shown in Table 1, the quick-cooked black rice in Comparative Example 1 and Example 1 exhibited higher enzyme activity, which is beneficial for decomposing the outer layer of the cooked sample, thereby increasing starch water absorption and reducing cooking time. The significantly increased water absorption rate of quick-cooked rice in Example 1 is not only due to the solid-state fermentation process but may also be due to the damage to the cell walls, cell membrane contraction, mesosome disintegration, and leakage of internal cell contents after far-infrared irradiation, which further facilitates the rapid migration of water into the interior of the black rice sample.

[0093] Furthermore, compared with the black rice in Comparative Example 1, the hardness, stickiness, elasticity, cohesiveness, chewiness, and resilience of the quick-cooked black rice in Comparative Example 1 were significantly reduced (p<0.05). These properties of the quick-cooked black rice in Example 1 were further reduced. This may be because cellulase and pectinase produced during fermentation (see enzyme activity data in Table 1) damage the fibrous cortex, and proteases hydrolyze the proteins in the black rice, reducing the protein network structure and thus affecting the textural properties of the black rice.

[0094] Furthermore, compared with the black rice in Comparative Example 1, the peak viscosity, trough viscosity, disintegration value, final viscosity, and recovery value of the quick-cooked black rice in Comparative Example 1 were significantly reduced (p<0.05), while these properties of the quick-cooked black rice in Example 1 were further reduced (p<0.05). This may be due to the amylase produced during solid-state fermentation (see data in Table 1) causing the starch granules to hydrolyze into short starch chains and small molecule sugars. At the same time, the reduction in protein content also reduces the gelatinization viscosity, and further verifies that pre-treatment with far-infrared light can promote the solid-state fermentation process.

[0095] (3) Crystal structure determination

[0096] The long-range crystal structure was determined using a wide-angle X-ray diffractometer (XRD). The diffraction intensity scan range was 5°–35° (2θ), and the scan rate was 0.02° / sec. Diffraction patterns were acquired using Jade 6.0 software, and the relative crystallinity (%) was calculated based on the ratio of the crystalline region area to the total area of ​​the diffraction pattern.

[0097] The short-range ordered structure of the sample was analyzed using Fourier transform infrared spectroscopy (FTIR). The infrared spectral scanning range was 4000–400 cm⁻¹. -1 The resolution is 4cm. -1 After 64 scans, a 1200cm diameter was observed. -1 up to 800cm -1 The spectrum within the range is subjected to baseline correction, smoothing, and deconvolution processing.

[0098] Calculate 1047 / 1022cm -1 With 1020cm -1 / 995cm -1The absorbance ratio (Method: Berti S, Jagus RJ, Flores S K. Effect of rice bran addition on physical properties of antimicrobial biocomposite films based on starch[J]. Food and Bioprocess Technology, 2021, 14(9): 1700-1711.).

[0099] The crystal structures of the quick-cooked black rice in Example 1 and Comparative Example 1 are as follows: Figure 2 As shown in Table 3, the crystallization characteristics of the quick-cooked black rice in Example 1 and Comparative Example 1 and the black rice in Control Example 1 are as follows.

[0100] Table 3 Crystallization characteristics of quick-cooking black rice and regular black rice

[0101]

[0102] Note: Data are expressed as mean ± standard deviation. Different letters in the same row indicate significant differences (p < 0.05).

[0103] See Figure 2 A. In Control Example 1, the black rice (BR) exhibited a typical starch type A crystallographic pattern, with strong diffraction peaks near 15° and 23°, a continuous unseparated doublet near 17° and 18°, and a weak diffraction peak near 20°. In contrast, the quick-cooked black rice (FBR) in Comparative Example 1 and the quick-cooked black rice (Fermented black rice stabilized by far infrared radiation) in Example 1 showed similar crystal characteristics. This indicates that solid-state fermentation or combined treatment with far infrared radiation has no effect on the crystal form of black rice starch. As shown in Table 3, the relative crystallinity of the black rice in Control Example 1 was 22.94%, while the relative crystallinity of the quick-cooked black rice in Comparative Example 1 and Example 1 decreased to 20.18% and 14.94%, respectively.

[0104] See Figure 2 As shown in B, in the infrared spectrum, 1047 cm⁻¹ -1 1020cm -1 and 995cm -1 The nearby absorption peaks correspond to the hydrogen bond structures formed between starch crystalline regions, starch amorphous regions, and hydroxyl groups, respectively, at (1047 / 1020) cm⁻¹. -1 and (1020 / 995)cm-1 The peak intensity ratio can reflect the degree of order on the surface of starch granules, i.e., (1047 / 1020) cm. -1 The larger the peak intensity ratio, the higher the (1020 / 995)cm -1 The smaller the peak intensity ratio, the more ordered the short-range structure of the starch. Compared with black rice (BR) in Comparative Example 1, the quick-cooked black rice (FBR) in Comparative Example 1 and the quick-cooked black rice (FFIR stabilized by far infrared radiation) in Example 1 have a peak intensity ratio of (1047 / 1020) cm⁻¹. -1 The ratio gradually decreases, reaching (1020 / 995) cm. -1 The ratio gradually increases, indicating that the short-range order of the quick-cooked black rice in Comparative Example 1 and Example 1 is less than that in Control Example 1, suggesting that solid-state fermentation or combined treatment with far-infrared irradiation can reduce the short-range structural order of black rice.

[0105] (4) Microstructure determination

[0106] The sample was broken by applying pressure along the middle of the sample with a blade, and the sample was fixed to the base with the broken surface facing upwards using conductive double-sided tape. The surface was then sprayed with gold and measured using a scanning electron microscope (SEM) at an accelerating voltage of 5 kV, with a magnification of 1500x.

[0107] Furthermore, following a slight modification of the method described by Huang K, Zhang S, Guan X, et al., Effect of the oat β-glucan on the development of functional quinoa (Chenopodium quinoa wild) milk[J]. Food Chemistry, 2021, 349:129-201, the fiber in black rice was determined. The method is as follows: 100 mg of the pulverized sample was weighed, and fluorescent whitening agent (1:2, m / v) was added for staining for 1 h. Excess dye was washed away with distilled water, and the sample was observed under 10x magnification using a laser confocal scanning microscope (CLSM). The excitation and emission wavelengths of the fluorescent whitening agent were 405 nm and 455 nm, respectively.

[0108] The microstructures of the quick-cooked black rice in Example 1 and Comparative Example 1 are as follows: Figure 3 As shown.

[0109] See Figure 3As shown in A1, because the bran layer of black rice is relatively dense and does not easily absorb water, and because microorganisms require fewer nutrients to grow within it, therefore, see... Figure 3 As shown in A2, the Aspergillus oryzae hyphae in Comparative Example 1 did not easily extend into the interior of the black rice, and Aspergillus oryzae growth was relatively low; see also Figure 3 As shown in A3, Aspergillus oryzae in Example 1 multiplied in large quantities, and the Aspergillus oryzae hyphae extended into the interior of the black rice grains. This is because after the black rice was treated with far-infrared irradiation, some of the starch granules were gelatinized and degraded into small molecules, which provided the black rice with the carbon source required for microbial growth. At the same time, after the treatment with far-infrared irradiation, the water absorption of the black rice increased (see the water absorption rate data in Table 2), which provided a basis for the growth of Aspergillus oryzae. In addition, the hydrolytic enzymes produced by Aspergillus oryzae during its growth and metabolism can hydrolyze the large molecules in the black rice into small molecules, which become the carbon and nitrogen sources for its own growth.

[0110] See Figure 3 As shown in B1, black rice has a relatively dense structure; see also... Figure 3 As shown in B2, the black rice sample in Comparative Example 1, after solid-state fermentation with Aspergillus oryzae, exhibited a porous structure; see also... Figure 3 As shown in B3, the black rice sample treated with a combination of far-infrared radiation and solid-state fermentation in Example 1 exhibited the most porous structure. This is mainly because Aspergillus oryzae reduced the enzymatic activity of cell walls, proteins, and starch granules during solid-state fermentation. Simultaneously, far-infrared radiation treatment enhanced the fermentation effect of Aspergillus oryzae, and the far-infrared treatment itself also caused cell wall damage in the black rice, thus facilitating starch hydration, increasing water absorption, and making the starch granules easier to gelatinize, thereby improving cooking performance (as shown in Table 2).

[0111] See Figure 3 As shown in C1, the fibrous structure of black rice is dense, and its fluorescent staining is relatively concentrated; see also Figure 3 As shown in C2, after solid-state fermentation, the black rice in Comparative Example 1 showed a gradual loosening of fiber and the appearance of more fiber fragments. This may be because the cellulase and pectinase produced during solid-state fermentation can degrade the cell wall, resulting in a looser fiber structure and partial fiber degradation. See also... Figure 3 As shown in C3, the black rice sample from Example 1, after being treated with a combination of far-infrared and solid-state fermentation, showed higher fiber distribution and more fiber fragments, which further verifies that far-infrared can enhance the fermentation effect of solid-state fermentation.

[0112] (5) Nutritional quality determination (total phenols, total flavonoids and DPPH antioxidant properties determination)

[0113] The total phenols, total flavonoids, and antioxidant activity were determined according to the method described in Zeng Z, Luo S, Liu C, et al. Phenolic retention of brown rice after extrusion with mesophilic α-amylase[J]. Food Bioscience, 2018, 21: 8-13.

[0114] Polyphenol extraction: Accurately weigh 1.00 g of black rice flour, add 30 mL of acidified ethanol (95% ethanol: 1 mol / L HCl = 85:15, v / v), mix and extract for 30 min, centrifuge at 3000 r / min for 5 min, collect the supernatant, and repeat the extraction 3 times. Combine the supernatants, evaporate at 45℃, and re-dilute to 10 mL with 50% chromatographic methanol, and store at -20℃ for analysis within one week.

[0115] Total phenol content was determined using the Folin-Ciocalteu method, and the results are expressed as micrograms of gallic acid per gram of dry weight of sample (μg GAE / g DW).

[0116] The total flavonoid content was determined by the AlCl3 method, and the results are expressed as micrograms of catechin per gram of dry weight of sample (μgCE / gDW).

[0117] Its antioxidant capacity was evaluated by free radical scavenging activity (DPPH and ABTS assays) and iron ion reducing power (FRAP assay). The antioxidant capacity results were expressed as μg Trolox equivalent per gram of dry weight sample (μg Trolox / g DW).

[0118] The polyphenol composition and antioxidant properties of the quick-cooked black rice in Example 1 and Comparative Example 1 are shown in Table 4.

[0119] Table 4 Nutritional quality of quick-cooking black rice and regular black rice

[0120]

[0121] Note: Data are expressed as mean ± standard deviation. Different letters in the same row indicate significant differences (p < 0.05).

[0122] As shown in Table 4, compared with the black rice in Control Example 1, the total phenol and total flavonoid content of the quick-cooked black rice after solid-state fermentation in Control Example 1 was reduced. This is mainly because the addition of water during solid-state fermentation caused the dissolution of soluble components such as anthocyanins in the black rice sample, resulting in a decrease in the DPPH antioxidant capacity of the quick-cooked black rice. Although solid-state fermentation can increase the phenols and flavonoids in the grains, the amount of active substances that may be dissolved is relatively large, and the increase in active substances by solid-state fermentation is insufficient to compensate for the dissolution loss.

[0123] As shown in Table 4, compared with the black rice in Control Example 1, the total phenolic and total flavonoid content and DPPH antioxidant capacity of the quick-cooked black rice in Example 1, which underwent combined far-infrared and solid-state fermentation treatment, were significantly increased. This is mainly because the changes in free and bound phenolic substances during grain fermentation largely depend on the various enzymes produced during microbial fermentation. In addition to soluble phenols existing in free form in cell vacuoles, some phenols in grains are bound to cell wall structures, hemicellulose, cellulose, and proteins in an insoluble form. Esterases and xylanases secreted during the growth and metabolism of Aspergillus oryzae can release bound phenolic substances, while pectinase, cellulase, and protease secreted during growth and metabolism can hydrolyze plant cell walls, hemicellulose, cellulose, and proteins, making it easier for esterases and xylanases to act on covalent bonds to release bound phenols. At the same time, the secreted α-amylase can catalyze the production of glucose during fermentation, serving as a carbon source for its own growth, thereby maximizing the release of polyphenols. Therefore, as shown in Table 1, the various enzyme activities produced by Aspergillus oryzae after the combined treatment with far-infrared and solid-state fermentation in Example 1 are stronger, which further verifies that far-infrared treatment can enhance the fermentation effect of solid-state fermentation.

[0124] (6) Determination of storage properties

[0125] The rancidity of oils in colored rice mainly occurs in two forms: hydrolysis and oxidation, which produce fatty acids and peroxides, respectively. Therefore, the degree of rancidity of oils is reflected by measuring the content of fatty acid value and peroxide value. Generally speaking, the higher the content of both, the worse the edible quality.

[0126] Accelerated storage experiment: Black rice (BR) from Control Example 1, Fermented black rice (FBR) from Comparative Example 1, and Fermented black rice stabilized by far infrared radiation (FFIR) from Example 1 were packaged in resealable bags and placed in a constant temperature and humidity chamber at 37°C and 75% relative humidity. Samples were taken at 0, 10, 20, 30, 40, and 50 days to determine their fatty acid value and peroxide value. The results are as follows: Figure 4 As shown.

[0127] Fatty acid value determination: Refer to the national standard GB / T 5510-2011 with slight modifications. The method is as follows: Take 1g of the crushed sample into a centrifuge tube, add 30mL of petroleum ether, and seal the centrifuge tube. Shake at 20℃ and 100r / min for 10min on a shaker, then centrifuge. Take 20mL of the supernatant into an Erlenmeyer flask, add 75mL of 50% ethanol and 5 drops of phenolphthalein solution. Using a microburette, titrate the lower ethanol solution with KOH standard titration solution (0.01mol / L) until a light pink color appears, continuing for about 3s. Replace the sample supernatant with 20mL of ethanol to perform a blank titration test. The result is expressed as mg / 100g (calculated as KOH), and the calculation formula is as follows:

[0128]

[0129] In the formula: c is the concentration of KOH standard titration solution (mol / L); m is the dry basis mass of the sample (g DW); V1 is the volume of KOH standard titration solution consumed by the sample (mL); V0 is the volume of KOH standard titration solution consumed by the blank (mL); 8415 is a constant represented by KOH, i.e. 56.1×1.5×100.

[0130] Peroxide value determination: The method was slightly modified from that of Wu J, McClements DJ, Chen J, et al. Improvement of storage stability of lightly milled rice using superheated steam processing[J]. Journal of Cereal Science, 2016, 71:130-137. 0.1 g of the pulverized sample was added to 9.8 mL of chloroform-methanol (7:3, v / v) solution, stirred in a 45℃ water bath for 10 min, cooled to room temperature, and centrifuged. All supernatant was diluted to 10 mL, 50 μL of ferric chloride solution was added and shaken well, followed by 50 μL of 30% potassium thiocyanate solution. After standing for 5 min, the absorbance was immediately measured at 500 nm. Using Fe... 3+ A standard curve is plotted using the solution as a standard, and the calculation formula is as follows:

[0131]

[0132] In the formula: A s A represents the absorbance of the sample. b λ is the blank absorbance; m is the reciprocal of the slope of the standard curve; m0 is the dry basis mass of the sample (g DW); 55.84 is the relative atomic mass of iron (g / mol).

[0133] See Figure 4As shown, during storage, the fatty acid values ​​of the three samples (e.g.) Figure 4 A) and peroxide value (e.g.) Figure 4 B) All values ​​gradually increased, with the fatty acid and peroxide values ​​of black rice (BR) in Control Example 1 increasing the fastest, followed by quick-cooked black rice (FBR) in Control Example 1. This may be because the organic acids produced by Aspergillus oryzae during fermentation inhibited lipase activity, thereby reducing the increase in fatty acid value. Among the three samples, the fatty acid and peroxide values ​​of quick-cooked black rice (FSS) in Example 1 increased the slowest, indicating that far-infrared combined solid-state fermentation can effectively improve the storage stability of black rice.

[0134] While embodiments of the present invention have been described in detail above, it will be apparent to those skilled in the art that various modifications and variations can be made to these embodiments. However, it should be understood that such modifications and variations fall within the scope and spirit of the invention as set forth in the claims. Furthermore, the invention described herein may have other embodiments and can be implemented or carried out in various ways.

Claims

1. A method for preparing quick-cooking rice using far-infrared combined solid-state fermentation, characterized in that, Includes the following steps: After inoculating the culture medium with Aspergillus oryzae, a suspension of Aspergillus oryzae spores was obtained. The selected Aspergillus oryzae was Aspergillus oryzae M6, which was deposited at the China Center for Type Culture Collection on April 12, 2023, with accession number CCTCC NO: M2023533, and the deposit address is Wuhan University, Wuhan, China. The colored rice sample was irradiated with far-infrared rays for 470-490 seconds and then cooled to room temperature to obtain the colored rice processed product; the irradiation temperature of the far-infrared rays was 150℃-170℃; the colored rice sample was black rice. After mixing the colored rice with nutrients, a suspension of Aspergillus oryzae spores was added for solid-state fermentation. During solid-state fermentation, Aspergillus oryzae can simultaneously metabolize α-amylase, protease, cellulase, carboxymethyl cellulase, and pectinase. The ratio of the volume of the Aspergillus oryzae spore suspension to the weight of the colored rice was 0.05-0.15 L / kg. The solid-state fermentation environment was maintained at a temperature of 36℃-38℃ and a humidity of 88%-92%, with a fermentation time of 70-74 hours. Nutritious rice is obtained by drying the fermentation products at low temperature.

2. The method for preparing quick-cooking rice using far-infrared combined solid-state fermentation according to claim 1, characterized in that, The Aspergillus oryzae spore suspension contained (2-3)×10 6 cfu / mL.

3. The method for preparing quick-cooking rice using far-infrared combined solid-state fermentation according to claim 1, characterized in that, During the solid-state fermentation process of mixing black rice with nutrients and adding Aspergillus oryzae spore suspension, the activities of α-amylase, protease, cellulase, carboxymethyl cellulase and pectinase produced by Aspergillus oryzae metabolism reached 334.5 U / g, 350.9 U / g, 7.5 U / g, 22.8 U / g and 36.1 U / g, respectively.

4. The method for preparing quick-cooking rice using far-infrared combined solid-state fermentation according to claim 1, characterized in that, In the solid-state fermentation process of mixing colored rice with nutrients and adding Aspergillus oryzae spore suspension, the ratio of Aspergillus oryzae colony-forming units in the Aspergillus oryzae spore suspension to the quality of the colored rice is controlled to be (2-3)×10. 8 cfu / kg.

5. The method for preparing quick-cooking rice using far-infrared combined solid-state fermentation according to claim 1, characterized in that, After mixing the colored rice product with nutrients, add Aspergillus oryzae spore suspension for solid-state fermentation. During this process, the solid-state fermentation product is turned over every 24 hours, and the total number of times it is turned over is controlled to be 2-3 times.

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