Method for improving extraction rate of anthocyanin in black soybean hulls by utilizing ultrasonic technology

Abstract

The invention discloses a method for improving the extraction rate of anthocyanin in black soybean hulls by using an ultrasonic technology, which is characterized in that black soybean hulls which are low in cost and are black soybean processing byproducts are used as raw materials, black soybean hull powder is obtained after pretreatment, the black soybean hull powder is crushed, sieved and dissolved in an ethanol-water solution with the concentration of 70% according to the solid-to-liquid ratio of 1: 10, and the black soybean hull powder is obtained. Putting the mixture into an ultrasonic cell crusher and carrying out ultrasonic treatment for 40 minutes at the power of 300W; centrifuging the mixed solution, repeatedly extracting the filter residue twice, combining the supernate, concentrating, and freeze-drying to obtain the black bean peel anthocyanin. According to the method, by optimizing the extraction conditions, the anthocyanin content of the extracted black bean peel reaches 55.5 mg / g, and compared with the anthocyanin content (31.4 mg / g) of a group without ultrasonic treatment, the anthocyanin extraction rate is increased by 76.8%. Meanwhile, the problem that the environment is influenced by byproducts in the black bean processing process is solved, and the method has theoretical guiding significance for further improving the comprehensive utilization value of the black beans.

Description

Technical Field

[0001] The invention relates to a method for improving the extraction rate of anthocyanins from black soybean hulls by utilizing ultrasonic technology, and belongs to the field of resource utilization of black soybean waste. Background Art

[0002] Black beans are a traditional agricultural product in my country, known for its medicinal and edible properties. Rich in nutrients such as protein, fat, and vitamins, they possess high nutritional and medicinal value. The Compendium of Materia Medica states that "regular consumption of black beans can prevent a wide range of illnesses," and that they have the potential to nourish the liver and kidneys and soften blood vessels. However, during the processing of black bean products, the bean hulls are often discarded as waste, their potential value overlooked. However, with the growing development of the black bean processing industry, awareness of their potential economic value has emerged, leading to research into their bioactive components, such as anthocyanins, phenols, polysaccharides, and vitamins.

[0003] Anthocyanins are a key component of black soybean hulls, responsible for their black color and possessing a range of biological activities. Currently, over ten anthocyanin species have been identified in black soybean hulls, including cyanidin 3-O-glucoside, delphinidin 3-O-glucoside, and pelargonidin 3-O-glucoside. These anthocyanins possess antioxidant, anticancer, and antidiabetic properties, and can also improve obesity and insulin resistance. Furthermore, anthocyanin-rich black soybean hulls can inhibit cancer cell growth by suppressing oxidative stress and inflammation, potentially showing potential in reducing tumor growth in various organs, including the intestine, breast, and ovary.

[0004] Numerous methods exist for extracting anthocyanins from black soybean hulls, including acid-solvent extraction, microwave-assisted extraction, ultrasonic extraction, supercritical fluid-assisted extraction, microbial fermentation, or enzymatic hydrolysis. Acid-solvent extraction typically uses methanol, ethanol, acetone, water, or a mixture of solvents, with a certain concentration of formic acid or hydrochloric acid added to prevent degradation of non-acylated anthocyanins. However, this method results in low extraction yields and high solvent consumption. Microwave-assisted anthocyanin extraction offers advantages such as time-saving, high efficiency, and energy conservation, but the extraction yield is relatively low, making large-scale extraction unfeasible. Supercritical fluid-assisted extraction is more suitable for extracting lipophilic, less polar pigments from natural plants. However, the extraction yield is lower for hydrophilic, highly polar, and high-molecular-weight pigments. Microbial fermentation or enzymatic hydrolysis utilizes microorganisms or enzymes to break down plant cell walls, allowing the anthocyanins within the cells to penetrate. However, the extracted anthocyanin solution contains relatively low levels of macromolecules such as polyethylene glycol and protein.

[0005] With the rapid development of biotechnology, ultrasound-assisted extraction (UAE) has become a new extraction method, gaining widespread application in the extraction of Chinese herbal medicine components, polyphenols, vegetable oils, aromatic compounds, polysaccharides, and other functional ingredients. It offers advantages such as simple experimental equipment, short extraction time, ease of operation, high efficiency, and high anthocyanin yield. Currently, while there is considerable research on the extraction of anthocyanins from fruits, vegetables, and flowers, there is limited research on the extraction of anthocyanins from black soybean hulls, particularly regarding the use of ultrasound-assisted ethanol extraction. Therefore, the use of ultrasound-assisted ethanol extraction of anthocyanins from black soybean hulls may offer the advantage of high yield and hold considerable research significance. Summary of the Invention

[0006] The present invention aims to provide a method for improving the extraction rate of anthocyanins from black soybean hulls by utilizing ultrasonic technology. The method comprises the following steps: pre-treating low-cost black soybean hulls, crushing the hulls, sieving the hulls, extracting the hulls with an ethanol-water solution, performing ultrasonic-assisted crushing and extraction with the ethanol-water solution, centrifuging the hulls, repeatedly extracting the residues, filtering, concentrating, and freeze-drying the hulls to obtain anthocyanins from black soybean hulls. The extraction conditions are optimized to screen out the extraction conditions with the highest anthocyanin content.

[0007] The object of the present invention is achieved through the following technical solutions:

[0008] A method for improving the extraction rate of anthocyanins from black soybean hulls by utilizing ultrasonic technology comprises the following steps: selecting black soybean hulls, pretreating, crushing, leaching with ethanol-water solution, ultrasonic-assisted extraction with ethanol-water solution, centrifuging, extracting the residues twice, filtering, concentrating, and freeze-drying to obtain anthocyanins from black soybean hulls.

[0009] The present invention optimizes the extraction conditions of anthocyanins from black soybean hulls, explores the effects of solid-liquid ratio, ethanol concentration, ultrasonic power, and ultrasonic time on the anthocyanin content in black soybean hulls during the extraction process, and determines the optimal level combination through single-factor and orthogonal experimental methods. BRIEF DESCRIPTION OF THE DRAWINGS

[0010] Figure 1 Effect of ethanol concentration on anthocyanin content in black soybean hulls;

[0011] Figure 2 Effect of solid-liquid ratio on anthocyanin content in black soybean hulls;

[0012] Figure 3 Effect of ultrasonic power on anthocyanin content in black soybean hulls;

[0013] Figure 4 Effect of ultrasound time on anthocyanin content in black soybean hulls; DETAILED DESCRIPTION

[0014] The technical solution of the present invention is further described below with reference to the accompanying drawings, but is not limited thereto. Any modification or equivalent replacement of the technical solution of the present invention that does not depart from the spirit and scope of the technical solution of the present invention should be included in the scope of protection of the present invention.

[0015] 1. Material selection

[0016] Black soybean husk was selected as the raw material for preparing anthocyanins.

[0017] 2. Pretreatment of black bean skin

[0018] Black bean hulls without impurities and of high quality are manually selected, crushed, and filtered through a 60-mesh sieve to obtain black bean hull powder.

[0019] 3. Ethanol-water solution extraction

[0020] Weigh 10g of black soybean hull powder, add 50-500mL of 40-90% ethanol-water solution, and extract in the dark at 4°C for 12h.

[0021] The mixed solution is placed in an ultrasonic cell disruptor with a power of 200 to 600 W and treated for 20 to 40 minutes.

[0022] 5. Centrifugation

[0023] The ultrasonic mixture was centrifuged at 5000 r / min for 10 min.

[0024] 6. Repeated extraction of filter residue

[0025] The supernatant after centrifugation was temporarily stored in the dark at 4°C, and the filtrate was added with ethanol-water solution again to repeat the above extraction process twice.

[0026] 7. Filter

[0027] The supernatants obtained from the three extractions were combined and filtered through filter paper.

[0028] 8. Concentration

[0029] The filtered filtrate was evaporated to remove ethanol in the dark at 4°C to achieve the purpose of concentration.

[0030] 9. Freeze drying

[0031] The concentrated anthocyanin extract is freeze-dried to obtain black soybean hull anthocyanin.

[0032] 10. Single-factor experiment

[0033] Figure 1The results show the effect of different ethanol concentrations on the anthocyanin content of black soybean hulls. As ethanol concentration increases, the anthocyanin content in black soybean hulls initially increases and then decreases. This is because when the ethanol concentration is low, the degree of binding between the reactants and the ethanol is relatively low, which is not conducive to the rapid extraction reaction and results in a low anthocyanin content. At higher ethanol concentrations, the ethanol may damage the plant's cell walls, reducing the effective penetration of anthocyanins and lowering the anthocyanin content. The highest anthocyanin content is achieved when the ethanol concentration is 80%.

[0034] Figure 2 The results show the effect of different solid-liquid ratios on anthocyanin content. As the solid-liquid ratio increases, the anthocyanin content in black soybean hulls increases first and then decreases, and there are significant differences in the anthocyanin content between different solid-liquid ratios (P<0.05). When the solid-liquid ratio is 1:20, the anthocyanin content in black soybean hulls is the highest.

[0035] Figure 3 The results show the effect of different ultrasonic powers on the anthocyanin content of black soybean peel. With the increase of ultrasonic power, the anthocyanin content of black soybean peel showed a trend of first increasing and then decreasing. When the ultrasonic power was 400W, the anthocyanin content of black soybean peel was the highest.

[0036] Figure 4 The results show the effect of different ultrasound times on the anthocyanin content of black soybean peel. With the increase of ultrasound time, the anthocyanin content of black soybean peel showed a trend of first increasing and then decreasing. When the ultrasound time was 50 minutes, the anthocyanin content of black soybean peel was the highest. However, when the ultrasound time was 40 minutes, the difference between the anthocyanin content of black soybean peel and the content at 50 minutes was not significant, so the ultrasound time of 40 minutes was selected.

[0037] Table 1 shows the optimal extraction conditions of anthocyanins from black soybean peels determined by orthogonal test. An orthogonal matrix with four factors and three levels was designed for a total of nine experimental groups. Based on the values of K1, K2, K3, k1, k2, k3, and R, the optimal extraction conditions were determined to be: 70% ethanol concentration, a solid-liquid ratio of 1:10, an ultrasonic power of 300W, and a sonication time of 40 minutes. Under these conditions, the anthocyanin content of the black soybean hulls prepared reached 55.5 mg / g; compared to 31.4 mg / g obtained without ultrasonic extraction.

Claims

1. A method for improving the extraction rate of anthocyanins from black soybean hulls using ultrasonic technology, characterized in that The method steps are as follows: Step 1: Material selection: Select black bean hulls as a by-product of black beans as the raw material for the preparation of anthocyanins. Step 2: Pretreatment: Manually pick high-quality black bean hulls to remove impurities. Step 3: The black bean hulls obtained in step 2 are crushed by a grinder for 10 seconds and then rest for 5 seconds, and the total crushing time is 60 seconds. Step 4: The black bean hull crushed material obtained in step 3 is sieved to obtain black bean hull powder, which is stored at -25°C for later use. Step 5: Ethanol-water extraction: The black bean hull powder obtained in step 4 is added to different concentrations of ethanol-water solutions (40% to 90%) at different solid-liquid ratios (1:5 to 1:50, mg / mL) and first extracted for 12 hours. Step 6: Ultrasonic-assisted extraction Ethanol-water extraction: The solution obtained in step 5 is placed in an ultrasonic cell crusher and treated for different times (20 to 60 minutes) under different ultrasonic powers (200W to 600W). Step 7: Centrifugation: Place the solution obtained in step 6 in a centrifuge and centrifuge at 5000r / min for 10 minutes, and save the supernatant. Step 8: Repeat the ultrasonic-assisted extraction ethanol-water solution extraction: Repeat the above extraction process twice with the same solid-liquid ratio and ethanol-water solution concentration. Step 9: Filtration: Combine the supernatants obtained in steps 7 and 8 and filter through filter paper. Step 10: Concentration: Place the filtrate obtained in step 9 in the dark at 4°C on a magnetic stirrer and stir for 4 hours to remove ethanol to obtain anthocyanin extract. Step 10: Freeze-drying: Vacuum freeze-dry the anthocyanin extract obtained in step 9 to obtain black bean anthocyanins.

2. The preparation method according to claim 1, characterized in that The mesh size of the sieve described in step 3 is 60 meshes.

3. The preparation method according to claim 1, characterized in that The concentration of the ethanol-water solution in step 4 is 70%.

4. The preparation method according to claim 1, characterized in that The solid-liquid ratio described in step 4 is 1:

10.

5. The preparation method according to claim 1, characterized in that The ultrasonic power described in step 4 is 300W.

6. The preparation method according to claim 1, characterized in that The ultrasonication time described in step 4 is 40 min.

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