What Are the Extraction Methods of Black Rice Extract?

Black rice is an important food crop. It contains a large amount of nutrients, such as high levels of protein, essential amino acids, vitamins and essential minerals. In addition, black rice also contains an important bioactive ingredient, black rice anthocyanin. Many studies have shown that black rice extract is an anthocyanin pigment, which is a type of polyphenol. Analysis has shown that black rice anthocyanin is mainly paeoniflorin-3-glucoside and cornflower-3-glucoside [1-5]. In recent years, many scholars at home and abroad have conducted a large number of studies on black rice extracts. This paper discusses the extraction process and physiological activity function research of black rice extracts.

1 Research progress on the extraction process of black rice extract

1.1 Solvent extraction of black rice anthocyanin

The general process flow of solvent extraction of black rice anthocyanin is as follows: black rice → impurity removal, crushing → black rice powder → solvent extraction → suction filtration → extraction solution → distillation → black rice pigment concentrate → drying → crude black rice anthocyanin.

Studies have shown that many factors affect the extraction of black rice anthocyanin by the solvent method, including the choice of extraction solvent, solvent concentration, extraction temperature, extraction time, liquid-to-material ratio, pH value, and degree of black rice pulverization. Different solvents have very different effects on the extraction of anthocyanin from black rice. Zhong Yan et al. showed that anthocyanin from black rice is soluble in water, acetone, methanol, ethanol, ether, n-butanol, isopentanol and other solvents, with the best extraction results obtained in methanol and ethanol. The experiment also showed that black rice pigment is insoluble in ethyl acetate and chloroform [6].

Niu Juanjuan et al. showed that the yield of the solvent ethanol extraction method is not significantly different from that of the solvent water extraction method, and that the extraction temperature is lowered and the extraction time is reduced [7]. Therefore, ethanol is generally selected as the extraction solvent for black rice anthocyanin extraction. The process parameters for the extraction of black rice pigments using solvents vary among different researchers, mainly because of the inconsistency in the raw materials used and the experimental design parameters. For example, Zhang Fudi et al. found that the highest extraction rate of black rice melanin was achieved when the extraction temperature was 80 °C, the extraction time was 30 min, the liquid-to-material ratio was 1:10, and the solvent was 50% ethanol (by volume) [8].

Wu Suling's research concluded that the optimal process parameters for black rice anthocyanin are: 50% ethanol (by volume), black rice powder fineness 50 mesh, temperature 80°C, pH 3, liquid-to-material ratio 1:5, extraction time 30 min [9]. Ji Yunqi et al. used an alcohol-soluble method to extract black rice anthocyanin. The optimal extraction process parameters are: 95% ethanol (volume fraction), liquid-to-material ratio 1:50, temperature 60 °C, time 90 min, pH 1.8. At this time, the extraction rate of black rice pigment is as high as 2. 71% [10]. Guo Mei et al. showed that the optimal extraction process parameters for black rice anthocyanin are: 95% ethanol (volume fraction), liquid-to-material ratio 1:45, pH 3.0, extraction temperature 80°C, extraction time 90min [11]. Alessandra et al. used response surface analysis to obtain the process parameters for the solvent extraction of black rice anthocyanin: acidified ethanol (ethanol: 1 mol/L citric acid 1:80), extraction temperature 34.7°C, time 81 min, liquid-to-material ratio 1:30. Under these conditions, the anthocyanin extraction yield of black rice was 116.58 mg/100 g [12]. The solvent method of extracting black rice pigment requires less investment, but the black rice pigment product has disadvantages such as solvent residue and low product purity.

1.2 Microwave-assisted extraction of black rice anthocyanin

The general process flow of microwave-assisted extraction of black rice anthocyanin is: black rice → impurity removal, crushing → black rice powder → addition of extraction agent → microwave-assisted extraction → suction filtration → extraction solution → distillation → black rice anthocyanin concentrate → drying → crude black rice anthocyanin.

Studies have shown that compared with the organic solvent method, the use of microwave-assisted extraction technology can help to improve the extraction rate of black rice extract, shorten the extraction time, and improve the quality of black rice anthocyanin products. The factors affecting the microwave-assisted extraction of black rice anthocyanin mainly include microwave power, microwave time, material-liquid ratio, extraction solvent concentration and pH, etc. Zhang Jixiang et al. studied the process parameters for the extraction of black rice pigments using microwaves. The results showed that the most critical factor affecting the extraction of black rice anthocyanins using microwaves was the concentration of the extraction agent ethanol. The optimal process parameters for the extraction of black rice anthocyanins using microwaves were 80% ethanol (volume concentration), microwave power 480 W, microwave time 120 s, and liquid-to-solid ratio 1:20. Under these conditions, the anthocyanin extraction rate of black rice is as high as 3.2% [13]. Yuan Bo et al. established a regression statistical model for the extraction of black rice anthocyanin. The study found that the microwave time had no significant or minor effect on the extraction rate of black rice anthocyanin, which may be due to the small range of microwave time selection when designing the experiment [14, 15].

Ma Ping et al. established a multiple regression statistical model for the extraction of anthocyanin from black rice and analyzed that the optimal combination for extracting anthocyanin from black rice is 80% ethanol (volume fraction), a liquid-to-material ratio of 1:18, and a microwave time of 9s. At this time, the anthocyanin extraction rate of black rice extract is 4.97% [16]. Wang Xinzhe et al. used distilled water as the extraction solvent, and the yield of anthocyanin in black rice as the index. Using orthogonal experiments, they found that the optimal combination for microwave-assisted extraction of anthocyanin from black rice was: microwave power 600 W, microwave time 7.5 min, pH 5, liquid-to-solid ratio 1:4. At this time, the yield of anthocyanin from black rice was 1. 28% [17]. The above research shows that microwave-assisted extraction is a better method for extracting anthocyanins from black rice.

1.3 Ultrasonic-assisted extraction of black rice anthocyanin

The general process flow of ultrasonic-assisted extraction of black rice anthocyanin is as follows: black rice → impurity removal and crushing → black rice powder → addition of extraction agent → ultrasonic-assisted extraction → suction filtration → extraction solution → distillation → black rice pigment concentrate → drying → crude black rice anthocyanin.

Ultrasonic-assisted extraction is also a good method for extracting natural active substances. It is energy-saving, environmentally friendly and highly efficient. Ultrasonic-assisted extraction technology can be used for the extraction of black rice anthocyanin. The main factors affecting the ultrasonic-assisted extraction of black rice anthocyanin are ultrasonic power, ultrasonic extraction time, extraction temperature, material-liquid ratio, solvent concentration and pH. Chen Xiaoquan et al. compared the differences between the solvent method and ultrasonic-assisted extraction of anthocyanin from black rice, and found that the ultrasonic-assisted extraction method has the characteristics of high extraction efficiency, energy saving, and does not affect the basic properties of black rice pigments. The optimal combination process for ultrasonic-assisted extraction of anthocyanin from black rice is: ultrasonic extraction time 5 min, ultrasonic power 70 W, temperature 60 °C, and liquid-to-solid ratio 1:8 [18]. Sun Xiaoxia et al. used black rice husks as raw material to study the ultrasonic-assisted extraction process of black rice husk extract, and determined the ultrasonic-assisted extraction process conditions of black rice husk extract: 80% ethanol (volume fraction), pH 1.5, liquid-to-material ratio 1:40, extraction temperature 50°C, ultrasonic extraction time 60 min, and ultrasonic power 128 W [19]. Ultrasonic-assisted extraction of black rice anthocyanins is a convenient and fast process, and is a good method for extracting black rice extracts [20].

1.4 Enzyme-assisted extraction of black rice anthocyanin

The general process flow of enzyme-assisted extraction of black rice anthocyanin is: black rice → impurity removal, crushing → black rice powder → enzyme addition and hydrolysis → solvent extraction → suction filtration → extract → distillation → black rice pigment concentrate → drying → crude black rice anthocyanin.

The advantage of enzymatic extraction is that the reaction conditions are relatively mild. Pretreatment of black rice powder with biological enzymes (usually cellulase or complex biological enzymes) can break down the cell walls of black rice and quickly dissolve black rice anthocyanins, thereby increasing the extraction rate of black rice anthocyanins. During the enzymatic hydrolysis of black rice powder, the type of enzyme, the amount of enzyme, the temperature, the duration and the pH value are important factors affecting the extraction of black rice anthocyanin by enzymatic methods. Liu Yongji et al. used black rice husks as raw material and found that the optimal combination of process parameters for the extraction of black rice anthocyanin with cellulase as an aid is: The optimal process parameters for the extraction of black rice anthocyanin with cellulase-assisted solvent extraction are: 2.0% cellulase, 128.8 min enzymatic hydrolysis time, 38.7 ℃ enzymatic hydrolysis temperature, 80% ethanol (volume fraction), 1:10 liquid to solid ratio, 50 ℃ extraction temperature, 40 min extraction time. At this time, the extraction yield of black rice anthocyanin was 21.9 mg/g [21].

1.5 Supercritical CO2 extraction of black rice anthocyanin

The general process flow of supercritical CO2 extraction of black rice anthocyanin is as follows: black rice → impurity removal, crushing → black rice powder → sample loading into extraction vessel → supercritical CO2 extraction → addition of ethanol to collect extract → suction filtration → extract → distillation → black rice pigment concentrate → drying → crude black rice anthocyanin.

Supercritical CO2 extraction of black rice anthocyanin has the advantages of high product purity and simple process, but the initial investment is relatively large. In the supercritical CO2 extraction process, the extraction temperature, flow rate, pressure, and time are the main factors affecting the extraction of black rice anthocyanin by supercritical CO2 extraction. Liu Yan's research concluded that the optimal process parameters for extracting black rice anthocyanin by supercritical CO2 extraction are: extraction temperature 38 ° C, extraction pressure 45 MPa, extraction time 3h, fluid flow rate 20kg/h. Under these conditions, the content of black rice anthocyanin stock solution reaches 86.8% [22].

1.6 Synergistic extraction process for extracting black rice anthocyanin

Ultrasonic synergistic microwave technology extraction can shorten the extraction time and improve the extraction rate. Factors affecting the synergistic extraction process for extracting black rice anthocyanin include ultrasonic power, ultrasonic time, microwave power, microwave time, solvent concentration, etc. Liu Zhenchun et al. used ultrasonic microwave technology to extract black rice proanthocyanidins, and the research obtained the process parameters for the extraction of black rice proanthocyanidins using ultrasonic microwave technology: 82% acetone (volume fraction), ultrasonic action time 54 min, microwave action power 480 W, and the extraction rate of black rice proanthocyanidins at this time was 2. 411% [23].

2. Research progress on the main bioactive functions of black rice extract

In recent years, research on the main bioactive functions of black rice extract has mainly focused on antioxidant activity, free radical scavenging, blood lipid lowering, liver and kidney protection, cancer prevention, and anticancer activity.

2.1. Possesses antioxidant activity and free radical scavenging ability

The development of natural active substances with antioxidant activity and free radical scavenging ability has always been one of the research hotspots in the field of natural product chemistry. Sun Ling et al. obtained in vitro simulation that black rice anthocyanin has a strong scavenging ability against the free radical O2 - · and the greater the concentration, the better the scavenging effect [24]. Zhang Mingwei, Shi Juan, Zhu Xiaoli and others confirmed through in vivo experiments that ingesting a certain amount of black rice anthocyanin can improve the total antioxidant capacity of the liver, glutathione peroxidase activity and superoxide dismutase activity, and can significantly reduce malondialdehyde levels [25-27]. Li Xinhua et al. showed that black rice anthocyanins have a very good scavenging effect on ·OH and DPPH free radicals, and also have a certain scavenging effect on O2 · [28]. Wang Qiao'e et al. compared the antioxidant activity of anthocyanin from different parts of purified black rice, and found that the water-soluble part had the strongest antioxidant capacity[29] . Huang et al. also believe that the substance that plays an antioxidant role in black rice bran is mainly black rice anthocyanin[30] . These studies show that black rice extract can be used as an ideal natural source of antioxidants.

2.2 Ability to lower blood lipids

In recent years, the number of people with high blood lipids has been increasing, which has seriously affected people's health. There is an urgent need to develop and research natural, side-effect-free substances that lower blood lipids. Many studies have shown that black rice anthocyanins have a good ability to lower blood lipids. Zhang Mingwei et al. showed that ingesting a certain amount of black rice anthocyanins can significantly lower the blood lipid levels of rats with hyperlipidemia, and the atherosclerosis index also decreased significantly. Qin Yu et al. found that patients taking black rice anthocyanins can lower total cholesterol and triglyceride levels, and that it has a good ability to lower blood lipids [31]. Liu Mingda et al. showed through animal experiments that black rice anthocyanins can significantly lower animal blood lipids, with decreases in the main serum indicators total cholesterol, triglycerides, and low-density lipoprotein cholesterol, and an increase in high-density lipoprotein cholesterol [32]. Jang et al. found that ingestion of a certain amount of black rice extract can significantly lower the total cholesterol and triglyceride levels in mouse serum [33]. Therefore, black rice pigments can be used as an ideal natural hypolipidemic substance.

2.3 Liver and kidney protection

Some studies have reported that black rice extract has liver and kidney protection functions. For example, Hou Fangli et al. found that after consuming a certain amount of black rice skin anthocyanin, liver damage caused by carbon tetrachloride was significantly improved. The mechanism may be that black rice melanin has better antioxidant activity and free radical scavenging ability [34]. Hou Zhaohua's research found that black rice anthocyanins can alleviate alcoholic liver damage, and that they can significantly reduce the activities of serum glutamic oxaloacetic transaminase, alanine aminotransferase and γ-glutamyl transpeptidase [35]. Niu Juanjuan's research also found that black rice anthocyanins have a certain liver protection function, which can significantly reduce the activities of glutamyl aminotransferase and alanine aminotransferase in the blood serum. Jang et al. found that ingesting a certain amount of black rice extract can significantly reduce the symptoms of fatty liver degeneration in mice. Hao Jie's research shows that black rice anthocyanin extract can play a protective role against oxidative stress-induced nephrotoxicity and hepatotoxicity by scavenging free radicals [36]. These studies show that black rice extract can also be used in the development of liver and kidney protection products.

2.4 Other biological activities

Some studies have reported that black rice anthocyanins also have anti-asthmatic effects. Niu Haifeng and Wang Tianyu et al. found that black rice anthocyanins can effectively inhibit the infiltration of inflammatory cells in the lungs of asthmatic mice, so black rice anthocyanins have certain anti-asthmatic effects [37, 38]. In addition, some studies have reported that black rice extract can effectively inhibit the growth and proliferation of many types of cancer cells, and has anti-cancer and anticancer functions [39, 40]. Dong Bo et al. believe that black rice cornflower pigment also has preventive and therapeutic effects on diabetes [41].

3 Prospects

China is rich in black rice resources, and the deep processing and comprehensive utilization of black rice has broad prospects. Black rice extract has good physiological activity and is an ideal natural pigment. Therefore, the food and pharmaceutical industries have an increasing demand for black rice anthocyanins. However, research and application of black rice extract still requires further efforts from relevant scientific researchers. For example, research and development of better extraction, separation and purification techniques for black rice anthocyanin, improving the quality of black rice pigments through physical, chemical and biological techniques, further research on the biological activity, structure-activity relationship and mechanism of action of black rice extract, and development of high-quality black rice extract functional foods and medicines.

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