What Is the Character and Benefit of Black Rice Extract Anthocyanin?

There are currently 397 varieties of black rice, and more than 90% of them are found in China, mainly distributed along the Yunnan-Guizhou Plateau[1]. Black rice is rich in amino acids, vitamins and trace elements such as Fe, Zn, Mn and Cu, and has high nutritional and medicinal value. It is known as “medicine rice”. The author refers to the research results of the past 20 years and provides a review of the chemical composition, physical and chemical properties, stability, and biological effects of black rice pigments, with the aim of providing guidance for the further development and utilization of black rice pigment resources.

1 Chemical composition of black rice extract

Black rice anthocyanin is a natural plant pigment extracted from the pericarp of black rice grains and belongs to the anthocyanin class of compounds. Liu Xiaogeng et al. [2] used paper chromatography (PC) to isolate two pigment bands, which were analyzed and found to be cyanidin-3-glucoside and cyanidin-3-rutinoside. Zhong Liyu et al. [3] used BAM (n-butanol ice: acetic acid water: = 4) chromatography, which resulted in five bands, indicating that black rice pigment is composed of five anthocyanin compounds. The structures of two of the wider bands (96% of the total) were analyzed and determined to be cyanidin-3-rhamnoside and peonidin-3-arabinoside. Cao Xiaoyong et al. [4] believe that black rice extract contains anthocyanidin-5-glucoside,anthocyanidin-3-rhamnoside,malvidin-3-galactoside,

anthocyanidin-3-glucoside, and methyl anthocyanidin-3-glucoside. Zhang Fudi et al. [5] used BAM analysis to identify the two main components of cyanidin-3-glucoside and cyanidin-3-rhamnoside. In summary, the pigment composition and chemical structure differ among different black rice varieties. It can be confirmed that the main component of black rice extract is anthocyanin, and the main sugar ligand is glucose.

2. Physicochemical properties of black rice extract [1, 4, 6]

2.1 Solubility

Black rice anthocyanins are soluble in polar solvents such as methanol, ethanol and propylene glycol; they are insoluble or hardly soluble in non-polar solvents such as petroleum ether, benzene and toluene. The hydrophobic black rice pigment has a wider solubility and is soluble in low-polarity solvents such as acetone, chloroform, and ethyl acetate [7].

2.2 Color and acidity

Black rice anthocyanin appears red or reddish purple under visible light; it appears blue after being smoked with ammonia and yellow-orange after being treated with concentrated sulfuric acid. The pH has a significant effect on the color of the black rice extract solution. It is bright red in an acidic medium (pH < 4), dark red or purple-red in a neutral or slightly alkaline medium, and blue-yellow in a strongly alkaline medium. When the pH is > 11, the pigment opens the ring and changes to a chalconone structure, giving an orange-yellow color.

2.3 Spectral absorption characteristics

Black rice anthocyanin has characteristic absorption in the visible light region at 535 nm and in the ultraviolet region at 270 nm in an ethanol and hydrochloric acid methanol medium.

2.4 Chelation reaction with metal ions

Anthocyanin has a hydrochloric acid-magnesium powder reaction, a neutral lead acetate precipitation reaction and a ferric chloride color development reaction.

2.5 Reaction with sugar ligands

2.5.1 Morish reaction

In a dilute ethanol solution of the pigment, add Morish reagent (an ethanol solution of α-naphthol), mix well, tilt the test tube, and slowly (without shaking) add concentrated sulfuric acid along the wall of the test tube. After standing for a while, a purple ring will appear at the interface of the two phases.

2.5.2 Perlin reaction

In a solution of black rice extract in water, add an equal mixture of the two reagents, Perrin A and Perrin B. Heat in a boiling water bath, and a red precipitate will form. Add concentrated hydrochloric acid to the filtrate dropwise to acidify, heat in a water bath to hydrolyze, add 10% NaOH to neutralize to neutral, and then carry out the Perrin reaction again, and a trace of dark red precipitate will form.

2.5.3 Saccharification

Concentrated hydrochloric acid was added dropwise to an aqueous solution of black rice extract, and the mixture was heated and acidified for a few minutes. Then, phenylhydrazine reagent was added, and the mixture was heated with boiling water for about 0.5 h. A small amount of yellow precipitate was produced. After cooling, the precipitate was filtered and crystallized, and microscopic observation showed that the crystals were like broom-like polyacicular needles.

3 Biological activity of black rice extract

3.1 Free radical scavenging and antioxidant effects

Lonsen studied the antioxidant effect of black rice extract using a chemiluminescence analysis method [8]. Black rice extract can scavenge H2O2, superoxide anion radicals (O2-) and hydroxyl radicals (·OH). Black rice extract has a strong inhibitory effect on whole blood chemiluminescence and a dose-response relationship. It also has a certain scavenging capacity for reactive oxygen species produced by intracellular or non-cellular systems. Black rice extract can be used as a better free radical scavenger in foods. Xiao Xiang et al. [6] used black rice anthocyanin, black soybean skin pigment and copper algae pigment to test the inhibitory effect of the three pigments on the Fe2+ induced egg yolk lipoprotein PUFA peroxidation system and the TBAS (thiobarbituric acid reactive substance) production system, as well as their scavenging effect on O2-. It was found that the three pigments had a good scavenging effect on both O2- and ·OH, and could also significantly inhibit the peroxidation of yolk lipoprotein PUFA. In 2006, Zhang Mingwei et al. also proved through experiments that black rice skin anthocyanin has an antioxidant effect and a certain protective effect on the peroxidation damage of vascular endothelial cells [9].

3.2 Lowering blood lipids and preventing atherosclerosis

Ling et al. reported that dietary black rice and black rice bran can lower blood lipid levels and inhibit the occurrence and development of atherosclerosis (AS) in experimental animals. Zhang Mingwei et al. [10] used black rice bran anthocyanin (the total anthocyanin content was determined by HPLC to be 72.83%, of which 38.75% was cyanidin-3-glucoside, Cyanidin-3,5-diglucoside 13.47 %, Pelargonidin-3,5-diglucoside 12.53 %, Malvinidin-3-glucoside 8.08 %) Tests were carried out on rats with hyperlipidemia to determine the total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C) and oxidized low-density lipoprotein (ox-LDL). The results showed that black rice bran anthocyanin significantly reduced the levels of TC and LDL-C in the serum of test rats, had a positive effect on HDL-C, and could inhibit the production of ox-LDL.

Yu Xiaoping et al. [11] also investigated the effect of black rice bran anthocyanin extract on the stability of atherosclerotic (As) plaques and blood lipids in Apolipoprotein E-deficient (ApoE-/-) mice. The results showed that black rice skin anthocyanin can significantly reduce the serum TC and LDL-C levels of ApoE-/- mice, and the frequency of unstable atherosclerotic plaques in the nameless artery is reduced, and the plaque area is reduced. Xia Xiaodong et al. [12] used black rice anthocyanin extract to conduct the same experiment, which proved that black rice anthocyanin extract can significantly reduce the levels of mouse serum TC and LDL-C, reduce the area of As plaques at the unnamed artery, reduce the content of matrix metalloproteinase-1 in the plaques, and increase the stability of As plaques, thereby making them less likely to break and produce blood clots. Yu Xiaoping et al. [13] also established an animal model of unstable atherosclerotic plaques (lipoprotein E knockout mice), and speculated that black rice anthocyanin promotes plaque stability, which is related to the down-regulation of inducible nitric oxide synthase expression in unstable plaques. It was found that black rice anthocyanin extract can significantly inhibit the expression of inducible nitric oxide synthase mRNA in unstable plaques in the unnamed artery, but the mechanism is unclear.

3.3 Protection of vascular endothelial cells

Zhang Mingwei et al. [9] studied the protective effect of four types of black rice anthocyanins on vascular endothelial damage caused by ox-LDL, and analyzed the effect of black rice anthocyanins on the morphology and viability of ox-LDL-induced vascular endothelial cells, as well as the production of malondialdehyde (MDA) and free radicals by endothelial cells. The results showed that black rice anthocyanin can significantly reduce the damage to the morphology of endothelial cells caused by ox-LDL, restore the normal shape, reduce the inhibitory effect of ox-LDL on cell proliferation, and significantly reduce the increase in MDA in cells. Black rice anthocyanins have a significant protective effect on peroxidative damage to vascular endothelial cells, which may be related to the scavenging of free radicals by black rice anthocyanins and the reduction of damage caused by oxidative stress. It may also be related to the high Zn content in the pigment [14].

3.4 Improves iron deficiency anemia

Tests have shown that black rice is rich in trace minerals Fe, Zn and Cu, and in particular, it is high in Fe content [15]. Xu Fei et al. fed black rice to rats with iron deficiency anemia, and the results showed that after 30 days, the anemia of the rats improved significantly, and the hemoglobin content of the anemic rats was increased. Fang Junjie et al. studied the effect and mechanism of action of black rice pigment extract (OSP) on the recovery of bone marrow hematopoiesis in mice after radiation and chemical treatment. The results showed that OSP promoted the proliferation of hematopoietic cells in the bone marrow inhibited by cyclophosphamide and accelerated hematopoietic recovery; for mice with severe damage to bone marrow hematopoietic cells, black rice can also help restore hematopoietic function, and it is speculated that the protective and restorative effect of OSP against radiation damage may be achieved by enhancing the renewal of hematopoietic stem cells and differentiation into progenitor cells [16]. Zhao Zesheng and others also achieved good results using black rice porridge to treat anemia during pregnancy. In summary, black rice has the effect of improving iron deficiency anemia, which is related to its high Fe content and low content of factors that interfere with Fe absorption [17].

3.5 Other biological effects

According to the relevant regulations of the United Nations Joint Food and Agriculture Organization/World Health Organization on food additives, black rice extract can be used in food production without pharmacological testing. There is very little pharmacological information on black rice extract in China, but for a long time, people have had a relatively consistent understanding of black rice: black rice can promote the growth of white blood cells in the body, improve immunity and tolerance to hypoxia, prevent aging, have hair-darkening effects, and inhibit the growth of cancer cells. Hu Qiulin[18] used black rice extract to conduct anti-fatigue and hypoxia tolerance tests on mice, and the results showed that feeding mice with an aqueous solution of black rice extract can significantly improve the animals' hypoxia tolerance and fatigue resistance, especially the hypoxia tolerance. Black rice extract can improve the vitality of the body. Li Jing et al. [14] also discussed the biological mechanism of black rice extract in exercise. The antitumor effect of black rice extract is positive, but there is a lack of literature on this aspect.

4 Conclusion

Black rice extract is a complex mixture. With the improvement of separation and purification methods, especially the application of chromatography-mass spectrometry technology, the study of the chemical composition of black rice pigments will surely be promoted. Black rice is rich in nutrients, and black rice extract has good biological effects. With further research, black rice extract will show broad development prospects in the development and application of natural pigments in foods, cosmetics and pharmaceuticals.

References

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