Polyphenols in whole rice grain: Genetic diversity and health benefits

Food Chemistry 180 (2015) 86–97

Contents lists available at ScienceDirect

Food Chemistry journal homepage: www.elsevier.com/locate/foodchem

Review

Polyphenols in whole rice grain: Genetic diversity and health benefits Yafang Shao a,b, Jinsong Bao b,⇑ a b

China National Rice Research Institute, Hangzhou 310006, China Institute of Nuclear Agricultural Sciences, Zhejiang University, Huajiachi Campus, Hangzhou 310029, China

a r t i c l e

i n f o

Article history: Received 2 November 2014 Received in revised form 2 February 2015 Accepted 5 February 2015 Available online 14 February 2015 Keywords: Polyphenols Nutraceutical property Genetics Health benefits Rice (Oryza sativa L.)

a b s t r a c t Polyphenols, such as phenolic acid, anthocyanin and proanthocyanidins, have both nutraceutical properties and functional significance for human health. Identification of polyphenolic compounds and investigation of their genetic basis among diverse rice genotypes provides the basis for the improvement of the nutraceutical properties of whole rice grain. This review focuses on current information on the identification, genetic diversity, formation and distribution patterns of the phenolic acid, anthocyanin, and proanthocyanidins in whole rice grain. The genetic analysis of polyphenol content and antioxidant capacity allows the identification of several candidate genes or quantitative trait loci (QTL) responsible for polyphenol variation, which may be useful in improvement of these phytochemicals by breeding. Future challenges such as how to mitigate the effects of climate change while improving nutraceutical properties in whole grain, and how to use new technology to develop new rice high in nutraceutical properties are also presented. Ó 2015 Elsevier Ltd. All rights reserved.

Contents 1. 2.

3.

4.

5. 6. 7.

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Phenolic acids . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.1. Chemistry and bioactivity. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.2. Extraction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.3. Diversity. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.4. Formation and distribution pattern . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Anthocyanins and proanthocyanidins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.1. Chemistry and bioactivity. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.2. Extraction, diversity and distribution pattern . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Genetics of polyphenols. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4.1. Biosynthesis pathway . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4.2. Regulatory factors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4.3. Improvement of polyphenols and antioxidant capacity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Applications and health benefits . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Future challenges . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Conclusion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

86 87 87 88 88 90 90 90 91 91 91 92 93 94 95 96 96

1. Introduction

⇑ Corresponding author. Tel.: +86 571 86971932; fax: +86 571 86971421. E-mail address: [email protected] (J. Bao). http://dx.doi.org/10.1016/j.foodchem.2015.02.027 0308-8146/Ó 2015 Elsevier Ltd. All rights reserved.

Rice (Oryza sativa L.) is a staple food in many parts of the world especially in Asian countries, where it is the most important crop. Rice consumption in Africa and Latin America is increasing faster

87

Y. Shao, J. Bao / Food Chemistry 180 (2015) 86–97

than that of any other staple food in the past decade, mainly due to urbanization and changes in eating habits. In addition, European, US and Australian citizens are eating more rice nowadays, possibly due to an increased interest in Asian cuisine. With the growth of population, our world faces some major global health challenges, such as malnutrition and increased incidence of chronic diseases like heart disease, Type II diabetes, obesity, and cancers (Dipti et al., 2012). Obviously, malnutrition is a major health burden in the developing countries. However, increased incidence of chronic diseases is a health problem in both developing and developed countries. Many interventional and epidemiological studies have shown that consumption of whole grains can reduce the risks of chronic diseases, such as cardiovascular diseases, type II diabetes, obesity and some cancers. The 2010 Diet Guidelines for Americans suggests that individuals should intake three or more ounce-equivalent of whole grain products per day. Whole rice grain is becoming popular in western countries due to its health benefits, and is more gradually accepted in developing countries with the improvement of living standards (Shao et al., 2011). Whole-grain rice is the unpolished version of the grains consisting of the germ, bran, and endosperm, and is also called brown rice. The colors of the whole grain rice range from white to red, and black (dark purple). The health benefits of whole grain are mainly contributed by one of its major constituents, the polyphenols. Polyphenols in rice grain can be classified into three subgroups: (1) phenolic acids, which is the most common secondary metabolites in cereal grains; (2) anthocyanins, which only exist in black or dark purple grains; and (3) proanthocyanidins, which mainly consist of catechin and epicatechin block unit in red rice and are considered to be the most effective antioxidants in nature (Gunaratne et al., 2013; Qiu, Liu, & Beta, 2010). This review considers the rapidly accumulated new data on polyphenols in rice, such as phenolic acids, anthocyanins, and proanthocyanidins. The contents of polyphenols in rice grain and rice-based foods are described, as well as the most pressing issues of genetic basis and associated genes of the biosynthetic pathway in rice. Finally, the current methods of improving polyphenol content and antioxidant capacity and the potential contributions of these compounds to the health effects are discussed, which could provide solutions to some of the grand challenges facing the riceconsuming countries.

ycinnamic acids contain caffeic, p-coumaric, sinapic, ferulic, and isoferulic acids. The content of hydroxybenzoic acids is very low in rice grain, while the content of hydroxycinnamic acid is high with ferulic acid the most abundant followed by p-coumaric acid and sinapic acid, both of which are present as cis- and trans-isomers in rice grain (Park et al., 2012; Shao et al., 2014a). Sinapic acid and p-hydroxybenzoic acid are significantly reduced with increase of CO2 concentration in brown and white rice grain (Goufo et al., 2014). Protocatechuic and caffeic acid could help the rice plant absorb and utilize precipitated apoplasmic Fe from root surface (Bashir et al., 2011). Besides a series of monomeric phenolic acids, phenolic dehydrodimers are formed with cross-linking to the cell walls during lignification (Renger & Steinhart, 2000). In rice grain, most dehydrodimers are connected by C–C links in linear form, and a few are linked in aryltetralin form or benzofuran form. Dehydrodiferulic acid with 5-50 coupled in wheat is the first identified phenolic dehydrodimer (Geissmann & Neukom, 1973). In 1990s, a number of dehydrodiferulic acids were isolated from various cell walls, e.g. 8-50 , 8-80 , 8-O-40 , 5-50 , 8-50 benzofuran form isolated from wheat bran, maize cell suspensions, and cell walls of wheat straw (Garcia-Conesa, Plumb, Waldron, Ralph, & Williamson, 1997; Grabber, Hatfield, Ralph, Zon, & Amrhein, 1995; Waldron, Parr, Ng, & Ralph, 1996). In rice, 8-80 aryl form, 8-50 benzofuran form, 8-80 , 8-50 , 8-O-40 , 5-50 have been detected by alkaline treatment with NaOH (Qiu et al., 2010; Renger & Steinhart, 2000; Zhang, Shao, Bao, & Beta 2015). Apart from ferulic acid, dehydrodimers of sinapic acid and sinapate-ferulate heterodimers have been identified in cereal dietary fiber (Bunzel et al., 2003). In wild rice, both dehydrodiferulic acids and dehydrodisinapic acids are detected by HPLC-QTOF-MS/MS (Qiu et al., 2010). Aryltetralin 8-80 -coupled

(a)

2. Phenolic acids 2.1. Chemistry and bioactivity Phenolic acids in rice grain are present in two forms: (1) soluble form, including free and conjugated forms, the former of which can be directly extracted by solvent, such as aqueous methanol, ethanol, and acetone, and the latter can be hydrolyzed from soluble phenolics by alkali; (2) insoluble form, also called bound phenolics, which esterify to the cell walls (Shao, Xu, Sun, Bao, & Beta, 2014b). Among these three phenolic fractions, insoluble bound phenolic acids are the most abundant, followed by soluble conjugated phenolic acids with soluble free phenolic acids the least (Park et al., 2012; Shao, Xu, Sun, Bao, & Beta, 2014a). Phenolic acids mainly exist in rice bran with trace amounts in endosperm in different rice genotypes (Shao et al., 2014a). There are two groups of phenolic acids in rice grain, derivates of hydroxybenzoic acids and hydroxycinnamic acids (Fig. 1a), which can be detected at the wavelength of 260–280 nm and 320– 325 nm, respectively (Irakli, Samanidou, Biliaderis, & Papadoyannis, 2012; Jun, Song, Yang, Youn, & Kim, 2012). Hydroxybenzoic acids contain gallic, p-hydroxybenzoic, salicylic, gentisic, protocatechuic, vanillic, and syringic acids, and hydrox-

(b)

(c) Fig. 1. Structures of polyphenols: (a) hydroxybenzoic acid (left) and hydroxycinnamic acid (right); (b) anthocyanin; (c) the block unit of proanthocyanidins.

88

Y. Shao, J. Bao / Food Chemistry 180 (2015) 86–97

dehydrodisinapic acid (thomasidioic acid) seems not to be present as a natural product in cereal grains, but might be derived from air oxidation during alkaline hydrolysis (Cai, Arntfield, & Charlton, 1999). However, Qiu et al. (2010) suggests it to be a natural compound. Therefore, further research on formation mechanisms of dehydrodiphenolic acid is still needed. Many studies indicate that soluble free, soluble conjugated, and insoluble bound phenolic acids have different potential health benefits. Hydroxycinnamic acids have potential antioxidant activity which is highly associated with hydroxylation and methylation. The antioxidant activities of hydroxycinnamic acids in rice are in the order caffeic acid > sinapic acid > ferulic acid > p-coumaric acid. The soluble phenolic acids would be rapidly absorbed in stomach and small intestine and spread to the whole body with the health benefits such as inhibition against oxidation of low-density lipoprotein (LDL) cholesterol and liposome (Chandrasekara & Shahidi, 2011). The soluble phenolics of red rice grain display a greater inhibitory effect on angiotension I-converting enzyme than that of non-pigmented rice (Massaretto, Madureira Alves, Mussi de Mira, Carmona, & Lanfer Marquez, 2011). The bound phenolic acids, especially the bound ferulic acid, would be released enzymatically in colon because the fibrous materials are hard to be digested in stomach and intestine, which may play an important role in prevention of colon cancer, as well as anti-inflammatory effects (Dipti et al., 2012). Therefore, the consumption of different forms of phenolic acids results in different health benefits. 2.2. Extraction Phenolic acids are common secondary metabolites in rice grains. Different extractants and extraction methods affect the extraction rate. Previously, the extraction of phenolics was conducted by organic solvents such as methanol, ethanol, acetone, and acetonitrile. It is reported that extractions with 40% acetone from black, red, green and brown bran exhibit higher TPC (64.9– 708.6 lg GAE (gallic acid equivalent)/ml extract) among 0–100% aqueous acetone, ethanol and methanol, respectively (Jun et al., 2012). Because hydrochloric acid could increase the extraction of total phenolics, many studies use the acidified organic solvent as extractants (Irakli et al., 2012; Park et al., 2012). Recently, a microwave-assisted extraction method has been tried and optimized by the response surface methodology, which can be adapted to investigate presence of phenolic compounds in a wide variety of rice grains (Setyaningsih, Saputro, Palma, & Barroso, 2015). Because three forms of phenolic acids exist in the rice grain, many extraction methods have emerged. Normally, aqueous organic solvents are used to extract the soluble free phenolic acids (Shen, Jin, Xiao, Lu, & Bao, 2009). The soluble conjugated or insoluble bound phenolic acids are usually extracted by sodium hydroxide with different concentrations which can break the chemical bonds between phenolic acids and soluble components or insoluble cell walls (Park et al., 2012; Shao et al., 2014b; Waldron et al., 1996). It is reported that 65–67% ethanol extraction for 40–45 min at 51–54 °C is an optimal ultrasonic-assisted extraction for rice bran (Tabaraki & Nateghi, 2011). The separation of phenolic components of cereal grains is mainly performed by reverse phase HPLC with C18 column as stationary phase and acetic acid/ methanol/ water or trifluoroacetic acid/ acetonitrile/ water as mobile phase, detected by photo diode array (PDA) detector, and confirmed by mass spectroscopy (Goufo et al., 2014; Shao et al., 2014a, 2014b). 2.3. Diversity Much literature data on phenolic acids is given as total amount of soluble free or insoluble bound phenolics assayed by Folin–Cio-

calteu reagent. The soluble free TPC is 44, 58–62, 72–82, 697 and 240–547 mg GAE/100 g, while the insoluble bound TPC is 61, 46– 63, 48–57, 86, 90–95 mg GAE/100 g for white, light brown, brown, red and black rice, respectively (Min, Gu, McClung, Bergman, & Chen, 2012). Rice bran has the TPC of 92 mg GAE/ 100 g, which is higher than other cereals and pseudocereals (Gorinstein et al., 2007). The soluble free TPC of white and black bran ranges from 441.0 to 530.2 and from 2086 to 7043 mg GAE/100 g, respectively, and the bound TPC of white and black bran ranges from 188.5 to 213.1 and from 221.2 to 382.7 mg GAE/100 g, respectively (Zhang, Zhang, Zhang, & Liu, 2010). The soluble free TPC in 481 rice accessions including 423 white rice, 52 red rice, and 6 black rice displays a wide range of 108.1–251.4, 165.8–731.8, and 841.0– 1244.9 mg GAE/ 100 g, respectively, which correlates well with grain color, size and weight (Shen et al., 2009). Detailed data of soluble free, soluble conjugated and insoluble bound phenolic acids are listed Table 1. Among non-pigmented, red and black rice, free protocatechuic acid is the highest and only detected in red and black rice with contents of 2.32 and 3.54 mg/ 100 g, respectively, and free cinnamic acid is only detected in non-pigmented rice with 0.29 mg/100 g, but free sinapic acid is not detected in any of the genotypes. For the soluble free phenolic acids of white rice, ferulic acid is the most abundant, followed by pcoumaric or (+)-catechin acid, and trace amounts of p-hydroxybenzoic and gallic acid are also detected (Irakli et al., 2012). Vanillic acid could be detected in both non-pigmented and pigmented rice grain with black rice grain (2.5 mg/100 g) higher than white (0.35 mg/100 g) and red rice grain (0.30 mg/100 g) (Irakli et al., 2012). Protocatechuic acid is the most abundant free phenolic acid in red and black rice, followed by ferulic acid for the former and vanillic acid for the latter (Irakli et al., 2012). However, Park et al. (2012) found that ferulic acid is the most abundant in Korean red and black rice, followed by p-coumaric acid without the detection of protocatechuic acid. The content of ferulic acid is almost the same in red and black rice, and gallic, p-hydroxybenzoic, caffeic, syringic, and p-coumaric acid are detected at very low levels (Irakli et al., 2012). For the soluble conjugated fraction, vanillic and protocatechuic acids are only present in black rice, both of which are the highest with around 4.5 mg/100 g, and sinapic acid is only detected in red and black rice with 1.0 and 0.6 mg/100 g, respectively (Shao et al., 2014a). Using methanol and sodium hydroxide to release the soluble conjugated phenolic acids, it is found that ferulic acid is the most abundant in white and red rice, followed by syringic acid (Table 1). When extracted by acidified methanol and sodium hydroxide, it is found that sinapic acid is the most abundant with 5.4–7.4, 6.8–9.5, and 6.6–16.6 mg/100 g in white, red, and black rice grain, respectively, followed by ferulic acid with the content of 5.1–6.4, 5.7–8.3, and 5.4–10.5 mg/100 g, respectively (Park et al., 2012). The bound phenolic acids, especially ferulic acid and p-coumaric acid, serve as substrates to form the basic skeleton of all flavonoid derivates. In black rice grain, ferulic acid is the most abundant, ranging from 114.5 to 166.4 mg/100 g, which is followed by the contents of protocatechuic and p-hydroxybenzoic acid ranging from 45.3 to 53.9 and from 43.4 to 79.0 mg/100 g, respectively (Laokuldilok, Shoemaker, Jonqkaewwattana, & Tulyathan, 2010). Isoferulic acid is found in white and pigmented rice genotypes with almost the same content (Shao et al., 2014a). Phenolic dehydrodimers are cell wall bound and only appeared in insoluble bound form in many cereal grains after hydrolysis of 4 M NaOH (Qiu et al., 2010; Renger & Steinhart, 2000). In rice grain, 8-80 aryl dehydrodiferulic acid had the highest contents, up to 57 lg/g (Renger & Steinhart, 2000). In commercial wild rice, 8-O-40 dehydrodiferulic acid was the most abundant (34 lg/g), and the content of 8-80 disinapic acid was 19 lg/g (Qiu et al., 2010). In white, light-

Table 1 Concentrations of phenolic acids in rice whole grain, milling fraction and products (mg/100 g dry weight).a Extractant Acidified acetonitrile 80% MeOH 85% MeOH containing 2 g/L Acidified acetonitrile 85% MeOH containing 2 g/L Acidified acetonitrile 80% MeOH 85% MeOH containing 2 g/L 80% MeOH and 4 M NaOH 85% MeOH containing 2 g/L 80% MeOH and 4 M NaOH 80% MeOH and 4 M NaOH 4 M NaOH (residue) 4 M NaOH (residue) 4 M NaOH (residue) Microwave-assisted

FA

of of

of of

0.79 0.34–1.51 BHA 0.59–0.77 1.24 BHA 0.53–0.87 1.4 0.2–0.5 BHA and 5 M NaOH 4.97–6.41 1.11 BHA and 5 M NaOH 5.4–10.5 1.6 2.1 4.28–7.79 5.7–7.7 17.9–18.6 1.13–3.81

p-CA

IFA

SRA

VA

SNA

CA

CTA

p-HA

PA

References

0.54 0.07–0.20 0.29–0.42 0.37 0.33–0.56 0.4 0.07–0.34 0.65–0.91 0.4 0.9–1.2 0.4 0.6 1.36–2.68 1.1–1.8 2.0–2.1 0–3.77

– – – – – – – – 0.4 – 0.41 1.2 1.20–1.59 0.8–1.3 1.0–1.3 –

0.30 – 0.022–0.03 0.37 0.03–0.05 0.2 – 0.08–0.12 1.08 0.1–0.3 0.8 – 0.60–0.61 0.50–0.52 0.63–0.67 0–0.60

0.35 – 0.041–0.05 0.30 0.04–0.41 2.5 1.2–3.2 0.82–0.85 – 0.8–4.5 – 4.5 – – 4.9–5.4 0.77–3.81

– – 0.05–0.09 – 0.07–0.22 – – 5.35–7.44 – 6.6–16.3 1.0 0.6 – – 0.4 0–3.77

0.29 0.12 – 0.20 – 0.2 3.0–3.2 – – – – – – – – TR

0.29 0.32–0.33 – – – – 0.4–1.4 – – – – – – – – –

0.25 – 0.046–0.053 0.23 0.05–0.08 0.3 0.2–6.0 0.72–1.21 – 0.8–3.3 – – – – – 0–4.63

– – – 2.32 – 3.54 – – – – – 4.5 – 0.2–1.3 0.76–4.48 0–3.77

Irakli et al. (2012) Huang and Ng (2012) Park et al. (2012) Irakli et al. (2012) Park et al. (2012) Irakli et al. (2012) Huang and Ng (2012) Park et al. (2012) Shao et al. (2014a) Park et al. (2012) Shao et al. (2014a) Shao et al. (2014a) Shao et al. (2014a, 2014b) Shao et al. (2014a, 2014b) Setyaningsih et al. (2015)

Rice milling fractions White bran 80% methanol Colored bran 80% methanol Bran 40% acetone Bran 80% MeOH and 4 M NaOH Embryo 80% MeOH and 4 M NaOH Embryo 4 M NaOH (residue) White bran 1 M NaOH Red bran 1 M NaOH Black bran 1 M NaOH Bran 4 M NaOH (residue)

0.7–5.7 3.5–6.8 0.18–0.44 5.4–7.6 37. 8–53.2 41.2–47.7 131.5 111.5 114.5–166.4 68.5–179.3

0.5–1.0 2.46–3.97 0.05–0.20 1.8–4.0 1.9–5.3 13.5–17.1 42.1 27.8 14.8–25.4 13.1–65.3

– – – 1.9–3.8 8.9–12.9 3.6–4.6 – – – 3.5–8.8

– – 0.004–0.056 3.7–6.2 5.3–5.5 2.4–3.7 – – – 3.1–4.4

0.2–1.7 9.48–16.6 0.052–0.11 31.7 only black 6.6 only black 11.0 only black – – – 1.6–35.0

– – 0.02–0.08 4.2–7.3 – 0.79–0.81 25.9 21.0 15.5–28.6 1.6–5.6

0.2–0.8 7.8–25.3 – – – – – – – –

1.3–2.4 5.8–48.7 – – – – – – – –

– 5.9–18.8 0.007–0.058 – – 0.42 only red 6.9 5.3 43.4–79.0 0.9–1.6

– – 0.001–0.098 34.2 only black – 0.40 only red 1.4 8.2 45.3–53.9 0.9–3.2

Huang and Ng (2012) Huang and Ng (2012) Jun et al. (2012) Shao et al. (2014a) Shao et al. (2014a) Shao et al. (2014a) Laokuldilok et al. (2010) Laokuldilok et al. (2010) Laokuldilok et al. (2010) Shao et al. (2014a)

Rice products Wines b Fermented

0.4–1.8

0.6–4.3

–

5.6–30.8

1.2–1.9

–

–

4.0–36.1

–

–

Que, Mao and Pan (2006)

Y. Shao, J. Bao / Food Chemistry 180 (2015) 86–97

Whole grain White White White Red Red Black Colored White White Red Red Black White Red Black Whole grian

a

Range or mean value. Values express as lg/ml. BHA, butylated hydroxyanisole; FA, ferulic acid; p-CA, p-coumaric acid; IFA, isoferulic acid; SRA, syringic aicd; VA, vanillic acid; SNA, sinapic acid; CA, caffeic acid; CTA, (+)-catechin acid; p-HA, phydroxybenzoic acid; PA, protocatechuic acid; TR, trace amount. b

89

90

Y. Shao, J. Bao / Food Chemistry 180 (2015) 86–97

purple, and black rice grain, all the forms of diferulic acid have similar contents (Zhang et al., 2015). Because of the existence of isoferulic acid, the structure of diferulic acid needs to be further clarified. Most recently, Wang et al. (2015) isolated a new compound, p-hydroxy methyl benzoate glucoside, together with nine known compounds as cycloeucalenol cis-ferulate, cycloeucalenol trans-ferulate, trans-ferulic acid, trans-ferulic acid methyl ester, cis-ferulic acid, cis-ferulic acid methyl ester, methyl caffeate, vanillic aldehyde and p-hydroxy benzaldehyde from rice bran. All of them exhibit high antioxidant activity, but their concentrations in whole rice grains needs to be further quantified.

decrease steadily during grain development. The bound phenolic in rice husk is unlike that in rice grain. The p-coumaric acid is the most abundant, followed by ferulic acid (Goufo et al., 2014). The p-hydroxybenzoic, vanillic and syringic acids exhibited the highest content at maturity or fully ripe stage (Butsat, Weerapreeyakul, & Siriamornpun, 2009).

2.4. Formation and distribution pattern

Red and black (purple) pericarp have higher antioxidant activities than white pericarp. The colored pericarp contains a unique subgroup of polyphenols. Specifically, anthocyanins are present in black bran and proanthocyanidins are present in red bran (Min, McClung, & Chen, 2011). Anthocyanins, as a large group of secondary metabolites, are water-soluble flavonoids, and may appear red, purple, or blue depending on pH. They have the structure of C6 (A ring) -C3 (C ring) -C6 (B ring) (Fig. 1b). Cyanidin 3-glucoside (Cy-3-Glu), cyanidin 3-galactoside (Cy-3-Gal), cyanidin 3-rutinoside (Cy-3-Rut), cyanidin 3-gentiobioside (Cy-3-Gent), cyanidin 3-arabidoside (Cy-3-Arab), cyanidin 3, 5-diglucoside (Cy-3,5-dGlu), cyanidin 3(600 -O-p-coumaryl)glucoside, peonidin 3-glucoside (Pn-3-Glu), peonidin 3-rutinoside (Pn-3-Rut), peonidin 3-(600 -O-pcoumaryl)glucoside, malvidin 3-glucoside (Mv-3-Glu), malvidin 3-galactoside (Mv-3-Gal), pelargonidin 3-glucoside (Pg-3-Glu), pelargonidin 3, 5-diglucoside (Pg-3,5-dGlu) have been identified from black rice with Cy-3-Glu significantly higher than others (Bordiga et al., 2014; Shao et al., 2014a, 2014b; Pereira-Caro, Cros, Yokota, & Crozier, 2013; Sompong, Siebenhandl-Ehn, Linsberger-Martin, & Berghofer, 2011). Anthocyanins have diverse bioactivity and act as an important component of traditional herbal medicines used by many countries (Konczak & Zhang, 2004). They can be derived from dried leaves, fruits, vegetables, storage roots or stems, or seeds. Up to now there are more than 500 different anthocyanins described and most of them contain glucose as a glycosylation sugar. However, free aglycone seems to have more bioactivity than the glycosides because of its hydrophobicity and small molecular size, which makes it more likely to penetrate the epithelial layer (He & Giusti, 2010). The digestion of anthocyanins begins from chewing in mouth and absorption in the stomach. The content of anthocyanins in blood rapidly increases after absorption by stomach. However, large amounts of anthocyanins are absorbed in the small intestine. Anthocyanins would release the aglycones by action of b-glucosidases, or be absorbed by the sodium-dependent glucose transport (He & Giusti, 2010). Because of the large molecular size and the lack of a free sugar moiety for transporter binding, the intact form of acylated anthocyanins are considered non-absorbable in small intestine (He et al., 2006). The unabsorbed anthocyanins travel down to the colon, where many microbes are located, and catalysis, hydrolysis and absorption occur. All the absorbed anthocyanins show high inhibition to the mobility and invasion ability of some cancers in human (Chen et al., 2006), and beneficial against cardiovascular diseases, type II diabetes and obesity (Dipti et al., 2012). However, the detailed mechanisms of their action need to be further clarified. Proanthocyanidins are a group of polymeric phenolic compounds consisting mainly of flavan-3-ol units such as afzelechin, epiafzelechin, catechin, epicatechin, gallocatechin, and epigallocatechin (Jaiswal, Jayasinghe, & Kuhnert, 2012). More complex proanthocyanidins, having the same polymeric building block, form the group of tannins. Proanthocyanidins can be A-type or Btype structure with flavan-3-ol units doubly linked by C4-C8 and

The botanical parts of whole rice grain consists of bran, embryo, and endosperm, which account for about 10–15%, 2.0–2.5%, and 80–90% of whole grain (Shao et al., 2014a). Rice bran, containing 0.9% (w/w) oil, 12.8% moisture, 12.0% ash, 17.0% protein and 58.1% carbohydrate has animal feed uses or is discarded as agricultural waste in rice milling process, and only a little is used for extraction of the edible oil (Chiou, Kobayashi, & Adachi, 2013). Rice bran is rich in polyphenols with high antioxidant activity and positive biological functions. Embryo, also called germ, is a good source of vitamin E and oryzanol. Endosperm, the largest part of whole rice grain, contains starch, protein, minerals, and vitamins and serves as the food supply for the germ and provides energy for the rest of the plant (Jonnalagadda et al., 2011). The distribution of phenolic acids in different genotypes is almost the same with bran having the most abundant and endosperm having trace amounts (Shao et al., 2014a). In the whole rice grain, the bran and the embryo contribute 59.2% of total phenolics and exhibit varietal differences (Ti et al., 2014a). Bound phenolic acids account for 88%, 89%, and 91% of the total phenolic acids in white, red, and black rice bran, respectively (Shao et al., 2014a). Despite the distribution of phenolic acids in different rice milling fractions is in an order of bran > embryo > whole grain > endosperm, their relative contents remain similar in rice grains and in different botanical parts. Geminated brown rice is valued as a functional food because it is good in digestion and absorption, and contains nutrients such as gamma-aminobutyric acid and ferulic acid in abundance compared to ordinary brown rice (Sakamoto et al., 2007). The phenolic acids at different germination stages of brown rice showed dynamic changes. Germination significantly increased TPC by 63.2% in the stages from 0 to 48 h, and the percentage contribution of bound phenolics to total (soluble free and insoluble bound) was 42.3% at 0 h and decreased slightly to 37.6% at 48 h (Ti et al., 2014b). The soluble free and insoluble bound phenolic acids detected by HPLC show an upward tendency, especially for the levels of ferulic, coumaric, syringic, and caffeic acids (Ti et al., 2014b). The accumulation pattern of phenolics in rice grains at different stages after flowering is different from that during germination. TPC of white and red rice was significantly higher at 1-week stage, but TPC of black rice was highest at maturity (Shao et al., 2014b). For the bound phenolic acids, protocatechuic acid, detected in black and red rice with the former significantly higher than the latter (Irakli et al., 2012), had relatively high levels at 1-week stage and maturity, respectively. Vanillic acid could be detected only in black rice with the mature stage having the highest and 1-week stage having the lowest content among four different development stages. Ferulic acid displays an inconsistent trend in white, red and black rice grain but peaked at the mature stage of black rice, and pcoumaric acid had similar content among all the development stages except the 1-week stage of red and black rice grain which had higher levels (Shao et al., 2014b). In rice husk, ferulic acid is the major soluble phenolic acid at all stages, and its concentration

3. Anthocyanins and proanthocyanidins 3.1. Chemistry and bioactivity

91

Y. Shao, J. Bao / Food Chemistry 180 (2015) 86–97

C2-O7 or C4-C6 and C2-O7 for the former, and linked mainly through C4-C8 or C4-C6 for the latter (Gu et al., 2003). B-type proanthocyanidin is very common in nature. In red rice, the block unit of proanthocyanidin consists of catechin and epicatechin (Gunaratne et al., 2013; Qiu, Liu, & Beta, 2009), and exists in B-type (Fig. 1c). However, whether C4-C8 or C4-C6 form has not been identified yet, and could be clarified by nuclear magnetic resonance (NMR) or other chemical analysis methods in future studies. Recently, Pereira-Caro et al. (2013) found that both A and B type of dimer, trimer, tetramer, pentamer, and hexamer of proanthocyanins coexist in both black and red rice grain. This might be attributed to the different rice genotypes used as materials. Because of high degree of polymerization and galloylation, proanthocyanidins are considered as the most effective natural antioxidants. It is reported that red rice wine possesses great anticholesterol oxidation capability, which may contribute to health benefits in preventing cardiovascular diseases, and the catechin in the red wine may play a more important role than resveratrol in inhibiting cholesterol oxidation (Tian, Wang, Abdallah, Prinyawiwatkul, & Xu, 2011).

Proanthocyanidins mainly exist in red rice grain with the total content ranging from 7.16 to 2270 lg/g dry weight (Table 3). In red rice, hexamer is the most abundant which accounts for about 16.9% for the total polymers, followed by heptamer, pentamer, and octamer which account for 10.1%, 9.7%, and 9.6%, respectively, and monomer is the lowest which account for only 0.4% (Min et al., 2012). In a different species of wild rice, the content of polymers was dramatically different; some varieties only had monomers and trace amount of dimers whereas some varieties had extremely high contents of trimers or pentamers (Qiu et al., 2009). Interestingly, a few black rice have both anthocyanins and proanthocyanins with the total content of 2050 and 153.1 lg/g, respectively (Finocchiaro, Ferrari, & Gianinetti, 2010). Anthocyanins mainly accumulate in black bran in free form which accounts for more than 95% of total anthocyanins. Less than 5% is distributed in the embryo, but it is not detectable in the endosperm (Shao et al., 2014a; Yoshimura, Zaima, Moriyama, & Kawamura, 2012; Zhang et al., 2010). The Cy-3-Glu and Pn-3-Glu content in bran accounted for 98% and 93% of the total content of Cy-3-Glu and Pn-3-Glu in black grain, and those in embryo accounted for about 2% and 7%, respectively (Shao et al., 2014a). Because of the trace amount of the other anthocyanin components, some can only be detected in the bran sample but cannot be detected in the whole grain flour sample, such as Cy-3-Rut. The distribution of proanthocyanidins was mainly in the bran which is more than 10 times higher than the whole grain red rice (Gunaratne et al., 2013). However, there is no report about the distribution of proanthocyanidins in embryo of red rice. The conjugated flavones in rice grain show a very stable profile during rice seed germination (Galland et al., 2014). During rice grain development after flowering, Cy-3-Glu and Pn-3-Glu have significantly higher contents at 2- and 3-weeks than at mature stage with the lowest at 1-week stage, which depend on the time of onset of anthocyanin accumulation and the different rates of development of bran outer layers and endosperm since anthocyanins are present in the former but not latter (Shao et al., 2014b). However, the changes of proanthocyanidins in red rice grain during germination and the different development stages after flowering have not been studied yet.

3.2. Extraction, diversity and distribution pattern In order to maintain the activity of the anthocyanins, their extraction is carried out in the acid conditions mostly using acidified solution as extractant, eg., methanol: 1 M HCl (85:15, v/v), water/acetic acid (99:1, v/v). From pH 3 to 6, anthocyanins change into colorless carbinol pseudobase or chalcone, while in neutral and slightly acidic pH, they turn into neutral quinonoidal bases showing purple to violet color (He & Giusti, 2010). For proanthocyanidins, organic solvent has been used as extractant, such as 80% methanol, acetic acid acidified acetone (Gunaratne et al., 2013; Min et al., 2012; Qiu et al., 2009). Many kinds of anthocyanins exist in rice grains. Cy-3-Glu is the most abundant ranging from 0.05 to 2.86 mg/g in different genotypes of black rice, followed by Pn-3-Glu ranging from 0 to 0.50 mg/g (Table 2). Anthocyanin content differs greatly among different genotypes with some genotypes without detectable Pn3-Glu. Besides those abundant anthocyanins, trace amounts of Cy-3-Rut, Mv-3-Glu, and Pn-3-Rut were found in some black rice (Bordiga et al., 2014). Cy-3-Rut is detectable with a high content in black rice bran and trace amount in the whole grain black rice (Shao et al., 2014a). Few red rice contain trace amounts of Cy-3Glu and cyanidin 3-O-(600 -O-p-coumaryl)glucoside (Pereira-Caro et al., 2013).

4. Genetics of polyphenols 4.1. Biosynthesis pathway A large number of genetic studies of anthocyanins have been attempted in rice, leading to a hypothesis of the CAP complemen-

Table 2 Concentrations of anthocyanins in black rice whole grain, milling fraction and products (mg/g dry weight).a Extractant

TAC

Cy-3-Glu

Pn-3-Glu

Cy-3-Rut

Cy-3-Gent

Mv-3-Glu

Pn-3-Rut

Acidified Acidified Acidified Acidified Acidified Acidified Acidified Acidified

0.87–0.88

0.14–0.16 0.06–0.12 0.14–0.47 0.11–0.13 0–0.08 – 0.009 –

0.06 0.04–0.06

0.038–0.042

0.032–0.033

0.032–0.037

2.86 0.19 0.003

0.52–0.56 0.51–1.00 0.7–2.3 0.2–1.4 0.05–1.6 0.5 0.18 –

–

–

–

0.02

–

–

–

–

Rice milling fractions Black bran Acidified methanol Black embryo Acidified methanol

6.28 0.34

3.58 0.12

0.70 0.09

Rice products Mead

28–35b

Whole grain Black rice Black rice Black rice Black rice Black rice Black rice Red rice Red rice

a

methanol methanol methanol methanol methanol water 3.47 methanol water 0.004

Fermented

1.1–2.6

0.28

References Shao et al. (2014a, 2014b) Bordiga et al. (2014) Laokuldilok et al. (2010) Sompong et al. (2011) Lee (2010) Pereira–Caro et al. (2013) Laokuldilok et al. (2010) Pereira-Caro et al. (2013) Shao et al. (2014a) Shao et al. (2014a) Katoh et al. (2011)

Range or mean value. Values express as lg/ml. TAC, total anthocyanin content; Cy-3-Glu, cyanidin 3-glucoside; Pn-3-Glu, peonidin 3-glucoside; Cy-3-Rut, cyanidin 3-rutinoside; Cy-3-Gent, cyanidin 3-gentiobioside; Mv-3-Glu, malvidin 3-glucoside; Pn-3-Rut, peonidin 3-rutinoside. b

– –

17

–

23

–

29

–

36(A); 22(B)

–

31(A); 29(B)

–

19(A); 22(B)

–

11(A); 41(B)

Rice milling fractions Red bran 80% methanol

–

–

25(B) Black riceb Black rice

a Range or mean value. TPAC, total proanthocyanidin content; Mono-, monomer; Di-, dimmer; Tri-, trimer; Tetra-, tetramer; Penta-, pentamer; Hexa-, hexamer; Hepta-, heptamer; Octa-, octamer; Nona-, nonamer; Deca-, decamer; Poly-, polymer. b The A and B in parenthesis indicate the A-type and B-type proanthocyanidin, respectively.

Gunaratne et al. (2013) 11950– 24700

0.2

–

–

Finocchiaro et al. (2010) 153.1 Methanol or acetone/water

Pereira-Caro et al. (2013) 20 325

Finocchiaro et al. (2010) Pereira-Caro et al. (2013) 42(B)

12(B)

2(A); 4(B)

7(A); 5(B)

9(A); 4(B)

8

4

2

–

– 92 2050 191

Methanol or acetone/water Acetone/water/acetic acid (70:29.5:0.5, v/v/ v) Same as above Red rice Red riceb

Qiu et al. (2009) – – – – – – 0–61.6 0–52.7 0–69.2 7.2– 18.2 7.16–239.2

0– 42.4

– – 56.9 – – – – 122.0 81.7 – – 128.2 – – 215.6 – – 123.7 – – 87.5 – – 52.5 – – 25.5 – – 5.3 1070–2270 491–2273 1272.9

Hepta- Octa- Nona- Deca- PolyHexaPentaTetraTriDiMonoTPAC Extractant

Whole grain Red rice 80% methanol Red rice 80% methanol Red rice Acetone/water/acetic acid (70:29.5:0.5, v/v/ v) Wild rice Acetone/water/acetic acid (70:29:1, v/v/v)

Table 3 Concentrations of proathocyanidins in red rice whole grain, milling fraction and products (lg/g dry weight).a

– Gunaratne et al. (2013) – Shao et al. (2014a, 2014b, 2014c) 374.1 Min et al. (2012)

Y. Shao, J. Bao / Food Chemistry 180 (2015) 86–97

References

92

tary genetic system which consists of three basic genes as C (chromogen), A (activator), and P (distributor) (Setty & Misro, 1973). The gene C governs the production of chromogen, the gene A converts or oxidizes chromogen into anthocyanins, and the regulatory gene P determines the temporal and spatial regulation of pigmentation (Setty & Misro, 1973). Currently, the biosynthetic pathway of polyphenols has become much clearer, being controlled by many structural and regulatory genes derived from L-phenylalanine (Fig. 2). By the catalysis of PAL (phenylalanine ammonia lyase) and C4H (cinnamate 4-hydroxylase), L-phenylalanine transforms into 4-coumarate which could develop into the groups of hydroxybenzoic and hydroxycinamic acids and the flavonoid derivant: 4-coumaryl CoA. Caffeic acid is produced by the catalysis of C3H (4-hydroxycinnamate 3hydroxylase) which adds hydroxyl group to C3 of the benzoic ring. By the effects of methyltransferase, caffeic and 5-hydroxyferulic acid are transferred into ferulic and sinapic acid, respectively. Anthocyanidin is produced by the catalysis of a series of enzymes, such as CHS (chalcone synthase), CHI (chalcone isomerase), F3H (flavonoid-3-hydroxylase), F30 H (flavonoid-30 -hydroxylase), DFR (dihydroflavonol 4-reductase), and ANS (anthocyanidin synthase), from 4-coumaryl-CoA and malonyl CoA. By the effects of glycosyl and methyl transferase, anthocyanidin is transformed into cyanidin-3-O-glucoside and peonidin 3-O-glucoside, respectively. Meanwhile anthocyanidin could be reduced into epi-flavan-3-ols by ANR (anthocyanidin reductase), and epi-flavan-3-ols and catechin could be transformed into proanthocyanidin by the catalysis of CPE (chain-polymerising enzyme). In the biosynthesis of flavonoids, narigenin is a very important intermediate product, which can be converted into flavones including chrysoeriol, tricin and isovitexin by a series of enzymes (Galland et al., 2014; Shih et al., 2008), or converted into dihydrokaempferol, eriodictyol, and phlobaphenes by the action of F3H, F30 H, and DFR, respectively, and also acts as the substrate of the transformation to flavans and taxfolin. The genes that participate in the biosynthesis of polyphenols interact with each other and have special functions in the accumulation of polyphenols in rice grain. It is reported that OsCHS, OsCHI, OsF3H, OsF30 H, OsDFR, and OsANS could change the seed coat of Arabidopsis from yellow to purple, and the direct interaction of OsCHS1 with OsF3H, OsF30 H, OsDFR, and OsANS1 were observed in yeast two-hybrid analysis (Shih et al., 2008). When DFR was transferred to Rcrd rice mutant, the seed color changes from brown to red, indicating that Rd encodes the dihydroflavonol 4-reductase (Furukawa et al., 2007). ANS, which succeeds DFR in the pathway, shows significant protein sequence similarity with CHS, CHI, and DFR (Turnbull et al., 2004). Over expression of ANS results in an increased accumulation of a mixture of flavonoids and anthocyanins, but with a decrease in proanthocyanidins, which might be due to a block in the anthocyanin biosynthesis pathway at the ANS mediated conversion of leucoanthocyanidin to anthocyanidin (Reddy, Reddy, Scheffler, Wienand, & Reddy, 2007). Some genes, such as CHS, F3H, DFR and ANS, have higher expression in the seeds at 13 days after flowering than any other tissues (Kim et al., 2007). Before the completion of germination, the expressions of some key flavone synthetic genes increase strongly (Galland et al., 2014). 4.2. Regulatory factors Biosynthesis of polyphenols is controlled by many structural genes, and also regulated by many regulatory genes which could up or down regulate the polyphenol accumulation. In rice, there are two kinds of regulatory genes that act as transcriptional activators: (1) R/B gene family and their homologues encode proteins with a basic helix-loop-helix (bHLH) region which is a domain present in MYC-like protein; (2) C1/Pl gene family and their homologues encode proteins that are homologous to DNA-binding

Y. Shao, J. Bao / Food Chemistry 180 (2015) 86–97

93

Fig. 2. The potential biosynthetic pathway of polyphenols in rice (redrawn based on Mustafa & Verpoorte, 2007; Shih et al., 2008). ACC: acetyl-CoA-carboxylase; A30 M: Sadenosyl-L-methionine; ANR: anthocyanidin reductase; ANS: anthocyanidin synthase; C4H: cinnamate-4-hydroxylase; CHI: chalcone isomerase; CHS: chalcone synthase; 4CL: 4-coumarate-CoA ligase; COMT: caffeic acid O-methyltransferase; CPE: chain-polymerising enzyme; DFR: dihydroflavonol 4-reductase; FGT: flavonoid 3-Oglycosyltransferase; F3H: flavonoid-3-hydroxylase; F30 H: flavonoid 30 -hydroxylase; F30 50 H: flavone 30 50 -hydroxylase; F5H: Ferulic acid 5-hydroxylase; FNS I or FNS II: flavone synthases I or flavone synthases II; LAR: leucoanthocyanidin reductase; MT: O-methyltransferase; OGT: O-glucosyltransferase; PAL: phenylalanine ammonia lyase; ROMT: rice O-methyltransferase.

domains of MYB-like proteins. Both families encode similar proteins with different expression patterns. It is the interaction between regulatory proteins and enzymes participating in the polyphenol biosynthesis pathway that induce the pigmentation in different parts of rice plant. Many regulatory genes involved in the anthocyanin pigmentation are first found in maize, but those similar genes are responsible for anthocyanin pigmentation in rice. Genetic analyses have demonstrated that the combination of R and C1 is responsible for pigmentation in the kernel, while the combination of B and Pl is responsible for pigmentation in mature tissues of the plant, such as the husk and leaves. Os9BGlu31, a glycoside hydrolase family GH1 transglycosidase in rice, acts to transfer glucose between phenolic acids, phytohormones, and flavonoids (Luang et al., 2013). Classical genetic analyses have found that two loci, Pb (Prp-b) and Pp (Prp-a), which are located on chromosome 4 and 1, respectively, are associated with anthocyanin pigmentation in black rice seed coat. Pb locus might be allelic to the OsB1 (Sakamoto et al., 2001) or Ra (Wang & Shu, 2007), which has a 2 bp deletion within exon 7. The genes responsible for black pigments formation in black rice await further characterization and function confirmation at molecular levels. Rc (brown pericarp and seed coat) and Rd (red pericarp and seed coat) are domestication-related genes that are required for red pericarp formation in rice. When present together, they produce red seed color. Rc in absence of Rd produces brown

seeds, and Rd alone has no phenotype (Sweeney, Thomson, Pfeil, & McCouch, 2006). Three Rc alleles have been cloned: Rc, which produces brown spots on a reddish-brown background; Rc-s, which exhibits a light red color; and rc, which is a null allele (Furukawa et al., 2007; Sweeney et al., 2006). The dominant red allele differed from the recessive white allele is caused by a 14 bp deletion within exon 6 on chromosome 7 that knock out the bHLH domain of the protein (Sweeney et al., 2006). Rd, encoding dihydroflavonol 4-reductase, has also been cloned (Furukawa et al., 2007). Kim et al. (2010) detected 82 transcription factor genes by microarray which might be associated with anthocyanin accumulation in black rice, and found 12 putative genes identified from comparison between the white cultivar and two black cultivars. They also found 15 genes with nine up-regulated genes and six down-regulated genes that play a regulatory role or relate to anthocyanin metabolism in the biosynthesis of polyphenols (Kim et al., 2011). However, the function of these transcription factors, especially in the biosynthetic pathway of polyphenols, needs to be further studied. 4.3. Improvement of polyphenols and antioxidant capacity With the economic development and the improvement of living standards, the overweight or obesity is not only happening in

94

Y. Shao, J. Bao / Food Chemistry 180 (2015) 86–97

adults, but also rising in young children. Overweight is also associated with additional risk factors for cardiovascular diseases, such as raised blood cholesterol and elevated blood pressure, and with type II diabetes and other potential psychological health issues. Because of unhealthy diet and lack of physical exercise, childhood obesity will likely continue to rise. Therefore, production of rice varieties with nutraceutical applications is a large challenge for rice-eating countries. Many epidemiological studies show the positive effects of polyphenols in rice grain against many chronic diseases, such as cardiovascular diseases, type II diabetes and some cancers (Dipti et al., 2012). The improvement of polyphenols and antioxidant capacity in rice grain is of significant importance and could be conducted by special breeding projects. Conventional plant breeding is a simple way to breed new varieties high in polyphenols and antioxidant capacity in rice grain. Some advanced breeding lines made from the cross between the black and white rice as parents have higher total phenolic content, total anthocyanin content and antioxidant activities than the black parent (Zhang, Shao, Bao, & Beta, 2015). It is also reported that red pericarp introgression lines derived from interspecific crosses of rice exhibit significant positive differences in the content of phenolic constituents and antioxidant properties with good grain quality characteristic over their parents (Sharma, Kaur, Mangat, & Singh, 2014). Molecular breeding which includes identificantion of quantitative trait loci (QTL) and marker assisted selection is becoming more and more popular. Association mapping has revealed some molecular markers highly associated with phenolic, flavonoid content and antioxidant capacity (Shao et al., 2011, 2014c). All of the QTLs provide potential markers for rice breeders to improve nutritional quality by marker-assisted selection or help scientists to further identify genes that are responsible for polyphenol content in rice whole grain. The markers developed from the major genes responsible for pigmentation formation in red and black rice are particularly useful (Shao et al., 2011). For example, Rc of red pericarp has a 14 bp deletion within exon 6, which caused proanthocyanidin accumulation in the rice bran. The putative Ra gene of black pericarp has a 2 bp deletion. Transgenesis is a process of introducing an exogenous gene or genes into a living organism, where the organism will show a new characteristic and transmit it to the offspring. There are some successful cases to improve flavonoid and antioxidant capacity in rice. When IFS (isoflavone synthase) from soybean is transformed to rice, the rice plant could produced many kinds of flavonoids (Sreevidya, Rao, Sullia, Ladha, & Reddy, 2006). The introduction of Rc into rcrd rice alters the coloration of seed from white to brown in transgenic rice, and the introduction of Rd into Rcrd rice changes the seed color from brown to red, both of which suggest that some genes are associated with the biosynthesis of proanthocyanidins in rice (Sweeney et al., 2006). Production of purple anthocyanin pigments has also been observed in the transformed callus and plants upon the introduction of maize regulatory genes C1 (colored-1), R (red) and the structural gene C2 (colore-2, encoding chalcone synthase) (Gandikota et al., 2001). Transgenic rice overexpressing anthocyanidin synthase accumulates a mixture of flavonoids leading to an increased antioxidant potential (Reddy et al., 2007). Therefore, in order to breed a variety that contain high nutraceutical properties, introduction of the homologous genes or regulator genes that control the complex nutraceutical properties of rice is an effective way to enhance the functional components of rice grain.

5. Applications and health benefits Whole rice grain is recommended as part of a healthful diet. It is popular in western countries, and will be gradually more accepted

in developing countries, because of its valuable sources of nutrition and high antioxidant activity that play important roles in disease prevention, such as reduction of the risks of cardiovascular disease, type II diabetes, obesity and cancer. In consideration of the nutrition, tasting, and diversity of whole rice grain food, processors are trying to develop good-tasting food with high nutritional quality. Whole grain rice can be utilized in a wide range of foods such as crisped rice, puffed rice, rice crackers, cereal bars, rice flour, rice noodles, fermented condiments (miso and mirin), rice malt, rice wine. The major traditional rice grain products have generally been produced from polished rice. With the understanding of the health benefits of whole rice grain, more and more traditional type whole rice grain food products with higher functional factors may emerge in the market. Mead, a kind of honey wine, has thousands years of history in Europe and Africa. If it is made from honey and black rice grain, it has higher total phenolic content and anthocyanins than that made from honey and glucose (Katoh, Koguchi, Saigusa, & Teramoto, 2011). The wine made from red rice possesses great anti-cholesterol oxidation capability, and the catechin in the red wine may play a more important role than resveratrol in inhibiting cholesterol oxidation (Tian et al., 2011). Beverages made from uncooked black rice would have higher anthocyanin content and antioxidant capacity than that made from cooked black rice, because of the hydrothermal effect of the anthocyanins (Hiemori, Koh, & Mitchell, 2009). Red rice vinegar, which is an acetic acid fermentation-derived traditional product, has significantly higher total phenolics and antioxidant capacity than unfermented red rice (Hsieh, Lu, Lin, Lai, & Chiou, 2013). The fermented rice sap, especially the fermented purple grain sap, could be further developed as functional foods and cosmetics (Manosroi, Ruksiriwanich, Kietthanakorn, Manosroi, & Manosroi, 2011), because cyanidin 3glucoside, caffeic and ferulic acid in the rice bran are useful in the control of skin aging. Many epidemiological and intervention studies show the positive effects of consuming whole rice grain (Table 4). The studies indicate that the whole black rice grain, specifically the anthocyanins, cyanidin 3-glucoside and peonidin 3-glucoside, contribute to inhibiting the invasion and mobility ability of human hepatocellular carcinoma (SKHep-1) cells with a reduced expression of matrix metalloproteinase-9 and urokinase-type plasminogen activator (Chen et al., 2006). The anthocyanin extract from black rice also play an important role in reducing hypertriglyceridemia and adverse effects of alcohol (Hou, Qin, & Ren, 2010; Yang et al., 2011). The phenolic acids from brown rice bran show putative breast and colon cancer chemopreventive properties (Hudson, Dinh, Kokubun, Simmonds, & Gescher, 2000). When dyslipidemic rats were fed with high fat diets supplemented with anthocyanin extracted from black rice, the platelet hyperactivity and body weight gain was significantly lower than in those fed with only high fat diet, suggesting that dietary intake of AEBR facilitates the maintenance of optimal platelet function in dyslipidemic rats induced by high fat diets (Yang et al., 2011). Recently, it was reported that the extract from red rice grain has potential anti-inflammatory effect in a dose-dependent manner (Niu et al., 2013). Because whole rice grain is harder to chew and less tasty according to traditional sensory perception, polished rice is usually consumed in major rice-eating countries. Pre-germinated rice is becoming popular because of its functional components and taste. It is produced by soaking brown rice in water to induce slight germination, and has higher nutritional value than ungerminated brown rice. It is reported that pre-germinated brown rice could enhance mental health and immunity and protect against diabetic deterioration (Sakamoto et al., 2007; Usuki et al., 2007). In order to have a healthy life, whole rice grain, especially red and black rice grain could be used as an additive to the polished

95

Y. Shao, J. Bao / Food Chemistry 180 (2015) 86–97 Table 4 Intervention studies on health effects of whole rice grain, rice bran, germinated brown rice or related extracts in various target groups. Reference diets

Target group

Design and dietary intervention

Effects

References

Anthocyanin extract from black rice Anthocyanin extract from black rice (AEBR)

Human cells

Inhibit the invasion and motility of various cancer cells AEBR could reduce platelet hyperactivity, hypertriglyceridemia, and body weight gain, and maintain optimal platelet function in dyslipidemic rats induced by high fat diets.

Chen et al. (2006) Yang et al. (2011)

Anthocyanin-rich extract from black rice (AEBR)

Male Wistar rats with the weight of 150 ± 20 g

Treated with extract, Cy-3-Glu, Pn-3-Glu in different concentrations, respectively Randomly divided into 3 groups and fed with AIN-93G normal diet (control), AIN-93G diet containing high fat and cholesterol, and high fat supplemented with AEBR (5 g/kg diet), respectively Randomly divided into 5 groups and fed with distilled water (3.7 g/kg body weight), ethanol (3.7 g/kg), ethanol (3.7 g/kg) + AEBR (0.5 g/kg), ethanol (3.7 g/kg) + AEBR (0.25 g/kg), and ethanol (3.7 g/kg) + AEBR (0.125 g/kg), respectively

Hou et al. (2010)

Red rice extracts

RAW 264.7 mouse macrophage cells

Administration of AEBR (500 mg/kg) + alcohol significantly decreased the activities of liver enzymes, MDA levels and concentrations of serum, hepatic triglyceride and total cholesterol, which indicated that AEBR has a beneficial effect in reducing the adverse effect of alcohol red rice extracts suppressed LPS stimulated IL-1b, IL-6 and COX-2 mRNA expressions in a dosedependent manner, which suggested that red rice have potential anti-inflammatory effect

Whole grain diet containing barley or whole wheat and brown rice Rice bran and oat bran

21 Non-hypertensive men with elevated plasma cholesterol levels 11 human subjects with moderatelyhigh blood cholesterol Human breast and colon cancer cell lines 41 Breast-feeding mothers

Increasing whole grain foods in a healthy diet can reduce cardiovascular risk

Hallfrisch, Scholfield, and Behall (2003) Hegsted, Windhauser, Morris, and Lester (1993) Hudson et al. (2000)

Eight phenolic acids from brown rice bran Pre-germinated brown rice and white rice Pre-germinated brown rice (PR), brown rice (BR), and white rice (WR)

Male dyslipidemic rats (n = 36), aged 8– 9 wk and weighing 180–220 g

Male Wistar rats

Treated with cell media containing 10 lg/ml or 100 lg/ml of red rice extracts for 24 h, and then added lipopolysaccharide (LPS) at initial concentration of 10 ng/ml, and incubated at 37 °C with 5% CO2 American Heart Association step 1 diet for 2-wk, and then consumed diets with brown rice/whole wheat, barley, or a combination for 5-wk in a Latin square A control diet without bran, 100 g/day stabilized rice bran or 100 g/day oat bran for two 3-wk periods in a crossover design

Rice and oat bran reduce the blood cholesterol levels with equally effective

Bran extract and its fractions at 100 lg/ml on cell viability and colony forming ability

Brown rice and bran may contain compounds with putative cancer chemopreventive properties

Randomly divided into two groups, took pregerminated brown rice and white rice (control) as their staple diet for 2-wk The diabetic rats (n = 31) and non-diabetic rats (n = 22) were divided into 4 groups with the diets of WR, BR, PR, and AIN93G as control, respectively

The pre-germinated brown rice could reduce depression, anger-hostility, and fatigue, and increase Ig-A level and resistance to stress PR treatment could protect diabetic deterioration and physiological parameters of diabetic neuropathy

rice grain in order to increase the functional component in our diets. Pre-germinated rice, whole rice grain beverages or soups have more nutraceuticals and taste better than whole grain rice alone; these foods are good choices to prevent chronic diseases. 6. Future challenges Although great progress has been made in understanding the polyphenols in whole grain rice, there are challenges that should be considered for better use of whole grain in the near future. Some of these issues have been mentioned above and need to be addressed in the future, for example, the specific and rare polyphenol compounds in different rice genotypes, the components with specific protective effects on diseases, the mechanisms for consumption of whole grain rice to prevent chronic diseases, and development of new food high in nutraceutical components. In addition, there are three challenges that need to be further considered carefully. Global climate change has a dramatic impact on crop production. The global concentration of atmospheric CO2 (carbon dioxide) increased from 290 to 375 lmol/ml during the last 100 years, and it might reach to 550 lmol/ml by the middle of the current century. The increase of CO2 concentration in the air will affect the growth of crops, yield, and even the quality of the product. When rice plants were grown in two levels of atmospheric CO2 (375 and 550 lmol/ml), the total phenolic, and total flavonoid content of all rice milling fractions in the higher CO2 level decreased about 3–18%, and 8–14%, respectively, with the highest reduction of sinapic acid (167%), p-hydroxybenzoic acid (100%) and tricin (12%) in brown rice, white rice and rice bran, respectively, compared to

Niu et al. (2013)

Sakamoto et al. (2007) Usuki et al. (2007)

those in normal CO2 level (Goufo et al., 2014). How to mitigate the effects of climate change while improving nutraceutical property in whole grain rice needs to be further addressed. Food safety is a serious problem in our daily life. How to produce the rice grains with high nutraceutical property without hazardous composition is a big challenge. Rice plant uptakes heavy metals, such as Cd, As, Hg, Pb and Cr from polluted soil, and these metals may be transported to the grain, and mostly accumulate in bran. The pesticides applied to rice plant might remain in the rice grain as residues. These problems occur in rice cultivation, and influence global food safety. Polyphenols occur mainly in the bran layer of the whole grain rice, so the whole grain rice would harm our health if it is polluted with heavy metals and pesticide residues. Organic farming is a form of agriculture that relies on techniques such as crop rotation, green manure, and biological pest control. Rice grain cultivated from organic farming has significantly higher antioxidant values than that from conventional farming (Kesarwani, Chiang, Chen, & Su, 2013). Therefore, organic rice grain production may produce rice with higher nutraceutical property without pesticide residue and heavy metal pollution. New technologies are challenged to produce rice with high nutraceutical property. First, the genes controlling the black rice pigment formation need to be cloned and functionally confirmed at the molecular level. Second, breeding of rice varieties with high nutraceutical property could also be conducted with the aid of molecular markers. For example, some black rice contains both anthocyanins and proanthocyanidins (Finocchiaro et al., 2010), and these kinds of black rice are visually difficult to be identified, but are easily identified with molecular markers. It is possible to pyramid the genes for black and red pigmentation together. Third,

96

Y. Shao, J. Bao / Food Chemistry 180 (2015) 86–97

new rice with multiple functionalities could be developed, for example, to develop black or red rice with large embryo conferring new rice high in c-aminobutyric acid (GABA) which has chemopreventive effects, or with low glutelin-content mutation to confer new rice suitable for consumers with kidney disease. Fourth, using current knowledge of anthocyanin biosynthesis, how to breed rice with black endosperm so that the anthocyanins could be accumulated in the endosperm needs to be addressed by the discovery of important genes specifically contributing to the accumulation of anthocyanins in the rice endosperm.

7. Conclusion Phenolic and flavonoid compounds exist in whole rice grain. Phenolic acids in rice grain are present in soluble form including free and conjugated forms, and in insoluble form (also called bound phenolics). The insoluble form accounts for about 90% of total phenolic acids. In the whole rice grain, ferulic, p-coumaric, isoferulic, syringic, vanillic, sinapic, caffeic, (+)-catechin, p-hydroxybenzoic, and protocatechuic acid have been identified, of which ferulic acid is the most abundant bound phenolic acid. In some colored grain, protocatechuic acid is highest among all the soluble phenolic acids, followed by ferulic and vanillic acids. The total phenolic content is highest in the seed at one week after flowering in white and red rice but at the mature stage in black rice. The soluble and insoluble phenolic acids may have different health benefits. Soluble phenolic acid may play an important role in inhibition activities against oxidation of low-density lipoprotein (LDL) cholesterol and liposome, and insoluble phenolic acid could be preventative against colon cancer, as well as have anti-inflammatory effects. The pigmentation of black and red grain is due to the accumulation of anthocyanins and proanthocyanidins in the pericarp and aleurone layers. In black rice, a total of 14 kinds of anthocyanin have been identified with Cy-3-Glu the most abundant. The anthocyanins accumulate to the highest content at 2–3 weeks stage after flowering. In red rice, proanthocyanidins have been identified from monomer to decamer with hexamer the most abundant, followed by pentamer, heptamer, octamer, and the monomer the least. Proanthocyanidins in rice are usually present in B type, but researchers find that both A and B type proanthocyanidins coexist in some red and black rice. The anthocyanins and proanthocyanidins are distributed mainly in bran layer; they are good to reduce the risk of cardiovascular diseases, type II diabetes and obesity, and also play important roles in inhibiting the mobility and invasion ability of some cancers and cholesterol oxidation. However, the specific mechanisms of whole gain foods in disease prevention need to be further clarified. The biosynthetic pathway and the genetic basis are also reviewed in this paper, and some challenges need to be addressed in further studies. How to produce the rice varieties with high accumulation of nutraceutical property components by breeding or by varying agronomic practices is an important priority. Climate-proof rice varieties that could adapt to changing environment need to be bred and produced, and appropriate processing methods which could improve sensory properties of whole rice grain need to be explored.

References Bashir, K., Ishimaru, Y., Shimo, H., Kakei, Y., Senoura, T., Takahashi, R., et al. (2011). Rice phenolics efflux transporter 2 (PEZ2) plays an important role in solubilizing apoplasmic iron. Soil Science and Plant Nutrition, 57, 803–812. Bordiga, M., Gomez-Alonso, S., Locatelli, M., Travaglia, F., Coïsson, J. D., HermosinGutierrez, I., et al. (2014). Phenolics characterization and antioxidant activity of six different pigmented Oryza sativa L. cultivars grown in Piedmont (Italy). Food Research International, 65, 282–290.

Bunzel, M., Ralph, J., Kim, H., Lu, F. C., Ralph, S. A., Marita, J. M., et al. (2003). Sinapate dehydrodimers and sinapate-ferulate heterodimers in cereal dietary fiber. Journal of Agricultural and Food Chemistry, 51, 1427–1434. Butsat, S., Weerapreeyakul, N., & Siriamornpun, S. (2009). Changes in phenolic acids and antioxidant activity in Thai rice husk at five growth stages during grain development. Journal of Agricultural and Food Chemistry, 57, 4566–4571. Cai, R., Arntfield, S. D., & Charlton, J. L. (1999). Structural changes of sinapic acid during alkali-induced air oxidation and the development of colored substances. Journal of the American Oil Chemists Society, 76, 757–764. Chandrasekara, A., & Shahidi, F. (2011). Bioactivities and antiradical properties of millet grains and hulls. Journal of Agricultural and Food Chemistry, 59, 9563–9571. Chen, P. N., Kuo, W. H., Chiang, C. L., Chiou, H. L., Hsieh, Y. S., & Chu, S. C. (2006). Black rice anthocyanins inhibit cancer cells invasion via repressions of MMPs and u-PA expression. Chemico-Biological Interactions, 163, 218–229. Chiou, T. Y., Kobayashi, T., & Adachi, S. (2013). Characteristics and antioxidative activity of the acetone-soluble and -insoluble fractions of a defatted rice bran extract obtained by using an aqueous organic solvent under subcritical conditions. Bioscience Biotechnology and Biochemistry, 77, 624–630. Dipti, S. S., Bergman, C., Indrasari, S. D., Herath, T., Hall, R., Lee, H., et al. (2012). The potential of rice to offer solutions for malnutrition and chronic diseases. Rice, 5, 1–18. Finocchiaro, F., Ferrari, B., & Gianinetti, A. (2010). A study of biodiversity of flavonoid content in the rice caryopsis evidencing simultaneous accumulation of anthocyanins and proanthocyanidins in a black-grained genotype. Journal of Cereal Science, 51, 28–34. Furukawa, T., Maekawa, M., Oki, T., Suda, I., Iida, S., Shimada, H., et al. (2007). The Rc and Rd genes are involved in proanthocyanidin synthesis in rice pericarp. Plant Journal, 49, 91–102. Galland, M., Boutet-Mercey, S., Lounifi, I., Godin, B., Balzergue, S., Grandjean, O., et al. (2014). Compartmentation and dynamics of flavone metabolism in dry and germinated rice seeds. Plant and Cell Physiology, 55, 1646–1659. Gandikota, M., Kochko, A. D., Chen, L., Ithal, N., Fauquet, C., & Reddy, A. R. (2001). Development of transgenic rice plants expressing maize anthocyanin genes and increased blast resistance. Molecular Breeding, 7, 73–83. Garcia-Conesa, M. T., Plumb, G. W., Waldron, K. W., Ralph, J., & Williamson, G. (1997). Ferulic acid dehydrodimers from wheat bran: isolation, purification and antioxidant properties of 8-O-4-diferulic acid. Redox Report, 3, 319–323. Geissmann, T., & Neukom, H. (1973). On the composition of the water-soluble wheat flour pentosans and their oxidative gelation. Lebensmittelhygiene Wissenschaft Technology, 6, 59–62. Gorinstein, S., Vargas, O. J. M., Jaramillo, N. O., Salas, I. A., Ayala, A. L. M., ArancibiaAvila, P., et al. (2007). The total polyphenols and the antioxidant potentials of some selected cereals and pseudocereals. European Food Research and Technology, 225, 321–328. Goufo, P., Pereira, J., Figueiredo, N., Oliveira, M. B. P. P., Carranca, C., Rosa, E. A. S., et al. (2014). Effect of elevated carbon dioxide (CO2) on phenolic acids, flavonoids, tocopherols, tocotrienols, gamma-oryzanol and antioxidant capacities of rice (Oryza sativa L.). Journal of Cereal Science, 59, 15–24. Grabber, J. H., Hatfield, R. D., Ralph, J., Zon, J., & Amrhein, N. (1995). Ferulate crosslinking in cell-walls isolated from maize cell-suspensions. Phytochemistry, 40, 1077–1082. Gu, L. W., Kelm, M. A., Hammerstone, J. F., Beecher, G., Holden, J., Haytowitz, D., et al. (2003). Screening of foods containing proanthocyanidins and their structural characterization using LC–MS/MS and thiolytic degradation. Journal of Agricultural and Food Chemistry, 51, 7513–7521. Gunaratne, A., Wu, K., Li, D., Bentota, A., Corke, H., & Cai, Y. Z. (2013). Antioxidant activity and nutritional quality of traditional red-grained rice varieties containing proanthocyanidins. Food Chemistry, 138, 1153–1161. Hallfrisch, J., Scholfield, D. J., & Behall, K. M. (2003). Blood pressure reduced by whole grain diet containing barley or whole wheat and brown rice in moderately hypercholesterolemic men. Nutrition Research, 23, 1631–1642. He, J., & Giusti, M. M. (2010). Anthocyanins: Natural colorants with healthpromoting properties. Annual Review of Food Science and Technology, 1, 163–187. He, J., Magnuson, B. A., Lala, G., Tian, Q., Schwartz, S. J., & Giusti, M. M. (2006). Intact anthocyanins and metabolites in rat urine and plasma after 3 months of anthocyanin supplementation. Nutrition and Cancer, 54, 3–12. Hegsted, M., Windhauser, M. M., Morris, S. K., & Lester, S. B. (1993). Stabilized rice bran and oat bran lower cholesterol in humans. Nutrition Research, 13, 387–398. Hiemori, M., Koh, E., & Mitchell, A. E. (2009). Influence of cooking on anthocyanins in black rice (Oryza sativa L. japonica var. SBR). Journal of Agricultural and Food Chemistry, 57, 1908–1914. Hou, Z., Qin, P., & Ren, G. (2010). Effect of anthocyanin-rich extract from black rice (Oryza sativa L. Japonica) on chronically alcohol-induced liver damage in rats. Journal of Agricultural and Food Chemistry, 58, 3191–3196. Hsieh, C. W., Lu, Y. R., Lin, S. M., Lai, T. Y., & Chiou, R. Y. Y. (2013). Stability of monacolin K and citrinin and biochemical characterization of red-koji vinegar during fermentation. Journal of Agricultural and Food Chemistry, 61, 7276–7283. Huang, S. H., & Ng, L. T. (2012). Quantification of polyphenolic content and bioactive constituents of some commercial rice varieties in Taiwan. Journal of Food Composition and Analysis, 26, 122–127. Hudson, E. A., Dinh, P. A., Kokubun, T., Simmonds, M. S., & Gescher, A. (2000). Characterization of potentially chemopreventive phenols in extracts of brown rice that inhibit the growth of human breast and colon cancer cells. Cancer Epidemiology Biomarkers & Prevention, 9, 1163–1170.

Y. Shao, J. Bao / Food Chemistry 180 (2015) 86–97 Irakli, M. N., Samanidou, V. F., Biliaderis, C. G., & Papadoyannis, I. N. (2012). Simultaneous determination of phenolic acids and flavonoids in rice using solid-phase extraction and RP-HPLC with photodiode array detection. Journal of Separation Science, 35, 1603–1611. Jaiswal, R., Jayasinghe, L., & Kuhnert, N. (2012). Identification and characterization of proanthocyanidins of 16 members of the Rhododendron genus (Ericaceae) by tandem LC–MS. Journal of Mass Spectrometry, 47, 502–515. Jonnalagadda, S. S., Harnack, L., Liu, R. H., McKeown, N., Seal, C., Liu, S., et al. (2011). Putting the whole grain puzzle together: Health benefits associated with whole grains summary of American Society for Nutrition 2010 Satellite Symposium. Journal of Nutrition, 141, 1011S–1022S. Jun, H. I., Song, G. S., Yang, E. I., Youn, Y., & Kim, Y. S. (2012). Antioxidant activities and phenolic compounds of pigmented rice bran extracts. Journal of Food Science, 77, C759–C764. Katoh, T., Koguchi, M., Saigusa, N., & Teramoto, Y. (2011). Production and antioxidative activity of mead made from various types of honey and black rice (Oryza sativa var. Indica cv. Shiun). Food Science and Technology Research, 17, 149–154. Kesarwani, A., Chiang, P. Y., Chen, S. S., & Su, P. C. (2013). Antioxidant activity and total phenolic content of organically and conventionally grown rice cultivars under varying seasons. Journal of Food Biochemistry, 37, 661–668. Kim, B. G., Kim, J. H., Min, S. Y., Shin, K. H., Kim, J. H., Kim, H. Y., et al. (2007). Anthocyanin content in rice is related to expression levels of anthocyanin biosynthetic genes. Journal of Plant Biology, 50, 156–160. Kim, C., Kikuchi, S., Kim, Y., Park, S., Yoon, U., Lee, G., et al. (2010). Computational identification of seed-specific transcription factors involved in anthocyanin production in black rice. Biochip Journal, 4, 247–255. Kim, C. K., Cho, M. A., Choi, Y. H., Kim, J. A., Kim, Y. H., Kim, Y. K., et al. (2011). Identification and characterization of seed-specific transcription factors regulating anthocyanin biosynthesis in black rice. Journal of Applied Genetics, 52, 161–169. Konczak, I., & Zhang, W. (2004). Anthocyanins—More than nature’s colours. BioMed Research International, 2004, 239–240. Laokuldilok, T., Shoemaker, C. F., Jonqkaewwattana, S., & Tulyathan, V. (2010). Antioxidants and antioxidant activity of several pigmented rice brans. Journal of Agricultural and Food Chemistry, 59, 193–199. Lee, J. H. (2010). Identification and quantification of anthocyanins from the grains of black rice (Oryza sativa L.) varieties. Food Science and Biotechnology, 19, 391–397. Luang, S., Cho, J. I., Mahong, B., Opassiri, R., Akiyama, T., Phasai, K., et al. (2013). Rice Os9BGlu31 is a transglucosidase with the capacity to equilibrate phenylpropanoid, flavonoid, and phytohormone glycoconjugates. Journal of Biological Chemistry, 288, 10111–10123. Manosroi, A., Ruksiriwanich, W., Kietthanakorn, B. O., Manosroi, W., & Manosroi, J. (2011). Relationship between biological activities and bioactive compounds in the fermented rice sap. Food Research International, 44, 2757–2765. Massaretto, I. L., Madureira Alves, M. F., Mussi de Mira, N. V., Carmona, A. K., & Lanfer Marquez, U. M. (2011). Phenolic compounds in raw and cooked rice (Oryza sativa L.) and their inhibitory effect on the activity of angiotensin Iconverting enzyme. Journal of Cereal Science, 54, 236–240. Min, B., Gu, L., McClung, A. M., Bergman, C. J., & Chen, M. H. (2012). Free and bound total phenolic concentrations, antioxidant capacities, and profiles of proanthocyanidins and anthocyanins in whole grain rice (Oryza sativa L.) of different bran colours. Food Chemistry, 133, 715–722. Min, B., McClung, A. M., & Chen, M. H. (2011). Phytochemicals and antioxidant capacities in rice brans of different color. Journal of Food Science, 76, C117–C126. Mustafa, N. R., & Verpoorte, R. (2007). Phenolic compounds in catharanthus roseus. Phytochemistry Reviews, 6, 243–258. Niu, Y. G., Gao, B. Y., Slavin, M., Zhang, X. W., Yang, F., Bao, J. S., et al. (2013). Phytochemical compositions, and antioxidant and anti-inflammatory properties of twenty-two red rice samples grown in Zhejiang. LWT – Food Science and Technology, 54, 521–527. Park, S. Y., Ha, S. H., Lim, S. H., Jung, J. Y., Lee, S. M., Yeo, Y., et al. (2012). Determination of phenolic acids in Korean rice (Oryza sativa L.) cultivars using gas chromatography-time-of-flight mass spectrometry. Food Science and Biotechnology, 21, 1141–1148. Pereira-Caro, G., Cros, G., Yokota, T., & Crozier, A. (2013). Phytochemical profiles of black, red, brown, and white rice from the Camargue region of France. Journal of Agricultural and Food Chemistry, 61, 7976–7986. Qiu, Y., Liu, Q., & Beta, T. (2009). Antioxidant activity of commercial wild rice and identification of flavonoid compounds in active fractions. Journal of Agricultural and Food Chemistry, 57, 7543–7551. Qiu, Y., Liu, Q., & Beta, T. (2010). Antioxidant properties of commercial wild rice and analysis of soluble and insoluble phenolic acids. Food Chemistry, 121, 140–147. Que, F., Mao, L., & Pan, X. (2006). Antioxidant activities of five Chinese rice wines and the involvement of phenolic compounds. Food Research International, 39, 581–587. Reddy, A. M., Reddy, V. S., Scheffler, B. E., Wienand, U., & Reddy, A. R. (2007). Novel transgenic rice overexpressing anthocyanidin synthase accumulates a mixture of flavonoids leading to an increased antioxidant potential. Metabolic Engineering, 9, 95–111. Renger, A., & Steinhart, H. (2000). Ferulic acid dehydrodimers as structural elements in cereal dietary fibre. European Food Research and Technology, 211, 422–428. Sakamoto, S., Hayashi, T., Hayashi, K., Murai, F., Hori, M., Kimoto, K., et al. (2007). Pre-germinated brown rice could enhance maternal mental health and immunity during lactation. European Journal of Nutrition, 46, 391–396.

97

Sakamoto, W., Ohmori, T., Kageyama, K., Miyazaki, C., Saito, A., Murata, M., et al. (2001). The Purple leaf (Pl) locus of rice: The Plw allele has a complex organization and includes two genes encoding basic helix-loop-helix proteins involved in anthocyanin biosynthesis. Plant and Cell Physiology, 42, 982–991. Setty, M., & Misro, B. (1973). Complementary genic conlplex for anthocyanin pigmentation in the apiculus of rice (Oryza sativa L.). Canadian Journal of Genetics and Cytology, 15, 779–789. Setyaningsih, W., Saputro, I. E., Palma, M., & Barroso, C. G. (2015). Optimization and validation of the microwave-assisted extraction of phenolic compounds from rice grains. Food Chemistry, 169, 141–149. Shao, Y. F., Jin, L., Zhang, G., Lu, Y., Shen, Y., & Bao, J. S. (2011). Association mapping of grain color, phenolic content, flavonoid content and antioxidant capacity in dehulled rice. Theoretical and Applied Genetics, 122, 1005–1016. Shao, Y. F., Xu, F., Sun, X., Bao, J. S., & Beta, T. (2014a). Identification and quantification of phenolic acids and anthocyanins as antioxidants in bran, embryo and endosperm of white, red and black rice kernels (Oryza sativa L.). Journal of Cereal Science, 59, 211–218. Shao, Y. F., Xu, F., Sun, X., Bao, J. S., & Beta, T. (2014b). Phenolic acids, anthocyanins, and antioxidant capacity in rice (Oryza sativa L.) grains at four stages of development after flowering. Food Chemistry, 143, 90–96. Shao, Y. F., Tang, F., Huang, Y., Xu, F., Chen, Y., Tong, C., et al. (2014c). Analysis of Genotype  environment interactions for polyphenols and antioxidant capacity of rice by association mapping. Journal of Agricultural and Food Chemistry, 62, 5361–5368. Sharma, N., Kaur, R., Mangat, G. S., & Singh, K. (2014). Red pericarp introgression lines derived from interspecific crosses of rice: physicochemical characteristics, antioxidative properties and phenolic content. Journal of the Science of Food and Agriculture, 94, 2912–2920. Shen, Y., Jin, L., Xiao, P., Lu, Y., & Bao, J. S. (2009). Total phenolics, flavonoids, antioxidant capacity in rice grain and their relations to grain color, size and weight. Journal of Cereal Science, 49, 106–111. Shih, C. H., Chu, H., Tang, L. K., Sakamoto, W., Maekawa, M., Chu, I. K., et al. (2008). Functional characterization of key structural genes in rice flavonoid biosynthesis. Planta, 228, 1043–1054. Sompong, R., Siebenhandl-Ehn, S., Linsberger-Martin, G., & Berghofer, E. (2011). Physicochemical and antioxidative properties of red and black rice varieties from Thailand, China and Sri Lanka. Food Chemistry, 124, 132–140. Sreevidya, V., Rao, C. S., Sullia, S., Ladha, J. K., & Reddy, P. M. (2006). Metabolic engineering of rice with soybean isoflavone synthase for promoting nodulation gene expression in rhizobia. Journal of Experimental Botany, 57, 1957–1969. Sweeney, M. T., Thomson, M. J., Pfeil, B. E., & McCouch, S. (2006). Caught redhanded: Rc encodes a basic helix-loop-helix protein conditioning red pericarp in rice. Plant Cell, 18, 283–294. Tabaraki, R., & Nateghi, A. (2011). Optimization of ultrasonic-assisted extraction of natural antioxidants from rice bran using response surface methodology. Ultrasonics Sonochemistry, 18, 1279–1286. Ti, H., Li, Q., Zhang, R., Zhang, M., Deng, Y., Wei, Z., et al. (2014a). Free and bound phenolic profiles and antioxidant activity of milled fractions of different indica rice varieties cultivated in southern China. Food Chemistry, 159, 166–174. Ti, H., Zhang, R., Zhang, M., Li, Q., Wei, Z., Zhang, Y., et al. (2014b). Dynamic changes in the free and bound phenolic compounds and antioxidant activity of brown rice at different germination stages. Food Chemistry, 161, 337–344. Tian, L., Wang, H., Abdallah, A. M., Prinyawiwatkul, W., & Xu, Z. (2011). Red and white wines inhibit cholesterol oxidation induced by free radicals. Journal of Agricultural and Food Chemistry, 59, 66453–66458. Turnbull, J. J., Nakajima, J. I., Welford, R. W., Yamazaki, M., Saito, K., & Schofield, C. J. (2004). Mechanistic studies on three 2-oxoglutarate-dependent oxygenases of flavonoid biosynthesis anthocyanidin synthase, flavonol synthase, and flavanone 3b-hydroxylase. Journal of Biological Chemistry, 279, 1206–1216. Usuki, S., Ito, Y., Morikawa, K., Kise, M., Ariga, T., Rivner, M., et al. (2007). Effect of pre-germinated brown rice intake on diabetic neuropathy in streptozotocininduced diabetic rats. Nutrition and Metabolism, 4, 25. Waldron, K. W., Parr, A. J., Ng, A., & Ralph, J. (1996). Cell wall esterified phenolic dimers: Identification and quantification by reverse phase high performance liquid chromatography and diode array detection. Phytochemical Analysis, 7, 305–312. Wang, C., & Shu, Q. (2007). Fine mapping and candidate gene analysis of purple pericarp gene Pb in rice (Oryza sativa L.). Chinese Science Bulletin, 52, 3097–3104. Wang, W., Guo, J., Zhang, J., Peng, J., Liu, T., & Xin, Z. (2015). Isolation, identification and antioxidant activity of bound phenolic compounds present in rice bran. Food Chemistry, 171, 40–49. Yang, Y., Andrews, M. C., Hu, Y., Wang, D., Qin, Y., Zhu, Y., et al. (2011). Anthocyanin extract from black rice significantly ameliorates platelet hyperactivity and hypertriglyceridemia in dyslipidemic rats induced by high fat diets. Journal of Agricultural and Food Chemistry, 59, 6759–6764. Yoshimura, Y., Zaima, N., Moriyama, T., & Kawamura, Y. (2012). Different localization patterns of anthocyanin species in the pericarp of black rice revealed by imaging mass spectrometry. PLoS One, 7, e31285. Zhang, H., Shao, Y. F., Bao, J. S., & Beta, T. (2015). Phenolic compounds and antioxidant properties of breeding lines between the white and black rice. Food Chemistry, 172, 630–639. Zhang, M. W., Zhang, R. F., Zhang, F. X., & Liu, R. H. (2010). Phenolic profiles and antioxidant activity of black rice bran of different commercially available varieties. Journal of Agricultural and Food Chemistry, 58, 7580–7587.