Aspectos Teóricos y Teorías relacionadas al tema

Clifford Hall III and Bin Zhao

CONTENTS

Introduction ... 22 Phytochemicals-Structural Characteristics ... 22 Monophenols and Phenolic Acids ... 22 Tocopherols and Tocotrienols ... 22 Phenolic Acids ... 23 Alkylresorcinols and Alkenylresorcinols ... 25 Flavonoids ... 26 Antioxidant Activity... 27 Health Benefits ... 28 Other Phytochemicals ... 29 Carotenoids ... 29 Phytosterols ... 30 Summary ... 30 Phytochemicals from Cereals and Pseudocereals ... 30 Defining Cereals and Pseudocereals ... 30 Cereals ...31 Barley ...31 Phenolics ... 32 Corn ... 35 Oats ...41 Rice ... 45 Rye ... 49 Wheat ... 55 Pseudocereals ... 59 Amaranth and Quinoa ...61 Phytochemicals from Pulses: Edible beans and Legumes ... 62 Dry Peas ... 63 Tocopherol and Carotenoids ... 63 Phenolic Compounds ... 63 Other Components ... 64 Antioxidant Activity... 64 Dry Bean ... 65 Tocopherol ... 65 Phenolic Compounds ... 65 Other Components ... 66 Antioxidant Activity... 66 Future Direction ... 67 References ... 67

Introduction

Phytochemicals are simply bioactive plant substances that provide a health benefit. Many of these compounds at one time were considered antinutrients. However, an extensive study of the phytochemicals (e.g., phenolics, carotenoids, tocopherols) has resulted in the discovery of many health benefits.

Furthermore, the usefulness of these components as food additives has been demonstrated.

In this chapter, the phytochemicals from cereals (and pseudocereals) and pulses (e.g., legumes and edible beans) will be presented. Due to the diverse functionality and chemical and structural makeup of the phytochemicals, only a small number of phytochemicals will be highlighted in this chapter. The phytochemicals of interest include simple phenols, polyphenolics, phenolic acid, carotenoids, and sterols.

Specific focus on the composition of phytochemicals from the various sources, effects of processing on the phytochemicals, and antioxidant activity of the phytochemicals will be highlighted. In addition, infor-mation will be presented regarding structural features of the general classes of phytochemicals. Methods for the isolation and characterization of the phytochemicals will not be presented in detail in this chapter.

The author suggests that the review of the referenced literature will be of value in this regard.

Important references prior to 2000 will be presented; however, the chapter material will cover research primarily from 2000 to 2007. Hall (2001, 2003) reported reviews on phytochemicals prior to 2000, and recent reviews by Awika and Rooney (2004) and Dykes and Rooney (2006) highlighted phytochemicals in several cereals, thus the reader is directed to these reviews. The authors of this chapter recognize the efforts of many researchers in the phytochemical area; however, not all of the research could be reported in this review.

Phytochemicals-Structural Characteristics Monophenols and Phenolic acids

Tocopherols and Tocotrienols

Tocopherols and tocotrienols (tocols; Figure 3.1) are a group of monophenols that have vitamin E and anti-oxidant activities. The antianti-oxidant activity of the tocols has been widely documented and will not be exten-sively described in this chapter. However, the phenolic hydrogen at the C₆ position can participate in chain breaking mechanisms, including radical scavenging (Figure 3.2). Tocopherols and tocotrienols have been well characterized as antioxidants (Yoshida, Niki, and Noguchi 2003). The research on the health benefits of tocopherols and tocotrienols is conflicting. However, some studies have supported the health benefits.

The role of tocols in disease prevention has been attributed to the antioxidant activity where the tocot-rienols appear to have the most benefit (Qureshi et al. 1997, 2000; McIntyre et al. 2000; Packer, Weber,

O

Figure 3.1 The monophenols tocopherol and tocotrienols.

24 Fruit and Cereal Bioactives: Sources, Chemistry, and Applications

para substitutions in the phenolic acids give mixed antioxidant results. The quinic acid substitution (i.e., chlorogenic acid) at the para position was equally effective as caffeic acid in controlling oxida-tion. Structurally, the only difference between the molecules was the para substitute; thus, the authors concluded that the acid proton of caffeic acid had little effect on the antioxidant activity of the cinnamic acid derivatives (Pratt and Birac 1979). In contrast, vinyl substituted phenolic acids (i.e., cinnamic acid derivatives) were more effective as antioxidants then the benzoic acid derivatives (Pratt and Hudson 1990; Cuvelier, Richard, and Berset 1992). Cuvelier, Richard, and Berset (1992) suggested that the vinyl

OH

Figure 3.4 Intramolecular hydrogen bonding of ortho substituted phenols. (Adapted from Baum, B., and Perun, A., Soc. Plastics Eng. Trans., 2, 250–7, 1962.)

COOH

Figure 3.3 Common phenolic acids in cereals, pseudocereals, and legumes, including examples of diferulic com-pounds associated with cell walls. (Adapted from Bunzel, M., Ralph, J., Marita, J., Hatfield, R., and Steinhart, H., J. Agric.

Food Chem., 48, 3166–9, 2000; Bunzel, M., Ralph, J., Marita, J., Hatfield, R., and Steinhar, H., J. Sci. Food Agric., 81, 653–60, 2001.)

and Rimbach 2001; Wu et al. 2005; Nakagawa et al. 2007). Halliwell, Rafter, and Jenner (2005) reported that the benefits might be related to the affects of these components in the gastrointestinal tract (GI) and the prevention of radical species formation in the GI tract. However, these authors did state that the mechanisms of action were still not clear. The anticarcinogenic activity of tocotrienols has been reported (Mizushina et al. 2006). For additional information on the health benefits of tocotrienols from rice see Hall (2003).

Phenolic Acids

Similar to tocols, the phenolic hydrogen(s) of phenolic acids (Figure 3.3) contribute antioxidant activity.

Phenolic acids tend to be located on the out layers (aleurone, pericarp) of cereals (Sosulski, Krygier, and Hogge 1982; Hutzler et al. 1998; Naczk and Shahidi 2006) in contrast to the higher tocol levels in the germ (Barnes 1983). Thus, the benefits of phenolic acid would be realized if the outer portions of the grain were not removed prior to the consumption.

Phenolic acids can act as antioxidants through a number of different mechanisms. The chain breaking mechanisms, which include hydrogen donation and radical acceptor (i.e., radical scavenging activity;

Scott 1985), are the most likely means by which phenolic acids act as antioxidants (Figure 3.2). Variations in the antioxidant activity of individual phenolic acids have been documented (Pratt and Birac 1979;

Pratt and Hudson 1990; Cuvelier, Richard, and Berset 1992). These authors observed key structure-activity relationships that accounted for the differences in antioxidant activities. The dihydroxy forms of the phenolic acids have better antioxidant activity due the addition of a second hydroxyl group in the ortho position. This statement can be supported by the observation of Pratt and Birac (1979) that caffeic acid had better antioxidant than the monohydroxy phenolic acids (i.e., ferulic acid and ρ-coumaric acid). The improved antioxidant activity of caffeic was likely due to the intramolecular hydrogen bond-ing (Figure 3.4) that can occur in ortho substituted phenols (Baum and Perun 1962). A third hydroxyl group further enhances the antioxidant activity as trihydroxybenzoic acid (i.e., gallic acid) and is a bet-ter antioxidant than 3,4-dihydroxy-benzoic acid (i.e., protocatechuic acid; Pratt and Birac 1979). The

LOOtrapping O

CH₃

CH₃

CH₃ HO

R₂ R₃ R₁

R₂ R₃ R₁

R₂ R₃ R₁

O O

Hydrogen abstraction

O O

OOL

Figure 3.2 Hydrogen donation and radical scavenging activity of monophenols.

Phytochemicals in Cereals, Pseudocereals, and Pulses 25

group could enhance the resonance stability of the phenoxyl radical whereby improving the antioxidant activity. Thus, by understanding the above relationships one can predict the antioxidant potential of a plant material containing phenolic acids.

alkylresorcinols and alkenylresorcinols

Alkylresorcinols and alkenylresorcinols have a 1,3-dihydroxybenzene base structure and an aliphatic substitution at carbon five of the ring (Figure 3.5). The aliphatic group typically has between 17 and 25 carbons (Kozubek and Tyman 1995, 1999; Ross et al. 2001; Ross, Kamal-Eldin, and Aman 2004).

When the aliphatic group is unsaturated, the compounds are generically referred to as alkenylresorci-nols. However, the alkylresorcinols (i.e., saturated aliphatic group) are the most common. Furthermore, these compounds are concentrated in the bran fractions of many cereal grains and may contribute to the health benefits attributed to whole grain consumption.

The interest in this group of compounds stems from the reported anticarcinogenic, antimicrobial, and antioxidant properties (Singh et al. 1995; Gasiorowski et al. 1996; Kozubek and Tyman 1999; Slavin et al.

2001). For a summary of the reported benefits, see the review by Ross, Kamal-Eldin, and Aman (2004).

The bioavailability of the alkylresorcinols shows that about 60% are absorbed by the human ileostomy (Ross et al. 2003a), but only small amounts are present in the plasma (Linko et al. 2002). However, higher alkylresorcinols concentrations were present in erythrocyte membranes, which appear to be a site for alkylresorcinol storage, than plasma membranes (Linko and Adlercreutz 2005). These authors also noted that the longer chained alkylresorcinols were incorporated into the erythrocyte membrane at higher concentrations than short-chained alkylresorcinols. Much of the intact alkylresorcinols and metabolites 3-(3,5-dihydroxyphenyl)-1-propanoic acid and 1,3-dihydroxybenzoic acid were found in urine (Ross, Aman, and Kamal-Eldin 2004). The reader is encouraged to read the review written by Ross et al. (2004c) for more information on alkylresorcinol structural chemistry, including metabolites.

The antioxidant function of alkylresorcinols and alkenylresorcinols has not been fully characterized.

Compounds with the substitutions at the meta position to the hydroxyl on the benzene ring typically have poor antioxidant activity (Miller and Quackenbush 1957). Yet, several researchers have reported antioxi-dant effects of the alkylresorcinols in model test systems (Nienartowicz and Kozubek 1995; Winata and Lorenz 1996; Hladyszowski, Zubik, and Kozubek 1998; Litwinienko, Kasprzycka-Guttman, and Jamanek 1999). Kamal-Eldin et al. (2001) evaluated hydrogen donating and peroxy radical scavenging activity of these compounds and found very poor antioxidant activities. In fact, based on the adherence to general antioxidant definition that the compounds must be effective at low concentrations, they concluded that

HO OH

Figure 3.5 Alkyl- and alkenylresorcinols found in cereals. (Adapted from Mattila, P., Pihlava, J.-M., and Hellström, J., J. Agric. Food Chem., 53, 8290–95, 2005; Ross, A., Shepherd, M., Schüpphaus, M., Sinclair, V., Alfaro, B., Kamal-Eldin, A., and Åman, P., J. Agric. Food Chem., 51, 4111–18, 2003.)

the alkylresorcinols were not effective antioxidants (Kamal-Eldin et al. 2001) in the DPPH and sunflower triacylglycerol systems. The conversion of the alkylresorcinols to trihydroxy derivatives was proposed as a reason for the antioxidant activity (Kozubek and Tyman 1999) and not the original alkylresorcinols.

Flavonoids

Flavonoids are polyphenolic compounds characterized by a C6–C3–C6 configuration (Figures 3.6 through 3.8). Flavones, flavonols, flavanones, anthocyanadins, and anthocyanins make up the largest and most diverse groups among the flavonoids. Although fruits and vegetables are the primary dietary sources

O

a Dihydroquercetin has an additional H at the C-3 position due to the loss of the double bond at the C-2:C-3 position.

b Rutin is a glycoside in which the C-3 position contains a o-rutinose.

c Hesperidin contains a o-rutinose at the C-7position. C-3 position contains a rhamnoglucose

unit.

b Fustin lacks a C-5 OH.

Figure 3.6 General flavonoids isolated from cereals, pseudocereals, and legumes.

Phytochemicals in Cereals, Pseudocereals, and Pulses 27

of flavonoids, cereals, legumes, and beans can contribute to the daily intake. Flavonoids are a group of compounds that have been well documented as hydrogen donors, radical scavengers, and metal chelators (Dziedzic and Hudson 1983; Torel, Cillard, and Cillard 1986; Husain, Cillard, and Cillard 1987; Bors et al.

1990; Das and Pereira 1990; Salah et al. 1995; Foti et al. 1999; Rice-Evans, Miller, and Paganga 1996;

Cao, Sofic, and Prior 1997). Flavonoids as food antioxidants and health promoters have been reviewed extensively (Hall and Cuppett 1997; Middleton 1998; Pietta 2000; Nijveldt et al. 2001; Rice-Evans 2001;

Yanishlieva and Heinonen 2001; Hall 2003; Valko et al. 2006).

Antioxidant Activity

As with phenolic acids, the antioxidant activity of flavonoids is dependent on the number and location of the hydroxyl groups. Hydroxyl groups on ring B play a significant role in the hydrogen donating activity. Hydroxyl groups at the 3′, 4′, and 5′ positions on the ring B have the greatest activity followed by flavonoids with ortho hydroxyl groups on ring B (Dziedzic and Hudson 1983; Hudson and Lewis 1983; Rice-Evans, Miller, and Paganga 1996). The hydrogen donating activity greatly diminishes in flavonoids with only one B ring hydroxyl group. Similar structural features were important for radical scavenging activity (Husain, Cillard, and Cillard 1987; Bors et al. 1990; Cao, Sofic, and Prior 1997;

Foti et al. 1999). Like other flavonoids, ortho hydroxyl configuration enhances radical scavenging

O+

Apigeninidin 5-glucoside Cyanidin 3-glycoside Delphinidin 3-glycoside Delphinidin 3-glucose

glucose Cyanidin 3-galactose

glucose Delphinidin 3-rutinoside

Petunidin 3-glucoside Malvidin 3-glucoside Pelargonidin 3-glucoside

galactose rutinose

Figure 3.7 Anthocyanins isolated from pigmented corn, rice, wheat, and legumes.

O

Figure 3.8 Common isoflavones in edible legumes.

activity of anthocyanidins (Yoshiki, Okubo, and Igarashi 1995; Rice-Evans, Miller, and Paganga 1996; Wang, Cao, and Prior 1997). However, the presence of the hydroxyl group at the 5′ position did not improve anthocyanidin antioxidant activity (Rice-Evans, Miller, and Paganga 1996; Wang, Cao, and Prior 1997).

The hydroxyl substitutions at the 5,8 and 7,8 positions on ring A improved flavonoids antioxidant activ-ity. However, ring A dihydroxy substitutions at 5,7 positions did not influence the antioxidant activities of flavonoids. In contrast, the 7 position on the A ring did not affect the antioxidant activity of isoflavones.

The addition of a hydroxyl group at the 5 position on ring A did improve the antioxidant activity (Hu et al. 1995; Wei et al. 1995).

The presence of a hydroxyl group at the 3 position on ring C enhances the antioxidant activity of the flavonoids. The flavonols are generally better antioxidants than flavanones due to the presence of the hydroxyl group at the 3 position. In addition, the presence of sugar moieties on the three location of ring C diminishes the antioxidant activity of the flavanones (Das and Pereira 1990; Nieto et al. 1993). In contrast, the radical scavenging activity of the anthocyanins (glycoside form) was better than the antho-cyanidins (Satué-Gracia, Heinonen, and Frankel 1997; Wang, Cao, and Prior 1997). Thus, the greater antioxidant activity of the flavones over the anthocyanidins was attributed to the carbonyl at position 4 of ring C in conjunction with the double bond at carbons 2 and 3 of ring C (Cao, Sofic, and Prior 1997;

Wang, Cao, and Prior 1997).

The metal chelating activity (Figure 3.9) of flavonoids can occur at two regions of the molecule. The 3′,4′-dihydroxy configuration is an important structural feature that accounts for the metal chelating properties of anthocyanins and anthocyanidins, whereas the ring C quinone at position 4 of flavones and flavonols was essential (Crawford, Sinnhuber, and Aft 1961; Pratt and Watts 1964; Letan 1966; Hudson and Lewis 1983). A loss in metal chelating activity of the flavones and flavonols was observed after the double bond at positions 2 and 3 on ring C was hydrogenation (Crawford, Sinnhuber, and Aft 1961; Letan 1966). The flavonoids have a very diverse function as a food antioxidant and these effects might contrib-ute to the health benefits of the flavonoids.

Health Benefits

The anti-inflammatory, anticarcinogenic, and antitumor activities of flavonoids have been reported (Hollman and Katan 1998; Middleton 1998; Waladkhani and Clemens 1998; Agarwal, Sharma, and Agarwal 2000). Hirano, Gotoh, and Oka (1994) reported that flavonoids had cytostatic activity against human breast carcinoma cells but did not find a structure-activity relationship. Sánchez et al. (2001) found that flavonoids lacking the C-8 methoxy substitutions had little cytotoxicity against Rhesus monkey kid-ney cells and rat glial tumor cells, whereas the C2’ and C5’ were an important structural feature. The anti-17beta-hydroxysteroid dehydrogenase activity was dependent on the C-7 hydroxyl group whereas flavonoids with C-7 methoxy or C-8 hydroxyl groups had only antiaromatase activity (Bail et al. 1998).

O

Figure 3.9 Metal chelate complexes of flavonoids. (Adapted from Hudson, B. and Lewis, J., Food Chem., 10, 47–55, 1983.)

Phytochemicals in Cereals, Pseudocereals, and Pulses 29

Bomser et al. (1999) and Zhao et al. (1999) observed antitumor activity of procyanidin B5-3′-gallate (Zhao et al. 1999). Quercetin, myricetin, and epicatechin inhibited the growth and altered the enzyme activities of MCF7 human breast cancer cells (Rodgers and Grant 1998).

Flavonoids also inhibit oxidation of LDL (Meyer et al. 1997; Meyer, Heinonen, and Frankel 1998;

Meyer, Jepsen, and Sórgensen 1998; Brown and Rice-Evans 1998; Heinonen, Meyer, and Frankel 1998;

Hwang, Hodis, and Sevanian 2001; Porter et al. 2001) and inhibit cholesteryl ester synthesis (Borradaile, Carroll, and Kurowska 1999). Naringenin and hesperetin reduce acyl CoA:cholesterol acyltransferase activity, inhibit the activity and expression of microsomal triglyceride transfer protein, and increase LDL receptor mRNA that promote the reduction in plasma cholesterol (Wilcox et al. 2001). The inhibitions of thromboxane synthase and prostaglandin production are the reasons for the anti-inflammatory activity of flavonoids (Ishiwa et al. 2000; Skaltsa et al. 2000).

Other Phytochemicals

Carotenoids and phytosterols are the final phytochemicals to be covered in this chapter. However, com-pounds specific to cereals or pulses will be presented under that section related to specific materials.

Avenanthramides in oats, oryzanols in rice, and policosanols in sorghum are a few of examples of health promoting phytochemicals.

Carotenoids

Carotenoids (Figure 3.10) are a group of compounds characterized by a conjugated polyene system.

The singlet oxygen quenching characteristics of carotenoids has been well documented (Foote, Chang, and Denny 1970; Burton and Ingold 1984; Terao 1989). The presence of nine or more double bonds and oxo groups at the 4(4′) position in the β-ionone ring in the carotenoid structure greatly enhanced the singlet oxygen quenching activity (Terao 1989). The carbonyl present on the ring enhanced the stability

HO

HO

OH β-carotene

β−cryptoxanthin

Lutein α-carotene

OH

OH

Zeaxanthin Figure 3.10 Carotenoids found in corn and wheat.

of trapped radicals; therefore, reducing the tendency of carotenoids to promote radical reactions. The polyene system can also trap radicals, thus providing additional protective activity. These activities are believed to be the cause of the health benefits of carotenoids. However, controversy also exists around the negative impact of high carotenoid levels in some populations. Dutta, Chaudhuri, and Chakraborty (2005) and Krinsky and Johnson (2005) have recently reviewed carotenoids.

In the context of this chapter, the carotenoids are responsible for the yellow color in corn and durum wheat. In cereals, for example, carotenoids exist as carotenes (α- and β-carotene) and xanthophylls (β-cryptoxanthin, lutein, and zeaxanthin), where xanthophylls are typically in the highest concentrations.

Lutein and zeaxanthin have attracted much attention due to the possible role in preventing cataracts (Knekt et al. 1992) and age-related macular degeneration, a condition that results in irreversible vision loss (Gale et al. 2003; Mozaffarieh, Sacu, and Wedrich 2003; Moeller et al. 2006; Trieschmann et al.

2007). Thus, grain consumption can contribute to the total dietary intake of carotenoids.

Phytosterols

Phytosterols and phytosanols (saturated form of the sterol; Figure 3.11) are widely present in grains (Piironen, Toivo, and Lampi 2002). These compounds exist as free sterols, fatty acid, or phenolic esters, and steryl glycosides (Toivo et al. 2001; Moreau, Whitaker, and Hicks 2002). The phytosterols have limited antioxidant activity and those esterified to phenolic acids can act as chain breaking antioxidants similar to phenolic compounds. However, the ferulate esters were found to have less activity than the fer-ulic acid (Xu and Godber 2001). In contrast, the phytosterols were effective in controlling the oxidation of frying oils Kamal-Eldin et al. (1988) and prevention of oil polymerization (Sims, Fioriti, and Kanuk 1972; Boskou and Morton 1976; Gordon and Magos 1983; White and Armstrong 1986).

The role of phystosterols in health is probably more significant than the antioxidant effects. The phy-tosterols have been shown to effectively reduce blood cholesterol (Fernandez et al. 2002; Gylling and Miettinen 2005), prostatic hyperplasi (Berges et al. 1995; Berges, Kassen, and Senge 2000), and colon cancer (Awad and Fink 2000). In addition, an enhanced immune function has been reported (Bouic and Lamprecht 1999). For a complete review of the benefits of phytosterols, see the recent review by Kritchevsky and Chen (2005).

Summary

A varied diet of foods would be required to achieve the health benefits of the phytochemicals previ-ously described. However, in some cases the components can be concentrated via physical methods or by solvent extractions. Thus, one must remember that in the following discussions for low levels of a component in a grain, or pulse is not necessarily a negative if the phytochemical is consumed as part of a varied diet or in a concentrated form.

Phytochemicals from Cereals and Pseudocereals Defining Cereals and Pseudocereals

Cereals and pseudocereals are plant materials that have similar end uses as flours for bakery prod-ucts. However, these plants are different botanically as cereals are grasses whereas pseudocereals are broadleaf plants. All of these plant materials have a cultivar of phytochemical constituents and are of interest to researchers in the health and medical fields. The cereals that have garnered attention include barley (Hordeum vulgare), corn (Zea mays), millet (Panicum milliaceum), oats (Avena sativa), rice (Oryza sativa), rye (Secale cerale), and wheat (Triticum spp). Pseudocereals of interest include amaranth (Amaranthus caudatus, A. cruentus), buckwheat (Fagopyrum esculentum), and quinoa (Chenopodium quinoa). Regardless of the plant materials, the hull or bran is usually the main source of the phytochemi-cals; however, the germ is also a valuable source of the lipid soluble phytochemicals. Thus, the benefit of whole grain consumption is related to the consumption of the aforementioned grain fraction.

Phytochemicals in Cereals, Pseudocereals, and Pulses 31

Cereals Barley

In human foods, barley is most often used in the brewing industry. However, barley consumption as a food source has recently increased due to the reported health benefits. Barley has a number of different phy-tochemicals that include tocols (Peterson 1994; Goupy et al. 1999), Δ⁵-avenasterol (Dutta and Appelqvist

HO

Figure 3.11 Phytosterolas, phytostanols, and sterol ferulates found in cereals and pseudocereals.

1996), flavonoids (Tamagawa et al. 1999), phenolic acids (Van Sumere et al. 1972; Slominski 1980; Mattila, Pihlava, and Hellström 2005), and alkylresorcinols (Mattila, Pihlava, and Hellström 2005).

Tocols

The content of tocopherols varies widely among cultivars. Goupy et al. (1999) reported an average tocopherols content of 25.1 mg/kg among nine barley cultivars. The tocopherols ranged from 9.7 to

The content of tocopherols varies widely among cultivars. Goupy et al. (1999) reported an average tocopherols content of 25.1 mg/kg among nine barley cultivars. The tocopherols ranged from 9.7 to