Cereal grain by-products as natural antioxidants

TABLE OF CONTENTS

1-INTRODUCTION

2-AIM OF INVESTIGATION

3-REVIEW OF LITERATURE
3.1. Antioxidants in cereal grains:
3.1.1. The phenolic content:
3.1.1.1. Phenolic compounds in cereal grains:
3.1.1.2. Wheat grains as source of phenols:
3.1.1.3. Sorghum grains as a source of phenols:
3.1.1.4. Corn grains as a source of phenols:
3.1.2. Phytate content in cereal grains:
3.1.2.1. Wheat grains as source of phytate:
3.1.2.2. Sorghum grains as source of phytate:
3.1.2.3. Corn grains as source of phytate:
3.1.3. Scavenging capacity of cereal grains:
3.2. Effect of domestic processing on antioxidant activity of cereal grains:
3.2.1. Effect of soaking on antioxidant content:
3.2.2. Effect of soaking on antioxidant activity:
3.2.3. Effect of germination on antioxidant content:
3.2.4. Effect of germination on antioxidant activity:
3.3. Using cereal grains milling fractions as natural source of antioxidant to improve meat products quality during storage:
3.3.1. The effect of phenolic compounds content on oxidative stability:
3.3.2. The effect of phytic acid on oxidative stability:
3.3.3. Beef meat products:
3.3.4. Chicken meat products:

4-MATERIALS AND METHODS
4.1. Materials:
4.1.1. Cereal grains:
4.1.2. Meat and burger ingredients:
4.1.3. Chemicals:
4.2. Processing treatments:
4.2.1. Soaking:
4.2.2. Germination:
4.2.3. Milling:
4.2.4. Preparation of chicken and beef burger:
4.3. Analytical methods:
4.3.1. Chemical analyses:
4.3.1.1. Gross chemical composition:
4.3.1.2. Extraction of total antioxidants:
4.3.1.3. Determination of total phenolics content:
4.3.1.4. Determination of antioxidant activity using the DPPH radical scavenging method:
4.3.1.5. Determination of phytic acid:
4.3.2. Evaluation of burger:
4.3.2.1. pH value:
4.3.2.2. Peroxide value (PV):
4.3.2.3. Thiobarbituric acid values (TBA):
4.4. Sensory evaluation:
4.5. Statistical analysis:

5-RESULTS AND DISCUSSION
5.1. Gross chemical composition
5.1.1. Gross chemical composition of sorghum milling fractions:
5.1.2. Gross chemical composition of wheat milling fractions:
5.1.3. Gross chemical composition of corn milling fractions:
5.2. Effect of soaking and germination processes
5.2.1. Effect of soaking and germination processes on gross chemical composition of sorghum milling fractions:
5.2.2. Effect of soaking and germination on gross chemical composition of wheat milling fractions:
5.2.2.1. Effect of soaking and germination on gross chemical composition of corn milling fractions:
5.3. Total phosphorus and phytic acid contents in grains milling fractions:
5.3.1. Total phosphorus and phytic acid content of sorghum milling fractions:
5.3.2. Total phosphorus and phytic acid contents of wheat milling fractions:
5.3.3. Total phosphorus and phytic acid contents of corn milling fractions:
5.4. Effect of soaking and germination on total phosphorus and phytic acid contents in studied grains:
5.5. Total phenolic compounds (TPC) of sorghum, wheat and corn grains:
5.6. Effect of soaking and germination on phenolic compounds content of cereal grains:
5.7. Total antioxidant activity of cereal grains:
5.8. Effect of soaking and germination on total antioxidant activity of cereal grains:
5.9. Effect of cereal grains by-products addition on meat products properties during storage at 5±2˚C:
5.9.1. Effect of cereal grains by-products addition on thiobarbituric acid value (TBA) of beef burger:
5.9.2. Effect of cereal grains by-products addition on thiobarbituric acid value (TBA) of chicken burger:
5.9.3. Effect of cereal grains by-products addition on peroxide value (PV) of beef burger:
5.9.4. Effect of cereal grains by-products addition on pH value of beef burger:
5.9.5. Effect of cereal grains by-products addition on pH value of chicken burger:
5.10 Sensory evaluation:
5.10.1. Sensory evaluation of beef burger formulas:
5.10.2. Sensory evaluation of chicken burger formulas:

6-SUMMARY

7-REFERENCES

Acknowledgement

First of all, thanks to GOD by the grace and care of whom indeed the complete of this work was possible.

I would like to express my deepest thanks and sincerest gratitude to Prof. Dr. Bolbol. R. Ramadan

Professor of Food Science and Technology Department, Fac. of Agric., Assiut Univ ., for his supervision, trustful help, unfailing advice and given me the power to complete this work. reviewing the manuscript and continuous help during the course of experimental work.

A special appreciation and my deepest thanks are due to Prof. Dr. Mohamed. A. Sorour

Professor and head of Food Science Department, Fac. of Agric., Sohag Univ., for his kindly suggesting the problem, direct supervision, planning the research program, reviewing the manuscript and continuous help during the course of experimental work.

Acknowledgement is also extended to Prof. Vincenzo Fogliano Napoli University, Dr. Amal amin, National Research Center and Dr. hosam elgepaly , , Agricultural Research Center, for their help to finish this work.

Also I would thanks all staff members of Food Science Department, Faculty of Agriculture, Assiut University, staff members of Food Science Department, , Faculty of Agriculture, Sohag University and staff members of Upper Egypt Flour Mills Company Sohag and any person who help me to complete this work.

Mohamed kelany

LIST OF TABLES

1 Basel constituents of burger
2 Additional ingredients used in burger formulation
3 Gross chemical composition of wheat, corn and sorghum grains fractions
4 Effect of soaking and germination processes on gross chemical composition of wheat, corn and sorghum grains
5 Total phosphorus and phytic acid contents in sorghum, wheat and corn milling fractions
6 Effect of soaking and germination processes on total phosphorus and phytic acid contents in milling fractions of sorghum, wheat and corn grains
7 Total phenolic compounds of grains milling fractions as mg GAE/100g
8 Effect of soaking and germination on phenolic content in cereal grains as mg (GAE)/100g
9 %DPPH scavenging activity of cereal grains fractions
10 Effect of soaking and germination on the %DPPH scavenging activity in wheat, corn and sorghum
11 Changes in thiobarbiturc acid values (mg malonaldehyde/kg) of different beef burger formulas during storage for 15 days at 5±2˚C
12 Changes in thiobarbiturc acid values (mg malonaldehyde/kg) of different chicken burger formulas during storage for 15 days at 5±2˚C
13 Changes in peroxide values (m. equv./kg fat) of different beef burger formulas during storage for 15 days at 5±2˚C
14 Changes in peroxide values (m. equv./kg fat) of different chicken burger formulas during storage for 15 days at 5±2˚C
15 Changes in pH values of different beef burger formulas during storage for 15 days at 5±2˚C
16 Changes in pH values of different chicken burger formulas during storage for 15 days at 5±2
17 Sensory evaluation of different beef burger formulas
18 Sensory evaluation of different chicken burger formulas

LIST OF FIGURES

1 Chemical structures of phenolic acids
2 Structure of phytic acid
3 Structure of phytic acid with the different possibilities to interact with both metal cations (minerals) as with protein residues
4 Total phenolic compounds of grains milling fractions as mg GAE/100g
5 Effect of soaking and germination time in phenolic content in cereal grains as mg (GAE)/100g
6 The %DPPH scavenging activity of cereal grains fraction
7 Effect of soaking and germination time on % DPPH scavenging activity in cereal grains
8 Changes in thiobarbiturc acid values (mg malonaldehyde/kg) of different beef burger formulas during storage for 15 days at 5±2˚C
9 Changes in thiobarbiturc acid values (mg malonaldehyde/kg) of different chicken burger formulas during storage for 15 days at 5±2˚C
10 Changes in peroxide values (m. equv./kg fat) of different beef burger formulas during storage for 15 days at 5±2˚C
11 Changes in peroxide values (m. equv./kg fat) of different chicken burger formulas during storage for 15 days at 5±2˚C
12 Changes in pH values of different beef burger formulas during storage for 15 days at 5±2˚C
13 Changes in pH values of different chicken burger formulas during storage for 15 days at 5±2˚C

1-INTRODUCTION

Antioxidants are an inhibitor of the process of oxidation, even at relatively small concentration and thus have diverse physiological role in the body. Antioxidant constituents of the plant material act as radical scavengers, and helps in converting the radicals to less reactive species. A variety of free radical scavenging antioxidants is found in dietary sources like fruits, vegetables and tea, etc. (Murthy, 2001 and Mandal et al., 2009).

The antioxidant can also be defined as "A compound capable of inhibiting oxygen mediated oxidation of diverse substances from simple molecule to polymer and complex bio-systems"(Chi-Tang, 1994).

Free radicals represent a class of highly reactive intermediate chemical entities whose reactivity is derived from the presence of unpaired electron in their structure, which are capable of independent existence for very brief interval of time (Cui et al., 2004).

Free radicals and other reactive species are derived either from normal essential metabolic processes or from external sources, such as exposure to x-rays, ozone, cigarette smoking, air pollutants, industrial chemicals etc. (Kumar et al., 2011).

Consumption of foods containing rich antioxidant activity substances, such as grains, vegetables, and fruits, may prevent many diseases and promote good health (Willet, 1994 and Temple, 2000). Although the cereal grains and legumes are not very rich sources of antioxidant compounds in practice, only phenolic compounds can be found there, they are mentioned because of these sources huge share in diet. The chemical structure as well as concentration of these compounds influences their antioxidant activity.

Among polyphenols enclosed in cereal grains, phenolic acids play important role, and especially ferulic acid is dominating in grains (wheat and rye, first of all). Phenolic acids are present in two forms: ester and glycoside ones. Besides these compounds, vanillic and p -coumaric acids play important role, even though they are present in smaller amounts. In the case of oats, the presences of other polyphenols called avertramidin were recorded, while rutin is the main polyphenol of buckwheat (Peterson et al., 2001 and Holasova et al., 2002). Grains in particular, are a major source of antioxidants in our daily diets. The main antioxidative components in grain are classified as phenolic compounds such as anthocyanins, tannins, and ferulic acid, and other substances (Tome´ et al., 2004). Whole grain products are recommended for healthy diets as being recognized sources of dietary fiber and antioxidant substances (Ragaee et al., 2006).

Health advantages of diets rich in antioxidant plant compounds include lowering the risk of cardiovascular disease, certain cancers and the natural degeneration of the body associated with the aging process. Recent scientific evidence is demonstrative of these advantages. The cause of the health problems listed above is the free radical molecule. Free radicals are unstable molecules formed when the body uses oxygen for energy. The instability of these molecules can damage tissues, alter DNA and change cell structure ( Oufnac, 2006).

In order to minimize the potentially damaging effects of oxidative reactions, biological systems including grains have developed multifunctional antioxidant defense systems. These defense systems control reactive oxygen species, oxidation catalysts and oxidation products in both lipid and aqueous environments by several different mechanisms, including metal chelation, free radical scavenging and peroxide inactivation ( Decker, 1998).

Whole grains, including wheat, contain several compounds that are capable of minimizing the damaging effects of oxidation reactions. These include phytate, proteins, polysaccharides, phenolics, lignans and tocopherols. Since grains contain significant concentrations of antioxidative compounds, it is possible that they could contribute to dietary antioxidant intake if these antioxidants are present in active forms in wheat-based foods ( Baublis et al., 2000a).

Numerous studies have evaluated the antioxidant activity of phenolics from fruits, vegetables, soybeans, herbs, teas, wines and medicinal plants. Much less attention has been focused on the antioxidant potential of phenolics from whole grains ( Kanner et al., 1994; Decker, 1998 and Pietta et al., 1998 ).

2-AIM OF INVESTIGATION

The aim of this work to investigate the changes in total phenolic compounds content, phytate content and free radical scavenging abilities during soaking and germination of three cereal grains; wheat (Sids 1), corn (H310 White) and sorghum (Giza 15). On the other hand, the present investigation was carried out to study the possibility of using the grain fractions to preparation the beef and chicken burger and improve it's quality and storagability.

Accordingly, This investigation was designed to study each of the following, topics:-

1. Determination of gross chemical composition and mineral content in milling fractions of studied grains.
2.Determination of phenolic compounds and phytate content in milling fractions of studied grains samples.
3.Extraction and determination of antioxidant activity using DPPH radical scavenging method.
4.Using milling fractions as antioxidant ingredients to improve the quality of beef and chicken burger during storage under refrigeration.
5.Estimation of thiobarbiturc acid, peroxide and pH values of beef and chicken burgers during storage at 5±2˚C up to 15 days.

3-REVIEW OF LITERATURE

Cereals can be defined as a grain or edible seed of the grass family (Bender and Bender , 1999). Cereals are grown for their highly nutritious edible seed, which are often referred to as grain. Some cereals have been staple foods both directly for human consumption and indirectly via livestock feed since the beginning of civilization (BNF, 1994).

The major cereals consumed worldwide are wheat, rice, maize, barley, oats, rye, millet and sorghum. Apart from being an important part of diet, these cereals are also rich in various health promoting components (Slavin, 2003). Cereals are staple foods providing major sources of carbohydrates, proteins, B vitamins and minerals for the world’s population. Cereals contain a range of substances which may have health promoting effects, these substances are often referred to as phytochemicals or plant bioactive substances (Goldberg, 2003). These substances are beneficial to human health but are not essential for the human body (Kris-Etherton, et al., 2002).

The majority of bioactive compounds of whole-grains are present in the bran/germ fraction of cereal-grains. These fractions of whole-grain may therefore help in reducing the risk of chronic diseases. Bioactive compounds in whole-grain cereals have not received as much attention as in fruits and vegetables. Epidemiological studies have shown that regular consumption of whole grains and it's products is associated with reduced risks of various types of chronic diseases such as cardiovascular diseases (Anderson, 2003 and Okarter and Liu, 2010).

3.1. Antioxidants in cereal grains:

Significant levels of antioxidants have been detected in grains and grain-based cereal products (Baublis et al., 2000b). Furthermore, the antioxidant potential and bioavailability of cereal antioxidants may depend on the species and varieties of grains, fractions of the grain (bran, flour, or whole grain), and processing conditions (Zielinski and Kozlowska, 2000).

Wheat, sorghum and corn are foods rich in antioxidants, which make them excellent sources for increased health benefits ( Oufnac, 2006).

3.1.1. The phenolic content:

Plant foods have phenolic compounds, which affect their: appearance, taste, odor and oxidative stability. In cereal grains, these compounds are located mainly in the pericarp (Naczk and Shahidi, 2004).

Polyphenols are compounds that have more than one phenolic hydroxyl group attached to one or more benzene rings. Phenolic compounds or polyphenols are ubiquitous in plants with more than 8000 structures reported (Bravo, 1998). The antioxidative potential of phenolic compounds can be attributed to their strong capability to transfer electron to (Reactive Oxygen Species) ROS/free radicals, chelating metal ions and to activate antioxidant enzymes . (Cos et al., 1998).

Folin-Ciocalteu (F-C) method has for many years been used as a means to determine total phenolics in natural products (Prior et al., 2005). The reaction that takes place is an oxidation/reduction one and because this reaction is the basic mechanism, F-C can also be considered an antioxidant capacity method. This assay has many variations. Different reagent concentrations and timing of additions and incubation are frequently varied (Prior et al., 2005). Moreover , many studies show the recommended reference standard (gallic acid) being replaced with tannic, caffeic, vanillic acids and catechin equivalents, among others. Phenolic compounds can be found in flavonoides, phenolic acids, hydroxycinnamic acid derivatives and lignans.

Sun et al. (2006) reported that methanol was the best solvent in extracting phenolic compounds from oat bran. That was in line with Oufnac (2006), they found that total phenolic compound content obtained by different conventional solvent extractions from low to high was hexane, acetone, and methanol extraction and 59.1, 88.0, and 241.3 μg catechin equivalent /g of fresh wheat bran, respectively. In addition, several studies showed that 80% methanol is an effective solvent in extracting phenolics and other polar substances in cereals (Przybylski et al., 1998 and Zielinski and Kozlowska, 2000).

3.1.1.1. Phenolic compounds in cereal grains:

Studies have shown that dietary phenolics have high antioxidant activity, which may contribute to their health benefits. In cereals, the predominant phenolic acid is ferulic acid, representing up to 90 % of total polyphenols. Other phenolic acids including vanilic, syringic, chlorogenic, p -coumaric, m -coumaric and OH-cinnamic acid have also been reported in cereals (Zhou and Yu , 2004; Mattila et al., 2005; Holtekjolen et al., 2006 and Hosseinian and Mazza, 2009). while phytoestrogens and flavonoid are presented in small quantities (Dordević et al., 2010).

Total amount of polyphenols in cereals is highly variable both in whole grain and in bran and also depends on the cereal variety and milling procedure ( Adom et al., 2003 and Adom et al., 2005).

The phenolic compounds of whole-grains including lignans, alkylresorcinols and phenolic acids have been shown to be metabolized and absorbed in humans and are among the major compounds inducing physiological changes underlying the protective effects (Andreasen et al., 2001 and Jacobs et al., 2002).

The major bioactive compounds in whole-grain cereals are phenolic compounds, phytosterols, tocols, dietary fibers (mainly beta-glucan), lignans, alkylresorcinols, phytic acid, γ-oryzanols, avenanthramides, cinamic acid, ferulic acid, inositols and betaine (Zieli et al., 2000).

Rice-Evans et al. (1995). Indicated that the antioxidant activity of phenolics is mainly due to their redox properties, which allow them to act as reducing agents, hydrogen donators, and singlet oxygen quenchers. In addition, they have a metal chelation potential.

phenols are presumed to be responsible for the beneficial effects derived from the consumption of whole grains, fruits, and vegetables. Phenolic compounds have strong in vitro and in vivo antioxidant activities associated with their ability to scavenge free radicals, break radical chain reactions, and chelate metals. Moreover, high phenol consumption had been correlated with a reduced risk of cardiovascular diseases and certain cancers Bonoli et al. (2004).

Phenolic compounds are secondary metabolites which synthesize in plants. They posses biological properities such as: antioxidant, antiapoptosis, anti-aging, anticarcinogen, anti-inflammation, anti-artherosclerosis, cardiovascular protection, improvement of the endothelialfunction, as well as inhibition of angiogenesis and cell proliferation activity. Most of these biological actions have been attributed to their intrinisc reducing capabilities (Han et al., 2007).

3.1.1.2. Wheat grains as source of phenols:

Wheat is a wild grass native to arid countries of western Asia. Almost 600 genera of wheat have been evolved and a number of classifications have been made. Owing to the presence of unique elastic protein complex, it is the only grain suitable for leavened bread and thus may be regarded as an important part of daily diet (Mazza, 2003). Owing to high nutritional values of wheat, it became a leading cereal crop long ago. The major producers of wheat are Europe, Pakistan, India, China, South and North America, and Australia. Almost 600 million tons of wheat are produced in the world per annum, out of which more than 19 million tons are produced in Pakistan (Soomro, 2003).

Similar to other cereal grains, wheat bran contains many different types of phenolic antioxidant compounds, such as ferulic, vanillic, caffeic, coumaric, and syringic acids (Li et al., 2005 and Kim et al., 2006) and relatively high levels of carotenoids, tocopherols and phytosterols (Nystrom et al ., 2005 and Zhou et al., 2005).

The ferulic acid content of wheat grain is near about to 0.8 - 2 g/kg dry weight basis, which may represent 90% of total polyphenols (Lempereur et al., 1997). Ferulic acid is the most abundant hydroxycinnamic acid found in cereal grains. It is esterified to the arabinoxylans of the grain cell wall. Wheat bran is the good source of ferulic acid which is esterified to the hemicelluloses of the cell walls (Dewanto et al., 2002).

Yu et al. (2004) evaluated the phenolic contents of flour extracts from three winter wheat varieties (Trego, Akron and Platte) grown at five testing locations. They found the phenolic contents of the flour samples Pearson Correlation tests had not detect any significant correlation between a single antioxidant property of each wheat variety and a selected environmental factor, including total solar radiation, daily average solar radiation, or the hours exceeding 32 °C.

According to Cheng et al. (2006) a previous phytochemical investigation established that phenolic acids (ferulic, syringic, vanillic and p-coumaric acids; Fig.1), tocopherols, and carotenoids are the natural antioxidants present in wheat grain and its fractions.

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Fig. 1. Chemical structures of phenolic acids

The phenolic composition of a wide variety of grains and grain-based products was determined using reversed phase (RP)-high performance liquid chromatography (HPLC) with a diode array detector (DAD), (Mattila et al., 2005).

In whole wheat flour, the bran/germ fraction contributed 83% of total phenolic content . (Liu, 2007).

Moore et al. (2005) studied the total phenolic content and phenolic acid composition of eight Maryland-grown varieties of soft wheat was determined using an RP-HPLC-DAD method , Total phenolic content ranged from 40-80 mg GAE/100 g wheat. Vanillic, syringic, p -coumaric, and ferulic acids were found in all soft wheat samples.

Liyana-Pathirana and Shahidi (2007) studied The effects of milling on the phenolic content and antioxidant capacity of two wheat cultivars, namely CWAD (Canadian Western Amber Durum; Triticum turgidum L. var. durum) and CWRS (Canadian Western hard red spring; Triticum aestivum L.). They found the milling of wheat afforded several fractions, namely bran, flour, shorts and feed flour. In addition, semolina was the end-product of durum wheat milling. Among different milling fractions the bran had the highest phenolic content while the endosperm possessed the lowest amount and this was also reflected in free radical and reactive oxygen species (ROS) scavenging capacity

Iqbal et al. (2007) determined the antioxidant activity of bran extracts from five wheat varieties indigenous to Pakistan, i.e. Punjab-96, Bhakkar-2002, Uqab-2000, SH-2002, and Pasban-90. They found all the bran extracts exhibited appreciable total phenolic content (2.12–3.37 mg gallic acid equivalent/g bran)

As in all plant foods, the phytochemical composition of wheat is greatly affected by genetics, environmental factors, and processing conditions. The total phenolic content of the bran that contains mainly the phenolic acids and polyphenols is 3362–3967 mg/g gallic acid equivalent, in which only 15% was from the methanol/water extractable fraction, i.e., 85% of the total phenolic content in wheat bran was in bound form, bound phenolic acids and other polyphenols may be more important in terms of the health benefits. Despite the large volume of studies on the phenolic acids in wheat, information on other polyphenol content of wheat has been lacking. However, available literature indicates lignans and flavonoids are found in wheat and they may have contributed to the many health benefits of wheat. (Yu, 2008).

Hirawan et al. (2010) Showed that Whole wheat spaghetti exhibited significantly higher levels of total phenolic content (1389 µg/g) than regular wheat spaghetti (865 µg/g); however, TPC (total phenolic contents) in both regular and whole wheat spaghetti was 48–78% of the original content after cooking .

Vaher et al. (2010) determined the total phenolic content of the bran layer; flour made from endosperm and whole grain of wheat. Fifteen different wheat samples of ten spring and five winter wheat varieties were analyzed. The spring wheat varieties were grown in both conventional and organic conditions. They found that total phenolic content of the bran layer was the highest (1258-3157 μg/g), followed by that of grains (168 - 459 μg/g) and the lowest of flour (44 - 140 μg/g).They noticed that Ferulic acid was a major compound among phenolic acids found in wheat varieties.

High levels of total phenolic acids were found in wheat bran (4527 mg/kg) and whole-wheat flour (1342 mg/kg). The total phenolic acid content of refined wheat flour (167 mg/kg) was much lower than that of whole wheat flour (1342 mg/kg) (Neal, 2012).

When antioxidant activity is compared at the free phenolic acid concentrations found in wheat, effectiveness is in the order of ferulic acid > vanillic acid > p -coumaric acid (Baublis et al., 2000b).

Ivanišová et al. (2012). evaluate total phenolic and flavonoid content of cereal extracts on four milling fractions of selected cereals grew in the year 2009 and 2010, they found that flour fractions (break flour and reduction flour) showed the lower proportion of the total antioxidant potential than bran fractions (fine bran and coarse bran), which was balanced in observed years while, extract from barley had the highest values of antioxidant activity and polyphenol content.

3.1.1.3. Sorghum grains as a source of phenols:

Sorghum (Sorghum bicolor L. Moench) is one of the world’s most important stable cereal crops. The top five sorghum producers in 2008 were the United States, Nigeria, India, Sudan and Australia, with the United States and Australia being the top two exporting countries in 2009 (FAO, 2011). In much of Africa and Asia sorghum is an important human food, whereas in countries such as the Unites States and Australia it is used primarily as animal feed. Interest is increasing in potential new food applications of sorghum, particularly due to sorghums resilience against the high temperature and drought conditions that may arise due to climate change ( Oria et al., 2000 and Taylor et al., 2006).

Besides the antioxidant enzymes, sorghum seeds are provided with antioxidant substances that are able to scavenger radical products. These compounds include lipid-soluble products, such as tocopherols and, water-soluble substances such as ascorbic acid and thiols. On the other hand, the chemical components of the sorghum grain cuticle are flavonoids, anthocyanidins and tannins (Kaluza et al., 1988).

Awika et al. (2002) studied the phenolic content of three sorghum varieties; brown (sumac and high tannins), black (Tx430), red (Tx 2911), white, and red wheat bran and found the brown sorghum and there barns had the highest phenols and while white sorghum had the lowest.

All sorghums contain phenolic acids and most contain flavonoids. Only varieties with a pigmented testa have condensed tannins. The types and quantities of phenols present in the grain are genetically controlled. There are a number of myths about sorghum that exist in the scientific community. These myths significantly and adversely affect sorghum use in food and industrial products. (Dykes and Rooney, 2006).

Rey et al. (1993) have identified apigeninidin as a major anthocyanidin present in sorghum. Regarding its physiological function, apigeninidin showed a high fungicidal activity in sorghum (Aida et al., 1996), and it was reported that apigeninidin effectively quenched ascorbyl radical and lipid radicals when supplemented with doses up to 200 mg/ml-1 (Boveris et al., 2001).

Sorghum is a major cereal food crop in many parts of the world; however, it is particularly important as a human food resource and folk medicine in Asia and Africa (Ryu et al., 2006). Sorghum is rich in phytochemicals known to significantly affect human health, such as tannins, phenolic acids, anthocyanins, phytosterols, and policosanols (Awika and Rooney, 2004). According to Choi et al. (2006), sorghum has antioxidant activity.

Young et al. (2009) studied the antioxidant activities of sorghum extracts prepared from 25 cultivars from South Korea. They summarized that the effects of various fractions from different cultivars of sorghum were examined for radical-scavenging activity by DPPH methods and for antioxidant activity by determining reducing power. With the exception of Kkachisusu, Kkachisusu (daerip), and Bitjarususu, all sorghum cultivars examined showed high antioxidant activity.

3.1.1.4. Corn grains as a source of phenols:

Corn is considered an important crop worldwide. There have been some studies in the identification and quantification of phenolic compounds from yellow corn including p -hydroxybenzoic acid, vanillic, protocatechuic, syringic, p -coumaric, ferulic, caffeic and sinapic acids (Adom and Liu, 2002). other phenolic compounds present in purple corn in addition to anthocyanins. Pedreschi, and Cisneros-Zevallos (2007) believed, these phenolic compounds might be involved in health-related properties and could have been missed in previous studies in which bioactive properties were attributed to anthocyanins only.

Pozo-Insfran, et al. (2006) compared polyphenolic and antioxidant content among corn genotypes and confermed that acidifecation post-nixtamalization could reduce polyphenolic and antioxidant losses.

Hodzic et al. (2009) studied total phenolics for corn whole grains. They found that ranged between (9.82 - 11.29) and (14.51- 16.56) mg GA/L at 20 and 40 ˚C, respectively.

3.1.2. Phytate content in cereal grains:

Phytic acid is a common plant constituent, comprising 1–5% by weight of edible legumes, cereals, oil seeds, pollens and nuts (Graf et al., 1987). Phytic acid (myoinositol hexa-phosphoric acid, IP6) is the major phosphorus storage compound of most seeds and cereal grains, it may account for more than 70% of the total phosphorus. Phytic acid has a strong ability to chelate multivalent metal ions, specially zinc, calcium and iron. The binding can result in very insoluble salts with poor bioavailability of minerals (Rhou and Erdman, 1995). The accumulation site of phytic acid in monocotyledonous seeds (wheat, millet, barley, rice, etc.) is the aleurone layer, particularly the aleurone grain. (Okazaki and Katayama, 2005).

Conversely high phytate intake can be a factor in reducing breast and prostate cancer in man (Vucenik and Shamsuddin, 2003). It exerts major antioxidants properties due to its relatively high binding affinity for iron (Graf and Eaton, 1990). Phytic acid is capable of forming an iron–phytate chelate that is totally inert in the Fenton reaction, because it occupies six coordination sites of iron ion and displaces all of the water molecules coordinated in the Fe3+-phytate complex (iron participation in hydroxyl radical formation requires at least one coordination site opened or occupied by water or other readily dissociable ligand). Moreover, phytic acid causes the rapid removal of the Fe2+ without the simultaneous production of hydroxyl radical. As for Fe3+, it was proven relatively inert, even when oxygen and polyunsatu-rated lipids were present in the reaction medium (Graf et al., 1987). Due to its antioxidative potential, phytic acid arouses great interest as a potential food preservative.

Besides its well-known negative properties IP6, by complexing iron, may bring about a favorable reduction in the formation of hydroxyl radicals in the colon (Graf and Eaton, 1993), also positive effect against carcinogenesis have been shown with in vitro cell culture systems, mice, rats and guinea pigs, but the mechanism of action is not understood (Harland and Morris, 1995).

Lee et al. (1998) found that phytic acid was effective for inhibition of the oxidative changes in a model beef system.

The structure of phytic acid is shown in Fig. 2 and then Fig. 3 shows the structure of phytic acid with the different possibilities to interact with both metal cations (minerals) as with protein residues.

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Fig. 2: Structure of phytic acid

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Fig. 3: Structure of phytic acid with the different possibilities to interact with both metal cations (minerals) as with protein residues

Phytic acid has a structure similar to that of myo-inositol which has been demonstrated to reduce hepatic lipid levels. Onomi et al. (2004).

It contains the mineral phosphorus tightly bound in a snowflake-like molecule. In humans and animals with one stomach, the phosphorus is not readily bioavailable. In addition to blocking phosphorus availability, the arms of the phytic acid molecule readily bind with other minerals, such as calcium, magnesium, iron and zinc, making them unavailable as well. Phytic acid molecule contains 12 dissociable hydrogens.

Lasztity and Lasztity (1988) found that, depending on the pH of the solution different phytic acid anions may be formed having different degree of protonation. Except at the pH corresponding to the isoelectric point, protein molecules are also charged. The terminal amino groups like lysyl, histidyl and arginyl can be positively charged at a low pH below the isoelectric point of proteins; any of these groups can directly form a complex with a negatively charged phytate anion. One phytate anion can interact with two charged groups of protein in the normal steric condition. According to the number of positively charged groups and conformational conditions, the protein molecule can bind more phytate anions at the same time. Also at intermediate pH values, the lysyl and arginyl groups are only positively charged, so in this case a slight possibility of electrostatic interactions exists between these groups and phytate anions. The interaction between phytic acid and protein is reduced when the pH is very high. It should also be noted that the ternary complexes can be formed when the polyvalent cations are present. In this case the cation forms a bond between the phytate anion and a negatively charged group of the protein.

Tompson (1986) showed that ternary complexes of protein, phytic acid and carbohydrate might form; the digestion rate of starch is affected in this case. In addition to being an element chelator of important minerals, phytic acid also inhibits the enzymes that we need to digest our food, namely pepsin, amylase and trypsin that are, respectively necessary for protein degradation in stomach, the starch into sugars and proteins degradation in the small intestine.Phytic acid is mainly located in the bran fraction of whole-grain cereals, especially within the aleurone layer (Gani et al., 2012).

Phytic acid from whole-grain cereals has long been considered to be nutritionally negative, since it chelates minerals such as Zn, Fe, Ca and /or Mg, thus limiting their intestinal bioavailability (Lopez et al., 2002).

However phytic acid is also a strong antioxidant in vitro, phytic acid also reduces the incidence of colonic cancer by suppressing the oxidative damage caused to the gut epithelium, particularly in the colon where bacteria also yield oxygenated radicals (Graf and Eaton, 1990). Phytic acid also inhibits xanthine oxidaseinduced superoxide-dependent DNA damage (Muraoka and Miura 2004).

3.1.2.1. Wheat grains as source of phytate:

Garca-Estepa et al. (1999) Assess total phytate content of cereal samples by a complexometric measure of residual iron after phytic acid precipitation and showed the complexing capacity in processed foods (breads) to estimate the phytic acid intake from some products in Spain.

They were determined the phytic acid in cereal (brans, flours and milled wheat products) and breads. The method was based on com- plexometric titration of residual iron (III) after phytic acid precipitation. The cereal flours showed values ranged between 3-4 mg/g for soft wheats, 9 mg/g for hard wheat and 22 mg/g for whole wheat. Wheat brans had wide ranges (25-58 mg/g). The milling products (semolinas) from hard wheat exhibited 10 mg/g and soft wheat a mean of 23 mg/g. The breads made with single or mixture cereal flours exhibited ranges between 1.5 and 7.5 mg/g.

3.1.2.2. Sorghum grains as source of phytate:

Kayodé (2006) stuidied Seventy-six farmers’ varieties of sorghum from Benin, they found the phytate concentration of the grain ranged from 0.4 to 3.5% with a mean of 1.2%.

Abdel Rahman and Osman (2011). Studied the phytic acid content of three three sorghum cultivars named, Fetarita, Safra and Ahmer, widely used in the Sudan. They found that the phytic acid content varied significantly among the three cultivars. It varied from 267.3 mg/100 g in Fetarita to 369.1 mg/100 g in Safra, with Safra variety showing the highest phytic acid content while the Fetarita the lowest one. The phytic acid content of the three cultivars falls within the reported range of phytic acid in sorghum (Doherty et al., 1982), they found phytate-P of thirty varieties of sorghum ranged between 0.17-0.38% (dry whieght). The phytic acid content of Fetarita was comparable to that found by Mahgoub and El-Hag (1998).

3.1.2.3. Corn grains as source of phytate:

Garca-Estepa et al. (1999) assess that total phytate of corn, millet and sorghum flours reported a mean of 10 mg/g and oat, rice, rye and barley between 4 and 7 mg/g.

In corn, IP6 is mostly found in the germ (Febles, et al., 2002; Vucenik, and Shamsuddin, 2006 and Zielinski, et al., 2008). According to Egli et al. (2002) the phytic acid content of the untreated sweet maize was 1.6 g/100 g dry mater.

Corn differs from other cereals, as more than 80% of phytic acid is concentrated in germ. Phytic acid content of cereals varies from 0.5 to 2.0%. Phytate is most known as a substance known to decrease mineral absorption however, it has also been looked at as a possible beneficial vitamin-like substance (Okazaki and Katayama, 2005).

3.1.3. Scavenging capacity of cereal grains:

Antioxidants have been defined in foods as substance that in small amounts are able to prevents or realty retard the oxidation of easily oxidisable material such as fats. Quenching agents may be involved to minimize the progress or the activity of singlet oxygen at several stages in the oxidation of foods.

Frimer (1985) stated that, because singlet oxygen induces and accelerates oxidation of biological molecules, protecting from singlet oxygen is very important. A large number of compounds either natural or biological compounds are known as singlet oxygen quenchers.

Thomas et al. (1990) mentioned that, free radical scavenger that react more rapidly with peroxyl radicals than poly unsaturated fatty acids by donating an electron to the peroxyl radical of fatty acid, and thus stop the propagation steps. The ability of compounds as hydrogen donating can measure by standard 1-electron reduction potentials of these compounds. On the other hand in biological system the antioxidants has been defined as, any substance that, when present at low concentration compared to those of an oxidisable compounds, delays or prevents oxidation of that compounds (Halliwell and Gutteridge, 1990).

Zhou et al. (2005) suggested that antioxidants may amend cellular oxidative status and prevent biologically significant molecules such as DNA, proteins, and membrane lipids from oxidative damage and as a result lessen the risk of several chronic diseases including cancer and cardiovascular disease.

DPPH is a stable organic radical, which bears no similarity to the highly reactive and transient peroxyl radicals involved in various oxidative reactions in vivo (Wu et al., 2004).

Empson et al. (1991) and Graf and Panter, (1991) reported that phytic acid was more effective than ascorbic acid, BHT, or EDTA for lowering TBA values in fresh beef homogenates incubated for 60 min at 37˚C (Lee and Hendricks, 1995).

Liyana-Pathirana and Shahidi (2007) determined The effects of milling on the antioxidant capacity of two wheat cultivars, namely CWAD (Canadian Western Amber Durum; Triticum turgidum L. var. durum) and CWRS (Canadian Western hard red spring; Triticum aestivum L.) they found that among different milling fractions the bran had the highest free radical and reactive oxygen species (ROS) scavenging capacity while the endosperm possessed the lowest amount.

3.2. Effect of domestic processing on antioxidant activity of cereal grains:

Several pre-treatments exist to improve the quality of cereal. Nout wrote that it is the simple traditional household technologies have been used to process the cereal in order to improve the nutritional quality (Nout, 1993).

Phytic acid is hydrolyzed, enzymatically by phytases, or chemically to lower inositol phosphates such as inositol pentaphosphate (IP5), inositol tetraphosphate (IP4), inositol triphosphate (IP3) and possibly the inositol di- and monophosphate during storage, fermentation, germination, food processing and digestion in the human gut (Burbano et al., 1995). Only IP6 and IP5 have a negative effect on a bioavailability of minerals, the other hydrolytic products formed have a poor capacity to bind minerals, or the complexes formed are more soluble (Sandberg et al., 1989).

The possibility to increase phytase activity and/or reduce the phytic acid content by soaking and germination was investigated in a wide range of grains and seeds, but not found to be effective. Germination, but not soaking, increased phytase activity 3 to 5-fold in some cereal grains and legume seeds, while the influence on phytic acid content was insignificant in most materials tested (Egli et al., 2002).

The profiles and quantities of polyphenols and tannins in foods are affected by processing due to their highly reactive nature, which may affect their anti-oxidant activity and the nutritional value of foods (Dlamini et al., 2009).

Soaking, germination and pressure-cooking proved to be effective household strategies to reduce the levels of polyphenols and tannins in grains (Shweta et al., 2010).

3.2.1. Effect of soaking on antioxidant content:

In general, soaking period was reported to have pronounced effects on the vitamin levels and anti-nutritional factors present in natural foods (Fadahunsi, 2009). Soaking is widely applied at both household and industrial scale. It is the most important operation in the germination or fermentation process of cereals. (Lestienne et al., 2005a,b). Soaking of sorghum flour (80% extraction) at room temperature for 24 h reduced phytic acid levels by 16-21% (Mahgoub and El-Hag, 1998).

Soaking of pounded maize for 1 hr at room temperature already led to a reduction of phytic acid by 51% (Hotz et al., 2001). Although mostly focused on grains and beans, research has shown soaking to be quite effective for the reduction of phytic acid in as little as 12 h as well as the subsequent increase in mineral availability (Mbithi- Mwikya et al., 2000; Egli, et al., 2002 and Perlas and Gibson, 2002). Glennie (1983) Studied the effect of soaking process on sorghum. He reported that concentration of total the phenolics of white sorghum ranged from 80 to 100 mg/100 g.

Yang (2009) reported that the total phenols content of non-tanin sorghums ranged from 90-1820 mg gallic acid equivalent (GAE)/100 g sample.

Afify et al. (2012a) investigated the changes in total phenols, and other phytochemicals, during soaking of three white sorghum varieties. They found that the losses of total phenols ranged between 21.97% to 28.30 in sorghum after soaking.

3.2.2. Effect of soaking on antioxidant activity:

According to Afify et al. (2012b), the DPPH scavenging activity in raw sorghum varied from 21.72 to 27.69% during soaking of three white sorghum varieties. Several studies demonstrated that soaking is a biological process of significant impact on phytate and phenolic compounds (Bvochora et al., 2005).

Khandelwal et al. (2010) studied Effect of soaking, germination and pressure cooking of Indian pulses. They reported that the processing reduced the concentrations of polyphenols by 19–59% and of tannins by 22–59%. A trend was observed in the degree to which processing reduced polyphenol and tannin contents (germination > pressure-cooking > soaking). Soaking, germination and pressure-cooking proved to be effective household strategies to reduce the levels of polyphenols and tannins in pulse-based foods, thereby enhancing the bioavailability of pulse protein

3.2.3. Effect of germination on antioxidant content:

The process of cereal seeds germination has been used for centuries for the purpose of softening the kernel structure, improving its nutritional value, and reducing anti-nutritional effects. In fact, the germination process is also one of methods used to improve the functionality of oat seed protein (Kaukovirta-Norja et al., 2004).

Egli, et al. (2002) found that the phytic acid content of sorghum and corn grains decreased during germination, except buckwheat. They concluded that neither soaking nor germination was a useful approach to increase phytase activity or phytic acid content in grains and seeds.

Cevallos-Casals and Cisneros-Zevallos (2010) analyzed 13 edible seeds for the levels of phenolic compounds and total antiradical capacity (TAC) at different germination states (dormant, imbibed and 7d sprouts). Accumulated phenolics (mg chlorogenic acid equivalent, CAE) and TAC (μg Trolox equivalent) on dry basis (DB) showed the general trend distribution of 7d sprouts > dormant seeds > imbibed seeds. Phenolic contents of 7d sprouts (DB) ranged from 490 (lentil) to 5676 (mustard) mg CAE 100 g-1. Increases in phenolics (DB) from dormant seed to 7d sprout differ among seeds from 2010% (mung bean) to -11% (kale), while increases in TAC (DB) ranged from 1928% (mung bean) to 0% (lentil). This study showed that germinated edible seeds are an excellent source of dietary phenolic antioxidants.

3.2.4. Effect of germination on antioxidant activity:

According to Frias et al. (2004) during germination, various reactive oxygen species (ROS) are generated as byproducts of metabolism. This group of ROS includes superoxide radicals (O2-), hydrogen peroxide radicals (pO2.-), and hydroxyl radicals (OH.-). The formation of these oxygen radicals results in the accumulation of lipid hydroperoxides by radical chain oxidation via phospholipids peroxy radicals within membranes. Therefore, it was hypothesized that this could be related to the increase of antioxidant activity in large unilamellar vesicles observed in germinated seeds. Lopez-Amoros et al. (2006) studied the effects of varying germination conditions for beans, lentils and peas, at semi-pilot scale, on bioactive compounds. They found that peas and beans undergo a significant increase in antioxidant activity after germination, whereas lentils show a decrease.

Fernandez-Orozco et al. (2009) investigated the effect of germination to improve the antioxidant properties of chickpeas. The results indicated that germination caused an increment in total phenolic content, increased peroxyl radical-trapping capacity (16-55%) and trolox equivalent antioxidant capacity (TEAC) (12-23%). A slight inhibition of lipid peroxidation inhibition was observed. Total phenolic compounds highly contributed to total antioxidant capacity. Results indicated that with the germination, the antioxidant properties of chickpea flours are enhanced and they can be used as desired ingredients for new functional food formulations

Tarzi et al. (2012) studied the germination effect on phenolic content and antioxidant activity of chickpea seeds. They found that the germination process causes various changes in the phenolic compounds and modifies the antioxidant activity. Acetone extracts of chickpea sprouts which contained higher phenolic compounds showed better antioxidant activities.

3.3. Using cereal grains milling fractions as natural source of antioxidant to improve meat products quality during storage:

The oxidative stability of meat depends upon the balance between anti- and pro-oxidants, including the concentration of polyunsaturated fatty acids. Meats containing high contents of polyunsaturated fatty acids, as in pasture feeding, should be more prone to oxidation than those with more saturated ones. But these two diets may also have differences in anti-oxidant content especially vitamins, carotenoids and/or flavonoids which can protect meat against oxidation (Wood and Enser, 1997). These anti-oxidants cannot be synthesized in animals and higher concentrations are found in green herbage compared to cereals (Daly et al., 1999).

The oxidation of lipids begins at the moment of slaughter and then occurs during storage, processing, heating and further storage of meat. Being the basic cause of fat rancidity, it leads to deterioration of quality or even spoilage of raw materials and food products. Heating of meat, followed by a low temperature storage, always results in the development of warmed-over flavours (WOF). The most important process in oxidation of lipids in meat is peroxidation of polyunsaturated fatty acids from cell membranes.

In meat, amino acids with reactive side chains, for example, amino and sulfhydryl groups, are especially susceptible to oxidation. Formation of carbonyls is one of the most important changes in oxidized proteins. Carbonyl compounds can be generated via direct oxidation of amino acid side chains, fragmentation of the peptide backbone, reaction with reducing sugars, or binding to nonprotein carbonyl compounds ( Xiong, 2000) .

Lipid as well as protein oxidation can be inhibited by using antioxidants. In meat, food additives, such as ascorbic acid and phosphates, increase the antioxidant protection. The antioxidant activity of plant extracts containing phenolic compounds against lipid oxidation has been investigated earlier, and thus plant extracts (or their phenolic extracts such as rosemary) might provide an alternative to food additive use.(Karpin˜ska et al., 2000 and Mc-Carthy et al., 2001).

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