Selected Powerful Natural Antioxidants : Structure , Food Sources , Antioxidant Activities , and Important Health Benefits

Numerous epidemiological studies indicate that consumption of antioxidant-rich foods is beneficial to human health and reduces the risks of dangerous diseases and premature aging. Among natural antioxidants some stand out for their powerful activity and health benefits and they are epigallocatechin gallate, quercetin, curcumin, resveratrol, hydroxytyrosol, astaxanthin, lycopene, dihydroquercetin, and lignans. Preclinical, clinical, and therapeutic studies of these antioxidants in their pure form or in their combination have shown positive health benefits. This review highlights basic information and interesting findings with their source, structure, antioxidant properties, and potential health benefits to human.


Introduction
Epidemiological studies have demonstrated that consumption of antioxidant rich foods and beverages reduces the risks of dangerous diseases and premature aging in human (Preedy, 2010;Klein, 2014;Preedy, 2014;Waltson, Preedy, & Zibadi, 2014;Preedy, 2014;Yashin, Vedenin, & Yashin, 2016;Chu, 2012).Under excessive oxidative stress, the natural human antioxidant system is not able to neutralize the deleterious effects of free radicals which could damage cellular DNA, lipids, proteins, and other biomolecules that led to the development of chronic diseases and premature aging.Antioxidants are required to scavenge free radicals and prevent their actions in vivo to protect cells and tissues.
Several dietary antioxidants are primarily originated from fruits, vegetables, berries, spices, nuts, tea, coffee, cocoa, and red wine.Food databases of the total antioxidant content in various foods have been created during recent years and the intake of natural antioxidants in different countries have been evaluated (Yashin et al., 2010;Nemzer, Yashin, & Yashin, 2013;Wu et al., 2004).Halvorsen et al., (2006) evaluated and ranked 1120 food samples from USDA for total antioxidant activity and found that top 50 antioxidant rich foods are occupied by spices, fruits, vegetables, berries, chocolate, and nuts (Halvorsen et al., 2006).Along with proteins, fats, carbohydrates, vitamins, and trace elements, natural antioxidants are recognized as an important component of a healthy diet.In fact, antioxidant potential plays significant role in the marketing of antioxidant rich superfoods.
An antioxidant functions by transferring electrons, donating hydrogen, reducing peroxides, quenching singlet oxygen and superoxide, and chelating metal ion (Brewer, 2011;Nimese & Pal, 2015).Several in vitro antioxidant assays have been developed in continuation of the searching and identifying natural and safe antioxidants.Due to their differences in structures and reaction mechanisms of free radical several assays have been used to measure the antioxidant activities.For instance, ORAC (oxygen radical absorbance capacity), DPPH (diphenylpicrylhydrazyl), ABTS (2,2-azobis-(3-ethyl-benzothiazoline-6-sulfonate), (FRAP (ferric-reducing antioxidant power), superoxide anion radical scavenging, phospholipid peroxidation assays have been widely used as in vitro tests to determine the antioxidant potential for synthetic and natural products.In most of the studies multiple assays were adopted to test the antioxidant capacity of number of plant extracts and phytochemicals as single antioxidant assay may not give conclusive information on antioxidant potential.
Antioxidant studies of fruits, vegetables, beverages, and functional foods have been attracting continuously for long time and the number of reports in this area increasing every year.Most of these reports covered the polyphenols content and their effect on antioxidant activities with their mechanism of action using appropriate antioxidant assays.These studies indicated the strong antioxidant activities arose due to the presence of certain polyphenolic and/or synergism of several antioxidant compounds (Brewer, 2011;Wu et al., 2004;Oroian & Escriche, 2015).Plant extracts and food matrices contain complex mixtures of polyphenols.Likewise, numerous compounds from these extracts have been reported to have potential antioxidant activities, many plant extract and food matrices are poorly characterized.However, among these natural antioxidant compounds very few have strong antioxidant activities.In this review we describe an overview of selected most powerful natural antioxidant compounds with their structure, source, bioavailability, antioxidant properties, and potential health benefits.

Powerful Natural Antioxidants
Antioxidants are subgrouped into three major groups such as polyphenols, vitamins, and carotenoids.In addition to these general group of antioxidant compounds some other nonphenolic compounds also showed promising antioxidant potential.Polyphenols are ubiquitous in plant species and more than 8000 phenolic compounds have been reported, many of which are major constituents of foods.Classification of polyphenols are based on the number of phenol rings and the nature of binding the rings to one another.Major groups of polyphenols constitute flavonoids, phenolic acids, phenolic alcohol, stilbenes, and lignans.Among them most common polyphenols are flavonoids that provide color and flavor to fruits and vegetables.Flavonoids are subgrouped into flavones (e.g., luteolin, tangeritin), flavonols (e.g., catechin, quercetin), flavanones (e.g., butin, naringin), flavanols (e.g., catechin, epicatechin gallate, epigallocatechin), anthocyanidins (e.g., petunidin pelargonidin, malvidin), and isoflavones (e.g., genistein, daidzein).Natural flavonoids exist as glycosides (sugar is attached) as well as aglycones forms.Most of the polyphenols exist in conjugated forms with one or more sugar residue linked to hydroxyl group.
The antioxidant potential of natural antioxidant depends on the structural composition of free hydroxyl (-OH) group in the flavonoid skeleton.Flavonoids with multiple hydroxyl groups show higher antioxidant activities compared to flavonoids with single hydroxyl group.However steric relationship and lipid/hydrophilic phase equilibrium also plays critical role towards the antioxidant activities (Brown & Kelly, 2007).The general mechanism of antioxidant activities of polyphenolic compounds undergoes inactivation of free radicals via transfer of hydrogen atom and electron.However, efficiency of in vivo and in vitro antioxidant activities depends on various factors such as structure, source, bioavailability of antioxidants, genetic, environmental, and physiological conditions.
Based on the reports on antioxidant activities of natural antioxidants very few have super strong antioxidant activity and have been used by consumers worldwide.We have listed nine such natural antioxidant compounds in the Table1 including their structure and major sources.

Epigallocatechin Gallate (EGCG) and other Catechins
Catechins are one of the major flavonoid compounds that constitute strong antioxidant activities.The common structural unit of catechin composed of a C15 (C6-C3-C6) skeleton containing 3 ring (A, B, and C) phenolic compounds where a double ring attached to a third ring (each phenyl ring has multiple hydroxyl group) by a single bond.In addition to the significant antioxidant properties tea catechins, they have the novel characteristic of trapping reactive carbonyl species.The ring A of the catechins is responsible for reactive carbonyl species trapping, and ring B contribute towards antioxidation (Wang & Ho, 2009).
Green tea is the major source of catechins containing15 to 20% of dry weight (Yashin, Vedenin, & Yashin, 2016;Juneja, Kapoor, Okubo, & Rao, 2013).Black tea and oolong tea which are fully and partially fermented tea respectively, also contain a significant level of catechin compounds.However, due to differences in manufacturing process polyphenol contents in black tea and oolong tea are different from green tea.Nonfermented green tea products contain higher levels of total phenolics and catechins.ECCG is reported as the main active constituent of green tea which comprises 50 to 80% of the total catechin content.In addition to ECCG epicatechin gallate (ECG), epigallocatechin (EGC), epicatechin (EC) and catechin (C) are some other catechin available in tea.Other sources of catechins include grapes, red wine, cocoa, and dark chocolate.Dry red wine contains catechin (191 mg/L) and epicatechin (82 mg/L) at levels that are 4 to 6 times higher than in white wine.Cocoa and dark chocolate contain catechin and epicatechin only.In cocoa they make up to 40% of the total polyphenol content.Catechins are also found in white and yellow tea which are not fermented.
Number of studies indicated catechin extract and purified fraction from all these natural resources exhibited significant antioxidant activity in vivo, in vitro, and in clinical studies.Recently He et al., (2018) studied the antioxidant potential of some catechins and reported that epigallocatechin gallate had the highest radicalscavenging activity and significant role to retard the ROS production because of the presence of hydroxyl and galloyl groups (He, Xu, Yang, & Wang, 2018).Grazseik et al., (2018) reported in a study that catechins showed the highest activities in ABTS-scavenging capacity and FRAP assay (Grazseik, Naparlo, Bartosz, & Sadowska-Bartosz, 2018).
Quercetin is highly abundant in fruits and vegetables.Onion is reported to have the highest level of quercetin (approx.300 mg/kg) (Beecher, 1999).Other vegetables, such as broccoli, kale, apples, cranberries, yellow bell peppers, celery, tomatoes, blueberries, blackberries, raspberries, black currants, cherries, and apricots also have significant range of quercetin.Processing and storage could change the content of quercetin in food matrices.Ikou et al., (2001) found that frying and boiling cooking methods reduces the level of quercetin due to the thermal degradation and leaching to the water (Ikou et al., 2001).Long-term storage of foods was found to change their quercetin content.Efforts have been directed for enhancing nutraceutical bioaccessibility for quercetin.In order to improve the bio-accessibility of this hydrophobic nutraceutical formulated emulsion-based excipient food were prepared (Chen et al., 2018).Quercetin encapsulated in nanoparticle, for instance Zein/chitosan nanoparticles (ZCPs-Q) have also been prepared which improved its stability and water solubility and intracellular antioxidant activities (Ma, Yu, Yin, Tang, & Yang, 2018).

Curcumin
Curcumin is a polyphenolic compound with the molecular formula of C 21 H 20 O 6 containing α, β -diketo group, carbon-carbon double bonds, and phenyl rings with various hydroxyl and methoxy groups.It exhibits keto enol tautomerism.It is major component of turmeric and soluble in lipids.It is a unique antioxidant with significant antioxidant activity and reported to be an excellent free radical scavenger.Antioxidant activity of curcumin related strongly with the phenolic OH group.Studies on structure-activity relationship(s) for the antioxidant effects of curcumin are controversial.Whereas few group researchers claimed that the strong activity of curcumin is because of the presence of hydroxyl moiety, some other researchers proposed the involvement of the carbonyl groups.Barclay et al., (2000) reported that curcumin could donate H-atoms from the phenolic group (Barclay et al., 2000).In another study it was reported that curcumin is a strong H atom donor from the methylenic group rather than from the phenolic group (Jovanovic, Steenken, Boone, & Simic, 1999).Priyadarsini et al., (2003) have also proposed that the phenolic group plays active role for the free-radical-scavenging activity indicating the increased activity by the methoxy group (Priyadarsini et al., 2003).Thus, curcumin features unique chain breaking antioxidant ability which is correlated to the phenolic OH group and CH 2 group of the diketone moiety.
Curcumin is found in turmeric in significant amounts.Turmeric is one of the components of the curry spice mixture widely used in cooking.In Asian medicine, curcumin has been used for more than 4,000 years.It is non-toxic even at high doses.Consumption of up to 12 g/day is well tolerated.Curcumin can be detected in blood serum in 1.5-2 hours after consumption of turmeric.One major problem with curcumin is its lower bioavailability.To address this problem several formulations with nanoparticles, liposomes, micelles, and phospholipid complexes have been prepared (Peng et al., 2014;pawar et al., 2012;Ghosh et al., 2012, Yallapu, Dobberpuhl, Maher, Jaggi, & Chauhan, 2012;Gong et al., 2013;Pandelidou, Dima, Georgopoulos, Hatziantoniou, & Demetzos, 2011).The recommended consumption level of curcumin is 100 mg/day.Significant loss of spice ingredient was observed when the spices were subjected to heat treatments in cooking methods (Jayashree et al., 2016).

Resveratrol
Resveratrol is one of the major nonflavonoids phenolic compounds where two aromatic rings are joined by an ethane linkage.Resveratrol exists in monomeric, oligomeric or polymeric form of stilbenes.It is a natural phytoalexin in the fruits and vegetables such as peanuts, mulberries, grapes, red cabbage, spinach, and red wine.The French paradox of wellness and wine consumption is attributed to the effect of resveratrol (Ferreiers, 2012;Renaud, de Lorgeril, 1992;Vidalayur, Otani, Singal, & Maulik, 2006).
Resveratrol is currently receiving considerable attention worldwide because of its beneficial effects on the human health including significant antioxidant activity.Number of studies reported that resveratrol has strong antioxidant and radical scavenging capacity against DPPH, ABTS, DMPD, and superoxide radical and metal chelating activities.In the study Gülç in, (2010) found that compared to standard antioxidant resveratrol inhibited higher lipid peroxidation (89.1%) of the lipid peroxidation of linoleic acid emulsion than BHA, BHT, α-tocopherol, and trolox (approximately 80%) at the concentration of 30 µg/ml.(Gülç in, 2010) Table 1.Food sources of strong natural antioxidants

Antioxidant
Structural Formula Food Sources Analytical techniques Cooking process such as roasting, boiling, and frying reduces the level of resveratrol.One of the big challenges for resveratrol is the low availability even after it is well absorbed and thus result in low plasma concentrations.

Epigallocatechin
In number of studies it was reported resveratrol as effective "therapeutic agent", however clinical and in vivo animal model studies indicated inconsistent results due to its low bioavailability showing sometime even 0% (Thazhath et al., 2016, De Vries, Strydom, Steenkamp, 2018).To enhance the bioavailability, better permeability and resistant to hydrolysis various formulation and alternative oral rout have been prepared (Watkins, Wu, Zhang, Davis, & Xu, 2015, Ansari, Vavia, Trotta, & Cavalli, 2011).

Hydroxytyrosol
Hydroxytyrosol is one of most potent natural antioxidant compounds with o-diphenols compounds under the group of phenolic alcohol compounds.Compared to the tyrosol, hydroxytrosol has one more OH group in the benzene ring and it contributes towards higher free radical scavenging and antioxidant activity.It is an amphipathic compound which has a water-soluble and fat-soluble component with alipophilic end and hydrophilic end.This unique structural feature makes hydroxytyrosol a good transporter of substances and penetrate the cellular membrane in human body (Ferná ndez-Bolaños, López, López-Garcí a, & Marset, 2012, Martí nez, Ros, & Nieto, 2018).
Hydroxytyrosol is present in olives and olive oil, as well as in other vegetable oils.It attributes intense flavor and aroma.Being reported hydroxytyrosol, tyrosol, and oleuropein as main polyphenolic antioxidants in olive oil, hydroxytyrosol has strongest antioxidant effects.Hydroxytyrosol is is fifteen and three times more antioxidant than green tea coenzyme Q10 respectively (Richards, 2012).The antioxidant properties of hydroxytyrosol has been explained based on various molecular, cellular and animal model studies.With respect to superoxide anion Visioli et al., (1998) proved that hydroxytyrosol has potent scavenging activity of superoxide formation (Visioli, Bellemo, & Galli, 1998).
Virgin olive oil phenolic compounds are highly absorbed and showed bioavailability upto 70 to 99% depending on the age, hormonal status or gender of the patients.After ingestion they are absorbed in a dose-dependent manner.In a study with the administration of solution of hydroxytyrosol and tyrosol with olive and water Tuck et al., (2002) found higher bioavailability with olive oil compared to water (99% and 98% respectively in olive oil solution, 75% and 71% in aqueous solution) (Tuck & Hayball, 2002).Hydroxytyrosol is resistant to gastric juices and significantly bioavailable (Khymenets et al., 2016, Martí nez, Ros, & Nieto, 2018).

Astaxanthin
Astaxanthin is one of the strong natural carotenoid pigments and known as "the king of the carotenoids,".Astaxanthin is a member of the xanthophylls and consists of two terminal rings joined by a chain of conjugated double bonds.Astaxanthin occurs as stereoisomers, geometric isomers, free and esterified forms because of the presence of the polyene bonds.In this oxygenated derivative molecule, it has two rings with hydroxyl and keto moieties with two asymmetric carbons in both ends and exist in esterified (one or both) forms by reacting with a fatty acid, protein or lipoprotein (Ambati, Phang, & Ravi, 2014).
Astaxanthin are naturally available in genus algae, bacteria, fungi, yeast, salmon, lobster, trout, krill, shrimp, and crayfish.The astaxanthin content in common salmon products has been measured by the U.S. Food and Drug Administration that ranges from 5.4 mg/kg to 40.4 mg/kg.Astaxanthin is also found in red fish roe.Another source of astaxanthin is a yeast, Phaffia, which grows on the bark of certain trees.Astaxanthin is found in its highest natural concentration in wild Pacific sockeye salmon in the range of 26-38 mg/kg flesh.Heat treatment reduce the level of astaxanthin while it is stable at 70-90 °C.Improved storage condition at 4 °C and 25 °C were developed by mixing the astaxanthin with cyclodextrin-water mixture (Yuan, Du, Jin, & Xu, 2013).Moreover lipid-based formulation of astaxanthin enhanced its bioavailability in human study (Olson, 2004).
Due to the unique structure of astaxanthin containing double bond, hydroxyl and keto group, it showed high antioxidant activity and even higher in comparison to other carotenoids such as lutein, lycopene, α-carotene and β-carotene (Naguib, 2004).In another study it was found that antioxidant activity of astaxanthin was 10 times higher than carotenoids including zeaxanthin, lutein, canthaxanthin, β-carotene and compared to α-tocopherol it was 100 times higher (Liu & Osawa, 2007).Whereas the oxo groups in astaxanthin result in high antioxidant activities the polyene chain as well as the terminal ring in astaxanthin traps radicals and scavenge radical's activity (Martin, Jagger, Ruck, & Schmidt, 1999).Astaxanthin protect the cell membrane from lipid peroxidation by scavenging the radicals at the outer and inner cell membranes (Ambati, Phang, & Ravi, 2014).In a study Augusti et al., (2012) found the elevation of antioxidant enzymes activities of superoxide dismutase and thioredoxin reductase in serum when astaxanthin were fed to rabbits (Augusti et al., 2012).Similar observations were reported Kamath et al., (2008) when ethanol-induced gastric ulcer rats were treated with astaxanthin (Kamath, Srikanta, Dharmesh, Sarada, & Ravisankar, 2008).

Lycopene
Lycopene is one of the major carotenoids among other 600 different carotenoids which could play crucial role in scavenging peroxyl radicals.It is an acylic isomer of beta carotene and contain a straight chain hydrocarbon with 11 conjugated and two non-conjugated double bonds.Due to the presence of these double bonds lycopene exist in cis-trans isomers.
Lycopene is found mainly in red fruits and vegetables such as tomatoes, watermelons, wolfberry, papaya, pink-grapefruits, apricots, pink-guavas, and peppers.In most of the natural resources it exists as trans isomerism.However, it can undergo cis-trans isomerisation with light heat and chemical reactions.Processing could significantly affect the bioavailability of lycopene by the trans to cis transformation.In a study it was found that lycopene from processed tomato juice in trans form is more bioavailable than raw tomato juice (Stahl & Sies, 1992).In human plasma trans lycopene are more bioavailable than cis isomers (Clinton et al., 1996).
During lipid peroxidation peroxyl radical are generated and that could damage the lipids in the cell wall.

Dihydroquercetin
Dihydroquercetin, also known as taxifolin is a flavanonol compounds which has two hydroxyl phenolic groups at the meta and para-positions in the phenyl rings in the flavonoid's skeleton.
Taxifolin is considered as antioxidant-rich functional food.It is highly available in citrus fruits and onion.Dihydroquercetin has a high antioxidant activity with approximately 60,000 ORAC units (Yashin et al., 2018;Pillow, et al., 1999).Due to its conjugation behavior and resonance structure stability of phenolic rings it provides powerful radical-scavenging activity.In a study Topal et al., (2016) found that taxifolin had remarkable radical scavenging with DMPD, ABTS, superoxide, and DPPH radicals, and metal chelating activities.It has been reported that scavenging activity of taxifolin against superoxide anion is more than that of BHA, BHT, α-tocopherol, and Trolox (Topal et al., 2016).

Lignans
Lignans are the group of phenolic compounds that is derived from the propylbenzene (C6-C3) unit.
Flaxseed (Linum usitatissimum L.) is a major source of of lignans.However other seeds, nuts, fruits, vegetables, coffee, tea, and wine also contain substantial amounts of lignans.Secoisolariciresinol diglucoside (SDG) is the major lignan present in flaxseed and it exist as component of a linear ester-linked complex.It has been observed that SDG is converted into enterolactone (ED) and enterodiol (EL) after its metabolism via the formation of secoisolariciresinol (SECO), a plant aglycon formed by the hydrolysis of SDG.
Flaxseed lignan secoisolariciresinoldiglucoside (SDG) and mammalian lignans enterodiol (ED) and enterolactone (EL) have been reported to play important role in protection from DNA damage and lipid peroxidation.In a study Hu et al., (2007) found that efficacy of lignans with the order of SDG > SECO>>ED>EL for DNA damage and SDG>SECO=ED=EL for liposome lipid peroxidation (Hu, He, Jiang, & Xu, 2007).Strong antioxidant activity of SDG and SECO were observed due to the presence of 3-methoxy-4-hydroxyl substituents in SDG while the mammalian lignan had only single hydroxyl group.The strong antioxidant activity of the mammalian lignans in an aqueous environment were appeared due to the ability of the benzylic hydrogen abstraction and resonance stabilized phenoxyl radicals.

Conclusion
The review describes an overview of selected most powerful natural antioxidant compounds with their structure, source, bioavailability, antioxidant properties, and key healthcare effects.In addition to these selected antioxidant compounds several other antioxidant compounds have been reported with significant antioxidant activity such as phenolic acids in coffee and berries and anthocyanins in berries and fruits, isoflavones in soy, and proanthocyanidins in cranberries.From the recent studies on the powerful antioxidant compounds it was found that antioxidant activity of a natural antioxidant depends on the number of hydroxyl group in the phenylring of the phenolic compounds as well as carbon carbon bonds in aliphatic skeleton.For few decades, efforts have been made by researchers to fill the knowledge gap between consumers and food nutrition in search for healthy diet.Information on such natural oxidants is expected to insight valuable source since current dietary guidelines recommend increasing the antioxidant rich foods.Despite the evidence of efficacy and safety of antioxidant compounds it has been found that most of the antioxidant compounds possessed limited bioavailability.Further studies to improve the bioavailability and medicinal value are needed for potential application.