Antioxidants in Tea

Reactive oxygen species (ROS) and free radicals (FR) are highly reactive molecules that have the potential to damage other molecules in the human body. These could be naturally generated in the body and could also enter the body through inhalation of polluted air, smoking and contaminated food. Presence of excessive amounts of ROS and FR is called as ‘oxidative stress’.

Prolonged oxidative stress has been found to be one of the key factors in the development of non-communicable diseases (NCD’s) such as heart disease, stroke, cancer, diabetes and neurodegenerative diseases. The human body has protective mechanisms against oxidative stress through antioxidant enzymes and antioxidant molecules. In addition, antioxidant molecules derived from the diet also play an important role in the protective mechanism against oxidative stress. Antioxidants in the diet includes Vitamin C, Vitamin E, carotenoids and plant flavonoids.

Tea has been identified as a plant with uniquely high flavonoid content. Therefore, all types of tea have high antioxidant capacity compared to other food. Antioxidant activity of tea flavonoids has been demonstrated in both in vitro and in vivo experiments. Tea flavonoids have the ability to neutralize or scavenge the FR and ROS in the human body. Research has indicated that tea flavonoids are capable of exerting antioxidant activity through other mechanisms, in addition to scavenging of FR and ROS. Therefore, regular tea consumption would have beneficial effects through the antioxidant activity of flavonoids.

Antioxidants

In recent times, antioxidants have become a ‘buzzword’ in relation to food. People have become interested in antioxidants in food, as antioxidants have the ability to effectively limit the damage caused to our bodies by certain molecules, which are either produced internally, or enter from outside. These harmful molecules are referred to as free radicals (FR) and reactive oxygen species (ROS).

Free Radicals and Reactive Oxygen Species

Most molecules in our body are fairly stable and unreactive. They would undergo reactions only when they enter natural metabolic processes, essential for normal functioning, that are taking place in our body.

In contrast, free radicals are highly unstable molecules. In order to stabilize, they would react with any other molecule in the vicinity. This process, whilst stabilizing the free radical, would damage the molecule that it reacts with often making it a free radical. The newly generated free radical would react with another molecule to stabilize itself thereby initiating a chain reaction. Thus the presence of free radicals in our body, could initiate a chain of reactions that would damage other important molecules in the human body and, unchecked, has the potential to disturb its normal functioning. Further, damage to certain types of molecules could play an important role in the development of NCD’s, such as heart disease, stroke and cancer. Reactive oxygen species (ROS), are highly unstable reactive molecules that contain oxygen, which is present in the body. Over production and accumulation of ROS could also initiate chain reactions that could potentially lead to NCD’s. Presence of excessive quantities of FR and ROS will subject the body to ‘oxidative stress’.

The accumulation of FR and ROS in the human body could take place, either through production in the body itself or through exposure to FR and ROS from outside.

Production of FR and ROS could take place in the normal metabolic processes that take place in our body. For example, energy required for the functioning of the body is obtained by utilization of energy stored in carbohydrates, fat or proteins, a process called ‘cellular respiration’. In cellular respiration, oxygen reacts with glucose (or other stored energy producing molecules), producing water, carbon dioxide and energy. This is similar to burning fuel to obtain energy. When firewood or petroleum products are burned energy in the form of heat is produced. However, production of toxic compounds, which are in the smoke, cannot be avoided in the process.

Energy production in the body is also a process of burning fuel, which are carbohydrates, fat or protein that come from our diet. Production of toxic molecules such as FR and ROS cannot be avoided in the cellular respiration also. However, the process inside the body is much more controlled and amounts of toxic material produced is far less than in burning of firewood or petroleum

products. In addition to cellular respiration, many other biochemical processes are taking place inside the body which could also produce FR and ROS.

FR and ROS, and other toxic material, could enter the human body from outside, through the inhalation of polluted air or in the food consumed. Another common method of exposure to FR, ROS and other toxic compounds is smoking. Smokers voluntarily expose themselves to the toxic material in tobacco smoke. Diet could also contain toxic molecules due to contamination and adulteration. Common contaminants are pesticide residues in agricultural products and aflatoxins due to fungal contamination, though all contaminants are not either FR or ROS.

The human body always tries to excrete toxic molecules as quickly as possible through urine, by converting the toxins to water soluble compounds. Sometimes FR and ROS could be produced in the detoxification process. Exposure to ionizing radiation such as ultra violet (UV) light could also produce FR and ROS in the body, which often leads to skin cancer. Further, heavy metals could also facilitate the generation of FR and ROS in the body.

Free Radicals and Reactive Oxygen Species in the Development of Non Communicable Diseases

Oxidative stress has been implicated in the development of many NCD’s. For example, reaction of FR and ROS with DNA (Deoxyribonucleic acid), the genetic material in our body which carries the genetic instructions used in the growth, development, functioning and reproduction, have the potential to disrupt some of these processes. More importantly, DNA damage could lead to carcinogenesis.1

Through similar mechanisms prolonged oxidative stress could potentially lead to the development of other NCD’s such as heart disease, stroke, neurodegenerative diseases and diabetes. 2, 3, 4

Antioxidants and Antioxidant Mechanisms on FR and ROS

FR and ROS in the body should be neutralized to avoid prolonged oxidative stress, which could predispose the individual to NCD’s. Antioxidants are capable of neutralizing (or scavenging) FR and ROS.

The human body has natural antioxidant defense systems which harness enzymes and other molecules with the capability of neutralizing the FR and ROS. Super Oxide Dismutase (SOD), Catalase (CAT), Glutathione Peroxidase (GTPx), Glutathione Transferase (GTS) are the important antioxidant enzymes in the human body, Glutathione being one of the most important.5

In addition to the above, certain molecules, such as Vitamin C (Ascorbic Acid) and Vitamin E (α-Tocopherol), play an important role in the anti-oxidant defence mechanism. Carotenoids and plant flavonoids are such important antioxidant molecules.

Plant Flavonoids

Plants produce flavonoids for various reasons and play an important role in the interaction of plants with other organisms in the environment. They have anti-feeding activity against insect pests and anti-fungal activity towards pathogenic fungi. Therefore, plant flavonoids are important in the plant defense systems. Thus all plants contain varying amounts of flavonoids, distributed in all parts of the plant.6 As such, food derived from plant sources would contain varying amounts of flavonoids.

Flavonoids in Tea

Fresh tender shoots of tea, which are used for processing into different types of tea such as Black tea, Green tea and Oolong tea, contain significantly high amounts of flavonoids compared to other plants. Therefore, a serving of any type of tea would provide significantly higher amounts of flavonoids than a serving of other flavonoid containing food.

Fresh tea shoots contain flavanols as the major water soluble component (18 - 32%) and flavonols (3 - 4%). During black tea processing most of the flavanols are converted to other types of flavonoids (theaflavin and thearubigins) while in green tea processing they remain unchanged. Flavonols remain unchanged during processing in both types of tea. (See chapter 2 for details on flavonoids in tea)

Due to the high content of flavonoids in tea, antioxidant activity of all types of tea would be significantly higher than any other food. Dietary surveys on the total flavonoid intakes have shown that tea is the major source of flavonoids, even in countries which are not considered as major tea consuming countries, emphasizing the high flavonoid content in tea. Zutphen elderly study in the Netherlands and dietary survey on flavonoid intake of US adults are two such studies. 7, 8

Comparison of Antioxidant capacity of tea and common vegetables

FIGURE 4.1 Total antioxidant capacity of tea and common vegetables9

Green tea Black tea Garlic Kale Spinach Brussels sprouts Alfalfa sprouts Broccoli flowers Beets Red bell pepper Onion Corn Egg-plant Cauliflower Potato Sweet potato Cabbage Leaf lettuce String bean Carrot Yellow squash Iceberg lettuce Celery Cucumber

0 200 400 600 800 1000

ORAC as micromol of Trolox equivalent/g of dry matter

Health and Antioxidant Activity of Tea

The majority of the health benefits of tea are due to the antioxidant activity of flavonoids in tea. Therefore, early research on health benefits of tea consumption had mainly focused on establishing antioxidant activity of tea flavonoids. Antioxidant activity of tea flavonoids and tea brew had been demonstrated, both in vitro (outside a living organisms) and in vivo (inside a living organisms) in these early experiments. Later these studies have been extended to find the mechanisms through which antioxidant flavonoids in tea could reduce the risk of NCD’s.

Strong antioxidant activity of tea extracts and individual flavonoids in tea had been well established, in vitro, using Trolox Equivalent Antioxidant Capacity (TEAC), Oxygen Radical Absorbance Capacity (ORAC) and many other assay methods.10, 11 The next question that arises, is whether these tea flavonoids are absorbed from the digestive tract and whether they could act as antioxidants in vivo too. Bio-availability (whether a compound is absorbed from the digestive tract and reaches the blood stream to make it available) of plant flavonoids and especially tea flavonoids has been established in many studies.12

It has been found, that micro-flora living in the digestive tract of humans, aid the absorption of flavonoids by converting them into easily absorbable forms. 13 In vivo antioxidant activity also has been well established using both animal model and human studies.14, 15 Comparison of in vivo antioxidant potential of catechins (in green

tea) and theaflavins (in black tea) revealed that they are equally effective antioxidants.16 In vivo antioxidant activity of thearubigin fraction of black tea also has been established.17

Often milk is added to tea by many consumers in different parts of the world. Milk proteins and tea flavonoids are known to interact with each other.

• Reactive oxygen species (ROS) and free radicals (FR) are highly reactive molecules that could damage other molecules in the human body potentially leading to non-communicable diseases such as heart disease, stroke, cancer, diabetes and neurodegenerative diseases. • Presence of excessive amounts of ROS and FR is known as ‘Oxidative stress’. • Antioxidants derived from the diet play an important role in reducing the oxidative stress. • Tea contains very high amounts of antioxidant flavonoids that could contribute to reduce oxidative stress. • Regular tea consumption could have beneficial effects on human health through its antioxidant flavonoids.

Therefore, there was a doubt whether addition of milk would reduce the antioxidant activity of tea. It has been found that addition of milk does not alter the antioxidant activity and tea, with or without milk, increases the antioxidant activity of humans by similar amounts.18 Research has shown that in addition to scavenging FR and ROS, tea may exert antioxidant activity through other mechanisms too. Tea flavonoids, after entering the human body, have the ability to enhance the activity of antioxidant enzymes such as superoxide dismutase (SOD), and the enzymes in the glutathione system improving the natural antioxidant defenses of the body.19

There are certain other enzymes in the human body, called pro-oxidant enzymes, such as Nitric oxide synthase, lipoxygenase, and cyclooxygenase, whose activity could result in the generation of oxidant molecules. Tea flavonoids have the ability to inhibit such enzymes.20 Further, excessive amounts of free metal ions in blood and other body fluids would facilitate the activity of oxidant (FR and ROS) molecules in the body. Tea flavonoids could bind with free metal ions in blood and body fluids and thus contribute to the antioxidant activity in vivo. 21

Certain molecules in the human body act as messengers and play an important role in the biological processes that take place inside. Some of those messengers could promote pro-oxidant activity. Studies have found that tea flavonoids could inhibit some pro-oxidant messengers. Inhibition of nuclear factor κB (NF κB) being one example.22

REFERENCES

1. Cheng Guo, Xiaofen Li, Rong Wang, Jiekai Yu, Minfeng Ye, Lingna Mao, Suzhan Zhang, and Shu Zheng, 2016, Association between

Oxidative DNA Damage and Risk of Colorectal Cancer: Sensitive Determination of Urinary 8-Hydroxy-2’-deoxyguanosine by UPLC-

MS/MS Analysis., Science Reports, 6, 32581. 2. Nageswara R. Madamanchi, Aleksandr Vendrov, Marschall S. Runge, 2005, Oxidative Stress and Vascular Disease, Arteriosclerosis,

Thrombosis and Vascular Biology, 25, 29-38 3. Raynoo Thanan, Shinji Oikawa, Yusuke Hiraku, Shiho Ohnishi, Ning Ma, Somchai Pinlaor, Puangrat Yongvanit, Shosuke Kawanishi and

Mariko Murata, 2015, Oxidative Stress and Its Significant Roles in Neurodegenerative Diseases and Cancer, International Journal of

Molecular Sciences, 16(1), 193-217 4. Stadler, K. 2012, Oxidative stress in diabetes, Advances in Experimental Medicine and Biology, Springer International Publishing AG, pp 272-287 5. Esra Birben, Umit Murat Sahiner, Cansin Sackesen, Serpil Erzurum and Omer Kalayci, 2012, Oxidative Stress and Antioxidant Defense,

WAO Journal, 5, 9 - 19 6. Manthey J.A., Buslig B.S and Baker M.E. Eds. 2002, Flavonoids in cell function, Advances in Experimental medicine and Biology, Vol 505,

Kluwer Academic/Plenum Publishers, New York. 7. M.G.L Hertog, E.J.M Feskens, D Kromhout, P.C.H Hollman, M.B Katan, 1993, Dietary antioxidant flavonoids and risk of coronary heart disease: the Zutphen Elderly Study, The lancet, 342 (8878), 1007-1011 8. Ock Kyoung Chun, Sang Jin Chung and Won O. Song, 2007, Estimated Dietary Flavonoid Intake and Major Food Sources of U.S. Adults,

The Journal of Nutrition, 137, 1244 - 1252 9. Cao G, Sofic E and Prior R.L., 1996, Antioxidant capacity of tea and common vegetables. Journal of Agricultural and Food Chemistry, 44, 3426 - 3431 10. G. Paganga; N. Miller; C. A. Rice-Evans, 1999, The polyphenolic content of fruit and vegetables and their antioxidant activities. What does a serving constitute? Free Radical Research, 30, 153 - 162. 11. S. A. Wiseman; D. A. Balentine; B. Frei, 1997, Antioxidants in tea, Critical Reviews in Food Science and Nutrition, 37, 705 - 718. 12. P. C. H. Hollman; L. B. M. Tijburg; C. S. Yang, 1997, Bioavailability of flavonoids from tea, Critical Reviews in Food Science and Nutrition, 37 (8), 719 - 738. 13. M. R. Olthof; P. C. Hollman; M. N. Buijsman; J. M. van Amelsvoort; M. B. Katan, 2003, Chlorogenic acid, quercetin-3-rutinoside and black tea phenols are extensively metabolized in humans, Journal of Nutrition, 133 (6), 1806 - 1814. Erratum in Journal of Nutrition, 2003, 133 (8), 2692. 14. Balz Frei and Jane V. Higdon, 2003, Antioxidant Activity of Tea Polyphenols In Vivo: Evidence from Animal Studies, Journal of

Nutrition, 133: 3275S - 3284S. 15. Alexander Yashin1, Yakov Yashin, Boris Nemzer, 2011, Determination of Antioxidant Activity in Tea Extracts, and Their Total

Antioxidant Content, American Journal of Biomedical Science, 3(4), 322-335 16. Lai Kwok Leung, Yalun Su, Ruoyun Chen, Zesheng Zhang, Yu Huang and Zhen-Yu Chen, 2001, Theaflavins in Black Tea and Catechins in

Green Tea Are Equally Effective Antioxidants, Journal of Nutrition, 131, 2248 - 2251. 17. Yoshino K, Hara Y, Sano, M, Tomita I, 1994, Antioxidative effects of black tea thearubigins and theaflavins on lipid peroxidation of rat liver homogenates induced by tert-butyl hydroperoxide. Biological and Pharmaceutical Bulletin, 17(1), 146-149 18. R Leenen, AJC Roodenburg, LBM Tijburg and SA Wiseman, 2000, A single dose of tea with or without milk increases plasma antioxidant activity in humans, European Journal of Clinical Nutrition. 54, 87-92. 19. Yu-Li Lin, Chong-Yurn Cheng, Ya-Ping Lin, Yong-Wei Lau, I-Ming Juan, and Jen-Kun Lin, 1998, Hypolipidemic Effect of Green Tea Leaves through Induction of Antioxidant and Phase II Enzymes Including Superoxide Dismutase, Catalase, and Glutathione S-Transferase,

Journal of Agricultural and Food Chemistry, 46 (5), 1893–1899 20. Sarkar, A. and Bhaduri, A., 2001, Black tea is a powerful chemopreventor of reactive oxygen and nitrogen species: comparison with its individual catechin constituents and green tea. Biochemical and Biophysical Research Communications, 284, 173–178. 21. Brown, J. E., Khodr, H., Hider, R. C. and Rice-Evans, C. A., 1998, Structural dependence of flavonoid interactions with Cu2+ ions: implications for their antioxidant properties, Biochemical Journal, 330, 1173–1178. 22. Yang, C. S., Maliakal, P. and Meng, X., 2002, Inhibition of carcinogenesis by tea. Annual Review of Pharmacology and

Toxicology, 42, 25–54