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Tea plant was characterized as an evergreen shrub. It can bloom yellow-white flowers and long elliptic, serrate, coriaceous leaves. Flowers are axillary , solitary , or up to three in a cluster, the small little flowers have six to eight petals. Outer petals were spoiled and inner petals are obviate to broadly obovate. Young leaves have fine white hairs on their underside, terminal buds are silvery gray and sericeous. The young leaves of C. Sickness generally sprouts starting from March thought October followed by blooming of flowers till February and fruiting occurs from August. (Chang and Bartolomew, 1984) .
The domestication process of C. Sinensis changed it Phytochemical and genetic characteristic depending on the interaction between Artificial selection and environment factors. Some cases , cultivated tea plants have decreased heterozygosity compare to their nearest relatives ( Kaundun and Matsumoto , 2003), whereas other cultivated C. Sinensis var Assamica population has heterozygosity under selection on the basis of their increased leaf area , plucking shoots weight, and caffeine and polyphenolic content compared to the rest of the cultivated tea plants and their wild relatives ( Chen et.al, 2005, Ahmed 2011).
The soft seedling develops in a nursery for ten months, which protect them from harsh climate conditions. After they have strengthened, the tea plant continues to develop in open fields that are sheltered by the wide tree shade.
Plucking of tea leaves is still conducted using handpicked method, this is to ensure that the quality of the tea collected is the best. Usually this tea picking season in Asia country starts with the beginning of spring and continues from May till August,
They are various types of tea leaves which are commonly grouped into 4 groups such as Black tea, Green tea , Oolong tea , and White tea. All of the varieties were collected from the same tea plant. In fact it was the tea processing procedure which differentiated them. The Fresh tea leaves are filled with 80% of liquid and it dried till less than 10% before it can be readily sold to the consumers. Hence drying is an important stage in the tea process, because reducing the concentration of the leaves and thereby strengthening and preserving the tea leaf.
( Citation : http://www.wtea.com/about-tea_growth.aspx , Wissotzky Tea 2007.)
2.1.1 Fresh tea flush
Young shoots of the tea plant are generally known as tea flush that consist if the terminal bud with adjacent leaves. In the fresh tea flush it contains high values of non-volatile compounds such as polyphenols, flavanols, phenolic acid , amino acids, chlorophyll, other pigments and nutrient. The chemical composition of the tea leaves depends upon the leafage, the clone that being examined, soil and climate conditions are agronomic practices.
The total polyphenol in the tea flush is between 20-30 %, and divided mainly into six groups of compounds. Among them catechins have the highest content, there are 4 types of catechins such as (-) -epigallocatechin-3-gallate (EGCG), (-) -epigallocatechin (EGC), (-) -epicatechin-3-gallate (ECG), (-) -epicatechin (EC), (+) -gallocatechin (GC), and (+) -catechin (C). It is widely investigating for their bioactivities and utilization.
(Citation : Yong -Su Zhen(edi), 2002, Tea, Bioactivity and its therapeutic Potential , )
Caffeine, theobromine, and theophylline, the principal alkaloids, account for about 4% of the dry weight. . A cup (200 ml) of green tea (Gun Powder, Hangzhou, China) contains about 142 mg EGCG, 65 mg EGC, 28 mg ECG, 17 mg EC, and 76 mg caffeine.
The soluble part of leaf includes about thirty polyphenolic bodies, over twenty amino acid, a dozen sugar and half a dozen organic acids. The total ash content of the tea leaf is about 5.5 percents including both soluble and insoluble portions. Polyphenol comprises about 30 percents of solid matter in the tea set. Most of it belongs to a group named flavanols, colorless plant pigments that readily turns brown
( Source : Belizt el at , 2009 ).
Â Teas are a major source of flavan-3-ols in the diet. The flavan-3-ol subclasses are ranked by degree of polymerization. The catechins are monomers (catechin, epicatechin, epicatechin gallate, epigallocatechin, and epigallocatechin gallate), the theaflavins are dimers (theaflavin, theaflavin 3-gallate, theaflavin 3'-gallate, theaflavin 3,3'digallate), and the derived tannins. These compounds are theorized to be responsible for the positive health effects of tea. As levels of fermentation increased from green to oolong to black tea, the major flavan-3-ol profiles changed. Total catechins were 13.6 g/100 g in dry weight green tea and 4.2 g/100 g dry weight in black tea.
(Citation : , Major Flavonoids in Dry Tea, USDA , Researcher from TURF University )
Tea trend around the world , Asia and Asean
Tea has rise in popularity in society especially among younger generation. Some of the most recent in tea trends is using tea as topping or ingredient in daily favorite food and drinks. Various product made up of tea has been seen widely from mixed drink, ice-cream, facial product and cosmetic.
Another tea trend is that of serving loose leaf tea rather than the standard tea bags. Many restaurant and individual are looking into purchasing loosing leaf tea for brewing. Many times, loose leaf tea will unfold once submerged in wateradn produce a pleasant asthetic sight.
Tea drink in the market had widely developed, other than plain tea, the introduction of various flower , herbs and even fruit in tea made it become flavored , scented and herbal tea , greatly increases popularity among teenagers. Now young generation prefer drinking more tea product than soda and sugar product.
For many years, " Teh Tarik" was popular among Malaysian and had been a symbolism drink for tourist milk was very popular. Now with all types of teas,Ice tea is another tea trend that can be enjoyed in many flavors. This is and excellent way for tea drinkers to enjoy a nice cool drink on a hot summer day. They maybe available in powder forms , as well as readily drink bottle version.
With the changing society of today's world, tea trend are constantly changing, They are also very different in different parts of the world. However tea trend comes about it is increasily becoming one of the hottest drink
Below was a statistical analysis conducted on the 7 countries that are the covers almost 80% of the world tea production. As can be seen below, the growth rate is quite impressive and almost constant.
The world production averaged in these 47 years to nearly 2.2 millions of tonnes per year with only 983,825 tons being produced in 1961 against nearly 3,896 millions in 2008.
As illustrated, we can see that China remain the biggest tea producing countries from 3.3 % to 39%. While india shares drop starting from 2006 from 36% to 20.7 %.
1.2 Black tea
Tea leaves are coming from the same plant just that the processing procedures in tea processing were different. Black tea , commonly known as fully fermented tea had covered about 80% of the tea markets. It was developed in the mid-17th century in Chongan Country, Fujian Province of China. The first black tea produce is named as Xiao Zhong Black tea.
Once the fresh tea leaves picked, they are left to wither for several hours under the sun or drying the leaves is by blowing currents of warm air over them. This step is essential for the end quality of the tea water is released to allow the fermentation and oxidation of the leaf to begin. Rolling step comes next when the moisture content is reduced to about 60 %. There are few types of rolling techniques such as orthodox roller or rotor vane, or CTC ( crushing, tearing, and curling) machine. This step is crucial to break the cell and release the enzyme , included polyphenol oxidase and peroxidase, and initiate the fermentation process with the introduction of oxygen. The smaller the tea leaf is crushed the wider the surface exposed. The four cathechins are enzymatically oxidize and undefgoes complex chemical changes.
After the leaves are rolled by hand or machine a unique compound is formed between the enzymes and the polyphenols. The fermentation process can then take place.Â Fermentation conditions (time, temperature, and moisture) determine the final flavor and aromas of black teas. The last step consists of drying of the tea leaves on the drying beds with forced hot air between 200F-250F (93.3-121 Celsius) flowing around it. When the leaves are about 80 percent dried, theyÂ complete their drying over wood fires.
The resulting product is reddish-brown in color and is sorted according to size. The sorted largerÂ tea leaveÂ is considered theÂ "leaf grade" and the smaller one asÂ "broken grade". These two gradesÂ are sold as loose-leaf teas. The "fanning" or "dust" grade isÂ usually used in tea bags.Â The final product will have a moisture content of about 4%.Â
When the fermentation occurs, the monomeric flavan-3-ols undergo polyphenol oxidase-dependent oxidative polymerization leading to the formation of bi-flavanols, thea-flavins, thearubigins, and other oligomers in a process commonly known as "oxidation." Theaflavins (about 1%-2% of the total dry matter of black tea), including theaflavin, theaflavin-3-O-gallate, theaflavin-3'-O-gallate, and theaflavin-3,3'-O-digallate, possess benzotropolone rings with dihydroxy or trihydroxy substitution systems, which give the characteristic color, and taste of black tea. About 10%-20% of the dry weight of black tea is due to thearubigens, which are even more extensive oxidized and polymerized, have a wide range of molecular weights, and are less well characterized. (Chung S. Yang, Zhi-Yuan Wang )
An approximate of the black tea beverage mean percentages of components of solid extracts are: catechins (10-12%), theaflavins (3-6%), thearubigins (12-18%), flavonols (6-8%), phenolic acids and depsides (10-12%), amino acids (13- 15%), methylxanthines (8-11%), carbohydrates (15%), proteins (1%), mineral matter (10%), and volatiles (<0.1%) (Graham, 1992)
Figure 2.1.2 Oxidation of individual catechin
2.2 Antioxidant activity
2.2.1 Antioxidant properties and mechanism of the tea polyphenol
Antioxidants act as protection against the damaging effects of free radical produces naturally in body. The globalization of the world, development of science had bring benefit to us, yet there are also adversed effects such as environemt pollution, smoking or physiological disorder which overhelmed the function of the antioxidants, interrupt the It's system in our body.Free radical production cause damage to proteins and DNA within the cells.
Recent studies on tea suggest that the comsupmtion of tea can help to prevent tumor formation in humans, This is because tea leaf contains abundant group ot polyphenols. Polyphenols also suggested to have antimutagenic and anticarcinigenic properties.It also have beneficiall protective properties against cholesterol (Maron and other,2003)
In fresh tea, major matabolites in the tea leaf includes polyphenoloc catechins and theaflavins, and alkaloids caffeine and theobromine catechins cover 80% of the total polyphenols in the tea leaf Inactivation of phenol oxidase in green tea orevent oxidations of the catechins, whereas enzyme phenolase-catalyzed the formation of dimeric theaflavins and polymeric thearubigin, which differentiate the black color to black tea ( Schwimmer 1981: Shahidi and Narczk 2004).
In this experiment there are 7 types of standard are comparable, which belongs to Catechins ( Catechin , Epicatechin-3-gallate, Epicatechin , Epigallocatechin, ) and Theaflavin , Theobromine, Gallic acid,and Caffeine.
Catechin is a polyphenolic flavonoid which has been isolated from a variety of natural sources including tea leaves, chocolates, apples, and the wood and bark of trees such as acacia. It was building blocks for procyanidins or known as condensed tannins. Tea catechins exist as two geometrical isomers: Trans-catechins and cis- catechins. Depending on the stereochemicla confriguration of the 3', 4' - dihydroxyphenyl and hydroxyl group at the 2'- and 3'- position of the C- ring, Each of the isomers can also exist as two optical isomers (+) or (-) catechins.(Friedman,2005). Cathechin from enzymatic oxidation will forms theaflavin and which further oxidized to thearubigins There are many kinds of catechins in this study we only study the following compound :
Epicatechin , EC
EGCG is the most abundance catechin, which account for 65%of the total catechins in green tea. However black tea catechins was undergoes that undergoes fermentation process, had convert catechins into polymer such as theaflavin and thearubigins. The level of catechins decrease from 35%-50% in fresh tea leaf to 10% in black tea. Below was the comparision of the total catechins in green tea and black tea.
Type of tea
Brewed black tea
Brewed Green tea
(Citaion : Karrie Heneman, PhD, Sheri Zidenberg-Cherr,Nutrition and Health info sheet, PhD UC Cooperative Extension Center for Health andNutrition Research )
Function of catechins
Catechinsis a more potent antioxidant than ascorbate or Î±-tocopherol in certainÂ in vitroÂ assays of lipid peroxidation.Â Catechin inhibits the free radical-induced oxidation of isolated LDL by AAPH.Â It has also been reported to act as an inhibitor of COX-1 with an IC50Â of about 80 ÂµM.3Â Catechin and other related procyanidin compounds have antitumor activity when tested in a two-stage mouse epidermal carcinoma model employing topical application.
Theaflavin is a chemical made up ofÂ polyphenols. Commonly found inÂ black tea, the plant-produced chemical has been clinically proven to prevent the accumulation of free radicals in the body. As a result, theaflavin is a popular compound in cancer research, as well as in other disease studies and treatments
WhenÂ green teaÂ is fermented to become black tea, three different theaflavins are created. CalledÂ theaflavin-3-gallate, theaflavin-3'-gallant and theaflavin-3,3'-digallate, they can also be found inÂ Oolong tea. The presence of these antioxidants alters both the hue as well as the flavor of the tea.
Theaflavins have been useful in slowing the growth of human immunodeficiency virus, or HIV, cells, which could help scientists discover a cure for the disease. Some studies suggest that regular consumption of theaflavin may help lower one's levels of low-density lipoprotein, or LDL,Â cholesterol. People who regularly drank theaflavins for 90 days in one study were able to lower their cholesterol by nearly 12 percent. Levels of LDL cholesterol specifically were reduced by 16 percent. Participants in the study were also able to raise their levels of high-density lipoprotein, orÂ HDLÂ cholesterol that the body needs, slightly.
Several other health benefits may result from the tea antioxidants. An antiviral agent, theaflavin may help lower blood sugar levels. One study shows that it may help enhance the functions of the liver, as well as reduce the amount of lipids and body weight of an organism. Most of these possible benefits remain under review by the scientific community. Many physicians also point out that black tea consumed in conjunction with a low-fat diet yields the most health benefits.
To drink the benefits of theaflavins, five cups of black tea are generally recommended daily. While this remedy is considered safe for most people, women who are pregnant or nursing should avoid it.
( Citation : Ness AR, Powles JW. Fruit and vegetables, and cardiovascular disease: a review. Int J Epidemiol1997;Â 26:1-3. )
Medical Researched on Tea
According to research, black tea was found to lower blood pressure, thereby reducing the risk of heart disease. It may also cut levels of "bad" LDL cholesterol and blood sugar. Again, given that heart disease is one of the major causes of death in industrialized countries, the regular cuppa is starting to look like an increasingly promising and unobtrusive solution to keep your health in check.
From a business point of view, tea manufacturers operate under two long-held beliefs: that the 30-50 year old female is the current primary customer and that conquering the young and hip generation of teens and twentysomethings is the only way to expand the category in the years to come. Both points are certainly valid, but they somewhat fail to reflect the economic and demographic realities of today. That is because the future, at least for the next couple of decades at least, actually belongs an entire population of baby-boomers that is currently entering retirement.Â
With vast amounts of wealth and legitimate concerns about ageing and health, this target should not be forgotten by tea businesses. Yes, that population may be not as gastronomically sophisticated or as creatively minded as others, yet no manufacturer or retailer can afford to neglect its impact on the bottom line, especially given that tea is so relevant to its health concerns. Indeed, most of the prominent health benefits of tea, such as reducing the incidence of cardiovascular diseases, cancer, Alzheimer's and obesity, to name but a few, are obviously more relevant to a baby boomer than a college student. Add to that increased health and medical awareness, more free time to enjoy meals (and brew proper tea), as well as the spending power that the elderly possess and you've got an offer difficult to refuse.
Viewed in this light, these medical studies are more than just about tentative scientific findings, because they may point to a somewhat new direction for the entire tea industry. Despite limitations on health claims in marketing, they offer new ways of thinking about the relevance and potential of a population that goes unnoticed far too often.
2.2.2 Test on antioxidant activity
22.214.171.124 Test of Total Polyphenol Content
Folin - Ciocalteu Assay
Folin & Ciocalteu's phenol or can be called theÂ Gallic AcidÂ Equivalence methodÂ (GAE), is a mixture ofÂ phosphomolybdateÂ andÂ phosphotungstateÂ used for the colorimetricÂ assayÂ of phenolic andÂ polyphenolic antioxidants. Â It is named afterÂ Otto Folin,Â VintilÄƒ Ciocâlteu, andÂ Willey Glover Denis.
The mixture of the phosphomolydate and phosphotungstate with acid and reflux will produce complexes with structure in the solution.
Folin & Ciocalteu's phenol reagent does not contain phenol. Rather, the reagent will react with phenols and non - phenolic reducing substances to form chromogens that can be detected spectrophotometric technique. The color development is due to the transfer of electrons on the basic pH to reduce the phosphomolybdic /phosphotungstic acid complexes to form chromogens in which the metals have lower valence.
The most common usage of this reagent is in the Lowry method for determining protein concentration.
In this method, protein is pre - treated with copper (II) in a modified biuret reagent (alkaline copper solution stabilized with sodium potassium tartrate). Folin & Ciocalteu's phenol reagent generates chromogens that give an increasing absorbance between 550 NM and 750 NM. Normally, absorbance at the peak (750 nm) or shoulder (660 nm) are used to quantify protein concentrations between 1-100 Âµg/ml while the absorbance at 550 nmis used to quantitate higher protein concentrations. In the absence of copper, color intensity
would be determined primarily by the tyrosine and tryptophan content of the protein, and to a lesser extent by cysteine, and histidine. Copper (II) enhances color formation by chelation with the peptide backbone, thus facilitating the transfer of electrons to the chromogens. Copper (II) has no effect on color formation by tyrosine, tryptophan, or histidine, but reduces that due to cysteine.
To control for the effect of these compounds on color development and, thus, on the calculated protein concentration of the sample, it is essential that the blank and standards be made up in the same medium as the samples.
(Citation : Fluka Analytical ,Sigma-Aldrich Chemie GmbH Â·)
Reversed Phase HPLC
Reversed phase chromatography has found both analytical and preparative applications in the area of biochemical separation and purification. Molecules that possess some degree of hydrophobic character, such as proteins, peptides and nucleic acids, can be separated by reversed phase chromatography with excellent recovery and resolution. Separation mechanism in reverse phase chromatography depends on the hydrophobic binding interaction between the solute molecules in the mobile phase and was immobilized hydrophobic ligand, (stationary phase ).