3 Catechin C 4 Epicatechin EC 5 Epigallocatechin gallate EGCG 6 Gallocatechin gallate Materials

71

24. 3 Catechin C

trt N Subset 1 2 3 4 3, 200C 2 1.2450 3, 180C 2 1.3200 1.3200 3, 220C 2 1.3400 1.3400 6, 180C 2 1.3950 1.3950 6, 200C 2 1.4250 1.4250 1.4250 6, 220C 2 1.4800 1.4800 1.4800 9, 200C 2 1.5900 1.5900 9, 180C 2 1.5900 1.5900 9, 220C 2 1.6400 Sig. .059 .086 .075 .083 Means for groups in homogeneous subsets are displayed. Based on observed means. The error term is Mean SquareError = ,006.

24. 4 Epicatechin EC

trt N Subset 1 2 3, 200C 2 1.6550 6, 220C 2 1.7300 6, 180C 2 1.7400 6, 200C 2 1.7500 3, 180C 2 1.7650 3, 220C 2 1.8250 1.8250 9, 200C 2 1.8700 1.8700 9, 180C 2 1.9900 1.9900 9, 220C 2 2.3750 Sig. .229 .060 Means for groups in homogeneous subsets are displayed. Based on observed means. The error term is Mean SquareError = ,057. 72

24. 5 Epigallocatechin gallate EGCG

trt N Subset 1 2 3 6, 200C 2 4.6150 6, 220C 2 5.4250 5.4250 6, 180C 2 5.6150 5.6150 9, 220C 2 5.7100 5.7100 9, 180C 2 5.8900 5.8900 3, 180C 2 6.4900 6.4900 3, 200C 2 6.5200 6.5200 3, 220C 2 7.0150 9, 200C 2 7.0550 Sig. .069 .075 .059 Means for groups in homogeneous subsets are displayed. Based on observed means. The error term is Mean SquareError = ,247.

24. 6 Gallocatechin gallate

trt N Subset 1 9, 220C 2 1.4100 6, 200C 2 1.4500 9, 180C 2 1.4600 3, 200C 2 1.5300 9, 200C 2 1.5750 6, 180C 2 1.6000 3, 180C 2 1.6050 6, 220C 2 1.6100 3, 220C 2 1.6350 Sig. .126 Means for groups in homogeneous subsets are displayed. Based on observed means. The error term is Mean SquareError = ,015. 73

24. 7 Epicatechin gallate

trt N Subset 1 2 6, 200C 2 .7350 6, 180C 2 .8550 .8550 9, 180C 2 .9100 .9100 6, 220C 2 .9250 .9250 3, 200C 2 .9850 .9850 3, 180C 2 1.0150 1.0150 9, 220C 2 1.0500 3, 220C 2 1.0600 9, 200C 2 1.1000 Sig. .058 .090 Means for groups in homogeneous subsets are displayed. Based on observed means. The error term is Mean SquareError = ,014. 1

I. INTRODUCTION

A. BACKGROUND

Tea is globally one of the most popular and lowest cost beverages, next only to water. Tea is consumed by a wide range of age groups in all levels of society. The tea plant Camellia sinensis has been widely used for over 5000 years for its specific aroma, taste, and putative positive physiological functions. According to statistics from the Food and Agricultural Organization FAO of United Nations 2008, production and consumption of tea are steadily increasing. The worldwide production of tea in 2006 reached up to 3.60 million ton and the worldwide consumption reached up to 3.64 million ton. Over past decade, world tea consumption has increased by 2.7 annually. The main tea-producing countries are China, India, Sri Lanka, Kenya, Turkey, Indonesia, and Vietnam, which accounted for 28.73, 25.93, 8.60, 8.59, 5.49, 5.15, and 3.65, respectively, of the 2006 output of total global tea production Hicks, 2008. Freshly harvested tea leaves are processed differently to produce specific types of tea such as green, oolong, and black tea. Of all the tea consumed in the world, 78 is black tea, 20 is green tea, and 2 is oolong tea. The green tea consumption in Indonesia was 3.13 thousand tons in 2005, while black tea consumption was more than green tea consumption, 67.9 thousand tons in 2005. FAO projected that world green tea production would grow at a faster rate than black tea by 2.0 annually, to reach 1097.7 thousand tons by 2016 Ho et al, 2005. Green tea is heated and dried to avoid enzymatic oxidation. Green tea contains polyphenols, and most of the green tea polyphenols GTPs are flavonols, commonly known as catechins. Tea polyphenols have been known for their antioxidant activity and antimutagenic and anticarcinogenic properties Yang et al 2007. Traditionally, tea is prepared from its dried young leaves and leaf buds, made into a beverage by steeping the leaves in boiling water. But today, tea powder is being developed because it has many advantages such as more practical, simple transport economics, and simply to prolong product’s shelf-life. Tea powder could be applied as functional food and as non food product, like handbody, shampoo, and toothpaste. Food products being developed are tea-rice, tea-noodles, tea-cake, tea-biscuits, tea-wine, tea-candy, tea-ice cream Hicks 1998. There are several method for produce tea powder, one of them is spray drying method. Spray drying is one-step continuous processing operation that can transform feed from a fluid state into a dried form by spraying the feed into a hot drying medium Okos et al, 2007. A short processing time, usually between three and thirty seconds and controlled operational conditions make the spray drying as an effective and unique method for various products, especially heat sensitive products and its retaining the high quality properties such as color, flavor, and nutrients. The quality of a food powder is judged by the amount of physical and chemical degradation occuring during the dehydration process. There are many researchs were did by researchers to know the effect of spray drying on powder characteristic. At this research, different of feed concentrations and spray dryer inlet air temperatures were used to evaluate its effects on physicochemical properties of the spray-dried green tea extract. This information 2 however is necessary to establish processing conditions to produce value-added powder green tea as there is an increasing demand for herbal tea products in the market.

B. OBJECTIVES

The objectives of this research were to investigate the effect of total solid concentration in feed and inlet air temperatures on physical and chemical properties of green tea powder which was produced by spray dryer. 3

II. LITERATURE REVIEW

A. TEA

Tea, one of the most popular beverages consumed worldwide, is a processed product from the leaves of tea plant Camellia sinensis. The taxonomy of tea is shown in Table 1 and the figure of tea plant shown in Figure 1. Table 1. The classify the taxonomy of tea Common name: Tea Kingdom: Plantae Division: Spermatophyta Subdivision: Angiospermae Class: Dicolyledone Ordo: Guttiferales Family: Theaceae Genus: Camellia Species: Camellia sinensis The worldwide production of tea in 2006 reached up to 3.60 million ton and the worldwide consumption reached up to 3.64 million ton. Over past decade, world tea consumption has increased by 2.7 annually. Tea Production of Indonesia in 2006 reached up to 187.9 thousand of tonnes. Hicks, 2008. Indonesia, a country with more than 222 million people, produces more than 150,000 tons of tea per year, exporting 80 of it, with the balance consumed by domestic people. The large population provides a ready workforce, as well as a promising market for tea consumption. Tea products are usually classified as white tea, green tea, oolong tea, and black tea, categorized by manufacturing process as shown in Figure 2. Figure 1. Tea plant Camellia sinensis 4 Tea is consumed in different parts of the world as white, green, black, or oolong tea. White and green tea are known as unfermented tea. The polyphenol oxidase enzyme of green tea is inactivated by steaming. Oolong tea is produced by withering and half fermenting the leaves. Thus oolong tea is called “semi-fermented tea”. Black tea is known as fermented tea because the leaves are fermented, allowing enzymic oxidation of the polyphenols. Processing tea differently results in variation of chemical component in tea Hara, 2000. The chemical component of tea are presented in Table 2. Table 2. Composition of green tea, black tea, infusion Compound Green Tea Black Tea Infusion Protein 15 15 Trace Amino Acids 4 4 3.5 Fiber 26 26 Other carbohydrates 7 7 4 Lipids 7 7 Trace Pigments 2 2 Trace Mineral 5 5 4.5 Phenolic compounds 30 5 4.5 Oxidixed phenolic compounds 25 4.5 Data refer to dry weight of tea leaves Chako et al, 2010 Fresh leaves Withering Dr ying A B Fermenting Rolling by tea roller, rotor vane or CTC Fresh leaves Withering Drying Fresh leaves Solar withering Pan firing Indoor withering and rolling Rolling Mass breaking Drying Fresh leaves Rolling Primary drying- rolling Steaming Secondary drying-rolling Final drying- rolling Drying C D Figure 2. The manufacturing process of tea: A white tea, B green tea, C oolong tea, and D black tea Hara, 2000 5 In process of green tea production, tea leaves are steamed immediately after harvesting and the enzymes are inactivated at the initial stage. Therefore, the composition of green tea is simple and similar to that in the fresh tea leaves. Green tea contains polyphenols, which include flavanols, flavandiols, flavonoids, and phenolic acids; these compounds may account for up to 30 of the dry weight. Most of the green tea polyphenols GTPs are flavonols, commonly known as catechins accounting for up to 30 of the dry weight of the leaves,. which are composed of eight kinds of catechins and their derivatives slightly deviates depending on the species of tea plant and the season of harvesting. There are eight major catechins in green tea: +-catechin C, --epicatechin EC, -- gallocatechin GC, --epigallocatechin EGC, --catechin gallate CG, --gallocatechin gallate GCG, --epicatechin gallate ECG, and --epigallocatechin gallate EGCG. The major Epigallocatechin gallate EGCG is the major component of the polyphenolic fraction of green tea, it makes up about 10-50 of total green tea catechins. EGCG is also most potent antioxidant of polyphenol type of tea, is at least 100 times more effective than vitamin C and 25 times more effective than vitamin E. The antioxidant activity increase in the following order: ECECGEGCEGCG Meterc et al, 2007. The percentage of major polyphenols in tea are shown in Table 3. Table 3. The percentage of major polyphenols in tea Compound Green Tea Oolong Tea Black Tea EC 0.74 – 1.00 0.21 – 0.33 – ECG 1.67 – 2.47 0.99 – 1.66 0.29 – 0.42 EGC 2.60 – 3.36 0.92 – 1.08 – EGCG 7.00 – 7.53 2.93 – 3.75 0.39 – 0.60 Yamanishi, 1995 Caffeine 1,3,7-trimetylxantine, C8H10N4O2 is a plant alkaloid, one of the few plant products which the general public is readily familiar because of its occurence in beverages such as coffee and tea, as well as various soft drinks. Caffeine extract from tea is added to some pain- relief medicines. Caffeine compound is well known for its stimulant effect and is present at 2-4 of dried tea leaf weight, depending on the types and quality of teas Yoshida et al, 1999. Studies using animal models show that green tea catechins provide some protection against degenerative diseases. Green tea catechins could also act as antitumorigenic agents Roomi et al, 2007 and as immune modulators in immunodysfunction caused by transplanted tumors or by carcinogen treatment. Green tea consumption has also been linked to the prevention of many types of cancer, including lung colon, esophagus, mouth, stomach, small intestine, kidney, pancreas, and mammary glands Koo, 2004. This beneficial effect has been attributed to the presence of high amounts of polyphenols, which are potent antioxidants. In particular, green tea may lower blood pressure and thus reduce the risk of stroke and coronary heart disease. Some animal’s studies suggested that green tea might protect against the development of coronary heart disease by reducing blood glucose levels and body weight Tsuneki et al, 2004. 6 Figure 3. Structures of the major catechin and caffeine in tea Zuo et al, 2002 The stability of a functional ingredient is fundamental to elaborate a nutraceutical product ecause changes in the ingredient may affect its nutritional value e.g. antioxidant capacity, composition, and bioavailability. The stability of the grape seed extract GSE was evaluated based on changes in their main individual phenolic compounds, as well as changes in their antioxidant activity and browning. pH affects the stability of polyphenolic compounds and that a between 4 and 5 confers more stability to catechins and their isomers and polymers than more alkaline or acidic valuesTabart, 2009. The concentration of catechins was more stable than the concentration of the rest of compounds. According Pardo et al 2011, the catechins and antioxidant activity in grape seed extract generally showed decreases after the thermal. The -- Epicatechin gallate ECG -- Epigallocatechin EGC -- Epigallocatechin gallate EGCG -- Catechin gallate CG -- Catechin C -- Epicatechin EC Caffeine CAF 7 decrease in ECG and EGCG may have affected the antioxidant activity of the extracts because these phenolic compounds are known to have more scavenging power than the flavan-3-ols that clearly increased: gallic acid, gallocatequin, and catechin, due to their stearic conformation and the presence of the gallate group joined to the C ring treatments,but they were not always significant p0.05.

B. FOOD POWDER

A major reason for production in powder form is simply to prolong shelf-life of the ingredient by reducing water content; otherwise the ingredient would be degraded in its natural biological environment. Another important reason is simple transport economics, because reducing water content reduces mass and costs of the ingredient to be transported Gustavo et al, 2010. Food ingredient powders must possess a number of functionalities which can be broadly classified as: powder handling capability; reconstitution recombination ability and ingredient functionality in the food product to be consumed. Poor handling during manufacture, storage and transport causes many problems which are quite common, such as no or irregular flow out of hoppers and silos and problems associated with stickiness and caking of powders. Production and processing will determine the properties of particles and powder, such as particle size distribution, shape, surface properties and moisture content. They will also influence ingredient functionality, for example, higher temperatures may cause denaturation of proteins and coating may prevent the ingredient functionality from being destroyed by oxidation Aguilera et al, 2008. It is well known that ingredient functionality in powder form may degrade over time between manufacture and final application. This depends on the sensitivity of the individual ingredient and its exposure over time to temperature, moisture and oxygen in the air. Some ingredients are encapsulated and some powders are coated in an effort to prevent its degradation and protect its functionality outlinedsome of the functional properties of food powders and particulates Lillford, 2002. Powders are important ingredients in a large variety of food formulations and they are responsible for the development important product characteristics such as texture, flavour, colour and nutritional value. Most of the powders will be used in some sort of wet formulation and therefore their functionality will depend on their interaction with water. Because the influence of drying parameters is not the same for all materials, optimal drying conditions vary depending on the final objective: volatile retention, preservation of enzymatic activity and avoidance of protein denaturation, fat oxidation or crystallisation. Usually, the resulting powder is made of dry particles with an average size of 30 microns and mean water activity around 0.2. The powder outlet temperature is typically less than 100°C and the residence time is of seconds Huntington, 2004. All these characteristics will have some effect on handling properties of powders such as: bulk and tapped densities, particle density, mixing with other powders, storage; wettability and solubility, porosity, specific area rehydration, instantisation; flowability size, surface asperities, friability and creationexistence of dust, stability in specific atmosphere and medium oxidation, humidification, active component release Huntington, 2004. Study on quality evaluation instant green tea powder showed that the important quality attributes for a green tea sample was rated as taste flavor color strength. Among the quality 8 attributes, taste was the strongest attribute for both instant tea and green tea granules produced, and strength was the weakest attribute. Sinija, 2011.

C. FREEZE CONCENTRATION

Concentration of fluid foods by freezing involves lowering the temperature of the product in a sufficiently controlled manner to partially freeze the product, resulting in a slurry of ice crystals in a fluid concentrate. If formed under the appropriate conditions, these ice crystals will be very pure. That is, very little product will be incorporated within the ice crystals. The ice crystals are then removed in some way with a minimum of liquid carryover, resulting in a concentrated product. The basic components of a freeze concentration system, as shown in Figure 4. Freeze concentration is appliable to many food concentration, such as citrus fruit juices, vinegar, coffee, tea, sugar syrups, dairy product, and aroma extract. The major advantage of using a freeze concentration process as opposed to evaporation or reverse osmosis are related to the low temperature operation suitable for sensitive food products without the loss of product quality. In addition, the solid-liquid separation in freeze concentration results in no losses of the more volatile flavors and aromas, as occur in evaporation. The disadvantages of freeze concentration compared to evaporation and reverse osmosis have include higher capital cost, higher operating costs, and excessive loss of product during the ice separation Hartel, 1992.

D. SPRAY DRYING

Spray drying is one-step continuous processing operation that can transform feed from a fluid state into a dried form by spraying the feed into a hot drying medium. The product can be a single particle or agglomerates. The feed can be a solution, paste, or a suspension. This process has become one of the most important methods for drying liquid foods to powder form. The principal of spray drying as shown in Figure 5. Feed Crystal Nucleation Crystal Growth Separation Crystal Slurry Concentrate Ice Figure 4. Schematic of a basic freeze-concentration process Hartel, 1992 9 The main advantages of spray drying are the following: • Product properties and quality are more effectively controlled • Heat-sensitive foods, biologic products, and pharmaceuticals can be dried at atmospheric pressure and low temperatures. Sometimes inert atmosphere is employed. • Spray drying permits high tonnage production in continuous operation and relatively simple equipment • The product comes into contact with the equipment surfaces in an anhydrous condition, thus simplifying corrosion problems and selection of material of construction • Spray drying produces relatively uniform, spherical particles with nearly the same proportion of nonvolatile compounds as in the liqiud feed. The principal disadvantages of spray drying are as follows: • Spray drying generally fails if a high bulk density product is required • In general it is not flexible. A unit designed for fine atomization may not be able to produce a coarse product, and vice versa. • For given capacity, evaporation rates larger than other types of dryers are generally required due to high liquid content requirement. The feed must be pumpable. Pumping power requirement is high • There is a high initial investment compared to other types of continuous dryers. • Product recovery and dust collection increases the cost of drying Xin and Mujumdar, 2010 Spray drying consist of four process stages: 1. Atomization of feed into a spray The formation of spray and the contacting of the spray with air, are the characteristic features of spray drying. The selection and operation of the atomizer is of supreme importance in achieving economic production of top quality products. The selection of the atomizer type depend upon the nature of the feed and desire characteristics of the dried product. In all atomizer types, increased amounts of energy available for liquid atomization result in sprays having samLler droplet sizes. If, the available atomization energy is held constant but the feed rate is increased, sprays having Figure 5. Spray Dryer. 1, feed reservoir; 2, feed pump; 3, product feed pipeline; 4, atomizer; 5, drying chamber; 6, air fan; 7, air heater; 8, hot air duct; 9, a mixture of dried product and air- carrying duct; 10, cyclone separator; 11, heavy powder falling down; 12, product tank; 13,exhaust air Sharma et al, 2000. 10 larger droplet sizes will result. Rotary atomizers are used to produce a fine to medium coarse product mean size 30-130 µm, while nozzle atomizers are used to produce a coarse product mean size 120-250 µm. 2. Spray-air contact mixing and flow Product and air pass through the dryer in co-current flow, they pass through the dryer in the same direction. This arrangement is widely used, especially if heat-sensitive products are involved. Spray evaporation is rapid, the drying air cools accordingly, and evaporation times are short. The product is not subject to heat degradation. 3. Drying of spray moisture volatiles evaporation As soon as droplet of the spray come into contact with the drying air, evaporation takes place from saturated vapour film which is quickly established at the droplet surface. The temperature at the droplet surface approximate to the wet-bulb temperature of the drying air. A substantial part of the droplet evaporation takes place when the droplet surfaces are saturated and cool. Drying chamber design and air flow rate provide a droplet residence time in the chamber, so that the desired droplet moisture removal is completed and product removed from dryer before product temperatures can rise to the outlet drying air temperature of the chamber. Hence, there is little likehood of heat damage to the product. 4. Separation of dried product from the air Total recovery of dried product takes place in the separation equipment. This system places great importance on the separation efficiency of the equipment. Separation of dried product from the air influences powder properties by virtue of mechanical handling involved during the separation stage. Axcessive mechanical handling can produce powders having a high percentage of fines. Master, 1991 There are many variables in spray dryer that give an effect on powder product, such as inlet temperature, feed solid content, drying temperature difference, and feed temperature. Increase of inlet temperature can decrease the heat requirement of the dryer for producing a given product rate because product dried quickly. Increase in feed solids for a given production rate from 50 to 60 reduces the heat load by nearly 50. Spray drying is an expensive method of evaporating volatiles and thus to obtain optimum heat utilization condition the spray dryer should always fed with the maximum solids feedstock possible. The higher the temperature difference ie. Inlet drying air temperature minus outlet drying air temperature, the lower the heat requirement to produce a unit weight of product of constant residual moisture content from a constant solid feedstock. Feed temperature, particularly in existing plants, can also be optimized. Increasing feed temperature reduces the heat required to produce a unit weight of dried product. Preheating of feed is normally carried out to reduce feed viscosities, thereby improving atomization performance and to present feed crystallization that can cause atomizer blokage Master, 1991. 11

III. RESEARCH METHODOLOGY

A. MATERIALS AND INSTRUMENTS

1. Materials

Dried green tea var. Oolong No 12 was supplied by Boonrod Tea Factory Thailand. Chemical reagents with analytical grade such as folin-ciocalteu 10 vv and gallic acid were supplied by Fluka Buchs, Switzerland, anhydrous sodium carbonate and potassium hexacyanoferrate [K3FeCN6] were purchased from Merck Darmstadt, Germany, standard HPLC of caffeine and catechins were purchased from Sigma-Aldrich St. Louis, Missouri, USA, acetonitrile, trifluoroacetic acid TFA and methanol HPLC-grade were purchased from Fluka Buchs, Switzerland, trolox ±-6-Hydroxy-2,5,7,8- tetramethylchromane-2-carboxylic acid and DPPH 2,2-diphenyl-1-picryhydrazyl were purchased from Aldrich Steinheim, Germany. monosodium phosphate monohydrate, Disodium phosphate heptahydrate and trichloroacetic acid TFA were purchased from Fluka Buchs, Switzerland, citric acid food grade, potassium mitrute salt food grade. Then, distilled water, filter paper No 1 and No 4.

2. Instruments