Effects of Inlet Temperature and Feed Concentration on Quality of Spray Dried Instant Green Tea Powder

EFFECTS OF INLET TEMPERATURE AND FEED

CONCENTRATION ON THE QUALITY OF SPRAY DRIED

INSTANT GREEN TEA POWDER

SKRIPSI

PRADHINI DIGDOYO

F24080005

FACULTY OF AGRICULTURAL ENGINEERING AND

TECHNOLOGY

BOGOR AGRICULTURAL UNIVERSITY

BOGOR

2012

ii

EFFECTS OF INLET TEMPERATURE AND FEED CONCENTRATION ON

QUALITY OF SPRAY DRIED INSTANT GREEN TEA POWDER

Pradhini Digdoyo1, Adil basuki Ahza1, Natthawuddi Donlao2, Puwanart Fuggate2

1_{Departement of Food Science and Technology, Faculty of Agricultural Engineering and}

Technology, Bogor Agricultural University, IPB Darmaga Campus, PO. BOX 220, Bogor, West Java, Indonesia

2_{School of Agro-Industry, Mae Fah Luang University, Muang, Chiang Rai 57100, Thailand}

ABSTRACT

Tea is a popular drink because of its unique aroma and characteristic flavor which is controlled by key chemical components which are volatile terpenes, teaine, organic acids, and polyphenols. Currently, the quality ofinstant green tea powder has been improved. It has some advantages like increase solubility, shelf life, and flavor. Instant green tea powder is produced trough drying method like spray drying.This process, unfortunately, potentially has deleterious effects on some component in green tea, like polyphenol compounds and volatile compounds.

The objective of this research is to study effect of inlet air temperature and feed concentration in feed on quality of green tea powder. Main material used in this research was green tea leaf which was supplied by Boonrod Tea Factory. The drying method used in this research was spray drying. There are three level of inlet temperature used in this research, which were 180 oC, 200 oC, and 220 oC. This research also used three level of feed concentration which were 3%, 6%, and 9%. Parameters studied in this research were volatile compounds, chemical compound, and physical properties of green tea powder.

The result shows that inlet temperature and feed concentration significantly affected (p<0.05) the quality of green tea powder. Generally, increased temperature (up to 200 oC inlet temperature) decreases amount and type of volatile compound, particularly ketone, total polyphenol, teaine content, solubility, and bulk density (p<0.05). On the other hand, increased feed concentration increases L value, b value, gallic acid, teaine, and catechin content (p<0.05). The best green tea powder is the one with 6% total solid in feed and 180 0C inlet temperature. It has highest amount of terpene and also contains hydrocarbon and aldehide.It also has low moisture content (4.17 ± 0.02 %), high total polyphenol content (26.42 ± 0.07 %), high teaine content (7.06 ± 0.01 %), and highest catechin content (26.16 ± 0.13 %). Physical properties analysis shows it has low water activity (0.21 ± 0.02), lowest a value (3.80 ± 0.35) low b value (18.87 ± 0.14) high hygroscopicity (9.09 ± 0.94 %) high solubility (98.79 ± 0.53 %) and low bulk density (0.51 ± 0.18 g / mL).

iii

Pradhini Digdoyo. F24080005. Effects of Inlet Temperature and Feed Concentration on Quality of Spray Dried Instant Green Tea Powder. Supervised by Adil BasukiAhza, Natthawuddhi Donlao, Puwanart Fuggate 2012.

SUMMARY

Tea has been consumed worldwide for years and one of the most popular drinks. Tea is popular because of its characteristic flavor which is controlled by some key chemical components like volatile terpenes, teaine, organic acids, and polyphenols. Recently, popularity of tea has increased due to its potential health benefitsmostly from activities of antioxidant flavonoids present in tea. As the development of technology, instant green tea powder has been produced because it has some advantages, such as prolong shelf-life, reduce transportation and storage cost, also more practical. The objectives of this research is to study effect of inlet temperature and feed concentration on quality of instant green tea powder which was produced by spray dryer.

In the preliminary research, chemical and volatile compounds of raw materials include were analyzed. In experiment I, production of concentrated green tea was done through freeze concentration process using freeze concentrating machine. There were three feed concentration used in this research, which were 3%, 6%, and 9%.In experiment II, production of green tea powder was done through spray drying process using JMC-minilab spray dryer. There were three inlet temperature used, which were 180 o_{C, 200} o_{C, and 220} o_{C. The instan green tea powder were}

then analyzed for its volatile compound, physical properties (such as water activity, bulk density, color, solubility, and hygroscopicity), and chemical compounds (such as moisture content, total polyphenol content, gallic acid, catechins, and teaine contain).

The result of chemical compound analysis of raw material shows that green tea leaf has moisture content 6.91± 0.02 %, total polyphenol content 28.80 ± 0.18 (g/100 g db), gallic acid 0.45 ± 0.02 (g/100 g db), teaine 1.21 ± 0.02 (g/100 g db), and total catechin 3.72 ± 0.0 8(g/100 g db). Volatile compound analysis shows that green tea leaf contains ketones, aldehide, alcohol, and terpenes. Spray drying process caused change in volatile and chemical compounds of green tea. Inlet temperature and feed concentration significantly affected quality of green tea powder.

Analysis of physical characteristic of green tea powder shows that green tea powder have water activity in range of 0.18 ± 0.01 to 0.26 ± 0.00, L value in range of 56.48 ± 0.33 to 64.11 ± 0.07, a value in range of 3.80 ± 0.35 to 4.22 ± 0.01, b value in range of 18.40 ± 0.06 to 20.39 ± 0.63, solubility in range of 95.77 ± 0.47% to 99.54 ± 0.29%, hygroscopicity in range of 7.24 ± 0.97% to 12.33 ± 0.58%, and bulk density in range of 0.43 ± 0.00 g/mL to 0.60 ± 0.01 g/mL. Analysis of chemical compound of green tea powder shows that green tea powder have moisture content in range of 4.04 ± 0.01% to 4.41 ± 0.02%, total polyphenol content in range of 20.65 ± 0.01 g/100 g to 31.22 ± 0.11 g/ 100 g, gallic acid content in range of 2.21 ± 0.10g/ 100 g to 2.68 ± 0.07g/ 100 g, teaine content in range of 6.20 ± 0.01g/ 100 g to 7.41 ± 0.00 g/ 100 g, and total catechin in range of 15.17 ± 0.02 g/ 100 g to 26.20 ± 0.04 g/ 100 g. Analysis of volatile compound shows that green tea powder contain alcohol, ketone, hydrocarbon, acid, aldehide, azole, and terpene.

Generally, increased temperature(up to 200 oC inlet temperature) decreases amount and type of volatile compound, particularly ketone, total polyphenol, teaine content, solubility, and

iv

bulk density. On the other hand, increasing feed concentration increases L value, b value, gallic acid, teaine, and catechin content. The best green tea powder is the one with 6% feed concentration and 180 0_{C inlet temperature. It has the highest amount of terpene and also contains alcohol and}

hydrocarbon, which resembles to green tea leaf. It also has low moisture content (4.17 ± 0.02 %), high total polyphenol content (26.42 ± 0.07 g/ 100 g), highest total catechin content (26.16 ± 0.13 g/ 100 g), and high teaine content (7.06 ± 0.01 g/ 100 g). Physical properties analysis shows it has low water activity (0.21 ± 0.02), lowest a value (3.80 ± 0.35), highsolubility (98.79 ± 0.53 %), low hygroscopicity (9.09 ± 0.94 %), and low bulk density (0.51 ± 0.18 g / mL).

EFFECTS OF INLET TEMPERATURE AND FEED

CONCENTRATION ON QUALITY OF SPRAY DRIED INSTANT

GREEN TEA POWDER

SKRIPSI

Submitted as a partial fulfillment of the requirement for degree of

SARJANA TEKNOLOGI PERTANIAN

at the Departement of Food Science and Technology

Faculty of Agricultural Engineering and Technology

Bogor Agricultural University

By:

PRADHINI DIGDOYO

F24080005

FACULTY OF AGRICULTURAL ENGINEERING AND

TECHNOLOGY

BOGOR AGRICULTURAL UNIVERSITY

BOGOR

Title : Effects of Inlet Temperature andFeed Concentrationon Quality of Spray Dried Instant Green Tea Powder

Name : Pradhini Digdoyo Student ID : F24080005

Approved by, Advisor

(Adil Basuki Ahza, PhD) NIP 19521021.197903.1.001

Acknowledged by:

Head of Departement of Food Science and Technology,

(Dr. Ir. Feri Kusnandar, M.Sc) NIP 19680526.199303.1.004

vii

STATEMENT LETTER OF SKRIPSI AND SOURCES OF

INFORMATION

Hereby I genuinely stated that the manuscript entitled Effects of Inlet Temperature and Feed Concentration on Quality of Spray Dried Green Tea Powderis an authentic work of mine under supervision of academic counselor and never being presented in any forms and universities. All the information taken and quoted from published or unpublished works of other writers had been mentioned in the texts and attached in the bibliography at the end of this manuscript.

Bogor, December 7, 2012 The undersigned,

Pradhini Digdoyo

F24080005

viii

AUTHOR BIOGRAPHY

Pradhini Digdoyo was born in Banyuwangi, East Java on December 7,1989. She was graduated from SD Negeri Cluring 1 elementary school in 2002, SMP Negeri 1 Cluring junior high school in 2005, and senior high school SMA Negeri 1Genteng in 2008. In the same year, she joined Bogor Agricultural University through Undangan Seleksi Masuk IPB (USMI) and she was graduated her bachelor degree of major Food Science and Technology in 2012. She was much involved in student activities such as Ikatan Mahasiswa Jawa Timur, she also was active as committee in HACCP (2010), Lomba Cepat Tepat Ilmu Pangan (2010), BAUR (20010) and IFBQC (2011). She also was active as laboratory assistant in Food Biochemistry Practice and Food Microbiology Practice.

Pradhini got some scholarships and achievements during her study at Bogor Agricultural University. She won as second winner in short story writing competition in Bogor Agricultural University (2009). And in 2010 she won as first winner in General Science Competition in Faculty of Agricultural Engineering and Technologi IPB. And the last, in 2012, she was selected to be the Indonesian delegates for the Credit Transfer MIT (Malaysia-Indonesia-Thailand) Student Exchange Program in Mae Fah Luang University, Thailand. Under such program, and did her undergraduated research project entitled “Effects of Inlet Temperature and Feed Concentration on Quality of Green Tea Powder” supervised by Aj. Natthawuddhi Donlao, Dr. Puwanart Fuggate, and Adil Basuki Ahza, PhD.

ix

FOREWORD

The author would like to thank God for the blessings, guidance and protection upon the writer throughout the whole research work and skripsi preparation. The research project entitled “Effects of Inlet Temperature and Feed Concentration on The Quality of Spray Dried Instant Green Tea Powder” was conducted at the School of Agro-Industry, Mae Fah Luang (MFU, Thailand) under Credit Transfer-MIT Program. The thanks and extensive gratitude of the author to:

1. My lovely Mom (Ida), Dad (Syahril), sister (Fika), and Dading Zainul Gusti, who has always supported me, for their love and their understanding in my life.

2. My research advisor in Bogor Agricultural University, Dr. Ir. Adil Basuki Ahza, MS for his advice and support.

3. My academic advisor in Mae Fah Luang University, Natthawuddhi Donlao B. Sc, M Eng., and my co-advisor DR. Puwanart Fuggate, B. Sc, M. Sc., thanks for the considerable advice. 4. The committee of MIT exchange program in Departement of Food Science and Technology,

Bogor Agricultural University and Mae Fah Luang University.

5. Directorate General of Higher Education- Ministry of National Education of the Republic Indonesia, which facilitated a student mobility program and for full financial support during MIT program.

6. The Ambassador of the Republic of Indonesia, Mr. H.E. Lutfi Rauf and Mr. Yunardi, The Attache of Education of the Republic of Indonesia, for the support and help during in Thailand 7. The commitee in School of Agroindustry who have evaluated my research during presentation. 8. All the laboratory staffs in S4 and S2 laboratory in Mae Fah Luang University

9. My friends in MFU University, Beam, Ben, Far, Nang, and others., for their kindness and friendship

10. All my classmates in ITP 45, especially for Ariesta, Taufiq, Annisa, Dini, Kamaliah, Virza, Chairul, Stefani, Sarinah, Misran, Hafiz, and Bangkit.Thanks for the sweet friendship.

Finally, I wish this report will be useful for anyone who read.

Bogor, December 7, 2012

x

TABLE OF CONTENTS

Contents

FOREWORD ... ix

TABLE OF CONTENTS ... x

LIST OF FIGURES ... xii

LIST OF TABLES ... xiii

APENDICES ... xiv

I. INTRODUCTION ... 1

A.

BACKGROUND ... 1

B.

OBJECTIVE ... 2

II. LITERATURE REVIEW ... 3

A.

TEA ... 3

B.

SPRAY DRYING ... 5

C.

FOOD POWDER ... 6

III. RESEARCH METHODOLOGY... 8

A.

MATERIALS AND INSTRUMENTS ... 8

1.

Materials ... 8

2.

Instruments ... 8

B.

EXPERIMENTAL DESIGN ... 8

1.

Production of concentrated green tea extract ... 8

2.

Production of green tea powder ... 9

C.

METHOD OF ANALYSIS ... 10

1.

Moisture content (AOAC 2000) ... 10

2.

Bulk density (Bhandari et al 1992) ... 10

3.

Color ... 11

4.

Water activity ... 11

5.

Higroscopicity (Jaya and Das 2004 modification) ... 11

6.

Solubity (Sanpakdhee 2007) ... 11

7.

Volatile compound (Pripdeevech and Matchan 2011) ... 12

8.

Total polyphenol content ... 12

9.

Teaine and catechin ... 12

xi

11.

Statistical analysis ... 13

IV. RESULTS AND DISCUSSION ... 14

A.

CHEMICAL COMPOSITION OF GREEN TEA LEAF (Camellia

sinensis) ... 14

B.

PRODUCTION OF CONCENTRATED GREEN TEA EXTRACT ... 16

C.

PRODUCTION OF GREEN TEA POWDER... 19

1.

Physical properties ... 20

2.

Chemical compound ... 23

3.

Volatile compound ... 25

D.

EFFECT OF TEMPERATURE AND FEED CONCENTRATION ON

QUALITY OF INSTANT GREEN TEA POWDER ... 26

V. CONCLUSION AND RECOMMENDATION... 28

A.

CONCLUSION ... 28

B.

RECOMMMENDATION ... 28

xii

LIST OF FIGURES

Figure 2. 1 Green tea plant ... 3

Figure 3. 1 Production of concentrated green tea extract ... 9

Figure 3. 2 Production of green tea powder ... 10

Figure 4. 1. Dried green tea leaf milling ... 14

Figure 4. 2 Pressing machine ... 16

Figure 4. 3. Freeze concentrating machine ... 17

Figure 4. 4 Centrifuge machine ... 17

Figure 4. 5 Relationship between

o

brix (refractometer) and total solid (oven

method) ... 17

Figure 4. 6 Volatile compounds in green tea extract ... 18

Figure 4. 7 Spray drying process ... 19

Figure 4. 8 Green tea powder ... 20

xiii

LIST OF TABLES

Table 2. 1 Taxonomy of tea plant ... 3

Table 2. 2 Aspects from food powder from consumer’s point of view ... 6

Table 4. 1 Chemical composition of dried green tea leaf ... 14

Table 4. 2Volatile compound in dried green tea leaf ... 15

Table 4. 3 Teaine and catechin content of green teas from different areas ……...15

Table 4. 4 Chemical composition of green tea extract... 18

Table 4. 5 Physical propertiesof green tea powder ... 20

xiv

APENDICES

Appendix 1. Data of volatile compounds ... 35

Appendix 2. Data of volatile compounds (cont’d) ... 36

Appendix 3. Data of moisture content of green tea powder ... 38

Appendix 4. Output of analysis of variance of moisture content ... 38

Appendix 5. Duncan’s test analysis result of moisture content... 39

Appendix 6. Data of total polyphenol content of green tea powder ... 40

Appendix 7. Output of analysis of variance of total polyphenol content of green

tea powder ... 41

Appendix 8. Duncan’s test analysis result of total polyphenol content of green tea

powder ... 41

Appendix 9. Data of gallic acid content of green tea powder of green tea powder

... 43

Appendix 10. Output of analysis of variance of gallic acid content of green tea

powder ... 43

Appendix 11. Duncan’s test analysis result of gallic acid content of green tea

powder ... 44

Appendix 12. Data of teaine content of green tea powder ... 45

Appendix 13. Output of analysis of variance of teaine content of green tea

powder ... 46

Appendix 14. Duncan’s test analysis result of teaine content of green tea powder

... 46

Appendix 15. Data of total catechin content of green tea powder... 48

Appendix 16. Output of analysis of variance of total catechin of green tea powder

... 48

Appendix 17. Duncan’s test analysis result of total catechin of green tea powder

... 49

Appendix 18. Data of water activity of green tea powder ... 50

Appendix 19. Output of analysis of variance of water activity of green tea powder

... 51

Appendix 20. Duncan’s test analysis result: Effect of interaction between feed

concentration and inlet temperature on water activity of green tea powder ... 51

Appendix 21. Data of L value of green tea powder ... 52

Appendix 22. Output of analysis of variance of L value of green tea powder ... 52

Appendix 23. Duncan’s test analysis result of L value of green tea powder ... 53

Appendix 24. Data of a value of green tea powder ... 54

Appendix 25. Output of analysis of variance of a value of green tea powder ... 55

Appendix 26. Duncan’s test analysis result of a value of green tea powder ... 55

Appendix 27. Data of b value of green tea powder ... 56

Appendix 28. Output of analysis of variance of b value of green tea powder ... 57

Appendix 29. Duncan’s test analysis result of b value of green tea powder... 57

Appendix 30. Data of solubility of green tea powder ... 59

Appendix 31. Output of analysis of variance of solubility of green tea powder .. 59

Appendix 32. Duncan’s test analysis result of solubility of green tea powder .... 60

xv

Appendix 34. Output of analysis of variance of hygroscopicity of green tea

powder ... 62

Appendix 35. Duncan’s test analysis result of hygroscopicity of green tea powder

... 62

Appendix 36. Data of bulk density of green tea powder ... 64

Appendix 37. Output of analysis of variance of bulk density of green tea powder

... 64

Appendix 38. Duncan’s test analysis result of bulk density of green tea powder 65

1

I.INTRODUCTION

A.

BACKGROUND

Tea has been consumed worldwide for years and is one of the most popular beverages. Tea is popular because of its unique aroma and characteristic flavor. The taste and flavor of tea are controlled by key chemical components which are volatile terpenes, caffeine, organic acids, and polyphenol (Borse et al. 2002). Recently, popularity of tea has increased due to its potential health benefits against cardiovascular diseases and cancer as well as pharmaceutical activities such as antihypertensive, antiateriosclerotic, hipocholesteroladmic, and hypolipidemic properties mostly from activities of antioxidant flavonoids present in tea (Cheng 2006).

Based on tea manufacturing (fermentation) process, teas from the genus Camellia can be divided into three categories: green tea (unfermented), oolong tea (partially fermented), and black tea (fully fermented). Among all of these, however, the most significant effects on human health have been observed with the consumption of green tea (Cabrera et al. 2006). For green tea manufacturing, freshly plucked tea leafs are immediately steamed or pan-fired to inactivate polyphenol oxidase and native microflora that initiates and catalyses the aerobic oxidation of tea catechins during tea fermentation. Whereas, fresh tea leafs are crushed and allowed to wither to induce oxidation as a part of tea fermentation process prior to drying for the black tea manufacturing process (generally more than 80% fermented). The characteristic reddish-black color, reduced bitterness and astringency, and removal of leafy and grassy flavor are derived from this oxidation process giving black tea a marked distinction from green tea (Cheng. 2006)

Green tea is commonly consumed in form of dried leaf. However, as the development of technology, green tea powder has been improved. Green tea powder has some advantages like easy to be served, easy to handle, and compact form. Production of green tea powder is done through drying process such as spray drying. Spray-drying is a unit operation by which a liquid product is atomized in a hot gas current to instantaneously obtain a powder. The gas generally used is air or more rarely an inert gas as nitrogen. The initial liquid feeding the sprayer can be a solution, an emulsion or a suspension. Spray-drying produces, depending on the starting feed material and operating conditions, a very fine powder(10–50 lm) or large size particles (2–3 mm) (Gharsallaouiet al. 2007).

Decreasing water content and water activity, spray-drying is generally used in food industry to ensure a microbiological stability of products, avoid the risk of chemical and/or biological degradations, reduce the storage and transport costs, and finally obtain a product with specific properties like instantaneous solubility. (Gharsallaouiet al. 2007). However, the drying process can cause some changes in food, such as phytochemical compounds and physicochemical properties which can affect quality of the product.

The quality of a food powder is judged by the amount of physical characteristic and chemical compoundof the product which is greatly affected by drying process. There are many researches were did by researchers to know the effect of spray drying on powder characteristic. At this research, different of spray dryer inlet temperature and feed concentrations were used to evaluate its effects on quality of spray dried instant green tea

2

powder. This information 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.

OBJECTIVE

The objective of this research is to study effects of inlet temperature and feed concentration on quality of instant green tea powder which was produced by spray drying process.

3

II. LITERATURE REVIEW

A.

TEA

Tea is leaf of Camellia sinensis plant. The taxonomy of tea plant is shown in Table

2.1 and the figure of tea plant is shown in Figure 2.1.

Figure 2. 1. Green tea plant

Table 2. 1. Taxonomy of tea plant

Common name Tea

Kingdom Plantae

Division Spermatophyta

Subdivision Angiospermae

Class Dicotyledone

Ordo Guttiferales

Family Theaceae

Genus Camellia

Species Camellia sinensis

Tea (Camellia sinensis) is the most consumed drink in the world next to water and the amount of consumption well exceeds coffee, beer, wine, and soft drinks (Rietveld &2003). The reasons for the worldwide popularity of tea are unique aroma and characteristic flavor.But recently, its popularity has increased due to its potential health benefits against cardiovascular diseases and cancer as well as pharmaceutical activities such as antihypertensive, antiateriosclerotic, hipocholesteroladmic, and hypolipidemic properties mostly from activities of antioxidant flavonoids present in tea (Cheng 2006).

Catechin is major polyphenol in green tea. Sutherland et al. (2005) reported some beneficial properties of catechin. Catechins chelate metal ions such as copper(II) and iron(III) to form inactive complexes and prevent the generation of potentially damaging free radicals. Another mechanism by which the catechins exert their antioxidant effects is through the ultrarapid electron transfer from catechins to ROS-induced radical sites on DNA

4

(Anderson et al. 2001). A third possible mechanism by which catechins scavenge free radicals is by forming stable semiquinone free radicals, thus, preventing the deaminating ability of free radicals (Guo et al. 1996). In addition, after the oxidation of catechins, due to

their reaction with free radicals, a dimerized product is formed, which has been shown to have increased superoxide scavenging and iron-chelating potential (Yoshino et al. 1999)

Epigallocatechin gallate has shown evidence of modulatinapoptotic pathways to protect against oxidative stress.Koh et al. (2003) showed that after hydrogen peroxideexposure in PC12 cells, EGCG inhibited many points ofthe apoptotic sequence, including caspase 3, cytochromec release, poly(ADP-ribose) polymerase cleavage, theglycogen synthase kinase-3 pathway and modulated cellsignaling by activating the phosphatidyl inositol-3 kinase(PI3K)/Akt pathway (which promotes cell survival). Furtherstudies have confirmed this by showing that after 3-HKexposure in SH-SY5Y human neuroblastoma cells, apoptosisand caspase 3 activity were inhibited by EGCG (Jeong et al.

2004). Catechins can also modulate apoptosis by altering the expression of antiapoptotic and proapoptotic genes. Epigallocatechin gallate prevented the expression of proapoptotic genes Bax, Bad and Mdm2 while inducing the antiapoptotic genes Bcl-2, Bcl-w and Bcl-xL to protect SH-SY5Y cells from 6-OHDA-induced apoptosis (Leviteset al. 2002).

Indonesia is one of the countries that produce tea. Tea production in Indonesia reached 150.342 tons in 2010 (Ministry of Agriculture). Indonesia’s tea production rate is 1.516 kg per hectare in a year which is lower than other tea producer countries. The government has taken some actions to increase tea production rate. These include uses of superior variety and education to tea farmer (Ministry of Agriculture). Other countries that produce tea are India, China, Sri Langka, Myanmar, also Thailand. Tea produced by these countries has each characteristic.

Tea from the genus Camellia can be divided into three categories based on tea manufacturing (fermentation) process: green tea (unfermented), oolong tea (partially fermented), and black tea (fully fermented). Green tea has been regarded as a rich source of catechin and its derivatives called tea catechins or flavan 3-ols including catechin (C), epicatechin (EC), epigallocatechin (EGC), epicatechin gallate (ECG), epigallocatechin gallate (EGCG), and gallocatechin gallate (GCG) that contribute to antioxidant capacity and organoleptic properties (Fernández et al. 2000). For green tea manufacturing, freshly plucked tea leaf is immediately steamed or pan-fired to inactivate polyphenol oxidase and native microflora that initiate and catalyses the aerobic oxidation of tea catechins during tea fermentation. Then it is allowed to whitering and drying process.

Green tea is popular in the Southeast Asian countries. The origin of green tea is the southwestern plateau region of China and golden delta area located partially in China, Vietnam, and Myanmar (Burma). Green teas differ depending on the type of process, whether it is steamed or pan-fired. Generally, green tea is steamed in Japan and pan-fired in the other Southeast Asian countries. Green tea has light yellowish green to green color when it is brewed. High-quality green tea has a slight greenish flavor given by certain terpenes such as linalool. Generally, concentrations of linalool and hexanal seem to play an important role in the quality of green teas (Kato and Sibamoto 2001).

Green tea contains some compounds like teaine, cathecin, and organic acid that affect flavor and aroma of green tea. Those compounds also bring beneficial health to the human. Teaine (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 occurrence in beverages

5

such as coffee and tea, as well as various soft drinks. Teaine extract from tea is added to some pain-relief medicines. teaine 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 and Cho 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).

B.

SPRAY DRYING

Spray drying is one of the drying methods that commonly used to dry food product. There are several reasons why food products are dried. First is lower transport cost, the total volume of the product decrease considerably if the product has been dried. Second is lower packing cost, also the packing cost become lower by the smaller of volume, but these costs become also lower because one can be enough with simple packing resources. For liquid products one must apply fluid-dense packing. The last reason is longer durability. Longer durability has to do with bacterium increase in liquid products. One of the most important factors for bacterium to be able to increase is water. If water has been removed, the bacteria will no longer grow (Weernink 2000).

Spray dryer have been used massively in food industry. Spray dryer can handle product with higher viscosity well. The viscosity stipulates for an important degree the atomizing system that can be used in a spray dryer. The two most used atomizing systems are wheel atomizing and high pressure systems with nozzle atomizing (Weernink 2000).

The spray drying process takes place in a large chamber. The product is atomized in very small drops in hot air. These drops will dry very rapidly, and dry particles fall to the lower part of dryer. Drying droplets is a physical process, it simply is water evaporation. The speed of evaporation depends on the ability of the air to take up the vapour. The important items for evaporation are the water absorption capacity of air, the contact area between air and solution, and the temperature of the solution. The water absorption capacity of air can increase by raising the temperature of air. The contact area between air and solution can be increased by making the droplets as small as possible during atomizing (Weernink 2000).

During drying, it is possible to agglomerate the product. This means that particles which have been atomized as small as possible are sticked together again. Because of this during drying larger particles are formed which have better dissolving properties. The agglomeration takes place in the top of the dryer. By placing the nozzles in a such way that the particles will hit each other a moment when they are not entirely dry, these particles will stick to each other and agglomerates will be formed. If also the fine particles from the cyclones or the bag filters are send back to this area, these particles will also stick to the almost dry particles in the top and result a better soluble product will be produced. Small

6

particles dissolve difficulty because they float on a water and as a result are difficult to moisten (Weernink 2000).

Spray drying is widely used in food industry. There are some studies that have been done to analyze effect of spray drying to quality of the food product. Some changes that are generally happened during process are decrease in water activity, glass transition, crystallization, melting of fat, and migration or retention of components, whether volatile or not (Bonazzi & Dumoulin 2011). A study on spray drying of Roselle extract indicated that some volatile compounds were lost and some others were generated due to a degradation process (Gonzales-Palomares 2009). Study on tomato powder by Santos de Sousa et al. (2008) showed that all the samples became significantly darker and less red with an increase of the variables under study. A low atomization speed (25000 rpm) and lower inlet air temperature (220 o_{C) produced the powders with a higher color index (a/b) and less}

darkening. Thankitsunthorn et al. (2009) reported that as inlet drying temperature increased, the moisture content and subsequentlythe water activity of indian goosebery powder decreased significantly at 0.05 level. The results also show that an increase in the inlet drying temperature results in a lower vitamin C content. Study on Agave angustifolia Haw by Fabela-Moron et al. (2011) indicated that increasing inlet air temperature increases the

particle size, average time of rehydration, and decreases the bulk density and moisture content of the powder. The powder characteristics of agave can be described through the effect of the dryer operating conditions on physical-chemical properties of powder.

C.

FOOD POWDER

Food powder can be produced through the drying process like spray drying. Food powder produced through spray drying process has some advantages, such as low transport cost, low packing cost, and long durability. Beside, during spray drying process, there is agglomeration process that results better soluble product (Weernink 2000).Some aspects from food powder from consumer’s point of view is shown on Table 2.2.

Table 2. 2. Aspects from food powder from consumer’s point of view

Appearance Color, brightness

Taste Taste itself, aroma, texture

Ease of use Rehydration or dissolving rate (cold or hot water) Apparent density (variation for proportioning) Stability overtime

Packaging enabling use and re-use of the product without any quality loss

No agglomeration or sticking to the packaging Composition Reproducibility

Additives origin and concentration

Energetic value (calories), fat content, vitamins, and so om

No allergens

No nitrate or pesticide residuals

Microbiology Limited number of microorganism with absence of pathogens

7

(Bonazzi & Dumoullin 2011)

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 cause 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 and Lillford 2008).

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 creation/existence 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 attributes, taste was the strongest attribute for both instant tea and green tea granules produced, and strength was the weakest attribute. (Sinija et al. 2011).

8

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% v/v) and gallic acid were supplied by Fluka (Buchs, Switzerland), anhydrous sodium carbonate and potassium hexacyanoferrate [K3Fe(CN)6] were purchased from Merck (Darmstadt, Germany). Standard HPLC of caffeine and cathecinsare purchased from Sigma-Aldrich (St. Louis, Missouri, USA), acetonitrile, trifluoroacetic acid (TFA), Monosodium phosphate monohydrate, disodium phosphate heptahydrate, and methanol (HPLC grade) were purchased from Fluka(Buchs, Switzerland). Other materials were citric acid (food grade), ammonium sulphate, dichloromethane, distilled water, and Whatman filter paper No 1.

2.

Instruments

The main instruments are JMC-miniLAB spray dryer (Euro Best Technology, ltd, Thailand), freeze concentrating machine (March Cool Industry Co.ltd, Thailand), centrifuge, hydrolic press (Owner Food Machinery Co.ltd, Thailand), hand refractometer (1-32o_{Brix ATAGO Model N-2E, Japan), a}

spectrophotometer (UV Vis. Biochrom/Libra S22, England), oven, disc mill, analytical balance, pHmeter, vacuum pump, SPME holder, SPME fiber 50/30 µm DVB/CAR/PDMS StableFlex/SS, round bottom flask, vial 250 mL, and HP model 6890 gas chromatograph equipped with an HP-5MS (5% phenyl-polymethylsiloxane) capillary column (30 m, 0.25 mm i.d., film thickness 0.25 µm; Agilent Technologies, USA) interfaced to an HP model 5973 mass-selective detector.

B.

EXPERIMENTAL DESIGN

This research is divided into two parts. The preliminary research is investigation of the chemical and volatile compounds of raw material (dried green tea leaf). Experiment I is production of concentrated green tea made from green tea extract and increase its concentration using freeze concentration machine, and then analysis of volatile compounds. Experiment II is production of green tea powder made from concentrated green tea using spray dryer, and then analysis of instant green tea powder characteristics.

1.

Production of concentrated green tea extract

Dried green tea leaf was milled with hammer mill resulted in milled green tea leaf. Then, milled green tea was extracted by dissolved in hot water (90o C) with regarding green tea: water 1:20 (w / v). The time of extraction was 60 minutes

9

and pH value was 5.0 (pH value was adjusted using citric acid). After that, extracted tea was filtered using clothes sheet and the residu was pressed by pressing machine to obtain extract of green tea. Total Dissolved Solid (TDS) of green tea extract was analyzed by refractometer and oven method. Then, green tea extract was concentrated using freeze concentration method, which was done by ice maker machine.

2.

Production of green tea powder

Green tea powder was produced from concentrated green tea extract using spray dryer. The operational conditions of the spray drying process were as follow: inlet air temperatures are 180, 200, and 220o _{C, outlet air temperature was}

controlled about 80o _{C, and blower speed was adjusted in 2500 rpm. The air}

pressure and feed rate need to be increased or decreased in order to control outlet temperature at 80o _{C. The air pressure and feed rate are affected by inlet air}

temperature. As the inlet air temperature increases, the air pressure and feed rate increase. Then, the final product was analyzed to determine volatile compounds, chemical compounds,and physical properties of dried green tea powder.

Extraction

Filtering

Green tea extract

Freeze concentrating

Concentrated green tea extract Milling

Dried green tea leaf

Figure 3. 1.Production of concentrated green tea extract

10

C.

METHOD OF ANALYSIS

1.

Moisture content (AOAC 2000)

Moisture content of the sample was determined using oven method. The moisture can was cleaned and dried in hot air oven for 12 hours, then cooled in desiccators and weighed using digital balance. Samples were weighed and placed into moisture can then were dried in hot air oven at 105o _{C overnight or until the}

constant weight accomplished. Then, the moisture can containing sample was cooled in desiccators and was weighed using digital balance. Dried sample was also weighed to determine its moisture content.

% M.C (wet basis) = ( ) ( ) 100

% M.C (dry basis) = ( ) ( ) 100

2.

Bulk density(Bhandari et al 1997)

Bulk density was determined by the tapping method. Two grams of powder are loosely weighed into 10 mL graduate cylinder. The cylinder containing the powder was tapped on a flat surface to a constant volume. The final volume was recorded and bulk density was calculated by dividing the sample weight by the volume.

Figure 3. 2.Production of green tea powder

Analysis of volatile compounds Spray drying Concentrated green tea

extracts (3, 6, 9 %)

11

3.

Color

Color values of dried samples (L, a, and b) were measured by using Color Quest XE/Hunter Lab (USA).

4.

Water activity

Water activity of dried samples were measured using Awmeter Novasina

5.

Higroscopicity (Jaya and Das 2004 modification)

A saturated solution of ammonium sulphate salt (equilibrium relative humidity is 80% at 20 ºC) was kept in glass wash bottle having two passage for air inlet and outlet. A diaphragm type vacuum pump was used to suck the air through the salt solution. Take filter paper in pump and weigh it until constant, then add powder sample 0.5 grams and it was spread in the filter paper. The increase in weight of the sample at every 15 min was noted. This measurement was continued till the difference between two weightings not exceed by 0.5%. The entire operation was carried out in a room maintained at 20 ºC.

Figure 3.3. Higroscopicity measurement equipment

6.

Solubity (Sanpakdhee 2007)

Weigh powder sample 0,5 g and mixed with 50mL of distilled water (25˚C)

in an 100mL beaker glass. Then it was agitated using a magnetic stirrer (diameter 2 mm and length 7mm) at a speed of 600rpm. The residue was filtrated on a filter paper No 4 and using the vacuum pump. The filter paper with an insoluble solid was placed in an oven set at 105 ± 2˚C until the weight is constant. The solubility (%) is calculated by using the following equation:

( %) = 1− 1− 2 100%

Where m1 is weight of filter paper and insoluble solid after dried by oven, m2 is weight of dried filter paper, and m is weight of powder sample.

12

7.

Volatile compound (Pripdeevech and Matchan 2011)

7.1.Extraction

The extraction was done using SPME fiber. Sample was weighed 20 gram and was placed into 250 mL vial with rubber inserted cover. Then SPME needle was inserted into vial. Vial and SPME holder was then placed in the oven at 50o C temperature 30 minutes.

7.2.Gas Chromatography –Mass Spectophotometry analysis

The volatile compound constituents of each sample were analyzed by an HP model 6890 gas chromatograph equipped with anHP-5MS (5% phenyl-polymethylsiloxane) capillary column (30 m x 0.25 mmi.d., film thickness 0.25 µm; Agilent Technologies,USA) interfaced to an HP model 5973 mass-selective detector. The oven temperature was initially held at 60o _{C and then}

increased by 2o _{C/min to 250}o _{C. The injector and detector temperatures were}

250o _{C and 280}o _{C, respectively. Purified helium was used as the carrier gas at}

a flow rate 1 mL/min. EI mass spectra were collected at70 eV ionization voltages over the range of m/z 29–300. The electron multiplier voltage was 1150 V. The ion source and quadrupolete temperatures are set at 230 and 150o

C, respectively. Identification of the volatile components was performed by comparison of the mass spectra of individual components with the reference mass spectra in the Wiley 275 and NIST 98 databases.

8.

Total polyphenol content

The total polyphenol content was determined by spectrophotometry, using gallic acid as standard, according to the method described by the International Organization for Standardization (ISO) 14502-1. Briefly, 1.0 mL of the diluted sample extract (50-100 fold dilution) was transferred in duplicate to separated tubes containing 5.0 mL of a 1/10 dilution of Folin-Ciocalteu’s in water. Then, 4.0 mL of a sodium carbonate solution (7.5% w/v) was added. The tubes were then allowed to stand at room temperatures for 60 min before absorbance at 765nm was measured against water. The total polyphenol was expressed as gallic acid equivalents (GAE) in g/100g material. The concentration of polyphenols in samples was derived from a standard curve of gallic acid ranging from 10 to 100

μg/mL.

9.

Teaine and catechin

Preparation of sample

Add to the instant tea (0.500±0.001) g in the flask approximately 25 mL of hot water (max 50ºC). The sample was mixed in room temperature. After that, add 5.0 mL acetonitrile and it was mixed again.

Preparation of Standards

Use the % purity from the certificate to prepare the stock standard solution. HPLC analysis

HPLC analysis of standards and samples was conducted on Water 966 high performance liquid chromatography comprising vacuum degasser, quaternary pump, auto-sampler, thermostatted column compartment, and photo diode array detector. The column used was a Platinum EPS C18 reversed phase, 3μm (L 53 x

13

i.d 7mm). Mobile phase eventually adopted for this study was water/acetonitrille (87:13) containing 0.05% (v/v) trifluoroacetic acid (TFA) with the flow rate of 2 mL/min. Absorption wavelength was selected at 210 nm. The column is operated at 30ºC. The sample injection was 20 μL. Peaks were identified by comparing their retention times and UV spectra in the 190-400 nm range with standards. The standard was injected before sample for made calibration curves. The caffeine content was calculated using its respective calibration curves.

10.

Gallic acid

Preparation of Standards

Weight Gallic acid 0.0100 g and dissolved substances 1 ml acetronitrile and

0.5 ml 10 μg/ml EDTA. Then, adjust the volume with distilled water 10 ml. Use

the 100% purity from the certificate to prepare the stock standard solution. HPLC analysis

HPLC analysis of standards and samples was conducted on Water 966 high performance liquid chromatography comprising vacuum degasser, quaternary pump, auto-sampler, thermostatted column compartment, and photo diode array

detector. The column used was a Platinum EPS C18 reversed phase, 3μm (L 53 x

i.d 7mm). Mobile phase eventually adopted for this study is water/acetonitrille (87:13) containing 0.05% (v/v) trifluoroacetic acid (TFA) with the flow rate of 2 mL/min. Absorption wavelength is selected at 210 nm. The column is operated at 30ºC. The sample injection is 10 μL. Peaks are identified by comparing their retention times and UV spectra in the 190-400 nm range with standards. The standard was injected before sample for made calibration curves. The Gallic acid content is calculated using its respective calibration curves.

11.

Statistical analysis

Data are analyzed by ANOVA using SPSS software application on univariate mode for full factorial. The post hoc test that used was Duncan’s test.

14

IV. RESULTS AND DISCUSSION

A.

CHEMICAL COMPOSITION OF GREEN TEA LEAF (Camellia

sinensis)

Dried green tea leaf (var. Oolong No. 12) was supplied by Boonrod Tea Factory (Thailand). Dried green tea leaf was milled using hammer mill as shown in Figure 4.1. There were some chemical compounds that have been observed in this research, which were moisture content, total polyphenol compound, teaine content, gallic acid, and cathecin. Beside, volatile compound in dried green tea leaf was also analyzed. Chemical composition on green tea leafis shown in Table 4.1. and volatile compound in green tea leaf is shown in Table 4.2.

Figure 4. 1. Dried green tea leaf milling

Table 4. 1Chemical composition of dried green tea leaf

Chemical composition Amount (Mean ± SD)

Moisture content (% w.b.) 6.91 ± 0.02

Total polyphenol (g/100g d.b.) 28.80 ± 0.18

Gallic acid (g/100g d.b.) 0.45 ± 0.02

Caffeine (g/100g d.b.) 1.21 ± 0.02

Total catechin (g/100g d.b.) 3.72 ± 0.08

GC (g/100g d.b.) 0.33 ± 0.00

EGC (g/100g d.b.) 0.97 ± 0.08

C (g/100g d.b.) 0.40 ± 0.02

EC (g/100g d.b.) 0.31 ± 0.01

EGCG (g/100g d.b.) 1.24 ± 0.11

GCG (g/100g d.b.) 0.23 ± 0.01

15

Table 4. 2Volatile compound in dried green tea leaf

Number Volatile compound Peak area/gram

1 2-propanone 116389.20

2 2-thiapropane 26040.51

3 2-methyl-butanal 10610.79

4 Trimethyl cyclohexanone 13404.64

5 Linalool oxide 49258.85

6 4-methyl-3-oriranemethanol 21376.33

7 Azulene 12510.03

Plants, including tea leaf from Camelia sinensis (Beecher 2003), produce secondary metabolites, organic compounds that are involved in the defense of the plants against invading pathogens, including insects, bacteria, fungi, and viruses. In the case of tea leaf, these metabolites include polyphenolic catechins and theaflavins and the alkaloids caffeine and theobromine.

Previous study by Graham (1992), Chu and Juneja (1997) said that the catechin content is up to 30% of the dry weight, whereas, the content of teaine is up to 5% of the dry weight. But, this result show a lower amount, total catechin content is 3.72 ± 0.08 (g/100g db) and caffeine content is 1.21 ± 0.02 (g/100g db). This can be explained by several factor such us season, varieties, and agricultural practice which are known to influence the content (Balentine et al. 1998 and Yao et al. 2009). Table 4.3. shows a result from Baptista et al.(1998)who studied some green teas from different area.

Table. 4.3. Teaine and catechin content of green tea from different area

Type of tea Teaine (% w/w) % Total Green Tea Polyphenol (w/w) Epicatechin Derrivate EGCG Azorean green tea

(Gorreana) 8.01 74.5 47.9

Chinese green tea

(Fuijian) 2.68 73.9 47.2

Chinese green tea

(Uncles Lee’s) 4.49 71.9 39.9

Japanese green tea

(Yamamoto) 2.66 59.5 37.4

Taiwan green tea

(Lung Ching) 2.50 66.8 42.6

(Baptista et al. 1998)

Based on Table 4.2., volatile compound analysis shows the highest constituent is 2-propanone, 2-propanone is the simplest form of ketone, other ketones are trimethyl cyclohexanone. Other constituents are alcohol, which are linalool oxide and 4-methyl-3-oriranemethanol, aldehide which is 2-methyl-butanal, hydrocarbon which is azulene, and sulfide, which is 2-thiapropane. These compounds are organic compounds that commonly found in plants (Pripdeevech and Matchan 2011). Kato and Shibamoto (2001) reported

16

that generally, concentrations of linalool and hexanal seem to play an important role in the quality of green tea.

B.

PRODUCTION OF CONCENTRATED GREEN TEA EXTRACT

Dried green tea leaf was extracted by dissolving in hot water (90o _{C) and then was}

filtered using pressing machine as shown in Figure 4.2to obtain green tea extract.

Figure 4. 2.Pressing machine

Green tea extract from previous step was then concentrated using freeze concentrating machine as shown in Figure 4.3. The principle of this machine is resulting slurry of ice crystals in a fluid concentrate. The ice crystals were then removed in some way, in this study it used centrifuge machine, as shown in Figure 4.4, for separating the ice crystals and a concentrated product. The total solid in fluid concentrate will increase as longer of freeze concentration time. There are three determined total solid, 3%, 6%, and 9%. The main advantage of freeze concentration process is it does not use high temperature to increase concentration of extract. On the other hand, it has some disadvantages, such us limited degree of final concentration achieved, long time process, and some compound can be disposed together with the ice crystal, as the suspended material can be a nucleus on crystallization process.

The concentration of extract was measured using refractometer and was confirmed using oven method. Hand refractometer is an equipment to measure TDS that content in fruit, food product that is contained fruit, and sucrose solution (Nielsen, 1996). Nowadays, hand refractometer is used in process of made a solution in the industry, like milk industry and beverage industry. The principle of hand refractrometer is measure the index of refraction from the food that contain of carbohydrate. The unit of refratometer is

˚Brix that equal with percentage of sucrose solution (g sucrose/ 100 g sample). Because

of this research used green tea extract as the sample, which it is non-sucrose sample, the calibration of hand refractometer measurement with oven method must be done. The relationship between total solid measurement using refractometer and total solid measurement using oven method is shown in Figure 4.5. The figure shows linear relationship between obrix and total solid that means refractometer can be used to measure concentration in green tea extract.

17

Figure 4. 3. Freeze concentrating machine

Figure 4. 4.Centrifuge machine

Figure 4. 5.Relationship between obrix (refractometer) and concentration (oven method)

y = 0,73x + 1,333 R² = 0,980

0 1 2 3 4 5 6 7

0 2 4 6 8

Concentration (%)

Brix

18

The chemical compositions and volatile compounds of concentrated green tea extract have been studied. The chemical composition is shown in table 4.4. and the volatile compounds are shown in figure 4.6.

Table 4. 3.Chemical composition of green tea extract

Chemical composition Sample

3% 6% 9%

Total polyphenol content (mg/100 mL) 25.19 ± 0.06a _{33.54 ± 0.01}b _{41.37 ± 0.05}c

Gallic acid (µg/ mL) 1.44 ± 0.02a _{2.45 ± 0.01}b _{4.45 ± 0.02}c

Caffeine (µg/ mL) 5.09 ± 0.04a 8.56 ± 0.04b 14.37 ± 0.05c

Total catechin (µg/ mL) GC (µg/ mL) EGC (µg/ mL) C (µg/ mL) EC (µg/ mL) EGCG (µg/ mL) GCG (µg/ mL) ECG (µg/ mL)

12.02 ± 0.12a

2.12 ± 0.01a 4.64 ± 0.01a

1.34 ± 0.03a 1.22 ± 0.02a

1.93 ± 0.02a

0.50 ± 0.04a

0.27 ± 0.01a

26.74 ± 0.68b

4.03 ± 0.01b 9.38 ± 0.04b

2.78 ± 0.00b 2.51 ± 0.00b

5.49 ± 0.00b

1.52 ± 0.03b

1.00 ± 0.07b

47.95 ± 0.06c

7.13 ± 0.01c 16.35 ± 0.05c

5.03 ± 0.01c 4.50 ± 0.01c

10.37 ± 0.01c

2.83 ± 0.01c

1.75 ± 0.00c

Value in a column followed by different letters are significantly (p<0.05) different.

Based on the Table 4.4., the result shows the highest amount of all the chemical composition is found in extract green tea 9%. This can be explained by extract 9% has the highest total solid of all samples. The highest catechin constituent in all samples is EGC then followed by EGCG, GC, C, EC, GCG, and ECG. This is different from green tea Leafs composition in which EGCG is the highest catechin constituent. According to Labbe et. al. (2008), EGCG extraction is more time/temperature dependent than EGC which means EGCG is more sensitive than ECG.

Figure 4. 6.Volatile compounds in green tea extract

There are three main groups of volatile compounds in green tea extract; alcohol, hydrocarbon, and terpene. Pripdeevech and Machan (2011) also found those compounds

0 10000 20000 30000 40000 50000 60000

Volatile compounds

Ext ract 3% Ext ract 6% Ext ract 9%

19

in green tea. The Extract 3% contains the highest amount of those compounds, followed by extract 6% and extract 9%. Alcohol is the biggest constituent in extract 3% and 6%, but it was not found in extract 9%. The biggest part of alcohol component is ethanol, the biggest part of hydrocarbon component is dichloromethane, and the biggest part of terpene is azulene. This composition is different with green tea leaf which means there is reaction occurred during extraction and freeze concentration process that caused change in volatile compound composition. Kim et al. (2007) also reported change in flavor of green tea liquor during heating caused by change in volatile compound composition.

C.

PRODUCTION OF GREEN TEA POWDER

Production of green tea powder was done through the spray drying process. There are some factors that can influence the drying process, such as inlet air temperature, outlet air temperature, blower speed, inlet and outlet humidity, and in feed condition, such as feed concentration, feed viscosity, feed temperature, and feed flow. This research used inlet temperature as the parameter. Inlet temperatures that have been used were 180 o_C,

200 oC, and 220 oC. According to Kim et al. (2009), heat sensitive product can be dried within the recommended temperature of 180 – 220o _{C inlet temperature. The outlet}

temperature was controlled at 80o C and the blower speed was adjusted in 2500 rpm. To control the outlet temperature, feed rate was need to be adjusted.

Green tea powder resulted from this process was then analyzed to observe physicalproperties, chemical composition, and volatile compound. The result is shown in Table 4.5.

20

Figure 4. 8.Green tea powder

1.

Physical properties

Table 4. 4.Physical propertiesof green tea powder

Total solid (%)

Inlet temp.

(0_C)

Aw L a b Solubility

(%)

Higroscopi city (%)

Bulk Density (g/mL)

3 180 0.2602 ±

0.0010d

58.38 ± 0.05b

4.00 ± 0.11c

18.62 ± 0.13a,b

98.65 ± 0.77b,c,d

8.51 ± 0.13a,b

0.6043 ± 0.0077f

3 200 0.2068 ±

0.0110b

64.11 ± 0.07d

3.98 ± 0.04b,c

19.45 ± 0.04e,f

99.21 ± 0.57c,d

10.10 ± 0.15c,d

0.5433 ± 0.0181d,e

3 220 0.2158 ±

0.0010b

57.80 ± 0.19a,b

4.10 ± 0.11c,d

18.40 ± 0.06a

97.22 ± 1.69a,b

12.56 ± 0.53e

0.5865 ± 0.1269f

6 180 0.2075 ±

0.0163b

57.92 ± 0.25a,b

3.80 ± 0.35a

18.87 ± 0.14b,c,d

98.79 ± 0.53b,c,d

9.09 ± 0.94b,c

0.5143 ± 0.1766c

6 200 0.2584 ±

0.1396d

56.48 ± 0.33a

4.10 ± 0.03c,d

18.46 ± 0.02a,b

96.51 ± 1.08a

11.16 ± 1.08d,e

0.5596 ± 0.0002e

6 220 0.2285 ±

0.0092b,c

58.76 ± 0.89b

3.98 ± 0.03b,c

19.11 ± 0.33d,e

97.57 ± 0.02a,b,c

7.24 ± 0.97a

0.5320 ± 0.0050c,d

9 180 0.1796 ±

0.0058a

59.42 ± 1.75b,c

3.82 ± 0.07a,b

18.96 ± 0.37c,d

99.54 ± 0.29d

11.36 ± 0.56d,e

0.5467 ± 0.1361d,e

9 200 0.2158 ±

0.0166b

60.71 ± 0.26c

4.01 ± 0.10c

19.56 ± 0.08f

97.15 ± 0.26a,b

12.31 ± 1.47e

0.4752 ± 0.0038b

9 220 0.2438 ±

0.0267c,d

63.69 ± 0.28d

4.22 ± 0.01d

20.39 ± 0.63g

95.77 ± 0.47a

12.33 ± 0.58e

0.4272 ± 0.0024a Value in a column followed by different letters are significantly (p<0.05) different

1.1. Water activity

Water activity (Aw) is an important index for spray-dried powder because it can greatly affect the shelf life of the powder. It is defined as the ratio of vapor pressure of water in a food system to vapour pressure of pure

21

water at the same temperature (Fennema OR 1996).Water activity is different from moisture content as it measures the availability of free water in a food system that is responsible for any biochemical reactions, whereas the moisture content represents the water composition in a food system. High water activity indicates more free water available for biochemical reactions and hence, shorter shelf life. Generally, food with Aw< 0.6 is considered as microbiologically stable and if there is any spoilage occurs, it is induced by chemical reactions rather than by micro-organism (Bonazzi and Dumoullin 2011).

The results shows water activity of green tea powder were in range of 0.18- 0.26. This means that the spray-dried powders were relatively stable microbiologically. However, the storage conditions also played an important role in this matter. Based on statistical analysis (appendix 18), feed concentration did not significantly affect water activity, neither inlet temperature. However, interaction between total solid in feed and inlet temperature affected water activity of green tea powder resulting significant difference among the samples. The highest water activity (is belong to green tea powder with 3% total solid in feed and 180 o_{C inlet temperature, the lowest}

water activity belong to green tea powder with 9% feed concentration and 180

o_{C inlet temperature, but it is not significantly different with other samples in}

the same subset).

1.2. Color

Color is one of the important sensory attributes of food and a major quality parameter in dehydrated food. During drying, color may change because of chemical or biochemical reaction. Enzymatic oxidation, Maillard reactions, caramelization, and ascorbic acid browning are some of the chemical reaction that can occur during drying and storage. Discoloration and browning during air drying may be result of various chemical reactions including pigment destruction (Farias and Ratti, 2009). The attributes as indicator in determining color are L, a, b, and hue values. L value indicates the brightness of sample with range 0 (black) to 100 (white). The a value indicates a micture colors of red and green. The +a value indicates red color with range 0-100, while –a value indicates green color with range 0-(-80). The b value indicates a combination of yellow and blue. +b range for 0-70 indicates yellowness while –b range for 0-(- 70) for blueness.

Based on Table 4.5., the result shows colors among samples are significantly different. The highest L value, which means the brightest color, belong to powder with 3% total solid in feed and 200o _{C inlet temperature but}

it is not significantly different with powder 9% feed concentration and 220o _C

inlet temperature. On the other hand, the darkest color, or the lowest L value belong to sample with 6% feed concentration and 200o C inlet temperature, but it is not significantly different with other samples in the same subset. The highest a value belongs to sample with 9% feed concentration and 220o _{C inlet}

22

6% feed concentration and 180o _{C inlet temperature has the lowest a color}

which means this sample has the most greenish color. Powder that has the highest b color or the most yellowish is sample with 9% feed concentration and 220o _{C inlet temperature, and the sample that has the lowest b value or the}

most bluish color is the one with 3% feed concentration and 180o C inlet temperature.

The result shows (Figure 4.8.) that green tea powder has greenish yellow color. Generally, increasing temperature increase L, a, and b value. High temperature makes shorter drying time, it caused browning reaction occurs faster. Besides, The Maillard reaction may occur in this research because green tea contains carbohydrate about 7% dry weight of tea leaf (Chako et al, 2010). However, freeze concentration also gives the effect of reducing color in the feed preparation and it results in different color of origin.

1.3. Solubility

Solubility is the most reliable criterion to evaluate the behavior of powder in aqueous solution. This parameter is attained after the powder undergoes dissolution steps of sinkability, dispersability and wettability. Solubility describe feasibility of powder to be dissolved in water (Chen and Patel, 2008). Based on Table 4.5., the result shows solubility of samples is in range of 95 – 99 %, this result is agreement with Nadeem et. al. (2011) who reported solubility of spray dried mountain tea are mostly between 98.5 and 99.5 %. The highest temperature belong to sample with 9% total solid in feed and 180o _{C inlet}

temperature, the lowest solubility is belong to powder with 9% total solid in feed and 220o _{C inlet temperature.}

Based on statistical analysis (appendix 31.), concentration did not significantly affect solubility, otherwise temperature did. Interaction between concentration and temperature significantly affect solubility of green tea powder. A hard surface layer might be formed over the powder particle at higher inlet temperature. This could prevent water molecules from diffusing through the particle. Consequently, decreased the wettability of the particle and reduced the solubility of the powder (Chegeni and Ghobadian 2005).

1.4. Hygr oscopicity

Hygroscopicity is the ability of food powder to absorb moisture from high relative humidity environment. In the case of fruit powders, glucose and fructose are responsible for strong interaction with the water molecule due to the polar terminals present in these molecules (Slade and Levine 1991).

The results shows the highest hygroscopicity belong to sample with 3% feed concentration and 220 oC inlet temperature, but it is not significantly different with other samples in the same subset. The lowest hygroscopicity belong to green tea powder with 6% feed concentration and 180o _{C inlet}

23

subset. Hygroscopicity of sample is varying among samples in range of 7.24 – 12.56%. This is higher than reported by Jaya and Das (2004), higroscopicity in instant coffe powder which is in range 9.09–10.32 %. But the result is lower than higroscopicity of spray dried mango powder which has higroscopicity 16.5 %. This variation can be explained by difference of material and condition of drying process. High higroscopicity indicates its strong capacity to attract water molecules when in contact with the surrounding air. This can lead to cacking and agglomeration in powder. Adding carrier, such us maltodextrin can help to lower higroscopicity (Ahmed et al 2010; Rodríguez-Hernández et al. 2005; Cai and Corke 2000).

1.5. Bulk density

Bulk density is one of food powder properties. The result shows bulk density is significantly different among samples in range 0.4272 ± 0.0024- 0.6043 ± 0.0077g/mL. The highest bulk density belong to sample with 3% feed concentration and 180o _{C inlet temperature, and the lowest bulk density belong}

to sample with 9% feed concentration and 220 oC inlet temperature, but both of them are not significantly different with other samples in the same subset. Higher total solid in feed decreased the bulk density, as well as higher temperaturedecreased the bulk density. This result is in agreement with Goula and Adamopoulos 2010, increased inlet air temperature causes a reduction in bulk density, as evaporation rates are faster and products dry to a more porous or fragmented structure. Walton (2000) reported that increasing the drying air temperature generally produces a decrease in bulk and particle density, and there is a greater tendency for the particles to be hollow. Also, the higher moisture content of powder, the more particles tend to stick together, leaving more interspace between them and consequently resulting a larger bulk volume (Goula and Adamopoulos 2005).

2.

Chemical compound

Table 4. 5The result of chemical analysis of green tea powder

Total solid (%)

Inlet temp.

(o_C)

Moisture content (% wb)

Total polyphenol (%

db)

Gallic acid (%

db) Teaine (% db)

Catechin (% db) 3 180 4.35 ± 0.02c 31.22 ± 0.11h 2.21 ± 0.10a 6.50 ± 0.02c 16.53 ± 0.28b 3 200 4.41 ± 0.02c _{30.22 ± 0.15}g _{2.38 ± 0.01}b,c _{6.33 ± 0.06}b _{15.17 ± 0.02}a

3 220 4.31 ± 0.04c 27.86 ± 0.04f 2.72 ± 0.01d 7.38 ± 0.02f 21.79 ± 0.07c 6 180 4.17 ± 0.02a _{26.42 ± 0.07}e _{2.38 ± 0.00}b,c _{7.06 ± 0.01}d _{26.16 ± 0.13}f

6 200 4.16 ± 0.11a _{23.82 ± 0.20}d _{2.46 ± 0.07}c _{6.47 ± 0.02}c _{16.18 ± 0.31}b

6 220 4.04 ± 0.01a _{23.96 ± 0.27}d _{2.31 ± 0.14}a,b _{6.20 ± 0.01}a _{23.77 ± 0.07}d

9 180 4.29 ± 0.01b _{22.40 ± 0.10}c _{2.62 ± 0.01}d _{7.41 ± 0.00}f _{24.75 ± 0.15}e

9 200 4.06 ± 0.03a 21.07 ± 0.07b 2.68 ± 0.07d 7.06 ± 0.06d 24.47 ± 0.14e 9 220 4.10 ± 0.00a _{20.65 ± 0.01}a _{2.66 ± 0.04}d _{7.26 ± 0.00}e _{26.20 ± 0.04}f

24

2.1. Moistur e content

Moisture content describes water composition in food. The result shows moisture content of green tea powder varies in range of 4.06 – 4.41 %, a small variation means that the drying process was done uniformly for all samples. Statistical analysis shows higher temperature lower the moisture content. This result is in agreement with Quek et al. (2006) who reported the moisture content

of the spray-dried powders decreased with the increased in inlet and outlet air temperature. This is because at higher inlet temperature, the rate of heat transfer to the particle is greater, providing greater driving force for moisture evaporation. Consequently, powders with reduced moisture content are formed.

2.2. Total polyphenol content

Total polyphenol content was analyzed using gallic as standard. The result (Table 4.6.) shows vary amount of total polyphenol content in sample in range 20.65 – 31/22 %. This result is higher than total polyphenol content in instant tea reported by Sinija et al. (2007). The highest amount of total polyphenol is found at green tea powder with 3% feed concentration and 180 o_C

inlet temperature, and the lowest one is found at sample with 9% feed concentration and 220 o_{C inlet temperature, but both of them are not}

significantly different with other samples in the same subset. Increasing total solid results decreased polyphenol content. Increasing inlet temperature also results decreased polyphenol content. This can be occurred because higher feed concentration means longer time in freeze concentration process in which can release polyphenol content during process, higher temperature caused degradation of polyphenol. Georgetti et al. (2008) also reported a slight decrease in the total polyphenol content of the spray-dried soybean extract by increasing the inlet air temperature. Monica et al. (2009) reported that increased temperature caused degradation on polyphenol content in apricot, especially epicatechin and quercetin.

2.3. Gallic acid

Gallic acid was analyzed using HPLC. Based on Table 4.6., the result shows vary amount among sample. Gallic acid of green tea powder is in range of 2.21% – 2.26% db. The highest gallic acid content is found at sample with 3% total solid temperature and 220 o_{C inlet temperature but it is not significantly}

different with other samples in the same subset. Yet the lowest gallic acid content is found at sample with 3% total solid in feed and 180 o_{C inlet}

temperature. Generally, higher total solid and higher temperature result higher gallic content. It can be explained by high total solid in feed and short time drying process.

25

2.4. Teaine

Teaine is one of the components that affect flavor of green tea. Based on Table 4.6., teaine content in sample varies among samples in range 6.20 – 7.41 %. This result is higher than teaine content in instant tea as reported by Shinja et al. (2007). The highest teaine content is found at sample with 3% feed concentration and 220 o_{Cinlet temperature, and the lowest is found at the sample}

with 6% feed concentration and 220 o_{C inlet temperature (but not significantly}

different with other samples in the same subset). Interaction between feed concentration and inlet temperature significantly affectedteaine content.

2.5. Catechin

Catechin is the important group of tea leaf compound that affects tea flavor. Catechin was analyzed using HPLC. Based on Table 4.6., catechin content of green tea powder varies among samples in range of 15.17 – 26.20 %. The highest catechin content is found at the sample with 9% feed concentration and 220 o_{C inlet temperature, And the lowest catechin content is found at the}

sample with 3% feed concentration and 200 o_{C inlet temperature (but it is not}

significantly different with other samples in the same subset). Increasing feed concentration results increased catechin.The highest catechin amount was found in sample with 9 % feed concentration 220 o_{C inlet temperature and 6% feed}

concentration and 180 o_{C inlet temperatuure. It can be explained by high total}

solid content in feed and short drying time.

3.

Volatile compound

Figure 4. 9.Volatile compounds in green tea powder

Volatile compound analysis was done by GC-MS equipment and the extraction was done by SPME. There are seven major groups of volatile compounds found in green

0 50000 100000 150000 200000 250000 300000

3% 180 3% 200 3% 220 6% 180 6% 200 6% 220 9% 180 9% 200 9% 220

P e a k a re a / g ra m Treatment

Green tea powder

Alcohol Ket one Hidrocarbon Acid Aldehide Azole Terpene

61

Solubility

Duncan

trt N

Subset

1 2 3 4

9 2 95.7703

5 2 96.5120

8 2 97.1496 97.1496

3 2 97.2249 97.2249

6 2 97.5734 97.5734 97.5734

1 2 98.6523 98.6523 98.6523

4 2 98.7859 98.7859 98.7859

2 2 99.2102 99.2102

7 2 99.5415

Sig. .064 .088 .084 .318

Means for groups in homogeneous subsets are displayed. Based on observed means.

The error term is Mean Square(Error) = .621.

Appendix 33. Data of hygroscopicity of green tea powder

Treatment Total solid (%) Temperature (o_{C) Hygroscopicity (%)}

1 3 180 8.51 ± 0.13a,b

2 3 200 10.10 ± 0.15c,d

3 3 220 12.56 ± 0.53e

4 6 180 9.09 ± 0.94b,c

5 6 200 11.16 ± 1.08d,e

6 6 220 7.24 ± 0.97a

7 9 180 11.36 ± 0.56d,e

8 9 200 12.31 ± 1.47e

9 9 220 12.33 ± 0.58e

62

Appendix 34. Output of analysis of variance of hygroscopicity of green tea powder

Tests of Between-Subjects Effects

Dependent Variable:Higroscopicity

Source

Type III Sum of

Squares df Mean Square F Sig.

Corrected Model 57.488a _{8 7.186 21.283 .000}

Intercept 1991.937 1 1991.937 5.900E3 .000

Concentration 24.324 2 12.162 36.021 .000

Temperature 7.362 2 3.681 10.902 .004

Concentration *

Temperature 25.802 4 6.451 19.105 .000

Error 3.039 9 .338

Total 2052.464 18

Corrected Total 60.527 17

a. R Squared = .950 (Adjusted R Squared = .905)

Appendix 35. Duncan’s test analysis result of hygroscopicity of green tea powder

Appendix 35. 1. Effect of feed concentration on hygroscopicity of green tea powder Hygroscopicity

Duncan Concen

tration N

Subset

1 2 3

6% 6 9.1657

3% 6 10.3891

9% 6 12.0042

Sig. 1.000 1.000 1.000

Means for groups in homogeneous subsets are displayed. Based on observed means.

63

Appendix 35. 2. Effect of inlet temperature on hygroscopicity of green tea powder

Hygroscopicity

Duncan Tempera

ture N

Subset

1 2

180 C 6 9.6586

220 C 6 10.7105

200 C 6 11.1898

Sig. 1.000 .187

Means for groups in homogeneous subsets are displayed.

Based on observed means.

The error term is Mean Square(Error) = .338.

Appendix 35. 3. Effect of interaction between feed concentration and inlet temperature on hygrospicity content of green tea powder

Hygroscopicity

Duncan

trt N

Subset

1 2 3 4 5

6 2 7.2425

1 2 8.5143 8.5143

4 2 9.0945 9.0945

2 2 10.0978 10.0978

5 2 11.1600 11.1600

7 2 11.3670 11.3670

8 2 12.3118

9 2 12.3337

3 2 12.5553

Sig. .056 .344 .118 .066 .054

Means for groups in homogeneous subsets are displayed. Based on observed means.

64

Appendix 36. Data of bulk density of green tea powder

Treatment Total solid (%) Temperature (0_{C) Bulk Density (g/mL)}

1 3 180 0.6043 ± 0.0077f

2 3 200 0.5433 ± 0.0181d,e

3 3 220 0.5865 ± 0.1269f

4 6 180 0.5143 ± 0.1766c

5 6 200 0.5596 ± 0.0002e

6 6 220 0.5320 ± 0.0050c,d

7 9 180 0.5467 ± 0.1361d,e

8 9 200 0.4752 ± 0.0038b

9 9 220 0.4272 ± 0.0024a

Value in a column followed by different letters are significantly (p<0.05) different

Appendix 37. Output of analysis of variance of bulk density of green tea powder Tests of Between-Subjects Effects

Dependent Variable:BulkDensity

Source

Type III Sum of

Squares df Mean Square F Sig.

Corrected Model .048a _{8 .006 49.015 .000}

Intercept 5.097 1 5.097 4.198E4 .000

Concentration .027 2 .014 111.787 .000

Temperature .005 2 .003 21.008 .000

Concentration *

Temperature .015 4 .004 31.632 .000

Error .001 9 .000

Total 5.145 18

Corrected Total .049 17

65

Appendix 38. Duncan’s test analysis result of bulk density of green tea powder Appendix 38. 1. Effect of feed concentration on bulk density of green tea powder

BulkDensity

Duncan Concen

tration N

Subset

1 2 3

9% 6 .483053

6% 6 .535290

3% 6 .578012

Sig. 1.000 1.000 1.000

Means for groups in homogeneous subsets are displayed. Based on observed means.

The error term is Mean Square(Error) = .000.

Appendix 38. 2. Effect of inlet temperature on bulk density content of green tea powder BulkDensity

Duncan Tempera

ture N

Subset

1 2

220 C 6 .515228

200 C 6 .526033

180 C 6 .555093

Sig. .124 1.000

Means for groups in homogeneous subsets are displayed.

Based on observed means.

66

Appendix 38. 3. . Effect of interaction between feed concentration and inlet temperature on

bulk density content of green tea powder BulkDensity

Duncan

trt N

Subset

1 2 3 4 5 6

9 2 .427252

8 2 .475237

4 2 .514331

6 2 .531950 .531950

2 2 .543276 .543276

7 2 .546669 .546669

5 2 .559587

3 2 .586483

1 2 .604278

Sig. 1.000 1.000 .144 .233 .190 .141

Means for groups in homogeneous subsets are displayed. Based on observed means.