research for building the production process of soluble tea from anoectochilus setaceus blume

UNIVERSITY OF AGRICULTURE AND FORESTRY NGUYEN THI HONG HANH RESEARCH FOR BUILDING THE PRODUCTION PROCESS OF SOLUBLE TEA FROM ANOECTOCHILUS SETACEUS BLUME BACHELOR THESIS Study Mode : Ful

UNIVERSITY OF AGRICULTURE AND FORESTRY

NGUYEN THI HONG HANH

RESEARCH FOR BUILDING THE PRODUCTION PROCESS OF

SOLUBLE TEA FROM ANOECTOCHILUS SETACEUS BLUME

BACHELOR THESIS

Study Mode : Full-time

Major : Food Technology

Faculty : Advanced Education Program Batch : K48

Thai Nguyen, September 2021

DOCUMENTATION PAGE WITH ABSTRACT

Thai Nguyen University of Agriculture and Forestry

Thesis Title Research for building the production process of soluble tea

from Anoectochilus setaceus Blume

Supervisor’s Signature

Abstract:

The report aims to analyze the initial chemical composition of

Anoectochilus setaceus Blume and create instant tea products with Flanovoid s

extract from Anoectochilus setaceus Blume Thereby creating a premise for

establishing a process for extracting beneficial bioactive extracts from plants and

further studying the composition and biological activities of Anoectochilus setaceus Blume The initial chemistry of Anoectochilus setaceus Blume underwent 3

experiments: moisture determination by drying; Determination of ash content and quantification of total Flanovoids after extraction For the extraction process of total

Flanovoids from Anoectochilus setaceus Blume, four experiments were used: study

of solvent selection, time, temperature and selection of Flanovoid s extraction materials Finally, will study to create instant tea products from total Flanovoid s

extract from Anoectochilus setaceus Blume After a period of research, a nu mber of

results have been obtained: the factors of temperature, time, and proportion of raw materials all affect the Flanovoids content 60% ethanol solvent, temperature 50, time 90 minutes and solvent content 1/15 (g/ml) give the best performance for extr action of Flanovoids-rich fractions

Keywords:

Anoectochilus setaceus Blume

Flanovoids Temperature Time

Solvent Material Number of pages: 64

Date of Submission:

1.12.2021

ACKNOWLEDGEMENT

First of all, I would like to thank the Board of Directors of Thai Nguyen University of Agriculture and Forestry, related departments, Office of Advanced Programs of Thai Nguyen University of Agriculture and Forestry for teaching and guiding me to have knowledge today In order to complete this graduation thesis, besides my own efforts I have still received much help from individuals and groups I would like to express my deep gratitude to Master Dinh Thi Kim Hoa- Faculty of Biotechnology - Food Technology, who enthusiastically instructed and created the best conditions for me to complete this graduation thesis I also would like to express my sincere thanks to the teachers of Thai Nguyen University of Agriculture and Forestry for helping me to complete this graduation thesis I would like to sincerely thank you for your help in the group of students who interns at the laboratory of the Faculty of Biotechnology - Technology and Technology for helping me throughout the project implementation I would like to thank the Faculty of Biotechnology - Biotechnology for providing the internship place for me to complete my graduation thesis Despite great efforts, the graduatation thesis may be unable to avoid its shortcomings I hope to receive your valuable sharing an d comments from teachers and friends

Thai Nguyen, September 2021

NGUYEN THI HONG HANH

CHAPTER II LITERATURE REVIEW 7

2.1 Anoectochilus setaceus Blume Overview 7

2.1.1 Origin and botanical characteristics of Anoectochilus setaceus Blume 7

2.1.2 Biological characteristics 9

2.1.3 Chemical composition of Anoectochilus setaceus Blume 9

2.1.4 Material production area of Anoectochilus setaceus Blume 14

2.1.5 The value of Anoectochilus setaceus Blume 14

2.1.6 Some products from Anoectochilus setaceus Blume 17

2.3 Overview of the extraction process of biologically active

substances from plants 19

2.3.1 Definition of extraction process 19

2.3.2 Extraction Methods 19

2.3.3 Factors affecting the extraction process 20

2.4 Overview of soluble tea products 21

2.4.1 The classification definition of tea products 21

2.4.2 Some production process of tea product 27

2.4.3 Materials for the production of soluble tea 29

2.4.4 Some soluble tea products on the market 29

CHAPTER III METHODOLOGY 31

3.1 Object and study scope 31

3.1.1 Research Object 31

3.1.2 Tool, equipment and research chemicals 32

3.1.3 Location and time of study 33

3.2 Research contents 33

3.2.1 Analysis of the initial chemical composition of materials 33

3.2.2.Study on extraction process of total polysaccharide from

Anoectochilus setaceus Blume 33

3.2.3 Research on creating instant tea products from total Flavonoids

extraced from Anoectochilus setaceus Blume 34

3.3 Research Methods 34

3.3.1 Chemical composition analysis of materials 34

3.3.2 Study on extraction process of total Flavonoids from Anoectochilus

setaceus Blume 37

3.3.3 Research on creating instant tea product from total Flavonoids

extracted from Anoectochilus setaceus Blume 41

3.3.4 Sensory evaluation method 45

3.4 Data Analyzing Method 45

CHAPTER IV RESULT AND DISCUSSION 46

4.1 Analytical results of initial chemical compositions of raw material Anoectochilus setaceus Blume 46

4.2 Research results on extraction process of total Flavonoids from Anoectochilus setaceus Blume 47

4.2.1 Research results on the selection of solvents for extraction of

Flavonoids from Anoectochilus setaceus Blume 47

4.2.2 Results of study on extraction time of Flavonoids from

Anoectochilus setaceus Blume 48

4.2.3 Research results of extraction temperature on the total amount of Flavonoids 50

4.2.4 Results of the study on selection of material/solvent ratio on

the extraction efficiency of Flavonoids in Anoectochilus setaceus Blume 52

4.3 Research results on selecting the ratio of auxiliary materials for

producing instant tea from Anoectochilus setaceus Blume 54

4.4 Preliminary cost calculation for 100g soluble tea product from

Anoectochilus setaceus Blume 57

CHAPTER V CONCLUSION AND RECOMENDATION 58

5.1 Conclusion 58 5.2 Recomendation 59 REFERENCES 60 APPENDIX

LIST OF FIGURES

Figure 2.1 Anoectochilus setaceus Blume 8

Figure 2.2: Cardocorz tablets for health protection 18

Figure 2.3: Pictures of black tea 22

Figure 2.4: Pictures of green tea 23

Figure 2.5: Pictures of oolong tea 24

Figure 2.6: Pictures of herbal teas 25

Figure 2.7: Pictures of tea bag 25

Figure 2.8: Technological process of producing instant balck tea 27

Figure 2.9: Technological process of producing green tea bottle 28

Figure 2.10: Pictures of some soluble tea products 30

Figure 3.1: Anoectochilus setaceus Blume Plant 31

Figure 3.2: Planned production process of soluble tea from

Anoectochilus setaceus Blume Plant 42

Figure 4.1: Graph showing the effect of solvent concentration on the extraction efficiency of Flavonoids in Anoectochilus setaceus Blume 48

Figure 4.2: Graph showing the effect of extraction time of Flavonoids from Anoectochilus setaceus Blume 49

Figure 4.3: Graph showing the effect of temperature on extraction efficiency of total Flavonoids obtained 51

Figure 4.4: Graph showing the effect of raw materials/solvent on the extraction efficiency of Flavonoids in Anoectochilus setaceus Blume 53

LIST OF TABLES

Table 3.1: Tool, equipment and research chemicals 32Table 4.1 Research results on the effect of solvent selection on the

extraction efficiency of Flavonoids in Anoectochilus setaceus Blume 47

Table 4.2 Effect of extraction time in extractiong total Flavonoids from

Anoectochilus setaceus Blume 49

Table 4.3 Effect of temperature on extraction efficiency of total

Flavonoids obtained 51Table 4.4 Effect of material/solvent ratio on extraction efficiency

of Flavonoids in Anoectochilus setaceus Blume 52

Table 4.5 Mixing ratio to produce soluble tea from

Anoectochilus setaceus Blume 54

Table 4.6 Results of sensory evaluation scores 56

CHAPTER I INTRODUCTION 1.1 Rationale

Anoectochilus setaceus Blume is also known as Orchid

(Orchidaceae), Orchidaceae (Orchidales) Orchid crawls on the ground 10 20cm high, the young part is sparsely hairy Leaves oval or ovate, rounded at base, blade 3-4cm long, 2-3cm wide, upper surface dark brown with yellow streaks in the middle and pale pink on veins, light brown underside, petiole 1-2cm long, wide at the base to form a sheath hugging the body The inflorescence is 5-7cm long, with 5-10 large pink flowers (about 2.5cm long) The lip is 15mm long, bearing 6-8 whiskers on each side, the tip of the lip is split into 2 oblong lobes Gourds 13mm long, sparsely hairy Flowering season in October-December Regeneration mainly by shoots of rhizomes Externally, the stems and leaves are purple, with 3 to 5 vertical stripes on each leaf Orchid Kim Tuyen lives on limestone slopes, along streams, under large trees in humid forests at an altitude of 500-1600m

-Around the world, Anoectochilus setaceus Blume is distributed in

China (Yunnan, Guangdong), India, and Indonesia In Vietnam, they are distributed narrowly in a number of localities such as Hai Phong, Lao Cai,

Tam Dao, Quang Tri, Kon Tum, Gia Lai, Son La, Yen Bai, Anoectochilus

setaceus Blume grows scatteredly but in numbers In each place, there are

not many and they are over-exploited (in the form of cutting down the

whole tree) to export across the border to China (under the name Kim

Tuyen) Anoectochilus setaceus Blume is known to have very high

economic value (1.5-2.5 million VND/kg) Especially in some provinces near the border, they have been hunted, searched, exploited, sold to private

traders to buy and export abroad This puts Anoectochilus setaceus Blume

species in the wild at risk of depletion and extinction if urgent measures are not taken for effective, sustainable exploitation and effective conservation

Anoectochilus setaceus Blume is a rare genetic resource classified in

group IA: Forest plants and animals prohibited from exploitation and use for commercial purposes of Decree 32/2006/ND-CP on the management of forest plants endangered and rare forest animals In the Vietnam Red Book - Forest Plants section 2007, this orchid is classified as endangered: EN A1a,c,d

1.2 Research Objectives

1.2.1 General Objectives

Researching the production process of soluble tea from

Anoectochilus setaceus Blume has the effect of supporting the treatment

of cardiovascular diseases, being convenient, safe in use and contributing to the diversification of tea products on the market

1.2.2 Specific Objectives

‒ Analysis of the initial chemical composition of materials

‒ Study on extraction process of total Flavonoids from Anoectochilus

setaceus Blume

‒ Research on creating soluble tea products from total Flavonoid

extracts from Anoectochilus setaceus Blume

1.3 Research Value

1.3.1 Scientific Value

The research topic was to provide optimal parameters for the

extraction of total Flanovoid from Anoectochilus setaceus Blume, initially to evaluate the heavy metal toxins present in the extract from Anoectochilus

setaceus Blume This is the original scientific basis for further studies on

the composition and biological activities of Anoectochilus setaceus Blume

Seeking the right parameters for the production process of instant tea

from Anoectochilus setaceus Blume at the laboratory scale

From the results of research as well as recommendations will be the basis for future research

1.3.2 Practical Value

Creating a basis for establishing a process to extract beneficial

biologically active extracts from Anoectochilus setaceus Blume

Diversifying tea products on the market, improving the use value of

Anoectochilus setaceus Blume

CHAPTER II

LITERATURE REVIEW

2.1 Anoectochilus setaceus Blume Overview

2.1.1 Origin and botanical characteristics of Anoectochilus setaceus Blume

Anoectochilus setaceus Blume is the scientific name, with

synonyms such as: Chrysobaphus roxburghii Wall 1826; Anoectochilus

roxburghii Wall ex Lindl 1840 and Anoectochilus regalis Blume 1858

It is a herbaceous plant, growing in soil with long stems and roots There are many soft hairs on the body The plant has 2-4 leaves that grow close to the ground Leaves are ovate, nearly rounded at base, slightly p ointed and short-nosed, varying in color with usually lighter veins (dark green with light green veins or brown-red with yellow-green veins) or pink); petiole 2-3 cm long Inflorescence 10-15 cm long, bearing 4 - 10 flowers sparsely The bracts are ovate, with a sharp tip, 8 -10mm long, pink Flowers are usually white, 2.5-3 cm long; perianth pieces about 6mm long; Lips up to 1.5 cm long, 6-8 narrow bands on each side of base, broad apex, deeply bisected, bile cavity 7mm long, gourd 1.3cm long, green, with many soft hairs

Anoectochilus regalis Blume has a flower season in

February-April Regeneration by shoots from rhizomes and seeds, few and very slow growth Grows under the forest canopy, on the slopes of granite,

riolite, and shale rocks, at an altitude of 500 - 1600m, scattered in groups of a few trees on moist soil, rich in humus and fallen leaves [1]

Science name: Anoectochilus setaceus Blume

Gender: Plantae Ordo: Asparagales Familia: Orchidaceae Subfamilia: Orchidoideae Tribus: Cranichideae Subtribus: Goodyerinae Genus: Anoectochilus Species: A setaceus

Figure 2.1 Anoectochilus setaceus Blume

2.1.2 Biological characteristics

It is a herbaceous plant that grows in soil with long stems and roots There are many soft hairs on the body The plant has 2-4 leaves that grow close to the ground Leaves are ovate, nearly rounded at base, slightly pointed and short-nosed, varying in color with usually lighter veins (dark green with light green veins or brown-red with yellow-green veins) or pink); petiole 2-3 cm long Inflorescence 10-15 cm long, bearing 4 - 10 flowers scattered The bracts are ovate, with a sharp spike, 8-10 mm long, pink Flowers are usually white, 2.5-3 cm long; perianth pieces about 6 mm long; Lips up to 1.5 cm long, 6-8 narrow bands on each side of base, broad apex, deeply bisected, bile cavity 7 mm long, gourd 1.3 cm long, green, with many soft hairs

Anoectochilus regalis Blume has a flowering season in

February-April Regeneration by shoots from rhizomes and seeds, few and very slow growth Grows under the forest canopy, on the slopes of granite, riolite, and shale rocks, at an altitude of 500 - 1600 m, scattered in groups of a few trees on moist soil, rich in humus and fallen leaves [1]

2.1.3 Chemical composition of Anoectochilus setaceus Blume

Modern analytical methods today have allowed to determine the chemical composition as well as precious active ingredients with

extremely important medicinal effects in Anoectochilus setaceus Blume,

including: Flavonoid group (Quercetin, Isorhamnetin), group Steroids,

Polysaccharides 13.32%, Oligosacharides 11.24% and simple sugars 9.73%, alkaloids and many other trace elements and minerals [2,3,12,13,15]

From the ethanolic extract of Anoectochilus roxburghii (Wall,

Lindl) identified eight compounds isolated from ethyl acetate and n butanol soluble fractions Based on spectroscopic methods, 8 compounds have been identified including: quercetin-7-O-β-D-[6''-O-(trans-feruloyl)]-glucopyranoside (compound 1), 8-Cp- Hydroxybenzylquercetin (compound 2), isorhamnetin-7-O-β-Dglucopyranoside (compound 3), isorhamnetin-3-O-β-D- glucopyranoside (compound 4), kaempferol-3-O-β-Dglucopyranoside (compound 4) substance 5), kaempferol-7-O-β-D-glucopyranoside (compound 6), 5-hydroxy-3',4',7- trimethoxyflavonol-3-O-β-D-rutinoside (compound 7), and isorhamnetin-3-O-β-D-rutinoside (compound 8) Among them quercetin-7-O-β-D-[6''-O-(trans-feruloyl)]-glucopyranoside (compound 1) is a yellow amorphous solid that gives positive test results Molish and Mg-HCl The molecular formula of compound 1 has the form C31H28O15, the UV spectrum in methanol shows characteristic absorption at 372 nm (band 1) and at 333 nm and 252 nm (band 2) After hydrolysis in acid, compound 1 obtained D -glucozo and querctin by thin plate chromatography The properties of quercetin-7-O-β-D-[6''-O-(trans-feruloyl)]-glucopyranoside also indicate that it is a flavonoid glucoside with potent biological activity and is

-expressed through reacts with 1,1-diphenyl-2-picrylhydrazyl free radical with IC50 value reaching 31 g/ml [3]

The anti-inflammatory and isulin resistance characteristics are also

investigated with aqueous extract of Anoectochilus setaceus Blume The

extraction process was carried out at 121oC for 30 minutes according to the method of Kim & Jang (2011) The study showed the antioxidant capacity of the extract through DPPH free radical reaction up to 70% (P<0.05) with an IC50 value of 3.39 ± 0.23 mg/ml Similar to ABTS free radicals, IC50 value reached 0.38 ± 0.02 mg/mL The above results show that the extract of Lan Kim Tuyen has antioxidant and anti-inflammatory properties The insulin resistance activity was also demonstrated in the glucose uptake during tumor treatment (TNF)-a with 3T3-L1 In addition, extracts at concentrations up to 200 g/ml were not toxic to human peripheral blood mononuclear cells (PBMCs) and were not mutagenic [13]

In particular, quercetin is one of the Flavonoids that have the function of improving heart health, reducing cholesterol, and preventing atherosclerosis Prevent stroke and some related cardiovascular diseases [6,7,16] In addition, it is also a very good active ingredient for patients with diabetes and high blood pressure With its anti-inflammatory ability as well as the ability to stabilize blood vessels against aging vascular

accidents [11,13] The study showed the effect of Quercetin on J774 macrophage cells with IC50 value ̴ 25µM

Quercetin also acts as an effective cancer prevention and treatment

agent, the high curative activity of Anoectochilus formosanus is due to

its secondary metabolites, especially the main kinsenosides and

Flavonoids [5] Its effects in Anoectochilus setaceus Blume have

confirmed the role of disease prevention and treatment of this medicinal plant In the study of the authors Fu-sheng Zang et al (2013) investigated the promoting role of F-23 fungus (genus Mycena) on the biomass and

content of kinsenosides and Flavonoids present in Anoectochilus

setaceus Blume Research results showed that after 10 weeks of

symbiotic cultivation, shoot height, dry weight and leaf count increased by 16.6%, 31.3% and 22.5%, respectively; increased the content of kinsenosides, isorhamnetin-3-O-β-D-rutinoside, and isorhamnetin-3-O-β-D-glucopyranoside by 85.5%, 226.1% and 196.0%, respectively; some hyphae in epidermal cells are stained red and/or reddish brown by safranine; and, significantly reduced the number of starch granules in cortical cells Furthermore, F-23 mushroom significantly improved the

kinsenoside and Flavonoid content of Anoectochilus setaceus Blume

These findings support reports that endophytes can alter the p roduction of secondary metabolites in their plant hosts, although physiological, genetic, and ecological analyzes are still warranted guaranteed

Meanwhile, another active ingredient in the flavonoid group, isorhamnetin, has the ability to prevent damage to cells in the body, acting as a powerful antioxidant Isorhamnetin is considered an anti-cancer agent because of its ability to inhibit the growt h of cancer cells [9,10] According to the author Le Dinh Chac (2019), the dry extract of

Anoectochilus setaceus Blume obtained from Vietnam was first made by

fermentation from yeast Saccharomyces strain cerevisiae and using high performance liquid chromatography method HPLC) to analyze the content of several compounds in this dry extract The results of the study determined that the quercetin, isorhamnetin and ferulic acid content in

the dry extract of Anoectochilus setaceus Blume produced by

fermentation, the quercetin content ranged from 105.5 to 109.3 µg/ml, average is 106.8 µg/ml; isorhamnetin content ranged from 180.1 to 191.8 µg/ml, averaged at 187.2 µg/ml and ferulic acid content ranged from 50.5 to 54.0 µg/ml, averaged 51 ,9 µg/ml This proves that the dry

extract of Anoectochilus setaceus Blume has very high pharmacological

effects in supporting the treatment of human diseases

In another study on the anti-proliferative effect of isorhamnetin on lung cancer cells in vitro and in vivo The results o f the study indicated that isorhamnetin markedly inhibited the growth of A549 cells with the induction of apoptotic changes Isorhamnetin at 20 μg/ml, can induce A549 cell apoptosis, upregulate the expression of the apoptosis genes

Bax, Caspase-3 and P53, and downregulate the expression of Bcl-2 protein, cyclinD1 and PCNA Tumors in isorhamnetin-treated tumor-bearing mice were significantly smaller than in the control group Outcomes of apoptosis-related genes, PCNA, cyclinD1 and other protein expression levels of transplanted Lewis cells were the same as those of A549 cells in vitro [10]

This is also a valuable active ingredient that plays an important role in the prevention and treatment of diabetes and its complications [2,7,11,13]

2.1.4 Material production area of Anoectochilus setaceus Blume

Domestically, Anoectochilus setaceus Blume is usually distributed

in Lao Cai (Sapa: Phang Xi Pang, Van Ban: Liem Phu), Ha Tinh (Huong Son: Rao An), Quang Tri, Kontum (Dak Glei: Ngoc Linh mountain, Sa Thay: Chu Mom Ray mountain), Dak Lak (Krong Bong: Chu Yang Sinh mountain), Lam Dong (Lac Duong: Bi Dop mountain) In the world, it is mainly cultivated in India, Nepal, Butan, China, Myanmar, Thailand, Laos, Cambodia, Malaysia, Indonesia

2.1.5 The value of Anoectochilus setaceus Blume

Anoectochilus setaceus Blume is a valuable medicinal herb used in

traditional remedies of ethnic minority communities in Vietnam The species of orchid have a wide distribution, but the number of individuals in the wild is not much due to the slow regeneration ability, requiring

strict living conditions, in addition, Anoectochilus setaceus Blume is

being over-exploited due to its many medicinal properties precious Facing that situation, scientists have now propagated this plant by seeds, by seedlings, by cuttings However, these methods are simple but have

many limitations such as Anoectochilus setaceus Blume in the wild,

often rare, the number of seeds is very small, or the cutting process takes a long time, not to mention the quality of the seedlings is not high or damaged pests In addition to the above measures, there is another propagation method, which is in vitro propagation, which is the p roduct of plant cell technology [14] The study was conducted to build an in

vitro multiplication process of Anoectochilus setaceus Blume species

through protocorm-like bodies (PLBs) with the aim of contributing to the conservation of rare orchid species The rate of induction of PLBs was 86.25% with the sample source being a cross section of the shoot body The mean fresh weight of PLBs after 30 days of culture was 0.23 grams Clusters of PLBs were grown on phylogenetic and shoot growth media in 30 days The formed shoots are separated and moved to the rooting stage The rooting process consists of 2 stages, each stage lasts for 30 days After 60 days, 100% of shoots produced roots with an average of 3.20±0.67 roots/bud, the average plant height was 4.57±0.85 cm with 4.21±0.42 leaves/plant The shoot multiplier of the whole procedure was 29.66±4.04 buds/cluster of PLBs Seedlings obtained from the procedure

have a high survival rate (80%-100%), seedlings grow normally when transferred to the nursery With the ability to multiply quickly under sterile conditions and the ability to provide a large number of seedlings to meet the needs of expanding this species

In parallel with the expansion of planting Anoectochilus setaceus

Blume, the application of scientific and technical advances to improve

the value of special medicinal products of Anoectochilus setaceus Blume

is also interested by domestic and foreign scientists so many, so much Among them, the extraction of high fractions rich in Flavonoid

compounds from the biomass of Anoectochilus setaceus Blume tree

creates the premise for the production of a number of functional products to take care of human health, which is currently being studied by scientists Studying in the country places a lot of attention

According to a study by Do Thi Gam (2017) [4] Extracting and

quantifying total Flavonoids from 3 species of Anoectochilus setaceus

Blume of Vietnam, the results showed that total Flavonoid content was

highest in Anoectochilus roxburghii (Wall.) Lindl (1.345%), then

Anoectochilus lylei Rolfe ex Downiex (1.044%) and the lowest species

Anoectochilus aff anamensis Aver (0.903%) Flavonoid compounds are

mainly accumulated in the leaves of Anoectochilus setaceus Blume

Conducting thin layer chromatography of total Flavonoid preparations

obtained from Anoectochilus setaceus Blume species in the solvent

system Ethylaxetat: Toluene: Formic acid: Water = 7:3:1.5:1 (v:v:v:v) From 9-14 components were found, the components separated clearly and separately and had the characteristic qualitative characteristics of the Flavonoid group Flavonoid compounds extracted from 3 species of Orchids all showed good antioxidant activity through indigocarmin oxidation reaction by enzyme peroxidase on blood group O Antioxidant

order of species Anoectochilus setaceus Blume in the study as follows: following: Anoectochilus roxburghii (Wall.) Lindl > Anoectochilus lylei Rolfe ex Downiex > Anoectochilus aff anamensis Aver

According to a study by Le Dinh Chac (2019) [9] on the chemical composition and role of quercetin, isorhamnetin and ferulic acid in

supporting human health improvement in Anoectochilus setaceus Blume

in Vietnam The results showed that the content of quercetin, isorhamnetin and ferulic acid were 106.8 µg, 187.2 µg and 51.9 µg, respectively, in the dry extract

2.1.6 Some products from Anoectochilus setaceus Blume

Anoectochilus setaceus Blume is a very rare medicinal plant

Anoectochilus setaceus Blume has been included in the Vietnam Red Book,

is an orchid that needs to be protected in the wild Besides, Anoectochilus

setaceus Blume is an extremely valuable material to support the treatment

of human diseases

Figure 2.2: Cardocorz tablets for health protection

[Source: https://cardocorz.vn/cardocorz.html]

For cancer patients: 25g fresh brocade orchid, 35g black musk

Wash and drink water daily

Anoectochilus setaceus Blume treatment of liver disease: 25g fresh

brocade orchids, 35g vines Wash and drink water daily to help detoxify and treat hepatitis and cirrhosis

Treatment of diabetes: 25g fresh brocade orchid, drinking water after

meals

Treatment of depression and insomnia: 25g fresh brocade orchid,

15g dry paint, 8g lotus heart, 20g resolution, 8g ground licorice, 10g natural flower, 12g jasmine flower, 10g black ginseng, 8g ox sock Sac drink within 1 month Drink continuously after 1 week will feel the effect

2.3 Overview of the extraction process of biologically active substances from plants

2.3.1 Definition of extraction process

Extraction is the process of selectively dissolving one or more components present in a sample of raw materials by exposing the material to a solvent The driving force of the extraction process is the difference in the concentration of the component in the raw material and in the solvent [1] [13]

2.3.2 Extraction Methods

Currently, there are two main extraction methods: liquid -liquid extraction method and liquid-solid extraction method Liquid ‒ solid extraction is the process of separating one or several substances dissolved in a solid by another liquid called a solvent Because the purpose of the study is to extract biologically active substances of Orchidaceae, namely Flavonoids, the method of liquid ‒ solid extraction will be p resented For this method the following requirements must be met:

Solvent requirements: Solvents must be selectively soluble, non-toxic, non-corrosive to equipment, cheap and readily available

Dissolution mechanism: Solvent penetrates through the capillaries into the medicinal cells, the permeation time depends on the diameter, length of the capillary, the nature of the solvent, etc The dissolution process depends on the chemical nature of solutes and solvents [1][13]

2.3.3 Factors affecting the extraction process

- The size of the material: The smaller the size of the material, the larger the contact surface area between the material and the solvent Therefore, it becomes easier to extract the components from the raw materials into the solvent However, if the size of the material is too small, the cost of the process [16]

- The degree of disruption of the material's cell structure: It is the basic factor that promotes the rapid and complete extraction process, creating conditions for the material to be fully exposed to the solvent [16]

- Extraction powder temperature: When the temperature is increased, the diffusion process will be enhanced because the viscosity of the oil in the raw material decreases, which increases the velocity of the oil into the solvent However, the temperature rise must also have a certain limit, if the temperature is too high, it will cause a lot of solvent loss and cause oil denaturation

- Moisture of extracted powder: Increasing the amount of moisture will slow down the diffusion process and increase the cohesion of extracted powder particles because moisture in the extracted powder will interact with proteins and other hydrophilic substances to prevent permeation The depth of solvent into the interior of the extracted p owder p articles slows down the diffusion process

- Velocity: The movement of the solvent in the extracted powder layer affects the diffusion process Increasing the solvent movement speed will shorten the extraction time, thereby increasing the productivity of the device

- Solvent-to-material ratio: Affects extraction rate, the more p owder extracted, the more solvent is required However, the amount of solvent has a great influence on the device size [16]

2.4 Overview of soluble tea products

2.4.1 The classification definition of tea products

Tea is a universal beverage Wherever on earth, the human culture has blossomed, tea has been with them either as a medicinal drink or as a magical energiser In the present time, the medicinal, as well as magical aspects of the tea, are all disputed But, tea has been assimilated into the modern culture so well, that it is nowadays perceived as a kind of formality and necessity on every occasion

Tea gets its significant flavour and influential quality from three

metabolites in ‘Carmelia Sinensis’ leaves They are catechin, theanine and

caffeine Catechin is a flavonoid and a natural antioxidant Different levels of oxidization of tea leaves make different types of teas On the basis of preparation, teas are classified as given below:

Black Tea

Black tea is perhaps the most popular tea type Even though it is called ‘black’ tea, the prepared tea drink has a reddish colour, and thus it is known

in China as ‘hongcha’, which means ‘red tea’ Tea leaves are withered and

oxidized more than other types of tea leaves, to make black tea

Figure 2.3: Pictures of black tea

To make black tea, fresh tea leaves are sun-dried and crushed which follows a process of oxidization and then shaping it into rolls and finally

dried to make them fit for black tea

Green Tea

Green tea is perhaps the oldest tea ever existed in China To make

green tea, ‘Carmelia Sinensis’ leaves do not need to go through the

sophisticated oxidisation process Instead, fresh tea leaves are steamed, and

hand-rolled, which then are dried to make them suitable for brewing into green tea Green tea is said to have several health advantages, like weight loss, prevention of cancer, although none of these has been scientifically proved The only proven health benefit is that regular intake of green take can reduce the risk of death due to cardiovascular disease by 5%

The culture of green tea is widespread in East Asian countries In China, green tea leaves are shaped like curled up snails and are called ‘Biluochun’ Tea leaves shaped like gunpowder pellets are named ‘gunpowder tea’ Some other Chinese green tea varieties are Chun Mee, Maojian tea, Longjian, etc

Figure 2.4: Pictures of green tea Oolong Tea

The literal translation of ‘Oolong tea’ is ‘black dragon tea’ Oolong tea is the tea variety which undergoes the most complex procedure to prepare It requires the sun drying of fresh tea leaves, followed by semi-

oxidization, and then either panning or baking, and then rolling for shaping The rolled tea leaves dried and fired further to make oolong tea variety

In China, there are several styles of Oolong tea flavours based on regions, like Fujian, Guangdong, and Taiwan Oolong tea can also be made into different grades of flavours, by changing the level of oxidization.

Figure 2.5: Pictures of Oolong tea

Herbal Teas

Herbal teas, also known as herbal infusions, are typically a blend of herbs, flowers, spices, and dried fruit This blend of ingredients is then brewed in the same way as your favorite traditional tea, either loose or in tea sachets or bags By combining quality ingredients, blends can be created that calm, invigorate, or treat minor ailments Colors and flavors range from light and fruity to vibrant and spicy, to match your mood

Figure 2.6: Pictures of herbal teas

Beside, the classification above, the tea product are also devided to several types based on the technical features They are tea bag product, instant

product, tea bottle product

Tea bag

A tea bag, or the compound teabag, is a small, porous, sealed bag or packet, typically containing tea leaves or the leaves of other herbs, which is immersed in water to steep and make an infusion Originally used only for tea (Camellia sinensis), they are now made with other tisanes ("herbal teas") as well

Figure 2.7: Pictures of tea bag

Instant tea

Instant tea is a powder in which water is added, in order to r econstitute it into a cup of tea The earliest form of instant tea was developed in the United Kingdom in 1885  A patent was granted for a paste made of concentrated tea extract, sugar, and evaporated milk, which became tea when hot water was added However, no notable developments were made until spray drying technology allowed for drying the tea concentrates at a temperature which did not damage the flavors of the product

Tea bag products is a product in the dried powder form of extracts obtained from fresh tea leaves or from semi-finished teas

Tea is considered a popular drink in the world, ranking 2nd after water Tea originated in China and has been used as a drink since 2500 BC, then spread to Japan and other Asian countries

Along with the development of society and the trend of economic

integration, the tea processing industry in the country and the world has also changed in terms of types and improved in quality Many types of herbs have been used for processing into teas with special p roperties that support the health of consumers These products are put on the market in the form of teas such as jasmine tea, passion fruit tea, stevia tea, etc Due to the busy work pressure, many people have very little time and space to making tea so manufacturers have processed teas that are convenient Two types of breakthoughs inconvenient tea use are bags and instant tea [4]

2.4.2 Some production process of tea product

Figure 2.8: Technological process of producing instant balck tea

[Source: Blog - Zen Teahouse (weebly.com)]

Figure 2.9: Technological process of producing green tea bottle

[Source:

http://mnb.com.vn/vi/day-chuyen-san-xuat-nuoc-giai-khat-tra-xanh-dong-chai-2.4.3 Materials for the production of soluble tea

Maltodextrin filler is used in the mixing process before spray drying to increase the dry matter concentration of the tea after concentration, satisfying the alcohol concentration required for the spray drying process

Sweeteners participate in making one of the three main flavors of instant tea (sweet - sour - acrid)

Sour substances are used in tea to help tea have a mild sour taste when brewed In production, some food acids are often used: tartric acid, citric acid, In which, citric acid is most commonly used in production instant tea, thanks to its sour taste, pleasant peach aroma

Coloring agent: In the production of instant tea, caramel color is used to increase the sensory color of the water mixed with tea powder Caramel is a colorant formed during caramelization, which is intense at the melting point of sugar

Fragrances are used to obtain the desired characteristic aroma, often by combining several odorants together

2.4.4 Some soluble tea products on the market

Tam That herbal tea, Ha Thu O herbal tea, Ginger herbal tea, Atiso herbal tea, Lipton Ice Tea with peach flavor, robinson farm soluble tea with squash flavor, Cozy Lemon Flavored Instant Tea, In addition to helping refreshment, body heat, some herbal tea products also have the effect of nourishing or treating different diseases depending on the materials used If herbs are to be mixed together to produce instant tea, they must be pharmacologically compatible

Some pharmacological soluble tea products:

Soluble Ginger tea is used in the treatment of diseases such as colds, abdominal pain, digestive support, cough,

Soluble tea from Chamomile has the ability to calm stress, boost the immune system and is effective in the treatment of diabetes

Ha Thu O tea has the ability to support digestion, is used in the treatment of diabetes, hyperlipidemia, nerve tonic effect

Atiso tea helps to improve acne-prone, less-than-fresh skin, has a cooling effect, filters the liver, helps the body eliminate toxins effectively, and reduces

Figure 2.10: Pictures of some soluble tea products

CHAPTER III METHODOLOGY

3.1 Object and study scope

3.1.1 Research Object

Research Object: Anoectochilus setaceus Blume

Study scope: Research on production of instant tea from Anoectochilus setaceus Blume at laboratory scale

Figure 3.1: Anoectochilus setaceus Blume Plant

[Source: https://duoclieusontra.com/cay-lan-kim-tuyen/]

3.1.2 Tool, equipment and research chemicals

Table 3.1: Tool, equipment and research chemicals List of materials and equipment