Influence of drying methods on the content of bioactive compounds and antioxidant activities of curculigo orchioides

MINISTRY OF EDUCATION AND TRAINING
HO CHI MINH CITY UNIVERSITY OF TECHNOLOGY AND EDUCATION
FACULTY FOR HIGH QUALITY TRAINING

CAPSTONE PROJECT
FOOD TECHNOLOGY

INFLUENCE OF DRYING METHODS ON THE CONTENT
OF BIOACTIVE COMPOUNDS AND ANTIOXIDANT
ACTIVITIES OF CURCULIGO ORCHIOIDES

LECTURER: PhD. HOANG VAN CHUYEN
STUDENT: PHAM TRUONG TIEU LAM

SKL 0 0 8 4 4 9

Ho Chi Minh City, December, 2021


HO CHI MINH CITY UNIVERSITY OF TECHNOLOGY AND EDUCATION

FACULTY FOR HIGH QUALITY TRAINING

GRADUATION PROJECT
CODE: 2021-17116015

INFLUENCE OF DRYING METHODS ON THE
CONTENT OF BIOACTIVE COMPOUNDS AND
ANTIOXIDANT ACTIVITIES OF CURCULIGO
ORCHIOIDES

Major: FOOD TECHNOLOGY
Advisor: HOANG VAN CHUYEN, PhD.
PHAM TRUONG TIEU LAM
Student ID: 17116015

Ho Chi Minh City, December 2021


GRADUATION PROJECT ASSIGNMENT

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DISCLAIMER

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ACKNOWLEDGEMENTS
I sincerely thank the supervisor Hoang Van Chuyen who has taught me during the
process of studying. He guided me enthusiastically during my study, he also trained me for
theorical knowledge and laboraty skills. He was also carefully told us when receiving
samples and how to prepare it; moreover, he supported us with the experimental supplies
and materials to help us complete the thesis better.
Besides, I would like to thank Ms. Tran Thi Thu Trang, who supported me in terms of
the facilities system and regulations in the laboratories. Moreover, I would like to express
my sincerest thanks to the lecturers, who have trained me step by step not only in
specialized knowledge but also self-study skills during my time at the university.
Finally, I would like to express my deep gratitude to my family, who have always
encouraged me from the time I worked on the research project until the time I finished the

experiments.

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ADVISOR’S EVALUATION SHEET

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PRE-DENFENSE EVALUATION SHEET

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EVALUATION SHEET OF DEFENSE COMMITTEE
MEMBER

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TABLE OF CONTENTS
GRADUATION PROJECT ASSIGNMENT ................................................................... i
DISCLAIMER ................................................................................................................... ii
ACKNOWLEDGEMENTS ............................................................................................. iii
ADVISOR’S EVALUATION SHEET ............................................................................ iv
PRE-DENFENSE EVALUATION SHEET ................................................................... vi
EVALUATION SHEET OF DEFENSE COMMITTEE MEMBER ......................... vii
TABLE OF CONTENTS .................................................................................................. x
LIST OF FIGURES ......................................................................................................... xii
LIST OF TABLES .......................................................................................................... xiii
LIST OF ACRONYMS .................................................................................................. xiv
ABSTRACT ..................................................................................................................... xv
CHAPTER 1: INTRODUCTION .................................................................................... 1
1.1. Research rationale ................................................................................................... 1
1.2. Research objectives ................................................................................................. 2
1.3. Research contents .................................................................................................... 2
1.4. Significance of the research results ....................................................................... 2
CHAPTER 2: LITERATURE REVIEW ........................................................................ 3
2.1. Overview of Curculigo orchioides .......................................................................... 3
2.2. Overview of Polyphenol .......................................................................................... 6
2.3. Overview of drying methods ................................................................................ 10
2.4. Literature review of of the studies on Curculigo orchioides .............................. 12
CHAPTER 3: MATERIALS AND METHODS ........................................................... 14
3.1. Materials, Chemicals and Equipments ............................................................... 14
3.2. Experimental design ............................................................................................. 18
3.3. Extraction of samples ............................................................................................ 20
3.4. Analytical methods ................................................................................................ 22
3.4.1. Determination of moisture content ............................................................... 22
3.4.2. Analysis total phenolic content ...................................................................... 23
3.4.3. Determination of DPPH free radical scavenging activity ........................... 25

3.4.4. Determination of ABTS free radical scavenging activity ........................... 27
3.4.5. Determination of Ferric reducing antioxidant power (FRAP) ................... 29
3.5. Statistical analysis ................................................................................................. 30
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CHAPTER 4: RESULTS AND DISCUSSION ............................................................. 31
4.1. Moisture content, TPC and antioxidant activities of the fresh Curculigo
orchioides rhizome (COR) ........................................................................................... 31
4.2. Effect of drying methods on the drying time and moisture content of the COR
slices with different thickness ...................................................................................... 31
4.3. Effect of drying methods on total phenolic content of the dried COR slices .. 35
4.4. Effect of drying methods on antioxidant activities ............................................ 38
4.4.1. Effect of drying methods on DPPH free radical scavenging capacity of the
dried COR slices........................................................................................................ 38
4.4.2. Effect of drying methods on ABTS free radical scavenging activity ......... 40
4.4.3. Effect of drying methods on Ferric reducing antioxidant power............... 42
CHAPTER 5: CONCLUSION AND RECOMMENDATIONS ................................. 45
5.1. Conclusion .............................................................................................................. 45
5.2. Recommendations ................................................................................................. 45
REFERENCES ................................................................................................................ 46
APPENDIX....................................................................................................................... 54

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LIST OF FIGURES
Figure 2.1. Curculigo orchioides plant [10] ....................................................................... 3
Figure 2.2. Curculigo orchioides tuberous root [11] .......................................................... 4
Figure 2.3. Cinamic acid and Benzoic acid derivatives ..................................................... 7

Figure 2.4. Chemical structure of Flavonoids .................................................................... 7
Figure 2.5. Chemical structure of Stilbenes ....................................................................... 8
Figure 2.6. Lignan biosynthesis .......................................................................................... 9
Figure 2.7. Examples of various lignan skeletal units ........................................................ 9
Figure 2.8. An example of Lignan molecule .................................................................... 10
Figure 3.1. Convective dryer (Memmert, Germany) ........................................................ 15
Figure 3.2. Vacuum dryer (Memmert, Germany) ............................................................ 15
Figure 3.3. Infrared drying (DS.IR – 03, HCMC University of Technology and
Education, Vietnam) .......................................................................................................... 16
Figure 3.4. Heat pump drying (cold drying system) (DSL – P – L – T – 02, HCMC
University of Technology and Education, Vietnam) ......................................................... 16
Figure 3.5. Analytical balance 10mg (LT 2200C, Precisa, Switzerland) ......................... 17
Figure 3.6. Analytical balance 1mg (LS3200C and LS320, Precisa, Switzerland) ......... 17
Figure 3.7. Powder grinder machine (China) ................................................................... 17
Figure 3.8. Spectrophotometers UV-Vis (UH 5300, Hitachi, Japan) ............................... 17
Figure 3.9. Diagram of the experimental design .............................................................. 18
Figure 3.10. Curculigo orchioides after cutting hairy roots ............................................. 19
Figure 3.11. Curculigo orchioides in size 2mm and size 5mm ........................................ 19
Figure 3.12. Diagram of extraction of samples ................................................................ 21
Figure 3.13. Reaction of evaluation of total Polyphenol content ..................................... 23
Figure 3.14. The experiment of determining total Polyphenol content ............................ 24
Figure 3.15. Reaction of DPPH free radicals scavenging ................................................ 26
Figure 3.16. The experiment of DPPH free radical scavenging ....................................... 26
Figure 3.17. Reaction of ABTS free radical scavenging .................................................. 28
Figure 3.18. The experiment of ABTS free radical scavenging ....................................... 28
Figure 3.19. Reaction of Ferric reducing antioxidant power............................................ 29
Figure 3.20. The experiment of Ferric reducing anitoxidant power ................................ 30
Figure 4.1. Effect of different drying methods on the moisture content of the dried COR
slices with the thickness of 2 mm (a) and 5mm (b)........................................................... 33
Figure 4.2. Effect of different drying methods on total phenolic content of the dried COR

slices with the thickness of 2 mm (a) and 5mm (b)........................................................... 36
Figure 4.3. Effect of different drying methods on DPPH free radical scavenging capacity
of the dried COR slices with the thickness of 2 mm (a) and 5mm (b) .............................. 39
Figure 4.4. Effect of different drying methods on ABTS radical scavenging capacity of
the dried COR slices with the thickness of 2 mm (a) and 5mm (b) .................................. 42
Figure 4.5. Effect of different drying methods on FRAP antioxidant capacity of the dried
COR slices with the thickness of 2 mm (a) and 5mm (b) ................................................. 44
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LIST OF TABLES
Table 2.1. Table of Taxonomical hierarchy is presented in ............................................... 3
Table 2.2. Quality of Curculigo orchioides crude drug samples collected from different
places ................................................................................................................................... 4
Table 3.1. Preparation of Gallic acid concentrations in the standard curve construction 24
Table 4.1. Characterization of the fresh Curculigo orchioides rhizome ........................... 31
Table 4.2. The require drying time for the 2mm and 5mm COR slices ........................... 32

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LIST OF ACRONYMS
COR: Curculigo orchioides rhizome
CD: convective drying
CD60: convective drying at 60oC
CD80: convective drying at 80oC
CD100: convective drying at 100oC
VD: vacuum drying
VD60: convective drying at 60oC
VD80: convective drying at 80oC

VD100: convective drying at 100oC
IRD: infrared drying
HPD: heat pump drying
SD: sun drying
TPC: total phenolic content
DPPH, ABTS: free radicals
FRAP: Ferric reducing antioxidant power

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ABSTRACT
Curculigo orchioides is one of the precious medicinal plants in mountainous areas of
the Northern Vietnam, its rhizome contains many crucial chemical compounds, especially
polyphenols and brings a variety of health benefits to the consumers. Currently, sun drying
is the most common method used for long-term preservation of Curculigo orchioides but
this drying method may cause severe loss of its bioactive ingredients. In this study, effects
of different drying methods on moisture content, total phenolic content and antioxidant
activity of Curculigo orchioides were investigated to determine the most suitable drying
method for this material. The results showed that the drying processes and material
thickness greatly impacted on total phenolic content and antioxidant powers of the dried
Curculigo orchioides. In particular, the infrared drying samples had the highest values of
TPC analysis (76.82% and 80.04%), DPPH radical scavenging activity (48.52% and
50.9%), ABTS antioxidant activity (67.31% and 67.94%), FRAP assay (47.02% and
50.45%) in order of layers 2mm and 5mm, while the convective drying at 100oC of DPPH
and ABTS tests, also the vacuum drying at 40oC of TPC and FRAP tests, were given to be
the weakest antioxidant capacity in this study. Overall, the infrared drying were found to
produce the finished medicinal products in good quality than the other thermal drying
techniques.

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CHAPTER 1: INTRODUCTION
1.1. Research rationale
Curculigo orchioides is a medicinal plant commonly grown in the mountainous
provinces in the North of Vietnam. According to the traditional experiences recorded in
the book “Vietnamese medicinal plants and remedies” [1]. Curculigo orchioides (COR),
also known as Sam Cau and Tien Mao, is a kidney tonic, a physiological enhancer, which
can be used as a decoction to drink or soak in wine. Additionally, it also has a number of
other benificial effects such as: anti-inflammatory activity, antiasthmatic activity,
hepatoprotective activity, anti-chronic fatigue syndrome… [2]. Curculigo orchioides has
been reported to be rich in bioactive ingredients through the research works [3], [4], [5].
The most bioactive group of COR is polyphenols, especially the phenolic glycosides,
which act as antioxidant agents to prevent the pro-oxidation process or biological oxidative
damages in the living cells [6].
Drying is one of the most convenient methods for preserving products that has
appeared for a long time, to prolong the shelf-life and retain the bioactive compositions
and beneficial nutrients of the raw materials for use in a long period [7]. Many drying
methods have been investigated and applied to preserve plant materials with different
structures, shapes and sizes in order to make sure the color, chemical compounds and
nutrients of the food products. The drying system for agricultural products is currently
being favored by many customers, playing an important role in the industries of food
processing, agricultural goods production, biological products, medicines, and even
exportation.
Although COR has been reported to be rich in valuable bioactive ingredients, it is
currently sun dried has been reported to be rich in bioactive ingredients for the long-term
preservation. However, sun drying may cause significant loss of its bioactive ingredients
and medicinal quality. Therefore in this study, the effects of different drying methods
including sun drying, convective drying, infrared drying, vacuum drying and heat pump

drying on drying time, moisture content, total phenolic content and antioxidant activities
of COR were investigated to determine the most suitable drying method for this medicinal
material. In addition, the influence of the thickness of the COR slices on the drying time,
physicochemical properties and antioxidant activities of the dried products was also
investigated.

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1.2. Research objectives
- Investigate the effect of thickness of the drying material on the drying time,
physicochemical properties and antioxidant activities of the dried products.
- Determine the most suitable drying method for drying of Curculigo orchioides rhizome
for maximizing the retaining of bioactive ingredients and antioxidant activities of the dried
COR.
1.3. Research contents
- Analysis of the moisture content, physicochemical properties and antioxidant activities
of the fresh COR.
- Investigate effects sun drying and the thickness of the COR slices on the drying time,
moisture content, total phenolic content and antioxidant activities of the dried COR.
- Investigate effects convective drying and the thickness of the COR slices on the drying
time, moisture content, total phenolic content and antioxidant activities of the dried COR.
- Investigate effects vacuum drying and the thickness of the COR slices on the drying time,
moisture content, total phenolic content and antioxidant activities of the dried COR.
- Investigate effects infrared drying and the thickness of the COR slices on the drying time,
moisture content, total phenolic content and antioxidant activities of the dried COR.
- Investigate effects heat pump drying and the thickness of the COR slices on the drying
time, moisture content, total phenolic content and antioxidant activities of the dried COR.
1.4. Significance of the research results
- The experimental results from the study of drying processes on the COR may be used as

the scientific basis for further studies on the drying processes at an industrial scale for COR
and other parts of this herbal plant as well as other herbs.
- The results from drying methods for COR on retaining beneficial bioactive characteristics
are useful knowledge for the development of functional foods and healthy pharmaceutical
products.

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CHAPTER 2: LITERATURE REVIEW
2.1. Overview of Curculigo orchioides
2.1.1. General introduction
- Curculigo orchioides rhizome has skin in black or brown color, white flesh and has a
distinctive aroma when be exposed to the sunlight. Ethnic minorities at the Northern
mountainous areas of Vietnam usually use this rootstock as a useful medicine so that it
called Sam in Vietnam, and the leaves are the same as Cau leaves in Vietnam and this plant
is named Sam Cau. In addition, it is also known as Tien Mao, Ngai Cau, Co Noc Lan [8].
- The scientific name of this plant is Curculigo orchioides, which belongs to
Amaryllidaceae family, with the English name Golden eye grass or Black Musale, the
Hindi name is Kali Musli [2], [9].
Table 2.1. Table of Taxonomical hierarchy is presented in [2]

Kingdom
Division
Class
Order
Family
Genus
Species

Plantae
Magnoliophyta
Monocotyledon
Liliales
Amaryllidacae
Curculigo
Orchioides

Figure 2.1. Curculigo orchioides plant as illustrated in [10]

- Parts used as medicine: Tuberous root or rhizome.
- Plant type: Herbal medicine.
2.1.2. Morphology
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- Curculigo orchioides, is a plant which stays a long time, the height of 20 – 30 cm. The
rhizome is cylindrical and long, straight, slender at the ends, with accessory roots which
are short or long, coarse blackish brown bark and white flesh [8].
- The leaves grow in clusters from the rhizome, folded like Cau leaves.
- There is a flower cluster growing on the leaves, the leaf sheath is large and long.
- Fruit capsule, oblong, about 1.2 - 1.5 cm long. Each fruit contains from 1 to 4 capsules.

Figure 2.2. Curculigo orchioides tuberous root [11]
Table 2.2. Quality of Curculigo orchioides crude drug samples collected from different places

Place (%)
Kottayam
Muvattupuzha
Ernakulam

Thrissur

Ash
(%)
2.92
5.02
4.90
3.93

Protein
(%)
4.90
4.94
4.73
5.36

Starch
(%)
15.17
34.08
17.41
23.49

Fibre
(%)
19.97
20.33
22.08
23.62

Na
(%)
0.784
0.790
0.756
0.857

P
(%)
0.101
0.110
0.076
0.180

K
(%)
0.479
0.838
0.903
0.725

Ca (%)
1.521
1.227
1.668
1.129

Source: M.Asif (2012) [3]

2.1.2. Distribution

Curculigo orchioides distributed mainly in tropical and subtropical Asia such as some
southern provinces of China, Laos, Vietnam and a few other countries in Southeast Asia
area.
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This is a plant that likes light and moisture, especially can tolerate shade, so they are
often found in the fertile soils of the valley, along the fields or at the foot of limestone
mountains in the northern provinces of Vietnam such as Lai Chau, Tuyen Quang, Cao
Bang.
This species was listed less likely to be threatened by IUCN (The International Union
for Conservation of Nature and Natural Resources) in 1999, which was due to
overexploitation and destroy the habitat; moreover, it has also low quality of seed and
germination is significantly declined in wild plant density [5].
2.1.3. Medicinal effects and applications [9]
- According to traditional medicine, this herb is spicy and warm.
+ Reduce rheumatism, strengthen tendons.
+ Treat impotence and aphrodisiac.
+ Help with diuresis.
+ Treatment of cold in the back, hands and feet.
+ Additionally, Curculigo o. also cures hemorrhoids, abdominal pain, cough, jaundice.
- According to modern medicine, some studies have indicated that Curculigo o. has many
health benefits:
+ Boost immunity.
+ Be able to replace testosterone – male sex hormone because it contains steroids.
+ Support the body's ability to adapt to the environment without oxygen or in high
temperature condition.
+ Control heart activity, dilate coronary vessels, protect liver.
+ Anti-inflammatory, anti-fungal, against blood-related disorders.
+ Enhance muscle contraction, treat convulsions, relieve pain.

+ Anti-aging, improve skin.
+ Prevent diabetes, cancer.
- The dosage of the herb:
+ The usage level of adults: 5 - 10 gram.
+ From the ages of 5 and 16: one half of adult usage.
+ From 1 to 5 years old: a quarter of adult usage.
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2.2. Overview of Polyphenol
Polyphenol compounds are organic benzene rings binding one or more hydroxyl
functional groups [12] and were heat-labile substances [13].
There are over 8000 phenolic compounds found in most species of flora. These
compounds in plants originated in an intermediate substrate, phenylalanine, or a precursor
and shikimic acid. They manifest in a conjugated shape, have the linkage of one or more
sugar radicals to hydroxyl functional groups, in spite of direct bonds of sugars
(polysaccharide or monosaccharide) to an arene occurs [14].
It acts as preservatives, take n-propyl gallate as an example, it was approved by the
FDA [15].
Polyphenols can be composed of different groups according to the number of rings that
make up a phenol and the structural principles that bind them together. The main categories
of polyphenol below are phenolic acids, flavonoids, stilbenes and lignans.
2.2.1. Phenolic acids
The molecular structure of this group generally consists of only one carboxylic acid
group [16].
Phenolic acids usually exhibit colour, organoleptic qualities, nutrients and antioxidant
characteristics in foods [17].
Additionally, the food factories have conducted on the analysis on profile and dosage
of phenolic acids as well as their role on fruit repening, enzymatic control and have used
them as preservatives [18] [19] [20].

Currently, citations have pointed to high antioxidant content in fruits and vegetables
with inhibitory mechanism on diseases caused by oxidative damage such as cancer,
cerebral accident, coronary heart disease [21] [22].
Phenolic acids displace two derivatives: cinnamic and benzoic acid [23].
They are discovered in many species of plants: Cinnamic and benzoic acid derivatives
appear in most of edible plants (examples in fruits, vegetables, and grains) and are spread
in kernels, leaves, stems and roots of the plant [24] [18].
Phenolic acids are relatively resistant biochemical species; they undergo metabolism in
the soil for some microorganisms can consume them as carbon sources [25].
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Besides, the natural presence of phenolic acids usually occurs in dried fruit, horse gram
plants, mushroom Basidiomycetes and soil humus [26].

Figure 2.3. Cinamic acid and Benzoic acid derivatives

2.2.2. Flavonoids
Chemically, flavonoids are formed on 15 carbon frames, containing 2 aromatic rings,
A and B, connected through a pyran called C as an illustration below. [27].

Figure 2.4. Chemical structure of Flavonoids

Normally, flavonoids are synthesized at specific parts in plants and contribute to the
color and fragrance of flowers. For fruis, flavonoids play a role to attract pollinators and
thus helping with dispersal during seed and spore germination and seedling growth and
development [28]. Flavonoids include many subgroups such as: flavones, flavonols,
flavanones, flavanonols, flavanols or catechins, anthocyanins and chalcones [29].
2.2.3. Stilbenes

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Stilbenes have the structure of a 1,2-diphenylethylene nucleus with hydroxyls attached
to the aromatic hydrocarbons and appear as monomers or oligomers; beside that, most of
the compounds studied are trans-resveratrol [30].
Stilbenes are phytoalexins generated by plants in response to disease, injury or stress
symptoms [31].

Figure 2.5. Chemical structure of Stilbenes

Stilbene compounds were formed in many crops including sorghum (Sorghum bicolor),
peanut (Arachis hypogaea), grape wine (Vitis vinifera) and many trees such as
Pinus and Picea [32].
Commercially, sources of stilbenes include a variety of flora grown in parts of Asia as
folk drugs, are Polygonum cuspidatum, Rhodomyrtus tomentosa, Rheum undulatum,
Melaleuca leucadendron, and Euphorbia lagascae. For pterostilbene, which is mostly
discovered in blueberries, bilberries and other kinds in grapes and juice pulp. [33].
2.2.4. Lignans
The lignans are constituted of two or three mono-lignols at the C position of 6-3 and
the catalyst of the reaction is an oxidizing enzyme. At units of 6-3, a lignan is formed by
the dimerization, which is derived from cynamyl groups [34].

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