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JFS C: Food Chemistry
Physicochemical Properties and Antioxidant
Capacity of 3 Polysaccharides fromGreen Tea,
Oolong Tea, and Black Tea
HAIXIA CHEN, ZHISHUANG QU, LINGLING FU, PENG DONG, AND XIN ZHANG
ABSTRACT: Three polysaccharide-rich fractions named GTPS, OTPS, and BTPS were isolated from green tea, oo-
long tea, and black tea, respectively. Chemical characteristics, glycosidase inhibitory effects, and antioxidant prop-
erties of the 3 fractions were compared. Monosaccharides of GTPS were composed of D-rhamnose, L-arabinose,
D-xylose, D-mannose, D-galactose, and D-glucose. But there were no xylose and mannose detected in OTPS and
BTPS. The molecular weight distributions were decreased from 9.2 to 251.5 KDa to 3.8 to 32.7 KDa with the fer-
mentation of the tea fromgreen tea to black tea. BTPS showed the highest -glucosidase inhibitory activity, antiox-
idant activities on hydroxyl radicals and DPPH radicals. The differences in antioxidant activities and glycosidase
inhibitory properties among the 3 polysaccharide-rich fractions appeared to be related to differences in monosac-
charide composition and molecular weight distribution of the polysaccharide.
PRACTICAL APPLICATION: Diabetes mellitus (DM) is one of the primary threats to human health due to its increas-
ing prevalence, chronic course, and disabling complications. Control of postprandial hyperglycemia and inhibition
of oxidative stress are suggested to be important in the treatment of diabetes. Many efforts had been made to search
for effective and safe -glucosidase inhibitors and antioxidants from natural materials to develop a physiological
functional food or lead compounds for curing diabetes. Coarse tea was used to cure diabetics in people in China and
Japan. The hypoglycemic activity increased with the contents of polysaccharide in coarse tea. Many studies have
focused on the hypoglycemic activities of tea polysaccharides, but little is known about the glycosidase inhibitory
effects of tea polysaccharide. The aim of this study was to find a tea polysaccharide with the best potential for ex-
ploitation in curing diabetes.
Keywords: antioxidant activities, composition, glycosidase inhibitory effects, tea polysaccharide
Introduction
D
iabetes mellitus (DM) is one of the primary threats to human
health due to its increasing prevalence, chronic course, and
disabling complications. Control of postprandial hyperglycemia
and inhibition of oxidative stress are suggested to be important in
the treatment of diabetes. Many efforts hadbeenmade to searchfor
effective and safe -glucosidase inhibitors and antioxidants from
natural materials to develop a physiological functional food or lead
compounds for curing diabetes (Hays and others 2008; Kwon and
others 2008).
Tea (Camellia sinensis L.) is the 2nd most consumed bever-
age in the world next to water. The tea plant has been widely
used for centuries by ancient cultures for its medicinal properties.
Tea is popularly consumed in unfermented (green tea), semifer-
mented (oolong tea), and fermented (black and pu-erh or red)
forms (Zhu and others 2002). The chemical composition of tea in-
cludes proteins, polysaccharides, polyphenols (catechins or flavan-
3-ols, theaflavins, thearubigins, and proanthocyanidins), chloro-
phyll, minerals and trace elements, volatile compounds, amino
and organic acids, lignins, and alkaloids (caffeine, theophylline,
and theobromine) (Seeram and others 2006). Tea was found to
MS 20090126 Submitted 2/12/2009, Accepted 5/4/2009. Authors are with
Tianjin Key Laboratory for Modern Drug Delivery & High-Efficiency,
School of Pharmaceutical Science and Technology, Tianjin Univ., Tian-
jin, 300072, P.R. China. Direct inquiries to author Chen (E-mail:
chennhxx@yahoo.com.cn).
have bioactivities including antioxidant (Yen and others 1997),
improving immune response (Bhattaxharyya and others 2004),
anti-atherosclerosis (Curin and Andriantsitohaina 2005), antihy-
pertension (Hodgson and others 2005), anti-infectious diseases
(Weber and others 2003), and antidiabetic properties (Anderson
and Polansky 2002) in recent studies. The chemical compositions
of the tea in different forms were different and induced the change
of the bioactivities (Yanagimoto and others 2003).
The control of postprandial hyperglycemia is critical in the early
therapy for diabetes. One therapeutic approach to decrease post-
prandial hyperglycemia is to retard absorption of glucose through
inhibition of carbohydrate hydrolyzing enzymes, for example, -
amylase and -glucosidase, in the digestive organs (Saito and oth-
ers 1998). In addition, several -glucosidases have been recently
screened and developed from natural sources (Lee and Lee 2001;
Kim and others 2004). Coarse tea was used to cure diabetics in
people in China and Japan. The hypoglycemic activity increased
with the contents of polysaccharide in coarse tea. Many studies
have been focused on the hypoglycemic activities of tea polysac-
charides (Chen and others 2005; Zhou and others 2007). But little
was known about the glycosidase inhibitory effects of tea polysac-
charide. In our previous studies, the chemical properties and the
antioxidant, hypoglycemic activities of the polysaccharide from
green tea were investigated (Chen and others 2004; Chen and oth-
ers 2008). But there was no report on the comparative study of the
composition and the bioactivities of the polysaccharides from dif-
ferent forms of tea. The objective of this study was to isolate the
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2009 Institute of Food Technologists
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Vol. 74, Nr. 6, 2009JOURNAL OF FOOD SCIENCE C469
doi: 10.1111/j.1750-3841.2009.01231.x
Further reproduction without permission is prohibited
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Physicochemical properties and antioxidant capacity. . .
polysaccharide-rich fraction from green tea, oolong tea, and black
tea, characterize the chemical composition, and evaluate the gly-
cosidase inhibitory effects and antioxidant activities of the 3 kinds
of tea polysaccharides (TPS).
Materials and Methods
Materials and chemicals
Green tea, oolong tea, and black tea were obtained from the
local tea market (They were produced by the Huangshan Moun-
tain Tea Factory, Anhui, China). -Glucosidase fromSaccharomyces
cerevisiae, p-Nitrophenyl -D-glucopyranoside as a synthetic sub-
strate of -glucosidase and pepsin were purchased from Sigma
Chemical Co. (St. Louis, Mo., U.S.A.). Trifuoroacetic acid (TFA)
1, 1- Diphenyl-2-picrylhydrazyl (DPPH), 2-deoxy-ribose, the stan-
dard monosaccharides (D-rhamnose, L-arabinose, D-galactose, D-
xylose, D-mannose, and D-glucose) and T-series Dextran standards
were obtained from China Sigma-Aldrich (Shanghai, China). All
other chemicals and reagents were purchased locally and were of
analytical grade.
Extraction and isolation of tea polysaccharide
Polysaccharides from green tea (GTPS), oolong tea (OTPS), and
black tea (BTPS) were obtained in the same procedure. Briefly, dry
meshed tea powders (100 g) were placed in a 3-L round-bottom
flask with 1.0 L of ethanol (80%, v/v) for 24 h to remove most of
the polyphenols and monosaccharide. After the supernatant was
removed, the residues were dried in air and then extracted with
hot water at 70

C for 60 min 3 times. The aqueous extracts were
concentrated and then precipitated with 4-fold volumes of 95%
ethanol. The precipitate that formed was collected by centrifuga-
tion at 3000 g for 10 min and repeatedly washed sequentially with
ethanol, acetone, and ether, respectively. The precipitate was dis-
solvedinhot water, andexcludedproteinwithSevag method(Staob
1965), and dialyzed against distilled water for 48 h with dialysis tub-
ing (molecular weight cut-off, 8000 Da) to remove low-molecular
weight matters, and then concentrated and precipitated with 4-
fold volumes of 95%ethanol to obtain the polysaccharide-enriched
fraction. The yield of tea polysaccharides from green tea, oolong
tea, and black tea were about 4.0, 4.6, and 4.2 g, respectively.
Composition analysis
Total sugar content was determined by the phenol-sulfuric
acid analysis using D-glucose as standard (Dubois and others
1956). Uronic acid content was determined by the carbazole-
sulfuric acid method using galacturonic acid as standard (Bitter
and Muir 1962). Protein was analyzed by the method of Brad-
ford (1976) using bovine serum albumin as the standard. Assay
of polyphenolic compounds in tea polysaccharide was conducted
according to Bonvehis method (Bonvehi and Coll 1997). The
composition of neutral monosaccharide was measured by gas
chromatography after converting them into acetylated derivatives
(Chaplin and Kennedy 1994). Briefly, 10 mg of polysaccharide were
hydrolyzed in a sealed glass tube with 2 Mtrifluoroacetic acid (TFA)
at 105

C for 10 h. The hydrolysate was evaporated to dryness.
The acid was removed under reduced pressure by repeated co-
evaporations withmethanol. The hydrolysates were thenconverted
into alditol acetates according to conventional procedures. Gas
chromatography was performed on a Shimadzu GC-14B instru-
ment with OV-1701capillary column (30 m 0.32 mm 0.5 m).
The temperature program was set to increase from 150 to 240

C at
an increment of 5

C/min and N
2
was the carrier gas. The standard
monosaccharides were used as references and they were measured
following the same procedure as the sample.
Determination of the molecular weight of tea
polysaccharide
For molecular weight determination of tea polysaccharides, high
performance gel permeation chromatography was performed on a
Shimadzu LC-20A chromatograph equipped with a Shodex OH pak
SB-804 column(7.8 300 mm; Showa Denko, Kawasaki, Japan) and
an RI detector (Polymer Lab. Ltd., Tokyo, Japan). Twenty microliters
of the polysaccharide were injected and eluted at a buffer (0.2 M
NaAc-AcOH, pH 6.0) flow rate of 0.5 mL/min at 35

C. The HPGPC
system was precalibrated with T-series Dextran standards (T-10, T-
40, T-70, T-110, and T-500).
Tea polysaccharide imaging with atomic
force microscopy
Tea polysaccharide was imaged with atomic force microscopy
according to the method of Zhang and others (2007). A stock so-
lution (1 mg/mL) was prepared by adding tea polysaccharide into
double distilled water. The solution was diluted to the final con-
centration of 1.0 g/mL. About 5 L of diluted tea polysaccharide
solution were dropped on the surface of mica sample carrier, al-
lowed to dry, and then imaged in air at room temperature. The
atomic force microscopy used in this study was an Ambios D3100
instrument (Ambios, Santa Cruz, Calif., U.S.A.) and was operated in
the tapping-mode. The resulting imaging force was estimated to be
0.05 to 3 nN and the resonant frequency was about 2 KHz.
Uv-vis and IR-FTR spectrophotometric analysis
The Uv-vis spectrophotometric analysis was conducted accord-
ing to the method of Yang and others (2008). Each tea polysac-
charide sample (1.0 mg) was dissolved in 10 mL of distilled water.
The absorbance of each sample solution was read over the range
of 190 to 900 nm, using a uv-2450 PC UV-visible spectrophotome-
ter (Shimadzu, Tokyo, Japan). FT-IR spectra (in KBr pellets, 2 mg
sample/200 mg KBr) were measuredusing the Bio-RadMerlinspec-
trophotometer operating at 4 cm
1
resolution.
Protease treatment of 3 polysaccharide fractions
Fifty micrograms of TPS were dissolved in PBS buffer (pH = 6)
and treated with 3 mg of pepsin (800-2500 U/mg protein, Sigma) at
55

C for 240 min. The reaction was stopped by increasing the tem-
perature to 90

C for 60 min to make protease inactive. After dial-
ysis with distilled water for 24 h, the polysaccharide solution was
concentrated, precipitated with 95% of ethanol, and freeze-dried.
The enzyme hydrolyzed fractions of TPS were obtained and tested
the scavenging effects on DPPH radicals at the concentration of 1
mg/mL. The same amount of pepsin treated with the same proce-
dure was used as the control.
Inhibition assay for -glucosidase activity
Inhibition assay of tea polysaccharide on -glucosidase activity
was performed according to Kim and others (2004). -Glucosidase
(0.25 unit) was premixed with tea polysaccharides at various con-
centrations, and 2 mM p-nitrophenyl -D-glucopyranoside as a
substrate in phosphate buffer was added to the mixture to start
the reaction. The reaction was incubated at 37

C for 30 min and
stopped by adding 2 mL of 0.1 M Na
2
CO
3
. -Glucosidase activ-
ity was determined by measuring release of p-nitrophenol from p-
nitrophenyl -D-glucopyranoside at 400 nm. Acarbose was used as
positive control.
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Physicochemical properties and antioxidant capacity. . .
Scavenging effects on hydroxyl radicals
Scavenging effects of tea polysaccharide on hydroxyl radicals
were performed as described by Halliwell and others (1987). Re-
action mixtures in a final volume of 1 mL contained deoxyribose
(60 mM), KH
2
PO
4
-KOH buffer (pH 7.4, 20 mM), FeCl
3
(100 M),
EDTA (100 M), various concentrations of tea polysaccharide sam-
ples, H
2
O
2
(1 mM), and ascorbic acid (100 M). After incubation at
37

C for 1 h, the color was developed by adding 1 mL of 1% thio-
barbituric acid (TBA) (w/v) and 1 mL of 25% (v/v) HCl, which was
then heated in a boiling water bath for 15 min. The absorbance of
resulting solution was measured spectrophotometrically at 532 nm
and butylated hydroxyl anisole (BHA) was used as positive control.
Scavenging effects on DPPHradicals
One hundred microliters of various concentration of the tea
polysaccharide were mixed with 2900 L of DPPH solution
(120 M) in ethanol and incubated in darkness at 37 C for 30 min.
The absorbance was recorded at 517 nm. Percentage (%) inhibition
of tea polysaccharide on free radical production by DPPH was cal-
culated (Liu and others 2007). Ascorbic acid was used as positive
control, and all tests were carried out in triplicate.
Inhibitory effects on nonenzymatic lipid
peroxidation induced by Fe
2+
/ascorbate
The inhibition of lipid peroxidation was assayed by the method
of Anup and others (2006) with slight modifications. Kunming mice
weighing 20 2 g (purchased from Beijing Weitonglihua Co., Bei-
jing, China) were housed under conventional conditions and were
allowed free access to food and water. All experiments were carried
out according to the guidelines for the care and use of experimental
animals and approved by state veterinary administration.
The mice were anesthetized using diethyl ether and the ab-
domen was opened and the liver was quickly removed. The livers
were then cut into small pieces and homogenized in phosphate
buffer (50 mM, pH 7.4) with a homogenizer to give a 10% (w/v)
liver homogenate. The liver homogenate was further centrifuged
at 3000 g for 10 min. The supernatant of the liver homogenate
was collected and the amount of protein was determined by the
method of Bradford (1976). The extent of lipid peroxidation was
evaluated by measuring the product of thiobarbituric acid-reactive
substances (TBARS) in the rat liver homogenate. The reaction mix-
ture was composed of 0.5 mL of tissue homogenate, 0.9 mL of phos-
phate buffer (50 mM, pH7.4), 0.25 mL of FeSO
4
(0.01 mM), 0.25 mL
of ascorbic acid (0.1 mM), and 0.1 mL of different concentration of
the extract and the standard sample. The reaction mixture was in-
cubatedat 37

Cfor 30 minandthe reactionwas thenterminatedby

adding 1 mL of TCA (20%, w/v) to the mixture. After centrifugation
at 3000 g for 15 min, the supernatant was incubated with 1 mL of
TBA (0.67%) at 100

C for 15 min. The absorbance of the complex
was measured at 532 nm. Tocopherol was used as positive control.
Statistical analysis
Values were expressed as means standard deviation (SD) of 3
replicates, and Students t test was used for the statistical analysis.
Results and Discussion
Composition analysis of tea polysaccharide
Although the green tea, oolong tea, and black tea were obtained
from the same region, there were significant differences existed
in the yield and composition of 3 kinds of tea polysaccharides
(Table 1). The yields of GTPS, OTPS, and BTPS from green tea,
oolong tea, and black tea were about 4.0%, 4.6%, and 4.2%,
respectively. The protein concentration in the polysaccharide-rich
fraction (BTPS) was about 38%, which was higher than that of GTPS
(32.6%) and OTPS (32.7%). The uronic acid and neutral sugar con-
centrations in BTPS were 16.1% and 18.8%, respectively, which was
lower than those of GTPS (20.8% and 27.3%) and OTPS (25.4%,
26.5%). There was no polyphenol detected in the 3 polysaccharide-
rich fractions. Inthe monosaccharide compositional analysis, there
was a significant difference among the composition and content of
the monosaccharide existed in GTPS, OTPS, and BTPS. The green
tea polysaccharide was composed of D-rhamnose, L-arabinose, D-
xylose, D-mannose, D-galactose, and D-glucose in the molecular
ratio of 7.8:41.8:7.1:7.3:18.7:17.0. But there were no xylose and man-
nose detected in the polysaccharides from oolong tea and black
tea. There was a percentage of about 20% for GTPS, and about the
same for the other 2 was not illustrated in Table 1. This was because
the water and the ash existed in the polysaccharide samples. Rare
earth fertilizer is widely applied in China to increase the yield and
the quality of crops including tea. In the study of Wang and others
(2003), an amount of 9.15% to 17.45% of rare earth elements in tea
infusion was bound to polysaccharide. So in the component analy-
sis of tea polysaccharides, there was mineral existed (Chen and oth-
ers 2007).
Molecular weight distribution of tea polysaccharide
To characterize differences in molecular weight, GTPS, OTPS,
and BTPS were analyzed by HPGPC. Because there was no fur-
ther purification, all 3 polysaccharide fractions had a wide distri-
bution. The molecular weight distribution of green tea polysac-
charide was from 9.2 to 251.5 KDa. But the molecular weight
distributionof oolong tea polysaccharide andblack tea polysaccha-
ride were 5.3 to 100.9 and 3.8 to 32.7 KDa, respectively. The pro-
portion of low molecular weight fractions (polysaccharides) was
higher in the molecular weight profile of BTPS which proved that
BTPS hada higher content of lowmolecular weight polysaccharides
than GTPS and OTPS. This difference of molecular weight distri-
bution in the 3 polysaccharides may be due to the different pro-
cessing procedure of the 3 kinds of tea. The green tea, oolong tea,
and black tea were belonged to unfermented tea, semifermented
tea, and fermented tea. The molecular weight distribution was re-
duced with the increase of the fermentation degree. During the
fermentation/oxidation of tea, the components were changed and
resulted in the different activities. According to Yanagimoto and
others (2003), significant antioxidative activities were obtainedonly
from the volatile extracts of green tea and roasted green tea among
the extracts tested. To compare caffeine and catechins in the same
Table 1--- Composition analysis of polysaccharide from
green tea, oolong tea, and black tea.
GTPS OTPS BTPS
Yield (%) 4.0 0.3a 4.6 0.2b 4.2 0.3a
Protein (%) 32.6 1.3a 32.7 1.6a 38.0 1.7a
Uronic acid (%) 20.8 0.4a 25.4 1.3b 16.1 0.7c
Neutral sugar (%) 27.3 1.0a 26.5 1.3a 18.8 0.8b
Neutral sugar composition (mol ratio%)
D-rhamnose 7.8 0.2a 16.2 0.7b 14.4 0.7b
L-arabinose 41.8 0.8a 43.7 1.0b 36.4 0.8c
D-xylose 7.1 0.7a --- ---
D-mannose 7.3 0.3 --- ---
D-galactose 18.7 1.2a 18.0 0.5a 19.7 1.0a
D-glucose 17 0.4a 21.9 0.5b 29.4 0.72c
Values are means SD of 3 parallel measurements.
Means with the same letter are not signicantly different (P > 0.05).
GTPS = green tea polysaccharide; OTPS = oolong tea polysaccharide;
BTPS = black tea polysaccharide.
Vol. 74, Nr. 6, 2009JOURNAL OF FOOD SCIENCE C471
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Physicochemical properties and antioxidant capacity. . .
tea but manufactured by different fermentation processes, the re-
sults of Lin and others (2003) showed that the level of caffeine in
different manufactured teas was in the order black tea > oolong
tea > green tea > fresh tea leaf, but the levels of EGCG and to-
tal catechins were in the order green tea > oolong tea > fresh tea
leaf > black tea. In this study, the length and the molecular weight
of the 3 kinds of tea polysaccharides were reduced with the increase
of the fermentation degree. This may be due to the hydrolysis of the
carbohydrate by the enzyme existed in the tea leaves.
Tea polysaccharide imaging with atomic
force microscopy
During its maturation, atomic force microscopy has devolved
into a versatile platform for experiments with individual molecules
(Li and others 1999). The technique has been used to image cell-
wall polysaccharides to characterize the contour length distribu-
tion of polysaccharides, to study polysaccharide association and
to detect branching of polysaccharide structures (Kirby and others
2008). The tea polysaccharide samples were deposited onto mica
at room temperature then the material appeared to collect as large
aggregates on the substrate. But in the whole visible fields there
were also some molecules appeared to spread as extended, stiff
chains. The shapes of the green tea polysaccharides and oolong
tea polysaccharides were like high-branched lines, but that of the
black tea polysaccharide was like balls. The lengths of the green tea
polysaccharide, oolong tea polysaccharide, and black tea polysac-
charide were 620 to 1526, 430 to 1296, and 430 to 1130 nm, respec-
tively. The length of the 3 kinds of polysaccharides was reduced
with the increase of the fermentation degree, which was well in
agreeing with the molecular weight distribution.
Spectrophotometric analysis of tea polysaccharide
UV-vis absorption is an effective tool for chemical characteriza-
tion because an adequate analysis of UV-vis spectra may provide
very interesting information on the chemical structure of an ana-
lyte. No apparent differences existed among the 3 spectra at a range
from190 to 900 nm(Figure 1.), which indicated that the mainstruc-
ture groups in the polysaccharides were similar. The UV-vis spec-
tra of these 3 samples showed a predominant peak at 199 to 200
nm while a slowly decreasing profile at 260 to 280 nm might in-
dicate the occurrence of proteins. FT-IR spectroscopy can also be
Figure 1--- UV spectra of 3 kinds of
tea polysaccharides.
GTPS = green tea polysaccharide;
OTPS = oolong tea polysaccharide;
BTPS = black tea polysaccharide;
concentration = 0.5 mg/mL, scan
range: 190 to 900 nm.
used for approximate identification of polysaccharides and pro-
teins when combined with chemical analyses data. The protein in
the polysaccharide was confirmed by its FT-IR spectrum (Figure 2)
which showed strong absorption bands of protein at 1650 cm
1
(amide I) and 1550 cm
1
(amide II), and stretching vibrations of
alkyl groups at 2958 cm
1
. The bands in the region 1500 to 1200
cm
1
are assigned to deformation of CH
2
and angular deforma-
tion of CCHand HCO. Those in the region 1200 to 950 cm
1
are
explained by stretching modes of CC and CO (Bureau and oth-
ers 2009). The variations observed in the spectral region range from
1500 to 900 cm
1
were shown for different polysaccharides due to
their different monosaccharide composition and content.
Inhibitory effects on -glucosidase activity
Glucosidase inhibitors are currently of interest owing to their
promising therapeutic potential in the treatment of disorders
such as diabetes, human immunodeficiency virus (HIV) infec-
tion, metastatic cancer, and lysosomal storage diseases. Glucosi-
dase inhibitors have also been useful in probing biochemical
pathways and understanding structureactivity relationship pat-
terns required for mimicking the enzyme transition state. Among
the various types of glucosidase inhibitors, disaccharides, imi-
nosugars, carbasugars, thiosugars, and nonsugar derivatives have
received great attention (de Melo and others 2006). Many studies
focused on the research for effective and safe -glucosidase in-
hibitors from natural materials to develop a physiological func-
tional food or lead compounds for use in antidiabetes (Kim and
others 2004; Li and others 2005). The inhibitory effects on alpha-
glucosidase activity of polysaccharide fractions were tested and
the reaction between alpha-glycosidase and polysaccharides was
detected by DNS reagent (1% 3,5-dinitrosalicylic acid and 12%
sodium potassium tartrate in 0.4 M NaOH). There was no glu-
cose released, which showed that there was no glucose presented
as side chains of polysaccharide linked trough alpha bonds. Black
tea polysaccharide had a dose-dependent effect on -glucosidase
inhibitory activity (Table 2). The inhibition rose from 14.3% to
91% with the concentration increased from 25 to 200 g/mL. But
there was no inhibition of green tea polysaccharide and oolong tea
polysaccharide on -glucosidase activity when the concentration
increasedfrom25 to 100 g/mL. Whenthe concentrationincreased
to 200 g/mL, the inhibition of green tea polysaccharide and
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Physicochemical properties and antioxidant capacity. . .
Figure 2--- IR spectra of 3 kinds of
tea polysaccharides.
GTPS = green tea polysaccharide;
OTPS = oolong tea polysaccharide;
BTPS = black tea polysaccharide.
Table 2--- Inhibitory effects of 3 kinds of polysaccharides
on -glycosidase activity (%).
Concentration (g/mL) GTPS OTPS BTPS
25 --- --- 14.3 2.2
50 --- --- 65.6 1.6
100 --- --- 83.2 0.7
200 11.2 1.8 22.3 1.4 91.1 3.2
Values are means SD of 3 parallel measurements.
GTPS = green tea polysaccharide; OTPS = oolong tea polysaccharide;
BTPS = black tea polysaccharide.
Table 3--- Antioxidant activities of 3 kinds of polysaccha-
rides.
Lipid peroxidation
induced by
DPPH radicaIs HydroxyI radicaIs Fe
2+
/ascorbate
IC
50
(g/ mL) IC
50
(g/ mL) IC
50
(g/mL)
GTPS 23.0 2.9a 424.3 13.6a 74.6 4.5a
OTPS 53.7 3.3b 537.3 9.0c 126.6 7.0b
BTPS 20.3 2.6a 352.3 12.0b 93.6 7.5a
Values are means SD of 3 parallel measurements.
Means with the same letter are not signicantly different (P > 0.05).
GTPS = green tea polysaccharide; OTPS = oolong tea polysaccharide;
BTPS = black tea polysaccharide.
oolong tea polysaccharide were only reached to 11.2% and 22.3%.
This may be due to the different conformation and composition
of the polysaccharides, which resulted in the different interaction
between aminoglycoside and the hydrophobic pocket of the en-
zyme. The mechanism of the -glycosidase inhibitory activities of
tea polysaccharides needs further study.
Antioxidant activities of tea polysaccharide
The uncontrolled production of oxygen derived free radicals was
involved in onset of many diseases such as cancer, hypoglycemic,
rheumatoid arthritis, and atherosclerosis as well as in degenera-
tive processes associated with aging (Wang and Luo 2007). So many
chemicals were detected for the antioxidant activities. As can be
seen from the data in Table 3, all the 3 kinds of polysaccharides ex-
hibited antioxidant activities on the DPPH radicals, hydroxyl rad-
icals, and lipid peroxidation. There was a distinguished difference
among the 3 polysaccharides on the antioxidant activities with the
different IC
50
values (P < 0.05). The scavenging effects of green
tea polysaccharides and black tea polysaccharide were more effec-
tive than that of oolong tea polysaccharides with the IC
50
value
a a
a a
a a
0
10
20
30
40
50
60
70
80
GTPS OTPS BTPS
Type of tea polysaccharides
S
c
a
v
e
n
g
i
n
g

r
a
t
e

(
%
)
0 min
240 min
Figure 3--- Antioxidant activities of TPS before and after
the hydrolysis of protease. Means with the same letter
are not signicantly different (P < 0.05).
of 23, 20, and 54 g/mL, respectively. In the study of scaveng-
ing effects on the hydroxyl radicals generated in a Fenton reac-
tion, black tea polysaccharides showed the highest activity (IC
50
value of 352.3 g/mL) and the oolong tea polysaccharides showed
the lowest (IC
50
value of 537.3 g/mL). The highest inhibition of
lipid peroxidation was observed in green tea polysaccharides with
the IC
50
value of 75 g/mL. The differences in antioxidant activ-
ities among the 3 polysaccharide-rich fractions appeared to be re-
lated to differences in monosaccharide composition and molecular
weight distribution of the polysaccharide. According to the stud-
ies of Chen and others (2008) and Zhang and others (2001), the
molecular weight of polysaccharides played an important role in its
bioactivity including antioxidant properties and antitumor bioac-
tivity.
Antioxidant activities of tea polysaccharide before
and after protease treatment
Scavenging rates of polysaccharide GTPS, OTPS, and BTPS on
DPPH radicals were 47.9%, 23%, and 61.7%, respectively (Figure 3).
After the treatment of pepsin, the antioxidant activities of GTPS,
OTPS, and BTPS on DPPH radicals were increased to 51.4%, 26.5%,
and 64.2%, respectively, but there was no significant difference
(P > 0.05). Pepsin is an enzyme which is released by the chief cells
in the stomach and which degrades food proteins into peptides
(Viera and others 1995). After the treatment of pepsin, the protein
Vol. 74, Nr. 6, 2009JOURNAL OF FOOD SCIENCE C473
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Physicochemical properties and antioxidant capacity. . .
bounded to the tea polysaccharides were hydrolyzed and excluded
by the dialysis. The results showed that there was no significant in-
fluence of protein in polysaccharide-rich fractions on the antioxi-
dant activities on DPPH radicals.
Conclusions
T
here are considerable differences between the yield and com-
position of the polysaccharide from green tea, oolong tea, and
black tea. The 3 polysaccharides also differed in their molecular
weight distribution, -glucosidase inhibitory activity, and antioxi-
dant properties. As a result, black tea polysaccharide consisted of a
high proportion of lowmolecular weight fractions, which was asso-
ciated with high bioactivities. There is a potential for exploitation of
black tea polysaccharide in curing diabetes.
Acknowledgment
The authors are grateful for the financial support of this study from
the Natl. Natural Science Foundation of China (grant nr 30600470).
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