Green tea preparation and its influence on the content of bioactive compounds
Drazˇenka Komes
*
, Dunja Horzˇic´, Ana Belšcˇak, Karin Kovacˇevic´ Ganic´, Ivana Vulic´
Faculty of Food Technology and Biotechnology, University of Zagreb, Pierottijeva 6, 10000 Zagreb, Croatia
a r t i c l e
i n f o
Article history:
Received 24 April 2009
Accepted 22 September 2009
Keywords:
Antioxidant capacity
Bioactive composition
Green tea
HPLC
Preparation
a b s t r a c t
The effect of different extraction conditions and storage time of prepared infusions on the content of bio-
active compounds of green teas and their antioxidant capacity were investigated. The content of total
phenols, total flavonoids and total non-flavonoids in green teas was determined spectrophotometrically,
while 7 flavan-3-ols, 6 phenolic acids and 3 methylxanthines were identified and quantified by using high
performance liquid chromatography (HPLC–PDA). Among the tested green teas bagged green tea Twinings
of London was recognized as the richest source of phenolic compounds (3585 mg/L GAE of total phenols).
The most abundant phenolic constituents of green tea were flavan-3-ols, of which EGCG was prevailing in
all teas (94.54–357.07 mg/L). The highest content of caffeine, as the most abundant methylxanthine, was
determined in powdered green tea. The findings of this investigation suggest that extraction efficiency of
studied bioactive compounds from green tea depends on the extraction conditions and that maximum
extraction efficiency is achieved during aqueous extraction at 80 °C, for 5
0
(powder), 15
0
(bagged) and
30
0
(loose leaf). In order to determine the antioxidant capacity of teas the DPPH, ABTS and FRAP assays
were applied. Regardless of the extraction conditions all green teas exhibited significant antioxidant
capacity in vitro, which was in correlation with their phenolic content, confirming that green tea is one
of the best dietary sources of antioxidants.
Ó 2009 Elsevier Ltd. All rights reserved.
1. Introduction
Freshly prepared infusion of dried leaves and buds of the plant
Camellia sinensis has been consumed for thousands of years as a
desirable beverage (
Scharbert, Holzmann, & Hofmann, 2004
). It is
appreciated because of its attractive aroma and taste characteris-
tics as well as beneficial health effects. Tea plant grows in a form
of an evergreen shrub in areas with suitable cultivation conditions
(optimal temperature in the range of 15–20 °C, high relative
humidity and annual rainfall) (
). Tea leaves are
manually harvested after each flush (the sprouting of the top two
leaves and bud) and then processed. In traditional green tea man-
ufacturing process the leaves of Camellia sinensis are harvested,
withered, rolled, and quickly steamed or heated to prevent enzy-
matic degradation of polyphenols. The most important biochemi-
cal process in harvested tea leaves is enzymatic oxidation, which
starts as soon as the integrity of the cells is broken, but other types
of enzymes, such as esterases, glycosidases and decarboxylases
may also catalyze transformations and degradations of polypheno-
lic compounds (
). Heat or steam inactivation of en-
zymes responsible for degradation retains a considerable amount
of the original biologically active components of green tea and also
produces some unique products.
Tea’s beneficial health effects are thought to stem from poly-
phenols with antioxidant properties. Green tea contains polyphe-
nols which include flavanols, flavandiols, and phenolic acids (up
to 30% of dry weight). The most important flavonoids are catechins,
which are present at about 10% of the dry weight basis (
). Six major catechins known to display
biological activity are (+)-catechin (C), ()-epicatechin (EC),
()-epigallocatechin
(EGC),
()-gallocatechin
gallate
(GCG),
()-epigallocatechin gallate (EGCG) and ()-epicatechin gallate
(ECG). These native phenolic compounds are strong antioxidants
with antimutagenic, anticarcinogenic, hypocholesterolemic, anti-
bacterial, antiallergic and other clinically relevant activities.
found that among green tea polyphenols
especially
()-epigallocatechin
gallate
(EGCG)
significantly
reduced food intake, body weight, blood levels of testosterone,
estradiol, leptin, insulin, insulin-like growth factor I, LH, glucose,
cholesterol and triglyceride, as well as growth of the prostate,
uterus and ovary.
Green tea also contains methylxanthines: caffeine (1,3,5-trim-
ethylxanthine) and two minor isomeric dimethylxanthines, theo-
bromine and theophylline, which are responsible for mildly
stimulant effects of the tea (
Anaya, Cruz-Ortega, & Waller, 2006;
). Besides being very important factors in tea
quality, these compounds are also reported to exhibit beneficial
health effects. Selected studies describe the use of caffeine to en-
hance mental activity, running performance, and to treat apnoea
and migraine headaches, and implementation of theophylline
0963-9969/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:
*
Corresponding author. Tel.: +385 1 4826252; fax: +385 1 4826251.
E-mail address:
(D. Komes).
Food Research International 43 (2010) 167–176
Contents lists available at
Food Research International
j o u r n a l h o m e p a g e : w w w . e l s e v i e r . c o m / l o c a t e / f o o d r e s
and theobromine in asthma and bradycardia treatment (
Levin, Choi, Kozukue, & Kozukue, 2006
Besides phenolic compounds and methylxanthines, tea provides
a significant dietary source of polysaccharides, vitamin B complex,
vitamin C, vitamin E, R-aminobutyric acid, fluoride, as well as min-
erals and trace elements such as K, Mn, Cr, Ni and Zn (
Pablos, Martın, & Gonzalez, 2002
).
The main objectives of the present study were: (A) to determine
the effect of different extraction conditions (temperature of water,
extraction time and multiple extraction) on the content of phenolic
compounds and methylxanthines in different green teas; (B) to
determine the antioxidant capacity of green teas depending on
extraction conditions and storage time of prepared infusion; and
(C) to compare the content of studied bioactive compounds in
loose leaf, bagged and powdered green teas.
2. Materials and methods
2.1. Chemicals
Folin–Ciocalteu, formic acid, ammonium peroxodisulphate,
sodium carbonate, sodium acetate trihydrate, acetic acid, hydro-
chloric acid, ferric chloride hexahydrate and ferric sulphate hepta-
hydrate were of analytical grade and supplied by Kemika (Zagreb,
Croatia). Formaldehyde was obtained from Alkaloid (Skopje,
Macedonia). DPPH (2,2-diphenyl-1-picrylhydrazyl) was supplied
by Fluka (Buchs, Switzerland) and methanol (HPLC grade) was
purchased from J.T. Baker (Deventer, Netherlands). Trolox
(6-hydroxy-2,5,7,8-tetramethylchromane-2-carboxylic acid), TPTZ
(2,4,6-tripyridyl-S-triazine),
ABTS
(2,2
0
-azino-bis(3-ethylbenz-
thiazoline-6-sulphonic acid) diammonium salt) as well as caffeine
(CF), theobromine (TB), theophylline (TP), ()-epicatechin (EC),
()-epigallocatechin
gallate
(EGCG),
()-epicatechin
gallate
(ECG), (+)-gallocatechin (GC), ()-gallocatechin gallate (GCG), (+)-
catechin (C), gallic acid (GA), protocatehuic acid (PcA), chlorogenic
acid (ChlA), ferulic acid (FA), p-coumaric acid (p-coumA) and vanil-
lic acid (VA) were obtained from Aldrich (Sigma–Aldrich Chemie,
Steinheim, Germany).
2.2. Sample preparation
Eleven green teas in powdered (Matcha), loose leaf (Gyokuro,
Sencha
J
, Bancha, Kukicha, Longjing, Yunnan, Sencha
CH
and Gunpow-
der) and bagged (Twinings of London and Franck) forms and two
blends of green and herbal teas in loose leaf (Rose of the Orient)
and bagged (Taylors of Harrogate) forms were purchased from a
specialized tea store. In order to simulate beverage brewing, teas
were prepared using an aqueous extraction procedure. In order
to study the effect of different extraction condition on the polyphe-
nol and methylxanthine content of tea samples, following extrac-
tion procedures were applied:
(a) Different water temperature – tea samples (2.0 g) were
extracted in 200 mL of distilled water (60, 80 and 100 °C)
and stirred with a glass rod for 3 min.
(b) Different extraction time – samples were extracted for 3, 5,
10, 15 and 30 min, with water heated to 80 °C.
(c) Multiple extractions – each tea sample was extracted three
times under the same conditions (80 °C/3 min) to produce
first, second and third extract.
After extraction, the infusions were filtered through a tea
strainer.
The changes related to the content of bioactive compounds as
well as antioxidant capacity of prepared infusions (80 °C/3 min)
were also studied during storage at the room temperature for 1,
2, 4, 6 and 24 h. To determine the effect of milk or lemon juice
on the antioxidant capacity of tea samples, 5 mL of milk or freshly
squeezed lemon juice was added to 100 mL of tea extract.
2.3. Determination of total phenols and flavonoids
Total phenol content (TPC) in tea extracts was determined spec-
trophotometrically according to a modified method of
Hosnedl, Pivec, and Orsák (1998)
, with Folin–Ciocalteu’s reagent.
Briefly, 0.5 mL of the sample was added into a 50 mL volumetric
flask containing 2.5 mL of Folin–Ciocalteu’s reagent, 30 mL of dis-
tilled water and 7.5 mL of 20% Na
2
CO
3
, and filled up to the mark
with distilled water. Two hours later, the absorbance of blue color-
ation was measured at 765 nm against a blank sample. Gallic acid
was used as the standard and the results are expressed as mg/L of
gallic acid equivalents (GAE).
In order to determine the content of flavonoids (TFC), precipita-
tion with formaldehyde was employed. Addition of formaldehyde
leads to the reactions with C-6 or C-8 on 5,7-dihydroxy flavonoid
and formation of methyl derivates that further react with other fla-
vonoid compounds also at positions C-6 and C-8. The condensed
products of these reactions are removed by filtration and the
remaining non-flavonoid phenols (TNC) are determined as previ-
ously described. Flavonoid content was calculated as the difference
between total phenol and non-flavonoid content (
). All measurements were performed in triplicate.
2.4. Determination of antioxidant capacity
2.4.1. Free radical scavenging ability by the use of DPPH radical
The samples were analyzed according to the method reported
by
Brand-Williams, Cuvelier, and Berset (1995)
. This method is
based on the reduction of a stable DPPH (2,2-diphenyl-1-pic-
rylhydrazyl) radical by methanolic solution of antioxidants, which
alters the purple color of the DPPH radical solution to a bright
yellow. Change of the coloration causes absorbance decrease,
which can be measured spectrophotometrically (
). In
brief, a volume of 3.8 mL of methanolic DPPH solution, c(DPPH) =
0.094 mmol/L, was added to 200
l
L of diluted sample. Free radical
scavenging capacity of the sample was evaluated by measuring the
absorbance at 517 nm immediately after the addition of DPPH
(t = 0) and at 1 min intervals until the reaction, i.e. corresponding
absorbance, reached its steady state. Antioxidant capacity was ex-
pressed as mmol/L Trolox equivalents, using the calibration curve
of Trolox (0–1000
l
M), a water soluble vitamin E analogue. All
determinations were performed in triplicate.
2.4.2. Free radical scavenging ability by the use of ABTS radical cation
Determination of antioxidant capacity of tea infusions was con-
ducted by Flow Injection Analysis (FIA) with electrochemically
generated ABTS radical cation (
Ivekovic´, Milardovic´, Roboz, & Gra-
). This method is based on the scavenging of stable
blue–green ABTS radical cation (ABTS*
+
), formed either by chemi-
cal (
Re et al., 1999; Van der Berg, Haenen, Van der Berg, & Bast,
) or enzymatic (
Arnao, Cano, & Acosta, 2001; Lemanska
) oxidation of ABTS several hours prior to the analysis.
The amount of ABTS radical cation scavenged by antioxidants is
measured by monitoring the decrease of absorbance of ABTS radi-
cal cation, and compared with the decrease of absorbance pro-
duced by the addition of a known amount of Trolox, a water
soluble vitamin E analogue. In order to avoid the time consuming
step of ABTS radical cation preparation, FIA modification of TEAC
method was employed. FIA is based on the electrochemical pro-
duction of ABTS radical cation in the electrolysis flow-cell forming
the part of FIA system (
). A 0.1 mol/L phosphate
168
D. Komes et al. / Food Research International 43 (2010) 167–176
buffer solution (pH = 7.40) was used as a carrier stream, and the
solution of ABTS radical cation generated electrochemically was
employed as a second (reagent) stream. Both streams were
pumped at a flow rate of 0.5 mL/min. The carrier and reagent
streams were mixed by passing through a mixing coil and the
absorbance at 734 nm was monitored by a detector placed
immediately after the mixing coil. For the analysis an aliquot of
20
l
L of diluted sample was injected into the carrier stream. The
height of the FIA peak obtained by the sample injection (
D
A
sample
)
was compared with the height of the peak produced by the
injection of 0.6 mmol/L Trolox solution (
D
A
Trolox
). The TEAC values
were then determined according to the formula: TEAC =
D
A
sample
c(Trolox)/
D
A
Trolox
, and expressed as equivalent concentration of
Trolox.
2.5. Determination of ferric reducing/antioxidant power (FRAP assay)
The ferric reducing/antioxidant power (FRAP) assay was carried
out according to
. This assay measures the
change in absorbance at 593 nm owing to the formation of a blue
colored Fe
II
-tripyridyltriazine compound from the colorless oxi-
dized Fe
III
form by the action of electron donating antioxidants
). FRAP reagent was prepared by
mixing acetic buffer, TPTZ and FeCl
3
6H
2
O (20 mM water solu-
tion) at a ratio of 10:1:1. Briefly, to a volume of 200
l
L of tea ex-
tract 3.8 mL of FRAP reagent was added. After 4 min the
absorbance of blue coloration was measured against a blank sam-
ple. A standard curve was prepared using different concentrations
(100–1200
l
mol/L) of Fe
2+
. All measurements were performed in
triplicate.
2.6. HPLC analysis of phenolic compounds and methylxanthines
Varian HPLC system (Varian, Walnut Creek, USA) consisting of
Pro Star Solvent Delivery System 230 and Pro Star 330 photodiode
array detector (PDA) and controlled by Star Chromatography
Workstation Version 5 software was used for HPLC analysis of
samples. Separation was performed using a reversed-phase Pinna-
cle II C-18 column (Restek, USA) (250 mm 4.6 mm, 5
l
m i.d.).
The samples were filtered through a 0.45
l
m membrane filter (Ny-
lon Membranes, Supelco, USA) and 20
l
L of each sample was in-
jected for HPLC analysis. The mobile phase consisted of 3%
formic acid (solvent A) and HPLC grade methanol (solvent B) at a
flow rate of 1 mL/min. The elution was performed with a gradient
starting at 2% B to reach 32% B at 20 min, 40% B at 30 min and 95%
B at 40 min, and becoming isocratic for 5 min. Chromatograms
were recorded at 278 nm. PDA detection was performed by record-
ing the absorbance of the eluate between 200 and 400 nm, with a
resolution of 1.2 nm (
). Phenolic compounds
were identified by comparing the retention times and spectral data
with those of authentic standards. All analyses were repeated
three times.
2.7. Statistical analysis
All measurements were carried out in triplicate and the results
are statistically analyzed using the Statistica 6.0 program to deter-
mine the average value and standard error. ANOVA and Duncan’s
multiple range test were performed to determine the effect of
extraction conditions (tea form, water temperature, extraction
time, multiple extraction) and storage time on the content of bio-
active compounds and antioxidant capacity of green teas. Correla-
tion analysis was also performed using the same statistical
package.
3. Results and discussion
The effect of different extraction conditions (tea form, water
temperature, extraction time and multiple extractions) and storage
time of prepared infusions on the content of phenolic compounds
and methylxanthines of green teas and their antioxidant capacity
were studied. Green tea is a very popular beverage because of its
attractive aroma, taste and beneficial effects. Nowadays, hundreds
of teas are produced. To make green tea, the leaves of Camellia sin-
ensis are chopped, rolled and quickly steamed or heated to inacti-
vate polyphenol oxidase. These processes retain a considerable
amount of original catechins of green tea, but they also result in
a development of some unique compounds (
). The consumption of green tea in western countries is a re-
cent trend and the market is continuously growing so the most
common commercially available types of green tea were tested
in this study.
Total phenol content (TPC) and total flavonoids content (TFC) of
13 green teas and their blends were determined spectrophotomet-
rically and presented on
. As can be seen, all tested teas pres-
ent a rich source of phenolic compounds, the content of which
depends on the extraction conditions. The highest TPC and TFC
were detected in bagged green tea Twinings of London (2560 and
1920 mg/L GAE, respectively) and in powdered green tea Matcha
0
500
1000
1500
2000
2500
3000
Matcha
Gyokuro
Sencha
Bancha
Kukicha
Longjing
Yunnan
Sencha
Gunpowder Rose of the
Orient
Twinings of
London
Taylors of
Harrogate
Franck
POWDER
LOOSE LEAF
BAGGED
mg GAE/l
Total non flavonoids
Total flavonoids
J
CH
Fig. 1. Total non-flavonoid and flavonoid content in green teas extracted at 80 °C/3 min.
D. Komes et al. / Food Research International 43 (2010) 167–176
169
(2230 and 1630 mg/L, respectively), while the lowest TPC and TFC
were detected in the green tea blend Rose of the Orient (880 and
440 mg/L, respectively). These results confirm the unique features
of each green tea analyzed in our study, which is in agreement with
the findings of
, who stated that the
composition of tea is influenced by the season, the age of the leaf,
climate, horticultural practices and a bit less by the effect of
locality.
In order to study the effect of different extraction conditions
(water temperature, time, multiple extraction) and storage time
on the polyphenol and methylxanthine content of tea samples,
powder (Matcha), loose leaf (Gyokuro) and bagged (Twinings of Lon-
don) green teas were tested. The effect of water temperature (60,
80 and 100 °C) and multiple extractions (1st, 2nd and 3rd) on
TPC and TFC of three different forms of green tea is shown on
. Among the tested teas, bagged green tea was recognized
as the richest source of phenolic compounds (TPC and TFC), which
reached their maximum values when water heated to 100 °C was
used for the extraction (3585 and 2865 mg/L GAE, respectively).
The obtained results confirmed previously published data (
) that water at higher temperature ex-
tracted higher TPC and TFC. Although the first extract of green
teas is often discarded due to excessive astringency, it is concluded
that the practice of reusing tea samples is not a good way of drink-
ing since most of health enhancing compounds (flavonoids)
decompose (
Astill, Birch, Dacombe, Humphrey, & Martin, 2001
Our results confirmed these statements, since in comparison with
1st extraction, 2nd and 3rd extractions of all three forms of green
tea at 80 °C (
) yielded significantly (p < 0.05) lower TPC and
TFC.
All tea extracts showed considerable fluctuations of TNC and
TFC, with regard to prolonged extraction time (
). In powder
and loose leaf green teas the prolonged extraction time (5
0
, 10
0
and 15
0
) resulted with higher TNC, while the TNC of 30
0
extracts
was decreasing. Bagged green tea extracts showed minor fluctua-
tions in TNC, and yielded the highest TNC in 30
0
extract. The TFC
of powdered green tea decreased in 5
0
extract, but increased in
all subsequent extracts, reaching the maximum in 30
0
extract.
The TFC of loose leaf and bagged green tea increased with pro-
longed extraction time, being the highest in 10
0
and 15
0
extracts
and then decreasing in 30
0
extracts, which indicates that prolonged
extraction time at high temperatures can lead to catechin
0
500
1000
1500
2000
2500
3000
3500
4000
60°C 80°C (1st)
2nd
3rd
100°C
60°C 80°C (1st)
2nd
3rd
100°C
60°C 80°C (1st)
2nd
3rd
100°C
POWDER
LOOSE LEAF
BAGGED
mg GAE/L
Total non flavonoids
Total flavonoids
Fig. 2. Total non-flavonoid and flavonoid content in powder (Matcha), loose leaf (Gyokuro) and bagged (Twinings of London) teas affected by water temperatures (60, 80 and
100 °C) and multiple extractions at 80 °C/3 min.
0
500
1000
1500
2000
2500
3000
3
5
10
15
30
3
5
10
15
30
3
5
10
15
30
POWDER
LOOSE LEAF
BAGGED
minutes
mg GAE/L
Total non flavonoids
Total flavonoids
Fig. 3. Total non-flavonoid and flavonoid content in powder (Matcha), loose leaf (Gyokuro) and bagged (Twinings of London) teas affected by extraction time (3, 5, 10, 15 and
30 min) at 80 °C.
170
D. Komes et al. / Food Research International 43 (2010) 167–176
degradation (
Cheong, Park, Kang, Ko, & Seo, 2005
). The obtained re-
sults are in accordance with those reported by
who observed that catechins tend to degrade during pro-
longed extraction time, as well as by using higher water
temperatures.
The content of non-flavonoids and flavonoids in all three differ-
ent forms of green tea was changed during storage time (
Namely, considerable fluctuations of TNC and TFC were observed,
especially after 2 and 4 h of storage, when TNC and TFC increased,
and then gradually decreased after 6 and 24 h of storage at room
temperature. Although the initial increase was followed by a de-
cline in TNC and TFC, at the end of 24-h storage, all three forms of
green tea exhibited satisfying stability during storage of tea extract.
The main classes of polyphenols present in green tea are flava-
nols and flavonols. These compounds constitute 16–30% of the dry
weight of fresh leaf (
). Catechins (flavan-3-ols) are
the major polyphenolic constituents of tea. These compounds are
colorless and water soluble and contribute to bitterness and astrin-
gency of green tea infusions (
). Due to a complex
bioactive composition of tea infusions it is very important to deter-
mine their methylxanthine content as well as polyphenolic profile
of teas, which is associated with their significant antioxidant prop-
erties. In this study, previously developed HPLC procedure (
) was performed and the content of flavan-3-ols, pheno-
lic acids and methylxanthines are summarized in
respectively. A comparison of the results, presented in these
Tables, showed that the most abundant bioactive constituents of
green tea were flavan-3-ols (ranging from 350.45 mg/L to
1073.15 mg/L), followed by methylxanthines (109.34–340.63
mg/L) and phenolic acids, which were determined in much lower
content (1.87–53.18 mg/L). As can be seen in
, EGCG was
the prevailing phenolic compound in all teas, ranging from 94.54
mg/L (in loose leaf green tea Sencha
CH
) to 357.07 mg/L (in bagged
green tea Taylors of Harrogate). The content of catechins is in agree-
ment with the previously published reports by
, who determined EGC (32.8–58.0 mg/g),
EGCG (70.9–118.0 mg/g), GCG (4.0–8.5 mg/g) and ECG (14.9–
24.5 mg/g) in Long Jing green tea, extracted during 5, 15 and
30 min. The detected values of catechins in the analyzed samples
are also similar to those reported by
Reto, Figueira, Filipe, and Al-
, who found 398.0–1127.0 mg/L of total catechins in
various green tea infusions, prepared with boiling water and
reported a little lower range of 33.4–
846.0 mg/L of catechins in different green tea samples prepared
also with boiling water. Considering the fact that water extraction
at 80 °C during 3
0
was performed for the purpose of present HPLC
analysis, the range of total flavan-3-ol content in tested green tea
extracts was in accordance with the total catechins content
(1067.9 mg/L) obtained by
Labbé, Tremblay, and Bazinet (2006)
,
who performed extractions with water at 80 °C during 5
0
. The com-
parison of the results is sometimes difficult due to the lack of uni-
formity in the conditions used to prepare tea infusions (ratio
leaves/water, extraction time, water temperature) as well as anal-
ysis methods.
Sencha
J
, Bancha and Kukicha are all Japanese tea types in loose
leaf forms, which are generally classified according to their regio-
nal origin, type of cultivation, processing and harvesting methods,
and composition. Sencha
J
is harvested in the early summer, Bancha
in the early fall, while Kukicha consists of a blend of leaves with the
stems and stalks normally discarded in the production of Sencha
J
and Gyokuro. Among the afore mentioned teas Sencha
J
contained
the highest content of some flavan-3-ols (169.27 mg/L of EGC,
208.71 mg/L of EGCG and 74.08 mg/L of ECG) and methylxanthines
(25.77 mg/L of TB and 5.92 mg/L of TF). The content of polyphenols
and methylxanthines in Kukicha green tea is closely similar to Sen-
cha
J
tea, even higher in case of EC, C, FA and CF, probably due to the
fact that Kukicha tea is a blend of Sencha
J
and Gyokuro teas, which
posses the highest content of these bioactive compounds. Bancha
tea had the lowest content of polyphenols (424.53 mg/L of total fla-
van-3-ols and 7.50 mg/L of total phenolic acids) and total methyl-
xanthines (125.04 mg/L).
Yunnan, Sencha
CH
and Gunpowder are Chinese teas, also in loose
leaf form, which differ in the processing method and tea prepara-
tion procedure. Yunnan originates from southwestern China and
it is produced from hand-rolled unopened tea buds, Sencha
CH
is
processed by steaming of the tea leaves, and Gunpowder is com-
posed of leaves that had been hand-rolled into tiny pellets. Gener-
ally, Sencha
CH
contained the lowest content of total flavan-3-ols
(350.45 mg/L), total phenolic acids (1.87 mg/L) and total methyl-
xanthines (109.34 mg/L), while Yunnan contained the highest con-
tent of these groups of bioactive compounds (886.13 mg/L,
40.56 mg/L and 316.8 mg/L, respectively).
Among the green tea blends, Franck and Twinings of London
(bagged forms) were analyzed in the present study. These blends
contain similar average flavan-3-ols, phenolic acids and methyl-
xanthines content, as the previously described pure green teas. In
0
500
1000
1500
2000
2500
1
2
4
6
24
1
2
4
6
24
1
2
4
6
24
POWDER
LOOSE LEAF
BAGGED
hours
mg GAE/L
Total non flavonoids
Total flavonoids
Fig. 4. Total non-flavonoid and flavonoid content in powder (Matcha), loose leaf (Gyokuro) and bagged (Twinings of London) teas affected by extract storage time (1, 2, 4, 6 and
24 h).
D. Komes et al. / Food Research International 43 (2010) 167–176
171
comparison with Franck tea, in which a lower content of total fla-
van-3-ols, phenolic acids and methylxanthines (683.29 mg/L,
10.38 mg/L and 280.74 mg/L, respectively) was detected, Twinings
of London tea was characterized with higher content of these com-
pounds (1073.15 mg/L, 53.18 mg/L and 280.74 mg/L, respectively).
Also, two blends of green and herbal teas; Taylors of Harrogate
(bagged form) and Rose of the Orient (loose leaf form) were studied.
Taylors of Harrogate is a blend of various green teas and lemon-
grass, while Rose of the Orient tea is green tea blended with jas-
mine, rosebuds, marigold and cornflower blossoms. The content
of extracted flavan-3-ols, phenolic acids and methylxanthines from
these blends followed the similar trend as the content of the same
compounds extracted from pure green teas. Therefore, total flavan-
3-ol content in these tea blends varied between 422.12 mg/L in
Rose of the Orient and 845.49 mg/L in Taylors of Harrogate, total
phenolic acid content is ranging from 27.23 mg/L in Rose of the Ori-
ent to 53.18 mg/L in Taylors of Harrogate, while total methylxan-
thines content varied between 125.96 mg/L in Rose of the Orient
and 340.63 mg/L in Taylors of Harrogate.
Green tea is an important source of methylxanthines, especially
caffeine. The leaf age is a very important factor because old leaves
contain less caffeine but more EGCG and total catechins than
young ones (
). CF, TF and TB are the main
methylxanthines constituting the tea alkaloids, hence, they are
one of the most important factors of the quality of teas. The con-
tent of CF in tested green tea (
) was between 99.21 mg/L
in Sencha
CH
(loose leaf green tea) and 300.00 mg/L in Matcha (pow-
dered green tea). The detected values are in agreement with previ-
ous findings by
, who found 141.0–338.0 mg/L of
caffeine in various green tea infusions prepared with boiling water
during 10
0
, while
reported a content of
305.4 mg/L of CF in green tea extracts obtained during 5
0
at
Table 1
Content of flavan-3-ols [()-gallocatechin (GC), ()-epigallocatechin (EGC), ()-epigallocatechin gallate (EGCG), ()-epicatechin, (+)-catechin, ()-gallocatechin gallate (GCG),
()-epicatechin gallate (ECG) in green teas extracted at 80 °C/3 min.
Flavan-3-ols
Total
()-GC
()-EGC
()-EGCG
()-EC
()-C
()-GCG
()-ECG
Powder tea
Matcha
70.49 ± 1.03
324.88 ± 18.01
345.55 ± 21.23
120.07 ± 2.38
25.75 ± 1.21
6.62 ± 0.81
102.67 ± 2.89
996.03
Loose leaf teas
Gyokuro
51.10 ± 0.92
c
279.87 ± 12.10
e
324.54 ± 18.76
e
123.43 ± 5.44
c
19.70 ± 1.43
b
3.90 ± 0.45
a
108.55 ± 6.13
a
911.09
f
Sencha
J
44.34 ± 0.87
bc
169.27 ± 13.23
d
208.71 ± 11.89
d
70.37 ± 4.81
b
12.80 ± 0.92
b
n.d.
*
74.08 ± 6.23
c
579.57
d
Bancha
53.73 ± 1.21
c
159.55 ± 14.11
d
103.28 ± 8.15
a
68.83 ± 3.89
b
6.56 ± 0.84
a
n.d.
*
32.58 ± 2.51
a
424.53
b
Kukicha
31.39 ± 0.73
b
159.39 ± 11.86
d
163.22 ± 9.23
c
76.20 ± 4.11
b
14.62 ± 1.37
b
2.73 ± 0.23
a
50.11 ± 4.15
b
497.66
c
Longjing
17.94 ± 0.36
a
69.71 ± 2.12
a
209.95 ± 13.71
d
62.50 ± 5.26
ab
43.72 ± 2.25
c
n.d.
*
n.d.
*
403.82
a
Yunnan
88.95 ± 2.54
d
266.24 ± 8.70
e
178.80 ± 12.1
c
169.16 ± 10.23
d
50.98 ± 3.81
c
n.d.
*
310.80 ± 18.34
e
886.13
e
Sencha
CH
47.47 ± 1.11
bc
124.47 ± 10.35
c
94.54 ± 7.62
a
55.42 ± 3.71
a
n.d.
*
n.d.
*
28.55 ± 2.06
a
350.45
a
Gunpowder
89.09 ± 1.87
d
93.35 ± 9.76
b
116.25 ± 6.45
a
48.80 ± 3.42
a
6.48 ± 4.11
a
n.d.
*
39.43 ± 1.76
a
393.40
a
Rose of the Orient
46.90 ± 0.85
bc
122.40 ± 7.45
c
130.30 ± 6.13
b
52.39 ± 2.71
a
10.81 ± 0.97
b
n.d.
*
59.32 ± 4.50
b
422.12
b
Bagged teas
Twinings of London
192.12 ± 16.31
c
235.94 ± 18.73
b
326.80 ± 25.55
b
114.98 ± 9.18
b
27.58 ± 2.41
a
9.31 ± 0.73
b
166.42 ± 7.55
b
1073.15
c
Franck
114.37 ± 11.24
a
184.33 ± 11.37
a
191.59 ± 12.18
a
89.43 ± 7.21
a
24.99 ± 1.64
a
5.23 ± 0.47
a
75.35 ± 4.28
a
683.29
a
Taylors of Harrogate
137.23 ± 12.62
b
n.d.
*
357.07 ± 19.64
bc
141.94 ± 6.54
c
31.14 ± 3.95
a
n.d.
*
178.11 ± 9.64
b
845.49
b
n.d.
*
= not detected.
Values are expressed as means in mg/L ± SD. The same letters (a, b, c, d, e, f) denote the flavan-3-ols, whose content is not significantly (p > 0.05) different between green teas
in loose leaf form, as well as in bagged form.
Table 2
Content of phenolic acids [protocatehuic acid (PCA), ferulic acid (FA), gallic acid (GA), p-coumaric acid (p-coumA), chlorogenic acid (ChlA) and vanillic acid (VA)] in green teas
extracted at 80 °C/3 min.
Phenolic acids
Total
PcA
FA
GA
p-coumA
ChlA
VA
Powder tea
Matcha
0.94 ± 0.05
15.24 ± 0.89
1.73 ± 0.21
0.93 ± 0.09
n.d.
*
n.d.
*
18.84
Loose leaf teas
Gyokuro
n.d.
*
n.d.
*
1.07 ± 0.09
b
6.63 ± 0.72
e
2.92 ± 0.05
c
n.d.
*
10.62
bc
Sencha
J
0.99 ± 0.08
b
7.03 ± 0.91
a
1.14 ± 0.11
b
n.d.
*
n.d.
*
n.d.
*
9.16
b
Bancha
1.27 ± 0.11
c
5.43 ± 0.63
a
0.73 ± 0.06
a
0.07 ± 0.01
a
n.d.
*
n.d.
*
7.50
b
Kukicha
1.35 ± 0.09
c
13.66 ± 1.01
b
1.16 ± 0.21
b
0.69 ± 0.03
c
0.65 ± 0.10
a
n.d.
*
17.51
c
Longjing
0.83 ± 0.05
b
11.28 ± 0.94
b
6.49 ± 0.52
d
0.63 ± 0.04
c
0.63 ± 0.08
a
1.00 ± 0.09
a
20.86
c
Yunnan
1.30 ± 0.09
c
14.29 ± 0.67
b
2.69 ± 0.18
c
0.01 ± 0.0
a
2.48 ± 0.06
c
9.79 ± 1.23
b
40.56
d
Sencha
CH
0.99 ± 0.10
b
n.d.
*
0.88 ± 0.22
a
n.d.
*
n.d.
*
n.d.
*
1.87
a
Gunpowder
0.36 ± 0.03
a
10.89 ± 0.73
ab
9.39 ± 0.19
d
1.22 ± 0.03
d
1.28 ± 0.13
b
n.d.
*
23.14
c
Rose of the Orient
0.98 ± 0.06
b
24.82 ± 0.51
c
1.24 ± 0.07
b
0.19 ± 0.01
b
n.d.
*
n.d.
*
27.23
c
Bagged teas
Twinings of London
n.d.
*
n.d.
*
5.77 ± 0.32
b
4.46 ± 0.53
b
4.23 ± 0.21
a
n.d.
*
14.46
a
Franck
0.87 ± 0.07
a
7.10 ± 0.11
a
2.41 ± 0.24
a
n.d.
*
n.d.
*
n.d.
*
10.38
a
Taylors of Harrogate
0.70 ± 0.03
a
9.78 ± 0.83
b
14.70 ± 1.63
c
1.44 ± 0.09
a
4.16 ± 0.34
a
22.40 ± 0.04
53.18
b
n.d.
*
= not detected.
Values are expressed as means in mg/L ± SD. The same letters (a, b, c, d, e) denote the phenolic acids, whose content is not significantly (p > 0.05) different between green teas
in loose leaf form, as well as in bagged form.
172
D. Komes et al. / Food Research International 43 (2010) 167–176
80 °C. For the healthy adult population, moderate daily caffeine in-
take at a dose content up to 400 mg/kg (6 mg/kg of body weight) is
not associated with adverse effects (
shows the total flavan-3-ols, phenolic acids and methyl-
xanthines of green teas influenced by different water temperature,
multiple extractions, extraction time and storage time. The highest
total flavan-3-ols, phenolic acids and total methylxanthines in
powder green tea were determined for 3
0
and 5
0
extracts, in bagged
green tea the highest total content of these bioactive compounds
was obtained for 15
0
extracts, while in loose leaf green tea for 30
0
extracts. Considering the extraction dynamics of different tea
forms,
observed higher extraction yield of
tea in the powdered form (at 100 °C), in comparison to loose leaf
tea, due to larger specific area of powdered tea, thus allowing a
faster extraction efficiency of various tea constituents. The results
of
indicate a higher methylxanthines
content of green teas in loose leaf form, in relation to green teas
in bagged form. According to our results, bagged green tea
yielded the highest content of both total flavan-3-ols and methyl-
xanthines, while powder tea yielded the highest content of
phenolic acids.
The obtained results suggest that higher water temperature and
short extraction time, as well as lower water temperature and
longer extraction time are the best combination for the extraction
of bioactive compounds of green tea, allowing the best extraction
efficiency. According to some previously conducted studies there
is a dependence of catechin content to time and/or temperature.
Sharma, Gulati, and Ravindranath (2005)
observed that catechins,
especially EGCG, EGC and EC, showed marked differences when ex-
tracted at different temperatures. In a study on green tea,
found that the best results in extraction of catechins
are achieved at 90 °C during 80 min.
ex-
plained the effect of temperature on catechin extraction by a dif-
ference in their molecular weight.
Table 3
Content of methylxanthines [caffeine (CF), theobromine (TB) and theophyline (TP)] in green teas extracted at 80 °C/3 min.
Methylxanthines
Total
CF
TB
TP
Powder tea
Matcha
300.00 ± 21.12
12.18 ± 0.91
10.12 ± 0.67
322.30
Loose leaf teas
Gyokuro
296.86 ± 19.75
d
15.41 ± 0.28
b
6.49 ± 0.53
b
318.76
d
Sencha
J
162.50 ± 13.63
a
25.77 ± 1.17
c
5.92 ± 0.36
b
194.19
c
Bancha
113.96 ± 12.28
a
8.22 ± 1.23
a
2.86 ± 0.21
a
125.04
b
Kukicha
174.68 ± 9.71
b
17.80 ± 0.98
b
4.33 ± 0.36
ab
196.81
c
Longjing
248.38 ± 9.62
cd
63.53 ± 2.11
d
15.71 ± 1.35
c
327.62
d
Yunnan
201.62 ± 10.73
c
86.18 ± 4.16
d
29.08 ± 2.14
d
316.88
d
Sencha
CH
99.21 ± 6.54
a
7.55 ± 0.28
a
2.58 ± 0.46
a
109.34
a
Gunpowder
155.93 ± 9.11
b
26.04 ± 0.91
c
5.56 ± 0.45
b
187.53
c
Rose of the Orient
112.21 ± 5.47
a
10.82 ± 0.86
ab
2.93 ± 0.17
a
125.96
b
Bagged teas
Twinings of London
202.29 ± 16.51
b
58.65 ± 3.80
b
19.80 ± 1.01
b
280.74
b
Franck
151.73 ± 13.78
a
21.58 ± 1.11
a
8.57 ± 0.87
a
181.88
a
Taylors of Harrogate
246.71 ± 20.05
b
66.91 ± 0.94
b
27.01 ± 0.36
b
340.63
b
Values are expressed as means in mg/L ± SD. The same letters (a, b, c, d) denote the methylxanthines, whose content is not significantly (p > 0.05) different between green teas
in loose leaf form, as well as in bagged form.
Table 4
Changes in the content of total flavan-3-ols, phenolic acids and methylxanthines in powder, loose leaf and bagged green teas, depending on extraction conditions.
Total flavan-3-ols
Total phenolic acids
Total methylxanthines
Powder
Loose leaf
Bagged
Powder
Loose leaf
Bagged
Powder
Loose leaf
Bagged
Water temperature
60 °C
662.31 ± 32.02
730.16 ± 21.00
990.89 ± 21.81
17.05 ± 1.50
d
10.61 ± 1.04
g
18.12 ± 2.31
j
277.39 ± 23.51
253.41 ± 24.02
284.81 ± 19.11
80 °C
996.02 ± 36.24
911.15 ± 27.17
1073.22 ± 20.54
18.82 ± 2.31
d
10.68 ± 0.98
g
14.50 ± 1.91
j
322.33 ± 24.10
318.75 ± 18.93
280.73 ± 21.35
100 °C
972.89 ± 36.31
998.78 ± 18.97
1371.32 ± 39.21
19.53 ± 2.62
d
9.05 ± 1.10
g
16.33 ± 1.91
j
310.33 ± 22.42
330.18 ± 21.62
337.83 ± 25.61
Multiple extractions
1st extract
996.02 ± 36.24
911.15 ± 27.17
1073.22 ± 20.54
18.82 ± 2.31
10.68 ± 0.98
14.50 ± 1.91
322.33 ± 24.10
318.75 ± 18.93
280.73 ± 21.35
2nd extract
342.83 ± 24.15
337.56 ± 22.73
502.06 ± 34.08
6.38 ± 0.91
4.31 ± 0.32
2.76 ± 0.22
86.10 ± 19.46
135.64 ± 11.77
77.33 ± 10.48
3rd extract
247.26 ± 34.72
296.82 ± 19.05
212.16 ± 16.19
5.74 ± 0.15
2.35 ± 0.12
2.34 ± 0.56
28.09 ± 7.01
48.89 ± 8.26
72.49 ± 9.00
Extraction duration
3 min
996.03 ± 21.05
a
911.12 ± 23.61
1073.23 ± 22.85
18.80 ± 1.31
e
10.64 ± 0.96
h
14.56 ± 1.23
322.37 ± 23.13
l
318.76 ± 25.14
n
280.79 ± 22.16
5 min
997.82 ± 26.79
a
984.05 ± 25.48
1177.53 ± 36.91
18.86 ± 1.02
e
10.19 ± 0.83
h
13.92 ± 1.20
303.91 ± 24.05
l
291.53 ± 24.08
n
330.07 ± 23.75
10 min
992.81 ± 23.83
a
1166.14 ± 32.88
1400.68 ± 40.12
16.86 ± 1.28
e
10.54 ± 0.87
h
20.95 ± 1.46
295.07 ± 21.06
l
318.52 ± 27.21
n
367.93 ± 31.00
15 min
932.06 ± 34.18
a
1229.48 ± 36.01
1567.33 ± 42.35
16.63 ± 0.81
e
11.69 ± 1.04
h
26.56 ± 1.93
291.73 ± 25.81
l
335.46 ± 26.81
n
398.36 ± 32.86
30 min
566.55 ± 18.64
a
1431.89 ± 38.42
1006.40 ± 28.78
17.10 ± 1.13
e
12.47 ± 1.15
h
21.61 ± 1.57
229.65 ± 19.23
l
345.15 ± 29.03
n
331.91 ± 28.94
Storage time
1 h
848.04 ± 15.86
b
656.73 ± 22.07
904.15 ± 20.12
c
20.12 ± 1.66
f
9.02 ± 0.52
i
14.08 ± 1.14
k
292.09 ± 22.18
m
263.37 ± 18.48
o
267.72 ± 22.05
p
2 h
799.58 ± 19.71
b
748.72 ± 18.76
1030.62 ± 32.46
c
19.70 ± 1.34
f
9.02 ± 0.74
i
17.69 ± 1.29
k
286.34 ± 24.34
m
244.78 ± 21.04
o
306.00 ± 25.31
p
4 h
935.48 ± 24.32
b
198.99 ± 9.42
660.61 ± 19.65
c
19.44 ± 1.56
f
7.67 ± 0.61
i
9.45 ± 1.10
k
317.86 ± 19.96
m
167.35 ± 12.64
o
220.87 ± 17.94
p
6 h
767.09 ± 16.22
b
568.55 ± 11.93
1094.58 ± 31.54
c
15.61 ± 0.97
f
6.65 ± 0.84
i
15.84 ± 0.99
k
269.69 ± 20.85
m
236.06 ± 21.86
o
339.45 ± 30.57
p
24 h
829.96 ± 19.55
b
232.65 ± 14.30
1009.83 ± 30.98
c
18.89 ± 1.03
f
6.25 ± 0.27
i
14.13 ± 5.43
k
294.74 ± 28.10
m
172.97 ± 13.37
o
304.10 ± 32.80
p
Values are expressed as means in mg/L ± SD. The same letters (a, b, c, d, e, f, g, h, i, j, k, l, m, n, o, p) denote the total flavan-3-ols, phenolic acids and methylxanthines, whose
content is not significantly (p > 0.05) affected by the extraction conditions.
D. Komes et al. / Food Research International 43 (2010) 167–176
173
Repeated extractions of powder, loose leaf and bagged green
teas showed significant influence (p < 0.05) on the content of fla-
van-3-ols, phenolic acids and methylxanthines. As with the TPC
and TFC (
), the content of flavan-3-ols, phenolic acids and
methylxanthines (
) also gradually decreased in subsequent
extractions, so the lowest content was obtained in third extracts of
all green tea forms.
The changes in the composition of polyphenols and methylxan-
thines during 24 h storage of green tea extracts (at room tempera-
ture) is difficult to interpret and compare, due to the lack of similar
studies on tea extracts. As could be seen in
, high variations
of total flavan-3-ols, phenolic acids and methylxanthines content
was observed, in the period of 24-h storage time, but finally after
24 h storage the content of polyphenols and methyxanthines in
all tea extracts decreased. The variation might be due to a great
abundance and variability of all tea constituents, which participate
in various reactions during storage of a tea infusion in the presence
of oxygen, such as polymerization, or even degradation of some tea
compounds.
Because individual catechins have different antioxidative and
health properties it is important to know the content of each one
of them in the infusions.
observed that the anti-
oxidant activity of green tea is higher in teas that contain higher
content of EGCG and EGC. According to previous findings by
Evans, Miller, and Paganga (1996)
who studied the antioxidant
activity of the polyphenolic constituents of green tea in relation
DPPH
0
3
6
9
12
mmol/L
T
rolox
ABTS
0
4
8
12
16
20
mmol/L
T
rolox
FRAP
0
5
10
15
20
25
Matcha
Gyokuro
Sencha
Bancha
Kukicha
Longjing
Yunnan
Sencha
Gunpowder Rose of the
Orient
Twinings of
London
Taylors of
Harrogate
Franck
POWDER
LOOSE LEAF
BAGGED
mmol/L
Fe2+
J
CH
Fig. 5. Antioxidant capacity of green teas prepared at 80 °C/3 min evaluated by using ABTS, DPPH and FRAP assays.
174
D. Komes et al. / Food Research International 43 (2010) 167–176
to their relative compositions, the order of contribution to the anti-
oxidant effectiveness in green tea was EGC EGCG ECG = EC > C.
The scavenging abilities of EGCG and ECG are stronger even if the
steric hindrance of EGC, GC, EC and (+)-C is smaller than that of
EGCG and GCG, indicating that the presence of a gallate group at
the 3 position plays the most important role in their abilities to
scavenge free radicals (
).
Antioxidant capacity of green teas prepared at 80 °C/3
0
and
determined with DPPH, ABTS and FRAP assays is shown on
.
As can be seen, Twinings bagged green tea exhibits the highest anti-
oxidant capacity (10.58 mmol/L Trolox), while Gunpowder loose
leaf green tea the lowest (4.36 mmol/L Trolox). As previously de-
scribed, Gunpowder is made up of leaves that had been hand-rolled
into tiny pellets, so the 3
0
extraction time is most likely insufficient
to extract a higher antioxidant content.
The order of antioxidant capacities of green teas influenced by
different water temperatures (
) is in accordance with the
TPC determined by the Folin–Ciocalteu method, which is con-
firmed by a high linear correlation obtained between the results
(r
DPPH/TPC
= 0.876, r
FRAP/TPC
= 0.595 and r
ABTS/TPC
= 0.912). In all
green teas an increase in antioxidant capacities was consistent
with the increase of water temperature used for the extraction. A
high correlation was as well observed by correlating the antioxi-
dant capacities with the TPC in subsequent extracts of green teas
(r
DPPH/TPC
= 0.928, r
FRAP/TPC
= 0.902 and r
ABTS/TPC
= 0.882), which im-
plies that the antioxidant potential of green tea is directly related
to their phenolic content.
Prolongation of the extraction time of bagged and loose leaf
green tea, also significantly (p < 0.05) affected the antioxidant
capacity, while in the case of powder tea extraction time do not
have significantly (p > 0.05) effect on its antioxidant capacity (
). The antioxidant capacities of 3
0
, 5
0
and 10
0
extracts gradually
increased, reaching maximum value after 15
0
and 30
0
extraction,
depending on the type of tea and the applied antioxidant assay.
Also, during 24-h storage at room temperature, fluctuations in
the antioxidant capacities of all green teas were observed, as
shown in
. In the first 6 h, the antioxidant capacity of the
analyzed green tea extracts increased and after 6 h of storage time,
almost all studied extracts exhibited the highest antioxidant
capacity. After the 24-h storage, the final antioxidant capacities de-
creased, but were still higher than the initial values for loose leaf
and powder green tea. Bagged green tea exhibited lower antioxi-
dant capacity after 24-h, in comparison with the initial values.
According to
, the in-
crease in the antioxidant capacity could be explained by the strong
tendency of polyphenols to undergo polymerization reactions.
When the degree of polymerization exceeds a critical value, the in-
creased molecular complexity and steric hindrance reduce the
availability of hydroxyl groups in reactions with radicals, which
causes decrease in the antiradical capacity. This may explain the
observed decrease in antioxidant capacity of studied extracts,
which occurred after the initial increase.
The addition of milk and lemon to tea extracts exhibited diverse
impact on the antioxidant capacity of tea in our study (
), as
well as in the literature survey.
Hollman, Van Het Hof, Tijburg, and
found that the addition of milk does not alter the
antioxidant properties of tea, while
stated
otherwise, claiming that due to the flavonoid-binding capacity of
milk proteins, the addition of milk can decrease the antioxidant po-
tential of black tea preparations. The addition of milk and lemon to
powdered Matcha tea mildly decreased its antioxidant capacity,
while in Gyokuro and Twinings the opposite was observed. The
antioxidant capacity of all three green tea forms increased with
the addition of lemon juice, which is directly attributed to the syn-
ergistic effect of ascorbic acid on the polyphenols present in tea ex-
tract (
Majchrzak, Mitter, & Elmadfa, 2004
). According to
Anese, Parpinel, Franceschi, and Lerici (1997)
various environmen-
tal impacts such as cultivation conditions, industrial processing,
Table 5
Influence of the extraction conditions (water temperature, duration, multiple extractions, and additions) and extract storage time on the antioxidant capacity of powder loose leaf
and bagged green teas.
Powder form
Loose leaf form
Bagged form
DPPH
FRAP
ABTS
DPPH
FRAP
ABTS
DPPH
FRAP
ABTS
mmol/L Trolox
mmol/L Fe
2+
mmol/L Trolox
mmol/L Trolox
mmol/L Fe
2+
mmol/L Trolox
mmol/L Trolox
mmol/L Fe
2+
mmol/L Trolox
Water temperature
60 °C
9.84 ± 0.49
15.65 ± 0.73
9.72 ± 0.64
6.74 ± 0.46
4.6 ± 0.18
8.12 ± 0.43
8.10 ± 0.57
13.55 ± 0.36
13.63 ± 0.84
80 °C
9.77 ± 0.45
a
18.55 ± 0.69
11.43 ± 0.89
7.97 ± 0.35
g
6.1 ± 0.27
h
9.77 ± 0.54
10.58 ± 0.83
i
14.65 ± 0.97
16.99 ± 1.10
n
100 °C
12.54 ± 0.83
20.4 ± 0.64
11.88 ± 0.36
10.81 ± 0.72
10.4 ± 0.53
10.47 ± 0.62
14.49 ± 0.52
15.35 ± 0.67
19.17 ± 1.47
Multiple extractions
1st extract
9.77 ± 0.45
18.55 ± 0.69
11.43 ± 0.89
7.97 ± 0.35
6.1 ± 0.27
9.77 ± 0.54
10.58 ± 0.83
14.65 ± 0.97
16.99 ± 1.10
2nd extract
2.91 ± 0.17
3.37 ± 0.29
1.82 ± 0.12
3.01 ± 0.12
1.65 ± 0.10
4.95 ± 0.26
6.45 ± 0.25
6.35 ± 0.63
9.62 ± 0.62
3rd extract
1.72 ± 0.09
0.62 ± 0.03
0.48 ± 0.01
1.89 ± 0.05
0.55 ± 0.02
1.74 ± 0.07
3.73 ± 0.12
2.35 ± 0.06
1.97 ± 0.05
Additions
Tea + lemon
8.86 ± 0.82
a
7.55 ± 0.23
9.79 ± 0.53
8.54 ± 0.36
g
6.00 ± 0.09
h
6.21 ± 0.72
11.97 ± 1.02
i
11.00 ± 0.93
16.99 ± 1.23
n
Tea + milk
8.69 ± 0.45
a
8.65 ± 0.49
8.59 ± 0.87
8.16 ± 0.53
g
8.00 ± 0.71
h
6.34 ± 0.43
11.87 ± 0.91
i
17.30 ± 1.13
15.09 ± 0.83
n
Extraction duration
3 min
9.77 ± 0.33
b
18.55 ± 1.23
d
11.43 ± 0.75
e
7.97 ± 0.45
6.10 ± 0.42
9.77 ± 0.83
10.58 ± 0.93
j
14.65 ± 1.37
l
16.99 ± 0.73
o
5 min
10.15 ± 0.41
b
22.87 ± 0.93
d
13.34 ± 1.10
e
9.73 ± 0.17
21.95 ± 1.12
12.19 ± 0.54
14.13 ± 1.85
j
21.65 ± 1.85
l
10.08 ± 0.31
o
10 min
10.31 ± 0.23
b
23.45 ± 0.64
d
12.10 ± 0.98
e
11.66 ± 0.27
22.22 ± 1.71
13.16 ± 1.03
14.45 ± 1.13
j
20.40 ± 2.01
l
18.44 ± 1.34
o
15 min
10.93 ± 0.81
b
21.42 ± 0.48
d
15.33 ± 0.67
e
12.77 ± 0.91
24.80 ± 1.26
15.77 ± 0.77
16.00 ± 1.37
j
20.25 ± 0.27
l
21.75 ± 1.94
o
30 min
10.45 ± 0.46
b
10.66 ± 0.52
d
12.50 ± 1.34
e
13.45 ± 0.68
23.78 ± 1.86
18.37 ± 1.08
15.38 ± 0.33
j
20.20 ± 0.83
l
21.86 ± 1.39
o
Storage time
1 h
11.97 ± 0.93
c
7.40 ± 0.36
11.38 ± 0.25
f
8.83 ± 0.54
7.59 ± 0.74
9.54 ± 0.64
14.79 ± 0.36
k
17.20 ± 0.46
m
16.51 ± 0.83
p
2 h
9.54 ± 0.62
c
7.86 ± 0.41
13.22 ± 0.53
f
8.96 ± 0.23
7.87 ± 0.45
11.79 ± 0.38
13.48 ± 0.84
k
14.20 ± 0.73
m
17.02 ± 0.62
p
4 h
8.33 ± 0.54
c
18.67 ± 1.67
12.26 ± 0.63
f
9.75 ± 0.37
13.00 ± 0.12
10.19 ± 0.75
11.76 ± 0.37
k
14.65 ± 0.83
m
16.71 ± 0.73
p
6 h
10.85 ± 0.71
c
18.80 ± 2.07
14.02 ± 1.12
f
10.23 ± 0.41
15.37 ± 0.99
13.44 ± 0.74
14.99 ± 0.98
k
15.40 ± 0.75
m
16.90 ± 0.18
p
24 h
10.08 ± 0.78
c
7.87 ± 0.66
13.64 ± 1.22
f
9.49 ± 0.68
8.65 ± 0.71
11.48 ± 0.38
12.54 ± 0.32
k
14.40 ± 0.62
m
16.41 ± 0.49
p
Values are expressed as means in mg/L ± SD. The same letters (a, b, c, d, e, f, g, h, i, j, k, l, m, n, o, p) denote the antioxidant capacity of teas, whose content is not significantly
(p > 0.05) affected by the extraction conditions.
D. Komes et al. / Food Research International 43 (2010) 167–176
175
packaging and storaging can influence the antioxidant capacity of
tea. The antioxidant activities of putative antioxidants have been
attributed to various mechanisms. Among these are prevention of
chain initiation, binding of transition metal ion catalysts, decom-
position of peroxides, prevention of continued hydrogen abstrac-
tion and radical scavenging (
).
4. Conclusion
This research is a contribution to the characterization of the sta-
bility of bioactive compounds of green teas during its preparation
and storage. The obtained results suggest significant differences in
the chemical composition of green teas commercially available in
Europe. Although green tea is a rich source of bioactive compounds
(polyphenols and methyxanthines) the extraction efficiency of
these compounds strongly depends on the extraction conditions.
The findings of this investigation suggest that maximum extraction
eficiency of studied bioactive compounds from green tea is
achieved during aqueous extraction at 80 °C, for 5
0
(powder), 15
0
(bagged) and 30
0
(loose leaf). Antioxidant capacity of all tested teas
was in correlation with the total phenol content, depending on the
extraction conditions.
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D. Komes et al. / Food Research International 43 (2010) 167–176
Document Outline
- Green tea preparation and its influence on the content of bioactive compounds
- Introduction
- Materials and methods
- Results and discussion
- Conclusion
- References
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