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Determination of Caffeine Content in Tea and Maté Tea by Using Different Methods

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Caffeine-containing products have been consumed for hundreds of years for their pleasent flavor and stimulating effects. In recent years, caffeine received increasing attention in food and pharmaceutical industries, due to its pharmacological properties which comprise stimulation of the central nervous system, peripheral vasoconstriction, relaxation of the smooth muscle and myocardial stimulation. The aim of this study was to determine the content of caffeine in five types of tea (white, yellow, green, oolong, black) and two types of maté tea (green maté and roasted maté tea). The content of caffeine was determined by using four different methods: extraction with chloroform, micro-method, method with lead-acetate and high performance liquid chromatography method (HPLC-PDA). The antioxidant capacity of teas as well as of the extracted („ raw“ ) caffeine was determined by using two methods: reactions with 2, 2`-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) radical (ABTS assay) and Ferric reducing antioxidant power (FRAP assay). The content of caffeine has been associated to plant origin and growth conditions, as well as processing conditions. By applying all four methods, the highest content of caffeine was determined in white tea, whereas maté and roasted maté tea were characterised with the lowest content of caffeine. The results of the micro-method and the method with lead-acetate have a good correlation with the results of HPLC method. Micro-method has proven to be the best alternative to the HPLC method. The highest antioxidant capacity was determined in yellow tea, while the lowest was determined in roasted maté tea. In comparison to the antioxidant capacity of teas, the antioxidant capacity of extracted („ raw“ ) caffeine is almost negligible, and does not contribute to the overall antioxidant properties of tea.
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Czech J. Food Sci. Vol. 27, 2009, Special Issue
Determination of Caffeine Content in Tea and Maté Tea
by using Different Methods
D. KOMES*, D. HO, A. BELŠČAK, K. KOVAČEVIĆ GANIČ and A. BALJAK
Faculty of Food Technology and Biotechnology, University of Zagreb, HR-10002 Zagreb,
Croatia, *E-mail: dkomes@pbf.hr
Abstrac t: Caffeine-containing products have been consumed for hundreds of years for their pleasant flavor and stimu-
lating effects. In recent years, caffeine received increasing attention in food and pharmaceutical industries, due to its
pharmacological properties which comprise stimulation of the central nervous system, peripheral vasoconstriction,
relaxation of the smooth muscle and myocardial stimulation. The aim of this study was to determine the content of
caffeine in five types of tea (white, yellow, green, oolong, black) and two types of maté tea (green maand roasted
maté tea). The content of caffeine was determined by using four different methods: extraction with chloroform,
micromethod, method with lead-acetate and high performance liquid chromatography method (HPLC-PDA). The
antioxidant capacity of teas as well as of the extracted (“raw ”) caffeine was determined by using two methods: reac-
tions with 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) radical (ABTS assay) and Ferric reducing antioxidant
power (FRAP assay). The content of caffeine has been associated with plant origin and growth conditions, as well
as processing conditions. By applying all four methods, the highest content of caffeine was determined in white tea,
whereas maté and roasted maté tea were characterised with the lowest content of caffeine. Spectrophotometric micro-
method has proven to be the best alternative to the HPLC method. The highest antioxidant capacity was determined
in yellow tea, while the lowest was determined in roasted maté tea. In comparison to the antioxidant capacity of teas,
the antioxidant capacity of extracted (“raw”) caffeine is almost negligible, and does not contribute to the overall anti-
oxidant properties of tea.
Keywords: antioxidant capacity; caffeine; HPLC; maté tea; tea
INTRODUCTION
Caffeine (1,3,5-trimethylxanthine) and two of its
minor isomeric dimethylxanthines, theobromine
and theophyline, belong to a group of methyl-
xantines. The widespread occurrence of caffeine
in a variety of plants played a major role in the
long-standing popularity of caffeine-containing
products. The most imp ortant sources of caf-
feine are coffee (Coffea spp.), tea (Camellia sinen-
sis), guarana (Paullinia cupana), maté (Ilex para-
guariensis), cola nuts (Cola vera), and cocoa (Theo-
broma cacao). The amount of caffeine found in
these products varies – the highest amounts are
found in guarana (4–7%), followed by tea leaves
(3.5%), maté tea leaves (0.89–1.73%), coffee beans
(1.1–2.2%), cola nuts (1.5%), and cocoa beans
(0.03%) (C et al. 1990).
Caffeine has pharmacological effects on central
nervous system, heart, peripheral and central
vasculature, renal, gastrointestinal and respiratory
system. Due to the widespread consumption of
methylxanthines, it is important to collect pre-
cise information on their content in foods. Most
research activities have been focused on chroma-
tographic methods, however, spectrophotometric
determination is preferred because of its rapidity,
high accuracy and reproducibility. Therefore, it is
important to develop more reliable, simpler and
faster methods for the determination of caffeine
from different sources in order to find a more pre-
cise relationship between the amounts of consumed
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Vol. 27, 2009, Special Issue Czech J. Food Sci.
caffeine and its physiological effects (G et
al. 2001; D A et al. 2005).
The aim of this study was to compare four dif-
ferent methods for determination of caffeine and
two methods for antioxidant capacity estimation
in tea (Camellia sinensis) and ma (Ilex para-
guariensis) products. Results of this study could
be used to facilitate selection of the appropriate
method in order to obtain satisfactory data on
caffeine content in plant materials.
MATERIALS AND METHODS
Sample preparation. Five types of tea in loose
leaf form: Pai Mu Tan-superior (white tea), Sencha
(green tea), Formosa Fine Oolong (oolong tea),
Lingia (black tea), Yin Zhen (yellow tea) and two
types of loose leaf maté tea (maté tea and roasted
matea) were purchased on a local market. In
order to simulate household brewing conditions,
teas were prepared using an aqueous extraction. Tea
samples (2.5 g) were poured with 200 ml of boiling
water and stirred for 10 minutes. Extracts were
filtered through a cotton wool, cooled at a room
temperature, diluted to 250 ml with distilled water,
and used for spectrophotometric analyses.
Caffeine isolation with chloroform. The caffeine
isolation procedure was performed according to
a modified method described by R (1994).
Briefly, 20 g of tea and 90 ml of distilled water was
refluxed for 30 min, and filtered under vacuum. The
residue was again refluxed and filtered. Obtained
filtrates were combined, 12.5 ml of Pb(CH3COO)2
solution was added, boiled (5 min), and filtered
through a Büchner funnel with silica gel layer. The
filtrate was extracted four times with chloroform
(40 ml). Combined chloroform phases were washed
with KOH solution and then with distilled water.
Chloroform was removed from extracts by rotary
evaporator. After evaporation, extracted caffeine
was weighed and expressed in mg/l.
Caffeine determination using the lead acetate
solution. This procedure is based on international
standards with some modifications (Y et al.
1992, 1993). Tea extract was treated with HCl so-
lution (5 ml), Pb(CH3COO)2 and H2SO4 solution.
Absorbance of obtained extracts was measured
at 274 nm. The content of caffeine (mg/l) was
calculated using a standard cur ve derived from
caffeine (0–250 mg/l). All measurements were
performed in triplicate.
The micromethod for the determination of
caff eine. The teas were also analysed for their
caffeine content according to the method reported
by G (1978). Briefly, tea extracts (pH =
8–9) were extracted with benzene and H2SO4.
Absorbance of extracts was read at 273 nm against
a blank (H2SO4). Results, obtained from triplicate
analyses, were calculated using a standard curve
and expressed as mg/l.
HPLC analysis of caffeine. Filtered tea extracts
and caffeine solutions were injected for HPLC
analysis according to the method reported in our
previous study (H et al. 2009). Equipment
used consisted of a Varian Pro Star Solvent Deliv-
ery System 230 and a Photodiode Array detector
Varian Pro Star 330 (Varian, Walnut Creek, USA)
with a reversed-phase column Pinnacle II C-18
(Restek, USA) (250 × 4.6 mm, 5 m i.d.). Caffeine
was identified by comparing the retention times
and spectral data with those of authentic standards.
All analyses were repeated three times.
Antioxidant capacity. Antioxidant capacity of
tea extracts and caffeine solutions (100 mg/l) was
determined using the ABTS radical scavenging
assay according to the method reported by Ret
al. (1999), and ferric reducing/antioxidant power
(FRAP) assay, carried out according to the original
method by B and S (1996).
RESULTS AND DISCUSSION
Caffeine content of five teas and two maté teas
determined by the extraction procedure with chlo-
roform ranged from 0.69% (black tea) to 1.33%
(white tea) (Figure 1). Aand K
(1986) stated that caffeine biosynthesis is the most
active in young tea leaves and buds, which implies
that white tea, produced from the youngest tea
buds should contain the highest caffeine content.
Our results confirm this thesis, since white tea
contains the highest caffeine content, and black
tea, the most processed tea type, contains the
lowest caffeine content.
In comparison to all methods used in this study
for the determination of caf feine content , the
micromethod with benzene exhibited the high-
est results. White tea contained the highest caf-
feine content (4.55%) and maté tea the lowest
(1.05%). The content of caffeine in green, black
and oolong tea ranged from 2.04% (green tea) to
3.86% (black tea), which is in agreement with the
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Czech J. Food Sci. Vol. 27, 2009, Special Issue
results obtained by G (1978), who found
2.47% caffeine in black tea, 3.45% in green tea
and 1.79% in oolong tea. Although there were no
significant differences between the order of caf-
feine content among the tested teas, the method
with lead acetate exihibited significantly lower
results with regard to other methods applied for
the determination of caffeine. Reversed-phase and
normal phase HPLC methods with UV detection
have been successfully applied for the separation
and determination of methylxanthines in a wide
range of samples. Some of the reported methods
require tedious pretreatment or do not allow sepa-
ration and quantitation of different derivatives in
the same sample (G et al. 2001). The gra-
dient method developed in this study has shown
to be appropriate for an efficient simultaneus
analysis of both methylxanthines and polyphenols
in tea (H et al. 2009). The caffeine content
of tested teas determined by the HPLC analysis
was decreasing in the following order: white tea
(3.62%) > yellow tea (3.18%) > black tea (2.79%) >
oolong tea (2.77%) > green tea (2.35%) > roasted
maté tea (1.13%) > maté tea (1.02%). According to
these results the caffeine content obtained with
HPLC analysis is the most comparable to the ones
obtained by the micromethod with benzene.
Earlier studies revealed that caffeine content is
associated to origin, genetic and environmental
variability, harvest time and processing manner
of plant material (A et al. 2000), and can
range from 24% to 40%. The displayed results
confirmed that caffeine content depends on the
age of tea leaves and processes involved in the
production of tea. Since the caffeine content was
the lowest in both types of analysed maté tea, it
is also obvious that caffeine content depends on
plant origin, indicating that the use of maté tea as
a beverage provides a milder effect in comparison
to other tea types.
Table 1. e antioxidant capacity of tea extracts and caffeine evaluated by two antioxidant assays
ABTS (mM Trolox) FRAP (mM Fe(II))
tea extract extracted caffeine tea extract extracted caffeine
White tea 6.66 ± 0.12 0.05 ± 0.01 7.77 ± 0.02 0.03 ± 0.00
Yellow tea 11.18 ± 0.12 0.07 ± 0.01 14.98 ± 0.10 0.05 ± 0.00
Green tea 8.85 ± 0.11 0.06 ± 0.01 10.20 ± 0.15 0.02 ± 0.00
Oolong tea 8.51 ± 0.12 0.07 ± 0.01 4.16 ± 0.06 0.02 ± 0.00
Black tea 4.46 ± 0.14 0.07 ± 0.01 9.02 ± 0.04 0.05 ± 0.01
Maté tea 3.23 ± 0.07 0.08 ± 0.01 5.69 ± 0.04 0.04 ± 0.00
Roasted maté tea 2.43 ± 0.10 0.07 ± 0.01 3.25 ± 0.06 0.05 ± 0.00
0
1
2
3
4
5
a b c d a b c d a b c d a b c d a b c d a b c d a b c d
White tea Yellow tea Green tea Oolong tea Black tea Maté tea Roasted Maté
% of c affeine
Figure 1. Caffeine contents of teas and maté teas determined by four different methods (a – chloroformic isolation;
b – micromethod; c – method with lead acetate; d-HPLC)
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Vol. 27, 2009, Special Issue Czech J. Food Sci.
The 2,2'-azino-bis(3-ethylbenzthiazoline-6-sul-
fonic acid) (ABTS) radical scavenging assay and
Ferric reducing antioxidant power (FRAP) assay
were applied for the evaluation of antioxidant
capacity of tea extracts and the extracted caf-
feine (determined by the isolation method with
chloroform). As could be seen in Table 1, yellow
tea exhibited the highest antioxidant capacities, in
both assays. The antioxidant capacities of extracted
caffeine was almost negligible in comparison to
the antioxidant capacities of tea, indicating that
caffeine itself does not contribute to the overall
antioxidant properties of tea.
CONCLUSION
The content of caffeine varies depending on tea
type, which is directly attributed to their process-
ing and leaf maturity. White tea, made from the
youngest tea leaves contained the highest caffeine
content, and maté and roasted maté teas the low-
est. The spectrophotometric micromethod proved
to be the best alternative for the determination of
caffeine content, exhibiting the most similar results
to the HPLC analysis. According to the obtained
results, all studied teas exhibited high antioxidant
capacity, as opposed to caffeine, indicating that the
contibution of caffeine to the antioxidant proper-
ties of these beverages is irrelevant.
References
A H., K H. (1986): Patterns of adenine
metabolism and caffeine biosynthesis in different parts
of tea seedlings. Plant Physiology, 68: 275–281.
A M.L., C G.C., S E.P. (2000):
Caffeine and theobromine in epicuticular wax of
Ilex paraguariensis A. St. Hil. Phytochemistry, 55:
853–857.
B I.F., S J.J. (1996): The ferric reducing abil-
ity of plasma (FRAP) as a measure of ‘antioxidant
power’: the FRAP assay. Analytical Biochemistry, 239:
70–76.
C M.N., R-M J.R. (1990): Chlo-
rogenic acids and purine alkaloids contents of mate
(Ilex paraguariensis) leaf and beverage. Food Chem-
istry, 35: 13–21.
D A N.M., V M.C.C., B M.S., F-
 S.L.C.,  A J.B. (2005): Multivariate
optimisation of the experimental conditions for deter-
mination of three methylxanthines by reversed-phase
high-performance liquid chromatography. Talanta,
67: 1007–1013.
G K.A., S V.F., P I.N.
(2001): Use of novel solid-phase extraction sorbent
materials for high-performance liquid chromatog-
raphy quantitation of caffeine metabolism products
methylxanthines and methyluric acids in samples of
biological origin. Journal of Chromatography B, 759:
209–218.
G D.S. (1978): A study of caffeine in tea. I. A new
spectrophotometric micro-method. II. Concentration
of caffeine in various strenghts, brands, blends, and
types of teas. The American Journal of Clinical Nutri-
tion, 31: 1727–1731.
H D., K D., B A., K G
K., I D., K D. (2009): The composi-
tion of polyphenols and methylxanthines in teas and
herbal infusions. Food Chemistry, 115: 441–448.
R V. (1994): Postupci priprave i izolacije organskih
spojeva. Školska knjiga, Zagreb.
R R., P N., P A., P A.,
Y M., R-E C. (1999): Antioxidant activity
applying an improved ABTS radical cation decolouri-
sation assay. Free Radical Biology & Medicine, 26:
1231–1237.
Y L.H., C C., C Y., L Y. (1992): The ki-
netics of green tea infusion. Journal of Food Science,
13: 3–6.
Y L.H., C Y., C C., L Y. (1993): The kinet-
ics of black tea infusion. Journal of Food Fermentation
and Industries, 19: 38–44.
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Background Herbal medication has been used for thousands of years. Although synergic efficiency of compound herbal medicine to humans have been explored for years, detailed mechanisms behind this "cocktail" medication still required open for discussion. Method With electrochemical inspections, this feasibility study tended to decipher combined interactions of synergy and antagonism of Levodopa (L-DOPA) and tea extracts of Camellia sinensis (L.) Kuntze (green tea) and Camelliaboreali-yunnanica (black tea) extracted from different temperatures via total phenolics concentrations, DPPH free radical scavenging ability, bioelectricity-generating capability of microbial fuel cells for bioenergy expression. Significance of findings According to replacement series and concentration addition (CA) methods, the most significant synergistic effect was quantitatively disclosed at the ratio of L-DOPA: tea extract at 80:20. This work provided the novel means revealing synergic efficacy of herbal medicine via the perspective of bioenergy extraction for medication. The rankings of power-stimulating efficiency in DC-MFC modules were HG2 > CG2 > HR2 > CR2. The replacement-series method indicated that both L-DOPA and tea extracts did not own tendency to out-compete each other. The CA method revealed that both L-DOPA and tea extracts were dis-similar and synergistic in action of bioenergy extraction. The most promising ratio of interactive synergy is L-DOPA: tea extract at 80:20.
Conference Paper
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Tea is one of the most popular beverages in the world. Beside its sensorial properties such as pleasant taste, aroma and flavour, it is consumed also because of its health-promoting effects, coming from the presence of various bioactive compounds including methylxanthines. Our study was aimed at characterisation of green and black tea samples based on occurrence and variability of selected methylxanthines (caffeine, theobromine and theophylline) in green tea. The content of methylxanhines was determined in green tea infusions by HPLC method. The content of caffeine varied from 19,78 to 27,22 mg/g DW. Theobromine was found in the concentrations between 1,22 and 6,04 mg/g and theophilline in concentrations between 0,52 and 1,28 mg/g DW. Statistical analysis showed as the important factors of differences between green and black tea samples content of teophylline and teobromine. Less important was content of caffeine, which values were varying in the similar range in black and green samples.
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Black tea is a popular and widely consumed beverage in Iran. Due to the possibility of transferring heavy metals and mycotoxins to the food chain, its potential hazards should be considered. In this study, forty samples of black tea imported to Bushehr port were collected. The physicochemical and microbial properties and their contamination with heavy metals including copper, lead, mercury, arsenic, and cadmium, as well as Aflatoxin B1 were evaluated. Moreover, to evaluate the health effects of exposure to heavy metals, both Hazard Index (HI) and Hazard Quotient (HQ) values were calculated. The concentration of lead, cadmium, arsenic and copper were: 0.63, 0.061, 0.01, 29.93 mg kg⁻¹ in Sri Lankan, 0.65, 0.063, 0.11, 21.3 mg kg⁻¹ in Indian, 0.61, 0.055, 0.085, 14.82 mg kg⁻¹ in Kenyan and 1.71, 0.06, 0.1, 37.87 mg kg⁻¹ in Vietnamese tea samples, respectively. No Hg contamination was observed. Aflatoxin B1 was only detected in Kenyan and Vietnamese samples (24.33 and 42.66 ng g⁻¹, respectively). The calculation of HQ value for all heavy metals and HI value for all brands, which all were below one, showed that there is no non-carcinogenic adverse health effect through consuming these black teas for the adult consumers.
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A method for the screening of antioxidant activity is reported as a decolorization assay applicable to both lipophilic and hydrophilic antioxidants, including flavonoids, hydroxycinnamates, carotenoids, and plasma antioxidants. The pre-formed radical monocation of 2,2'-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS*+) is generated by oxidation of ABTS with potassium persulfate and is reduced in the presence of such hydrogen-donating antioxidants. The influences of both the concentration of antioxidant and duration of reaction on the inhibition of the radical cation absorption are taken into account when determining the antioxidant activity. This assay clearly improves the original TEAC assay (the ferryl myoglobin/ABTS assay) for the determination of antioxidant activity in a number of ways. First, the chemistry involves the direct generation of the ABTS radical monocation with no involvement of an intermediary radical. Second, it is a decolorization assay; thus the radical cation is pre-formed prior to addition of antioxidant test systems, rather than the generation of the radical taking place continually in the presence of the antioxidant. Hence the results obtained with the improved system may not always be directly comparable with those obtained using the original TEAC assay. Third, it is applicable to both aqueous and lipophilic systems.
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Contents of purine alkaloids in different parts of tea (Camellia sinensis L. cv. Yabukita) seedlings, seeds and tissue cultures were determined with high-performance liquid chromatography. More than 99% of the caffeine detected was in the leaves of the 4-month-old seedlings. The amount expressed per g fresh weight was higher in older leaves. Theobromine, a precursor of caffeine biosynthesis, was found only in younger leaves. Zero or only trace amounts of theophylline, a degradation product of caffeine, were found in the seedlings. Almost all the caffeine in tea seeds was found in the seed coats. Theobromine and theophilline could not be detected in any part of the seeds. Tracer experiments using [8-14C]-adenine indicate that (i) caffeine biosynthesis from [8-14C]-adenine occurs only in younger leaves,(ii) “salvage” of [8-14C]-adenine for nucleic acid synthesis takes place in all parts of the seedlings, (iii) considerable degradation of [8-14C]-adenine by conventional purine degradation pathway via uric acid takes place in roots and lower parts of stem tissue. The results strongly suggest that caffeine is synthesized in younger leaves and accumulated within the leaves. Both caffeine contents and its synthetic activity from adenine were extremely low in tissue culture of tea.
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Five samples of Maté leaf have been analysed for chlorogenic acids, caffeine and theobromine, and found to comprise two distinct categories of product. Category 1 produced brown extracts and infusions with relatively small contents of alkaloids and total chlorogenic acids whereas Category 2 produced green extracts and infusions with larger total alkaloids contents and much larger total chlorogenic acids contents. The green Maté samples contained a quercetin glycoside, and were a convenient source from which to isolate 3-caffeoylquinic acid and 3,5-dicaffeoylquinic acid. Neither condensed nor hydrolysable tannins were detected.
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Caffeine (1,3,7-trimethylxanthine), theobromine (3,7-dimethylxanthine) and theophylline (1,3-dimethylxanthine) are the most important naturally occurring methylxanthines. Caffeine is a constituent of coffee and other beverage and included in many medicines. Theobromine and theophylline are formed as metabolites of caffeine in humans, and are also present in tea, cocoa and chocolate products. In order to improve the chromatographic resolution (R(s)) with a good analysis time, experimental designs were applied for multivariate optimisation of the experimental conditions of an isocratic reversed-phase high-performance liquid chromatographic (RP-HPLC) method used for the simultaneous determination of caffeine, theobromine and theophylline. The optimisation process was carried out in two steps using full three-level factorial designs. The factors optimised were: flow rate and mobile phase composition. Optimal conditions for the separation of the three methylxanthines were obtained using a mixture of water/ethanol/acetic acid (75:24:1%, v/v/v) as mobile phase and a flow rate of 1.0mLmin(-1). The RP-HPLC/UV method was validated in terms of limit of detection (LOD), limit of quantitation (LOQ), linearity, recovery and the precision, calculated as relative standard deviation (R.S.D.). In these conditions, the LOD was 0.10mugL(-1) for caffeine, 0.07mugL(-1) for theobromine and 0.06mugL(-1) for theophylline. The proposed method is fast, requires no extraction step or derivatization and was suitable for quantification of these methylxanthines in coffee, tea and human urine samples.
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A new spectrophotometric micro-method for the determination of caffeine in tea is described. This method is then used to evaluate the caffeine content of a variety of brands and blends of bagged and loose hot tea prepared in different strengths and by different brewing methods. In addition, the caffeine content of instant tea, ice tea, and Mr. Coffee automatic tea is evaluated.
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A simple, automated test measuring the ferric reducing ability of plasma, the FRAP assay, is presented as a novel method for assessing "antioxidant power." Ferric to ferrous ion reduction at low pH causes a colored ferrous-tripyridyltriazine complex to form. FRAP values are obtained by comparing the absorbance change at 593 nm in test reaction mixtures with those containing ferrous ions in known concentration. Absorbance changes are linear over a wide concentration range with antioxidant mixtures, including plasma, and with solutions containing one antioxidant in purified form. There is no apparent interaction between antioxidants. Measured stoichiometric factors of Trolox, alpha-tocopherol, ascorbic acid, and uric acid are all 2.0; that of bilirubin is 4.0. Activity of albumin is very low. Within- and between-run CVs are <1.0 and <3.0%, respectively, at 100-1000 micromol/liter. FRAP values of fresh plasma of healthy Chinese adults: 612-1634 micromol/liter (mean, 1017; SD, 206; n = 141). The FRAP assay is inexpensive, reagents are simple to prepare, results are highly reproducible, and the procedure is straightforward and speedy. The FRAP assay offers a putative index of antioxidant, or reducing, potential of biological fluids within the technological reach of every laboratory and researcher interested in oxidative stress and its effects.
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A method for the screening of antioxidant activity is reported as a decolorization assay applicable to both lipophilic and hydrophilic antioxidants, including flavonoids, hydroxycinnamates, carotenoids, and plasma antioxidants. The pre-formed radical monocation of 2,2'-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS*+) is generated by oxidation of ABTS with potassium persulfate and is reduced in the presence of such hydrogen-donating antioxidants. The influences of both the concentration of antioxidant and duration of reaction on the inhibition of the radical cation absorption are taken into account when determining the antioxidant activity. This assay clearly improves the original TEAC assay (the ferryl myoglobin/ABTS assay) for the determination of antioxidant activity in a number of ways. First, the chemistry involves the direct generation of the ABTS radical monocation with no involvement of an intermediary radical. Second, it is a decolorization assay; thus the radical cation is pre-formed prior to addition of antioxidant test systems, rather than the generation of the radical taking place continually in the presence of the antioxidant. Hence the results obtained with the improved system may not always be directly comparable with those obtained using the original TEAC assay. Third, it is applicable to both aqueous and lipophilic systems.
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Caffeine and theobromine were identified and quantified in leaf epicuticular waxes of Ilex paraguariensis A. St.-Hil. (Aquifoliaceae). The total epicuticular leaf wax content was ca. 0.5% on average of dry leaf weight. Epicuticular caffeine and theobromine contents varied from 0.16 to 127.6 microg/mg and from 0 to 9.5 microg/mg of wax, respectively. For some selected samples, the intracellular methylxanthine concentration was also determined. A positive correlation was found between inner and epicuticular caffeine contents.
An automated reversed-phase high-performance liquid chromatographic (RP-HPLC) method, using a linear gradient elution, is described for the simultaneous analysis of caffeine and metabolites according to their elution order: 7-methyluric acid, 1-methyluric acid, 7-methylxanthine, 3-methylxanthine, 1-methylxanthine, 1,3-dimethyluric acid, theobromine, 1,7-dimethyluric acid, paraxanthine and theophylline. The analytical column, an MZ Kromasil C4, 250 x 4 mm, 5 microm, was operated at ambient temperature with back pressure values of 80-110 kg/cm2. The mobile phase consisted of an acetate buffer (pH 3.5)-methanol (97:3, v/v) changing to 80:20 v/v in 20 min time, delivered at a flow-rate of 1 ml/min. Paracetamol was used as internal standard at a concentration of 6.18 ng/microl. Detection was performed with a variable wavelength UV-visible detector at 275 nm, resulting in detection limits of 0.3 ng per 10-microl injection, while linearity held up to 8 ng/microl for most of analytes, except for paraxanthine and theophylline, for which it was 12 ng/microl and for caffeine for which it was 20 ng/microl. The statistical evaluation of the method was examined performing intra-day (n=6) and inter-day calibration (n=7) and was found to be satisfactory, with high accuracy and precision results. High extraction recoveries from biological matrices: blood serum and urine ranging from 84.6 to 103.0%, were achieved using Nexus SPE cartridges with hydrophilic and lipophilic properties and methanol-acetate buffer (pH 3.5) (50:50, v/v) as eluent, requiring small volumes, 40 microl of blood serum and 100 microl of urine.