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Food Additives & Contaminants: Part B: Surveillance Publication details, including instructions for authors and subscription information: http://www.tandfonline.com/loi/tfab20
Chromium level and intake from Chinese made tea ab
c
a
a
b
b
Wei-hao Li , Hai-ping Zhou , Ning Li , Sai-di Wang , Xiao-juan Liu , Zeng-jun Jin , Yan-zhen d
Bu & Zhi-xue Liu
a
a
School of Life Science and Technology, Tongji University, Shanghai, China
b
Handan Municipal Centre for Disease Control and Prevention, Handan, China
c
Handan Municipal Health Bureau, Handan, China
d
College of Life Science, Henan Normal University, Xinxiang, China Accepted author version posted online: 09 Jul 2013.Published online: 07 Aug 2013.
To cite this article: Wei-hao Li, Hai-ping Zhou, Ning Li, Sai-di Wang, Xiao-juan Liu, Zeng-jun Jin, Yan-zhen Bu & Zhi-xue Liu (2013) Chromium level and intake from Chinese made tea, Food Additives & Contaminants: Part B: Surveillance, 6:4, 289-293, DOI: 10.1080/19393210.2013.822934 To link to this article: http://dx.doi.org/10.1080/19393210.2013.822934
PLEASE SCROLL DOWN FOR ARTICLE Taylor & Francis makes every effort to ensure the accuracy of all the information (the “Content”) contained in the publications on our platform. However, Taylor & Francis, our agents, and our licensors make no representations or warranties whatsoever as to the accuracy, completeness, or suitability for any purpose of the Content. Any opinions and views expressed in this publication are the opinions and views of the authors, and are not the views of or endorsed by Taylor & Francis. The accuracy of the Content should not be relied upon and should be independently verified with primary sources of information. Taylor and Francis shall not be liable for any losses, actions, claims, proceedings, demands, costs, expenses, damages, and other liabilities whatsoever or howsoever caused arising directly or indirectly in connection with, in relation to or arising out of the use of the Content. This article may be used for research, teaching, and private study purposes. Any substantial or systematic reproduction, redistribution, reselling, loan, sub-licensing, systematic supply, or distribution in any form to anyone is expressly forbidden. Terms & Conditions of access and use can be found at http:// www.tandfonline.com/page/terms-and-conditions
Food Additives & Contaminants: Part B, 2013 Vol. 6, No. 4, 289–293, http://dx.doi.org/10.1080/19393210.2013.822934
Chromium level and intake from Chinese made tea Wei-hao Lia,b, Hai-ping Zhouc, Ning Lia, Sai-di Wanga, Xiao-juan Liub, Zeng-jun Jinb, Yan-zhen Bud* and Zhi-xue Liua* a School of Life Science and Technology, Tongji University, Shanghai, China; bHandan Municipal Centre for Disease Control and Prevention, Handan, China; cHandan Municipal Health Bureau, Handan, China; dCollege of Life Science, Henan Normal University, Xinxiang, China
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(Received 25 March 2013; final version received 3 July 2013) Tea is a popular drink around the world. It is also one of the sources of metal intake. The objectives of this study were to assess chromium (Cr) intake from popular green, oolong, black and Pu-erh tea. In total, 128 Chinese made teas were analysed and concentration differences among four types of tea were explored. Black tea contained highest total Cr, which varied between 0.63 and 17.60 mg/kg. The lowest content was found in the green tea samples, between 0.26 and 1.30 mg/ kg. Cr(III) and Cr(VI) in black tea were higher than in other types of tea. Cr(III), Cr(VI) and total Cr concentration in different tea infusions were also analysed. The results suggest that drinking tea is an effective way for Cr intake and the risk of adults and children being chronically intoxicated by tea infusions is low. Keywords: chromium; tea; tea infusion; intake; safety
Introduction Chromium (Cr) is widely used in the chemical industry for different applications such as pigments, metal plating or leather tanning, and in chemical production such as chemical synthesis and as catalysts (Fibbi et al. 2012). As a result, different species of Cr can be released into the environment (soil, surface and ground water) and are available to plants and humans (Mukherjee 1998; Rowbotham et al. 2000; Redondo-Gomez et al. 2011). Cr species exist in the environment mainly in two oxidation states, Cr(III) and Cr(VI) (Kotas & Stasicka 2000; Unceta et al. 2010), which have contrasting physiological effects. Cr(III) is considered an essential micronutrient in the human diet and is widely used as a nutritional supplement for humans and animals. It plays an important role in human physiology by stimulating glucose metabolism, controlling blood cholesterol levels, stimulating the synthesis of protein, increasing resistance to pain and suppressing hunger pain (Nickens et al. 2010). In contrast, Cr(VI) is toxic and carcinogenic to humans. It is suspected of being extremely toxic after inhalation and oral exposure with effects on the respiratory tract, liver, kidney, gastrointestinal and immune systems, possibly on blood and dermal exposure may cause dermatitis, sensitivity and ulceration of the skin (Debiasi et al. 2001). Cr(VI) has been recognised as a class I human carcinogen by the International Agency for Research on Cancer (IARC). Tea, a product made from leaf and bud of the plant Camellia sinensis, is the second most consumed beverage in the world next to water, well ahead of coffee, beer, wine and carbonated soft drinks (Schlesier et al. 2012). *Corresponding author. Email: [email protected] © 2013 Taylor & Francis
Originating from China, tea has gained the world’s taste in the past 2000 years. Since tea is known to contain several minerals, trace elements and antioxidants, it is considered a healthy beverage (McKay & Blumberg 2002; Yu et al. 2007). However, some undesirable elements, such as Pb, Hg, Cd and Cr, are of concern for tea consumers (Han et al. 2005), since tea (Camellia sinensis) can accumulate heavy metals. Rapid industrialisation and urbanisation in China over the last two decades has resulted in a heavy metal burden to the environment (Zheng et al. 2011). The concentration of Cr in tea was reported in several recent studies (Karak & Bhagat 2010; Braganca et al. 2011). However, the data described in these studies focused only on total Cr and Cr(VI) (Wrobel et al. 2000; Seenivasan et al. 2008; Mandiwana et al. 2011). According to the contrasting effect of Cr(III) and Cr(VI), Cr in tea may have both beneficial and adverse effects on human health. Therefore, speciation of Cr as Cr(III) and Cr(VI) in tea and tea infusion is essential. On the basis of their degree of fermentation, Chinese made tea can be categorised into four different types (Wang et al. 2010): “non-fermented” green tea (produced by drying and steaming fresh leaves to inactivate the polyphenol oxidase and, thus, no oxidation occurs); “semi-fermented” oolong tea (produced when fresh leaves are subjected to a partial fermentation stage before drying); “full-fermented” black tea (which undergoes a fermentation stage by polyphenol oxidase before drying and steaming) and “post-fermented” Pu-erh tea (which undergoes a longer post-harvest fermentation stage by polyphenol oxidase and microorganisms). In this study, a survey
290
W.-h. Li et al.
was conducted on the speciation of Cr in Chinese made tea and tea infusion, to understand the benefit and risk of Cr intake from Chinese made tea.
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Materials and methods Reagents and standards The working standards of Cr were prepared by serial dilution of 1000 mg/L Cr(VI) and 1000 mg/L Cr(III) solutions (National Standard Center, China). The standard tea sample GBW07605 (National Standard Center, China) was applied as control sample (Cr content = 0.80 ± 0.03 mg/kg). Ultra-pure water (resistivity18.3 MΩ cm), obtained from a Milli-Q water purification system (Millipore, USA), was used to prepare all standards and samples. Supra-pure HNO3 and H2O2 (National Medicines, China) were used to digest tea samples in a microwave before Cr determination. Sample collection and preparation According to the fermentation degree, 128 tea samples (200 g each) were collected in China during 2011–2012, including 35 green tea, 32 oolong tea, 32 black tea and 29 Pu-erh tea. The samples were dried in an oven at 105°C for 2 hours and grounded into a 40 mesh powder with a grinder prior to use. To determine the Cr content consumed by tea infusion, an amount of 2.0 g of tea was added to a glass tea-maker and 200 mL of boiling deionised water was poured in to simulate the traditional way of tea making (Lung et al. 2003). After 5 min, tea infusion was filtered for analysis. Digestion Approximately 0.25 g tea sample was weighed into a Teflon coated tube and 3.0 mL of concentrated HNO3 and 2.0 mL concentrated H2O2 were added. The mixtures were allowed to predigest for 15 min at room temperature and then transferred to the microwave oven. Digestion was carried out at 150°C for 10 min at a maximum pressure of 800 psi. Samples were cooled to room temperature and diluted to 25 mL with ultra-pure de-ionised water. A blank sample was digested accordingly. Separation of Cr(III) and Cr(VI) Nowadays, the most prevalent method for determining Cr(VI) in soils, sediments and sludges is the EPA revised method 3060A (US Environmental Protection Agency 1995), which is based on an alkaline digestion procedure according to the findings of Mandiwana et al. (2011). In this study, a modified separated method based on EPA Method 3060A was employed for extraction of
Cr(III) and Cr(VI): 0.25 g tea sample (or 2.5 mL tea infusion) is digested using 25 mL of a solution of 0.28 mol/L Na2CO3 and 0.5 mol/L NaOH (pH about 12) and 100 mg MgCl2 and boiled for 5 min. Under these conditions, Cr(VI) is transferred into the solution due to the high alkalinity of the media while the remaining Cr(III) species are in a state of insoluble carbonate, hydroxide or oxide compounds. The amount of Cr(III) can be calculated by subtracting Cr(VI) from the total Cr. Analysis by atomic absorption spectrometry All measurements were carried out using a Hitachi Z5000 atomic absorption spectrometer (Hitachi, Japan) with Zeeman-effect background correction equipped with a chromium hollow cathode lamp operating at 9 mA. Measurements were performed at 359.3 nm with a spectral band pass of 1.3 nm. Argon with a purity of 99.99% (Praxair, USA) was used as purge and protective gas with a flow rate of 30 mL/min during all stages, except atomisation when the gas flow was stopped. Integrated absorbance (peak area) was used for signal evaluation and quantification. The limits of detection (LOD) and quantification (LOQ) were calculated from 15 measurements of analytical blanks. The values of LOD (calculated as 3 × SD) and LOQ (calculated as 10 × SD) were 0.1 μg/L and 0.3 μg/L, respectively. With respect to the tea samples, assuming that 0.25 g of tea was used for preparation of 25.0 mL of tea infusion, LOD and LOQ were 0.01 mg/kg and 0.03 mg/kg, respectively. Calibration curves with standards were used for all determinations. The graphite furnace temperature programme is given in Table 1. The accuracy of the measurements was assessed by applying the standard addition method in tea and tea infusion samples. Recoveries of spiked samples were 97.4% from tea and 95.3% from tea infusion. The method repeatability was evaluated by the relative standard deviation (RSD), which was 3.7% and 4.3% (n = 7) in tea and tea infusion, respectively. Statistical analysis Statistical analyses were performed using SPSS 18.0 software packages (SPSS Inc., Chicago, IL, USA). All determinations were carried out in triplicate. Means and Table 1.
Temperature programme for the determination of Cr. Temperature (°C)
Time (sec)
Stage
Start
End
Ramp
Drying Ashing Atomising Cleaning
80 700 2800 2800
140 700 2800 2800
40 20
Hold
5 4
Food Additives & Contaminants: Part B standard error of means are presented in the tables as an indication of variation. One-way ANOVA test was used to compare the means of different groups and p < 0.01 was considered to point out a significant difference between the compared groups.
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Results and discussion Cr in Chinese made tea The analytical results of tea samples are summarised in Table 2. Among the tea varieties, black tea contained most Cr, statistically and significantly (p < 0.01) different from other types of tea. The content of Cr(III) and Cr(VI) in black tea was accordingly higher than other types of tea. It suggested that the content of Cr(III) and Cr(VI) were positively correlated with the level of total chromium in China made tea. Cr transfer into tea infusion Cr(III), Cr(VI) and total Cr concentration in tea infusion were summarised in Table 3. The transfer ratio of Cr(III) and Cr(VI) from tea leaves into infusions was determined by comparing Cr content between teas and infusions. The results are shown in Figure 1. The Cr(III) transfer level in green tea was significantly lower (p < 0.01) than in other types. Contrary to green tea, the black tea, oolong tea and Pu-erh tea are all fermented under controlled conditions. Such a difference in leaf processing method causes different reactions in the leaf material, e.g. oxidation of phenolics such as catechols or tannins to give polyphenols (Lin et al. 1998; Wang et al. 2012). Consequently, Cr(III) can form soluble complexes with organic ligands during the
Table 2.
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fermentation process, thus enhancing the transfer ratio in tea infusion. The transfer ratio of Cr(VI) was 37.8% in green tea, 37.4% in oolong tea, 38.7% in black tea and 38.2% in Pu-erh tea. There was no significant difference (p > 0.01) between the various types of tea. Cr(VI) transfer was significantly higher (p < 0.01), almost twofold to threefold, than that of Cr(III), indicating that solubility of Cr(VI) was higher than that of Cr(III).
Estimation of chromium intake from tea ATI (Adequate Daily Intake) is defined as the amount of tea infusions that provide adequate intake of total Cr every day and ADI (Acceptable Daily Intake) is the amount of tea infusions that can be consumed every day without detectable chronic effects. ATI and ADI can be estimated based on RNI (Recommended Nutrient Intake) and UL (Tolerable Upper Intake Level), respectively: ATIð1=dayÞ ¼ RNI=Cavg ðμg=lÞ
(1)
ADIð1=dayÞ ¼ UL=Cavg ðμg=lÞ
(2)
where RNI is 10 μg/day for children and 50 μg/day for adults, UL is 100 μg/day for children and 250 μg/day for adults; Cavg is the mean total chromium concentration of tea infusions. ATI and ADI were calculated for different tea infusions, both for adults and children. The results are given in Table 4 and show that the chance of adults and children being chronically and acutely intoxicated by drinking tea was low. On the other hand, the results suggested that
Cr(III), Cr(VI) and total Cr concentrations in tea samples (mg/kg).
Type Green tea Oolong tea Black tea Pu-erh tea
Cr(III)
Number of samples ± SD
Range
35 32 32 29
0.23–1.18 0.30–1.60 0.45–14.73 0.28–2.12
Cr(VI)
Mean ± SD 0.79 0.90 4.18 0.96
± ± ± ±
0.36 0.35 3.06 0.52
Range 0.03–0.12 0.03–0.25 0.18–2.87 0.04–0.31
Total Cr
Mean ± SD 0.08 0.16 1.16 0.21
± ± ± ±
0.02 0.07 0.56 0.04
Range 0.26–1.30 0.33–1.85 0.63–17.60 0.32–2.43
Mean ± SD 0.87 1.02 5.38 1.23
± ± ± ±
0.38 0.42 3.62 0.56
Table 3. Cr(III), Cr(VI) and total Cr concentrations in tea infusions (µg/L).
Type of tea Green tea Oolong tea Black tea Pu-erh tea
Number of samples 35 32 32 29
Cr(III) Range 0.2–1.2 0.5–2.5 0.8–22.0 0.4–3.4
Cr(VI)
Mean ± SD 0.8 1.4 7.1 1.6
± ± ± ±
0.2 0.4 3.8 0.8
Range 0.1–0.6 0.1–1.0 0.7–11.4 0.2–1.2
Total Cr
Mean ± SD 0.3 0.6 4.5 0.8
± ± ± ±
0.2 0.3 2.6 0.4
Range 0.3–1.8 0.6–3.5 1.5–33.4 0.6–4.6
Mean ± SD 1.2 2.0 11.3 2.3
± ± ± ±
0.4 0.7 6.4 1.2
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Figure 1.
Transfer ratios of Cr(III) and Cr(VI) from tea leaves into infusions.
Table 4.
Estimated total Cr’s ATI and ADI for tea infusions. ATI (l/day)
Table 5.
ADI (l/day)
Type of tea
Adult
Children
Adult
Children
Green tea Oolong tea Black tea Pu-erh tea
42 25 4 22
8 5 1 4
208 125 22 109
83 50 9 43
Cr(III) and Cr(VI) intake per cup of tea (µg). Cr(III)
Type of tea
Number of samples
Range
Green tea Oolong tea Black tea Pu-erh tea
35 32 32 29
0.04–0.24 0.10–0.50 0.16–4.40 0.08–0.68
drinking tea is an effective way for Cr intake. Tea drinkers who like more concentrated tea would prepare the tea for a longer period of time (>5 min) or use more tea leaves. In those cases, more Cr would be infused in water due to longer preparation time or use of more tea leaves, thus increasing more Cr consumption than reported here. Furthermore, this work was conducted with deionised water. Thus, intake concentrations of Cr in actual tea infusion with tap water could be higher than the current measurements. Literature data for Cr mainly focuses on total Cr rather than Cr speciation. Until now, there is no defined
Cr(VI) Mean ± SD 0.16 0.28 1.42 0.32
± ± ± ±
0.04 0.08 0.76 0.16
Range 0.02–0.12 0.02–0.2 0.14–2.28 0.04–0.24
Mean ± SD 0.06 0.12 0.90 0.16
± ± ± ±
0.04 0.06 0.52 0.08
maximum acceptable concentration of Cr(III) and Cr(VI). As indicated earlier, Cr(III) and Cr(VI) have contradicting physiological effects. In order to discuss Cr(III) and Cr (VI) intake from daily tea consumption, this study gives average Cr(III) and Cr(VI) content per cup (200 µg/mL tea infusion), presented in Table 5, which can be evaluated to draft limits for chromium in tea. Conclusion Cr level is significantly higher in black tea than in other types. From the database which was generated in this
Food Additives & Contaminants: Part B study to estimate the Cr intake from tea, it could be concluded that drinking tea is a reliable and effective way for Cr intake, the risk of adults and children being chronically intoxicated by tea infusions being low. Acknowledgement This work was supported by the National Natural Scientific foundation of China (Grant 31140037).
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References Braganca VLC, Melnikov P, Zanoni LZ. 2011. Trace elements in different brands of yerba mate tea. Biol Trace Elem Res. 144:1197–1204. Debiasi R, Nicolau M, Buzica D, Mitran E. 2001. Certified reference material usable in controlling the environment pollution with hexavalent chromium, manganese, chloride and nitrogen/nitrites dioxide. Rev Chim-Bucharest. 52:240–244. Fibbi D, Doumett S, Lepri L, Checchini L, Gonnelli C, Coppini E, Del Bubba M. 2012. Distribution and mass balance of hexavalent and trivalent chromium in a subsurface, horizontal flow (SF-h) constructed wetland operating as post-treatment of textile wastewater for water reuse. J Hazard Mater. 199:209–216. Han WY, Shi YZ, Ma LF, Ruan JY. 2005. Arsenic, cadmium, chromium, cobalt, and copper in different types of Chinese tea. B Environ Contam Tox. 75:272–277. Karak T, Bhagat RM. 2010. Trace elements in tea leaves, made tea and tea infusion: a review. Food Res Int. 43:2234–2252. Kotas J, Stasicka Z. 2000. Chromium occurrence in the environment and methods of its speciation. Environ Pollut. 107:263–283. Lin JK, Lin CL, Liang YC, Lin-Shiau SY, Juan IM. 1998. Survey of catechins, gallic acid, and methylxanthines in green, oolong, pu-erh, and black teas. J Agric Food Chem. 46:3635–3642. Lung SC, Hsiao PK, Chiang KM. 2003. Fluoride concentrations in three types of commercially packed tea drinks in Taiwan. J Expo Anal Environ Epidemiol. 13:66–73. Mandiwana KL, Panichev N, Panicheva S. 2011. Determination of chromium(VI) in black, green and herbal teas. Food Chem. 129:1839–1843. McKay DL, Blumberg JB. 2002. The role of tea in human health: an update. J Am Coll Nutr. 21:1–13.
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Mukherjee AB. 1998. Chromium in the environment of Finland. Sci Total Environ. 217:9–19. Nickens KP, Patierno SR, Ceryak S. 2010. Chromium genotoxicity: a double-edged sword. Chem Biol Interact. 188:276–288. Redondo-Gomez S, Mateos-Naranjo E, Vecino-Bueno I, Feldman SR. 2011. Accumulation and tolerance characteristics of chromium in a cordgrass Cr-hyperaccumulator, Spartina argentinensis. J Hazard Mater. 185:862–869. Rowbotham AL, Levy LS, Shuker LK. 2000. Chromium in the environment: an evaluation of exposure of the UK general population and possible adverse health effects. J Toxicol Env Heal B. 3:145–178. Schlesier K, Kuhn B, Kiehntopf M, Winnefeld K, Roskos M, Bitsch R, Bohm V. 2012. Comparative evaluation of green and black tea consumption on the iron status of omnivorous and vegetarian people. Food Res Int. 46:522–527. Seenivasan S, Manikandan N, Muraleedharan NN. 2008. Chromium contamination in black tea and its transfer into tea brew. Food Chem. 106:1066–1069. Unceta N, Seby F, Malherbe J, Donard OF. 2010. Chromium speciation in solid matrices and regulation: a review. Anal Bioanal Chem. 397:1097–1111. [USEPA] US Environmental Protection Agency. 1995. Alkaline digestion of hexavalent chromium, Method 3060A, in test methods for evaluating solid waste – physical/chemical methods, SW-846, update 3. Washington (DC): Government Printing Office. Wang D, Xiao R, Hu XT, Xu KL, Hou Y, Zhong Y, Meng J, Fan BL, Liu LG. 2010. Comparative safety evaluation of Chinese Pu-erh green tea extract and Pu-erh black tea extract in Wistar rats. J Agric Food Chem. 58:1350–1358. Wang YF, Shao SH, Xu P, Chen H, Lin-Shiau SY, Deng YT, Lin JK. 2012. Fermentation process enhanced production and bioactivities of oolong tea polysaccharides. Food Res Int. 46:158–166. Wrobel K, Wrobel K, Urbina EMC. 2000. Determination of total aluminum, chromium, copper, iron, manganese, and nickel and their fractions leached to the infusions of black tea, green tea, Hibiscus sabdariffa, and Ilex paraguariensis (mate) by ETA-AAS. Biol Trace Elem Res. 78:271–280. Yu F, Sheng JC, Xu J, An XX, Hu QH. 2007. Antioxidant activities of crude tea polyphenols, polysaccharides and proteins of selenium-enriched tea and regular green tea. Eur Food Res Technol. 225:843–848. Zheng HR, Li RN, Cao SX. 2011. Comment on “chromium contamination accident in China: viewing environment policy of China.” Environ Sci Technol. 45:9822–9822.
Food Additives & Contaminants: Part B: Surveillance Publication details, including instructions for authors and subscription information: http://www.tandfonline.com/loi/tfab20
Chromium level and intake from Chinese made tea ab
c
a
a
b
b
Wei-hao Li , Hai-ping Zhou , Ning Li , Sai-di Wang , Xiao-juan Liu , Zeng-jun Jin , Yan-zhen d
Bu & Zhi-xue Liu
a
a
School of Life Science and Technology, Tongji University, Shanghai, China
b
Handan Municipal Centre for Disease Control and Prevention, Handan, China
c
Handan Municipal Health Bureau, Handan, China
d
College of Life Science, Henan Normal University, Xinxiang, China Accepted author version posted online: 09 Jul 2013.Published online: 07 Aug 2013.
To cite this article: Wei-hao Li, Hai-ping Zhou, Ning Li, Sai-di Wang, Xiao-juan Liu, Zeng-jun Jin, Yan-zhen Bu & Zhi-xue Liu (2013) Chromium level and intake from Chinese made tea, Food Additives & Contaminants: Part B: Surveillance, 6:4, 289-293, DOI: 10.1080/19393210.2013.822934 To link to this article: http://dx.doi.org/10.1080/19393210.2013.822934
PLEASE SCROLL DOWN FOR ARTICLE Taylor & Francis makes every effort to ensure the accuracy of all the information (the “Content”) contained in the publications on our platform. However, Taylor & Francis, our agents, and our licensors make no representations or warranties whatsoever as to the accuracy, completeness, or suitability for any purpose of the Content. Any opinions and views expressed in this publication are the opinions and views of the authors, and are not the views of or endorsed by Taylor & Francis. The accuracy of the Content should not be relied upon and should be independently verified with primary sources of information. Taylor and Francis shall not be liable for any losses, actions, claims, proceedings, demands, costs, expenses, damages, and other liabilities whatsoever or howsoever caused arising directly or indirectly in connection with, in relation to or arising out of the use of the Content. This article may be used for research, teaching, and private study purposes. Any substantial or systematic reproduction, redistribution, reselling, loan, sub-licensing, systematic supply, or distribution in any form to anyone is expressly forbidden. Terms & Conditions of access and use can be found at http:// www.tandfonline.com/page/terms-and-conditions
Food Additives & Contaminants: Part B, 2013 Vol. 6, No. 4, 289–293, http://dx.doi.org/10.1080/19393210.2013.822934
Chromium level and intake from Chinese made tea Wei-hao Lia,b, Hai-ping Zhouc, Ning Lia, Sai-di Wanga, Xiao-juan Liub, Zeng-jun Jinb, Yan-zhen Bud* and Zhi-xue Liua* a School of Life Science and Technology, Tongji University, Shanghai, China; bHandan Municipal Centre for Disease Control and Prevention, Handan, China; cHandan Municipal Health Bureau, Handan, China; dCollege of Life Science, Henan Normal University, Xinxiang, China
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(Received 25 March 2013; final version received 3 July 2013) Tea is a popular drink around the world. It is also one of the sources of metal intake. The objectives of this study were to assess chromium (Cr) intake from popular green, oolong, black and Pu-erh tea. In total, 128 Chinese made teas were analysed and concentration differences among four types of tea were explored. Black tea contained highest total Cr, which varied between 0.63 and 17.60 mg/kg. The lowest content was found in the green tea samples, between 0.26 and 1.30 mg/ kg. Cr(III) and Cr(VI) in black tea were higher than in other types of tea. Cr(III), Cr(VI) and total Cr concentration in different tea infusions were also analysed. The results suggest that drinking tea is an effective way for Cr intake and the risk of adults and children being chronically intoxicated by tea infusions is low. Keywords: chromium; tea; tea infusion; intake; safety
Introduction Chromium (Cr) is widely used in the chemical industry for different applications such as pigments, metal plating or leather tanning, and in chemical production such as chemical synthesis and as catalysts (Fibbi et al. 2012). As a result, different species of Cr can be released into the environment (soil, surface and ground water) and are available to plants and humans (Mukherjee 1998; Rowbotham et al. 2000; Redondo-Gomez et al. 2011). Cr species exist in the environment mainly in two oxidation states, Cr(III) and Cr(VI) (Kotas & Stasicka 2000; Unceta et al. 2010), which have contrasting physiological effects. Cr(III) is considered an essential micronutrient in the human diet and is widely used as a nutritional supplement for humans and animals. It plays an important role in human physiology by stimulating glucose metabolism, controlling blood cholesterol levels, stimulating the synthesis of protein, increasing resistance to pain and suppressing hunger pain (Nickens et al. 2010). In contrast, Cr(VI) is toxic and carcinogenic to humans. It is suspected of being extremely toxic after inhalation and oral exposure with effects on the respiratory tract, liver, kidney, gastrointestinal and immune systems, possibly on blood and dermal exposure may cause dermatitis, sensitivity and ulceration of the skin (Debiasi et al. 2001). Cr(VI) has been recognised as a class I human carcinogen by the International Agency for Research on Cancer (IARC). Tea, a product made from leaf and bud of the plant Camellia sinensis, is the second most consumed beverage in the world next to water, well ahead of coffee, beer, wine and carbonated soft drinks (Schlesier et al. 2012). *Corresponding author. Email: [email protected] © 2013 Taylor & Francis
Originating from China, tea has gained the world’s taste in the past 2000 years. Since tea is known to contain several minerals, trace elements and antioxidants, it is considered a healthy beverage (McKay & Blumberg 2002; Yu et al. 2007). However, some undesirable elements, such as Pb, Hg, Cd and Cr, are of concern for tea consumers (Han et al. 2005), since tea (Camellia sinensis) can accumulate heavy metals. Rapid industrialisation and urbanisation in China over the last two decades has resulted in a heavy metal burden to the environment (Zheng et al. 2011). The concentration of Cr in tea was reported in several recent studies (Karak & Bhagat 2010; Braganca et al. 2011). However, the data described in these studies focused only on total Cr and Cr(VI) (Wrobel et al. 2000; Seenivasan et al. 2008; Mandiwana et al. 2011). According to the contrasting effect of Cr(III) and Cr(VI), Cr in tea may have both beneficial and adverse effects on human health. Therefore, speciation of Cr as Cr(III) and Cr(VI) in tea and tea infusion is essential. On the basis of their degree of fermentation, Chinese made tea can be categorised into four different types (Wang et al. 2010): “non-fermented” green tea (produced by drying and steaming fresh leaves to inactivate the polyphenol oxidase and, thus, no oxidation occurs); “semi-fermented” oolong tea (produced when fresh leaves are subjected to a partial fermentation stage before drying); “full-fermented” black tea (which undergoes a fermentation stage by polyphenol oxidase before drying and steaming) and “post-fermented” Pu-erh tea (which undergoes a longer post-harvest fermentation stage by polyphenol oxidase and microorganisms). In this study, a survey
290
W.-h. Li et al.
was conducted on the speciation of Cr in Chinese made tea and tea infusion, to understand the benefit and risk of Cr intake from Chinese made tea.
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Materials and methods Reagents and standards The working standards of Cr were prepared by serial dilution of 1000 mg/L Cr(VI) and 1000 mg/L Cr(III) solutions (National Standard Center, China). The standard tea sample GBW07605 (National Standard Center, China) was applied as control sample (Cr content = 0.80 ± 0.03 mg/kg). Ultra-pure water (resistivity18.3 MΩ cm), obtained from a Milli-Q water purification system (Millipore, USA), was used to prepare all standards and samples. Supra-pure HNO3 and H2O2 (National Medicines, China) were used to digest tea samples in a microwave before Cr determination. Sample collection and preparation According to the fermentation degree, 128 tea samples (200 g each) were collected in China during 2011–2012, including 35 green tea, 32 oolong tea, 32 black tea and 29 Pu-erh tea. The samples were dried in an oven at 105°C for 2 hours and grounded into a 40 mesh powder with a grinder prior to use. To determine the Cr content consumed by tea infusion, an amount of 2.0 g of tea was added to a glass tea-maker and 200 mL of boiling deionised water was poured in to simulate the traditional way of tea making (Lung et al. 2003). After 5 min, tea infusion was filtered for analysis. Digestion Approximately 0.25 g tea sample was weighed into a Teflon coated tube and 3.0 mL of concentrated HNO3 and 2.0 mL concentrated H2O2 were added. The mixtures were allowed to predigest for 15 min at room temperature and then transferred to the microwave oven. Digestion was carried out at 150°C for 10 min at a maximum pressure of 800 psi. Samples were cooled to room temperature and diluted to 25 mL with ultra-pure de-ionised water. A blank sample was digested accordingly. Separation of Cr(III) and Cr(VI) Nowadays, the most prevalent method for determining Cr(VI) in soils, sediments and sludges is the EPA revised method 3060A (US Environmental Protection Agency 1995), which is based on an alkaline digestion procedure according to the findings of Mandiwana et al. (2011). In this study, a modified separated method based on EPA Method 3060A was employed for extraction of
Cr(III) and Cr(VI): 0.25 g tea sample (or 2.5 mL tea infusion) is digested using 25 mL of a solution of 0.28 mol/L Na2CO3 and 0.5 mol/L NaOH (pH about 12) and 100 mg MgCl2 and boiled for 5 min. Under these conditions, Cr(VI) is transferred into the solution due to the high alkalinity of the media while the remaining Cr(III) species are in a state of insoluble carbonate, hydroxide or oxide compounds. The amount of Cr(III) can be calculated by subtracting Cr(VI) from the total Cr. Analysis by atomic absorption spectrometry All measurements were carried out using a Hitachi Z5000 atomic absorption spectrometer (Hitachi, Japan) with Zeeman-effect background correction equipped with a chromium hollow cathode lamp operating at 9 mA. Measurements were performed at 359.3 nm with a spectral band pass of 1.3 nm. Argon with a purity of 99.99% (Praxair, USA) was used as purge and protective gas with a flow rate of 30 mL/min during all stages, except atomisation when the gas flow was stopped. Integrated absorbance (peak area) was used for signal evaluation and quantification. The limits of detection (LOD) and quantification (LOQ) were calculated from 15 measurements of analytical blanks. The values of LOD (calculated as 3 × SD) and LOQ (calculated as 10 × SD) were 0.1 μg/L and 0.3 μg/L, respectively. With respect to the tea samples, assuming that 0.25 g of tea was used for preparation of 25.0 mL of tea infusion, LOD and LOQ were 0.01 mg/kg and 0.03 mg/kg, respectively. Calibration curves with standards were used for all determinations. The graphite furnace temperature programme is given in Table 1. The accuracy of the measurements was assessed by applying the standard addition method in tea and tea infusion samples. Recoveries of spiked samples were 97.4% from tea and 95.3% from tea infusion. The method repeatability was evaluated by the relative standard deviation (RSD), which was 3.7% and 4.3% (n = 7) in tea and tea infusion, respectively. Statistical analysis Statistical analyses were performed using SPSS 18.0 software packages (SPSS Inc., Chicago, IL, USA). All determinations were carried out in triplicate. Means and Table 1.
Temperature programme for the determination of Cr. Temperature (°C)
Time (sec)
Stage
Start
End
Ramp
Drying Ashing Atomising Cleaning
80 700 2800 2800
140 700 2800 2800
40 20
Hold
5 4
Food Additives & Contaminants: Part B standard error of means are presented in the tables as an indication of variation. One-way ANOVA test was used to compare the means of different groups and p < 0.01 was considered to point out a significant difference between the compared groups.
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Results and discussion Cr in Chinese made tea The analytical results of tea samples are summarised in Table 2. Among the tea varieties, black tea contained most Cr, statistically and significantly (p < 0.01) different from other types of tea. The content of Cr(III) and Cr(VI) in black tea was accordingly higher than other types of tea. It suggested that the content of Cr(III) and Cr(VI) were positively correlated with the level of total chromium in China made tea. Cr transfer into tea infusion Cr(III), Cr(VI) and total Cr concentration in tea infusion were summarised in Table 3. The transfer ratio of Cr(III) and Cr(VI) from tea leaves into infusions was determined by comparing Cr content between teas and infusions. The results are shown in Figure 1. The Cr(III) transfer level in green tea was significantly lower (p < 0.01) than in other types. Contrary to green tea, the black tea, oolong tea and Pu-erh tea are all fermented under controlled conditions. Such a difference in leaf processing method causes different reactions in the leaf material, e.g. oxidation of phenolics such as catechols or tannins to give polyphenols (Lin et al. 1998; Wang et al. 2012). Consequently, Cr(III) can form soluble complexes with organic ligands during the
Table 2.
291
fermentation process, thus enhancing the transfer ratio in tea infusion. The transfer ratio of Cr(VI) was 37.8% in green tea, 37.4% in oolong tea, 38.7% in black tea and 38.2% in Pu-erh tea. There was no significant difference (p > 0.01) between the various types of tea. Cr(VI) transfer was significantly higher (p < 0.01), almost twofold to threefold, than that of Cr(III), indicating that solubility of Cr(VI) was higher than that of Cr(III).
Estimation of chromium intake from tea ATI (Adequate Daily Intake) is defined as the amount of tea infusions that provide adequate intake of total Cr every day and ADI (Acceptable Daily Intake) is the amount of tea infusions that can be consumed every day without detectable chronic effects. ATI and ADI can be estimated based on RNI (Recommended Nutrient Intake) and UL (Tolerable Upper Intake Level), respectively: ATIð1=dayÞ ¼ RNI=Cavg ðμg=lÞ
(1)
ADIð1=dayÞ ¼ UL=Cavg ðμg=lÞ
(2)
where RNI is 10 μg/day for children and 50 μg/day for adults, UL is 100 μg/day for children and 250 μg/day for adults; Cavg is the mean total chromium concentration of tea infusions. ATI and ADI were calculated for different tea infusions, both for adults and children. The results are given in Table 4 and show that the chance of adults and children being chronically and acutely intoxicated by drinking tea was low. On the other hand, the results suggested that
Cr(III), Cr(VI) and total Cr concentrations in tea samples (mg/kg).
Type Green tea Oolong tea Black tea Pu-erh tea
Cr(III)
Number of samples ± SD
Range
35 32 32 29
0.23–1.18 0.30–1.60 0.45–14.73 0.28–2.12
Cr(VI)
Mean ± SD 0.79 0.90 4.18 0.96
± ± ± ±
0.36 0.35 3.06 0.52
Range 0.03–0.12 0.03–0.25 0.18–2.87 0.04–0.31
Total Cr
Mean ± SD 0.08 0.16 1.16 0.21
± ± ± ±
0.02 0.07 0.56 0.04
Range 0.26–1.30 0.33–1.85 0.63–17.60 0.32–2.43
Mean ± SD 0.87 1.02 5.38 1.23
± ± ± ±
0.38 0.42 3.62 0.56
Table 3. Cr(III), Cr(VI) and total Cr concentrations in tea infusions (µg/L).
Type of tea Green tea Oolong tea Black tea Pu-erh tea
Number of samples 35 32 32 29
Cr(III) Range 0.2–1.2 0.5–2.5 0.8–22.0 0.4–3.4
Cr(VI)
Mean ± SD 0.8 1.4 7.1 1.6
± ± ± ±
0.2 0.4 3.8 0.8
Range 0.1–0.6 0.1–1.0 0.7–11.4 0.2–1.2
Total Cr
Mean ± SD 0.3 0.6 4.5 0.8
± ± ± ±
0.2 0.3 2.6 0.4
Range 0.3–1.8 0.6–3.5 1.5–33.4 0.6–4.6
Mean ± SD 1.2 2.0 11.3 2.3
± ± ± ±
0.4 0.7 6.4 1.2
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W.-h. Li et al.
Figure 1.
Transfer ratios of Cr(III) and Cr(VI) from tea leaves into infusions.
Table 4.
Estimated total Cr’s ATI and ADI for tea infusions. ATI (l/day)
Table 5.
ADI (l/day)
Type of tea
Adult
Children
Adult
Children
Green tea Oolong tea Black tea Pu-erh tea
42 25 4 22
8 5 1 4
208 125 22 109
83 50 9 43
Cr(III) and Cr(VI) intake per cup of tea (µg). Cr(III)
Type of tea
Number of samples
Range
Green tea Oolong tea Black tea Pu-erh tea
35 32 32 29
0.04–0.24 0.10–0.50 0.16–4.40 0.08–0.68
drinking tea is an effective way for Cr intake. Tea drinkers who like more concentrated tea would prepare the tea for a longer period of time (>5 min) or use more tea leaves. In those cases, more Cr would be infused in water due to longer preparation time or use of more tea leaves, thus increasing more Cr consumption than reported here. Furthermore, this work was conducted with deionised water. Thus, intake concentrations of Cr in actual tea infusion with tap water could be higher than the current measurements. Literature data for Cr mainly focuses on total Cr rather than Cr speciation. Until now, there is no defined
Cr(VI) Mean ± SD 0.16 0.28 1.42 0.32
± ± ± ±
0.04 0.08 0.76 0.16
Range 0.02–0.12 0.02–0.2 0.14–2.28 0.04–0.24
Mean ± SD 0.06 0.12 0.90 0.16
± ± ± ±
0.04 0.06 0.52 0.08
maximum acceptable concentration of Cr(III) and Cr(VI). As indicated earlier, Cr(III) and Cr(VI) have contradicting physiological effects. In order to discuss Cr(III) and Cr (VI) intake from daily tea consumption, this study gives average Cr(III) and Cr(VI) content per cup (200 µg/mL tea infusion), presented in Table 5, which can be evaluated to draft limits for chromium in tea. Conclusion Cr level is significantly higher in black tea than in other types. From the database which was generated in this
Food Additives & Contaminants: Part B study to estimate the Cr intake from tea, it could be concluded that drinking tea is a reliable and effective way for Cr intake, the risk of adults and children being chronically intoxicated by tea infusions being low. Acknowledgement This work was supported by the National Natural Scientific foundation of China (Grant 31140037).
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