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The analysis and identification of charred suspected tea remains unearthed from Warring State Period Tomb

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Recently, a bowl containing charred suspected tea remains unearthed from the early stage of Warring States period tomb in Zoucheng City, Shandong Province, China. To identify the remains is significant for understanding the origin of tea and tea drinking culture. Scientific investigations of the remains were carried out by using calcium phytoliths analysis, Fourier transform infrared spectroscopy (FTIR), Gas Chromatograph Mass Spectrometer (GC/MS) and Thermally assisted hydrolysis—methylation Pyrolysis Gas Chromatography Mass Spectrometry (THM-Py-GC/MS) techniques. Modern tea and modern tea residue were used as reference samples. Through phytoliths analyses, calcium phytoliths identifiable from tea were determined in the archeological remains. The infrared spectra of the archaeological remains was found similar as modern tea residue reference sample. In addition, the biomarker compound of tea—caffeine was determined in the archaeological remains by THM-Py-GC/MS analysis. Furthermore, through GC/MS analysis, some compounds were found both in the archeological remains and the modern tea residue reference samples. Putting the information together, it can be concluded that the archaeological remains in the bowl are tea residue after boiling or brewing by the ancient.
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
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The analysis and identication
of charred suspected tea remains
unearthed from Warring State
Period Tomb
Jianrong Jiang1, Guoquan Lu2*, Qing Wang2 & Shuya Wei1*
Recently, a bowl containing charred suspected tea remains unearthed from the early stage of Warring
States period tomb in Zoucheng City, Shandong Province, China. To identify the remains is signicant
for understanding the origin of tea and tea drinking culture. Scientic investigations of the remains
were carried out by using calcium phytoliths analysis, Fourier transform infrared spectroscopy (FTIR),
Gas Chromatograph Mass Spectrometer (GC/MS) and Thermally assisted hydrolysis—methylation
Pyrolysis Gas Chromatography Mass Spectrometry (THM-Py-GC/MS) techniques. Modern tea and
modern tea residue were used as reference samples. Through phytoliths analyses, calcium phytoliths
identiable from tea were determined in the archeological remains. The infrared spectra of the
archaeological remains was found similar as modern tea residue reference sample. In addition,
the biomarker compound of tea—caeine was determined in the archaeological remains by THM-
Py-GC/MS analysis. Furthermore, through GC/MS analysis, some compounds were found both in
the archeological remains and the modern tea residue reference samples. Putting the information
together, it can be concluded that the archaeological remains in the bowl are tea residue after boiling
or brewing by the ancient.
China is the rst country in the world to discover and cultivate tea. In Chinese legend, tea was rst discovered
as an antidote by Emperor Shen Nung in 2737 , according to the rst monograph on Chinese herbal medicine
Shennong’s Classic of Materia Medica (神农本草经)1. e rst mention of tea planting is believed to occur in
the Xiaxiaozheng (夏小正), a Chinese earliest almanac recording traditional agricultural aairs, probably writ-
ten in the Warring States Period (475–221 ). According to the literature, in the Spring and Autumn Period
(770–476 ), tea had been used as a sacrice and vegetable, in the Warring States period and the early Western
Han Dynasty, tea cultivation, tea making techniques and tea drinking custom in Sichuan province began to
spread to other places2.
e physical evidence of tea is very important to conrm the origin, development, function and culture of tea.
As archaeological plant leaves remains have been buried for many years, most of them have rotted or charred,
it is dicult to nd archaeological plant leaves remains in archeological excavation. e rst tea remains were
found in Northern Song tomb of Lu’an, Anhui Province3. e oldest physical evidence of tea remains are from
other two funerary sites: the Han Yangling Mausoleum in Xi’an, Sha’anxi Province, and the Gurgyam Cemetery
in Ngari district, western Tibet, revealing that tea was used by Han Dynasty emperors as early as 2100year BP
and had been introduced into the Tibetan Plateau by 1800year BP4, but whether the tea was used as beverage,
food, medicine is unclear. Recently, some charred suspected tea remains (CST) were found in a bowl unearthed
from tomb No. 1 at Xigang in the Ancient Capital City Site of the Zhu Kingdom in Zoucheng City (e early
stage of Warring States, approximately 2400years ago), Shandong Province (Fig.1)5. If the remains could be
determined as tea, that would be the direct evidence for tea drinking in the ancient time.
Previously, the researchers commonly used for the identication of plant remains were mainly based on
the morphology of the plant, however, most of archaeological plant remains have rotted or charred due to the
interference of various microorganisms, oxidation and other factors in the buried environment for many years,
the morphology of the plants also changed dramatically, therefore, to identify the plant species by morphology
is not applicable to the sample CST. Recently calcium phytoliths (calcium oxalate plant crystals), biomarkers
OPEN
Institute of Cultural Heritage and History of Science & Technology, University of Science and Technology Beijing,
             *email:

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(caeine and theanine) were identied in archaeological tea remains by using Gas Chromatography/Mass
spectrometry (GC/MS) and Ultra-performance Lquid Chromatography and Mass Spectrometry (UPLC–MS)
techniques4. For modern tea study, Fourier transform infrared spectroscopy (FTIR)69, Spectrophotometry, er-
mospray–LC–MS10, Head space solids-phase microextraction in combination with gas chromatography–mass
spectrometry (HS-SPME/GC–MS)11, Gas chromatography–mass spectrometry (GC–MS)12,13 are the mainly
techniques applied.
In this study, methods of calcium phytoliths analysis, Fourier transform infrared spectroscopy (FTIR), Gas
Chromatography/Mass spectrometry (GC/MS) and thermally assisted hydrolysis–methylation Pyrolysis Gas
Chromatography/Mass Spectrometry (THM–Py–GC/MS) were chosen for the identication of the sample CST
found in the Warring State tomb. In the meantime, modern reference samples were studied by using the same
analytical methods as the archaeological sample for comparison.
Materials and methods
Archaeological sample: the Archaeological sample CST take from the residues which poured out from the bowl
unearthed from the Tomb No.1 at Xigang (Fig.2).
Reference samples: since the sample CST was unearthed in a bowl, therefore, it is not excluded that the sample
is tea residue le aer boiling or brewing by the ancient, so in this study, modern tea and modern tea residue
were used as reference samples. Modern tea residue was prepared from brewing tea with water for several times,
then dry thoroughly and nally grinded into powders. e modern tea samples were bought from tea shop in
Beijing, which are black tea produced from Shangdong Laoshan.
Analysis of calcium phytoliths. Calcium phytoliths experiment was performed according to the proce-
dure described in the literature4. Identication of calcium phytoliths was performed under a LEICA DM2700P
microscope.
Fourier transform infrared spectroscopy (FTIR). For FTIR analysis, Nicolet 6700 Advanced Fourier
transform infrared spectrometer (America ermo Fisher Scientic) was used. e spectra were collected over
Figure1. e map shows (a) Location of Shandong Province in China; (b) e Ancient Capital City Site of the
Zhu Kingdom in Zoucheng City; (c) e plan of the tomb; (d) e plan and prole of tomb No. 1 at Xigang.
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the 4000–500 cm−1 region, using attenuated total reectance (ATR) for the measurements, the spectral resolu-
tion is 4 cm−1 and the number of scans is 64. Each sample was scanned at 25 and the data acquisition system
used was OMNIC.
Gas chromatograph–mass spectrometer (GC/MS). GC–MS analysis was performed using Agilent
GC–MS-QP2010Ultra (Shimadzu, Japan). A capillary column Ultra-5MS (5% diphenyl/95% dimethyl silox-
ane), 0.25mm internal diameter, 0.25m lm thickness and 30m length [Frontier lab, Japan] was used for the
separation. Temperature programmed: initially keeping the column at 120°C for 2min, followed by a gradient
of 5°C/min to 270°C and hold for 10min. e injector temperature was set to 240°C. 30:1 split ratio. e car-
rier gas used was Helium (purity 99.999%). e electronic pressure control was set to a constant ow of 1ml/
min; Electron ionization (EI) temperature was at 280°C; Transmission lines temperature was at 220°C; Range
of Scanning: 35 ~ 510m/z.
Analysis procedure: A sample (20mg) was weighed, transferred into a sampling vial with ultrapure water,
boiled the sample for 10min in a water bath, extracted under sonication at 60°C for 30min and then centrifuged
for 10min. Aerwards, the supernatant was transferred to a sample vial, then it was evaporated and dried in a
stream of N2 at 60°C. Finally, the dried tea extract was dissolved in solvent acetonitrile (ACN) and derivatized
with N-(tert-Butyldimethylsilyl)-N-methyltriuoroacetamide (MTBSTFA, 1: 1 to ACN, v/v) at 110°C for 30min,
transfer the mixed uid into an auto sampling vial for GC/MS analysis.
Pyrolysis gas chromatography mass spectrometry (Py-GC/MS). For Py-GC/MS analysis, a Multi-
Shot pyrolyzer, type EGA/PY-3030D, made by Frontier Lab, Japan, and a gas chromatograph mass spectrometer,
GC–MS-QP2010 Ultra (Shimadzu, Japan). Shimadzu GC–MS real time analysis soware was used for GC–MS
control, peak integration and mass spectra evaluation.
e pyrolysis was performed at 550°C for 12s. e pyrolyser interface was set to 290°C and the injector was
set to 250°C. A capillary column SLB-5MS (5% diphenyl /95% dimethyl siloxane), 0.25mm internal diameter,
0.25m lm thickness and 30m length [Supelco] was used in order to provide an adequate separation of the
components. e chromatographic conditions were as follows: the oven initial temperature was set to 35°C for
5min, followed by a gradient of 60°C/min to 100°C, for 3min, 14°C/min to 240°C , then 6°C/min to 315°C
and hold for 1.5min, the carrier gas was Helium (He, purity 99.999%). e electronic pressure control was set to
a constant ow of 0.92ml/min, in split mode at 1:20 ratios. Ions were generated by electron ionization (145.3eV)
in the ionization chamber of the mass spectrometer. e mass spectrometer was set from m/z 35 to 750. EI mass
spectra were acquired by total ion monitoring mode. e temperatures of the interface and the source were
280°C and 200°C, respectively.
NIST14 and NIST14s Library of Mass Spectra were used for identifying the compounds.
Analysis procedure. About 50g sample was placed in a sample cup, 3 L of 25% aqueous TMAH (analytical
pure, Sinopharm Chemical Reagent Co., Ltd) solution were injected into the sample cup, the cup was placed
on top of the pyrolyzer at ambient temperature and then pyrolyzed immediately, aerwards the temperature
program for the GC/MS analysis was started. During the process of analysis, blank tests were conducted before
each sample.
Figure2. e map shows (a) Tomb No. 1 at Xigang; (b) Burial objects in the ware box; (c) e unearthed bowl;
(d) e residues which poured out from the bowl; (e) e sample CST take from the residues.
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Statement. e use of plants parts in the present study complies with National standards of the People’s
Republic of China on black tea (GB/T13738).
Results and discussion
Analysis of calcium phytoliths. Calcium phytolith analyses were carried out according to the procedure
described in the literature4. e morphology observation of sample CST under microscope is depicted in Fig.3,
which reveals that the sample contains abundant calcium phytoliths, including the crack, druses and trichome
base, these calcium phytoliths also match the genus Camellia, especially druse and trichome base are the most
distinctive crystals in tea plants4.
FTIR analysis. e infrared spectra of modern tea, modern tea residue and the sample CST are shown in
Fig.4. e vibrations of the functional groups of the compounds in tea and their corresponding infrared absorp-
tion characteristic peaks are consistent with the literatures2,68. Taking infrared spectrum of modern tea as an
example, the band assignment to chemical bonds for the vibrational FTIR spectra of modern tea is summarized
in Table1.
e peak shapes and the positions of main absorption peaks (1632, 1041 cm−1) of the archaeological sample
CST are consistent with the modern tea reference samples (Fig.4), so it is speculated that the sample CST is
most likely ancient tea.
e intensity of some absorption peaks in modern tea residue decreased signicantly in comparison with
modern tea, such as the peaks at 1516, 1454, 1237 cm−1, indicating some compounds in the tea may be dissolved
in water. Sample CST has been buried for hundreds of years, microorganisms and other factors in the burial
environment may cause chemical changes, resulting in the subtle dierences in infrared spectra between the
archaeological sample CST and reference samples. To conrm whether the sample CST is ancient tea or not,
further study by other techniques were carried out as following.
Biomarker analysis. THM‑Py‑GC/MS analysis. e chromatograms of modern tea, modern tea residue
and the sample CST obtained by THM-Py-GC/MS are shown in Fig.5. ree parallel analyses were conducted
for each sample, the analysis results are consistent with each other.
Figure3. Photographs under microscope of calcium phytoliths from sample CST. (a) crack-type calcium
phytoliths; (b) druse-type calcium phytoliths; (c) trichome-type calcium phytoliths.
Figure4. Infrared spectra of A—modern tea; B—modern tea residue; C—sample CST.
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Comparing retention time and mass spectrum of the main chromatographic peaks in the sample CST and
the reference samples (modern tea and modern tea residue), the main peaks found in the sample CST are also
present in the reference samples (peak No. 1–8), which are 1,3,5-trimethoxy-benzene, 2,4,6-trimethoxytoluene,
3,4-dimethoxy-benzoic acid methyl ester, 3,4,5-trimethoxy-benzoic acid methyl ester, caeine, hexadecanoic
acid methyl ester, 9-octadecenoic acid methyl ester, methyl stearate, the peak 1, 2, 3, 4 belong to methoxyben-
zene compounds which are the characteristic components of tea aroma14,15, and hexadecanoic acid methyl ester,
9-octadecenoic acid and methyl stearate are common fatty acids in tea16. Especially the biomarker compound
of tea—caeine was identied (peak No. 5, RT 10.5min) in CST sample, the mass spectra are shown in Fig.6.
Caeine is easily soluble in water, most of the caeine in tea was leached out in the process of brewing tea, there-
fore, the content of caeine in modern tea residue is signicantly lower than that in modern tea (Fig.5). Peak 3
is one of the most intense for CST but not signicant in the modern sample, probaly due to two reasons: rstly,
the relative contents of the chemical components in dierent tea are dierent; secondly, it is probably due to the
eect of the burial environment. In order to see the inuence of the burial environment, soil samples from the
area where the bowl was found were analyzed by Py-GC/MS, small amount of fatty acids were found, which will
not aect the conclusion for sample CST.
GC/MS analysis. e modern tea reference sample, modern tea residue reference sample and the sample CST
were pretreated according to the procedure described in a previous section in this document. e chromato-
grams of them obtained by GC/MS analyses are shown in Fig.7. ree parallel analyses were conducted for each
sample, the analysis results are consistent with each other.
e main amino acids contained in tea were detected in modern tea sample aer derivatized by MTBSTFA,
which is consistent with the literature17,18. Especially the tea marker compound-theanine, two derivatized peaks
of theanine were detected in modern tea reference sample (labeled as T1 and T2 in Fig.7). In the modern tea
Table 1. Band assignments for the FTIR spectra obtained from modern tea.
Wavenumber (cm−1) Vibrational mode assignment Absorption peak intensity: s (strong)/m
(medium)/w (weak)
3420 –OH stretching vibration of tea-polyphenols and tea-
polysaccharides s
2923, 2852 Saturated C–H stretching vibration m, w
1646 C=C stretching vibration peak of sugars and avonoids s
1516 –NO2 stretching vibration peak of aromatic compounds
in tea w
1454 saturated C–H deformation vibration w
1362 –NO2 stretching vibration peaks of aliphatic compounds w
1237 C–O stretchingvibrationpeak in amides w
1146 C–O–C antisymmetric stretching vibration w
1035 O–H in-plane deformation vibration s
Figure5. TIC chromatogram obtained by THM-Py-GC/MS of A—modern tea; B—modern tea
residue; C—sample CST; e peak numbers are corresponding to: (1): 1,3,5-trimethoxy-benzene; (2):
2,4,6-trimethoxytoluene; (3): 3,4-dimethoxy-benzoic acid, methyl ester; (4): 3,4,5-trimethoxy-benzoic acid,
methyl ester; (5): caeine; (6): hexadecanoic acid, methyl ester; (7): 9-octadecenoic acid, methyl ester; (8):
methyl stearate.
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residue sample, only a trace of theanine (T1) was found, but not found in the CST sample. eanine is the main
free amino acid in tea, its concentration is signicantly decreased aer tea was brewed due to its good solubility
in water, therefore, the content of theanine in modern tea residue and sample CST is so low that even cannot be
detected. However, there are some other compounds were detected both in modern tea residue and sample CST,
which are listed in Table2, most of these components are organic acids, which are the common components in
tea, and as a water-soluble substance, which can be also leached out in the process of brewing tea19. Although
some of the compounds were not identied what they were, but they were both present in modern tea residue
and sample CST (peak No. 1, 4, 10, 12, 13 in Fig.7), indicating those compounds origin from tea, which provide
additional evidence that the archaeological sample CST is most likely tea residue le aer boiling or brewing.
e relative intense of peak 4 is higher in CST sample in comparison with the reference samples, probably due
to ageing.
Conclusions
In this study, Calcium phytoliths analysis, Fourier transform infrared spectroscopy (FTIR), Gas Chromato-
graph Mass Spectrometer (GC/MS) and ermally assisted hydrolysis-methylation pyrolysis-gas chromatog-
raphy/mass spectrometry (THM–Py-GC/MS) techniques were applied for the identication of archaeological
remains–charred suspected tea (CST) excavated from the early stage of Warring State Period tomb in Shandong
Province. e experimental results show that the sample CST contains abundant calcium phytoliths identi-
able as tea, e FTIR spectra of CST sample are similar with that of the modern tea residue. Moreover, caf-
feine, methoxybenzene compounds, organic acids, 2-Amino[1,3]thiazolo[4,5-d]pyrimidine-5,7-diol and several
Figure6. e mass spectrum of caeine (peak 5 in Fig.5) of A—modern tea; B—modern tea residue; C—
sample CST by THM Py-GC/MS analysis.
Figure7. TIC chromatogram obtained by GC/MS of A—modern tea; B—modern tea residue; C—sample CST;
T1, T2: two derivatized peaks of theanine in modern tea sample (A); e peak numbers are corresponding to
the numbers in Table2.
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unidentied compounds were detected in both the sample CST and the reference sample (modern tea residue) by
THM–PY-GC/MS and GC/MS. By putting the information together, it can be concluded that the archaeological
remains in the bowl are tea residue aer boiling or brewing by the ancient.
Tea drinking is one of the most representative traditional cultures in China, since ancient times, the Chinese
people have always had the habit of drinking tea, but there is no physical evidence to prove when tea actually
appeared, until the discovery of tea in the Han Yangling Mausoleum, which proved that Chinese tea has a history
of at least 2150years, which has earned recognition from Guinness World Records as the oldest tea in 20164. e
identication of the tea remains at the Ancient Capital Site of the Zhu Kingdom in Zoucheng (the early stage of
Warring States, approximately 2400years ago) has advanced the origin of tea by nearly 300years. Furthermore,
the tea was found in a small bowl, providing additional evidence of the usage of tea. e results of this study
indicate that tea drinking culture may start as early as in Warring State period.
Received: 1 April 2021; Accepted: 20 July 2021
References
1. Gu, G. G. Shennong’s Classic of Materia Medica Vol. 79 (Academy Press, 2007).
2. Li, X. S. A brief history of Chinese tea. J. Anhui. Agric. Sci. 12, 7559–7560 (2011).
3. Fan, W. Q., Gong, D. C., Yao, Z. Q. & Li, D. W. Identication analysis of carbonized suspected tea from Lu’an tomb in the northern
song dynasty. Agric. Archaeol. 2, 212–217 (2012).
4. Lu, H. Y. et al. Earliest tea as evidence for one branch of the Silk Road across the Tibetan Plateau. Sci. Rep. 6, 18955 (2016).
5. Wang, Q., Chen, R. X., Liu, C., Chen, Z. L. & Lang, J. F. e 2017 excavation of the ancient capital city site of the Zhu Kingdom in
Zoucheng, Shandong Province. Southeast Cult. 3, 37–56 (2019).
6. Yang, Q. & Wang, Y. L. Detecting dierent teas by Fourier transform infrared spectroscopy. Las. Optoelect. Prog. 4, 117–120 (2010).
7. Fan, S. Z., Han, Y. D., Hou, D. Y., Shan, L. & He, C. B. Review on application of infrared spectrometry in analysis of tealeaf. Chem.
Res. 23, 107–110 (2012).
8. Cai, J. X., Wang, Y. F., Xi, X. G., Li, H. & Wei, X. L. Using FTIR spectra and pattern recognition for discrimination of tea varieties.
Int. J. Biol. Macromol. 78, 439–446 (2015).
9. Li, X. W., Wang, T. T., Zhang, M. Z., Zhang, R. X. & Li, G. Eect of dierent storage years on the quality of unzymic and zymic
Pu-er tea by FTIR combined with feature extraction. Int. Conf. Multimed. Technol. 1, 2450–2453 (2011).
1 0. Kiehne, A. & Engelhardt, U. H. ermospray-LC-MS analysis of various groups of polyphenols in tea. Z. Lebensmittel‑Untersuchung
Forschung. 202, 48–54 (1996).
11. Chen, L. et al. Aroma proling of oolong tea by SDE and HS-SPME in combination with GC-MS. J. Tea. Sci. 6, 692–704 (2019).
12. Xu, C. H. & Wang, Y. X. Discrimination of three famous tea in Jiangxi based on GC-MS combined with chemometrics. Food Sci.
20, 1–17 (2020).
13. Wang, M. Q. et al. Characterization of the key aroma compounds in Longjing tea using stir bar sorptive extraction (SBSE) combined
with gas chromatography-mass spectrometry (GC-MS), gas chromatography- olfactometry (GC-O), odor activity value (OAV),
and aroma recombination. Food Res. Int. 130, 1–11 (2020).
14. Guan, X. X., Hu, W. Z., Wang, Y., Li, C. B. & Jiang, A. L. Advances in research on tea aroma chemistry. in Proc. of 15th Conf. organ‑
ized by Chinese Inst. Food Sci. and Technol., 471–472 (Shandong Province, Qingdao city, 2018).
15. Cao, Y. N. & Liu, T. X. Analysis of lerpenes and methoxybenzene components in aroma composition of pu-erh raw tea and ripe
tea. Sci. Technol. Food. Ind. 33, 128–129 (2012).
16. Guo, L. et al. Analysis of fatty acid compositions and contents in oolong tea from Fujian Province. J. Tea Sci. 39, 611–618 (2019).
17. Zhang, J., Wang, C. P. & Ruan, J. Y. Determination of main free amino acids in tea by gas chromatography-mass spectrometry
(GC-MS) and gas chromatography-ame ionization detector (GC-FID). J. Tea Sci. 30, 445–452 (2010).
18. Wang, L. et al. Analysis of free amino acids in Chinese teas and ower of tea plant by high performance liquid chromatography
combined with solid-phase extraction. Food. Chem. 4, 1259–1266 (2010).
Table 2. GC/MS analysis result of modern tea residue and sample CST.
Peak no. RT (min) Main ions (m/z) Compounds identied
1 18.04 115, 147, 173 Unidentied
259, 386
2 19.04 147, 189, 221 Boric acid, 3TMS derivative
263, 355
3 20.54 211, 269, 383, 425 Phosphoric acid, tris(tert-butyldimethylsilyl) ester
4 22.56 147, 221, 263, 337 Unidentied
5 27.53 117, 131, 313 Palmitic acid, TBDMS derivative
6 30.54 129, 337 Linoelaidic acid, tert.-butyldimerthylsilyl ester
7 30.59 129, 339, 381 Petroselinic acid, TBDMS derivative
8 30.68 129, 339 .alpha.-Linolenic acid, TBDMS derivative
9 31 117, 129, 341 Stearic acid, TBDMS derivative
10 32.92 117, 143, 237 Unidentied
252, 359
11 33.54 223, 339, 455, 469 2-Amino[1,3]thiazolo[4,5-d]pyrimidine-5,7-diol
12 34.15 185, 241, 256, 359 Unidentied
13 37.14 238, 323, 397 Unidentied
439, 495
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19. Liu, P. et al. Study on organic acids contents in tea leaves and its extracting characteristics. J. Tea Sci. 33, 405–410 (2013).
Acknowledgements
e authors gratefully acknowledge Professor Yan Wu (Key Laboratory of Vertebrate Evolution and Human
Origins of Chinese Academy of Sciences, Institute of Vertebrate Paleontology and Paleoanthropology, Chinese
Academy of Sciences, Beijing) and Dr Xiumin Xia (National Museum of China) for the help of calcium phyto-
liths analysis.
Author contributions
J.J. performed all experiments, analyzed the data and produced gures. J.J. and S.W. draed the manuscript. G.L.
and Q.W. provided the archaeological data, materials and reviewed the manuscript.
Funding
Funding was provided by Scientic research and protection of ancient organic cultural relics (Grant No.
06102070) and the National Key Research and Development Program of China. No. 2020YFC1522402.
Competing interests
e authors declare no competing interests.
Additional information
Correspondence and requests for materials should be addressed to G.L.orS.W.
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