3.The Assembly of a Novel Enzyme Biosensor for Aflatoxin B_1 Detection
Da-Ling LIU ; Yi SHEN ; Jing ZHANG ; Dong-Sheng YAO ;
China Biotechnology 2006;0(03):-
A novel biosensor for aflatoxin B1 detecting has been reported. The biosensor electrode for AFB1 detecting was assembled by immobilized aflatoxin-oxidoreductase using open-ended multi-walled carbon nanotubes as matrix. Its linear range was between 0.16?M and 3.2?M. And if the specific anti-aflatoxin B1 antibody and aflatoxin oxidoreductase were both immobilized on the electrode with Multi-Walled carbon nanotubes, the detection limit of the modified electrode could be 16 nM with a 10 times improved sensitivity. The aflatoxin enzyme biosensor assembled this way strode one step forward its practical application.
4.Troubleshooting of bioinequivalence of compound valsartan tablets.
Da SHAO ; Yi-Fan ZHANG ; Yan ZHAN ; Xiao-Yan CHEN ; Da-Fang ZHONG
Acta Pharmaceutica Sinica 2014;49(4):524-529
The study aims to evaluate the bioequivalence of valsartan hydrochlorothiazide tablets, and to investigate the potential cause of bioinequivalence. This was a single-center study with an open, randomized double-way crossover design. Test and reference preparations containing 160 mg of valsartan and 25 mg of hydrochlorothiazide were given to 36 healthy male volunteers. Plasma concentrations of valsartan and hydrochlorothiazide were determined simultaneously by LC-MS/MS. The pharmacokinetic parameters and relative bioavailability were calculated, while the bioequivalence between test and reference preparations were evaluated. The dissolution profiles of test and reference preparations in four different mediums were determined via dissolution test and HPLC. The similarity was investigated according to the similarity factors (f2). The F(o-t) and F(0-infinity) were (139.4 +/- 65.2)% and (137.5 +/- 61.2)% for valsartan of test preparations. It led to get the conclusion that test and reference preparations were not bioequivalent for valsartan. A significant difference was observed between test and reference tablets in the valsartan dissolution test of pH 1.2 hydrochloric acid solution. The key factor of the bioinequivalence might be that dissolution of valsartan in acid medium has marked difference between two preparations.
Administration, Oral
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Adolescent
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Adult
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Angiotensin II Type 1 Receptor Blockers
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administration & dosage
;
adverse effects
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blood
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pharmacokinetics
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Antihypertensive Agents
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administration & dosage
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adverse effects
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blood
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pharmacokinetics
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Area Under Curve
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Chromatography, Liquid
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Cross-Over Studies
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Drug Liberation
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Humans
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Hydrochlorothiazide
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administration & dosage
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adverse effects
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blood
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pharmacokinetics
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Male
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Tablets
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Tandem Mass Spectrometry
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Therapeutic Equivalency
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Valsartan
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administration & dosage
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adverse effects
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blood
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pharmacokinetics
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Young Adult
5.Cloning and expression analysis of a zinc-regulated transporters (ZRT), iron-regulated transporter (IRT)-like protein encoding gene in Dendrobium officinale.
Gang ZHANG ; Yi-Min LI ; Biao LI ; Da-Wei ZHANG ; Shun-Xing GUO
China Journal of Chinese Materia Medica 2015;40(1):42-47
The zinc-regulated transporters (ZRT), iron-regulated transporter (IRT)-like protein (ZIP) plays an important role in the growth and development of plant. In this study, a full length cDNA of ZIP encoding gene, designed as DoZIP1 (GenBank accession KJ946203), was identified from Dendrobium officinale using RT-PCR and RACE. Bioinformatics analysis showed that DoZIP1 consisted of a 1,056 bp open reading frame (ORF) encoded a 351-aa protein with a molecular weight of 37.57 kDa and an isoelectric point (pI) of 6.09. The deduced DoZIP1 protein contained the conserved ZIP domain, and its secondary structure was composed of 50.71% alpha helix, 11.11% extended strand, 36.18% random coil, and beta turn 1.99%. DoZIP1 protein exhibited a signal peptide and eight transmembrane domains, presumably locating in cell membrane. The amino acid sequence had high homology with ZIP proteins from Arabidopsis, alfalfa and rice. A phylogenetic tree analysis demonstrated that DoZIP1 was closely related to AtZIP10 and OsZIP3, and they were clustered into one clade. Real time quantitative PCR analysis demonstrated that the transcription level of DoZIP1 in D. officinale roots was the highest (4.19 fold higher than that of stems), followed by that of leaves (1.12 fold). Molecular characters of DoZIP1 will be useful for further functional determination of the gene involving in the growth and development of D. officinale.
Amino Acid Sequence
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Cloning, Molecular
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Dendrobium
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chemistry
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classification
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genetics
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metabolism
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Gene Expression Regulation, Plant
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Iron
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metabolism
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Membrane Transport Proteins
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chemistry
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genetics
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metabolism
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Molecular Sequence Data
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Phylogeny
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Plant Proteins
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chemistry
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genetics
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metabolism
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Plants
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chemistry
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classification
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genetics
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Sequence Alignment
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Zinc
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metabolism
6.Isolation and differential expression of a novel MAP kinase gene DoMPK4 in Dendrobium officinale.
Gang ZHANG ; Yi-Min LI ; Ben-Xiang HU ; Da-Wei ZHANG ; Shun-Xing GUO
Acta Pharmaceutica Sinica 2014;49(7):1076-1083
Mitogen-activated protein kinases (MAPKs) are important signaling transduction components well conserved in eukaryotes and play essential roles in various physiological, developmental and hormonal responses in plant. In the present study, a MAPK gene, designated as DoMPK4 (GenBank accession No. JX297597), is identified from a rare endangered medicinal orchid species D. officinale using the reverse transcription polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE) methods. The full length cDNA of DoMPK4 is 1 518 bp in length and encoded a 369 aa protein with a molecular weight of 42.42 kD and an isoelectric point of 5.55. DoMPK4 protein contained a serine/threonine protein kinase active site (158-170), a MAP kinase site (71-174), and eight conserved motifs. DoMPK4 had a transmembrane (214-232) but no signal peptide. Multiple sequence alignment showed that DoMPK4 shared high identities (74.9%-80.6%) with MAPK proteins from various plants. Phylogenetic analysis demonstrated that DoMPK4 belonged to group A of the MAPK evolutionary tree, and is closely related to monocots. Real time quantitative PCR (qPCR) analysis revealed that DoMPK4 is differentially expressed among the five organs including leaf, stem, root, seed, and protocorm-like body (PLB). The transcription level of DoMPK4 is the highest in the PLBs with 17.65 fold, followed by seeds, roots, and stems with 5.84, 2.28, and 1.64 fold, respectively. The progressive enhancement of DoMPK4 transcripts in the developing PLBs compared to that in the germinating seeds, suggests a role of DoMPK4 during the development of embryogenic PLBs formation in D. officinale.
Amino Acid Sequence
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DNA, Complementary
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genetics
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DNA, Plant
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genetics
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Dendrobium
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enzymology
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genetics
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Gene Expression Regulation, Plant
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Mitogen-Activated Protein Kinases
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genetics
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metabolism
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Phylogeny
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Plant Leaves
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metabolism
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Plant Proteins
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genetics
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metabolism
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Plant Roots
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metabolism
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Plant Stems
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metabolism
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Seeds
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metabolism
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Sequence Alignment
7.Usefulness of Ankle Brachial Index to Predict the Extent of Coronary Stenosis
Shi-wei YANG ; Da-yi HU ; Xian WANG ; Shouyan ZHANG ; Jian ZHANG ; Shi GUO
Chinese Journal of Rehabilitation Theory and Practice 2006;12(6):461-463
ObjectiveTo examine the relationship between ankle brachial index (ABI) and the extent of coronary stenosis and evaluate the usefulness of ABI to predict the extent of coronary stenosis in old patients.Methods118 patients with coronary angiography were examined by ABI and hemostatic factors evaluation in addition to history collection.ResultsABI was inversely and significantly associated with Gensini score. ABI reduced significantly (P<0.001) in the patients with 3-vessel or left main coronary artery disease (CAD). But there were no significant differences in ABI among the patients with no CAD, 1-vessel or 2-vessel CAD. The corresponding area under the ROC curve was 0.75±0.045, with 95% CI=0.67~0.84 (P<0.001) in ABI in 3-vessel or left main CAD. When ABI≤0.9, it had a relatively high specificity (89.1%) and sensitivity (55.6%) for predicting the presence of 3-vessel disease or left main CAD.ConclusionIn the old patients, ABI is inversely and significantly associated with the extent of coronary stenosis, and ABI≤0.9 has a relatively high specificity and sensitivity for predicting the presence of 3-vessel or left main CAD.
8.Experimental study of platelet-rich plasma optimizing mesenchymal stem cells culture.
Chinese Journal of Hematology 2006;27(6):403-405
OBJECTIVETo explore a new method for in vitro expansion of human mesenchymal stem cells (MSC) by using platelet-rich plasma (PRP) in place of fetal calf serum (FCS).
METHODSBone marrow(BM) samples were obtained from the proximal femurs of patients with normal haematopoietic function undergoing total hip arthroplasty. 2 x 10(5)/cm2 BM nucleated cells were seeded in 25 cm2 flasks for MSC cultivation containing one of the 3 mediums: complete Dexter medium with 12.5% FBS and 12.5% horse serum (medium1), alpha-MEM with 10% FCS (medium2) and alpha-MEM with 5% PRP(medium3). At the same time, 1 x 10(6) nucleated BM cells and same amount of nucleated cells from iliac aspirate were seeded in 25 cm2 tissue flasks, colony forming unit-fibroblast (CFU-F) assay.
RESULTSCulture-expanded cells from proximal femurs presented a typical fibroblast-like morphology. Cells were positive for SH2 (CD 105), SH3 (CD73), Thy-1 (CD90), while negative for CD34 and CD45. On induction, these cells could differentiate into osteoblasts. A significantly higher proliferative capacity of MSCs expanded in medium3 was observed in comparison to those in mediuml or 2 without alteration of the phenotype and the differentiation property. CFU-F assays indicated that bone marrow from the proximal femoral contained significantly more CFU-F than that from iliac aspirate.
CONCLUSIONPlatelet-rich plasma can be used in place of FCS to provide a safer and more effective culture condition to expand MSCs for clinical purpose. The proximal femur BM cells can be obtained in hip surgeries.
Blood Platelets ; Cell Culture Techniques ; methods ; Cells, Cultured ; Humans ; Mesenchymal Stromal Cells ; cytology ; Plasma
9.Expression of SDF-1 gene in bone marrow mesenchymal stem cells of patients with myelodysplastic syndrome.
Journal of Experimental Hematology 2006;14(2):281-284
This study was aimed to investigate the expression level of stromal cell derived factor-1 gene (SDF-1) in bone marrow mesenchymal stem cells (MSC) of patients with myelodysplastic syndrome (MDS). The MSC from bone marrow samples of MDS patients were isolated, cultured and expanded, the morphology and immunophenotype of MSC were analyzed. The expression levels of SDF-1 and internal reference GAPDH in MSC of MDS patients were detected by real-time quantitative reverse transcriptase polymerase chain reaction (RQ-RT-PCR) method and were compared with expression levels of healthy donors. The results showed that the expression levels of SDF-1 in MDS patients were significantly different from those in healthy donors (1.53 +/- 0.92 vs 5.51 +/- 0.99) (P < 0.01). SDF-1 gene expression levels in bone marrow MSC of MDS patients were significantly higher than that in MSC derived from healthy donors. It is concluded that the abnormal expression of SDF-1 gene in MSC may influence the regulation of hematopoiesis of the bone marrow microenvironment in MDS patients and it is worthy of further investigation for new clue on etiological mechanism and treatment of MDS.
Adult
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Aged
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Bone Marrow Cells
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metabolism
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pathology
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Cells, Cultured
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Chemokine CXCL12
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biosynthesis
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genetics
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Female
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Humans
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Male
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Mesenchymal Stromal Cells
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metabolism
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pathology
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Middle Aged
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Myelodysplastic Syndromes
;
genetics
;
metabolism
;
pathology
10.Quantitative analysis of theophylline and its metabolites in urine of Chinese healthy subjects after oral administration of theophylline sustained-release tablets.
Ying LIU ; Yan ZHAN ; Yi-Fan ZHANG ; Xiao-Yan CHEN ; Da-Fang ZHONG
Acta Pharmaceutica Sinica 2014;49(7):1039-1043
To study the metabolite excretion of theophylline, a rapid and specific method by liquid chromatography with heated electrospray ionization tandem mass spectrometry (LC-HESI/MS/MS) method for simultaneous determination of theophylline, 1, 3-dimethyluric acid (1,3-DMU), 3-methylxanthine (3-MX) and 1-methyluric acid (1-MU) in human urine was developed using theophylline-d6 and 5-fluorouracil as internal standards. Selected reaction monitoring (SRM) with heated electrospray ionization (HESI) was used in the negative mode for mass spectrometric detection. After diluted with methanol and centrifuged, the analytes and ISs were separated on a XDB-Phenyl (150 mm x 4.6 mm, 5 microm) column with a mixture of water-methanol-formic acid (30 : 70 : 0.15) as mobile phase at a flow rate of 0.6 mL x min(-1). The linear calibration curves for theophylline, 1, 3-DMU, 3-MX and 1-MU were obtained in the concentration range of 1.0-250 microg x mL(-1), separately. The method herein described is effective and convenient, and can be used for determination of theophylline and its three metabolites. The results showed that urinary excretion ratio of theophylline, 1,3-DMU, 3-MX and 1-MU is approximately 1 : 3 : 1 : 2 in Chinese subjects, which is similar to the reported excretion pattern in Caucasian.
Administration, Oral
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Asian Continental Ancestry Group
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Calibration
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Chromatography, Liquid
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Delayed-Action Preparations
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metabolism
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Healthy Volunteers
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Humans
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Spectrometry, Mass, Electrospray Ionization
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Tablets
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Tandem Mass Spectrometry
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Theophylline
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metabolism
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urine
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Uric Acid
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analogs & derivatives
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urine
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Xanthines
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urine