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Objective To establish an UHPLC-MS/MS method for the determination of uracil (U) and dihydrouracil (UH2) in human plasma. Methods A positive ion detection mode was adopted on the Agilent 6460A mass spectrometer. Chlorouracil was used as the internal standard. 3% bovine serum albumin was used as surrogate plasma matrix. The pretreatment of plasma sample was completed based on liquid-liquid extraction with ethyl acetate. The chromatographic separation was achieved on an Agilent Poroshell 120 SB-Aq (2.1 mm×100 mm, 2.7 μm) column with gradient elution. The mobile phase was 5 mmol/L ammonium acetate aqueous solution and acetonitrile solution. The flow rate was 0.3 ml/min. The column temperature was 30°C. The injection volume was 5 μl. Results The linear range of uracil and dihydrouracil was 10.0-1500.0 ng/ml. Both of uracil and dihydrouracil had good linear relationship with correlation coefficient (r)>0.990. Both of inter- and intra-day precision was <15%. Conclusion The established method is simple, selective, and suitable for the determination of U and UH2 in human plasma.
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Since the last decade, hybrid drug strategies have attracted many researchers for their improved anti-cancer potential incomparison to single drug components. Complying to this approach, 28 novel Uracil–Coumarin hybrids with differentsized linkers (2–5 carbon atoms) and substituents were designed to occupy the active site of protein epidermal growthfactor receptor (EGFR) tyrosine kinase (Protein Data Bank ID: 1M17). Molecular docking studies were performedfor all ligands (A1-D7) to identify the potential candidate using Schrödinger software. The relative binding affinity ofhybrids toward EGFR was compared with standard Erlotinib on the basis of gScore and Emodel score. Positively, allthe hybrids docked inside the cavity and showed significant interactions, compounds A6, A2, and A7 with short-chainlinker (two carbon atoms) and halogen substituents were found to have more interactions and better docking score thanstandard Erlotinib. The visualization results depicted that compound A6 showed the highest affinity and formed thebest binding pose to the target EGFR with gScore = −8.891 kcal/mol and Emodel score = −100.744 in comparison tostandard Erlotinib (gScore of −8.538 kcal/mol and Emodel score = −80.588). Moreover, a molecular dynamics studyalso reveals that ligand A6 forms a stable complex with root mean square deviation (RMSD) of 0.3 nm and the plateauphase started just after 10 ns (time). Hence, the present research provides computational insights of Uracil–Coumarinhybrids as potential ligands against EGFR tyrosine kinase and in future in vitro investigations of these hybrids mayprove their therapeutic potential against cancer.
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Cancer is the most dreadful disease and the second main cause of death worldwide. The continuous developments havebeen going on in order to design potent molecules such that this leading cause of death can be dealt with. In order todecrease the level of toxicity and to improve the selectivity of drugs toward cancer targets, the development of hybridmolecules has become the center of research, and scientists are doing timeless efforts to generate such a hybrid whichhas got no comparison with the previous developments. The heterocyclic moiety Uracil and many of its derivativeswere already exposed as promising anticancer agents. Moreover, coupling of Uracil and 5-Fluorouracil (5-FU) withdifferent pharmacophores has been proven to be an excellent strategy against cancer. Hence, the present review is aneffort to collectively represent all the earlier and recent developments of Uracil and 5-FU hybrids reported to have asignificant anticancer profile. Expectantly, we can assure that this article can serve as the basis for further developmentsin Uracil and 5-FU hybrids and will surely motivate the medicinal chemists for producing unique anticancer drug
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Objective To establish a UPLC method for the simultaneous determination of uracil, adenosine, guanosine, and uridine in Aconiti Lateralis Radix Praeparata (ALRP) and Aconiti Radix (AR), and to compare the content difference of four nucleosides in the samples based on multivariate statistical analysis. Methods Nucleosides in ALRP and AR were extracted with purified water by ultrasonic extraction. The UPLC method was performed on a BEH C18 (100 mm × 2.1 mm, 1.7 μm) through a gradient elution of acetonitrile and water at a flow rate of 0.2 mL/min with column temperature at 30 ℃. The injection volume was set at 4.0 μL, and the detection wavelength was set at 260 nm. Difference significance analysis, hierarchical cluster heatmap analysis, principal component analysis, and TOPSIS analysis were used for data processing to comprehensively evaluate the quality of ALRP and AR. Results The method was in accordance with the regulations, the quantitative evaluation of uracil, adenosine, guanosine, and uridine was in good linear range (r2 > 0.999 3), and the average recovery was 99.86%, 99.14%, 99.74%, and 98.71% respectively, and the RSDs were all less than 2.0%. Taking the four nucleosides as indexes, the samples No. 18 of ALRP and the samples No. 23 of AR were the best in quality. Conclusion The established method is simple, accurate, and reliable with good precision, repeatability, and stability, which can be used for the simultaneous determination of four nucleosides in ALRP and AR, and it might provide the scientific basis for the study of water-soluble constituents in ALRP and AR.
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AIM To establish the HPLC fingerprints of Kangfuxin Liquid (extract of Periplaneta americana L.) and to determine the contents of six constituents.METHODS The analysis of this drug was performed on a TOSOH TSK-GEL ODS column (250 mm × 4.6 mm,5 μm),with the mobile phase comprising of acetonitrile-water (containing 0.07％ acetic acid) flowing at 1.0 mL/min in a gradient elution manner,and the detection wavelength was set at 280 nm.RESULTS There were twenty-four common peaks in the fingerprints of ten batches of samples (Ⅰ-Ⅹ) with the similarities of 0.932-0.993 (except for sample Ⅰ).Uracil,hypoxanthine,xanthine,inosine,protocatechuic acid and Cyclo (Gly-Tyr) showed good linear relationships within the ranges of 3.460-173.0,3.960-198.0,3.596-179.8,1.338-66.9,3.672-183.6 and 3.552-177.6 μg/mL,whose average recoveries (RSDS) were99.8％ (2.65％),98.0％ (2.55％),99.7％ (1.59％),100.7％ (2.80％),102.0％ (2.09％) and 99.6％ (1.88％),respectively.CONCLUSION This accurate,stable and simple method can be used for the quality control of Kangfuxin Liquid.
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OBJECTIVE: To establish a quantitative method for determining the contents of uracil, cytidine, hypoxanthine, xanthine, uridine, inosine, and guanosine in Holotrichia diomphalia Larvae by HPLC. METHODS: The HPLC analysis was performed on Waters HSS T3 column(4.6 mm×250 mm, 5 μm). The mobile phase was composed of acetonitrile(A) and water(B), and gradient elution was carried out. The flow rate was 1.0 mL•min-1. The column temperature was maintained at 30 ℃. The detection wavelength was 260 nm. RESULTS: The correlation coefficients of the seven components ranged from 0.999 1 to 1.000 0. The average recovery rates(n=6) were between 91.2% and 97.4%, and the RSDs were between 1.5% and 2.3%. CONCLUSION: The proposed method is simple, accurate and reliable, thus providing basis for comprehensive quality control of Holotrichia diomphalia Larvae.
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OBJECTIVE: To establish a preparation process of pivotal intermediate of SKI2496, which is low-cost, environmental-friendly and suitable for industrialization as well. METHODS: 1-(2-Fluoro-6-(trifloromethyl)benzyl)urea(2) was synthesized from 2-fluoro-6-(trifluoromethyl)benzylamine(1) with urea,followed by aminolysis with t-butyl acetoacetate and cyclization to give 1--6-methylpyrimidine-2,4(1H,3H)-dione(4).Finally,the title product was obtained via bromation and condensation reaction with piperazine. RESULTS: The synthetic process included four steps with an overall yield of 44.6%(based on compound 1) and its structure was confirmed by 1H-NMR and MS. CONCLUSION: The process is easy to operate, safe and suitable for industrial production.
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<p><b>Objectives:</b> To evaluate the efficacy of tegafur–uracil (UFT), a prodrug of 5-fluorouracil, plus cisplatin and dexamethasone in patients with docetaxel-refractory prostate cancers.</p><p><b>Methods:</b> Twenty-five patients with docetaxel-refractory prostate cancer were administered oral UFT plus intravenous cisplatin (UFT-P therapy) and dexamethasone. Treatment responses were assessed monthly via prostate-specific antigen (PSA) level measurements. Treatment-related adverse events and overall survival were also assessed.</p><p><b>Results:</b> UFT-P therapy resulted in decreased PSA levels in 14 (56%) patients and increased PSA levels in 11 (44%). In patients with increased PSA levels, 7 (64%) of the 11 patients displayed decreased PSA doubling times. The UFT-P therapy response rate was 84% (21/25 patients). Imaging studies revealed that tumor shrinkage during UFT-P therapy occurred in 1 patient in whom bilateral hydronephrosis caused by lymph node metastasis improved. The median survival time from docetaxel initiation was 36 months. In UFT-P-treated patients, the median PSA progression and overall survival times were 6 and 14 months, respectively. UFT-P treatment-related adverse events were mild diarrhea, general fatigue, and anorexia. Treatment was not discontinued for any of the patients. UFT-P therapy did not cause serious hepatic or renal dysfunction or pancytopenia.</p><p><b>Conclusions:</b> UFT-P therapy is a safe and effective treatment for patients with docetaxel-refractory prostate cancer, although large-scale, multicenter, prospective studies are needed to validate these findings.</p>
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Objective:To study the HPLC fingerprint of Ningxinbao capsules, and establish a method for the simultaneous content determination of uracil, uridine, adenine and adenosine. Methods: The separation was carried out on an Agilent Zorbax SB-Aq C18 column(250 mm × 4. 6 mm, 5μm) with gradient elution using methanol-0. 05 mol·L-1 potassium dihydrogen phosphate at 30℃ and at a flow rate of 1. 0 ml·min-1 , the detection wavelength was 212 nm for fingerprint and 260 nm for the determination of the four nu-cleosines. Totally 10 batches of samples were analyzed with the developed HPLC fingerprint and the determination method, the data calculation was performed with similarity evaluation system in the chromatographic fingerprint of TCM. Results: In the fingerprint, 10 common peaks were marked and the separation of the four nucleosines was good. Conclusion:The method is simple and reliable. The HPLC fingerprint and contents of the four nucleosines in Ningxinbao capsules can be used for the quality control.
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OBJECTIVE:To establish a method for the simultaneous determination of uracil,hypoxanthine,xanthine,uridine and adenosine in Eisenia foetida. METHODS:HPLC was performed on the column of Venusil XBP C18 with mobile phase of 50%methanol-0.01 mol/L potassium dihydrogen phosphate solution(gradient elution)at a flow rate of 1.0 ml/min. The detection wave-length was 254 nm with a column temperature at 30 ℃,and the injection volume was 10 μl. RESULTS:The linear range was 0.05-1.60 μg for uracil(r=0.9991),0.05-1.60 μg for hypoxanthine(r=0.9993),0.002-0.010 μg for xanthine(r=0.9991), 0.0075-0.24 μg for uridine(r=0.9999)and 0.0250-0.80 μg for adenosine(r=0.9990);RSDs of precision,stability and accuracy tests were lower than 3%;recoveries were 98.30%-100.76%(RSD=0.8%,n=9),98.68%-100.96%(RSD=0.8%,n=9), 95.16%-97.67%(RSD=0.9%,n=9),96.15%-99.57%(RSD=1.5%,n=9)and 96.39%-101.93%(RSD=1.8%,n=9),respective-ly. CONCLUSIONS:The method is simple and stable with good reproducibility,and can be used for the simultaneous determina-tion of uracil,hypoxanthine,xanthine,uridine and adenosine in E. foetida.
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Objective: To study the chemical constituents from the stems and leaves of Cirsium henryi. Methods: The chemical constituents were isolated by various chromatography techniques and their structures were elucidated on the basis of spectroscopic analysis. Results: Sixteen compounds were isolated and identified to be stearic acid (1), 2,3-dihydroxypropyl hexadecanoate (2), palmitic acid (3), taraxasterol (4), pseudo taraxasterol (5), taraxa-sterol acetate (6), β-sitosterol (7), daucosterol (8), protocatechuic acid (9), uracil (10), linarin (11), apigenin (12), quercitrin (13), acacetin (14), acacetin-7-O-β-D-glucoside (15), and 4-hydroxy- 3,5-dimethoxy benzoic acid (16), respectively. Conclusion Compounds 3 and 10-16 are isolated from the plant for the first time, and compounds 10, 13, and 16 are isolated from the plants of Cirsium Mill. Emend. Scop. for the first time.
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Here, we describe a uracil-DNA glycosylase (UNG)-treated reverse transcription loop-mediated isothermal amplification (uRT-LAMP) for the visual detection of all subtypes of avian influenza A virus (AIV). The uRT-LAMP assay can prevent unwanted amplification by carryover contamination of the previously amplified DNA, although the detection limit of the uRT-LAMP assay is 10-fold lower than that of the RT-LAMP without a UNG treatment. To the best of our knowledge, this is the first successful application of deoxyuridine triphosphate/UNG strategy in RT-LAMP for AIV detection, and the assay can be applied for the rapid, and reliable diagnosis of AIVs, even in contaminated samples.
Subject(s)
Animals , Deoxyuridine , Diagnosis , DNA , Influenza in Birds , Limit of Detection , Reverse Transcription , Uracil-DNA GlycosidaseABSTRACT
Objective: To develop a rapid high-performance liquid chromatography coupled with diode array detection (HPLC-DAD) method for the simultaneous determination of six polar compounds in Ophiocordyceps sinensis. Methods: A poroshell SB Aq column (50 mm × 4.6 mm, 2.7 μm) and gradient elution were used; The detection wavelength of compounds was set at 260 nm. The chromatographic peaks of the six investigated compounds in sample were identified by comparing their retention times with reference compounds. Results: All calibration curves showed good linearity (r > 0.999) within the tested ranges. The intra- and inter-day precisions of the six analytes were less than 0.8% and 2.1%, respectively, and the recoveries of the six analytes were between 95% and 103%. The validated method was successfully applied to the determination of six polar compounds in O. sinensis samples. Conclusion: The poroshell SB Aq column is suitable for the rapid analysis of polar components in Chinese materia medica on conventional HPLC system and the developed HPLC method is also helpful to the quality control of O. sinensis. © 2014 Tianjin Press of Chinese Herbal Medicines.
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Objective: To initially establish HPLC fingerprints for analysis of 17 batches of the introdued jujube (Ziziphus jujuba) varieties in order to lay the foundation for scientific evaluation and production quality control of jujube. Methods: TOSOH TSKgel ODS-100V C18 column (150 mm × 4.6 mm, 3 μm) was used. The mobile phase was as follows: methanol and 0.3% KH2PO4 in gradient elution mode. The detective wavelength was 260 nm, the flow rate was 1.0 mL/min, and the separation was performed at 40 ℃. Similarity Evaluation System for Chromatographic Fingerprint of Chinese Materia Medica 2004A published by the state Pharmacopeia Committee of China was adopted for the fingerprint analysis on the 17 batches of introdued jujube varieties in Germplasm Nursery of Tarim University. Results: The HPLC characteristic fingerprint of jujube has been established. A total of 22 common peaks were characterized, the peaks appeared uniform and were well-separated. The uracil, hypoxanthine, adenine, cytidine, uridine, cAMP, and cGMP were determined by HPLC method. Conclusion: HPLC chromatographic fingerprint analysis technology can be used for the identification of jujube and its quality control.
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This article was aimed to study the effect of pinellia combined with prunella vulgarisin of different ratios on the extraction yield of trigonellin and uracil, in order to investigate the changing disciplines and reasonable com-bination of Shuangxia Soup. A Welch Ultimate AQ-C18 column (4.6 mm×250 mm, 5 μm) was used in the study. The mobile phase, consisting of solvent A (water) and solvent B (acetonitrile) was programmed as a linear gradi-ent. The flow rate was 1.0 mL·min-1. And the detection wavelength was 265 nm. The column temperature was 25℃. HPLC was used in the determination of extraction yield of trigonellin and uracil for the comparison among different compatibilities. The results showed that the linearity was achieved in the range of 0.872-4.36 ng for trigonelline, and 10.8-50.4 ng for uracil. The average recoveries were 96.53%-101.48%, with RSD of 0.18%-4.26%. Com-pared with pinellia, the extraction yield of trigonelline was reduced and the extraction yield of uracil was increased after the compatibility. It was concluded that different compatibilities of Shuangxia Soup had effects on the extrac-tion yield of trigonelline and uracil. The best compatibility ratio still requires in-depth study.
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Objective To establish an effective method for the determining hypoxanthine, xanthine, uridine and uracil in earthworm in Shanghai Pheretima and Pheretima aspergillum (E. Perrier), contributing to quality control of the medicinal material. Methods Hypoxanthine, xanthine, uridine and uracil were extracted from the earthworms with 0.9% NaCl by ultrasonic and determined by HPLC. The chromatographic conditions were: SORBAX SB-Aq column (250 mm×4.6 mm, 5 μm, Aglient Co.,Ltd), 5 mmol/L KH2PO4 (pH 2.9) as the mobile phase with a flow rate of 1 mL/min,the detection wavelength was 254nm, the column temperature was set at 30℃, and the injection volume was 10 μL. Hypoxanthine, xanthine, uridine and uracil were determined by HPLC at the same time. Results The linear range was 0.5-100 μg (r=0.999 9) for hypoxanthine, with the average recovery being 99.37%, RSD=1.36% (n=6). The linear range was 0.5-100 μg (r=0.993 1) for xanthine, with the average recovery being 91.57%, RSD=1.40% (n=6). The linear range was 0.5-100 μg (r=0.999 9) for uridine, with the average recovery being 95.31%, RSD=1.64% (n=6). The linear range was 0.5-100 μg (r=0.999 9) for uracil, with the average recovery being 100.21%, RSD=1.98% (n=6). Conclusion The current method is reproducible and has satisfactory recovery, and it can be used to determine hypoxanthine, xanthine, uridine and uracil in earthworm medicinal material.
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Objective: To study the chemical constituents from Leonurus japonicus Injection. Methods: The constituents were purified by macroporus resin, silica gel, Sephadex LH-20, and recrystallization. The structures were identified by spectroscopic methods and spectroscopic analysis. Results: Fifteen compounds were isolated from L. japonicus Injection, including four amino acids and their derivatives such as N-isobutyl valine (1), valine (2), alanine (3), L-pyroglutamic acid methyl ester (4), nine alkaloids: stachydrine (5), choline (6), trigonelline (7), uracil (8), 5-methyluracil (9), 2', 3'-O-isopropylidene uridine (10), 3-hydroxypyridine (11), 5-hydroxyl-2-hydroxymethylpyridine (12), and 2-methyl-3-hydroxypyridine (13); and two furoic acids such as furan-2- carboxylic acid (14) and 5-hydroxymethyl-2-furancarboxyic acid (15). Conclusion: In addition to compounds 5 and 8, the others are isolated from the plants of Leonurus Linn. for the first time. Among them, compound 1 is a natural product. Its NMR data, undoubtedly assigned by 2D-NMR, have been reported for the first time.
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Objetivos: Diseñar una estrategia alternativa por PCR para el genotipado de secuencias ricas en citosinas, basada en modificación nucleotídica. Material y métodos: Se modificó el gen FMR1 nativo de ocho individuos clínicamente no afectados por el Síndrome X frágil, cambiando las citosinas por uracilos, empleando bisulfito de sodio. El ADN modificado fue purificado y cuantificado por espectrofotometría. Las estructuras alternativas y potenciales islas CpG que adopta el microsatélite inestable fueron simuladas con los programas MFOLD y CpGplot. Se generaron cebadores específicos que hibriden tanto con el microsatélite modificado (Primer T) y con una secuencia modificada de las islas CpG (Primer M), utilizando el programa MethPrimer. Finalmente, ambas secuencias fueron amplificadas por PCR y los amplicones fueron separados por electroforesis en gel de poliacrilamida (PAGE por sus siglas en inglés) al 6% y visualizados con tinción de nitrato de plata. Resultados: La modificación del ADN fue evidenciada por espectrofotometría al uracilo. Las estructuras observadas en la simulación fueron las horquillas encontrándose dos potenciales islas CpG. La amplificación con los cebadores T, confirmó el diseño in silico desarrollado para abordar la estructura en horquillas. La amplificación con los cebadores M permitió detectar metilación de la primera isla CpG del gen FMR1.Conclusión: Se propone un diseño alternativo para amplificación de secuencias de microsatélite que contengan citosinas metiladas y no metiladas. Se requieren estudios posteriores con muestras de ADN que contengan microsatélites muy expandidos para validar su aplicación para diagnóstico molecular. (AU)
Objectives: To design an alternative strategy for genotyping cytosine-rich sequences using PCR and nucleotide modification. Methods: The FMR1 gene wild type was modified in the DNA obtained from eight individuals clinically unaffected for Fragile X Syndrome; cytosines were replaced by uracils using sodium bisulfite. Modified DNA was purified and quantified by spectrophotometry. Alternative structures and potential CpG islands of the unstable microsatellite were simulated using MFOLD and CpGplot tools. Specific primers were generated to hybridize with both the modified microsatellite (Primer G) and a modified sequence of CpG islands (Primer M) using the MethPrimer software. Finally, both sequences were amplified by PCR and the amplicons were separated by electrophoresis in silver-stained PAGE 6% gels. Results: The DNA modification was evidenced by spectrophotometry to uracil. We found two potential CpG islands. The amplification with T primers confirmed the "in silico" design developed to engage hairpin structures. The amplification with M primers detected methylation of the first CpG island in the FMR1 gene. Conclusion: We propose an alternative design for amplifying microsatellite sequences that contain methylated and unmethylated cytosine bases. Further studies are required with DNA samples containing expanded microsatellites to validate its molecular diagnostic application. (AU)
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Humans , Male , Female , Uracil , Polymerase Chain Reaction , Cytokines , Fragile X SyndromeABSTRACT
Objective: To study the chemical constituents from the roots and rhizomes of Acorus tatarinowii. Methods: Using different chromatographic methods to isolate and purify the constituents of A. tatarinowii, and their structures were identified by physicochemical properties and spectroscopic technology. Results: Thirteen compounds were isolated and identified as fumaric acid (1), nicotinic acid (2), p-hydroxybenzonic acid (3), uracil (4), N-trans-feruloyltyramine (5), thymine (6), variecolorquinone A (7), butanedioic acid (8), tatarol (9), tataroside-12-β-D-glucoside (10), β-sitosterol (11), 2, 5-dimethoxy- benzoquinone (12), and 5-hydroxymethyl-2-furaldehyde (13). Conclusion: Compounds 1-7 are isolated from the plants in Acorus L. for the first time.
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Objective: To study the chemical constituents from Scolopendra multidens. Methods: Compounds were isolated and purified by a combination of chromatographic techniques including silica gel, ODS, Sephadex LH-20 column chromatography, middle and low pressure preparative chromatograms, and pre-HPLC. The structures were elucidated on the basis of physicochemical properties and spectroscopic analyses. Results: Ten compounds were separated and identified as uracil (1), 7, 8-dimethyl-isoalloxazine (2), indole-3-acetamide (3), N-(2-phenylethyl) acetamide (4), (3S)-1, 2, 3, 4-tetrahydro-β-carboline-3-carboxylic acid (5), cyclo-(L-Ile-L-Pro) (6), cyclo-(L-Leu-L-Pro) (7), cyclo-(L-Phe-L-Pro) (8), cyclo-(L-Phe-L-Tyr) (9), and cyclo-(L-Val-L-Pro) (10). Conclusion: All the compounds are isolated from S. multidens for the first time.