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21.
基于尿嘧啶作为一种碱基,具备一定的分子识别能力,制备了一种新颖的尿嘧啶共价修饰电极,用X射线光电子能谱和电化学方法进行了表征,并研究了酪氨酸、色氨酸、儿茶酚胺(如多巴胺,肾上腺素,去甲肾上腺素)及相关的化合物尿酸、抗坏血酸在该电极上的电化学行为,获得相应的氧化电位、电流灵敏度、线性范围和检测限等信息。其中,色氨酸检测线性范围:1.8 - 120 mM,检测限(s/n=3):0.8 mM;酪氨酸检测线性范围:1.8 - 89mM,检测限(s/n=3):0.8 mM。实验表明,尿嘧啶修饰电极能催化氧化上述电活性物质,但催化能力不同,据此,我们讨论了尿嘧啶与上述物质的相互作用,详细探讨了催化机理,扩展了对基于分子识别的传感器的研究。 相似文献
22.
LI Wan-nan ZHUANG Yan LI He SUN Ying FU Yao WU Xiao-xia ZHAO Zhi-zhuang FU Xue-qi . Edmond H. Fischer Signal Transduction Laboratory College of Life Sciences . Key Laboratory for Molecular Enzymology & Engineering Ministry of Education Jilin University Changchun P. R. China 《高等学校化学研究》2008,24(5):592-596
This study is focused on the expression of an SH2 domain-truncated form of protein tyrosine phosphatase SHP-1(designated ΔSHP-1) and the preparation of its polyclonal antibodies. A cDNA fragment encoding ΔSHP-1 was amplified by PCR and then cloned into the pT7 expression vector. The recombinant pT7-ΔSHP-1 plasmid was used to transform Rosetta(DE3) E. coli cells. ΔSHP-1 was distributed in the exclusion body of E. coli cell extracts and was purified through a two-column chromatographic procedure. The purified enzyme exhibited an expected molecular weight on SDS-gels and HPLC gel filtration columns. It possesses robust tyrosine phosphatase activity and shows typical enzymatic characteristics of classic tyrosine phosphatases. To generate polyclonal anti-ΔSHP-1 antibodies, purified recombinant ΔSHP-1 was used to immunize a rabbit. The resultant anti-serum was subjected to purification on ΔSHP-1 antigen affinity chromatography. The purified polyclonal antibody displayed a high sensitivity and specificity toward ΔSHP-1. This study thus provides the essential materials for further investigating the biological function and pathological implication of SHP-1 and screening the inhibitors and activators of the enzyme for therapeutic drug development. 相似文献
23.
在波长200~400 nm范围内,测定酪氨酸、色氨酸和苯丙氨酸混合体系的吸光度,用连续小波变换(CWT)对光谱数据进行预处理,再用支持向量回归(SVR)方法进行建模,建立了支持向量回归紫外分光光度法同时测定酪氨酸、色氨酸和苯丙氨酸的方法,用所建方法对模拟样品进行了测定。结果表明,酪氨酸、色氨酸和苯丙氨酸预测结果的回收率在98%~102%之间,测定结果准确。 相似文献
24.
Elena V. Suprun Svetlana A. Khmeleva Yana Y. Kiseleva Sergey P. Radko Alexander I. Archakov Victoria V. Shumyantseva 《Electroanalysis》2016,28(9):1977-1983
The tyrosine based electrochemical analysis of synthetic amyloid‐β (Aβ) peptide – an analog of natural peptide implicated in Alzheimer's disease pathogenesis – was applied for a quantitative estimation of peptide aggregation in vitro. The analysis was carried out by square wave voltammetry (SWV) on carbon screen printed electrodes (SPE). The electrooxidation peak current (Ip) for Aβ42 peptide in different aggregation states was directly compared with the size and structure of Aβ42 aggregates occurring in the analyzed sample. Dynamic light scattering (DLS) and thioflavin T (ThT) based fluorescence assay were employed to estimate the size and structure of Aβ42 aggregates. The Ip was found to decrease in a linear fashion when the average diameter of aggregates and the relative ThT fluorescence in Aβ42 solutions exceeded 35 nm and 3, respectively, while being nearly constant below these values. It was suggested that the electrooxidation current is mostly generated by peptide monomers and that a depletion of the monomer pool due to inclusion of Aβ42 molecules in aggregates is responsible for the decrease of electrooxidation current. The direct electrochemistry is emerging as a method complementary to methods based on aggregates’ detection and commonly employed for monitoring Aβ aggregation. The work further enlarges the basis for application of the cost‐effective and rapid electrochemical techniques, such as SWV on carbon SPE, to in vitro studies of Aβ aggregation. 相似文献
25.
V. B. Gavrilov A. V. Lychkovskii E. P. Shostak S. V. Konev 《Journal of Applied Spectroscopy》1998,65(3):379-384
A more simple and sensitive assay for the quantitative determination of tyrosine in blood plasma is developed on the basis
of a modification of the method of S. Udenfriend (1962). A volume of 0.2 ml of plasma is used in the analysis; after its deproteinization
with trichloroacetic acid, it is reacted with sodium nitride, nitrous acid, and 1-nitroso-2-naphthol at 65°C, and the content
of the reaction product is measured by the fluorescence at λex/λem=460/570 nm. The consumption of plasma and reagents is reduced by a factor of 5–10 compared to the original method; in addition,
the stage of extraction with an organic solvent is excluded. The linear dependence of the fluorescence signal on the tyrosine
concentration within the range of 2–60 μg/ml, high specificity, accuracy, and reproducibility of the assay are shown. The
importance of tyrosine determination in monitoring of glucocorticoid therapy and protein catabolism is discussed.
Institute of Photobiology, National Academy of Sciences of Belarus, 27, Akademicheskaya St., Minsk, 220072, Belarus. Translated
from Zhurnal Prikladnoi Spektroskopii, Vol. 65, No. 3, pp. 366–371, May–June, 1998. 相似文献
26.
A rapid, selective, and sensitive liquid chromatography–tandem mass spectrometry method was developed and validated for the simultaneous determination of unbound sunitinib and its active metabolite N‐desethyl sunitinib in plasma. Plasma and post‐dialysis buffer samples were extracted using a liquid–liquid extraction procedure with acetonitrile–n‐butylchloride (1:4, v/v). Chromatographic separation was achieved on a Waters X‐Terra® MS RP18 column with a mobile phase consisting of acetonitrile and water (60:40, v/v) containing formic acid (0.1%, v/v) using an isocratic run, at a flow‐rate of 0.2 mL/min. Analytes were detected by electrospray tandem mass spectrometry in the selective reaction monitoring mode. Linear calibration curves were generated over the ranges 0.1–100 and 0.02–5 ng/mL for sunitinib and 0.2–200 and 0.04–10 ng/mL for N‐desethyl sunitinib in plasma and in phosphate‐buffered solution, respectively. The values for both within‐day and between‐day precision and accuracy were well within the generally accepted criteria for analytical methods. The analytical range was sufficient to determine the unbound and total concentrations of both analytes. The method was applied for measurement unbound concentrations in addition to total concentrations of sunitinib and its metabolite in plasma of a cancer patient receiving 50 mg daily dose. Copyright © 2012 John Wiley & Sons, Ltd. 相似文献
27.
Mushroom tyrosinase was immobilized on modified polystyrene—polyaminostyrene (PSNH) and polymethylchloridestyrene (PSCL)—to
produce l-DOPA from l-tyrosine. Glutaraldehyde was used as an activating agent for the PSNH to immobilize the tyrosinase and 10% (w/v) glutaraldehyde
was optimal in conferring the highest specific activity (11.96 U/g) to the PSNH. Methylchloride on the PSCL was directly linked
with the tyrosinase, and 1.5 mmol of Cl/g was optimal in attaining the specific activity of 17.0 U/g. The temperature and
optimal acidity were, respectively, 60°C and pH 5.5 for the PSNH, and 70°C and pH 3.0 for the PSCL. In a 50-mL batch reactor
working over 36 h, the l-DOPA production rate at 30°C was 1.44 mg/(L·h) for the PSNH and 2.33 mg/(L·h) for the PSCL. The production rate over 36 h
was 3.86 mg/(L·h) for the PSNH at 60°C and 5.54 mg/(L·h) for the PSCL at 70°C. Both of the immobilized enzymes showed a remarkable
stability with almost no change in activity after being stored wet. The operational stability study indicated a 22.4% reduction
in l-DOPA production for the PSNH and an 8.63% reduction for the PSCL over seven runs (each run was for 144h at 30°C) when the
immobilized enzymes were used under turnover conditions. The immobilized tyrosinase was more stable on the PSCL than on the
PSNH. 相似文献
28.
29.
SHI Dong-lei DONG Hong-bo ZHU Zhi-cheng SUN Mei SU Ji-quan ZHANG Xing-yi XU Yue-chi FU Xue-qi 《高等学校化学研究》2007,23(2):204-207
This study focuses on the expression of human protein tyrosine phosphatase 1B(PTP1B) catalytic domain (△PTP1B) and preparation of polyclonal antibody against △PTP1B. △PTP1B gene was PCR amplified with the cDNA of human PTP1B as the template, and cloned into the pT7 expression vector. The recombinant pT7-△PTP1B was expressed in E. coli Rosetta( DE3 ) host cells and purified. The antiserum was prepared by immunizing rabbit with purified recombinant △PTP1B. The polyclonal antibody against △PTP1B was purified by PVDF immobilized antigen affinity chromatography. △PTP1B was correctly cloned, expressed, and purified as confirmed by PCR, DNA sequence ratio) and 0. 1 ng, respectively. This study provides an important basis for further studying the biological function of PTP1B and its relationship with human diseases. 相似文献
30.
酪氨酸激酶在生物分子的信号转导中起着非常重要的作用,目前除抗体技术外尚无有效的化学方法能够实现对酪氨酸磷酸化蛋白或多肽的选择性富集,然而抗体通常成本较高,而且往往会有模体序列的选择性识别。本文发展了一种基于化学反应的酪氨酸磷酸化肽段的选择性富集,该方法利用了β消除反应只能发生在丝氨酸和苏氨酸磷酸化多肽的特性,以反相选择方法,从而实现对酪氨酸磷酸化肽段的选择性富集。以标准多肽对其反应效率和回收率进行了考察,20分钟内丝氨酸磷酸化多肽的β消除反应效率可达99%以上,而同时酪氨酸磷酸化肽段可保持70%的回收率。进一步以六种标准蛋白混合物的酶解产物对其进行考察,经β消除反应和亲和富集之后,只有酪氨酸磷酸化多肽可以被检测出来,该方法为蛋白质酪氨酸磷酸化的分析提供了一种新的手段。 相似文献