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41.
42.
A simple procedure is described for the determination of the photosensitizing potency of drugs, using three leukemic cell lines, two of lymphocytic origin, L1210 and P388 and one of erythroid type, Friend-745. The procedure allows one to investigate several aspects of the photosensitization properties of tested compounds such as cellular localization and direct (trypan blue exclusion) or delayed (clonogenicity) photomediated toxicities.The method was assessed using crude hematoporphyrin derivative (HPD) as well as dihematoporphyrin ether (DHE) or commercially available Photofrin II. Results were compared to those obtained with normal cells, e.g spleen lymphocytes and erythropoietic stem cells (CFU-e), and discussed in the light of the relative response of normal versus transformed cells.Abbreviations DHE
Dihematoporphyrin Ether
- FCS
Fetal Calf Serum
- HPD
Hematoporphyrin Derivative
- PDT
Photodynamic Therapy 相似文献
43.
Summary Binding of azide to a series of copper(II) complexes has been investigated by absorption, CD and EPR spectroscopy. Axial binding of azide to Cu(II) can be differentiated from equatorial binding through the lower intensity and lack of optical activity of the LMCT band. The affinity of azide for Cu(II) increases with the overall positive charge of the complex. The preliminary data on thiocyanate binding to Cu(II) seem to agree with the trends observed for the corresponding azide adducts. 相似文献
44.
Alberto Alcázar Elena Martin Juan López-Fando Dr. Matilde Salinas 《Neurochemical research》1988,13(9):829-836
A simple and short purification procedure applicable to casein kinase II has been developed, for fully characterizing the enzyme from calf cerebral cortex cytosol. The procedure consists of four chromatographic steps: DEAE-cellulose, phosphocellulose, phosvitin-Sepharose and ATP-agarose which yields 87% pure casein kinase II. The purified enzyme shows three major bands with apparent molecular masses of 42, 38, and 27 kDa by polyacrylamide gel electrophoresis in sodium dodecyl sulfate and is self-autophosphorylated on its 27 kDa polypeptide. The enzyme shows all the characteristics described for casein kinase II from other sources: it is independent of cyclic nucleotides, calcium/phospholipids, and double-stranded poly(I).poly(C); it can utilize both ATP and GTP as phosphoryl donors and can phosphorylate both casein and phosvitin but not histone. The kinetic studies establish that theK
m for ATP is 12.5 M and 25.1 M when using phosvitin and casein respectively as phosphoryl acceptors. TheK
m for phosvitin is 0.91 mg/ml and for casein 1.43 mg/ml, while theV
max is 315 nmol/min/per mg protein and 479 nmol/min/per mg protein for phosvitin and casein respectively. The activity of the kinase is highly stimulated by KCl or NaCl, and almost completely inhibited by heparin concentrations of 1 g/ml (92%). This inhibition is reduced to only 33% in the presence of optimal KCl concentrations (150 mM). Spermine stimulates enzyme activity, whilst hemin produces a slight inhibition. 相似文献
45.
It has been known for some time that bicarbonate reverses the inhibition, by formate under HCO3
--depletion conditions, of electron transport in thylakoid membranes. It has been shown that the major effect is on the electron acceptor side of photosystem II, at the site of plastoquinone reduction. After presenting a historical introduction, and a minireview of the bicarbonate effect, we present a hypothesis on how HCO3
- functions in vivo as (a) a proton donor to the plastoquinone reductase site in the D1-D2 protein; and (b) a ligand to Fe2+ in the QA-Fe-QB complex that keeps the D1-D2 proteins in their proper functional conformation. They key points of the hypothesis are: (1) HCO3
- forms a salt bridge between Fe2+ and the D2 protein. The carboxyl group of HCO3
- is a bidentate ligand to Fe2+, while the hydroxyl group H-bonds to a protein residue. (2) A second HCO3
- is involved in protonating a histidine near the QB site to stabilize the negative charge on QB. HCO3
- provides a rapidly available source of H+ for this purpose. (3) After donation of a H+, CO3
2- is replaced by another HCO3
-. The high pKa of CO3
2- ensures rapid reprotonation from the bulk phase. (4) An intramembrane pool of HCO3
- is in equilibrium with a large number of low affinity sites. This pool is a H+ buffering domain functionally connecting the external bulk phase with the quinones. The low affinity sites buffer the intrathylakoid [HCO3
-] against fluctuations in the intracellular CO2. (5) Low pH and high ionic strength are suggested to disrupt the HCO3
- salt bridge between Fe2+ and D2. The resulting conformational change exposes the intramembrane HCO3
- pool and low affinity sites to the bulk phase.Two contrasting hypotheses for the action of formate are: (a) it functions to remove bicarbonate, and the low electron transport left in such samples is due to the left-over (or endogenous) bicarbonate in the system; or (b) bicarbonate is less of an inhibitor and so appears to relieve the inhibition by formate. Hypothesis (a) implies that HCO3
- is an essential requirement for electron transport through the plastoquinones (bound plastoquinones QA and QB and the plastoquinone pool) of photosystem II. Hypothesis (b) implies that HCO3
- does not play any significant role in vivo. Our conclusion is that hypothesis (a) is correct and HCO3
- is an essential requirement for electron transport on the electron acceptor side of PS II. This is based on several observations: (i) since HCO3
-, not CO2, is the active species involved (Blubaugh and Govindjee 1986), the calculated concentration of this species (220 M at pH 8, pH of the stroma) is much higher than the calculated dissociation constant (Kd) of 35–60 M; thus, the likelihood of bound HCO3
- in ambient air is high; (ii) studies on HCO3
- effect in thylakoid samples with different chlorophyll concentrations suggest that the left-over (or endogenous) electron flow in bicarbonate-depleted chloroplasts is due to left-over (or endogenous) HCO3
- remaining bound to the system (Blubaugh 1987).Abbreviations DCMU
3-(3,4-dichlorophenyl)-1, 1-dimethylurea (common name: diuron)
- PSII
photosystem II
- QA
first plastoquinone electron acceptor of PSII
- QB
second plastoquinone acceptor of PS II 相似文献
46.
Terry L. Riss Kenneth P. Karey B. Daniel Burleigh David Farker David A. Sirbasku 《In vitro cellular & developmental biology. Plant》1988,24(11):1099-1106
Summary A serum-free clonal density growth assay was developed for the quantification of the biological activity of human recombinant
insulin-like growth factor I (IGF-I). The assay measures IGF-I stimulated growth of Balb/c 3T3 cells cultured over 4 d on
poly-d-lysine-coated plastic surfaces in a serum-free medium formulation composed of a 1∶1 (vol/vol) mixture of Ham's F12 and Dulbecco's
modified Eagle's media, supplemented with 3.0 ng/ml bovine basic fibroblast growth factor (bFGF), 10 μg/ml human transferrin,
100 μg/ml ovalbumin, and 1.0 μM dexamethanose. Low-temperature trypsinization of serum-supplemented stock cultures combined with the use of poly-d-lysine-coated plates made it unnecessary to use serum or fibronectin to promote cell attachment and survival. Serum-free
growth conditions were optimized with respect to the concentrations of the supplements. Addition of IGF-I resulted in 3.5-fold
more cells than control cultures without IGF-I after 4 d. Deletion of bFGF resulted in no IGF-I stimulation of growth. The
concentrations of various preparations of IGF-I required to achieve one-half maximal stimulation of cell number (ED50), ranged between 1.25 and 4.7 ng/ml. In parallel assays, IGF-I was 6.6 times more potent than human recombinant insulin-like
growth factor II and 32 times more potent than insulin. When cells were seeded into medium containing IGF-I, transferrin,
ovalbumin, and dexamethasone but no bFGF, growth was minimal. Dose-response addition of bFGF showed an ED50, of 0.9 ng/ml. The methods reported are useful to monitor the biological potency of recombinant and natural-source growth
factors as well as providing a new means of studying the multiple growth factor requirements of Balb/c 3T3 cells in cultures.
This work was supported by a contract from IMCERA Bioproducts, Inc. 相似文献
47.
Purification and partial characterization of two azoreductases from Shigella dysenteriae Type 1 总被引:1,自引:0,他引:1
Two azoreductases (I and II) were purified to homogeneity from extracts of Shigella dysenteriae (type 1). Azoreductase I was a dimer of identical subunits of M(r) 28,000, whereas azoreductase II was a monomer of 11,000 M(r). Both were flavoproteins, each containing 1 mol of FMN per mol enzyme. Both NADH and NADPH functioned as electron donors for the azoreductases. Azoreductase I used Ponceau SX, Tartrazine, Amaranth and Orange II as substrates. Azoreductase II utilized all the dyes except Amaranth. 相似文献
48.
The role of phytosiderophores in acquisition of iron and other micronutrients in graminaceous species: An ecological approach 总被引:19,自引:2,他引:17
V. Römheld 《Plant and Soil》1991,130(1-2):127-134
Phytosiderophores (PS) are released in graminaceous species (Gramineae) under iron (Fe) and zinc (Zn) deficiency stress and are of great ecological significance for acquisition of Fe and presumably also of Zn. The potential for release of PS is much higher than reported up to now. Rapid microbial degradation during PS collection from nutrient solution-grown plants is the main cause of this underestimation. Due to spatial separation of PS release and microbial activity in the rhizosphere a much slower degradation of PS can be assumed in soil-grown plants. Concentrations of PS up to molar levels have been calculated under non-sterile conditions in the rhizosphere of Fe-deficient barley plants.Besides Fe, PS mobilize also Zn, Mn and Cu. Despite this unspecific mobilization, PS mobilize appreciable amounts of Fe in calcareous soils and are of significance for chlorosis resistance of graminaceous species. In most species the rate of PS release is high enough to satisfy the Fe demand for optimal growth on calcareous soils.In contrast to the chelates ZnPS and MnPS, FePS are preferentially taken up in comparison with other soluble Fe compounds. In addition, the specific uptake system for FePS (translocator) is regulated exclusively by the Fe nutritional status. Therefore, it seems appropriate to retain the term phytosiderophore instead of phytochelate. 相似文献
49.
50.