RESUMO
Temperature measurements in a laser heated diamond anvil cell (DAC) are currently limited to temperatures above 1000 K using optics and detectors in the visible range. We have built a pyrometer in the IR range and expanded the lower limit of temperature detection to 400 K. The pyrometer is designed for very low thermal radiation intensities, measured sequentially through a set of bandpass filters in the range of 1.2-3.4 microm using very efficient IR photodetectors. The thermal radiation from the center of the cw Nd:YAG laser heated spot is least square fitted to a Planck curve, using a gray body approximation. Melting is detected by changes in the light scattering picture of an auxiliary He-Ne laser from the surface of the hot spot, and by a change in slope in the plot of hot spot temperature versus laser power. In this work we demonstrate measurement of the melting curve of zinc up to 25 GPa. The melting curve is in very good agreement with previous results which were taken up to 6 GPa in a large volume press.
RESUMO
The synthesis of the fluorescent derivative of adenosine, by reaction with 5-(dimethylamino)naphthalene-1-sulfonyl chloride in dry pyridine at low temperature, yielding 3'-O-[5-(dimethylamino)naphthalene-1-sulfonyl]adenosine (3'-O-dansyladenosine), is here described. 3'-O-Dansyladenosine is partially soluble in water (approximately 10(-4) M) and upon excitation at 325 nm exhibits maximum fluorescence emission at 516 +/- 22 nm (corrected) in buffered aqueous solution at pH 7.6 with a quantum yield of 0.21 and a lifetime of 11.8 +/- 0.2 ns. The fluorescence of 3'-O-dansyladenosine is sensitive to the polarity of its solvent: in pyridine, a quantum yield of 0.61 at the emission maximum of 435 nm was observed. 3'-O-Dansyladenosine is a reversible competitive inhibitor of adenosine deaminase with a moderate inhibitive dissociation constant K1 = (1.54 +/- 0.13) X 10(-5) M. The enzyme-substrate analogue association constant was determined by equilibrium dialysis to be K = (0.69 +/- 0.05) X 10(5) M-1, very close to KI-1. The hydrophobic nature of its binding site in adenosine deaminase is evident from the strong blue shift of the fluorescence emission maximum to 440 nm, the 4-fold increase in fluorescence quantum yield, and the longer lifetime of 15.8 +/- 0.2 ns; the tight, rigid nature of the complex is evident from its high fluorescence polarization value, 0.23. An 85% decrease in the fluorescence emission intensity of the adenosine deaminase-3'-O-dansyladenosine complex in the presence of adenosine indicates the selective binding to the enzyme active site. Correlation between the conformation of the probe, either when free in various solvents or when bound to the enzyme, and its fluorescence quantum yield is noted. 3'-O-Dansyladenosine is suitable for fluorescent labeling of adenosine deaminase in cell systems.
