EFFICIENCY OF PHOTOAFFINITY LABELING DNA HOMOPOLYMERS AND COPOLYMERS WITH ETHIDIUM MONOAZIDE *

Abstract— Photoaffinity labeling of synthetic DN As with ethidium monoazide was studied to determine if the efficiency of adduct formation was related to DNA sequence. Equilibrium drug binding to DNA homopolymers and copolymers was quanitified by phase partition techniques. The amount of drug bound to a deoxypolymer at equilibrium was then compared to the fraction of ethidium analog covalently-linked following photoactivation at the same drug/DNA input ratio. There were significant sequence-related differences in the ability of the photoaffinity probe to label DNA covalently. The efficiency of covalent-adduct formation decreased in the order poly(dG-dC)^. poly(dG-dC)> poly-(dG). poly(dC)poly(dA-dT). poly(dA-dT)poly(dA). poly(dT). Ethidium monoazide was about 2-fold more efficient in labeling deoxyhomopolymers and deoxycopolymers composed of G-C pairs than the A-T base counterparts. In low ionic buffers (0.015 M Na⁺), the efficiency of photoactivation decreased with increasing ethidium monoazide concentrations. However. the base sequence effect was observed over a 40-fold range of drug concentrations. Therefore, the amount of ethidium monoazide bound to a DNA site after irradiation does not appear to represent the true affinity of the drug for that site.     

IRRADIATION IN THE FROZEN STATE: A TECHNIQUE FOR DIRECT PHOTOAFFINITY LABELING

UV irradiation of rabbit muscle phosphofructokinase (PFK) in the presence of adenosine 3',5'-cyclic phosphate (cAMP) resulted in the covalent attachment of this ligand molecule to the enzyme protein. Irradiation in the frozen ice state enhanced the rate of this incorporation more than 10-fold above that achieved in aqueous solution, without significantly affecting the rate of photodestruction of the protein. [³H]-cAMP and [³²P]-cAMP were each incorporated into PFK at identical rates in the frozen state. Rates of photoincorporation in the frozen and liquid states were both half-maximal at a free ligand concentration approximately equal to the dissociation constant of cAMP and PFK. Adenosine diphosphate (ADP) and adenosine monophosphate (AMP), both of which are known to compete for cAMP binding to PFK, inhibited photoincorporation of cAMP. Guanosine monophosphate (GMP), inosine monophosphate (IMP), and guanosine 3',5'-cyclic phosphate (cGMP), which do not compete for cAMP binding, had no effect on photoincorporation of cAMP. Irradiation of [³H]-AMP or [³H]-ADP resulted in photoincorporation into PFK at 0°C, with enhancement at — 77°C similar to that noted with cAMP.     

THE USE OF SPIN LABEL OXIMETRY IN THE STUDY OF PHOTODYNAMIC INACTIVATION OF CHINESE HAMSTER OVARY CELLS

Spin label oximetry has been used to study the effects of photosensitizer hematoporphyrin on oxygen consumption in Chinese hamster ovary cells. These measurements demonstrate that, in the presence of the sensitizer, there occurs (i) an increase in total oxygen consumption during irradiation and (ii) a decrease in oxygen consumption following irradiation. These results are attributed to the effects of photosensitized oxygen consumption and decreased cellular respiration due to cell inactivation. Thus the inhibition of oxygen consumption after irradiation was shown to correlate with decreased cell survival measured in cell culture experiments. Possible mechanisms of inactivation and extensions of the oximetric approach are discussed.     

THE PHOTOCHEMISTRY OF 3-NITROBENZOYL AND 4-NITROBENZOYL AZIDES: POSSIBLE REAGENTS FOR PHOTOAFFINITY LABELING

Abstract— The photochemistry of 3- and 4-nitrobenzoyl azides was studied to examine their suitability for use in photolabeling procedures. These aroyl azides absorb light in the near UV-spectral region. Their irradiation leads both to photo-Curtius rearrangement with formation of the analogous isocyan-ates and to generation of nitrenes by loss of nitrogen. The chemical and spectroscopic properties of the aroyl nitrenes indicate that they are singlets in their ground states. These singlet nitrenes insert rapidly into unactivated, aliphatic carbon-hydrogen bonds, trap nucleophiles, and add to olefins to form aziridines with retention of stereochemistry. Photolabeling agents based on these azides may offer some advantages over conventional agents for the labeling of lipophilic sites.     

THE USE OF THE INTERNAL STANDARD IN REFINING THE UNIT CELL PARAMETERS OF SINGLE CRYSTALS

Journal of Structural Chemistry - The procedure of using the internal standard in refining unit cell parameters of single crystals is described. The study is carried out on single crystal Bruker...     

烯丙基保护基在多羟基化合物中的应用

烯丙基保护基由于能够在温和的反应条件下,在羟基、氨基等基团中引入和脱除,并且具有很好的区域选择性和化学选择性,因此被广泛应用于有机合成中。本文介绍了烯丙基保护基的发展历史,综述了烯丙保护基对多羟基化合物中不同羟基的选择性保护应用,并阐述了烯丙保护基的引入和脱除的机理及方法。     

偶氮胂-DCS作为钙镁络合滴定指示剂的研究及其应用

研究了利用光度显色剂偶氮胂－ＤＣＳ作为钙镁联合滴定指示剂的可能性及使用该指示剂滴定钙镁的准确性,与常用的钙镁络合滴定指示剂进行了比较,并成功地应用于水泥试样中钙镁含量测定。结果表明,偶氮胂－ＤＣＳ是一种良好的钙镁络合滴指示剂。     

SOME PROBLEMS IN THE ABSOLUTE MEASUREMENT OF GERMICIDAL ULTRAVIOLET RADIATION: THE USE OF ''PEN RAY'' LAMPS AS A CALIBRATION STANDARD

Abstract— Problems of absolute measurement of the dose rate of ultraviolet radiation of germicidal lamps in energetic units were studied. Irradiance at 254 nm generated by three different Pen Ray SC-1 low-pressure mercury lamps was measured independently in different laboratories using different instruments: (a) Westinghouse SM-600 Meter, (b) General Electric Germicidal Meter, and (c) large-surface thermopile with a Bäckström filter. These lamps were then used as secondary standards of absolute irradiance at wavelength 254 nm and compared with the Latarjet dosimeter and the International Light IL-254 Germicidal Photometer. Mutual agreement of calibration coefficients of three calibrated Pen Ray SC-1 lamps was roughly within ±5 per cent error. This calibration uncertainty indicates limits of the usefulness of Pen Ray lamps as standards. A direct radiometric calibration using an FT-16 Schwarz-Hilger vacuum thermopile, with interference filter NB-254 or UVR-250, was in agreement with the above comparison. On the basis of above radiometric calibration absolute D₃₇ lethal doses were determined equaling 2·7. J/m² for the bacteriophage T2 and 11·0 J/m² for the bacteriophage φX-174, the values being read from exponential survival curves.     

二甘醇二苯醚作为气相色谱固定液的研究

以二甘醇二苯醚作为气相色谱的固定液，对其色谱性能、分离效率作了初步研究，它对醇类、酯类、含氮类、芳香类化合物、氯代烃及高沸点有机混合物，其分离度大于1．5，分离效率达99．7％，分离效果好，显示了较高的柱效能和广谱特性。     

ARYL AZIDES AS PHOTOAFFINITY LABELS. A PHOTOCHEMICAL STUDY OF SOME 4-SUBSTITUTED ARYL AZIDES

Abstract— The photochemistry of a series of aryl azides used in, or closely related to some of those used in photoaffinity labeling studies, has been investigated by flash photolysis, continuous irradiation and low-temperature irradiation with IR and UV spectroscopic analysis. Results indicate that labeling preferentially takes place via singlet species. The reactive intermediates appear to be cycloazaheptatetraenes and/or the corresponding benzazirines. The reactive intermediates have τ_½ of the order of milliseconds or shorter and oxygen may act as a quencher, thereby interfering with the affinity labeling.     

THE EFEFCT OF MAGNETIC FIELD ON POLYMERIZATION OF STYRENE IN THE PRESENCE OF POLYVINYL NAPHTHALENE AND POLYVINYL BENZOPHENONE

In the process of bulk photopolymerization of styrene initiated by AIBN decomposition polyvinyl benzophenone (PVB) can supply an effective cage for triplet-triplet energy transfer between PVB macromolecules and small molecules of AIBN to influence the molecular weight of polystyrene in weak magnetic field (less than 0.035T), that was different from the case of polyvinyl naphthalene (PVN) which supplied cages for this system only in the stronger magnetic field (more than 0.2 T) studies. It was found that in the same conditions, PVN could exert more tremendous influences on the bulk photopolymerizatiou system of styrene than PVB because in the stronger magnetic field the triplet PVN had much longer life time than PVB.     

水溶性二苯酮类光敏剂中的羟基对光谱和光聚合的影响

有许多公司和研究所在从事光聚合的研究.本文报导两种不同分子结构的水溶性光敏剂的紫外光谱、磷光光谱和在不同溶剂中的量子产率,比较它们在水溶性预聚体中的光聚合效应. 两种水溶性光敏剂为:     

USE OF POTASSIUM IODIDE AS A CHEMICAL ACTINOMETER

Abstract Aqueous solutions of KI were examined for use as chemical actinometers to measure 254 nm (germicidal) radiation. Irradiation results in electron ejection from iodide such that aqueous electrons and iodine atoms are formed. In the presence of N₂0, an electron scavenger, recombination of these two reactive species is eliminated and stoi-chiometric formation of triiodide occurs. The absorbance increase due to triiodide was followed and the quantum yield determined using either a radiometer or ferrioxalate actinometry to estimate the amount of energy absorbed by the KI solution. The quantum yield ( ø ) at 25°C was determined to be 0.26 using radiometry, and 0.224–0.233 using actinometry, depending on the radiation conditions. The following expression was used to measure the incident Huence rate for 254 radiation at a given temperature T for irradiation over a time interval Δt (s) fluence rate (W/m²) = 4. 96 × 10⁶ΔOD(λ)/ e(λ) [0.23+0.004(T-25)] δ t cm⁻¹
At temperatures other than 25°C, the temperature dependence of the quantum yield (0.004/°C) is taken into account by the term shown in the denominator. Because KI remains relatively blind to longer wavelengths such as those found in normal room light, measurements can be made in the presence of room light without having to work in a darkened room.     

应用光亲和分子进行PEG的光照修饰及蛋白质的光照偶联

合成了一种光活性标记分子-对叠氮苯甲酸,将其偶联到具有双羟基的碳酸酯与乳酸的共聚物P (LA-co-DHP)上,获得了具有光反应活性的可生物降解共聚物P(LA-co-DAP),在光照条件下,可以将蛋白质方便快捷地共价偶联到P(LA-co-DHP)聚合物纤维上.在溶液中进行PEG与对叠氮苯甲酸的光照反应,通过核磁共振光谱...     

5—Br—PAN—S作指示剂配位滴定耐火材料原料中三氧化二铝

研究了以水溶性5-Br-PAN-S为指示剂,配位滴定耐火材料原料中的三氧化二铝的方法。试验结果表明,5-Br-PAN-S是一种EDTA配位滴定法的良好指示剂。     

A PRELIMINARY INVESTIGATION OF THE USE OF LIMONENE FOR DETECTING SINGLET OXYGEN AS A COMPONENT OF POLLUTED AIR

Abstract— Exploratory experiments using simulated conditions indicate that the terpene, (+)-limonene can serve to detect O₂(¹Δ_g) in polluted atmospheres. ¹O₂ attack on limonene results in the formation of specific oxidation products in a specific distribution; quantitative identification of these products is a highly specific test for ¹O₂.     

LIGHT-INDUCED LEAKAGE OF SPIN LABEL MARKER FROM LIPOSOMES IN THE PRESENCE OF PHOTOTOXIC PHENOTHIAZINES

Abstract— Liposomes prepared from dipalmitoyl lecithin, cholesterol and dicetyl phosphate and containing a trapped spin label marker were exposed to long wavelength UV light in the presence of a series of phenothiazine tranquilizers. EPR spectroscopy was used to detect spin label marker released from liposomes, taking advantage of the disappearance of line broadening from electron spin exchange which occurred on spin label release. The minimum effective phototoxic dose in mice of these phenothiazines was also determined. Kinetic studies of light-induced spin label release from phenothiazine-sensitized liposomes showed that membrane damage was rapidly induced and that the damaging species were short-lived. The damage process was oxygen dependent and could be temporarily prevented by cysteamine or α-tocopherol added immediately before irradiation. Only those phenothiazines which mediated light-dependent liposomal membrane damage had phototoxic activity in mice and the degree of photosensitization was parallel in the two systems. In both photosensitization phenomena, the nature of the substituent at the phenothiazine 2-position was more important than the phenothiazine side chain.     

QUANTUM EFFICIENCY FOR THE PHOTOCONVERSION OF BACTERIORHODOPSIN AT VERY LOW TEMPERATURES

Abstract— Quantum efficiencies for photoconversion of bacteriorhodopsin (trans-bR) to a bathochromic product (batho-bR^t) and its photoreversion in purple membrane at 77 and 9 K were investigated with low temperature spectrophotometry. The kinetics of the photoconversion and its photoreversion cannot be expressed by a single exponential curve. The photoconversions at 77 and 9 K showed the same slope in the early stage. The kinetics of the photoreversions were identical at the two temperatures. These results indicate that the quantum efficiencies for the conversion of trans-bR to batho-bR^t or for its photoreversion are identical at the two temperatures.
The fact that the photoreversion cannot be expressed by a single exponential curve suggests the existence of several conformational states of batho-bR^t due to the trimer structure of the purple membrane.