ESR自旋捕集技术的发展和在生物医学研究中的应用

ESR是研究自由基最直接和最有效的技术,但是这些自由基必须是相对稳定的,而且要达到一定浓度才能用ESR技术检测和研究. 而生物体系中产生的自由基大部分是不稳定的,这是常规ESR波谱仪无法检测的. 为了克服ESR技术的这一局限性,发展了自旋捕集技术,这是目前研究生物和医学体系中活泼自由基应用最多也是最成功的方法, 每年都有新的自旋捕集剂合成和大量在生物医学应用的报道,为自由基生物医学的研究和发展做出了巨大贡献.  作者建立和发展了捕捉超氧阴离子自由基、羟基自由基、脂质过氧化产生的脂类自由基、一氧化氮自由基和单线态氧及一氧化氮和氧自由基同时检测技术. 利用这些技术开展了细胞、组织产生自由基机理,天然抗氧化剂的筛选及一些重大疾病如炎症、心脏病、老年痴呆症、帕金森综合症,中风,辐射损伤,蛋白质氧化,植物光合系统中产生活性自由基和植物的发病机理研究中的应用,取得了一系列研究结果.     

用ESR研究缺血再灌注组织和多形核白细胞产生的NO自由基

总结了近年来我们实验室用低温顺磁共振(ESR)技术在生物体系,特别是在组织缺血再灌注和多形核白细胞中研究NO自由基的技术,波谱解析和所得到的结果.     

烷烃蒸汽微波放电分解反应活泼自由基的ESR研究

本文用前文^[5]报道的气相有机化合物微波放电装置、自旋捕捉剂苯亚甲基叔丁基氮氧化物PhCH=NC↑^O(CH₃)₃(PBR)与ESR相结合,系统研究了十种烷烃蒸汽微波放电分解产生的活泼自由基。从ESR谱的超精细结构及其随PBN与放电腔距离的变化,不仅证实了上述体系放电分解既有H·生成,又有碳中心自由基R·产生,而且还找到了PBN与R·H·生成加合物比值变化的规律。由PBN与放电腔距离的不同,得出的碳中心自由基R·加合物的ESR波谱参数,证明了PBN捕捉不同的自由基R·。     

多自由基体系溶液ESR谱的数据分析

将ER-200D ESR波谱仪ASPECT 3000计算机采集的ESR图谱传输到IBM PC/XT计算机上,通过自行设计的ESR图谱模拟程序ESRSIMU,同屏显示比较,方便、灵活地对多自由基体系溶液ESR一级近似谱进行了模拟,获取了各个自由基的ESR波谱参数和体系中各类自由基的相对浓度等信息.对单个自由基ESR谱的模拟,ESRSIMU的运行速度比BRUKER EPR 3002快近100倍.     

二苯甲酮与烃的光化夺氢反应中产生的活泼自由基的ESR研究

本文用具立体阻碍的自旋捕捉剂2,4,6-三-特丁基亚硝基苯(TBN)与ESR相结合的方法,研究了十种烷烃及四种芳烃与二苯甲酮光化夺氢反应中产生的活泼自由基。结果表明,R₃R₂R₁Ċ和TBN形成烷氧苯胺自由基加合物。R-ĊH-R'与TBN形成烷氧苯胺自由基加合物和氮氧自由基加合物。TBN可用于区分有、无空间位阻的烷烃,芳烃自由基。自旋加合物的ESR波谱可表明自由基中未偶电子所在碳原子附近的有效质子数目。     

卷烟烟气中活性自由基的自旋捕获ESR研究

卷烟烟气中的瞬态自由基对人体健康是相当有害的，而检测活性自由基最常用的方法是自旋捕获方法. 本研究中使用高效新型捕捉剂DEPMPO能够直接在水相中捕捉到烷基自由基与羟基自由基，并未发现烷氧自由基加合物. 考虑到DEPMPO对活性自由基的捕捉能力和加合物ESR谱图解析特征性都优于PBN与DMPO，且在有机溶剂中溶解较多的氧分子. 因此认为以往文献中所捕获的烷氧自由基来源于烷基自由基氧化后的次级自由基产物.     

110keV铁离子束辐照L(+)–半胱氨酸的分子改性研究

用110keVFe离子注入L(+)-半胱氨酸薄膜样品,然后通过傅里叶变换红外光谱、紫外-可见光谱和核磁共振氢谱分析表明,L(+)-半胱氨酸在接受Fe离子束辐照后受到了严重损伤,在样品中产生了新的分子基团.ESR波谱分析表明在辐照产物中存在一种长寿命自由基.ESI质谱分析进一步支持了低能离子束辐照所致生物分子改性的发生.     

TX-100胶束中光解蒽醌的TR-ESR研究

用时间分辨电子自旋共振(TRESR)波谱仪研究了光解蒽醌自由基的化学诱导动态电子极化(CIDEP).实验测量表明,在蒽醌/乙二醇/TX-100体系中,蒽半醌自由基AQH·有很强极化ESR信号,在激光激发后~0.8μs,检测到蒽醌负离子基AQ·^-的信号,表明AQH·自由基部分地离解为负离子基AQ·^-.     

吖啶与脂肪醇的光化夺氢反应活泼自由基的ESR研究

本文用自由基捕捉技术与ESR联用的方法,研究了吖啶与各种脂肪醇光化夺氢反应中产生的活泼自由基,并讨论了夺氢反应机理。     

茶多酚及其单体结构自由基的ESR研究

通过ESR检测方法研究了邻苯三酚、茶多酚及其四种单体的自氧化醌类自由基的ESR波谱信号,系统阐明了这几种物质的内在结构联系.     

Improved method to detect nitric oxide in biological systems

In this work an improved method is described for using organic solvent extracting to detect nitric oxide. The partition coefficients of the diethylthiocarbamate (DETC)-Fe²⁺ complex between different organic solvents and water, the signal intensity of the same NO trapping complex concentration in different organic solvents, and the extracting abilities of the organic solvents were determined. It was found that ethyl acetate was the optimal organic solvent. With ethyl acetate as extracting solvent, the (DETC)₂-Fe²⁺-NO complex was extracted from water phase to organic phase, and low concentration of nitric oxide in large volume could be detected by electron spin resonance (ESR) at room temperature. The ESR signal intensity had a good linear relationship with the concentration of nitric oxide, at a signal-to-noise ratio of 2. The detection threshold of this method was improved to lower than 50 nM, which was more sensitive than the usually used method. The (DETC)₂-Fe²⁺-NO complex was stable in the dark at 0–4°C, and there was little change after days. Nitric oxide produced by cardiomyocytes cultured in media and other biological systems was firstly detected with this method.     

ESR study of the nitric oxide production in tissues of animals under an external influence on the functioning of the cardiovascular and nervous systems

Electron spin resonance (ESR) of the ternary (DETC)₂-Fe²⁺-NO complex has been applied to determine the nitric oxide production in tissues of rats and snails. A preliminary ESR study of the NO content in tissues of rats before and after artificially induced acute myocardial infarct was performed. The analysis of the obtained results shows that the nitric oxide production during the first hour after the moment of inducing myocardial infarct decreases. It is also demonstrated that ESR may be useful in the study of the influence of the long-term sensitization of snails on the nitric oxide production in their body. The changes in the NO production after the external influences in both cases are discussed.     

Snapback应力引起的90 nm NMOSFET's栅氧化层损伤研究

实验结果发现突发击穿(snapback)，偏置下雪崩热空穴注入NMOSFET栅氧化层，产生界面态，同时空穴会陷落在氧化层中.由于栅氧化层很薄，陷落的空穴会与隧穿入氧化层中的电子复合形成大量中性电子陷阱，使得栅隧穿电流不断增大.这些氧化层电子陷阱俘获电子后带负电，引起阈值电压增大、亚阈值电流减小.关态漏泄漏电流的退化分两个阶段：第一阶段亚阈值电流是主要成分，第二阶段栅电流是主要成分.在预加热电子(HE)应力后，HE产生的界面陷阱在snapback应力期间可以屏蔽雪崩热空穴注入栅氧化层，使器件snapback开态和关态特性退化变小. 关键词： 突发击穿 软击穿 应力引起的泄漏电流 热电子应力     

GC-MS-gestützte Synthese und Qualitätskontrolle von 15N-markiertem Stickstoffmonoxid mit hoher 15N-Häufigkeit für umweltrelevante Tracer-Untersuchungen

Abstract

[¹⁵N]nitric oxide for investigations on the effect of nitric oxide in biological systems can be synthesized economically from the commercially available [¹⁵N]nitric acid. Difficulties arise, however, for the preparation of highly enriched ¹⁵N-labelled nitric oxide on a lower mmol scale. These difficulties are caused by oxygen and result in both, yield depression and insufficient long-time stability of the NO-containing gas mixture due to its oxidation to NO₂. Therefore, a sensitive in situ oxygen detection during synthesis was carried out by on line coupling of the synthesis apparatus with a GC-MS. Additionally, this equipment offers the advantage that both, conversion and ¹⁵N enrichment may be measured almost continuously and simultaneously. In this way, the yield can be increased up to 85% without increasing by-products. The gas mixture prepared from this [¹⁵N]nitric oxide showed an adequate stability for several months at a concentration of about 100 ppm(v) NO.     

Ein „F-Zentrum” als paramagnetischer Donator in Zinkoxid

The ESR line atg?1.96 in zinc oxide single crystals has been reinvestigated. It is found that the defect responsible for the resonance can be described as anF-center, i.e., an electron trapped in an oxygen vacancy. TheF-centers act as donors and due to overlapping of their wave functions give rise to donor band conduction. This model has been confirmed by studying various methods to produce the centers and by ESR and conductivity measurements down to liquid helium temperature.     

Effects of chinonin on nitric oxide free radical, myocardial damage and arrhythmia in ischemic-reperfusion injury in vivo

With sodium nitrite NaNO₂ as a standard source of nitric oxide, we compared the correlation coefficients obtained by three measuring methods used currently in the determination of the NOFe²⁺ (DETC)₂ complex (DETC, N,N-diethyldithiocarbamate) with that of the measuring method suggested in this study. The result showed that measuring the total height of triplet signals was the best linear correlation to the concentration of NO compared with other methods used in this system. With this method, we observed the effect of chinonin on the NOFe²⁺ (DETC)₂ complex in myocardial ischemic-reperfusion injury in vivo. The hearts of Wistar rats were subjected to 30 min of ischemia and 10 min of reperfusion in vivo. Different doses of chinonin (5, 10, 25, 50 mg/kg intraperitoneally) were administered to the ischemic-reperfusion rats. Chinonin increased the signal intensity of the NOFe²⁺ (DETC)₂ complex, inhibited the formation of thiobarbituric acid reaction substance and release of creatine kinase, and mitigated the incidence of ventricular arrhythmia in a dose-dependent way. Chinonin has cardiovascular protective effects by means of adjusting the level of NO and inhibiting oxygen free-radical-induced lipid peroxidation in myocardial ischemic-reperfusion injury in vivo.     

动态应力下超薄栅氧化层经时击穿的可靠性评价

实验发现动态电压应力条件下,由于栅氧化层很薄,高电平应力时间内隧穿入氧化层的电子与陷落在氧化层中的空穴复合产生中性电子陷阱,中性电子陷阱辅助电子隧穿.由于每个周期的高电平时间较短(远远低于电荷的复合时间),隧穿到氧化层的电子很少,同时低电平应力时间内一部分电荷退陷,形成的中性电子陷阱更少.随着应力时间的累积,中性电子陷阱达到某个临界值,栅氧化层突然击穿.高电平时形成的陷阱较少和低电平时一部分电荷退陷,使得器件的寿命提高. 关键词： 超薄栅氧化层 斜坡电压 经时击穿     

A Comparative ESR Study on Blood and Tissue Nitric Oxide Concentration during Renal Ischemia-Reperfusion Injury

Electron paramagnetic resonance (EPR) spin trapping technology is a sensitive and unambiguous method for detection of nitric oxide (NO). Due to the short lifetime, NO must be trapped before EPR measurement. There are two EPR spin trapping techniques used currently, including the detections of EPR signals of diethyldithiocarbamate-iron-nitric oxide (DETC₂-Fe²⁺-NO) and nitrosyl hemoglobin (HbNO). In this study, we firstly investigated the kinetics of the EPR signal of DETC₂-Fe²⁺-NO in normal and ischemia-reperfused kidneys. In normal rat kidneys, the signal of DETC₂-Fe²⁺-NO was found at 5 min after the spin trappers Fe²⁺/DETC were administrated, the peak concentration was at 15 min and the period with relatively stable signal intensity was at the time range from 15 to 70 min. In the ischemia-reperfused rat kidneys, the signal of DETC₂-Fe²⁺-NO was increased at 30 min of ischemia and decreased at 60 min of ischemia after the occlusion of renal artery (corresponding to the time course of 60 and 90 min after Fe²⁺/DETC injection respectively). We then investigated the EPR signal of HbNO in blood. No characteristic HbNO signal was found in the rats of the sham control and 30 min of ischemia. An HbNO signal occurred in the rats exposed to 60 min of ischemia and it became pronounced with increased duration of reperfusion. The signal intensity reached a plateau at 150 min of reperfusion. The results suggest that the DETC₂-Fe²⁺-NO signal can be only suitable for the NO measurement in the short-term ischemia-reperfusion model, whereas the HbNO signal can be applied to represent NO in the relatively long-term ischemia-reperfusion model. In addition, N^G-nitro-L-arginine (L-NAME) and allopurinol were used to identify the source of NO. By detecting the HbNO signal, we demonstrated that the activation of xanthine oxidase is an important source of NO formation at the long-term period of ischemia and reperfusion. Authors' address: Jiangang Shen, School of Chinese Medicine, University of Hong Kong, 10 Sassoon Road, Hong Kong SAR, China     

ESR spectroscopy applied to the study of radiation mechanisms

ESR spectroscopy is a powerful tool for detecting radicals or radical-ions trapped in solids, and in many cases “primary” electron-gain and-loss centres can be identified and their structures elucidated. Sometimes these primary centres are trapped in pairs, separated byca. 5 to 15 Å giving triplet state ESR spectra. The significance of pair-trapping in relation to current theories for the action of ionizing radiation will be discussed with reference to several recent examples. Generally, the primary centres are well separated and ESR does not give useful information about spurs or local regions of high radical densities. Reasons for trapping will be summarized with specific reference to DNA and proteins. The question of electron-return will be discussed, and several examples of this will be reported.