哌嗪乙氧基雌酮的电子电离质谱和快原子轰击电离(－)质谱测定及裂解途径

哌嗪乙氧基雌酮是一种可望用于防治骨质疏松症的全新合成的雌激素类化合物，本文采用EI-MS、FAB (－) -MS 、ESI-MS三种质谱技术分别对其结构和裂解途径进行了研究。采用FAB(－) -MS，获得m/z 404［M+Na-H］^-、382［M］^-、381［M-H］^-质谱峰；采用EI-MS, 获得m/z 382 M^+•；采用ESI-MS, 获得m/z 383［M+H］⁺。分子离子m/z 382 M^+•和主要特征子离子与其结构特征相符，并解释了其中的主要特征子离子，对其结构和裂解途径进行了确证。     

水产品中碱性嫩黄O残留量的液相色谱-串联质谱测定

建立了水产品中碱性嫩黄O工业染料残留的高效液相色谱 电喷雾串联质谱测定方法。样品经甲醇超声提取、旋转蒸发浓缩、过滤后,在正离子电喷雾离子源(ESI+),多反应监测(MRM)模式下进行测定。方法线性范围为2.5~100 μg•L^-1,在添加水平为5.0~20.0 μg•kg^-1时的平均回收率为78.9%~92.1%,相对标准偏差小于11%,定量限为0.5 μg•kg^-1。     

质谱多反应监测技术（MRM）及其在蛋白质细胞器定位与定量研究中的应用

肝脏中的蛋白质只有处于特定的亚细胞中才能发挥其功能。研究特定蛋白质的亚细胞定位对于肝脏功能研究有重要意义[1]。对于多定位的蛋白质,确定其在不同细胞器中量的变化,也是肝脏蛋白表达谱研究的重要内容之一。目前,蛋白亚细胞定位分析方法主要包括实验方法和理论预测方法两大类。其中实验方法主要有绿色荧光蛋白（GFP）-激光共聚焦显微镜法、免疫电镜胶体金标记法、差速离心-免疫印迹法等。理论预测方法主要根据蛋白质的氨基酸组成、氨基酸对组成以及位置特异性打分矩阵等分类特征,采用模糊k近邻、支持向量机及分组重量编码法等方法进行理论预测。实验方法不仅分析通量低,还需要特异的抗体,并且在定位的同时难以实现定量分析。对于不同的理论预测方法而言,往往会得出不同的结论,准确度差。因而需要建立一种可以实现通量化、准确地进行蛋白亚细胞定位及定量分析的方法。 近年来,质谱多反应监测技术（MRM）在蛋白质定量分析方面的应用越来越广泛^[2]。该技术通过特异性的检测预先设置的母离子和子离子,在母离子和子离子两个水平排除其它离子的检测干扰, 从而增强检测的特异性。同时结合同位素稀释技术可以同时实现对多种目标蛋白质的绝对定量。由于该技术采用特异性检测预设的母子离子对的方法,从而保证了检测的重现性。而依据特定子离子和相应的同位素标记子离子对峰面积进行绝对定量的方法保证了定量的可靠性。同时可以批量化同时进行多个蛋白质的亚细胞定量及定位分析。因而本研究利用MRM技术,用于目标蛋白在肝脏细胞器中定位及定量分析研究。 鉴于本研究样品采用蔗糖密度梯度离心法制备,首先对细胞器样品的分离效果进行评价。选用文献报道^[3]的正向和反向免疫评价抗体蛋白,包括线粒体Marker蛋白COX4、VDAC和Cytc,细胞核Marker蛋白Lamin B,质膜Marker蛋白Na＋K＋-ATPase等。利用MRM技术分析蛋白在不同细胞器样品中的含量,从而对细胞器样品纯度进行评价。同时考察方法的灵敏度,重现性以及动态范围。 在确定了细胞器样品纯度的基础上,利用MRM质谱技术对选定的部分目标蛋白进行了定位分析。根据这些蛋白在不同细胞器样品中的含量,判断其在不同细胞器样品中的定位。并选择其中一些蛋白进行GFP-激光共聚焦显微镜分析,与基于MRM技术的定位分析结果进行对比验证。     

EGC/GC和EGCG/GCG的ESI-IT-TOF质谱裂解规律研究

利用离子阱飞行时间质谱仪的高质量准确度、高分辨率、多级测定的性能，对表没食子儿茶素/没食子儿茶素(EGC/GC)和表没食子儿茶素没食子酸酯/没食子儿茶素没食子酸酯(EGCG/GCG)（二组对映异构体）的质谱裂解规律进行了研究，并利用氢/氘交换的方法对其裂解方式进行了确证。研究发现，儿茶素对映异构体间具有相同的质谱裂解途径，即使在多级质谱中也无明显区别，仅碎片离子的相对丰度存在差异。二级质谱中， EGC/GC在B环丢失CO₂，且其A, B环都可失去C₂H₂O。^1,4A^-、^1,3A^-、^1,2A^- 和[M-H-B环]^-四个碎片离子是EGC/GC的特征性离子，通过此四离子质荷比的变化，可推测A环上的取代情况。因EGCG/GCG的结构上都含有没食子酸的取代基，在二级质谱中均可见m/z 169的特征性离子峰，此离子可用于EGCG/GCG和EGC/GC的区分。     

苯环壬酯及其代谢物的电子轰击和电喷雾电离质谱分析

采用电子轰击质谱（EI-MS）、电喷雾电离质谱（ESI-MS）两种技术分别对苯环壬酯结构和裂解途径进行了研究。采用ESI-MS获得m/z 358［M+H］⁺、156、342等质谱峰，并对大鼠尿中的代谢产物结构进行了确证；采用EI-MS获得m/z 357［M］^＋、138、175、154、215等质谱峰，并解释了其中的主要特征碎片离子。     

LC-MS/MS法测定人全血中环孢素A浓度及环孢素眼用乳剂健康人体药代动力学研究

建立液相色谱 质谱联用法测定人全血中环孢素A浓度,选用Kromasil-C₁₈色谱柱（20 mm×4.6 mm×5 μm）,以甲醇1 mmol•L^-1甲酸铵溶液为流动相,采用梯度洗脱进行分离,样品用沉淀蛋白法处理后进样,流速1.1 mL•min^-1,柱温60 ℃,进样量20 μL。选用3200 QTrap型三重四极杆串联质谱仪的多重反应监测（MRM）扫描方式进行检测。环孢素A的线性范围为0.2~20.00 μg•L^-1,定量下限为0.2 μg•L^-1。准确度与精密度结果显示,方法日间、日内变异均小于15%,相对偏差为-12.60%~12.80%,方法提取回收率为（98.1±4.7）%,稳定性较好。所建立的方法快速、灵敏、专属性强、重现性好,可用于环孢素眼用乳剂健康人体药代动力学研究。     

基于离子/中性复合物中间体介导的气相离子反应研究

离子/中性复合物（Ion-neutral complex,INC）是单分子气相反应的重要中间体。近20年来,各国科学家已经相继报道了多类离子/中性复合物参与的质谱裂解反应,包括质子转移反应、阳离子转移反应以及多种重排反应等^[1-4]。 在质子化芳基醚、苯磺酰胺等几类化合物的多级质谱中,观察到一系列以离子/中性复合物为中间体的气相离子裂解反应,包括一类新的电子转移反应^[5],这些裂解反应形成产物离子的途径、以及产物离子的丰度均与取代基效应有关。我们通过理论计算和氘代实验系统研究了这些反应。     

高效液相色谱-串联质谱法在他克莫司临床血药浓度监测中的应用

建立高效液相色谱-质谱联用法监测人全血中他克莫司浓度。选用Shim-pack VP-ODS色谱柱，以甲醇10 mmol•L^-1乙酸铵为流动相，采用梯度洗脱进行分离，样品用乙腈进行蛋白沉淀后进样，用3200Qtrap型质谱仪的多重反应监测（MRM）扫描方式进行检测。他克莫司的线性范围为0.5~50.0 μg•L^-1，定量下限和最低检测限分别为0.5、0.1 μg•L^-1。准确度与精密度结果显示方法日间、日内变异均小于6.8%，相对偏差为-2.99%~2.23%，方法提取回收率均接近100.0%，稳定性较好。该方法快速、灵敏、专属性强、重现性高，适用于他克莫司治疗药物监测（TDM）工作。     

舒它西林的电喷雾-四极杆-飞行时间串联质谱裂解规律研究

为阐明药物舒它西林（氨苄西林/舒巴坦）的质谱裂解规律,采用电喷雾-四极杆-飞行时间串联质谱(ESI-Q-TOF-MS/MS)法在正离子模式下,对其离子碎片结构进行分析。通过质谱的高分辨能力及同位素氘代信息推断出舒它西林[M+H]⁺碎片离子断裂途径主要为氨苄西林部分的β-内酰胺环断裂,[M+Na]⁺裂解可观察到较多碎片离子峰,包括氨苄西林部分β-内酰胺环断裂、双酯连接处断裂和舒巴坦部分四元环断裂,此外有脱去CO₂、SO₂、CO等中性分子的峰出现。两种分析模式可为舒它西林的结构解析、体内代谢物质或环境中降解产物的研究提供快速鉴别理论依据。     

超高效液相色谱-串联质谱法检测动物源食品中8种氨基糖苷类药物残留

建立了动物源食品中大观霉素,链霉素,双氢链霉素,阿米卡星,卡那霉素,安普霉素,庆大霉素和新霉素等8种氨基糖苷类药物残留检测的超高效液相色谱 串联质谱（UPLC-MS/MS）方法。结果表明：8种氨基糖苷类药物在0.05～2 mg•L^-1浓度范围内呈现良好的线性关系,相关系数R²均大于0.998;方法检测限为5 μg•kg^-1,定量限为10 μg•kg^-1;从100、200和300 μg•kg^-1 3个添加浓度检测结果可以看出,方法平均回收率为83.0%～114.1％,批内RSD为1.1%～11.4％,批间RSD为2.1%～8.9％,可以满足食品安全有关法规的要求。     

质谱在生物标志物研究中的应用

生物标志物是一个指示剂,可以对其进行测定并用以解释正常生理过程、病理过程及治疗药物的药理学效应,广泛的应用于药物研发和临床诊断中。生物标志物的研究涉及多种技术平台和方法,其中基于质谱的研究方法无论在生物标志物的发现还是确证过程中都有着广泛的应用。本文讨论了基于质谱的研究方法在通常的生物标志物以及阿尔茨海默氏症研究中的进展。     

A review of current applications of mass spectrometry for biomarker/molecular tracer elucidation

Mass spectrometry, especially coupled with gas chromatography or tandem, is the analytical method of choice in elucidation of biomarker compounds present in organic mixtures extracted from geological, environmental, or biological samples. This review describes the biomarker concept; i.e., the precursor natural products to the geological/environmental derivatives, and their application as tracers in the geosphere and ambient environment. The mass spectrometric methods currently utilized for such analyses are reviewed, and typical examples of applications are described with a general key to the literature.     

An automated template‐based adaptive threshold approach for measuring ventricular volume enlargement in mouse brain MR microscopy

Mouse models for human diseases play an important role in developing therapeutic measures and understanding disease development and mechanisms. Ventricular enlargement is an objective and sensitive biomarker of neuropathological change associated with mild cognitive impairment (MCI) and Alzheimer's disease (AD). Thus, imaging based volumetric measures of mouse brain ventricle can be used as a biomarker for detecting and monitoring pathology. However, till now most mouse ventricular segmentation is still based on manual tracing, and current region of interests (ROIs) labelling approaches on mouse brain don't work well on small brain structure like ventricle. In this paper, an automated template‐based method was developed to evaluate for the identification of a ventricular ROIs in mouse brain imaging studies. Monte Carlo simulation was applied to evaluate the efficiency of this approach in detecting computer‐simulated ventricle in laboratory mice. The method demonstrated a satisfactory performance with consistent high correlations between the detected and simulated volume changes. This approach can be used to investigate the ventricular volume changing in transgenic mice and other putative animal models of AD.     

STRATEGIES AND CHALLENGES IN METHOD DEVELOPMENT AND VALIDATION FOR THE ABSOLUTE QUANTIFICATION OF ENDOGENOUS BIOMARKER METABOLITES USING LIQUID CHROMATOGRAPHY‐TANDEM MASS SPECTROMETRY

Metabolomics is a dynamically evolving field, with a major application in identifying biomarkers for drug development and personalized medicine. Numerous metabolomic studies have identified endogenous metabolites that, in principle, are eligible for translation to clinical practice. However, few metabolomic‐derived biomarker candidates have been qualified by regulatory bodies for clinical applications. Such interruption in the biomarker qualification process can be largely attributed to various reasons including inappropriate study design and inadequate data to support the clinical utility of the biomarkers. In addition, the lack of robust assays for the routine quantification of candidate biomarkers has been suggested as a potential bottleneck in the biomarker qualification process. In fact, the nature of the endogenous metabolites precludes the application of the current validation guidelines for bioanalytical methods. As a result, there have been individual efforts in modifying existing guidelines and/or developing alternative approaches to facilitate method validation. In this review, three main challenges for method development and validation for endogenous metabolites are discussed, namely matrix effects evaluation, alternative analyte‐free matrices, and the choice of internal standards (ISs). Some studies have modified the equations described by the European Medicines Agency for the evaluation of matrix effects. However, alternative strategies were also described; for instance, calibration curves can be generated in solvents and in biological samples and the slopes can be compared through ratios, relative standard deviation, or a modified Stufour suggested approaches while quantifying mainly endogenous metabolitesdent t‐test. ISs, on the contrary, are diverse; in which seven different possible types, used in metabolomics‐based studies, were identified in the literature. Each type has its advantages and limitations; however, isotope‐labeled ISs and ISs created through isotope derivatization show superior performance. Finally, alternative matrices have been described and tested during method development and validation for the quantification of endogenous entities. These alternatives are discussed in detail, highlighting their advantages and shortcomings. The goal of this review is to compare, apprise, and debate current knowledge and practices in order to aid researchers and clinical scientists in developing robust assays needed during the qualification process of candidate metabolite biomarkers. © 2019 John Wiley & Sons Ltd. Mass Spec Rev     

吐哈盆地中下侏罗统煤系地层源岩和油中饱和烃的GC/MS分析

Saturated hydracarbon of coal,carbonaceous mudstone and oils from the Lower Jurassic coal measures in the Turpan basin were studied,and biomarker characteristics and coal thermal maturity analyzed to draw the following conclusions. There are many similar biomarker characteristics between oil from middl-lower Jurassic of Turpan Basin and coal and carbonaceous mudstone in the same strata. They all contain specific r-lupane, I-norbietane, C24-tetracyclic and high content of C29-steranes. These characteristics suggest that they have similar matter source of the organic matter derived from matter with abundant high plants. Meanwhile, biomarkers often used to indicate depositional environments characterized by high Pr/Ph ratio, little or no gammacerane and high abundance dibenzofurans, such biomarker distributions are indicative of suboxic and freshwater environment. Although coal and carbonaceous mudstone remain in lower thermal maturity (Ro=0.47-0.53), but C29-ββ/(αα ββ) sterane ratio (0.294-0.489) and bezohopane are detected. Because these ferture are related to bacterial activity, bacterial degrdation of organic matter maybe take an important role in coal-derived oil.     

人尿液蛋白质N-糖肽富集和质谱鉴定的新方法研究

蛋白质的N-糖基化修饰参与了许多重要的生理过程,是研究多种重大疾病诊断标志物的热点。与其他体液活检样本相比,尿液可以无创、大量获取,并且不受稳态调节的影响,能够在一定程度上反映整个机体的生理和病理状态。因此,人尿液蛋白质N-糖基化的规模化研究对疾病诊断标记物的筛选和治疗靶点的发现均具有重大意义。由于人尿液中的N-糖蛋白含量有限,修饰比例较低,在质谱分析时N-糖肽易被高丰度的非糖肽所掩盖,难以鉴定。因此,发展高效、高选择性的富集材料是实现尿蛋白N-糖基化深度覆盖的必要前提。本研究利用巯基-烯点击化学反应,使用［3-(甲基丙烯酰氨基)丙基］二甲基(3-硫代丙基)氢氧化铵内盐（SPP）制备了两性离子修饰亲水硅胶材料（SPP-SiO₂）。该材料成功地应用于标准糖蛋白和健康人尿蛋白N-糖肽的富集和质谱检测,共鉴定了包含1 065个位点的633个尿液糖蛋白,比文献报道的鉴定规模提高了12.2%,证明了该亲水材料在尿液糖蛋白质组研究中的应用潜力。     

Glycoprotein analysis using protein microarrays and mass spectrometry

Protein glycosylation plays an important role in a multitude of biological processes such as cell–cell recognition, growth, differentiation, and cell death. It has been shown that specific glycosylation changes are key in disease progression and can have diagnostic value for a variety of disease types such as cancer and inflammation. The complexity of carbohydrate structures and their derivatives makes their study a real challenge. Improving the isolation, separation, and characterization of carbohydrates and their glycoproteins is a subject of increasing scientific interest. With the development of new stationary phases and molecules that have affinity properties for glycoproteins, the isolation and separation of these compounds have advanced significantly. In addition to detection with mass spectrometry, the microarray platform has become an essential tool to characterize glycan structure and to study glycosylation‐related biological interactions, by using probes as a means to interrogate the spotted or captured glycosylated molecules on the arrays. Furthermore, the high‐throughput and reproducible nature of microarray platforms have been highlighted by its extensive applications in the field of biomarker validation, where a large number of samples must be analyzed multiple times. This review covers a brief survey of the other experimental methodologies that are currently being developed and used to study glycosylation and emphasizes methodologies that involve the use of microarray platforms. This review describes recent advances in several options of microarray platforms used in glycoprotein analysis, including glycoprotein arrays, glycan arrays, lectin arrays, and antibody/lectin arrays. The translational use of these arrays in applications related to characterization of cells and biomarker discovery is also included. © 2010 Wiley Periodicals, Inc., Mass Spec Rev 29:830–844, 2010     

基于生物质谱的蛋白质组学绝对定量方法研究进展

蛋白质组学定量方法包括相对定量和绝对定量两部分。相对定量蛋白质组学（也称比较蛋白质组学）是指对不同生理病理状态下细胞、组织或体液蛋白质表达量的相对变化进行比较分析;绝对定量蛋白质组学是测定细胞、组织或体液蛋白质组中每种蛋白质的绝对量或浓度。目前蛋白质组学的绝对定量方法主要有基于内标法的蛋白质组学绝对定量方法和基于质谱数据统计分析的非标记方法。蛋白质组学定量的信息可以帮助了解蛋白质在相互作用网络中所起的作用;通过对临床生物标记物绝对量的分析,可以帮助直观地判断疾病的发生和发展过程,这对于临床诊断和疾病治疗都具有现实的指导意义。     

天然气MID／GC／MS检测及其应用

建立了天然气重烃富集及MID／GC／MS分析方法。从各类天然气中检测出甾萜烷、芳烃和金刚烷等一系列生物标志化合物,并进行了天然气生标的应用研究探索。分析应用结果表明本方法可作为天然气勘探研究的重要手段之一,故有重要的推广应用价值。