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1.
The structural elucidation of advanced glycation end-product (AGE)-modified proteins and quantitative analysis of free AGEs have been successfully performed, by use of mass spectrometry (MS) in plasma and tissues of patients with AGE-related diseases, such as diabetes mellitus, uremia, cataract, and liver cirrhosis. Matrix-assisted laser desorption/ionization (MALDI)-MS made it possible to directly analyze the AGE-modified proteins such as albumin and IgG. However, because the direct structural analysis of intact AGE-modified proteins is often not easy due to the formation of broad and poorly resolved peaks, peptide mapping after enzymatic hydrolysis was introduced into the analysis of AGE-modified proteins and the site-specific analysis of defined AGEs by MALDI-MS. Liquid chromatography/electrospray ionization mass spectrometry (LC/ESI-MS) has been employed not only for the structural elucidation of enzymatically hydrolyzed AGEs-modified peptides but also for simultaneous quantification of free AGEs in plasma and tissues of patients. Based on many studies that use MS for the analysis of AGEs, there is no doubt as to the important role of protein-linked AGEs in several diseases.  相似文献   

2.
Among post-translational modifications of proteins, non-enzymatic glycation is one of the less frequently studied by experts in proteomics. However, the relevance of protein glycation has been widely shown up in several pathological conditions. In fact, non-enzymatic glycation has been strongly related to hyperglycemic conditions and, thus, to chronic complications associated to diabetes mellitus and renal failure as well as degenerative changes occurring in the course of aging. Two different glycation levels are distinguished whether the structure of the protein is seriously damaged or not. The biochemical and clinical significance of both glycations have been already described. Several reasons have contributed to the lack of highly sensitive and selective methods for identification and quantitation of glycated proteins. These are mainly (1) the low concentration of glycated proteins in humans due to the low efficiency of the glycation process, (2) the modification of enzymatic digestion patterns, (3) the low ionization efficiency of glycated peptides, and (4) the lack of software including tools to identify this post-translational modification. The aim of this review is to provide the analytical guidelines required to succeed in the analysis of glycated proteins. For this purpose, different analytical approaches are considered to solve the main drawbacks derived from this gap in the proteomics field. Some challenges are finally proposed to be taken into account in future research.  相似文献   

3.
Mass spectrometry has been applied successfully to the study of non-enzymatic protein glycation, which is a topic of wide interest in the diabetes field. Low- and high-resolution mass spectra, GC/MS, and collisional activation spectroscopy allow the structural identification and quantitative evaluation of advanced glycation end-products, which represent important molecules for monitoring diabetes. More recently available techniques, such as ESI and MALDI/MS, have had a significant impact on analytical problems in diabetes. In particular, MALDI has been applied to the study of protein glycation in in vitro and in vivo conditions, because the number of glucose molecules that condense onto the protein can be easily determined by this approach. In the former case, glycation kinetics have been studied in various sugars and sugar concentrations, proteins, and buffer concentrations; in the latter, comparisons of MALDI spectra of circulating proteins from healthy and diabetic subjects determine the exposure of patients to high glucose levels. The method has been applied to an evaluation of the glycation level of immunoglobulins, and indicates that glycation takes place preferentially on the Fab fragment of the protein; data are relevant in relating immunological impairment with glycation-induced changes in the functionality of immunoglobulins.  相似文献   

4.
Age and diabetes have long been known to induce an oxidative reaction between glucose and collagen, leading to the accumulation of advanced glycation end-products (AGEs) cross-links in collagenous tissues. More recently, AGEs content has been related to loss of bone quality, independent of bone mass, and increased fracture risk with aging and diabetes. Loss of bone quality is mostly attributed to changes in material properties, structural organization, or cellular remodeling. Though all these factors play a role in bone fragility disease, some common recurring patterns can be found between diabetic and age-related bone fragility. The main pattern we will discuss in this viewpoint is the increase of fibrillar collagen stiffness and loss of collagen-induced plasticity with AGE accumulation. This study focused on recent related experimental studies and discusses the correlation between fluorescent AGEs content at the molecular and fibrillar scales, collagen deformation mechanisms at the nanoscale, and resistance to bone fracture at the macroscale.  相似文献   

5.
Electrochemistry (EC) is a mature scientific discipline aimed to study the movement of electrons in an oxidation–reduction reaction. EC covers techniques that use a measurement of potential, charge, or current to determine the concentration or the chemical reactivity of analytes. The electrical signal is directly converted into chemical information. For in‐depth characterization of complex electrochemical reactions involving the formation of diverse intermediates, products and byproducts, EC is usually combined with other analytical techniques, and particularly the hyphenation of EC with mass spectrometry (MS) has found broad applicability. The analysis of gases and volatile intermediates and products formed at electrode surfaces is enabled by differential electrochemical mass spectrometry (DEMS). In DEMS an electrochemical cell is sampled with a membrane interface for electron ionization (EI)‐MS. The chemical space amenable to EC/MS (i.e., bioorganic molecules including proteins, peptides, nucleic acids, and drugs) was significantly increased by employing electrospray ionization (ESI)‐MS. In the simplest setup, the EC of the ESI process is used to analytical advantage. A limitation of this approach is, however, its inability to precisely control the electrochemical potential at the emitter electrode. Thus, particularly for studying mechanistic aspects of electrochemical processes, the hyphenation of discrete electrochemical cells with ESI‐MS was found to be more appropriate. The analytical power of EC/ESI‐MS can further be increased by integrating liquid chromatography (LC) as an additional dimension of separation. Chromatographic separation was found to be particularly useful to reduce the complexity of the sample submitted either to the EC cell or to ESI‐MS. Thus, both EC/LC/ESI‐MS and LC/EC/ESI‐MS are common. © 2013 The Authors. Mass Spectrometry Reviews published by Wiley Periodicals, Inc. Mass Spec Rev 34:64–92, 2015  相似文献   

6.
7.
大气压电离液质联机的应用   总被引:3,自引:0,他引:3  
杨树民 《质谱学报》1998,19(1):66-80
本文以我们几年来的实际工作为素材,对电喷雾(ESI)和大气压化学电离(APCI)这一近年发展起来的软电离质谱接口技术做了应用性介绍。文中列举了该技术用于药物代谢、蛋白质构象以及热不稳定化合物分析的几个实例,对一些典型化合物在离子源内经碰撞诱导解离(CID)形成的质谱开裂产物进行了解析,对流量匹配、脱溶剂的技术特点、离子在溶液中的预形成、溶剂的选择、仪器的噪声等实践中遇到的问题做了较为详细的讨论。  相似文献   

8.
A complete biochemical understanding of the mechanisms by which hyperglycemia causes vascular functional and structural changes associated with the diabetic milieu still eludes us. In recent years, the numerous biochemical and metabolic pathways postulated to have a causal role in the pathogenesis of diabetic vascular disease have been distilled into several unifying hypotheses. These involve either increased reductive or oxidative stress to the cell, or the activation of numerous protein kinase pathways, particularly protein kinase C and mitogen-activated protein kinases. As detailed below, there is tremendous crosstalk between these competing hypotheses. We propose that increased tissue glucose levels alter cytosolic coenzyme balance by increased flux of glucose through the sorbitol pathway increasing free cytosolic NADH levels. Increased NADH levels can generate reactive oxygen species via numerous mechanisms, lead to the formation of intracellular advanced glycation end products, and induce growth factor expression via mechanisms involving protein kinase C activation. The elevation in growth factors, particularly vascular endothelial growth factor (VEGF), is responsible for the vascular dysfunction via numerous mechanisms reported here in detail.  相似文献   

9.
张建丽  王小兵 《质谱学报》2009,30(4):219-222
采用LC/MS和LC-MS/MS法同时检测保健品中非法添加的伐地那非。用乙酸乙酯提取,以10 mmol•L-1的甲酸铵(pH 3.5)和乙腈为流动相,分别用Agilent Zorbax SB C18(150 mm×2.1 mm×5 μm)和Agilent Zorbax SB C18(100 mm×2.1 mm×3.5 μm)色谱柱分离,采用电喷雾离子源,正离子扫描方式进行分析检测。该方法简便、快捷、可靠,适用于保健品中非法添加伐地那非的常规检测。  相似文献   

10.
BOTao  ZHANGZheng-xiang 《质谱学报》2009,30(Z1):133-133
Tandem quadrupole-time of flight(QTOF) mass spectrometry can provide the high accurate mass (<2×10-6) of parent and product ions for small and large molecules with excellent sensitivity and high mass resolution. Especially for the trace analysis in complex matrix(e.g. food, serum, fluid, environmental matrix), HPLC-QTOF plays an important role in identification and determination. In this study, a 250-illigal drug database based on accuracy mass was established, and a process for rapid screening and confirming of non-target illegal drugs was presented using urine and oral fluid preparation. The real samples were rapidly screened for illegal drugs using the molecular feature Extraction(MFE) algorithm of the Agilent Mass Hunter software, the established drug database, and accurate mass spectra using liquid-chromatography(LC), electrospray(ESI), quadrupole-time-of-flight mass spectrometry(Q-TOF LC/MS). The TOF data was processed by MFE, based on which the ions from noise and background were greatly removed. Using the MFE algorithm, a compound list containing interesting ions were contracted and the related chemical formula were calculated based on accuracy mass and isotope ratio calculation. Accurate masses of the ions detected and identified by MFE were compared to the exact masses of compounds in the database. Positive compounds were then further confirmed using the MS/MS(Q-TOF LC/MS) mode. Finally, the product ions from MS/MS mode are used for the determination of illegal drugs in the positive samples, and the methodology(linearity, repeatability and sensitivity) were investigated.  相似文献   

11.
The determination of disulfide bonds is an important aspect of gaining a comprehensive understanding of the chemical structure of a protein. The basic strategy for obtaining this information involves the identification of disulfide-linked peptides in digests of proteins and the characterization of their half-cystinyl peptide constituents. Tools for disulfide bond analysis have improved dramatically in the past two decades, especially in terms of speed and sensitivity. This improvement is largely due to the development of matrix-assisted laser desorption/ionization (MALDI) and electrospray ionization (ESI), and complementary analyzers with high resolution and accuracy. The process of pairing half-cystinyl peptides is now generally achieved by comparing masses of non-reduced and reduced aliquots of a digest of a protein that was proteolyzed with intact disulfide bonds. Pepsin has favorable properties for generating disulfide-linked peptides, including its acidic pH optimum, at which disulfide bond rearrangement is precluded and protein conformations are likely to be unfolded and accessible to cleavage, and broad substrate specificity. These properties potentiate cleavage between all half-cystine residues of the substrate protein. However, pepsin produces complex digests that contain overlapping peptides due to ragged cleavage. This complexity can produce very complex spectra and/or hamper the ionization of some constituent peptides. It may also be more difficult to compute which half-cystinyl sequences of the protein of interest are disulfide-linked in non-reduced peptic digests. This ambiguity is offset to some extent by sequence tags that may arise from ragged cleavages and aid sequence assignments. Problems associated with pepsin cleavage can be minimized by digestion in solvents that contain 50% H(2) (18)O. Resultant disulfide-linked peptides have distinct isotope profiles (combinations of isotope ratios and average mass increases) compared to the same peptides with only (16)O in their terminal carboxylates. Thus, it is possible to identify disulfide-linked peptides in digests and chromatographic fractions, using these mass-specific markers, and to rationalize mass changes upon reduction in terms of half-cystinyl sequences of the protein of interest. Some peptides may require additional cleavages due to their multiple disulfide bond contents and/or tandem mass spectrometry (MS/MS) to determine linkages. Interpretation of the MS/MS spectra of peptides with multiple disulfides in supplementary digests is also facilitated by the presence of (18)O in their terminal carboxylates.  相似文献   

12.
For interrogation of peptides with diverse modifications, no other instrument is as versatile as the Fourier-transform mass spectrometer (FTMS). Particularly using electrospray ionization (ESI), many intact proteins and their proteolytic products harboring post-translational and chemical modifications (PTMs) have been studied by high resolution tandem mass spectrometry (MS/MS). The widely touted analytical figures of merit for FTMS in fact have translated into clarity when analyzing PTMs from phosphorylations to disulfides, oxidations, methylations, acetylations, and even exotic PTMs found in the biosynthesis of antibiotics and other natural products. A top down approach to PTM detection and localization is proving extensible to an increasing variety of PTMs, some of which are stable to MS/MS at the protein level but unstable to amide bond cleavage by threshold dissociations at the level of small peptides <3 kDa. In contrast, MS/MS using electron capture dissociation (ECD) allows precise localization of even labile PTMs given enough sample and abundant molecular ions. Finally, this brief synopsis of recent literature highlights specific PTMs that perturb the protein backbone therefore altering MS/MS fragmentation patterns. Thus, FTMS will continue its expansion into more laboratories in part because of its ability to detect and deconvolute the regulatory mechanisms of biology written in the language of PTMs.  相似文献   

13.
液相色谱-电喷雾质谱联用技术分析人参皂苷   总被引:11,自引:4,他引:11  
应用液相色谱 -电喷雾质谱 ( L C-MS)联用技术 ,对 8种人参皂苷单体进行了探讨。发现在电喷雾负离子检测方式下 ,在离子阱和四极杆这两种不同质量分析器的质谱仪中 ,8种皂苷分子所产生的准分子离子峰均为 [M+ 45 ]-峰 ;并根据离子阱多级质谱扫描功能产生的碰撞诱导裂解 ( CID)谱 ,研究了该类化合物分子在电喷雾质谱、负离子检测方式下的裂解特征 ,有助于该类化合物的分子量确定和结构鉴定 ;同时建立了液相色谱 -质谱联用法 ,可用于该类化合物的定性分析  相似文献   

14.
液相色谱-质谱法检测肝脏中5种β-受体激动剂   总被引:1,自引:0,他引:1  
白凌  陈大舟  李蕾 《质谱学报》2008,29(1):10-12
建立了合理、可靠的液相色谱-质谱联用测定β-受体激动剂的方法。样品经5%高氯酸溶液提取,反相C18小柱净化,采用Agilent Zorbax Eclipse Plus C18柱分离,以0.1%乙酸水溶液(A)-乙腈(B)为流动相进行等度洗脱。通过液相色谱-质谱以MRM方式检测,测得5种 β-受体激动剂的检测限为0.4~1.2 μg•kg-1。对肝脏进行添标回收实验,平均回收率为77.2%~95.6%。  相似文献   

15.
肖晓萍  郭正光  成洁  孙伟 《质谱学报》2017,38(2):187-194
分别应用20、100和200 W超声功率对牛血清白蛋白以及人尿蛋白组的胶内条带进行胶内酶切,通过比较不同功率鉴定的多肽数和蛋白数,确定最优的超声功率。同时比较超声辅助酶切与传统过夜酶切方法,并分析超声辅助胶内酶切方法的特点。结果表明,超声功率为20 W时对胶粒破坏最小,样品损失最少,在牛血清白蛋白样品中鉴定的多肽数最多为65。在人尿蛋白组2个条带中,20 W方法鉴定的蛋白数分别为168和199,鉴定效果最优。通过对多肽误切率和不完全酶切多肽的定性和定量分析发现,超声酶切方法的误切率高于传统方法,但不完全酶切率较低。通过分析多肽误切位点发现,传统过夜酶切法和超声辅助酶切法的主要误切类型分别为脯氨酸(P)和天冬氨酸/谷氨酸(D/E),3种超声方法之间各种误切类型所占比例均无明显差别。超声辅助酶切可显著缩短蛋白质胶内酶切时间,其中20 W功率超声酶切的效果最佳。与传统过夜酶切法相比,超声辅助酶切多肽的鉴定结果更好,其多肽的误切率更高,不完全酶切率较低,可用于发现蛋白质组学研究。  相似文献   

16.
This article reviews recent literature on current methodologies based on chromatography coupled to mass spectrometry to analyze phenolic compounds with endocrine‐disrupting capabilities. For this review we chose alkylphenol ethoxylates, bisphenol A, bisphenol F, and their degradation products and halogenated derivatives, which are considered important environmental contaminants. Additionally, some related compounds such as bisphenol diglycidylethers were included. Growing attention has been paid to the mass spectrometric characterization of these compounds and the instrumentation and strategies used for their quantification and confirmation. The current use of gas chromatography–mass spectrometry (GC–MS) and liquid chromatography–mass spectrometry (LC–MS) methodologies with different mass spectrometers and ionization and monitoring modes is discussed. Practical aspects with regards to the use of these analytical techniques, such as derivatizing reagents in GC–MS, ion suppression in LC–MS, and the most problematic aspects of quantification, are included in the discussion. © 2009 Wiley Periodicals, Inc., Mass Spec Rev 29:776–805, 2010  相似文献   

17.
Macrolides are a group of antibiotics that have been widely used in human medical and veterinary practices. Analysis of macrolides and related compounds in food, biological, and environmental matrices continue to be the focus of scientists for the reasons of food safety, pharmacokinetic studies, and environmental concerns. This article presents an overview on the primary biological properties of macrolides and their associated analytical issues, including extraction, liquid chromatography-mass spectrometry (LC-MS), method validation, and measurement uncertainty. The main techniques that have been used to extract macrolides from various matrices are solid-phase extraction and liquid-liquid extraction. Conventional liquid chromatography (LC) with C18 columns plays a dominant role for the determination of macrolides, whereas ultra-performance liquid chromatography (UPLC) along with sub-2 microm particle C18 columns reduces run time and improves sensitivity. Mass spectrometry (MS), serving as a universal detection technique, has replaced ultraviolet (UV), fluorometric, and electrochemical detection for multi-macrolide analysis. The triple-quadrupole (QqQ), quadrupole ion trap (QIT), triple-quadrupole linear ion trap, time-of-flight (TOF), and quadrupole time-of-flight (QqTOF) mass spectrometers are current choices for the determination of macrolides, including quantification, confirmation, identification of their degradation products or metabolites, and structural elucidation. LC or UPLC coupled to a triple-quadrupole mass spectrometer operated in the multiple-reaction monitoring (MRM) mode (LC/MS/MS) is the first choice for quantification. UPLC-TOF or UPLC-QqTOF has been recognized as an emerging technique for accurate mass measurement and unequivocal identification of macrolides and their related compounds.  相似文献   

18.
用原位点击化学方法合成新型乙酰胆碱抑制剂   总被引:1,自引:0,他引:1  
采用高效液相色谱-质谱联用技术,分析加入叠氮和炔构建块的乙酰胆碱酯酶(AChE)孵育液。找到4个乙酰胆碱酯酶诱导叠氮和炔构建块发生Husigen[3+2]环加成反应的产物,然后通过常规化学合成及活性筛选证明该4种产物(syn-TZ2/QA5、syn-6TZ2/QA5、syn-8TZ2/QA5和syn-6TZ2/BA6)具有很强的乙酰胆碱酯酶抑制活性。  相似文献   

19.
原油及其馏分油,包括各种石油产品中所含元素对石油炼制、加工和产品质量有很大影响,快速准确测定其含量具有重要意义。文章对油品元素分析工作中样品的预处理方法和检测方法进行了综述,其中前处理方法包括萃取法、湿法消解法、干灰化法、压力溶弹法、微波消解法、有机直接进样法和乳液直接进样法,检测方法包括比色法、化学滴定法、X射线荧光光谱法(XRF)、原子吸收法(AAS)、电感耦合等离子体原子发射光谱法(ICP-OES)、电感耦合等离子体质谱法(ICP-MS),并对各方法的优缺点和适用范围做出了评价。  相似文献   

20.
建立测定人血浆中银杏内酯B的含量的LC-MS/MS方法。取正常人血浆50μL,加入2.5mol/L盐酸溶液10μL酸化,再加入内标2μg/mL银杏内酯C20μL,以1mL乙醚萃取后,取提取液500μL挥干溶剂,然后用30%的乙腈500μL溶解,取10μL进行LC-MS/MS测定。LC条件:色谱柱为ZorbaxC18柱(4.5*150mm,5μm);流动相:乙腈-水梯度洗脱;流速:0.6mL/min,柱温:35℃。质谱条件:ESI电离源;负离子模式;用于定量分析的离子对分别为m/z423.1→m/z367.2(银杏内酯B)和m/z439.1→383.2(内标:银杏内酯C)。该方法血浆中银杏内酯B的最低定量限为5ng,线性范围为5ng/mL~0.5μg/mL。本方法操作简便,灵敏度高,结果准确可靠,可用于该药物的含量测定,同时也可为临床药代动力学研究提供参考。  相似文献   

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