辅助电极对纳升电喷雾影响的研究

电喷雾质谱作为一种有效的软电离技术易于和各种液相分离技术如高效液相色谱以及毛细管电泳等技术实现在线联用,已越来越广泛地用于生物大分子的分析鉴定,但常规电喷雾技术要求分析试剂流速一般为1～10μL／min,耗样量大．因而低样品消耗、高灵敏的纳升电喷雾技术的研究越来越受到重视。纳升电喷雾技术因其喷雾头口径较小,产生的泰勒锥体也相应减小,样品分子的去溶剂化和离子化效率大大提高,有文献报道,利用纳升电喷雾技术可检测到10^-18mol级蛋白样品。     

表面修饰探针纳升电喷雾质谱脂质组学对大型溞和蚤状溞的快速鉴别

采用表面修饰探针敞开式纳升电喷雾质谱技术建立了脂质组学方法, 以实现溞属(Daphnia)生物大型溞和蚤状溞的快速鉴别. 采用微尺度表面修饰探针直接从Daphnia体内萃取和富集出脂质化合物, 然后在常压敞开条件下进行纳升电喷雾质谱分析, 利用高分辨率质谱仪获得脂质指纹图谱并进行脂质种类和结构鉴定. 采用该方法获得了大型溞和蚤状溞的脂质指纹图谱, 并研究了大型溞和蚤状溞体内的脂质组成特征. 通过结合化学计量学手段实现了大型溞和蚤状溞的快速区分, 并鉴定了相关的生物标志物. 结果表明, 所建立的微萃取探针纳升电喷雾质谱脂质组学方法具有分析速度快、 灵敏度高的优点, 可用于Daphnia属生物的快速鉴定和识别.     

偶氮胂Ⅲ的电喷雾质谱行为

采用电喷雾质谱法研究了偶氮胂Ⅲ离子化过程. 结果发现,偶氮胂Ⅲ易于在电喷雾电离过程中通过去质子化作用形成带有单电荷或双电荷负离子. 考察了酸度和醋酸根等离子对偶氮胂Ⅲ电喷雾电离的影响. 结果表明,偶氮胂Ⅲ在弱酸性条件下有利于形成负离子. 提出了偶氮胂Ⅲ在电喷雾电离源中通过去质子化作用形成负离子的可能机理,并用串联质谱法研究了所形成不同价态离子的裂解规律. 结果表明,电喷雾串联质谱法能方便快捷地研究偶氮胂Ⅲ的电离行为和所形成不同价态离子的性质,其检测限达0.74×10 -15 g.     

苦参生物碱电喷雾质谱分析

用电喷雾离子阱质谱(ESI-ITMS)研究了苦参碱和氧化苦参碱一级质谱电离规律和二级质谱裂解规律。实验结果显示,苦参碱和氧化苦参碱电喷雾一级质谱中均易产生聚合及加合离子,二级质谱中易产生环逐步裂解碎片离子,即结构相似的苦参碱和氧化苦参碱具有相似的电喷雾质谱行为。苦参碱的特征碎片为m/z148和150,氧化苦参碱的特征碎片为m/z248([M H-17] )、m/z247([M H-18] )以及m/z148、m/z150。对苦参对照药材的甲醇提取物进行了电喷雾质谱分析,由一级质谱的准分子离子及其二级质谱碎裂信息对各已知组分进行了结构认证。     

新型纳升级毛细管刻蚀电喷雾质谱接口研究

对普通石英毛细管表面使用氢氟酸刻蚀技术进行刻蚀, 并与商品化鞘流液毛细管电泳-质谱接口(Sheathflow CE-MS interface)结合, 将其改装成一种新型的纳升级电喷雾质谱接口. 玻璃膜接口部分呈多孔结构, 壁厚约10 μm. 以细胞色素c对新型接口加以评价, 样品的流量最低可达到20 nL/min; 在50~500 nL/min流量范围内刻蚀接口具有较高的响应信号. 考察了接触电解质溶液对样品电离的影响; 比较微升级不锈钢接口和新型接口的蛋白质检测结果发现, 在流速为200 nL/min时, 检测灵敏度可以提高3.6倍.     

间歇电喷雾的初步研究

设计了一种高灵敏的质谱离子化方法-间歇电喷雾，并介绍了间歇电喷雾的基本原理、稳定性和应用在蛋白质分析上的优势。这种质谱离子化方法能够显著地改善信噪比，降低对样品溶解度的要求，提高样品的离子化效率，减少样品的消耗和仪器的污染，可以预言间歇电喷雾的方法在蛋白质的分析上将会有广泛的应用前景。     

电喷雾离子源原理与研究进展

电喷雾离子源(ESI)是蛋白质组学研究采用的LC-MS/MS中最常用的接口之一,其作为一种软电离技术,具有可直接测定热不稳定化合物、形成多电荷离子等特征,在蛋白质组学研究中具有独特优势.本文介绍了电喷雾离子源(ESI)的工作原理与研究进展,并对各种新型离子化方法与应用进行了系统评述.     

蛋白质非共价化合物的电喷雾质谱研究

介绍了近几年来利用电喷雾离子化质谱技术研究蛋白质的基本构象及蛋白质非共价化合物的基本化学信息的进展。许多研究表明,电喷雾质谱在研究蛋白质的复杂结构和功能方面有十分广阔的前景。文中列举了若干实例说明了电喷雾离子化质谱技术在分析蛋白质非共价化合物的应用和一般方法。     

合成多肽的电喷雾质谱研究

利用电喷雾多极串联质谱对3种合成多肽进行了系统的鉴定和分析研究。首先通过全扫描模式测定了其分子量，然后选择[M H]^ 或[M 2H]^2 离子通过串联质谱(MS／MS)得到碎片离子，采用y离子和b离子互补的方法测定了多肽序列。利用文献数据对这种方法进行了验证，实验结果表明，该方法简便、快速、实用。     

脱水淫羊藿素的电喷雾多级串联质谱研究

研究了脱水淫羊藿素的电喷雾质谱行为，结果表明：该化合物在正、负离子模式下，均可得到较好的电喷雾质谱信息，且在负离子模式下电喷雾质谱分析的灵敏度较高；运用电喷雾碰撞诱导解离技术，发现正离子和负离子模式下的裂解方式虽有相似之处但并不完全相同；分别阐明了该化合物在正、负离子模式下的电喷雾质谱碎裂规律。     

Polyaniline-coated nanoelectrospray emitters: performance characteristics in the negative ion mode

The increasing tendency to miniaturize analytical techniques has resulted in the widespread use of nanoelectrospray ionization mass spectrometry. A new polyaniline-coated nanoelectrospray emitter has shown increasing promise as a more durable and stable alternative to traditional metal-coated emitters. In this report, the utility of polyaniline-coated nanoelectrospray emitters in the negative ion mode is investigated. Here, oligonucleotides and peptides have been ionized in the negative mode using polyaniline-coated nanoelectrospray emitters. The emitters were found to be durable for at least an hour in the negative ion mode, during which time the signal was stable. The high amount of electrical discharge usually associated with negative ion mode nanoelectrospray was not problematic with the polyaniline-coated emitters. These characteristics make possible the reliable coupling of low-flow separations to negative ion nanoelectrospray without the worry of emitter failure during the course of the experiment.     

Native Mass Spectrometry from Common Buffers with Salts That Mimic the Extracellular Environment

Nonvolatile salts are essential for the structures and functions of many proteins and protein complexes but can severely degrade performance of native mass spectrometry by adducting to protein and protein complex ions, thereby reducing sensitivity and mass measuring accuracy. Small nanoelectrospray emitters are used to form protein and protein complex ions directly from high‐ionic‐strength (>150 mm ) nonvolatile buffers with salts that mimic the extracellular environment. Charge‐state distributions are not obtained for proteins and protein complexes from six commonly used nonvolatile buffers and ≥150 mm Na⁺ with conventionally sized nanoelectrospray emitter tips but are resolved with 0.5 μm tips. This method enables mass measurements of proteins and protein complexes directly from a variety of commonly used buffers with high concentrations of nonvolatile salts and eliminates the need to buffer exchange into volatile ammonium buffers traditionally used in native mass spectrometry.     

Silanization of inner surfaces of nanoelectrospray ionization emitters for reduced analyte adsorption

During the course of nanoelectrospray ionization (nanoESI) of substance P, an unusual type of signal reduction was observed with flow rates <10 nL/min. This reduction in signal appears to be induced by the adsorption of positively charged analytes onto negatively charged free silanol groups on the inner surface of emitters; analytes with higher pI values (such as substance P) exhibit greater tendency for adsorption. Support for this hypothesis is demonstrated by the decrease in signal reduction in the presence of concentrated salts or for emitters whose internal silanols have been covalently silanized. Emitters treated with hexamethyldisilazane or 3-aminopropyltriethoxysilane showed higher analyte signals for substance P than untreated emitters, suggesting a reduction of analyte adsorption onto the inner walls of silanized emitters. The efficacy of reduced peptide adsorption was demonstrated for emitters silanized with 3-aminopropyltriethoxysilane using a simple peptide mixture as well as a more complex peptide mixture (a tryptic digest of horse hemoglobin).     

A simple nanoelectrospray arrangement with controllable flowrate for mass analysis of submicroliter protein samples

A simple arrangement for nanoelectrospray ionization using a conventional syringe pump connected to a pulled unmodified capillary has been evaluated. This arrangement avoids several disadvantages associated with metal-coated nanoelectrospray emitters. The relatively large orifice (approximately 9 microns) at the pulled capillary tip reduces sample clogging and the use of the pump minimizes spray disruption due to gas bubbles. Subattomole detection limit was achieved with nanomolar protein sample solutions at 5-10 nL/min flowrates using an LCQ mass spectrometer. Submicroliter samples can be loaded from the tip orifice and stored inside the capillary to virtually eliminate any dead volume, and then be electrosprayed for extended periods at well-controlled flowrates.     

Two-dimensional microfabricated sources for nanoelectrospray

The idea of a novel two-dimensional (2D) nanoelectrospray ionization emitter tip with the shape of a nib is explored here. This novel planar design is studied as an alternative to the needle-like standard emitter tips that suffer from a lack of reproducibility and robustness and from an inherent incompatibility with high-throughput analysis. The composition of the micro-nib sources is analogous to the working of a simple fountain pen, with a liquid reservoir linked to a micro-nib tip from which the sample is electrosprayed via a capillary slot. The micro-nib prototypes described here were fabricated using microtechnology techniques and using the epoxy-based negative photoresist SU-8. The resulting free-standing micro-nib structure was supported by a silicon wafer. We present here two series of such micro-nib sources, the latter series exhibiting improved characteristics such as a 8 micro m source width of the nib tip. They were tested in mass spectrometry experiments on an ion trap mass spectrometer (LCQ Deca XP+, Thermo Finnigan) using standard peptide samples having concentrations down to 1 micro M and with a high voltage (HV) supply around 1 kV for the second series of micro-nib sources. In addition to the stability of the spray, the obtained mass spectra showed the reliability of these sources for peptide analysis; the signal of the spectra was as intense and the signal-to-noise ratio (S/N) as high as that obtained with the use of standard emitter tips.     

Needlelike bicrystalline GaN nanowires with excellent field emission properties

Large-yield and crystalline GaN nanowires have been synthesized on a Si substrate via a simple thermal evaporation process. The majority of the GaN nanowires has bicrystalline structures with a needlelike shape, a triangular prism morphology, and a uniform diameter of approximately 100 nm. Field-emission measurements show that the bicrystalline GaN nanowires with sharp tips have a lower turn-on field of approximately 7.5 V/microm and are good candidates for low-cost and large-area electron emitters. It is believed that the excellent filed emission property is attributed to the bicrystalline structure defects and sharp tips.     

Multiple electrosprays generated from a single polycarbonate microstructured fibre

Electrospray ionization (ESI) has been invaluable to the mass spectrometric detection of biomolecules, due largely to the sensitivity afforded by the ionization technique. Lower flow rates, e.g. in the nanoelectrospray regime, result in smaller initial electrosprayed droplets, leading to higher ionization efficiency and greater signal. One approach to improving sensitivity without lowering flow rate is to generate multiple electrosprays (MESs) from the same sample, essentially splitting one larger flow into smaller flows in the nanoESI regime. Presented here is a series of novel MES emitters in the form of polycarbonate fibres. Based on microstructured fibre (MSF) technology whereby a set of homogeneous parallel channels are formed in a heat‐drawn fibre intended to conduct light, a custom design was fabricated in which 3, 6, 9 and 12 holes were arranged in a radial pattern to prevent inhomogeneities in the electric field. The MSFs have dimensions that are compatible with current standards in nanoESI equipment, and the tip is more compatible with standard MS orifices than other larger multielectrospray emitters. By measuring the spray current provided by the various emitters under the same solvent/voltage/total flow rate conditions, a plot was obtained clearly demonstrating the expected dependence on the square root of the number of holes, i.e. the number of independent electrosprays. With this firm proof of principle using this design/format, further effort is justified in developing similar emitters in alternative materials that better prevent surface wetting and allow greater hole density, ultimately leading to greater signal enhancement. Copyright © 2012 John Wiley & Sons, Ltd.     

Atmospheric molded poly(methylmethacrylate) microchip emitters for sheathless electrospray

Disposable poly(methylmethacrylate) (PMMA) sheathless electrospray microchip emitters were prepared for the first time using the atmospheric molding fabrication protocol. A sheathless electrospray from uncoated channel outlets, machined to cone-shaped three-dimensional tips, is demonstrated utilizing a simple cross design with an on-chip liquid junction to obviate the need for external unions to voltage electrodes, thus reducing the dead volume effects as well as the complexity of fabrication. The fast replication of microchip emitters was performed by molding prepolymeric methylmethacrylate solutions into silicon-master/aluminum-spacer/glass-plate molds followed by UV-initiated free radical polymerization. The performance of the new microchip emitters was demonstrated for mass spectral measurements of methionine enkephalin, adrenocorticotropic hormone and insulin peptide/protein mixtures. The samples were infused through capillary connections using hydrodynamic pumping. The polymeric emitters prepared by this flexible fabrication route offer an easy way of operation and high stability, without a need for attachment of external voltage unions or metallizing the emitter tips. The new approach should provide a useful low-cost tool for widespread coupling of mass spectrometry to chip systems.     

微纳加工技术在超微电极制备中的应用

从电极材料、绝缘材料、薄膜的图形化等方面,评述了微纳加工技术在单超微电极和超微电极阵列制备中的应用。在单超微电极的制备中,随着微纳加工技术的引入,可以实现具有规则几何形状和微小电极尖端的单超微电极的重复性制备。     

Characterization of micromachined hollow tips for two-dimensional nanoelectrospray mass spectrometry

In this work an improved design of chip-based nanoelectrospray nozzles is reported. Two-dimensional matrices of out-of-plane 10 microm i.d. silicon dioxide tips with a tapered shape were manufactured using deep reactive ion etching technology. Using a peptide sample, six micromachined tips and six commercially pulled silica capillary tips were compared employing an ion trap mass spectrometer. At a flow rate of 100 nL/min, the detectability obtained was approximately the same for the two types of tips. The relative standard deviation of the signal-to-noise ratio for the peptides between six different tips was on average 22% for the micromachined tips and 45% for the pulled capillary tips. The usefulness of the micromachined tips for analysis of non-covalent protein-ligand complexes was demonstrated by the analysis of a sample of RNase A and cytidine 2'-monophosphate. In another test, analyzing a tryptic digest of 1 pmol/microL cytochrome C, 18 peptides corresponding to a 82% sequence coverage were detected. Using MS/MS, the whole sequence of an 11 amino acid cytochrome C fragment was obtained. Computer simulations were performed on the shape and magnitude of the electrical field around micromachined and pulled capillary tips. To reach the threshold electric field density at the tip apex required to initiate an electrospray, a higher electrospray voltage was needed for the chip-based tips compared with pulled capillary tips. This is due to the influence of the chip base.