On-line CE-MS using pressurized liquid junction nanoflow electrospray interface and surface-coated capillaries

A simple and cost-effective laboratory-made liquid junction interface was used for coupling of CE with MS. In this device the capillary column and the spray tip were positioned in the electrode vessel containing appropriate spray liquid. The electrospray potential was applied on the electrode inside the liquid junction. A stable electrospray was produced at nanoliter per minute flow rates generated in the emitter tip without using an external pump. This arrangement provided high durability of the spray tip and independent optimization of the CE separation (use of coated capillaries) and ESI conditions. CE-MS analysis of mixtures of drugs, peptides, tryptic digests of proteins and biological fluids was optimized with respect to the effects of the distance between the separation capillary and electrospray tip and pressure applied on the liquid junction. The sensitivity of the system, in terms of the LOD (base peak monitoring) was below 10 ng/mL for the beta-blocker drugs and below 200 ng/mL for peptide analysis.     

Micro-flow injection analysis system: on-chip sample preconcentration, injection and delivery using coupled monolithic electroosmotic pumps

A micro-fluidic chip, within which two monolithic electroosmotic pumps are utilised for sample preconcentration, injection and delivery is presented. The monolithic pumps were capable of producing stable and bubble free flow rates at applied voltages below 2 kV, with a current <10 microA. Electrokinetic (EK) sample injection, down to low nano-litre volumes, was quantitatively controlled through applied voltage and injection times, whilst the sample pump delivered a carrier solution to indirectly dispense the sample. A nano-flow sensor (NFS) was used to continuously monitor the flow rate stability of each pump, showing response times of <5-10 s for changes in applied voltage. A capacitively coupled contactless conductivity detector (C(4)D), as an off-chip on-capillary detector, was used to complete the micro-flow injection analysis (FIA) system. A monolithic electroosmotic pump (EOP), modified with an anionic surfactant, was used to demonstrate a novel approach to on-chip cation preconcentration and elution.     

Characterization of the atmospheric pressure ionization mass spectrometric process obtained using a fused-silica emitter with the high voltage applied upstream

The atmospheric pressure ionization process obtained when a mixture of methanol and water (90:10, v/v) also containing 50 microM sodium hydroxide is dispersed from a fused-silica emitter was studied. A combination of a high electric field and a nebulizer gas with the high voltage applied upstream in the liquid flow was utilized to facilitate the spray process. By comparing the dependences of the spray current and ion signals on the spray potential, it was found that electrical corona discharges were obtained for potentials higher than about 2.6 kV, which resulted in a mixed electrospray and chemical ionization process. By introducing vapour from a solvent, such as benzene or toluene, with a low ionization energy into the nebulizing gas, it was found that the appearance of the corresponding molecular ion was correlated with a change in the slope of the spray current-potential curve. This indicates that the breakpoints in the spray current-potential curves observed were correlated with the onsets of corona discharges. It was shown that the mixed ionization process gives rise to increased amounts of protonated solvent molecules and assists in the formation of sodiated adduct ions from an uncharged fatty acid methyl ester.     

Selection of the optimum electrospray voltage for gradient elution LC-MS measurements

Changes in liquid composition during gradient elution liquid chromatography (LC) coupled to mass spectrometry (MS) analyses affect the electrospray operation. To establish methodologies for judicious selection of the electrospray voltage, we monitored in real time the effect of the LC gradient on the spray current. The optimum range of the electrospray voltage decreased as the concentration of organic solvent in the eluent increased during reversed-phase LC analyses. These results and related observations provided the means to rationally select the voltage to ensure effective electrospray operation throughout gradient-elution LC separations. For analyses in which the electrospray was operated at constant voltage, a small run-to-run variation in the spray current was observed, indicating a changing electric field resulting from fouling or degradation of the emitter. Algorithms using feedback from spray current measurements that can maintain the electrospray voltage within the optimum operating range throughout gradient elution LC-MS were evaluated. The electrospray operation with voltage regulation and at a constant, judiciously selected voltage during gradient elution LC-MS measurements produced data with similar reproducibility.     

电渗泵中电渗流的控制

电渗泵是利用载流的电渗驱动原理，结合电色谱(EC)、毛细管电泳(CE)、液相色谱柱技术制作的输液微泵，是新颖的流体和样品输送技术。电渗泵中电渗流(EOF)控制方法与EC和CE等文献中的电渗流控制方法是相同的。本文对EC和CE等文献中有关EOF控制方法作了总结，并对电渗泵的研究现状和应用作一些前瞻分析。     

Comparison between different sheathless electrospray emitter configurations regarding the performance of nanoscale liquid chromatography-time-of-flight mass spectrometry analysis

Four different sheathless electrospray ionization (ESI) configurations were investigated for a nano liquid chromatography (LC) system. The studied configurations were: a column with an integrated emitter, with the ESI potential applied before or after the column, and a column with separate emitter, with the ESI voltage applied at a union before the emitter or at the emitter tip. The results indicates that the efficiency of the LC system is rather independent of the configuration when using 95 microm i.d. columns, acetic mobile phase and standard peptides as a sample. Introduction of post column dead volume seems not to be a critical issue at least with flow rates down to 600 nl/min.     

A multilayer poly(dimethylsiloxane) electrospray ionization emitter for sample injection and online mass spectrometric detection

An ESI emitter made of poly(dimethylsiloxane) interfaces on-chip sample preparation with MS detection. The unique multilayer design allows both the analyte and the spray solutions to reside on the device simultaneously in discrete microfluidic environments that are spatially separated by a polycarbonate track-etched, nanocapillary array membrane (NCAM). In direct spray mode, voltage is applied to the microchannel containing a spray solution delivered via a syringe pump. For injection, the spray potential is lowered and a voltage is applied that forward biases the membrane and permits the analyte to enter the spray channel. Once the injection is complete, the bias potential is switched off, and the spray voltage is increased to generate the ESI of the injected analyte plug. Consecutive injections of a 10 microM bovine insulin solution are reproducible and produce sample plugs with limited band broadening and high quality mass spectra. Peptide signals are observed following transport through the NCAM, even when the peptide is dissolved in solutions containing up to 20% seawater. The multilayer emitter shows great potential for performing multidimensional chemical manipulations on-chip, followed by direct ESI with negligible dead volume for online MS analysis.     

Pulsed dual electrospray ionization for In/In reactions

A pulsed dual electrospray ionization source has been developed to generate positive and negative ions for subsequent ion/ion reaction experiments. The two sprayers, typically a nano-electrospray emitter for analytes and an electrospray emitter for reagents, are positioned in a parallel fashion close to the sampling orifice of a triple quadrupole/linear ion trap tandem mass spectrometer (Sciex Q TRAP). The potentials applied to each sprayer are alternately pulsed so that ions of opposite polarity are generated separately in time. Ion/ion reactions take place after ions of each polarity are sequentially injected into a high-pressure linear ion trap, where axial trapping is effected by applying an auxiliary radio frequency voltage to the end lenses. The pulsed dual electrospray source allows optimization of each sprayer and can be readily coupled to any spray interface with no need for instrument modifications, provided the potentials required to transmit the ion polarity of interest can be alternated in synchrony with the emitter potentials. Ion/ion reaction examples such as charge reduction of multiply charged protein ions, charge inversion of peptides ions, and protein-protein complex formation are given to illustrate capabilities of the pulsed dual electrospray source in the study of gas-phase ion/ion chemistry.     

Optimization of a pressurized liquid junction nanoelectrospray interface between CE and MS for reliable proteomic analysis

A pressurized liquid junction nanoelectrospray interface was designed and optimized for reliable on-line CE-MS coupling. The system was constructed as an integrated device for highly sensitive and selective analyses of proteins and peptides with the separation and spray capillaries fixed in a pressurized spray liquid reservoir equipped with the electrode for connection of the electrospray potential. The electrode chamber on the injection side of the separation capillary and the spray liquid reservoir were pneumatically connected by a Teflon tube filled with pressurized nitrogen. This arrangement provided precisely counterbalanced pressures at the inlet and outlet of the separation capillary. The pressure control system was driven by an electrically operated valve and maintained the optimum flow rate for the electrospray stability. All parts of the interface being in contact with the CEBGE, spray liquid and/or sample were made of glass or Teflon. The use of these materials minimized the electrospray chemical noise often caused by plastic softeners or material degradation. During optimization, the transfer of the separated zones between the separation and electrospray capillaries was monitored by UV absorbance and contactless conductivity detectors placed at the outlet of the separation capillary and inlet of the electrospray tip, respectively. This arrangement allowed independent monitoring of the effects of pressure, CE voltage and geometry of the liquid junction on the spreading and dilution of the separated zones after passage through the interface.     

A Study of Electrospray Ionization Emitters with Differing Geometries with Respect to Flow Rate and Electrospray Voltage

The performance of several electrospray ionization emitters with different orifice inside diameters (i.d.s), geometries, and materials are compared. The sample solution is delivered by pressure driven flow, and the electrospray ionization voltage and flow rate are varied systematically for each emitter investigated, while the signal intensity of a standard is measured. The emitters investigated include a series of emitters with a tapered outside diameters (o.d.) and unaltered i.d.s, a series of emitters with tapered o.d.s and i.d.s, an emitter with a monolithic frit and a tapered o.d., and an emitter fabricated from polypropylene. The results show that for the externally etched emitters, signal was nearly independent of i.d. and better ion utilization was achieved at lower flow rates. Furthermore, emitters with a 50 μm i.d. and an etched o.d. produced about 1.5 times more signal than etched emitters with smaller i.d.s and about 3.5 times more signal than emitters with tapered inner and outer dimensions. Additionally, the work presented here has important implications for applications in which maximizing signal intensity and reducing frictional resistance to flow are necessary. Overall, the work provides an initial assessment of the critical parameters that contribute to maximizing the signal for electrospray ionization sources interfaced with pressure driven flows.     

Electrospray ionization source with a rear extractor

A new electrospray source design is introduced by having an extractor electrode placed at 1 to 2 mm behind the emitter tip. The extractor was integrated into the sprayer body as a single device. An insulating tube was used to isolate the emitter from the extractor and to deliver the sheath gas for the electrospray. The electric field strength at the emitter was primarily determined by the relative position and the potential between the needle and the extractor; therefore, the spraying condition was insusceptible to the change of sprayer position or orientation with respect to the ion sampling inlet. Such design allowed the use of much lower operating voltage and facilitated the optimization of sprayer position by keeping the electric field parameter constant. Using an emitter capillary of 150 and 310 μm in inner and outer diameters, strong ion signal could still be acquired with 2‐kV emitter potential even if the distance between the emitter and ion inlet was extended to >70 mm. Charge reduction of protein ions using 2 extractor‐based electrosprays of opposite emitter polarities was also demonstrated.     

Robust monolithic silica-based on-chip electro-osmotic micro-pump

A robust, compact, on-chip, electro-osmotic micro-pump (EOP) for micro-flow analysis, based on parallel, encased, 10 x 0.1 mm I.D. monolithic silica capillary columns has been developed. A 15 x 40 x 2 mm poly(methyl methacrylate) (PMMA) chip, containing a total of nine parallel EOP systems was fabricated, allowing the use of single, double or triple monolithic columns to produce increased flow as required. The monolithic silica was compatible with both aqueous and organic solvents without swelling or shrinking problems, with the triple column EOP capable of generating flow of up to 0.6 microL min(-1) under zero pressure load and over 0.1 microL min(-1) with an applied pressure of ca. 2.4 bar using an applied voltage of just 2 kV. Current generated at the 2 kV applied voltage for a 2 mM acetate buffer solution (pH 4.5) was under 4 microA, allowing stable, bubble-free flow. The developed triple column EOP was incorporated within a micro-fluidic chip (5.0 x 2.0 x 0.4 cm) integrated with a second single 10 x 0.1 mm column EOP, for combined sample injection and simple on-chip micro-flow analysis.     

A smart and portable micropump for stable liquid delivery

Precise and reliable liquid delivery is vital for microfluidic applications. Here, we illustrate the design, fabrication, characterization, and application of a portable, low cost, and robust micropump, which brings solution to stable liquid delivery in microfluidic environment. The pump is designed with three optional speeds of different pumping flow rates, and it can be simply actuated by spring‐driven mechanism. The different flow rates of the pump are realized via passive microvalves in a compact microfluidic chip, which is installed in the pump. Importantly, the membrane structures of the microvalves allow accurate liquid control, and stable flow rates can be achieved via a spring setup. The proposed pump is applied to continuously and stably infuse microbead suspension into an inertial microfluidic chip, and good particle focusing is realized in the spiral channel of the inertial microfluidic chip. The proposed portable, self‐powered, and cost‐efficient pump is crucial for microfluidic lab‐on‐a‐chip system integration, which may facilitate microfluidic application for precise liquid delivery, control, measurement, and analysis.     

A robust sheath-flow CE-MS interface for hyphenation with Orbitrap MS

The hyphenation of capillary electrophoresis with high-resolution mass spectrometry, such as Orbitrap MS, is of broad interest for the unambiguous and exceptionally sensitive identification of compounds. However, the coupling of these techniques requires a robust ionization interface that does not influence the stability of the separation voltage while coping with oxidation of the emitter tip at large ionization voltages. Herein, we present the design of a sheath-flow CE-ESI-MS interface which combines a robust and easy to operate set-up with high-resolution Orbitrap MS detection. The sheath liquid interface is equipped with a gold coated electrospray emitter which increases the stability and overall lifetime of the system. For the characterization of the interface, the spray stability and durability were investigated in dependence of the sheath-flow rate, electrospray voltage, and additional gold coating. The optimized conditions were applied to a separation of angiotensin II and neurotensin resulting in LODs of 2.4 and 3.5 ng/mL.     

Characteristics of probe electrospray generated from a solid needle

Probe electrospray ionization (PESI) has recently been developed, in which the electrospray was generated from a solid needle instead of by using a capillary. In this paper, the characteristics of probe electrospray ionization were studied based on the measurement of spray current, optical microscopy, and PESI mass spectrometry. In the experiment, the solid needle was moved up and down a vertical axis, and a small amount of sample was repeatedly loaded to the needle when the tip of the needle touched the surface of the liquid sample at the lowest position. After the application of high voltage, a liquid droplet was formed on the tip of the solid needle probe, with its size was determined by the size of the needle tip. The liquid flow rate to the tip, as indicated by the spray current, depends on the voltage applied to the needle as well as the loaded liquid amount. Stable electrospray can be maintained until the total consumption of liquid sample. The kilohertz current pulsation takes place in the case of overloading the sample to the needle. The influences of the applied voltage and the liquid flow rate on the PESI mass spectra were also examined.     

Current-controlled nanospray ionization mass spectrometry

The hypothesis that direct determination of electrospray current would provide a viable method for maintaining spray stability to enable optimal nanospray analysis was tested by building a feedback apparatus capable of reading the current and readjusting the emitter voltage in real time. The apparatus consists of a current-sensing circuit that reads the voltage drop across a resistor located between the high-voltage power supply and the nanospray emitter. A low voltage proportional to the observed current is generated and sent to a data acquisition card. The information is used by a proportional-derivative-integral (PID) algorithm to calculate the magnitude of a low-voltage signal that is used to control the power supply output. Any variation of current across the sensing resistor is thus counteracted by an opposite-direction variation of the high voltage applied to the nanospray emitter. In this way, the apparatus adjusts the emitter voltage to achieve a preset value of current, which it strives to maintain over time in spite of any possible variation of the parameters influencing the spray regime. Preliminary results have shown that the feedback apparatus is capable of establishing and maintaining stable spray for samples that are usually considered challenging in traditional voltage-controlled analysis, such as those consisting of nucleic acid solutions with high salt loads. For these types of samples, the total ion count recorded in current-controlled mode was significantly more stable than that observed in voltage-controlled mode. At the same time, overall signal intensities and signal-to-noise ratios were also significantly improved. Setting the target nanospray current to a predefined value and letting the apparatus reach the target without operator intervention enabled the acquisition of viable data from solutions containing up to 2. 5 M ammonium acetate, which are ordinarily difficult by traditional manual tuning. A deeper understanding of the current-voltage relationships for samples of very different compositions is expected to enable one not only to predict the target current that should be used for a certain analysis, but also to devise algorithms to change such target as a function of predictable variations of sample properties and analytical conditions. This will allow for optimal performance to be maintained during on-line gradient chromatography in which the nature of the sprayed solution may vary very widely during the course of the analysis.     

A sheath-flow nanospray interface for capillary electrophoresis/mass spectrometry

We have developed a novel sheath-flow interface for low-flow electrospray ionization mass spectrometry (ESI-MS) and capillary electrophoresis/electrospray mass spectrometry (CE/ESI-MS). The interface is composed of two capillaries. One is a tapered fused-silica ESI emitter suitable for microliter and nanoliter flow rate electrospray and the other is a tail-end gold-coated CE separation column that is inserted into the emitter. A sheath liquid is supplied between the column and the emitter capillaries. The gold coating and the sheath liquid are used as the conducting media for ESI and the CE circuit. This novel design was initially evaluated by an infusion ESI-MS analysis of the most common antiretroviral dideoxynucleosides, followed by CE/MS coupling analysis of several antidepressant drugs. With infusion studies, the effects of the sheath liquid and the sample flow rates on detection sensitivity and signal stability were investigated. For an emitter with an internal diameter of 30 microm, the optimum flow rates for the sheath and the sample were 200 and 300 nL/min, respectively. The main improvement of this approach in comparison with conventional sheath liquid approaches using an ionspray interface is the gain in sensitivity. Sensitivities were three times better for dideoxynucleosides analyzed by infusion and 12 times higher for antidepressant drugs analyzed by CE/MS with this interface compared with ionspray. The emitter is durable, disposable, and simple to fabricate.     

Rapid determination of aliphatic amines in water samples by pressure-assisted monolithic octadecylsilica capillary electrochromatography-mass spectrometry

A pressure-assisted capillary chromatography-mass spectrometry method based on the use of a monolithic octadecylsilica (ODS) capillary is proposed for the determination of aliphatic amines. A 25 mM citric acid buffer containing 10% methanol is used as running electrolyte. Separation is achieved by simultaneously applying a capillary electrophoresis (CE) voltage of 13 kV and an overimposed pressure of 8 bar. The use of pressure is required to ensure stable electrospray conditions. Analysis times are reduced by using a capillary column consisting of a 30 cm long monolithic silica capillary column bound with ODS and a fused-silica capillary column also 30 cm long. The proposed method was successfully applied to the determination of low-molecular-weight aliphatic amines in tap and river water. The analysis of real samples requires cleanup and preconcentration, which can be performed automatically by inserting a minicolumn in the replenishment system of the commercial instrument.     

Laser spray: electric field-assisted matrix-assisted laser desorption/ionization

A new liquid chromatography/mass spectrometry interface, the laser spray, has been developed. Explosive vaporization and mist formation occur when an aqueous solution effusing out from the tip of the stainless-steel capillary is irradiated from the opposite side of the capillary by a 10.6 microm infrared laser. Weak ion signals could be detected when the plume was sampled through the ion sampling orifice. When a high voltage (3-4 kV) was applied to the stainless-steel capillary, strong ion signals appeared. The ion abundances were found to be orders of magnitude greater than those obtained by conventional electrospray ionization in the case of aqueous solutions. The present method is regarded as an electric-field assisted form of matrix-assisted laser desorption/ionization in which the liquid chromatographic solvent (water, etc.) acts as a liquid matrix. Laser spray ionization is expected to become a versatile method for biological mass spectrometry because this method is compatible with the natural solvent, water.     

Atmospheric pressure ion lens extends the stable operational region of an electrospray ion source for capillary electrophoresis-mass spectrometry

An atmospheric ion lens incorporated into an electrospray ion source for capillary electrophoresis-mass spectrometry (CE-MS) is found to extend the stable operational regions for both flow rates and electrospray ionization (ESI) voltages. The stable operating conditions for the ESI source with and without the ion lens were characterized. The results showed that the stable operation region was widest when the voltage difference between the sprayer and the ion lens ranges from 2.6 to 2.8 kV, and under these condition, the CE-MS interface can be adapted to a broader range of electroosmotic and modifier flow rates. Modeling of the electric field in the electrospray ion source with the ion lens suggests that the extension of the stable region is attributed to the flatter equipotential surfaces around the sprayer tip and higher electric field strengths in the rest of the interface region.