Low flow electrospray

Very low flow electrospray is called nanoelectrospray [26] where the samples are infused into the mass spectrometer at the nanoliter flow rate range. The infusion of a few microliters will result in a stable signal for more then 30 min, using pulled capillaries or chip-based emitters [27]. With infusion, signal averaging allows to improve the limit of detection in tandem mass spectrometry. Nanoelectrospray is particularly important in combination with nanoflow liquid chromatography or chip-based infusion for the analysis of peptides and proteins. 

For all of the above good reasons, few publications have come out on combinatorial mixture analysis in this review period. Mixture analysis of peptides by MS and MS/MS has continued as an active field [2,59,64], Nanospray (very low flow electrospray) has become the method of choice for studies of mixtures of peptides by MS/MS [65,66], 2D NMR has been used to assure that unusual amino acids were correctly incorporated in a mixed synthesis [59]. 

In principle, two types of electrospray interfaces must be distinguished. Most attention is paid to the high-flow devices applied for routine LC-MS. However, an extreme low flow electrospray needle device, i.e., the so-called nanoelectrospray, has been developed as well, especially by Mann in the mid-1990s. Nanoelectrospray is especially important in... 

Electrospray ionization mass spectrometry can be carried out over a wide range of flow rates. Technically, the term electrospray can be applied to any flow rate (it refers to the method of ionization), but additional terms have been coined to describe low-flow electrospray. Microspray describes electrospray conducted at intermediate flow rates ( 0.1-10pL/ min), and nanospray refers to very low flow electrospray (typically < lOOnL/min). 

A limitation with operation of ESTMS at high flow rates is the requirement that the spray capillary be placed far back from the inlet aperture, which means that only a fraction of the spray is transferred into the mass spectrometer, and the detection limit of the technique suffers. This limitation can be overcome with the use of microspray or nanospray, provided that smaller inner-diameter capillaries are employed. The smaller inner-diameter capillaries and lower flow rates employed with nanospray and microspray result in the production of smaller droplets, which allows the capillary to be placed closer to the inlet aperture than is possible with high-flow operation. Consequently, a larger fraction of the analyte may enter the mass spectrometer, and the mass detection limit may improve. Another advantage of low-flow electrospray is that it often eliminates the need for nebulizing gas. 

A number of investigators have observed that there are differences in performance of nanospray versus electrospray ionization. The very different conditions (temperature, capillary diameter, use of nebulizer gas) typically employed for conventional and low-flow electrospray make absolute comparisons of sensitivity at widely different flow rates difficult. The current perception, however, is that very low flow rate systems have greater mass sensitivity than higher-fiow-rate systems. 

The use of low flow rates allows the production of ions from mobile phases containing high percentages of water which, as noted previously, usually presents a particular problem because of its high surface tension and conductivity. The production of electrospray spectra from aqueous solution is possible at flow rates of less than 50 tLlmin ... 

In general terms, electrospray ionization is considered to be concentration-sensitive at low flow rates and mass-flow-sensitive at high flow rates, while APCI is considered to be mass-flow-sensitive. Low and high are both subjective terms and require investigation as part of method validation. 

Column size is another important consideration. For equipment designed for most routine laboratory HPLC situations the relative sensitivity of APTelectrospray instruments is better at low flow rates (0.2-0.8 mL/min) whereas the relative sensitivity of APCI instruments is enhanced at high flow rates (0.5-2 mL/min). As a result, small columns are appropriate for API-electrospray/MS and, if only one or two compounds of interest are found in a particular sample, high-resolution separations are not necessary. For APTelectrospray analysis of complex samples, 150 mm x 4.1 mml.D., 3 pm columns (flow 0.5-1.0 mL/min) are usually sufficient. For drug quantification involving analysis of single or low numbers of compounds, small columns such as 30 mm x 2.1 mm I.D., 3.5 pm columns (flow rate 0.2-0.4 mL/min) provide sufficient separation and a saving in both column cost and solvent utilization. The reduced injection volume required for the small columns often results in better resolution and increased sensitivity. 

Chen, Y. R., Tseng, M. C., Chang, Y. Z., and Her, G. R. (2003). A low-flow CE/electrospray ionization MS interface for capillary zone electrophoresis, large-volume sample stacking, and micellar electroklnetlc chromatography. Anal. Chem. 75, 503 — 508. 

Wetterhall, M., Nilsson, S., Markides, K. E., and Bergquist, J. (2002). A conductive polymeric material used for nanospray needle and low-flow sheathless electrospray ionization applications. Anal. Chem. 74, 239-245. 

Electrospray ionization can be considered as an electrolysis cell (Fig. 1.11) where, in the positive mode, cations are enriched at the surface of the solution and negative ions move inside the capillary. Oxidation of the analyte has been observed at certain occasions, in particular at very low flow rates. Also in the case of... 

DC Gale, RD Smith. Small-volume and low-flow-rate electrospray ionization-mass spectrometry of aqueous samples. Rapid Commun Mass Spectrom 7 1017-1021, 1993. 

Nanoelectrospray ionization (nanoESI), also known as nanospray, nanoflow electrospray, and micro-electrospray, is a low flow/high sensitivity approach to ESI. NanoESI15 is a slight variation on ESI such that the spray needle has been made very small and is positioned close to the entrance of the vacuum of the mass spectrometer and the mass analyzer (Figure 6). This greatly reduces required sample amounts allowing nanoliter flow rates and femto-mole sample consumption. The end result is increased efficiency since the flow rates for... 

Electrospray in the mid 1980s revolutionized biological mass spectrometry, in particular in the field of protein and peptide sequence analysis. Electrospray is a concentration-dependent, rather than a mass-dependent process, and maximum sensitivity is achieved at low flow rates with high-concentration, low-volume samples (Griffiths 2000). Joint NMR, x-ray diffraction, electrophoresis, and chromatography techniques with mass spectrometry (MS) techniques would be a trend in the future. 

D. R. Barnidge, S. Nilsson, and K. E. Markides, A design for low-flow sheathless electrospray emitters, Anal. Chem., 71 (1999) 4115-4118. 

Two factors are driving the market for precise, very-low-flow HPLC pumping systems extremely limited sample sizes in biotechnology and the electrospray and nanospray interfaces that are concentration and flow-rate dependent. It is very difficult to get precise flow and gradient formation from pumps that have a 5- to 10-/iL plunger displacement, even using 3200-step stepper motor drives. This has forced manufacturers to resurrect a very old concept from the earliest days of HPLC, the syringe pump. 

Recently, the word nano has become a trend in science and technology and some of us think that it is the new generation but, as mentioned above, its root is about 22 years old. NLC and NCE are gaining importance day by day. They are very useful and effective tools for samples of low quantities or having low concentrations of the analytes. Columns of low internal diameter are ideal for use in NLC and NCE, especially with detectors requiring very low flow rates, such as electrospray liquid chromatography/mass spectroscopy (LC/MS). Besides, these columns offer high sensitivity due to their low... 

The choice of interface is dependent on both the particular analysis and the instrumentation available. Some interfaces require the use of very low flow rates and therefore necessitate the use of either microbore or capillary LC equipment, or a sample splitter if standard-bore equipment is used. Thermospray ionization is the most frequently quoted interface, owing to compatibility with standard-bore instruments. However, the upper molecular weight limit for thermospray ionization is low, and the electrospray interface is becoming popular. The maximum flow rates for different interfaces are listed in Table 3.8.54... 

The development of the first CE-MS was prompted by the early reports on electrospray ionization (ESI-MS) by Fenn and co-workers in the mid-1980s [1], when it was recognized that CE would provide an optimal flow rate of polar and ionic species to the ESI source. In this initial CE-MS report, a metal coating on the tip of the CE capillary made contact with a metal sheath capillary to which the ESI voltage was applied [5]. In this way, the sheath capillary acted as both the CE cathode, closing the CE electrical circuit, and the ESI source (emitter). Ideally, the interface between CE and MS should maintain separation efficiency and resolution, be sensitive, precise, linear in response, maintain electrical continuity across the separation capillary so as to define the CE field gradient, be able to cope with all eluents presented by the CE separation step, and be able to provide efficient ionization from low flow rates for mass analysis. 

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