Stable gradient nanoflow LC-MS

Abstract

This paper demonstrates improved nanoflow LC-MS performance on a QqTOF instrument with the incorporation of a heated nanoflow interface (particle discriminator) and a nebulizer assisted sprayer. It is shown that the nebulizer broadens the usable range of electrospray potentials, simplifying the tuning procedure, particularly for negative mode nanoflow gradients. The improved desolvation capability with the particle discriminator results in signal/noise improvements of approximately 3.5ϫ for negative ion mode samples prepared in predominantly acidified water as well as increased ion current stability. For nanoLC applications, the combined desolvation capabilities of a counter-current gas and heated laminar flow chamber provide reduced background, increased signal stability, reduced background drift, and improved protein sequence coverage when compared with data generated with only a counter-current gas for desolvation. This system is capable of subfemtomole nanoflow LC-MS sensitivity in both positive and negative ion mode across the solvent gradient. and electrospray ionization-mass spectrometry (ESI-MS) has evolved substantially since its initial implementation in the 1980s [1]. Traditional LC systems operated at solvent flow rates that were orders of magnitude higher (hundreds of L/min to mL/min) than those used for mass spectrometry (tens of L/ min), resulting in the need for solvent flow splitters [2]. Some of the subsequent improvements were focused on the use of nebulizer gases [3] and heating [4,5] for stabilization of the electrospray process to accommodate higher LC solvent flows.An alternate approach involves modification of the LC system to accommodate separations within the nanoflow regime [6,7]. Nanoflow LC-MS offers benefits in terms of sensitivity, solvent consumption, and sample consumption when compared with higher flow rate LC-MS, however, it can be much more difficult to operate. Nanoflow sprayers are typically fabricated from tapered fused silica capillary resulting in the need for some type of surface coating [8,9], inserted electrode [10,11], conductive union [6, 12, 13], or dialysis membrane [14] for application of the electrospray potential. Alternatively, sprayers can be fabricated from conductive materials, such as stainless steel, [15], however, tapered stainless steel sprayers are much more difficult to fabricate than fused silica sprayers. An additional benefit associated with fused silica sprayers is that the analytical column may be packed directly into the tapered sprayer, eliminating any post-column dead volume [16,17].A necessary but understated requirement for gradient elution in nanoLC-MS is an electrospray system that is capable of stable operation over a wide range of solvent compositions, particularly for samples containing both hydrophilic and hydrophobic components. Typical nanoflow electrospray systems display good stability for samples consisting of 20 to 100% organic solvent. However, stability can be an issue when running predominantly aqueous solvents because of the h...

Nano and rapid resolution liquid chromatography–electrospray ionization–time of flight mass spectrometry to identify and quantify phenolic compounds in olive oil

“…Finally, the B content was decreased to the initial conditions (20%) within 2 min and the column rinsed with these conditions for 5 min. As it can be observed, the gradient is limited between 20 and 100% of organic solvent, as in most of the nanoLC-MS systems described in the literature [14,27,28], in order to improve spray stability that can be an issue when predominantly aqueous solvents are used (due to the highsurface tension of water). Different flow rates were tested: 200, 300 and 400 nL/min (the maximum flow rate supported by the column is 600 nL/min).…”

Calibrant delivery for mass spectrometry

“…All nanoflow electrospray experiments were carried out using the standard MicroIonSpray II assembly [4] located in close proximity to the inlet of the curtain plate. Tapered fused silica sprayers (New Objective Inc., Woburn, MA) were used with inner, outer, and tapered tip diameters of 20, 360, and 10 m, respectively, to limit the dead-volume after the conductive liquid junction.…”

Ion sampling effects under conditions of total solvent consumption

“…Five roughing pumps (four Varian HS 602 pumps and one Leybold S25B pump) were required for the largest orifice diameter. To desolvate the spray, a heated laminar flow chamber (particle discriminator interface (PDI)7, 8) modified to handle higher temperatures was used. Four different orifice plates were tested with aperture diameters of 0.25, 0.33, 0.6, and 0.88 mm.…”

“…The gas flow was set to 0.2 L/min providing both atomization and cooling of the liquid inside the sprayer electrode. A MicroIonSpray II assembly7 was used to mount the nebulizer and sprayer electrodes. An electrospray potential of (±) 2800–3400 V was applied through the internal low dead volume union 5.2 cm upstream of the sprayer tip.…”

“…The electrospray capillary tip geometry and position proved to be critically important to achieve these high efficiencies with sprayer‐to‐sprayer measurements ranging from less than 1% to 12% overall efficiency. Much of the work to date has focused on improving sprayer‐to‐sprayer reproducibility,4, 5 ion current stability,4–6 for operation under conditions of high aqueous solvents, and flows in the hundreds of nL/min utilizing heated chambers and nebulizer gases,7–9 and general adaptability to liquid chromatography separations 7–13…”