Analytica Chimica Acta 406 (2000) 37–52 Challenges in achieving a fundamental model for ESI Terri L. Constantopoulos, George S. Jackson, Christie G. Enke ∗ Department of Chemistry, University of New Mexico, Albuquerque, NM, USA Received 14 August 1998; accepted 14 February 1999 Abstract The relative intensities of the ions in an electrospray ionization (ESI) mass spectrum typically do not reflect the relative concentrations of the ions in solution. A partitioning equilibrium model was developed to explain in quantitative detail many of the behaviors observed in ESI in terms of matrix effects. The fundamental experiments required to validate this model presented several challenges in terms of instrumentation, interpretation, and application. The most basic requirement is a stable spray current that can be measured with accuracy and precision since several factors depend on the spray current. Modifications to the instrument used for these experiments resulted in a stable spray current that could be measured accurately, minimum drifting of the total ion current (TIC), and removal of risking damage to the instrument if arcing occurs. The presence of multiple analyte and electrolyte peaks due to fragment and cluster ions, in addition to solvent and impurity ion peaks, require care in distinguishing and accounting for each component in the solution. The dependence of the spray current on several parameters (analyte and electrolyte concentrations, flow rate, needle voltage, and needle position) as well as the interdependencies among these parameters presented other challenges. The limited the range of variability of each parameter due to these interdependencies was extended by extending the range of stability of the spray current by replacing the metal needle with a glass capillary needle. Overcoming these challenges has enabled us to carry out the quantitative fundamental experiments required to establish a working ESI model with broad applications. ©2000 Elsevier Science B.V. All rights reserved. Keywords: Electrospray ionization; Sodium ion; Tetrapentylammonium ion; Working curve; Interference 1. Introduction The ability of electrospray ionization (ESI) to ex- tend the m/z range of mass analyzers with upper mass-to-charge limits has helped with the identi- fication, mechanistic interpretation, and structural elucidation of biomolecules [1–3]. Furthermore, the sensitivity of the technique has increased with the introduction of nanospray [4] and microspray [5]. ∗ Corresponding author. Tel.: +505-277-3159; fax: +505-277-2609. E-mail address: enke@unm.edu (C.G. Enke). Despite the high level of interest in and the large volume of work done with ESI, it is still very dif- ficult to predict the effects of solvent, analyte, and electrolyte on the sprayed solution. For many singly- and doubly-charged ions, the relative intensities of the ions in the spectrum do not reflect the relative concentrations of the ions in solution [6], sometimes by several orders of magnitude. This potentially high selectivity has limited the capabilities of ESI and avoiding matrix effects has become something of an art. However, the application of ESI could be opti- mized on fundamental grounds if the mechanism of selectivity is understood. 0003-2670/00/$ – see front matter ©2000 Elsevier Science B.V. All rights reserved. PII:S0003-2670(99)00600-5