1. pH can change the retention because the analyte charge could be suppressed, but it can also affect MS response.
2. We want to adjust pH to promote the charged state of the analyte in the electrospray ionization mode.
3. The concentration of the buffer, or acid or base used to adjust/control the pH, should be as low as possible for ES.
4. Ammonium acetate and ammonium formate are generally applicable at pH 7 but concentrations should be kept to a minimum. …

Optimization of the Selection of Mobile Phase #

The selection of mobile phase is important to obtain a good chromatographic separation, but it also affects the analyte ionization and the sensitivity of the mass spectrometer. For example, analyte charge should be suppressed by manipulation of the mobile-phase pH for optimum retention but this can have a detrimental affect on MS response. Contrary to conditions for RP LC retention, for optimized electrospray (ES) ionization, the pH should be adjusted to promote the charged state of the analyte over its neutral species as ionization takes place in the liquid phase. In contrast, for optimized atmospheric pressure chemical ionization (APCI), in which ionization takes place in the gas phase, formation of the neutral species is favored due to the higher volatility of the neutral versus charged species and hence better vaporization. A compromise can be sought by experimentation but this is particularly difficult when a wide range of pesticides, with differing properties, have to be analyzed.

The situation is further complicated if polarity switching is employed so that positive and negative ions are periodically sampled throughout the analytical run. Alternatively, the target pesticides are divided into anionic and cationic groups and analysis performed separately. For routine application, even designs of orthogonal nebulizers for atmospheric pressure ionization interfaces are still restricted to the use of volatile buffers.

The concentration of the buffer, or acid or base used to adjust/control the pH, should be as low as possible for ES. If not, competition between analyte and electrolyte ions for conversion to gas-phase ions decreases the analyte response. If a species is in large excess, it will cover the droplet surface and prevent other ions to access the surface, and thus to evaporate. A species in large excess will also catch all charges available and prevent the ionization of other molecules present at much lower concentration.

Ammonium acetate and ammonium formate are generally applicable at pH 7 but concentrations should be kept to a minimum.

RP LC separations are sometimes improved at acidic pH, using acetic acid or formic acid, as such or in combination with ammonium acetate or ammonium formate.

The addition of reagents postcolumn can be used to generate pH conditions optimum for ionization without changing chromatographic separation but this approach is rarely implemented for routine analyses.

Methanol and/or acetonitrile are used as organic modifiers. Low surface tension and a low dielectric constant of the solvent promote ion evaporation, which favor the ionization process.

The gas-phase basicity (proton affinity) and gas-phase acidity (electron affinity) are also important solvent properties in the positive and negative ionization modes, respectively. Those features encourage the use of methanol versus acetonitrile.

Methanol is also preferred over acetonitrile when MS with ES is coupled to gradient LC because the lower eluotropic strength of methanol causes compounds to elute at a higher percentage of organic solvent, where ES sensitivity is increased. In most cases, more pesticides appear to elute in the middle of the analytical run.

Given the extra demand on MS acquisition in terms of obtaining sufficient data points across a peak and a long enough dwell time for sensitivity, it is surprising that few authors have reported efforts to optimize the gradient conditions so that pesticides exhibit a wider elution profile.

p76, Analysis of Pesticides in Food and Environmental Samples, CRC Press, 2008, edited by José L. Tadeo