Effect of large volume injection of hydrophobic solvents on the retention of less hydrophobic pharmaceutical solutes in RP‐LC

Abstract

Injection of large volumes of samples in solvents other than mobile phase composition has been proved for some less hydrophobic compounds. Thus, the retention behavior of several compounds of pharmaceutical interest (isosorbide-2-nitrate, isosorbide-5-nitrate, tropicamide, pentoxifylline, and methyl p-hydroxybenzoate) was studied by using different hydrophobic solvents (n-hexane, n-heptane, or i-octane) as sample solvents. Two types of stationary phases were used: octyl and octadecyl modified silica (both of Zorbax Eclipse type). The experiments showed a linear dependence between capacity factor of each solute and sample injection volume, up to maximum volume values of about 680 microL for C8 stationary phase and 580 microL for C18 stationary phase, when the solutes are no longer retained in stationary phase. Injection of large volumes of these hydrophobic solvents is thus possible in RP-LC with a gradual reduction of retention and peak efficiency. Two major conditions are however necessary in order to apply such an injection approach: the solutes must have a proper solubility in hydrophobic solvents and meanwhile they have to be less hydrophobic than the sample solvent in order to avoid competition with solvent molecules in partitioning between mobile and stationary phases.

Sample preparation for large‐scale bioanalytical studies based on liquid chromatographic techniques

“…A new possibility is injecting directly a high volume of solvent phase (up to 500 μL) for achieving high sensitivity of chromatographic detection. This approach was checked in several theoretical studies (David, Galaon, & Aboul‐Enein, ; Udrescu, Medvedovici, & David, ), and successfully applied for bioequivalence studies for some drugs (Medvedovici, Udrescu, Albu, Tache, & David, ; Udrescu et al, ). The procedure becomes safer for the technical personal involved in sample preparation when the organic solvent is environmentally friendly, although all organic solvents have adverse effects on the environment (Filippou, Bitas, & Samanidou, ), or more pleasant (limonene) when manipulated for SP (Medvedovici, Udrescu, & David, ).…”

Peak compression induced by large volume injection of hydrophobic alcohols in reversed-phase liquid chromatography

“…A new possibility avoiding the solvent evaporation for sample concentraion is to inject directly high volumes of the organic solvent in order to load larger (detectable) amounts of extracted species. 13 Examples of non-miscible sovents usefull in LLE and applied in liquid chromatography by large volume injection are aliphatic [14][15][16] or aromatic 17 hydrocarbons, aliphatic alcohols, 18,19 chlorinated solvents, 20 or "green" solvents. 21 The principle of this approach applied to the reversed-phase liquid chromatography (RP-LC) relies on the adsorption of the sample diluent (solvent) onto the surface of the stationary phase at the head of the chromatographic column, while the dissolved analytes participate to the retention process on the rest of the stationary phase.…”

“…21 The principle of this approach applied to the reversed-phase liquid chromatography (RP-LC) relies on the adsorption of the sample diluent (solvent) onto the surface of the stationary phase at the head of the chromatographic column, while the dissolved analytes participate to the retention process on the rest of the stationary phase. 13,14 The sample solvent also elutes eventually from the column head and act as a "compressing" eluent for the analytes. Consequently, by high volume injection the analytes elute at smaller retention time than resulted from the injection of typical injection volumes (i.e., 1-10 μL).…”

Use of a green (bio) solvent – limonene – as extractant and immiscible diluent for large volume injection in the RPLC‐tandem MS assay of statins and related metabolites in human plasma

“…Large volume injection of samples obtained in diluents stronger than and not miscible with the mobile phase is strictly forbidden by the commonly accepted practices in liquid chromatography (Dolan, ). However, recent publications (Medvedovici et al ., ; Udrescu et al ., , ) indicate that such an approach is feasible if some conditions are fulfilled. Applications of the concept in bioanalysis are already available for determination of indapamide in whole blood (Udrescu et al ., ) and fenspiride in plasma (Medvedovici et al ., ).…”