A new HPLC method developed for the analysis of oxygen heterocyclic compounds in Citrus essential oils

In 2009, a temporary crisis in the world production of acetonitrile caused the abnormal increase of the price of this substance, commonly used as a solvent and in high-pressure liquid chromatography (HPLC) separation as a mobile phase. For this reason, a new HPLC method was developed to analyze the oxygen heterocyclic components of Citrus oils, substituting acetonitrile with methanol. The new method described in this article was validated, new components were isolated and characterized by nuclear magnetic resonance (NMR), and others were synthesized, to obtain pure standards, not commercially available. The results are evaluated in comparison with methods previously developed. Seven genuine cold-pressed Citrus essential oils – lemon (C. limon (L.) Burm.), Key lime (C. aurantifolia (Christm.) Swing.) type B, bergamot (C. bergamia), grapefruit (C. paradisi Macf.), bitter orange (C. aurantium L.), sweet orange (C. sinensis (L.) Osbeck) and mandarin (C. deliciosa Ten.) – were analyzed under identical conditions. In conclusion, it was observed that the replacement of acetonitrile with methanol in the mobile phase and the increase of the column temperature (from 30 to 40°C) led to a better separation in terms of resolution and total analysis time. Limit of detection (LOD) and limit of quantification (LOQ) were comparable with methods previously developed with acetonitrile in the mobile phase.