Rapid and accurate analysis of rosmarinic acid in infusions of Melissa officinalis by LC-MS/MS

ARCE, SILVIA; ACOSTA, GIMENA; LLABOT, JUAN; MARTINEZ, LUIS DANTE; GOMEZ, MARÍA ROXANA; CERUTTI, SOLEDAD

Congreso; Reunión Internacional de Ciencias Farmacéuticas RICiFa; 2012

INTRODUCTION Infusions and preparations from Melissa officinalis L. (?Melisa? or ?Lemon balm?) are traditionally administered for the symptomatic treatment of gastrointestinal disturbances, functional dyspepsia, nervous disorders and in the therapy of Herpes simplex infections (1, 2). Lemon balm is rich in terpenoids, flavonoids and cinnamic acid derivatives, with rosmarinic acid (RA) as the main component, usually quantified in the quality control assays. However, it is well known that phenolic acids are unstable and sensitive to many different factors such as high temperature, exposure to UV radiation or air oxygen, and metal ions [3, 4]. Stability studies of RA have been restricted so far to analyses performed under the daylight [5], but temperature effect above certain limits has not been assessed. Therefore, the aim of this work was to establish a rapid and sensitive method for the analysis of RA in M. officinalis leaves. Prior determinations per se, different sample treatments were evaluated in terms of RA recovery. MATERIALS AND METHODS Organic cultured leaves of M. officinalis L. were used for analyses. Plants were grown in their natural habitat in the province of Entre Rios (Argentina). Air-dried plant material was ground, homogenized, and stored in darkness at room temperature. Analyzed samples consisted in infusions prepared according to FNA VI ed. (10% w/v). In order to compare, water extracts were both analyzed directly and mechanically dried on a Mini-spray BÜCHI Dryer 290, inlet temperature ranging between 90 and 140 °C, with Aerosil ® as carrier, and subsequently reconstituted. The determination of RA was carried out in a liquid chromatographic system (Acquity, Waters) coupled to a tandem mass spectrometer (Quattro Premier XE, Waters) equipped with an electrospray ionization source configured in a positive mode (LC-MS/MS). Thus a fast, sensitive, selective, and reliable methodology was developed. RESULTS Opposite to most of the time consuming methods reported in the literature and, as a consequence of using an ultra high performance system, the chromatographic resolution and detection of RA was accomplished in less than two minutes. The methodology was validated and applied to the aforementioned samples. To achieve a reliable quantification of RA and to determine its variation among samples, the effect of the sample matrix on the analyte signal was assessed. Infusions treated under different conditions were evaluated and the best results were obtained for those previously manipulated under the mechanical procedure, without observing negative changes in the RA content. In fact, the RA levels were higher in these samples. CONCLUSIONS A rapid, sensible, and reliable methodology for the evaluation of RA content in M. officinalis material was developed. To the best of our knowledge, this was the first time than evaluation of the matrix effect on the RA quantification signal was assessed when detection by a mass analyzer was performed. The proposed methodology allowed the monitoring of RA in M. officinalis infusions. Thus samples with and without spray-drying procedures were compared in order to establish the final RA intakes. Although the mechanical approach has the effect of increasing the amount of RA detected, it could also have a detrimental effect regarding the presence of degradation products; this is an open field of study that requires to be exhaustively examined. The results obtained have important implications for potential RA-based pharmaceutical formulations.