Whitening Agents from Reseda luteola L. and Their Chemical Characterization Using Combination of CPC, UPLC-HRMS and NMR
Skin whitening agents occupy an important part of the dermo-cosmetic market nowadays. They are used to treat various skin pigmentation disorders, or simply to obtain a lighter skin tone. The use of traditional skin bleachers (e.g., hydroquinone, corticoids) is now strictly regulated due to their side effects. When considering this and the growing consumers' interest for more natural ingredients, plant extracts can be seen as safe and natural alternatives. In this perspective, in vitro bioassays
... in vitro bioassays were undertaken to assess cosmetic potential of Reseda luteola, and particularly its promising whitening activities. A bioguided purification procedure employing centrifugal partition chromatography, Ultra Performance Liquid Chromatography-Mass Spectrometry (UPLC-HRMS) and NMR was developed to isolate and identify the whitening agents (i.e., luteolin and apigenin) from aerial parts of R. luteola. UPLC-HRMS also enabled the characterization of acetylated luteolin-and apigenin-O-glycosides, which occurrence is reported for the first time in R. luteola. One can observe a boom in the development of natural whitening agents over the last decade: the search for natural tyrosinase inhibitors in recent years has demonstrated that plant extracts could be potential sources of new whitening ingredients [1, 5] . Reseda luteola L. (Resedaceae family) is a biennial species native from western Asia and is naturalized in the Mediterranean basin that grows up to 1.5 m by 0.5 m in sunny exposure  . Commonly named 'dyer's weld', the aerial parts of this herbaceous species were used to produce a mordant dye, especially for dyeing wool and silk during Medieval and early modern times: the word 'luteola' meaning 'yellowish' actually refers to the color of this dye [7, 8] . The substances responsible for producing this color are two flavones: luteolin, principally concentrated in all of the upper green parts of the plant, e.g., the flowers and the seeds, on one hand, and apigenin on the other hand [7, 8] . Furthermore, O-glycoside-forms of these two flavones are also inventoried in R. luteola: usually hydrolyzed to the parent aglycone flavone in the dyebath, they indirectly participate to the yellow color  . Displaying interesting agronomic characteristics, as well as good dyeing properties, R. luteola's potential to be a promising new crop was investigated as part of the PrisCA project in Italy . This little-demanding species is nowadays considered as a non-food crop, supporting the growing demand for natural colorants in the building (constituting an alternative to toxic paints) and DIY (Do It Yourself) sectors , as well as in the plastic industry  , and is therefore cultivated in numerous small plots in the Mediterranean basin. When considering this and the growing consumers' interest for more natural dermo-cosmetics, the anti-oxidant potential, as well as the whitening and anti-inflammatory activities of R. luteola extract were hence assessed using in vitro bioassays. From an extensive literature survey, it appeared that research on the chromatographic separation and quantification of flavonoids from R. luteola has already been undertaken    . However, no study on the preparative isolation of its bioactive agents by preparative chromatography technique has been reported to date, even given the widespread use over the last decade of countercurrent chromatography (CCC) to isolate natural bioactive, and often chemically sensitive, compounds [15, 16] . These reasons prompted the use of CPC (Centrifugal Partition Chromatography), applied in elution-extrusion mode, and combined with UPLC-HRMS (Ultra Performance Liquid Chromatography-Mass Spectrometry) to separate, purify, and identify the major bioactive flavonoids from an ethyl acetate extract of R. luteola displaying promising whitening properties.