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DEVELOPMENT AND VALIDATION OF AN HPLC-DAD METHOD TO ESTIMATE TOTAL KNOWN AND UNKNOWN FUROCOUMARIN CONTENT
EP25670
DEVELOPMENT AND VALIDATION OF AN HPLC-DAD METHOD TO ESTIMATE TOTAL KNOWN AND UNKNOWN FUROCOUMARIN CONTENT
Submitted on 03 Apr 2017

Nicholas Housel (1), Ivan Domicio da Silva Souza (2), Rocco Burgo (1)
1. INOLEX Inc.; 2. Rowan University
This poster was presented at PittCon 2017, Chicago, USA
Poster Views: 1,766
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Poster Abstract
Furocoumarins are a family of natural chemicals with phototoxic potential, commonly found in plant extracts and fragrances. The European Cosmetics Directive 76/768/EEC limits the total content of furocoumarins in cosmetics to 1 ppm. Current methods define “total content” based on a certain number of external standards (markers), overlooking other unknown furocoumarins. An accurate total content analysis is unpractical, but we aimed to develop a method to estimate total content of both known and unknown furocoumarins. As a family of chemicals, different furocoumarins have similar UV absorption spectrum (see Fig. 1). Therefore, we propose an HPLC method based on external markers and the use of DAD-UV to identify unknown furocoumarins. Eight markers were studied: Bergapten; Bergamottin; Imperatorin; Isoimperatorin; Isopimpinellin; Oxypeucedanin; Xanthotoxin and 8-Geranyloxypsoralen. Elution gradient comprised water, acetonitrile and tetrahydrofuran through a C18 column. We obtained well resolved peaks (purity factors all above 999.9). Linearity proved to be satisfactory (R2= 0.9977 to 0.9998) in the range of 0.1 µg/mL to 2.5 µg/mL). Intra-day precision was highly noticed (RSD= 0.0865% to 1.73%) and inter-day precision was further confirmed (RSD= 0.0539% to 3.92%), except for Bergamottin and 8-Geranyloxypsoralen. Quantification limits ranged from 345.2 pg to 3421.4 pg. Percent recovery was determined using a plant extract not known to contain furocoumarins, and results were acceptable (85.68% to 114.03%), except for Bergamottin and 8-Geranyloxypsoralen. Robustness was studied by changing flow rate, oven temperature and concentration of mobile phase. The method was fully validated for 6 markers. DAD-UV detector allows us to identify unknown furocoumarins in between the markers and estimate their content based on linear regression of equivalent markers.

Frérot, E.; Decorzant, E. Quantification of Total Furocoumarins in Citrus Oils by HPLC Coupled
with UV, Fluorescence, and Mass Detection. J. Agric. Food Chem. 2004, 52, 6879–6886.
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