Purification of coumarins, including meranzin and pranferin, from grapefruit by solvent partitioning and a hyphenated chromatography

Abstract

Grapefruit heterocyclic oxygenated compounds have demonstrated antimicrobial and cytotoxic properties; purifying these compounds is essential for systematic biological studies to elucidate their bioactive properties. In the current study, a new method was used to isolate and purify coumarins from grapefruit oil and peels. Grapefruit oil was fractionated into volatile and non-volatile fractions (NVFs) by vacuum distillation. The NVF was further partitioned into hexane soluble and hexane insoluble fractions (HIFs). Then, the NVF–HIF composite was subjected to a hyphenated chromatography and separations were performed using a C-18 reversed phase column to yield several compounds. Additionaly, grapefruit peel was extracted with hexane and five compounds were purified using a normal phase silica column using hyphenated chromatography. Structures of the purified compounds were elucidated by 1H and 13C attached proton test (APT) NMR spectral data and identified as pranferin (1), meranzin (2), bergapten (3), dihydroxybergamottin (4), and marmin (9). Compounds (5–8) were later identified as heptamethoxy flavone, auraptene, osthol and nobilitin by comparing retention times in HPLC and mass spectra. With this method, it was possible to selectively separate major compounds from minor bioactives prior to hyphenated chromatography. This approach of solvent partitioning and rapid hyphenated chromatography proved highly effective in isolating several compounds, including pranferin from grapefruit oil.