Isolation, characterization, and HPTLC-quantification of compounds with anticancer potential from Loranthus acaciae Zucc.

Abstract

The cytotoxic effects of the crude extract of Loranthus acaciae Zucc. and its n-hexane, chloroform, and n-butanol fractions were assessed against three cancer cell lines using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay. Cell apoptosis was determined using an annexin V-phycoerythrin/7-aminoactinomycin kit. We observed that the L. acaciae n-hexane extract (LAHE) could inhibit cancer cell growth, particularly of MCF7 and A549 cells. Chromatographic purification of LAHE and nuclear magnetic resonance analysis led to the identification of two compounds from this plant species, namely, betulinic acid and β-sitosterol, for the first time. Flow cytometry study suggested that betulinic acid induced cell death via apoptosis, as a distinguished marked enhancement in the early and late apoptosis of human lung (A549) and breast (MCF-7) cancer cell lines. The isolated compounds were further estimated concurrently in LAHE using a validated high-performance thin-layer chromatographic (HPTLC) method on a 10 × 10 cm2 HPTLC plate with chloroform, methanol, and glacial acetic acid (97:2:1, v/v/v) as the mobile phase and a λmax of 540 nm. The amounts of betulinic acid and β-sitosterol in LAHE were 69.46 and 135.53 µg/mg of dried weight of extract, respectively. The excellent cytotoxic effect of LAHE could be attributed to the presence of ample amounts of betulinic acid.