Biomedical Research

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Quantitative determination of resveratrol in Polygonum cuspidatum and its anti-proliferative effect on melanoma A375 cells

Objective To establish HPLC method for determination of resveratrol content in Polygonum cuspidatum, and to study its inhibitory effect on human melanoma A375 cells. Methods HPLC with an Agilent column (model Angilent ZORBAX Eclips Plus C18, 4.6 × 250 mm, 5 μm) is used; mobile phase: methanol-water (75:25); flow rate: 1.0 mL/min; UV detection wavelength: 300 nm; injection volume: 5 μL; and column temperature: 25 ?. Anti-melanoma activity of resveratrol is analyzed by observing changes in cell morphology under inverted microscope and by MTT assay. Results Resveratrol shows a good linearity within a 1.248-12.48 μg range, and its recovery is 98. 58%, with a RSD of 1.62% (n = 6). MTT assay results show that the viability of A375 cells decreases with increasing concentration of resveratrol; and A375 cell inhibition rate of resveratrol exhibits rather evident dose-response relationship within the experimental concentration range. At high magnification, cells in the control group have clear nuclear membranes; a large number of mitochondria are visible within cytoplasm; double membranes and ridge structure are all clear. A small number of rough endoplasmic reticula, free ribosomes and lysosomes are seen, and cell structure is intact. Cells treated with resveratrol change rather greatly, with swollen organelles, increased number of intracellular granules, and even cell membrane disintegration, liquefaction degeneration cytoplasm, substantial disappearance of mitochondria and endoplasmic reticulum structure, release of cellular contents and occurrence of apoptosis. Cell morphology is not intact, and number of adherent cells is reduced. Conclusion HPLC method is accurate, simple and reproducible, which is suitable for quantitative analysis of resveratrol in Polygonum cuspidatum. Resveratrol can effectively inhibit the proliferation of human melanoma A375 cells.

Author(s): Ying Sun, Yan. Qi, Zhen Mu, Keyu Wang