Determining the Intracellular Concentrations of Flavonoids in MDA-MB-231 Cells Using HPLC-Coupled Mass Spectrometry

Abstract

Flavonoids are polyphenolic compounds widely distributed in fruits, vegetables and beverages. In recent years, flavonoids have been found to be beneficial to human health due to their anti‐oxidant and anti‐tumor activities. Our lab has investigated the ability of flavonoids to induce cytotoxicity in human breast cancer cell lines. We have discovered that some compounds can induce cytotoxicity very readily whereas others are not. We hypothesize that this difference is attributed to the compounds ability to transverse the cell membrane and enter the cytosol. Therefore, we have used HPLC‐Mass Spectrometry (LC‐MS) analysis to quantitate the amount of flavonoids that are able to enter into MDA‐MB‐231 cells, a human breast cancer cell line. We tested kaempferol, quercetin, myricetin, daidzein, and chrysin. MDA‐MB‐231 cells were plated, and treated with 100μM flavonoids for 24 and 48 hours. After treatment, cells were counted and extracted using 40% methanol. Flavonoids from extracts were characterized and quantitated by LC‐MS using an Agilent QTOF6520 tandem mass spectrometer and an Agilent 1100 HPLC. Our results indicate as intracellular flavonoid concentration increase, cellular cytotoxicity also increases. We could not detect quercetin and myricetin in cell extracts. However, quercetin was found to be a moderate inducer of cytoxicity. We find that quercetin dimerizes over time in cell medium only in the presence of serum suggesting an enzymatic mechanism. The quercetin dimer could not be detected in cellular extracts suggesting that it could not cross the membrane.

In Vitro Cytotoxic Activity of Origanum vulgare L. on HCT-116 and MDA-MB-231 Cell Lines

“…In addition, several studies have indicated high cytotoxic and anticancer activity of flavonoids [15,16]. Studies have shown that flavonoids may reduce breast cancer cell proliferation [17,18] and human colon carcinoma cell proliferation [19]. Flavonoid anticancer activities include inhibition of cell growth, inhibition of protein kinase activities, induction of apoptosis [20].…”

Flavanonas de Citrus - mecanismos de proteção de células β-pancreáticas submetidas ao estresse oxidativo

“…Por isso, elas podem ser mais expostas aos metabólitos de flavanonas no meio de cultura. Já in vivo, a quantidade dos metabólitos expostos às células pode não ser tão alta quanto in vitro, devido à superfície de crescimento limitada no ambiente celular (Pham, 2012).…”