Apigenin mediated gold nanoparticle synthesis and their anti-cancer effect on human epidermoid carcinoma (A431) cells

Abstract

We report that the flavonoid, apigenin (4', 5, 7,-trihydroxyflavone) is able to form apigenin linked gold nanoparticles (ap-AuNPs) at RT with apigenin itself acting as the stabilizing agent.The synthesized ap-AuNPs have been characterised by UV-Visible spectroscopy, HR-TEM, DLS, FTIR and TGA analyses. The biocompatible nature of ap-AuNPs is shown by its nontoxicity towards normal epidermoid cells (HaCat). Additionally, it is shown to exhibit anticancer activity towards epidermoid squamous carcinoma cells (A431). The uptake of ap-AuNPs into A431 cells is seen by TEM. Apoptosis induced by ap-AuNPs in these cells is observed through EtBr/AO and DAPI staining. Flow cytometric results also reveal the occurrence of early apoptotic cells, late apoptotic cells and the presence of sub-G1 population. DNA prepared from ap-AuNPs treated A-431 cells reveals a ladder like pattern in agarose gel electrophoresis indicating oligonucleosomal cleavage which is a hallmark of apoptosis. The ap-AuNPs also inhibit angiogenesis as shown by Chick chorioallantoic membrane (CAM) assay. Taking the results together, we believe that ap-AuNPs show promise in treatment of skin cancer. The ap-AuNPs also have cytotoxic potential against the human cervical squamous cell carcinoma cell line -SiHa.indicating that almost a four-fold increase in capacity to kill the cancer cells is realized by the ap-AuNPs conjugation.The effect of ap-AuNPs on SiHa cells has also been studied. The results show a concentration and time dependent decrease in viability. On treatment with 200 µg / mL of ap-AuNPs for 24 h, the viability is found to be 52.7 ± 2.6 %, which further decreases to 44.2 ± 3.1 % and 31.8 ± 1.1 % after 48 h and 72 h respectively (Fig. S5a). Gallic acid conjugated gold nanoparticles (150µM) are reported to induce cell death in SiHa cells. 48 The viability results are comparable to that of A431 cells indicating that the ap-AuNPs exhibit comparable cytotoxicity towards these two types of epidermoid squamous carcinoma cell lines. Microscopy Analysis EtBr/AO and DAPI stainingThe EtBr/AO staining of HaCat and A431 cells treated with different concentrations (50, 100, 150 and 200 µg / mL) of ap-AuNPs for 48 h has been carried out. Fig. 5a shows that the control and treated HaCat cells possess similar morphology and fluoresce green, which indicates that they are viable. No significant difference is observed even in the presence of a higher concentration of ap-AuNPs (200 µg / mL). But at the same time, A431 cells treated at a concentration of 50 µg / mL of ap-AuNPs show the presence of a few dead cells (red fluorescence). The initial stage of membrane damage and reddish-orange colour stained cells are observed in the presence of 100 µg / mL of ap-AuNPs. The loss of membrane integrity with most of the cells fluorescing red post-treatment at concentrations of 150 and 200 µg / mL of ap-AuNPs has also been observed. Under these conditions, some cells are also found to detach from the plate (Fig. 5b).The concentration of Au in the ap-AuNPs was determined by atom...

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“…Moreover, the internal mechanism of the conversion of ketones to carboxylic acids in flavonoids is likely to be involved in Au 3+ ion reduction [32]. The presence of such mechanisms may indeed explain the ability of flavonoids to be adsorbed onto the surface of a nascent nanoparticle implying that they are involved in the nucleation and aggregation stages of nanoparticle synthesis, as well as in the bio reduction stage [32,41].…”

“…Isolated flavonoids and flavonoid glycosides have been reported to have the ability to induce the formation of metal nanoparticles as in the case of Apiin (apigenin glycoside) extracted from Lawsonia inermis. This glycoside was used in the synthesis of anisotropic gold and quasi-spherical silver nanoparticles having an average size of 21-30 nm [32,41].…”

Plant Flavone Apigenin: an Emerging Anticancer Agent

“…The apigenin-AuNPs also exhibit decent antiangiogenic property. Hence, apigenin-AuNPs is a promising candidate for use in skin cancer treatment [234]. Further studies on nano-apigenin using poly (lactic-co-glycolide) (PLGA) was effective in oral and topical application for skin cancer, achieving a higher efficacy and potency with reduced toxicity [235].…”

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“…The 3-day-old chicks were treated with MoO 3 nanoparticles and were observed for 72 h. Figure 12 clearly displays that the control samples showed good gradual development and thickening of capillaries marking the events of angiogenesis. Whereas the membranes treated with MoO 3 did not show any development or thickening of vessels [28]. The control samples showed an increase in the capillaries after 24 h. These capillaries thickened uniformly into prominent blood vessels after 72 h. In the CAM cultured with MoO 3 nanoparticles, there was no significant increase in the capillaries.…”