Antioxidant and anti-inflammatory activity of Ocimum labiatum extract and isolated labdane diterpenoid

Background: Plants from the genus Ocimum are used as folk medicine for treating various diseases including inflammatory and immune-related diseases. Numerous reports have suggested plant extracts and their constituents as possible anti-inflammatory agents. Here, in vitro evidence of Ocimum labiatum's immune-enhancing and antioxidant properties is presented for the first time. Methods: The anti-inflammatory effect of O. labiatum ethanolic extract and an isolated diterpenoid was determined using a cytometric bead array (CBA) technique. The effect on phytohemagglutinin (PHA)-induced nitric oxide (NO) production in peripheral blood mononuclear cells (PBMCs) was also assessed. A battery of antioxidant assays were used for detecting antioxidant activity while the anti-inflammatory mechanism was evaluated using an ELISA-based activator protein (AP-1) (c-Jun) assay. Cytotoxicity was determined on TZM-bl and PBMCs using a tetrazolium dye and confirmed by a novel label-free real-time assay. Results: A 25 mu g/mL non-cytotoxic concentration of O. labiatum extract significantly (p < 0.05) inhibited the production of pro-inflammatory cytokines; IL-2, IL-4, IL-6 and IL-17A. Except for the dual acting pro-or anti-inflammatory cytokine, IL-6, which was upregulated, a non-cytotoxic 50 mu M concentration of the isolated labdane diterpenoid compound significantly (p < 0.05) decreased the production of all the pro-inflammatory cytokines. In the anti-inflammatory pathway studies, the compound also inhibited AP-1 significantly (p < 0.05) at 50 mu M. The extract demonstrated strong, dose dependent antioxidant activity with IC50 values ranging from 13 +/- 0.8 to 54.86 +/- 1.28 mu g/mL while the terpene had no antioxidant property. The extract and diterpenoid decreased the production of the inflammatory mediator NO, at non-cytotoxic concentrations. The CC50 of the extract in TZM-bl and PBMCs was 62.6 +/- 0.6 and 30.1 +/- 0.4 mu g/mL while that of the compound was 112.6 +/- 0.2 and 70 +/- 0.4 mu M respectively. The real time studies confirmed tetrazolium dye assessed viability and also detected a unique growth pattern for the plant materials compared to untreated cells. Conclusions: O. labiatum extract demonstrated promising anti-inflammatory and antioxidant properties while the terpenoid showed anti-inflammatory but no antioxidant activity. The anti-inflammatory mechanism of the terpene was a result of inhibition of AP-1. These data represents promising first steps towards the development of naturally derived anti-inflammation drugs.