Anti-Inflammatory Activity β-Patchoulene Isolated From Patchouli Oil In Mice

ObjectivesPogostemonis Herba, known as "Guang-Huo-Xiang" in Chinese, is dried aerial part of Pogostemon cablin (Blanco) Benth. (Labiatae). Clinically, it has been traditionally used for inflammatory disorders. In our previous investigation, patchouli oil, patchouli alcohol and pogostone were demonstrated to exert attractive anti-inflammatory activity in various animal and cell models.β-patchoulene (β-PAE, C15H24), one of the representative hydrocarbon sesquiterpenoids from patchouli oil. However, as one of the major principle of patchouli oil, the biological activity of β-PAE has not been explored so for. Additional, it is a kind of azulene compounds, which have strong anti-inflammatory activity. In the present work, we managed to isolate β-PAE and investigated whether oral application of β-PAE could exhibit in vivo anti-inflammatory effect.Methods1. The anti-inflammatory effect of β-PAETo establish an efficient method to obtain P-PAE from patchouli oil and performed the acute toxicity. In the present work, we managed to isolate β-patchoulene and investigated whether oral application of β-PAE could exhibit in vivo anti-inflammatory effect by using three common inflammatory animal models, i.e. xylene-induced ear edema in mice, acetic acid-induced vascular permeability in mice and carrageenan-induced paw edema in mice. To further elucidate the possible anti-inflammatory action and the underlying mechanisms, the effect of β-PAE on histological alternation, oxidative indicators such as MDA and MPO level, inflammatory mediators such as TNF-α, IL-1β, IL-6, PGE2 and NO, expression of inducible iNOS and COX-2, and NF-κ B signaling pathway were probed in the carrageenan-induced mice paw.2. The effect of β-PAE against LPS-induced acute lung injuryTo investigate the potential therapeutic effect of β-PAE (2.5,5 and 10 mg/kg) on lipopolysaccharide (LPS)-induced acute lung injury (ALI), mice were pretreated with β-PAE prior to LPS exposure. After LPS challenge, the lungs were excised and the histological changes, wet to dry weight ratios, MPO activity reflecting neutrophil infiltration, and MDA activity reflecting oxidative stress were examined. The inflammatory cytokines in the BALF were determined by ELISA assay. Moreover, the activation of NF-κ B p65 subunit, phosphorylated IκBα, Keapl and Nrf2 in the nucleus were measured by Western blot analysis, and meanwhile the dependent genes of NF-κB and Nrf2 were assessed by RT-qPCR.Results1. The anti-inflammatory effect of β-PAEThe β-PAE were abtained by fractional distillation and the purity was> 98%. The LD50 value was greater than lOg/kg, which indicated that the β-PAE had a relatively wide margin of safety. The results showed that β-PAE evoked a significant dose-dependent inhibition of ear edema induced by xylene, paw edema induced by carrageenan and suppressed the increase of vascular permeability elicited by acetic acid. Histopathological analysis indicated that β-PAE could markedly decrease the cellular infiltration in paw tissue. β-PAE was also shown to significantly decrease the malondialdehyde (MDA) level and myeloperoxidase (MPO) activity in edema paw. In addition, carrageenan-induced production of some pro-inflammatory cytokines including TNF-α, IL-1β, IL-6, PGE2 and NO, were suppressed in a dose-dependent manner in mice subjected to β-PAE pretreatment, and it also significantly down-regulated the protein expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). Further analysis revealed that β-PAE also inhibited the translocation of nuclear factor-κ B (NF-κ B) from the cytoplasm to the nucleus and stabilize the conversion of nuclear factor-κ Ba (I κ Ba) level.2. The effect of β-PAE against LPS-induced acute lung injuryThe results showed that pretreatment with β-PAE markedly improved survival rate, attenuated the histological alterations in the lung, decreased the wet/dry weight ratio of lungs, reduced the MPO and MDA levels, down-regulated the level of pro-inflammatory mediators including TNF-α, IL-1β and IL-6. Furthermore, pretreatment with β-PAE enhanced the Nrf2 dependent genes including GCLC, NQO-1 and H0-1 but suppressed NF-κ B regulated genes including TNF-a, IL-1β and IL-6.ConclusionTo the best of our knowledge, this is the first report investigating the anti-inflammatory property of β-PAE, and the observed anti-inflammatory action of β-PAE might be mediated through regulation on oxidation stress and inflammatory cytokines. Furthermore, administration of β-PAE improved survival rate, and the mechanism behind the protective effect was correlated with its regulation on the balance between Nrf2 and NF-κ B signaling pathways.