Natural products are used as traditional medicines for the treatment or prevention of a wide variety of human diseases due to their extensive pharmacological properties. As a kind of relatively safe natural product, plant-derived essential oils are more easily accepted by consumers. To reduce substance abuse, the use of essential oils to relieve or treat diseases has also been developed as an alternative therapy. Z. striolatum has traditional medicinal value, but the anti-inflammatory effect of its essential oil has not yet been reported. In the present work, we mainly analyzed the composition of EOZS from seven regions and evaluated their suppressive effect on inflammatory response and oxidant stress. Sixteen common chemical compounds were identified from the EOZS of seven regions, suggesting that these characteristic peaks may be used as the index components for quality control of Z. striolatum. From the phytochemical classification, Z. striolatum from seven locations could be divided into three clusters. The factors causing these differences may be related to the geographical location, climatic conditions and soil environment. Although the main chemical components and relative contents in S1–S5 were similar, the cytotoxicity of S1 was stronger than other samples. This indicated that it may be caused by minor or trace components in the S1.

Our research showed that EOZS from seven locations exhibited a certain degree of DPPH scavenging activity. Furthermore, they also significantly inhibited the production of NO and ROS in LPS-induced RAW 264.7 cells. Interestingly, EOZS from Huaihua City (S2), Zhangjiajie City (S4) and Enshi City (S5) showed a stronger inhibitory effect on NO production in LPS-induced RAW 264.7 cells. From the results of PCA and HCA, it can be seen that the main difference between cluster III (S1–S5) and other clusters included β-phellandrene, β-pinene, α-pinene and α-humulene. Lee et al. identified β-phellandrene as the main component of Zanthoxylum schinifolium essential oil, and found that Zanthoxylum schinifolium essential oil significantly inhibited the mRNA transcription level of inducible nitric oxide synthase. There was direct evidence that α-pinene and α-humulene suppressed NO produced by activated macrophages or monocytes. In addition, as the main component of the essential oil of Xylopia parviflora, β-pinene has been shown to reduce the NO production in LPS-induced RAW 264.7 cells. The above evidence may partly explain the anti-inflammatory activity of S2, S4 and S5. S1 was not compared with other samples in cluster III at the same concentration because of its stronger cytotoxicity. However, the main components of S3 and S2, S4 and S5 were similar, whilst the effect of S3 was weaker, indicating that there may be an antagonistic effect between different components in S3.

Transcriptomics is a method to study the expression of RNA and the regulation of transcription in cells. In recent years, transcriptomics has been widely used to reveal differences in the expression of genes in biological functions and to discover the mechanisms or targets of drugs. By using transcriptomics analysis, Li et al. found that Rebaudioside A could protect Caenorhabditis elegans from oxidative stress via regulating TOR and PI3K/Akt pathways. RNA sequencing results of this study indicated that the MAPK and NF-κB pathways may be involved in the potential anti-inflammatory mechanism of EOZS. The MAPK signaling pathway plays an important role in extracellular signaling to the inner nucleus, and it can be activated by external environmental stress or stimulation by inflammatory mediators. It has been shown that the conditional knockdown of the p38α MAPK gene results in a significant reduction in TNF-α production by LPS-induced microglia in mice, thus preventing the neuronal damage produced by LPS induction. NF-κB, as a core regulator of inflammatory response, has been a popular research target in the field of inflammation and immunity. When canonical NF-κB signaling pathway is activated, IκB is phosphorylated via the IκB kinase (IKK) complex and subsequently polyubiquitinated and degraded by the 26S proteasome. NF-κB is then phosphorylated and enters the nucleus, leading to the initiation and protein expression of pro-inflammatory genes such as TNF-α and IL-6. All of this evidence suggests that the activation status of MAPK and NF-κB signaling pathways has a critical impact on the production of downstream pro-inflammatory mediators. Therefore, the inhibition of the excessive activation of MAPK and NF-κB signaling pathways is a strategy for the treatment of inflammation-related diseases.

During inflammation, cells damaged by irritation recruit immune cells to migrate towards the site of inflammation by releasing cytokines and chemokines, while immune cells produce more inflammatory mediators through activated MAPK and NF-κB signaling pathways, creating a positive feedback loop that may further exacerbate the progression of inflammation. In this study, we demonstrated that EOZS inhibited the activation of macrophage and production of pro-inflammatory mediators through the regulation of MAPK and NF-κB signaling pathways (Figure 1). In vivo experiments also showed that EOZS could reduce the infiltration of immune cells to the site of inflammation to alleviate inflammation. Indeed, ample evidence suggests that plant volatile oils possess anti-inflammatory pharmacological activity in vivo and in vitro.