## 3 Discussion  
### Substance Basis Difference between B. marginatum var. stenophyllum and Other Four Species  
Modern pharmacological studies have shown that the volatile oil of _Bupleurum_ has antipyretic, anti-inflammatory and other effects, and it is considered the pharmacodynamic material basis for Bupleuri Radix (\"_Chailhu_\" in Chinese) to mitigate fever [17]. _Chailhu injection_ made from volatile oil of Bupleuri Radix is a commonly used medicine in the treatment of fever, which is listed in the national standard for Chinese patent drugs (NSCPD). The sources of Bupleuri Radix (\"_Chailhu_\" in Chinese) listed in the _Pharmacopoeia of the People's Republic of China_ are _B. chinense_ DC. and _B. scorzonerifolium_ Wild. (State Pharmacopoeia Commission, 2020) and recognized as \"_North Bupleurum_\" and \"_South Bupleurum_\". In addition, _B. marginatum_ Wall. ex DC. and _B. smithii_ Wolff are collected by multiple local standards. The dried roots of _B. marginatum_ var. _stenophyllum_ are neither included in Pharmacopoeia nor in local standards. At present, there is a phenomenon of misusing _B. marginatum_ var. _stenophyllum_ as Bupleuri Radix in the market. _B. marginatum_ var. _stenophyllum_ is a variant of _B. marginatum_. In the heatmap, there were obvious differences between _B. marginatum_ var. _stenophyllum_ and other samples in chemical composition. And our PCA results also showed that _B. marginatum_ var. _stenophyllum_ are different from the other six samples (Figure 3A,B). Figure 8: The microscopic structure of the root cross-sections of the five species/varieties. The samples were stained by 1% safranine and 0.5% fast green solution; the red part represents the nucleus and the lignified cell walls; the blue part represents the cytoplasm and the cellulose containing cell walls. The slides were photographed using an Olympus BX51 light microscope. (**A**) BcR; (**B**) BsR; (**C**) BmsR; (**D**) BmR; (**E**) BsmR. In the Chinese tradition, _B. scorzonerifolium_ Willd contains a high content of volatile oil, which gives it a strong odor and is called \"_XiangChainu_\" [21]. Our results show that the volatile oil content of _B. scorzonerifolium_ Willd and its wild species was the highest among all the samples, which was consistent with the results of [PERSON] and [PERSON] et al. [21; 22]. In our results, the relative contents of lauraldehyde, \(\alpha\)-pinene, \(\beta\)-pinene, and sabinene in BsR and BsR_W were significantly higher than other samples. [PERSON]'s study suggested that lauraldehyde might be the effective components of the volatile oil of the _B. scorzonerifolium_ Willd [23], and that sabinene, \(\beta\)-pinene, and \(\alpha\)-pinene all had good anti-inflammatory activities [24; 25; 26]. In particular, the relative content of caproaldehyde in _B. chinense_ DC. was higher than that in other samples. One of the effects of Bupleuri Radix recorded in the _Pharmacopoeia of the People's Republic of China_ is its mitigation of fever. Some studies have shown that the two chemical components, caproaldehyde and heptaldehyde, are closely related to the pharmacological effect of the volatile oil of Bupleuri Radix in mitigating fever. Some studies also use caproaldehyde and heptaldehyde as the basis for quality control of _Chaihu injection_, _Chaihu oral liquid_, and _Chaipin soft capsules_[27; 28; 29]. This may be the reason why the antipyretic effect of the two species is better than that of _B. marginatum_ Wall. ex DC. and _B. smithii_ [PERSON]. The content of lauraldehyde in _B. smithii_ [PERSON] was much lower than that in _B. scorzonerifolium_ [PERSON], but the relative contents of \(\alpha\)-pinene, \(\beta\)-pinene, and sabinene in _B. smithii_ [PERSON] were significantly higher than in the other samples, which may be the reason why it was used for mitigating fever [30]. Although the volatile oil content of _B. marginatum_ Wall. ex DC.