Abstract

   Background/Objectives: Bupleuri Radix is a plant in the Apiaceae family
   Bupleurum Chinense DC. or Bupleurum scorzonerifolium Willd. root. The
   dissimilarities in the metabolite profiles of plants directly correlate
   with the disparities in their clinical efficacy. Methods: Therefore,
   the wild Bupleurum Chinense DC. (YBC) and wild Bupleurum
   scorzonerifolium Willd. (YNC) were used as research materials. They
   were analyzed using the UPLC-MS/MS and the similarities and differences
   were uncovered based on differential metabolites. Results: Our results
   proved that the differences in clinical efficacy between YBC and YNC
   may be attributed to their distinct metabolite profiles, as follows:
   (1) a total of 12 classes of 2059 metabolites were identified in the
   roots, with phenolic acids, terpenoids, and flavonoids being the most
   abundant metabolic products, with 2026 shared components between the
   two, 2045 in YBC, and 2040 in YNC; (2) a total of 718 differential
   metabolites were identified, accounting for 35.44% of the shared
   metabolites. Among them, YBC had 452 metabolites with higher content
   relative to YNC, representing 62.95%, and 266 components with lower
   content, representing 37.05%; (3) the KEEG enrichment analysis results
   show that the differential metabolic pathways are flavone and flavonol
   biosynthesis, linoleic acid metabolism, arachidonic acid metabolism,
   sesquiterpenoid and triterpenoid biosynthesis, and linolenic acid
   metabolism. Conclusions: These new findings will serve as a foundation
   for further study of the BR biosynthetic pathway and offer insights
   into the practical use of traditional Chinese medicine in clinical
   settings.

   Keywords: Bupleurum chinense DC., Bupleurum scorzonerifolium Willd.,
   differentially accumulated metabolites, Chinese medicine, UPLC/ESI-Q
   TRAP-MS/MS

1. Introduction

   Bupleuri Radix (BR, or Chaihu in Chinese) is a plant in the Apiaceae
   family Bupleurum Chinense DC. (BC) or Bupleurum scorzonerifolium Willd.
   (NC) dried root. According to their different characters, they were
   called “Northern Bupleurum” and “Southern Bupleurum” and had a
   medicinal history of more than 2000 years [[40]1]. BR contains
   saponins, flavonoids, volatile oils, polysaccharides, lignans and other
   components [[41]2]. It has the effect of clearing heat deficiency
   [[42]3], soothing the liver and relieving depression [[43]4], and
   lifting Yang Qi [[44]5], and was often used for cold fever, cold and
   heat exchanges, chest and hypochondriac pain, uterine prolapse,
   menstrual irregularity, insomnia, and dreaminess [[45]6,[46]7]. On the
   other hand, it has anti-tumor, anti-depression [[47]8],
   anti-inflammatory, anti-viral, and other effects [[48]9]. It was a
   commonly used clinical medication for relieving exterior symptoms and
   has a large market demand.

   In the clinical application, BC is mainly used to treat typhoid fever
   and NC is mainly used to clear liver fever [[49]10]. Over the past
   decade, in most areas of our country, there was no distinction and
   there was confusion about its use. Some parts of China, such as
   Beijing, required that when using two kinds of BR, the doctors should
   prescribe them separately as needed. It is traditionally believed that
   NC has better quality than BC because of its medicinal material
   properties, color, odor, and habitat [[50]11,[51]12,[52]13]. However,
   the quality of medicinal materials is uneven, high-quality BR resources
   were relatively scarce, and BC was the mainstream in the north of
   commodity circulation. Currently, the medicinal materials mainly come
   from cultivated bupleurum, mainly from Gansu, Shanxi, and Shaanxi
   provinces.

   Studies have shown that there were differences in pharmacological
   effects between them. Through the study of the saikosaponin (SS)
   content in NC and BC, the results showed that the total SS in BC and
   SSa were more than those in NC, and some studies showed that the total
   SS in BC was about four times that in NC [[53]14]. In addition, through
   the study of the volatile oil components in BR, it was found that the
   volatile oil content of BC was much lower than that of NC, and there
   were obvious differences in the chemical composition [[54]15]. Not only
   were there many differences in the proportion of common components, but
   NC also contained some unique sesquiterpene components, such as
   piperene, A-cedrene, b-elemene, etc. Guo et al. [[55]16] compared the
   antidepressant efficacy of different varieties of Xiao Yao powder
   composed of BR to explore its influence on the changes in endogenous
   metabolites in rats, and found that both had obvious antidepressant
   effects, but the antidepressant effect and onset time of Xiaoyao powder
   composed of NC was slightly better than that of BC, indicating that NC
   had a unique advantage in pharmacological activity.

   Metabolomics technology can comprehensively analyze metabolites in
   organisms, provide strong technical support for the study of natural
   products, and provide a more comprehensive perspective for the study of
   natural products [[56]17]. It’s a newly developed important subject of
   bioomics and also is an important component of systems biology
   [[57]18,[58]19,[59]20]. In this study, we selected the wild Bupleurum
   chinense DC. (YBC) and Bupleurum scorzonerifolium Willd. (YNC) as
   research materials, and the metabolite types of them were detected by
   using widely targeted metabolomics methods and ultra-high-performance
   liquid chromatography tandem mass spectrometry (UPLC/ESI-Q TRAP-MS/MS)
   technology, and their differences were compared and analyzed. From the
   perspective of different metabolites, the similarities and differences
   between them were revealed, which provided references for the