Hemostatic action of lotus leaf charcoal is probably due to transformation of flavonol aglycons from flavonol glycosides in traditional Chinses medicine
Graphical abstract
Introduction
Driven by the growing market demand for traditional Chinese medicine (TCM) widely occurs from North America and East Asia, more and more attention has been drawn to the functional component contained in the herbal medicines as well as their bioactivities. However, the processing of Chinese herbal medicine, which represents a crucial part of the traditional Chinese pharmacology, has been largely untapped. Chinese herbal medicines were processed before use depending on their properties and clinical needs. Some of the raw materials are soaked, fried, steamed, boiled, carbonized, fermented or specially processed to either enhance or change their bioactive functions as recorded in the China Pharmacopoeia (2015 edition). The processing of TCM has been empirically performed with a long history. However, scientific evidence of these processing methods, in another word, the transformation of the bioactive components during processing as well as the criterion of the operating conditions remain elusive.
Nelumbo nucifera Gaertn, commonly known as lotus, is a well-known aquatic plant. All of the organs of lotus are eaten or used in China with a long history. The seeds and roots of lotus are edible, while its leaves, stamens, seedpods, and flowers can be taken as herbal medicines (Mukherjee et al., 2009). Lotus leaves contain flavonols and alkaloids, both of which contribute to several potential pharmacological activities such as antioxidant (Huang et al., 2010), anti-obesity (Ono et al., 2006) and antiviral effects (Kashiwada et al., 2005). The applications of lotus leaves in TCM fall into two categories. Ethanol extracts of raw lotus leaves (RLL) are primarily used to produce patent antilipemic agents such as Hedan Pian and Xuezhining Wan. Besides, lotus leaves are carbonized to charcoals to produce hemostatic styptics such as Heye Wan.
Many charcoal medicines are recorded in the China Pharmacopoeia e.g. Cirsii Japonici Herba Carbonisata, Schizonepetae Spica Carbonisata, Pollen Typhae Carbonisata and Sophorae Flos charcoal. Such charcoal medicines are mainly served as hemostatic styptics. Oral application of the charcoal medicines showed a therapeutic effect on hemoptysis, hematemesis and hemorrhage in clinic (Gao et al., 2019). A drug combination of ten charcoal medicines called Shihui San is proved to affect disseminated intravascular coagulation (DIC)-caused gastrointestinal bleeding (Xu, 2010) and idiopathic thrombocytopenic purpura (Zhang, 2002) in clinic. After being carbonized, the hemostatic activity of some herbal materials can be produced or enhanced. Several studies have been carried out to find the mechanism of this phenomenon. The most recent studies reported that the carbon dots derived from Pollen Typhae Carbonisata and Schizonepetae Spica Carbonisata showed promising hemostatic effect by decreasing activated partial thromboplastin time (APTT) as well as increasing fibrinogen (Fbg) and platelet count (PLT) (Sun et al., 2018; Yan et al., 2017). As reported by Wang et al., calcium oxalate crystals reduced in size and decreased in number during the carbonizing process (Wang and Lu, 1988). At high temperatures, insoluble calcium oxalate crystals were converted into soluble calcium ion, which involved in the acceleration of fibrin polymerization and thus conducive to promoting blood clotting (Boyer et al., 1972). In addition, flavonoid was found to be a significant component in most raw materials used as charcoal medicines, e.g. Sophora japonica L. flower (Paniwnyk et al., 2001), Cacumen biotae (Lu et al., 2006), Typha pollen (Tao et al., 2011) and lotus leaves (Li et al., 2014). The observation that quercetin in heated Sophora flower buds showed dose-dependent antihemorrhagic activity suggests that flavonoid could be another pivotal factor in the hemostatic charcoal medicines (Ishida et al., 1987).
Although efforts have been made on the chemical conversions in the charcoal medicines, the hemostatic principle of charcoal medicines and their associated production conditions remain unclear and lack of standardization. Therefore, the aim of this study is to find the hemostatic principle in lotus leaf charcoals (LLC), investigate the transformation of phytochemical compounds after carbonization, and to develop an evaluation criterion to optimize the processing conditions to LLC.
Section snippets
Plant materials and chemicals
N. nucifera leaves were collected from Guangchang, Jiangxi Province, China in May 2017, prior to the fresh samples being air-dried under shade. A voucher specimen has been deposited in the State Key Laboratory of Food Science and Technology, Nanchang University (accession number TZY1708). The dried lotus leaves were served as RLL. Half of the RLL samples were fine ground with a grinder. All of the samples were stored at 4 °C for further applications.
Quercetin, hyperoside, isoquercitrin,
Calcium content in RLL and LLC
Calcium content in RLL and LLC materials and their water infusions were determined by ICP-MS. As shown in Table 1, both the total calcium content in the materials and the soluble calcium content in the infusions showed no significant differences among the three groups. Although it has been reported that the transformation of calcium oxalate crystal into soluble calcium ion played a role in the hemostatic mechanism of the charcoal medicines (Wang and Lu, 1988), it may not be the hemostatic
Conclusion
Although the heating process did not increase the soluble calcium content in lotus leaves, the transformation of phytochemicals was observed in LLC. The flavonoids and alkaloids were found to be degraded during carbonization when comparing the phytochemical profiles of RLL and LLC obtained at different temperatures. Flavonol glycosides in RLL first degraded to their corresponding aglycons at 150 °C and then further decomposed to the hydroxybenzoic acids at 220 °C. EAE and 150 °C LLC which were
Author contributions
The animal experiments were designed by Yuhuan Chen and Zeyuan Deng, and performed by Yuhuan Chen. The detection of blood coagulation parameters and platelet count were performed by Fan Sun and Xiaozhong Wang. The basic concepts were clarified by Qiwen Chen. The manuscript was written by Yuhuan Chen and Qiwen Chen. The manuscript was revised and polished by Yawei Fan, Xiaoru Liu and Hongyan Li.
Declaration of competing interest
The authors declare no conflict of interest.
Acknowledgments
The authors acknowledge the National Natural Science Foundation of China (Grant 31671853 and 31860430) and Free Inquiry Project of State Key Laboratory of Food Science and Technology, Nanchang University (SKLF-ZZB-201709) for supporting this project.
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