| Zaoxintu,also known as Terra Flava Usta,is the soil block on the inner wall of the stove after being smoked and burned by firewood for a long time.It was first published in "Supplementary Records of Famous Physicians",and its clinical application has a history of more than 2,000 years.Zaoxintu has the effects of warming and drying dampness,relieving vomiting,hemostasis,and antidiarrheal.In addition to being directly used as medicine,it can also be used as a classic adjuvant of the traditional method of loess frying to increase the "spleen-warming" effect of medicinal materials.Huangtu Decoction,a classic recipe with Zaoxintu as the monarch,was first published in "The Synopsis of the Golden Chamber" by Zhang Zhongjing,a famous medical scientist in the Eastern Han Dynasty.It is a representative recipe for warming the spleen and stopping bleeding.This study took Zaoxintu and Huangtu Decoction as the objects to explore the efficacy and processing principle of "with the texture of loess and the properties of fire",and to study its modern processing technology.ObjectiveTo sort out the history and research status of Zaoxintu and Huangtu Decoction,preliminarily clarify the processing principle and mechanism of Zaoxintu "with the texture of loess and the properties of fire" in Huangtu Decoction,clarify the medicinal effect of Zaoxintu to warm the spleen and stop diarrhea,from affecting the intestinal flora and host metabolism,preliminary research on the physical and chemical properties of Zaoxintu and modern processing technology.Method(1)Literature research: Consult the classic ancient books of TCM and modern research documents,and sort out the historical evolution,processing method,quality standard,chemical composition,pharmacological effect,clinical application and Chinese patent medicine of Zaoxintu,the history,clinical application and modern research situation of Huangtu Decoction,analyze the problems existing in the current situation,and provide relevant theoretical basis for the follow-up research.(2)Research on the processing principle of Zaoxintu in Huangtu Decoction: LC-MS/MS technology was used to analyze the chemical components in Huangtu Decoction,and the network pharmacology method was used to analyze the mechanism of Huangtu Decoction’s "warming the spleen and stopping bleeding" to ulcerative colitis,compare and verify the efficacy of Zaoxintu and uncalcined loess in Huangtu Decoction,analyze the processing principle of Zaoxintu in Huangtu Decoction.The immunohistochemical,WB and q RT-PCR method were adopted to verify predicted mechanism of network pharmacology,and to clarify the processing connotation of Zaoxintu.(3)Study on the effect and mechanism of spleen-warming and anti-diarrhea of Zaoxintu:A mouse model of "spleen-yang deficiency and diarrhea" established to verify and compare the efficacy of Zaoxintu and uncalcined loess by indicators such as body weight,diarrhea rate,loose stool rate,stomach and small intestine pathological sections and scores.It was further explored through the gene sequencing of intestinal flora and metabolomic methods to explore the mechanism of Zaxintu.(4)Research on physicochemical properties and processing technology of Zaoxintu: The representative samples of Zaoxintu and loess from different areas in China were collected,and carry out character identification,physical and chemical identification,p H,bulk density,water-soluble extracts,infrared absorption spectrum,adsorption force measurement,scanning electron microscope and X-ray diffraction analysis,to find the difference between Zaoxintu and loess;at the same time,the calcination method was selected to simulate the formation conditions of Zaoxintu,and optimized by the response surface method,to acquire the calcining process parameters.Results(1)Literature research: Zaoxintu is a traditional Chinese medicine with a history of more than 2,000 years.Huangtu Decoction is a commonly used classic recipe for the clinical treatment of dysentery and blood in the stool,and both have clear clinical efficacy.However,modern research is insufficient.In the follow-up,we can carry out systematic research on the material basis and mechanism of Zaoxintu and Huangtu Decoction,and the modern processing technology of Zaoxintu,so as to provide clinical drugs with controllable quality and accurate efficacy,in order to realize the inheritance and innovation of traditional Chinese medicine.(2)47 compounds were separated and identified by LC-MS/MS in Huangtu Decoction,11 components were closely related to 18 key target genes of ulcerative colitis,and the mechanism of warming the spleen to stop bleeding in the treatment of UC may be related to the Matrix Metalloproteinases pathway.HTD(Zaoxintu based recipe)can reduce the weight loss of DSS-induced UC mice,improve the bloody stool symptoms and colon pathological damage,and reduce the serum IL-1β and IL-6 levels in UC mice,which was better than HTD(Loess based recipe).It showed that Zaoxintu had "the properties of fire" after been calcined,and enhanced the warm spleen and hemostasis effect of Huangtu Decoction.The mechanism of Huangtu Decoction is to inhibit the protein expression and m RNA level of MMP1,MMP3 and MMP9 in colon.(3)Zaoxintu has dose-related effect of warming the spleen and stopping diarrhea,which could significantly reduce the weight loss trend,diarrhea rate and loose stool rate,increasing the spleen and thymus index of mice,and improving the pathological damage of colon and gastric mucosa in the model mice of "spleen-yang deficiency and diarrhea",while no antidiarrheal effect of uncalcined loess found on model mice.The results of 16 s r RNA sequencing showed that the relative abundance of Erysipelotrichaceae and Allobaculum was significantly reduced,while Lachnospiraceae and Lachnospiraceae＿NK4A136＿group were increased after administration of Zaoxintu.The results of non-targeted serum metabolomics identified 12 biomarkers related to the spleen-warming and antidiarrheal effect of Zaoxintu.The KEGG pathway enrichment results showed that Zaoxintu may affect HIF-1 signaling pathway,vitamin digestion and absorption pathway and ascorbic acid and aldonate metabolism pathway by down-regulating L-ascorbic acid,affect platelet activation pathway,5-hydroxytryptamine synaptic pathway and arachidonic acid metabolism pathway by up-regulated prostaglandin G2,affect metabolic pathway of α linolenic acid by downregulated Jasmonic acid.(4)Through a series of studies on physical and chemical properties,it was found that there are significant differences between Zaoxintu and loess in the bulk density,watersoluble extract,infrared spectrum,and microscopic characteristics under the scanning electron microscope.X-ray diffraction shows that the main phase is hexagonal quartz(synthetic)phase both in Zaoxintu and loess,and it is believed that long-term calcination does not cause changes in the main chemical components in loess,which means Zaoxintu has "the texture of loess".Taking the bulk density and water-soluble extract as indicators,combined with the results of single-factor investigation,the BBD method was used to screen the calcination temperature,time and thickness of the modern processing technology of Zaoxintu.The optimal calcination process parameter was determined as at the temperature of 625 ℃,thickness of 2 cm,time of 75 h.The results of the process verification indicated that the optimal calcination process was basically reasonable and feasible,which provided a reference for the in-depth study of Zaoxintu.ConclusionZaoxintu "with the texture of the loess,the nature of fire" can enhance the effect of warm the spleen and stop bleeding of Huangtu Decoction in the treatment of UC,which mechanism is to inhibit the protein expression and m RNA level of MMP1,MMP3,and MMP9 in the colon.The "fire-like" warm spleen antidiarrheal effect of Zaoxintu is stronger than that of uncalcined loess,which can be used for the treatment of diarrhea due to spleenyang deficiency,and the mechanism may be related to increase the expression of beneficial bacteria and reduce the expression of harmful bacteria in the intestine,while regulating various metabolic pathways.No significant change found in main chemical composition between Zaoxintu and uncalcined loess,indicating that Zaoxintu has "the texture of loess".The calcination method can be used for the modern processing of Zaoxintu. |
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