[Quality control of Tianwang Buxin Pills based on UPLC fingerprint and multi-component quantification].

Tianwang Buxin Pills have demonstrated therapeutic effects in clinical practice, whereas there is a serious lack of comprehensive quality control to ensure the safety and effectiveness of clinical medication. In this study, ultra-performance liquid chromatography(UPLC) was employed to establish the fingerprint and the method for simultaneously determining the content of seven components of Tianwang Buxin Pills. Furthermore, chemometrics was employed to identify the key factors for the stable quality, which provided a reference for the comprehensive quality control and evaluation of this preparation. There were 25 common peaks in the UPLC fingerprints of 15 batches of Tianwang Buxin Pills, from which thirteen compounds were identified. A quantitation method was established for seven pharmacological components(α-linolenic acid, salvianolic acid B, glycyrrhetinic acid, schisandrin A, ß-asarone, 3,6'-disinapoylsucrose, and ligustilide). The principal component analysis(PCA) and partial least square discriminate analysis(PLS-DA) were performed to determine the key pharmacological components for controlling the quality stability of Tianwang Buxin Pills, which included 3,6'-disinapoylsucrose, α-linolenic acid, and ß-asarone. The established fingerprint and multi-component content determination method have strong specificity, stability, and reliability. In addition, 3,6'-disinapoylsucrose, α-linolenic acid, and ß-asarone are the key pharmacological components that ensure the quality stability between batches and can be used to comprehensively control the quality of Tianwang Buxin Pills. The findings provide a scientific basis for the quality evaluation and standard establishment of Tianwang Buxin Pills.

[Effect of origin species, body weight, and gender on quality of Bufonis Venenum].

Based on standard sampling for Bufonis Venenum, this study analyzed the effect of the origin, and body weight and gender of Bufo bufo gargarizans on the quality of Bufonis Venenum. To be specific, mass spectrometry(MS) and the content determination methods in Chinese Pharmacopoeia(2020) were adopted. First, MS was performed on 76 Bufonis Venenum samples collected from 40 cities/counties in 17 provinces/autonomous regions which were derived from B. bufo gargarizans and B. melanostictus. Based on content determination, the body weight and gender of B. bufo gargarizans, which influenced the quality of Bufonis Venenum, were evaluated. Multivariate statistical analysis suggested huge difference in the material basis of the medicinal material derived from B. bufo gargarizans and B. melanostictus, and 9 differential compounds were identified. The content of components specified in Chinese Pharmacopoeia was higher in the medicinal material derived from B. bufo gargarizans than in the medicinal material derived from B. melanostictus. The content of the components specified in Chinese Pharmacopoeia was low in Bufonis Venenum derived from heavy B. bufo gargarizans, and higher in the Bufonis Venenum produced by male B. bufo gargarizans than in that produced by female B. bufo gargarizans irrespective of time and geographic location. In summary, this study provide new ideas and reference for the quality control of Bufonis Venenum, collection and processing of Bufonis Venenum, artificial breeding of B. bufo gargarizans, and biosynthesis mechanism of Bufonis Venenum.

[Distribution characteristics of Bufo bufo gargarizans resources in China based on quality evaluation of Bufonis Venenum].

On the premise of strictly controlling the harvesting conditions of Bufonis Venenum, we studied the relationship between the quality and resource distribution of Bufonis Venenum in China, aiming to provide the data for comprehensively understanding the geographical distribution and characteristics of Bufonis Venenum in China. In this study, 105 samples of Bufonis Venenum were collected from 42 counties and cities in 19 provinces in China, and the material basis and index components were determined by mass spectrometry and high performance liquid chromatography. The obtained data formed the quality database of Bufonis Venenum from different producing areas in China. The analysis of the material basis showed that Bufonis Venenum was mainly produced in two characteristic regions(north area and south area) divided by Qinling Mountains, northern edge of Huaiyang hills and the connecting area of Huang-Huai Plain, Huangshan Mountains, and Tianmu Mountains. Eight differential components were identified in the Bufonis Venenum samples from the south area and the north area. All the Bufonis Venenum samples from the north area showed the content of index components above the requirements of Chinese Pharmacopoeia(2020 edition), while those from the south area had the content of index components lower than the standards of Chinese Pharmacopoeia(2020 edition). The quality evaluation showed uneven distribution of Bufonis Venenum quality, which was high in the north and low in the south. The results provided a research basis for the breeding base selection of Bufo bufo gargarizans.

Shexiang Baoxin Pill, a Proprietary Multi-Constituent Chinese Medicine, Prevents Locomotor and Cognitive Impairment Caused by Brain Ischemia and Reperfusion Injury in Rats: A Potential Therapy for Neuropsychiatric Sequelae of Stroke.

Ischemic stroke is a common type of cerebrovascular event and also the leading cause of disability. Post-stroke cognitive impairment occurs frequently in stroke survivors. Shexiang Baoxin Pill (SBP) is a proprietary Chinese medicine, initially used to treat cardiovascular diseases. Herein, we aim to explore the effects of SBP on oxygen glucose deprivation and reoxygenation (OGD/R) in neuronal cells (CATH.a) and cerebral ischemia/reperfusion injury induced post-stroke cognitive impairment in middle cerebral artery occlusion (MCAO) rat model. MCAO rats received two doses of oral SBP treatment (28 or 56 mg/kg) after 1 h of operation and once daily for 2 weeks continuously. Behavioral tests, immunoblotting, and immunofluorescence were examined after 14 days. Current data suggest that SBP enhanced cell viability and downregulated apoptosis via activating the PI3K/Akt signaling pathway in CATH. a cells. Furthermore, 14 days of SBP treatment promoted the recovery of learning and locomotor function in the MCAO rats. SBP up-regulated the expression of p-Akt, p-GSK3ß, as well as the expression of NMDAR1, PSD-95, and AMPAR. Also, SBP down-regulated the expression of p-CaMKII. These results indicated that long-term SBP treatment might be a potential option for cognitive impairment induced by the ischemic stroke.

Shexiang Baoxin Pill, a Traditional Chinese Herbal Formula, Rescues the Cognitive Impairments in APP/PS1 Transgenic Mice.

BACKGROUND: Shexiang Baoxin Pill (SBP), a formulated traditional Chinese medicine (TCM), has been widely used to treat cardiovascular diseases for years. This herbal mixture has been shown to promote differentiation of cultured neuronal cells. Here, we aimed to investigate the effects of SBP in attenuating cognitive impairment in APP/PS1 transgenic mice. METHODS: Ethanol and water extracts of SBP, denoted as SBPEtOH and SBPwater, were standardized and applied onto cultured rat pheochromocytoma PC12 cells. The potential effect of SBPEtOH extract in attenuating the cognitive impairments in APP/PS1 transgenic mice was shown by following lines of evidence: (i) inhibition of Aß fibril formation, (ii) suppression of secretions of cytokines, and (iii) improvement of behavioral tests by Morris water maze. RESULTS: SBPwater and SBPEtOH inhibited the formation of ß-amyloid fibrils and protected the Aß-induced cytotoxicity in cultured PC12 cells. In APP/PS1 transgenic mice, the treatment of SBPEtOH inhibited expressions of NO, NOS, AChE, as well as aggregation of Aß. Besides, the levels of pro-inflammatory cytokines were suppressed by SBP treatment in the transgenic mice. Importantly, the behavioral tests by Morris Water maze indicated that SBP attenuated cognitive impairments in APP/PS1 transgenic mice. CONCLUSION: The current result has supported the notion that SPB might ameliorate the cognitive impairment through multiple targets, suggesting that SBP could be considered as a promising anti-AD agent.

Shexiang Baoxin Pill, a Formulated Chinese Herbal Mixture, Induces Neuronal Differentiation of PC12 Cells: A Signaling Triggered by Activation of Protein Kinase A.

Background: Shexiang Baoxin Pill (SBP) is a well-known composite formula of traditional Chinese medicine (TCM), which is commonly used today in treating cardiovascular diseases. SBP consists of seven materials thereof, including Moschus, extract of Ginseng Radix et Rhizoma, Bovis Calculus Artifactus, Cinnamomi Cortex, Styrax, Bufonis Venenum, and Borneolum Syntheticum. Here, we are investigating the potential roles of SBP in inducing neuron differentiation, i.e., seeking possible application in neurodegenerative diseases. Methods: Water and ethanol extracts of SBP, denoted as SBPwater and SBPEtOH, respectively, as well as its individual herbal materials, were standardized and applied onto cultured rat pheochromocytoma PC12 cells. The potential effect of SBP extracts in neuronal differentiation was suggested by following parameters: (i) induction of neurite outgrowth of PC12 cells, (ii) increase of neurofilament expression, and (iii) activation of transcription of neurofilament. Results: The treatments of SBPwater and SBPEtOH, or extracts from individual herbal materials, with or without low concentration of nerve growth factor (NGF), could potentiate the differentiation of cultured PC12 cells. The differentiation was indicated by increase of neurite outgrowth, as well as expression of neurofilaments. In addition, application of H89, a protein kinase A (PKA) inhibitor, suppressed the SBP-induced neurofilament expressions, as well as the phosphorylation of cAMP-responsive element binding protein (CREB) in cultures. Conclusion: SBP is proposed to possess trophic activity in modulating neuronal differentiation of PC12 cells, and this induction is shown to be mediated partly by a cAMP-PKA signaling pathway. These results indicate the neurite-promoting SBP could be useful in developing potential drug in treating or preventing neurodegenerative diseases.

Protective effect of Danning tablet on acute livery injury with cholestasis induced by α-naphthylisothiocyanate in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Danning tablet, as a composite prescription of traditional Chinese medicine, has been used clinically to relieve liver and gallbladder diseases in China. However, the mechanisms involved are still unclear. AIM OF THE STUDY: The present investigation was designed to assess the effects and possible mechanisms of Danning tablet on α-naphthylisothiocyanate (ANIT)-induced liver injury with cholestasis. MATERIALS AND METHODS: Danning tablet (3, 1.5 or 0.75g/kg body weight/day) was intragastrically (i.g.) given to experimental rats for seven days before they were treated with ANIT (60mg/kg daily via i.g.) which caused liver injury. Serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), γ-glutamyltranspeptidase (γ-GTP), total bilirubin (T-Bil), direct bilirubin (D-Bil), total bile acid (TBA) and bile flow were measured to evaluate the protective effect of Danning tablet at 48h after ANIT treatment. Furthermore, protective mechanisms of Danning tablet against ANIT-induced liver injury were elucidated by assays of liver enzyme activities and component contents including myeloperoxidase (MPO), superoxide dismutase (SOD), glutathione peroxidase (Gpx), catalase (CAT) and glutathione S-transferase (GST), as well as liver lipid peroxide (LPO) and glutathione (GSH). The biochemical observations were supplemented by histopathological examination. Phytochemical analysis of Danning tablet was performed by UPLC-MASS. RESULTS: Obtained results demonstrated that high dose (3g/kg) of Danning tablet significantly prevented ANIT-induced changes in bile flow (P<0.01), and serum levels of ALT, AST, ALP, γ-GTP, T-Bil, D-Bil (P<0.01) and TBA (P<0.05). In addition, ANIT-induced increases in hepatic MPO, GST activities and GSH, LPO contents were significantly (P<0.01) reduced, while SOD, Gpx, CAT activities in the liver tissue which were suppressed by ANIT were significantly (P<0.01) elevated in the groups pretreated with Danning tablet at the dose of 3g/kg B.W. Histopathology of the liver tissue showed that pathological injuries were relieved after Danning tablet (3g/kg) pretreatment. The results also showed that medium dose (1.5g/kg) of Danning tablet exhibited partially protective effect on ANIT-induced liver injury with cholestasis by reversing part of biochemical parameters and histopathological changes. Low dose (0.75g/kg) of Danning tablet did not show any protective effect on ANIT-induced liver injury with cholestasis. Phytochemical analyses revealed the presence of anthraquinones, flavonoids and stilbene in the Danning tablet. CONCLUSION: These findings indicate that Danning tablet exerts a dose-dependently protective effect on ANIT-induced liver injury with cholestasis in rats, and the possible mechanism of this activity is likely due to its attenuation of oxidative stress in the liver tissue and neutrophil infiltration.