[Chemical constituents and mechanism of Chuanzhi Tongluo Capsules based on UPLC-Q-Exactive Orbitrap-MS and network pharmacology].

The chemical constituents of Chuanzhi Tongluo Capsules were analyzed and identified using ultra-high performance liquid chromatography-quadrupole/electrostatic field orbitrap high-resolution mass spectrometry(UPLC-Q-Exactive Orbitrap-MS) to clarify the pharmacological substance basis. In addition, network pharmacology was employed to explore the mechanism of Chuanzhi Tongluo Capsules in the treatment of cerebral infarction. Gradient elution was performed using acetonitrile and 1% acetic acid in water as the mobile phase. Mass spectrometry was performed in positive and negative ion modes. Xcalibur 4.2 software was used for compound analysis, including accurate mass-to-charge ratio and MS/MS fragment information, combined with the comparison of reference standards and literature data. A total of 152 compounds were identified, including 32 organic acids, 35 flavonoids and their glycosides, 33 diterpenes, 13 phthalides, 12 triterpenes and triterpene saponins, 23 nitrogen-containing compounds, and 4 other compounds, and their fragmentation patterns were analyzed. SwissTargetPrediction, GeneCards, DAVID, and other databases were used to predict and analyze the core targets and mechanism of Chuanzhi Tongluo Capsules. Protein-protein interaction(PPI) network topology analysis identified 10 core targets, including TNF, VEGFA, EGFR, IL1B, and CTNNB1. KEGG enrichment analysis showed that Chuanzhi Tongluo Capsules mainly exerted their effects through the regulation of lipid and atherosclerosis, glycoproteins in cancer, MicroRNAs in cancer, fluid shear stress, and atherosclerosis-related pathways. Molecular docking was performed between the key constituents and core targets, and the results demonstrated a strong binding affinity between the key constituents of Chuanzhi Tongluo Capsules and the core targets. This study comprehensively elucidated the chemical constituents of Chuanzhi Tongluo Capsules and explored the core targets and mechanism in the treatment of cerebral infarction based on network pharmacology, providing a scientific reference for the study of the pharmacological substance basis and formulation quality standards of Chuanzhi Tongluo Capsules.

[Proteomic study of Jingfang Mixture on urticaria based on label-free quantitative proteomics technology].

This study aims to explore the effect of Jingfang Mixture on the protein expression of urticaria in mice and explain the mechanism of Jingfang Mixture in the treatment of urticaria. Twenty-seven male Kunming mice were randomly divided into a normal group, a model group and a Jingfang Mixture group according to body weight. Except for the normal group, mice in the model group and the Jingfang Mixture group were injected with the mixture of ovalbumin and Al(OH)_3 gel for the first immunization, and the second immunization was performed on the 10 th day to induce the urticaria model. Mice in the Jingfang Mixture group started to be administered on the 6 th day after the initial immunization, and was administered continuously for 21 days. The normal group and the model group were replaced with the same amount of purified water. Twenty-four hours after the last administration, an appropriate amount of skin was taken, and label-free quantitative proteomics technology was used to detect the differences in protein expression in skin tissue. The signaling pathways involved in the differential proteins was further analyzed. The results of proteomics indicated that seventy-six proteins were involved in the intervention of Jingfang Mixture on mice with urticaria, and the differential proteins were mainly enriched in biological process(BP), molecular function(MF), and cellular component(CC). Kyoto Encyclopedia of Genes and Genomes(KEGG) analysis showed that the signaling pathways regulated by Jingfang Mixture mainly involved carbon metabolism, metabolic pathways, glucagon signaling pathway, glycolysis/gluconeogenesis, pentose phosphate pathway, hypoxia inducible factor-1(HIF-1) signaling pathway, purine metabolism, adherens junction, calcium signaling pathway, leukocyte transendothelial migration, and inflammatory mediator regulation of transient receptor potential(TRP) channels, which were involved in skin tissue energy metabolism and immune regulation. The findings of this study showed that the protective effect of Jingfang Mixture on mice with urticaria was closely related to the regulation of immune disorders, and the regulatory effect on immune system may be achieved through the regulation of energy metabolism by Jingfang Mixture.

[Critical quality attribute assessment of big brand traditional Chinese medicine: online NIR quality control research on boiling time during extraction process].

Assessment of the status property(boiling time) is a challenge for the quality control of extraction process in pharmaceutical enterprises. In this study, the pilot extraction process of Phellodendron chinense was used as the research carrier to develop an online near-infrared(NIR) quality control method based on the status property(boiling time). First, the NIR spectra of P. chinense were collected during the two pilot-scale extraction processes, and the status property(boiling time) was assessed by observing the state of bubbles in the extraction tank using a transparent window during the extraction process, which was then used as a reference standard. Based on the moving block standard deviation(MBSD) algorithm, the assessment model using online NIR spectra for boiling time during extraction process was established. In addition, the model was optimized as follows: standard normal variable(SNV) for spectral pretreatment, modeling band of 800-2 200 nm, and window size of 4. The results showed that, with 0.002 0 as the MBSD model threshold, the boiling time can be accurately assessed using online NIR spectra during extraction process. Furthermore, the principal component analysis-moving block standard deviation(PCA-MBSD) model was developed by our group to reduce the influence of online NIR spectral noise and background signal on the model, and the number of principal components was optimized into 2 in the PCA-MBSD model. The results showed that, with 0.000 075 as the PCA-MBSD model threshold, the boiling time can be accurately assessed using online NIR spectra during extraction process, with improved reliability. This study can provide a assessment method for boiling time during extraction process using online NIR spectra, which can replace the empirical judgment in manual observation, and realize the digitalization of the extraction process for big brand traditional Chinese medicine.

[Critical quality attribute assessment of big brand traditional Chinese medicine: quality control method of ginkgo leaves extract material based on powder physical properties].

The physical properties of ginkgo leaves extract(GLE) are the critical quality attributes for the control of the manufacturing process of ginkgo leaves preparations. In this study, 53 batches of GLE with different sources from the real world were used as the objects to carry out the research from 3 levels. First, based on micromeritics evaluation method, a total of 29 physical attribute quality parameters in five dimensions were comprehensively characterized, with a total of 1 537 data points. Further, with use of physical fingerprinting technology combined with similarity evaluation, the powder physical properties of 53 batches of GLE showed obvious differences from an overall perspective, and the similarity of the physical fingerprints was 0.876 to 1.000. Secondly, hierarchical clustering analysis(HCA) and principal component analysis(PCA) models were constructed to realize the reliable identification and differentiation of real-world materials produced by GLE from different sources. Multivariate statistical process control(MSPC) model was used to create GLE material Hotelling T~2 and squared prediction error(SPE) control charts. It was found that the SPE score of B_(21) powder exceeded the 99% confidence control limit by 22.495 9, and the SPE scores of A_1 and C_(10) powder exceeded the 95% confidence control limit by 16.099 2, realizing the determination of abnormal samples in the materials of GLE from the production in real world. Finally, the physical quality control method of GLE in the production process of ginkgo leaves preparations was established in this study, providing a reference for the quality control methods of ginkgo leaves preparations in their manufacturing process.

[A new quinoline alkaloid from Scolopendra subspinipes mutilans].

Three compounds, including scolosprine C(1), uracil(2) and hypoxanthine(3), were isolated and purified from the ethyl acetate fraction of centipede by silica gel normal-phase column chromatography, reversed-phase medium pressure preparation chromatography, and high-pressure semi-preparative HPLC. The structure was elucidated through a combination of spectroscopic analyses [such as nuclear magnetic resonance(NMR) and mass spectrometry(MS)] and literature review. Among them, compound 1 was a new quinoline alkaloid. In previous reports, we have described the isolation and structure elucidation of one new and two known quinoline alkaloids. In this paper, we would report the isolation and structure elucidation of scolosprine C in detail.

[Study on therapeutic effect of Chaiyin Particles on combining disease with syndrome of human coronavirus pneumonia with pestilence attacking lung syndrome based on regulation of immune function].

According to the classification of traditional Chinese medicine syndromes of coronavirus disease 2019 by the national competent authority, this study determined that human coronavirus 229 E(HCoV-229 E) was infected in a mouse model of cold and dampness syndrome, so as to build the human coronavirus pneumonia with pestilence attacking lung syndrome model. The model can simulate the traditional Chinese medicine treatment of common disease syndromes in Coronavirus Disease 2019 Diagnosis and Treatment Program(the sixth edition for trial). Specific steps were as follows. ABALB/c mouse model of cold and dampness syndrome was established, based on which, HCoV-229 E virus was infected; then the experiment was divided into normal control group, infection control group, cold-dampness control group, cold-dampness infection group(the model group), high-dose Chaiyin Particles group(8.8 g·kg~(-1)·d~(-1)), and low-dose Chaiyin Particles group(4.4 g·kg~(-1)·d~(-1)). On the day of infection, Chaiyin Particles was given for three consecutive days. Lung tissues were collected the day after the last dose, and the lung index and inhibition rate were calculated. The nucleic acid of lung tissue was extracted, and the HCoV-229 E virus load was detected by Real-time fluorescent quantitative RT-PCR. Blood leukocytes were separated, and the percentage of T and B lymphocytes was detected by flow cytometry. Lung tissue protein was extracted, and IL-6, IL-10, TNF-α and IFN-γ contents were detected by ELISA. High and low-dose Chaiyin Particles significantly reduced the lung index(P<0.01) of mice of human coronavirus pneumonia with pestilence attacking the lung syndrome, and the inhibition rates were 61.02% and 55.45%, respectively. Compared with the model control group, high and low-dose Chaiyin Particles significantly increased cross blood CD4~+ T lymphocytes, CD8~+T lymphocytes and total B lymphocyte percentage(P<0.05, P<0.01), and reduced IL-10, TNF-α and IFN-γ levels in lungs(P<0.01). In vitro results showed that TC_(50), TC_0, IC_(50) and TI of Chaiyin Particles were 4.46 mg·mL~(-1), 3.13 mg·mL~(-1), 1.12 mg·mL~(-1) and 4. The control group of in vitro culture cells had no HCoV-229 E virus nucleic acid expression. The expression of HCoV-229 E virus nucleic acid in the virus control group was 1.48×10~7 copies/mL, and Chaiyin Particles significantly reduced HCoV-229 E expression at doses of 3.13 and 1.56 mg·mL~(-1), and the expression of HCoV-229 E nucleic acid was 9.47×10~5 and 9.47×10~6 copies/mL, respectively. Chaiyin Particles has a better effect on the mouse model with human coronavirus pneumonia with pestilence attacking the lung syndrome, and could play a role by enhancing immunity, and reducing inflammatory factor expression.

Inhibitory effect of arctigenin on lymphocyte activation stimulated with PMA/ionomycin.

This study investigated the effect of arctigenin (Arc) on the cell activation, cytokines expression, proliferation, and cell-cycle distribution of mouse T lymphocytes. Mouse lymphocytes were prepared from lymph node and treated with Phorbol-12-myristate-13-acetate (PMA)/Ionimycin (Ion) and/or Arc. CD69, CD25, cytokines, proliferation and cell cycle were assayed by flow cytometry. The results showed that, at concentrations of less than 1.00 micromol x L(-1), Arc expressed non-obvious cell damage to cultured lymphocytes, however, it could significantly down-regulate the expression of CD69 and CD25, as well as TNF-alpha, IFN-gamma, IL-2, IL-4, IL-6 and IL-10 on PMA/Ion stimulated lymphocytes. At the same time, Arc could also inhibit the proliferation of PMA/Ion-activated lymphocytes and exhibited lymphocyte G 0/G1 phase cycle arrest. These results suggest that Arc possesses significant anti-inflammatory effects that may be mediated through the regulation of cell activation, cytokines expression and cell proliferation.

Quantification of binding capacity of natural products to target proteins by sensors integrating SERS labeling and photocrosslinked molecular probes.

Natural products-based screening of active ingredients and their interactions with target proteins is an important ways to discover new drugs. Assessing the binding capacity of target proteins, particularly when multiple components are involved, presents a significant challenge for sensors. As far as we know, there is currently no sensor that can accomplish high-throughput quantitative analysis of natural product-target protein binding capacity based on Raman spectroscopy. In this study, a novel sensor model has been developed for the quantitative analysis of binding capacity based on Surface-Enhanced Raman Spectroscopy (SERS) and Photocrosslinked Molecular Probe (PCMP) technology. This sensor, named SERS-PCMP, leverages the high throughput of molecular probe technology to investigate the active ingredients in natural products, along with the application of SERS labelling technology for target proteins. Thus it significantly improves the efficiency and accuracy of target protein identification. Based on the novel strategy, quantitative analysis of the binding capacity of 20 components from Shenqi Jiangtang Granules (SJG) to α-Glucosidase were completed. Ultimately, the binding capacity of these active ingredients was ranked based on the detected Raman Intensity. The compounds with higher binding capacity were Astragaloside IV (Intensity, 138.17), Ginsenoside Rh2 (Intensity, 87.46), Ginsenoside Rg3 (Intensity, 73.92) and Ginsenoside Rh1 (Intensity, 64.37), which all exceeded the binding capacity of the positive drug Acarbose (Intensity, 28.75). Furthermore, this strategy also performed a high detection sensitivity. The limit of detection for the enzyme using 0.1 mg of molecular probe magnetic nanoparticles (MP MNPs) was determined to be no less than 0.375 µg/mL. SERS-PCMP sensor integrating SERS labeling and photocrosslinked molecular probes which offers a fresh perspective for future drug discovery studies. Such as high-throughput drug screening and the exploration of small molecule-target protein interactions in vitro.

Synergistic and attenuated effects and molecular biological mechanisms of Shouhui Tongbian capsule in the treatment of slow transit constipation based on UPLC-MS/MS, network pharmacology and animal experimental validation.

BACKGROUND: Shouhui Tongbian capsule (SHTB) has been widely used for the treatment of constipation. There are few studies on SHTB at present. The current study aimed to explore the effects of multi-components compatibility of SHTB for efficacy enhancement and toxicity reduction and evaluate its molecular biological mechanisms in the treatment of slow transit constipation (STC). METHODS: Ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) was used to quantify 17 anthraquinone components in different compatible systems of SHTB. Network pharmacological analysis was used to probe the potential mechanisms of SHTB in treating STC. In addition, an animal experiment combined with western blot analysis was performed to further validate the predicted results. RESULTS: After compatibility, the dissolution of 13 components with good effects in treating constipation increased, while the dissolution of 3 components with hepatotoxicity decreased. Overall, 145 common targets of 13 synergistic components and constipation were identified. A synergistic component-target-disease network showed that chrysoobtusin, obtusifolin, emodin, obtusin and 2-hydroxyl emodin-1-methyl ether were the potential key synergistic components. A protein-protein interaction network analysis identified 91 targets, and an analysis of topological characteristics was conducted to confirm the core targets. Gene Ontology function revealed that the 13 synergistic components for the treatment of STC mainly played roles via protein phosphorylation, positive regulation of phosphorylation, phosphotransferase activity, kinase activity and protein kinase activity, and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis showed that these components were enriched in pathways in cancer, MAPK signaling pathway, IL-17 signaling pathway, NF-κB signaling pathway, etc. The results of animal experimental validation showed that SHTB significantly reduced the expression levels of p-p38 and p-ERK proteins in the colon tissue of the STC rats. CONCLUSION: This study preliminarily demonstrated that efficacy enhancement and toxicity reduction of SHTB could be achieved after compatibility, which expounded the connotation of compatibility theory of traditional Chinese medicine from the perspective of chemical composition, reflecting the rationality and scientificity of compatibility theory. Meanwhile, the study also revealed the core targets and potential molecular biological mechanisms of SHTB in the treatment of STC, which may serve as a reference for subsequent studies and clinical applications of SHTB.

Mailuo Shutong pills inhibit neuroinflammation by regulating glucose metabolism disorders to protect mice from cerebral ischemia-reperfusion injury.

ETHNOPHARMACOLOGICAL RELEVANCE: Mailuo Shutong Pill (MLST), a traditional Chinese medicine (TCM), has been widely used for clearing heat and detoxifying, eliminating stasis and dredging meridians, dispelling dampness and diminishing swelling. Earlier study found that MLST could improve cerebral ischemic-reperfusion injury, however, the potential mechanism has not been well evaluated. AIM OF STUDY: In this study, a well established and widely used mice model of middle cerebral artery occlusion/reperfusion (MCAO/R) was preformed to evaluate the protective function of MLST on cerebral ischemic-reperfusion injury and further discuss the potential pharmacological mechanisms. MATERIALS AND METHODS: Chemical profiling of MLST was analyzed based on Ultra-high-performance liquid chromatography electrospray ionization orbitrap tandem mass spectrometry. ICR mice were challenged by MCAO/R surgery. The protective effect of MLST on MCAO/R injury was evaluated by neurological deficit score, cerebral infarct rate, brain water content, H&E and nissl staining. The blood-brain barrier (BBB) integrity was detected by Evans blue staining. The potential pharmacological mechanism of MLST in treating MCAO/R injury was further elucidated by the methods of proteomics, central carbon targeted metabolomics, as well as Western blot. Immunohistochemistry was used to detect the microglia infiltration, enzyme linked immunosorbent assay (ELISA) kit was explored to evaluate the content of IL-1ß, TNF-α and IL-6 in brain tissue, and Western blot was used to detect proteins expression in brain tissue. RESULTS: A total of 76 chemical compounds have been determined in MLST. MLST effectively protected mice from MCAO/R injury, which was confirmed by lower neurological deficit score, cerebral infarct rate, brain water content and nissl body loss, and improved brain pathology. Meanwhile, MLST upregulated the expression of ZO-1, Occludin and Claudin 5 by downregulating the ratio of TIMP1/MMP9 to suppress the entrance of Evans blue to brain tissue, indicating that MLST maintained the integrity of BBB. Further studies indicated that MLST inhibited the inflammatory level of brain tissue by inhibiting microglia infiltration and downregulating NLRP3 inflammasome signaling pathway. The results of proteomics, Western blot, and central carbon targeted metabolomics confirmed that MLST regulated Glycolysis/Gluconogenesis, Pyruvate metabolism and TCA cycle in brain tissue of mice with MCAO/R. CONCLUSION: MLST inhibits neuroinflammation by regulating glucose metabolism disorders to interfere with immune metabolism reprogramming and inhibit the NLRP3 inflammasome signaling pathway, and finally improve cerebral ischemia-reperfusion injury. This study confirms that MLST is a potential drug for treating Cerebral ischemic stroke.

Integrated serum pharmacochemistry, 16S rDNA sequencing, and metabolomics to reveal the material basis and mechanism of Shouhui Tongbian capsule against diphenoxylate-induced slow transit constipation in rats.

BACKGROUND: Slow transit constipation (STC) is highly prevalent and has rising incidence. Shouhui Tongbian capsule (SHTB) is a traditional Chinese Medicine formula with extensive and highly efficacious usage in STC treatment, however, its mechanism of action, especially the regulation of microbiome and lipid metabolites, remains unclear. METHODS: After quality control of SHTB using LC‒MS to obtain its material basis, we tried to elucidate the cohesive modulatory network of SHTB against STC using hyphenated methods from microbiomics, lipidomics, mass spectrometry imaging (MSI) and molecular methods. RESULTS: SHTB could repair intestinal barrier damage, reduce systemic inflammation and increase intestinal motility in a diphenoxylate-induced STC rat model. Based on 16S rDNA sequencing results, SHTB rehabilitated the abnormal changes in Alloprevotella, Coprococcus, Marvinbryantia, etc., which were associated with STC symptoms. Meanwhile, microbial functional prediction showed that lipid metabolism was improved with SHTB administration. The differential lipids, including fatty acids, lysophosphatidylcholine, phosphatidylcholine, sphingomyelin triglyceride and ceramide, that are closely related to STC disease and SHTB efficacy. Furthermore, SHTB significantly reversed the abnormal expression of these key target enzymes in colon samples, including CTP-phosphocholine cytidylyltransferase, CTP-phosphoethanolamine cytidylyltransferase, phosphatidic acid phosphatase, acid sphingomyelinase etc. CONCLUSIONS: Combined analysis demonstrated that SHTB reducing lipid accumulation and recovery of intestinal microbial homeostasis was the critical mechanism by which SHTB treats STC.

Integration of UPLC-MS/MS-based metabolomics and desorption electrospray ionization-mass spectrometry imaging reveals that Shouhui Tongbian Capsule alleviates slow transit constipation by regulating bile acid metabolism.

Slow transit constipation (STC) is a common intestinal disorder. Some studies reported that Shouhui Tongbian Capsule (SHTB) can effectively mitigate STC symptoms. A detailed understanding of the changes in the endogenous metabolite profile of rats is crucial for a more accurate comprehension of the molecular pathological characteristics of SHTB in treating STC. In the present study, a method integrating metabolomics based on Ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) and Desorption electrospray ionization (DESI)-mass spectrometry imaging (MSI) was proposed to investigate serum, feces and colon tissue metabolic alterations of STC rats induced by diphenoxylate and the effect of SHTB treatment on metabolism. Then, Enzyme-linked immunosorbent assay (ELISA) and Western blot (WB) analysis for verifying the potential mechanism of SHTB in treating STC. As a result, we first indicated that SHTB significantly improved intestinal peristalsis and low fecal water content in STC rats. Furthermore, after treatment with SHTB, the thickness of muscle layers was increased, demonstrated SHTB's effectiveness in reducing intestinal injury in STC rats. Besides, bile acid (BA) metabolomics based on UPLC-MS/MS revealed significant increase in serum levels of Cholic acid (CA), Deoxycholic acid (DCA), Chenodeoxycholic acid (CDCA), Ursodeoxycholic acid (UDCA), and Glycolithocholic acid (GLCA), whereas the contents of CA and DCA in feces were significantly decreased in STC rats. Nonetheless, they returned to the control levels after the SHTB administration. ELISA results showed that SHTB significantly hindered the excessive reabsorption of BAs by inhibiting apical sodium-dependent bile acid transporter (ASBT), organic solute transporter alpha (OSTα) and organic solute transporter beta (OSTß) in the ileum tissue of STC rats. Furthermore, the DESI-MSI analysis revealed that SHTB remarkably enhanced DCA in the colon tissue of STC rats. The WB results indicated that SHTB reinstated Takeda G-protein-coupled receptor 5 (TGR5) expression, a receptor for BAs and a key regulator of colonic motility. Consequently, DCA exerted its effects on TGR5, leading to the promotion of colonic motility. This study provided more comprehensive and detailed information about the BA metabolomics in the serum, feces and colon of STC rats. These findings highlighted the promising potential of metabolomics based on UPLC-MS/MS and DESI-MSI method for application in the study of STC diseases.

Jingfang granules protects against intracerebral hemorrhage by inhibiting neuroinflammation and protecting blood-brain barrier damage.

Intracerebral hemorrhage (ICH) can induce intensive oxidative stress, neuroinflammation, and brain cell apoptosis. However, conventional methods for ICH treatment have many disadvantages. There is an urgent need for alternative, effective therapies with minimal side effects. Pharmacodynamics experiment, molecular docking, network pharmacology, and metabolomics were adopted to investigate the treatment and its mechanism of Jingfang Granules (JFG) in ICH. In this study, we investigated the therapeutic effects of JFG on ICH using behavioral, brain water content and Magnetic resonance imaging experiments. However, the key active component and targets of JFG remain unknown. Here we verified that JFG was beneficial to improve brain injury after ICH. A network pharmacology analysis revealed that the anti-inflammatory effect of JFG is predominantly mediated by its activation of the phosphatidylinositol 3-kinase (PI3K)/AKT pathway through Luteolin, (+)-Anomalin and Phaseol and their targeting of AKT1, tumor necrosis factorα (TNF-α), and interleukin-1ß (IL-1ß). Molecular docking analyses revealed an average affinity of -8.633 kcal/mol, indicating a binding strength of less than -5 kcal/mol. Metabolomic analysis showed that JFG exerted its therapeutic effect on ICH by regulating metabolic pathways, such as the metabolism of taurine and hypotaurine, biosynthesis of valine, leucine, and isoleucine. In conclusion, we demonstrated that JFG attenuated neuroinflammation and BBB injury subsequent to ICH by activating the PI3K/Akt signaling pathway.

Icaritin inhibits oxidative stress in murine astrocytes by binding to Orai1 to block store-operated calcium channel.

Previous study has shown that icaritin (ICT) has meaningful protective effect on cerebral ischemic stroke, and this study aimed to investigate its mechanism from the aspect of protecting astrocytes from oxidative stress. Murine primary astrocytes were pretreated by ICT and exposed to H2 O2 to induce oxidative stress. The results indicated that ICT inhibited H2 O2 -induced astrocytes apoptosis, decreased Bax and cleaved caspase-3, and increased Bcl-2. In addition, ICT inhibited H2 O2 -induced oxidative stress, increased mitochondrial membrane potential (ΔΨm ), and maintained mitochondrial morphology. ICT decreased the synthesis of malondialdehyde and increased the activity of glutathione peroxidase, catalase, and superoxide dismutase. Moreover, ICT suppressed the transient and resting intracellular Ca2+ overload. Further investigation revealed that ICT could target the combination with Orai1 to block store-operated calcium channel induced by H2 O2 . However, ICT did not enhance the protective effect of RO2959, a selective blocker of Orai1. These results indicate that ICT can play a neuroprotective role against oxidative stress injury by binding to Orai1 to block SOCC.

Jingfang Granules improve glucose metabolism disturbance and inflammation in mice with urticaria by up-regulating LKB1/AMPK/SIRT1 axis.

ETHNOPHARMACOLOGICAL RELEVANCE: Jingfang Granule (JFG) is a Traditional Chinese Medicine prescription to empirically treat skin disease such as urticaria in clinical practice. However, the potential mechanisms of JFG on urticaria are not fully defined. AIM OF STUDY: The aim of this study is to investigate the mechanisms of JFG in treating urticaria through an OVA/aluminum hydroxide induced urticaria mice model. MATERIALS AND METHODS: KM mice were injected intraperitoneally (i.p.) with OVA/aluminium hydroxide to establish the model with urticaria. After the mice were administered JFG, itching degree and hematoxylin and eosin (H&E) staining were used to assess the protective effect of JFG on mice with urticaria. The regulatory networks were investigated by proteomics and central carbon metabolomics. Spleen T lymphocyte subsets were detected by flow cytometry. Peripheral blood cytokines were detected using ELISA kits or Cytometric Bead Array (CBA) kits. The protein expression of skin tissue was detected by western blot or immunohistochemical staining. RESULTS: JFG significantly relived skin tissue lesions and skin pruritus in mice with urticaria. Meanwhile, JFG significantly decreased IgE, IL-1ß, IL-6, IL-4, TNF-α and IL-17A levels and increased IFN-γ levels in the serum of urticaria mice by inhibiting the expression of inflammation associated proteins including TLR4 and p-NF-κB p65, p-ERK1/2, p-JNK and p-p38, NLRP3, ASC and cleaved caspase-1. The results of proteomics, central carbon metabolomics, western blot and immunohistochemical staining confirmed that JFG inhibited Glycolysis/Gluconeogenesis and Pentose phosphate pathway in the skin tissue of urticaria mice by activating the LKB1/AMPK/SIRT1 axis and then downregulating the protein expressions of Glut1, TORC2, p-CREB, PEPCK, HNF4α and G6Pase. CONCLUSION: The current study demonstrates that JFG is effective in treating OVA/aluminum hydroxide-induced skin lesions and inflammation in mice, and JFG exhibits the clinical benefits via modulating LKB1/AMPK/SIRT1 axis, which in turn inhibits Glycolysis/Gluconeogenesis and Pentose phosphate pathway.

Differences in the chemical composition of Panax ginseng roots infected with red rust.

ETHNOPHARMACOLOGICAL RELEVANCE: Ginseng rusty root (GRR) is a commonly occurring disease that affects the continuous farming and economic value of mountain cultivated ginseng (MCG). Previous studies have demonstrated a generally smaller level of total ginsenoside in GRR tissue, but differences in individual ginsenosides or changes between rusty and healthy MCG with a higher age have not been investigated. AIM OF THE STUDY: This research aimed to identify differences in the chemical components in the roots of rusty compared with healthy MCG harvested at 20-years of age. MATERIALS AND METHODS: Differences between rusty and healthy MCG roots in individual ginsenosides were evaluated using a non-targeted metabonomic-based ultrahigh-performance liquid chromatography quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS) technique. Chemical markers and the principal constituents were then quantified by ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). Furthermore, total ginsenosides, total polysaccharides, and the elemental composition were evaluated separately using spectrophotometry and inductively coupled plasma optical emission spectrometer (ICP-OES). RESULTS: There was no significant difference in the levels of total ginsenosides or total polysaccharides between the rusty and healthy groups. However, the concentrations of pivotal individual ginsenosides, including ginsenoside Rc, ginsenoside Ro, and ginsenoside Rd were significantly lower in the rusty group. In addition, concentrations of Fe and Al were higher in the rusty group compared with the healthy group. CONCLUSIONS: The results suggest that GRR affects the synthesis of ginsenosides of 20-year-old MCG, which further establishes reference data and the basis for exploration of the mechanisms causing metabolic changes in ginseng resulting from GRR.

Therapeutic efficacy and safety of Yinzhihuang granules with phototherapy in neonatal pathologic jaundice: An updated systematic review and meta-analysis.

BACKGROUND: Yinzhihuang granule, consisting of extracts of Artemisia capillaris Thunb., Gardenia jasminoides Ellis, Lonicera japonica Thunb., and Scutellaria baicalensis Georgi is a well-known traditional Chinese patent medicine for patients with liver injury in China. However, the effects and safety of its use for pathologic jaundice in newborns require further systematic evaluation. PURPOSE: To systematically evaluate the efficacy and safety of Yinzhihuang granules for the treatment of neonatal pathologic jaundice and to provide clinical evidence. METHODS: Chinese databases (China Network Knowledge Infrastructure, Wan Fang Database, and VIP Database) and English databases (PubMed, EmBase, and the Cochrane Library) were thoroughly investigated through screening randomized controlled trials on Yinzhihuang granules for neonatal pathologic jaundice from the establishment of all databases to November 18, 2021. A meta-analysis was performed for selected data using STATA software. TSA software was used for trial sequential analyses of the total effective rate and adverse reactions. RESULTS: A total of 19 trials and 2,221 newborns with pathologic jaundice were included in this study. Outcome measures of clinical efficacy in the experimental group were higher than in controls, including total bilirubin (WMD = -30.34, 95% CI = -35.44 to -25.23, p < 0.001), direct bilirubin (WMD = -15.03, 95% CI = -23.54 to -6.52, p < 0.001), indirect bilirubin (WMD = -11.22, 95% CI = -17.50 to -4.95, p < 0.001), recovery time (WMD = -2.96, 95% CI = -3.92 to -2.00, p < 0.001), hospitalization time (WMD = -3.83, 95% CI = -4.89 to -2.76, p < 0.001), and liver function indices. There were statistically significant differences between the two groups. Likewise, the incidence of adverse reactions, including diarrhea, erythra, and fever decreased remarkably in the trial group (RR = 0.44, 95% CI =  0.33 to 0.59, p < 0.001). Publication bias did not exist. We verified the efficacy and safety of Yinzhihuang granules with phototherapy for pathologic jaundice in newborns according using TSA analysis. CONCLUSION: Yinzhihuang granules with phototherapy for neonatal pathologic jaundice are more effective than phototherapy alone. The incidence of ADRs does not increase with the application of Yinzhihuang granules. Due to the heterogeneity across the included studies, additional multicenter clinical trials with follow-ups are needed to confirm our findings.

Multi critical quality attributes monitoring of Chinese oral liquid extraction process with a spectral sensor fusion strategy.

The traditional Chinese medicine (TCM) extraction process is a complicated dynamic system with many variables and disturbance. Therefore, multi critical quality attributes (CQAs) monitoring is of great significance to understand the whole process. Spectroscopy is a powerful process analytical tool used for process understanding. However, single senor sometimes could not provide comprehensive information. Sensor fusion is a very practical method to overcome this deficiency. In this study, the extraction process of Xiao'er Xiaoji Zhike Oral Liquid (XXZOL) was carried out in pilot scale, where near infrared (NIR) spectroscopy and mid infrared (MIR) spectroscopy were collected to determine the concentrations of seven CQAs (synephrine, arecoline, chlorogenic acid, forsythoside A, naringin, hesperidin and neohesperidin) during extraction process. Based on fused data blocks, fusion partial least squares (PLS) models were established. Two fusion data blocks are obtained from the concatenation of original spectra (low-level data fusion) and the concatenation of characteristic variables based on band selection (mid-level data fusion) respectively. The results indicated that for all seven analytes, the mid-level data fusion models were superior to the single spectral models, with the prediction performance significantly improved. Specifically, the coefficients of determination (Rp2 and Rt2) of NIR, MIR and fusion quantitative models were all higher than 0.95. The relative standard errors of prediction (RSEP) values were all within 10%, except for models of neohesperidin, which were 10.76%, 12.39%, 12.05%, 10.03% for NIR, MIR, low-level and mid-level models respectively. These results demonstrate that it is feasible to monitor the extraction process of Xiao'er Xiaoji Zhike Oral Liquid more accurately and rapidly by fusing NIR and MIR spectroscopy, and the proposed approach also has vital and valuable reference value for the rapid monitoring of the mixed decoction process of other TCM.

Simultaneous Determination of Saponins and Lignans in Rat Plasma by UPLC- MS/MS and its Application to a Pharmacokinetic Study of Shenqi Jiangtang Granule.

BACKGROUND: Shenqi Jiangtang Granule (SJG), a classical prescription of traditional Chinese medicine, is widely used to treat diabetes and its complications. Although, the clinical efficacy of SJG, is sufficient, the pharmacokinetic behavior of various substances in the plasma of SJG is unknown. OBJECTIVE: The aim of this study was to investigate the plasma pharmacokinetics during absorption of SJG after oral administration in rats. METHODS: A rapid and accurate ultra-high performance liquid chromatography/tandem mass spectrometry (UPLC- MS/MS) method was developed for the simultaneous determination of eight analytes in SJG, including gomisin D, schisandrin A, schisandrin B, schizandrol A, schizandrol B, ginsenoside Rd, ginsenoside Re and notoginsenoside Ft1. The analysis was carried out on a BEH C18 column (2.1 mm × 50 mm, 1.7 µm) with gradient elution at a flow rate of 0.2 mL/min in a mobile phase consisting of 0.1% formic acid water and acetonitrile. In addition, lignans and saponins were detected in positive ion mode and negative ion mode, respectively. RESULTS: Eight analytes in SJG, including gomisin D, schisandrin A, schisandrin B, schizandrol A, schizandrol B, ginsenoside Rd, ginsenoside Re and notoginsenoside Ft1, showed good linearity (R2 in the range of 0.9955 ~ 0.9999). The lower limit of quantification (LLOQ) was 5, 0.8, 0.8, 8, 0.8, 5, 0.6 and 10 ng/mL. The accuracy and precision of all analytes were at ±15%. Matrix effect and average extraction recovery were > 85%. All analytes performed well under four storage conditions. CONCLUSION: The results showed that in vivo absorption and exposure of gomisin D and ginsenoside Rd were better than other analytes, while schizandrol B and notoginsenoside Ft1 were poorly absorbed. This approach could be applied to study the pharmacokinetic characteristics of various analytes in plasma after oral administration of SJG in rats.

An integrated approach to discriminate the quality markers of Traditional Chinese medicine preparation based on multi-dimensional characteristic network: Shenqi Jiangtang Granule as a case.

BACKGROUND: Due to the complexity of traditional Chinese medicine (TCM), the current quality evaluation of TCM are difficult to associate with clinical efficacy. Shenqi Jiangtang Granule (SJG), a classical TCM formula, is proven as a therapy for treatment of type II diabetes mellitus (DM) and complications while the substantial basis of the therapeutic effects is not clear. PURPOSE: The present study proposed an integrated approach to discriminate the quality markers (Q-markers) based on multi-dimensional characteristic network for quality control of TCM. METHODS: The multi-dimensional characteristic network was established by "Spider-web" mode, which was comprehensively integrating "compatibility-content-activity- efficiency-stability" of the candidate ingredients. The activity dimension was evaluated by the inhibitory activity of SJG on α-glucosidase and aldose reductase. The efficacy dimension was assessed through the association between the compounds and the target pathway of diabetic nephropathy (DN) based on integrated pharmacology platform. Each dimension for the feature network was quantified by multivariate statistical analysis, and regression area of the candidate compounds was constructed in the network. Finally, the candidate compounds were sorted comprehensively by the regression area. RESULTS: A total of 30 chemical compounds with effective hypoglycemic activity were identified as the potential Q-markers. From the data analysis, three dimensions of activity, efficacy and content performed a greater impact on the regression area of the characteristic network. Among these compounds, ginsenoside Re, ginsenoside Rd, ginsenoside Rg1, calycosin, ginsenoside Rb1, formononetin, astragaloside IV, ginsenoside Rf, ginsenoside Rc, notoginsenoside Fe, schisandrol A, gomisin D were screened out as the candidate Q-markers of SJG. CONCLUSION: The multi-dimensional characteristic network integrating compatibility, content, activity, efficiency and stability is efficient to discriminate the potential Q-markers of TCM prescription. Our results demonstrated that 12 candidate compounds from Panax Ginseng, Radix Astragali and Schisandrae Chinensis might select as Q-markers for qualitative evaluation of SJG.