Can the heptapeptide ASSIVSF of the ß2-adrenoceptor recognize ephedrine and pseudoephedrine epimers in a complex system?

Epimer separation is crucial in the field of analytical chemistry, separation science, and the pharmaceutical industry. No reported methods could separate simultaneously epimers or even isomers and remove other unwanted, co-existing, interfering substances from complex systems like herbal extracts. Herein, we prepared a heptapeptide-modified stationary phase for the separation of 1R,2S-(-)-ephedrine [(-)-Ephe] and 1S,2S-(+)-pseudoephedrine [(+)-Pse] epimers from Ephedra sinica Stapf extract and blood samples. The heptapeptide stationary phase was comprehensively characterized by scanning electron microscopy, X-ray photoelectron spectroscopy, and Fourier transform infrared spectroscopy. The separation efficiency of the heptapeptide column was compared with an affinity column packed with full-length ß2-AR functionalized silica gel (ß2-AR column). The binding affinity of the heptapeptide with (+)-Pse was 3-fold greater than that with (-)-Ephe. Their binding mechanisms were extensively characterized by chromatographic analysis, ultraviolet spectra, circular dichroism analysis, isothermal titration calorimetry, and molecule docking. An enhanced hydrogen bonding was clearly observed in the heptapeptide-(+)-Pse complex. Such results demonstrated that the heptapeptide can recognize (+)-Pse and (-)-Ephe epimers in a complex system. This work, we believe, was the first report to simultaneously separate epimers and remove non-specific interfering substances from complex samples. The method was potentially applicable to more challenging sample separation, such as chiral separation from complex systems.

Flavonoids and phenols from the stems of Ephedra equisetina.

Twelve undescribed compounds, including five flavonoids and seven phenols, were isolated from the stems of Ephedra equisetina Bunge. Their structures were elucidated by spectroscopic methods, including NMR spectroscopy and HRESIMS analysis. Their absolute configurations were elucidated by comparing their experimental and calculated ECD spectra. In the in vitro bioactive assay, all compounds were tested for their anti-asthmatic activities by releasing ß-Hex in C48/80-induced RBL-2H3 cells. The ß-Hex release rates of compounds 3, 8, 10, and 11 were 0.8502 ± 0.0231, 0.8802 ± 0.0805, 0.7850 ± 0.0593, and 0.8361 ± 0.0728, respectively, suggesting that compounds 3, 8, 10, and 11 have potential anti-asthmatic activities.

A comprehensive study of Ephedra sinica Stapf-Schisandra chinensis (Turcz.) Baill herb pair on airway protection in asthma.

ETHNOPHARMACOLOGICAL RELEVANCE: Ephedra sinica Stapf (Mahuang) and Schisandra chinensis (Turcz.) Baill (Wuweizi) are commonly utilized in traditional Chinese medicine for the treatment of cough and asthma. The synergistic effect of Mahuang-Wuweizi herb pair enhances their efficacy in alleviating respiratory symptoms, making them extensively employed in the management of respiratory disorders. Although previous studies have demonstrated the therapeutic potential of Mahuang-Wuweizi in pulmonary fibrosis, the precise mechanism underlying their effectiveness against asthma remains elusive. AIM OF THE STUDY: The objective of this study is to investigate the mechanism underlying the preventive and therapeutic effects of Mahuang-Wuweizi herb pair on asthma progression, focusing on airway inflammation and airway remodeling. MATERIALS AND METHODS: The active constituents and potential mechanisms of Mahuang-Wuweizi in the management of asthma were elucidated through network pharmacology analysis. Liquid chromatography tandem mass spectrometry (LC-MS/MS) was used to detect the main components of Mahuang-Wuweizi decoction. A rat model of bronchial asthma was established, and the effects of Mahuang-Wuweizi were investigated using hematoxylin-eosin (HE) staining, immunohistochemistry (IHC) staining, enzyme-linked immunosorbent assay (ELISA), Western blotting (WB), and real-time reverse transcription polymerase chain reaction (RT-qPCR). RESULTS: The results of network pharmacological prediction showed that Mahuang had 22 active components and Wuweizi had 8 active components, with 225 potential targets. 1159 targets associated with asthma and 115 targets that overlap between drugs and diseases were identified. These include interleukin-6 (IL-6), tumor necrosis factor (TNF), Tumor Protein 53, interleukin-1ß (IL-1ß), as well as other essential targets. Additionally, there is a potential correlation between asthma and Phosphatidylinositol 3 kinase (PI3K)/Protein Kinase B (AKT) signaling pathway, calcium ion channels, nuclear factor-kappa B (NF-κB) signaling pathway, and other signaling pathways. The animal experiment results demonstrated that treatment with Mahuang and Wuweizi, in comparison to the model group, exhibited improvements in lung tissue pathological injury, reduction in collagen fiber accumulation around the airway and proliferation of airway smooth muscle, decrease in concentration levels of IL-6, TNF-α and IL-1ß in lung tissue, as well as alleviation of airway inflammation. Furthermore, Mahuang and Wuweizi suppressed the expression of phospholipase C (PLC), transient receptor potential channel 1 (TRPC1), myosin light chain kinase (MLCK), NF-κB P65 protein in ovalbumin (OVA)-sensitized rat lung tissue and downregulated the mRNA expression of PLC, TRPC1, PI3K, AKT, NF-κB P65 in asthmatic rats. These findings were consistent with network pharmacological analysis. CONCLUSION: The results show that the synergistic interaction between Mahuang and Wuweizi occur, and they can effectively reduce airway remodeling and airway inflammation induced by inhaling OVA in bronchial asthma rats by inhibiting the expression of PLC/TRPC1/PI3K/AKT/NF-κB signaling pathway. Therefore, Mahuang and Wuweizi may be potential drugs to treat asthma.

[Mahuang (herbaceous stem of Ephedra spp.): chemistry, pharmacodynamics, and pharmacokinetics].

The Chinese medicinal herb Mahuang is herbaceous stem of Ephedra sinica, E. intermedia, or E. equisetina(Family, Ephedraceae). In China, Mahuang has been used, all the way over a millennium, as a key component herb of many herbal medicines for management of epidemics of acute respiratory illness and is also used in officially recommended herbal medicines for COVID-19. Mahuang is the first-line medicinal herb for cold and wheezing and also an effective diuretic herb for edema. However, Mahuang can also exert significant adverse effects. The key to safety and effectiveness is rational and precise use of the herb. In this review article, we comprehensively summarize chemical composition of Mahuang and associated differences in pharmacognosy, pharmacodynamics and pharmacokinetics of Mahuang compounds, along with the adverse effects of Mahuang compounds and products. Based on full understanding of how Mahuang is used in Chinese traditional medicine, systematic research on Mahuang in line with contemporary standards of pharmaceutical sciences will facilitate promoting Chinese herbal medicines to become more efficient in management of epidemic illnesses, such as COVID-19. To this end, we recommend research on Mahuang of two aspects, i.e., pharmacological investigation for its multicompound-involved therapeutic effects and toxicological investigation for clinical manifestation of the adverse effects, chemicals responsible for the adverse effects, and conditions for safe use of the herb and the herb-containing medicines.

Combination of Ephedra sinica stems and Terminalia chebula fruits produces new ephedrine derivatives in vivo that diminish the permeability to BBB while retaining airway dilation and hepatoprotective effects.

ETHNOPHARMACOLOGICAL RELEVANCE: The stems of Ephedra sinica and the fruits of Terminalia chebula are combined using in traditional Mongolian medicine formula "Gurigumu-7" for liver diseases. E. sinica stems contains ephedrine with broncho-dilatory activity. However, ephedrine can pass through the blood-brain barrier (BBB) and excite the central nervous system (CNS) to cause insomnia and restlessness. AIM OF THE STUDY: The present study was to investigate the structures and bioactivities of new compounds formed in vivo after co-administration of E. sinica stems and T. chebula fruits. MATERIALS AND METHODS: Pharmacokinetic investigation was carried out in rats. A parallel artificial membrane permeability measurement system was used to determine BBB permeability. Ex vivo experiments using tracheal rings of guinea pig was performed to examine the tracheal relaxation effect. In vivo hepatoprotective tests were carried out in Tg (fabp10a: dsRed) liver transgenic zebrafish. The fluorescent probe, 2,7-dichlorodihydrofluorescein diacetate, was used to measure reactive oxygen species, and UHPLC-MS was used to determine glutathione concentrations after derivatization with N-ethylmaleimide. RESULTS: New ephedrine derivatives (1 and 2) formed in vivo and reached their maximum serum concentrations at 0.5 h after administration of the two herbal drugs. Compounds 1 and 2 showed lower BBB permeability than ephedrine, suggesting that they have less adverse effects on the CNS. Compounds 1 and 2 relaxed the tracheal rings and had strong hepatoprotective effect on transgenic zebrafish with liver specific expression of RFP. Compounds 1 and 2 significantly reduced the level of reactive oxygen species while increasing that of glutathione in thioacetamide-treated zebrafish, which might be the hepatoprotective mechanism. CONCLUSION: These results provided evidences that the chemical constituents in various herbal drugs in a medicinal formula can interact to generate new compounds with fewer side effects and increased or additive bioactivity.

Stability and Reproducibility of Ephedra sinica Ephedrine Alkaloid Content and Terrestrial Stem Dry Weight.

Dried terrestrial stems of Ephedra sinica are known as 'Ephedra herb.' The pharmacological effects are mainly related to two major ingredients, (-)-ephedrine and (+)-pseudoephedrine (total alkaloids which are defined in Japanese Pharmacopoeia, TA). In this study, in order to aid in cultivation and breeding, the stability of TA content and stem dry weight of 46 E. sinica genets was evaluated from the first year of transplantation to the sixth year. TA content and composition ratio of these genets were stable after the second year, and dry weight was stable after the fourth year. These traits showed high inter-genet variability but low annual variability for each genet. Additionally, rank correlation coefficients of each trait among the genets were high. There was no significant correlation between these traits. Furthermore, to assess the reproducibility of these traits in clones, we evaluated TA content and dry weight of three clonal lines with high TA contents. TA content and composition ratio of the clonal lines were also stable after the second year of transplantation, and dry weight of the clonal lines was also stable after the fourth year. Moreover, TA content and composition ratio in each clonal line were comparable with those of each original genet after the second year. These results suggested that ephedrine alkaloids content and dry weight of E. sinica plants are stable, and that these traits are highly reproducible in clones. Therefore, selection breeding of E. sinica using vegetative propagation can be effective for high and stable quality of Ephedra herb.

Non-ephedrine constituents from the herbaceous stems of Ephedra sinica.

Three new flavonoids, ephedroside A (1), ephedroside B (2), ephedroside C (3), together with fifty-four known compounds 4-57 were isolated from the EtOH extract of the herbaceous stems of Ephedra sinica. The structures of these compounds were elucidated by spectroscopic techniques, as well as by comparison with literature data. Thirty-eight of these compounds were isolated from the genus Ephedra for the first time. The antimicrobial activities of eight compounds were tested by measuring the minimum inhibitory concentrations (MIC) against bacteria (both Gram positive and Gram negative) and fungi, and were found to be in the range of 0.105-0.926 mM. Among them, compound 2 showed the best antimicrobial activity against Pseudomonas aeruginosa with MIC value of 0.105 mM.

Therapeutic effects of Gambi-jung for the treatment of obesity.

Obesity is known as metabolic syndrome and it affects many tissues including adipose tissue, liver, and central nervous system (CVS). Gambi-jung (GBJ) is a modified prescription of Taeumjowi-tang (TJT), which has been used to treat obesity in Korea. GBJ is composed of 90% Ephedra sinica Stapf (ES). Therefore, the present study was designed to assess the antiobesity effects of GBJ and to compare the effects of GBJ and ES on obesity. GBJ administration remarkably reduced the body weight, Body mass index (BMI), and body fat percentage compared to the ES administration in human subjects. GBJ-treated mice had lower white adipose tissue (WAT) amounts than ES-treated mice. GBJ and ES administration enhanced adenosine monophosphate-activated protein kinase (AMPK) expression in 3T3-L1 adipocytes, epididymal WAT and liver of HFD-induced obese mice. Moreover, GBJ and ES reduced food intake by suppressing the mRNA levels of orexigenic peptides, agouti-related protein (AgRP) and neuropeptide-Y (NPY), as well as AMPK in the brain of HFD-induced obese mice. Furthermore, GBJ-treated mice had dramatically lower expression of macrophage marker F4/80 in epididymal WAT than those of ES-treated mice. Based on these results, we suggest the use of GBJ as a natural drug to control weight gain.

Active components in Ephedra sinica stapf disrupt the interaction between ACE2 and SARS-CoV-2 RBD: Potent COVID-19 therapeutic agents.

ETHNOPHARMACOLOGICAL RELEVANCE: Ephedra sinica Stapf is a widely used folk medicine in Asia to treat lung diseases, such as cold, cough and asthma. Many efforts have revealed that some traditional Chinese medicine (TCM) prescriptions containing Ephedra sinica could effectively alleviate the symptoms and prevent the fatal deterioration of COVID-19. AIM OF THE STUDY: The present study aims to discover active compounds in Ephedra sinica disrupting the interaction between angiotensin-converting enzyme 2 (ACE2) and the SARS-CoV-2 spike protein receptor-binding domain (SARS-CoV-2 RBD) to inhibit SARS-CoV-2 virus infection. MATERIALS AND METHODS: The ethanol extracts of Ephedra sinica were prepared. Activity guided isolation of constituents was carried out by measuring the inhibitory activity on ACE2-RBD interaction. The structures of active compounds were identified by HPLC-Q-TOF-MS/MS and NMR. To testify the contribution of main components for the inhibitory activity, different samples were prepared by components knock-out strategy. The mechanism of compounds inhibiting protein-protein interaction (PPI) was explored by competition inhibition assays, surface plasmon resonance (SPR) assays and molecular docking. SARS-CoV-2 S protein-pseudoviruses were used to observe the viropexis effect in cells. RESULTS: Ephedra sinica extracts (ESE) could effectively inhibit the interaction between ACE2 and SARS-CoV-2 RBD (IC50 = 95.01 µg/mL). Three active compounds, 4,6-dihydroxyquinoline-2-carboxylic acid, 4-hydroxyquinoline-2-carboxylic acid and 4-hydroxy-6-methoxyquinoline-2-carboxylic acid were identified to inhibit ACE2-RBD interaction (IC50 = 0.58 µM, 0.07 µM and 0.15 µM respectively). And knock-out the three components could eliminate the inhibitory activity of ESE. Molecular docking calculations indicated that the hydrogen bond was the major intermolecular force. Finally, our results also showed that these compounds could inhibit the infectivity of SARS-CoV-2 S protein-pseudoviruses to 293T-ACE2 (IC50 = 0.44-1.09 µM) and Calu-3 cells. CONCLUSION: These findings suggested that quinoline-2-carboxylic acids in Ephedra sinica could be considered as potential therapeutic agents for COVID-19. Further, this study provided some justification for the ethnomedicinal use of Ephedra sinica for COVID-19.

Screening and evaluation of anti-SARS-CoV-2 components from Ephedra sinica by ACE2/CMC-HPLC-IT-TOF-MS approach.

Traditional Chinese medicines played an important role in the treatment of COVID-19 in 2020. Ephedra sinica, one of the major constituent herbs of multi-component herbal formula, has been widely used to treat COVID-19 in China. However, its active components are still unclear. The objectives of this study are to screen and evaluate active components from the traditional Chinese medicine Ephedra sinica for the treatment of COVID-19. In our study, we established an ACE2/CMC bioaffinity chromatography model, and then developed an ACE2/CMC-HPLC-IT-TOF-MS system for the active compounds screening and identification from Ephedra sinica extract. We performed molecular docking and surface plasmon resonance (SPR) assays to assess the binding characteristics (binding mode and KD value). We used CCK-8 staining to assess the toxicity of screened compounds, and also used SARS-CoV-2 pseudovirus to observe the viropexis effect of screened compounds in ACE2h cells. In this current work, one fraction was fished out, separated and identified as ephedrine (EP), pseudoephedrine (PEP), and methylephedrine (MEP). Binding assays showed that the three compounds could bind with ACE2 in a special way to some amino acid residues, similar to the way SARS-CoV-2 bound with ACE2. Additionally, the three compounds, especially EP, can inhibit the entrance of SARS-CoV-2 spike pseudovirus into ACE2h cells because they can reduce the entrance ratio of pseudovirus in the pseudovirus model. Overall, the ACE2/CMC-HPLC-IT-TOF-MS system was established and verified to be suitable for ACE2-targeted bioactive compound screening. EP, PEP, and MEP with ACE2-binding features were screened out from Ephedra sinica, and acted as blockers inhibiting SARS-CoV-2 spike pseudovirus entering ACE2h cells.

Network Pharmacology-Based Analysis on the Mechanism of Action of Ephedrae Herba-Cinnamomi Ramulus Couplet Medicines in the Treatment for Psoriasis.

BACKGROUND This study explored the mechanism of action of Ephedrae Herba-Cinnamomi Ramulus couplet medicine (MGCM) at the pharmacological level in the treatment of psoriasis. MATERIAL AND METHODS The active ingredients in MGCM were mined through literature retrieval and the BATMAN-TCM database, and potential targets were predicted. In addition, targets associated with psoriasis were acquired using multiple disease-related databases. Thereafter, an interaction network between candidate MGCM targets and the known psoriasis-associated targets was constructed based on the protein-protein interaction (PPI) data, using the STRING database. Then, the topological parameter degree was determined for mining the core targets for MGCM in the treatment of psoriasis, which also represented the major hubs within the PPI network. In addition, the core networks of targets and ingredients were constructed using Cytoscape software to apply MGCM in the treatment for psoriasis. These core targets were then analyzed for Gene Ontology biological processes and Kyoto Encyclopedia of Genes and Genomes pathway enrichment using OmicShare. RESULTS The ingredient-target core network of MGCM for treating psoriasis was constructed; it contained 52 active ingredients and corresponded to 19 core targets. In addition, based on enrichment analysis, these core targets were majorly enriched for several biological processes (immuno-inflammatory responses, leukocyte differentiation, energy metabolism, angiogenesis, and programmed cell death) together with the relevant pathways (Janus kinase-signal transducer and activator of transcription, toll-like receptors, nuclear factor kappaB, vascular endothelial growth factor, and peroxisome proliferator-activated receptor), thus identifying the possible mechanism of action of MGCM in treating psoriasis. CONCLUSIONS The present network pharmacology study indicated that MGCM alleviates various pathological factors of psoriasis through multiple compounds, multiple targets, and multiple pathways.

Herb pair of Ephedrae Herba-Armeniacae Semen Amarum alleviates airway injury in asthmatic rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Ephedrae Herba (EH, Ephedra sinica Stapf.) and Armeniacae Semen Amarum (ASA, Prunus armeniaca L. var. ansu Maxim.) have been used to treat asthma, cold, fever, and cough in China for thousands of years. AIM OF THE STUDY: In this study, we aimed to investigate the optimal ratio of EH and ASA compatibility (EAC) to reduce airway injury in asthmatic rats and its possible mechanism. METHODS: Rats were sensitized with a mixture of acetylcholine chloride and histamine bisphosphate 1 h before sensitization by intragastric administration of EAC or dexamethasone or saline for 7 days. Subsequently, the ultrastructure of rat airway epithelial tissue changes, apoptosis of the airway epithelial cells, and the expression of mRNA and protein of EGRF and Bcl-2 were detected. RESULTS: Transmission electron microscope: EAC (groups C and E) had the most prominent effect on repairing airway epithelial cells' ultrastructural changes in asthmatic rats. TUNEL: dexamethasone and EAC (groups B、C、E and F) inhibited the apoptosis of airway epithelial cells in asthmatic rats (P < 0.05). In situ hybridization: EAC (group E) inhibited the overexpression of EGFR and Bcl-2 mRNA (P < 0.05).Western Blotting: EAC (groups A、B、C、E and F) inhibited the upregulation of airway epithelial EGFR and Bcl-2 protein expression (P < 0.01). CONCLUSIONS: Our findings indicate that EAC can inhibit abnormal changes in airway epithelial structure and apoptosis of airway epithelial cells, thereby alleviating airway injury. In this study, the best combination of EH and ASA to alleviate airway epithelial injury in asthmatic rats was group E (EH: ASA = 8: 4.5).

A new application of acetylation for analysis of acidic heteropolysaccharides by liquid chromatography-electrospray mass spectrometry.

A novel ultra-high-performance liquid chromatography-electrospray mass spectrometry (UHPLC-ESI-MS) method was proposed for simultaneous determination of neutral and acidic monosaccharaides by employing acetylation derivatization that is specialized for gas chromatography-mass spectrometry (GC-MS) analyses. The UHPLC-ESI-MS was carried out in a positive ionization mode with selective ion monitoring (SIM) of sodium adducts at m/z 385 (Ara, Xyl and Rib), 399 (Rha and Fuc), 443 (Glc, Gal and Man) and 457 (GlcA and GalA). The separation was achieved on a Cortecs C18+ (2.1 mm × 150 mm, 1.6 µm) within 8 min using acetonitrile water as the mobile phase without additional buffer salt added. Furthermore, quantitative analysis of the multi-monosaccharaides by single marker (QAMS) was developed and validated using a relative correction factor. The UHPLC-ESI-MS and QAMS were compared and successfully applied to analyze the monosaccharide contents of a purified acidic heteropolysaccharide previously isolated from Ephedra sinica. The established UHPLC-ESI-MS and QAMS approach possesses high precision, stability and repeatability, and can be widely used for high-throughput analysis of the monosaccharide compositions of plant polysaccharides.

[Antipyretic effect of active components of Mahuang Decoction and its correlation with pharmacokinetics in febrile rats].

To investigate the antipyretic effect of active components of Mahuang Decoction in febrile rats, and explore its correlation with pharmacokinetics at different time points. The feverished rat models were induced by dry yeast, and intragastrically administered with the effective components of Mahuang Decoction with different orthogonal compatibility ratios. At different time points after administration, body temperature was measured; blood was taken from orbital vena plexus, and the contents of interleukin-6(IL-6), interleukin-1ß(IL-1ß), and tumor necrosis factor-α(TNF-α) in rat serum were determined with the kits. Combined with the pharmacokinetic data of the seven effective components in Mahuang Decoction, PK-PD(pharmacokinetics-pharmacodynamics) data fitting was conducted by using the analysis method of non-atrioventricular model, and then the pharmacodynamic parameters were calculated to determine the optimal binding model. The results showed that the effective components of Mahuang Decoction inhibited the release of heat-causing factors IL-6, IL-1ß and TNF-α, and reduced the increase of body temperature. There was a significant lag between drug effect and blood drug concentration, which was consistent with Sigmoid-E_(max) model. The model fitting value showed a good correlation with mea-sured data, which could be used to evaluate and predict the correlation between PK and PD in Mahuang Decoction, and further applied to the multiple-indicator and multiple-effect study of PK-PD in other compound traditional Chinese medicines.

Protective effect of gui zhi (Ramulus Cinnamomi) on abnormal levels of four amino acid neurotransmitters by chronically ma huang (Herb Ephedra) intoxicated prefrontal cortex in rats treated with a ma huang-gui zhi herb pair.

ETHNOPHARMACOLOGICAL RELEVANCE: The Herb Ephedra (Ma Huang in Chinese)-Ramulus Cinnamomi (Gui Zhi in Chinese) herb pair is a classic traditional Chinese herb pair used to treat asthma, nose and lung congestion, and fever with anhidrosis. In previous study, we found that chronic administration of ma huang induced obvious neurodegeneration in rat brains, with the prefrontal cortex showing the greatest effect. Gui zhi decreased hyperactivity produced by repeated ma huang administration, and attenuated oxidative stress in rat prefrontal cortex induced by ma huang. AIM OF THE STUDY: The study was aimed to investigate the protective effect of gui zhi on ma huang-induced abnormal levels of four amino acid neurotransmitters in rat prefrontal cortex. MATERIALS AND METHODS: All ma huang and ma huang-gui zhi herb pair extracts were prepared using methods of traditional Chinese medicine and were normalized based on the ephedrine content. Two-month-old male Sprague-Dawley rats (6 rats/group) were administered ma huang or ma huang-gui zhi herb pair extracts for 1, 3, 5 or 7 days (ephedrine = 48 mg/kg). The contents of ephedrine, glutamate (Glu), aspartic acid (Asp), glycine (Gly), and gamma-aminobutyric acid (GABA) in the prefrontal cortex were determined using ultra-high-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) at 0.5, 1.0, 5.0 h after administration. RESULTS: Ma huang significantly enhanced the levels of GABA, Gly, Glu and Asp in the prefrontal cortex, while gui zhi partially abolished the effects. CONCLUSIONS: Ma huang-induced neurotoxicity may be associated with its effects on amino acid neurotransmitters. Gui zhi is a promising neuroprotective agent against for ma huang-induced neurotoxicity. The information presented in this study will help supplement the available data for future ma huang-gui zhi herb pair compatibility studies.

[Pharmacokinetics of compatible effective components of Mahuang Decoction in febrile rats].

In the present paper,after the febrile rat model was prepared by injecting yeast,orthogonally compatible effective components from prescription drugs of Mahuang Decoction( Ephedra sinica total alkaloids,Cinnamomum cassia essential oil,amygdalin,Glycyrrhiza uralensis total flavonoids+G. uralensis total saponins) with nine different dosage ratios were given by gavage administration.The plasma concentrations of main active ingredients including ephedrine hydrochloride,pseudoephedrine hydrochloride,methylephedrine hydrochloride,cinnamic acid,amygdalin,liquritin and glycyrrhizin at different time points were analyzed by liquid chromatograph mass spectrometer( LC-MS). Based on the pharmacokinetic parameters of non-compartmental model,the area under curve of total quantum( AUCt) and the mean chromatographic retention time of total quantum( MRTt) were further calculated,in order to evaluate the effect of compatibility on the total statistical moment parameters. The results showed that the pharmacokinetic characteristics of main active components in febrile rats were significantly different after treatment with orthogonally compatibility of E. sinica total alkaloids,C.cassia essential oil,amygdalin,G. uralensis total flavonoids and G. uralensis total saponins. Orthogonal analysis confirmed that different compatibility components had different effects on the total statistical moment parameters. The contribution of effective components of Mahuang Decoction to AUCtwas as follows in a descending order: E. sinica total alkaloids>C. cassia essential oil>amygdalin>G. uralensis total flavonoids+G. uralensis total saponin,while the contribution to MRTtwas: E. sinica total alkaloids >G. uralensis total flavonoids+G. uralensis total saponin>amygdalin>C. cassia essential oil. The E. sinica total alkaloid had the greatest effects on both of the above parameters,and the optimal combination was A_3B_3C_2D_1 for AUCt,and A_1B_1C_1D_1 for MRTt.

Anti-neuroinflammatory effects of Ephedra sinica Stapf extract-capped gold nanoparticles in microglia.

Background: Combination therapy remains a promising strategy for treating neurodegenerative diseases, although green synthesis of gold nanoparticles for treating chronic neuroinflammation and studying their efficacy in treating neuroinflammation-mediated neurodegenerative diseases is not well assessed. Results: Here, Ephedra sinica Stapf (ES) extract was used as the reducing, capping, and stabilizing agent for gold nanoparticle synthesis. We developed ES extract-capped gold nanoparticles (ES-GNs) and investigated their anti-neuroinflammatory properties in microglia. ES-GNs displayed maximum absorption at 538 nm in ultraviolet-visible spectroscopy. Dynamic light scattering assessment revealed that ES-GN diameter was 57.6±3.07 nm, with zeta potential value of -24.6±0.84 mV. High resolution-transmission electron microscopy confirmed the spherical shape and average diameter (35.04±4.02 nm) of ES-GNs. Crystalline structure of ES-GNs in optimal conditions was determined by X-ray powder diffraction, and elemental gold presence was confirmed by energy-dispersive X-ray spectroscopy. Fourier transform-infrared spectroscopy confirmed gold nanoparticle synthesis using ES. Anti-neuroinflammatory properties of ES-GNs on production of pro-inflammatory mediators (nitric oxide, prostaglandin E2, and reactive oxygen species) and cytokines (tumor necrosis factor-α, IL-1ß, and IL-6) in lipopolysaccharide (LPS)-stimulated microglia were investigated by ELISA and flow cytometry. ES-GNs significantly attenuated LPS-induced production of pro-inflammatory mediators and cytokines, which was related to suppressed transcription and translation of inducible nitric oxide synthase and cyclooxygenase-2, determined by RT-PCR and western blotting. ES-GNs downregulated upstream signaling pathways (IκB kinase-α/ß, nuclear factor-κB, Janus-activated kinase /signal transducers and activators of transcription, mitogen-activated protein kinase , and phospholipase D) of pro-inflammatory mediators and cytokines in LPS-stimulated microglia. Anti-neuroinflammatory properties of ES-GNs were mediated by ES-GNs-induced AMP-activated protein kinase)-mediated nuclear erythroid 2-related factor 2 /antioxidant response element signaling. Conclusion: Collectively, these findings provide a new insight on the role of ES-GNs in treating chronic neuroinflammation-induced neurodegenerative diseases.

[Chinese classical formulas Ephedra associated prescriptions for treatment of skin diseases].

Chinese classical formulas, firstly recorded in Shanghan Zabing Lun by Zhang Zhongjing in the Han dynasty, have been widely used by traditional Chinese medicine physicians for thousands of years due to its good compatibility, strict structures, significant clinical efficacy and less adverse effects. Ephedra associated prescriptions, widely used in the treatment of skin diseases, are famous representative prescriptions in Chinese classical formulas. The commonly used Ephedra associated prescriptions include Mahuang Lianqiao Chixiaodou decoction, Maxing Shigan decoction, Maxing Yigan decoction, Mahuang decoction, Mahuang Jiazhu decoction, Guizhi Mahuang Geban decoction, and Guizhi Eryue Biyi decoction. Skin allergy is an allergic reaction, manifested as erythema, papules, and itching. It has been identified that Guizhi Mahuang Geban decoction (1/2 Guizhi decoction + 1/2 Mahuang decoction) is effective in dispersing wind and cold, relieving exterior syndrome and dredging the collaterals, so it can be used to treat skin allergy caused by pathogenic wind-cold. Guizhi Eryue Biyi decoction (1/4 Guizhi decoction + 1/8 Yuebi decoction) is effective in dispersing wind and cold, relieving exterior syndrome and clearing heat, so it is mainly used for the syndrome of wind-cold tightening superficies and heat transformed from wind-cold. Additionally, Mahuang Lianqiao Chixiaodou decoction can relieve exterior syndrome, clear heat and remove dampness, mainly for the dampness and heat depression in the muscle surface. It is found that Guizhi Mahuang Geban decoction and Guizhi Eryue Biyi decoction have certain efficacy for skin allergies, while Mahuang Lianqiao Chixiaodou decoction has significant efficacy for urticaria, acute and chronic eczema, etc. It has been identified that if the conditions of Ephedra associated prescriptions are well mastered by traditional Chinese medicine physicians, they could be widely applied to treat exogenous diseases and miscellaneous diseases.

Antipyretic Effect of Herba Ephedrae-Ramulus Cinnamomi Herb Pair on Yeast-Induced Pyrexia Rats: A Metabolomics Study.

OBJECTIVE: To investigate the antipyretic mechanism of Herba Ephedrae (Eph)-Ramulus Cinnamomi (RC) herb pair on yeast-induced pyrexia in rats. METHODS: Totally 30 qualified male SD rats were randomly assigned to the normal control (NC) group, the pyrexia model (model) group, the Eph, RC and Eph-RC treatment groups by a random digital table, 6 rats in each group. Each rat received a 20% aqueous suspension of yeast (10 mL/kg) except the NC group. The 3 treatment groups were administered 8.1, 5.4 and 13.5 g/kg Eph, RC and Eph-RC respectively at 5 and 12 h after yeast injection, the NC group and the model groups were administered equal volume of distilled water. Rectal temperatures were measured at 0, 6, 8, 10, 12, 15, 18, 24 and 30 h and urine was collected prior to yeast injection and at 6, 10, 18, 24, 30, and 36 h after yeast injection. Then urine metabolomic profiling by gas chromatography tandem mass spectrometry, coupled with multivariate statistical analysis and pattern recognition techniques were used to explore the antipyretic effects of Eph-RC. Partial least squares discriminate analysis was used to analyze the metabolomics dataset including classification and regression in metabolomics plot profiling. RESULTS: Compared with the NC group, rectal temperatures were significantly higher in the model group (P<0.01), while 3 treatment groups decreased significantly compared with the model group (P<0.05 or P<0.01). Rectal temperatures of Eph-RC-treated rats started to go down at 6 h, and markedly decreased at 8, 12, 15, 18 and 24 h (P<0.05 or P<0.01), while those of the Eph and RC groups had decreased firstly at 8 h and were markedly lower at 12 h (P<0.05 or P<0.01). Seventeen potential biomarkers related to pyrexia were confirmed and identified, including pyruvic acid, L-phenylalanine, L-tyrosine, phenylacetic acid, hippuric acid, succinic acid, citrate and so on. Eight potential alterations of metabolic pathways including phenylalanine metabolism, citrate cycle, tryptophan metabolism, biosynthesis of valine, leucine and isoleucine, were identified in relation to the antipyretic effects of Eph-RC using MetPA software. CONCLUSION: The antipyretic effect of Eph-RC herb pair on yeast-induced pyrexia in rats involved correction of perturbed amino acid, fatty acid, and carbohydrate metabolism according to the metabolic pathway analysis with MetPA.