Comparison of Quality Change During Processing Process of Achyranthis Bidentatae Radix from Different Origins Based on Color-component Correlation Analysis / 中国实验方剂学杂志

ObjectiveTaking Achyranthis Bidentatae Radix（ABR） from different origins as samples， to quantitatively analyze the chemical composition and chromaticity of ABR with different processing degrees， and clarify the correlation and change law between color and composition in the processing process of ABR， so as to provide reference for the quality evaluation of processed products of ABR. MethodThe colorimeter is used to measure the chromaticity values of three kinds of processing degrees of ABR in different origins to show the color value change trend during the processing process， and the color parameters of wine-processed and salt-processed products of ABR with different processing degrees were analyzed by principal component analysis（PCA）， orthogonal partial least squares-discriminant analysis（OPLS-DA） and other analysis methods. The contents of eight representative components of ABR were measured by high performance liquid chromatography（HPLC）， the correlation between chromaticity and each representative component was analyzed by Pearson correlation analysis， and the applicability of the selected eight representative components was further verified by Fisher linear discriminant analysis， and the wine-processed and salt-processed products of ABR with different processing degrees were grouped according to the degree of processing， and 48 samples of wine-processed and salt-processed products with different processing degrees were used as training samples. Taking the contents of 5-hydroxymethylfurfural， polypodine B， β-ecdysterone， 25R-inokosterone， 25S-inokosterone， ginsenoside Ro， chikusetsusaponin Ⅳa and polysaccharides as variables， the discriminant function was established respectively， and 12 samples of wine-processed and salt-processed products of ABR with different processing degrees were back-tested to verify the discriminant function and test the reliability of the function. ResultPCA and OPLS-DA results showed that ABR samples with different processing degrees were classified into clusters， and the results could significantly distinguish different processed products. During the process of wine and salt processing， the contents of 5-hydroxymethylfurfural， ginsenoside Ro， and chikusetsusaponin Ⅳa gradually increased with the deepening of the processing degree， while the contents of polypodine B， β-ecdysterone， 25R-inokosterone， 25S-inokosterone and polysaccharides showed a gradual decreasing trend， indicating these 8 components increased and decreased to different degrees in the process of wine and salt processing. The results of Pearson correlation analysis showed that the 5-hydroxymethylfurfural content of the samples with different processing degrees of wine-processed and salt-processed products were negatively correlated with the brightness value（L*） and the total color difference value（E*ab）（P<0.01）， and positively correlated with the red-green value（a*） and the yellow-blue value（b*）（P<0.01）， and that the content of polypodine B and polysaccharides were positively correlated with L* and E*ab（P<0.01）. The discriminant functions of wine-processed and salt-processed products of ABR were established by Fisher linear discriminant analysis， and their accuracy rates in the training samples were 93.75% and 95.83%， respectively. Twelve test samples of wine-processed and salt-processed products with different processing degree were back substitution， and the correct rate was 100%. ConclusionThe trend of composition and color changes of ABR with different processing degrees in different production areas is relatively consistent， and the color value can better distinguish ABR with different processing degrees， and the color of ABR is related to some representative components in the processing process， indicating that the color can provide reference for the identification of the processing degree of ABR and the prediction of component content.

Effect of Yunvjian with or Without Achyranthis Bidentatae Radix on Glucose and Lipid Metabolism and Inflammatory Response in Diabetic Rats with Syndrome of Yin Deficiency and Internal Heat / 中国实验方剂学杂志

ObjectiveTo investigate the different effects of Yunvjian with or without Achyranthis Bidentatae Radix on glucose and lipid metabolism and inflammatory response in diabetic rats with the syndrome of Yin deficiency and internal heat. MethodThe rat model of diabetes due to Yin deficiency and internal heat was established by feeding with a high-sugar and high-fat diet and injection of thyroxine and streptozotocin. The successfully modeled rats were randomized into model control， Yunvjian without Achyranthis Bidentatae Radix （11.8 g·kg-1）， Yunvjian with Achyranthis Bidentatae Radix （12.8 g·kg-1）， and Achyranthis Bidentatae Radix （1.0 g·kg-1） groups （n=10）， and another 10 rats were taken as the normal control group. Each group was administrated with corresponding drugs or saline by gavage for 28 days. The fasting blood glucose （FBG）， fasting insulin （FINS）， total cholesterol （TC）， triglycerides （TG）， low-density lipoprotein cholesterol （LDL-C）， and high-density lipoprotein cholesterol （HDL-C） in rats were measured. Enzyme-linked immunosorbent assay was employed to determine the levels of cyclic adenosine monophosphate （cAMP）， cyclic guanosine monophosphate （cGMP）， triiodothyronine （T3）， thyroxine （T4）， interleukin （IL）-1β， IL-6， tumor necrosis factor （TNF）-α， and C-reactive protein （CRP） in the serum. The histopathological changes of the liver were observed. The expression of lipoxygenase-2 （COX-2） was detected by immunofluorescence. The mRNA levels of nuclear transcription factors-κB （NF-κB）， monocyte chemotactic protein-1 （MCP-1）， and intercellular adhesion molecule-1 （ICAM-1） were determined by real-time polymerase chain reaction （Real-time PCR）.Western blot was employed to determine the protein levels of NF-κB in hibitory protein（IκB） kinase β （IKKβ）， IκBα， and phosphorylated IκBα （p-IκBα） in the liver and the protein levels of NF-κB in the cytoplasm and nucleus. ResultCompared with the normal group， the model group showed elevated levels of FBG， FINS， insulin resistance index， TC， TG， LDL-C， cAMP， T3， T4， IL-1β， IL-6， TNF-α， and CRP， up-regulated mRNA levels of NF-κB， MCP-1， and ICAM-1， and up-regulated protein levels of COX-2， p-IκBα， and nuclear NF-κB （P<0.01）. Compared with the model group， Yunvjian without Achyranthis Bidentatae Radix lowered the levels of FBG， FINS， insulin resistance index， TC， TG， LDL-C， cAMP， T3， T4， IL-1β， IL-6， TNF-α， and CRP， down-regulated the mRNA levels of NF-κB， MCP-1， and ICAM-1， and down-regulated the protein levels of COX-2， p-IκBα and nuclear NF-κB （P<0.05， P<0.01）. Compared with the Yunvjian without Achyranthis Bidentatae Radix， Yunvjian with Achyranthis Bidentatae Radix showed lowered levels of FBG， FINS， insulin resistance index， and inflammatory cytokines， down-regulated mRNA levels of NF-κB， MCP-1， and ICAM-1， and down-regulated protein levels of p-IκBα and nuclear NF-κB （P<0.05， P<0.01）. ConclusionAchyranthis Bidentatae Radix can enhance the performance of Yunvjian in reducing blood glucose and inhibiting inflammation in diabetic rats with the syndrome of yin deficiency and internal heat by down-regulating the IKK/IκB/NF-κB signaling pathway.

Pharmacokinetics and tissue distribution of four alkaloids in Ermiao Pills and Sanmiao Pills in normal and arthritic model rats / 中国中药杂志

This work aimed to investigate the differences of pharmacokinetics and tissue distribution of four alkaloids in Ermiao Pills and Sanmiao Pills in normal and arthritic model rats. The rat model of arthritis was established by injecting Freund's complete adjuvant, and ultra-high performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS) in the positive ion multiple reaction monitoring(MRM) mode was used for the determination of four alkaloids in plasma and tissues of normal and arthritic rats after administration of Ermiao Pills and Sanmiao Pills, respectively. The differences in pharmacokinetics and tissue distribution of the four active components were compared, and the effect of Achyranthis Bidentatae Radix on the major components of Sanmiao Pills was explored. This study established an UPLC-MS/MS for simultaneous determination of four alkaloids, and the specificity, linearity, accuracy, precision, and stability of this method all met the requirements. Pharmacokinetics study found that as compared with normal rats, the AUC and C_(max) of phellodendrine, magnoflorine, berberine and palmatine in model rats were significantly decreased after administration of Ermiao Pills, the clearance rate CL/F was significantly increased, and the distribution and tissue/plasma concentration ratio of the four alkaloids in the liver, kidney, and joint were significantly reduced. Achyranthis Bidentatae Radix increased the AUC of phellodendrine, berberine, and palmatine, reduced the clearance rate, and significantly increased the distribution of the four alkaloids in the liver, kidney, and joints in arthritic rats. However, it had no significant effect on the pharmacokinetics and tissue distribution of the four alkaloids in normal rats. These results suggest that Achyranthis Bidentatae Radix may play a guiding role in meridian through increasing the tissue distribution of effective components in Sanmiao Pills under arthritis states.

Effect of Achyranthis Bidentatae Radix-Paeoniae Radix Alba on Neuroinflammation and RhoA/ROCK2 Signaling Pathway in Parkinson's Disease Mouse Model / 中国实验方剂学杂志

ObjectiveTo investigate the protective effect and mechanism of Achyranthis Bidentatae Radix-Paeoniae Radix Alba on dopaminergic neurons in Parkinson's disease mouse model with the syndrome of ascendant hyperactivity of liver Yang. MethodThe C57BL/6 mice were randomly assigned into normal group， a model group， low-， medium， and high-dose （3.25， 6.5， 13 g·kg-1） Achyranthis Bidentatae Radix-Paeoniae Radix Alba groups， and a selegiline group （0.01 g·kg-1）. The mouse model of Parkinson's disease with the syndrome of ascendant hyperactivity of liver yang was established by intragastric administration of Fuzitang combined with intraperitoneal injection of 1-methyl-4-phenyl-1，2，3，6-tetrahydropyridine （MPTP）. The behavioral changes were evaluated by rotarod test and pole test. The protein levels of Ras homolog gene family member A （RhoA）， Rho-associated coiled-coil containing protein kinase 2 （ROCK2）， myosin light chain 1 （MLC1）， and α-synuclein in the substantia nigra were determined by Western blot. Real-time fluorescence quantitative PCR （Real-time PCR） was employed to determine the mRNA levels of RhoA， ROCK2， and MLC1 in the substantia nigra. Enzyme-linked immunosorbent assay （ELISA） was used to measure the levels of tumor necrosis factor-α （TNF-α） and interleukin-1β （IL-1β）. The ultrastructural changes of mouse neurons were observed under a transmission electron microscope. ResultCompared with the normal group， the modeling shortened the latency to fall， increased the average total time in the pole test （P<0.01）， and up-regulated the levels of RhoA， ROCK2， MLC1， TNF-α， α-synuclein， and IL-1β in the substantia nigra （P<0.05）. Compared with the model group， different doses of Achyranthis Bidentatae Radix-Paeoniae Radix Alba and selegiline prolonged the latency to fall， shortened the average total time in the pole test （P<0.05， P<0.01）， and down-regulated the levels of ROCK2， MLC1， α-synuclein， TNF-α， and IL-1β in a dose-dependent manner （P<0.05）. Further， the modeling decreased the number of cytoplasmic organelles and caused mitochondrial swelling and abnormal shape of endoplasmic reticulum compared with the normal group. The neurons in high-dose Achyranthis Bidentatae Radix-Paeoniae Radix Alba and selegiline groups showed intact nuclei， clear cell boundary， and normal endoplasmic reticulum shape. ConclusionThe combination of Achyranthis Bidentatae Radix and Paeoniae Radix Alba may improve the motor coordination ability of Parkinson's disease mouse model with the syndrome of ascendant hyperactivity of liver yang by inhibiting the neuroinflammation mediated by the RhoA/ROCK2 signaling pathway in the brain.

Integrated smart hyperspectral imaging and CARS-based characteristic band selection for rapid determination of SO_2 content in sulphur-fumigated Achyranthis Bidentatae Radix / 中国中药杂志

In order to realize the rapid and non-destructive detection of SO_2 content in sulphur-fumigated Achyranthis Bidentatae Radix, this paper first prepared the sulphur-fumigated Achyranthis Bidentatae Radix samples with the usage amount of sulphur being 0, 2.5%, and 5% of the mass of Achyranthis Bidentatae Radix pieces. The SO_2 content in different batches of sulphur-fumigated Achyranthis Bidentatae Radix was determined using the method in Chinese Pharmacopoeia, followed by the acquisition of their hyperspectral data within both visible-near infrared(435-1 042 nm) and short-wave infrared(898-1 751 nm) regions by hyperspectral imaging. Meanwhile, the first derivative, AUTO, multiplicative scatter correction, Savitzky-Golay(SG) smoothing, and standard normal variable transformation algorithms were used to pre-process the original hyperspectral data, which were then subjected to characteristic band extraction based on competitive adaptive reweighted sampling(CARS) and the partial least square regression analysis for building a quantitative model of SO_2 content in sulphur-fumigated Achyranthis Bidentatae Radix. It was found that the accuracy of the quantitative model built depending on the visible-near infrared spectra was high, with the determination coefficient of prediction set(R■) reaching 0.900 1. The established quantitative model has enabled the rapid and non-destructive detection of SO_2 content in sulphur-fumigated Achyranthis Bidentatae Radix, which can serve as an effective supplement to the method described in Chinese Pharmacopeia.

Stimulation quantitation of saponins and sterones from Achyranthis Bidentatae Radix by high-performance liquid chromatography with double external standards calibration method / 药学学报

Saponins and sterones are two main characteristic components in Achyranthis Bidentatae Radix. In order to control the quality of Achyranthis Bidentatae Radix more effectively, a high-performance liquid chromatography (HPLC) method was established by using double external standards calibration method (DESCM) for simultaneous determination of the contents of achyranthoside C, achyranthoside D, β-ecdysterone, 25R-inokosterone and 25S-inokosterone in Achyranthis Bidentatae Radix. Chromatographic separation was achieved on an Agilent Poroshell 120 EC-C18 (150 mm × 4.6 mm, 2.7 µm) using 0.1% phosphoric acid in water and 0.1% phosphoric acid in acetonitrile as mobile phase. The flow rate was 0.8 mL·min-1 and the column temperature was set as 35 ℃, the injection volume was 5 μL and the total analytical time was 30 min. β-Ecdysterone was used as the reference to calculate the relative correction factors (RCF) and relative retention time (RRT) of 25R-inokosterone and 25S-inokosterone, achyranthoside D was used for achyranthoside C. The RCFs of 25R-inokosterone, 25S-inokosterone, and achyranthoside C were 1.116, 1.056, and 0.888 1, respectively. The double external standards calibration method (DESCM) and external standard method (ESM) were used to calculate the contents of five ingredients in Achyranthis Bidentatae Radix samples from different sources and the variation between the results was within acceptable limits (RE ≤ 5%). The results showed that the contents of two saponins and three sterones of Achyranthis Bidentatae Radix were 0.597%-1.916% and 0.044%-0.150% respectively. The total content of saponins was about 10 times that of sterones. In conclusion, the established DESCM allowed simultaneous determination of five ingredients (achyranthoside C, achyranthoside D, β-ecdysterone, 25R-inokosterone, and 25S-inokosterone) in Achyranthis Bidentatae Radix, providing a scientific and feasible overall quality evaluation method for Achyranthis Bidentatae Radix.

Efficacy of Achyranthis Bidentatae Radix Against Cartilage Degeneration in Osteoarthritis： An Exploration Based on Computer Simulation / 中国实验方剂学杂志

Objective:To screen the active components of sovereign medicinal Achyranthis Bidentatae Radix in Rongjin Niantong formula based on bioinformatics and network pharmacology and observe their effects on therapeutic targets of osteoarthritis （OA） in <italic>in vivo</italic> and <italic>in vitro</italic> animal experiments. Method:The main active components and therapeutic targets of Achyranthis Bidentatae Radix were retrieved from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform （TCMSP）， and the differentially expressed genes relevant to OA from the Gene Expression Omnibus （GEO） database for cross analysis. The effects of main active components in Achyranthis Bidentatae Radix on enriched therapeutic targets of rats with OA <italic>in vivo </italic>and <italic>in vitro</italic> were detected by real-time polymerase chain reaction （Real-time PCR） and Western blot. Result:There were 20 active components for Achyranthis Bidentatae Radix against OA， with quercetin being an important one. Among the three target genes， osteopontin （OPN） and plasminogen activator inhibitor-1（PAI-1） were the key ones in the network. Gene Ontology （GO） analysis yielded 227 related terms， involving the regulation of physiological response to trauma （GO： 1903034）， negative regulation of trauma response （GO： 1903035）， etc. Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway analysis revealed 12 related pathways， involving extracellular matrix receptor interaction （hsa04512） and so on. In animal experiments， compared with the normal group， the model group exhibited increased gene and protein expression of OPN and PAI-1. Compared with the model group， the quercetin group displayed decreased gene and protein expression of OPN and PAI-1 （<italic>P</italic><0.05）. In cell experiments， the OPN and PAI-1 protein expression levels in the model group were increased as compared with those in the normal group， while the Collagen Ⅱ protein expression was decreased. The OPN and PAI-1 protein expression levels in the quercetin group and the inhibitor group were down-regulated in contrast to those in the model group， whereas the Collagen Ⅱ protein expression levels were up-regulated significantly （<italic>P<</italic>0.05）. Conclusion:Achyranthis Bidentatae Radix<italic> </italic>inhibits cartilage degeneration and exerts the preventive and therapeutic effects against OA， which is possibly due to the efficacy of its active component quercetin in down-regulating the expression of OPN and PAI-1 in chondrocytes and up-regulating the Collagen Ⅱ protein expression.

Network pharmacology-based study on material basis and mechanism of Achyranthis Bidentatae Radix in treatment of Alzheimer’s disease / 中草药

Objective This study was designed to predict the main targets of Alzheimer’s disease (AD) in Achyranthis Bidentatae Radix based on network pharmacology and molecular docking methods, and to explore its “multi-component, multi-target, and multi- pathway” mechanism. Methods According to the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP), the library of chemical constituents of Achyranthis Bidentatae Radix was established by referring to Chinese and foreign literature reports and collecting targets for treating AD in DrugBank. The Discovery Studio 3.5 software was used to carry out molecular docking, virtual screening of the chemical composition set of Achyranthis Bidentatae Radix combined with AD target, and KEGG database was used to enrich and analyze the key target of virtual screening. The active compounds of Achyranthis Bidentatae Radix with anti-AD activities were yielded to Discovery Studio 3.5 software and molecular docking to predict the poteneial proteins and carry out related KEGG pathways notation separately. Finally, the network of “active compound-target proteins-pathway” was built and analyzed using the Cytoscape 3.2.1 software. Results The 58 active compounds were selected from Achyranthis Bidentatae Radix, of which were mostly small alkanes, esters, and carboxylic acids followed by flavonoids and terpenoids. These active ingredients may regulate 36 potential target proteins such as CaMK-IIα, CaMK-IIβ, CaMK-IIγ, Akt1, and TNF-α to play a role in the pathogenesis of AD. The results also suggested that 12 signaling pathways were involved in the pathogenesis of AD, such as MAPK signaling pathway, Wnt signaling pathways and so on. Conclusion This research method initially revealed that the active ingredients of flavonoids and glycosides in Achyranthis Bidentatae Radix are the material basis for the treatment of AD. Its mechanism of action involved anti-inflammatory, anti-apoptosis and so on.

Analysis on Fingerprint of Achyranthis Bidentatae Radix / 中国实验方剂学杂志

Objective:To study HPLC fingerprints of Achyranthis Bidentatae Radix from different origins,compare different specifications in the same origin,and explore the effect of origin and specifications on the quality of Achyranthis Bidentatae Radix and relationship between the specifications and the internal quality of Achyranthis Bidentatae Radix, in order to provide basis for the identification of its origin. Method:The HPLC fingerprints of Achyranthis Bidentatae Radix from different origins and with different specifications in the same origin were collected. The similarity analysis,cluster analysis and principal component analysis were adopted to analyze the fingerprints,the differences in fingerprints of Achyranthis Bidentatae Radix from different origins and with different specifications in the same origin were compared. Result:Analysis of different origins and principal component analysis could be used to distinguish Achyranthis Bidentatae Radix from five producing areas,and the identification results of origin analysis was better than those of cluster analysis and similarity analysis. Analysis of different specifications, similarity analysis or principal component analysis could not distinguish Achyranthis Bidentatae Radix with different specifications. Conclusion:There are significant differences in chemical composition and peak height among Achyranthis Bidentatae Radix from different origins,with less differences in chemical composition and peak height of Achyranthis Bidentatae Radix with different specifications, the principal component analysis could be used to identify origins of Achyranthis Bidentatae Radix.

Correlation between surface features and hygroscopicities of Achyranthis bidentatae Radix extract powders added with different excipients / 中成药

AIM To investigate the correlation between surface features and hygroscopicities of Achyranthis bidentatae Radix extract powders added with different excipients.METHODS Achyranthis bidentatae Radix aqueous extract was added with crospovidone CL-SF,maltodextrin DE11-14,β-cyclodextrin,Arabic gum,dextrin,mannitol,soluble starch and lactose,respectively,after which extract powders were prepared,and hygroscopicities and morphological changes were determined.Then the surface features of various extract powders were compared by double exponential model.RESULTS Compared with other excipients,soluble starch and β-cyclodextrin could reduce the saturated moisture content of the extract powders more obviously,and crospovidone CL-SF exhibited the strongest inhibitory effect on powder synechia after moisture absorption.No direct relationship between hygroscopicities and specific surface area or pore volume was observed,but specific surface area,pore volume and average particle size demonstrated positive correlations with model parameter A1.CONCLUSION Addition of proper excipients can improve the surface features and hygroscopicities of Achyranthis bidentatae Radix extract powders.

Research progress on meridian-guiding theory of traditional Chinese medicine / 中国中药杂志

Ancient materia medica and medical formularies were consulted to illustrate the development history of meridian-guiding theory of traditional Chinese medicine (TCM). The influences of various meridian-guiding drugs (Achyranthis Bidentatae Radix, borneol, Bupleuri Radix, Platycodon Radix) on the pharmacokinetic and pharmacodynamic characteristics of other drugs were summarized. Meridian-guiding drugs can promote the absorption and targeted distribution of other drugs and enhance the efficacy of injured tissues. The possible mechanisms of meridian-guiding are related with changing the component of cell membrane, inhibiting the efflux of P-gp, opening physiological barriers, modulating the levels of biochemicals, promoting microcirculation and adjusting the pH of targeted tissues. The chemical components of meridian-guiding drugs are the substance basis of meridian-guiding. The aim of exploring meridian-guiding chemicals is to find a natural targeted delivery system. At the present time, some progress has been made in the research on meridian-guiding field. However, further studies are required for the meridian-guiding theory of TCM.

Effects of Tongsaimai Prescription and Tongsaimai Prescription without Achyranthis Bidentatae Radix on ischemic brain injuries in rats / 中草药

Objective: To investigate the effects of Tongsaimai Prescription (TSMP) and TSMP without Achyranthis Bidentatae Radix (ABR) on ischemic brain injuries in rats. Methods: The male SD rats were divided into Sham, model, Nimodipine (32.4 mg/kg), TSMP (crude drug 31.74 g/kg), and TSMP without ABR (crude drug 27.77 g/kg) groups. Except the Sham and model groups, the rats were ig administered once daily, for 3 d. After the treatment, except the Sham group, the cerebral ischemic stroke model was established. After 24 h, the blood and brain tissue were taken, the brain tissue was stained with TTC, and the cerebral infarction ratio was measured. The pathological changes were observed, and the contents of HMGB1, TNF-α, and IL-1β in serum were detected. The expression of NF-κB in ischemia tissue was observed by immunohistochemisty method. Results: The expression of HMGB1, TNF-α, and IL-1β increased obviously (P < 0.05). TSMP and TSMP without ABR could decrease the contents of HMGB1, TNF-α, and IL-1β, inhibit the expression of NF-κB (P < 0.05), and the effect of TSMP without ABR on decreasing IL-1β was stronger. Conclusion: Both TSMP and TSMP without ABR could inhibit the activation of HMGB1-related NF-κB signal pathway with the function of anti-imflammation, alleviated encephaledema, and protecting nerve cells. TSMP without ABR has the similar effect on the ischemic brain injuries and the better effect on inhibiting the secretion of inflammatory factor IL-1β.