Effects of Rehmanniae Radix and Rehmanniae Radix Praeparata on proteomics and autophagy in mice with type 2 diabetes mellitus induced by high-fat diet coupled with streptozotocin / 中国中药杂志

To compare the pancreatic proteomics and autophagy between Rehmanniae Radix-and Rehmanniae Radix Praeparata-treated mice with type 2 diabetes mellitus(T2DM). The T2DM mouse model was established by high-fat diet coupled with streptozotocin(STZ, intraperitoneal injection, 100 mg·kg~(-1), once a day for three consecutive days). The mice were then randomly assigned into a control group, low-(5 g·kg~(-1)) and high-dose(15 g·kg~(-1)) Rehmanniae Radix groups, low-(150 mg·kg~(-1)) and high-dose(300 mg·kg~(-1)) catalpol groups, low-(5 g·kg~(-1)) and high-dose(15 g·kg~(-1)) Rehmanniae Radix Praeparata groups, low-(150 mg·kg~(-1)) and high-dose(300 mg·kg~(-1)) 5-hydroxymethyl furfuraldehyde(5-HMF) groups, and a metformin(250 mg·kg~(-1)) group. In addition, a normal group was also set and each group included 8 mice. The pancreas was collected after four weeks of administration and proteomics tools were employed to study the effects of Rehmanniae Radix and Rehmanniae Radix Praeparata on protein expression in the pancreas of T2DM mice. The expression levels of proteins involved in autophagy, inflammation, and oxidative stress response in the pancreatic tissues of T2DM mice were determined by western blotting, immunohistochemical assay, and transmission electron microscopy. The results showed that the differential proteins between the model group and Rehmanniae Radix/Rehmanniae Radix Prae-parata group were enriched in 7 KEGG pathways, such as autophagy-animal, which indicated that the 7 pathways may be associated with T2DM. Compared with the control group, drug administration significantly up-regulated the expression levels of beclin1 and phosphorylated mammalian target of rapamycin(p-mTOR)/mTOR and down-regulated those of the inflammation indicators, Toll-like receptor-4(TLR4) and Nod-like receptor protein 3(NLRP3), in the pancreas of T2DM mice, and Rehmanniae Radix showed better performance. In addition, the expression levels of inducible nitric oxide synthase(iNOS), nuclear factor erythroid 2-related factor 2(Nrf2), and heine oxygenase-1(HO-1) in the pancreas of T2DM mice were down-regulated after drug administration, and Rehmanniae Radix Praeparata demonstrated better performance. The results indicate that both Rehmanniae Radix and Rehmanniae Radix Praeparata can alleviate the inflammatory symptoms, reduce oxidative stress response, and increase the autophagy level in the pancreas of T2DM mice, while they exert the effect on different autophagy pathways.

Mechanism Analysis of Strengthening Anti-cerebral Ischemia Injury of Chuanxiong Rhizoma Processed with Wine Based on UHPLC-Q-Orbitrap HRMS and Integrated Network Pharmacology / 中国实验方剂学杂志

ObjectiveTo analyze the differential components in water extract of Chuanxiong Rhizoma before and after processing with wine， and to explore the molecular mechanism of Chuanxiong Rhizoma processed with wine in enhancing anti-cerebral ischemia injury. MethodUltra high performance liquid chromatography tandem quadrupole orbitrap high resolution mass spectrometry （UHPLC-Q-Orbitrap HRMS） was used to qualitatively analyze the main chemical components in water extract of Chuanxiong Rhizoma based on the spectral information of compound， comparison of reference substance and references. The chemical pattern recognition method was used to screen the differential components of Chuanxiong Rhizoma before and after processing. Based on these differential components， the potential targets of differential components were predicted by online databases， and the related targets of cerebral ischemia were searched. Cytoscape 3.6.0 was used to establish the network diagram of differential components-action targets-diseases of Chuanxiong Rhizoma processed with wine. The protein-protein interaction （PPI） network of intersection targets was constructed by STRING 11.5. The potential targets of differential components against cerebral ischemia were analyzed by Gene Ontology （GO） and Kyoto Encyclopedia of Genes and Genomes （KEGG） enrichment analysis through DAVID 6.8. At the same time， the chemical compounds with high relative content and increased peak area after wine processing were docked with their corresponding targets to verify the mechanism of enhanced effect after wine processing. ResultA total of 71 chemical components were identified from Chuanxiong Rhizoma， 34 differential components and 603 potential targets were screened out. At the same time， a total of 769 disease targets and 60 intersection targets were obtained. Seven key targets were identified through PPI network analysis， including JUN， signal transducer and activator of transcription 3 （STAT3）， mitogen-activated protein kinase 3 （MAPK3）， interleukin-1β （IL-1β）， vascular endothelial growth factor A （VEGFA）， Caspase-3 （CASP3） and mtrix metalloproteinase 9 （MMP9）. Tumor necrosis factor （TNF） signaling pathway was the main differential signaling pathway. The results of molecular docking showed that differential components （senkyunolide K， senkyunolide F， 3-n-butylphthalide， Z，Z′-6，8′，7，3′-diligustilide， ferulic acid and Z-ligustilide） and corresponding targets had good binding activities. ConclusionThe synergistic mechanism of Chuanxiong Rhizoma processed with wine may be related to the enhanced inhibitory effect of inflammatory reaction.

Effect of Guilu Erxiangao on Alzheimer's Disease and Its Mechanism Based on Kidney-brain Correlation / 中国实验方剂学杂志

ObjectiveTo investigate the effect and mechanism of Guilu Erxiangao on Alzheimer's disease （AD） model rats induced by hydrocortisone and amyloid β-protein（Aβ） based on the theory of kidney-brain correlation. MethodIntraperitoneal injection of hydrocortisone and intracerebroventricular injection of Aβ were performed to induce AD in rats， and different concentrations of Guilu Erxiangao were used for intervention. The indexes of hippocampus， kidney and adrenal gland were measured， and the spatial learning and memory ability of AD rats was observed by Morris water maze experiment. The levels of testosterone （T） and corticosterone （CORT） in serum samples were determined by enzyme-linked immunosorbent assay （ELISA）. Liquid chromatography-mass spectrometry （LC-MS） was used to collect and analyze the serum metabolic data of model rats. The active components and corresponding targets of Guilu Erxiangao were collected using Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform（TCMSP）， a Bioinformatics Analysis Tool for Molecular mechANism of Traditional Chinese Medicine （BATMAN-TCM） and Traditional Chinese Medicine Integrated Database（TCMID）. GeneCards and Online Mendelian Inheritance in Man （OMIM） were retrieved to obtain AD-related targets， and protein-protein interaction （PPI） network was constructed to perform gene ontology （GO）and Kyoto Encyclopedia of Genes and Genomes （KEGG） enrichment analysis. The expression level of interleukin-6 （IL-6） in the hippocampus of rats was detected by Western blot. ResultCompared with the model group， the low-， medium- and high-dose groups of Guilu Erxiangao exhibited significantly increased hippocampus index， kidney index and adrenal gland index， reduced CORT levels in serum and down-regulated IL-6 levels in hippocampal tissues. According to the results of water maze experiment， as compared with the model group， the platform crossing times of rats was significantly increased in the low- and high-dose groups of Guilu Erxiangao， with evidently prolonged distance traveled in quadrant Ⅲ （%） and time in quadrant Ⅲ （%）. A total of 24 serum differential metabolites associated with AD were identified by LC-MS， and 50 high-frequency common compounds and 187 high-frequency common targets for AD treatment were screened by network pharmacology method. Results demonstrated phosphatidylinositol 3-kinases（PI3K）/protein kinase B（Akt） signaling pathway plays an important role in the complex AD pathological mechanism. ConclusionGuilu Erxiangao can significantly improve the cognitive dysfunction of AD model rats induced by hydrocortisone and Aβ， reduce serum CORT levels and IL-6 levels in hippocampal tissues， and regulate the metabolic level， which provides a reference for its clinical application.

Rehmanniae Radix and Rehmanniae Radix Praeparata improve diabetes induced by high-fat diet coupled with streptozotocin in mice through AMPK-mediated NF-κB/NLRP3 signaling pathway / 中国中药杂志

This study investigated the differential mechanisms of Rehmanniae Radix and Rehmanniae Radix Praeparata in improving diabetes in mice through AMPK-mediated NF-κB/NLRP3 signaling pathway. The diabetic mouse model was established with high-fat diet coupled with streptozotocin(STZ, intraperitoneal injection, 100 mg·kg~(-1), once a day for three consecutive days), after which the mice were randomly divided into model group, low-dose(5 g·kg~(-1)) and high-dose(15 g·kg~(-1)) Rehmanniae Radix groups, low-dose(5 g·kg~(-1)) and high-dose(15 g·kg~(-1)) Rehmanniae Radix Praeparata groups, catalpol group(250 mg·kg~(-1)), 5-hydroxymethylfurfural(5-HMF) group(250 mg·kg~(-1)), metformin group(250 mg·kg~(-1)), with the normal group also set. The organ indexes of heart,liver, spleen, lung, kidney and pancreas were calculated after four weeks of administration. The pathological changes and fibrosis of pancreas, kidney and liver in mice were observed by hematoxylin-eosin(HE) staining and Masson staining. Western blot was used to determine the expression levels of Toll-like receptor-4(TLR4), nuclear factor-κB(NF-κB), Nod-like receptor protein 3(NLRP3),interleukin-1β(IL-1β), adenosine monophosphate-activated protein kinase(AMPK), phosphorylated AMPK(p-AMPK) in the pancreas, kidney and liver of mice. Compared with the model group, the administration groups witnessed significant decrease in the liver,spleen, kidney, pancreas and fat indexes of diabetic mice, and there was no significant difference in heart and lung indexes. The pathological states and fibrosis of pancreatic, kidney and liver tissues were significantly improved after administration. Additionally, the expression levels of TLR4, NF-κB and NLRP3 in pancreas, kidney and liver of diabetic mice were significantly lowered. The expression levels of p-AMPK/AMPK were enhanced significantly in kidney and liver of mice in Rehmanniae Radix group while in pancreas, kidney and liver in Rehmanniae Radix Praeparata group. This suggests that Rehmanniae Radix and Rehmanniae Radix Praeparata differ in the mechanism of regulating energy metabolism of multiple organs and thereby exerting anti-inflammatory effects to alleviate symptoms of diabetic mice.

Blood-cooling and hemostasis effects of Rehmanniae Radix before and after carbonizing / 中国中药杂志

To compare the blood-cooling and hemostasis effects of Rehmanniae Radix before and after carbonizing on rats with blood heat and hemorrhage syndrome. The blood heat and hemorrhage syndrome rat model was established. Indexes including rectal temperature,whole blood viscosity,plasma viscosity,thrombin time(TT),activated partial thromboplastin time(APTT),prothrombin time(PT),fibrinogen content(FIB),red blood cell(RBC),hemoglobin(Hb),hematocrit(HCT),blood platelet count(PLT),mean platelet volume(MPV),serum IL-1,serum IL-6 and lung histopathology were detected to investigate the blood-cooling and hemostasis effects of Rehmanniae Radix and its carbonized products. Compared with the blank control group,the rectal temperature was significantly increased with rise of the high,middle and low whole blood viscosities and plasma viscosity(P<0.05); both the high and low whole blood restore viscosity and the high and low whole blood relative viscosity were increased significantly(P< 0.05); TT,APTT and PT were notably prolonged with the increase in FIB content(P<0.05); RBC,Hb and HCT increased significantly(P< 0.05); concentrations of serum IL-1 and IL-6 were also increased(P< 0.05) in model group. Additionally,obvious hemorrhages in lung and stomach were observed in rats of the model group. Rehmanniae Radix and its carbonized products can significantly reduce rectal temperature,high middle and low whole blood viscosities and plasma viscosity(P<0.05). TT and APTT were shortened,with lower expression of FIB in group of Rehmannia Radix and its carbonized products. Hemorrhages of lung and stomach were improved by Rehmannia Radix and its carbonized products. The results indicated that Rehmannia Radix before and after carbonizing had the hemostasis and blood-cooling effects by promoting coagulation,improving blood rheology and inhibiting expressions of IL-1 and IL-6.

Analysis of Mechanism of Phytolaccae Radix and Its Processed Products on Doxorubicin-induced Nephropathy in Rats / 中国实验方剂学杂志

Objective: To compare the protective effect of Phytolaccae Radix and its processed products on nephropathy induced by doxorubicin (DOX) in rats, and explore its mechanism. Method: A rat model of nephropathy was established by a single tail intravenous injection of DOX hydrochloride. Content of esculentoside A (EsA) in Phytolaccae Radix and its processed products was determined by HPLC-ELSD. Contents of serum total protein (TP), albumin (Alb), urea nitrogen (BUN), serum creatinine (SCr), total cholesterol (TC) and urine protein (UP) were determined by enzyme-linked immunosorbent assay (ELISA). The mRNA and protein expression of transforming growth factor-β (TGF-β) in renal tissue of rats was examined by real-time quantitative polymerase chain reaction (Real-time PCR) and immunohistochemistry. Result: A single intravenous injection of DOX could induce a severe nephrotic syndrome associated with decreased serum TP, Alb and elevated serum BUN, SCr, TC, and a high urinary excretion of protein (Pβ in renal tissue of model group rats was significantly higher than that of blank group (PPPConclusion: Phytolaccae Radix and its processed products can improve the symptoms of DOX nephropathy model rats in different degrees, among which the vinegar prepared products have the strongest effect, and this effect may be related to the reduction of TGF-β expression in renal tissue.

Preliminary investigation on processing mechanism of Kansui Radix by liquorice based on thermal analysis technology / 中国中药杂志

In the thermal analysis, the pyrolysis characteristics of crude Kansui Radix, alcohol extract of Kansui Radix, petroleum ether extract, chloroform extract, ethyl acetate extract, n-butanol extract, and licorice vinegar were analyzed with simulated air (N₂-O₂ 4:1) as the carrier gas, at a temperature increase rate of 10 °C·min⁻¹ and a volume flow rate of 60 mL·min⁻¹, respectively. The results showed that due to the different polarity of the extraction solvent, the type and quantity of the chemical components contained in each polar part were different, and with the increase in the amount of solid powder of licorice, the peak of the maximum heat loss rate occurred in advance. For petroleum ether, chloroform, and ethyl acetate fractions, (157.40±1.06), 3.50, (25.83±1.66) °C in advance respectively, but the weight loss rate of the chloroform fraction was increased by (2.62±5.19) °C, while decreased by (33.90±1.72), (19.28±1.11) °C for the petroleum ether and ethyl acetate fractions. So we can conclude that with the addition of licorice, the pyrolysis rate of the petroleum ether and chloroform fractions in the toxic part of Kansui Radix was increased; the temperature point at the peak of the maximum weight loss rate was decreased, and the ethyl acetate fraction (effective part) showed a decrease in temperature rising process, but its overall ratio of weight loss and weight loss rate were relatively small, retaining the effect of medicinal ingredients. This proved the mechanism of licorice system Kansui Radix on attenuating toxicity after processing and the scientificity and rationality of licorice system Kansui Radix. At the same time, as the proportion of glycyrrhizin was increased, the peak of the maximum heat loss rate of petroleum ether, chloroform and ethyl acetate fractions occurred in advance; the peak temperature was decreased, with easy pyrolysis. Among them, the thermogravimetric rate of the mixture of petroleum ether and chloroform fractions (10:1) was relatively large, with a low peak temperature, while ethyl acetate fraction showed opposite results. This conclusion has certain guiding significance for the ratio of gansui to licorice.

Processing of calamine with modern analytical techniques: Processed with Huanglian Decoction () and Sanhuang Decoction () / 中国结合医学杂志

<p><b>OBJECTIVE</b>To determine the pyrolysis characteristics of calcined and processed calamine, qualitatively and quantitatively compare the contents of related elements, morphology and functional groups of the pyrolysis products dried at different heating temperatures and explore the critical temperature and the optimal drying temperature for the process of calamine with Huanglian Decoction (HLD, ) and San Huang Decoction (SHD, ).</p><p><b>METHODS</b>Pyrolysis products were prepared by programmable and constantly heating the calcined and processed calamine to or at different heating temperatures. Thermogravimetry (TG) was used to test their pyrolysis characteristics. Fourier transform infrared spectroscopy and scanning electron microscopeenergy dispersive spectrometer were used to determine their morphology, functional groups and element contents. Page model was used to investigate the constant drying kinetics of processed calamine.</p><p><b>RESULTS</b>The adding of HLD or SHD to calcined calamine (CC) can slow its weight loss in drying pyrolysis process. The temperature ranges where HLD and SHD can affect its weight loss were 65-150 °C and 74-180 °C, respectively. The drying temperature was optimized as 90 °C. The drying kinetic for the processed calamine fits Page model shows good linearity.</p><p><b>CONCLUSIONS</b>Conclusions: The critical temperature and the optimal drying temperature where HLD and SHD can affect the weight loss rate in the process of calamine were explored using the theories and methods of both biophysical chemistry and processing of Chinese materia medica. This work provides a good example for the study of the process of other Chinese medicines using modern analytical techniques.</p>

Pyrolysis characteristics of radix rhizoma rhei, cortex moudan radicis, and radix sanguisorbae and correlations with the carbonizing process of Chinese herbs / 中国天然药物

AIM@#The aim of the work is to study the pyrolysis characteristics of radix rhizoma rhei, cortex moudan radicis, and radix sanguisorbae in an inert atmosphere of argon (Ar), and to investigate the mechanism of the carbonizing process of the three traditional Chinese herbs.@*METHODS@#The pyrolysis characteristics of the crude materials and their extracts were studied by thermogravimetry-mass spectrometry (TG-MS) in a carrier gas of argon, coupled with Fourier transform infrared spectrometry (FTIR) and scanning electron microscopy (SEM) methods. Correlation of the pyrolysis behaviors with the carbonizing process by stir-frying of traditional Chinese medicines was made.@*RESULTS@#Within the temperature range of 200-300 °C, which is the testing range for the study of the carbonizing process of Chinese herbs, the temperatures indicated by the maximum weight loss rate peak of the above three extracts were taken as the upper-limit temperatures of the carbonizing process of the herbs, and which were 200, 240 and 247 °C for radix rhizoma rhei, cortex moudan radicis, and radix Sanguisorbae, respectively. The ion monitoring signal peaks detected by the TG-MS method corresponded with reports that the level of chemical components of traditional Chinese medicinal materials would decrease after the carbonizing process. It was confirmed by Fourier transform infrared spectrometry (FTIR) and scanning electron microscopy (SEM) methods that better results of "medicinal property preservation" could be obtained by heating at 200 °C for radix rhizoma rhei, at about 250 °C for cortex moudan radicis, and radix sanguisorbae, as the relative intensity values of the common peaks were among the middle of their three carbonized samples by programmed heating.@*CONCLUSION@#The upper-limit temperatures of the carbonizing process for radix rhizoma rhei, cortex moudan radicis and radix sanguisorbae were 200, 240 and 247 °C respectively. It is feasible to research the mechanism and technology of the carbonizing process of traditional Chinese medicinal materials using thermogravimetry, Fourier transform infrared spectrometry, and scanning electron microscopy methods.

Study on thermoanalysis-based calamina processing / 中国中药杂志

Thermogravimetry (TG), TG-MS, Fourier transform infrared spectroscopy (FTIR), scanning electron microscope (SEM)-energy dispersive spectrometer(EDS) were adopted to investigate the pyrolysis characteristics of calamina. According to the findings of the qualitative and quantitative studies on the changes in the content of relevant elements, the whole shape, the functional groups, and the volatile components of calamina before and after being pyrolyzed, the 200-360, 580-750 degrees C were two sensitive temperature ranges related to the changes in effective component during calamina processing. Thermal weight loss was observed for ZnCO3, Zn(OH)2 and ZnCO3-2Zn(OH)2-H2O under 200-360 degrees C and for CaCO3 under 580-750 degrees C. The results of studies on chemical reaction kinetics showed good linear relations. This experiment integrated relevant methods and theories of physical chemistry and science of traditional Chinese medicine processing, and interpretes calamina processing techniques and mechanism, in order to provide a good example for modem studies on other traditional Chinese medicine processing.

Research on processing mechanism of Zushima which was stir-fried with licorice based on TG-DTG / 中国中药杂志

<p><b>OBJECTIVE</b>To investigate the processing mechanism of Zushima which was stir-fried with licorice.</p><p><b>METHOD</b>Study of pyrolysis characteristics for extraction of Zushima effective part, petroleum ether, chloroform, ethyl acetate, n-butanol parts and the mixture of Zushima effective parts and licorice solid powder according to the proportion of 10: 1, 10: 2, 10: 3 was carried out in the thermogravimetric analyzer, the simulation of air (N2-O2 4:1) was chosen as carrier gas and heating rate was 5 degrees C x min(-1).</p><p><b>RESULT</b>Compared with TG-DTG curve of Zushima effective parts, the major weightloss temperature range of petroleum ether extraction which has strong stimulation was 320-390 degrees C, 0.69% x min(-1) weightlessness rate peak gradually moved forward with the addition of licorice powder, finally it was merged with the peak around 265 degrees C. In addition, effective department of Zushima at 291, 516 degrees C for 2.38% x min(-1) and 2.42% x min(-1) thermal weightlessness rate peak shift to lower temperature, the moving range were about 20-26, 19-50 degrees C, the former was significantly reduced, the latter was significantly increased.</p><p><b>CONCLUSION</b>In the course of programmed temperature heating, petroleum ether department was easy to lose with the addition oflicorice solid powder. At the same time, the main efficacy components of Zushima had a slow loss rate, which supported the processing mechanisms of TG-DTG method to research traditional Chinese medicine, and verified irritating characteristics that stimulus reduced after stir-fried with licorice.</p>