Analysis on Antidepressant Mechanism of Verbascoside Based on RNA-Seq Technology / 中国实验方剂学杂志

Objetive： To investigate the mechanism of antidepressant effect of verbascoside based on high-throughput sequencing technology （RNA-Seq），and to explore the possible targets and signaling pathways. MethodsForty C57BL/6 mice were randomly divided into control group，model group，fluoxetine group and verbascoside group，10 mice in each group. Except for the control group，all the other three groups were constructed with chronic unpredictable mild stimulation （CUMS） combined with solitary feeding for four weeks. Control group，fluoxetine group and verbascoside group were administered by gavage once daily for three weeks during the second week of modeling. The mice were assessed by sugar-water preference test，forced swimming test，open field test，elevated cross maze test，and water maze test. Enzyme linked immunosorbent assay（ELISA） was performed to detect the levels of major neurotransmitters and inflammatory factors in mice serum，and mRNA high-throughput sequencing was performed in the nucleus accumben and colon to screen differentially expressed genes and perform pathway enrichment analysis. Real-time polymerase chain reaction（Real-time PCR） was used to detect the mRNA expression of Gad，Slc32a1（VGAT） and brain-derived neurotrophic factor (BDNF） in nucleus accumbens. ResultCompared with the control group，the anxiety and depression-like behaviors in the model group increased，while the learning and memory ability decreased significantly. The content of neurotransmitter in serum decreased significantly. The content of pro-inflammatory factors increased significantly. The mRNA expression of Gad and Slc32a1（VGAT） in nucleus accumbens decreased significantly，while that of BDNF increased significantly. Compared with the model group，the anxiety and depression-like behavior of mice in verbascoside group was significantly relieved. The neurotransmitter content increased significantly，and the pro-inflammatory factors decreased significantly. The mRNA expression of Gad and Slc32a1（VGAT） in nucleus accumbens increased significantly，while that of BDNF decreased significantly.A total of 48 differentially expressed genes in nucleus accumbens and 43 differentially expressed genes in colon were screened by high-throughput sequencing. Differential genes in nucleus accumbens mainly focus on neuroactive ligand-receptor interaction， γ-aminobutyric acid（GABA）ergic synapse，synaptic vesicle cycle and other pathways. Colonic differential genes are mainly concentrated in GABAergic synapses，synaptic vesicle circulation，cyclic adenosine monophosphate（cAMP） signal pathway and other signal pathways. Compared with the control group，the mRNA expression of Gad and Slc32a1（VGAT） in nucleus accumbens of model group decreased significantly，while the mRNA expression of BDNF increased significantly. ConclusionVerbascoside has significant antidepressant effects. Its antidepressant effect may be related to the increase of monoamine neurotransmitters，the decrease of pro-inflammatory factors and the restoration of neurotransmitter homeostasis by increasing GABA，and it mainly acts through the signaling pathways such as neuroactive ligand-receptor interaction，GABAergic synapses，synaptic vesicle cycle and cAMP signaling pathway.

Preparation of Rehmanniae Radix Juice for Baihe Dihuangtang Based on UPLC-Q-TOF-MS / 中国实验方剂学杂志

ObjectiveTo compare the four preparation methods of Rehmanniae Radix juice described in ancient literature and find the method that is most suitable for the preparation of Rehmanniae Radix juice used in Baihe Dihuangtang. MethodThe ancient medical books record four methods for preparing Rehmanniae Radix juice： crushing fresh Rehmanniae Radix for juice， steaming fresh Rehmanniae Radix for juice， boiling fresh Rehmanniae Radix for juice， and boiling dry Rehmanniae Radix for juice. Ultra performance liquid chromatography-quadrupole time-of-flight mass spectrometry （UPLC-Q-TOF-MS） was employed to detect the compounds in the four juice samples， followed by principal component analysis （PCA）. Result① Totally 27 compounds were identified in the juice samples， including 10 iridoid glycosides， 14 phenylethanoid glycosides， 2 phenolic acids， and 1 irisone. Among them， 15 common compounds were shared by the four juice samples， including 7 iridoid glycosides， 7 phenylethanoid glycosides， and 1 phenolic acid. ② Five common compounds in the four juice samples can be matched with the reference standards， which were catalpol， aucubin， rehmannioside D， ajugol， and purpureaside C. ③ Verbascoside and isoacteoside were not detected in the juice prepared by crushing fresh Rehmanniae Radix， while it was detected in the other three juice samples， which indicated that the two components were produced after heating rather than being the original components in fresh Rehmanniae Radix. ④ The comparison of the ion fragments demonstrated that verbascoside was produced from purpureaside C after the cleavage of the glycosidic bond and removal of a molecule of mannose. ⑤ Isoacteoside could be isomerized from verbascoside， and its relative content increased with the extension of heating time. However， the relative content of verbascoside and purpureaside C did not decrease significantly. Therefore， it was hypothesized that purpureaside C was produced from its upstream component. ConclusionThe juice prepared by crushing fresh Rehmanniae Radix has the chemical composition significantly different from the juice samples prepared with the other 3 methods， while the latter 3 juice samples had similar chemical composition. Although all the four methods can be used， it is more suitable to prepare Rehmanniae Radix juice by steaming fresh Rehmanniae Radix， boiling fresh Rehmanniae Radix， and boiling dry Rehmanniae Radix.

Pharmacokinetic study of Erlong Zuoci Pill in rats / 中国药科大学学报

@#To establish a quantitative LC-MS/MS method for the simultaneous detection of components of Erlong Zuoci Pill in rat plasma: verbascoside, oxypaeoniflorin, echinacoside and benzoylpaeoniflorin, and to evaluate the pharmacokinetic characteristics of Erlong Zuoci Pill in rats, plasma samples were purified by protein precipitation using methanol as a protein precipitant.Methanol was used as the organic phase and aqueous solution containing 0.1% formic acid was used as the water phase.The quantitative analysis method of verbascoside, oxypaeoniflorin, echinacoside and benzoylpaeoniflorin was established in negative ion mode, and the validation of bioanalytical method was carried out.Healthy SD rats were selected, and 20 mL/kg (equivalent to the original drug 10 g/kg dose) of Erlong Zuoci Pill extract was administered by intragastric administration.The plasma concentration of the target compounds at different time intervals after administration was determined using the established method, and the pharmacokinetic parameters was calculated by the Phoenix WinNonlin8.3 software using the non-compartmental model.The method validation results showed that verbascoside (r = 0.993 7) and oxypaeoniflorin (r = 0.994 6) had good linear relationship in the concentration range of 0.5-50 ng/mL, echinacoside (r = 0.993 6) and benzoylpaeoniflorin (r = 0.992 6) had good linear relationship in the concentration range of 1-100 ng/mL.The relative standard deviations of the inter- and intra- batch precision of the four compounds were all less than 15%, and the inter- batch and intra- accuracies were between 85% and 115%.Extraction recovery, matrix effect and stability met the relevant requirements.After a single gavage of Erlong Zuoci Pill extract in rats, all the four compounds were rapidly absorbed and eliminated.Oxypaeoniflorin, echinacoside, and benzoylpaeoniflorin showed two peaks in their drug concentration-time curves.Compared with the other three compounds, oxypaeoniflorin has the highest concentration in rat plasma with cmax1 of (24.40 ± 4.78) ng/mL and cmax2 of (22.50 ± 2.70) ng/mL. The results show that the validation results of this method are in line with the guiding principles of biological sample analysis methods, and it can be used to evaluate the pharmacokinetic characteristics of Erlong Zuoci Pill extract in rats.

HPLC Fingerprint Analysis and Multi-component Determination of Substance Benchmark of Yiweitang / 中国实验方剂学杂志

Objective:To establish high performance liquid chromatography （HPLC） fingerprint and multi-component determination for the substance benchmark of Yiweitang， and to evaluate its quality in combination with chemical pattern recognition method. Method:Fifteen batches of substance benchmark of Yiweitang were prepared， the "Chinese medicine chromatographic fingerprint similarity evaluation system" （2012 edition） was used to calculate similarity. Cluster analysis， principal component analysis and orthogonal partial least squares-discriminant analysis were employed to handle the common peaks for evaluating the quality difference among 15 batches of the substance benchmark. The contents of catalpol， verbascoside and methylophiopogonanone A were determined with mobile phase system of acetonitrile-phosphoric acid solution at detection wavelengths of 210 nm and 334 nm. Result:There were 22 common peaks in HPLC fingerprint of the substance benchmark， among them， peaks 1， 9， 12， 14-17， 19 and 20 belonged to Rehmanniae Radix， peaks 3， 4， 6， 7 and 21 belonged to Glehniae Radix， peaks 5 and 22 belonged to Ophiopogonis Radix， peaks 2 and 18 belonged to Polygonati Odorati Rhiaoma， peak 8 was the common peak of Ophiopogonis Radix and Rehmanniae Radix， peak 10 was shared by Ophiopogonis Radix， Polygonati Odorati Rhiaoma<italic> </italic>and<italic> </italic>Rehmanniae Radix， peak 11 was the common peak of these four herbs， and peak 13 was shared by Polygonati Odorati Rhiaoma and Rehmanniae Radix. The similarities between HPLC fingerprints of 15 batches of the substance benchmark and the control fingerprint were all >0.90， the samples could be divided into four categories by three chemical pattern recognition methods. Quantitative analysis showed that the contents of catalpol， verbascoside and methylophiopogonanone A among 15 batches of samples ranged from 0.37% to 1.14%， 0.002% to 0.054% and 0.016% to 0.079%， respectively. Conclusion:The established fingerprint and determination for the substance benchmark of Yiweitang have good separation and high accuracy， which reflect the overall chemical composition characteristics of Yiweitang， and can provide experimental basis for the further development and quality control of the compound preparations of this famous classical formula.

Determination, Isolation, And Identification Of Aucubin And Verbascoside In The Leaves Of Iraqi Plantago Lancoleta L. Using Different Detecting Methods

Objective: Plantago lanceoleta L. (ribwort plantain) is one of the important medicinal herbs which is widespread fortune available in Iraq, that have a wide range of medicinal properties. The aim of this work was to determine, isolate and identify verbascoside and aucubin in Iraqi P. lanceoleta L. by using different chromatographic and spectrometric methods. Methods: Verbascoside and aucubin were isolated and quantified by preparative TLC, and then they were determined by the high-performance thin-layer chromatography (HPTLC) fingerprinting. Aucubin and catalpol in the plant extract were analyzed by liquid chromatography-mass spectrometry (LC-MS); aucubin and verbascoside that isolated from the plant sample were examined by fourier-transform infrared spectroscopy (FT-IR) and LC-MS, respectively. Results: The result showed that the Iraqi P. lanceoleta L. contains 1.74 percent (verbascoside) and 0.24 percent (aucubin) of dry powdered leaves. Each TLC-isolated compound showed a single spot on the HPTLC plate, which give an idea about the purity of the isolated compound. Aucubin (with catalpol) and verbascoside both are detected by LC-MS in different ionization mode. Many functional groups were identified in the TLC-isolated aucubin by FT-IR. Conclusion: The Iraqi P. lanceoleta L. showed a high content of verbasoside, and it is a very rich source for this compound, which can be easily isolated by TLC and subjected to many pharmacological studies. The extract of the young leaves of this plant gave a little amount of aucubin, and it is easy to obtain a higher content from the older leaves.

Optimization of extraction process of Modified Yangyin Qingfei Decoction by Box-Behnken response surface methodology based on analytic hierarchy process / 中草药

Objective: To optimize the extraction process of Modified Yangyin Qingfei Decoction (MYQD). Methods: The effects of ethanol concentration, extraction time and extraction times on the optimization of extraction process of MYQD were investigated by multiple indicators comprehensive scoring and Box-Behnken response surface methodology. The content of verbascoside, chlorogenic acid, paeonol, and glycyrrhizic acid was simultaneously determined by HPLC, and the method of analytic hierarchy process was used to calculate the weight coefficient. Results: The optimum extraction process was as follow: using 69% ethanol for once extraction for 68 min. Conclusion: The optimum extraction process is simple and feasible, and the extraction efficiency of components is high, which can provide reference for the extraction process of MYQD.

Extraction parameters and drying methods of Cistanche deserticola / 中草药

Objective: The optimum extraction process parameters of Cistanche deserticola were selected to study the effects of different drying methods on five phenylethanoid glycosides. Methods: Single factor screening combined with Box-Behnken response surface method was used to optimize the extraction process. After optimal conditions were extracted, HPLC method was used to detect the content of echinacoside, cistanche A, verbascoside, isoacteoside, and 2’-acetylacteoside in different drying methods, and one-way ANOVA, cluster analysis, principal component analysis and close value analysis were used to analyze the content of five phenylethanoid glycosides to choose the best drying method. Results: Optimal extraction process was as following: methanol volume fraction was 55.14%, liquid to material ratio was 46.39, extraction time was 38.50 min. Cluster analysis, principal component analysis, and close value analysis showed that the quality of C. deserticola obtained by freeze-drying method was the best, followed by drying at 80 ℃ and the lowest at 40 ℃. Conclusion: Using this process to extract C. deserticola, the five phenylethanoid glycosides are completely and fully extracted. Although the freeze-drying method of C. deserticola has the highest active ingredient retention, from the production point of view, the 80 ℃ drying method can achieve a balance of cost and efficiency.

Advances in biosynthesis and chemical synthesis of phenylethanoid glycosides / 中草药

Phenylethanoid glycosides (PeGs) are natural active ingredients of many plants at home and abroad. They possess a spectrum of beneficial activities, such as anti-oxidant, hepatoprotective, whitening, and neuroprotective. However, the content of PeGs in food or medicinal materials is low and unstable. During the past decade, studies on biosynthesis and metabolic regulation of PeGs have been extensively carried out. Here, the recent achievements in biosynthesis and metabolic regulation of PeGs in plants and microorganisms are reviewed, as well as in total synthesis and semi-synthesis of PeGs. Hopefully, this work done so far will provide reference for elucidating the biosynthesis mechanism of PeGs, stabilizing and improving the content of PeGs in herbal materials, improving the quality, and synthesizing PeGs by microbial metabolic engineering or chemical synthesis.

Ameliorate effect and mechanism of verbascoside on memory impairment in hypoxic mice / 药学实践杂志

Objective To investigate the effects and mechanism of verbascoside on hypoxia-induced memory impairment.Methods The eight-arm maze was used to train mice′s spatial memory ability.After successful training, mice were randomly divided into five groups:a normoxic control group (distilled water, 0.1ml/10g), hypoxic model group (distilled water, 0.1ml/10g), the verbascoside low dose group (50 mg/kg), medium dose group (150 mg/kg), and high dose group (300mg/kg) were administered orally once a day for a total of 7days.After administration on the fourth day, except for the normoxic control group was placed in the animal room (1 500m), the remaining four groups were placed in a large-scale hypobaric chamber to simulate the hypoxic environment of the plateau (7 500m, 3days).Eight-armed maze test (4 000m) was used and the plasma and brain tissues were dissected out and measured for reactive oxygen species (ROS) in the brain, malondialdehyde (MDA), reduced glutathione (GSH) and total superoxide dismutase (T-SOD) activity in plasma and brain.Results Compared with the normoxic control group, the indexes of the eight-armed maze, ROS and MDA in the brain, MDA in the plasma of the hypoxia model group were significantly increased, and the GSH and T-SOD enzyme activities in the brain and plasma were notably decreased.Compared with the hypoxic model group, the indexes of the eight-armed maze, ROS and MDA in the brain, MDA in the plasma in the various groups of verbascoside were reduced more or less, the GSH and T-SOD enzyme activities in the brain and plasma slightly were increased.Conclusion Verbascoside could ameliorate the hypoxic memory impairment at high altitude, which might be related to the stabilization of the body′s antioxidant enzyme system balance and

Optimization of the Extraction Technology of Verbascoside from Cistanche tubulosa by Box-Behnken Response Surface Me- thodology / 中国药房

OBJECTIVE： To optimize the extraction technology of verbascoside from Cistanche tubulosa， and to provide reference for further development and comprehensive utilization of C. tubulosa. METHODS： The content of verbascoside in C. tubulosa was determined by HPLC. The determination was performed on  Inertsil-ODS-3V  column with  mobile phase consisted of methanol-0.2％ formic acid aqueous solution （40 ∶ 60， V/V） at  the flow rate  of  1 mL/min. The column temperature was 30 ℃， the detection wavelength was 330 nm， and the sample size  was 10 μL. Using extraction rate of verbascoside as index， soaking time， ethanol concentration， liquid-solid ratio， extraction time and extraction times were investigated by single factor tests. According to the results of above tests， ethanol concentration， liquid-solid ratio and extraction time were optimized by Box-Behnken response surface methodology. The verification tests were carried out on the optimized extraction technology. RESULTS： The linear range of verbascoside was 18.65-932.4 μg/mL. The optimal extraction technology included that ethanol concentration 63％， liquid-solid ratio 8 ∶ 1 （mL/g）， soaking for 2 h， extraction time 1.5 h， extracting for 2 times. The extraction rates of verbascoside in the three parallel verification tests were 78.21％， 76.95％， 79.34％， respectively. The relative errors of those to predicted value 76.76％ were 1.89％， 0.25％， 3.36％. CONCLUSIONS： The optimized extraction technology of verbascoside from C. tubulosa is stable and feasible， and is suitable for the extraction of verbascoside.

In vivo anti-arthritic and antioxidant effects from the standardized ethanolic extract of Moussonia deppeana

ABSTRACT Moussonia deppeana (Schltdl. & Cham.) Klotzsch ex Hanst., Gesneriaceae, known as tlachichinole, is a Mexican medicinal plant used for treatment of chronic inflammation-related diseases such as arthritis. In this paper, the main metabolite verbascoside was quantified in ethanolic extract; anti-arthritic and antioxidant activities were also evaluated in Complete Freund's Adjuvant induced arthritis in mice, with complete hematological evaluation, and oxidative stress measure in edema and ganglionic tissues on day 28. In popliteal ganglion, CD4+ lymphocytes and tumor necrosis factor alpha concentration were measured in addition to histological analysis. Ethanolic extract contained 79.2 mg of verbascoside/g extract, and this extract at 450 mg/kg generated an inhibition of 24% over paw edema development and increased body weight gain on final day. For hematological parameters, same dose decreased total leukocytes and lymphocytes, as well as decreased oxidation rate over biomolecules in edema and ganglionic tissues, and increased antioxidant enzyme activity. In ganglionic tissue, CD4+ lymphocytes and tumor necrosis factor alpha level showed no differences at any tested dose compared to complete Freund's adjuvant untreated group. Histological analysis of popliteal ganglion revealed moderate reduction of follicular hyperplasia, leukocyte infiltration and lipid inclusions at 450 mg/kg dose. Ethanolic extract of M. deppeana possesses anti-edematous activity associated to a moderate reduction in follicular hyperplasia, with immune-modulatory and antioxidant effects during experimental arthritis in mice.

Simultaneous Determination of Seven Components in Shenrong Guben Tablets by HPLC Wavelength Switching Method / 中国药师

Objective:To develop an HPLC wavelength switching method for the determination of seven components in Shenrong Guben tablets simultaneously.Methods:A Waters Sunfire C18column (250 mm ×4.6 mm,5 μm) was adopted. The column temperature was set at 30 ℃. The mobile phase was acetonitrile-0.1% phosphate solution with gradient elution. The flow rate was 0.9 ml·min-1. The detection wavelength were set at 330 nm for verbascoside and martinoside,230 nm for albiflorin and paeoniflorin,and 203 nm for ginsenoside Rg1, ginsenoside Re and ginsenoside Rb1. Results:The linear range of verbascoside, martinoside,albiflorin, paeoniflorin, ginsenoside Rg1, ginsenoside Re and ginsenoside Rb1was 6.38-159.50 μg·ml-1(r =0.999 3),3.19-79.75 μg·ml-1(r =0.999 9),4.37-109.25 μg·ml-1(r =0.999 5),14.26-356.50 μg·ml-1(r =0.999 4), 1.95-48.75 μg·ml-1(r = 0.999 8), 2.21- 55.25 μg·ml-1(r = 0.999 7) and 2.09- 52.25 μg·ml-1(r = 0.999 1), respectively. The average recovery and the corresponding RSD was 98.24% (1.11%),97.64% (1.43%),99.23% (0.80%), 100.13% (0.65%),96.99% (1.56%),98.10% (1.24%) and 97.75%(1.37%),respectively. Conclusion:The developed method can determine the contents of verbascoside, martinoside, albiflorin, paeoniflorin, ginsenoside Rg1, ginsenoside Re and ginsenoside Rb1in Shenrong Guben tablets simultaneously,which can be applied in the quality control of Shenrong Guben tablets.

Study of anti-oxidants of Rehmanniae Radix and Rehmannia Radix Praeparata by HPLC-UV-DPPH method / 中草药

Objective To develop an HPLC-UV-DPPH method to compare anti-oxidants of Rehmanniae Radix and Rehmanniae Radix Praeparata sample from different manufactories and to provide an effective method for the processing and quality control of traditional Chinese medicine. Methods HPLC in HPLC-UV-DPPH system was performed on Aglient Extend C18 (250 mm × 4.6 mm, 5 μm) column with gradient elution of acetonitrile-0.1% acetic acid at the flow rate of 1.0 mL/min, and the detection wavelength was at 334 nm. The column temperature was 30 ℃. Flow rate of DPPH solution is 0.5 mL/min, and detection wavelength was set at 517 nm. The total activities of the samples and the contribution rate of each component to the total activity were evaluated by the “quantity-effect” research idea, and verbascoside was regarded as a positive reference. The main anti-oxidants in original and processed herbs were identified by HPLC-FT-MS and were compared. Results The detection of HPLC-UV-DPPH method showed that there were nine anti-oxidants in Rehmanniae Radix extract, while 13 anti-oxidants were found in Rehmanniae Radix Praeparata. There were eight common anti-oxidants in the two herbs. The anti-oxidants were obviously different before and after Rehmanniae Radix processed. The activities of the antioxidants in different samples were markedly different. Anti-oxidants with higher contributions were mussaenosidic acid, echinacoside, jionoside A1/A2, verbascoside, and isoverbascoside, respectively. Conclusion The HPLC-UV-DPPH method is stable, sensitive, reproducible, and suitable for rapid screening of anti-oxidants and quality evaluation of Rehmanniae Radix and Rehmannia Radix Praeparata.

Quality standard for Pogostemon auricularius / 中成药

AIM To establish the quality standard for Pogonostemon auricularius (L.) Hassk..METHODS The original,characteristic and microscopic features of P.auricularius were identified,after which TLC was used for qualitative identification,HPLC was adopted in the content determination of verbascoside,and the contents of water,total ash,acid-insoluble ash,extract were determined by Chinese Pharmacopoeia methods.RESULTS The clear TLC plots displayed good reproducibility.Verbascoside showed a good linear relationship within the range of 0.102-2.04 μg (r =0.999 98),whose average recovery was 97.99％ with the RSD of 2.02％.In four-teen batches of samples,the content ranges of water,total ash,acid-insoluble ash,extract were 6.47％-12.07％,7.59％-12.35％,0.23％-3.55％,13.28％-26.82％,respectively.CONCLUSION This stable and reliable method can be used for the quality control of P.auricularius

Decoction Amount Changes of Verbascoside and Gallic Acid in Single Decoction and Combined Decoction of Rehmanniae Radix Preparata,Corni Fructus and Poria / 中国药房

OBJECTIVE:To investigate the decoction amount changes of verbascoside,gallic acid in single decoction and com-bined decoction of rehmanniae radix preparata,corni fructus and poria,and provide scientific basis for further study of effective substance basis. METHODS:The single decoction,combined decoction with each 2 medicine and combined decoction with the 3 medicines of rehmanniae radix preparata,corni fructus and poria,were respectively prepared. HPLC was adopted to detect and com-pare the decoction amount of active component verbascoside in rehmanniae radix preparata and active component gallic acid in cor-ni fructus in each group. RESULTS:Compared with single decoction of rehmanniae radix preparata,the decoction amount of ver-bascoside was decreased in combined decoction of poria+rehmanniae radix preparata(P<0.01),decoction amount of verbascoside was increased in combined decoction of corni fructus+rehmanniae radix preparata or combined decoction with the 3 medicines(P<0.01). Compared with single decoction of corni fructus,the decoction amount of gallic acid in each combined decoction was de-creased (P<0.01). CONCLUSIONS:The combined decoction of rehmanniae radix preparata,corni fructus and poria has certain promotion and inhibition effects on the decoction of verbascoside in rehmanniae radix preparata and inhibition effect on the decoc-tion of gallic acid in corni fructus. It is speculated verbascoside may be one of the main components in combined decoction playing the role of effectiveness.

Study on the Extraction Technology for Active Constituents of Tibetan Medicine Pedicularis kansuensis / 中国药房

OBJECTIVE:To optimize the extraction technology of Tibetan medicine Pedicularis kansuensis and compare con-tent of verbascoside and isoverbascoside differences in P. kansuensis from various habitats. METHODS:Using verbascoside and iso-verbascoside and dry paste yield as comprehensive evaluation indexes,single factor test and orthogonal test were used to investigate the extraction solvent,solvent dosage,extraction time and times to optimize extraction technology,and the verification test was conducted. Contents of the 2 constituents verbascoside and isoverbascoside in P. kansuensis from Gansu,Qinghai and Sichuan were compared. RESULTS:The optimal extraction technology was as follows as 8-fold 50% ethanol,extraction for 3 times,90 min each time. The verification results showed that the average contents of verbascoside and isoverbascoside were 3.49%(RSD=1.28%,n=3),1.26%(RSD=1.32%,n=3),and average dry paste yields were 37.99%(RSD=1.97%,n=3). The contents of verbascoside and isoverbascoside in P. kansuensis from Qinghai were relatively higher. CONCLUSIONS:Optimized extraction tech-nology is reasonable,stable,feasible;the contents of index constituents in P. kansuensis from different habitats have certain differ-ences. The study can provide scientific evidence for the development and utilization of extraction,and the in-depth study of quality evaluation for medicinal material.

Reduction of iron deposition by verbascoside in HO-1 overexpression rats astrocytes / 中国药学杂志

OBJECTIVE: To investigate the effect of verbascoside on iron deposition of astrocyte cell overexpressing heme oxygenase-1 gene, and to seek new drug in treating Alzheimer's disease. METHODS: Primary cultured astrocyte cells of rats were cultured and purified in vitro. With gene recombination and transfer techniques, transfect HO-1 gene into primary cultured astrocyte cells. HO-1 transfected astrocyte were then treated by different concentrations of verbascoside. Proliferation of cell was measured by CCK-8 assay and expression of HO-1 protein was analyzed by Western blotting. Iron deposition was also visualized by Perl's Prussian blue stain. RESULTS: Immunofluorescence staining showed that most of the cells were GFAP-postitive and the purity was over 95%. The expression of HO-1 protein was increased in HO-1 injury model of astrocyte. Treatment of verbascoside with variable concentration caused the decrease of HO-1 protein expression more or less and reduction of iron deposition in transfected cells(20 μmol·L-1 reducing 47.69%, 40 μmol·L-1 reducing 51.63%). CONCLUSION: Verbascoside could enhance astrocyte cell resistance against injury induced by overexpression of heme oxygenase-1 gene.

Photoprotective effect of verbascoside from Buddleja cordata in SKH-1 mice exposed to acute and chronic UV-B radiation / Efecto fotoprotector del verbascósido de Buddleja cordata en ratones SKH-1 expuestos de forma aguda y crónica a laradiación UV-B

Ultraviolet radiation (UVR) is involved in both sunburn and the development of skin cancer, which has a high incidence worldwide. Strategies to reduce these effects include the use of photoprotective substances. The aim of this work was to investigate the photoprotective effect of verbascoside isolated from the methanolic extract of Buddleja cordata (BCME) in SKH-1 mice exposed to acute and chronic UV-B radiation. The mouse dorsal area was evaluated macroscopically and microscopically for diagnosis; verbascoside penetration into mouse skin was investigated in vivo by the tape stripping method. After acute UV-B exposure, 100 percent of irradiated mice that had been protected with verbascoside showed no signs of sunburn or of inflammatory processes. After chronic exposure, 100 percent of unprotected mice showed skin carcinomas; in contrast, in mice topically treated with either BCME or verbascoside, the presence of lesions was decreased by 90 percent. These results prove that verbascoside penetrates through the skin of mice and suggest that verbascoside and BCME may potentially prevent photodamage on mice’s skin after acute and chronic UVR exposure.

La radiación ultravioleta (RUV) provoca quemaduras solares y el desarrollo de cáncer de piel. El objetivo de este trabajo fue investigar el efecto fotoprotector del verbascósido obtenido del extracto metanólico de Buddleja cordata (EMBC) en ratones SKH-1 expuestos a RUV-B de manera aguda y crónica. El diagnóstico histológico se llevó a cabo en la piel de la zona dorsal de los ratones. La penetración del verbascósido fue cuantificada mediante la técnica de la cinta adhesiva. En el experimento agudo, el 100 por ciento de los ratones protegidos con verbascósido no evidenciaron signos de quemadura ni procesos inflamatorios. En el experimento crónico los ratones sin protección e irradiados presentaron carcinomas cutáneos. En contraste en los ratones protegidos con EMBC o verbascósido las lesiones disminuyeron un 90 por ciento en ambos grupos. El verbascósido penetró en la piel del ratón. Los resultados sugieren que el EMBC y el verbascósido previenen el fotodaño en la piel de ratones expuestos de forma aguda o crónica a la RUV.

Preparation technology of verbascoside and isoverbascoside from Pedicularis kansuensis / 中草药

Objective: To establish a preparation method for verbascoside and isoverbascoside with high purity from Pediculariskansuensis. Methods: With the contents of verbascoside and isoverbascoside as indexes, static and dynamic adsorption-desorption were used to select the best macroporous resin from 01A1, D101, HPD100, AB-8, XAD-6, DM130, DM-301, DM-201, YWD06B, and YWD06C. The purification technology parameters of total phenylethanoid glycosides (PhGs) from P. kansuensis were optimized by selected macroporous resin. Medium pressure column chromatography was further used to purify verbascoside and isoverbascoside. The chemical structures were identified on the basis of the spectral data. Results: D101 type resin showed the best purifying profile, its optimum technology conditions were as follows: Sample injection concentration was 20 mg/mL, sample flow rate was 3 BV/h, then washing off the impurity with 3 BV deionized water, and being eluted by 5 BV 50% ethanol with the desorption rate at 2 BV/h. The 50% ethanol fraction was evaporated and dried with cry desiccation to obtain the total PhGs. The contents of verbascoside and isoverbascoside in total PhGs were 42.29% and 28.51%, respectively. After purified by medium pressure column chromatography, the contents of prepared verbascoside and isoverbascoside reached 98.3% and 99.2%, respectively. Conclusion: This method can be used for the large-scale preparation of verbascoside and isoverbascoside with high purity, which is rapid, simple, and practicable with high efficiency.

Content Determination of Verbascoside in Naosaitong Honeyed Pill by HPLC / 中国药房

OBJECTIVE:To establish a method for the content determination of verbascoside in Naosaitong honeyed pill. METHODS:HPLC was performed on the column of Zorbax ODX with mobile phase of acetonitrile-0.1% phosphoric acid(16∶84, V/V)at a flow rate of 1.0 ml/min,the detection wavelength was 334 nm,column temperature was 30 ℃,and the injection volume was 10 μl. RESULTS:The linear range of verbascoside was 0.200-1.000 μg(r=0.999 8);RSDs of precision,stability and repro-ducibility tests were lower than 1%;recovery was 95.16%-99.78%(RSD=1.80,n=6). CONCLUSIONS:The method is simple and accurate,and suitable for the content determination of verbascoside in Naosaitong honeyed pill.