ABSTRACT
OBJECTIVE To investigate the impacts of isorhynchophylline （IRN） on airway inflammation in asthmatic mice by regulating the monocyte chemotactic protein-1 （MCP-1）/CC chemokine receptor 2 （CCR2） signaling pathway. METHODS The asthmatic mice model was established by injecting and inhaling ovalbumin. The successfully modeled mice were randomly grouped into asthma group， IRN low-dose group （IRN-L， intragastric administration of 10 mg/kg IRN）， IRN high-dose group （IRN-H， intragastric administration of 20 mg/kg IRN）， IRN-H+CCL2 group [intragastric administration of 20 mg/kg IRN+intraperitoneal injection of 7.5 ng CC chemokine ligand 2 （CCL2）] and positive control group （intraperitoneal injection of 2 mg/kg dexamethasone）. The mice injected and inhaled with sterile phosphate-buffered solution were included in the blank control group， with 10 mice in each group. The mice in administration groups were given relevant medicine once a day， for consecutive 2 weeks. The levels of airway hyperreactivity indexes such as enhanced （Penh） value， tumor necrosis factor-α （TNF-α），interleukin-13 （IL-13） and IL-4 in serum， the number of eosinophil （EOS）， lymphocyte （LYM） and neutrophils （NEU） in alveolar lavage fluid and the protein expressions of MCP-1 and CCR2 in lung tissue were observed in each group； the pulmonary histopathological changes were observed， and inflammatory cell infiltration score was evaluated. RESULTS Compared with the blank control group， the infiltration of inflammatory cells in the lung tissue of mice was more significant in the asthma group， and there was swelling and shedding of cells； inflammatory infiltration score， Penh value， the levels of IL-4， IL-13 and TNF-α， the number of EOS， NEU and LYM， the protein expressions of MCP-1 and CCR2 were increased significantly （P＜0.05）. Compared with the asthma group， the pathological injuries of the IRN-L group， IRN-H group and positive control group were improved， and the above quantitative indexes were decreased significantly （P＜0.05）. Compared with the IRN-L group， the above quantitative indexes of the IRN-H group and positive control group were decreased significantly （P＜0.05）. There was no statistical significance in the above quantitative indexes between the IRN-H group and the positive control group （P＞0.05）. Compared with the IRN-H group， the above quantitative indexes of the IRN-H+CCL2 group were increased significantly （P＜0.05）. CCL2 reversed the protective effect of high-dose IRN on asthmatic mice. CONCLUSIONS IRN may reduce the release of airway inflammatory factors in asthmatic mice by inhibiting the activation of the MCP-1/CCR2 signaling pathway， so as to achieve the purpose of improving asthma.
ABSTRACT
ObjectiveTo determine the contents of four kinds of indole alkaloids （rhynchophylline， isorhynchophylline， corynoxeine， isocorynoxeine） in Uncaria by high-performance liquid chromatography-mass spectrometry-automatic internal standard （HPLC-MS-AIS）. MethodsChromatographic separation was performed using C18 column （3.0 mm×50 mm， 3.3 μm）， and the mobile phase， comprising 0.1% formic acid aqueous solution-acetonitrile （82∶18， V/V）， was eluted at a flow rate of 0.5 mL/min and column temperature of 30℃. Mass spectrometric detection was performed using an electrospray ionization source and positive multiple-reaction monitoring mode at a voltage capillary of 4 000 V. The mass-to-charge ratio （m/z） transition was 385.25/160.10 for rhynchophylline， 385.30/160.10 for isorhynchophylline， 383.25/160.15 for corynoxeine and 383.25/160.15 for isocorynoxeine， respectively. The injection volume was kept constant at 2 μL. ResultsThe linear concentration ranges of rhynchophylline， isorhynchophylline， corynoxeine and isocorynoxeine were 2.30~600.00 ng/mL （r=0.999 3）， 2.30~600.00 ng/mL （r=0.999 2）， 2.47~650.00 ng/mL （r=0.999 4） and 2.47~650.00 ng/mL （r=0.999 2）， respectively. The relative standard deviation （RSD） of precision and stability were all lower than 5.00%， the accuracy ranged from 92.40% to 104.10%， and the average recovery was 95.90%~104.60%. ConclusionHPLC-MS-AIS method is simple and accurate for the determination of four kinds of alkaloids in Uncaria， and can be used as a new method for quality control of Uncaria.
ABSTRACT
ObjectiveTo develop a two-dimensional high-performance liquid chromatography （2D-HPLC） for simultaneous determination of the contents of four kinds of Uncaria alkaloids： rhynchophylline， isorhynchophylline， corynoxeine and isocorynoxeine. MethodsThe 2D-HPLC apparatus was comprised of a first chromatographic column in version Aston SC2 （3.5 mm×25 mm， 5 μm）， an intermediate column in version Aston SH C18 （3.5 mm×10 mm， 5 μm）， and an analytical column in version Aston SCB （4.6 mm×125 mm， 5 μm）. The mobile phase of the first and second liquid chromatography system were CAA-1 and mixed mobile phase （V BPI-1 basic mobile phase ∶ V MPI-1 mobile phase ∶ V OPI-1 organic mobile phase = 45∶14∶41）. The chromatographic parameters included a flow rate of 1.0 mL/min， a column temperature of 40℃， a wavelength of 254 nm， an injection volume of 500 μL and a detection time of 9.5 min. ResultsThe linear ranges of rhynchophylline， isorhynchophylline， corynoxeine and isocorynoxeine were 9.77~10 000.00 ng/mL （r=0.999 6）， 10.74~11 000.00 ng/mL （r=0.999 7）， 10.74~11 000.00 ng/mL （r=0.999 7）， 10.74~11 000.00 ng/mL（r=0.999 6）， respectively. The relative standard deviation （RSD） of precision， stability and repeatability were all less than 5.00%. The accuracy was 95.20%~104.01%， and the recovery rate was 93.63%~101.38%. ConclusionThe 2D-HPLC developed for simultaneous determination of four kinds of alkaloids in Uncaria is simple and accurate， which can be used as a new method for quality control of Uncaria.
ABSTRACT
Gastrodiae Rhizoma-Uncariae Ramulus cum Uncis is the most frequently used herbal pair in the treatment of Parkinson's disease(PD). Gastrodin and isorhynchophylline are important components of Gastrodiae Rhizoma-Uncariae Ramulus cum Uncis herb pair with anti-Parkinson mechanism. This study aimed to investigate the effect of gastrodin combined with isorhynchophylline on 1-methyl-4-phenylpyridinium(MPP~+)-induced apoptosis of PC12 cells and their antioxidant mechanism. The leakage of lactate dehydrogenase(LDH) from cells to media was analyzed by spectrophotometry. Apoptotic cells were labeled with Annexin V-fluorescein isothiocyanate(FITC) and propidium iodide(PI) and analyzed by flow cytometry. The cell cycle was analyzed using propidium iodide(PI) staining. Lipid peroxidation(LPO) level was analyzed by spectrophotometry. The mRNA expression of caspase-3 was examined by Real-time RT-PCR. The protein expressions of heme oxygenase 1(HO-1) and NADPH: quinoneoxidore-ductase 1(NQO-1) were determined by Western blot. Gastrodin combined with isorhynchophylline reduced the percentage of Annexin V-positive cells and cell cycle arrest in MPP~+-induced PC12 cells. Gastrodin combined with isorhynchophylline down-regulated the mRNA expression of caspase-3, up-regulated the protein expressions of HO-1 and NQO-1, and reduced LPO content in MPP~+-induced PC12 cells. PD98059, LY294002 or LiCl could partially reverse these changes pretreated with gastrodin combined with isorhynchophylline, suggesting that gastrodin combined with isorhynchophylline inhibited MPP~+-induced apoptosis of PC12 cells and oxidative stress through ERK1/2 and PI3 K/GSK-3β signal pathways. Our experiments showed that gastrodin combined with isorhynchophylline could down-re-gulate the mRNA expression of caspase-3 and up-regulate the protein expressions of HO-1 and NQO-1, so as to reduce oxidative stress and inhibit apoptosis.
Subject(s)
Animals , Rats , 1-Methyl-4-phenylpyridinium/toxicity , Antioxidants , Apoptosis , Benzyl Alcohols , Cell Survival , Glucosides , Glycogen Synthase Kinase 3 beta , Oxindoles , PC12 CellsABSTRACT
OBJECTIVE： To establish a method to determine the contents of rhynchophylline and isorhynchophylline in Uncaria rhynchophylla. METHODS： The separation degree of ionic liquid 1-butyl-3-methylimidazolium chloride （C4mimCl） as mobile phase additive was compared with that of mobile phase without additives and with traditional additive triethylamine （which damaged the chromatographic column）. The optimum concentration of C4mimCl was screened and the contents of rhynchophylline and isorhynchophylline in U. rhynchophylla from 4 habitats in Jiangxi province were determined by the newly established method. The determination was performed on Dikmatech Diamonsil Plus C18 column， the mobil phase was acetonitrile-buffer （0.1％ phosphoric acid+3.0 mmol/L C4mimCl）， gradient elution. UV detection wavelength was set at 245 nm and the flow rate was 1 mL/min. Sample size was 10 μL. RESULTS： When mobile phase had no additives or 3.0 mmol/L triethylamine and 3.0 mmol/L C4mimCl were added as additives， the separation of rhynchophylline from the front peak was 1.02， 1.23 and 1.72， and the separation from the back peak was 1.06， 6.00 and 4.25， respectively. The symmetry factors were 0.81， 0.86 and 1.13， respectively. The separation of isorhynchophylline from the front peak was 0.96， 3.89 and 4.05， and the separation from the back peak was 1.02， 2.34 and 2.36， respectively. The symmetry factors were 0.88， 0.81 and 0.96， respectively. The linear range of rhynchophylline and isorhynchophylline were 4.93-157.76 （r＝0.999 9） and 4.98-159.50 μg/mL （r＝1.000）， respectively. The quantitative limits were 0.486 4， 0.793 6 μg/mL， respectively. RSDs of precision， repeatability， stability and durability tests were all less than 5％ （n＝6）. The recovery rates were 102.9％-107.8％ （RSD＝1.7％，n＝6） and 95.4％-106.3％ （RSD＝3.9％，n＝6）， respectively. The content of rhynchophylline and isorhynchophylline in U. rhynchophylla from 4 habitats were 0.758-1.343 and 1.511-1.823 mg/g， respectively. CONCLUSIONS： Addition of C4mimCl into mobile phase can enhance its separation. Established HPLC method is rapid， accurate and reproducible， which can be used for content determination of rhynchophylline and isorhynchophylline in U. rhynchophylla.
ABSTRACT
Objective To explore the effect of rhynchophylline and isorhynchophylline on headshakes behavior and levels of monoamine neurotransmitter in model rats with Tourette syndrome.Methods 40 Wistar rats were randomly divided into DOI-induced head-shakes rats (HSR group),haloperidol group,rhynchophylline group and isorhynchophylline group with 10 in each group.The inhibitory effects of rhynchophylline and isorhynchophylline were estimated by observing the HSR behavior.Dopamine (DA) and 5-hydroxytryptamine(5-HT) in the rat striatum were detected by the enzyme-linked immunosorbent assay (ELISA) method.The 5-HT2A receptor mRNA expression in prefrontal lobe cortex of the rats was measured by real-time PCR.Results Compared with HSR group,the head shakes of the rats in haloperidol group and isorhynchophylline group were significantly decreased(P＜0.01),and no change of head-shakes number was observed in rhynchophylline group (P＞0.05).There was no significant difference of head-shakes number between the haloperidol group and isorhynchophylline group(P＞0.05).Compared with HSR group,DA levels in the rat striatum were significantly decreased in isorhynchophylline group and haloperidol group((152.35± 5.80) μ~L vs (111.19±4.30) μg/L,(152.35±5.80) μg/L vs (126.42±3.17) μg/L,P＜0.01),while DA levels in the rat striatum in rhynchophylline group were not changed ((152.35±5.80) μg/L vs (142.71±5.51) μg/L,P＞0.05).There was no significant change of 5-HT2A receptor mRNA expression in rat prefrontal lobe cortex in every group(P＞0.05).Conclusion Isorhynchophylline may have an inhibitory effect on rats with DOI-induced HSR.Isorhynchophylline may decrease the DA levels in the rat stratum with DOI-induced HSR.Rhynchophylline has no significant inhibitory effect on head-shakes behavior and DA levels in the rat stratum with DOI-induced HSR.
ABSTRACT
Isorhynchophylline ( IRN) is an alkaloid isolated from Uncaria rhynchophylla .IRN mainly acts on cardiovascular and central nervous systems diseases including hypertension , brachycardia and sedation .In recent years , it has been found that IRN has widespread pharmacological effects in central nervous disease .This review aim at reviewing neuroprotective activities of Isorhynchophyl-line on Alzheimer′s disease and other central nervous systems diseases via improving learning and memory impairments , inhibiting Tau protein hyperphosphorylation , antagonizing β-Amyloid-iInduced neurotoxicity , and reducing glia-secretory inflammatory factors .
ABSTRACT
Objective To study the content of isorhynchophylline from different regions and different harvest time.Methods The content of isorhynchophylline in Uncaria rhynchophylla from different regions and different harvest time was tested by HPLC. The Waters Symmetry C18 color (4.6 mm× 250 mm, 5 μm), mobile phase of methanol - 0.01 mol/L ammonium acetate buffer (pH 8.0) (60:40), column temperature 25℃, 20 μl sample volume, velocity of 1.0 ml/min, detection wavelength of 246 nm were setted.Results The isorhynchophylline can be detected in different regions and different harvest time, with the highest content was found in Ningming county and from September to February of the next year.Conclusion The content of isorhynchophylline in Uncaria rhynchophylla varied from different regions and harvest times. The best harvest time is during September to February of the next year.
ABSTRACT
ObjectiveTo explore the content of isorhynchophylline in different medicinal parts of Ramulus Uncariae Cum Uncis.MethodsHPLC was adopted to determine 8 batches of different Ramulus Uncariae Cum Uncis of different origin and different medicinal parts. The Waters Symmetry C18 color(4.6 mm×250 mm, 5μm) was used with mobile phase of methanol -0.01 mol/L ammonium acetate buffer(pH 8.0)(60∶40), column temperature of 25℃ , 20μl sample volume, velocity of 1.0 ml/min, and detection wavelength of 246 nm. ResultsIsorhynchophylline can be detected in all 8 batchs of Ramulus Uncariae Cum Uncis in different regions, and different content were found among different origins. The content of isorhynchophylline in different parts of the same origin showed a decreasing sequence of the rhabd, stem with hook, stem without hook, and twig without hook and leaves.ConclusionMost of medicinal part of Ramulus Uncariae Cum Uncis contains isorhynchophylline, which provide a lab basis for exploring medicinal parts of this herbal medicine.
ABSTRACT
AIM : To establish the method for determining tetrahydropalmatine,rhynchophylline and isorhynchophylline in AnshenYangxue Oral Liquid (Ramulus uncariae cum uncis, stephania kwangsiansis, Radix polygoni multiflori praeparata cum succo glycinus sotae , Caulis Polygoni multiflori and Pine needle). METHODS: Tetrahydropalmatine,rhynchophylline and isorhynchophylline were determined by HPLC. Chromatographic condition was composed of Kromasil C_(18) column, a mixture of methanol and water(55: 45) as mobile phase with 0.01 mol/L triehthylamine, adjusted with acetic acid to pH of 7.5, UV detection wavelength of rhynchophylline and isorhynchophylline was set at 254 nm, UV detection wavelength of tetrahydropalmatine was set at 281 nm. RESULTS: The averagere recoveries of tetrahydropalmatine,rhynchophylline and isorhynchophylline were 98.47% ,99.04% and 98. 75% respectively; RSD were 0.95% ,2.6% and 1.6%, respectively. CONCLUSION: This method is simple, sensitive, accurate, and can be used for determining tetrahydropalmatine ,rhynchophylline and isorhynchophylline in Anshen Yangxue Oral Liquid.
ABSTRACT
Objective To establish a pretreatment mthod of the tissue sample of rat and a determination method of the concentration of isorhynchophylline in tissue of rats by RP-HPLC. Method The tissue sample were treated by adding methanol, and the supernatant was dried by nitrogen gas, the residue was redissolved in methanol, then detected by RP-HPLC directly. Waters? RP18 (4.6 mm?250 mm, 5 ?m) and pre-column Wondasil C18 (4.0 mm?10 mm, 5 ?m) were used as the stationary phase. Methanol-water (70∶30) added 0.03% triethylamine was taken as mobile phase with flow rate of 0.8 mL/min. The wavelength of detection was 254 nm. Results The standard curve of Isorhynchophylline concentraton in rat tissues:Y=48.558X+0.207 6 (n=6), r=0.999 6, it appears a good linear relationship within the range of 0.16～16.0 ?g/mL. The average recovery of was 99.66%, RSD was 1.56%. Conclusion This mehod is sensitive, rapid, simple, specific and reproducible.
ABSTRACT
Aim To investigate the effect of Isorhynchophylline(Isorhy)on platelet aggregation or thrombosis,and explore the mechanism of it's action.Methods Rat platelet aggregation was determined.cAMP contents were assessed by Born's method and radioimmunoassay respectively.The effect of Isorhy on rat's thrombosis was observed with the thrombogenesis model of artery-vein bypass.Results Isorhy(0.65 mmol?L-1,1.30 mmol?L-1)was shown to markedly inhibit the rat's platelet aggregation induced by ADP or thrombin in a concentration-dependent manner(P
ABSTRACT
Hypotensive effects of rhynchophylline, isorhynchophylline, total alkaloid and non-alka-loid fraction isolated from Uncaria rhynchophylla (Miq.) Jacks. were compared. Change in peripheralblood pressure was recorded by inserting a catheter into the right common carotid artery of anaethetizedrat, while each of the four hypotensive constituents was injected individually into the femoral vein by a mi-croinfusion pump. Results of the findings showed that the four constituents in U. rhynchophylla displayeddifferent hypotensive potency in the order of isorhynchophylline [lowering of mean arterial pressure(MAP) by 42.0%] > rhynchophylline (lowering of MAP by 32.1%) > total alkaloid (lowering of MAPby 21.3%) > non-alkaloid fraction (lowering of MAP by 12.4%). It was concluded that isorhyn-chophylline and rhynchophylline were the main hypotensive constituents in U. rhynchophylla.
ABSTRACT
AIM: To establish the method for determining tetrahydropalmatine、rhynchophylline and isorhynchophylline in AnshenYangxue Oral Liquid(Ramulus uncariae cum uncis, stephania kwangsiansis,Radix polygoni multiflori praeparata cum succo glycinus sotae,Caulis Polygoni multiflori and Pine needle). METHODS: Tetrahydropalmatine、rhynchophylline and isorhynchophylline were determined by HPLC.Chromatographic condition was composed of Kromasil C_18 column,a mixture of methanol and water(55∶45) as mobile phase with 0.01 mol/L triehthylamine,adjusted with acetic acid to pH of 7.5,UV detection wavelength of rhynchophylline and isorhynchophylline was set at 254 nm,UV detection wavelength of tetrahydropalmatine was set at 281 nm. RESULTS: The averagere recoveries of tetrahydropalmatine、rhynchophylline and isorhynchophylline were 98.47%、99.04%and 98.75% respectively;RSD were 0.95%、2.6%and 1.6%,respectively. CONCLUSION: This method is simple,sensitive,accurate,and can be used for determining tetrahydropalmatine、rhynchophylline and isorhynchophylline in Anshen Yangxue Oral Liquid.
ABSTRACT
Aim To study the effect of rhynchophylline and isorhynchophylline on the proliferation of rat vascular smooth muscle cell(VSMCs)induced by angiotensin Ⅱ(AngⅡ)and discuss it's mechanism.Methods A cell proliferating model of VSMC induced by AngⅡ was established.The proliferative activity of VSMC with rhynchophylline and isorhynchophylline was analyzed by MTT method.The effect of rhynchophylline and isorhynchophylline on cell cycle of VSMC was observed by flow cytometry.The effect of rhynchophylline and isorhynchophylline on protein of AT1R gene,NF-?B gene,Stat3 gene,c-Myc gene and c-Fos gene in VSMCs was observed by flow cytometry.The effect of rhynchophylline and isorhynchophylline on c-Myc mRNA and c-Fos mRNA in VSMCs was observed by RT-PCR.Results The proliferation of VSMC induced by AngⅡ stimulation was inhibited by rhynchophylline and isorhynchophylline,and the cell numbers of G0/G1 phase were increased and those of S phase were decreased markedly.At the same time,high expression of the protein of c-Myc gene,c-Fos gene,AT1R gene,NF-?B gene and stat3 gene stimulated by AngⅡ could be inhibited by rhynchophylline and isorhynchophylline significantly,and high expression of c-Myc gene mRNA and c-Fos gene mRNA stimulated by AngⅡ could also be inhibited by rhynchophylline and isorhynchophylline significantly.Conclusions The proliferation of VSMCs induced by AngⅡ can be inhibited by rhynchophylline and isorhynchophylline.The mechanisms are related to preventing VSMCs from G0/G1 phase entering S phase,and down regulating the expression of protein of AT1R gene,NF-?B gene,Stat3 gene,c-Myc gene and c-Fos gene.In addition,c-Myc gene mRNA and c-Fos gene mRNA expression are down regulated as well.
ABSTRACT
The effects of Rhynchophylline (Rhy) and Isorhychophyiline (Isorhy), the alkaloids abstracted from the Chinese traditional herb Uncaria rhynchophyllia (Miq) Jackson, on the 45Ca-influx and efflux were investigated in rabbit aorta. Both Rhy and Isorhy (10 ?mol? L-1) inhibited the 45Ca-influx induced by high K+(77. 0 mmol ? L-1), but neither significant-ly influenced the 45Ca-influx and efflux induced by noradrenaline (10 ?mol ? L-1). The results suggest that these alkaloids block the Voltage-dependent calcium channel.