Effects of salinomycin on proliferation and apoptosis of oral squamous cell carcinoma / 北京大学学报(医学版)

OBJECTIVE@#To investigate the effects of salinomycin on the proliferation and apoptosis of oral squamous carcinoma cells and to further understand the mechanisms of these effects.@*METHODS@#The human oral squamous carcinoma cell line CAL-27 was cultured in different concentrations of salinomycin and cisplatin. After co-culture with 0, 1, 2, 4, 8, 16 and 32 μmol/L salinomycin or 0, 1.25, 2.5, 5, 10, 20, 40 and 80 μmol/L cisplatin for 24 hours and 48 hours, the proliferation of oral squamous carcinoma cells were detected by cell counting kit-8(CCK-8) assay. After being exposed to 0, 2, 4, 8 μmol/L salinomycin and 0, 5, 10, 20 μmol/L cisplatin for 48 hours, the cell cycle of oral squamous carcinoma cells was detected by flow cytometry assay, and Western blot analysis was performed to analyze the expressions of cysteine-containing aspartate-specific proteases-3(Caspase-3), cysteine-containing aspartate-specific proteases-9(Caspase-9), poly ADP-ribose polymerase (PARP), protein kinase B (Akt) and phosphorylated protein kinase B (p-Akt) protein in oral squamous carcinoma cells.@*RESULTS@#Both salinomycin and cisplatin significantly inhibited the proliferation of oral squamous cell carcinoma CAL-27 cells in a time- and dose-dependent manner. However, compared with the first-line chemotherapeutic drug cisplatin, salinomycin showed stronger anti-proliferation activity in oral squamous carcinoma cells than cisp-latin (P < 0.001). After being exposed to 8 μmol/L salinomycin, CAL-27 cells exhibited markedly higher proportion in quiescent/ first gap phases (40.40%±1.99% vs. 64.46%±0.90%, P < 0.05), and had a significantly lower proportion in synthesis phases and second gap / mitosis phases (24.32%±2.30% vs. 18.73%±0.61%, P < 0.05; 35.01%±1.24% vs. 16.54%±1.31%, P < 0.05) compared with the dimethyl sulfoxide control group; moreover cisplatin didn't show cell-cycle specific effect on CAL-27. Western blot proved that salinomycin could up-regulate the expressions of Caspase-3 and Caspase-9 protein in oral squamous cell carcinoma CAL-27 cells (P < 0.05). At the same time, the levels of PARP, Akt and p-Akt protein were down-regulated (P < 0.05).@*CONCLUSION@#Compared with cisplatin, salinomycin has a better inhibitory effect on the proliferation of oral squamous carcinoma cells and blocks the cell cycle process at the quiescent / first gap phase. At the same time, salinomycin could trigger apoptosis of oral squamous carcinoma cells and the mechanism is associated with the Akt/p-Akt signaling pathway.

Isolation and identification of endophytic fungi producing harpagoside and harpagide from Scrophularia ningpoensis / 中国中药杂志

The endophytic fungi from root,main stem,branch and leaf of Scrophularia ningpoensis were isolated from Zhejiang,whether these strains could yield harpagide or harpagoside were tested by HPLC and LC-MS. According to the morphological characteristic and the similarity of the nucleotide sequence of internal transcribed spacer( ITS) between r DNAs,the strains producing harpagide or harpagoside were identified. The results showed that 210 strains were isolated from the samples,which were classified into 9 orders,13 families and 17 genera by morphological study. Harpagide was detected in endogenous fungi ZJ17 and harpagoside was detected in endogenous fungi ZJ25 by HPLC coupled with LC-MS. ZJ17 was identified as Alternaria alternate and ZJ25 was identified as A.gaisen by its morphology and authenticated by ITS( ITS4 and ITS5 regions and the intervening 5. 8 S rDNA region).

Overexpression of autophagy-related gene 3 promotes autophagy and inhibits salinomycin-induced apoptosis in breast cancer MCF-7 cells / 南方医科大学学报

OBJECTIVE@#To study the effects of the overexpression of autophagy-related gene 3 (ATG3) on autophagy and salinomycin-induced apoptosis in breast cancer cells and explore the underlying mechanisms.@*METHODS@#We used the lentivirus approach to establish a breast cancer cell line with stable overexpression of ATG3. Western blotting, immunofluorescence staining and transmission electron microscopy were used to analyze the effect of ATG3 overexpression on autophagy in breast cancer MCF-7 cells. Using the AKT/mTOR agonists SC79 and MHY1485, we analyzed the effect of AKT/mTOR signal pathway activation on ATG3 overexpression-induced autophagy. Western blotting and flow cytometry were used to analyze the effect of autophagy on apoptosis of the ATG3-overexpressing cells treated with salinomycin and 3-MA (an autophagy inhibitor).@*RESULTS@#In ATG3-overexpressing MCF-7 cells, ATG3 overexpression obviously promoted autophagy, inhibited the AKT/mTOR signaling pathway, significantly weakened salinomycin-induced apoptosis ( < 0.01), caused significant reduction of the levels of the pro-apoptotic proteins cleaved-caspase 3 ( < 0.01) and Bax ( < 0.05), and enhanced the expression of the anti-apoptotic protein Bcl-2 ( < 0.05). The inhibition of autophagy obviously weakened the inhibitory effect of ATG3 overexpression on salinomycin-induced apoptosis.@*CONCLUSIONS@#ATG3 overexpression promotes autophagy possibly by inhibiting the AKT/mTOR signaling pathway to decrease salinomycin-induced apoptosis in MCF-7 cells, suggesting that autophagy induction might be one of the mechanisms of drug resistance in breast cancer cells.

Salinomycin enhances doxorubicin sensitivity through reversing the epithelial-mesenchymal transition of cholangiocarcinoma cells by regulating ARK5

Chemotherapy response rates in patients with cholangiocarcinoma remain low, primarily due to the development of drug resistance. Epithelial-mesenchymal transition (EMT) of cancer cells is widely accepted to be important for metastasis and progression, but it has also been linked to the development of chemoresistance. Salinomycin (an antibiotic) has shown some potential as a chemotherapeutic agent as it selectively kills cancer stem cells, and has been hypothesized to block the EMT process. In this study, we investigated whether salinomycin could reverse the chemoresistance of cholangiocarcinoma cells to the chemotherapy drug doxorubicin. We found that combined salinomycin with doxorubicin treatment resulted in a significant decrease in cell viability compared with doxorubicin or salinomycin treatment alone in two cholangiocarcinoma cell lines (RBE and Huh-28). The dosages of both drugs that were required to produce a cytotoxic effect decreased, indicating that these two drugs have a synergistic effect. In terms of mechanism, salinomycin reversed doxorubicin-induced EMT of cholangiocarcinoma cells, as shown morphologically and through the detection of EMT markers. Moreover, we showed that salinomycin treatment downregulated the AMP-activated protein kinase family member 5 (ARK5) expression, which regulates the EMT process of cholangiocarcinoma. Our results indicated that salinomycin reversed the EMT process in cholangiocarcinoma cells by inhibiting ARK5 expression and enhanced the chemosensitivity of cholangiocarcinoma cells to doxorubicin. Therefore, a combined treatment of salinomycin with doxorubicin could be used to enhance doxorubicin sensitivity in patients with cholangiocarcinoma.

Sources of varieties and quality of circular Fructus Ligustri Lucidi / 中国天然药物

This study aimed to trace sources and quantitatively analyze the specnuezhenide content of circular Fructus Ligustri Lucidi for clinical use. Different specifications of Fructus Ligustri Lucidi were identified using DNA barcoding technology and the specnuezhenide content was analyzed by High Performance Liquid Chromatography (HPLC). The ITS sequence of circular Fructus Ligustri Lucidi was identical to that of standard privet, which was determined through botanical identification. ITS sequence similarity between circular Fructus Ligustri Lucidi and Fructus Ligustri Lucidi which was registered in NCBI ranged from 99.5% to 100%. The sequences of circular and other Fructus Ligustri Lucidi were clustered in a Neighbor-Joining tree with bootstrap value of 95, and these sequences could be distinguished from adulterants. Conforming to pharmacopoeia standard, the average specnuezhenide content of circular Fructus Ligustri Lucidi was higher than that of chicken waist Fructus Ligustri Lucidi. Circular Fructus Ligustri Lucidi derived from Ligustrum lucidum Ait. and the specnuezhenide content was higher in circular Fructus Ligustri Lucidi than that in chicken waist Fructus Ligustri Lucidi.

Simultaneous determination of alizarin and rubimaillin in Rubia cordifolia by ultrasound-assisted ionic liquid-reversed phase liquid chromatography / 中国中药杂志

Four kinds of ionic liquids were adopted to analyze the content of rubimaillin and alizarin in Rubia cordifolia roots with ultrasonic-assisted extraction coupled with HPLC. The chromatographic column, Purospher star RP-C18 (4.6 mm x 250 mm, 5 microm), was used. Methanol and 0.4% acetic acid-water as mobile phase with flow rate at 0.85 mL min(-1), gradient elution, detection wavelength at 250 nm, chromatographic column temperature was controlled at room temperature. The result showed that rubimaillin and alizarin had the highest extraction yield when the [ HMIM] PF6methanol solution concentration of 0.6 mol x L(-1) as extraction solvent and the conditions were solid-liquid ratio of 1:80 (g x mL(-1)). Under the optimal extraction conditions, the content of alizarin from 0.01 to 0.04 microg showed a good linearity (r = 0.9999), the average recovery was 97.12%, the content of rubimaillin from 0.41 to 1.35 microg showed a good linearity (r = 0.9999), the average recovery was 98.10%. This experiment adopted environmentally friendly reagent as extraction solvent, the extraction efficiency was improved, and the environmental pollution caused by organic solvent was avoided, the harm of human body aslo was reduced. This method was simple and reliable, its repeatability was also very good, which had an important significance in the study of traditional Chinese medicine active ingredient extraction methods.

Salinomycin Enhances the Apoptosis of T-cell Acute Lymphoblastic Leukemia Cell Line Jurkat Cells Induced by Vincristine / 中国实验血液学杂志

<p><b>OBJECTIVE</b>This study was aimed to investigate the effect of salinomycin combined with vincristine on the proliferation and apoptosis of Jurkat cells and its possible mechanisms.</p><p><b>METHODS</b>The proliferation of Jurkat cells was examined by CKK-8 assay. Flow cytometry was used to assess cellular apoptosis. Levels of BCL-2, caspase-3, and caspase- 8 were measured by Western blot.</p><p><b>RESULTS</b>The salinomycin or vincristine, either alone or in combination, inhibited the proliferation of Jurkat cells in a dose-dependent manner. Salinomycin combined with vincristine produced more obveous inhibition of cell proliferation than either compound used alone (P<0.05). Western blot analysis showed that the combined use of Sal and VCR reduced the expression of BCL-2 protein, and increased expression of caspase 3 and caspase 8 protein, more significantly. Furthermore, combination of Sal and VCR synergistally promoted apoptosis of the Jurkat cells (P<0.05).</p><p><b>CONCLUSION</b>The combination of salinomycin and vincristine synergistically inhibits proliferation and promotes apoptosis of T-cell acute lymphoblastic leukemia Jurkat cells.</p>

Evaluation of effect of oleuropein on skin wound healing in aged male balb/c mice

Olive oil and olive leaf extract are used for treatment of skin diseases and wounds in Iran. The main component of olive leaf extract is Oleuropein. This research is focused on the effects of Oleuropein on skin wound healing in aged male Balb/c mice. In this experimental study, Oleuropein was provided by Razi Herbal Medicine Institute, Lorestan, Iran. Twenty four male Balb/c mice, 16 months of age, were divided equally into control and experimental groups. Under ether anesthesia, the hairs on the back of neck of all groups were shaved and a 1 cm long full-thickness incision was made. The incision was then left un-sutured. The experimental group received intradermal injections with a daily single dose of 50 mg/kg Oleuropein for a total period of 7 days. The control group received only distilled water. On days 3 and 7 after making the incision and injections, mice were sacrificed, and the skin around incision area was dissected and stained by hematoxylin and eosin [H and E] and Van Gieson's methods for tissue analysis. In addition, western blot analysis was carried out to evaluate the level of vascular endothelial growth factor [VEGF] protein expression. The statistical analysis was performed using SPSS [SPSS Inc., Chicago, USA]. The t test was applied to assess the significance of changes between control and experimental groups. Oleuropein not only reduced cell infiltration in the wound site on days 3 and 7 post incision, but also a significant increase in collagen fiber deposition and more advanced re- epithelialization were observed [p<0.05] in the experimental group as compared to the control group. The difference of hair follicles was not significant between the two groups at the same period of time. Furthermore, western blot analysis showed an increased in VEGF protein level from samples collected on days 3 and 7 post-incision of experimental group as compared to the control group [p<0.05]. These results suggest that Oleuropein accelerates skin wound healing in aged male Balb/c mice. These findings can be useful for clinical application of Oleuropein in expediting wound healing after surgery

Comparation of gastrointestinal absorption studies of specnuezhenide with salidroside in rats / 中国中药杂志

The experiment was designed to study the mechanism of increasing efficiency of Ligustrum lucidum steamed with wine. Rats in vivo with gastrointestinal perfusion model were used. The contents of salidroside and specnuezhenide in the fluid of gastrointestinal perfusion of rats were measured by HPLC at different time points after dosing. Then the K(a) and absorption percentage were calculated. Specnuezhenide could be detected in the fluid of gastrointestinal perfusion of specnuezhenide. The K(a) of the specnuezhenide and salidroside in the fetal intestines are 0.055 3 and 0.144 2 h(-1) respectively and the total absorptivity are 24.46% and 60.14% respectively after 4 hours. The K(a) in the stomach are 5.70 and 8.26 h(-1) respectively and the total absorptivity are 34.21% and 47.23% respectively after 4 hours. The experiment proved that specnuezhenide can be metabolized into salidroside which is more beneficial for gastrointestinal absorption. The experiment proved that specnuezhenide can be metabolized into salidroside both in the rat's stomach and the fetal intestine and compared with the specnuezhenide salidroside is more conducive to gastrointestinal absorption. The results suggested that the increasing efficiency on liver and kidney of L. lucidum steamed with wine has business with the fact that Specnuezhe nide is more conducive to the body after it is changed into salidroside.

Studies on effects of Achyranthes bidentata on tongsaimai pellets main active ingredients chlorogenic acid, isoliquiritin, harpagoside and glycyrrhizin in vivo pharmacokinetics / 中国中药杂志

To study on the effects of Achyranthes bidentata on Tongsaimai pellets main active ingredients chlorogenic acid, isoliquiritin, harpagoside and glycyrrhizin in rats in vivo pharmacokinetic behaviors, a method for the simultaneous determination of chlorogenic acid, isoliquiritin, harpagoside and liquiritigenin in rat plasma was established by UPLC-MS/MS. The analysis was performed on a waters Acquity BEH C18 column (2.1 mm x 100 mm, 1.7 microm) with the mixture of acetonitrile and 0.1% formic acid/water as mobile phase, and the gradient elution at a flow rate of 0.3 mL x min(-1). The analytes were detected by tandem mass spectrometry with the electrospray ionization (ESI) source and in the multiple reaction monitoring (MRM) mode. It turned out that the analytes of Tongsaimai pellets groups C(max) and AUC(Q-infinity) values were higher than that with A. bidentata group, and the C(max) values of chlorogenic acid had significantly difference (P < 0.05), the AUC(0-infinity) values of chlorogenic acid and glycyrrhizin had significantly difference (P < 0.05); The T(max) and CL values of two groups had no significantly difference. Results showed that the established method was specific, rapid, accurate and sensitive for the studies of Tongsaimai pellets four main active ingredients in rat in vivo pharmacokinetic, and A. bidentata have varying degrees of effects on Tongsaimai pellets four main active ingredients in rat in vivo pharmacokinetic behaviors.

Simultaneous determination of seven constituents in Lamiophlomis rotata by HPLC / 中国中药杂志

<p><b>OBJECTIVE</b>To establish a new method for simultaneous determination of shanzhiside methyl ester, chlorogenic acid, 8-O-acetyl shanzhiside methylester, forsythiaside B, rutin, acteoside and galuteolin in Lamiophlomis rotata.</p><p><b>METHOD</b>Separation was performed on a Welchrom-C18 chromatographic column with acetonitrile-0.1% orthophosphoric acid as mobile phasewith gradient elution. The flow rate was 1.0 mL x min(-1). The column temperature was 30 degrees C, and the detection wavelength was set at 238 nm, 330 nm and 350 nm.</p><p><b>RESULT</b>The seven compounds were well separated with good linear correlations. The mean recoveries of seven compounds were 96.47%-102.2% (RSD 0.70%-2.2%).</p><p><b>CONCLUSION</b>There were good correlations among the seven compounds in the samples of aerial parts. The mean sum of shanzhiside methyl ester and 8-O-acetyl shanzhiside methylester in samples of aerial parts is 1.44%. The aerial parts have more kinds of composition and with higher content than that of underground parts in L. rotata, which was reasonable for the resonable use of the aerial part as medicinal part. The method was simple, repeatable and stable, which could be used for identification and quality evaluation of L. rotata.</p>

[Iridoid glycosides from Eremostachys azerbaijanica rech. f. root]

Eremostachys azerbaijanica [family Lamiaceae] is one of the 16 endemic Iranian herbs of the genus Eremostachys. In Iran, the root of E. azerbaijanica is traditionally used as local analgesic and anti-inflammatory. In this research, roots of E. azerbaijanica were phytochemically studied until perhaps by identification of chemical content of this plant, a step to be taken toward correct use from this natural product in treatment of diseases. Methanolic extract from the root of E. azerbaijanica was prepared by Soxhlet method and its three compounds were isolated by SPE method and reversed-phase preparative HPLC. Finally, the compounds have been elucidated by UV and 1D NMR spectroscopic analysis. Three iridoid glycosides, Lamalbide [Lamiridoside], Pulchelloside I and Sesamoside were isolated from the root of E. azerbaijanica. The comparison of the results obtained from the present study and former published results, shows that three iridoid glucosides which identified in this research, have been previously elucidated in some species of Eremostachys

The agonist binding mechanism of human CB2 receptor studied by molecular dynamics simulation, free energy calculation and 3D-QSAR studies / 药学学报

CB2-selective agonists have drawn attention in drug discovery, since CB2 becomes a promising target for the treatment of neuropathic pain without psychoactive or other CNS-related side effects. However, the lack of experimental data of the 3D structures of human cannabinoid receptors hampers the understanding of the binding modes between ligands and CB2 by traditional methods. In the present work, combinational molecular modeling studies including flexible docking, MD simulations and free energy calculations were performed to investigate the interaction modes and mechanism of CB2-unselective agonist CP55940 and CB2-selective agonist GW842166X, separately binding with the homology model of CB2 in a DPPC/TIP3P simulated membrane environment. The binding free energies calculated by MM-PBSA method give an explanation for the activity differences of the studied ligands. Binding free energies decomposition by MM-GBSA method shows that the van der Waals interaction is the dominant driving force during the binding process. Our MD simulations demonstrate that Phe197 could be a critical residue for the binding of CB2-selective agonists. Furthermore, by using the MD simulated binding conformer as a template, the 3D-QSAR studies were performed with the comparative molecular field analysis (CoMFA) approach on a set of GW842166X analogues. A combinational exploration of both CoMFA steric and potential contour maps for CB2 affinities and the MD studied interaction modes sheds light on the structural requirements for CB2 agonists and serves as a basis for the design of novel CB2 agonists.

Absorption and pharmacokinetics of radix rehmanniae in rats / 药学学报

In this paper, absorption and pharmacokinetic study of Radix Rehmanniae was studied by liquid chromatography coupled with mass spectrometry method after oral administration to rats. By comparing the chromatograms of ultraviolet, full scan, extracted ion and selective reaction monitoring (SRM) of standard solution, Radix Rehmanniae, blank plasma and rat plasma post drug administration, catalpol and ajugol were found to be the main compounds absorbed from Radix Rehmanniae. Plasma concentrations of aucubin, dihydrocatalpol, rehmannioside A (or rehmannioside B/ melittoside) and rehmannioside D were very low. Quantitative method for catalpol and aucubin and semi-quantitative method for other compounds in rat plasma were established. The pharmacokinetic study of those absorbed components was conducted after oral administration of 6 g x kg(-1) Radix Rehmanniae water extract to rats. Cmax, t(1/2) and AUC(0-infinity) of catalpol and ajugol were (2349.05 +/- 1438.34) and (104.25 +/- 82.05) ng x mL(-1), (0.86 +/- 0.32) and (0.96 +/- 0.37) h, (4407.58 +/- 2734.89) and (226.66 +/- 188.38) ng x h x mL(-1), respectively. tmax was at 1.00 h for catalpol and ajugol. Both catalpol and ajugol were absorbed and excreted rapidly.

Pharmacokinetics of 8-O-acetylharpagide and harpagide after oral administration of Ajuga decumbens extract in beagle dog / 中国中药杂志

8-O-acetylharpagide and harpagide are two kinds of effective component of Ajuga decumbens extract. A sensitive LC-MS/MS method has been established for pharmacokinetics of 8-O-acetylharpagide and harpagide in beagle dog after oral administration of from A. decumbens extract. Female beagle dogs received orally 12.9, 25.7 mg x kg(-1) p. o. Concentrations of 8-O-acetylharpagide and harpagide in plasma were determined by LC-MS/MS method at different time points and all pharmacokinetic parameters were estimated by non-compartment analysis. The mobile phase consisted of 0.1% formic acid in water (A) and acetonitrile (B), which was run at a flow rate of 0.3 mL x min(-1). Chromatographic separation was achieved on an Agilent ZORBAX XDB-C18 column (2.1 mm x 50 mm, 3.5 microm) using a gradient elution of 5% B at 0-2 min, 95% B at 2. 1-5 min and 5% B at 5. 1-10 min. All analytes, including the IS, were monitored under positive ionization conditions and quantified in MRM mode with transitions of m/z 429.2-369.2 for 8-O-acetylharpagide, m/z 387.2-207.2 for harpagide, and m/z 149.2-103.1 for IS. High purity nitrogen was employed as both the nebulizing and drying gas. Other parameters of the mass spectrometer were optimized as follows: drying gas flow 10.0 L x min(-1); drying gas temperature 300 degrees C; capillary voltage 4 000 V. Results showed that 8-O-acetylharpagide and harpagide showed a dose-dependence profile. T(max) of 8-O-acetylharpagide is 1.7 h, and T(max) of harpagide is 1.57 h, which was higher than T(max) of 8-O-acetylharpagide and harpagide after oral administration of from A. decumbens extract in rats. The different pharmacokinetic parameters may be due to the species differences of rat and beagle dog.

Effects of lovastatin, clomazone and methyl jasmonate treatment on the accumulation of purpurin and mollugin in cell suspension cultures of Rubia cordifolia / 中国天然药物

AIM@#To determine the IPP origin of the naphthoquinones (NQs) in Rubia cordifolia, and to evaluate the effects of methyl jasmonate (MeJA) treatment, MEP, and MVA pathway inhibitor treatment on the accumulation of anthraquinones (AQs) and NQs in cell suspension cultures of R. cordifolia.@*METHODS@#Cell suspension cultures of R. cordifolia were established. Specific inhibitors (lovastatin and clomazone) and MeJA were supplied to the media, respectively. Treated cells were sampled every three days. Content determination of purpurin (AQs) and mollugin (NQs) were carried out using RP-HPLC. The yield of the two compounds was compared with the DMSO-supplied group and the possible mechanism was discussed.@*RESULTS@#Lovastatin treatment increased the yield of purpurin and mollugin significantly. Clomazone treatment resulted in a remarkable decrease of both compounds. In the MeJA-treated cells, the purpurin yield increased, meanwhile, the mollugin yield decreased compared with control.@*CONCLUSION@#The IPP origin of mollugin in R. cordifolia cell suspension cultures was likely from the MEP pathway. To explain the different effects of MeJA on AQs and NQs accumulation, studies on the regulation and expression of the genes, especially after prenylation of 1,4-dihydroxy-2-naphthoic acid should be conducted.

Simultaneous determination of four compounds in Erzhiwan by HPLC / 中国中药杂志

<p><b>OBJECTIVE</b>To establish an HPLC method for simultaneous determination of nuzhenide, specnuezhenide, wedelolactone and oleanic acid in Erzhiwan.</p><p><b>METHOD</b>The DIKMA C18 (4.6 mm x 200 mm, 5 microm) column was adopted with acetonitrile and 0.1% phosphoric acid solution as the mobile phase and gradient elution. The flow rate was 1.0 mL x min(-1) and the volume of injection was 20 microL. The column temperature was maintained at 30 degrees C and the detective wavelength was set at 215 nm.</p><p><b>RESULT</b>There were good linear relationships between the peak area and concentration at the range of 2.008-80.32 (r = 0.999 6), 5.872-234.88 (r = 0.999 7) , 0.9-36 (r = 0.999 9), 13.24-529.6 mg x L(-1) (r = 0.999 6) for nuzhenide, specnuezhenide, wedelolactone and oleanic acid, respectively. The average recovery rates of nuzhenide, specnuezhenide, wedelolactone and oleanic acid were 99.25%, 98.70%, 96.23% and 101.5%, respectively, with RSD of less than 3%.</p><p><b>CONCLUSION</b>The established method was so easy, rapid and accurate that it can be used as an effective way for quality control of Erzhiwan.</p>

Study on intestinal absorption of mollugin and purpurin in rats / 中国中药杂志

<p><b>OBJECTIVE</b>To study the absorption kinetic characteristics of mollugin and purpurin in each intestinal segment of rats.</p><p><b>METHOD</b>The in situ single-way perfusion rat model was established to study absorption characteristics of mollugin and purpurin in each intestinal segment of rats. The volume of recirculation fluid was regulated by phenol red.</p><p><b>RESULT</b>Different quality concentrations (12.33, 24.66, 49.32 mg x L(-1)) of mollugin and (8.455, 16.91, 33.82 mg x L(-1)) purpurin showed a concentration gradient of absorption dose in each intestinal segment, with the osmotic coefficient increasing to more than 0.2 x 10(-4) cm x s(-1). In the same concentration, mollugin and purpurin showed an identical trend of P(eff) in each intestinal segment in the order of colon > duodenum > ileum > jejunum, with a significant difference (P < 0.05).</p><p><b>CONCLUSION</b>Mollugin and purpurin are highly permeable in rat intestinal segments, with absorption in each segment, while the specific absorption existed in the colon segment.</p>

Boiling extraction of oleuropein at low temperature and reduced pressure / 中国中药杂志

<p><b>OBJECTIVE</b>To look for optimum extraction techniques for oleuropein by boiling olive leaves at low temperature and reduced pressure.</p><p><b>METHOD</b>According to single factor experiment (SFE) design, the effects of seven factors, the impact of seven factors, type of solvent, temperature, time, ratio of material to liquid, ethanol concentration, vacuum degree and extraction times, on extraction yield of oleuropein were investigated. Based on the results of SFE, four more important factors, temperature, time, ratio of material to liquid and ethanol concentration, were selected in L9 (3(4)) orthogonal experiment (OE) to compare with those extracted with traditional methods.</p><p><b>RESULT</b>The optimum conditions for boiling extraction of oleuropein at low temperature and reduced pressure were as follows: temperature 60 degrees C, time 20 min, ratio of material to liquid 1:30 and ethanol concentration 85%. The conditions presented an extraction yield of 5.90%.</p><p><b>CONCLUSION</b>Compared with traditional extraction methods and the ultrasound assisted extraction method, boiling extraction techniques at low temperature and reduced pressure were so quick and efficient that it has a good application prospect.</p>

p53 and DNA-dependent protein kinase catalytic subunit independently function in regulating actin damage-induced tetraploid G1 arrest

We previously reported that the p53 tumor suppressor protein plays an essential role in the induction of tetraploid G1 arrest in response to perturbation of the actin cytoskeleton, termed actin damage. In this study, we investigated the role of p53, ataxia telangiectasia mutated protein (ATM), and catalytic subunit of DNA-dependent protein kinase (DNA-PKcs) in tetraploid G1 arrest induced by actin damage. Treatment with actin-damaging agents including pectenotoxin-2 (PTX-2) increases phosphorylation of Ser-15 and Ser-37 residues of p53, but not Ser-20 residue. Knockdown of ATM and DNA-PKcs do not affect p53 phosphorylation induced by actin damage. However, while ATM knockdown does not affect tetraploid G1 arrest, knockdown of DNA-PKcs not only perturbs tetraploid G1 arrest, but also results in formation of polyploidy and induction of apoptosis. These results indicate that DNA-PKcs is essential for the maintenance of actin damage induced-tetraploid G1 arrest in a p53-independent manner. Furthermore, actin damage-induced p53 expression is not observed in cells synchronized at G1/S of the cell cycle, implying that p53 induction is due to actin damage-induced tetraploidy rather than perturbation of actin cytoskeleton. Therefore, these results suggest that p53 and DNA-PKcs independently function for tetraploid G1 arrest and preventing polyploidy formation.