Zi Wang

Application of response surface methodology to optimise ultrasonic-assisted extraction of four chromones in Radix Saposhnikoviae.

INTRODUCTION Radix Saposhnikoviae is one of the most famous Chinese herbal medicines with many pharmacological activities towards inflammatory symptoms and antioxidation. Chromones are considered as one of the effective components. It is important to find a reasonable method to extract the chromones in S. divaricata. OBJECTIVE To develop an ultrasonic-assisted extraction (UAE) to extract chromones in Radix Saposhnikoviae and to optimise extraction conditions. METHODOLOGY Four chromones (prim-O-glucosylcimifugin, cimifugin, 5-O-methylvisammioside and sec-O-glucosylhamaudol) were extracted by the UAE method combined with response surface methodology (RSM). Box-Behnken design (BBD) was applied to evaluate the effects of three independent variables (ethanol concentration, extraction time and extraction temperature) on the chromones yield of Radix Saposhnikoviae. RESULTS Correlation analysis of the mathematical-regression model indicated that a quadratic polynomial model could be employed to optimise the extraction of chromones by UAE method. The optimal conditions to obtain the highest chromones yield of Radix Saposhnikoviae were a solvent of 75% ethanol, an extraction time of 48 min and an extraction temperature of 67°C. CONCLUSION Under these optimal conditions, the experimental values agreed closely with the predicted values. The analysis of variance indicated a high goodness of model fit and the success of RSM method for optimising chromones extraction in Radix Saposhnikoviae.

Ginsenoside Rg5 ameliorates cisplatin-induced nephrotoxicity in mice through inhibition of inflammation, oxidative stress, and apoptosis

Although cisplatin is an effective anti-cancer agent that is widely used for treating various types of malignant solid tumors, the nephrotoxicity induced by cisplatin severely limits its clinical application. The present study was designed to explore the potential protective effect of ginsenoside Rg5, a rare ginsenoside generated during steaming ginseng, on cisplatin-induced nephrotoxicity in a mouse experimental model. The possible mechanisms underlying this nephroprotective effect were also investigated for the first time. Rg5 was given at doses of 10 and 20 mg/kg for 10 consecutive days. On Day 7, a single nephrotoxic dose of cisplatin (25 mg/kg) was injected to mice. Cisplatin administration resulted in renal dysfunction as evidenced by increase in serum creatinine (CRE) and blood urea nitrogen (BUN) levels. In addition, cisplatin increased the level of malondialdehyde (MDA) and 4-hydroxynonenal (4-HNE), the makers of lipid peroxidation, and depleted glutathione (GSH) content and superoxide dismutase (SOD) activity in renal tissues. These effects were associated with the significantly increased levels of cytochrome P450 E1 (CYP2E1), 4-hydroxynonenal (4-HNE), tumor necrosis factor (TNF)-α, interleukin (IL)-1β, nuclear factor-kappa B (NF-κB) p65, and cyclooxygenase-2 (COX-2) in renal tissues. However, pretreatment with ginsenoside Rg5 significantly attenuated the renal dysfunction, oxidative stress and inflammation response induced by cisplatin. Furthermore, ginsenoside Rg5 supplementation inhibited activation of apoptotic pathways through increasing Bcl-2 and decreasing Bax expression levels. Histopathological examination further confirmed the nephroprotective effect of Rg5. Collectively, these results clearly suggest that Rg5-mediated alleviation of cisplatin-induced nephrotoxicity may be related to its anti-oxidant, anti-apoptotic and anti-inflammatory effects.

Determination of Kanamycin A in Animal Feeds by Solid Phase Extraction and High Performance Liquid Chromatography with Pre‐Column Derivatization and Fluorescence Detection

Abstract A selective method was developed for the determination of kanamycin A in medicated animal feeds by reversed‐phase high performance liquid chromatography (HPLC) with precolumn derivatization. Samples were extracted with 0.1 mol/L hydrochloric acid solution and cleanup was achieved with MCX solid phase extraction (SPE). The purified extract was derivatized with O‐phthaladehyde (OPA), separated on a reversed‐phase C18 column and the fluorescence detection was performed at the excitation and emission wavelengths of 230 and 389 nm, respectively. The run time was approximately 14 min, at the flow rate of 0.8 mL/min. This method provides the average recoveries 98.4%–106.0% in feeds spiked in the range of 10–2000 g/ton, with the coefficients of variation 1.17%–9.78%. The limit of detection in feeds was 5 g/ton and the limit of quantification in feeds was 10 g/ton, which are well below the effective dose.

Pressurized liquid extraction followed by LC‐ESI/MS for analysis of four chromones in Radix Saposhnikoviae

An efficient pressurized liquid extraction (PLE) technique was employed in extracting chromones from the roots of Saposhnikovia divaricata (Radix Saposhnikoviae). Chromones were quantified and analyzed by LC-ESI/MS. The PLE procedure was optimized, validated and compared with the other conventional extraction techniques. PLE gained the best result due to the highest extraction efficiency within the shortest extraction time. The optimal conditions of PLE were employing 50% ethanol as the extraction solvent at a temperature of 140°C and an extraction pressure of 1500 psi, using one extraction cycle with a static extraction time of 8 min. A good LC separation was achieved using a Hypersil ODS2 column and methanol/water as the mobile phase at a flow rate of 0.8 mL/min. MS coupling with an ESI interface in the positive ion mode was used as the detection technique. This is the first report on combining PLE with LC-ESI/MS for the extraction and quantification of chromones in Radix Saposhnikoviae. The elaborated PLE method also provided a good alternative for the chromone extraction from other plant substances.

Application of accelerated solvent extraction to the investigation of saikosaponins from the roots of Bupleurum falcatum.

Accelerated solvent extraction (ASE) was applied to the extraction of saikosaponin a, saikosaponin c and saikosaponin d from the roots of Bupleurum falcatum. Main extraction parameters such as the extraction solvents, extraction temperature and static extraction time were investigated and optimized. The optimized procedure employed 70% methanol as extraction solvent, 120 degrees C of extraction temperature, 10 min of static extraction time, 60% of flush volume and the extraction recoveries of the three compounds were near to 100% with one extraction cycle. The extracted samples were analyzed by HPLC with UV detector. The HPLC conditions were as follows: Hypersil ODS2 (4.6 mmx250 mm, 5 microm) column, acetonitrile and water as mobile phase, flow rate of 1.0 mL/min, UV detection wavelength of 204 nm and injection volume of 20 microL. Compared with the traditional methods including heat-reflux extraction and ultrasonic-assisted extraction, the proposed ASE method was more efficient and faster to be operated. The results indicated that ASE was an alternative method for extracting saikosaponins from the roots of B. falcatum.

Ameliorative Effects and Possible Molecular Mechanism of Action of Black Ginseng (Panax ginseng) on Acetaminophen-Mediated Liver Injury

Background: Frequent overdosing of acetaminophen (APAP) has become the major cause of acute liver injury (ALI). The present study aimed to evaluate the potential hepatoprotective effects of black ginseng (BG) on APAP-induced mice liver injuries and the underlying mechanisms of action were further investigated for the first time. Methods: Mice were treated with BG (300, 600 mg/kg) by oral gavage once a day for seven days. On the 7th day, all mice were treated with 250 mg/kg APAP which caused severe liver injury after 24 h and hepatotoxicity was assessed. Results: Our results showed that pretreatment with BG significantly decreased the levels of serum alanine aminotransferase (ALT) and aspartate transaminase (AST) compared with the APAP group. Meanwhile, hepatic antioxidant including glutathione (GSH) was elevated compared with the APAP group. In contrast, a significant decrease of the levels of the lipid peroxidation product malondialdehyde (MDA) was observed in the BG-treated groups compared with the APAP group. These effects were associated with significant increases of cytochrome P450 E1 (CYP2E1) and 4-hydroxynonenal (4-HNE) levels in liver tissues. Moreover, BG supplementation suppressed activation of apoptotic pathways through increasing Bcl-2 and decreasing Bax protein expression levels according to western blotting analysis. Histopathological examination revealed that BG pretreatment significantly inhibited APAP-induced necrosis and inflammatory infiltration in liver tissues. Biological indicators of nitrative stress like 3-nitrotyrosine (3-NT) were also inhibited after pretreatment with BG, compared with the APAP group. Conclusions: The results clearly suggest that the underlying molecular mechanisms of action of BG-mediated alleviation of APAP-induced hepatotoxicity may involve its anti-oxidant, anti-apoptotic, anti-inflammatory and anti-nitrative effects.

Anti-Tumor Effect of Steamed Codonopsis lanceolata in H22 Tumor-Bearing Mice and Its Possible Mechanism

Although previous studies confirmed that steaming and the fermentation process could significantly improve the cognitive-enhancement and neuroprotective effects of Codonopsis lanceolata, the anti-tumor efficacy of steamed C. lanceolata (SCL) and what mechanisms are involved remain largely unknown. The present study was designed to evaluate the anti-tumor effect in vivo of SCL in H22 tumor-bearing mice. The results clearly indicated that SCL could not only inhibit the tumor growth, but also prolong the survival time of H22 tumor-bearing mice. Besides, the serum levels of cytokines, such as interferon gamma (IFN-γ), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and interleukin-2 (IL-2), were enhanced by SCL administration. The observations of Hoechst 33258 staining demonstrated that SCL was able to induce tumor cell apoptosis. Finally, immunohistochemical analysis revealed that SCL treatment significantly increased Bax expression and decreased Bcl-2 and vascular endothelial growth factor (VEGF) expression of H22 tumor tissues in a dose-dependent manner. Moreover, LC/MS analysis of SCL indicated that it mainly contained lobetyolin and six saponins. Taken all together, the findings in the present study clearly demonstrated that SCL inhibited the H22 tumor growth in vivo at least partly via improving the immune functions, inducing apoptosis and inhibiting angiogenesis.

Platycoside N: A New Oleanane-Type Triterpenoid Saponin from the Roots of Platycodon grandiflorum

A new oleanane-type triterpenoid saponin, named platycoside N (1), together with six known saponins, was isolated from the roots of Platycodon grandiflorum. On the basis of acid hydrolysis, comprehensive spectroscopic data analyses and comparison with the spectral data of the known compounds, its structure was elucidated as 3-O-β-D-glucopyranosyl-(1→6)-β-D-glucopyranosyl-2β,3β,16α,23-tetrahydroxyolean-12-en-28-oic acid 28-O-β-L-rhamnopyranosyl-(1→2)-α-L-arabinopyranoside. The six known compounds were platycodin D (2), deapioplatycodin D (3), platycodin D3 (4), deapio- platycodin D3 (5), platycoside E (6) and deapioplatycoside E (7).

ISOLATION AND PURIFICATION OF SAPONINS FROM PLATYCODON GRANDIFLORUM BY SEMI-PREPARATIVE HIGH PERFORMANCE LIQUID CHROMATOGRAPHY AND LC/ESI-MS

In this work, the isolation and purification of saponins (platycosides) from the roots of Platycodon grandiflorum, including deapio-platycodin D, platycodin D, and polygalacin D, was investigated using semi-preparative high performance liquid chromatography (SP-HPLC). The chromatographic separation was carried out on a Zorbax Eclipse XDB C18 column (250 mm × 9 mm; 10 µm). The conversion of separation condition from analytical level to preparative level was successfully achieved by optimizing mobile phase, flow rate, and sample load. A satisfactory separation of platycosides was obtained with 3.5 mL/min of flow rate in less than 40 min. Chromatography was performed using binary mobile phase composed of methanol-water with gradient elution and a sample load of 100 mg. UV detection was set at 210 nm. With the optimal chromatographic conditions, the purity of each platycoside was more than 98.5%. Furthermore, the molecular weights of platycosides were identified by HPLC-ESI-MS analysis in both positive and negative modes, and then their structures were further elucidated by IR, UV, and 13C-NMR spectroscopy data. The present method is robust and suitable for preparing available quantities of pure platycosides from P. grandiflorum in conventional laboratories.

Maltol, a Maillard reaction product, exerts anti-tumor efficacy in H22 tumor-bearing mice via improving immune function and inducing apoptosis

The purpose of this study was to investigate the anti-hepatoma activity of maltol, a Maillard reaction product, in H22 tumor-bearing mice. The results demonstrate that maltol not only significantly inhibited the growth of hepatoma H22 transplanted in mice, but also prolonged the survival time of H22-bearing mice. Furthermore, the levels of serum cytokines in H22 tumor-bearing mice, such as interferon gamma (IFN-γ), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and interleukin-2 (IL-2), were enhanced by maltol treatment. Importantly, immunohistochemical and western blotting analysis clearly show that maltol treatment increased Bax and decreased Bcl-2 protein expression levels of H22 tumor tissues in a dose-dependent manner. Collectively, our findings in the present study clearly demonstrate that the maltol markedly suppressed the tumor growth of H22 transplanted tumors in vivo at least partly via improving the immune functions, inducing apoptosis, and inhibiting angiogenesis.