Jia Xb

Metabolite profiles of icariin in rat plasma by ultra-fast liquid chromatography coupled to triple-quadrupole/time-of-flight mass spectrometry

In this work, the metabolite profiles of icariin in rat plasma were qualitatively investigated, and the possible metabolic pathways of icariin were subsequently proposed. After oral administration of icariin, rat plasma samples were collected and pretreated by protein precipitation. Then these pretreated samples were injected into a Venusil ASB-C18 column with mobile phase consisted of 0.1% formic acid water and 0.1% formic acid acetonitrile and detected by Ultra fast liquid chromatography coupled to time-of-flight mass spectrometry (UFLC-TOF/MS). A total of 19 metabolites, namely, icariside I (M1), icaritin (M2), desmethylicaritin and its isomer (M3-M6), icaritin-3-O-gluA (M7), icaritin-7-O-gluA (M8), icariside II and its isomer (M9 and M10), icaritin-3,7-di-O-gluA (M11), 1,3-isoprene alcohol icaritin and its isomer (M12, M13 and M18), 1,3-isoprene alcohol icariside II (M14), allylic alcohol icaritin and its isomer (M15 and M16), 1,3-isoprene icariside II (M17) and icaritin-3-O-rha-7-O-gluA (M19) were detected and tentatively identified, and 9 of them, including M7 and M8 and M12-M18 were reported for the first time. The metabolites profiles in plasma revealed that glucuronide conjugates of isoflavonoids and flavonoid aglycones were the major circulating forms of icariin.

A novel drug–phospholipid complex enriched with micelles: preparation and evaluation in vitro and in vivo

Mixed micelles are widely used to increase solubility and bioavailability of poorly soluble drugs. One promising antitumor drug candidate is 20(S)-protopanaxadiol (PPD), although its clinical application is limited by low water solubility and poor bioavailability after oral administration. In this study, we developed mixed micelles consisting of PPD–phospholipid complexes and Labrasol® and evaluated their potential for oral PPD absorption. Micelles were prepared using a solvent-evaporation method, and their physicochemical properties, including particle size, zeta potential, morphology, crystal type, drug loading, drug entrapment efficiency, and solubility, were characterized. Furthermore, in vitro release was investigated using the dialysis method, and transport and bioavailability of the mixed micelles were investigated through a Caco-2 cell monolayer and in vivo absorption studies performed in rats. Compared with the solubility of free PPD (3 μg/mL), the solubility of PPD in the prepared mixed micelles was 192.41 ± 1.13 μg/mL in water at room temperature. The in vitro release profiles showed a significant difference between the more rapid release of free PPD and the slower and more sustained release of the mixed micelles. At the end of a 4-hour transport study using Caco-2 cells, the apical-to-basolateral apparent permeability coefficients (Papp) increased from (1.12 ± 0.21) × 106 cm/s to (1.78 ± 0.16) × 106 cm/s, while the basolateral-to-apical Papp decreased from (2.42 ± 0.16) × 106 cm/s to (2.12 ± 0.32) × 106. In this pharmacokinetic study, compared with the bioavailability of free PPD (area under the curve [AUC]0–∞), the bioavailability of PPD from the micelles (AUC0–∞) increased by approximately 216.36%. These results suggest that novel mixed micelles can significantly increase solubility, enhance absorption, and improve bioavailability. Thus, these prepared micelles might be potential carriers for oral PPD delivery in antitumor therapies.

Ursolic Acid Inhibits Cigarette Smoke Extract-Induced Human Bronchial Epithelial Cell Injury and Prevents Development of Lung Cancer

Cigarette smoking is the main cause of chronic obstructive pulmonary disease and lung cancer. The present study was aimed to explore the chemopreventive effect of ursolic acid (UA) on these diseases. In the CSE treated normal human bronchial epithelial cell model, UA alleviated cytotoxicity caused by CSE, recovered the intracellular redox balance, and relieved the stimulation of external deleterious factors as well. UA mitigated CSE-induced DNA damage through the Nrf2 (nuclear factor erythroid 2-related factor 2) pathway. Moreover, UA inhibited lung cancer development in the model established by A549 cells in nude mice in vivo. For the first time, our results indicate that UA could be developed as a potential lung cancer chemopreventive agent.

Reactive oxygen species-mediated mitochondrial pathway is involved in Baohuoside I-induced apoptosis in human non-small cell lung cancer.

Baohuoside I (also known as Icariside II) is a flavonoid isolated from Epimedium koreanum Nakai. Although Baohuoside I exhibits anti-inflammatory and anti-cancer activities, its molecular targets/pathways in human lung cancer cells are poorly understood. Therefore, in the present study, we investigated the usefulness of Baohuoside I as a potential apoptosis-inducing cytotoxic agent using human adenocarcinoma alveolar basal epithelial A549 cells as in vitro model. The apoptosis induced by Baohuoside I in A549 cells was confirmed by annexin V/propidium iodide double staining, cell cycle analysis and dUTP nick end labeling. Further research revealed that Baohuoside I accelerated apoptosis through the mitochondrial apoptotic pathway, involving the increment of BAX/Bcl-2 ratio, dissipation of mitochondrial membrane potential, transposition of cytochrome c, caspase 3 and caspase 9 activation, degradation of poly (ADP-ribose) polymerase and the over-production of reactive oxygen species (ROS). A pan-caspase inhibitor, Z-VAD-FMK, only partially prevented apoptosis induced by Baohuoside I, while NAC, a scavenger of ROS, diminished its effect more potently. In addition, the apoptotic effect of Baohuoside I was dependent on the activation of ROS downstream effectors, JNK and p38(MAPK), which could be almost abrogated by using inhibitors SB203580 (an inhibitor of p38(MAPK)) and SP600125 (an inhibitor of JNK). These findings suggested that Baohuoside I might exert its cytotoxic effect via the ROS/MAPK pathway.

enhanced bioavailability of apigenin via preparation of a carbon nanopowder solid dispersion

In this study, a novel carbon nanopowder (CNP) drug carrier was developed to improve the oral bioavailability of apigenin (AP). Solid dispersions (SDs) of AP with CNP were prepared, and their in vitro drug release and in vivo performance were evaluated. The physicochemical properties of the formulations were examined by differential scanning calorimetry, X-ray diffraction, and scanning electron microscopy. Drug release profiles showed that AP dissolution from the CNP-AP system (weight ratio, 6:1) after 60 minutes improved by 275% compared with that of pure AP. Moreover, the pharmacokinetic analysis of SD formulations in rats showed that the AP area under the curve0–t value was 1.83 times higher for the CNP-AP system than for pure AP, indicating that its bioavailability was significantly improved. In addition, compared with pure AP, SDs had a significantly higher peak and shorter time to peak. Preliminary intestinal toxicity tests indicated that there was no significant difference in the tissues of the rats treated with the CNP-AP system, rats treated with the CNP alone, and controls. In conclusion, CNP-based SDs could be used for enhancing the bioavailability of poorly water-soluble drugs while also improving drug safety.

Enhanced oral absorption of 20(S)-protopanaxadiol by self-assembled liquid crystalline nanoparticles containing piperine: in vitro and in vivo studies

Background 20(S)-protopanaxadiol (PPD), similar to several other anticancer agents, has low oral absorption and is extensively metabolized. These factors limit the use of PPD for treatment of human diseases. Methods In this study, we used cubic nanoparticles containing piperine to improve the oral bioavailability of PPD and to enhance its absorption and inhibit its metabolism. Cubic nanoparticles loaded with PPD and piperine were prepared by fragmentation of glyceryl monoolein (GMO)/poloxamer 407 bulk cubic gel and verified using transmission electron microscopy and differential scanning calorimetry. We evaluated the in vitro release of PPD from these nanoparticles and its absorption across the Caco-2 cell monolayer model, and subsequently, we examined the bioavailability and metabolism of PPD and its nanoparticles in vivo. Results The in vitro release of PPD from these nanoparticles was less than 5% at 12 hours. PPD-cubosome and PPD-cubosome loaded with piperine (molar ratio PPD/piperine, 1:3) increased the apical to basolateral permeability values of PPD across the Caco-2 cell monolayer from 53% to 64%, respectively. In addition, the results of a pharmacokinetic study in rats showed that the relative bioavailabilities of PPD-cubosome [area under concentration–time curve (AUC)0–∞] and PPD-cubosome containing piperine (AUC0–∞) compared to that of raw PPD (AUC0–∞) were 166% and 248%, respectively. Conclusion The increased bioavailability of PPD-cubosome loaded with piperine is due to an increase in absorption and inhibition of metabolism of PPD by cubic nanoparticles containing piperine rather than because of improved release of PPD. The cubic nanoparticles containing piperine may be a promising oral carrier for anticancer drugs with poor oral absorption and that undergo extensive metabolism by cytochrome P450.

Metabolite Profiling of Four Major Flavonoids of Herba Epimdii in Zebrafish

The zebrafish model organism was applied first in a metabolic study of icariin, baohuoside I, epimedin A and epimedin C, which are flavonoids in Herba Epimedii. Metabolites of these compounds in zebrafish after exposure for 24 h were identified by HPLC-ESI-MS, whereby the separation was performed with a Zorbax C-18 column using a gradient elution of 0.05% formic acid acetonitrile-0.05% formic acid water. The quasi-molecular ions of compounds were detected in simultaneous negative and positive ionization modes. Metabolic products of icariin and epimedin C via cleavage of glucose residue instead of rhamnose residues were found, which coincided with the results using regular metabolic analysis methods. In addition, the zebrafish model was used to predict the metabolism of the trace component epimedin A, whose metabolic mechanisms haven’t been clearly elucidated with the current metabolism model. The metabolic pathway of epimedin A in zebrafish was similar to those of its homologue icariin and epimedin C. Our study demonstrated that the zebrafish model can successfully imitate the current models in elucidating metabolic pathways of model flavonoids, which has advantages of lower cost, far less amount of compound needed, easy set up and high performance. This novel model can also be applied in quickly predicting the metabolism of Chinese herb components, especially trace compounds.

Characterization of metabolites of 20(S)-protopanaxadiol in rats using ultra-performance liquid chromatography/quadrupole-time-of-flight mass spectrometry.

In this study, ultra-performance liquid chromatography (UPLC)/quadrupole-time-of-flight mass spectrometry (QTOF-MS) was applied to the rapid analysis of 20(S)-protopanaxadiol (PPD) metabolites in rats after oral administration, enabling the structural characterization of 23 metabolites in plasma, bile, urine, and feces. 16 of these, including M1-M5, M9, and M11-M15, have not been previously reported. The results also indicated that demethylation, dehydration, dehydrogenation, oxidation, deoxidation, and glucuronidation were the major metabolic reactions of PPD in vivo. This study provides important information about the metabolism of PPD which will be helpful for fully understanding its mechanism of action. Furthermore, structural modification of PPD in vivo may aid in obtaining new chemical derivatives for pharmacological screening.

Metabolism of tanshinone IIA, cryptotanshinone and tanshinone I from Radix Salvia miltiorrhiza in zebrafish.

The study aimed to investigate the potential of zebrafish in imitating mammal phase I metabolism of natural compounds. Three diterpenoid quinones from Radix Salvia miltiorrhiza, namely tanshinone IIA (TIIA), cryptotanshinone (Cry) and tanshinone I (TI) were selected as model compounds, and their metabolites mediated by zebrafish were characterized using a high-performance liquid chromatography coupled ion-trap mass spectrometry (HPLC/IT-MSn) method with electrospray ionization in positive mode. The separation was performed with a Zorbax C-18 column using a binary gradient elution of 0.05% formic acid acetonitrile/0.05% formic acid water. According to the MS spectra and after comparison with reference standards and literature reports, hydroxylation, dehydrogenation or D-ring hydrolysis metabolites of TIIA and Cry but not of TI were characterized, which coincided with those reported using regular in vivo or in vitro metabolic analysis methods, thus verifying that zebrafish can successfully imitate mammalian phase I metabolism which instills further confidence in using zebrafish as a novel and prospective metabolism model.

Anti-Hepatitis B Virus Activity of Chickweed (Stellaria media (L.) Vill.) Extracts in HepG2.2.15 Cells

Stellaria media (Linn.) Villars is a traditional Chinese medicine that has been used for over 200 years, mainly for the treatment of dermatitis and other skin diseases. It has also been used as an anti-viral agent. All the fresh chickweed juice samples used in this study were prepared using macroporous resin and ultrafiltration technology. The anti-hepatitis B virus (HBV) activity of S. media was evaluated in vitro using the human HBV-transfected liver cell line HepG2.2.15. The concentrations of hepatitis B surface antigen (HBsAg) and hepatitis B e antigen (HBeAg) in HepG2.2.15 cell culture medium were determined by enzyme-linked immunosorbent assay (ELISA) after S. media-n (SM-n) treatment for 6 or 9 days. HBV DNA was quantified using transcription-mediated amplification and real-time polymerase chain reaction. In HepG2.2.15 cells, 30 μg/mL SM-3 effectively suppressed the secretion of HBsAg and HBeAg with inhibition rates of 27.92% and 25.35% after 6 days of treatment, respectively. Consistent with the reduction in HBV antigens, SM-3 also reduced the level of HBV DNA in a dose-dependent manner. The characterization and quantitation of the chemical composition of SM-3 showed the presence of flavonoid C-glycosides, polysaccharides, and protein, which exhibited diverse antiviral activities. In conclusion, our results demonstrate that SM-3 possesses potential anti-HBV activity in vitro. This is the first report demonstrating the anti-HBV effects of S. media, which is currently under early development as a potential anti-HBV drug candidate.