Preclinical safety assessment of antipyrine combined with lidocaine hydrochloride as ear drops.

This study was designed to assess the preclinical toxicity of antipyrine combined with lidocaine hydrochloride ear drops (ALED) and support the clinical trials of ALED in clinical settings in China. All the experiments including acute toxicity in rodents, skin sensitization toxicity in guinea pigs, skin irritation toxicity in rabbits and chronic toxicity in rats were performed according to China Food and Drug Administration guidelines. The maximum tolerated dose (MTD) of ALED administration for mice and rats was over (400 g antipyrine plus 100 g lidocaine hydrochloride)/kg and (240 g andtipyrine plus 60 g lidocaine hydrochloride)/kg, respectively. No obvious skin sensitization toxicity and skin irritation toxicity were observed. The main changes concentrated in chronic toxicity study in rats. For the chronic toxicity, rats were administrated once a day for 28 consecutive days, and a 14-day recovery period was followed. The side effects of ALED included decreased dietary intake in male rats, increased proportion of reticulocytes, decreased or even inversed granulocyte:erythrocyte ratio, fluctuated alanine aminotransferase and aspartate aminotransferase, and slightly increased relative weight of liver. Conclusively, blood system (especially erythrocyte system) and digestive system, including liver and gastrointestinal tract, might be the toxic targets of ALED.

Microbial transformation of hederagenin by Cunninghamella echinulate, Mucor subtilissimus, and Pseudomonas oleovorans.

The pentacyclic triterpenoid hederagenin (1) was subjected to biotransformation by Cunninghamella echinulate CGMCC 3.2000, Mucor subtilissimus CGMCC 3.2454 and Pseudomonas oleovorans CGMCC 1.1641. Three metabolites were obtained. On the basis of nuclear magnetic resonance and high-resolution mass spectral analyses, their structures were characterized as 3ß, 23-dihydroxyolean-12-en-28-oic acid 28-O-ß-D-glucopyranosyl ester (2), 3ß, 15α, 23-trihydroxyolean-12-en-28-oic acid (3), 1ß, 3ß, 23-trihydroxyolean-12-en-28-oic acid (4), and metabolite (3) was a new compound. This was the first report on the biotransformation of hederagenin.

Microbial transformation of the anti-diabetic agent corosolic acid by Cunninghamella echinulata.

The pentacyclic triterpenoid corosolic acid was metabolized by Cunninghamella echinulata CGMCC 3.2000 to its C-24 aldehyde group metabolite and five other hydroxylated metabolites: madasiatic acid (2), 2α, 3ß, 7ß-trihydroxyurs-12-en-28-oic acid (3), 2α, 3ß, 15α-trihydroxyurs-12-en-28-oic acid (4), 2α, 3ß, 6ß, 7ß-tetrahydroxyurs-12-en-28-oic acid (5), 2α, 3ß, 7ß, 15α-tetrahydroxyurs-12-en-28-oic acid (6), and 2α, 3ß,7ß-trihydroxy-24-al-urs-12-en-28-oic acid (7); compounds 3, 5, and 7 were new compounds. The α-glucosidase inhibitory effects of the metabolites were also evaluated.

Cordyceps sinensis Oral Liquid Inhibits Damage Induced by Oxygen and Glucose Deprivation in SH-SY5Y Cells.

CONTEXT: Cordyceps sinensis has been used in traditional Chinese medicine for thousands of years. It has been demonstrated to have a variety of biological activities, and an extract of it has been demonstrated to possess a protective effect in occlusion-induced focal cerebral ischemia of the middle cerebral artery in rats. It could be explored as an agent for treatment of ischemic stroke, and the mechanisms need to be studied further. OBJECTIVE: The study intended to investigate the protective effects of the Cordyceps sinensis oral liquid (CSOL) against damage induced by oxygen and glucose deprivation (OGD) in SH-SY5Y cells. DESIGN • The research team designed an in vitro study. SETTING: The study occurred at the Naval Medical Research Institute in Shanghai, China. INTERVENTION: SH-SY5Y cells were exposed to CSOL in doses of 0.01, 0.03, 0.10, 0.30, and 1.00 mg/mL, creating 5 intervention groups. The OGD condition was induced by transfer of the cells from high-glucose Dulbecco's Modified Eagle's medium (DMEM) in a box gassed with air containing 5% CO2 to glucose-free DMEM in a box gassed with 94% N2, 5% CO2, and 1% O2. Like the cells for the interventions groups, the cells for a model group were cultured with high-glucose DMEM and were transferred to the OGD, but they received no dose of COSL. Cells in a control group were cultured with high-glucose DMEM, were not transferred to the OGD condition, and did not receive any dose of COSL. OUTCOME MEASURES: Cell viability was assayed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method. The apoptosis and the mitochondrial membrane potential (MMP) were detected by flow cytometry, and the protein expression of caspase-3 was observed by western blot. RESULTS: After exposure to OGD, the cell viability of cells treated with 0.01, 0.03, 0.10, 0.30, and 1.00 mg/mL of CSOL increased in a dose-effect relationship. Compared with the cells in the model group, the treatment of CSOL at all the experimental concentrations significantly inhibited both the cell apoptosis (P < .01) and the capase-3 activation (P < .01). The MMP dissipation in the cells of the model group increased significantly compared with those of the control group (P < .01). The treatment with all doses of CSOL significantly inhibited the MMP dissipation (P < .01). CONCLUSIONS: CSOL protects against the damage induced by OGD through inhibiting the mitochondrial apoptosis pathway in SH-SY5Y cells.

Microbial transformation of asiatic acid.

Asiatic acid (1), a major pentacyclic triterpene of Centella asiatica, was subjected to transformation by Penicillium lilacinum ACCC 31890, Fusarium equiseti CGMCC 3.3658, and Streptomyces griseus CGMCC 4.18 strains. Incubation of asiatic acid with P. lilacinum ACCC 31890 and F. equiseti CGMCC 3.3658 gave an identical product: 2α,3ß,15α,23-tetrahydroxyurs-12-en-28-oic acid (2). Biotransformation of asiatic acid by S. griseus CGMCC 4.18 resulted in three derivatives: 2α,3ß,21ß,23-tetrahydroxyurs-12-en-28-oic acid (3), 2α,3ß,23-trihydroxyurs-12-en-28, 30-dioic acid (4), and 2α,3ß,23,30-tetrahydroxyurs-12-en-28-oic acid (5). The structures of those derivatives were deduced from their spectral data. Products (2), (3), and (4) were new compounds. In addition, the in vitro cytotoxicities of those derivatives along with 1 were evaluated with several human cancer cell lines.

[Effect of Cordyceps sinensis sporocarp on learning-memory in mice].

OBJECTIVE: To study the effect of extracts of Cordyceps sinensis sporocarp on learning-memory in scopolamine treated mice and the possible mechanism. METHODS: ICR mice were randomly divided into five groups: sham control, model, piracetam and CSE 0.5, 1 g/kg. Lotomotor activity was assessed. Morris water maze was used to evaluate the memory ability of mice 30 min later after ip scopolamine 1.0 mg/kg BW. Then acitivity of AchE was measured after behavioral test. RESULTS: CSE had no influence on lotomotor activity. However, CSE 0.5, 1 g/kg both shortened escape latency and increased times of come-crossing platform in Morris water maze, meanwhile activity of AchE in the brain was decreased by CSE. CONCLUSION: CSE can significantly improve the learning and memory impairment in mice induced by scopolamine, which may be correlated with the inhibition of activity of AchE.

The cytotoxic and mechanistic effects of aaptamine on hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is the fifth most common form of cancer and the third most frequent cause of cancer-associated mortality worldwide. We isolated aaptamine from the marine sponge Aaptos, and synthesized derivatives of this compound. Aaptamine and synthetic derivatives displayed various biological activities. This represents the first account of studies on the effects of aaptamine and its derivatives in hepatocarcinogenesis. In this study, Cell Counting Kit (CCK8) was used to evaluate the anti-proliferative effect of aaptamine on HCC in vitro. Additionally, a subcutaneous xenograft model was used to determine if aaptamine could inhibit hepatocellular carcinoma in vivo. We also used RT-PCR and Western blot to analyze the mechanisms behind these effects. Our results showed that aaptamine has anti-proliferation effects on the cell lines LM3 and HepG2. Aaptamine also suppressed the colony-formation ability of HCC cells. We found that aaptamine treatment led to cell cycle arrest in HCC cells, reduced the expression of SOX9 and CDK2. Significant anti-tumor effects were observed in aaptamine-administered tumor-bearing mice as compared to controls. However, structural changes made to aaptamine yielded two derivatives for which all the effects listed above were considerably reduced as compared to the original compound aaptamine. In conclusion, aaptamine is demonstrated for the first time to inhibit liver cancer progression. The aaptamine-induced cell cycle arrest was associated with the increased binding of p21 to Cdk2-cyclin D/E complexes, inhibition of Cdk2 kinase activity in HCC cells. Furthermore, aaptamine appears to be a promising and efficient treatment of liver cancer HCC-LM3 in vivo. We have also uncovered structural changes that might affect the biological activity. The work provides a promising drug candidate for HCC treatment.

SCAIF80, a Novel Inhibitor of Angiogenesis, and Its Effect on Tumor Growth.

A novel inhibitor of angiogenesis named SCAIF80 (shark cartilage-derived angiogenesis inhibitory factor) from shark cartilage has been isolated and characterized. SDS-PAGE analysis followed by silver staining revealed a single band with molecular weight (M(r)) of 80 kD. To determine whether this protein was capable of inhibiting angiogensis, it was assayed in endothelial cell (EC) proliferation and migration assay. The results showed that SCAIF80 significantly suppressed EC proliferation and migration in a dose-dependent manner. In the chick chorioallantoic membrane (CAM) assay, SCAIF80 also showed a potent inhibitory activity on angiogenesis in vivo. In animal tests, the growth of tumor was potently suppressed by SCAIF80 therapy. Lewis lung carcinoma was inhibited by 93.83 % at a dose of 5 mg/(kg.d). These findings suggest that shark cartilage may produce a novel protein with anti-angiogenic and anti-tumor activity.

Microbial transformation of the anti-diabetic agent corosolic acid.

Biotransformation of corosolic acid (1) by Cochliobolus lunatus and Streptomyces asparaginoviolaceus afforded four metabolites, which were identified by using (1)H NMR, (13)C NMR, DEPT, HSQC, HMBC and NOESY spectral data. Biotransformation of corosolic acid by C. lunatus R.R. Nelson & Haasis CGMCC 3.4381 produced three metabolites: 2α,3ß,21ß-trihydroxyurs-12-en-28-oic acid (2), 2α,3ß,7ß,21ß-tetrahydroxy-urs-12-en-28-oic acid (3) and 2α,3ß-dihydroxy-21-oxours-12-en-28-oic acid (4). Incubation of corosolic acid with growing cultures of S. asparaginoviolaceus CGMCC 4.0175 afforded metabolite 2α,3ß,30-trihydroxyurs-12-en-28-oic acid (5). All the metabolites were reported for the first time. The substrate and four metabolites, along with four products obtained previously, were evaluated for their inhibitory effects on α-glucosidase; all the triterpenes tested showed potent inhibitory effects.