Yan-Hua Lu

Chemical fingerprint and quantitative analysis of Salvia plebeia R.Br. by high-performance liquid chromatography

To control the quality of Salvia plebeia R.Br., a simple and reliable method of high-performance liquid chromatography coupled with photodiode array detector (HPLC-DAD) was developed both for fingerprint analysis and quantitative determination of seven bioactive compounds, namely caffeic acid, luteolin-7-glucoside, nepetin-7-glucoside, homoplantaginin, luteolin, nepetin and hispidulin. In fingerprint analysis, twelve peaks were selected as characteristic peaks. In quantitative analysis, seven compounds showed good regression (R2>0.9995) within test ranges and the recovery of the method was in the range of 91.7-103.2%. The content ranges (mg/g) of seven compounds in the collected samples of S. plebeia were 0.80-1.67 (hispidulin), 2.18-5.75 (homoplantaginin), 0.52-1.22 (nepetin), 1.56-3.48 (nepetin-7-glucoside), 0.12-0.24 (luteolin), 0.97-2.22 (luteolin-7-glucoside) and 0.21-0.44 (caffeic acid), respectively. From the results obtained, the content of homoplantaginin was the highest. In addition, luteolin and luteolin-7-glucoside were isolated for the first time from S. plebeia.

Tyrosinase inhibitory effects and inhibition mechanisms of nobiletin and hesperidin from citrus peel crude extracts

The inhibitory effects of nobiletin and hesperidin from citrus peel crude extracts on tyrosinase diphenolase activity are evaluated. IC50 of nobiletin and hesperidin is 1.49 mM and 16.08 mM, respectively and their inhibition mechanism is competitive type with Ki = 2.82 mM and noncompetitive with Ki = 9.16 mM, respectively. Crude extracts from citrus peel (C. unshiu Marc.) were extracted with 95% ethanol and fractionated by petroleum ether (PCPE). The ethanol phase (ECPE) was further desorbed from macroporous adsorption resin (FGRE). Their IC50 values were 8.09 mg/mL, 7.53 mg/mL and 4.80 mg/mL, respectively. Their inhibition on melanogenesis in B16 mouse melanoma cells was also evaluated. FGRE showed a significant inhibition (42.5% at 31.25 μg/mL, p < 0.01) while hesperidin showed almost no inhibition. Nobiletin and PCPE give efficacious antiproliferation effects on B16 mouse melanoma cell with IC50 values 88.6 μM and 62.96 μg/mL, respectively, by the MTT test. Hesperidin and other crude extracts showed very low cytotoxity to the B16 cell.

Neuroprotective Effects of Hypericum perforatum on Trauma Induced by Hydrogen Peroxide in PC12 Cells

The standard extracts of Hypericum perforatum L. (SEHP), a well-known medicinal plant, are used for the treatment of depression, exhibited upgrading and significant protective effects on the trauma of PC12 cells induced by 200 microM H2O2 in a dose-dependent manner within 24-hour treatment. Cell viability was assessed by the MTT method, and in situ cellular hydrogen peroxide (H2O2)-induced oxidative stress was examined by measurement of reactive oxygen species (ROS) formation using CDCFH procedures. Intra- and extra-cellular ROS levels decreased significantly to 71.9% and 50.0% of the control at a moderate concentration of 20 microg/ml, respectively, suggesting that SEHP could easily enter the cells and play important roles in reducing ROS levels. Our results were proved by detection of DNA fragmentation and inspection of cell morphology of PC12 cells. SEHP can obviously block DNA fragmentation and prevent the cells from shrinking and turning round of H2O2-induced apoptosis in PC12 cells at concentrations of 10 approximately 100 microg/ml. This data suggests SEHP may be a candidate for application in neurodegenerative diseases such as Alzheimers disease or Parkinsons disease.

Mechanism and inhibitory effect of galangin and its flavonoid mixture from Alpinia officinarum on mushroom tyrosinase and B16 murine melanoma cells

The whitening effects of the flavonoid constituents of Alpinia officinarum Hance were investigated on melanin biosynthesis in B16 mouse melanoma cells, tyrosinase inhibition and UV absorption. The melanin content was reduced to 1.276 μg /105cell for flavonoid mixture and 1.161 μg /105cell for galangin while the melanin control was 1.632 μg /105cell. Both flavonoid mixture and galangin reduced melanin production with an inhibition of 21.81% and 28.86% at a concentration of 26.5 μ g/mL and 29 μg /mL (107.4 μM), respectively. Tyrosinase inhibition by the flavonoid mixture and galangin were higher at lower concentrations and galangin showed competitive inhibition at a concentration less than 21.23 μg/mL which was soluble. In addition, the flavonoid mixture and galangin showed a broad absorption band at 270 ∼ 290 nm related to the UV-B area. These observations suggest that galangin may be a whitening agent and a promising candidate for prevention of skin cancer. This is the first full scale report on the evaluation of the whitening effect of galangin.

Tyrosinase inhibitory effect and inhibitory mechanism of tiliroside from raspberry

Tiliroside was found to inhibit both monophenolase and diphenolase activity of mushroom tyrosinase. The lag time of tyrosine oxidation catalyzed by mushroom tyrosinase was obviously lengthened; 0.337 mM of tiliroside resulted in the lag time extension from 46.7 s to 435.1 s. A kinetic analysis shown that tiliroside was a competitive inhibitor for monophenolase and diphenolase with Ki values of 0.052 mM and 0.26 mM, respectively. Furthermore, tiliroside showed 34.5% (p < 0.05) inhibition of intracellular tyrosinase activity and 54.1% (p < 0.05) inhibition of melanin production with low cytotoxicity on B16 mouse melanoma cells at 0.168 mM. In contrast, arbutin displayed 9.1% inhibition of cellular tyrosinase activity and 29.5% inhibition of melanin production at the same concentration. These results suggested that tiliroside was a potent tyrosinase inhibitor and might be used as a skin-whitening agent and pigmentation medicine.

Inhibitory effects of Schisandra chinensis extract on acne-related inflammation and UVB-induced photoageing.

Abstract Context: Schisandra chinensis (Turcz.) Baill. (Schisandraceae) fruit extract (SFE) has been reported to induce non-specific tissue protection against inflammation in vivo. However, the effects of SFE on Propionibacterium acnes-stimulated acne and UVB-irradiated photoageing have yet to be investigated. Objective: To systematically investigate the effects of SFE against P. acnes and photoageing in vitro. Materials and methods: Qualitative and quantitative analyses of SFE were performed by HPLC. SFE concentrations from 2.5 to 50 μg/mL were tested. Specifically, ELISA was used to examine the levels of pro-inflammatory cytokines in THP-1 cells as well as of collagen I and matrix metalloproteinases-1 in HDF cells. The anti-bacterial effect of SFE was determined using the microdilution broth method. Glutathione and malondialdehyde levels were examined using the colorimetric and TBA methods, respectively. The degree of ageing was determined by cytochemical staining. Results: SFE significantly inhibited P. acnes growth (MIC 0.5 mg/mL) and 50 μg/mL of SFE suppressed the production of interleukin-1β, interleukin-8 and tumour necrosis factor α, by 59.67%, 62.69% and 68.30%, respectively, in P. acnes-stimulated THP-1 cells. Additionally, 10 μg/mL of SFE suppressed photoageing in UVB-exposed fibroblasts by decreasing metalloproteinase levels by 88.4%, inducing collagen by 58.4% and activating the anti-oxidant defence system, by limiting lipid peroxidation by 51.1% and increasing glutathione production by 34.1% (2.5 μg/mL SFE). Discussion and conclusion: These results indicated that SFE could significantly ameliorate the inflammatory state in P. acnes-stimulated THP-1 and UVB-irradiated HDF cells, suggesting its potential as a novel agent for acne therapy and photoageing prevention.

Anti-photoageing and anti-melanogenesis activities of chrysin.

Abstract Context: Melanin plays an important role in preventing skin photoageing by blocking ultraviolet B (UVB). However, East Asian women prefer light and fair skin, therefore they want to keep their skin from photoageing and at the same time reduce the melanin in their skin. Chrysin is a kind of natural flavonoid with luxurious biological activities, which has a very promising effect on achieving this goal. Objective: To elucidate the effects and mechanisms of chrysin on photoageing and melanogenesis. Materials and methods: Human dermal fibroblasts (HDF) and B16 murine melanoma cells were incubated with chrysin (0–25 μM) for 48 h. Anti-photoageing activity was examined in HDF by assessment of synthesis/degradation of collagen I, antioxidative and antisenescent activities through ELISA and colorimetric method. Anti-melanogenesis activity was tested by assessment of melanin, tyrosinase (TYR), melanogenic proteins inhibition activities in B16 cells using colorimetric and ELISA method. Results: Chrysin increased collagen I secretion (50–121.54% at 6.25–25 μM) and chrysin showed anti-photoageing activity by decreasing the degradation of collagen I, repairing oxidation damage and reducing the rate of HDF senescence. Furthermore, chrysin exhibited inhibitory activities with 3.00–20.35% reduction of melanin content at 6.25–25 μM, and inhibited melanin synthesis through the inhibition of TYR activity and the suppression of melanogenic proteins (TYR, TYR-related protein-1/2 and microphthalmia-associated transcription factor) expressions. Discussion and conclusion: Chrysin may have potential for developing a functional cosmetic agent because of its anti-photoageing and anti-melanogenesis activities.

Protective Effects of Salidroside on Hypoxia/Reoxygenation Injury by Sodium Hydrosulfite in PC12 Cells

Abstract Hypoxia/reoxygenation causes cellular injury and death associated with many pathophysiologic conditions, including respiratory disorders, myocardial ischemia, and tumour progression diseases. Neuronal pheochromocytoma (PC12) cells are widely used as a model system for neurologic research and are subject to chemical hypoxia induced with sodium hydrosulfite (Na2S2O4), a common oxygen-consuming agent. Salidroside, which is the main active component of the famous traditional Chinese herb Rhodiola rosea. L. (Crassulaceae), has been proved to possess many bioactivities. In this article, we studied the protective effects of salidroside on hypoxia/reoxygenation injury in PC12 cells induced by Na2S2O4. Cultures of PC12 cells were exposed for 1 h to 10 mM Na2S2O4 for hypoxia, followed by reoxygenation for 2 h. The results showed that salidroside was very stable in medium and was not harmful to PC12 cells at the experimental concentrations of 0 ∼ 200 µg/mL. The cytoprotection by salidroside was dose-dependent, and the cell viability was 41.8 ± 5.7%, 62.4 ± 4.1%, and 92.2 ± 3.7% at 0, 50, and 100 µg/mL of salidroside, respectively. The level of released LDH significantly decreased from 513.5 ± 5.5% (without salidroside) to 258.1 ± 6.3% (with 100 µg/mL salidroside). Flow cytometry was performed to measure apoptotic rate. The results of flow cytometry assay indicated that the apoptotic rate was 17.0 ± 1.2% after hypoxia/reoxygenation injury. When the cells were treated with salidroside 12.5, 50 and 100 µg/mL, the apoptotic rate was 9.5 ± 0.9%, 7.4 ± 0.5%, and 4.5 ± 0.4%, respectively. In addition, our results were confirmed by inspection of cell morphology of PC12 cells. Treatment with salidroside (12.5, 50, 100 µg/mL) significantly prevented the cells from morphologic changes. All the above results showed salidroside could effectively protect PC12 cell against hypoxia/reoxygenation injury.

Activity of Schisandrin C Isolated from Schisandra chinensis against Human Cancer Cell Lines

Schisandrin C, a dibenzocyclooctadiene lignan isolated from the fruits of Schisandra chinensis (Turcz.) Baill. (Schisandraceae), was investigated for cytotoxicity and influence on three human cancer cell lines. Among human hepatocelluar carcinoma cells (Bel-7402), human breast cancer cells (Bcap37) and human nasopharyngeal carcinoma cells (KB-3-1), Bel-7402 cells were most sensitive with IC50 equal to 81.58 ± 1.06 μ M after treatment with schisandrin C for 48 h. Cytotoxicity of schisandrin C on tumor cells depends on cellular accumulation of the drug. The intracellular schisandrin C concentration was assayed by high-performance liquid chromatography (HPLC) and showed that schisandrin C could permeate the cell membrane. Staining with Hoechst 33258 showed fragmentation and condensation of chromatin in the cell treated with 75 μ M schisandrin C for 24 h. Flow cytometric analysis was performed to determine hypodiploid cells. The results of flow cytometry indicated that the percentage of hypodiploid Bel-7402 cells was 40.61 ± 1.43% after 24 h treatment with 100 μ M schisandrin C. The treatment resulted in the appearance of a hypodiploid peak (sub-G0/G1 region), probably due to the presence of apoptosing cells and apoptotic bodies with DNA content less than 2n. To our knowledge, this is the first report against human hepatocelluar carcinoma cells (Bel-7402) of schisandrin C.