Studies On Phase â…¡ Enzyme-Inducing And Anti-Inflammatory Activities Of Ginger Compounds

A wide variety of xenobiotic chemical carcinogens, can be absorbed by human body via various pathways, and lead to multiple diseases. The human body has developed complex systems to metabolize and detoxify these toxic substances. This primarily involved two sequential processes:Phase I and PhaseⅡreaction. Phase I reaction might convert some exogenous procarcinogens into more reactive carcinogen in some cases; Phase II reaction, catalyzed by phase II enzymes (QR, GST, HO-1, GCL and so on), can truly detoxify these carcinogen chemicals. It is well-documented that the induction of phaseⅡenzymes was correlated with celluar protection against toxic and reactive oxygen species (ROS) in animal and human bodies. Moreover, the evaluation of phaseⅡenzymes has been widely proposed as a major mechanism underlying chemopreventive actions of numerous plant phytochemicals. Since some synthetic phaseⅡenzyme-inducers exhibit undesirable consequences to enhance risk of carcinogenesis, this greatly initiated the efforts toward screening effective and safe inducers from plants. Up to date, many classes of plants or phytochemicals have been isolated and identified as potential chemopreventive agents, such as sulforaphane, curcumin, and certain withanolides. These inducers can upregulate phase II detoxification enzymes via activating Nrf2 signal pathway. These findings contribute a theoretical basis and a promising prospect for designing and developing effective chemopreventive agents against cancer in the future.Ginger (Zingiber Officinale Roscoe), has been utilized worldwidely as both a spice and a medical herb. In China, ginger has been grown for several thousand years. Numerous studies have demonstrated that ginger or its extracts present some degree of pharmacological activity including anti-inflammation, analgesic effect, anti-tumor and anti-oxidation activity. However, to the best of our knowledge, there are few literatures focusing on chemopreventive properties of ginger or ginger compunds, and on underlying mechanisms. In particular, no information is available regarding the phase II enzymes-inducing activity of ginger and/or ginger compounds. Under this interest, the following work was performed and the results are summarized:1. QR is used as a biomarker for phase II enzyme induction in cultured murine hepatoma cells (Hepa1c1c7). Guided by QR-inducing activity, frozen-dried ginger powder was firstly extracted with some solvents (ethyle acetate, ethanol and water), then fractionated with silica gel chromatography, sephadex LH-20 and thin layer chromatography, with the purpose of isolating potential phase II enzyme-inducing components. Four compounds were finally obtained and their structures were identified on the basis of ESI-MS、HR-ESI-MS and NMR spectroscopic methods. They were 1-dehydro-[6]-gingerdione (9.2mg)、6-shogaol (6.7 mg)、hexahydrocurcumin (3.9 mg) and 6-dehydroshogaol (6.9 mg)2. The in vitro antioxidant activity of four ginger compounds were evaluated for scavenging 1,1-diphenyl-2-picyrlhydrazyl (DPPH) and Trolox equivalent antioxidant capacity 2.2"-azinobis-(3-ethyl benzothiozoline-6-sulphonic acid) (ABTS) radicals. Results showed that four compounds could significantly scavenge DPPH radical at 150μM. Hexahydrocurcumin and 1-dehydro-[6]-gingerdione exhibited comparable higher potency in scavenging DPPH radicals at 60 min, with the inhibitory rate of 76.21±1.10% and 69.95±2.74%, respectively. The weakest inhibition (42.39±3.07%) was observed in 6-dehydroshogaol-treated group. TEAC test showed that four compounds were able to scavenge ABTS radicals in a concentration-dependent fashion (50-200μM). 1-Dehydro-[6]-gingerdione exhibited the strongest ABTS-scavenging activity, as indicated by its TEAC values of 95.18±1.53μM (50μM),188.61±3.39μM (100μM) and 375.45±7.11μM (200μM), then followed by hexahydrocurcumin,6-shogaol and 6-dehydroshogaol.3. In order to investigate the anti-inflammatory activity of four ginger compounds, lipopolysaccharide (LPS)-stimulated murine macrophage cells (RAW 264.7) were used. Results showed that 6-shogaol, 1-dehydro-[6]-gingerdione, and 6-dehydroshogaol was able to strongly suppress the NO production in LPS-stimulated RAW 264.7 cells. The anti-inflammatory activities of these compounds are ranked based on their IC50 values: 6-dehydroshogal (5.80±1.27μM),6-shogaol (7.50±0.44μM)> 1-dehydro-[6]-gingerdione (25.06±4.86μM)> hexahydrocurcumin (304.76±54.80μM). The cell viability was maintained> 80% for all compounds over the entire dose-range evaluated in comparison to control cells. The stronger NO inhibitory activity of 1-dehydro-[6]-gingerdione and 6-dehydroshogaol might be attributed to their down-regulating expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in LPS-stimulated macrophages.4. The induding potency of four ginger compounds on QR and GST, and underlying mechanisms were investigated using murine Hepa 1c1c7 cell bioassay. Results showed that 6-dehydroshogal, 1-dehydro-[6]-gingerdione and hexahydrocurcumin were able to significantly increase QR activty. The highest QR-inducing effect was observed in 6-dehydroshogaol-treated cells with a CD value of 9.23±0.22μM, followed by 1-dehydro-[6]-gingerdione (13.24±0.45μM), and then hexahydrocurcumin (68.81±3.90μM). Increasing QR activity was associated with elevated expression of NQO-1 protein in cultured cells.Incubation with 6-dehydroshogaol, 1-dehydro-[6]-gingerdione and 6-shogaol also lead to marked increases in GST activity in Hepa 1c1c7 cells. Further research indicated that 6-dehydroshogaol and 1-dehydro-[6]-gingerdione might up-regulate phaseⅡenzymes through Nrf2-Keap1 signaling pathway. Nrf2 was dissociated from Nrf2-Keap1 complex when stimulated by phaseⅡenzyme inducers, then translocated into the nucleus and bind ARE to enhance the expression of phaseⅡgenes.5. The effect of ginger extracts on endogenous antioxidant and phaseⅡenzymes was investigated in Wistar rats. The animals were fed with different doses of ginger extract (100, 200, and 400 mg/kg·BW) by gavage for 14 consecutive days and then half of animals were given 5 mg Benzo(a)pyrene intraperitoneally for 24 h. Results showed that after 14-day administration, catalase (CAT) activity in liver and blood serum of rats in all groups, glutathione peroxidase (GPx) activity in liver of rats in all groups, and superoxidase dismutase (SOD) activity in the liver of rats treated with Benzo(a)pyrene was significantly up-regulated (P< 0.05);QR and GST activity was markedly increased in liver, lung and kidney of rats that fed with ginger extracts, while uridine-5’-diphosphoglucuronosyl transferase (UGT) activity was partly up-regulated compared with the control.