Research On The Antioxidant,antihyperglycemic And Antiproliferative Activities Of Phenolics From Sugarcane(saccharum Officinarum L.)

For many years,chronic diseases such as diabetes and cancers have been considered as one of the main factors affecting health and the quality of human life and reactive oxygen species?ROS?play a key role in the development of these diseases.Currently many clinical drugs have side-effects.Thus the exploitation on the natural products to discover the agents with high antioxidant activity but low toxicity has been a popular research area.Sugarcane?Saccharum officinarum L.?has been considered as an important cash crop globally.After eating or extraction of juice,most of the remaining bagasse was used as fuel to power the mills.Several attempts have been conducted using bagasse as raw material including biomass and bioenergy product but as a kind of natural products,the health-care values of bagasse should not be ignored.This work is aimed to study the phenolic profile and the antioxidant,antihyperglycemic and antiproliferative activities of phenolics from sugarcane based on our previous work.First,the total phenolic content of the extracts which were previously prepared was measured.The glucose uptake activity of the very active fraction based on HepG2 cells was conducted.The correlation between total phenolic content and antioxidant/antihyperglycemic activity was analyzed.UHPLC-HR-TOFMS was used to determine the phenolic compounds in the active fraction.The results showed that the total phenolic content of sugarcane bagasse was 7.83 ± 0.24 mg GAE/g,including free phenolics?EF?7.35 ± 0.25 mg GAE/g and bound phenolics 0.48 ± 0.01 mg GAE/g.The fraction with the highest total phenolic contents was ethyl acetate fraction?EAF,241.42 ± 1.91 mg GAE/g?,followed by n-butanol fraction?BF?.The total phenolic content of hydroalcoholic extracts?30% – 70%E?followed the order: 30%E>50%E>70%E,ranging from 170.68 ± 3.25 mg GAE/g to 73.60 ± 0.67 mg GAE/g.The total phenolic content showed significant correlation with the antioxidant/antihyperglycemic activity.30%E showed no cytotoxicity against HepG2 cells at 0.05 – 0.1 mg/mL and showed significant activity to improve the glucose uptake on insulin-resistance HepG2 cell model.The glucose uptake improvement activity of 30%E at 0.1 mg/mL is significantly higher than the positive reference rosiglitazone?10 ?M?in this experimental condition.Then seven phenolic compounds existing in sugarcane were used to evaluate their antioxidant activity.Only quercetin,luteolin and protocatechuic acid showed antioxidant capacity in peroxyl radical scavenging capacity?PSC?assay.Tricin showed the highest activity in the oxygen radical absorbance capacity?ORAC?assay?9.76 ± 1.01 ?mol TE/?mol?.In cellular antioxidant activity?CAA?assay,quercetin and luteolin displayed antioxidant activity intracellularly to scavenge ROS but protocatechuic acid only had extracellular activity?on plasma membrane?.After analysis of the structures of seven phenolics and the different activities presented in CAA,it was confirmed with literature that the o-diphenolic group plays a key role in CAA assay and the lipophilicity and bioavailability can also influence the cellular antioxidant activity.Hep G2 cells and hydrogen peroxide?H2O2?were used to build up the oxidative damage model.Under the protection of medium and high concentrations of luteolin,the cell viability was significantly higher than that of the damaged group.The increase rate of ROS,the enzymatic activities of LDH,SOD and CAT and the apotosis rate of the protected groups were significantly lower than those of the damaged group.At last,quercetin,luteolin,apigenin,tricin,tricin 7-O-?-glucopyranoside?T7G?,p-coumaric acid and protocatechuic acid were used to evaluated the antiproliferative activity against human liver cancer HepG2 cells and human colon cancer Caco-2 cells;The research based on human breast cancer MCF-7 cells were conducted to study the combination effects among quercetin,luteolin,p-coumaric acid,tricin and apigenin on the proliferation and cytotoxicity of MCF-7 cells.Luteolin showed the best inhibitory activity against HepG2 cells with an EC₅0 value at 36.28 ± 0.30 ?M and the CC50 value of luteolin on HepG2 was higher than 113 ?M and the selective index?SI?was higher than 2.Compared to HepG2 cells,quercetin,luteolin and apigenin showed lower activity against the proliferation of Caco-2 cells and the EC₅0 values of Caco-2 were higher than those of HepG2.Apigenin had the best effect against Caco-2 cell proliferation with an EC₅0 value at 213.91 ± 2.95 ?M.The CC50 value of apigenin against Caco-2 cells was higher than 311 ?M and the SI value was higher than 1.55.The combination effect of apigenin and luteolin showed an additive effect with a CI value at 1.1 at EC₅0.The EC₅0 of luteolin and quercetin against MCF-7 cells was 28.45 ± 0.91 ?M and 161.30 ± 1.48 ?M,respectively and the values decreased to 14.91 ± 0.34 ?M and 61.46 ± 1.46 ?M,respectively,after the combination with tricin.The combination effect of quercetin and luteolin showed a moderate antagonistic effect with a CI value at 1.22 at EC₅0.Luteolin and p-coumaric acid showed good synergistic activity against the proliferation of MCF-7 cells in the indicated concentrations and the CI value decreased with the increase of concentrations.At EC₅0,the CI value was 0.80 ± 0.02 and the dose reduction index?DRI?of luteolin and p-coumaric acid was 1.80 ± 0.07 and 4.04 ± 0.11,respectively.When the inhibitory effect against MCF-7 cells was 22%,the CI value of the combination of quercetin and p-coumaric acid was 0.70 ± 0.14 which was a moderate synergistic effect.However,the CI value increased with the inhibition rate and the antagonistic effect arose.At EC₅0,the DRI of p-coumaric acid and quercetin was 1.29 ± 0.06 and 1.45 ± 0.17,respectively.And the CI value at EC₅0 was 1.47 ± 0.11.