Characterization of Indian honey and its anticancerous activity against colon cancer

Abstract

Four multifloral honey types from different Indian origin (named as Samples A, B, C and D) were analyzed for their phenolic acids using high performance liquid chromatography (HPLC). HPLC‐Mass Spectroscopy analysis revealed that dihydroxy benzoic acid, caffeic acid, ferulic acid and cinnamic acid were the major phenolic acid constituents of the tested honey. The total antioxidant potential and free radical scavenging ability varied among the honey samples and exhibited a significant correlation with their phenolic content. Further the honey samples were studied for rheology and thermal properties. Controlled shear rate viscometry indicated the Newtonian behavior (a drop in the viscosity as the temperature increases) in the honey samples. Differential Scanning Calorimetry (DSC) analysis revealed the inflection transition temperature (Tg) of the honey samples ranged between ‐ 51.40°C to −30.64°C. Thiazolyl blue tetrazolium bromide (MTT) assay revealed that the honey samples containing higher phenolic content showed significant antiproliferative effect against colon cancer cells. From the abovesaid experiments, Sample C among the four was found to be better based on its phenolic, antioxidant and antiproliferative effects and chosen for further studies. Honey induced the apoptotic signal via initial depletion of intracellular non protein thiols, consequently reducing the Mitochondrial Membrane Potential (MMP) and increased the Reactive Oxygen Species (ROS) generation. P53 up‐regulation and modulation of pro and antiapoptotic proteins were observed in the honey induced apoptosis. The work was extended further to study the constituents of honey that induced apoptosis. For this, we screened the antiproliferative effect of some reported phenolic constituents namely p‐coumaric acid, caffeic acid, gallic acid and eugenol in colon cancer cells. We had selected eugenol based on their better antiproliferative nature and elucidated the mechanism. Eugenol treatment resulted in reduction of intracellular non‐protein thiols and increase in the earlier lipid layer break. Further events like dissipation of mitochondrial membrane potential and generation of ROS were accompanied in the eugenol‐induced apoptosis. Finally we had studied the anticancer activity of honey samples (Samples B and C) and eugenol against Ehrlich ascites carcinoma. We found that Sample C remarkably inhibited the tumor growth by 40% compared to Sample B (12.17%) against Ehrlich ascites carcinoma when given 25% v/v intraperitoneally (i/p). We had found eugenol with a dose of 100 mg/Kg showed nearly 29% tumor inhibition.