INHIBITORY EFFECTS OF EXTRACTS OF CITRUS (CITRUS SPP.) PEELS ON KEY ENZYMES LINKED WITH ANGIOGENESIS AND METASTASIS IN COLON CANCER

Abstract

The burden of colon cancer is compounded by the cost of therapy, which is unbearable for low-income earners and patients in developing countries. This has led to a continual search for alternative and affordable therapy. This study investigated the effects of phenolic extracts from some Nigerian citrus peels [Grapefruit (Citrus paradisi), Orange (Citrus sinensis) and Shaddock (Citrus maxima)] on therapeutic targets [matrix metalloproteinases (MMP) and proteasome] relevant to colon cancer in Caco-2 (cell line isolated from a primary human colonic tumor), LoVo (metastatic human colorectal adenocarcinoma cell line) and LoVo ADR (drug resistant metastatic human colorectal adenocarcinoma cell line). The free phenolics were extracted with 80% acetone (v/v), while bound phenolics were extracted from the alkaline (NaOH) and acid (HCl) hydrolyzed residue with ethyl acetate. The total phenolic and flavonoid contents were determined and identified by high performance liquid chromatography coupled with a diode array detector (HPLC-DAD). The cellular antioxidant activities (CAA) of the extracts as well as their effect on glutathione reductase (GR) and glutathione peroxidase (GPx) activities in the colon cancer cells were determined. Furthermore, their reducing power, Fe2+- chelating ability and radicals [1,1 diphenyl-2- picrylhydrazyl (DPPH), hydroxyl (OH), nitric oxide (NO) and 2, 2-azinobis-3-ethylbenzo-thiazoline-6-sulfonate (ABTS)] scavenging abilities as well as inhibition of Fe2+ and sodium nitroprusside induced lipid peroxidation in rat colon were investigated. Bound total phenolics (27.4 mg/g) and flavonoids (6.9 mg/g) contents of orange were the highest, while shaddock bound total phenolics (11.6 mg/g) and flavonoids (3.4 mg/g) were the least. The free phenolic extracts had higher MMP inhibition than the corresponding bound phenolics in the colon cancer cells. Furthermore, free phenolics from orange had the strongest proteasome inhibition in Caco-2 and LoVo cell lysates, while free phenolics from grapefruit had the least. The extracts had dose-dependent inhibition of proteasome activity and had cellular antioxidant activities in the colon cancer and endothelial cells. In addition, the extracts increased the activities of intracellular GPx and GR and also displayed cell-free antioxidant activities. HPLC analysis revealed the presence of flavonoids (resveratrol, rutin, quercetin, quercetrin, naringin, kampferol, catechin, epicatechin), phenolic acids (caffeic acid, ellagic acid, chlorogenic acid) and tannin (gallic acid). The cellular antioxidant properties and inhibition of key enzymes relevant to colon cancer by the citrus peels' extracts make them good dietary/ nutraceutical means for the management and/or prevention of colon cancer.