Cytoprotective effect of a bilberry extract against oxidative damage of rat hepatocytes

Abstract

The effect of a bilberry extract (BE, 25% anthocyanins) against oxidative damage in primary cultures of rat hepatocytes, induced by tert-butyl hydroperoxide and allyl alcohol, was investigated. BE displayed cytoprotective effects at 100 and 500 μg/ml in the MTT viability test. It protected the cells against lactate dehydrogenase leakage and lipoperoxidation products formation. Maximum protection (58%) was noted using 500 μg/ml of BE and intoxication by allyl alcohol. The observed cytoprotective effect is probably due to the antioxidant properties of its constituents, mainly anthocyanins. BE scavenged DPPH (IC₅₀ 3.99 ± 0.14 μg/ml) and enzymatically generated superoxide radical with an activity equivalent to 108 ± 7.2 U of superoxide dismutase per mg of extract. Our results support the use of bilberry and bilberry extracts in functional foods and food supplements designed for the prevention of chronic diseases associated with oxidative stress.

Introduction

Vaccinium myrtillus L. (bilberry) is a member of the Eriacaceae family and is also known as European blueberry, huckleberry, whortleberry or blueberry. It is a shrubby perennial plant that can be found in the mountains and forests of Europe and in the northern United States. The fruit of the bilberry plant is a blue-black or purple berry and differs from the American blueberry in that the flesh of the berry is purple, rather than cream or white. Bilberry has been used as food for centuries, due to its high nutritive value, and today it represents a precious wild delicacy. Bilberry’s history of medicinal use dates back to the Middle Ages but it did not become widely known to herbalists until the 16th century, when it was used for treating bladder stones, biliary disorders, scurvy, coughs and lung tuberculosis. More recently, bilberry fruit extracts have been used for the treatment of diarrhea, dysentery, and mouth and throat inflammations (Anonymous, 2001).

The berries contain the anthocyanoside flavonoids (anthocyanins, Nakajima, Tanaka, Seo, Yamazaki, & Saito, 2004), stilbenoids (resveratrol, Lyons et al., 2003), vitamins, sugars and pectins. The anthocyanins are considered the most important of the pharmacologically active constituents. Anthocyanin concentration in the fresh fruit is approximately 0.1–0.5%, while concentrated bilberry extracts are usually standardized to 25% anthocyanins (Ichiyanagi et al., 2004, Zhang et al., 2004). Phenolic compounds, including anthocyanins, are known as strong antioxidants. While in vitro antioxidant activity of bilberry extracts has been studied in various non-cellular models (Laplaud et al., 1997, Martín-Aragon et al., 1998, Martín-Aragon et al., 1999), cytoprotective activity against oxidative damage has not yet been documented. The aim of our study was to investigate the effect of a bilberry extract (BE) against oxidative cell damage induced by tert-butyl hydroperoxide and allyl alcohol in primary cultures of rat hepatocytes.