Peroxisomes from pepper fruits (Capsicum annuum L.): purification, characterisation and antioxidant activity

Pepper is a vegetable of importance in human nutrition. Currently, one of the most interesting properties of natural products is their antioxidant content. In this work, the purification and characterisation of peroxisomes from fruits of a higher plant was carried out, and their antioxidative enzymatic and non-enzymatic content was investigated. Green and red pepper fruits (Capsicum annuum L., type Lamuyo) were used in this study. The analysis by electron microscopy showed that peroxisomes from both types of fruits contained crystalline cores which varied in shape and size, and the presence of chloroplasts and chromoplasts in green and red pepper fruits, respectively, was confirmed. Peroxisomes were purified by differential and sucrose density-gradient centrifugations. In the peroxisomal fractions, the activity of the photorespiration, beta-oxidation and glyoxylate cycle enzymes, and the ROS-related enzymes catalase, superoxide dismutase, xanthine oxidase, glutathione reductase and NADP(+)-dehydrogenases, was determined. Most enzymes studied had higher specific activity and protein content in green than in red fruits. By native PAGE and western blot analysis, the localisation of a Mn-SOD in fruit peroxisomes was demonstrated. The ascorbate and glutathione levels were also determined in crude extracts and in peroxisomes purified from both green and red peppers. The total ascorbate content (200-220 mg per 100 g FW) was similar in crude extracts from the two types of fruits, but higher in peroxisomes from red peppers. The glutathione concentration was 2-fold greater in green pepper crude extracts than in red fruits, whereas peroxisomes from both tissues showed similar values. The presence in pepper peroxisomes of different antioxidative enzymes and their corresponding metabolites implies that these organelles might be an important pool of antioxidants in fruit cells, where these enzymes could also act as modulators of signal molecules (O2*-, H202) during fruit maturation.