Lakhasly

1. Introduction
   Ingestion of dietary phytochemicals from fruits and vegetables has proven to play an important role in pre- venting diseases due to their potent antioxidant activ- ity. Screening of raw materials for identifying potential antioxidant activity has resulted in a large body of published works over the past three decades. However, there is a gap in knowledge about the health benefits, if they may either be ascribed to bioactive phytochemicals like vitamins C and E, carotenoids and polyphenolics or also attributed to chlorophylls. Nevertheless, in most research, chlorophylls had not been included in the trials despite being the most abun dant pigments in nature. The scarcity of information could be attributed to the difficulty to obtain chloro- phylls and their derivatives in a purified form and also to their instability, which triggers a degradation pro- cess. A research group in Japan (Endo, Usuki, & Kan- eda, 1985a, 1985b; Usuki, Endo, & Kaneda, 1984b; Usuki, Suzuki, Endo, & Kaneda, 1984) first suggested a pro-oxidant activity of chlorophylls under light, which could be understood as a transfer of the energy of singlet-excited chlorophyll to oxygen that would form reactive oxygen species. However, the same authors also reported that chlorophylls and pheophy- tins provide protection by preventing autoxidation of vegetable edible oils stored in the dark and suggested a hydrogen donating mechanism breaking the radical chain reactions. In addition, they stated that the intact chemical structure of porphyrin seems to be essential for antioxidant activity.

Wanasundara and Shahidi (1998) reported that the presence of chlorophyll in tea extracts was responsible for a pro-oxidant effect on the oxidation of marine oils. Hoshina, Tomita, and Shioi (1998) found that chloro- phylls were better antioxidants than their metal free derivatives and confirmed the importance of the porphy- rin ring on the inhibition of lipid autoxidation. Sakata et al. (1990) reported that a related compound to pheo- phorbide a, isolated from clam, appeared to be the most important antioxidant in the organism. Ferruzzi, Bo ̈hm, Courtney, and Schwartz (2002) provided information concerning in vitro antioxidant and antimutagenic activ- ities of water and lipid soluble chlorophyll derivatives as well as of metal chelated derivatives by free radical scav- enge and bacterial reverse mutagenesis assays, respec- tively. The antioxidant activities of the synthetic metallo-chlorophyll derivatives, especially Cu-chelated compounds were found to be much higher than those of natural chlorophylls and of Mg-free derivatives, which showed an almost insignificant antioxidant capac- ity. The highest activities were found for Cu-substituted chlorins. Previously, Sato et al. (1986) identified Cu- chlorin e4 as the major antioxidant found in commercial grade sodium Cu-chlorophyllin, by measuring its anti- oxidant activity in rat liver homogenates.
Questions have been raised about the natural chloro- phyll metabolites impact on health, due to their high content in some vegetables, widespread occurrence in nature, and their degradation during food processing and human digestion. As very few data are available on both the antioxidant activity and dose-dependent re- sponse, the aim of this work was to evaluate not only the protective action of six natural purified chlorophyll derivatives (chlorophyll a and b, pheophytin a and b, pheophorbide a and b) but also the synthetic Cu-chloro- phyllin against lipid oxidation employing two methods: the bleaching of b-carotene in a water/linoleic acid emulsion and the scavenging of the stable radical 2,2-di- phenyl-1-picrylhydrazyl (DPPH).