Cytotoxic and Antioxidant Activity of 4

Cytotoxic and Antioxidant Activity of 4-Methylthio-3-butenyl Isothiocyanate from Raphanus sativus L. (Kaiware Daikon) Sprouts

J. Agric. Food Chem.,56 (3),875-883,2008.10.1021/jf073123c
Alessio Papi,†Marina Orlandi,† Giovanna Bartolini,*† Jessica Barillari,§ Renato Iori,*§ Moreno Paolini,# Fiammetta Ferroni,Maria Grazia Fumo,⊥Gian Franco Pedulli,⊥and Luca Valgimigli*⊥

Dipartimento di Biologia Evoluzionistica e Sperimentale, Università di Bologna, via Selmi 3, 40126 Bologna, Italy; Consiglio per la Ricerca e la Sperimentazione in Agricoltura, Centro di Ricerca per le Colture Industriali (CRA-ISCI), via di Corticella 133, 4129 Bologna, Italy; Dipartimento di Farmacologia, Università di Bologna, via Irnerio 48, 40126 Bologna, Italy; and Dipartimento di Chimica Organica "A. Mangini", Università di Bologna, via San Giacomo 11, 40126 Bologna, Italy

Received October 24, 2007




There is high current interest in the chemopreventive potential of Brassica vegetables (cruciferae), particularly due to their content in glucosinolates (GL), which upon myrosinase hydrolysis release the corresponding isythiocyanates (ITC). Some ITCs, such as sulforaphane (SFN) from broccoli (Brassica oleacea italica), have been found to possess anticancer activity through induction of apoptosis in selected cell lines, as well as indirect antioxidant activity through induction of phase II detoxifying enzymes. Japanese daikon (Raphanus sativus L.) is possibly the vegetable with the highest per capita consumption within the Brassicaceae family. Thanks to a recently improved gram scale production process, it was possible to prepare sufficient amounts of the GL glucoraphasatin (GRH) as well as the corresponding ITC 4-methylthio-3-butenyl isothiocyanate (GRH-ITC) from its sprouts. This paper reports a study on the cytotoxic and apoptotic activities of GRH-ITC compared with the oxidized counterpart 4-methylsulfinyl-3-butenyl isothiocyanate (GRE-ITC) on three human colon carcinoma cell lines (LoVo, HCT-116, and HT-29) together with a detailed kinetic investigation of the direct antioxidant/radical scavenging ability of GRH and GRH-ITC. Both GRH-ITC and GRE-ITC reduced cell proliferation in a dose-dependent manner and induced apoptosis in the three cancer cell lines. The compounds significantly (p < 0.05) increased Bax and decreased Bcl2 protein expression, as well as producing caspase-9 and PARP-1 cleavage after 3 days of exposure in the three cancer cell lines. GRH-ITC treatment was shown to have no toxicity with regard to normal human lymphocytes (−15 ± 5%) in comparison with SFN (complete growth inhibition). GRH and GRH-ITC were able to quench the 2,2-diphenyl-1-picrylhydrazyl radical, with second-order rate constants of 14.0 ± 2.8 and 43.1 ± 9.5 M−1 s−1, respectively (at 298 K in methanol), whereas the corresponding value measured here for the reference antioxidant α-tocopherol was 425 ± 40 M−1 s−1. GRH reacted with H2O2 and tert-butyl hydroperoxide in water (pH 7.4) at 37 °C, with rate constants of 1.9 ± 0.3 × 10−2 and 9.5 ± 0.3 × 10−4 M−1 s−1 (paralleling recently developed synthetic antioxidants) being quantitatively (>97%) converted to GRE. It is demonstrated that GRH-ITC has interesting antioxidant/radical scavenging properties, associated with a selective cytotoxic/apoptotic activity toward three human colon carcinoma cell lines, and very limited toxicity on normal human T-lymphocytes.