Characterization, Interaction and Chemoprevention of Potent Constituents From Grape Cell Culture

Grape and its cell culture extracts are rich in flavonoids and stilbenes that are biologically active. The objectives of this study were (1) to evaluate catalytic inhibition of grape (a Vitis hybrid 'Bailey Alicant A') cell culture extract and subfractions on human DNA topoisomerase II for cancer chemoprevention; (2) to characterize constituents in the most potent fractions; (3) to investigate interactions between bioactive polyphenols present in the most potent fractions; and finally (4) to assess cytotoxicity of the subfractions using both cancerous and non-cancerous cells lines. The most potent topoisomerase II catalytic inhibitors from grape cell culture extract were Toyopearl (TP) fractions 4 and 6 (IC 50 = 0.28--0.29 mug/mL), which were significantly more potent than crude extract (1.02 mug/mL) or resveratrol (18.0 mug/mL), a recognized chemopreventive agent), on mug/mL bases. Using both HPLC and LC-ESI/MS, constituents in TP-4 and TP-6 were characterized, many of which have not been previously characterized from grape cell cultures. Epicatechin gallate (IC 50 = 0.029 muM), myricetin (0.39 muM), procyanidin B2 (PB2, 4.5 muM), and resveratrol (65.7 muM) were potent topoisomerase II catalytic inhibitors present in TP-4 from grape cell culture. Synergistic interaction between myricetin and resveratrol (at a molar ratio of 1:70) on topoisomerase II catalytic inhibition was confirmed with isobolographic analysis. Likewise, potentiating interactions between PB2 and other polyphenolic constituents were also clearly demonstrated. Both TP-4 and TP-6 were effectively cytotoxic to L1210 mouse leukemia cells, whereas only TP-6 inhibited HepG2 human liver cancer cells. Both fractions did not show any cytotoxicity to non-cancerous PK15 pig kidney cells. After further subfractionation of TP-6 (comprised primarily of oligomenc procyanidins) using Sephadex LH-20, the cytotoxicity of bioactive subfractions to HepG2 human liver cancer cells was in a descending order of TP-6-5, TP-6-6 (similar potency to TP-6), and TP-6-4, suggesting that cytotoxicity of procyanidin mixtures may be proportional to the degree of polymerization. Due to selective cytotoxicity to the carcinoma cell lines tested, TP-6 and its potent subfractions (TP-6-5 and TP-6-6) show more potential as antihepatoma agents than either resveratrol or etoposide, warranting further investigation of their potential as cost-effective chemopreventive agents, particularly in antihepatoma therapy.

Name thesis:degree_name

Ph.D.

Level thesis:degree_level

Dissertation

Grantor

University of Illinois at Urbana-Champaign

Year dc:date

2015

Author dc:creator

• Jo, Jeong-Youn

Contributors dc:contributor

• Lila, Mary Ann

• Health Sciences, Nutrition

Language dc:language

eng

Identifier

(MiAaPQ)AAI3202110

OAI identifier oai:identifier

oai:www.ideals.illinois.edu:2142/87876