Isolation of oligomeric proanthocyanidins from flavonoid-producing cell cultures

The extraction, fractionation, and chromatographic separation of a series o f proanthocyanidin monomers and oligomers pere facilitated using a flavonoi d-rich cell culture of Vaccinium pahalae Skottsberg as the donor tissue. Th e cell cultures, after exposure to light, readily accumulated anthocyanin p igments and other flavonoids in relatively large amounts, with minimal conc urrent production of pectins, enzymes, and complex sugars produced in field -grown Vaccinium berries. The absence of these interfering compounds greatl y simplified the isolation and purification of proanthocyanidins and other phenolic compounds from cell cultures, primarily using vacuum chromatograph y. Subsequently, the structures and molecular weights of several individual compounds and the general composition of unresolved fractions were establi shed with H-1- and C-13-NMR and MS. The initial extract of V. pahalae cell cultures was readily fractionated on silica gel to yield a series of fracti ons containing proanthocyanidin B-2, a series of increasingly polar proanth ocyanidin oligomers ranging from dimers to heptamers largely based on (-)-e picatechin structures (some with A-type linkages), a mixture of E- and Z-p- coumaric acid, the corresponding 4-O-glucoside, and other compounds contain ing E- and Z-p-coumaric acid moieties. Cell culture extracts demonstrated b road antioxidant capacity and significant ability to inhibit tumor promotio n in vitro, as indicated in an ornithine decarboxylase assay.