| Summary: | The diterpenoid taxol, a compound isolated from the bark of the Pacific Yew tree,
Taxus brevefolia, has shown remarkable success in the treatment of several types of
cancer. Limited supply of taxol, due to low concentrations produced naturally and
difficulty in synthesis of the drug, have spurred research attempts to develop alternative
methods of producing taxane compounds in vitro using suspension cell cultures.
Suspension cell culture propagated from the stems of Taxus media were grown in
airlift type bioreactors for 28 days using a modified B5 medium. The cells and broth from
multiple bioreactor experiments were combined and extracted with ethyl acetate and
methanol. The ethyl acetate extracts were roughly separated by partitioning with solvents
before high pressure liquid chromatography (HPLC) analysis was performed to determine
if taxol, baccatin III, 10 deacetylbaccatin, 14-hydroxy-10-deacetylbaccatin, 14-
hydroxybaccatin, cephlomannine, 7-epitaxol or 7-methyltaxol were present.
Additionally, the partitioned extract was checked agains the taxane standards by thin
layer chromatography (TLC). There was no indication that the taxane compounds used
as standards were present in the cell extract above the detectable limits for these methods.
To determine what types of compounds the cell culture was producing, the cell
extract was separated using column chromatography, preparative TLC and
recrystallization. Five compounds were purified and identified as 3-β-0[P- β -glucoside]
β -sitosterol, 3B-6B, 23-trihydroxyolean-28-oic acid, cyclo (L-proline-L- valine), by
comparing mass spectral (MS) and nuclear magnetic resonance (NMR) spectroscopy
with that published in the literature. Two novel compounds 9,10 epoxy, 11-hydroxy
12,13 linoleic acid and 4,6- dihydroxy,-2-methoxy, 8-methyl xanthone were found in
cell culture. They were identified using NMR and MS data and by comparison with
compounds of like structure in the literature.
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