New HPLC Methods to Quantitate Terpenoid Aldehydes in Foliage of Cotton (Gossypium)

The cotton plant (Gossypium) produces protective terpenoid aldehydes in lysigenous pigment glands. These terpenoids include hemigossypolone, hemigossypolone-6-methyl ether, gossypol, gossypol-6-methyl ether, gossypol-6,6'- dimethyl ether, heliocides H1, H2, H3 and H4, and heliocides B1, B2, B3 and B4. Quantitation of these compounds is important to understand host-pest interactions. The column used in our previous HPLC/DAD method is no longer manufactured. Thus, three new methods were developed that use a Scientific Glass Engineering ProteCol-GP-C18 column maintained at 30 °C with gradients of acetonitrile:ethanol:isopropanol:N,N-dimethylformamide:methanol:ethyl acetate as the organic solvent and 0.1% H3PO4 as the aqueous solvent. Method one separates hemigossypolone, hemigossypolone-6- methyl ether, gossypol, gossypol-6-methyl ether, gossypol-6,6'-dimethyl ether and heliocides H1, H2, H3 and H4; method two separates these compounds as well as heliocides B1 and B4; method three allows resolution of heliocides B1, B2, B3 and B4 as well as the other terpenoids. The new methods provide superior separation of the methylated gossypol derivatives as well as the heliocides H1 to H4 and the heliocides B1, B2 and B3. The chromatogram is monitored at 272 nm, and spectra are stored over 240-600 nm. These methods allow cotton breeders to track the terpenoid aldehyde composition in plant tissues as they develop new cotton germplasm. The method has been used to establish for the first time the introgression from the wild cotton G. sturtianum into commercial Upland cotton G. hirsutum.