Chromatography of acid phytohormones on columns of Sephadex LH-20 and insoluble poly-N-vinylpyrrolidone, and application to the analysis of conifer extracts

The chromatographic behaviour of abscisic acid (ABA), indole-3-acetic acid (IAA), phenylacetic acid (PAA), and gibberellins A₁, A₄, A₈, A₉, A₁₃ and A₂₀ on columns of Sephadex LH-20 and insoluble poly- N -vinylpyrrolidone (PVP) eluted with buffers of different pH values is described. PVP shows considerable batch differences that must be carefully checked. Chromatography of acidic ethyl acetate-soluble fractions of Scots pine ( Pinus sylvestris L.) extracts at pH 4.5 resulted in great losses of phytohormones, due to poor solubility of the extracts. If the extracts were applied to the column dissolved in buffer of pH 7.5, subsequent elution at pH 4.5 was possible with only small losses. The performance of the chromatographic column was strongly affected by the application volume. A combined PVP/Sephadex LH-20 column eluted at pH 4.5 allows remarkable purification of pine and spruce ( Picea abies (L.) Karst.) extracts, collection of IAA in a fraction that can be directly analyzed by e.g. the indolo-α-pyrone method, and collection of another fraction containing ABA, PAA and probably most of the known C₁₉-gibberellins; whereas the C₂₀-gibberellin A₁₃ is eluted later (with IAA).