Fractionation of gibberellins in plant extracts by reverse phase high performance liquid chromatography

In studies on endogenous plant gibberellins (GAs), reverse phase (Bondapak C(18)) high performance liquid chromatography (HPLC) has proved to be a useful method for the fractionation of plant extracts. The behavior of 18 authentic GAs in such a chromatographic system is described. The main factors determining chromatographic behavior are the degree and the position of hydroxylation of the GA. Generally, dihydroxylated GAs elute before monohydroxylated GAs, whereas 13-hydroxylated GAs elute before 3-hydroxylated GAs. The number of carboxyl groups and the degree of saturation of the A-ring have little effect. For 20-carbon GAs, the oxidation state at C-20 is only relevant insofar as GAs having a methyl group at this position elute later than those with other groups (lactone, aldehyde, or carboxyl).As an illustration of the use of reverse phase HPLC, the endogenous GAs of immature seeds of Pharbitis nil L., strain "Violet," were reinvestigated. The presence of gibberellins A(3), A(5), A(17), A(20), and A(29) was confirmed by gas-liquid chromatography-mass spectrometry. In addition, two other GAs, A(19) and A(44), were also identified in extracts of this material.