A method is described for the estimation of betaines and proline in plant extracts using 1H nuclear magnetic resonance. After partial purification of the crude extract on a simple cation-exchange column the technique enables the simultaneous determination of glycinebetaine, choline and other N-methylammonium-containing compounds as well as proline. The method is simple and the results obtained compare favourably with those determined by other methods.

Fifteen species of Melaleuca and two species of Callistemon from the field were examined to determine whether they accumulated nitrogen-containing compatible solutes and, if so, which. In addition to L-proline, N-methyl-L-proline (MP) (isolated for the first time from plants), trans-4-hydroxy-N-methyl- L-proline (MHP), and N, N'-dimethyl-trans-4-hydroxy-L-proline (DHP) were found in various combinations in the 15 Melaleuca species. M. lanceolata seedlings were subjected to water or salinity stress and M. uncinata to water stress under laboratory conditions. In both species significant reductions in leaf water potential (Ψw), osmotic potential (Ψs), turgor potential (Ψp), and relative water content (RWC) were observed in response to water stress. Salinised M. lanceolata plants showed considerable osmotic adjustment and maintained Ψp comparable to that of control plants; salinity, however, decreased RWC. In response to the imposed stresses under laboratory conditions, proline and MHP levels in M. lanceolata, and MHP and DHP levels in M. uncinata, increased. In addition to possible protective or osmotic roles in vivo, these proline analogues may be useful in chemotaxonomic investigations of Melaleuca species.

Australian native flora was examined with nuclear magnetic resonance (n.m.r.) techniques for its content of nitrogenous compatible solutes. Plants were sampled from four habitats: two arid, one subhumid, and one saline estuarine marsh. Eight and two of the 15 plants in the subhumid area accumulated proline and glycinebetaine, respectively, whereas many of the plants in the two arid habitats accumulated these solutes. With only two exceptions plants in the saline marsh could be described as either proline accumulators (six species) or glycinebetaine accumulators (eight species). Attempts to correlate the glycinebetaine and proline contents with the relative water content (RWC) were not successful. Some plants accumulate compounds other than, or in addition to, proline or glycinebetaine, such as trans- 4-hydroxy-N-methyl-L-proline, which was accumulated in Melaleuca lanceolata. Exocarpos aphyllus accumulated an as yet unidentified compound.

A rapid water stress, imposed by withholding water under a high transpirational demand or by the application of polyethylene glycol (PEG), resulted in the accumulation of glutamine, asparagine and valine in amounts equal to or greater than that of proline. However, progressive water stress imposition resulted in proline and glycinebetaine being the dominant accumulated compounds of those measured. PEG had an apparently specific effect on amino acid accumulation, particularly of the amide forms. With some exceptions, such as glycinebetaine, the stress-induced changes in the content of most solutes were reversible upon rewatering.

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