In these studies, the altered urinary biomarkers included dT, dU, and dX, N1-acetylspermidine, N- acetylglucosamine/galactosamine-6-sulfate, N-acetyltaurine, N-hexanoylglycine, taurine, and
isethionic acid.
The water-soluble cue was subsequently isolated and characterized as a complex between the sugar floridoside and isethionic acid (F-I complex; Williamson et al., 2000).
During further research on this system, we obtained inductive fractions that contained isethionic acid but not floridoside, and we were also unable to reproduce a synthetic F-I complex that induced settlement of larval H.
Peak 1 displayed the pattern of isethionic acid (Barrow et al., 1993), as determined by [.sup.1]H- and [.sup.13]C-NMR spectroscopy, as well as some additional signals that were not characteristic of floridoside (see next section).
Isethionic acid and taurine were the major compounds in peak 1, as determined by [.sup.1]H- and [.sup.13]C-NMR spectroscopy and comparison with synthetic samples.
[.sup.13]C-NMR spectroscopy analysis of this peak showed only [.sup.13]C-signals for floridoside and isethionic acid (Williamson et al., 2000).
Isethionic acid was prepared by passing a solution of sodium isethionate (Aldrich) in MilliQ water through a column of AG50WX2 ([H.sup.+] form) ion-exchange resin.
All compounds--[1], the synthetic floridoside-isethionic acid complex, floridoside [2], and isethionic acid--were made up in 10 mg [ml.sup.-1] stock solutions in MilliQ water, and aliquots were added to treatment dishes to give a final concentration in the bioassay of 200 [micro]M.
The [sup.1]H and [sup.13]C NMR data for [1] corresponded to previously published data for floridoside [[Alpha]-D-galactopyranosyl-(1-2.)-glycerol] (Karsten et al., 1993) and isethionic acid (Barrow et al., 1993) in 1:1 integral ratios (Fig.
No metamorphosis was observed in treatments containing floridoside or isethionic acid, or in the controls.
Through bioassay-guided fractionation, the cue for metamorphosis was demonstrated to be a noncovalently-bound complex of the sugar floridoside and isethionic acid.