Fluorine-Tagged 5-Hydroxytryptophan to Investigate Amino Acid Metabolism In Vivo

Auxin a plant growth hormone, has a metabolic pathway that includes molecules and enzymes like those in animal brains. In this study, tomato plant seedlings (Lycopersicon esculenta) were used to investigate the fate of fluorine-tagged 5-hydroxytryptophan (PF-5-HTP) being developed for fluorine spectroscopy and imaging. Seedlings were treated with high or low concentrations of 5-HTP or PF-5-HTP and compared with controls. Metabolites of the PF-5-HTP were quantified using a custom immunoassay for the tag. Serotonin (5-HT) levels were measured with spectrofluorometry and thin-layer chromatography. Plants in treatment conditions had serotonin levels five to six times higher than controls. PF-5-HTP served as a precursor for serotonin in a biosynthetic pathway in this plant model, providing evidence for the bioavailability of the novel molecule. The increase in serotonin in plants grown in media culture supplemented with 5-HTP or PF-5-HTP might have useful applications in pharmacology. 1. Introduction The use of plants as models in pharmacology research could reduce the use of live animals. This approach accords well with Congressional intent and guidelines from the Interagency Coordinating Committee on the Validation of Alternative Methods (ICCVAMs), a permanent committee representing 15 federal regulatory and research agencies that use, generate, and disseminate toxicology testing information. The method described here can be useful in both neuroscience and botany. In neuroscience, a host of disorders have been linked to the neurotransmitter serotonin. Investigating the serotonin (5-HT) pathways in the brain would be greatly facilitated by having a multiple-fluorine atom tagged version for magnetic resonance spectroscopy (MRS) and functional imaging (fMRI). As serotonin cannot cross the blood brain barrier, it is necessary to produce a tagged version of the precursor that is bioavailable in vivo. One of the metabolic pathways for the biosynthesis of indole-3-acetic acid (auxin) from tryptophan uses the intermediate tryptamine which is structurally like 5-hydroxytryptamine (5-HT). Indole-3-acetic acid (auxin) is an essential hormone for plant growth and development. We used the auxin metabolic pathway as a model for investigating the fate of a novel molecule created for use in brain imaging. Toxicologists are interested in auxin for another reason. Having an accurate method for measuring tryptophan derivatives in plant tissues is important because of the role these compounds play in ecosystems and their sensitivity to environmental stress. The complex