Pico‐tag analysis of arogenic acid and related free amino acids from plant and fungal extracts

Abstract

Free amino acids from various plant tissues and fungal mycelia were extracted with buffered ethanol (75%, v/v) in 0.5% 2-amino-2-methyl-1-propanol-hydrochloride pH 11.0. Following removal of proteins and other high molecular weight components by ultrafiltration, free amino acids in the extracts were dried, derivatized with phenylisothiocyanate and analysed by high performance liquid chromatography using a Waters Pico-Tag column. Arogenic acid, a non-aromatic amino acid precursor of phenylalanine and tyrosine, was not detected although both aromatic amino acids were quantitated in all plant and fungal extracts. Nevertheless, arogenic acid in the mycelium of a triply-blocked mutant of Neurospora crassu was readily detected. It is proposed that the absence of arogenic acid in plant extracts suggests that this intermediate does not accumulate in the metabolic pool thereby negating a possible regulatory role during aromatic amino acid biosynthesis.

G-Protein-Coupled Receptor 1, G-Protein Gα-Subunit 1, and Prephenate Dehydratase 1 Are Required for Blue Light-Induced Production of Phenylalanine in Etiolated Arabidopsis

“…Examination of the Arabidopsis genome reveals no copies of arogenate dehydratase or cyclohexadienyl dehydratase. Further, Razal et al (1994) were unable to isolate or detect arogenate from any plant grown under any light condition.…”

“…Amino acid analysis was conducted as described (Razal et al, 1994) with these modifications: Amino acid content was assessed in the aerial portions of wild-type Col and insertion mutant seedlings (pd1-1 and pd1-2) 2 h after BL irradiation treatment or mock treatment (no BL). Seedlings were grown and irradiated as described above, aerial portions harvested in liquid nitrogen, ground to a fine powder, and stored at 280°C until amino acid analysis could be performed.…”

“…It was proposed from these data that production of Phe only proceeds via an arogenate intermediate. Subsequent investigations have failed to confirm the presence of arogenate directly from plants in either light-or dark-grown seedlings (Razal et al, 1994).…”

“…Despite the fact that 30% to 40% of the mass of an individual plant can be derived from aromatic amino acids L-Phe and L-Tyr, little is known about their biosynthetic pathway in plants ( Lewis and Yamamoto, 1989;Davin et al, 1992;Razal et al, 1994Razal et al, , 1996. In light-grown seedlings, the Shikimate pathway links carbon assimilated via photosynthesis to the biosynthesis of aromatic amino acids, through the formation of chorismate.…”

“…To our knowledge, there are no such studies in etiolated seedlings. The Shikimate metabolic pathway does appear to function, albeit at low levels, in etiolated seedlings (Razal et al, 1994;Hemm et al, 2004). Jung et al (1986) reported the presence of arogenate in the chloroplasts of light-grown plant cells in culture.…”