Determination of β-N-methylamino-l-alanine (BMAA) in plant (Cycas circinalis L.) and animal tissue by precolumn derivatization with 9-fluorenylmethyl chloroformate (FMOC) and reversed-phase high-performance liquid chromatography

A high-performance liquid chromatography (HPLC) method is described for determining subpicomole concentrations of β-N-methylamino-l-alanine (BMAA) in plant and animal tissue. BMAA and other amino acids were reacted with 9-fluorenylmethyl chloroformate (FMOC) for 10 min under alkaline conditions to form highly fluorescent and stable derivatives. All amino acids, including BMAA, eluted from the column within 22 min. BMAA (t_r = 18.02 ± 0.07 min) was detected in Cycas circinalis L. seed and in serum, cerebrospinal fluid and brain tissue from BMAA-treated monkeys and rats. The primary amino acids glutamine, glutamic acid, aspartic acid, alanine, glycine and γ-aminobutyric acid (GABA) could also be detected since they were well resolved from BMAA. These amino acids and BMAA were linear over the concentration range of 0.15-7.5 μM with a relative standard deviation ranging from 2.1-6.7%. This method should prove useful in studies to determine the role of BMAA in the Western Pacific amyotrophic lateral sclerosis/Parkinsonism-dementia complex for which cycad seed is the principal etiological candidate.