Development of new method for simultaneous quantitative determination of five opines found in marine invertebrates

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Many groups of marine invertebrates produce unique imino acids so called 'opine'. Opines are considered as their metabolites of anaerobic glycolysis, and include meso- alanopine, D-strombine, D-tauropine, β-alanopine, and D-octopine. At present, several methods for quantitative determination of opines are available. However, there is no method capable of determining the five opines simultaneously. Such the situation makes the study of opine somewhat troublesome. Thus we attempted the development of a method determining the five opines simultaneously.Interfering primary amino acids were removed by o-phthalaldehyde (OPA) derivatization and the following fractionation using a Sep-Pak Plus PS-1 cartridge. This pre-treatment made possible to determine the five opines simultaneously by a cation-exchange HPLC with post-column NaClO/OPA detection. The sensitivity of this method is in the range 20 pmol - 1.0 nmol. The applicability of this method to biological samples was confirmed by analyzing the levels of opines in several marine animals.