Ultrasound-assisted enzymatic hydrolysis for iodinated amino acid extraction from edible seaweed before reversed-phase high performance liquid chromatography-inductively coupled plasma-mass spectrometry.

The combination of reverse phase high performance liquid chromatography (RP-HPLC) with inductively coupled plasma mass spectrometry (ICP-MS) was used for the determination of monoiodotyrosine (MIT) and diiodotyrosine (DIT) in edible seaweed. A sample pre-treatment based on ultrasound assisted enzymatic hydrolysis was optimized for the extraction of these iodinated amino acids. Pancreatin was selected as the most adequate type of enzyme, and parameters affecting the extraction efficiency (pH, temperature, mass of enzyme and extraction time) were evaluated by univariate approaches. In addition, extractable inorganic iodine (iodide) was also quantified by anion exchange high performance liquid chromatography (AE-HPLC) coupled with ICP-MS. The proposed procedure offered limits of detection of 1.1 and 4.3ngg(-1) for MIT and DIT, respectively. Total iodine contents in seaweed, as well as total iodine in enzymatic digests were measured by ICP-MS after microwave assisted alkaline digestion with tetramethylamonium hydroxide (TMAH) for total iodine assessment, and also by treating the pancreatin extracts (extractable total iodine assessment). The optimized procedure was successfully applied to five different types of edible seaweed. The highest total iodine content, and also the highest iodide levels, was found in the brown seaweed Kombu (6646±45µgg(-1)). Regarding iodinated amino acids, Nori (a red seaweed) was by far the one with the highest amount of both species (42±3 and 0.41±0.024µgg(-1) for MIT and DIT, respectively). In general, MIT concentrations were much higher than the amounts of DIT, which suggests that iodine from iodinated proteins in seaweed is most likely bound in the form of MIT residues.

Circulating diiodotyrosine: studies of its serum concentration, source, and turnover using radioimmunoassay after immunoextraction.

This report describes the application of a sensitive, specific, and reproducible RIA for diiodotyrosine (DIT) in human serum and metabolic studies on the source and kinetics of circulating DIT. Interference by cross-reactivity of T4 and other analogs was completely eliminated by isolation of DIT from serum with an efficient preparative immunoprecipitation technique. Mean (+/- SD) serum DIT levels were 161 +/- 133 pmol/liter (7.0 ng/100 ml) in 41 normal subjects, 64 +/- 30 pmol/liter in 46 pregnant women, 241 +/- 83 pmol/liter in the cord serum of 48 newborn infants, 542 +/- 494 pmol/liter in 22 hyperthyroid patients, and 101 +/- 71 pmol/liter in 15 hypothyroid patients. Mean values in pregnant, newborn and hyperthyroid subjects were significantly different from the normal mean. Very low DIT serum levels were found in four athyreotic patients during oral T4 substitution therapy, indicating that little DIT is formed by peripheral T4 degradation. In five normal subjects who received a single oral dose of 3 mg T4, serum DIT remained unchanged in one case and decreased in four cases. Radioimmunological measurements of DIT elimination from serum after the iv injection of 1 mg DIT in two normal volunteers gave MCRs of 103 and 133 liters/day and an average extrathyroidal DIT turnover rate of 19 nmol/day (8.2 microgram/day). These data indicate that circulating DIT arises predominantly from the thyroid, suggesting that peripheral formation of DIT is a minor metabolic pathway in the human.