Analysis of the amino acid indospicine in biological samples by high performance liquid chromatography

Abstract

Indospicine is a hepatotoxic amino acid that accumulates in the meat of horses that consume the legume Indigofera linnaei. A method to determine indospicine concentration in biological samples using an amino acid analyser has been reported, but the analysis time is long and therefore not suited to the analysis of large numbers of samples. A rapid and reliable method was developed for the analysis of indospicine in horsemeat and serum using High Performance Liquid Chromatography. Horsemeat and serum were extracted with either water or 0.01 N hydrochloric acid, respectively, and deproteinized by ultrafiltration. Precolumn derivatization of samples with phenylisothiocyanate was followed by separation of indospicine from other amino acids on a Pico-Tag C 18 column and UV detection at 254 nm. The calibration curves for indospicine in horsemeat extract were linear over the concentration range 0.4 microg ml(-1) to 20 microg ml(-1), while for indospicine in serum, the linear range was from 0.17 microg ml(-1) to 16.67 microg ml(-1). The mean recovery of indospicine in horsemeat extract was 87.2 +/- 6.8% and in serum was 97.3 +/- 9.9%. Analysis time for indospicine in horsemeat samples was 31 min and in serum samples was 36 min.

Hepatotoxicosis in dogs consuming a diet of camel meat contaminated with indospicine

“…An additional 15 dogs (dogs [5][6][7][8][9][10][11][12][13][14][15][16][17][18][19] were identified from clinical records as having also eaten the diet and their serum biochemistry (ALP, ALT, cholesterol and albumin) was determined. Dogs 5,7,8,11,12,15 and 19 had ceased the diet between 4 and 13 days before blood samples were collected ( Table 1). All other dogs were withdrawn from the diet on the day blood samples were taken.…”

The Occurrence and Toxicity of Indospicine to Grazing Animals

“…Indospicine is an unusual amino acid in that it is not incorporated into protein but exists in both plant and animal tissues as the free amino acid [42]. It is seemingly not readily metabolized by mammalian systems and accumulates as persistent residues in all tissues of animals consuming a diet containing the amino acid.…”

“…It is seemingly not readily metabolized by mammalian systems and accumulates as persistent residues in all tissues of animals consuming a diet containing the amino acid. Indospicine has been shown to accumulate in the tissues of horses [1,42,43], cattle [44] and goats [45] consuming I. linnaei, and in rabbits fed I. spicata seed [12]. The persistence of these residues has been demonstrated in experimental studies and tissue indospicine levels were still detectable several months after the cessation of feeding Indigofera plant material in horses [43], cattle [44], goats [45] and rabbits [12].…”

Towards a platform for the metabonomic profiling of different strains of Drosophila melanogaster using liquid chromatography–Fourier transform mass spectrometry

“…There is also a peak typical of CS which is absent from OR, which corresponds in elemental composition to indospicine. This compound occurs in nature as a hepatotoxic amino acid in the legume Indigofera linnaei and is an inhibitor of nitric oxide production from arginine [19]. Obtaining quantities of a standard would be necessary to make a definitive identification of this compound, but it is of interest that it relates to the same area of metabolism which appears to vary between CS and OR.…”