Determination of cysteine, homocysteine, cystine, and homocystine in biological fluids by HPLC using fluorosurfactant-capped gold nanoparticles as postcolumn colorimetric reagents.

We have demonstrated for the first time the suitability of fluorosurfactant-capped spherical gold nanoparticles as HPLC postcolumn colorimetric reagents for the direct assay of cysteine, homocysteine, cystine, and homocystine. The success of this work was based on the use of an on-line tris(2-carboxyethyl)phosphine reduction column for cystine and homocystine. Several parameters affecting the separation efficiency and the postcolumn colorimetric detection were thoroughly investigated. Under the optimized conditions, cysteine, homocysteine, cystine, and homocystine in human urine and plasma samples were determined. Detection limits for cysteine, homocysteine, cystine, and homocystine ranged from 0.16-0.49 µM. The accuracy in terms of recoveries ranged between 94.0-102.1%. This proposed method was rapid, inexpensive, and simple.

Cystathionine synthase in tissue culture derived from human skin: enzyme defect in homocystinuria.

Fibroblasts derived from normal human skin and from cells in amniotic fluid and grown in tissue culture have cystathionine synthase activity. Skin from homocystinuric patients gives rise to fibroblast lines with normal activities of methionine-activating enzyme, but with very low or undetectable cystathionine synthase activity. Thus, the enzyme lesion in homocystinuria is demonstrable in readily available human cells. Neither cystathionine synthase nor methionine-activating enzyme could be detected in intact normal skin.

Adaptation of a high-performance liquid chromatographic method for quantitative determination of homocysteine in urine.

A modification of the Bio-Rad total homocysteine HPLC-test is presented in order to enable not only plasma homocysteine measurements but also the quantification of homocysteine in urine samples using the same principle of measurement. Coelution of the internal standard provided in the test kit with an endogenous compound in urine demands for an alternative analytical procedure. Therefore, we introduced 3-mercaptopropionic acid as a substitute for the internal standard. The analytical method validation was performed for the matrix of urine specimens. The applicability of this method was demonstrated in a clinical study with volunteers after homocysteine thiolactone hydrochloride loading.

Neonatal urine screening for metabolic disease with auxotrophic strains of Bacillus subtilis.

A simplified method for neonatal urine screening for metabolic diseases is described. This procedure involves the use of two bacterial inhibition assays which can detect uracil, lysine, and homocystine, in combination with two other assays utilizing spores of amino acid auxotrophic mutants of Bacillus subtilis which can detect several amino acids and purines. These four tests have the capability to detect several treatable inherited metabolic diseases, including three disorders of the urea cycle, using a urine specimen on filter paper. This method results in savings of cost and time over chromatographic screening procedures, since only four agar trays (56 specimens per tray) are used, which can be processed with one cycle through a semi-automated punch index machine.

Homocysteine and cysteine loads in patients with homocystinuria due to cystathionine synthase deficiency: effects of vitamin B-6.

The metabolic response of patients with homocystinuria due to cystabhionine synthase deficiency to oral loads of homocysteine indicates: that even severely affected patients with homocystinuria have pools of cystine in their tissues; that control of sulfur amino acid metabolism favors increased concentrations of methionine rather than homocystine in the plasma; and that even patients who apparently are not B-6-responsive respond differently to the loads of homocysteine when challenged during B-6-treatment compared with their response before B-6 treatment. Loading tests with homocysteine indicate that B-6 treatment be of some benefit even in individuals who do not have an obvious biochemical response to such therapy.

Homocystinuria with congenital/developmental cataract.

The aim of the study is to screen patients for homocystinuria with and without cataract and analyse for homocystine and methionine. Fifty-eight samples from 29 patients, i.e., plasma and urine collected after overnight fasting were analysed by the screening test for homocystine, and paper chromatography for homocystine and methionine. Out of 29 homocystinuric patients, 24 had cataract. Only one had appreciable amounts of methionine in his serum. He also had mental retardation as expected and belongs to Type I. The other types did not have methionine but had only homocystine. There was no mental retardation or ectopia lentis. So they belonged to Types II, III or IV. As there is excess methionine in Type I, with low cystine, cataract may be due to deficiency of cysteine and reduced glutathione and might be averted by suitable therapy, i.e., high cystine-low methionine diet with B6. In other types with low methionine, cataract may be due to decreased availability of amino acids for the synthesis of lens proteins; the treatment of choice should be B12, and folate with methionine.