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1.
J Sep Sci ; 37(1-2): 30-6, 2014 Jan.
Article in English | MEDLINE | ID: mdl-24302617

ABSTRACT

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.


Subject(s)
Chromatography, High Pressure Liquid/methods , Cysteine/analysis , Cystine/analysis , Homocysteine/analysis , Homocystine/analysis , Chromatography, High Pressure Liquid/instrumentation , Cysteine/blood , Cysteine/urine , Cystine/blood , Gold/chemistry , Homocysteine/blood , Homocysteine/urine , Homocystine/blood , Homocystine/urine , Humans , Nanoparticles/chemistry
2.
Anal Chem ; 79(2): 666-72, 2007 Jan 15.
Article in English | MEDLINE | ID: mdl-17222035

ABSTRACT

Gold nanoparticles (GNPs) capped with nonionic fluorosurfactant molecules (Zonyl FSN) were synthesized, and with the colloidal solution as a probe reagent, a new postcolumn colorimetric detection method for HPLC assay of homocysteine (Hcy) has been developed. The FSN-capped GNPs exhibited excellent stability in aqueous solutions, even in the presence of high salt. The aggregation of the GNPs could be induced by either Hcy or cysteine, resulting in an absorption decrease of the colloidal solution at 525 nm and an absorption increase at longer wavelengths (600-700 nm); however, the GNPs did not respond to other amino acids and biomolecules such as glutathione, cysteinylglycine, and glucose. Under optimal conditions (i.e., high salt, neutral pH, and approximately 70 degrees C), the color change of the GNP solution could almost complete ( approximately 90%) within approximately 30 s upon the addition of Hcy. The high selectivity and very fast kinetics of the reaction make it a promising system for HPLC postcolumn detection. The new technique has been employed to determine total Hcy levels in human urine and plasma samples, and the results are satisfactory.


Subject(s)
Chromatography, High Pressure Liquid/methods , Gold/chemistry , Homocystine/analysis , Nanoparticles/chemistry , Surface-Active Agents/chemistry , Amino Acids/analysis , Homocystine/blood , Homocystine/urine , Humans , Organic Chemicals/chemistry
3.
Anal Bioanal Chem ; 385(5): 855-60, 2006 Jul.
Article in English | MEDLINE | ID: mdl-16791565

ABSTRACT

A new analytical method is proposed for simultaneous determination, by liquid chromatography, of the three main urinary thiols-cysteine, cysteinylglycine, and homocysteine. To measure the total amount of these thiols urine is reduced with sodium borohydride, to convert disulfides to thiols which are then derivatized with 2-chloro-1-methylquinolinium tetrafluoroborate. Separation and quantitation of the 2-S-quinolinium thiol derivatives formed were achieved by high-performance liquid chromatography with detection at 355 nm. Validation showed the method enabled reliable simultaneous determination of these aminothiols in urine. The calibration graphs for each analyte, obtained by use of normal urine spiked with increasing amounts of cysteine, cysteinylglycine, and homocysteine, were linear (R2 > or = 0.997) over the range covering most practical situations. The recovery of the assay was 98-100% and sensitivity was 0.12-0.25 micromol L(-1). The method was applied to 91 different samples of normal urine to establish reference values for the aminothiols, normalized on creatinine.


Subject(s)
Chromatography, High Pressure Liquid/methods , Cysteine/urine , Dipeptides/urine , Homocystine/urine , Creatine/urine , Cysteine/chemistry , Dipeptides/chemistry , Homocystine/chemistry , Humans , Hydrogen-Ion Concentration , Molecular Structure , Sensitivity and Specificity
4.
J Chromatogr B Analyt Technol Biomed Life Sci ; 828(1-2): 122-5, 2005 Dec 15.
Article in English | MEDLINE | ID: mdl-16203185

ABSTRACT

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.


Subject(s)
Chromatography, High Pressure Liquid/methods , Homocystine/urine , 3-Mercaptopropionic Acid/chemistry , Humans , Male , Reproducibility of Results
5.
Dtsch Med Wochenschr ; 130(43): 2439-43, 2005 Oct 28.
Article in German | MEDLINE | ID: mdl-16240242

ABSTRACT

HISTORY: Pronounced osteoporosis was discovered in a 44-year-old man presenting with an herniated intervertebral disk. He reported severe myopia, incomplete dislocation of the lenses and retinal detachment. He did not know of any thrombembolic events in the past. On physical examination Marfan-like appearance and a funnel chest were noted. Because of these findings Marfan syndrome was suspected. INVESTIGATIONS: Considering the findings of the x-rays, homocystinuria was suspected as a cause of osteoporosis, despite apparently normal cognitive functions. This diagnosis was confirmed by greatly increased values of serum homocysteine and a positive test for urinary homocystine. Since methionine and homocystine were both elevated, the diagnosis of cystathionin-beta-synthase deficiency was established. DIAGNOSIS, TREATMENT AND COURSE: After taking folate and vitamin B6, the homocysteine level decreased moderately. Betaine and subsequently vitamin B12 were added. Homocysteine values declined markedly on this therapeutic regimen. Two years later the patient was admitted again because of atypical angina. Coronary heart disease could be excluded by coronary angiography. CONCLUSION: Diagnosis of premature osteoporosis should prompt consideration of homocystinuria even in adults. Premature arteriosclerosis, thrombembolic diseases, dislocation of the lens and retinal detachment may give further clues.


Subject(s)
Homocystinuria/diagnosis , Osteoporosis/etiology , Adult , Age Factors , Angina Pectoris/etiology , Betaine/therapeutic use , Diagnosis, Differential , Drug Therapy, Combination , Folic Acid/therapeutic use , Homocysteine/blood , Homocystine/urine , Homocystinuria/complications , Homocystinuria/drug therapy , Humans , Intervertebral Disc Displacement/complications , Lens Subluxation/etiology , Male , Marfan Syndrome/diagnosis , Myopia/etiology , Retinal Detachment/etiology , Vitamin B 12/therapeutic use , Vitamin B 6/therapeutic use , Vitamin B Complex/therapeutic use
7.
Indian J Pediatr ; 67(10): 725-8, 2000 Oct.
Article in English | MEDLINE | ID: mdl-11105422

ABSTRACT

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.


Subject(s)
Cataract/etiology , Homocystinuria/complications , Adult , Cataract/congenital , Cataract/epidemiology , Child , Chromatography, Paper , Female , Homocystine/blood , Homocystine/urine , Homocystinuria/classification , Homocystinuria/metabolism , Homocystinuria/therapy , Humans , Male , Mass Screening , Metabolism, Inborn Errors/diagnosis , Methionine/blood , Pyridoxine/therapeutic use
8.
Biochem Int ; 12(1): 61-9, 1986 Jan.
Article in English | MEDLINE | ID: mdl-2868723

ABSTRACT

Exposure of sheep to 36% nitrous oxide for 8 days (2-hr per day) led to 90%, 82% and 74% inhibition of 5-methyltetrahydrofolate-homocysteine methyltransferase in the liver, heart and brain, respectively, while there was no significant decrease in the activity of methylmalonyl-CoA mutase. There was also no change of betaine-homocysteine methyltransferase activity. The level of plasma methionine in nitrous-oxide-exposed sheep fell to 30% of its initial value. S-Adenosylmethionine level was reduced to 50% of the control value in the liver, and was also significantly decreased in the heart, but not in the brain. Excretion of formiminoglutamic acid and homocystine was also observed in the urine of sheep exposed to nitrous oxide. These results demonstrate that inhibition of 5-methyltetrahydrofolate-homocysteine methyltransferase causes a pronounced perturbation of methionine metabolism in sheep, suggesting that dietary methionine plus methionine synthesized from the methyl groups of betaine are not sufficient to meet the methyl needs for biological methylation reactions in this species and, in turn, emphasizing the role of 5-methyltetrahydrofolate-homocysteine methyltransferase in methionine synthesis in the sheep.


Subject(s)
Brain/enzymology , Liver/enzymology , Methionine/metabolism , Myocardium/enzymology , Nitrous Oxide/pharmacology , Vitamin B 12/metabolism , 5-Methyltetrahydrofolate-Homocysteine S-Methyltransferase/metabolism , Animals , Betaine-Homocysteine S-Methyltransferase , Formiminoglutamic Acid/urine , Homocystine/urine , Male , Methionine/blood , Methylmalonyl-CoA Mutase/metabolism , Methyltransferases/metabolism , Orchiectomy , S-Adenosylmethionine/metabolism , Sheep
12.
Can Anaesth Soc J ; 29(3): 260-3, 1982 May.
Article in English | MEDLINE | ID: mdl-7074405

ABSTRACT

Research into the biotransformation of inhaled general anaesthetic agents, including nitrous oxide, has led to a better understanding of the underlying mechanisms. It is now known that nitrous oxide can react chemically with vitamin B12, oxidizing Cob(I)alamin to the inactive Cob(III) alamin form. Clinical and experimental evidence in mammals has confirmed that nitrous oxide toxicity, with symptoms suggestive of clinical vitamin B12 deficiency, occurs on exposure to nitrous oxide in a way which is dose and time related and reversible on withdrawal of the nitrous oxide. Nitrous oxide depresses the two known vitamin B12 dependent enzymes methylmalonyl CoA mutase and methionine synthetase by inactivation of their coenzymes adenosylcobalamin and methylcobalamin respectively. Methionine synthetase catalyses the conversion of homocystine to methionine, so interference with this reaction should cause methionine to be depleted and homocysteine to accumulate and to be excreted in the urine. We postulated that the detection of homocystinuria would therefore be an early indicator of nitrous oxide toxicity. Accordingly, we tested the first urine voided postoperatively of 41 patients undergoing nitrous oxide anaesthesia (17 neonates exposed to 50-66 per cent nitrous oxide for a mean of 3.0 hr, and 24 older patients exposed to 66 per cent nitrous oxide for a mean of 7.2 hr). None of these patients demonstrated homocystinuria.


Subject(s)
Anesthesia , Homocystine/urine , Nitrous Oxide , Adolescent , Adult , Child , Child, Preschool , Humans , Infant , Male , Nitrous Oxide/adverse effects , Vitamin B 12/metabolism
13.
Clin Chim Acta ; 119(3): 345-9, 1982 Mar 12.
Article in English | MEDLINE | ID: mdl-6802522

ABSTRACT

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.


Subject(s)
Bacillus subtilis/genetics , Metabolism, Inborn Errors/urine , Biological Assay/methods , Homocystine/urine , Humans , Infant, Newborn , Lysine/urine , Metabolism, Inborn Errors/diagnosis , Mutation , Uracil/urine
14.
Eur J Biochem ; 104(2): 419-23, 1980 Mar.
Article in English | MEDLINE | ID: mdl-7363899

ABSTRACT

Exposure of rats to a 50% N2O/oxygen mixture led to a rapid loss of methionine synthase activity in both liver and brain. This enzyme has vitamin B12 as a cofactor. There was impaired conversion of deoxyuridine to deoxythymidine by bone marrow cells and this defect followed loss of methionine synthase activity. There was no homocystinuria. Withdrawal of N2O was followed by a relatively slow recovery of methionine synthase activity over four days. The inactivation of vitamin B12 by N2O promises to be a valuable tool in the study of vitamin B12 metabolism.


Subject(s)
5-Methyltetrahydrofolate-Homocysteine S-Methyltransferase/antagonists & inhibitors , Brain/enzymology , Liver/enzymology , Methyltransferases/antagonists & inhibitors , Nitrous Oxide/pharmacology , Animals , Bone Marrow/metabolism , Brain/drug effects , DNA Replication/drug effects , Deoxyuridine/pharmacology , Homocystine/urine , Liver/drug effects , Rats , Thymidine/metabolism
15.
Clin Chim Acta ; 79(1): 197-210, 1977 Aug 15.
Article in English | MEDLINE | ID: mdl-890952

ABSTRACT

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.


Subject(s)
Cystathionine beta-Synthase/deficiency , Cysteine , Homocysteine , Homocystinuria/metabolism , Hydro-Lyases/deficiency , Pyridoxine/pharmacology , Cysteine/metabolism , Cystine/blood , Cystinuria , Disulfides/blood , Disulfides/urine , Female , Homocysteine/metabolism , Homocystine/blood , Homocystine/urine , Humans , Male , Pyridoxine/therapeutic use , Time Factors
17.
N Engl J Med ; 292(10): 491-6, 1975 Mar 06.
Article in English | MEDLINE | ID: mdl-1117892

ABSTRACT

Homocystinuria and homocystinemia without hypermthioninemia, but with reccurent episodes of folate responseive schizophrenic-like behavior, was documented in a mildly retarded adolescent girl who lacked the habitus associated with cystathionine synthase deficiency. Enzymes involved in homocysteine-methionine metabolism were demonstrated to be normal. A defect in the ability to reducte N-5-10--methylenetetrahydrofolate to 5-methyltetrahydrofolate was demonstrated. Methylenetetrahydrofolate reductase was 18 per cent of control values. Methyltetrahydrofolate is used for the methylation of homocysteine to methionine, and a deficiency of this compound could explain the homocystinemia and homocystinuria.


Subject(s)
Alcohol Oxidoreductases/deficiency , Folic Acid/therapeutic use , Homocystinuria/enzymology , Schizophrenia/enzymology , Adolescent , Diagnosis, Differential , Female , Folic Acid/blood , Folic Acid/metabolism , Homocysteine/metabolism , Homocystine/blood , Homocystine/urine , Homocystinuria/diagnosis , Homocystinuria/drug therapy , Humans , Methionine/blood , Methionine/metabolism , Methylation , Schizophrenia/diagnosis , Schizophrenia/etiology , Tetrahydrofolates , Vitamin B 12/blood
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