High-performance liquid chromatographic separation with electrochemical detection of amino acids focusing on neurochemical application

Canevari, Laura; Vieira, Raúl; Aldegunde, M.; Dagani, F.

doi:10.1016/0003-2697(92)90590-4

Cited by 31 publications

(10 citation statements)

References 36 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The concentration of cystathionine was determined following o-phthaldialdehyde derivatization, essentially as described previously (30,31). Briefly, the enzymatic reaction was stopped by the addition of an equal volume of 10% trichloroacetic acid, and the precipitated protein was removed by centrifugation.…”

Section: Enzyme Activity Assaysmentioning

confidence: 99%

H2S Biogenesis by Human Cystathionine γ-Lyase Leads to the Novel Sulfur Metabolites Lanthionine and Homolanthionine and Is Responsive to the Grade of Hyperhomocysteinemia

Chiku

Padovani

Zhu

et al. 2009

Journal of Biological Chemistry

437

424

View full text Add to dashboard Cite

Although there is a growing recognition of the significance of hydrogen sulfide (H 2 S) as a biological signaling molecule involved in vascular and nervous system functions, its biogenesis and regulation are poorly understood. It is widely assumed that desulfhydration of cysteine is the major source of H 2 S in mammals and is catalyzed by the transsulfuration pathway enzymes, cystathionine ␤-synthase and cystathionine ␥-lyase (CSE). In this study, we demonstrate that the profligacy of human CSE results in a variety of reactions that generate H 2 S from cysteine and homocysteine. The ␥-replacement reaction, which condenses two molecules of homocysteine, yields H 2 S and a novel biomarker, homolanthionine, which has been reported in urine of homocystinuric patients, whereas a ␤-replacement reaction, which condenses two molecules of cysteine, generates lanthionine. Kinetic simulations at physiologically relevant concentrations of cysteine and homocysteine, reveal that the ␣,␤-elimination of cysteine accounts for ϳ70% of H 2 S generation. However, the relative importance of homocysteinederived H 2 S increases progressively with the grade of hyperhomocysteinemia, and under conditions of severely elevated homocysteine (200 M), the ␣,␥-elimination and ␥-replacement reactions of homocysteine together are predicted to account for ϳ90% of H 2 S generation by CSE. Excessive H 2 S production in hyperhomocysteinemia may contribute to the associated cardiovascular pathology.H 2 S is the newest member of a growing list of gaseous signaling molecules that modulate physiological functions (1-3). Concentrations of H 2 S ranging from 50 to 160 M have been reported in the brain (4), where it appears to function as a neuromodulator by potentiating the activity of the N-methyl-Daspartate receptor and by altering induction of long term potentiation in the hippocampus, important for memory and learning (5). H 2 S levels in human plasma are reported to be ϳ50 M, and in vitro studies suggest that it functions as a vasodilator by opening K ATP channels in vascular smooth muscle cells (6).A recent in vivo study has demonstrated the efficacy of H 2 S in attenuating myocardial ischemia-reperfusion injury by protecting mitochondrial function (7). The role of H 2 S in inflammation is suggested by several studies (8 -11); however, the underlying mechanism is unknown. Remarkably, H 2 S can also induce a state of suspended animation in mice by decreasing the metabolic rate and the core body temperature presumably by inhibiting cytochrome c oxidase in the respiratory chain (12).Endogenous H 2 S is presumed to be generated primarily by desulfhydration of cysteine catalyzed by the two pyridoxal phosphate (PLP) 3 -dependent enzymes in the transsulfuration pathway: cystathionine ␤-synthase (CBS) and cystathionine ␥-lyase (CSE) (13,14). In fact, it is widely assumed, based on the reported absences of CSE in the brain (15) and of H 2 S in the brain of CBS knock-out mice (16), that CBS is the primary source of H 2 S in this organ, whereas CSE plays the...

show abstract

Section: Enzyme Activity Assaysmentioning

confidence: 99%

H2S Biogenesis by Human Cystathionine γ-Lyase Leads to the Novel Sulfur Metabolites Lanthionine and Homolanthionine and Is Responsive to the Grade of Hyperhomocysteinemia

Chiku

Padovani

Zhu

et al. 2009

Journal of Biological Chemistry

437

424

View full text Add to dashboard Cite

show abstract

“…Taurine concentrations were determined following o-phthaldialdehyde (OPA) derivatization, essentially as previously described (Canevari et al 1992;Alvarez et al 2006). SH-SY5Y cells were washed with 2 · 2 mL of isotonic buffer A and then lysed in 2 mL of ice-cold 6% TCA.…”

Section: Determination Of Taurine Massmentioning

confidence: 99%

Muscarinic receptor regulation of osmosensitive taurine transport in human SH‐SY5Y neuroblastoma cells

Foster

Vitvitsky

Banerjee

et al. 2008

Journal of Neurochemistry

View full text Add to dashboard Cite

The ability of G-protein-coupled receptors to regulate osmosensitive uptake of the organic osmolyte, taurine, into human SH-SY5Y neuroblastoma cells has been examined. When monitored under isotonic conditions and in the presence of physiologically relevant taurine concentrations (1-100 µM), taurine influx was mediated exclusively by a Na + -dependent, highaffinity (K m =2.5 µM) saturable transport mechanism (V max =0.087 nmol/mg protein/min). Reductions in osmolarity of >20% (attained under conditions of a constant NaCl concentration) resulted in an inhibition of taurine influx (>30%) that could be attributed to a reduction in V max , whereas the K m for uptake remained unchanged. Inclusion of the muscarinic cholinergic agonist, oxotremorine-M (Oxo-M), also resulted in an attenuation of taurine influx (EC 50~0 .7 µM). Although Oxo-M-mediated inhibition of taurine uptake could be observed under isotonic conditions (~25-30%), the magnitude of inhibition was significantly enhanced by hypotonicity (~55-60%), a result that also reflected a reduction in the V max , but not the K m , for taurine transport. Oxo-M-mediated inhibition of taurine uptake was dependent upon the availability of extracellular Ca 2+ but was independent of protein kinase C activity. In addition to Oxo-M, inclusion of either thrombin or sphingosine 1-phosphate also attenuated volume-dependent taurine uptake. The ability of Oxo-M to inhibit the influx of taurine was attenuated by 4-[(2-butyl-6,7-dichloro-2-cyclopentyl-2,3-dihydro-1-oxo-1H-inden-5-yl)oxy]butanoic acid (DCPIB), an inhibitor of the volume-sensitive organic osmolyte and anion channel. DCPIB also prevented receptormediated changes in the efflux and influx of K + under hypoosmotic conditions. The results suggest that muscarinic receptor activation can regulate both the volume-dependent efflux and uptake of taurine and that these events may be functionally coupled.

show abstract

“…However, the volume of sap obtained from a single Arabidopsis plant by this method is very limited (generally a few nanolitres). Although numerous methods have been developed for amino acids analysis over the past 50 years [3][4][5][6][7], there is not yet any method reported that is able to profile the concentrations of amino acids in the small volumes of sap obtained from individual Arabidopsis plants. Such resolution is crucial if functional genomics is to determine the effects of individual gene knockouts and to quantify both within-plant and plant-to-plant variation.…”

Section: Introductionmentioning

confidence: 99%

Amino acid analysis by micellar electrokinetic chromatography with laser‐induced fluorescence detection: Application to nanolitre‐volume biological samples from Arabidopsis thaliana and Myzus persicae

et al. 2005

View full text Add to dashboard Cite

Amino acids were derivatised with 4-fluoro-7-nitrobenzo-2,1,3-oxadiazol (NBD-F), separated by micellar electrokinetic chromatography (MEKC), and detected by argon-ion (488 nm) laser-induced fluorescence. The optimised MEKC background electrolyte conditions were: 40 mM sodium cholate, 5 mM beta-cyclodextrin in 20 mM aqueous borate buffer, pH 9.1, with 7% v/v acetonitrile. Using these conditions, 19 amino acids were separated within 17 min. The limits of detection were in the range of 7.6-42.2 pmol/mL and limits of quantitation from 0.05-0.14 nmol/mL. The method was systematically validated for injection volume error, migration time variation, calibration linearity, accuracy, precision, and recovery. Nanolitre volume samples of phloem sap of individual sieve element cells from the plant Arabidopsis thaliana and honeydew from the aphid Myzus persicae were directly analysed with this method. Quantitative amino acid concentrations in these two biological matrices were profiled for the first time. This method is particularly important because it allows the complete profile of the amino acids obtained from individual phloem elements, allowing cell to cell and plant to plant variation to be quantified, which to date has not been possible with Arabidopsis thaliana.

show abstract

High-performance liquid chromatographic separation with electrochemical detection of amino acids focusing on neurochemical application

Cited by 31 publications

References 36 publications

H2S Biogenesis by Human Cystathionine γ-Lyase Leads to the Novel Sulfur Metabolites Lanthionine and Homolanthionine and Is Responsive to the Grade of Hyperhomocysteinemia

H2S Biogenesis by Human Cystathionine γ-Lyase Leads to the Novel Sulfur Metabolites Lanthionine and Homolanthionine and Is Responsive to the Grade of Hyperhomocysteinemia

Muscarinic receptor regulation of osmosensitive taurine transport in human SH‐SY5Y neuroblastoma cells

Amino acid analysis by micellar electrokinetic chromatography with laser‐induced fluorescence detection: Application to nanolitre‐volume biological samples from Arabidopsis thaliana and Myzus persicae

Contact Info

Product

Resources

About