Rapid high-throughput assay for the measurement of amino acids from microdialysates and brain tissue using monolithic C18-bonded reversed-phase columns

Dawson, Lee A.; Organ, Andrew J.; Winter, Panida; Lacroix, Laurent; Shilliam, Claire S; Heidbreder, Christian; Shah, Ajit J.

doi:10.1016/j.jchromb.2004.04.012

Cited by 21 publications

(12 citation statements)

References 33 publications

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“…Previously published work using monolithic technology [15] has used naphthalene-2,3-dicarboxaldehyde in the presence of cyanide (NDA/CN − ) as the fluorogenic reagent for amino acid detection, which has been shown to form stable derivatives and offers improved sensitivity in comparison to OPA/2-mercaptoethanol (2-ME) derivatisation [16]. Our assay uses OPA/3-MPA derivatisation, which forms more stable isoindole derivatives that those produced by OPA/2-ME [17].…”

Section: Methods Verificationmentioning

confidence: 99%

“…In addition, the faster separation time of this method to some extent negates the issue of degradation. As stated previously, derivatisation with OPA/3-MPA involves a one-step process lasting 3 min, unlike NDA/CN − derivatisation, which requires the components to be prepared as separate solutions and requires 15 min incubation [15]. Therefore, higher throughput is possible with OPA/3-MPA derivatisation.…”

Section: Methods Verificationmentioning

confidence: 99%

See 1 more Smart Citation

Monolithic column-based reversed-phase liquid chromatography separation for amino acid assay in microdialysates and cerebral spinal fluid

Devall

Blake

Langman

et al. 2007

Journal of Chromatography B

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Section: Methods Verificationmentioning

confidence: 99%

Section: Methods Verificationmentioning

confidence: 99%

Monolithic column-based reversed-phase liquid chromatography separation for amino acid assay in microdialysates and cerebral spinal fluid

Devall

Blake

Langman

et al. 2007

Journal of Chromatography B

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“…In recent years, separation of the derivatives by pre-column derivatization and RP-HPLC has been widely used (Dawson et al 2004;Yang et al 2007). As most amino acids lack UV absorbance in 220-254 nm range, general method employs derivatization agent to transform the analytes into derivatives that can be separated and determinated more easily (Knapp 1979;Fan et al 1998;Kőrös et al 2008).…”

Section: Introductionmentioning

confidence: 99%

Determination of 20 Free Amino Acids in Asparagus Tin by High-Performance Liquid Chromatographic Method after Pre-Column Derivatization

Hou

Cao

et al. 2011

Food Anal. Methods

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A novel method was studied for determination of 20 free amino acids in asparagus tin by high-performance liquid chromatography with pre-column derivatization. Derivatization of the samples was performed with 4-chloro-3,5-dinitrobenzotrifluoride (CNBF), and the SPE cartridge was used for purification and enrichment of the analytes. The derivatization conditions and the influence of elution composition on the separation were investigated. The reaction of amino acids with CNBF was completed in pH 9.0 borate buffer. The separation of amino acids was achieved at room temperature within 60 min by gradient elution mode with triethylamine in mobile phase, and the flow rate is 0.32 mL min −1 constantly. The method correlation coefficient was from 0.9975 to 1.0000 in concentrations ranging from 20 to 2000 μmol L −1 , except asparagine (from 100 to 10000 μmol L −1 ). The detection limits of amino acids were from 1.2 to 6.0 μmol L −1 , with a signal-to-noise ratio of three times. The calculated recoveries of the proposed method were from 81.4% to 109.4%, and relative standard deviations were 0.48-3.94% in application to the quantitative determination of free amino acids in asparagus tin samples. The present method is reliable and sensitive that allows fast analysis of free amino acids in asparagus tin, which makes it suitable for further study of free amino acids in other asparagus foods.

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“…Besides, the four amino acids were all relatively stable within 60 min after derivatization which indicated that once OPA derivatization product was injected into the separation column, it was stable during a run time of 25 min without being degradated. Furthermore, to our knowledge, although another derivatizing agent, naphthalene-2,3-dicarboxaldehyde (NDA), which required potassium cyanide to produce a 1-cyanobenz[f]isoindole (CBI) derivative, was more widely used due to its greater stability than OPA derivatization but had the disadvantage of using the highly toxic substance (CBI) [25,26]. In addition, some other fluorescence derivatization reagents like dansyl (DNS), fluorescein isothiocyanate (FITC) and 7-fluoro-4-nitrobenzo-2-oxa-1,3-diazole (NBD) were reported in the previous studies [27,28], but they were not as convenient as OPA in the application, for instance, amino acids were derivatized with DNS or NBD by heating to 60°C for several minutes and FITC-derivatized samples generated a large background peak disturbing experimental analysis.…”

Section: Linearity Precision Recovery and Stabilitymentioning

confidence: 99%

RP-LC with Fluorescence Detection of Amino Acids in Rat Brain Synaptosomes

Mao¹,

Cao

Min³

et al. 2011

Chromatographia

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For the first time, a simple and validated reversed-phase liquid chromatography (RP-LC) with fluorescence detection has been developed for the simultaneous analysis of glutamate (Glu), c-aminobutyric acid (GABA), glycine (Gly) and taurine (Tau) in Wistar and tremor rats brain synaptosomes. The samples were separated on a C18 analytical column with gradient elution of methanol and 0.1 mol L -1 potassium acetate at a flow rate of 1 mL min -1 . Total run time was approximately 25 min. All calibration curves exhibited good linearity (r 2 [ 0.999) within test ranges. The reproducibility was estimated by intra-and inter-day assays and RSD values were less than 2.48%. The recoveries were between 96.32 and 105.21%. The method was successfully applied to the quantification of amino acids in Wistar and tremor rats brain synaptosomes. Through this developed protocol, the levels of Glu in hippocampal and prefrontal cortical synaptosomes of tremor rats were both significantly elevated than those of adult Wistar rats whereas significantly decreased concentrations of GABA and Gly were observed in the hippocampal region of tremor rats without evident difference in the prefrontal cortex between experimental and control groups. In addition, our studies also showed a marked elevation of Tau in tremor rats hippocampal synaptosomes although there was no pronounced difference in the prefrontal cortical region of Wistar and tremor rats.

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Rapid high-throughput assay for the measurement of amino acids from microdialysates and brain tissue using monolithic C18-bonded reversed-phase columns

Cited by 21 publications

References 33 publications

Monolithic column-based reversed-phase liquid chromatography separation for amino acid assay in microdialysates and cerebral spinal fluid

Monolithic column-based reversed-phase liquid chromatography separation for amino acid assay in microdialysates and cerebral spinal fluid

Determination of 20 Free Amino Acids in Asparagus Tin by High-Performance Liquid Chromatographic Method after Pre-Column Derivatization

RP-LC with Fluorescence Detection of Amino Acids in Rat Brain Synaptosomes

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