Rapid quantitative method for the detection of phenylalanine and tyrosine in human plasma using pillar array columns and gradient elution

Song, Yanting; Takatsuki, Katsuya; Sekiguchi, Tetsushi; Funatsu, Takashi; Shoji, Shuichi; Tsunoda, Makoto

doi:10.1007/s00726-016-2248-6

Cited by 22 publications

(22 citation statements)

References 24 publications

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“…The microchip fabrication has been described in previous studies [7][8][9][10]. The 20 mm × 20 mm microchip was composed of a cross-Tesla mixer, a separation channel, and a sample channel.…”

Section: Fabrication Of the Microchipmentioning

confidence: 99%

“…Six fluorescently labeled amino acids were analyzed within 100 s by isocratic elution using the pillar array column [7,8]. Furthermore, to separate the components of complex biological samples, a gradient elution system was developed to accelerate the elution of strongly retained solutes using pressure-driven liquid chromatography (LC) on a microchip [9,10].…”

Section: Introductionmentioning

confidence: 99%

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Retention and Bandwidth Predictions by Fast Gradient Elution Chromatography Using a Pillar Array Column

et al. 2016

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Previously, we have developed a gradient elution system for pillar array columns, which achieved faster separation than isocratic elution. In this study, we validated gradient elution in microchip liquid chromatography (LC) and investigated the retention and bandwidth predictions of fluorescently labeled aliphatic amines in fast gradient elution chromatography using semi-empirical retention models. The retention times and peak widths under different gradient elution programs were predicted by three solvent strength models and compared with the experimental results. The relative errors of prediction for the retention times and peak widths were below 14 % and 12 %, respectively. The results showed that the solvent strength models could be utilized for predicting the retention times and peak widths under gradient elution in the microchip LC system and that the gradient elution program for pillar array columns worked efficiently. The prediction by the retention model promises to be a potential tool for essential compound identification in biological samples.

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Section: Fabrication Of the Microchipmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Retention and Bandwidth Predictions by Fast Gradient Elution Chromatography Using a Pillar Array Column

et al. 2016

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“…Pillar array columns with perfectly ordered structures can shorten separation times significantly without lessening the quality of the separation. [9][10][11][12][13][14][15][16] However, pillar array columns filled with non-porous pillars have limited retention ability since only the outer surface of the pillars is available for separation. Therefore, until now, only fluorescent derivatives of hydrophobic amino acids, such as valine (Val), phenylalanine, and tyrosine, derivatized with the flurogenic reagent, 4-fluoro-7-nitro-2,1,3-benzoxadiazole (NBD-F), were retained and separated by pillar array columns.…”

Section: Introductionmentioning

confidence: 99%

“…Therefore, until now, only fluorescent derivatives of hydrophobic amino acids, such as valine (Val), phenylalanine, and tyrosine, derivatized with the flurogenic reagent, 4-fluoro-7-nitro-2,1,3-benzoxadiazole (NBD-F), were retained and separated by pillar array columns. [14][15][16] The analysis of hydrophilic amino acids still cannot be performed on pillar array columns because of their weak retention ability.…”

Section: Introductionmentioning

confidence: 99%

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Retention of Fluorescent Amino Acid Derivatives in Ion-pairing Reversed-phase Liquid Chromatography

Kuroki

Funatsu

et al. 2018

ANAL. SCI.

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Recent studies have shown that pillar array columns enable fast and quantitative analysis of amino acids. However, hydrophilic amino acids still cannot be retained on pillar array columns since they have limited retention ability. Ionpairing liquid chromatography is a promising means of increasing analyte retention. In this study, the effects of ionpairing reagents on the retention of eight hydrophilic amino acids (histidine, asparagine, glutamine, serine, arginine, aspartic acid, glycine, and glutamic acid) derivatized with 4-fluoro-7-nitro-2,1,3-benzoxadiazole (NBD-F) under reversedphase conditions on a conventional ODS column were studied. Among the ion-pairing reagents investigated, tetraheptylammonium bromide proved to be the most effective for increasing analyte retention. With a mobile phase consisting of 20 mM citrate buffer (pH 6.3)-acetonitrile (100:40, v/v) and 2 mM tetraheptylammonium bromide, the retention times of the eight NBD-amino acids-except NBD-arginine-were longer than 19.4 min, which was the retention time of NBD-valine when eluted without an ion-pairing reagent. As NBD-valine was well retained on pillar array columns, the chromatographic conditions may thus be applied in the analysis of hydrophilic amino acids using pillar array columns.

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Identification of methylated tubulin through analysis of methylated lysine

2018

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Post-translational modifications to tubulin such as acetylation and detyrosination play important roles in microtubule functions. Methylation is an important post-translational modification; however, to date, few methylated tubulins have been identified. In the present study, we developed a method for analyzing methylated lysine with the aim of identifying methylated tubulin. This method involves four steps: (1) acid hydrolysis of tubulin into amino acids, (2) selective extraction of methylated lysine using a monolithic-silica disk-packed spin column, (3) fluorescence derivatization of methylated lysine with 4-fluoro-7-nitro-2,1,3-benzoxadiazole (NBD-F), and (4) separation of NBD-methylated lysine on a column consisting of C18, cation and anion ligand, and fluorescence detection. Using the newly developed method, the dimethylation of lysine in tubulin was identified. This new method could be applied to searches for other methylated proteins.

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Rapid quantitative method for the detection of phenylalanine and tyrosine in human plasma using pillar array columns and gradient elution

Cited by 22 publications

References 24 publications

Retention and Bandwidth Predictions by Fast Gradient Elution Chromatography Using a Pillar Array Column

Retention and Bandwidth Predictions by Fast Gradient Elution Chromatography Using a Pillar Array Column

Retention of Fluorescent Amino Acid Derivatives in Ion-pairing Reversed-phase Liquid Chromatography

Identification of methylated tubulin through analysis of methylated lysine

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