High-sensitivity phenylthiohydantoin amino acid analysis using conventional and microbore chromatography

Cunico, Robert L.; Simpson, Richard J.; Correia, Lilian Cherubin; Wehr, C. Timothy

doi:10.1016/s0378-4347(00)85134-8

Cited by 29 publications

(6 citation statements)

References 9 publications

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“…Thus, coefficients with a variation from 8 -21% at no equilibration step (t = 0 min) range to lower than 0.64% when the equilibration time is 10 min or higher. These results are in agreement with those reported by Guitart et al [38] for several amino acid derivatives, but Asp and Glu derivatives show the best results here, 0.34 and 0.58%, respectively, against some 1 -3% in previously published data [18,24,28,41]. The equilibration time proposed here is much lower than the 60 min used by Beaver [42].…”

Section: Column Conditioningsupporting

confidence: 93%

“…From several columns assayed, we selected an Aminoquant 200 mm62.1 mm from Hewlett-Packard. Previous works based on microbore or narrow-bore columns [28,29,35,36] have shown poor results relating to the resolution of several pairs of amino acids like Thr -Ala, Glu -Ser and Ile -Leu even though they use longer columns than we have considered in this work. Furthermore, the optimization of chromatographic separation has been performed in terms of amino acid derivative resolution aiming to obtain an appropriate value higher than 1.5.…”

Section: Resultsmentioning

confidence: 86%

“…The PITC derivatization method has been widely applied and different modifications have been proposed; nevertheless, the proposed modifications mainly concerned previous steps in the method such as hydrolysis or amino acid derivatization [26,27], but few [28,29] concern scale reduction with an increase in analytical performance. In this work, we revisited this well known chromatographic method in order to implement it in a microbore column more appropriate to our objectives.…”

Section: Introductionmentioning

confidence: 99%

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Revisitation of the phenylisothiocyanate‐derivatives procedure for amino acid determination by HPLC‐UV

Checa-Moreno

Manzano

Mirón

et al. 2008

J of Separation Science

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A revisitation of the well known chromatographic procedure for the determination of amino acids as phenylthiocarbamyl derivatives (PTC) is performed. The method was developed for a microbore column that it is more appropriate to our later aims, the characterization of proteinaceous binders present in microsamples coming from the Cultural Heritage field. Several variables relating to chromatographic aspects were studied such as the pH and temperature of the mobile phase, buffer and modifier (triethylamine) concentrations in mobile phase and the stability of PTC-derivatives in solution. The calibration function was studied in depth. To prevent the heteroscedastic behaviour that it is observed, we used the weighed least squares fitting as the best strategy among other normalizing transformations, such as square root and logarithmic functions. Finally, the proposed method showed results similar to the traditional method in terms of efficiency, runtime, LODs and other characteristics, but with two additional advantages: a lower mobile phase consumption and the possibility of working with a lower sample volume. The usefulness of proposed method is checked against easel painting samples of Pictorial Heritage.

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Section: Column Conditioningsupporting

confidence: 93%

Section: Resultsmentioning

confidence: 86%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Revisitation of the phenylisothiocyanate‐derivatives procedure for amino acid determination by HPLC‐UV

Checa-Moreno

Manzano

Mirón

et al. 2008

J of Separation Science

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“…The individual amino acids were separated on a Waters HPLC system with a Waters C18-resolve column (5-pm spherical packing 3.9 mm x 15 mm stainless steel column) and detected by fluorometric measurements of the orthophtalaldehyde (OPA) derivatives (Jarrett et al 1986). The analyses were also performed on a Pico-tag system by Waters (Cunico et al 1984) and compared with those obtained with OPA derivatives. The analysis performed with the Pico-tag system gave similar results.…”

Section: Photosynthesis and I4c Feedingmentioning

confidence: 99%

Carbon partitioning in nodulated alfalfa in the presence of nitrate

Côté¹,

Bordeleau²,

Lapointe³

et al. 1989

Can. J. Bot.

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C 6~6 , R., BORDELEAU, L. M., LAPOINTE, J., and GRODZINSKI, B. 1989. Carbon partitioning in nodulated alfalfa in the presence of nitrate. Can. J. Bot. 67: 1998Bot. 67: -2004. Photosynthetic carbon partitioning from a fully expanded upper canopy leaf of nodulated alfalfa was examined in plants in which nitrogenase activity and leghemoglobin content were reduced by 70 and 50%, respectively, in the presence of 10 mM KNO,. The nitrite level in nodules of nitrate-treated plants was 12.5 + 0.5 pg Nlg FW and 0.3 f 0.4 pg Nlg FW in the control plants. Growth of axillary shoots and roots was markedly enhanced in treated plants, as were steady state leaf photosynthesis (54 f 4 to 113 f 8 pmol CO,/(mg chlorophyll. h)) and translocation speed (0.37 f 0.05 to 0.56 + 0.05 cm . min-I). Nodules received 5 % of the newly fixed carbon of the feed leaf 7 days after transfer to NO;, compared with less than 1 % for the control plants maintained on NO;-free medium. Sucrose was the major 14C-labelled photoassimilate, although small amounts of labelled raffinose-series sugars were detected in leaves of treated plants. The specific activity of asparagine in the leaf increased dramatically, consistent with a reduction in both nitrogen fixation and the import of unlabelled asparagine from nodules. Concentrations of alanine, y-amino butyric acid, glutamic acid, and aspartic acid were greater in all tissues of treated plants. However, pools of asparagine, glutamine, serine, glycine, threonine, and arginine were lower in leaf, shoot, and root tissue of the treated plants. The levels of all amides and amino acids in the nodules remained high in treated plants. The high levels of nitrite and amino acids in nodules of treated plants were correlated with reduced leghemoglobin content and nitrogenase activity, but this does not explain directly the mechanism of suppression of nitrogen fixation in alfalfa exposed to nitrate. Nevertheless, the ability of alfalfa nodules, which have an indeterminate growth pattern, to compete for carbon from upper leaves in the canopy may explain their longevity under NO; stress in the field.Key words: partitioning, photosynthesis, 14C0,, alfalfa, nitrate, nitrogenase, amino acids. C~T E , R., BORDELEAU, L. M., LAPOINTE, J., et GRODZINSKI, B. 1989. Carbon partitioning in nodulated alfalfa in the presence of nitrate. Can. J. Bot. 67 : 1998-2004.La rtpartition des composCs carbonks photosynthttisCs 2 partir d'une feuille mature de la partie suptrieure de la tige principale a Ctt CtudiCe chez la luzerne nodulte. L'activitt nitrogtnasique et la concentration en leghtmoglobine diminuent de 70 et 50%, respectivement, suite i une application de 10 mM KNO, pendant 7 jours. La concentration en nitrite dans les nodules des plants traitts est de 12,5 f 0,5 pg Nlg mf et de moins de 0,3 f 0,4 pg Nlg mf chez les plantes contrales. Chez les plants trait&, on observe une stimulation de la croissance des tiges axillaires et des racines de mCme qu'une augmentation du taux de photosynthkse 2 1'Cquilibre (54 + 4 2 113 + 8 pmol CO,/(mg ...

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“…TLC of PTH-amino acids has extensively been reviewed by Rosmus and Deyl (1972), Niederwieser (1972), Allen (1981), and Bhushan and Reddy (1987). The excellent resolution and reproducibility with high sensitivity of UV absorbance detectors have made HPLC a very successful method for identification and quantitation of PTH-amino acids using isocratic elution (Kin, 1982), binary gradient (Somack, 1980) and ternary gradient (Cunico et al, 1984). The excellent resolution and reproducibility with high sensitivity of UV absorbance detectors have made HPLC a very successful method for identification and quantitation of PTH-amino acids using isocratic elution (Kin, 1982), binary gradient (Somack, 1980) and ternary gradient (Cunico et al, 1984).…”

Section: Introductionmentioning

confidence: 99%

Liquid chromatographic separation of some PTH‐amino acids

Bhushan¹,

Saso²

1998

Biomed. Chromatogr.

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Liquid chromatographic studies on the separation of ten PTH-amino acids were carried out using normal phase untreated silica gel plates, C-18 RP precoated plates and RP-HPLC. Resolution of a complex mixture of PTH-amino acids was achieved using all the three types. Certain new successful solvent systems have been worked out in each case. HPLC was carried out with Lichrosphere 100 RP-18 (5 mm) column. Acetonitrile and sodium acetate buffer of pH 4.0 was used for reversed phase chromatography while for normal phase TLC combinations of chloroform-acetonitrile and chloroform-tetrahydrofuran were applied. #

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High-sensitivity phenylthiohydantoin amino acid analysis using conventional and microbore chromatography

Cited by 29 publications

References 9 publications

Revisitation of the phenylisothiocyanate‐derivatives procedure for amino acid determination by HPLC‐UV

Revisitation of the phenylisothiocyanate‐derivatives procedure for amino acid determination by HPLC‐UV

Carbon partitioning in nodulated alfalfa in the presence of nitrate

Liquid chromatographic separation of some PTH‐amino acids

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