Rapid assay for tryptophanase using reversed-phase high- performance liquid chromatography

Krstulović, Ante M.; Matzura, Cynthia

doi:10.1016/s0021-9673(00)85653-8

Cited by 17 publications

(9 citation statements)

References 7 publications

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“…For instance, the total dispersion of an elution peak is the sum of the individual contributions of the different parts of the chromatographic system [2].…”

Section: Extra-column Contributionsmentioning

confidence: 99%

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Measurement of the parameters of the mass transfer kinetics in high performance liquid chromatography

Miyabe

Guiochon²

2003

J of Separation Science

148

161

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Fundamental studies of the mass transfer kinetics are as essential as those of the retention equilibrium for a detailed understanding of the characteristics and the mechanisms of chromatographic separations. The acquisition of a large amount of reliable experimental data and of meaningful results is necessary for any further progress of our knowledge of kinetics. The main goal of this review is to provide information on the methods used to perform accurate measurements and on the data analysis procedures used for deriving the kinetic parameters characterizing mass transfer in HPLC. First, the general characteristics of several methods of determination of some kinetic parameters are briefly reviewed. Secondly, we give detailed explanations of the experimental conditions of the pulse on a plateau method (i.e., elution chromatography on a plateau of finite concentration or pulse response method) and of the data analysis procedures based on moment analysis. Thirdly, we explain some important requirements for the acquisition of appropriate experimental data and discuss corrections to be applied when deriving several kinetic parameters. Fourthly, we discuss the accuracy of the kinetic parameters derived from the pulse on a plateau method and from moment analysis. Finally, some results concerning the mass transfer kinetics in RPLC systems are demonstrated as examples.

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“…For instance, the total dispersion of an elution peak is the sum of the individual contributions of the different parts of the chromatographic system [2].…”

Section: Extra-column Contributionsmentioning

confidence: 99%

“…(21). For instance, the following equations provide the band spreading (variance) in a tube (r tub ), in volume unit [2,73].…”

Section: Extra-column Contributionsmentioning

confidence: 99%

Measurement of the parameters of the mass transfer kinetics in high performance liquid chromatography

Miyabe

Guiochon²

2003

J of Separation Science

148

161

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“…The indole production in an enzymic reaction was evaluated using reversed-phase HPLC as described by Krstulovic & Matzura (1979). The reaction mixture contained the following reagents in a final volume of 100 ml: 167 mM potassium phosphate buffer (pH 7.5), 0.165 mM PLP, 0.2 mM reduced glutathione, 0.25 mg BSA ml 21 , 2 mM L-tryptophan and 50 mg ml 21 crude extract of P. gingivalis strain W83, KO100 or RS101.…”

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confidence: 99%

Identification and molecular characterization of tryptophanase encoded by tnaA in Porphyromonas gingivalis

et al. 2009

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Indole produced via the b-elimination reaction of L-tryptophan by pyridoxal 59-phosphatedependent tryptophanase (EC 4.1.99.1) has recently been shown to be an extracellular and intercellular signalling molecule in bacteria, and controls bacterial biofilm formation and virulence factors. In the present study, we determined the molecular basis of indole production in the periodontopathogenic bacterium Porphyromonas gingivalis. A database search showed that the amino acid sequence deduced from pg1401 of P. gingivalis W83 is 45 % identical with that from tnaA of Escherichia coli K-12, which encodes tryptophanase. Replacement of the pg1401 gene in the chromosomal DNA with the chloramphenicol-resistance gene abolished indole production. The production of indole was restored by the introduction of pg1401, demonstrating that the gene is functionally equivalent to tnaA. However, RT-PCR and RNA ligase-mediated rapid amplification of cDNA ends analyses showed that, unlike E. coli tnaA, pg1401 is expressed alone in P. gingivalis and that the nucleotide sequence of the transcription start site is different, suggesting that the expression of P. gingivalis tnaA is controlled by a unique mechanism. Purified recombinant P. gingivalis tryptophanase exhibited the Michaelis-Menten kinetics values K m 50.20±0.01 mM and k cat 51.37±0.06 s "1 in potassium phosphate buffer, but in sodium phosphate buffer, the enzyme showed lower activity. However, the cation in the buffer, K + or Na + , did not appear to affect the quaternary structure of the enzyme or the binding of pyridoxal 59-phosphate to the enzyme. The enzyme also degraded S-ethyl-L-cysteine and S-methyl-L-cysteine, but not L-alanine, L-serine or L-cysteine.

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“…Some methods 7) have been proposed to explain the effect in chromatography of the eluent composition on the capacity factor. Yano and lanod0 8 ) have reported the hydrophobic interaction chromatography of n-alcohols on cross-linked dextran gels and showed that the transfer free energy of the alcohols from external water to internal water was entropic.…”

Section: Discussionmentioning

confidence: 99%

“…These phenomena seem to have been due to the non-linearity of the respective adsorption isotherms to the TMP-IOOPN gel, which may be favorable. 6,7) Despite this, examining the effect of methanol concentration in the eluent on the separability of an alkyl glucoside and its corresponding alcohol by HPLC with the TMP-I OP gel gives a good indication of the methanol concentration required of the eluent for conventional column chromatography with the TMP-IOOPN gel. More detaisl of the operating conditions for conventional column chromatography based upon the results obtained by HPLC require rate processes such as diffusion of the solute in the gel to be considered.…”

Section: Separation Of Alkyl Glucoside and Alcohol By Conventionalmentioning

confidence: 99%

Separation of Alkylβ-d-Glucosides andn-Alcohols by Using a Porous Trimethylolpropane Trimethacrylate Homopolymer Gel

Adachi

Panintrarux

Araki

et al. 1994

Bioscience, Biotechnology, and Biochemistry

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Rapid assay for tryptophanase using reversed-phase high- performance liquid chromatography

Cited by 17 publications

References 7 publications

Measurement of the parameters of the mass transfer kinetics in high performance liquid chromatography

Measurement of the parameters of the mass transfer kinetics in high performance liquid chromatography

Identification and molecular characterization of tryptophanase encoded by tnaA in Porphyromonas gingivalis

Separation of Alkylβ-d-Glucosides andn-Alcohols by Using a Porous Trimethylolpropane Trimethacrylate Homopolymer Gel

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