Abstract
The problems with separation of amino acid mixtures in reversed-phase mode are the result of their hydrophilic nature. The derivatisation of the amino group of mentioned above solutes leads to their solution. For this purpose, 9-fluorenylmethoxycarbonyl chloroformate (f-moc-Cl) as the derivatisation reagent is often used. In our study, the separation of some f-moc- amino acid derivatives (alanine, phenylalanine, leucine, methionine, proline and tryptophan) with the use of micellar systems of reversed-phase high-performance thin-layer chromatography (HPTLC) and pressurized planar electrochromatography (PPEC) is investigated. The effect of surfactant concentration, its type (anionic, cationic and non-ionic) and mobile phase buffer pH on the discussed above solute migration distances are presented. Our work reveals that the increase of sodium dodecylsulphate concentration in the mobile phase has a different effect on solute retention in HPTLC and PPEC. Moreover, it also affects the order of solutes in both techniques. In PPEC, in contrast to the HPTLC technique, the mobile phase pH affects solute retention. The type of surfactant in the mobile phase also impacts solute retention and migration distances. A mobile phase containing SDS improves system efficiency in both techniques. Herein, such an effect is presented for the first time.
Highlights
The problems with separation of amino acid mixtures in reversed-phase mode are the result of their hydrophilic nature
Our work reveals that the increase of sodium dodecylsulphate concentration in the mobile phase has a different effect on solute retention in high-performance thin-layer chromatography (HPTLC) and pressurized planar electrochromatography (PPEC)
It should be noted that in thin-layer chromatography, the micelle mobile phase can affect the demixing of eluent during plate development, resulting in two mobile phase fronts
Summary
The problems with separation of amino acid mixtures in reversed-phase mode are the result of their hydrophilic nature. With regard to other amino acid derivatives, separation of phenylthiocarbamyl utilizing TLC was presented in[12] These experiments were performed in both normal and reversed phase mode, while solute spot detection involved an in-situ iodine– azide reaction. Amino acid separations with the use of pressurized planar electrochromatography (PPEC) and TLC techniques and silica gel as the stationary phase and acetonitrile-aqueous buffer mixture as the eluent was compared in[14]. With regard to TLC and micellar eluent, Cerhati et al.[15] investigated the effect of a methanol-aqueous mobile phase containing surfactant (SDS or non-ionic Genapol O 80) on the retention of amino acids and a homologous series of peptides. An eluent with a mixture of surfactants (SDS and tridecylalcohol diglycolate) of various concentrations has been applied for investigating low molecular mass peptide interactions with a stationary phase of the RP-type (aluminium oxide impregnated with n-hexane paraffin oil mixture)[16]
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