Identification and Interaction of Amino Acids with Leucine-Anthracene Reagent by TLC and Spectrophotometry: Experimental and Theoretical Studies

Abstract

A new reagent has been synthesized by coupling anthracene moiety to L-leucine. The reagent is characterized by different analytical techniques. It is capable for easy identification of various amino acids on thin-layer chromatography plates by developing distinguishable colors (detection limit between 0.1-0.5 μg at cold condition and 0.1-0.4 μg after heating). This reagent also binds with different amino acids very strongly in solution (methanol). Estimation of equilibrium binding constants of this new reagent with different amino acids has also been carried out. The values of the binding constants are lowest for L-Tyrosine (6.86 × 10³ dm³ mol(-1)) and highest for L-Arginine monohydrochloride (8.86 × 10⁵ dm³ mol(-1)) at 25°C. A theoretical study (Hartree-Fock) has been performed to investigate the interaction of the reagent with a representative amino acid, glycine.

Modified Ninhydrin reagent for the detection of amino acids on TLC plates

“…The DDQ (Sinhababu et al, 1994) and 2,3-Dichloro-1,4-Naphthaquinone (Khawas et al, 2004) are the two important reagents that can be used alone instead of ninhydrin in this respect. Some complex reagents have also been developed by other workers (Das et al, 2010;Sahana et al, 2011) but their detection limits and color contrasts are not so good enough than some other reagents referred here. Recently, we have also introduced some p-haloderivatives as the modified spray reagents (Sinhababu et al, 2013).…”

“…Thin layer chromatography (TLC) finds its place when the relatively costly equipment required by other methods is unavailable. Thin layer chromatography is an important tool for the identification of amino acids by various spray reagents (Lorenz and Flatter, 1970;Devaux and Mesnard, 1971;Wolski et al, 1980;Distler and Fresenius, 1981;Laskar and Basak, 1988;1990;Basak and Laskar, 1990;Laskar et al 1991;2001a,b,c;Basak et al, 1993;Sinhababu et al, 1994;2013;Khawas and Laskar 2003a,b;Khawas et al, 2004;Samanta et al, 2004;Samanta and Laskar, 2006;Das et al, 2010;Sahana et al, 2011). Of the reagents in general use, ninhydrin is the most popular because of its remarkably high sensitivity (Stahl, 1969).…”