Short communication
Transition metal ion binding studies of carnosine and histidine: Biologically relevant antioxidants

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Abstract

Carnosine and histidine are biologically interesting antioxidants. In order to probe whether they exert their antioxidant effect through metal ion chelation, the Cu(II) ion chelating abilities of these compounds were measured by UV–vis spectroscopy. Both of these compounds showed 1:1 complexations with Cu(II) ions as shown by their Job's plot. The binding constants for histidine and carnosine, as determined by Benesi–Hildebrand method, at pH 7.84 ± 0.18, were found to be 71 and 1.1 M−1, respectively. The unexpectedly lower binding constant values of carnosine show the relatively minor role of the transition metal ion chelation in their antioxidant abilities.

Introduction

Age-related diseases including Parkinson disease and Alzheimer's disease are characteristically associated with transition metal ion catalyzed oxidative damage to lipids, proteins and nucleic acids. Transition metal ion catalyzed lipid peroxidation, through Fenton reactions, leads to the formation of a variety of toxic byproducts such as trans-4-hydroxy-2-nonenal (HNE) and trans-4-oxo-2-nonenal (ONE) [1], [2], [3]. The latter toxicants can potentially deplete intracellular levels of glutathione and related cellular antioxidants. Biologically interesting antioxidants based on histidine and its derived dipepides, and related analogues, play a major role in chelating transition metal ions [4], [5], [6]. Further they can exert their antioxidant properties through reaction with secondary toxicants such a HNE and acrolein, thus attenuating their deleterious activities. Histidine and its derivatives, in particulalr, carnosine (β-alanyl-l-histidine), is also postulated to be effective in trapping the Fenton reaction derived reactive oxygen species (ROS). Carnosine is also known to be antiglycating agent as it can trap the advanced glycation end products (AGEs) resulting from the Maillard reactions of protein amino groups with reactive carbonyl compounds [7], [8], [9].

Histidine derived naturally occurring antioxidants such as carnosine and its analogues, anserine and carcinine are known to be a superior in vivo antioxidant as compared to histidine, although the precise mechanism of its antioxidant activity remains to be established [10], [11], [12], [13], [14], [15], [16], [17] The antioxidants effects of histidine and its derivatives may be ascribed to (1) transition metal ion chelation, (2) trapping of ROS, (3) through reaction with cellular toxicants such as HNE, and suppressing the formation of AGEs [8], [18]. Chelation of transition metal ions also helps in attenuating the production of ROS. It would thus be interesting to explore if, indeed, transition metal ion chelation play a major role in the antioxidant properties of histidine and its derived peptides. The binding constants of amino acids and peptides are dependent on the pH values of their corresponding solutions. There have been reports of the binding constants of histidine and carnosine, although a close comparison for these two compounds could not be achieved as they are measured under varied experimental conditions [4], [5], [6], [11]. With that goal, we have now estimated the comparative binding constants of Cu(II) ions by histidine and carnosine at close to physiological pH range under similar experimental conditions.

Section snippets

Results and discussion

We have measured the binding constants of histidine and carnosine with Cu(II) ions in order to determine their relative metal ion chelating abilities in aqueous solutions maintained at pH 7.84 ± 0.18, which is in the range of biological pH. We have adopted the continuous variation method, as described by Karikari et al. for the measurement of binding constants [19], [20]. The relative volumes of the Cu(II) solution and the antioxidants are shown in Table 1. Thus the total volume of each of the

Reagents and stock solutions

l-Histidine (>98%) and CuCl2·2H2O were obtained from Alfa, and l-carnosine (>99%) was obtained from Sigma Chemicals, and were used as received. A stock solution of the Cu(II) chloride solution (2.00 × 10−2 M) was prepared by dissolving CuCl2·H2O (0.178 g) in distilled water (50 mL), immediately prior to use. The 2.00 × 10−2 M stock solutions of l-histidine (0.155 g in 50 mL H2O; pH 7.71) and l-carnosine (0.225 g in 50 mL H2O; pH 7.97) were prepared immediately prior to use.

Instrumentation

A Cary-50 UV/vis

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