Novel Approach of Sakaguchi's Reaction for Quantitative Determination of L-Arginine via Flow Injection Analysis-Merging Zones with Spectrophotometric Detection

Abstract

A new batch and flow injection analysis-merging zones method characterized by simplicity, accuracy and speed for determination of L-arginine in pure material and in pharmaceutical formulations. The methods was based on oxidation-condensation for L-arginine with a-naphthol was added in urea, in presence of sodium hydroxide as a medium. For amino acid reaction, using sodium hypobromite as oxidizing agent to form an intense red water-soluble dye that is stable and has a maximum absorption at 501 nm. The optimum parameters were 5.5 mL min^-1 flow rate using distilled water as a carrier, 42.19 μL sample volume (35 μg mL^-1), 43.175 μL [a-naphthol (1.4 × 10-3 M) in 10 % urea], 54.95 μL NaOBr (0.25 M) for L₁, L₂ and L₃ respectively and open value for the purge of sample segment and other chemicals. A graphs of absorbance versus concentration show that Beer’s law is obeyed over the concentration range of 1-45 and 3-1400 μg mL^-1 of L-arginine with detection limits of 0.5 and 0.1 μg mL^-1 of L-arginine for batch and flow injection analysis-merging zones methods respectively. The optimized flow injection analysis system is able to determine L-arginine through put 45 h^-1. The correlation coefficient (r) was 0.992 and percentage linearity (% r²) was 98.4 %, RSD % for the repeatability (n = 8) was 0.5-1.5 % for determination of arginine with concentration 15 and 30 μg mL^-1. The proposed method was applied successfully for determination of arginine in pharmaceutical formulations. The newly developed method has been statistically evaluated with official method and there were no significant differences between either methods. The proposed method can be accepted as an alternative analytical method for determination of L-arginine in pharmaceutical and biological samples.