Enzymatic oxidation in aqueous and micellar media based on horseradish peroxidase–hydrogen peroxide system using a SIA manifold

Abstract

Aqueous and micellar catalysis of horseradish peroxidase was studied in a sequential injection system through selecting the oxidation by hydrogen peroxide of two phenothiazines: the water-soluble chlorpromazine and perphenazine, a low water soluble, micellised in a Tween 80 medium. The coloured free-radical intermediate formed was monitored spectrophotometrically at 527 nm. The system enables the determination of chlorpromazine in water and perphenazine in micellar medium up to 1.25 × 10 −4 mol L −1, with quantification limits of 2 × 10 −5 and 1.25 × 10 −5 mol L −1, respectively. R.S.D. values were in both cases less than 1.6%. The SIA system optimized consumed 125 μL of sample, 1.5 μg of peroxidase and 5 × 10 −8 mol of hydrogen peroxide per determination, which guarantees economy in the use of sample and reagents with reduced residue production, in good agreement with the current recommendations of green chemistry. The developed system was applied to the determination of chlorpromazine and perphenazine in pharmaceutical preparations. Results revealed different reaction rates for aqueous and micellar media, which yielded determination frequencies of 17 and 31 determinations per hour, respectively. The potential effect of several compounds commonly used as excipients on analytical signals was studied and no interfering effect was noticeable. Statistical comparison of the results obtained with the proposed methodology and with the reference methods showed good agreement and indicate no significant difference at the 95% confidence level.