A kinetic-based stopped-flow DPPH^• method.

Area of Science:

• Analytical Chemistry
• Biochemistry
• Food Science

Background:

• Assessing antioxidant functionality requires studying reactions with free radicals.
• Fast-acting antioxidants, like ascorbic acid, pose challenges due to rapid reaction times.
• Existing methods struggle to accurately quantify the kinetics of these rapid interactions.

Purpose of the Study:

• To develop a rapid and sensitive method for determining the absolute rate constants of reactions between fast antioxidants and DPPH• (2,2-diphenyl-1-picrylhydrazyl).
• To characterize the kinetic parameters, including second-order kinetics (k1) and potential side reactions (k2).
• To compare the antioxidant activity of various compounds and food samples, particularly fruit juices.

Main Methods:

• Utilized a stopped-flow spectrophotometric system to monitor the decay of DPPH• upon reaction with antioxidants.
• Applied a kinetic-based reaction mechanism to fit experimental data and derive kinetic parameters.
• Quantified the reaction rate constants (k1 and k2) for ascorbic acid, eleven phenols, and seven fruit juices.

Main Results:

• Ascorbic acid exhibited the highest reaction rate constant (k1 = 21,100 ± 570 M⁻¹s⁻¹).
• Eleven other phenols showed k1 values ranging from 45 to 3070 M⁻¹s⁻¹.
• Several compounds, including catechin and quercetin, displayed a side reaction (k2 from 15 to 60 M⁻¹s⁻¹).
• Strawberry juice demonstrated the fastest antioxidant activity, while red plum juice was the slowest among the tested juices.

Conclusions:

• The proposed kinetic-based DPPH• method is a simple, rapid, and sensitive technique for evaluating antioxidant activity.
• The method is suitable for studying the capacity of diverse molecules and antioxidant-rich food samples, such as fruit juices.
• This approach provides accurate kinetic data for fast-acting antioxidants, overcoming limitations of traditional methods.