Chemiluminescence determination of the antioxidant capacity of honey samples and fruit juices using lab-on-a-chip device

Honey is well-known as a natural and healthy food containing, in addition to sugars, a wide range of substances including flavonoids, phenolic acids and lysozyme with antibacterial and antioxidant activity. Fruits and vegetables partially due to the possessing various antioxidants such as vitamins and polyphenols are very beneficial for the human health [1]. In general, antioxidants can be defined as artificial or natural chemical compounds that their presence even at low-level concentrations can hinder or slow the oxidation of the oxidisable substances [2]. Indeed, antioxidants can protect organisms or non-living systems from the adverse effects of free radical reactions involving reactive oxygen and nitrogen species (ROS and RNS). In general, for expression of antioxidant status and therapeutic potential of a food sample, the terms of “total antioxidant capacity (TAC)” is used. The assessment of TAC is very sample- and method-specific because the reaction mechanisms of TAC determination methods are totally different in terms of types of oxidants, reaction conditions and even results statement. However, the assessment of the TAC under precisely controlled conditions is an outstanding strategy for comparison and ranking of the antioxidant potential of food products. For instance, TAC assessment can be used for evaluating and characterization of honey samples. In general, a variety of methods have been developed for determination of TAC, most of which may fall in one of the three following major categories: (a) spectrophotometry methods: those are mainly based on the reaction of the colored reagents such as 2,2-diphenyl-1 picrylhydrazyl radical (DDPH•) radicals with antioxidants, leading to change of the color of reaction solution, (b) electrochemistry methods: those are usually based on the oxidation of antioxidants on the surface of an appropriate working electrode and (c) chemiluminescence (CL) methods: those are generally based on the enchasing or inhibitory effects of antioxidants on the CL reactions. In this research work, a new microfluidic system with chemiluminescence (CL) detection for the rapid assay of the total antioxidant capacity (TAC) of apple and pomegranate juices and honey samples was developed. The method exploited the green and simple CL reaction of NaHCO3-H2O2-Co2+. It was found that gallic acid (GA), catechin, caffeic acid, ferulic acid and rutin as selected phenolic and polyphenolic antioxidants could suppress the light emitted from the NaHCO3-H2O2-Co2+ CL reaction. The CL intensity was inversely related to the concentration of these antioxidants. After optimization of flow rate, concentrations of reagents and pH, the parameters influencing the CL emission, the calibration curve for each antioxidant was drawn. The linear range and limit of detection (LOD) for the antioxidants were as follows: 0.5-3 mg L-1 and 0.27 mg L-1 for GA, 0.2-5.0 mg L-1 and 0.17 mg L-1 for catechin, 0.03-2.0 mg L-1 and 0.03 mg L-1 for caffeic acid, 0.3-2.0 mg L-1 and 0.23 mg L-1 for ferulic acid and 0.3-4.0 mg L-1 and 0.15 mg L-1 for rutin, respectively. Of the tested phenolic and polyphenolic antioxidants, GA was chosen to state TAC of the apple and pomegranate juices and honey samples as GA equivalents (GAE). Moreover, the correlation study of the MF-CL method with the 2,2-diphenyl-1-picrylhydrazyl radical scavenging capacity assay (DPPH• assay) was performed. The results showed very good correlation between MF-CL and DPPH• methods. In addition, IC50 (an amount of antioxidant concentration needed to decrease the CL intensity by 50%) of the fruit juices and honey samples was also calculated.