Development of enzyme-linked immunosorbent assays for Sudan dyes in chilli powder, ketchup and egg yolk

Abstract

This study aimed at developing sensitive competitive enzyme-linked immunosorbent assays (ELISAs) for the banned Sudan dyes using polyclonal antibodies. Three different formats were developed and characterized in terms of sensitivity, selectivity and rapidity. A competitive indirect ELISA was developed, which showed an IC₅₀ of 3.8 µg l^–1. Two competitive direct ELISAs were also developed, in which the antibody was added before or simultaneously with the other reagents; the first showed an IC₅₀ of 8.3 µg l^–1 and the latter showed an IC₅₀ of 4.9 µg l^–1. Nevertheless, considering dilution of extracts which is needed to offset matrix interference, the limits of detection of the three formats were substantially the same (10 µg kg^–1). The antibodies in all three test formats were able to recognize Sudan I and partially Sudan II, III and IV; no cross-reactivity was observed with the five edible dyes. Twenty food samples, including chilli powder, paprika, ketchup, and egg, were extracted by a simple sample preparation and very limited dilution. Extracts were analyzed by the developed competitive direct ELISA with the simultaneous addition of reagents. A good correlation was observed (y = 1.19x–10.0, r ² = 0.991, n = 20) when the data was compared with that obtained through a conventional HPLC method.

Keywords:

• Sudan dyes
• ELISA
• chilli powder
• egg yolk
• ketchup

Acknowledgement

This work was funded by the Italian Ministry for Universities (PRIN, prot. n. 2005030782_001).