Determination of annatto in high‐fat dairy products, margarine and hard candy by solvent extraction followed by high‐performance liquid chromatography

Abstract

Utilizing solvents such as ethanolic aqueous ammonia, petroleum ether, hexane and chloroform, annatto components alpha- and beta-norbixin and alpha- and beta-bixin were extracted from cheese, butter, margarine and hard candy. After transferring the extract into a solution of aqueous acetic acid in methanol, bixin and norbixin were determined quantitatively using high-performance liquid chromatography (HPLC) and an absorbance detector set at 500 nm. Recovery of norbixin from spiked cheese samples averaged 92.6% over a range of 1 to 110 micrograms/g. Commercial cheese samples were found to contain 1.1-68.8 microgram/g total norbixin, and two samples also contained 5.1-5.6 micrograms/g total bixin. Samples of uncoloured butter were spiked with bixin and recovery averaged 93.2% over a range of 0.1 to 445 micrograms/g. Levels of 0.2 microgram/g total bixin and 0.91 microgram/g total norbixin were found in one commercial butter sample; the others contained trace levels of both compounds. Hard candies were prepared in the laboratory and recovery studies conducted. Recovery of norbixin averaged 88%.