Pectin esterase (PE, E.C. 3.1.1.11) is implicated in the cloud loss in citrus juices. This study describes a specific method of extracting pectin esterase from blood oranges and developing, contemporarily, an enzyme assay suitable for the pH of such juices. The PE showed an optimum extraction at pH 7.0, without addition of surfactants. The enzyme assay was carried out at pH 3.6, a condition typical of the orange juices, increasing sensitivity in determining PE compared to other methods proposed up until now.