Purification and Characterization of Soluble Cichorium intybus Var. silvestre Lipoxygenase

A water-soluble lipoxygenase enzyme (EC 1.13.11.12; LOX) occurring in the red cultivar produced
in the geographical area of Chioggia (Italy) of Cichorium intybus var. silvestre was isolated and
characterized. The molecular mass of the enzyme was estimated to be 74000 Da by sodium dodecyl
sulfate-polyacrylamide gel electrophoresis and gel filtration chromatography. The isoelectric point
was pH 6.85. The optimum values of pH, ionic strength, and temperature, shown by isoresponse
surface calculated by a randomized multilevel factorial design, were 7.58, 30 mM, and 38.5 °C,
respectively. The enzyme showed high specificity toward linoleic acid, and the study of the variation
of linoleic acid concentration between 30 and 300 íM, in the presence of Tween 20 at a concentration
lower than the critical micelle concentration (0.01 v/v), resulted in a typical Michaelis-Mentem curve
with KM and Vmax values of 1.49 10-4 M and 2.049 íM min-1 mg-1, respectively. The biochemical
properties, the kinetic parameters found, and the carotene-bleaching activity shown in aerobic
conditions seem to indicate that the isolated enzyme is a lipoxygenase type III according to the
indications given for soybean isoenzymes.