46E-10 |
Quantification of the activities of tyrosinase and dioxygenase in beet (Beta vulgaris) cell cultures |
M. E. JARAMILLO-FLORES1, E. Nava-Rodríguez, A. Jiménez-Aparicio3, L. Dorantes-Alvarez, and H. S. Humberto. (1) Depto. Ingeniería Bioquímica, Escuela Nacional de Ciencias Biológicas, Instituto Politécnico Nacional, Carpio y Plan de Ayala, Col. P.E. Calles, México, DF, CP. 11340, Mexico, (2) CeProBi - IPN, Yautepec, Mor, Mexico Color has a definite influence in food preference. Betalains are natural pigments with several applications in the food industry as color developers or enhancers. Plant biotechnology has been used successfully in the production of natural pigments, as well as in the study of their biosynthesis mechanisms. The purpose of this work was to identify and quantify the activity of two of the enzymes (tyrosinase and dioxygenase) possibly involved in the betalains biosynthesis pathway of the beet (Beta vulgaris) in cell culture. Three different phases of growth (lag, exponential and stationary) were analyzed in a semisolid culture of beet cells. The samples were extracted with 10 mM phosphate - 10% glycerol - 1 mM PMSF - 1% ascorbic acid buffer. The extracts were treated with 90% saturation ammonium sulfate and the precipitates were dialysed against the same buffer (without PMSF), concentrated and analyzed for tyrosinase and dioxygenase activities. The cells harvested during the exponential phase showed the highest activity for both enzymes. The specific activities (in activity units/ mg of protein) in the three phases were 27.3, 31.1 and 19.6 respectively for the tyrosinase. In the case of the dioxygenase, the specific activities (in D Abs @ 420 nm/ mg of protein/ min) were 0.37, 0.63 and 0.62 respectively. These results are very important in the study of the betalains biosynthesis pathway. These enzymes surely have regulation mechanisms that, when fully studied, could be used in the over-production of the pigments.
Session 46E, International
|