Biotecnología

 

Production and optimization of a chlorophyl-free leaf protein concentrate from alfalfa (Medicago sativa) through aqueous two-phase system

 

Producción y optimización de un concentrado de proteína de alfalfa (Medicago sativa) libre de clorofila mediante sistemas de dos fases acuosas

 

P. Vázquez-Villegas, E. Acuña-González, L.A. Mejía-Manzano, M. Rito-Palomares y O. Aguilar*

 

Centro de Biotecnología-FEMSA, Tecnológico de Monterrey, Campus Monterrey, Ave. Eugenio Garza Sada 2501 Sur, Monterrey, NL 64849, México. *Corresponding author. E-mail: alex.aguilar@itesm.mx Tel. 83-58-20-00, ext, 4821.

 

Received October 23, 2014
Accepted April 8, 2015

 

Abstract

There is a growing interest in producing protein from green crops, being alfalfa (Medicago sativa) one of the more studied models. In this work we develop an aqueous two-phase systems (ATPS) process to obtain a chlorophyl-free leaf protein concentrate. A two-level factorial experimental design in order to study five factors that interferes with recovery of total protein in ATPS is performed, employing two model proteins (BSA and Lyz). The factors of polymer concentration and total protein concentration were statistically significative. From here, we applied steepest descent and central composite design (CCD) methods in order to optimize chlorophyl-free, total protein recovery from alfalfa (Medicago sativa) green tissue. An extraction of more than 809% of total protein was reached for bottom phase LPC and 51% of protein in powder from the lyophilized LPC.

Key words: alfalfa, aqueous two-phase system (ATPS), central composite design, leaf protein concentrate.

 

Resumen

Existe un interés creciente en producir proteína a partir de tejido vegetal, siendo alfalfa (Medicago sativa) uno de los modelos más estudiados. En el presente trabajo, se desarrolla un proceso con sistemas de dos fases acuosas (ATPS, de sus siglas en ingles) para obtener un concentrado de proteína libre de clorofila, a partir de tejido verde. Se llevó a cabo un diseño experimental factorial de dos niveles para estudiar los factores que interfieren con la recuperación de proteína total, empleando dos proteínas modelo (albumina de suero bovino y lisozima). Los factores de concentración de polímero y concentración total de proteína fueron estadísticamente significativos. A partir de dichos resultados, se aplicó el método estadístico de búsqueda de descenso por gradiente (steepest descent) y un diseño central compuesto para optimizar la recuperación de proteína total libre de clorofila a partir de tejido verde de alfalfa (Medicago sativa). Un concentrado con mas de 80% y 51% de proteína se obtuvo en fase inferior líquida y liofilizada, respectivamente.

Palabras clave: alfalfa, sistema de dos fases aquosas (ATPS), diseño central compuesto, concentrado de proteína.

 

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Acknowledgements

The authors wish to acknowledge the financial support of Tecnológico de Monterrey, Bioprocesses and Synthetic Biology Group and CONACyT for the fellowships of E. Acuña-González and L.M. Mejía-Manzano, No. 332683 and No. 252731, respectively.

 

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