Giberelinas

Volumes 113–116 (12 Issues), Spring 2004, ISSN: 0273–2289

Applied Biochemistry and Biotechnology
Executive Editor: Ashok Mulchandani

Biotechnology for Fuels and Chemicals
The Twenty-Fifth Symposium
Editors

Mark Finkelstein James D. McMillan Brian H. Davison Barbara Evans

Copyright © 2004 by Humana Press Inc. Gibberellic Acid Production All rights of any nature whatsoever reserved.0273-2289/04/114/0381–0388/$25.00

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Gibberellic Acid Production by Free and Immobilized Cells in Different Culture Systems
ENRIQUE DURÁN-PÁRAMO,* HÉCTOR MOLINA-JIMÉNEZ, MARCO A. BRITO-ARIAS, AND FABIÁN ROBLES-MARTÍNEZ
Bioprocess Department, Unit of Biotechnology (UPIBI), National Polytechnic Institute, Ave. Acueducto s/n, La Laguna, Ticomán, 07340 Mexico, D.F., E-mail:eduran@acei.upibi.ipn.mx

Abstract
Gibberellic acid production was studied in different fermentation systems. Free and immobilized cells of Gibberella fujikuroi cultures in shakeflask, stirred and fixed-bed reactors were evaluated for the production of gibberellic acid (GA3). Gibberellic acid production with free cells cultured in a stirred reactor reached 0.206 g/L and a yield of 0.078 g of GA3/g biomass.Index Entries: Cell immobilization; gibberellic acid; Gibberella fujikuroi; shake flasks; free cells; immobilized cells.

Introduction
Gibberellic acid represents one of the most important plant growth regulators produced from microorganisms owing to its extensive practical uses in agriculture. At present, the production of gibberellic acid by submerged fermentation with the filamentous fungiGibberella fujikuroi can be a practical alternative to increase the production of vegetal biomass for the production of more and better foods, fuels, and chemicals. Gibberellic acid production has been widely studied with different strains of G. fujikuroi in different fermentation systems (1–8). In the present work, we studied gibberellic acid production using free and immobilized cells of G. fujikuroiNRRL-2278 in order to select the most suitable fermentation system using this particular strain.

*Author to whom all correspondence and reprint requests should be addressed.
Applied Biochemistry and Biotechnology

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Vol. 113–116, 2004

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Durán-Páramo et al.

Materials and Methods Microorganism
G. fujikuroi NRRL-2278 obtained from the US Department of Agriculture, maintainedon potato dextrose agar (PDA) slants, and stored at 4°C.

Growth Medium and Inoculum
The culture medium proposed by González et al. (4) with a slight modification was used. The composition was glucose (25.0 g/L), KH2PO4 (5.0 g/L), NH4NO3 (1.33 g/L), and MgSO4 (1.0 g/L). The culture medium was sterilized at 121°C for 20 min. The pH of the culture medium was about 4.0 after sterilization.Inoculum (200 mL) for submerged fermentations was obtained from a 48-h shake-flask culture (120 rpm) at 30°C. The seed medium was inoculated with mycelium grown 5 d in PDA slants.

Fermentation
The microorganism was grown at 38°C within agitation of 200 rpm and a 0.3-vvm aeration in a 5-L laboratory fermentor with a working volume of 3.5-L (Bioflo II; New Brunswick Scientific). For free-cell cultures,0.7 L of preinoculum was grown in Erlenmeyer flasks for 12 h at 38°C in a rotary shaker and used to inoculate 3.5 L of medium in the fermentor.

Immobilization
The mycelium biomass recovered from 200 mL of the seed culture was mixed with 200 mL of sterile 2% (w/v) κ-carrageenan (E407; Ceca, Paris, France) at 42°C. The mycelium-carrageenan suspension was then pumped through a couple of needlesand dropped into sterile KCl solution (0.3 M) to form gel beads with an average diameter of 3 mm. Gel beads were soaked for 30 min in the same KCl solution. All immobilization experiments were carried out under aseptic conditions.

Dissolution of Beads
To quantify the immobilized biomass, gel beads were dissolved by the addition of 0.1 M sodium citrate (9).

Analysis
Reducing sugars were...