Optimization of nutrient medium for biosynthesis of gibberellic acid

Abstract

A new method of optimization of the medium composition by biostatistical methods is reported. The well-known BOX-WILSON scheme combined with the method of optimum curves was applied for an exploration of the response surface. The primary goal has been to optimize the concentration of nutrients for the production of gibberellic acid by the fungus Gibberella fujikuroi with a minimum number of experiments. As the result, a nearly fivefold improvement yield of the was achieved performing only two experiments.Gibberellic acid (GA,) and some other biological active gibberellins are produced by the phytopathogenic fungus Gibberella fujikuroi (SAW) WOLL. (Fusarium moniliforme SHELD) when grown in submerged culture in nitrogen-limited media.Gibberellins are typical secondary metabolites. The phases of growth can be discerned, described and related to the nutritional and environmental state of the fermentation (BORROW etal., 1961, 1964, VASS and JEFFERYS, 1979. -Significant production ,of gibberellins starts only after nitrogen exhaustion. On the other hand, production is greater the higher the value of initial nitrogen over a wide range. Further increases in nitrogen concentrations lead to a decrease of the mycelium productivity (BORROW et al., 1964). Therefore, the selection of both the initial concentration of nitrogen and of an optimal C/N ratio are the most important factors.The object of this paper was the improvement of the medium composition by mathematical methods for planning the experiment.In an unconventional manner the Box-WILSON scheme is used for the optimization of a nutrient medium composition consisting of four nutrients -corn steep liquor (CL), ammonium sulfate (AS), sunflower oil (SO), and potassium dihydrogen phosphate (PP). Originally, the BOX-WILSON method was developed for the optimization of chemical processes (Box and WILSON 1951). To optimize a biological response such as the biosynthesis of gibberellins and other secondary metabolites, some modifications according design and analysis are needed to make the BOX-WILSON procedure more effectively. A combination with the method of optimum curves developed by SCHRODER and WEIDE (1973) seems to be very helpful to avoid an overdimensional expansion of experimental effort. In the following by the example of gibberellic acid formation an appropriate methodical basis is given for the application of factorial designs in fermentation. As the rule a consulting statistician solving problems in fermentation is confronted with four problems which are typical for the optimization of biological response. I The great variation of experimental results after repetition; I1 The bad reproducibility of fermentation experiments over the time; I11 The overdimensional effort of the factorial design (2k or 2k-p) as a consequence of the high-dimensional parameter space; IV The violation of the smoothness hypothesis underlying the response surface technique. 16'

Synthesis of Indoleacetic Acid, Gibberellic Acid and ACC-Deaminase by Mortierella Strains Promote Winter Wheat Seedlings Growth under Different Conditions

“…The concentration of IAA was calculated from the regression equation of a standard curve of pure indole-3-acetic acid (Sigma-Aldrich, St. Louis, MI, USA. To quantify the GA using the modified method of Brückner et al [ 35 , 54 ] and Hasan [ 44 ], the pH of filtrates was adjusted to 2.8 with 1 M HCl and extracted three times with equal volumes of ethyl acetate. The ethyl acetate fractions were collected and vacuum-evaporated (using Eppendorf Concentrator plus) at 20 °C.…”

Gibberellin and auxin-indole production by plant root-fungi and their biosynthesis under salinity-calcium interaction

“…GA and IAA were spectrophotometrically determined according to Bruckner et al [13] and Wohler [14], respectively. The ethyl acetate layer was dried over Na 2 SO 4 , and evaporated to dryness in a rotary evaporator.…”

Gibberellin and auxin production by plant root-fungi and their biosynthesis under salinity-calcium interaction

“…IAA showed violet-red colour in VL and orange in UV light after spraying. GA and IAA were spec- trophotometrically determined according to Bruckner et al (1991) and Wohler (1997), respectively. The values obtained were computed from the standard curves of GA 3 and IAA.…”