effects and interactions of glucose, cell and benzaldehyde initial concentrations. In this design, a set of 11
experiments, including three replicates at the central point, was performed. The range and the levels of the
variables herein investigated are given in Table 2. “STATISTICA” (version 7.0) software was used for
regression and graphical analyses of the data obtained.
3. Results and Discussion
Identification of important medium constituents was performed using Plackett–Burman design (PB-12).
This statistical tool was selected for the optimization of culture medium to increase the production of LPAC
and reduce producing costs by minimizing the salts added to the medium in shake flasks. Table 1
shows the results for the experimental design. A p-value inferior to 0.05 for the three variables viz Peptone
(X1), Na2HPO4.12H2O (X5) and citric acid (X6) indicates that these are significant variables for L-PAC
production, which can be visualised in the Pareto chart (Figure 2a). The results indicate that it is possible
to minimize the concentrations of peptone from 20 to 5 g/L, CaCl2.2H2O from 0.05 to 0.01 g L and Citric
acid from 10.7 to 2 g/L. The Pareto chart for L-PAC production (Figure 2a) shows that the concentration of
MgSO4.7H2O is not statistically significant. However, when 1 g / L of MgSO4.7H2O is used L- PAC
production is rather low (1.02 g/L) compared with production of L -PAC (2.07 g/L) with 5 g/L. Although the
statistical analysis shows that MgSO4.7H2O do not influence the production of L-PAC, the ion Mg+2 is very
important to favour the metabolic pathway to L-PAC production since it is required for the enzymatic
transformation of benzaldehyde and pyruvate in L- PAC, avoiding the formation of benzyl alcohol.
The optimal salt composition determined by PB- 12 was MgSO4.7H2O 5 g/L, CaCl2.2H2O 0.01 g/L,
Na2HPO4 .12 H2O 35 g/L, citric acid 2 g/L and benzaldehyde 4 g/L.