## 3 Results and Discussion
### Impact of Feeding Strategy  
Considering the importance of the glycerol concentration on biomass growth and 1,3-PD synthesis, two different feeding strategies were tested. As in all the previous experiments, the pH was maintained using an automatic addition of glycerol-potassium hydroxide mixture. An initial glycerol concentration of 40 g/L was used in both strategies. The difference between them lied in the glycerol concentration in the fermenting medium. The first strategy assumed maintaining a relatively high glycerol concentration (approximately 25 g/L) throughout the process. In the second strategy, the concentration of glycerol was allowed to decrease to a significantly lower value of less than 15 g/L. The kinetics of both processes are presented in Figure 10, and a summary of the most important process parameters is given in Table 4. \begin{table}
\begin{tabular}{l c c c c} \hline \hline \multicolumn{5}{c}{**Stirring Rate (rpm)**} \\ \cline{2-5}
**Metabolites** & **40** & **80** & **120** & **160** \\ \hline
1,3-PD (g/L) & 35.72 \(\pm\) 1.49 a & 43.30 \(\pm\) 2.07 a & 41.39 \(\pm\) 0.15 a & 36.35 \(\pm\) 1.83 a \\ Lactic acid (g/L) & 11.96 \(\pm\) 1.49 a & 10.82 \(\pm\) 0.05 a & 10.49 \(\pm\) 0.16 a & 11.35 \(\pm\) 0.11 a \\ Acetic acid (g/L) & 8.01 \(\pm\) 0.18 a & 9.65 \(\pm\) 0.04 b & 9.76 \(\pm\) 0.16 b & 8.69 \(\pm\) 0.39 ab \\ Succinic acid (g/L) & 4.18 \(\pm\) 0.42 a & 4.85 \(\pm\) 0.04 a & 5.47 \(\pm\) 0.15 a & 4.95 \(\pm\) 0.30 a \\ Ethanol (g/L) & 1.70 \(\pm\) 0.58 a & 1.86 \(\pm\) 0.09 a & 1.66 \(\pm\) 0.30 a & 1.32 \(\pm\) 0.03 a \\ DCW (g/L) & ND & 3.03 \(\pm\) 0.14 a & 3.19 \(\pm\) 0.03 a & 3.06 \(\pm\) 0.07 a \\
3-HPA (mmol/g DCW) * & ND & 4.12 \(\pm\) 0.34 a & 3.37 \(\pm\) 0.49 a & 2.94 \(\pm\) 0.59 a \\ \hline \hline \end{tabular}  
* maximum concentration of 3-HPA achieved; Means \(\pm\) SD with different superscript letters in the same row differ significantly (\(p<\) 0.05).  
\end{table} 
Table 3: Final concentrations of metabolites (1,3-propanediol (1,3-PD), lactic acid, acetic acid, succinic acid, ethanol) and dry cell weight (DCW) in fed-batch cultures of C. _freundii_ AD119 at various stirring rate values. Figure 9: Production of 3-hydroxypropionaldehyde (3-HPA) with C. _freundii_ AD119 in fed-batch cultures with a fixed temperature of 30 \({}^{\circ}\)C and pH value of 7.0 at various stirring rate values. With the use of the first feeding strategy, a maximum concentration of 1,3-PD of 47.48 g/L was obtained with a volumetric productivity of 0.33 g/(L-h) and yield of 38.91%. Glycerol was utilized in 82.05%. The by-products obtained were as follows: lactic acid (24.29 g/L), acetic acid (9.62 g/L), succinic acid (5.95 g/L), and ethanol (3.06 g/L). The dry cell weight at the end of the process was 2.82 g/L. Using the second feeding strategy, a higher concentration of 1,3-PD was obtained (53.44 g/L) with an improved volumetric productivity (0.44 g/(L-h). Higher concentrations of by-products, with the exception of lactic acid, were also obtained. The second strategy resulted in obtaining lactic acid in a decreased concentration. The production of lactic acid is generally unwanted during the microbial 1,3-PD fermentation as it decreases the conversion yield of glycerol to the diol by reducing the amount of available NADH\({}_{2}\)[5; 37; 41]. In both cases, the rate of 1,3-PD production decreased with time which may likely be a result of the accumulation of fermentation products or the impurities contained in crude glycerol used as the carbon source [12; 27]. So far, the highest 1,3-PD concentration was obtained using the _C. freundii_ FMCC-B 294 (VK-19) strain in a fed-batch culture, and this was equal to 68.1 g/L [33]. The microbial production of 1,3-PD still attracts the attention of researchers. New studies are published