The Impact of Process Parameters on 1,3-Propanediol Production and 3-Hydroxypropionaldehyde Accumulation in Fed-Batch Fermentation of Glycerol with _Citrobacter freundii_ AD119  
[PERSON]  
1  
[PERSON]  
2  
[PERSON]  
2  
[PERSON]  
1 Department of Biotechnology and Food Microbiology, Faculty of Food Science and Nutrition, Poznan University of Life Sciences, 60-624 Poznan, Poland  
2  
Footnote 10: emailthe pH-stat fed-batch culture, the addition of substrate is coupled to the delivery of a neutralizing agent used to maintain the pH of the broth.  
Many species of bacteria have the ability to produce 1,3-PD, including those belonging to the genera of _Clostridium_, _Klebsiella_, _Citrobacter_, _Enterobacter_, and _Lactobacillus_[8; 9; 10; 11; 12]. In 1,3-PD producers, the catabolism of glycerol consists of two coupled branches--one oxidative, the other reductive. In the oxidative branch, glycerol is transformed to dihydroxyacetone, which then undergoes glycolysis to form various end-products, such as lactic acid, acetic acid, ethanol, etc. In the reductive pathway, glycerol is converted to 3-hydroxypropionaldehyde (3-HPA), which is then reduced to 1,3-PD. The reductive pathway maintains the redox balance of the cell, as it uses NADH\({}_{2}\) and regenerates NAD\({}^{+}\)[4; 13; 14].  
It is worth emphasizing that the intermediate of the reductive branch, 3-HPA, belongs to a multi-component antimicrobial system called reuterin. It is a dynamic system which mainly consists of 3-HPA, its dimer, hydrate, and acrolein. The health-promoting properties of reuterin have been demonstrated, which include anti-infective, anti-inflammatory, and anti-carcinogenic activities. Moreover, the strong antimicrobial activity of this system makes it suitable for preserving various types of food [15]. However, the accumulation of 3-HPA during the microbial production of 1,3-PD leads to negative consequences which include cell growth limitation and premature cessation of the process. There are numerous reports on this phenomenon [1; 3; 16; 17; 18; 19]. It is known that 3-HPA induces oxidative stress in the cells, and its dimer inhibits the synthesis of DNA [18; 20]. [PERSON] et al. [17] noticed differences in the accumulation of 3-HPA in various microorganisms and its dependence on the concentration of glycerol. Premature cessation of fermentation was observed at elevated glycerol concentrations for _Enterobacter agglomerans_, _Klebsiella pneumoniae_, and _Citrobacter freundii_ strains. No accumulation of 3-HPA, however, was found to occur in cultures of _Cistoridium butyricum_[1].  
The stirring rate and pH of the medium can also influence the kinetics of 1,3-PD fermentation. [PERSON] et al. [19] noticed that by using a higher stirring rate, a higher 1,3-PD productivity and a decreased 3-HPA concentration can be achieved in _K. pneumoniae_ cultures. According to [PERSON] et al. [1], when a pH value of the culture medium of 8.0 was used, the production of 3-HPA in _E. agglomerans_ did not affect fermentation kinetics, as it was low and transient.  
There is scarce information, however, on the impact of culture conditions on 3-HPA and 1,3-PD production with _C. freundii_. For this reason, optimization of selected fermentation parameters such as temperature, stirring rate, and pH with respect to 1,3-PD production and 3-HPA accumulation with _C. freundii_ AD119 was conducted in this study. Additionally, two feeding strategies were examined in a pH-stat fed-batch culture of this microorganism performed under optimized fermentation conditions.