193
The aim of the present thesis was that to evaluate the use of selected Lactic Acid Bacteria (LAB) to improve quality (through the increasing in folic acid concentration) and safety against food fungal spoilage and foodborne causes, (especially in the dairy food), for the production of food product in the developing countries. For this reason different researches were performed, testing the ability of selected LAB in the production of folic acid and to protect food against the growth of filamentous fungi and food pathogens. Detailed researches were focused on the characterization of the inhibitory compound produced by LAB in milk, for characterize the bacteriocin production during the fermentation process. The economical and easily use of LAB during the food production represent other important further strengths for the food company. Finally together with what has just been stated, the use of bio-product during the food production, such as microorganisms like LAB, can be the answer for the higher demand posted by the consumers for the production of food without ingredients of synthetic origins.
The first part of the thesis was focused on the analysis of the production on folic acid in milk, by selective St. thermophilus and L. delbrueckii subsp. bulgaricus.
Milk is a food with naturally low free folate intake (average 26µg/L), but still play an important role in the diet. In the research it was demonstrated that the folate concentration in milk can been increased by using selected LAB strain with folic acid production ability. St. thermophilus represent the strain with the higher ability on the folate production during the fermentation process, but it was shown that the production is still strain dependent and it can change during the early hours of fermentation steps process. It was also demonstrate that the combination with strain of L. delbrueckii subsp. bulgaricus, for the production of fermented milk, can decrease significantly the folate concentration previously product by the St.
thermophilus strain, caused by a folate consumption by L. delbrueckii subsp.
bulgaricus. But the results are demonstrating that the pre-selection and an in vitro evaluation of the folate production by LAB represent the better system to find the
194
right combination of the bacteria to obtain the optimal increasing production of folic acid in milk. Finally, the combination of the two LAB strains can determine a higher final concentration in folate (average increased 100 time’s). In the last few years researches shown that the use of folate synthetically produced and use during the food enriching, can be responsible of disease and health problems. The use of LAB with ability to increase the folate amount in milk can represent the right solution for this problem.
The second part of the thesis was focused on the study of the inhibition ability of LAB against fungal food spoilage and pathogens growth in food. In vitro and fermented milk/yogurt assay were performed for the studies. Further studies showed the ability of some LAB to produce lactic acetic, carbon dioxide and other compound able to inhibit the growth of target microorganism. The researchers conducted in the present thesis were pointed on the identification of the production of probable antimicrobial compounds, such as bacteriocins. For that the pH was monitored and the growth rates of the single specie were collected. For the study selected LAB with hypothetical antimicrobial activity were use, Lactobacillus plantarum UC8491 and Lactobacillus rhamnosus UC8490. The first study was effectuated to evaluate the inhibition activity against two species of Penicillium spp., common filamentous fungi responsible of food spoilage. The results shown that the two LAB were able to inhibit the growth of the moulds, both if they was inoculated individually or in combination in fermented milk, showing an higher ability on the mould inhibition. Statistical analysis (probabilistic model of multinomial logistic regression) was made run and the solution demonstrate that all the results obtained were statistical positive, indicating the good inhibitory activity performed by the LAB. pH analysis and inhibition rate of the filamentous fungi give the possibility to speculate the presence of antimicrobial compound, produced by the two LAB strain.
195
Inhibitory test were performed against food pathogen to. The test were made using the LAB strain Lactobacillus plantarum UC8491 and Lactobacillus rhamnosus UC8490, used for the test against the fungal spoilage. Preliminary test with commercial fermented milk wad made, and resistance of the pathogens to the fermentation was evaluated. Results shown a different attitude on the resistance on the low pH obtained during the fermentation, from the pathogen strain used during the experiment. Survival rate of the pathogen were subsequently obtained on fermented milk obtained with the inoculum of the two LAB strains, showing a positive inhibitory activity in the first 12 hours of the fermentation process, emphasizing the antimicrobial ability of the two LAB.
The final part of the thesis were focused on the characterization of the antimicrobial compound produced by the LAB strain used during the inhibitory test, Lactobacillus plantarum UC8491 and Lactobacillus rhamnosus UC8490.
Proteomic approach on the genome sequence of the tow LAB was made. The use of different database software was necessary for the evaluation. The results confirm the hypothesis that the LAB selected for the antimicrobial test, have the ability to produce bacteriocin. Different types of bacteriocin were identified. Peptide homologs to plantaricin bacteriocin were identified, but other peptide belonging to bacteriocin not strictly related to the LAB was recorded. The hypothesis for the presence of this peptide was made, according to other authors, but lower is actually known about the presence of homologues peptide encoding for different bacteriocins. Results obtained are important to demonstrate that the LAB strain are able to produce bacteriocin and have the gene to encode peptide of several different bacteriocins with different spectrum of activity, but more studies are necessary to understand all the characteristics of the bacteriocins produced by the lactic acid bacteria.
For conclude, the thesis have shown the probiotic activity of selected LAB strain and their role for the human nutrition, through the production of extracellular