Yeast-Lactobacillus Co-Cultures as in situ Ethanol Producers for Flavor Ester Synthesis using Lipase in Fermented Milks

Nowadays, novel biotechnological methods are preferred for flavoring productions since traditional methods include disadvantages. The aim of this study was to assess in situ biosynthesis of natural fruity flavors in fermented milks using microbial co-cultures and lipase enzyme. Materials and Methods: Trans esterification of milk fats with ethanol was carried out to develop fruity flavors in ultra-high-temperature whole milks using lipase of Palatase coupled with ethanol fermentation. Kluyveromyces marxianus NCYC 1425 was used to produce in situ ethanol in co-cultures with Lactobacillus fermentum PCC or Lactobacillus paracasei L26. Effects of co-culturing on cell viability and fermentation progress were assessed using enumeration of viable cells and measurement of pH in samples at 0, 24 (Palatase addition) and 48 h (end of fermentation). Headspace solid phase microextraction-gas chromatography (SPME)-MS/FID was used for ester, ethanol and free fatty acid analyses at the beginning and end of the fermentation. Standard curve of ethanol was used to assess the products in terms of being Halal. Results and Conclusion: Kluyveromyces marxianus included synergistic effects on Lactobacillus paracasei growth as well as antagonistic effects on Lactobacillus fermentum growth. Antimicrobial effects were seen in Kluyveromyces marxianus-Lactobacillus paracasei co-cultures when Palatase was added. Palatase significantly increased ester levels of the fermented samples. The co-cultures did not include significant differences in shorter chain ester levels (esters of 4-7 carbon chain fatty acids); in contrast, Kluyveromyces marxianus- Lactobacillus fermentum resulted in higher levels of longer chain esters. Although the Kluyveromyces marxianus cultures resulted in higher ester levels compared to that its co-cultures did, the cultures can be used as appropriate adjunct cultures with Lactobacillus cultures to boost flavor ester synthesis. This flavor synthesis can be an appropriate alternative for artificial flavoring agents.