The significance of yeast inoculation in kombucha and the environmental effects on their fermentative properties

Formålet med dette projekt var at øge alkoholprocenten i kombucha. Normalt findes hard kombucha med en ABV (Alcohol By Volume) mellem 3,5-5,5%, men ønsket var, at dette skulle forhøjes yderligere. Hensigten var at ABV’en skulle kunne måle sig med vin, hvis ABV typisk ligger mellem 11-13%. Under overvejelserne for hvordan dette bedst muligt kunne gøres, blev gær, og dets evne til at omdanne sukrose til ætanol ved fermentering, besluttet at være den optimale løsning. Gærtype og kvantitet har en signifikant betydning for kombuchas fermentering, dets smag og dets surhedsgrad, derfor blev tre gærtyper, Saccharomyces cerevisiae, Brettanomyces bruxellensis og Lachancea thermotolerans, udvalgt til forsøget. De fik tilsat henholdsvis 10% eller 20% sukrose og herefter inkuberet ved henholdsvis 16°C, 22°C og 28°C over en periode på 14 dage. For at sammenligne udviklingen på de inokulerede prøver, blev der derudover lavet referencer for at matche alle faktorer. Sidst blev forsøget gentaget for yderligere sammenligning og samtidig akkumulere en større database til brug til fremtidige studier. Der blev udført pladespredning og taget målinger for OD (optisk densitet), med det formål at få indsigt i gærvæksten. HPLC (High-Performance Liquid Chromatography) blev benyttet til måling af sukrose-, ætanol-, og eddikesyrekoncentrationerne. Forsøg 2 blev ikke som forventet, men gav interessante data for et ændret miljøs indvirkning på gærens fermenterings evner. På baggrund af det, forgrenede projektet sig ud i overvejelser omkring gærs evne til at fermentere i et surt miljø samt optimering af forsøgsdesign. Af alle de tre typer gær skilte S. cerevisiae sig ud med henblik på hurtigst sukrose forbrug, lavest akkumulation af eddikesyre og højest ætanol produktion. Det lykkedes i det første forsøg at opnå en ætanolkoncentration på 7,16%.
The purpose of this project was to increase the alcohol percentage in kombucha. Normally, hard kombucha has an ABV (Alcohol By Volume) between 3.5-5.5%, but the goal was to elevate this further. The intention was for the ABV to be comparable to that of wine, which ABV typically ranges between 11-13%. During considerations on how to achieve this most effectively, yeast and its ability to convert sucrose into ethanol through fermentation were deemed the optimal solution. As the type and quantity of yeast significantly influence the kombucha fermentation, the overall flavour and the acidity of the beverage, three types of yeast, Saccharomyces cerevisiae, Brettanomyces bruxellensis, and Lachancea thermotolerans, were selected for the experiment. They were introduced with respectively 10% or 20% sucrose and subsequently, incubated at 16°C, 22°C, and 28°C over a period of 14 days. To compare the development in the inoculated samples, references were also created to match all factors. Finally, the experiment was repeated for further comparison and obtaining a larger database to base future studies on. Plate spreading and measurements of OD (optical density) were conducted to gain insight into yeast growth. HPLC (High-Performance Liquid Chromatography) was used to measure sucrose-, ethanol-, and acetic acid concentrations. Trial 2 did not proceed as expected but provided interesting data regarding the impact of a changed environment on yeast fermentation abilities. Based on this, the project branched out into considerations about yeasts ability to ferment in an acidic environment and the optimization of the experimental design. Among all three types of yeast, S. cerevisiae stood out in terms of fastest sucrose consumption, lowest acetic acid accumulation and highest ethanol production, reaching an ethanol concentration of 7.16%.