Purification and Identification of Bovine Cheese Whey Fatty Acids Exhibiting In Vitro Antifungal Activity

Inhibition of Candida albicans germination in different inducing media by bovine whey chain FFA-enriched fractions. Blastospores of the Candida albicans HWP1-LacZ strain (5×10⁵spores/mL) were induced to germinate at 37°C during 4h in dextrose-free Sabouraud, Spider, or Lee's media. Each of the FFA fractions was used at a concentration of 10μg/mL. (A) The percentage of germination represents the level of β-galactosidase expression, which is under the control of the HWP1 promoter expressed in hyphae, of cells incubated with samples to sample free control and corrected for biomass using XTT. *Indicates P<0.02 and ** indicates P<0.005 when compared with the sample-free control. (B) Microscopic observations of cells after 4h of incubation at 37°C in different inducing media.

Antifungal activity of bovine non-straight-chain chain FFA-enriched fraction. (A) Blastospores of Candida albicans (5×10³spores/mL) incubated at 37°C for 48h in dextrose-free Sabouraud liquid medium containing FFA, non-straight-chain FFA, or straight-chain FFA at the indicated concentrations. (B) Spores of A. fumigatus (5×10³spores/mL) were incubated at 37°C for 48h in Sabouraud liquid medium supplemented with the indicated concentrations of FFA fractions. The growth inhibition (%) represent the reduction of absorbance at 595nm of spores incubated with samples to sample free control. Error bars represent standard deviations for fourth experiments.

Separation of the non-straight-chain FFA enriched fraction using a reversed-phase Nova-Pak HR C18 semi-prep HPLC column (7.8×300mm, 6μm). Non-straight-chain FFA dissolved in 50% ethanol was injected manually onto the column and compounds eluting with an increasing concentration of acetonitrile were detected at 215nm (milli-arbitrary units, mAU). The flow rate was 8mL/min, and each detected peak was manually collected. Fractions exhibiting antifungal activity are indicated.