Saliva-Induced Emulsion Flocculation: Role of Droplet Charge - Food Colloids: Self-Assembly and Material Science - page 465

breakfast and brushing their teeth, donors refrained from eating and drinking,
with the exception of water, for 2 h before donation. Saliva was collected with
closed lips for a couple of minutes and then expectorated into ice-chilled
vessels. The samples were kept constantly on ice during both donation and
handling. Saliva was pooled, centrifuged to remove cellular debris, frozen in
liquid nitrogen and stored at -801C. This pooled saliva supernatant is indicated
in the text simply as saliva.
Storage did not affect the pH of the saliva, which ranged from 6.8 to 7.1.
Protein content determination of the saliva was performed according to the
BCA method, 15 with bovine serum albumin as standard. The determined
protein content varied from 1.2 to 1.6 mg mL 1 .
32.2.3 Preparation of Emulsions and their Mixtures with Saliva
The surfactants SDS, Tween 20 and CTAB were dissolved in demineralized
water for 2 h at room temperature. The proteins b-lg and lysozyme were
dissolved overnight at 41C in demineralized water and in 10 mM NaCl,
respectively. Pre-emulsions were prepared using an Ultra-Turrax T25 Basic
(IKA-Werke) and subsequently homogenized at room temperature with 10
passes through a Delta Lab-scale homogenizer (Delta Instruments) operating
at a pressure of 70 bar or (in the case of the lysozyme emulsion) 100 bar. The
different pressure was used to obtain similar mean droplet sizes. Oil-in-water
emulsions stabilized by SDS, Tween 20, CTAB and b-lg at pH ¼ 6.7 contained
40 wt% sunflower oil and 1 wt% emulsifier. The b-lg emulsion at pH ¼ 3.0 was
prepared by lowering the pH after homogenization. The lysozyme emulsion
(pH ¼ 6.7, 1 wt% protein, 10 mM NaCl) contained 20 wt% sunflower oil.
Droplet-size distributions were obtained as averages of three measurements
performed by laser diffraction with the Mastersizer Hydro 2000S (Malvern
Instruments, Southborough, UK). Dilution with water was applied during
analysis. Zeta potential measurements of emulsions were carried out using an
Electroacoustic Spectrometer model DT 1200 (Dispersion Technology, Bed-
ford Hills, NY, USA).
The emulsion + saliva mixtures containing 10 wt% oil phase and 0.6 mg
mL 1 of salivary proteins were prepared at room temperature by adding saliva
to diluted emulsions with 10 mM NaCl in the continuous phase. The pH of the
mixtures was around 6.8 for emulsions made at pH ¼ 6.7 and around 4.3 for
the b-lg emulsion made at pH ¼ 3.0. Unless specified otherwise, the pH in the
text refers to the pH of the emulsions.
32.2.4 Characterization of the Flocculation
Flocculation was studied by light microscopy, particle-size analysis and rheo-
logy. Light microscopy images were taken using an Olympus BX 60 microscope
equipped with an Olympus DP 70 camera (Olympus Nederland, Zoeterwoude,
the Netherlands). Size-distribution analysis of the emulsion + saliva mixtures