Preparation of Lipid emulsions.
A lipid mixture was prepared with 97% (w}w) triolein (95%pure; Sigma),2±5% egg phosphatidylcholine (95% pure; Prolabo, Paris, France) and 0±5%free cholesterol (99% pure; Sigma) at a ®nal concentration of 16% (w}w) inchloroform} methanol (2 :1, v}v). Aliquots of the lipid mixture solution (1 mleach) were evaporated to dryness under nitrogen in 7 ml glass tubes. The buffer (with or without ®bre) was added to a ®nal volume of 4 ml. Theaqueous mixture thus contained 4% (w}w) lipids and 0±2% ®bre, i.e. in the physiological range [5,6,23,24]. The tubes were stoppered, attachedhorizontally and shaken at 200 strokes}min for 2 h at 37 °C. These operatingconditions resulted from preliminary experiments designed to discover conditions that would generate lipid emulsions with droplet sizes in the rangefound in human and rat stomach during fat digestion.
Determination of the amount of emulsi®ed lipids The upper limit for emulsion droplet size was set at 100 lm, given the instability and negligibleinterfacial area of lipid droplets above this value . To allow accuratemeasurements, [
- "%C]triolein (98% pure; 69 mCi}mmol; CEA,Gif-sur-Yvette, France) was added in trace amounts to the lipid mixture.Radioactivity was measured by liquid scintillation counting using a Packard1600TR instrument (Packard, Meriden, CT, U.S.A.) from 100 ll aliquots of two fractions collected at the end of the emulsi®cation process. One fractionwas the ¯oating oily layer composed of unemulsi®ed lipids (oily material plus lipid droplets &100 lm) which collect above the aqueous solutionsurface when tubes are left to stand for a given time (0±5±10 min at 1 g)calculated from the Stoke's sedimentation equation for droplets &100 lmunder the present conditions, as previously reported. According to Stoke'ssedimentation law, the relation between sedimentation time and particlediameter is expressed in the following equation: where
is the particlediameter (m), g! is the viscosity coefficient of the solvent (N[s}m#), q is thedensity of the sample (kg}m$), q! is the density of the solvent (kg}m$),
isacceleration due to gravity (m[s−#),
is the sedimentation time (s) and