pubs.acs.org/JAFC Published on Web 06/17/2010 © 2010 American Chemical Society J. Agric. Food Chem. 2010, 58, 8085–8092 8085 DOI:10.1021/jf101325m New Mathematical Model for Interpreting pH-Stat Digestion Profiles: Impact of Lipid Droplet Characteristics on in Vitro Digestibility YAN LI AND DAVID JULIAN MCCLEMENTS* Biopolymers and Colloids Research Laboratory, Department of Food Science, University of Massachusetts, Amherst, Massachusetts 01003 The pH-stat method is commonly used to characterize the in vitro digestibility of lipids under simulated small intestine conditions. This method measures the fraction of free fatty acids (FFA) released from triacylglycerols over time. A new mathematical model has been developed to characterize the FFA versus time profiles generated by the pH-stat method, which can be used to quantify the influence of physicochemical parameters on the rate (k) and extent (φ max ) of lipid digestion. In this model, k is the amount of FFA produced per unit time per unit surface area, whereas φ max is the maximum fraction of digestible FFAs released. This model is used to quantify the influence of lipid droplet characteristics (size, concentration, composition, and emulsifier type) on the digestion of emulsified lipids. The rate (k) of lipid digestion increased with decreasing lipid content (from 2.5 to 0.5 wt %), increasing droplet diameter (from d = 200-15000 nm), and decreasing fatty acid molecular weight (MCT versus corn oil). The extent (φ max ) of lipid digestion was also considerably less for corn oil than for MCT. The rate and extent of lipid digestion did not depend strongly on initial emulsifier type: β-lactoglobulin, Tween 20, lecithin, or lyso-lecithin. These results are interpreted in terms of differences in the concentrations of reactants, products, catalysts and cofactors at the lipid droplet surfaces during digestion, for example, triacylglycerols, emulsifiers, FFA, lipase, and bile salts. This model provides a useful means of quantifying the influence of specific parameters on lipid digestion using the pH-stat method. KEYWORDS: Emulsions; lipid digestion; lipase; calcium; calcium binding; pH-stat INTRODUCTION There is growing interest in understanding and controlling the digestibility of emulsified lipids as they pass through the human gastrointestinal (GI) tract ( 1 -5 ). This knowledge is being used by the pharmaceutical industry to rationally design emulsion-based delivery systems to carry biologically active agents to specific locations within the GI tract and release them at a controlled rate ( 6 , 7 ). Analogous systems are also being developed by the food industry to encapsulate, protect, and deliver bioactive food components (nutraceuticals) ( 4 , 5 , 8 -11 ). These delivery systems have been proposed as a means of increasing the bioavailability of highly lipophilic nutraceuticals or as a means of delivering functional food ingredients to specific locations within the GI tract, such as the colon for chemopreventative agents. Emulsion- based delivery systems with controlled stability and digestibility within the GI tract have also been proposed to be an effective method of controlling appetite and therefore combating obesity ( 12 -16 ). In this case, studies have found that emulsions which remain stable to gravitational separation in the stomach and/or which have a delayed digestion in the small intestine can stimulate the release of gut hormones that induce satiety and reduce food intake. Consequently, there is great interest in the development of analytical tools that can be used to establish the major physicochemical and structural factors that affect lipid digestion and absorption under conditions that simulate the human GI tract. These tools range from physicochemical mea- surements using in vitro digestion tests, to cell culture models, to animal feeding studies, and ultimately to human trials ( 4 , 17 , 18 ). The pH-stat method is an analytical tool that is finding increasing use within pharmaceutical and food research for the in vitro characterization of lipid digestion under simulated small intestinal conditions ( 19 -22 ). The pH-stat method is based on measurements of the amount of free fatty acids (FFAs) released from lipids, usually triacylglcyerols (TAGs), after lipase addition at pH values close to neutral. The lipid-containing sample to be analyzed is placed within a temperature-controlled reaction chamber that contains simulated small intestinal fluid (SSIF). The SSIF typically includes appropriate concentrations of the major digestive components known to influence lipid digestion, such as lipase, colipase, bile salts, phospholipids, and mineral ions. The lipase in the SSIF catalyzes lipid digestion leading to the generation of two FFAs and one monoacylglycerol (MAG) per TAG molecule. The concentration of alkali (NaOH) that must then be titrated into the digestion cell to neutralize any FFA produced by lipid digestion, and thereby maintain the pH at the *Corresponding author [e-mail mcclements@foodsci.umass.edu; phone (413) 545-1019; fax (413) 545-1262].