FOM Institute for Atomic and Molecular Physics (AMOLF), Science Park 104, 1098 XG Amsterdam, The Netherlands.
J Am Chem Soc. 2010 Jun 23;132(24):8433-9. doi: 10.1021/ja102069d.
The digestion and absorption of lipophilic, bioactive molecules such as lipids, physiologically active nutrients (nutraceuticals), and drugs play a crucial role in human development and health. These molecules are often delivered in lipid droplets. Currently, the kinetics of digestion of these lipid droplets is followed by in vitro models that simulate gastrointestinal conditions, while phase changes within the lipid droplets are observed by light or electron microscopy. However real-time, spatially resolved information about the local chemical composition and phase behavior inside the oil droplet is not accessible from these approaches. This information is essential as the surface and phase behavior determine the local distribution of molecules in the oil droplets and thus may influence the rate of uptake, for example, by impairing the effective transfer of bioactive molecules to intestinal cells. We demonstrate the capability of multiplex coherent anti-Stokes Raman scattering (CARS) microspectroscopy to image the digestion process non-invasively, with submicrometer resolution, millimolar sensitivity, and without the need for labeling. The lipolysis of glyceryl trioleate emulsion droplets by porcine pancreatic lipase is imaged, and the undigested oil and the crystalline lipolytic products are distinguished by their different vibrational signatures. The digestion of droplets containing the phytosterol analogue ergosterol is also probed, and the crystals are observed to dissolve into the lipolytic products. The lipophilic drug progesterone and Vitamin D(3) are dissolved in glyceryl trioctanoate emulsion droplets, and the local concentration is mapped with millimolar sensitivity. The bioactive molecules are observed to concentrate within the droplets as the oil is hydrolyzed. This observation is ascribed to the low solubility of these molecules in the lipolytic products for this system. Neither the type of bioactive molecule nor the initial radius of the emulsion droplet had a large effect upon the rate of digestion under these conditions; lipolysis of the triglyceride by pancreatic lipase appears insensitive to the type of bioactive molecule in solution. These findings shed important new light on lipid digestion and open new possibilities for the chemical visualization of lipid digestion and phase changes in lipid droplets containing bioactive molecules, which in combination with other existing techniques will provide a full picture of this complex physicochemical process.
亲脂性、生物活性分子（如脂质、生理活性营养素（营养保健品）和药物）的消化和吸收在人类发育和健康中起着至关重要的作用。这些分子通常被递送到脂质滴中。目前，这些脂质滴的消化动力学是通过模拟胃肠道条件的体外模型来跟踪的，而脂质滴内的相变则通过光或电子显微镜来观察。然而，从这些方法中无法获得关于油滴内局部化学组成和相行为的实时、空间分辨信息。这些信息是必不可少的，因为表面和相行为决定了分子在油滴中的局部分布，从而可能影响吸收速度，例如，通过阻碍生物活性分子向肠道细胞的有效传递。我们展示了复用相干反斯托克斯拉曼散射（CARS）显微光谱技术的能力，该技术能够以亚微米分辨率、毫摩尔灵敏度、无需标记地非侵入性地对消化过程进行成像。通过猪胰腺脂肪酶对甘油三油酸酯乳液滴的脂解作用进行成像，并通过它们不同的振动特征区分未消化的油和结晶的脂解产物。还探测了含有植物固醇类似物麦角固醇的液滴的消化，观察到晶体溶解在脂解产物中。亲脂性药物孕酮和维生素 D(3)溶解在甘油三辛酸酯乳液滴中，并以毫摩尔灵敏度绘制局部浓度图。观察到随着油的水解，这些生物活性分子在液滴内浓缩。这种观察归因于在该体系中这些分子在脂解产物中的低溶解度。在这些条件下，生物活性分子的类型和乳液液滴的初始半径对消化速度都没有很大影响；胰脂肪酶对甘油三酯的脂解作用对溶液中生物活性分子的类型不敏感。这些发现为脂质消化提供了新的重要见解，并为含有生物活性分子的脂质滴的脂质消化和相变化的化学可视化开辟了新的可能性，这与其他现有技术相结合将为这一复杂物理化学过程提供全面的图景。
