To understand the uptake into the body of bioactive "healthy" molecules from fruits and vegetables it is necessary to know where they are located within the original plant structures and what path they take as the plant is processed before consumption. This means not only the location but also the physical state of the molecule, e.g., crystalline or solvated, as this affects uptake. This study demonstrates how confocal Raman microspectroscopy can be used to follow changes in the physical state of carotenoids in tomatoes. We show how the three main carotenoids found in tomatoes, lycopene, β-carotene, and lutein, are distinguished from each other and how their physical state can be determined. We then show how this information can be used in situ within tomato fruit organelles, in three different tomato varieties, to locate the different carotenoids and determine their physical status. The effect of processing on carotenoids in tomatoes is also shown. This type of information can be of great benefit in availability trials to help rationalize results and define the state in which foods should be presented to the body to maximize uptake.

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