Fruits and vegetables are generally considered as important contributors to a healthy diet and their intake is extremely helpful to reduce the risk of specific diseases like cancers, cardiovascular diseases, neural tube defects, and cataracts. Bioactive constituents from fruits and vegetables, such as carotenoids, folic acid and dietary fiber appear to play important roles in the prevention of these diseases. Carotenoids and their derivatives are versatile isoprenoids and play a vital role in plants and animals, starting from cellular antioxidant to gene regulation and so their importance at cellular and molecular level is well established. The most significant aspect of carotenoids in our diet is the antioxidant and provitamin A activity, and also the color that they impart to our food. The composition and bioavailability of carotenoids in food are significantly influenced by processing and other post-harvest technologies. This review discusses the theoretical aspects and recent developments in structural properties, biosynthesis and enhancement, processing, methods of analysis, composition in fruits and vegetables, and bioaccessibility and bioavailability of carotenoids. Additionally, future research challenges in this context are identified.