Light is a key factor that affects phytochemical synthesis and accumulation in plants. Due to limitations of the environment or cultivated land, there is an urgent need to develop indoor cultivation systems to obtain higher yields with increased phytochemical concentrations using convenient light sources. Light-emitting diodes (LEDs) have several advantages, including consumption of lesser power, longer half-life, higher efficacy, and wider variation in the spectral wavelength than traditional light sources; therefore, these devices are preferred for in vitro culture and indoor plant growth. Moreover, LED irradiation of seedlings enhances plant biomass, nutrient and secondary metabolite levels, and antioxidant properties. Specifically, red and blue LED irradiation exerts strong effects on photosynthesis, stomatal functioning, phototropism, photomorphogenesis, and photosynthetic pigment levels. Additionally, ex vitro plantlet development and acclimatization can be enhanced by regulating the spectral properties of LEDs. Applying an appropriate LED spectral wavelength significantly increases antioxidant enzyme activity in plants, thereby enhancing the cell defense system and providing protection from oxidative damage. Since different plant species respond differently to lighting in the cultivation environment, it is necessary to evaluate specific wavebands before large-scale LED application for controlled in vitro plant growth. This review focuses on the most recent advances and applications of LEDs for in vitro culture organogenesis. The mechanisms underlying the production of different phytochemicals, including phenolics, flavonoids, carotenoids, anthocyanins, and antioxidant enzymes, have also been discussed.