Lippia rotundifolia Cham. is in the family Verbenaceae and is endemic to the Cerrado. This species is aromatic and characterized by the presence of glandular trichomes on its leaves that are rich in monoterpenes. The objective of this study was to evaluate the growth, photosynthetic pigment production, and chemical composition of L. rotundifolia grown in vitro under different light wavelengths and intensities. The light intensities consisted of five treatments using cool white fluorescent lamps at 20, 54, 78, 88, and 110 μmol m s. The light quality consisted of six treatments using light-emitting diodes (LEDs) in different light wavelengths, namely, white, red, blue, and their interactions: 1R:1B, 2.5R:1B, and 1R:2.5B. After 45 days, the biometric parameters, photosynthetic pigment content, and volatile compounds were evaluated. The lower light intensities of 20 and 54 μmol m s generated higher growth, photosynthetic pigment content, and biomass accumulation. Myrcene and pentadecane were highest under light intensities of 88 and 110 μmol m s, respectively. The highest limonene and ocimenone levels were obtained at 20 and 54 μmol m s intensity, respectively, and the highest myrcenone content was obtained at 78 μmol m s intensity. Regarding the light wavelengths, the combination of red and blue spectra further stimulated plantlet growth, and the 2.5R:1B combination obtained the best biometric data and total chlorophyll content. The z-ocimenone chemical compound contents were highest under the 1R:2.5B light spectrum. The monochromatic blue spectrum increased the myrcene and limonene content but decreased the myrcenone content, which was increased by red light. The highest pentadecane contents were obtained with the white spectrum and the red and blue combinations.