Piper aduncum L. (Piperaceae) has great potential for economic exploration because of the proven use of its essential oil in the agriculture and in the human health. A technique that improves its propagation, as the tissue culture, becomes necessary. Some parameters must be determined for the successful cultivation in vitro. Thus, this study aimed to determine the salts concentration of MS medium, temperature, luminosity and light quality for in vitro culture of this species. The following treatments were conducted: 1/4MS, 1/2MS, MS and 2MS; 20℃, 25℃, 30℃ and 35℃; monochromatic blue, red and white lights and the combination of red and blue, using light emitting diodes (LEDs); luminosities of 17, 37, 48 and 73 μmol·m^-2·s^-1 obtained with cool white fluorescent lamp. After 35 days, the treatments were analyzed. To determine the salts concentration of MS, the variables were evaluated: number of shoots, leaves and roots, length and width of leaves, length and dry mass of root and shoots and percentage of death by necrosis. To the plantlets under different temperatures, length and dry mass of shoot and root, number of shoots, number, length and width of leaves and survival and rooting percentages were measured. The plantlets that were maintained under different luminosities were evaluated for length of shoot and root, dry mass of leaf and root. To evaluate the growth under the LEDs, the length and dry mass of shoot and root, number of shoots and roots, percentage of rooting and sprouting were assessed. The medium 1/4MS and the medium 1/2MS showed better responses for number and length of root, leaf width and shoot length. The temperature 25℃ provided the highest number of leaves, length of shoot and root, root dry mass and rooting percentage. The luminosity 73 μmol·m^-2·s^-1 provided higher values in length of root and dry mass of leaf and root. The red LED provided plantlets with greater growth. Thus, for the in vitro cultivation of Piper aduncum, 1/4MS, environment temperature of 25℃, light intensity of 73 %K Tissue Culture %K LEDs %K Monkey Pepper %U http://www.scirp.org/journal/PaperInformation.aspx?PaperID=51860