The experiment was conducted to study the influence of potting mixture on acclimatization and growth of in vitro developed plantlets of Zygopetalum intermedium, in vitro developed plants were transferred to ten different potting mixtures viz., White Moss + Brick, White Moss + Charcoal, White Moss + Tree Fern, White Moss + Coconut Husk, Brick + Charcoall Brick + Tree Fern, Brick + Coconut Husk, Charcoal + Tree Fern, Charcoal + Coconut Husk, Tree Fern + Coco Peat in hardening house for studying the growth in ex vitro conditions for acclimatization. Even though the plantlets transferred to pots with well developed roots got established within one month, the growth differed well with potting mixture used. The plantlets grown in potting mixture comprising White Moss + Charcoal (WM+CH) recorded maximum increase in root number and root length. Increase in leaf length and root diameter was recorded highest in White Moss + Tree Fern media (WM+TF), while maximum increase in shoot length, leaf width and leaf number was recorded in potting media comprising White Moss + Bricks (WM+BR). The present experiment emphasizes combination of White Moss with Charcoal, Bricks and Tree Fern replacing the other conventionally used hardening and growing medium like cocopeat, perlite and vermiculite for Zygopetalum intermedium.