Galangal (Kaempferia galanga L.) is a herbaceous plant with significant economic value and is widely cultivated in Indonesia. The increasing demand for galangal requires efforts to provide it with good quality and quantity. This can be achieved through the expansion of cultivation areas. However, the expansion is hindered by the limited potential cultivation area available. The development of galangal varieties resistant to environmental stress has been carried out through in vitro culture with the addition of the selective agent PEG 6000, followed by further drought stress tests in the field. The objective of this research is to determine the contribution of irrigation frequency in the drought stress test to the growth and biochemical properties of galangal seedlings resulting from in vitro culture. This study utilized a Completely Randomized Design (CRD) with a non-factorial design having 5 levels and 5 replications, namely P1 = Control / normal irrigation every 2 days, P2 = Irrigation every 7 days, P3 = Irrigation every 14 days, P4 = Irrigation every 21 days, and P5 = Irrigation every 28 days. The data were statistically analyzed using a simple linear model. The results showed that the irrigation frequency contributed to the growth and yield of galangal plants; a decrease in water supply resulted in a decline in variables such as the number of leaves, number of roots, fresh rhizome weight, dry rhizome weight, fresh plant weight, and dry plant weight. The irrigation frequency had a significant contribution to the biochemical content of galangal plants, such as phenols and flavonoids. A reduction in water supply increased the flavonoid content in galangal plants undergoing drought stress.