Polyamine (PA) metabolism was studied in liquid cultures of Scots pine (Pinus sylvestris L.) embryogenic cells. The focus of the study was on the metabolic changes at the interphase between the initial lag phase and the exponential growth phase. PA concentrations fluctuated in the liquid cultures as follows. Putrescine (Put) concentrations increased, whereas spermidine (Spd) concentrations decreased in both free and soluble conjugated PA fractions. The concentrations of free and soluble conjugated spermine (Spm) remained low, and small amounts of excreted PAs were also found in the culture medium. The minor production of secondary metabolites reflected the undifferentiated stage of the embryogenic cell culture. Put was produced via the arginine decarboxylase (ADC) pathway. Futhermore, the gene expression data suggested that the accumulation of Put was caused neither by an increase in Put biosynthesis nor by a decrease in Put catabolism, but resulted mainly from the decrease in the biosynthesis of Spd and Spm. Put seemed to play an important role in cell proliferation in Scots pine embryogenic cells, but the low pH of the culture medium could also, at least partially, be the reason for the accumulation of endogenous Put. High Spd concentrations at the initiation of the culture, when cells were exposed to stress and cell death, suggested that Spd may act not only as a protector against stress but also as a growth suppressor, when proliferative growth is not promoted. All in all, Scots pine embryogenic cell culture was proved to be a favourable experimental platform to study PA metabolism and, furthermore, the developed system may also be beneficial in experiments where, e.g., the effect of specific stressors on PA metabolism is addressed.