Figure 2 Determined concentrations of extracellular glucose, lactate, and NH4+ in CHO-K1 HP and CHO-K1 LP shake flask cultures. (●, black) LP constant-temperature, (■, pink) HP constant temperature, (▼, green) LP temperature downshift, and (▲, purple) HP temperature downshift. The temperature was downshifted from 37°C to 33°C on Day 5 (vertical dashed line). (a) Extracellular lactate concentration. (b) Significance analysis of extracellular lactate concentration on days 8, 10, and 12. (c) Extracellular NH4+ concentration. (d) Significance analysis of extracellular NH4+concentration on days 8, day 10, and day 12. (e) Extracellular glucose concentration. Samples were drawn at 12-h intervals and experimental values represent three biological replicates ± SD. Two-tailed Student’s t-test was conducted using GraphPad Prism 8.0 ((*) p < 0.05, (**) p < 0.01, (***) p < 0.001 and (****) p < 0.0001).