and considerable increase in culture volume, but the volume
of the workable supernatant (approximately 7 L after
biomass removal) hardly changed during the fermentation
process (Figure 3).Since a high percentage of the culture is biomass, the
appropriate way to express angiostatin production is in mg
per liter supernatant instead of mg per liter culture, the
average production (based on five consecutive runs) was
20 (+-5) mg L
supernatant.Inducing the culture at higher cell
densities resulted in significant problems with maintaining
the desired temperature and dissolved oxygen level because
the high amounts of oxygen, needed for methanol oxidation,
generate a large amount of heat during the
exponential growth phase on methanol.When the culture density reached approximately 150
OD at 600 nm (10% v/v), about 20 h after starting the fermentation,
methanol addition was commenced and its concentration
in the culture was kept at 2 g L
-1
using the
described methanol sensor.