The oxygen transfer rate as key parameter for the characterization of Hansenula polymorpha screening cultures

Screening cultures are usually non-monitored and non-controlled due to a lack of appropriate measuring techniques. A new device for online measurement of oxygen transfer rate (OTR) in shaking-flask cultures was used for monitoring the screening of Hansenula polymorpha. A shaking frequency of 300 rpm and a filling volume of 20 ml in 250-ml flasks ensured a sufficient oxygen transfer capacity of 0.032 mol (l h)(-1) and thus a respiration not limited by oxygen. Medium buffered with 0.01 mol phosphate l(-1) (pH 6.0) resulted in pH-inhibited respiration, whereas buffering with 0.12 mol phosphate l(-1) (pH 4.1) resulted in respiration that was not inhibited by pH. The ammonium demand was balanced by establishing fixed relations between oxygen, ammonium, and glycerol consumption with 0.245+/-0.015 mol ammonium per mol glycerol. Plate precultures with complex glucose medium reduced the specific growth rate coefficient to 0.18 h(-1) in subsequent cultures with minimal glycerol medium. The specific growth rate coefficient increased to 0.26 h(-1) when exponentially growing precultures with minimal glycerol medium were used for inoculation. Changes in biomass, glycerol, ammonium, and pH over time were simulated on the basis of oxygen consumption.