Stöckmann Christoph, Maier Ulrike, Anderlei Tibor, Knocke Christof, Gellissen Gerd, Büchs Jochen
Biochemical Engineering, RWTH Aachen University, Worringerweg 1, Sammelbau Biologie, 52056, Aachen, Germany.
J Ind Microbiol Biotechnol. 2003 Oct;30(10):613-22. doi: 10.1007/s10295-003-0090-9. Epub 2003 Oct 28.
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.
由于缺乏合适的测量技术，筛选培养通常不受监测和控制。一种用于在线测量摇瓶培养中氧传递速率（OTR）的新装置被用于监测多形汉逊酵母的筛选。在250毫升烧瓶中，300转/分钟的振荡频率和20毫升的装液量确保了0.032摩尔/（升·小时）的充足氧传递能力，从而保证呼吸不受氧限制。用0.01摩尔/升磷酸盐（pH 6.0）缓冲的培养基导致pH抑制的呼吸，而用0.12摩尔/升磷酸盐（pH 4.1）缓冲则导致不受pH抑制的呼吸。通过建立氧、铵和甘油消耗之间的固定关系，以每摩尔甘油0.245±0.015摩尔铵来平衡铵需求。用复杂葡萄糖培养基进行平板预培养会使后续使用最低甘油培养基培养时的比生长速率系数降至0.18小时-1。当使用最低甘油培养基的指数生长预培养物进行接种时，比生长速率系数增加到0.26小时-1。基于氧消耗模拟了生物量、甘油、铵和pH随时间的变化。
