71lent to 50% dissolved air saturation (DAT). The Eycoferm con-
troller displays the DAT and not the DOT.- It is not advisable to use plates that are more than 3 weeks old
because growth in the chemostat will be slow and cells will be
more clumped in culture. - The DOT level in the culture could be above 10% DOT (10%
DOT is equivalent to 50% DAT) and will need to drop to 10%
DOT as soon as possible. High DOT levels are indicative of
poor or slow growth. It is important for the stirrer speed to
increase to disperse the cells. Once the DOT has dropped to
the set point (10%), the Eycoferm controller will inform the
stirrer to increase its speed to maintain a DOT of 10%. A con-
sistent DOT level and a high stirrer speed are indicative of
active growth. - DOT levels may fluctuate during a transition from batch to
fed-batch. - A high flow rate too early on in continuous mode may lead to
culture “wash-out”. The flow rate is measured using a device
that is constructed using a glass pipette which has been inserted
into the tubing between the medium bottle and the medium
pump via a connector with a T junction in it. The pipette is
capped with a piece of tubing and a vent filter (Fig. 1 ). The
bottom of the pipette is normally clamped off. The clamp is
removed and medium is drawn up into the pipette using a
syringe attached to the vent filter at the top of the pipette. The
medium bottle is then clamped off so that the culture
subsequently draws the medium from the pipette and not from
the medium bottle. The speed at which the culture uses the
medium from the pipette is then measured. The flow rate and
dilution rates can then be calculated. Remember to remove the
clamp from the medium bottle and replace the clamp at the
bottom of the pipette once flow rate determinations have been
completed. - A flow rate of 5 mL/h will give a dilution rate of 0.01 h−^1 and
a mean generation time of 69.3 h (slow growth). Whereas a
flow rate of 15 mL/h will give a dilution rate of 0.03 h−^1 and
a mean generation time of 23.1 h (fast growth). The flow of
medium into the vessel (F) is related to the culture volume (V)
by the dilution rate (D) where D = F/V. The volume is
expressed in mL, the flow rate is expressed in mL/h, so that
the dilution rate is therefore expressed as h−^1. Under steady-
state conditions the biomass remains constant, therefore the
specific growth rate (μ) must equal the dilution rate, i.e., μ = D.
The dilution rate is related to the doubling time (Td) by the
equation Td = loge2/D. The equation, (X = Yx/s (So−S)), has
been derived from the material balance equation of the limit-
Application of Continuous Culture for Assessing Antibiotic Activity Against...