A good cysteine addition method that could increase the specific glutathione (GSH) production rate (Pc) was investigated and utilized to maximize total GSH production in fed-batch culture of Saccharomyce s cerevisiae. The single-shot addition of cysteine was a better method compared to a continuous method that maintained a constant cysteine concentration in the.. reactor. The shot method increased Pc about twofold compared to a culture without cysteine. The increase in Pc by the shot method can be achieved without growth inhibition if the cysteine dose is maintained at 0.7 mmol.g-1 cell or less. The positive effect on Pc (at every specific growth rate,/z) was saturated when the cysteine shot concentrations was 3 mM or more. A simple model was developed consisting of mass balance equations and the relationship between p and Pc, for the single cysteine shot addition method. From this model an optimal operating strategy was determined to maximize total GSH production in fed-batch culture. This optimal operation consisted of separating the process into phases of (1) cell growth and (2) GSH production, through a bang-bang profile control of/z, and a shot of cysteine just at the start of the GSH production phase. In other words, the cysteine shot time and the ~ switching time should be the same. For a total feeding time of 10 h, both the switching time of p and cysteine shot time were calculated to be about 6.4 h.

Regardless of interaction with less frequent imitative caregivers, infants can obtain the vowels of the caregivers' mother tongues by finding the correspondence between their own vowels and the caregivers' vowels. This paper proposes a learning method based on auto-mirroring bias (AMB) with a self-evaluation mechanism to find such correspondence. AMB is the robot's anticipation of being imitated by its caregiver and has a role of narrowing the candidates for the correspondence. The self-evaluation mechanism biased by AMB works as outlier (incorrect mapping) rejection, expecting that the outliers appear less consistently than the correct mappings do in the interaction. Results from several computer simulations with real sound wave recording from a human experimenter show that the robot could successfully achieve being imitated by the proposed method even if interacting with a caregiver who would seldomly imitate its utterances.

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A fuzzy logic controller (FLC) for the control of ethanol concentration was developed and utilized to realize the maximum production of glutathione (GSH) in yeast fedbatch culture. A conventional fuzzy controller, which uses the control error and its rate of change in the premise part of the linguistic rules, worked well when the initial error of ethanol concentration was small. However, when the initial error was large, controller overreaction resulted in an overshoot.An improved fuzzy controller was obtained to avoid controller overreaction by diagnostic determination of "glucose emergency states" (i.e., glucose accumulation or deficiency), and then appropriate emergency control action was obtained by the use of weight coefficients and modification of linguistic rules to decrease the overreaction of the controller when the fermentation was in the emergency state. The improved fuzzy controller was able to control a constant ethanol concentration under conditions of large initial error.The improved fuzzy control system was used in the GSH production phase of the optimal operation to indirectly control the specific growth rate mu to its critical value micro(c). In the GSH production phase of the fed-batch culture, the optimal solution was to control micro to micro(c) in order to maintain a maximum specific GSH production rate. The value of micro(c) also coincided with the critical specific growth rate at which no ethanol formation occurs. Therefore, the control of micro to micro(c) could be done indirectly by maintaining a constant ethanol concentration, that is, zero net ethanol formation, through proper manipulation of the glucose feed rate. Maximum production of GSH was realized using the developed FLC; maximum production was a consequence of the substrate feeding strategy and cysteine addition, and the FLC was a simple way to realize the strategy.

The physiological states with respect to cell growth and ethanol production in a yeast fed-batch culture expressed in linguistic form could be recognized on-line by fuzzy inferencing based on error vectors. The error vector was newly defined here in a macroscopic elemental balance equation. The physiological states for cell growth and ethanol production were characterized by error vectors using many experimental data from fed-batch cultures. Fuzzy membership functions were constructed from the frequency distributions of the error vectors and state recognition was performed by fuzzy inferencing. In particular, an unusual physiological state for a yeast cultivation, in which aerobic ethanol production was accompanied by very low cell growth, could be recognized accurately. According to the results of the state recognition, an energy parameter, the P/O ratio in the metabolic reaction model was adaptively estimated, and the cell growth was successfully evaluated with the estimated P/O. (c) 1995 John Wiley & Sons, Inc.