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Both the activity and conformational properties of α-amylase during the reversed micellar liquid-liquid extraction were investigated by the measurement of enzyme activity and circular dichroism (CD) probe analysis, respectively. The measurement of α-amylase activity indicated that only n -butanol, among the six n -alcohols ( n -butanol, n -pentanol, n -hexanol, n -heptanol, n -octanol, and n -decanol), was the most suitable cosolvent for Aliquat 336 reversed micelles to extract and recover α-amylase in a full reversed micellar extraction cycle. In this case, as high as 80% of the total activity of α-amylase in the initial aqueous phase could be recovered after the forward and backward extraction cycle. For the other five n -alcohols, however, the recovery of enzyme activity was relatively low. Meanwhile, the CD spectra analysis during the same extraction processes revealed some similar behaviors on the conformational change of α-amylase. In the case of n -butanol as the cosolvent, it was found that the conformation of α-amylase in the stripping solution could be restored almost completely to its native state after an extraction cycle; this implied that most of the enzyme activity would be recovered under this condition. For the other five cosolvents, it was found that the conformation of α-amylase could be restored partly to its native state after the full extraction cycle which demonstrated the fact obtained from the activity measurement experiments that only small parts of the total activity of α-amylase in the initial aqueous solution could be recovered at the end of an extraction cycle. Both the measurement of enzyme activity and CD spectra analysis proved that n -butanol was the most suitable cosolvent for Aliquat 336 reversed micelles to extract α-amylase. This observation may be helpful in developing a more effective method to monitor the process of protein reversed micellar extraction. |
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