When I use the supernatant directly without lyophilized I have the change of color by the DNS (red-brown) but whe I use the lyophilized supernatant don´t have reaction, the control and the sample have the same color (Yellow) after boil.
1. Too much salts/ unwanted chemicals/ other proteins after lyophilization of growth media interfering. So dilute it out.
2. Mixed well to solubility in DNS before boiling?
3. Usually enzymes have 'bound water' and after lyophilization, once these water molecules are lost, they do not get active ever- dead enzymes- and hence, no reactions. So, instead of lyophilization, use other salting out mechanisms to recover amylases or semi-purify them for activity measurements. Like ethanol, acetone, or other salt precipitation techniques. I guess this third one could be the major issue.
Thanks,
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