A bacterial strain having higher nitrile hydratase activity, named as $\underline{Brevibacterium}$ sp. CH2, was developed by repeating the cultivation of $\underline{Brevibacterium}$ sp. CHl in the broth containing 1.75%(v/v) acrylonitrile. The properties of the strain was greatly improved. The specific nitrile hydratase activity of the strain increased up to 36 units/mg of dry cell weight, more than 3.6 times higher than that of $\underline{Brevibacterium}$ sp. CHl and the enzyme was not inhibited by the substrate, acrylonitrile concentration of 6% (v/v).
The effects of temperature and pH on the activity of free and immobilized cells were investigated. The substrate specificity and the enzyme reaction kinetics such as the substrate and product inhibition were also studied. The conversion yield was nearly 100% with a trace amount of acrylic acid produced. The strain had high nitrile hydratase activity for acrylonitrile, but amidase activity for acrylamide was negligible.
In batch cultivation the maximum specific growth rate of 0.24 $h^{-1}$ was obtained at a glucose concentration of 15 g/L and the final cell density 15.4 g/L. In fed-batch culture the cell density increased up to 80 g of dry cell mass/L by regulatory substrate feeding.
Continuous production of acrylamide was carried out using the whole cells of $\underline{Brevibacterium}$ sp. CH2 immobilized with polyacrylamide in a two-stage packed bed reactor. The volumetric productivity of the reactor was 93.2 g/L.h with more than 50% conversion. For maximum acrylamide productivity, acrylonitrile concentration and the reaction temperature were 6%(v/v) and 4℃ respectively.
