Nitrilases catalyze nitrile hydrolysis reactions to carboxylic acids and ammonia, without the formation of free amide intermediates. For this reason, nitrilases can be used as a biocatalyst in conversion reactions (R)-4-chlorine-3-hydroxybutyronitrile (KHB) into (R)-4-chlorine-3-hydroxybutane acid, where spontaneous cycling of the product occurs in (R)-3-hydroxy-γ-butyrobactone (HBL). HBL is used in the synthesis of various polymers, solvents and pharmaceuticals as a chiral building block. Enzyme catalyzed processes are increasingly used for the synthesis of HBL because of the mild reaction conditions, high selectivity and can provide an economically viable synthesis process for this valuable chemical. While numerous, already existing chemical pathways of HBL synthesis are attempted to be used as little as possible because most of them have defects such as expensive or dangerous reactants, intense process conditions, and low utilization and selectivity, which causes the product extraction process to increase. In this work, a panel of 24 nitrilases was evaluated. During the study, the same initial conditions were used, the concentration of the substrate (KHB) and the mass concentration of nitrilase, while the reaction was monitored for 6 hours. The enzyme with the highest obtained product yield was chosen for the next step, i.e., kinetic research. Kinetic parameters of nitrilase catalyzed synthesis (R)-4-chlorine-3-hydroxybutanic acid from (R)-4-chlorine-3-hydroxybutironitrile were determined, from which the mathematical model was developed. It consists of a kinetic model and a balance of substances in the reactor. It was found that the enzyme has a low affinity for the substrate and that the reaction accompanies Michaelis-Menten's kinetics and the inhibition effect of the substrate and the product. The model was successfully validated by the implementation of the reaction in six different batch reactors with different initial conditions in the reactor. Samples of the reaction mixture were analyzed on a high-efficiency liquid chromatograph and a gas chromatograph. From these experiments, the deactivation constant was evaluated and it was concluded that the initial conditions in the reactor affect the stability of the enzyme.