Safety evaluation of a fungal beta-glucanase

Pig and poultry feed are often included with beta-glucanase to alleviate the anti-nutritional impacts of beta-glucans found in many cereal grains. Safety studies were conducted on beta-glucanase, including skin and eye irritation, dermal sensitization, a 90-day rat oral subchronic study, and genotoxicity, to evaluate the safety of a beta-glucanase for use as an animal feed additive. Beta-glucanase is produced by fermentation with a fungal (Trichoderma reesei) production strain expressing a wild-type fungal endoglucanase (egl2) gene to overexpress the endoglucanase (referred to as beta-glucanase throughout the paper) enzyme. Beta-glucanase was predicted to be non-irritant (No Category) to the skin or eyes in reconstructed human epidermis tissues (RhE) and reconstructed human cornea-like epithelium (RhCE), respectively. Beta-glucanase was tested in vitro in the direct peptide reactivity assay (DPRA), the KeratinoSens™ assay and the human cell line activation test (h-CLAT). Using the 2 out of 3 approach, beta-glucanase was not a dermal sensitizer based on the negative KeratinoSens™ and h-CLAT assays. In vitro genotoxicity testing confirmed beta-glucanase to be non-genotoxic. In the 90-day subchronic study, rats were administered beta-glucanase daily via oral gavage at dose-levels of 0 (Milli-Q® water), 250, 500 or 1000 mg total organic solids (TOS)/kg bodyweight (bw)/day (equivalent to 0; 42,102; 84,205 and 168,410 units (U)/kg bw/day, respectively). No test item-related adverse effects were observed. A no-observed-adverse-effect level (NOAEL) for beta-glucanase was established at 1000 mg TOS/kg body weight/day, the highest test concentration. Based on this NOAEL and an estimate of consumption determined from the proposed inclusion of the beta-glucanase in feed at the maximum recommended level (426 U/kg), a margin of safety value of 5681 was calculated based on highest expected daily feed intake in broilers, the target species with highest feed intake relative to body weight. The findings support the safe use of beta-glucanase as an animal feed additive.