ABSTRACT
A 90-day subchronic oral toxicity study was conducted to evaluate the safety of a consensus bacterial phytase variant 6-phytase (PhyG) for use as an animal feed additive. This phytase is produced by fermentation with a fungal (Trichoderma reesei) production strain expressing a biosynthetic variant of a consensus bacterial phytase gene assembled via ancestral reconstruction with sequence bias for the phytase from Buttiauxella sp. Rats were administered PhyG daily via oral gavage at dose-levels of 0 (distilled water), 250, 500 or 1000 mg total organic solids (TOS)/kg bodyweight (bw)/day (equivalent to 0, 112,500, 225,000 and 450,000 phytase units (FTU)/kg bw/day, respectively). No test article-related adverse effects were observed. A no-observed-adverse-effect level (NOAEL) for PhyG was established as 1000 mg TOS/kg bw/day, the highest test concentration. Based on this NOAEL and an estimate of broiler consumption determined from the proposed inclusion of the phytase in feed at the maximum recommended level (4000 FTU/kg), a margin of safety value of 1613 was calculated. Results of in vitro genotoxicity testing and in silico protein toxin evaluation further confirmed PhyG to be non-genotoxic and not likely to be a protein toxin upon consumption. These data support the safety of PhyG as an animal feed additive.
ABSTRACT
A safety assessment was conducted for a symthetic variant Cytophaga sp. α-amylase enzyme expressed in Bacillus licheniformis and formulated into two distinct product formats: whole broth (a preparation in which the production organism is completely inactivated, but containing residual cell debris) and clarified preparation (from which the production organism is completely removed). The enzyme was improved via modern biotechnology techniques for use in the endohydrolysis of starch, glycogen, related polysaccharides and oligosaccharides. Applications range from carbohydrate processing, including the manufacture of sweeteners, fermentation to produce organic acids, amino acids and their salts, and potable or fuel alcohol, with resulting co-products (distillers' grains and corn gluten feed/meal) destined for use in animal feed. The toxicological studies summarized in this article (90-day rodent oral gavage and in vitro genotoxicity studies) noted no test article-related adverse effects and thus substantiate the safety of the α-amylase in not only the clarified form but also as a whole-broth preparation. Consistent with the decision tree analysis for enzymes produced with modern biotechnology techniques, this paper provides supporting information that this variant amylase with homology to an amylase from a potentially pathogenic organism (Cytophaga sp.) can be safely produced in an expression host that belongs to a Safe Strain Lineage, for safe use as processing aid to manufacture human and animal food.
