<p style="text-align: justify;"><b>Dietary Exposure </b></p>

<p style="text-align: justify;">Little whole kernel or processed maize is directly consumed by humans in comparison to maize-based food ingredients. Maize is a raw material for the manufacture of starch, the majority of which is converted to a variety of sweetener and fermentation products, including high fructose syrup and ethanol. Maize oil is commercially processed from the germ. These materials are components of many foods including bakery and dairy goods, and the human food uses of grain from MON810 are not expected to be different from the uses of non-transgenic field maize varieties. As such, the dietary exposure to humans of grain from insect resistant hybrids will not be different from that for other commercially available field maize varieties.</p>

<p style="text-align: justify;"><b>Nutritional Data</b></p>

<p style="text-align: justify;">Data on fatty acid profiles, protein content, amino acid composition, crude fibre, ash, phytate, and moisture content were provided for samples of MON810 grown in field trials in various locations in the United States and Europe. Comparisons of these parameters between MON810 and a non-transgenic control maize line did not reveal any biologically significant differences. The observed variations in nutritional composition were judged to arise from normal variability rather than as a result of the inserted novel traits. As a percentage of dry weight, the component analyses for line MON810, are approximately: protein 13.1%; fat 3.0%; moisture 12.4%; calories 408 Kcal/100g; ash 1.6%; and carbohydrate 82.4%.</p>

<p style="text-align: justify;"><b>Toxicity</b></p>

<p style="text-align: justify;">The trypsin-resistant Cry1Ab protein core expressed in insect-protected MON810 was identical to the same form of the protein contained in microbial <i>Bt</i> spray formulations that have been safely used in agriculture for more than 30 years. The low potential for toxicity of plant-expressed Cry1Ab protein was further demonstrated by a lack of amino acid sequence homology with known protein toxins, rapid digestion in simulated gastric juices, and lack of toxicity in feeding studies with laboratory animals. An acute oral toxicity study was done to assess the potential mammalian toxicity of Cry1Ab protein purified from <i>Escherichia coli</i> transformed with the same <i>cry1Ab</i> gene used to produce MON810. Bacterial expressed protein was used in these studies because insufficient amounts could be purified from plant tissue. Data demonstrating the molecular equivalence of bacterial and plant-expressed Cry1Ab protein were provided. The Cry1Ab core protein was administered to groups of ten male and female CD-1 mice in doses up to 4000 mg/kg body weight. These doses were well above the level of expression found in insect-protected maize plants and represented a 200-1000 fold excess over the level of exposure that would be predicted based on consumption of MON810 grain. As a control, equivalent groups of mice were administered either 4000 mg/kg bovine serum albumin or 66.66 mg/kg sodium carbonate solution (vehicle control). Clinical observations were performed and body weights and food consumption were determined. One female mouse belonging to the vehicle control died during the test &mdash; on day 1. The death of the control female was considered a result of the intubation procedure. As there were no deaths in other treated mice, or at higher exposure levels, the death was not considered to be treatment related. Mice were observed up to 9 days after dosing and no treatment related effects on body weight, food consumption, survival, or gross pathology upon necropsy were observed for mice administered the Cry1Ab test protein.</p>

<p style="text-align: justify;"><b>Allergenicity</b></p>

<p style="text-align: justify;">The Cry1Ab protein was evaluated for potential allergenicity by examining: (1) physiochemical characteristics; (2) amino acid sequence homology to known protein allergens; (3) digestibility; and (4) history of safe use of microbial insecticides containing this protein. Although the molecular weight of the Cry1Ab trypsin-resistant core protein, 63 kDa, was within the size range of known protein allergens, unlike many of these allergens it was not glycosylated. A search for amino acid sequence homology between the Cry1Ab protein and the amino acid sequences of 219 known allergens, using a database assembled from the public domain databases GenBank, EMBL, Pir and SwissProt, did not reveal any significant matches. Maize products are an important alternative to wheat flour for individuals afflicted with celiac disease, an immune mediated food intolerance for which wheat gliadins have been implicated as the causal agent. In light of the importance of maize products to these individuals, a sequence similarity search was conducted and no amino acid sequence homologies between the Cry1Ab protein and gliadins were detected. The digestibility of Cry1Ab protein was determined experimentally using <i>in vitro</i> mammalian digestion models. Purified Cry1Ab trypsin-resistant core protein (63 kDa) was added to simulated gastric and intestinal fluids and incubated at 37&ordm;C. The degradation of the protein in the digestion fluid was assessed over time by Western blot analysis and insect bioassay. In simulated gastric fluid, more than 90% of the Cry1Ab protein was degraded after 2 minutes incubation, while in simulated intestinal fluid the trypsin-resistant Cry1Ab core protein was not further degraded after more than 19 hrs incubation. This latter result was expected as serine proteases, such as trypsin, are the predominant proteolytic components of intestinal fluid. The source of the <i>cry1Ab</i> gene has a long history of use on food crops as a biopesticide and no evidence of adverse effects. This fact, combined with the lack of amino acid sequence homology between Cry1Ab protein and known allergens, and the rapid degradation of Cry1Ab protein in acidic gastric fluids, were sufficient to provide a reasonable certainty of lack of allergenic potential.</p>

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