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Midwest Biomedical Student Forum

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Birth defects are a known cause of infant death in the United States with higher rates in the Midwest perhaps due to higher use of agrichemicals for farm activities. Nitrate and atrazine are two of the most prevalent agrichemicals contaminating drinking water supplies. When ingested together, in the acidic environment of the human stomach, they can react to form a nitrosamine, NNAT. Many nitrosamines are known to be mutagenic, carcinogenic and teratogenic.


Preliminary studies have suggested that NNAT is a teratogen and mutagen. Several other agrichemical compounds have been detected in Nebraska groundwater wells that have the potential to nitrosate and studies are underway to evaluate the risk of adverse birth outcomes associated with exposure to these compounds. In this study, we evaluated growth in chick embryos after exposure to NNAT.


We hypothesize that the embryo exposed to NNAT would have delayed development compared to unexposed embryos.

Experimental design

Three groups of eggs were observed in the study: the treatment group was treated with different doses of NNAT dissolved in Dimethyl sulfoxide (DMSO), the vehicle control group were treated with only DMSO and the untreated group (control). The air sac of the treatment groups was injected with varying concentrations of NNAT at one time to stimulate an acute response and at different times for a chronic response.


All analysis was done on SAS 9.4. The effect of treatments on the weight and survival of the embryos were determined using analysis of covariance (ANCOVA) and multiple logistic regression respectively. The effect of treatment was found to be significant on the embryo treated with NNAT after adjusting for the effect of the positions of the eggs during incubation (p<0.0001). Embryos that were untreated (controls) had the highest weight (weight=0.34g), and as the dose of NNAT was increased during treatment the weight of the embryo decreased significantly after incubation for 5 days : 0.245 µmol/l of NNAT (weight=0.15g, p=0.0009), 1.11 µmol/l of NNAT (weight=0.13g, p=0.004), 2.22 µmol/l of NNAT (weight=0.11g, p=0.01) and 3.33 µmol/l of NNAT (weight=0.12g, p=0.01). Also, it was found that the odds of survival among the embryo decreased linearly with increased dose when treated with NNAT as compared to the untreated embryos: 1.11 µmol/l of NNAT vs untreated (OR=0.250 95% C.I. 0.078-0.799), 2.22 µmol/l of NNAT vs. untreated (OR=0.120 95% C.I. 0.042-0.347) and 3.33 µmol/l of NNAT vs untreated (OR=0.106 95% C.I. 0.036-0.311)


This study has revealed that weight and odds of survival of embryo exposed to NNAT decreased after 5 days of incubation. Dose-effect of the treatment was also observed during the development of embryos exposed to NNAT. Additional studies, however, is needed to evaluate the toxic effects of DMSO on the chick’s embryo before a conclusion can be reached.


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Effect of Nitrosamine (NNAT) on Embryogenesis: Evidence from a Study Using Avian Embryos Exposed to NNAT

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