Synergistic interactions between commonly used food additives in a developmental neurotoxicity test

Toxicol Sci. 2006 Mar;90(1):178-87. doi: 10.1093/toxsci/kfj073. Epub 2005 Dec 13.


Exposure to non-nutritional food additives during the critical development window has been implicated in the induction and severity of behavioral disorders such as attention deficit hyperactivity disorder (ADHD). Although the use of single food additives at their regulated concentrations is believed to be relatively safe in terms of neuronal development, their combined effects remain unclear. We therefore examined the neurotoxic effects of four common food additives in combinations of two (Brilliant Blue and L-glutamic acid, Quinoline Yellow and aspartame) to assess potential interactions. Mouse NB2a neuroblastoma cells were induced to differentiate and grow neurites in the presence of additives. After 24 h, cells were fixed and stained and neurite length measured by light microscopy with computerized image analysis. Neurotoxicity was measured as an inhibition of neurite outgrowth. Two independent models were used to analyze combination effects: effect additivity and dose additivity. Significant synergy was observed between combinations of Brilliant Blue with L-glutamic acid, and Quinoline Yellow with aspartame, in both models. Involvement of N-methyl-D-aspartate (NMDA) receptors in food additive-induced neurite inhibition was assessed with a NMDA antagonist, CNS-1102. L-glutamic acid- and aspartame-induced neurotoxicity was reduced in the presence of CNS-1102; however, the antagonist did not prevent food color-induced neurotoxicity. Theoretical exposure to additives was calculated based on analysis of content in foodstuff, and estimated percentage absorption from the gut. Inhibition of neurite outgrowth was found at concentrations of additives theoretically achievable in plasma by ingestion of a typical snack and drink. In addition, Trypan Blue dye exclusion was used to evaluate the cellular toxicity of food additives on cell viability of NB2a cells; both combinations had a straightforward additive effect on cytotoxicity. These data have implications for the cellular effects of common chemical entities ingested individually and in combination.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Aspartame / toxicity*
  • Benzenesulfonates / toxicity*
  • Cell Line, Tumor
  • Coloring Agents / toxicity*
  • Dose-Response Relationship, Drug
  • Drug Combinations
  • Drug Synergism
  • Food Additives / toxicity*
  • Glutamic Acid / toxicity*
  • Mice
  • Neurites / drug effects*
  • Neurites / pathology
  • Neuroblastoma
  • Quinolines / toxicity*
  • Sweetening Agents / toxicity*


  • Benzenesulfonates
  • Coloring Agents
  • Drug Combinations
  • Food Additives
  • Quinolines
  • Sweetening Agents
  • quinoline yellow
  • Glutamic Acid
  • brilliant blue
  • Aspartame