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Exposure to Flavoring Chemicals

What are Flavorings?

Flavorings are often complex mixtures of natural and man-made substances. The U.S. Food and Drug Administration (FDA) evaluates flavoring ingredients to determine whether they are “generally recognized as safe” (GRAS) to be eaten. Even if they are safe to eat, these ingredients might still be harmful to breathe in the forms and amounts to which food and chemical industry workers may be exposed. Given the complexity of flavorings mixtures, and the lack of health data for many of the component materials, identifying the relative contributions of individual substances to causing flavoring-induced lung disease is a difficult challenge. As noted in the NIOSH Alert: Preventing Lung Disease in Workers Who Use or Make Flavorings, the flavorings industry has estimated that over a thousand flavoring ingredients have the potential to be respiratory hazards due to possible volatility and irritant properties (alpha, beta-unsaturated aldehydes and ketones, aliphatic aldehydes, aliphatic carboxylic acids, aliphatic amines, and aliphatic aromatic thiols and sulfides).


Diacetyl is a chemical that was found to be a prominent volatile constituent in butter flavoring and air at the microwave popcorn plant initially investigated by NIOSH. Diacetyl is also known as the alpha-diketone, 2,3-butanedione, or by its Chemical Abstracts Service (CAS) number, 431-03-08. NIOSH has published a comprehensive document describing diacetyl and 2,3 pentanedione entitled Criteria for a Recommended Standard: Occupational Exposure to Diacetyl and 2,3-Pentanedione.

Workers in microwave popcorn manufacturing are exposed to many materials besides diacetyl. Thus, NIOSH’s initial studies in a total of six microwave popcorn plants were not able to definitely determine if diacetyl exposure contributed to lung disease or was a marker for other hazardous substances that contributed to disease. Still, NIOSH studies in the initial plant documented a relationship between cumulative exposure to diacetyl vapor over time and having abnormal lung function as measured by spirometry. Higher cumulative exposure to diacetyl in this plant was associated with having a lower level of forced expiratory volume in 1 second (FEV 1), an important measure of lung function. Across all six microwave popcorn plants studied by NIOSH, working as a mixer of butter flavorings and heated soybean oil was associated with higher exposure to diacetyl vapor than working in other areas of the plants. People who had ever worked as mixers had more chest symptoms and poorer lung function as measured by lower FEV 1 than people who had never worked as mixers. People who had worked as mixers for more than 12 months had more shortness of breath with exertion and lower FEV 1 than people who had worked as mixers for less than 12 months.

Subsequent studies have helped to clarify the role of diacetyl in substance toxicity. Toxicology studies have shown that vapors from heated butter flavorings can cause damage to airways in animals (Hubbs et al. 2002). Studies in both rats and mice demonstrate that the cells lining the airways can be damaged by inhaling diacetyl vapors as a single agent exposure in both acute and sub-chronic studies (Hubbs et al. 2008, Morgan et al. 2008).  In mice, aspiration of diacetyl alone caused a pattern of injury that replicates some of the features of human obliterative bronchiolitis (Morgan et al. 2008).  In addition, inhaling either diacetyl or the related flavoring, 2,3-pentanedione can cause an obliterative bronchiolitis-like condition in rats (Morgan et al. 2016).

Furthermore, diacetyl reacts with proteins and a recent study demonstrating striking changes in protein homeostasis in the airways of diacetyl-exposed mice strongly implicates protein damage in airway epithelium as the mechanism for diacetyl-induced airway injury (Hubbs et al. 2016).  Dosimetry studies indicate that at a given exposure concentration, a much greater concentration of diacetyl can reach the deep lung of humans than reaches the deep lung of rats (Gloede et al. 2011Morris and Hubbs. 2009). These findings support the hypothesis that diacetyl vapors are an inhalation hazard in the workplace. Also, chemical workers in a plant that manufactured diacetyl and coffee workers exposed to diacetyl developed the same type of lung disease as microwave popcorn workers (van Rooy et al. 2007 and 2009). The chemical workers had less complicated exposures than microwave popcorn workers. Overall, current evidence points to diacetyl as one agent that can cause flavorings-related lung disease.


The alpha-diketone, 2,3-pentanedione, has received attention as a flavoring substitute for diacetyl. It is also known as acetyl propionyl or by CAS number 600-14-6. It is structurally very similar to diacetyl since 2,3-pentanedione is a 5-carbon alpha-diketone and diacetyl is a 4-carbon alpha-diketone. Recent mechanistic studies implicate the alpha-diketone functional group in the airway toxicity of diacetyl (Morgan et al. 2016, Hubbs et al. 2016).

Reports of 2,3-pentanedione toxicity were first published in 2010 (Hubbs et al. 2010a,  Morgan et al. 2010). A follow-up full length publication demonstrated that acute inhalation exposures to 2,3-pentanedione cause airway epithelial damage that is similar to diacetyl in laboratory studies (Hubbs et al. 2012). In longer 2-week inhalation studies in rats, researchers from the National Institute of Environmental Health Sciences (NIEHS) found that repeated exposures to either 2,3-pentanedione or diacetyl can cause airway fibrosis, including obliterative bronchiolitis-like changes, in rats (Morgan et al. 2016). In the acute inhalation study of 2,3-pentanedione, changes in gene expression were noted in the brain (Hubbs et al. 2012). Similarly, diacetyl can cause changes in gene expression and other markers of damage in the olfactory bulb of the mouse brain (Hubbs et al. 2016).

Other Alpha-diketones

As a group, these publications raise concerns that the toxicologic effects of diacetyl may be shared with close structural analogs used in food manufacturing such as 2,3-hexanedione and 2,3-heptanedione (Day et al. 2011). The 6-carbon alpha-diketone, 2,3-hexanedione is less chemically reactive than diacetyl or 2,3-pentanedione, and only 2 of 12 rats had bronchial fibrosis after a 2-week inhalation exposure to 2,3-hexanedione (Morgan et al. 2016).


Criteria for a Recommended Standard: Occupational Exposure to Diacetyl and 2,3-Pentanedione – DHHS (NIOSH) Publication Number 2016-111

Day G, LeBouf R, Grote A, Pendergrass S, Cummings K, Kreiss K, and Kullman G [2011]. Identification and measurement of diacetyl substitutes in dry bakery mix production. J Occ Env Hygiene 8(2):93-103.

Gloede, E., Cichocki, J.A., Baldino, J.B. and Morris, J.B. (2011). A validated hybrid computational fluid dynamics-physiologically based pharmacokinetic model for respiratory tract vapor absorption in the human and rat and its application to inhalation dosimetry of diacetyl. Toxicol Sci, 123, 231-246.

Hubbs AF, Battelli LA, Goldsmith WT, Porter DW, Frazer D, Friend S, Schwegler-Berry D, Mercer RR, Reynolds JS, Grote A, Castranova V, Kullman G, Fedan JS, Dowdy J, Jones WG [2002]. Necrosis of nasal and airway epithelium in rats inhaling vapors of artificial butter flavoring. Toxicol Appl Pharmacol 185(2):128–135.

Hubbs, A.F., Fluharty, K.L., Edwards, R.J., Barnabei, J.L., Grantham, J.T., Palmer, S.M., Kelly, F., Sargent, L.M., Reynolds, S.H., Mercer, R.R., Goravanahally, M.P., Kashon, M.L., Honaker, J.C., Jackson, M.C., Cumpston, A.M., Goldsmith, W.T., McKinney, W., Fedan, J.S., Battelli, L.A., Munro, T., Bucklew-Moyers, W., McKinstry, K., Schwegler-Berry, D., Friend, S., Knepp, A.K., Smith, S.L. and Sriram, K. [2016]. Accumulation of Ubiquitin and Sequestosome-1 Implicate Protein Damage in Diacetyl-Induced Cytotoxicity. The American journal of pathology, 186, 2887-2908.

Hubbs AF, Goldsmith WT, Kashon ML, Frazer D, Mercer RR, Battelli LA, Kullman GJ, Schwegler-Berry D, Friend S, Castranova V [2008]. Respiratory toxicologic pathology of inhaled diacetyl in Sprague-Dawley rats. Toxicol Pathol 36(2):330–344.

Hubbs AF, Moseley AE, Goldsmith WT, Jackson MC, Kashon ML, Battelli LA, Schwegler-Berry D, Goravanahally MP, Frazer D, Fedan JS, Kreiss K, and Castranova V [2010a]. Airway epithelial toxicity of the flavoring agent, 2,3-pentanedione. The Toxicologist CD — An official Journal of the Society of Toxicology 114:319.

Hubbs AF, Cumpston, AM, Goldsmith WT, Battelli LA, Kashon ML, Jackson MC, Frazer DG, Fedan JS, Goravanahally MP, Castranova V, Kreiss K, Willard PA, Friend S, Schwegler-Berry D, Fluharty KL, Sriram K [2012]. Respiratory and olfactory cytotoxicity of inhaled 2,3-pentanedione in Sprague-Dawley rats. Am J Pathol 181(3):829-844.

Morgan DL, Flake GP, Kirby PJ, Palmer SM [2008]. Respiratory toxicity of diacetyl in C57BL/6 mice. Toxicol Sci 103(1):169–180.

Morgan, D.L., Jokinen, M.P., Johnson, C.L., Price, H.C., Gwinn, W.M., Bousquet, R.W. and Flake, G.P. [2016]. Chemical Reactivity and Respiratory Toxicity of the alpha-Diketone Flavoring Agents: 2,3-Butanedione, 2,3-Pentanedione, and 2,3-Hexanedione. Toxicol Pathol, 44, 763-783.

Morris, J.B. and Hubbs, A.F. [2009]. Inhalation dosimetry of diacetyl and butyric acid, two components of butter flavoring vapors. Toxicol Sci, 108, 173–183.