Putting natto in the refrigerator does not freeze its aroma in place. In a 2012 study, storage at 4°C was associated with a tendency for fruity-smelling esters to decrease and for nutty pyrazines and cheesy branched-chain fatty acids to increase.
The researchers combined an examination of the principal odors of commercial natto with a time-course study of laboratory-made natto to explore the compounds behind aroma changes during refrigeration.
Examining commercial products and following freshly made natto
The initial samples were commercial stringy natto tested immediately after purchase and after one week of storage at 4°C. Aroma compounds were collected by steam distillation and solvent extraction, and the concentrated extract was diluted in stages. People smelled the instrumentally separated compounds to find key odors that remained detectable after dilution, and substances were identified using mass spectrometry and other evidence.
Commercial products varied considerably, however. Even within the same brand from the same manufacturer, products made on different dates differed in the dilution at which odors remained perceptible. For some compounds, the relative results immediately after purchase and after refrigeration were even reversed. The paper’s table presents one example from repeated investigations.
In a separate experiment, the researchers therefore made natto in the laboratory using specified soybeans, natto bacteria, and production conditions. After fermentation at 40°C for 20 hours, it was stored at 4°C. Compounds were followed immediately after fermentation and after 1, 2, 4, 8, and 16 days.
This experiment used solid-phase microextraction (SPME), which collects compounds from the air above a sample onto a thin sorptive material, and compared analytical peak areas relative to an internal standard. Sampling involved extraction at 50°C for 60 minutes, so the values were not direct measurements of air inside the refrigerator. Each sample was analyzed five times, but this does not mean there were five independently manufactured batches.
Natto also contained fruity aromas
The analysis of commercial products identified fatty-acid esters with fruity odors, including ethyl isobutyrate, ethyl 2-methylbutyrate, and ethyl isovalerate, as one group of principal odorants. The strong aroma characteristic of natto thus contains layers of different odor qualities.
In the laboratory-made product, these esters were detected after fermentation and tended to decrease during refrigerated storage. In the commercial products, some esters were also no longer detectable at the same high dilutions after one week.
The authors considered the decline in esters an important contributor to aroma changes during refrigeration. A shift in the overall impression may result not only from stronger odors increasing but also from fruity elements weakening.
Nutty and cheesy elements tended to increase
Another major group consisted of pyrazines, including 2,5-dimethylpyrazine and trimethylpyrazine. These had nutty odors when separated and smelled. They tended to increase during refrigerated storage of the laboratory-made product, and sensory observations of the whole natto also suggested that its nutty odor became stronger over time.
Branched-chain fatty acids with cheesy odors, including isobutyric acid, 2-methylbutyric acid, and isovaleric acid, also tended to increase during storage. Many aroma compounds originally derived from soybeans, meanwhile, decreased during fermentation or refrigeration and were considered to contribute less to the overall natto aroma.
These are trends for groups of compounds. Individual compounds did not necessarily increase continuously every day. The paper’s main text and summary differ in their descriptions of changes in acetic acid, and the figure does not show a one-directional change either. It is therefore not included here as a uniform example of a fatty acid that increases during refrigeration.
Even at low temperatures, the balance of formation and loss changes
For pyrazine formation, the authors discussed a pathway involving reactions between acetoin and other fermentation products and small amounts of ammonia. They considered amino-acid degradation relevant to branched-chain fatty acids. Esters may involve formation from these fatty acids and ethanol, followed by volatilization or breakdown.
The experiment examined these explanations by following increases and decreases in compounds; it did not directly trace reaction pathways using labeled substances or similar methods. It treats aroma during refrigeration as a combination of compounds that persist, decline, or newly form.
For the laboratory-made natto, the researchers reported that no ammonia odor was perceived even after longer refrigerated storage. Failure to perceive an ammonia odor, however, cannot establish that ammonia is absent or that the food is safe. This was not a study designed to determine shelf life or safety for consumption.
Summary
During refrigerated storage of stringy natto, fruity esters tended to decline, while nutty pyrazines and cheesy branched-chain fatty acids tended to increase. Soybean-derived compounds also declined, changing the balance of the overall aroma.
Both commercial and laboratory-made samples varied, however, so the same changes do not necessarily occur after the same number of days. The study shows natto aroma to be not a single natto odor but a combination of several aromas that continues to change during refrigeration.
At publication, the authors were affiliated with Kyoritsu Women’s Junior College, Kyoritsu Women’s University, the University of Niigata Prefecture, and Aikoku Gakuen Junior College. No separate funding or conflict-of-interest declaration section was identified.
References
Tanaka, T., Kimura, S., Kiuchi, K., Suzuki, A., Muramatsu, K., & Mitsuboshi, S. (2012). Changes in Aroma of Itohiki-Natto during Refrigeration. Nippon Shokuhin Kagaku Kogaku Kaishi, 59(4), 167–174. doi:10.3136/nskkk.59.167 (Full original article)
