Determination of compounds responsible for tempeh aroma Henryk Jelen ´ ⇑ , Małgorzata Majcher, Alexandra Ginja, Maciej Kuligowski Faculty of Food Science and Nutrition, Poznan ´ University of Life Sciences, Wojska Polskiego 31, 60-624 Poznan ´, Poland article info Article history: Received 19 November 2012 Received in revised form 11 February 2013 Accepted 13 March 2013 Available online 20 March 2013 Keywords: Tempeh AEDA Aroma analysis Flavour Gas chromatography–olfactometry GC–O Odour activity values OAV Gas chromatography/mass spectrometry GC/MS abstract Tempeh is a fermented food, popular mainly in south-east Asia, but also among vegetarians worldwide. It is produced by fermenting soybean or other beans with Rhizopus strains and usually eaten deep-fried, steamed or roasted. The flavour of tempeh depends upon the fermentation time, beans used and the (eventual) frying process. Our goal was to identify compounds responsible for the unique aroma of fer- mented and fried soy tempeh. Gas chromatography–olfactometry (GC–O) with the aroma extract dilution analysis (AEDA) approach, was used to determine key odorants after 1 and 5 days of fermentation and subsequent frying. Comprehensive gas chromatography–mass spectrometry (GC Â GC–ToF-MS) was used for their quantitation using stable isotope dilution analysis (SIDA) or standard addition (SA) methods. Odour activity values (OAV) were calculated for 19 out of 21 key odorants. Tempeh was fermented for 5 days and fried, and the main aroma compounds were found to be the following: 2-acetyl-1-pyrroline, (FD = 1024, OAV 1380), 2-ethyl-3,5-dimethylpyrazine (FD = 512, OAV 338), dimethyl trisulfide, (FD = 512, OAV 900), methional (FD = 512, OAV 930), 2-methylpropanal (FD = 512, OAV 311) and (E,E)-2,4-decadi- enal (FD = 512, OAV 455). The frying process induced the increase or appearance of the main key odorants in tempeh. Ó 2013 Elsevier Ltd. All rights reserved. 1. Introduction Tempeh (also known as tempe) is a traditional Indonesian food produced by the fermentation of soybeans using Rhizopus species. Apart from soybeans, other substrates used for fermentation have been utilised: chickpeas (Angulo-Bejarano et al., 2008), barley (Feng, Eriksson, & Schnürer, 2005) and beans (Ashenafi & Busse, 1991). The tempeh of a specific flavour is produced from pressed copra (Hachmeister & Fung, 1993). Tempeh is normally consumed fried, boiled, steamed or roasted. During the fermentation process, the enzymatic digestion of substrates, leads to an increased amount of free aminoacids, water-soluble nitrogen compounds, free fatty acids, and to the development of characteristic flavour. During the fermentation of tempeh, a decrease in the amount of crude lipids is observed, as lipids serve as the main source of en- ergy for the microorganisms during the fermentation process. The protein hydrolysis may amount to 25% of the initial soy protein (Sparringa & Owens, 1999). In addition to being a source of proteins and lipids in a vegetar- ian diet, tempeh became of interest due to its interesting nutri- tional/functional properties. It has been observed that fermented soybean products, such as tempeh, miso and natto, are more resis- tant to lipid oxidation than unfermented soybeans. The isoflavone levels in tempeh are relatively high compared to other soybean products. Raw tempeh contains the highest levels of daidzein and genestein, compared to tofu or soybean drinks. However, the pro- cess of tempeh deep frying significantly (up to 45%) reduces the to- tal isoflavones contents (Haron, Ismail, Azlan, Shahar, & Peng, 2009). The potential use of tempeh as a functional food has in- creased. Methods of production of c-aminobutyric acid enriched tempeh, which has antihipertensive effects (Aoki, Furuya, Endo, & Fujimoto, 2003a; Aoki et al., 2003b), as well as isoflavone-enriched tempeh (Nakajima, Nozaki, Ishihara, Ishikawa, & Tsuji, 2005) have been reported. The properties, described above, of tempeh are mainly a result of the microbial/enzymatic activity of microorganisms, used for its production and biotransformation of soy constituents. The main fungus used for the preparation of tempeh in Indonesia is Rhizopus oligosporus, which is considered a domesticated form of Rhiyopus microspores (Feng, 2006). More strains identified in tempeh, in- cluded Rhizopus formosaensis, R. and Rhizopus oryzae (Babu, Bhaky- araj, & Vidhyalakshmi, 2009). It has been postulated that the natural habitat for Rhizopus could be fresh leaves of the Hibiscus species, which is used to wrap cooked soybeans (Ogawa, Toku- masu, & Tubaki, 2004). Apart from Rhizopus species, various micro- organisms, including filamentous fungi, yeasts and bacteria, are found in the traditional tempeh. Tempeh co-inoculation with lactic acid bacteria (LAB) is performed to improve the safety of the prod- uct and this process contributes also to the fermentation process (Ashenafi & Busse, 1991). The growth of LAB (Lactobacillus plantarum), coexisting with R. oligosporus, especially with the 0308-8146/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.foodchem.2013.03.047 ⇑ Corresponding author. Tel.: +48 61 8487273; fax: +48 61 8487314. E-mail address: henrykj@up.poznan.pl (H. Jelen ´ ). Food Chemistry 141 (2013) 459–465 Contents lists available at SciVerse ScienceDirect Food Chemistry journal homepage: www.elsevier.com/locate/foodchem