Biotechnological production and characterization of natural flavors

Abstract

Biotechnological production of flavors is economically relevant and a key to enable a tasty and diverse food supply. Fungi are suitable biocatalysts for the production of flavor compounds and often more efficient than the classical flavor extraction from plants and foods. Fungi from the department of Basidiomycota have a powerful enzymatic toolbox to generate flavor compounds. Based on a broad screening of Basidiomycota, media optimization was performed to find suitable conditions for an efficient biotransformation of black currant pomace. During fermentation, the scents were investigated on a sensory and instrumental level via gas chromatography-mass spectrometry (GC-MS). W. cocos was grown on a solid-state medium composed of 30 g kg-1 pomace, 6.24 g kg-1 sodium aspartate-monohydrate and 30 g kg-1 agar-agar for 10 days. The flavor was characterized by means of a self-validated aroma dilution analysis (ADA), quantified, and proven with recombination experiments. Solid-state fermentation of pomace and aspartate led to a pleasant odor reminiscent of wild strawberries. Methyl anthranilate (2,206 μg kg-1, flavor dilution factor 210), (R)-linalool (1,879 μg kg-1, 211), 2-amino benzaldehyde (771 μg kg-1, 25), and geraniol (138 μg kg-1, 25) were determined as aroma-relevant for the wild strawberry like flavor in recombination experiments. The substrate-fungus combination made use of the nutrients provided by the black currant pomace leading to an appealing natural flavoring for food. In a second approach, 80 g L-1 pomace and 60 g L-1 sucrose (pH 4.4) were fermented in a submerged culture until the medium reached a pH-value of 3.5. Supernatants of the fermented product were evaluated for a potential use as a beverage. Submerged fermentation of black currant pomace and sucrose led to a pleasant flowery and honey-like odor and a fruity, slightly sour taste. The produced fruit acids and flavor compounds were characterized and quantified. The scaled-up fermentation product was processed and carbonized in cooperation with the Geisenheim University. A panel evaluated the carbonated fermented beverage on a hedonic scale, ranging from 0 to 10, with 8.0 ± 1.4, whereas the value for the non-inoculated medium was 2.5 ± 2.5. A rise of the oxalic acid concentration was detected, which led to the characteristic fruity taste in combination with citric acid (935 mg L-1 citric acid and 192 mg L-1 oxalic acid). Linalool (23 μg L-1), geraniol (42 μg L-1), phenylacetic acid (950 μg L-1), eugenol (23 μg L-1) and methyl phenylacetate (94 μg L-1) were detected with odor activity values above one, which imparted fruity and honey-like odors. Fermentation of black currant pomace led to a tasty beverage, which suits the current beverage trends. Upcycling of pomace with its physiologically valuable nutrients as a fermented beverage could contribute to a healthy human nutrition. In a third study, 28 potentially aroma active non-canonical terpenes were investigated in cooperation with the Dechema Research Institute. For twelve especially potent compounds, the odor thresholds (OT) were determined in comparison to their non-methylated equivalents. In addition to the classical approach according to Ullrich and Grosch (1987), a novel method including the individual OT determination of the internal standards was established. The odors of several non-canonical terpenes were described for the first time. Twelve compounds were identified as highly intense flavors, e.g., the flowery smelling 2-methyllinalool with an OT of 1.8 ng L-1 air, the earthy smelling 2-methyl-α-fenchol (3.6 ng L-1 air), and the flowery scent 2-methylgeraniol (5.4 ng L-1 air). The newly developed method for the OT determination appeared as an improvement compared to the method used so far. Methylated derivatives of linalool, citronellol, and geraniol were highly attractive due to their flowery and citrus-like scents, which are highly interesting for the cosmetic industry.