Michael Backes

Enzymatic conversion of flavonoids using bacterial chalcone isomerase and enoate reductase.

Flavonoids are a large group of plant secondary metabolites with a variety of biological properties and are therefore of interest to many scientists, as they can lead to industrially interesting intermediates. The anaerobic gut bacterium Eubacterium ramulus can catabolize flavonoids, but until now, the pathway has not been experimentally confirmed. In the present work, a chalcone isomerase (CHI) and an enoate reductase (ERED) could be identified through whole genome sequencing and gene motif search. These two enzymes were successfully cloned and expressed in Escherichia coli in their active form, even under aerobic conditions. The catabolic pathway of E. ramulus was confirmed by biotransformations of flavanones into dihydrochalcones. The engineered E. coli strain that expresses both enzymes was used for the conversion of several flavanones, underlining the applicability of this biocatalytic cascade reaction.

High-impact sulfur compounds: constitutional and configurational assignment of sulfur-containing heterocycles.

To unambiguously identify their structures and to evaluate their organoleptic properties, several constitutional und configurational isomers of dialkyl‐tetrathianes and dialkyl‐pentathiepanes were synthesized by two different synthetic protocols, and separated by preparative gas chromatography. Raman and NMR spectroscopy were used to differentiate between the constitutional 3,6‐dialkyl‐1,2,4,5‐tetrathiane and the 4,6‐dialkyl‐1,2,3,5‐tetrathiane isomers. Furthermore, cis‐ and trans‐isomers of 3,6‐dialkyl‐1,2,4,5‐tetrathianes were distinguished by temperature‐dependent NMR experiments. Static, quantum‐chemical simulations of the NMR spectra for these cis‐ and trans‐isomers were calculated in the gas layer in order to confirm our experimental assignments. In addition, the assignment of 4,7‐alkyl‐1,2,3,5,6‐pentathiepanes were deducted from their Raman spectra. Dialkyl‐tetrathianes and dialkyl‐pentathiepanes are interesting components to be used in flavor applications due to their unique olfactory impact and facets.

Synthesis and sensory studies of umami-active scaffolds.

The class of 2‐isopropyl‐5‐methylbicyclo[4.1.0]heptane‐7‐carboxamides, 1–4, has been identified as potent umami‐tasting molecules. A scalable synthesis of this challenging scaffold and new sensory insights will be presented. Interestingly, the umami characteristics differ remarkably, depending on constitutional and stereochemical features of the parent scaffold. During our studies, we could identify the carboxamide moiety as a crucial factor to influence the umami intensity of these scaffolds. In addition, the configuration of the cyclopropyl moiety exerts some influence, whereas the absolute configuration of the menthyl scaffold, at least the tested D‐ and L‐configuration, is less important.

Evaluation of unsaturated alkanoic acid amides as maskers of epigallocatechin gallate astringency.

Some foods, beverages, and food ingredients show characteristic long-lasting aftertastes. The sweet, lingering taste of high intensity sweeteners or the astringency of tea catechins are typical examples. Epigallocatechin-3-gallate (EGCG), the most abundant catechin in green tea, causes a long-lasting astringency and bitterness. These sensations are mostly perceived as aversive and are only accepted in a few foods (e.g., tea and red wine). For the evaluation of the aftertaste of such constituents over a certain period of time, Intensity Variation Descriptive Methodology (IVDM) was used. The approach allows the measurement of different descriptors in parallel in one panel session. IVDM was evaluated concerning the inter- and intraindividual differences of panelists for bitterness and astringency of EGCG. Subsequently, the test method was used as a screening tool for the identification of potential modality-selective masking compounds. In particular, the intensity of the astringency of EGCG (750 mg kg(-1)) could be significantly lowered by 18-33% during the time course by adding the trigeminal-active compound trans-pellitorine (2E,4E-decadienoic acid N-isobutyl amide 1, 5 mg kg(-1)) without significantly affecting bitterness perception. Further, structurally related compounds were evaluated on EGCG to gain evidence for possible structure-activity relationships. A more polar derivative of 1, (2S)-2-[[(2E,4E)-deca-2,4-dienoyl]amino]propanoic acid 9, was also able to reduce the astringency of EGCG similar to trans-pellitorine but without showing the strong tingling effect.

Rubemamine and Rubescenamine, Two Naturally Occurring N-Cinnamoyl Phenethylamines with Umami-Taste-Modulating Properties

Sensory screening of a series of naturally occurring N-cinnamoyl derivatives of substituted phenethylamines revealed that rubemamine (9, from Chenopodium album) and rubescenamine (10, from Zanthoxylum rubsecens) elicit strong intrinsic umami taste in water at 50 and 10 ppm, respectively. Sensory tests in glutamate- and nucleotide-containing bases showed that the compounds influence the whole flavor profile of savory formulations. Both rubemamine (9) and rubescenamine (10) at 10-100 ppm dose-dependently positively modulated the umami taste of MSG (0.17-0.22%) up to threefold. Among the investigated amides, only rubemamine (9) and rubescenamine (10) are able to directly activate the TAS1R1-TAS1R3 umami taste receptor. Moreover, both compounds also synergistically modulated the activation of TAS1R1-TAS1R3 by MSG. Most remarkably, rubemamine (9) was able to further positively modulate the IMP-enhanced TAS1R1-TAS1R3 response to MSG ∼ 1.8-fold. Finally, armatamide (11), zanthosinamide (13), and dioxamine (14), which lack intrinsic umami taste in vivo and direct receptor response in vitro, also positively modulated receptor activation by MSG about twofold and the IMP-enhanced MSG-induced TAS1R1-TAS1R3 responses approximately by 50%. In sensory experiments, dioxamine (14) at 25 ppm in combination with 0.17% MSG exhibited a sensory equivalent to 0.37% MSG.

Biotechnological Production of Methyl-Branched Aldehydes

A number of methyl-branched aldehydes impart interesting flavor impressions, and especially 12-methyltridecanal is a highly sought after flavoring compound for savory foods. Its smell is reminiscent of cooked meat and tallow. For the biotechnological production of 12-methyltridecanal, the literature was screened for fungi forming iso-fatty acids. Suitable organisms were identified and successfully grown in submerged cultures. The culture medium was optimized to increase the yields of branched fatty acids. A recombinant carboxylic acid reductase was used to reduce 12-methyltridecanoic acid to 12-methyltridecanal. The efficiency of whole-cell catalysis was compared to that of the purified enzyme preparation. After lipase-catalyzed hydrolysis of the fungal lipid extracts, the released fatty acids were converted to the corresponding aldehydes, including 12-methyltridecanal and 12-methyltetradecanal.