Jens-Michael Hilmer

Enantioselective Hydrolysis of d,l-Menthyl Benzoate to L-(–)-Menthol by Recombinant Candida rugosa Lipase LIP1

The stereoselective synthesis of L-menthol is an attractive process in the flavor and fragrance industry. One promising way to obtain optically pure menthol is the enantioselective hydrolysis of menthol esters under enzymatic catalysis. We developed an effective and highly enantioselective method for the synthesis of L-(−)-menthol (>99% EE) by hydrolyzing the key industrial starting compound, d, l-menthyl benzoate. The enzyme of choice was the lipase from Candida rugosa (CRL). While commercially available preparations of this lipase showed only minor selectivity (E=15), excellent enantiomeric purity (E>100) was achieved using the heterologously expressed isoenzyme LIP1.

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.

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.

Creation and Production of Liquid and Dry Flavours

While in ancient times, the sensorial properties of a flavour for foodstuffs were of major importance, modern flavours have to perform like multifunctional systems. Physical form, chemical and mechanical stability and controlled release mechanisms are meanwhile essential criteria for the flavour quality. All these properties have to be addressed by a flavourist in close cooperation with technologists. Therefore, knowledge about food product properties must lead to a careful and intelligent evaluation of the flavour system as an important driver for the success of the final product.

Quantification of Important Flavor Compounds in Beef Stocks and Correlation to Sensory Results by “Reverse Metabolomics”

The meaningful correlation of sensory data with analytical data is one of the most challenging tasks in flavor research. In beef stocks in particular, due to the presence of low levels of aroma-active compounds and the taste contribution of non-volatile molecules to the typical “juiciness” character, the consumer encounters a complex matrix situation. The goal of our study was to carry out a comprehensive analysis of all relevant flavor molecules and the correlation to human sensory data. A technique recently developed at the IPB and termed “reverse metabolomics” was used to link biological activity (i.e., sensory data) with variations in the metabolic profile (i.e., analytical data). We used this methodology for the first time to correlate sensorial attributes and GC-MS, LC-MS, and NMR data in culinary beef stocks. Reverse metabolomics was applied to study the link between sensory and chemical composition in a series of freshly prepared culinary beef stocks. A set of 10 different beef stocks was prepared. The degree of liking of the samples was recorded on a hedonic 1–9 scale. Analysis of the stocks was performed by LC-MS, GC-MS, and NMR. 1H-NMR data directly obtained from the meat stock were very complex. Analysis of this dataset by reverse metabolomics revealed some basic structural elements of the key taste compounds, such as carnosine or anserine. The reverse metabolomics correlation of quantitative data with partiality revealed the importance of a set of compounds. This relevance of these compounds has been confirmed by additional sensory experiments which showed an increase in perceived juiciness.

Steroide und Aromastoffe

Mikroorganismen besitzen nicht nur die Fahigkeit, eine grose Vielfalt von Substanzen aus einfachen Kohlenstoffquellen zu synthetisieren, sie sind auch in der Lage, viele organische Verbindungen zu modifizieren. Diese mikrobiellen Stoffumwandlungen erfolgen dank der dabei beteiligten Enzyme fast immer stereospezifisch und werden als Biotransformation bezeichnet. Besonders aufregende Erfolge wurden damit bei der Herstellung von Steroiden als Medikamente in der Humanmedizin erzielt (Abschn. 12.1). Aber auch bei der Gewinnung von Aromastoffen spielen verschiedene mikrobielle Biotransformationssysteme (ganze Zellen oder Enzyme) zunehmend eine wichtige Rolle (Abschn. 12.2).