Studies on the Sunlight Flavor of Beer

Three synthetic lupulone analogues, i.e., 1-acetyl-3,3,5-tri-allyl-, 1-acetyl-3,3,5-tri-n-propyl-and 1-acetyl-3,3,5-tri-n-butyl-hexadiene- (1,5) -diol- (2,6) -on- (4) did not cause sunlight flavor of beer when they were added to a fermented solution of sucrose and exposed to sunlight.

When methanol solutions of humulone and acetolupuphenonen were exposed to sunlight, changes in the UV-absorption spectra were observed. This fact indicates that humulone and acetolupuphenone underwent some photochemical reaction upon exposure to sunlight.

Prenyl (γ,γ-dimethylallyl) mercaptan was synthesized, which gave a typical sunlight flavor when it was added, in a trace amount, to a fermented solution of sucrose.     

Bildung und weiterreaktion von acylvorstufen der hopfenbitterstoffe

Isobutyric acid is incorporated into desoxycohumulone, cohumulone and colupulone and isovaleric acid is incorporated into desoxyhumulone, humulone and lupulone. The compound 1-isobutyryl-3-(3,3-dimethylallyl)-phloroglucinol (CoX) can be considered as an intermediate for the first three components of the hop resins while isovaleryl-3-(3,3-dimethylallyl)-phloroglucinol (X) is an intermediate in the biosynthesis of the parent compounds.     

Biosynthetic experiments with tall plants under field conditions. 18O2 incorporation into humulone from Humulus lupulus

Five segments of a large hop plant (Humulus lupulus var. Hallertauer Magnum) carrying several cones were enclosed in sealed glass vessels that were gassed with (18)O(2). After 14 days, the segments were harvested and humulone and cohumulone were analysed by NMR spectroscopy and mass spectroscopy. The oxygen atoms in position 6 of humulone and cohumulone showed 9% (18)O enrichment, respectively. It follows that the C-6 hydroxy groups were introduced by oxygenase catalysis.     

Einbau von [14C]-Essigsäure in hopfenbitterstoffe

Labeled acetic acids are incorporated by hops into desoxycohumulone, desoxyhumulone, humulone, colupulone and lupulone. The incorporation of acetic acid is considered in relation to the mixed isoprenoid-polyketide origin of these bitter principles.     

Structure elucidation and an investigation into the in vitro effects of hop acids on human cancer cells

An investigation into the keto-enol relationships in a number of hop acids has been undertaken. These provided samples to enable X-ray analyses of both natural 1 and unnatural humulone 5 to be carried out. Our results suggest that the configuration of humulone 1 may have been incorrectly assigned. An in vitro investigation into the biological activity of hop acids suggests that both humulone 1 and lupulone 2 exhibit significant activity against some human cancer cells. Our results provide evidence to show that they inhibit cell growth, induce caspase dependent apoptosis and inhibit adhesion of some cancer cells to bone tissue.     

HlPT-1, a membrane-bound prenyltransferase responsible for the biosynthesis of bitter acids in hops

Female flowers of hop (Humulus lupulus L.) develop a large number of glandular trichomes called lupulin glands that contain a variety of prenylated compounds such as α- and β-acid (humulone and lupulone, respectively), as well as xanthohumol, a chalcone derivative. These prenylated compounds are biosynthesized by prenyltransferases catalyzing the transfer of dimethylallyl moiety to aromatic substances. In our previous work, we found HlPT-1 a candidate gene for such a prenyltransferase in a cDNA library constructed from lupulin-enriched flower tissues. In this study, we have characterized the enzymatic properties of HlPT-1 using a recombinant protein expressed in baculovirus-infected insect cells. HlPT-1 catalyzed the first transfer of dimethylallyl moiety to phloroglucinol derivatives, phlorisovalerophenone, phlorisobutyrophenone and phlormethylbutanophenone, leading to the formation of humulone and lupulone derivatives. HlPT-1 also recognized naringenin chalcone as a flavonoid substrate to yield xanthohumol, and this broad substrate specificity is a unique character of HlPT-1 that is not seen in other reported flavonoid prenyltransferases, all of which show strict specificity for their aromatic substrates. Moreover, unlike other aromatic substrate prenyltransferases, HlPT-1 revealed an exclusive requirement for Mg(2+) as a divalent cation for its enzymatic activity and also showed exceptionally narrow optimum pH at around pH 7.0.     

Application of chitooligosaccharides as antioxidants in beer to improve the flavour stability by protecting against beer staling during storage

Objective

To improve beer flavour stability by adding chitooligosaccharides that prevent formation of staling compounds and also scavenge radicals in stale beer.

Results

Chitooligosaccharides, at 0.001–0.01%, inhibited the formation of staling compounds in forced aged beer. The formation of 5-hydroxymethylfurfural, trans-2-nonenal and phenylacetaldehyde were decreased by 105, 360 and 27%, respectively, when compared with those in stale beer without chitooligosaccharide addition. The capability of chitooligosaccharides to prevent staling compound formation depended on their molecular size (2 or 3 kDa). The DPPH/hydroxyl radical scavenging activity in fresh beer significantly lower than that in forced aged beer in the presence of chitooligosaccharides. When compared with stale beer without added chitooligosaccharides, the radical scavenging activity could be increased by adding chitooligosaccharides to forced aged beer.

Conclusions

Chitooligosaccharides play an active part in the prevention of beer flavour deterioration by inhibiting the formation of staling compounds and increasing radical scavenging activity.

     

Hydrolyzing Pathway,Substrate Specificity and Inhibition of Tannin Acyl Hydrolase of Asp. oryzae No. 7

Tannin acyl hydrolase (EC 3.1.1.20) of Asp. oryzae No. 7 hydrolyzes tannic acid to glucose and gallic acid. The intermediate hydrolyzates are 1,2,3,4,6-pentagalloyl glucose, 2,3,4,6-tetragalloyl glucose and two kinds of monogalloyl glucose.

The enzyme hydrolyzes ester compounds of gallic acid, but does not hydrolyze any other substrate analogues such as methyl-resorcyrate.

The enzyme reaction is inhibited competitively by substrate analogues which have phenolic hydroxyls with the exception that 2,6-dihydroxy benzoic acid inhibits noncompetitively. Therefore the binding site of the enzyme may be able to react with any kind of phenolic hydroxyl, although the substrate forming a true ES-complex must be an ester compound of gallic acid.     

Membrane leakage in Bacillus subtilis 168 induced by the hop constituents lupulone,humulone, isohumulone and humulinic acid

Summary The bactericidal activity of the hop constituents lupulone, humulone, isohumulone and humulinic acid against Bacillus subtilis 168 is shown to result from primary membrane leakage. This process leads to complete inhibition of active transport of -methyl-d-glucopyranoside and several amino acids into whole bacteria and isolated membrane vesicles within 5 min at 37°C at the corresponding minimal inhibitory concentrations of 1, 2, 25 and 250 g/ml, respectively. Inhibition of the respiratory chain, of protein, RNA and DNA synthesis seems to be a secondary event.     

Occurrence of α-acetolactate decarboxylases among lactic acid bacteria and their utilization for maturation of beer

Summary Acetolactate decarboxylase activity has been detected among three genera, nine species and 263 strains of lactic acid bacteria tested in the course of a screening for acetolactate decarboxylases amenable for use in brewing as maturation aid. Streptococcus diacetylactis strain FD-64-D was found to generate a decarboxylase exhibiting a satisfactory activity and an excellent stability at the pH prevailing in beer and wort. This decarboxylase could not be solubilized but enzymatically active, freeze-dried cells were effective for satisfactory flavour maturation of beer although difficulties were encountered during attempts to remove the applied cell material by filtration of the beer. Lactobacillus casei DSM 2547 was likewise found to produce a decarboxylase exhibiting a satisfactory activity and stability at the low pH of beer and which, in addition, was readily solubilized. A method has been developed for pilot scale production of preparations of this decarboxylase suitable for use in brewing.Abbreviations DSM Deutsche Sammlung von Microorganismen - EDTA Ethylene diaminetetra-acetic acid     

Flavour-active wine yeasts

The flavour of fermented beverages such as beer, cider, saké and wine owe much to the primary fermentation yeast used in their production, Saccharomyces cerevisiae. Where once the role of yeast in fermented beverage flavour was thought to be limited to a small number of volatile esters and higher alcohols, the discovery that wine yeast release highly potent sulfur compounds from non-volatile precursors found in grapes has driven researchers to look more closely at how choice of yeast can influence wine style. This review explores recent progress towards understanding the range of ??flavour phenotypes?? that wine yeast exhibit, and how this knowledge has been used to develop novel flavour-active yeasts. In addition, emerging opportunities to augment these phenotypes by engineering yeast to produce so-called grape varietal compounds, such as monoterpenoids, will be discussed.     

Monitoring the influence of high-gravity brewing and fermentation temperature on flavour formation by analysis of gene expression levels in brewing yeast

During fermentation, the yeast Saccharomyces cerevisiae produces a broad range of aroma-active substances, which are vital for the complex flavour of beer. In order to obtain insight into the influence of high-gravity brewing and fermentation temperature on flavour formation, we analysed flavour production and the expression level of ten genes (ADH1, BAP2, BAT1, BAT2, ILV5, ATF1, ATF2, IAH1, EHT1 and EEB1) during fermentation of a lager and an ale yeast. Higher initial wort gravity increased acetate ester production, while the influence of higher fermentation temperature on aroma compound production was rather limited. In addition, there is a good correlation between flavour production and the expression level of specific genes involved in the biosynthesis of aroma compounds. We conclude that yeasts with desired amounts of esters and higher alcohols, in accordance with specific consumer preferences, may be identified based on the expression level of flavour biosynthesis genes. Moreover, these results demonstrate that the initial wort density can determine the final concentration of important volatile aroma compounds, thereby allowing beneficial adaptation of the flavour of beer.     

<Emphasis Type="Italic">Humulus lupulus</Emphasis> L., a very popular beer ingredient and medicinal plant: overview of its phytochemistry,its bioactivity,and its biotechnology

Humulus lupulus L. (Cannabaceae), commonly named hop, is widely grown around the world for its use in the brewing industry. Its female inflorescences (hops) are particularly prized by brewers because they produce some secondary metabolites that confer bitterness, aromas and antiseptic properties to the beer. These sought-after metabolites include terpenes and sesquiterpenes, found in essential oil, but also prenylated phenolic compounds, mainly acylphloroglucinols (bitter acids) from the series of α-acids (humulone derivatives). These metabolites have shown numerous biological activities, including among others, antimicrobial, sedative and estrogenic properties. This review provides an inventory of hop’s chemistry, with an emphasis on the secondary metabolites and their biological activities. These compounds of biological interest are essentially produced in female inflorescences, while other parts of the plant only synthetize low quantities of them. Lastly, our article provides an overview of the research in plant biotechnology that could bring alternatives for hops metabolites production.     

Studies of the Sunlight Flavor of Beer

The recent findings^1~3) that prenylmercaptan (3-methyl-2-butene-1-thiol) is the major component of the sunlight flavor of beer has led us to investigate the pathway of its evolution. S-Prenyl-l-cysteine, S-(3-methyl-2-butenyl)-l-cysteine, was synthesized according to the general outline of A. Stoll et al.⁴⁾ from l-cyteine and prenylbromide, since it was considered as one of the precursors of the sunlight flavor of beer. S-Prenyl-l-cysteine was a colorless and odorless crystal, but this compound generated prenylmercaptan when the aqueous solution was exposed to sunlight. The addition of a small amount of riboflavin to the solution as a photosensitizer increased the mercaptan evolution. Prenylmercaptan formed by sunlight was isolated as its 2,4-dinitrophenyl derivative and identified by the comparison of melting point, chromatographic behavior and infrared spectrum with an authentic sample and by its elemental analysis.     

N-Coronafacoyl-L-isoleucine and N-coronafacoyl-L-alloisoleucine,potential biosynthetic intermediates of the phytotoxin coronatine

Two new naturally-occurring analogues of the phytotoxin coronatine have been isolated from liquid cultures of Pseudomonas syringae pv. glycinea. These have been identified as N-coronafacoyl-L-isoleucine and N-coronafacoyl-L-alloisoleucine by mass spectrometry and by studies of the products of acid hydrolysis of the two compounds. The compounds were purified as a mixture of ca 2:1 composition, but the two parent components were not preparatively separated. The possible significance of the two compounds, to the biosynthesis of coronatine, is discussed.     

β-Lactams: A New Class of Conformationally-Rigid Inhibitors of γ-Aminobutyric Acid Aminotransferase

Abstract

A structural similarity of several monobactams (2–4), 3-aminonocardicinic acid (6), 6-aminopenicillanic acid (7), 7-aminocephalosporanic acid (8), and 7-aminodesacetoxycephalosporanic acids (9, 10) to γ-aminobutyric acid (GABA) and to known inhibitors and substrates of GABA aminotransferase is described. Because of this, the above-mentioned compounds were tested as competitive inhibitors and as inactivators of pig brain GABA aminotransferase. All of the compounds were competitive inhibitors of GABA aminotransferase. On the basis of the inhibitory potency of these conformationally-rigid GABA analogues it is hypothesized that GABA is bound at the active site with its amino and carboxylate groups in a syn orientation. None of the compounds inactivates GABA aminotransferase. These β-lactam analogues represent the first examples of a new class of inhibitors of GABA aminotransferase.     

Vanillin production using metabolically engineered <Emphasis Type="Italic">Escherichia coli</Emphasis> under non-growing conditions

Background  

Vanillin is one of the most important aromatic flavour compounds used in the food and cosmetic industries. Natural vanillin is extracted from vanilla beans and is relatively expensive. Moreover, the consumer demand for natural vanillin highly exceeds the amount of vanillin extracted by plant sources. This has led to the investigation of other routes to obtain this flavour such as the biotechnological production from ferulic acid. Studies concerning the use of engineered recombinant Escherichia coli cells as biocatalysts for vanillin production are described in the literature, but yield optimization and biotransformation conditions have not been investigated in details.