The female flower of hop (Humulus lupulus var. lupulus) is an essential ingredient that gives characteristic aroma, bitterness and durability/stability to beer. However, the molecular genetic basis for identifying DNA markers in hop for breeding and to study its domestication has been poorly established. Here, we provide draft genomes for two hop cultivars [cv. Saazer (SZ) and cv. Shinshu Wase (SW)] and a Japanese wild hop [H. lupulus var. cordifolius; also known as Karahanasou (KR)]. Sequencing and de novo assembly of genomic DNA from heterozygous SW plants generated scaffolds with a total size of 2.05 Gb, corresponding to approximately 80% of the estimated genome size of hop (2.57 Gb). The scaffolds contained 41,228 putative protein-encoding genes. The genome sequences for SZ and KR were constructed by aligning their short sequence reads to the SW reference genome and then replacing the nucleotides at single nucleotide polymorphism (SNP) sites. De novo RNA sequencing (RNA-Seq) analysis of SW revealed the developmental regulation of genes involved in specialized metabolic processes that impact taste and flavor in beer. Application of a novel bioinformatics tool, phylogenetic comparative RNA-Seq (PCP-Seq), which is based on read depth of genomic DNAs and RNAs, enabled the identification of genes related to the biosynthesis of aromas and flavors that are enriched in SW compared to KR. Our results not only suggest the significance of historical human selection process for enhancing aroma and bitterness biosyntheses in hop cultivars, but also serve as crucial information for breeding varieties with high quality and yield.

The method based on the reaction of stable DPPH radical proved to be the best for the determination of antioxidant characteristics of hops and hop products. Antioxidant activity is expressed as the rate of decline in absorbance of the reaction environment and assessed in relative percents. Differences in the values of antioxidant activity were determined in Czech and foreign hop varieties. The highest antioxidant activities in the scope of 70 to 80% rel. were measured in Saaz and Spalter Select. Antioxidant activity in most of the varieties moved in the scope of 40 to 60% rel. A part of antioxidant activity of hops is irreversibly lost in the course of drying. The loss does not usually exceed 5% of the original RA DPPH value. Drying also resulted in a decrease of polyphenol compound contents. Drying in belt and chamber kilns is comparable from the point of view of preserving hop antioxidant properties. Results of determination of antioxidant activity in hot water extracts of raw hops and ground hops were comparable and statistically non-significant. The same held true for pelletizing of ground hops. The antioxidant activity of raw hops declined in the course of long-term storage in dependence on storage temperature. Storage temperature had no effect on the antioxidant activity of the hop pellets packed in a multi-layer foil without air access.

Wild hops (Humulus lupulus L.) are potential new germplasms to expand the variability of genetic resources for hop breeding. We evaluated Canadian (62 plants) and Caucasian (58 plants) wild hops by their chemical characteristics and with molecular genetic analyses using sequence-tagged site and simple sequence repeat markers, in comparison with European (104 plants) and North American (27 plants) wild hops. The contents of alpha and beta acids varied from 0.36% to 5.11% and from 0.43% to 6.66% in Canadian wild hops, and from 0.85% to 3.65% and from 1.22% to 4.81% in Caucasian wild hops, respectively. The contents of cohumulone and colupulone distinctly differed between European and North American wild hops: the cohumulone level in alpha acids was in the range 46.1%-68.4% among North American wild hops and in the range 13.6%-30.6% among European wild hops. The high content of myrcene and the low contents of humulene, farnesene, and selinenes were typical for wild hops from Canada, in contrast to wild hops from the Caucasus region. We compared the chemical characteristics with molecular genetic data. Chemical characteristics differentiated wild hops into North American and Eurasian groups. Molecular genetic analysis was able to separate Caucasian wild hops from European wild hops. We proved a hop phylogeny by means of wide molecular analysis.

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Market varieties of hops are classified to several groups according to their use in the brewing industry -aroma, bitter (dual-purpose), high-alpha ones. Saaz and other genetically related varieties form a separate group among the aromatic hops. The group called fine aroma hops has a low content of α-bitter acids (3-4% w/w), its content of β-bitter acids is in the range of 4-7% w/w and cohumulone ratio in the interval of 23-26% rel. The composition of hop oils is characterised by the content of β-farnesene in the range of 15-20% rel. and trans-α-bergamotene at the amount of ca. 1% rel. Most market varieties of hops are of hybrid origin. It holds true about the Czech varieties Sládek, Bor, Premiant and Agnus. The content of α-bitter acids in bitter varieties is in the range of 7-10% w/w while the content of α-bitter acids in highalpha hops is higher than 10% w/w.

Hop (Humulus lupulus L.) inflorescence, commonly known as 'hop cone', is valuable for contents of bitter acids, essential oils and polyphenols, used in brewing industry and biomedical application. These compounds are predominantly formed in glandular trichomes, known as 'lupulin glands'. In this study, we investigated chemical and morphological characteristics of lupulin glands by microscopy and HPLC analyses. Even though no significant correlations were found between lupulin glands and hop cones for contents of alpha and beta bitter acids, polyphenols, all ratios between individual metabolites (alpha/beta acids, X/alpha acids, X/DMX) were highly and significantly correlated. The average number of lupulin glands on bract/bracteole ranged from 115 to 713 with 28.5% of variability. Linear regression analyses confirmed that total volume of lupulin glands and total lupulin glands area on bract/bracteole surface highly correlated with contents of bitter acids and polyphenols in hop cone. Therefore, lupulin glands are unique and exclusive organs responsible for contents of bitter acids and polyphenols in hop cone. Chemical characteristics of lupulin glands J. Patzak et al. Significant correlations at 0.01 probability level are in bold. Chemical characteristics of lupulin glands J. Patzak et al. Chemical characteristics of lupulin glands J. Patzak et al.

Changes in the content and composition of hop secondary metabolites during storage are reflected in beer quality and in the economics of beer production. A 12-month storage experiment with T90 pellets of four hop varieties showed different dynamics of hop aging in relation to both storage conditions and hop variety. Negligible effects on the a-and b-acids were detected during storage without air access at +2C. Storage at +20 C resulted in a final loss of 20-25% a-acids, but the content of b-acids did not change significantly. Large decreases in a-acids (64-88%) and in b-acids (51-83%) were found in hops stored with access to air at +20 C. The rate of decline accelerated markedly after 6 months of storage. In terms of hop resin changes, Premiant and Sládek were the most and the least stable varieties, respectively. After 12 months, the content of the total polyphenols and flavonoids decreased by 30-40% and by 20-30%, respectively, irrespective of storage conditions. The rate of decline accelerated strongly after 6 months. The DPPH (1,1-diphenyl-2-picrylhydrazyl) antiradical potential decrease was significant only in hops stored under aerobic conditions. The depletion was 9-25% after 1 year; Saaz was the most stable variety.

The enzymatic properties of four chalcone synthase homologues CHS_H1, VPS, CHS 2 and CHS 4 from Humulus lupulus L. were investigated after heterologous expression in Escherichia coli. It was found that both VPS and CHS_H1 can utilize isovaleryl-CoA and isobutyryl-CoA as substrates producing compounds with positions in thin layer chromatography characteristic for phloroisovalerophenone and phloroisobutyrophenone. These reactions are accompanied by the formation of associated byproducts. The formation of naringenin chalcone can be catalyzed primarily by CHS_H1. Comparatively the ability of VPS to perform chalcone synthase reaction is very limited. Since only CHS_H1 has true chalcone synthase activity, this enzyme can be considered a key enzyme in prenylflavonoid biosynthesis. Both CHS 2 and CHS 4 utilize isovaleryl-CoA and isobutyryl-CoA as substrates, but the reactions were prematurely terminated. In comparison with VPS and CHS_H1, the optimum pH of CHS 2 was shifted to lower value. High expression of chalcone synthase-like genes were found in maturating hop cones of cultivars with high bitter acid content (Agnus, Magnum, Target) by Northern and Western blotting using probes specific for vps, chs_H1, chs 4 and polyspecific serum risen against recombinant protein CHS4, respectively. It was also found that these cultivars maintained expression of CHS homologues for a longer period of time during cone development in contrast to timelimited expression of CHS homologues in cultivars with low bitter acids content.