AUTOMATED REPORTING ON THE QUALITY OF HOPS AND HOP PRODUCTS

The quality of a hop variety or a hop product can readily be assessed by a fully automated sequence of selective extraction, fractionation and quantitative analysis. To illustrate the elegance of the method, nine hop varieties and three hop extracts were compared with respect to the content of important marker compounds in the hop oils and of the hop acids. Supercritical fluid extraction at different densities of carbon dioxide was applied to extract selectively, the hop oils and the hop acids, respectively. The hop oils were further fractionated into an apolar and a polar fraction by solid phase extraction and consecutive elution with n-hexane and ethyl acetate. Separation and identification were achieved by capillary gas chromatography coupled to mass spectrometry. Myrcene, β;-caryophyllene, α;-humulene and β;-farnesene in the apolar fraction, linalool, undecan-2-one, tridecan-2-one and humuladienone in the polar fraction were selected for quantitative evaluation of the respective hop oils. Sulphur-containing compounds were revealed by capillary gas chromatography using sulphur-selective atomic emission detection. Complete separation and quantification of all hop α;-acids and β;-acids was effected by microemulsion electrokinetic chromatography coupled to diode array detection .     

NON-IMPLICATION OF HOP α- AND β- ACIDS AS SOURCES OF SULPHUR COMPOUNDS IN BEER

Whereas hop oil terpenoids can give rise to organoleptically undesirable sulphur compounds in beer brewed using hops dressed on the bine with sulphur, the hop resin α- and β-acids and their transformation products appear incapable of reactions with sulphur under analogous conditions. The evidence indicates that the hop resins are not potential sources of sulphur compounds in beer     

IONIZATION BEHAVIOUR OF HOP COMPOUNDS AND HOP-DERIVED COMPOUNDS

The ionization behaviour of colupulone, (?)-humulone, trans-isohumulone and trans-humulinic acid was investigated using a number of techniques. When a potentiometric technique was employed, precise estimates of pKa could not be obtained for any of the compounds, since a drift in the observed pKa values occurred during the titrations. Approximate, or equilibrium pKa values for the most acidic function of each of the compounds were: colupulone, 6.1; (?)-humulone, 5.0; trans-isohumulone, 3.1; trans-humulinic acid, 2.7. Evidence from spectroscopic measurements (UV, ¹H-NMR, ¹³C-NMR), conductiometric experiments and measurements of the pH dependency of solvent partition behaviour and aqueous solubility of the compounds is consistent with the possibility that, in aqueous solution, compounds such as trans-isohumulone are present as covalent hydrates. It is likely that reversible hydration occurs at one or more carbonyl groups. As a consequence of this behaviour, no single pKa value is sufficient to describe the behaviour of any one hop compound or hop-derived compound. The potential significance of this phenomenon is discussed.     

ANALYSIS OF HOPS AND HOP PRODUCTS FOR α-ACIDS OR ISO-α-ACIDS

Two methods based on the resolution of mixtures of hop compounds by chromatography on Sephadex columns have been adopted by E.B.C. and A.S.B.C. as ‘International Methods’.     

QUANTITATIVE ANALYSIS OF HOPS AND HOP EXTRACTS BY HIGH PRESSURE ION EXCHANGE CHROMATOGRAPHY

A new analysis for hop acids in hops and hop extracts is described. It is based on recent developments in high pressure liquid chromatography using pellicular anion exchange column filling material. Quantitative evaluation of α-acids, β-acids and oxidation products in hops and hop extracts is carried out by standard addition of pure humulone. The α-acids are completely separated from other hop substances before quantitation. The results of α-acids determinations must therefore be more accurate than was formerly possible and they are compared with conductometric titration results which are systematically higher. This is to be expected as it becomes more and more obvious that paper strip and conductometric analysis are not selective enough and determine fractions as “α-acids” which are in fact oxidation products of the hop acids.     

ENZYMIC DEGRADATION OF α-ACIDS IN HOPS

Evidence is presented for the biological oxidation of α-acids in hop cones by an enzyme having molecular weight 17,000 daltons, K_m 6–0 and Mn as the active metal ion. The enzyme is located in bracts and bracteoles and not in lupulin glands. The presence of an endogenous inhibitor was demonstrated. The specific activity of protein in hop cones increased with maturity and this was probably associated with release of α-acids from lupulin glands.     

PREPARATION OF HOP EXTRACTS RICH IN PARTICULAR CONSTITUENTS*

When a column containing powdered hops is extracted with liquid carbon dioxide, chromatographic separation of hop components occurs. They are extracted in the order essential oils, β-acids, α-acids; and the separation is enhanced when finely milled hops are extracted. Early fractions (～0·5 hour) contain a high proportion of the available essential oils when hops are extracted at ?20°C and such extracts are suitable as a replacement for dry hops. Fractions can be obtained from extractions at ～7°C which are rich in α-acids and contain low levels of β-acids. Small amounts of fats and waxes are normally present in fractions collected towards the end of a run when seeded hops are extracted.     

SEPARATION AND QUANTIFICATION OF ALL MAIN HOP ACIDS IN DIFFERENT HOP CULTIVARS BY MICROEMULSION ELECTROKINETIC CHROMATOGRAPHY

Microemulsion electrokinetic chromatography has been applied to the analysis of hop acids in extracts of different hop cultivars. With this new analytical method, the 6 main hop bitter acids are completely separated within 10 minutes. Quantitative analyses of extracts from Saaz, Nugget and Wye Target cultivars have been compared to those obtained with reversed-phase high-performance liquid chromatography. The cohumulone and colupulone ratios are in good agreement. Microemulsion electrokinetic chromatography however is much faster and provides, in addition, data for the adhumulone and adlupulone ratios.     

HOP INDUSTRY ADVISORY COMMITTEE: ANALYSIS OF HOPS OF THE 1975 CROP

Analytical values for α-acid content of eleven varieties of hops from the 1975 crop are presented in relation to the district of production. For most of the varieties the variation within growths was lower than in the previous year. With amended sampling rates, the tolerance limits of ±0·5 % were achieved for all hop varieties and the results are acceptable for commercial purposes. Some α-acid values for seedless hops from the Seedless Hop Trials are included in the Report and comparisons with seeded hops are shown over a three year period.     

CAMBRIDGE PRIZE LECTURE. STUDIES ON THE SENSITIVITY OF LACTIC ACID BACTERIA TO HOP BITTER ACIDS

Hop bitter acids act as mobile-carrier ionophores. They inhibit the growth of beer-spoilage bacteria by dissipating the transmembrane pH gradient. Their activity is pH dependent. Low pH favours antibacterial activity but high pH reduces it. Resistance to hop bitter acids is a stable character, associated only with beer-spoilage lactic acid bacteria. Hop-resistant organisms can maintain a larger transmembrane pH gradient and ATP pool than can hop-sensitive organisms. Prior exposure of bacteria to trans-isohumulone does not influence the degree of resistance to hop bitter acids. However, in some strains, exposure to trans-isohumulone does induce the ability to spoil beer. The chemistry of these compounds is more complex than previously thought. In aqueous solutions, such as beer, hop acids bind to metal ions and may be covalently hydrated.     

ANALYSIS OF HOPS OF THE 1987 CROP

Analytical values for α-acid content, as measured by lead conductance value, of twelve varieties of hops from the 1987 crop are presented in relation to the district of production. Satisfactory variabilities have been achieved by Fuggle, Progress, Bramling Cross, Northern Brewer, Zenith, Wye Challenger and Omega. Wye Northdown has increased in variability, reaching the upper limit of acceptability despite an increase in sampling rate. Goldings, W.G.V. and Wye Target have continued to show higher than normal variabilities. This year Yeoman has moved in variability above the acceptable limit. The sampling rates have been considered and in view of the unusual growing conditions experienced with the 1987 crop, with hop yields considerably reduced and α-acids increased to very high levels, no changes have been recommended.     

SOLUBILITY OF HOP α- AND β-ACIDS IN LIQUID CARBON DIOXIDE*: ADDENDUM: SOLUBILITY OF PESTICIDES IN LIQUID CARBON DIOXIDE

A procedure is described for determining the solubility of hop α- and β-acids in liquid carbon dioxide. Results have shown that the optimum temperature range for the extraction of hops with liquid carbon dioxide is +5 to +10°C. A number of pesticides used by hop growers are appreciably soluble in liquid carbon dioxide.     

STUDIES ON THE EFFECT OF POLLINATION ON THE YIELD AND RESIN CONTENT OF HOPS (HUMULUS LUPULUS L.)

Pollination trials in otherwise male-free hop gardens are described. The results showed that a reduction in cone yield of up to 30% may be expected from seedless hops, but an increase in α-acid content of the cones more or less compensates for this yield reduction and the yield of α-acid per acre can be almost as high from seedless as seeded hops. Northern Brewer, in particular, performed better than other varieties when grown seedless but most of the high α-acid varieties gave reasonable results. It is suggested that the variation in response may depend on the number of resin glands present on the seeds of the different varieties, and differences in the distribution of resin glands on other parts of the cone.     

HOP INDUSTRY COMMITTEE ANALYSIS OF HOPS OF THE 1984 CROP

Analytical values for α-acid content of thirteen varieties of hops from the 1984 crop are presented in relation to the district of production. For all varieties, except Yeoman, the tolerance aim of ±0·5% was achieved. The production of Yeoman has almost doubled this year and the inclusion of young hops with mature hops in some growths has led to increased within growth variation and failure of this variety, by a small margin, to remain within the limits of the tolerance aim. Apart from this, the variability of the crop compared well with previous years and the results are acceptable for commercial purposes. The sampling rates have been considered and no changes have been recommended.     

HOP RESIN ANALYSIS BY HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY ON MICROPARTICULATE SILICA

A high-performance liquid chromatographic separation of hop resins on a commercially available microparticulate silica has been developed. The chromatography involves a new mechanism in hop resin separation. A solvent containing di-n-butylammonium acetate is used. The method gives good separation of α-acid, iso-α-acid, β-acid and humulinic acid, and has been used for the quantitative analysis of α-acid and iso-α-acid in hop extracts and isomerised hop extracts.     

RAPID SPECTROPHOTOMETRIC METHOD FOR DETERMINATION OF THE BITTERING VALUE OF ISOMERIZED HOP EXTRACTS

A method is described which allows the rapid determination of the bittering value of isomerized hop extracts. The method is also applicable in the case of isomerized hop extracts containing impurities such as α-acids and β-acids.     

IMPROVEMENT OF THE BITTERNESS OF HOPS: PHOTOREACTIONS OF ALPHA ACIDS

The phototransformation of the α-acids (humulone) of hop resins has been studied in order to improve their bitter properties and to establish the optimal experimental conditions.     

HOP INDUSTRY COMMITTEE: ANALYSIS OF HOPS OF THE 1977 CROP

Analytical values for α-acid content of thirteen varieties of hops from the 1977 crop are presented in relation to the district of production. For all varieties the tolerance aim of ±0.5% was easily achieved, the whole crop being less variable than in any previous year since these records were kept. The results are acceptable for commercial purposes. α-Acid values for seedless hops from Hampshire and from the Seedless Hop Trials are included and comparisons with seeded hops are shown over a three year period.     

HOP INDUSTRY COMMITTEE: ANALYSIS OF HOPS OF THE 1978 CROP

Analytical values for α-acid content of thirteen varieties of hops from the 1978 Crop are presented in relation to the district of production. For all varieties the tolerance aim of ±0.5% was achieved and this year the crop was no more variable than the average for the last five years. As before the results are acceptable for commercial purposes.     

HOP OIL CONSTITUENTS DERIVED FROM THE OXIDATION OF COLUPULONE

Several spiroacetal and furanone derivatives have been detected in the volatile oxidation products formed by heating colupulone with gaseous oxygen. This finding explains the occurrence of high levels of these compounds in hops which have undergone a prolonged period of storage.