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.     

THE DETERMINATION OF α- AND β-ACIDS IN HOPS AND HOP PRODUCTS USING HPLC

A rapid reversed phase HPLC method for the analysis for α- and β-acids in hops and hop products is described and has been evaluated. The method uses citric acid in the eluent as a complexing agent to overcome the irreversible adsorption effects shown by some columns, thus allowing optimum eluent pH to be selected. The precision of the method for analysis of hop extract has been determined giving relative standard deviations of 1·0% and 2·1% for α- and β-acids respectively. General agreement with results obtained using a polarimetric α-acids analysis method for hop extracts and hops has been demonstrated.     

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     

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 .     

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.     

STABILITY OF RESINS IN PELLETISED HOPS

The α-acids content and Lead Conductance Value of hop pellets, manufactured from seedless Pride of Ringwood hops grown in Australia and stored at 5°C, 20°C or 30°C, were monitored over a twelve month storage period. No decrease in α-acids content occurred during storage at 5°C, whereas at 20°C the decrease (ca. 7%) was approximately one-third that experienced for baled hops of the same variety. At both 5°C and 20°C the Lead Conductance Value decreased at approximately one-third the rate of that in baled hops. During twelve months storage at 30°C the α-acids content of the hop pellets decreased by approximately 40% and the Lead Conductance Value by 30%. The hop pellets stored at 20°C and 30°C developed rancid odours during the twelve months storage period.     

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.     

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.     

ESTIMATION OF α- AND β-ACIDS IN HOP EXTRACTS BY HPLC

A procedure relying on high performance liquid chromatography for the estimation of α-acids and β-acids in hop extracts has been collaboratively tested by members of a Sub-Committee of the Institute of Brewing Analysis Committee and is recommended for use. No significant differences were found between precision values obtained using peak height and peak area measurements. For α-acids, the mean repeatability (r₉₅) and reproducibility (R₉₅) values were 1-3 and 2-4% m/m respectively over the range 41–62% m/m. For β-acids they were 0-9 and 2-0% m/m respectively over the range 11 to 35% m/m. The precision values were judged to be independent of concentration.     

ON THE ANALYSIS OF HOP BITTER ACID

A High Performance Liquid Chromatography (HPLC) method for automated routine analysis of the hop bitter acids is described. It allows the simultaneous determination of α-acids, β-acids, cohumulone and colupulone. This analysis based on two chromatographic runs takes less than 1/2 h. With this new method as reference, the defects of existing conductometric titration techniques were studied and a new improved conductometric method based on toluene-buffer extraction was developed. The new methods are applicable to all forms of hop products containing α- and β-acids. Analysis for α-acids with ether-acid aqueous methanol extraction gives results which are too high because of several basic defects.     

ESTIMATION OF α-, β- AND ISO-α-ACIDS IN HOPS,HOP PRODUCTS AND BEER USING HIGH PERFORMANCE LIQUID CHROMATOGRAPHY

Methods are described in which high performance liquid chromatography (HPLC) is used to estimate α-, β- and iso-α-acids in hops, hop products and beer. The chromatography relies on an isocratic elution of components from a polythene ‘cartridge’ column, and the method is calibrated with the pure substances as primary standards. Using such a column over 1000 analyses have been carried out without any significant loss in resolution or precision. The procedures are sufficiently rapid for use in commercial transactions and for quality control purposes. For hops and hop extracts coefficients of variation (%) of 2·5 and 0·8 were obtained respectively for α-acids. Values of 0·9 and 0·3 were obtained for iso-α-acids in isomerised extracts and beers respectively. For some isomerised extracts it has been observed that peaks in addition to those given by the iso-α-acids are present on the chromatogram. The current method recommended by the EBC over estimates the iso-α-acid content since these other constituents are included in the analysis.     

老化的颗粒酒花对啤酒发酵抗氧化能力的影响

分析老化后酒花的主要成分,研究发现在酒花老化过程中α酸和β酸发生氧化降解,前期生成异α酸,随着老化的进行,继而产生葎草灵酮(humulinone)和希鲁酮(hulupones)等衍生氧化物。酒花中的总多酚含量逐渐减少,具有抗氧化性的酚酸也发生氧化聚合,使用液相色谱检测几乎无响应峰。并测定总多酚、DPPH清除率、TRAP值、TBA值等抗氧化指标来分析老化的酒花对麦汁和啤酒的抗氧化能力影响,实验发现老化的酒花对麦汁和啤酒的抗氧化能力影响较大,尤其是风味稳定性系数SI值,波动最为明显。     

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.     

THE EFFECT OF STORAGE TEMPERATURE ON THE STABILITY OF THE ALPHA-ACIDS CONTENT OF BALED HOPS

The α-acids content of baled hops (Pride of Ringwood) stored at ?20°C, ?5°C, +5°C and +22°C was monitored over a twelve month storage period. The α-acids content of the hops was found to decrease at a linear rate, i.e., by zero order kinetics, at each storage temperature. The relationship between reaction rate and storage temperature was found to accord with the predictions of classical reaction kinetics, enabling a prediction of the stability of hops stored at various temperatures to be made.     

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.     

DETERMINATION OF HOP ALPHA-ACIDS BY SEMI-AUTOMATIC CONDUCTOMETRIC TITRATION

The conductometric method for determination of α-acids in hops consists of extraction of the hops with organic solvent, followed by conductometric titration of α-acids in the extract. A method is now described for semi-automatic conductometric titration of the extracts.     

HOP RESINS AND BEER FLAVOUR V. THE SIGNIFICANCE OF OXIDIZED HOP RESINS IN BREWING

Oxidation products of α- and β-acids have been used to flavour beers. Ion-exchange chromatographic examinations of extracts of these beers are described. Examination of old hops has revealed the presence of certain oxidation products of hop resins.     

啤酒花活性成分与抗氧化活性的相关性

以3 个品种的萨斯特啤酒花为原料,比较了其石油醚、氯仿、乙酸乙酯、正丁醇和水相等不同极性溶剂提取物中α-酸、β-酸、总黄酮和总多酚的含量,采用?OH、1,1-二苯基-2-三硝基苯肼自由基、β-胡萝卜素-亚油酸3 种不同的抗氧化活性评价体系对各极性溶剂提取物的抗氧化活性进行了研究,并与其活性成分的含量进行了相关性分析。结果表明,啤酒花各极性部位均具有不同程度的抗氧化活性。α-酸、β-酸在石油醚相中含量较高,在3 个啤酒花样品中的含量范围分别为α-酸444.29～583.81 mg/g、β-酸131.83～293.19 mg/g,在氯仿相中含量相对较低,其余提取物中均未检出;相关性研究表明α-酸、β-酸是石油醚和氯仿相抗氧化活性的主要物质基础。中等极性和极性较大溶剂提取物中啤酒花总黄酮、总多酚含量较高,相关性研究也表明黄酮、多酚类物质是这类提取物抗氧化活性的主要物质基础;其中,3 个啤酒花样品中总黄酮、总多酚在乙酸乙酯相中含量最高,分别为20.43～24.13 mg/g和34.97～40.24 mg/g,正丁醇次之,分别为9.45～11.68 mg/g和12.12～19.14 mg/g。     

THE EFFECT OF TIME OF DRESSING ON THE FLOWERING BEHAVIOUR AND YIELD OF HOPS VARIETY LATE CLUSTER

The Late Cluster variety of hops when introduced to Kashmir (India) from USA gave two flowering flushes in a growing season, thus giving two harvests. By adjusting the date of dressing (root pruning) from December to May 15 the abnormal flowering behaviour of the plant could be altered from two flushes to only one. Plants dressed late in spring, April 15 to May 1, gave the highest cone yield which was expressed in only one harvest. This also produced the highest α-acids content.     

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.