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Technical Publications
Hop Polyphenols - do more than just cause turbidity in beer
Hopfenrundschau International, 68 74, 1999

A. Forster, B. Beck and R. Schmidt

Introduction
Even today polyphenols originating from barley-malt or hops are considered to be a nuisance by many farmers, as certain groups of polyphenols promote the formation of irreversible turbidity in the bottled beer. In order to attain a high physical stability they prefer to reduce the dosage of polyphenols or partially remove them. This can be done e.g. by using specially cultivated varieties of barley free of proanthocyanidine such as Caminant or by using pure-resin hop extracts free of polyphenol. Alternatively turbid-active polyphenols can be reduced by adsorption on polyvinylpolypyrrolidon (PVPP) during filtration. However this is contrary to an interesting development: In food and plant research increased efforts are being made to find natural antioxidants e.g. in herbs, tea or seeds. The effects of these antioxidants, which are often of polyphenolic character, are seen in two different ways. They can protect food such as fats from being spoiled by oxygen as far as the taste is concerned. On top of this it is thought that they are used to catch radicals in the human body thus exerting an anticarcinogenic effect. Examples of this are rosemary, tea and especially green tea, or red-wine. Now the hop is a plant with comparatively high contents of polyphenols (4-6 weight -%) which raises the question whether and how this group of substances can specifically be used.

Analytics
Until a few years ago, for the most part there were only unspecific colouring methods available for the analytical description of polyphenols. In the meantime considerably more informative methods exist using high-pressure liquid chromatography (HPLC) linked with a diode array detector (DAD). By this means it is possible to separate a polyphenol compound into its individual components. In this article analytical details such as the preparation of samples, chromatographical conditions, identification of peaks and their calibration will deliberately be avoided. Meanwhile about 100 single polyphenolic hop components can be separated. Chart 1 shows the main groups of substances with reference values on their respective contents. By way of the permanent spectral recording a DAD makes it possible to secure the identity of peaks and by comparing the spectren of known substances with those of test peaks enables them to be classified into substance groups. Furthermore, simultaneous representation is possible in several wave-lengths (e.g. with 265, 280, 310 and 360 nm). With a logically devised method the polyphenols can be analyzed analogue in worts and beer.

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Influences of hop varieties and production area

The analyses of several years relating to the influence of important hop varieties on the composition of the polyphenols will soon be completed. A detailed description of the results will not be prejudged here. In Diag. 1 just to illustrate the HPLC chromatograms (wave-length 360 nm) two hop varieties produced in the Hallertau i.e. Perle and Nugget are compared as finger-prints. This very comparison already clearly shows that the hop variety exerts a definite influence on the composition of polyphenols. Aroma hops contain larger amounts of low-molecule polyphenols. As far as fruit is concerned, e.g. paprika or oranges, it is already known that the production area with its climatic conditions exerts an influence on the composition of flavonoids. Here too analyses on hops are nearly finished and will soon be made public. Diag. 2 shows the HPLC chromatogram of the variety Nugget from the Hallertau and Yakima production areas. The peak distributions differ significantly. According to this climate and production area play a role in the composition of polyphenols in hops. A moderate climate such as in the Hallertau has a positive effect.

Brewing trials
Many brewers are prejudiced against polyphenols as they promote the formation of turbidity in beer and therefore impair its physical stability. Allegedly they also have a negative effect on colour, foam and taste of the beer. In order to clarify these matters the following brewing trials were carried out in the 120 l pilot brewery of the experimental station of the Swiss breweries in Zrich (5): Hop polyphenols in the form of a bracteole fraction, as produced by the mechanical concentration of hop powder type 45, was added to the brew hopped with pure-resin extract free of polyphenols. The comparison or neutral brew did not contain any of this hop polyphenol fraction (HPF). In the 1st series the dosage, except in the case of the comparison brew, was with 300 and 600 g/hl of a HPF from Saaz hops not only at the beginning of the boil but also at the end. In the 2nd series amounts between 50 and 300 g/hl HPF were used , besides the comparison brew. The factors hop variety and boiling time of the HPF in the wort were varied. The features foam, colour, physical stability, reduction power, polyphenol values and particularly taste were observed. The results of both series can be summarized as follows: Colour and foam were not influenced by the dosage of a HPF. The physical stability suffered especially when the HPF was boiled for a longer time. The reduction power only ascertained in the 2nd series in accordance with MEBAK method 2.20.1. improved from 45 (comparison brew) via 50 (approx. 100 g/hl HPF) up to 55 (150-300 g/hl HPF). The polyphenol ratios in the beers are shown in Chart 2. The beers were tested in a three-glass test with the comparison beer and with 11-15 tasters.

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HOPS ARE OUR WORLD
In the 1st series 4 beers were available: Comparison or neutral beer without hop polyphenols = 0 600 g/hl HPF hopped at beginning of the boil (BB) = 600 BB 300 g/hl HPF hopped at end of the boil (EB) = 300 EB 600 g/hl HPF hopped at end of the boil (EB) = 600 EB

Chart 3 shows the results which can be summarized as follows: There were significant findings with regard to differentiation and preference, by which the beers containing spent hops surprised the tasters with their pleasant, hoppy, slightly fruity aroma and taste. It is still not clear whether these impressions of taste can also be linked with the observations on glycosidic compounds in hops (1). Only where the 600 BB beer was concerned could a somewhat broader bittering be ascertained. The results were unchanged when the beer-tasting was repeated with the same samples. In the second round again the same tasters preferred neither the neutral beer nor the beers with HPF. According to this, definite sensory effects can be attained with the addition of a HPF which the majority of tasters judged positively but some negatively. This basic judgement did not change after 4 weeks storage at 27C. The beers had aged badly over 15 months at 22C. The beer-tasting again produced clear results. The comparison beer without HPF was totally aged and was undrinkable. This was followed by the beer with 600 g HPF, hopped at the beginning of the boil (600 BB). The beers with 300 and 600 g HPF - hopped at EB - had aged but were perfectly drinkable. However it must be taken into consideration that the freshly bottled beer from the pilot brewery had a total oxygen content of about 0.8 to 1.0 mg/l. Unfortunately negative effects offset these positive results (see Chart 4). The nitrate values of the beers correspond to the supplementary amounts hopped with the HPF and increased considerably. The physical stability also deteriorated correspondingly.

The beer-tasting results of the 2nd series with 50 up to 300 g HPF can be summarized as follows: The addition of a HPF can be seen with 50-100 g/hl and one-star significance, with 150 g/hl and two-star significance and from 300 g/hl upwards with three-star significance. The significance of the classification decreased with a longer boiling time and with bitter hops. Having tasted the beer several times a minority of the tasters preferred the comparison beer, a majority the beers with HPF. This preference was clearer when the boiling times were short and when aroma hops were added. The beers with HPF were designated typically hoppy, fruity, grassy. A so-called tannin bitter was only found in beers from 300 g/hl HPF upwards and with a long boiling time.

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From this the following conclusions for the dosage of hop polyphenols can be drawn: There is no negative influence on foam and colour of the beers. No hard bitters occur with shorter boiling times. The opposite tends to be more likely. Definite tastes can be achieved. The reducing power of the beers increases, the stability of taste improves. This applies at least for the pilot draughts with relatively high oxygen content. However the tendency to become turbid increases especially where there are longer boiling times. The nitrate content increases corresponding to the size of the dosage.

In practice this means a deliberate dosage of hop pellets in fact amounting to about 50 g/hl upwards. The actual expectations relating to the nitrate content in beer and the physical stability act as a brake for high dosages. They possibly limit them being used to a greater extent for qualitative or physiological reasons. Attempts can be made to make deliberate use of hop polyphenols up to these limits which apply for every brewery or for the individual beer type. Here the following recommendations can be given: Suitable hop varieties should be selected with a higher proportion of low-molecular polyphenols (e.g. aroma varieties from moderate climatic zones). Fresh hops or pellets should preferably be used instead of older ones, in order to maintain the low-molecular character of the polyphenols. The boiling time for the respective hopping in the wort-kettle should be limited. If e.g. the nitrate determines the possible amounts of pellets, pure-resin extract can be added at the beginning of the boil and pellets only after the boiling time is half-way. In order to keep the doses of polyphenols constant over the years, the annual fluctuations of the (-acids can be balanced out by means of a flexible mechanical concentration (variable type). Therefore the ratio (-acids to polyphenols) remains constant. A lupulin concentration makes sense as the hop spindle with a relatively low polyphenol content is discharged and therefore the result is a more favourable ratio of polyphenols to nitrate in the pellet.

Polyphenols during the brewing process

The dissolving behaviour of polyphenols in wort and beer is not the same for all components. Hydrophilic groups of substances such as hydroxybenzoin - or hydroxy cinnamic acids, flavanols or proanthocyanidins dissolve more easily than the more lipophil prenylflavonoids; flavonoids are in between. Supplementary to Chart 2, reference values can be calculated for the polyphenol yields from the 1st series of brewing trials. In this respect the unspecific EBC-colouring method was compared with the specific HPLC method. Chart 5 shows the results. The initial values of the HPF used were: 5.2 weight -% polyphenol content according to EBC 1.2 weight -% calibrated polyphenols with HPLC (without xanthohumols)

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The average yield of the low-molecular HPLC polyphenols is with 66% higher than that of the EBC polyphenols with 40%, which is due to the better solubility of the hydrophilic, low-molecular structure. Losses occur due to thermal conversions and eliminations with hot or cool sludge, yeast and dregs as well as during filtration. As far as the prenylflavonoids are concerned (main representative: xanthohumol) the ratios are different. Xanthohumol is not as soluble as all the other polyphenols and must first be isomerized similar to the (-acids. Chart 6 shows the development resulting from one trial. In this trial 5.0 mg xanthohumol were added per liter wort with an average boiling time of 35 min. For the most part xanthohumol is converted into isoxanthohumol. The high losses can be explained by poor solubility and adsorption. Even with long boiling times and reduced discharge due to turbidity and filtration only about 30% yield in the form of isoxanthohumol can be expected in relation to the xanthohumol used. Polyphenols causing turbidity are extracted from beer that is treated with PVPP to make it keep longer or improve its physical stability. The amount g/hl of PVPP added is primarily regarded as variable, but possibly the type of PVPP used is too. As an example Chart 7 shows the reduction of some polyphenol groups through a PVPP filtration. There were differences in a moderate and in a strong treatment. In fact there is a stronger adsorption of the proanthocyanidins and flavonols causing turbidity, but other polyphenols are reduced. The PVPP treatment is therefore not a strictly selective process of totally removing the polyphenols which cause turbidity.

Summary
The dosage of hop polyphenols does in fact go along with an increase in nitrate and an increased tendency to turbidity. On the other hand when a technologically specific procedure is used, their application is linked with interesting sensory effects. Therefore there is no reason to refrain from using hop polyphenols under the doctrine of low nitrate and high turbid stability. Recently modern selective HPLC methods facilitate the choice of suitable raw commodities, e.g. fresh aroma hops from temperate production zones. Preferably medium and short boiling times instead of long boiling times. Therefore neither the colour nor the foam on the beer suffers, the reduction power increases, taste and taste stability can be influenced positively. For brewing technologists, who want to develop a polyphenol philosophy there are sufficient findings and possibilities to produce one.

Literature: see previous article by M. Biendl

BARTH-HAAS GROUP
HOPS ARE OUR WORLD

BARTH-HAAS GROUP
HOPS ARE OUR WORLD

BARTH-HAAS GROUP
HOPS ARE OUR WORLD

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