Phenolic acids in beer are important because they can be decarboxyiated to phenols, which usually impart off-flavours. An improved high performance liquid chromatographic system was used to monitor phenolic acids and phenols during the brewing process. Ferulic acid was the most significant phenolic acid found in beers prepared from malted barley. Extraction of ferulic acid from malt involved an enzymatic release mechanism with an optimum temperature about 45°C. Mashing-in at 65°C significantly decreased the release of free ferulic acid into the wort. Wort boiling produced 4-vinyl guaiacol by thermal decarboxylation, in amounts (0.3 mg/L) close to its taste threshold, from worts that contained high contents of free ferulic acid (> 6 mg/L). The capacity of yeasts to decarboxylate phenolic acids (Pof* phenotype) was strong in wild strains of Saccharomycoa and absent in all lager brewing yeast and most ale brewing yeasts. Some top-fermenting strains, especially those used in wheat beer production, possessed a weak decarboxylating activity (i.e. Pof1). During storage of beers there were appreciable temperature-dependent losses of 4-vinyl guaiacol. These results indicated that the production of 4-vinyl guaiacol is amenable to close technological control.

Acetone: water (3 : 1) extracts of milled barley grains contained simple monomeric, dimeric and trimeric flavanols in addition to higher molecular weight flavanoid tannins.Whereas ( + )-catechin and the simple individual oligomeric proanthocyanidins were readily separated by h.p.1.c. and t.1.c. fine resolution of the tanning components was not accomplished. Adsorption chromatography on Sephadex LH-20 permitted group separation and measurement of tanning components which were characterised as polymers of (+)-catechin and (+)-gallocatechin. Using h.p.1.c. little change in the contents of simple flavanols was detected during the malting of barley, neither were there substantial differences in flavanol contents between five different varieties of barley. The tannins appeared to be formed in the grain prior to harvesting, possibly by oxidation of simple flavanols, and were not artefacts of post-extraction treatments.

The highest concentrations of complexed and polymeric flavanols were found in the early runnings from a small-scale mash tun. Formation of break during and after boiling was accompanied by the removal of flavanols from solution, especially the complexed and polymeric fractions. Variations in boiling procedure affected the amount of break formed, the amount of residual soluble flavanols and the colloidal stability of the resultant worts. Colloidal stability in beer was influenced by the contents of simple flavanols and the availability of air during storage. Dimeric and trimeric flavanols seemed especially susceptible to oxidation which implied an important role in haze formation. Flavanols found in commercial beers were mainly monomeric and complexed forms.

The flavanoid polyphenol extracts from barley and hops were each separated into six fractions by adsorption chromatography on Sephadex LH20. These fractions were further characterised by several analytical methods, including high-performance liquid chromatography and a colorimetric measurement of polymerisation index. The tanning powers of the fractions were graded according to their reactivities with cinchonine sulphate solution in a standardised turbidometric test. Whereas, almost 75% of the flavanols from Ark Royal barley were non-tanning oligomers almost 96% of the flavanols from Bullion hops were polymeric tannins. Reactivity of most of the barley flavanols with cinchonine sulphate was increased greatly by oxidation with peroxidase and hydrogen peroxide. Some effects of polymerisation, caused by enzyme action or by exposure to air, on oxidisable polyphenols (nontannins) were measured using ( + )-catechin. procyanidin B3 and prodelphinidin B3 in model systems. These, and other measurements on experimental and commercial beers indicated that oxidation of simple flavanols from barley produced polymers with tanning properties. In contrast, the hop flavanols when extracted apparently in their native forms, were capable of co-precipitating with polypeptides in beer. Treatment of beers with different stabilising agents, such as Polvelar AT and silica hydrogel, retarded haze formation by restricting 'proteinpolyphenol' interactions.Key words: flavanols, haze, high-performance liquid chromatography. oxidation, polyphenols. tannins. IntroductionAlthough it is recognised that non-biological haze in beer may be generated through several possible routes,30 the popular and time-honoured concept is that the tanning of polypeptides" by flavanoid39 polyphenols is the primary cause. Monomeric and dimcric flavanols14"1920"-43 such as are present in barley,39 have relatively weak tanning powers themselves but can be transformed19 either by oxidation or acid-catalysis into polymeric tannins. Modern chromatographic methods28-3"6 have confirmed earlier claims9-20 that beers invariably contain pools of flavanoid haze precursors. Fortunately, acceptable colloidal stability can be achieved by a variety of methods,2"'■4246-52 not the least of which is the prevention of oxidation.41 Owing, perhaps, to the suc cesses of contemporary stabilisation procedures interactions of the heterogeneous beer 'proteins' and polyphenols have not been studied in detail. While probing the chemical mechanism of some local processing features known to influence colloidal stability a need was seen for a realistic experimental model of protein-polyphenol interactions in wort and beer. Consequently, it was decided to elaborate on the investigations made with selected simple flavanols141619 and include more examples from the wide range of flavanoids"-36-18 obtainable from barley and hops.The work described herein preceded an investigation of protein-polyphenol interactions which will be reported later.37 At the outset it was necessary to isolate flavanols fr...

Four high-molecular weight proteose fractions from beer were precipitated to varying extents by four different tannin fractions, as shown turbidimetrically in a simple assay. According to this test the most acidic proteose fraction reacted least with all four tannins and the formation of insoluble proteose-tannin complexes was strongly dependent on pH. Moreover, tannins obtained from hops displayed greater tanning power with beer proteoses than did either a tannin isolated directly from barley or one prepared by aerial oxidation of barley prodelphinidin B3. The different reactivities of tannins to proteoses, however, did not correlate well with their reactivities to cinchonine sulphate (CS). Whereas the amounts of break precipitated on boiling a wort moderately were related to the amounts of hop tannins added, barley flavanols had little effect on break formation. In beer, about 50% of the proteoses were of low molecular weight (< 10,000 Daltons) and some were complexed with flavanols. In contrast, isolated fractions of high molecular weight beer proteoses (MW> 10,000 Daltons) were not associ ated with flavanols. The most acidic of these proteoses were the least stable in solution, being precipitated from beer by down-shifts in pH. The tanning power of unstabilised beer as measured by reaction with cinchonine sulphate was shown to arise from the cumulative effects of at least four different beer components, which should be considered when interpreting the effects of different haze-stabilisation treatments.