A-Acids in beer

The requirements for the analysis of the iso-a-acids are diverse and, to a large extent, dictate the resolving power required of the chromatography. The requirement to separate cis/trans pairs of the iso-a-acids is often driven by research projects. Hughes (16) used a multicomponent mobile phase and a NovaPak C18 cartridge to obtain resolution of the five iso-a-acids in beer within 7 minutes (Fig. 3). The separation of cw-isohumulone (often the major iso-a-acid in beer) from frans-isoadhumulone (most minor of the six iso-a-acids) usually proves to be difficult. Other workers have successfully resolved the six compounds, and micro HPLC has proved particularly effective (34). 

Fig. 3 Resolution of the six major iso-a-acids in beer into five bands by reverse-phase HPLC. Mobile phase acetonitrile water methanol 0.2 M magnesium acetate (aq) formic acid trifluoroacetic acid (840 490 480 24 3.6 1.2 v/v/v/v/v/v). Column NovaPak C18 5 /rm cartridge (100 X 8 mm i.d.) under radial compression. Mobile phase flow rate = 2 ml min-1. (Chromatogram redrawn from original).

R Szucs, J Vindevogel, P Sandra, LC Verhagen. Sample stacking effects and large injection volumes in micellar electrokinetic chromatography of ionic compounds direct determination of iso-a-acids in beer. Chromatographia 36 323-329, 1993. 

JG Clark, LJ Burroughs, JA Gusinski. High-performance liquid chromatography of tetrahydro-iso-a-acids in beer. J Am Soc Brew Chem 56 76-79, 1998. 

PH Todd, PA Johnson, LR Worden. The assay of pure iso-ar-acids and reduced iso-a-acids in beer. Proc Am Soc Brew Chem, 1972, pp 30-32. 

The ASBC [108] retain the second method of Ribgy and Bethune [110] as it provides a more accurate estimate of the iso-a-acids in beer. In this... 

Intelmann D., Hofmann T. On the autoxidation of bitter-tasting iso-a-acids in beer. Journal of Agricultural and Food Chemistry, 58 ... 

A device described by Sawyer and Dixon [13] was used for the determination of alcohol and acid in beer and stout. Attempts to improve the reliability of this method and to improve the signaTto-noise characteristics of the measurements prompted a critical design described by Lidzey et al. [14]. This unit overcomes many of the fluctuations in results observed with use of the first unit in this a number of possible sources of surging were indicated and these were not controlled owing to the varying conditions in the coil. In addition, the separation of the waste involatile material from the volatile phase took place outside the heated flask distillation unit. Air bubbles present in the segmented stream were also responsible for considerable surging. 

Nardini, M. Ghiselli, A. 2004. Determination of free and bound phenolic acids in beer. Food Chem. 84 137-143. 

Hops (Humulus lupulus L.) belong to the mulberry family (Moraceae). The leaf and flower cone with luplin (yellow secreted particles) are used to flavor beer, and to alleviate thirst as a nutraceutical. Air-dried hops contain a-acids in 2-12% yields [102]. The a-acids are found in a lead-acetate-precipitate derived from the methanol extract of hop. The chemical structures are shown in Fig. 14a [103,104]. Humulone is the most easily obtainable a-acid by repeated crystallization. Isomerization of the a-acids is the most important reaction in beer brewing, and is easily induced by boiling. Beer contains 20-25 mg/L of iso-a-acids, which mainly contribute to its bitterness. 

SieberL K.J., Modeling the flavor thresholds of organic acids in beer as a function of their molecular properties. Food Qual. Pref, 10, 129-137, 1999. 

Marshall, P A., Trenerry, V. C., and Thompson, C. O., The determination of total ascorbic acid in beers, wines, and fruit drinks by micellar electrokinetic capillary chromatography, J. Chmmatogr. ScL, 33, 426, 1995. 

The example cited is significant, for it illustrates the role of an SCF in influencing product distribution. Most other studies illustrate its role in changing the rate constant of the reaction. A good example of the latter is the conversion of hermalones (a-acids) in a mixture with lupidones ( 9-acids) to the trans-iso form via an oxidative ir-methane rearrangement (Andre et al., 1985). Both acids are extracted from hop and contribute to the bitterness of beer, but the a-acids must be converted to the iso-a form before addition to the beer. 

The level of a-acid in hops falls during storage but the bittering potential of the hops does not fall to the same extent (Fig. 13.5). This is because many of the oxidation products of both a- and p-acids discussed in Chapter 13 are capable of bittering beer. However, at the present time, the nature of... 

The estimation of iso-a-acids in wort, beer, and isomerized hop extracts is usually based on their light-absorption properties (Table 14.12). Measurements are made at 275 nm in acid solution or 255 nm in alkaline solution. Humulinic acids, which possess no bitterness, show similar light absorption to the iso-a-acids. Both a- and p-acids show appreciable absorption at the wavelengths mentioned so all these must be absent from the extract analysed. Hulupones exhibit 80-90% of the absorption of the iso-a-acids at these wavelengths but, since they are also bitter, it is often not regarded as so important to exclude them from the analysis. Beer produced by conventional wort boiling will contain only trace amounts of a-acids and humulinic acids, and iso-a-acids may be estimated directly on an isooctane extract of the beer. Worts, on the other hand, may also contain appreciable amounts of a-acids and isomerized hop extracts may contain a-acids, iso-a-acids, and humulinic acids. 

The nature of the other bittering principles in beei brewed from old hops is still largely unknown. These substances probably contain more oxygen than the iso-a-acids, are more polar, and incompletely extracted into isooctane. The abeo- %o-aL-2iC ds (p. 487) are such compounds, the concentration of which in lager beers was reported to be 88-160 mg/1. Subsequent work established the presence of polyphenols in the extracts analysed and found that the concentration of f6 o-iso-a-acids in English beers was less than 6 ppm [57]. Many other oxidation products of the hop resins have been detected in beers (see Chapter 12) but in most cases it has not been established that they are normal constituents. 

Of the individual iso-a-acids, isohumulone is concentrated into the foam to a greater extent than isocohumulone and unhopped beer bittered (21 0 BU) with isohumulone had a better head retention (2 = 132) than that bittered with isocohumulone (2 = 115) [146]. Addition of iso-a-acids to beer increases head retention but, in particular, the iso-a-acids are responsible for foam adhesion, cling or lacing. Unhopped beer does not show these effects. Traces of heavy metal ions (iron, cobalt, nickel and copper) also improve the head retention of beers but only in the presence of iso-a-acids [139]. 

Horak, T., Culik, J., Jurkova, M., Cejka, P, Kellner, V. (2008). Determination of free medium-chain fatty acids in beer by stir bar sorptive extraction. Journal of Chromatography A, 1196-1197, 96-99. http //dx.doi.Org/10.1016/j.chroma.2008.05.014. 

Gluconobacter (Acetomonas) G. oxydans. Produce acetic acid in beer sometimes giving a cidery note. 

Montanari L, Perretti G, Natella F, Guidi A, Fantozzi P (1999) Organic and phenolic acids in beer. LWT - Food Sci Technol 32 535-539... 

Following the publications of Stiittgen and Beer on the use of vitamin A acid in the treatment of ichthyosiform erythroderma, Thomson and Milne reported the treatment of a seven-year-old girl with ichthyosiform erythroderma (epidermolytic hyperkeratosis) (Figure 21.1) by topical and systemic retinoic acid. 

On the other hand, the hop bitter acids constitute such a small fraction of the beer cost that a gain in utilization yield is not relevant with respect to a change in beer quality. In our opinion there is little chance that isomerized extracts will ever conquer the market. However, isomerized extracts can help though to correct undesirable situations for instance to increase the bitter level of beers which, for some reason are not bitter enough. 

In our first paper (24) on the LC of hop bitter acids, no attempt at quantification was made. In 1980 we presented a first quantitative LC method for alpha acids with chalkone as internal standard (31). The adverse effect of trace metals in the chromatographic system, although small, was clearly demonstrated. In 1981 we described a method for the analysis of the iso-alpha acids in beer (32) using iso-octane extraction and with 2.6-di-t.butylphenol as internal standard. Direct injection of beer as such on a reversed phase LC column can simplify the analytical procedure, as described by our laboratory in 1982 (33). This development seems not to have... 

Iso-alpha acids need to be determined in beer, in wort, in ethanol hop extracts and in pre-isomerized hop extracts. Undoubtedly the quantitative determination of iso-alpha acids in beer is the most important. Analysis of iso-alpha acids in wort may also be worthwhile to establish the degree of alpha acids conversion into iso-alpha acids (isomerization yield, utilization yield). This can indeed be very different from one brewing installation to another and can also vary with the nature of, for instance, the hop extract used. Quantitative analysis of the iso-alpha acids content of worts may help solve problems in this respect. The issue of iso-alpha acids in ethanol hop extracts has already been mentioned in Chapter 16 on alpha acids analysis. Pre-isomerized hop extracts have of course to be analysed. The iso-alpha acids content claimed by the manufacturer must be checked and the level of iso-alpha acids must be controlled/followed in time. Our experience indicates that mostly the claimed figures for isomerized hop extracts are much too high. Such extracts can also show a dramatic decrease in iso-alpha acids content in only a few months. 

There is a small amount of alpha acids in beer, up to 4 ppm. It has been claimed that alpha acids in beer have an anti-gushing effect. Where do these alpha acids come from The simple answer is to believe that they are just dissolved from hops, but there are reasons to doubt this. When isohumulone is shaken in a two-phase system of iso-octane buffer pH 5.0, up to 10% humulone can be formed (9,10). This point would be interesting to study by LC. If correct, it would mean that the alpha acids found in beer are not unreacted material derived from hops, but could be formed in beer from the iso-alpha acids during storage or maybe on pasteurization. Could this be a quality criterion or a beer age criterion Is older beer less bitter because some iso-alpha acids have been transformed into alpha acids Monitoring this in stored beer by LC would probably answer the question. 

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