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Evaluating the methods

 

Unfortunately the methods described do not produce identical results when several analyses of a sample are made. In judging the methods Dr. Adrian Forster says in his publication "Wie analysiert man Hopfen" ("How to analyse hops")(1993):

How do you evaluate the methods?

A whole number of criteria can be applied here somewhat like the following:

1st Group: Accuracy, reproducibility, strength of evidence. These points embrace the quality of a method. How accurately can the desired substance be analysed? Do several assistants In a laboratory obtain the same result on different days or do they vary? How do the results vary if several laboratories examine the same types of samples? These points all come under the term "Reproducibility", by which it can be ascertained by statistical formulas what deviations must be expected when using a certain method.

By strength of evidence this means what information is obtained overall. There are methods which not only produce one value but several. Thus the complete Wšllmer analysis can produce the total content of bitter compounds (=total resin) and the aging degree (hard resin content) as well as the alpha-acids. The ASBC-method provides an aging index as well as the alpha-acids. The HPLC is the most informative method as characteristics of the specific variety can even be ascertained as well as alpha- and beta-acids and an indication on aging.

2nd Group: Apparatus required, time needed, capacity, use of solvents, other expenses. Last of all this group encompasses points which are included in the costs for an analysis. How expensive is the equipment? How long does it take until the result is available? Of course in processing hops it is important to reach a result quickly. If several analyses can be carried out simultaneously, how many analyses does a person/a team make per day? In other words: What use is the best analysis if only very few results are produced per employee and per day? Furthermore it is interesting to know how high the consumption of solvents and other material is. Finally it is also important what qualification the analyst must possess in order to cope with handling the equipment. Therefore all questions concerning the cost of an analysis are contained in this point.

3rd Group: Safety, health risks, environmental compatibility. Understandably this group has played an increasingly important role in recent years. Is a solvent which is required for the analysis explosive, similar to e.g. petrol? Are there any risks for human beings in handling it? Is a solvent harmful to health? Generally it must be said that all the solvents required for hop analytics are subject to certain reservations, anyway it is important to make sure that the laboratory is well ventilated.


 

However a few solvents are subject to special suspicions. They can cause headaches and nausea, some such as benzole are suspected of being carcinogenic. In any case solvents do have to be disposed of after use, but even so they should not cause any harm to the environment in the waste water etc.

To summarize this chapter, we have to recognise that many points contribute to the overall evaluation an analysis. Compromises often have to be met which is also the reason why there are several methods for one and the same measuring problems.

Survey for the most important analyses for bitter compounds


In the following survey the most important methods are listed together under the criteria: Reference, Abbreviation, Measuring Principle, Solvent and Evidence: :

Reference


EBC 7.4 & ASBC 6


EBC 7.5 &
Mebak 5.1.5.1

EBC 7.5 &
Mebak 5.1.5.1


ASBC 9.6


EBC 7.7
Abbreviation


1. EBC-Toluol 1)


2a Woellmer 1)


2b Volle Woellmer


3. Spectro 2)


4. HPLC 3)
Measuring Principle

Conductometry


Conductometry


Gravimetry +
Conductometry


Spectral photometry


High pressure liquid
chromatography
Solvent


Toluol + Methanol

Ether + Methanol

Ether, Methanol +
Hexan

Toluol + Methanol

Ether, Methanol

Statement


Alpha-acids


Alpha-acids


Total resins,
soft resins,
Hard resins
Alpha- + beta-acids +
aging index

Alpha- + beta-acids, indication
of variety and aging 


1) Unspecific value for alpha-acids
2) Unspecific value for alpha- and beta-acids
3) Specific value for alpha- and beta-acids


Here the terms "specific" and "unspecific" should be briefly explained. The result of an analysis can be considered specific when it is guaranteed that only the one desired substance has actually been recorded.
Therefore a separation process ( = chromatography) must generally be used. Figure 1 shows for example a HPLC chromatogram of the bitter acids of a fresh hop. With the alpha- and beta-acids two peaks can be seen in each case, i.e. co-alpha and n- + ad-alpha-acids as well as co-beta- and n- + ad-beta-acids. 
 

analytik6_1 analytik6_2


Fig. 1: Fresh Hallertauer Hersbrucker          Fig. 2: Aged Hallertauer Hersbrucker


The height of the co-peaks compared with the other two (n- and ad-peak) is lower in the case of aroma hops such as Hersbrucker, Tettnanger and Spalter than with bitter hops. 

Therefore with the HPLC we can be sure that we only measure alpha- or beta-acids.


As a comparison Figure 2 shows the chromatogram of a badly aged hop. The losses of alpha- and beta-acids can be seen immediately. In addition to this some new peaks are visible Ð substances which have resulted from separating the bitter acids.

On the other hand unspecific methods such as conductometry or spectral-photometry do without a separation (chromatography). They make use of specific characteristics of the alpha-acids Ð the spectral-photometric method e.g. the optic characteristics of the alpha-acids. On the other hand the conductometry (measuring conductibility) uses the property of the alpha-acids to bond with a salt. Besides the alpha-acids other hop components also possess the properties mentioned (light absorption in the spectral-photometry and salt formation in the conductometry). Consequently these components are also in the measurement. Therefore a somewhat higher measurement is obtained with unspecific methods than with specific ones.  

Principle of the conductometry

Humulone + lead acetate = humulate

The tannins extracted from the sample are titrated conductimetrically with Pb(AcO)2 (lead acetate).

The humulones react with the lead acetate and form lead salts. When all the humulones (or iso-humulones) have reacted, the conductometer value increases. What makes the analysis unspecific is the fact that other hop components can also cause interference during measuring.

The use of lead acetate is entered in a formula with which the alpha-acid content of the sample is calculated as a percentage.
In the case of fresh hops the results correlate with the actual alpha-acid content of the sample.

If on the other hand the hops or the hop product are old and oxidized, the aging components distort the result of the analysis.

Reaction curve EBC 7.5 
Reaction principle