Proceedings.

Annual meeting - American Society of

Brewing Chemists

ISSN: 0096-0845 (Print) 2576-1315 (Online) Journal homepage: https://www.tandfonline.com/loi/ujbc19

An Index of Deterioration in Hops (Humulus

Lupulus)

S.T. Likens, G.B. Nickerson & C.E. Zimmermann

To cite this article: S.T. Likens, G.B. Nickerson & C.E. Zimmermann (1970) An Index of
Deterioration in Hops (Humulus�Lupulus), Proceedings. Annual meeting - American Society of
Brewing Chemists, 28:1, 68-74, DOI: 10.1080/00960845.1970.12006963

To link to this article: https://doi.org/10.1080/00960845.1970.12006963

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68
An Index ofDeterioratlon in Hops
(Humulus Lu.pulus)'
S.T. LIKENS, G.B. NICKERSON, and C.E. ZIMMERMANN 2 , Oregon State
University, CorvaUis, Ore., and U.S. Department of Agriculture, Prosser,
Wash.
T he spectrophotometric method for
the determination of a- and (3-acids
in hops (3) is simple, rapid and accurate.
Consequently, it has replaced the gravi-
metric method and is now used almost
exclusively for the evaluation of hops in
the United States.
The gravimetric method (2) provided a
value for hard resins (mainly oxidation
products of a- and (3-acids) and thereby
gave an indication of the extent of deteri-
oration which had taken place prior to
examination. Since the gravimetric meth-
od was abandoned, analysts have had no
clear indication of the stage of deteriora-
tion of the hops at the time of evaluation.
This paper deals with an index of
deterioration derived from "the method of
Alderton et al. (1). The index is based on
the same spectral measurements that are
used in the spectrophotometric method
and no additional analytical procedures
are necessary.
Hop acids in alkaline methanol absorb
light much more strongly at 325 mlJ.
(A 3 2 S ) than at 275 ffiIJ. (A 2 7 S ) ' Con-
versely, their oxidation products absorb
more strongly at 275 ffiIJ. than at 325 mu.
Thus, as oxidation progresses, A 3 2 5 de-
1 Cooperative investigations of Crops Research
Division, Agricultural Research Service, U.S.
Department of Agriculture, and the Oregon
.Agricultural Experiment Station. 'Technical
paper No. 2888 O.A.E.S. Corvallis, Oregon.
This work was supported in part by the U.S.
~rewers Association,
Chemists, Crops Research Division, Agricul-
tural Research Service, U.S. Department of
Agriculture, Oregon Agricultural Experiment
Station, Corvallis, Oregon; and Plant Physiolo-
gist, Crops Research Division,'Agricultural Re-
search Service, U.S. Department of Agriculture,
Prosser, Washington, respectively.
HOP DETERIORATION
creases, A 2 75 increases, and the ratio
A 2 7 5 / A 3 2 5 becomes a sensitive indica-
tion of the extent of completion of the
reaction (Fig. 1).3
Alderton et al. (1) determined that the
specific absorption coefficients for hop
acids in alkaline methanol at 275 mlJ. and
325 mlJ. were 9.0 and 38.1, respectively,
for a-acids, and 3.7 and 33.1, respectively,
for (3-acids. A 2 75/ A 3 25 for the pure com-
pounds are 0.24 for a-acids and 0.11 for
(3-acids. A 2 75/A 3 25 for all proportions of
the two hop acids lies between 0.24 and
0.11, and can be calculated by:
A2 75/ A 3 25 for combinations of hop
9.0 X %a + 3.7 X %(3
acids
38.1 X %a + 33.1 X %(3
Any A 2 75/ A 3 2 5 in excess of that which
can be accounted for in the above equa-
tion is necessarily the result of absor-
bance by compounds other than a- and
(3-acids, but possibly related to them.
Experimental Results
Effect ofMaturation
Rates of synthesis of hop acids change
during the growing season (4). Beta-acids
synthesis is greater during the early
stages, and a-acids synthesis is greater
during the later stages of maturity. Conse-
quently, values for A 2 7 5 / A 32 5 would
increase during maturation until the pro-
portions of a- and (3-acids become con-
stant. A maturity series was run on the
variety Bullion to determine the extent to
which A 2 7 5 / A 3 2 5 was affected (Table I).
3 A3 5 5 could have been used to represent hop
acids as well as An s- The latter was selected
simply because it is associated with the more
valuable a-acid.
A.S.B.C. PROCEEDINGS 69

1.0

% (<<.,e>
LOST
0.8
100 2.4

...u
..
0.•
z
c

.
0
II:
II)

c 0.4
50

0.2 0 0.20

0.0 .....- - - - - - - -........- - - - - -.......- - - - ' - -....

275 325 355
WAVELENGTH (mill

Figure 1. Typical spectra ofalkaline methanol solutions obtained from

hops in various stages of deterioration. A 325 decreases as hop acids are
oxidized and A 2 7 5 increases as oxidation products accumulate, result-
ing in proportional increases in A 2 75/A325.

After August 9 there was no significant
change in the proportion of a- and {3:acids
or in A275/A325 which was attributable
to the two acids. Excess A2 75'1 A 325 was
0.01 to 0.06 through the entire series
which indicated very little contribution
by compounds other than hop acids. A
small amount of oxidation of hop acids
could be expected during drying, packag-
ing, and grinding of the samples in pre-
paration for analysis. Conversion of 0.2%
a- and 0.1% {3-acids to _their oxidation
products would account for the excess
A275/A325 found in Table 1.

TABLE I
Maturity Series of the Hop Variety Bullion a
mloill A275/A325
100g a-acids {j-acids Thear-
Date hops (%1 (%1 Found etical Excess

3Aug. '66 0.18 7.9 4.7 0.22 0.16 0.06

6Aug. '66 0.22 8.5 4.7 0.23 0.19 0.04
9 Aug. '66 0.42 10.1 5.4 0.22 0.20 0.02
12 Aug. '66 0.62 9.4 5.0 0.24 0.20 0.04
15 Aug. '66 1.15 11.3 6.0 0.22 0.21 0.01
18 Aug. '66 1.54 11.7 5.6 0.22 0.20 0.02
21 Aug. '66 1.71 10.8 4.3 0.22 0.20 0.02
25 Aug. '66 2.44 11.7 5.0 0.24 0.20 0.04
28 Aug. '66 2.67 10.9 5.6 0.26 0.20 0.06
31 Aug. '66 2.92 10.6 4.5 0.25 0.20 0.05
2 Sept. '66 3.30 11.3 5.6 0.25 0.21 0.04
aOil contents were determined by the method of Wright and Connery (61. a- and {j-acids were
determined by the spectrophotometric method (A.S.B.C.1. Results are adjusted to a common basis of
8% moisture in the hops.
70 HOP DETERIORATION

TABLE II
Loss of Hop Acids by Nine Varieties and Corresponding
Increases in A2 7 5 /A 3 2 5 During Storage at 72° F.
a-t{3
Months in a-acids (3-acids a+~ lost
Identification storage (%) (%) (%) (%)

Brewers Gold o 8.2 4.0 12.6 o 0.26

(seeded) 3 6.1 2.6 8.7 31 0.55
6 4.5 1.8 6.3 50 0.82
10 0.1 2.6 2.7 79 1.23
Brewers Gold o 10.9 5.1 16.0 o 0.25
3 7.2 3.3 10.5 34 0.56
6 4.9 1.9 6.8 58 0.93
10 4.6 0.6 5.2 68 1.43
Bullion o 11.2 5.8 17.0 o 0.24
3 8.8 3.7 12.5 26 0.47
6 5.1 2.1 7.2 58 0.94
10 2.0 1.7 3.7 78 1.39
Early Cluster o 7.4 5.0 12.4 o 0.22
(clone E-2) 3 7.2 4.8 12.0 3 0.24
6 6.2 5.0 11.2 10 0.28
10 6.1 4.0 10.1 19 0.32
Fuggle o 4.8 2.6 7.4 o 0.27
3 4.4 2.3 6.7 9 0.33
6 3.7 2.1 5.8 22 0.45
10 3.3 1.3 4.6 38 0.61
Talisman o 9.9 4.8 14.7 o 0.26
3 9.5 4.4 13.9 5 0.29
6 7.9 3.7 11.6 21 0.42
10 2.7 3.6 6.3 57 0.79
Yakima Cluster o 9.1 5.6 14.7 o 0.22
(clone L-1) 3 8.8 5.7 14.5 1 0.25
6 8.6 5.8 14.4 2 0.28
10 8.0 4.7 12.7 14 0.33
56008 (Expt'l) o 7.4 6.2 13.4 o 0.22
3 5.7 4.7 10.4 22 0.39
6 3.9 3.1 7.0 48 0.73
10 2.5 1.2 3.7 72 1.27
56013 (Expt'l) o 6.6 6.2 12.8 o 0.22
3 4.4 4.1 8.5 34 0.53
6 2.5 2.5 5.0 61 0.90
10 1.6 0.9 2.5 80 1.52
66030 (Expt'1) o 12.8 5.8 18.6 o 0.24
3 11.5 5.1 16.6 11 0.33
6 8.2 3.3 11.5 38 0.60
10 5.0 1.0 6.0 68 1.15

Relationship ofA27sIA32s to Disappear- analyzed spectrophotometrically shortly

ance of Hop Acids
A second experiment was run (a) to
establish the quantitative relationship of
observed A 2 75/ A 3 25 to the extent of
oxidation of the hop acids, and (b) to
exclude the possibility of interference in
A2 75/ A 3 25 by unrelated compounds
which might arise during storage. Nine
varieties, representing all degrees of stor-
age stability, were used. Each was
after harvest and subsamples were placed
into storage at 72°F. in a dark room for
subsequent analysis. Samples were re-
moved and analyzed after three, six, and
10 months (Table II).
The sum of a- and ~-acids ranged from
7.4% for the variety Fuggle to 18.6% for
66030, an experimental genotype. There-
fore, it was necessary to convert all values
for hop acids to "per cent lost" in order
A.S.B.C. PROCEEDINGS 71

TABLE III
Indicated Loss of Hop Acids From 62 Commercial Samples of Bullion
and Brewers Gold from the 1968 Crop. Analyzed Between
4 Sept., 1968 and 4 Oct., 1968.
Range No. Avg. Indicated
A275/A325 Samples A275/A325 % Hop Acids loss (%)a

<0.30 12 0.277 15.59 6

0.30-0.32 23 0.309 14.93 10
0.33-0.35 14 0.332 14.69 13
0.36-0.39 11 0.375 13.46 19
>0.39 2 0.440 11.45 25

aT;lken from Figure 2 for the average A275/A325 found for each group.

to put the extent of their deterioration apparent. The weighted-average loss of

on a common basis.
Rates of deterioration are varietal
characteristics and, as expected, were
very different for the nine varieties. For
example, 56013 (experimental) lost 80%
of its hop acids after 10 months, while
Yakima Cluster lost only 14% during the
same period. No interferences were evi-
dent, however, and A 2 75/ A 3 25 reflected
the extent of deterioration equally well
for all varieties.
Graphic representation revealed that
the relationship between A2 75/ A 3 2 5 and
the extent of deterioration of hop acids
was logarithmic (Figure 2). Extrapolation
of the regression to complete degradation
(100% lost) gave a value of approximately
2.5. Values from 2.4 to 2.6 were found
when samples of lupulin from several
varieties were oxidized in air at elevated
temperatures until no further change in
A2 75/ A 3 2 5 took place.
Indication of Deterioration in Com-
mercial Hops
A third set of data was compiled from
the official analyses of 62 samples of
commercially produced Bullion and
Brewers Gold during September and
October of 1968. A 2 75/ A 3 25 values for
many of the samples exceeded 0.30, indi-
cating that significant losses of hop acids
had occurred by the time of analyses.
Values found for A 2 75/ An 5 and hop
acids content were averaged for relatively
narrow ranges of observed A2 75/A 3 25
(Table III). The association of decreasing
hop acids with increasing A27dA325 is
hop acids was estimated to be 12% for
the 62 samples and may represent a sub-
stantial economic loss.
Accelerated Deterioration of Lupulin
Obtained from Male Hops
In a fourth experiment lupulin from
male hops was examined to determine the
applicability of A 2 75/A 3 2 5 to the evalua-
tion of relative storage stability of Q- and
l3-acids in male genotypes. Lupulin was
collected by gently agitating a water
slurry of male flowers with a low speed
blender, followed by sieving, washing, and
drying in a desiccator over CaS04' Single
samples (approximately 25 mg.) were
homogenized (10,000 r.p.m. in a 10 mI.
cup) with 5 mI. methanol, diluted appro-
priately with alkaline methanol, and hop
acids were measured spectrophoto-
metrically. A duplicate series (single sam-
ples of each) was held at 85°C. (±O.Olo)
for 16 hours and anlyzed as above.
The data for hop acids were converted
to "% lost" and compared with the cor-
responding values for A2 75/A 3 25 (Table
IV). The relationship between A 2 7 5
/ A 3 25 and oxidation of male hop acids
was very similar to that in the female
varieties shown in Table II, even though
conditions for the oxidation were very
different, and indicates that the same
oxidation products occurred in both
cases. If different products were pro-
duced it is very likely that their absorb-
ance would be different at one or both
wavelengths, in which case a slope differ-
ent from that in Fig. 2 would result.
72 HOPDETERIORATION
a.o r----.---,---,----.---,---,---,----y--..------,
completely
oxidized lupuli /'
2.0 /'
/'
/'

o
iII 1.0
0 .• o
~ 0.' o
- 0.7
} 0.'
:) 0.5

0.4

OQ o
o.a 00 0

o
0.2 o~---'-----'---L----'-------'----'-----'------J~--'---~
20 80 100

Figure 2. Relationship of deterioration of hop acids to increase of

A 2 7slA32s.Nine varieties stored 3, 6, and 10 months are represented.
The arrow at A27S/A32S .= 0.24 is the average initial value for all
varieties. Lupulin was held at elevated temperatures until no further
change took place in its absorbtion spectrum: A 27S/A 32s = 2.5 and
represents 100% loss.

TABLE IV
Loss of Hop Acids from Lupulin of 20 Male Genotypes and Corresponding
Increases in A 2 7 5/ A 3 2 5 After 16 Hours at 85° C (±0.01)
Initial After Storage Hop Acids
Identification a+(3 A2 7 S / A3 2 S a+(3 A2 7 S / A 3 2 S lost
1%) 1%) (%)

60028 M 81.1 0.21 43.4 0.67 46

6620-51 M 79.1 0.25 45.5 0.61 42
6616-32 M 72.5 0.26 36.7 0.74 49
6616-47 M 77.5 0.25 53.9 0.49 30
6775-05 M 75.7 0.24 41.9 0.64 45
54066 M 72.6 0.22 37.5 0.69 48
60026 M 73.4 0.25 30.7 0.82 58
6620-63 M 77.9 0.25 60.6 0.40 22
64030 M 81.2 0.21 49.8 0.48 39
6616-64 M 75.9 0.25 44.1 0.58 42
SY 325 M 82.5 0.25 48.7 0.57 41
63033 M 80.5 0.25 43.2 0.65 46
19054 M 71.3 0.22 55.1 0.38 23
SY 519 M 78.6 0.21 30.8 0.88 61
6801-26 M 79.6 0.24 41.2 0.69 48
6803-20 M 85.2 0.21 30.9 0.92 64
6802-37 M 81.7 0.22 32.6 0.89 60
SY 152 M 68.8 0.24 13.5 1.31 80
6803-60 M 78.1 0.22 46.2 0.60 41
6803-92 M 83.0 0.22 59.4 0.44 28
A.S.B.C. PROCEEDINGS 73

TABLE V
Statistical Data for Relationship of "% Hop Acids Lost" and
"Log A 2 7 5/A 3 2 5" for Hops (Female) and Lupulin (Male).
Correlationand Linear Regression
Female Male
Sample size 30 20
Mean of "log A275/A325 " -0.2400 -0.1925
Mean of "% lost" 37.37 45.65

Regression line (% lost =) 61.8 + 102 logA275 66.1 + 106 logA275

A325 A325
Standard error of constant 0.9601 0.9291
Standard error of slope 2.784 4.006
Correlationcoefficient 0.9897 0.9875
Analysis of Variance
Source of variation
Total degrees of freedom 29 19
mean square 670.7920 204.0289

Regression degrees of freedom 1 1

mean square 19053.6338 3779.8603

Error degrees of freedom 28 18

mean square 14.2619 5.3716

Regression Analyses amounts of a- and {3-acids present at the

Regression lines of "% lost" on "log
A 2 75/ A 3 25" for the two sets of data
(Fig. 2 and Table IV) are given in Table
V, using the standard form Y = A + BX.
The constants, A, were 61.8 for hops
(female) and 66.1 for lupulin (male), and
their standard errors were, respectively,
0.96 and 0.93, indicating a small, but
real, difference. The standard errors of
the slopes, 2.78 and 4.01 for the hops
and lupulin, indicate that the two slopes
are indistinguishable.
That the two curves are different only
by a small, constant amount across the
entire scale of oxidation probably results
from a contribution from the petals of
the hops which does not change during
the process of deterioration.
Discussion
Whitear (5) has offered convincing evi-
dence that hop acid values obtained at
harvest time are appreciably better guides
to hop usage than values found after
storage-deterioration of hop acids.
The most significant observation
which can be made from the present
study is that the spectrophotometric
method accounts, at once, for the
time of analysis and the amounts which
were present at harvest. For example:
A sample is analyzed and found to
have 6.0% a-acids, 4.0% /3-acids (a +
{3= 10.0%),andA275/A325 = 0.39.
Reference to Figure 2 indicates that
when A275/A325 = 0.39, 20% of
the original hop acids have been
lost and that the total hop acids
found, therefore, represent 80% of
those which were present at har-
vest: 10.070.8 = 12.5%.
There is little change in proportions
of a- and /3-acids until 30-40% have
been oxidized (Table II); therefore,
a-acids at harvest can be estimated
as (6.0 710.0) X 12.5 = 7.5%.
According to Whitear, 7.5% is the
better indication of the necessary hop-
ping rate to obtain a particular isohumu-
lone content in beer.
In addition to being useful for judging
the extent of deterioration which com-
mercially prepared hops have undergone,
A 2 75/A 32 5 can be used effectively in
screening large numbers of genotypes for
their relative storage stability (a varietal
characteristic). Although a great deal of'
latitude exists in selecting the conditions
74
for a comparative storage test, it is neces-
ary that all entries are exposed to the
same conditions for the same length of
time. It is helpful to include varieties with
known storage stabilities as controls and
as reference points. The advantages of
using single analyses (i.e. after deteriora-
tion only, rather than before and after)
for judging both hop acid content and
storage stability are obvious.
Summary
The ratio of absorbance of hop acids
(from extracts of hops or lupulin) in
alkaline methanol at 275 mil and 325 mil
(A27S/A32S) was shown to be loga-
rithmically related to the extent of oxida-
tion of the acids. It was demonstrated
that A2 75/A 32 5 can be used as a quanti-
tative estimate of the degree of deteriora-
tion of commercial hop samples at the
time of analysis. Furthermore, the hop
HOP DETERIORATION
acid content prior to any deterioration
can be estimated.
A27S/A32S was also shown to bear
the same relation to oxidation of hop
acids in lupulin from male hops as in
cones of the female plants. The relation-
ship was found to be reliable at elevated
'temperatures for short periods in an ac-
celerated storage test.
Literature Cited
1. Alderton, G., Bailey, G.F., Lewis, J.C., and
Stitt, F. Anal. Chern. 26: 983 (1954).
2. American Society of Brewing Chemists.
Methods of Analysis (6th ed.), (HOPS-4).
The Society: Glenview, IUinois (1958).
3. Ibid. (HOPS-5).
4. Burgess, A.H. Hops. Leonard Hill Books,
London, England (1964).
5. Whitear, A.L. J. lnst, Brewing 72: 177
(1966).
6. Wright, R.G., and Connery, F.E. Am Soc.
Brewing Chemists, Proc, 1951, p. 87.