Hop Flavor and Aroma: Proceedings of the 2nd International Brewers Symposium by Scientific Societies

CHAPTER 3

Cultivation Conditions of Saaz Hop for Promoting Desired Hop-Derived Taste and Aroma in Beer

Takako Inui

Suntory Beer Ltd, Beer Development Department, 5-2-5 Yamazaki, Shimamoto-cho, Mishima-gun, Osaka, 618-0 001 Japan Hiroo Matsui

Suntory Global Innovation Center Ltd, Research Institute, 8-1-1 Seikadai, Seika-cho, Soraku-gun, Kyoto, 619-0284 Japan

Introduction The taste, flavor, and aroma of hops originate from their bitter acids, such as humulones and their oxidized derivatives (Algazzali and Shellhammer, 2016; Haseleu et al., 2009), essential oil, and polyphenol constituents. Isohumulones, formed by isomerization of humulones during wort boiling (Intelmann et al., 2009) and linalool are well known as the key compounds related to the bitterness and floral aroma of kettle hopped beer (Hanke, 2009). Extensive studies have shown that different hop varieties have different aroma (Inui et al., 2013; Donaldson et al., 2012; Daniel et al., 2010; Steinhaus et al., 2007; Sakuma et al., 1991) and polyphenols compound profiles (Hasegawa and Inui, 2016). However, considerable differences in the amounts of humulones and linalool have been observed even within a single hop variety, resulting in aroma variations in the produced beer (Fig. 3.1). Therefore, in order to achieve a desirable, stable, and consistent hop aroma and flavor in beer, a fixed target compound profile at harvest is required for the hops. The variations in chemical profiles within one hop variety may be attributed to the following factors: 1.) season-dependent factors, such as temperature and rainfall; 2.) the history of the individual hop plant, such as root age, clone type, and viral infections; 3.) cultivation conditions, such as fertilization method, agrochemicals used, soil 25

26 Hop Flavor and Aroma: Proceedings of the 2nd International Brewers Symposium

Fig. 3.1. Humulone (%) and linalool (mg/kg) distribution in Czech Saaz hops grown in commercial

farms from 2010 and 2011. (From Inui, T., Matsui, H., Hosoya, T., Kumazawa, S., Fukui, N. and Oka, K. (2016). Effect of harvest time and pruning date on aroma characteristics of hop teas and related compounds of Saaz hops. J. Am. Soc. Brew. Chem. 74: 231–241. Reprinted by permission of the publisher, Taylor & Francis Ltd.)

type, pruning date, and harvest time; and 4.) the growing location (Matsui et al., 2013 and 2016; Jelinek et al., 2012; Bailey et al., 2009; Krofta et al., 2009; Murphey and Probasco, 1996). In general, hops harvested at different farms or maturity levels can be homogenized by pelletization to achieve a more consistent and target humulone content. However, the concentrations of other constituents, such as terpenoids and polyphenols, may not be able to be controlled by this process. To some extent, cultivation conditions can be manually controlled and could be important factors in controlling the taste and aroma of hops in beer. Consequently, we investigated the effect of root age, harvest time, and pruning date on the taste, aroma, and compound profiles of beer brewed with Saaz hops in some cultivation trials.

The Influence of the Root Age on the Quality of Hop Taste and Aroma in Beer Qualities of young hops are thought to be unstable and impart different taste and aroma characteristics when compared to older hops. Pluhackova et al. (2011) studied the relationship between hop root age, yield, and hop quality, but did not assess the impact of these factors on beer quality. Therefore the wide distribution of Saaz root age in 2011 (Fig. 3.2) was expected to have a significant impact on the hop taste and aroma in beer. To evaluate the impact of hop root age on hop taste, hop aroma, and hop chemical profiles, hop cones with varying root ages were collected from 8 hop gardens in the Saaz region during the 2010 and 2011 growing seasons (Table 3.1). The concentrations of humulones were higher in the younger hops (Fig. 3.3). The amount of essential oil and

Inui and Matsui 27

Fig. 3.2. Root age distribution of Saaz hops in Saaz region in 2011. This data was cited from UKZUZ that is one of the organizations of ministry of Agriculture in Czech Republic. (Courtesy T. H. Shellhammer and S. R. Lafontaine—© ASBC and MBAA)

Table 1 Test samples Table 3.1. Test samplesa,b

Area

Variety

Saaz in CZ

Saaz

Garden a b c d e f g h

Year of planting 2010 2009 2008 2007 2006 2001 1996 1991

2010 1 2 3 4 5 10 15 20

Age

2011 2 3 4 5 6 11 16 21

Reproduced, by permission, from Matsui, H., Inui, T., Ishimaru, M., Hida, Y., and Oka, K. (2013). The Influence of the age of hop plant on the quality of hop aromas in beer. Acta Hortic. 1010: 171–182. doi:10.17660/ ActaHortic.2013.1010.20

All hops are ‘virus free’ and consist of ‘Osvald’s clone 72’.

All hops are ‘virus free’ and consist of ‘Osvald’s clone 72’. Fig. 3.3. Content of humulone (%) in

hops harvested from 8 different gardens in 2 consecutive harvest years, 2010 and 2011. (Reproduced, by permission, from Matsui, H., Inui, T., Ishimaru, M., Hida, Y., and Oka, K. (2013). The Influence of the age of hop plant on the quality of hop aromas in beer. Acta Hortic. 1010: 171–182. doi:10.17660/ActaHortic.2013.1010.20)

the relative percentage of each terpene compound in the hop oil are also reported (Table 3.2). The relative concentration for each compound over the different hop root ages was compared to the average of about 30 different hop samples collected each harvest year (Fig. 3.4). In general, while root age did not seem to have a significant effect on hop total oil content, younger hops (under 5 years old) were found to contain higher levels of sesquiterpenes (known to contribute to a mild aroma to beer) and lower concentrations of monoterpenes (known to contribute to hoppy aromas to beer) with the exception of o-cimene. Brewing trials over the two harvest years were carried out using 4 randomly selected samples (‘b,’ ‘d,’ ‘g,’ and ‘h’ in Table 3.1). Significant differences were

28 Hop Flavor and Aroma: Proceedings of the 2nd International Brewers Symposium

Table 2 Amounts of essential oils and the relative % of each terpene in hops from 8 different gardens

Table 3.2. Amounts of essential oils and the relative % of each terpene in hops from 8 different gardensa a Oil amount Linalool Geraniol Myrcene Ocimene β-Pinene D-Limonene β-Farnesene Bergamotene α-Humulene β-Caryophyllene

2010

2011

ml/100g

0.64

0.49

0.62

0.52

0.54

0.75

0.53

0.54

1.04

0.44

0.61

0.64

0.58

0.71

0.57

0.58

rel. % rel. % rel. % rel. % rel. % rel. % rel. % rel. % rel. % rel. %

0.33 0.13 23.90 0.08 0.47 0.11 22.40 0.94 18.20 8.10

0.28 0.09 15.60 0.07 0.34 0.05 28.60 1.00 20.90 9.80

0.42 0.07 33.30 0.11 0.48 0.12 24.10 0.77 16.90 6.80

0.20 0.03 15.40 0.10 0.28 0.07 34.80 1.02 17.60 9.50

0.28 0.04 21.40 0.10 0.36 0.10 23.30 1.02 22.20 9.20

0.70 0.28 37.70 0.09 0.55 0.19 24.50 0.87 15.70 6.00

0.61 0.28 31.70 0.11 0.50 0.13 25.90 0.93 16.50 6.70

0.57 0.25 34.00 0.13 0.55 0.14 22.10 0.78 17.00 7.50

0.32 0.05 28.30 0.12 0.39 0.11 20.00 0.80 27.30 7.86

0.36 0.06 25.50 0.13 0.44 0.11 18.90 0.88 30.10 8.63

0.38 0.04 27.30 0.09 0.33 0.10 11.60 0.83 37.80 6.62

0.45 0.11 28.30 0.09 0.46 0.13 21.50 0.94 25.40 7.38

0.54 0.12 44.70 0.16 0.66 0.18 16.70 0.71 18.20 5.25

0.54 0.22 40.55 0.11 0.52 0.15 18.45 0.77 18.15 4.93

0.51 0.16 37.00 0.15 0.54 0.16 17.60 0.75 22.30 6.42

0.54 0.17 42.50 0.15 0.63 0.16 16.50 0.75 19.40 5.47

Fig. 3.4. Comparison of terpene profiles in hop samples. The 6 compounds indicated on the right-hand side of the chart are monoterpenes (linalool, geraniol, myrcene, ocimene, β-pinene and d-limonene). The 4 compounds indicated on the left-hand side of the chart are sesquiterpenes (β-farnesene, ber gamotene, α-humulene and β-caryophyllene). The charts show the relative content of each compound in each age of hop, compared to the average of about 30 different hop samples collected each harvest year. (Reproduced, by permission, from Matsui, H., Inui, T., Ishimaru, M., Hida, Y., and Oka, K. (2013). The Influence of the age of hop plant on the quality of hop aromas in beer. Acta Hortic. 1010: 171–182. doi:10.17660/ActaHortic.2013.1010.20)

observed in the hop taste and aroma as well as the concentrations of aroma compounds in these beers and hops (Figs. 3.5-3.8; Table 3.3). Similar to the hops, beers made with the younger hops contained lower concentrations of monoterpenes, higher concentrations of sesquiterpenes, and generally contributed lower aromas (Figs. 3.5 and 3.8). Though the beers made with the older hops had higher concentrations of monoterpenes and generally

Inui and Matsui 29

Fig. 3.5. Sensory results for hop aroma characteristics in beer. The charts show the average normalized

sensory score from 7 panelists. (Reproduced, by permission, from Matsui, H., Inui, T., Ishimaru, M., Hida, Y., and Oka, K. (2013). The Influence of the age of hop plant on the quality of hop aromas in beer. Acta Hortic. 1010: 171–182. doi:10.17660/ActaHortic.2013.1010.20)

Fig. 3.6. Sensory results for beer taste. The charts show the average normalized sensory score from 7 panelists. (Reproduced, by permission, from Matsui, H., Inui, T., Ishimaru, M., Hida, Y., and Oka, K. (2013). The Influence of the age of hop plant on the quality of hop aromas in beer. Acta Hortic. 1010: 171–182. doi:10.17660/ActaHortic.2013.1010.20)

Fig. 3.7. Terpene profiles in hop samples. The charts show the relative content of each terpene, compared to the maximum content found in each of the 4 different root ages. (Reproduced, by permission, from Matsui, H., Inui, T., Ishimaru, M., Hida, Y., and Oka, K. (2013). The Influence of the age of hop plant on the quality of hop aromas in beer. Acta Hortic. 1010: 171–182. doi:10.17660/ ActaHortic.2013.1010.20)

imparted more intense aroma to beer characterized by citrusy, floral, and fruity qualities. This supports the connection between aroma compounds measured in hops and the beer produced from them. The length of the vine, leaf size, stem diameter, and flowering timing were also monitored throughout the growing season to assess the impact of hop root age on the vegetative and reproductive growth of hops (Figs. 3.9 and 3.10). In general, for the Saaz variety,

30 Hop Flavor and Aroma: Proceedings of the 2nd International Brewers Symposium

Fig. 3.8. Terpene profiles in beer samples. The charts show the relative content of each terpene

compared to the maximum content found in each of the 4 different root ages. (Reproduced, by permission, from Matsui, H., Inui, T., Ishimaru, M., Hida, Y., and Oka, K. (2013). The Influence of the age of hop plant on the quality of hop aromas in beer. Acta Hortic. 1010: 171–182. doi:10.17660/ Table 3. Concentration of aroma compounds in beer (ppb) ActaHortic.2013.1010.20)

Table 3.3. Concentration of aroma compounds in beer (ppb)a Linalool Geraniol Geranyl acetate Myrcene D-Limonene β-Citronellol α-Terpineol Nerol Ethyl 2-methyl butanoate Isovaleric acid ethyl ester β- Caryophyllene Caryophyllene oxide β-Farnesene α-Humulene β-Ionone Nerolidol

b 8.27 3.96 0.40 4.42 1.50 6.47 2.63 1.77 0.24 0.45 0.49 1.68 13.46 2.38 0.26 0.69

2010 d g 7.40 24.80 3.59 9.91 0.36 1.24 3.36 7.03 1.32 2.37 6.54 15.27 2.58 4.29 1.61 2.49 0.27 0.28 0.44 0.42 0.52 0.60 1.48 1.34 11.21 12.10 5.19 2.14 0.24 0.13 0.65 0.43

h 18.66 6.04 0.65 5.91 2.03 10.70 4.23 2.43 0.24 0.35 0.83 2.39 12.90 2.54 0.19 0.75

b 9.62 3.15 0.18 5.50 1.91 6.51 2.96 1.27 0.36 0.58 0.29 0.13 8.15 1.28 0.15 0.27

2011 d 27.20 5.34 0.25 7.35 2.00 11.11 4.80 1.99 0.32 0.54 0.17 0.16 6.27 1.03 0.11 0.36

g 31.08 9.18 0.63 9.33 2.75 21.66 4.92 2.70 0.39 1.25 0.23 0.09 7.21 1.36 0.09 0.22

h 30.96 8.58 0.58 10.35 3.66 18.21 5.74 2.45 0.42 0.56 0.19 0.13 6.26 1.27 0.13 0.37

Fig. 3.9. Vegetative growth of hops: (x) height, (y) stem

diameter at 1.5 m, and (z) leaf size at 1.5 m above ground, just before harvest. (Reproduced, by permission, from Matsui, H., Inui, T., Ishimaru, M., Hida, Y., and Oka, K. (2013). The Influence of the age of hop plant on the quality of hop aromas in beer. Acta Hortic. 1010: 171–182. doi:10.17660/ActaHortic.2013.1010.20)

Fig. 3.10. Reproductive growth— days from pruning to cone formation. (Reproduced, by permission, from Matsui, H., Inui, T., Ishimaru, M., Hida, Y., and Oka, K. (2013). The Influence of the age of hop plant on the quality of hop aromas in beer. Acta Hortic. 1010: 171–182. doi:10.17660/ ActaHortic.2013.1010.20)

Inui and Matsui 31

pruning is performed in April to guide the twining of the new sprout above ground, and it lasts about 20 days. Harvest of hop cones occurs from the middle of August to the beginning of September, within about 25 days. In this study, when observing the number of days from pruning to cone formation, it was found that younger hops take longer to flower and form cones Based on these results, it was concluded that late flowering associated with the younger roots changed secondary metabolism and affected the generation of terpenes which altered the resulting aroma intensity and quality these hops attributed to beer.

The Influence of the Harvest Time and Pruning Date on the Quality of Hop Taste and Aroma in Beer The harvest window for Saaz hops, grown in Saaz, Czech Republic, can span around 25 days from mid-August to early September. This harvest window is one the longest amongst all hop varieties and has the potential to lead to large differences in hop quality. In addition, the relationship between the pruning date and the flowering time surveyed at more than 20 farms in the Saaz area (Fig. 3.11) and measured in a model cultivation test by Matsui et al. (2016), found that later pruning date can also impact hop quality because it results in a shorter period between pruning and flowering and duration of vegetative growth. Therefore, the impact of both harvest timing and pruning date on the aroma characteristics and chemical profiles of Saaz hops was evaluated (Inui et al., 2016). In this study, two cultivation trials (Table 3.4 (A) and (B)) were carried out. In one cultivation trial, Saaz hops were harvested from one growing area over four years (from 2011 to 2014) with harvest windows spanning up to 40 days, which is around twice the usual harvest window. In the other cultivation trial, Saaz hops were harvested from four different growing areas in 2012.

Fig. 3.11. Relationship between pruning and flowering dates for Czech Saaz hops grown in 22 commercial farms in 2012. It can be seen that the pruning date does not affect the flowering date, as the slope of the line corresponding to their correlation is very small. There was no correlation between the pruning and flowering dates. However, shorter pruning intervals corresponded to shorter blooming intervals. (From Inui, T., Matsui, H., Hosoya, T., Kumazawa, S., Fukui, N. and Oka, K. (2016). Effect of harvest time and pruning date on aroma characteristics of hop teas and related compounds of Saaz hops. J. Am. Soc. Brew. Chem. 74: 231–241. Reprinted by permission of the publisher, Taylor & Francis Ltd.)

32 Hop Flavor and Aroma: Proceedings of the 2nd International Brewers Symposium

Table 3.4. Design of cultivation (A) trial I and (B) trial IIa Table 3.4. Design of cultivation (A) trial I and (B) trial IIa (A) Cultivation trial

Harvest year Farm Pruning date Harvest time (days) (B) Cultivation trial Harvest year Farm Pruning date Harvest time (days) Harvest year Farm Pruning date Harvest time (days)

2011 Rybnany April 15 29 50

April 4 49 69

April 6 39 58

Lipenec 78

2012 Rybnany April 16 32 51

2013 Steknik April 20 37 53

2012

April 14 38 58

Rybnany April 16 32 51

April 4 38 58

2014 Steknik April 15 40 57

April 24 25 44

Blsany April 14 45 63

Krouvcova 67

2012 60

April 25 25 44

From Inui, T., Matsui, H., Hosoya, T., Kumazawa, S., Fukui, N. and Oka, K. (2016). Effect of harvest time and pruning date on aroma characteristics of hop teas and related compounds of Saaz hops. J. Am. Soc. Brew. Chem. 74: a Courtesy T. H. Shellhammer and S. R. Lafontaine—© ASBC and MBAA. 231–241. doi:10.1094/ASBCJ-2016-4628-01. Reprinted by permission of the publisher, Taylor & Francis Ltd.

While brewing trials are the most preferred technique to characterize how hop aroma, hop flavor, and hop compound profiles imparted to beer vary across different hop treatments, it is a time-consuming process to evaluate a large number of hop samples. Generally, the differences in hop aroma characteristics are evaluated based on the essential oils which are extracted from the hop via steam distillation. However, the aroma characteristics and components imparted to beer from hops do not necessarily match these essential oils profiles. Therefore, for this study, the hop aroma and compound profiles of the different hop treatments were evaluated using hop teas and hot hop solutions to imitate late hopping wort. In brief, pulverized hops were added to a heated citric acid buffer solution and left to stand for 5 min. Subsequently, the hop solution was cooled and diluted with water. A panel of 7 well-trained individuals was selected to perform quantitative descriptive analysis (QDA) of various hop aroma characteristics. Four sensory descriptors: floral, fruity, citrusy, and hay-like were used because the differences in their intensities between the samples were clearly distinguishable by the panelists. The intensities of the sensory descriptors were assigned scores, ranging from 0 to 3; and the scores were normalized to remove potential bias from each panelist’s score. In order to evaluate the compounds that contribute to the hop aroma in beer, hot water hop extractions were performed. In brief, pulverized hop was added to a citric buffer at the same concentration as that of beer, and left to stand at 100 degree Celsius for 5 min. This extracted solution was then loaded into a solid phase extraction column and eluted using dichloromethane. The collected solvent was subsequently concentrated via vacuum evaporation and injected into a GC×GC-TOF/MS system. The concentrations of humulones and the total oil content of hops significantly increased with later harvest timing over each harvest year in Trial I (Fig. 3.12 (A)). Yet, in Trial II, though the total oil content of the hops significantly increased with later harvest time, in general, the concentrations of humulones were not significantly impacted by harvest timing across the different farms (Fig. 3.12 (B)). Analysis of variance (ANOVA) results support these observations because significant effects were observed for both harvest time and harvest year with the concentrations of humulones and total oil content for Trial I (Table 3.5 (A)). However, in Trial II, farm was a more significant factor in influencing the concentrations of humulones whereas the oil contents were heavily determined by the harvest time (Table 3.6 (B)).

Inui and Matsui 33

Fig. 3.12. Humulone and oil content in hop samples at different harvest times by (A) harvest year or (B) farms from cultivation trial I and II. (From Inui, T., Matsui, H., Hosoya, T., Kumazawa, S., Fukui, N. and Oka, K. (2016). Effect of harvest time and pruning date on aroma characteristics of hop teas and related compounds of Saaz hops. J. Am. Soc. Brew. Chem. 74: 231–241. Reprinted by permission of the publisher, Taylor & Francis Ltd.)

Table 3.5.

Table 3.5. F-values and significance from two-way ANOVA of humulone and oil content in (A) trial I and (B) trail IIa (A)

Compounds Humulone (%) Oil contents (%)

Harvest time

Harvest year

30.45d 23.69c

12.12c 5.56b

(B) Compounds Humulone (%) Oil contents (%)

Harvest time

Farm

1.59

d 8.78

27.24d

3.89

Harvest time X Harvest year 1.45 1.29

Pruning date

Harvest time X Farm

Harvest time X Pruning date

0.07

0.35

3.21

0.07

0.95

0.21

From Inui, T., Matsui, H., Hosoya, T., Kumazawa, S., Fukui, N. and Oka, K. (2016). Effect of harvest time and pruning date on aroma characteristics of hop teas and related compounds of Saaz hops. J. Am. Soc. Brew. Chem. 74: 231–241. doi:10.1094/ASBCJ-2016-4628-01. Reprinted by permission of the publisher, Taylor & Francis Ltd.

Significance of F-values at the 95.0% level.

Table 3.6. of F-values at the 99.0% level. Significance

Significance of F-values at the 99.9% level.

(A)

Harvest time X year according to harvest time in Trials I and II ofHarvest hop teas Harvest year found floral, citrusy, tended to increase with the Floral three aroma characteristics: c 1.14 fruity, and 1.04 13.32 harvest time over the different harvest years and different farms (Fig. 3.13). However, Fruity 0.93 15.85 4.14bwas not dependent the score for the hay- likec characteristic on the harvest time. Again, Citrussy b 1.63 ANOVA results support significant effects were determined 9.87 these observations because 0.86 for harvest time and the and citrusy characteristics in both trials (Table 3.6 Hey-like 3.48floral, fruity,0.22 0.78 (A) (B) and (B)). No two-way interactions with harvest time and pruning date or farm were observed for any of the evaluated attributes. Harvest time X Harvest time X Attribute Harvest time Farm Pruning date Attribute Harvest time The organoleptic evaluation

Floral Fruity Citrussy

34.43d 34.08d 30.19d

0.55 1.52 0.53

12.02c 22.14d 14.38c

Farm 2.12

Pruning date 0.13

1.98

0.36

1.44

0.46

(%)

34 Hop Flavor and Aroma: Proceedings of the 2nd International Brewers Symposium

Table 3.6.

Table 3.6. F-values and significance from ANOVA of the organoleptic evaluation of hop teas in (A) trial I and (B) trail IIa (A)

Attribute Floral Fruity Citrussy Hey-like (B) Attribute Floral Fruity Citrussy Hay-like

Harvest time X Harvest year 1.04

Harvest time

Harvest year

13.32c c 15.85 b 9.87

1.14 b 4.14 1.63

0.93

3.48

0.22

0.78

Harvest time

Farm

Pruning date

34.43d 34.08d

0.55

12.02c 22.14d

d 30.19 0.32

1.52 0.53 0.53

0.86

c 14.38 3.57

Harvest time X Farm 2.12

Harvest time X Pruning date 0.13

1.98

0.36

1.44

0.46

1.76

1.72

From Inui, T., Matsui, H., Hosoya, T., Kumazawa, S., Fukui, N. and Oka, K. (2016). Effect of harvest time and pruning date on aroma characteristics of hop teas and related compounds of Saaz hops. J. Am. Soc. Brew. Chem. 74: 231–241. doi:10.1094/ASBCJ-2016-4628-01. Reprinted by permission of the publisher, Taylor & Francis Ltd.

Significance of F-values at the 95.0% level.

Significance of F-values at the 99.0% level.

Significance of F-values at the 99.9% level.

The impact of pruning date on the organoleptic evaluation of the hop teas in Trial I and II was also investigated (Fig. 3.13(C)). In this graph, the harvest times are classified as early, middle, late, and very late, based on the duration of the harvest time. Floral, fruity, and citrusy characteristics were more commonly observed for samples with later pruning dates. However, the score for the hay-like characteristic showed no dependence on the pruning date. Results from ANOVA support these observations and significant effects were observed between pruning date and the floral, fruity and citrusy aromas (Table 3.6). Based on spectral identification by GC×GC-TOF/MS, 63 compounds were identified in the hot water hop extractions. However, there was considerable variation in analytical value (percent coefficient of variation (%CV) >20) for some compounds, particularly for myrcene, limonene, and humulene. These large variations are thought to be caused by poor compound stability during the hot water extract and poor detection sensitivity. Therefore, the compounds with %CV >20 were removed as targets for statistical analysis and only 33 compounds were quantitated. Concentrations for each compound, obtained from calibration curves for the 33 compounds are reported (Table 3.7). Based on the ANOVA results from trial I, harvest time and harvest year obviously had a significant impact on some of these components, notably for linalool and linalool oxides (Table 3.8 (A)). Significant two-way interaction effects were also observed between harvest time and harvest year. The ANOVA results derived from Trial II (Table 3.8 (B)) showed that harvest time, farm source, and pruning date also had a significant impact on most of the chemical compounds. Again, some two-way interactions were observed between harvest time and both farm and pruning date for some of these compounds. Additionally, to construct a model to predict the response variables for the floral, fruity, and citrusy characteristics, a partial least squares (PLS) regression was performed on the data generated by QDA on the hop teas and the concentrations of aroma com-