Beer, together with Huangjiu and grape wine, is known as the world’s three major alcoholic beverages, and its life cycle has extended for thousands of years. With the development of the beer industry, due to differences in brewing process and production strains, modern beer is mainly divided into Ale beer (top-fermented beer) and Lager beer (bottom-fermented beer) [2]. Compared with Ale beer, Lager beer has a lower fermentation temperature and a longer cycle; it not only has delicate and rich foam and a refreshing and pure taste, but also has uniform quality and is suitable for industrial production at scale [3]. Therefore, Lager beer is deeply favored by consumers for its light and refreshing flavor and stable quality, and it has gradually replaced Ale beer as the mainstream product of modern industrial beer. Common volatile flavor compounds in beer include alcohols, esters, aldehydes, and ketones, etc. The quality and content of these compounds not only determine the taste and flavor of beer, but also affect the quality of beer to a certain extent [5]. In recent years, as consumers’ requirements for beer quality and flavor have become increasingly high, how to maintain and improve the flavor of beer has become the focus commonly concerned by many production enterprises [6]

Malt is the main raw material for brewing beer. As the “backbone” of beer, malt quality defects will directly lead to abnormalities in saccharification and filtration, reduce production efficiency, and increase product cost [7]. Therefore, the composition and quality of malt are related to beer flavor and quality. At the same time, in recent years, with the steady increase of beer output in China, the quality and yield of domestic malts have gradually declined, which has aggravated the demand of China’s beer industry for high-quality raw materials [8]. As for research on the impact of malt on beer quality, it is currently mainly focused on analysis of components such as β-glucan and protein [9], or on optimization of key brewing parameters such as fermentation temperature and mashing process [10], while studies on the selection and control of brewing raw materials at the source are relatively few.

In order to control beer production from the source, through a laboratory beer fermentation simulation system, this study explored the quality of three different types of malts and their effects on beer quality and brewing flavor. By measuring malt quality indices such as extract rate, α-amino acid nitrogen, saccharifying power, and Kolbach index, as well as physicochemical indices of the beer brewed such as alcohol content and original wort concentration, and by using headspace solid-phase microextraction (HS-SPME) combined with gas chromatography-mass spectrometry (GC-MS) to analyze differences in aroma components, the most suitable malt variety for beer fermentation was ultimately selected, in order to brew high-quality Lager beer and provide theoretical reference for its industrialized production.

1 Materials and Methods

1.1 Materials and Reagents

1.1.1 Strain

Brewing yeast (Saccharomyces cerevisiae) Lager 497: laboratory stock.

1.1.2 Chemical reagents

Yeast extract, peptone, agar powder (all biochemical reagents): Bioengineering (Shanghai) Co., Ltd.; anhydrous glucose, iodine, potassium iodide (all analytical grade): Sinopharm Group Chemical Reagent Co., Ltd.; 3-octanol (chromatographic grade): Sigma-Aldrich, USA; British malt (Eng), Canadian malt (Can), German malt (Ger), and SAAZ hops: imported commercial purchase.

1.1.3 Nutrient medium

Yeast extract peptone dextrose (yeast extract peptone dextrose, YPD) medium: yeast extract 10 g/L, peptone 20 g/L, anhydrous glucose 20 g/L, sterilize at 115 ℃ for 20 min.
Wort medium: self-made wort, sterilize at 105 ℃ for 10 min.

1.2 Instruments and equipment

DLEUW22050 malt mill: Bühler-Miag, Switzerland; BGT-8A saccharimeter: Hangzhou BoRi Technology Co., Ltd.; Trace1310-ISQ LT gas chromatography-mass spectrometry: TG-WAXMS A capillary column (60 m × 0.25 mm × 0.25 μm), TriPlus RSH automatic headspace sampler, headspace solid-phase microextraction fiber (50/30 μm DVB/Carboxen/PDMS): Thermo Scientific, USA.

1.3 Test methods

1.3.1 Establishment of the laboratory Lager beer fermentation simulation system

Weigh an appropriate amount of malt, add water for mashing, and after saccharification complete, filter and collect the clear wort. Boil for 60 min, during which an appropriate amount of hops is added. Filter again, adjust the wort concentration, obtain clear wort. Sterilize at 105 ℃ for 10 min, cool and dispense.

Yeast activation culture: inoculate a loopful of single colony into 1 mL wort medium, culture at 25 ℃, 220 r/min for 12 h; transfer the above culture into 9 mL wort medium, culture at 20 ℃, 220 r/min for 12 h; then transfer the above 10 mL culture into 90 mL wort medium, culture at 15 ℃, 220 r/min for 12 h. Let stand, take the yeast cell suspension; inoculate into wort medium at 1×10⁶ CFU/mL, then cap with a rubber stopper and seal aseptically [13].

Laboratory simulated fermentation: culture statically at 12 ℃ for 8 d. When the fermentation sugar degree is about 4 °Bx, primary fermentation ends. Place statically at 4 ℃ for 7 d; when there is no obvious diacetyl flavor in the wine, fermentation is completed [14].

1.3.2 Malt quality index testing method

Take an appropriate amount of malt, and determine quality indices such as extract rate, α-amino acid nitrogen, saccharifying power and Kolbach index (also called KI value) according to the analytical method in light industry standard QB/T 1686—2008 “Beer malt”.

1.3.3 Beer physicochemical index testing method

After fermentation, take samples for testing, and determine physicochemical indices of beer such as alcohol content and original wort concentration according to national standard GB/T 4928—2008 “Beer analysis methods”.

1.3.4 GC-MS detection method for beer flavor substances

(1) Chromatographic conditions:TG-WAXMS A capillary column (60 m × 0.25 mm × 0.25 μm), temperature program: hold at 40 ℃ for 1 min, raise temperature at 3 ℃/min to 180 ℃, then raise at 20 ℃/min to 230 ℃, hold for 15 min. Vaporization chamber temperature 250 ℃, carrier gas is high-purity helium (purity >99.99%), carrier gas flow 1.0 mL/min, no split injection, solvent delay time 1 min.Mass spectrometry conditions: EI ionization mode, ion source temperature 230 ℃, quadrupole temperature 150 ℃, electron energy 70 eV, emission current 34.6 μA, interface temperature 280 ℃, mass scan range 29～500 amu.
(2) Sample preparation: take 7.5 mL beer sample into a 20 mL headspace vial, add 0.5 mL 3-octanol internal standard solution (0.3 mg/L), seal tightly, crimp with aluminum cap. Keep in 60 ℃ water bath, headspace extract for 40 min; retract the fiber, insert into GC injector for desorption for 5 min [15].
(3) Qualitative and quantitative analysis
Through GC-MS analysis to detect the total ion chromatogram of flavor substances in the beer sample, use the National Center for Biotechnology Information (National Center of Biotech-nology Information, NIST) 2014 standard mass spectral library search, combined with retention index (retention index, RI) to qualify volatile flavor components in the fermentation liquid, and use peak area normalization method for quantitative analysis [16].

2 Results and analysis

2.1 Main quality indices of different types of malts

2.1.1 Kolbach index of different types of malts

The Kolbach index of beer malt mainly reflects the dissolution degree of malt protein during the malting process, and it is appropriate to be maintained at 40%～45% [17-18]. Among the three different types of malts, the Kolbach indexes of Can and Ger malts were moderate, 45% and 40% respectively; while the Kolbach index of Eng malt was relatively high (50%), and there was a significant difference among the three (P<0.05).

(figure note) Different letters indicate significant differences between different malts (P<0.05). Same below.
Fig. 1 Kolbach indexes of different types of malts

2.1.2 Saccharifying power of different types of malts

Saccharifying power is also called starch saccharifying enzyme activity; the higher the starch saccharifying enzyme activity, the higher the saccharifying power value, and the better the malt quality [19]. Malt saccharifying power is expressed in Windisch-Kolbach (WK) units: under conditions of 20 ℃ and pH 4.3, 100 g anhydrous malt decomposes soluble starch for 30 min to produce 1 g maltose, which is 1 WK. According to the analysis method in QB/T 1686—2008 “Beer malt”, the saccharifying powers of the three malt samples were 357 WK, 414 WK and 392 WK respectively, all significantly higher than the premium standard (>260 WK), and there was no significant difference among the three (P>0.05).

Fig. 2 Saccharifying power of different types of malts

2.1.3 Extract rate and α-amino acid nitrogen of different types of malts

Malt extract rate to a certain extent reflects the loss of substances during the germination process of barley. According to the analysis method of QB/T 1686—2008 “Beer malt”, the measured results of extract rate of different types of malts are shown in Fig. 3. It can be seen from Fig. 3 that the extract rate of Ger malt (in dry basis) is ≥79%, belonging to premium pale malt; the extract rate of Can malt (in dry basis) is ≥77%, belonging to first grade; the extract rate of Eng malt (in dry basis) is ≥75%, belonging to second grade.

Fig. 3 Extract rate and α-amino acid nitrogen contents of different types of malts

α-amino acid nitrogen reflects the degradation degree of malt protein and the activity of proteolytic enzymes, and determines the amount of free amino nitrogen in wort during the malting process. Generally, industrial production requires that the α-amino acid nitrogen content in wort during fermentation be ≥160 mg/L, and it is appropriate to control it at about 180～220 mg/L. It can be seen from Fig. 3 that among the different malt samples, except that the α-amino acid nitrogen content of Ger malt is relatively moderate (186 mg/L), the α-amino acid nitrogen contents of Eng and Can malts are relatively low, 125 mg/L and 118 mg/L respectively.

2.2 Effect of malt types on physicochemical indices of beer fermentation liquid

2.2.1 Effect of malt types on alcohol content and original wort concentration of beer fermentation liquid

Compare the alcohol content and original wort concentration of beer brewed by three different types of malts, and the results are shown in Fig. 4.

Fig. 4 Effect of different types of malts on alcohol content and original wort concentration of Lager beer

During beer brewing, yeast absorbs and metabolizes various fermentable sugars in wort, and finally produces ethanol and CO₂ [20]. It can be seen from Fig. 4 that among the three malt fermentation products, the beer brewed with Can malt has the highest alcohol content, 2.97%vol; the beer brewed with Eng malt is the second, with an alcohol content of 2.91%vol; the alcohol content of beer brewed with Ger malt is significantly lower than that of Eng and Can beers (2.19%vol) (P<0.05).

Original wort concentration refers to the percentage content of extract in wort. The higher the alcohol content of beer, the higher the original wort concentration [21]. It can be seen from Fig. 4 that the original wort concentration of Ger beer is low (7.56 °P), with stable quality, good storability; the original wort concentrations of Eng and Can beers are moderate (8.55 °P and 8.63 °P respectively), with full body and better quality.

2.2.2 Effect of malt types on actual concentration and fermentation degree of beer fermentation liquid

Beer concentration refers to the mass percentage of extract (sugars, amino acids and other soluble organic and inorganic substances) in beer or fermentation liquid, including actual concentration and apparent concentration [22]. Apparent concentration refers to the concentration obtained by directly measuring the concentration in the fermentation liquid; actual concentration refers to the concentration obtained by distilling out alcohol from the fermentation liquid and then measuring the concentration.

Fermentation degree refers to the percentage of wort extract consumed by yeast, and as an important index to judge whether fermentation is normal, it can be divided into apparent fermentation degree and real fermentation degree. Specifically, apparent fermentation degree is calculated by apparent concentration, while real fermentation degree is calculated by actual concentration (also called real fermentation degree) [23]. Generally, apparent fermentation degree is higher than real fermentation degree. Compare the concentrations and fermentation degrees of beer brewed by three different types of malts, and the results are shown in Fig. 5.

Fig. 5 Effect of different types of malts on the concentration (A) and fermentation degree (B) of Lager beer

It can be seen from Fig. 5A that among the beers brewed by three kinds of malts, the actual concentration of Ger beer (3.22%) is the highest, which is significantly different from Eng and Can (P<0.05), and there is no significant difference between Eng and Can (P>0.05). Among the beers brewed by three kinds of malts, the apparent concentration of Ger beer (1.64%) is the highest, which has no significant difference with Eng beer (P>0.05), while Can beer has the lowest apparent concentration, and there is a significant difference from the other two beers (P<0.05). Compared with Eng and Ger beers, Can beer has low extract content and a refreshing taste without sweetness.

It can be seen from Fig. 5B that among the beers brewed by three kinds of malts, the real fermentation degrees of Eng and Can beers are relatively high, 67.21% and 67.97% respectively, and there is no significant difference between them (P>0.05); the real fermentation degree of Ger beer is the lowest (57.44%), which is consistent with its high actual concentration (P<0.05). Among the beers brewed by three kinds of malts, the apparent fermentation degree of Can beer is the highest (86.41%), followed by Eng beer (81.21%), and Ger beer is the lowest (78.30%), and there is a significant difference among them (P<0.05). The results show that compared with Eng and Ger malts, yeast has a higher utilization rate of extract in wort prepared by Can malt, which is conducive to yeast growth and ethanol production, and is consistent with the results of alcohol content analysis.

2.3 Effect of malt types on beer flavor substances

Aroma components are key factors for judging beer quality. In this experiment, GC-MS analysis was carried out on the beer samples fermented by three different types of malts, and the results of aroma substances are shown in Table 1.

Table 1 Aroma components content of beer fermented by different types of malts

Sample	Eng beer relative content/%	Eng beer standard deviation	Can beer relative content/%	Can beer standard deviation	Ger beerrelative content/%	Ger beer standard deviation
Phenethyl alcohol	35.16	8.98	46.37	5.92	41.07	15.64
Isoamyl alcohol	21.90	6.08	39.01	0.92	27.01	9.18
Ethanol	0.20	0.25	8.41	4.22	3.62	0.79
Isobutanol	1.44	0.49	2.11	0.15	2.03	0.81
Octanol	0.70	0.13	1.24	0.04	0.52	0.12
1-Decanol	0.57	0.22	1.09	0.29	0.17	0.07
Linalool	0.51	0.09	0.88	0.04	0.52	0.21
trans-Nerolidol	0.49	0.14	0.93	0.07	0.79	0.33
Geraniol	0.24	0.04	–	–	–	–
n-Propanol	0.28	0.08	0.47	0.02	0.41	0.16
(R)-3,7-Dimethyl-6-octen-1-ol	0.17	0.05	0.31	0.01	0.40	0.16
n-Amyl alcohol	0.17	0.16	0.39	0.06	0.25	0.06
1-Hexadecanol	0.10	0.01	0.27	0.04	0.27	0.10
3-(Methylthio)-1-propanol	0.16	0.07	0.31	0.05	0.13	0.06
9-Hexadecen-1-ol	0.10	0.06	0.33	0.03	0.05	0.02
1-Butanol	0.16	0.02	0.27	0.02	0.11	0.04
Furfuryl alcohol	0.08	0.01	0.16	0.04	0.22	0.13
2,3-Butanediol	–	–	0.14	0.03	–	–
Linalool	0.07	0.02	0.08	0.01	0.13	0.04
1-Dodecanol	0.04	0.01	0.08	0.02	0.07	0.03
n-Butanol	0.02	0.01	0.02	0.00	0.05	0.01
2-Hexadecanol	0.02	0.01	–	–	0.02	0.01
Nerol	0.01	0.00	0.02	0.00	0.03	0.01
Benzyl alcohol	–	–	0.02	0.00	0.01	0.01
Total alcohols	62.59	16.93	102.91	11.98	77.88	27.99
Ethyl octanoate	20.64	1.49	34.52	8.22	13.01	6.98
Ethyl hexanoate	7.03	1.91	13.78	2.01	4.29	0.99
Ethyl acetate	3.65	0.93	7.16	0.46	4.34	1.35
Isoamyl acetate	2.59	0.55	7.96	1.32	2.13	0.61
Ethyl benzoate	2.69	0.28	6.09	1.70	3.35	1.50
Ethyl phenylacetate	1.80	0.08	4.41	1.25	5.71	3.63
Ethyl caprate (Ethyl decanoate)	1.14	0.26	3.62	1.20	1.53	0.66
Ethyl 9-hexadecenoate	1.57	0.76	2.15	0.85	1.12	0.86
Ethyl butyrate	0.42	0.11	0.77	0.06	0.29	0.06
Ethyl lactate	–	–	0.66	0.08	0.42	0.22
Ethyl butyrate (oil-like ester)	0.16	0.06	0.58	0.02	0.44	0.23
Ethyl octadecanoate	0.10	0.03	0.26	0.00	0.18	0.06
2,2,4-Trimethyl-1,3-pentanediol diisobutyrate	0.14	0.06	0.28	0.03	0.08	0.01
Ethyl tetradecanoate	0.06	0.01	0.20	0.02	0.09	0.05
Cyclohexyl sulfamic acid ester	0.06	0.04	0.26	0.08	0.10	0.02
Ethyl nonanoate	0.04	0.01	0.17	0.05	0.09	0.04
Ethyl 2-hexenoate	0.06	0.01	0.12	0.01	0.10	0.05
Isobutyl acetate	0.06	0.02	0.13	0.02	0.07	0.02
Ethyl 3-phenylpropionate	0.04	0.00	0.10	0.01	0.05	0.02
Dimethyl phthalate	0.06	0.02	0.10	0.01	0.04	0.01
Diethyl phthalate	0.05	0.01	0.07	0.02	0.08	0.03
Diisobutyl phthalate	0.05	0.01	0.07	0.00	0.04	0.01
Ethyl 9-hexadecenoate	0.02	0.01	0.10	0.03	0.06	0.02
Propyl acetate	0.04	0.01	0.07	0.00	0.05	0.01
Ethyl butyrate	0.02	0.00	0.08	0.01	0.05	0.02
Ethyl isovalerate	–	–	0.06	0.01	–	–
Ethyl acetate	–	–	0.06	0.01	–	–
Ethyl phenylacetate	–	–	0.06	0.00	–	–
Methyl benzoate	0.01	0.00	0.02	0.00	0.05	0.01
Isobutyl caprate	–	–	0.03	0.01	0.07	0.03
3-Methylbutyl 2-methylpropanoate	0.02	0.00	0.09	0.01	–	–
Ethyl 2-methylbutyrate	0.02	0.01	0.05	0.00	0.03	0.01
Ethyl linoleate (branched chain straight chain)	0.01	0.00	0.05	0.01	–	–
Ethyl phenylacetate	0.01	0.00	–	–	0.02	0.01
Methyl acetate	0.01	0.00	0.02	0.00	–	–
Amyl acetate	–	–	0.02	0.00	0.01	0.00
Total esters	42.57	6.68	84.11	17.5	37.89	17.52
Benzaldehyde	0.19	0.05	0.43	0.07	0.29	0.14
Acetaldehyde	0.10	0.09	0.23	0.04	0.46	0.25
Furfural	0.05	0.01	0.22	0.03	0.20	0.08
Valeraldehyde	0.07	0.01	0.12	0.03	0.06	0.01
Isovaleraldehyde	–	–	0.05	0.01	0.03	0.01
4-Hydroxy-3-methoxybenzaldehyde	0.03	0.01	0.05	0.00	–	–
Hexanal	–	–	0.03	0.00	0.02	0.00
Total aldehydes	0.44	0.17	1.13	0.18	1.06	0.49
2-Octanone	0.02	0.00	–	–	0.22	0.28
4,6-Dimethyl-2-heptanone	0.05	0.01	0.06	0.01	0.07	0.02
4-Methyl-2-heptanone	0.04	0.01	0.04	0.01	–	–
Acetone	0.02	0.00	0.04	0.00	0.03	0.00
Methyl octenone	0.01	0.00	0.02	0.00	0.02	0.00
2-Nonanone	0.01	0.00	0.02	0.01	0.01	0.00
2-Hexanone	0.01	0.00	0.01	0.00	0.01	0.00
Total ketones	0.16	0.02	0.19	0.03	0.36	0.30
Caproic acid	32.16	5.44	66.03	9.41	46.23	15.99
Octanoic acid	5.75	1.32	10.51	2.61	23.20	9.54
Hexanoic acid	2.55	0.48	5.04	0.73	3.20	0.93
9-Decenoic acid	3.32	1.40	0.78	0.84	4.33	1.78
Acetic acid	0.50	0.13	0.96	0.12	0.69	0.27
2-Methylbutyric acid	–	–	0.16	0.07	0.25	0.09
Butyric acid	0.11	0.06	–	–	0.06	0.03
Isobutyric acid	0.02	0.00	–	–	0.12	0.04
Valeric acid	–	–	–	–	0.04	0.01
Total acids	44.41	8.83	83.48	13.78	78.12	28.68

Note: “-” indicates not detected.

As can be seen from Table 1, the types of volatile aroma substances in the three beer samples are generally similar. A total of 82 aroma substances were detected, including 35 alcohols, 24 esters, 9 acids, 7 aldehydes, and 7 ketones. Among them, 71, 75, and 73 aroma substances were detected in Eng beer, Can beer, and Ger beer, respectively, and the total relative contents of volatile aroma components were 150.17%, 271.82%, and 195.31%, respectively.

A total of 24 alcohol substances were detected in the three beers. Eng beer, Can beer, and Ger beer each detected 22 alcohol substances, of which 20 alcohols were common; the total relative contents of alcohols were 62.59%, 102.91%, and 77.88%, respectively, accounting for 41.68%, 37.86%, and 39.88% of the total flavor substances of the corresponding beer samples. Among them, the main alcohol substances include phenethyl alcohol, isoamyl alcohol, ethanol, isobutanol, etc. Alcohols mainly come from yeast metabolism during fermentation, and secondly also come from the decomposition of sugars and ester substances in wort, etc. [24]. The fermentation results of different types of malts showed that phenethyl alcohol and isoamyl alcohol had the highest contents in the three beer samples, and the contents of the two accounted for 82.97%–91.16% of the total alcohol content.

As an important component of higher alcohols in beer, phenethyl alcohol and isoamyl alcohol endow alcoholic beverages with a full aroma and taste, and increase the harmony of the wine, constituting the main aroma of the beer body. Phenethyl alcohol is mainly synthesized from phenylalanine via the Ehrlich pathway under the action of aminotransferase, decarboxylase, and dehydrogenase [25], and it originates from the husk of malt, rather than the endosperm and the whole barley [26]. Among the three beer samples, the relative content of phenethyl alcohol was the highest in Can sample (46.37%), followed by Ger sample (41.07%), and the lowest in Eng sample (35.16%). Butanol, a higher alcohol, imparts a strong pungent taste to the beer body [27]. Compared with other beer samples, the relative content of isobutanol in Eng sample was the lowest, which has an effect on the beer flavor has a relatively small influence on beer flavor. Higher alcohols are precursor substances of certain ester compounds, increasing the types of esters formed during beer fermentation, which plays an important role in beer flavor and can improve the sensory quality of beer [28]. Can beer has a rich relative content of total ester substances, and Canadian malt is more suitable for brewing beer with higher alcohol content.

2.3.2 Comparative analysis of ester substances

Appropriate volatile ester compounds not only enrich the flavor and aroma coordination of beer, but also have a certain masking effect on beer aging substances [29]. A total of 35 ester substances were detected in the three beer samples, mainly ethyl octanoate, ethyl hexanoate, ethyl acetate, etc. Eng beer, Can beer and Ger beer detected 30, 34 and 30 ester substances respectively, of which 26 esters were common; the relative contents of total esters were 42.57%, 84.11% and 37.89% respectively, accounting for 28.35%, 30.94% and 19.40% of the total flavor substances of the corresponding beer samples.

Volatile esters are important flavor substances in beer and also the main carrier of beer aroma [30]. Appropriate amounts of esters give beer floral and fruity aromas, making the flavor harmonious [31]. Among them, ethyl octanoate, ethyl hexanoate, ethyl acetate, and isoamyl acetate are typical ester substances in beer [32]. In Eng beer, Can beer and Ger beer, the relative concentration of ethyl octanoate was the highest, accounting for 48.48%, 41.04% and 34.34% of total esters respectively, giving the beer body a sweet and mellow aroma. At the same time, ethyl hexanoate with a pineapple and apple aroma, ethyl acetate with a sweet aroma, and isoamyl acetate with a banana aroma all have relatively high contents, endowing the beer with a rich fruity aroma [33].

Among them, Can beer has the highest total relative content of esters and the richest ester types; at the same time it contains ethyl acetate with a relatively high concentration, which has apple and peach fruity aromas and pleasant fragrance, and can appropriately alleviate the irritability brought by phenethyl alcohol in beer.

Esters and alcohols are the most important components of beer aroma substances. The relative ester content is high or the ratio of esters to alcohols is suitable, and the beer will have a richer aroma; while high alcohol content can effectively relieve headaches, and the ratio of esters to alcohols is an important basis for evaluating beer flavor points [34]. Generally, an ester-to-alcohol ratio of 3～4 is more ideal, but at present most beers tend to be biased lower [35].

The results of ester-to-alcohol ratio in the three beer samples are shown in Table 2. As can be seen from Table 2, the ester-to-alcohol ratio of Ger beer is the highest, between 2～3, while the ester-to-alcohol ratios of the other two beer samples are both between 1～2. Therefore, it can be seen that different types of malts still have large differences in the ester-to-alcohol ratio of the brewed beer.

Table 2 Effect of different types of malts on the ratio of alcohols to esters

Sample	Total alcohols	Total esters	Ratio of alcohols to esters
Eng beer	62.59	42.57	1.47
Can beer	102.91	84.11	1.22
Ger beer	77.88	37.89	2.06

2.3.3 Comparative analysis of aldehyde substances

Aldehydes are the main class of aging compounds in beer. Aldehyde compounds in fresh beer are mainly formed during wort boiling or filtration; with the extension of storage period, the content of aging aldehyde compounds gradually increases, eventually causing beer flavor instability and deterioration [36]. In the three beer samples, a total of 7 aldehyde substances were detected, including benzaldehyde (bitter almond, cherry and坚果 aromas), acetaldehyde (green grass aroma), etc. Eng beer, Can beer and Ger beer detected 5, 7 and 6 aldehyde substances respectively, of which 4 aldehydes were common; the relative contents of total aldehydes were 0.44%, 1.13% and 1.06% respectively, accounting for 0.29%, 0.42% and 0.54% of the total aroma components.

Benzaldehyde is the low-volatility aldehyde with the highest content in beer, with a bitter-almond nutty aroma [37]. The relative content of benzaldehyde in Can sample (0.43%) was significantly higher than that in other beer samples. The benzaldehyde formed in fermentation will give the beer sample a green-grassy taste, shorten the shelf life. As a prominent aroma component feature in beer, the ethanol content of Ger sample is significantly higher than that of other beer samples, forming a prominent raw-wort taste.

Among the three beer samples, the relative content of aldehyde aroma substances in Eng beer is relatively low; the relative content of aldehyde aroma substances in Can beer is the highest, and total aldehydes are the highest, while maintaining the typicality of beer; Ger sample has a relatively low aldehyde aroma substance content compared with Can sample, but the percentage of this sample in total aroma components is the highest.

2.3.4 Comparative analysis of ketone substances

Among the three beer samples, a total of 7 ketone substances were detected. Eng beer, Can beer and Ger beer detected 6, 7 and 6 ketone substances respectively, of which 5 ketones were common; the total relative contents of ketones were 0.16%, 0.19% and 0.36% respectively, accounting for 0.11%, 0.07% and 0.18% of total aroma components.

Among the three beer samples, Eng sample has the richest types of ketone aroma substances, Ger sample has the highest relative content of ketone aroma substances. Although there is no significant difference in ketone content, the relative content in Ger sample is the highest (0.22%), giving the beer sample a strong “fermented aroma, cooked rice aroma,” and accompanied by the aromas of milk, cheese, and mushrooms, while no ketones were detected in Can beer.

2.3.5 Comparative analysis of acid substances

Among the three beer samples, 9 acid substances such as caproic acid, octanoic acid, and hexanoic acid were detected. Eng beer, Can beer and Ger beer detected 7, 6 and 9 acid substances respectively, of which 5 acids were common; the total relative contents of acids were 44.41%, 83.48% and 13.78%, accounting for 29.56%, 30.70%, and 39.99% of total aroma components.

In Eng beer, Can beer and Ger beer, the relative content of caproic acid was the highest, accounting for 72.42%, 79.10% and 59.18% of total acids respectively; octanoic acid was second, accounting for 12.95%, 12.59% and 29.70% of total acids respectively. Both are representative substances of “acidic off-flavor” in beer. They easily form rancid odor of the beer body, leading to a decline in beer quality, among which the relative content of caproic acid in Can sample (66.03%) and the relative content of octanoic acid in Ger sample (23.20%) are significantly higher than those in other beer samples. At the same time, hexanoic acid was detected in all three beer samples, and Can sample has the highest relative content (5.04%), which is easy to form a bitter astringent off-flavor of the beer body, increasing the sourness of the beer body. The total acid contents of Can and Ger samples are more than twice that of Eng sample, proving that differences in raw materials during the brewing process will directly cause significant differences in the content of free fatty acids in the finished beer.

Based on the comprehensive analysis results of aroma components, phenethyl alcohol (rose aroma), isoamyl alcohol (sweet aroma), ethyl octanoate (mellow aroma), and ethyl hexanoate (pineapple aroma, apple aroma) are the main components of beer body aroma. Compared with other beer samples, Can sample has the richest types of aroma substances (75 kinds), the highest relative content (271.82%), the richest aroma types, and a soft and round beer body. Specifically, the total alcohol and total ester contents are high, the beer body aroma is complex, mellow and harmonious, further proving that Canadian malt is more suitable for Lager beer brewing than other malts.

3 Conclusions

In this study, by measuring the wort extract, α-amino acid nitrogen, saccharifying power, Kolbach index and other quality indices of three different types of malts, as well as the alcohol content, original wort concentration, real extract, apparent extract and other physicochemical indices of the brewed beers, and by using GC-MS to analyze the differences in beer flavor components, the results showed that, compared with British malt and German malt, Canadian malt had moderate Kolbach index (45%) and extract rate (≥77%), higher saccharifying power (414 WK), and relatively low α-amino acid nitrogen (118 mg/L); meanwhile, the physicochemical indices of beer brewed by Canadian malt were moderate (alcohol content 2.97%vol, original wort concentration 8.63 °P), with lower extract (real extract 2.76%, apparent extract 1.17%) and higher fermentation degree (real fermentation degree 67.97%, apparent fermentation degree 86.41%). It was finally determined that Canadian malt is more suitable than other malts for Lager beer brewing/production in the laboratory simulated system.

References

SUN Kecheng, ZHAO Xinrui, GU Qianhui, XIE Tingting, LI Jianghua, DU Guocheng. Effect of different types of malts on the brewing flavor of Lager beer[J]. China Brewing, 2021, 40(4): 148-154.