Analysis of Some Alcohols, Aldehydes, and Esters in Distilled Spirits with the Agilent 8860 Gas Chromatograph

Application Note Spirits Analysis

Author Abstract Youjuan Zhang Agilent Technologies The Agilent 8860 GC configured with a flame ionization detector (FID) provides (Shanghai) Co. Ltd. Shanghai high sensitivity, good linearity, and stability for the analysis of alcohols, aldehydes, 200131 P. R. China organic acids, and esters in distilled spirits. The highly inert performance of the column ensures excellent peak shapes for most compounds. Introduction Experimental To achieve the required levels, six vials were made at each calibration level Distilled spirits constitute a complex Analyses were performed using an by spiking varying amounts of stock mixture of hundreds of flavor 8860 GC equipped with an FID. Sample solution in ethanol aqueous solution. compounds in an ethanol-water matrix, introduction was done using an The calibration standards were prepared including alcohols, aldehydes, organic Agilent 7693A automatic liquid sampler at standard concentrations of 10, 30, acids, and esters. The proportion of with a 5 µL syringe. Table 1 shows the 50, 100, 500, and 1,000 µg/mL. An these trace ingredients determines the instruments and conditions used. internal standard (IS) was spiked to each flavor and quality of the liquor. It is very calibration level, corresponding to an IS important for manufacturers to monitor Sample preparation concentration of 440 µg/mL. Mixed standard stock solutions were and test flavor compounds that originate The spirits samples were obtained prepared by adding defined amounts from the original raw materials and the locally from retail providers in China, and of each single standard compound subsequent processes of mashing, introduced into the GC as neat samples. fermentation, and distillation. with a microliter syringe. A stock solution of 10 compounds, each with Chinese liquor is a spirit famous a concentration of 1,000 µg/mL was worldwide for its strong flavor and long prepared in 60:40 ethanol:water solution aftertaste. According to flavor types, (v/v). Chinese liquor can be divided into Maotai, Luzhou, Fen, and rice flavors, among others. For a long time, the Table 1. Agilent J&W DB-FATWAX UI column for spirits method conditions. research on liquor focused primarily Parameter Value on the evolution of microorganisms, GC system 8860 GC/FID and the formation of flavor substances Split/splitless, 250 °C, split ratio 30:1 Inlet during fermentation. The influence of Liner: Ultra Inert (p/n 5190-2295) main aroma components such as esters, Column J&W DB-FATWAX Ultra Inert, 30 m × 0.25 mm, 0.25 µm (p/n G3903-63008) acids, alcohols, and aldehydes of the typical liquor style was also analyzed. Carrier Helium, 1 mL/min, constant flow The concentration and proportion of 40 °C (4 minutes), Oven then 5 °C/min to 100 °C, ethyl acetate, ethyl lactate, and ethyl then 10 °C/min to 200 °C (10 minutes) hexanoate play a decisive role in the 250 °C, Hydrogen: 30 mL/min, flavor of liquor. For the sake of the health FID Air: 300 mL/min, of the drinkers, the concentration of Make-up gas (N2): 25 mL/min methanol, isobutyl alcohol, and isoamyl Injection 0.5 µL alcohol should be strictly controlled in the production process of liquor. In this Application Note, some real samples were injected into the Agilent 8860 GC system, and certain liquor flavor types were analyzed. The 10 most common compounds in liquor were also quantitatively analyzed.

2 pA ISTD Ethanol Results and discussion 1. Ethyl acetate 35 2. Methanol To develop a reliable method for 3. 2-Butanol monitoring flavor substances in 4. Ethyl butyrate 30 5. n-Propanol distilled spirits, an 8860 GC configured ISTD. n-Butyl acetate 6. Isobutanol with automatic injection and an FID 25 7. 1-Butanol was used. Figure 1 shows a typical 8. Isoamyl alcohol chromatogram acquired by the system 20 6 9. Ethyl hexanoate 10. Ethyl lactate with the 10 standard compounds and one internal standard at a concentration 15 4 8 7 9 of 100 µg/mL. 3 5 10 10 Figures 2, 3, and 4 are example 1 2 chromatograms profiling some of 5 the major components that were found in different flavors of Chinese 0 2 4 6 8 10 12 14 16 18 liquor samples. The system shows Time (min) great resolution and peak shape for Figure 1. GC/FID chromatogram of 10 target compounds (100 μg/mL) using a J&W DB-FATWAX Ultra all alcohols, aldehydes, organic acids, Inert column. and esters. As shown in Figure 2, ethyl acetate, acetal, and methanol were 1. Acetaldehyde 10. Ethyl butyrate 19. 3-Hydroxyl-2-butanone 28. 1,2-Propanediol almost baseline-separated. Methanol is 2. Acetone 11. n-Propanol 20. Ethyl heptanoate 29. Butyric acid 3. Ethyl acetate 12. Isobutanol 21. Ethyl lactate 30. Valeric acid strongly polar and is prone to tailing, but 4. Acetal 13. Ethyl valerate 22. n-Hexanol 31. Hexanoic acid in this method, a sharp and symmetric 5. Methanol 14. 2-Pentanol 23. Ethyl octanoate 32. 2-Phenylethanol 6. Isovaleraldehyde 15. n-Butanol 24. Acetic acid 33. Heptanoic acid peak shape was obtained. 7. Ethanol 16. Isoamyl alcohol 25. Furfural 34. Octanoic acid 8. 2-Pentanone 17. Ethyl hexanoate 26. 2,3-Butanediol(levo) 35. Ethyl palmitate 9. 2-Butanol 18. n-Pentanol 27. 2,3-Butanediol(meso)

pA 7 17 31

30 16 21 25

20 3 4 1 11 15 12 24 29 10 2 8 15 28 10 22 13 25 27 30 32 5 26 33 6 9 14 19 34 35 5 18 20 23

0 5 10 15 20 25 30 Time (min)

Figure 2. GC/FID chromatogram of a Luzhou flavored Chinese liquor sample.

3 1. Acetaldehyde 10. Ethyl butyrate 19. n-Pentanol 28. 2,3-Butanediol(meso) 2. Acetone 11. n-Propanol 20. 3-Hydroxyl-2-butanone 29. 1,2-Propanediol 3. Ethyl acetate 12. n-Butyl acetate (IS) 21. Ethyl heptanoate 30. Butyric acid 4. Acetal 13. Isobutanol 22. Ethyl lactate 31. Valeric acid 5. Methanol 14. Ethyl valerate 23. n-Hexanol 32. Hexanoic acid 6. Isovaleraldehyde 15. 2-Pentanol 24. Ethyl octanoate 33. 2-Phenylethanol 7. Ethanol 16. n-Butanol 25. Acetic acid 34. Heptanoic acid 8. 2-Pentanone 17. Isoamyl alcohol 26. Furfural 35. Octanoic acid 9. 2-Butanol 18. Ethyl hexanoate 27. 2,3-Butanediol(levo) 35. Ethyl palmitate 22 pA 3 7 11 40

35 17 25 30

20 13 29 12 15 4 26 1 10 32 9 27 10 15 16 18 20 23 5 14 21 24 30 28 33 36 5 2 6 8 19 31 34 35

0 5 10 15 20 25 30 Time (min)

Figure 3. GC/FID chromatogram of a Maotai flavored Chinese liquor sample.

pA 4 1. Acetone 11 2. Ethyl acetate 3. Methanol 18 6 4. Ethanol 5. n-Propanol 6. -Butyl acetate (IS) 16 n 10 7. Isobutanol 8. n-Butanol 14 9 9. Isoamyl alcohol 10. Ethyl lactate 12 11. Acetic acid 12. 2,3-Butanediol(levo) 13. 2,3-Butanediol(meso) 10 14. Hexanoic acid 15. 2-Phenylethanol 8 5 7

12 6 3 1415 2 13 4 1 8

5 10 15 20 25 30 30 Time (min)

Figure 4. GC/FID chromatogram of a Fen flavored Chinese liquor sample.

4 The correlation coefficients (R2) for Table 2. R2, RSDs, and LODs for 10 target compounds. all 10 compounds were ≥0.9992 RSD (n = 6) for the range of 10 to 1,000 µg/mL. Table 2 shows detailed calibration Chinese MDL Name RT R2 100 µg/mL liquor (µg/mL) information, and Figures 5, 6, and 7 show the calibration curves of methanol, Ethyl acetate 4.51 0.9992 3 2.8 3 isobutanol, and ethyl lactate. Methanol 4.75 0.9998 1.3 1.2 5 2-Butanol 7.88 0.9998 1.3 1.4 2

Ethyl butyrate 8.09 0.9995 2.5 2.2 2

n-Propanol 8.26 0.9998 1.1 1.1 2

Isobutanol 9.96 0.9998 1.1 1.2 2

1-Butanol 11.43 0.9998 1.4 1.2 2

Isoamyl alcohol 13.25 0.9998 1.2 1.3 2

Ethyl hexanoate 13.88 0.9998 1.6 1.1 2

Ethyl lactate 16.93 0.9999 1.1 1.2 3

Methanol, FID1 A 1.4 Area ratio = 0.55615073*AmtRatio + 0.0059838 Rel. res%(6): –6.4427e-1 1.2 6 Correlation: 0.99985 1.0

tio 0.8 5 0.6 Area ra

0.4

0.2 4 3 1 2 0

0 0.5 1.0 1.5 2.0 Amount ratio

Figure 5. Methanol calibration curve (R2 = 0.99985).

Isobutanol, FID1 A Area ratio = 1.98236322*AmtRatio +0.0125253 Rel. res%(1): –16.627 6 4 Correlation: 0.99986

3 tio 5

Area ra 2

1 4 3 1 2 0

0 0.5 1.0 1.5 2.0 Amount ratio

Figure 6. Isobutanol calibration curve (R2 = 0.99986).

5 To verify the integrity of the system, Ethyl lactate, FID1 A repeatability was determined. Area ratio = 0.63500293*AmtRatio + 0.0031097 1.4 Rel. res%(1): –11.876 Repeatability tests were performed 6 Correlation: 0.99990 on both the standard and the real 1.2 samples. Table 2 illustrates that for 1.0 most compounds, the area %RSD is tio 5 well below 3 %. Figure 8 is an overlay 0.8 Area ra of six replicates of the Chinese liquor 0.6 injections. As the figure illustrates, the retention time and area stability were 0.4 excellent. 0.2 4 3 1 2 Signal-to-noise ratio (S/N) was used 0 for method detection limit (MDL) 0 0.5 1.0 1.5 2.0 Amount ratio calculation. A concentration of 5 µg/mL 2 standard solution was used to test the Figure 7. Ethyl lactate calibration curve (R = 0.99985). MDL, and Table 2 lists the values for all pA compounds. For all compounds, the MDLs were ≤5 µg/mL. 80

Conclusion 70

In this Application Note, the 8860 GC 60 configured with automatic injection and an FID provides a reliable and 50 economical solution for the analysis of alcohols, aldehydes, organic acids, and 40 esters in liquor. The EPC control and J&W DB-FATWAX UI column provides 30 excellent peak shape, resolution, and 20 great repeatability. 10 References 0 1. Kenneth L.; Zhou, Y. Analysis of Distilled Spirits Using an Agilent J&W 5 10 15 20 25 30 DB-WAX UItra Inert Capillary GC Time (min) Column, Agilent Technologies Figure 8. Overlaid GC/FID chromatograms of six repeat injections of the same Chinese liquor. Application Note, publication number 5991-6638EN, 2016. 2. Cai, X. Y.; Yin, J. J.; Hu, G. D. Determination of Minor Flavor Components in Chinese Spirits by Direct Injection Technique with Capillary Columns. Chin. J. Chromatogr. 1997, 15(5).

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