Off-Flavour Compounds in Beer with a HSS/GC/MSD System

Conditions and Instruments:

HP 7694 Headspace Sampler (Silcosteel):

8ml beer (4g Na2SO4) in 20 ml HSS-vial
Sample temp. 80°C for 15 min., high shake
1 ml sample loop
Analysis of Diacetyl
Before the analysis, each sample was heated 10 min. to 60°C (Butadione), Pentadiones,
and then vented for 1 min. to release most of the CO2
Sulphur Compounds,
HP 6890 GC: Alcohols and Esters in Beer
Split/splitless inlet 200°C, split 5:1 (3:1 for SIM), direct liner with HSS/GC/MSD
Oven: 36°C (2 min.) 8°C/min. to 220°C (5 min.)
Column: 60m x 0.32mm x 1.8µm HP-624
Normally 3 different analyses
are necessary:
necessary
HP5973 MSD, Scan: 1- GC/ECD analysis for
Interface/source/quad. temp.: 250/200/120°C
Tune: Autotune
Butadione and Pentadiones
Scan range: 25-300 amu 2- GC/FPD analysis for the
EM offset: Atune + 400 V Sulphur compounds
SIM: 3-GC/FID analysis for the
EM offset: Atune + 500 V
Low resolution Alcohols and Esters
Dwell time: 50 - 200 ms

1
 Off-Flavour Compounds in Beer with a HSS/GC/MSD System
Scan Analysis of 8 ml Beer + 4g Na2SO4, split 5:1
Abundance CO2 Ethanol Ethylacetate i-Pentanol
6000000 i-Butanol Isoamylacetate

5500000

5000000

4500000

4000000

3500000
1-Propanol
CS2
3000000 Acetaldehyde

2500000

2000000 Acetal
Ethyl-
1500000 hexanoate

1000000

50000
0

Time-->
4.00 6.00 8.00 10.0 12.00 14.00 16.00 18.00
0

To increase the sensitivity for the full scan analysis of the beer sample, about 4g Na2SO4 was added.
Already in the TIC, most of the alcohols and esters are visible.

2
 Off-Flavour Compounds in Beer with a HSS/GC/MSD System
Diones in 8 ml Beer + 4g Na2SO4, split 5:1
Extracted Ions 86 and 100

By extracting the ions 86 and 100, Butadione and the 2,3-Pentadione are visible.
The concentration of the Butadione is about 60ppb in this beer.
Ion 100.00 (99.70 to 100.30): BEER-9.D
Butadione
Ion 86.00 (85.70 to 86.30): BEER-9.D
Abundance (Diacetyl)
16000
15000
14000
13000
12000
11000
10000
9000 2,3-Pentadione
8000
7000
6000
5000
4000
3000
2000
1000

Time--> 8.00 9.00 10.00 11.00 12.00 13.00

3
Off-Flavour Compounds in Beer with a HSS/GC/MSD System
Scan Analysis of 8 ml Beer + 4g Na2SO4, split 5:1

4
Off-Flavour Compounds in Beer with a HSS/GC/MSD System
Sulphur Compounds in 8 ml Beer + 4g Na2SO4, split 5:1

Ion 62.00 (61.70 to 62.30): BEER-9.D

CS2
Abundance Ion 78.00 (77.70 to 78.30): BEER-9.D
120000 Ion 90.00 (89.70 to 90.30): BEER-9.D

110000

100000

90000
Dimethylsulfide
80000

70000

60000

50000

40000

30000

20000 Methylthioacetate
(Ethanethioicacid S-methylester)
10000

Time--> 6.00 7.00 8.00 9.00 10.00 11.00 12.00 13.0

0

5
Off-Flavour Compounds in Beer with a HSS/GC/MSD System
Scan Analysis of 8 ml Beer + 4g Na2SO4, split 5:1

6
Off-Flavour Compounds in Beer with a HSS/GC/MSD System
SIM Analysis of 8 ml Beer, split 3:1

Ethylacetate
Abundance i-Pentanol Isoamylacetate
i-Butanol
1200000
CS2
1100000

1000000
1-Propanol
900000
Acetaldehyde
800000

700000

600000

500000

400000

300000

200000 Dimethyl Ethylbutyrate Ethyloctanoate

Ethylhexanoate
sulfide Diacetyl
100000
~20 ppb

Time--> 6.00 8.00 10.00 12.00 14.00 16.00 18.00 20.00 22.00

7
Off-Flavour Compounds in Beer with a HSS/GC/MSD System
SIM analysis of 8 ml Beer, split 3:1 60m x 0.25mm x 0.5µm INNOWax column

Abundance Ethylacetate Iso-Amylalcohol

TIC: BEER-05.D
300000
0 Acetaldehyde
280000
0
260000
0
240000
0
220000
0
200000
0 CS2
180000
0
160000
0
140000
0 Iso-Amylacetate
120000
0
100000
0
80000
0 DMS
60000
0
2-Phenylethanol
40000 Propanol-1 Ethylhexanoate
0 Iso-Butanol
20000
0

Time--> 2.00 6.00 10.00 14.00 18.00 22.00

8
Off-Flavour Compounds in Beer with a HSS/GC/MSD System
Diones (SIM) in 8 ml Beer, split 3:1

Ion 100.00 (99.70 to 100.30): BEER-20.D

Abundance Ion 90.00 (89.70 to 90.30): BEER-20.D 2,4-Pentadione
3800
Ion 86.00 (85.70 to 86.30): BEER-20.D
3600
3400
Diacetyl ~20ppb
3200
3000
2800
2600
2400
2200
2000
1800
2,3-Pentadione
1600
Methylthioacetate
1400
1200
1000
800
600
400
200

Time--> 9.00 10.00 11.00 12.00 13.00 14.0 15.00

0

9
Off-Flavour Compounds in Beer with a HSS/GC/MSD System
Relative Standard Deviation (%) for the Diones and Methylthioacetate, 4 Analyses

RSD 2.7%
Abundance Diacetyl ~60ppb
Ion 100.00 (99.70 to 100.30): BEER-23.D
5000
Ion 86.00 (85.70 to 86.30): BEER-23.D RSD 2.5%
Ion 90.00 (89.70 to 90.30): BEER-23.D
Ion 100.00 (99.70 to 100.30): BEER-22.D 2,4-Pentadione
4500 Ion 86.00 (85.70 to 86.30): BEER-22.D RSD 4.5%
Ion 90.00 (89.70 to 90.30): BEER-22.D
Ion 100.00 (99.70 to 100.30): BEER-20.D Methylthioacetate
Ion 86.00 (85.70 to 86.30): BEER-20.D
4000 Ion 90.00 (89.70 to 90.30): BEER-20.D
Ion 100.00 (99.70 to 100.30): BEER-21.D
Ion 86.00 (85.70 to 86.30): BEER-21.D
Ion 90.00 (89.70 to 90.30): BEER-21.D
3500

3000
RSD 5.0%
2,3-Pentadione

2500

2000

1500

1000

500

Time--> 10.00 11.00 12.00 13.00

10
 Off-Flavour Compounds in Beer with a HSS/GC/MSD System
Acetaldehyde 15729825.99 1-Propanol 7719302.981 3-Methylbutanol 13367179.73 2,4-Pentadione 117354.72
Acetaldehyde 15341274.82 1-Propanol 7923033.760 3-Methylbutanol 13893632.59 2,4-Pentadione 123422.40
Acetaldehyde 14721893.88 1-Propanol 7428209.195 3-Methylbutanol 12882220.20 2,4-Pentadione 120364.16
Acetaldehyde 14695810.38 1-Propanol 7478456.869 3-Methylbutanol 13083777.11 2,4-Pentadione 123872.64
Average 15122201.27 7637250.701 13306702.41 121253.48
Stdev. 503075.94 228996.63 438947.75 3030.70
% RSD 3.33 3.00 3.30 2.50

Dimethylsulfide 2327403.593 Diacetyl 165029.402 2-Methylbutanol 42442052.81 Ethylbutyrate 2685026.226

Dimethylsulfide 2361272.455 Diacetyl 169227.593 2-Methylbutanol 43768634.10 Ethylbutyrate 2783782.171
Dimethylsulfide 2143758.171 Diacetyl 159985.739 2-Methylbutanol 41144350.37 Ethylbutyrate 2593955.535
Dimethylsulfide 2153378.128 Diacetyl 159911.968 2-Methylbutanol 41004121.82 Ethylbutyrate 2606203.715
Average 2246453.087 163538.676 42089789.78 2667241.912
Stdev. 113938.09 4485.62 1292947.29 87548.97
% RSD 5.07 2.74 3.07 3.28

CS2 23834861.24 Ethylacetate 24712133.91 Methylthioacetate 104140.563 3-Methylbutylacetate 18896570.02

CS2 22745171.40 Ethylacetate 25339672.34 Methylthioacetate 104097.010 3-Methylbutylacetate 18642382.66
CS2 23528797.65 Ethylacetate 23819146.95 Methylthioacetate 96167.281 3-Methylbutylacetate 17338112.26
CS2 25337386.63 Ethylacetate 23846162.55 Methylthioacetate 96353.152 3-Methylbutylacetate 18201991.27
Average 23861554.23 24429278.94 100189.502 18269764.05
Stdev. 1085638.27 735098.37 4537.82 684177.99
% RSD 4.55 3.01 4.53 3.74

Ethylformate 335660.255 i-Butanol 3145014.027 2,3-Pentadione 68543.04 2-Methylbutylacetate 2384360.161

Ethylformate 345895.404 i-Butanol 3216562.846 2,3-Pentadione 69739.75 2-Methylbutylacetate 2525903.050
Ethylformate 328526.972 i-Butanol 3006342.592 2,3-Pentadione 63412.80 2-Methylbutylacetate 2353445.675
Ethylformate 330028.435 i-Butanol 3034850.940 2,3-Pentadione 63492.24 2-Methylbutylacetate 2359332.033
Average 335027.767 3100692.601 66296.96 2405760.230
Stdev. 7868.91 97685.47 3320.77 81208.89
% RSD 2.35 3.15 5.01 3.38

Relative Standard Deviation for 4 Analyses

11
Off-Flavour Compounds in Beer with a HSS/GC/MSD System
Diones (SIM) in 8 ml Beer, split 3:1

Ion 100.00 (99.70 to 100.30): BEER-20.D

Diacetyl ~60ppb Ion 90.00 (89.70 to 90.30): BEER-20.D
Abundance

7500

7000

6500

6000

5500 2,4-Pentadione
5000

4500 2,3-Pentadione
4000

3500

3000

2500

2000

1500

1000

500

Time--> 9.00 10.00 11.00 12.00 13.00 14.00 15.00

12
Off-Flavour Compounds in Beer with a HSS/GC/MSD System
Diones and DMS (SIM) in 8 ml Beer, split 3:1 60m INNOWax column

Abundance DMS Abundance Diacetyl

Ion 86.00 (85.70 to 86.70): BEER-06.D

Ion 100.00 (99.70 to 100.70): BEER-06.D

50000 6000

Ion 62.00 (61.70 to 62.70): BEER-06.D 5500

45000

5000
40000
4500

35000
4000
2,3-Pentadione
30000
3500

25000 3000

2500
20000

2000
15000

1500

10000
1000

5000
500

Time--> 3.00 3.3 3.60 3.90 4.20 Time--> 7.80 8.20 8.60 9.00 9.40

13