perkinelmer: determining flavors and “defects” in beer by headspace trap/gas chromatography/mass...
DESCRIPTION
Beer is a popular beverage produced by the fermentation of hopped malt extracted from barley and other grains. Some compounds (flavors) have a positive effect on aroma (attributes) and some have a negative effect (defects). This presentation will focus on a new method that enables the investigation and characterization of flavors and defects of beer in one analysis using HS trap/GC/MS. Classically, this analysis is performed on four separate detectors. This new method employs one detector (MS) to provide these solutions required for the production and the testing of beer. The outcome is a more cost effective, accurate means to ensure the validity and the quality control of their product. Other benefits include enhanced productivity, attaining more information from a single analysis, and requiring less bench space. The following experiments and results will be discussed. • Quantitation of dimethyl sulfide (DMS), 2,3-butanedione (diacetyl), 2,3-pentandione and t,2-nonenal • Characterization of several types of beers • Fermentation profiling • Analysis of raw materials • Aging studies Originally presented at Pittcon 2012.TRANSCRIPT
11 © 2009 PerkinElmer© 2009 PerkinElmer© 2009 PerkinElmer© 2012 PerkinElmer
Determining Flavors and “Defects” in Beer by Headspace Trap/Gas
Chromatography/Mass Spectrometry
Andrew Tipler, Chromatography R&D ManagerLee Marotta, Field Application Scientist
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This presentation will describe a system that provides an almost comprehensive analysis of flavor compounds and defects in beer Theory Design and operation Beer Application
The system comprises the following components: A headspace trap sampling system A gas chromatograph A mass spectrometer
Content
33
Ethanol - FID
Defects Vicinal Diketones (‘butterscotch’) - ECD Dimethyl Sulfide (sulfury character) – FPD or SCD Aldehydes (oxidation ‘cardboard’ products) - FID Thiols (skunkiness) - SCD
Flavor Compounds – MS (characterization) Alcohols Ketones Esters Acids Turpenes
Fermentation Markers Diacetyl
Replacing Multiple Systems with One …
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The authors would like to thank Bill Yawney of the LongTrail Brewery, Vermont, for his advice, inspiration and some of the analytical data used in this presentation
Acknowledgement
55 © 2009 PerkinElmer
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Headspace Instrumentation – Theory, Design and Operating Principles
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Principles Behind Headspace Sampling
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Besides actually making beer, much of the fun is associated with drinking it!
It’s nutritious, it makes the world easier to live in and it tastes good.
Taste is obviously subjective but we beer connoisseurs generally consider the following when drinking a fine beer: Don’t drink out of the bottle Don’t cool the beer to Arctic temperatures Use an appropriately shaped glass Don’t fill the glass completely
Tasting Beer
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Besides actually making beer (of course), much of the fun is associated with drinking it!
It’s nutritious, it makes the world easier to live in and it tastes good.
Taste is obviously subjective, but beer connoisseurs will generally consider the following when drinking a fine beer: Don’t drink out of the bottle Don’t cool the beer to Arctic temperatures Use an appropriately shaped glass Don’t fill the glass completely These are all done to ensure that the beer
aroma is involved in the tasting process (beer aroma, or ‘nose’ as it’s called, is an important part of the formal beer-judging process)
Tasting Beer
99
Headspace sampling is a bit like smelling the aroma
Step 1 – put beer sample into a vial and seal it
Step 2 – heat the vial for a period of time at a constant temperature
Step 3 – extract some of the vapor and analyze it by gas chromatography
Headspace Sampling
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During the equilibration step, molecules distribute themselves according to their partition coefficients
Molecules with low partition coefficients favor the vapor (headspace) phase whereas molecules with high partition coefficients favor the liquid (sample) phase
Partition coefficients are reduced as the temperature is increased
At equilibrium, the concentration in the headspace phase is proportional to the original concentration in the sample
Determining the composition of the headspace phase enables the composition of the sample to be established.
Theory
Liquid Sample
Compound
K=Cl/Cv
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Key Components in the PerkinElmer Headspace System
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Since polar compounds in water (or beer) have very high partition coefficients – often less than 0.5% of the compound in the sample may pass into the headspace.
With headspace without the trap, only a small fraction of the total headspace vapor will enter the column
The headspace trap technique can enhance detection limits by 100 times by withdrawing the entire HS volume and enabling several injections from same vial to be focused on trap
Enhanced Sensitivity with the Headspace Trap
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Sample Vial Thermal Equilibration
column
Headspace Sampler Gas Chromatograph
detector
seal
vial
oven
valve
trap
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Vial Pressurization
column
Headspace Sampler Gas Chromatograph
detector
seal
valve
vial
oven
trap
column isolation
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Trap Load
column
Headspace Sampler Gas Chromatograph
detector
seal
valve
vial
oven
trap
1616
Vial Re-Pressurization
column
Headspace Sampler Gas Chromatograph
detector
seal
valve
vial
oven
trap
1717
Trap Re-Load
column
Headspace Sampler Gas Chromatograph
detector
seal
valve
vial
oven
trap
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Dry Purge
column
Headspace Sampler Gas Chromatograph
detector
seal
valve
vial
oven
trap
1919
Trap Desorption
column
Headspace Sampler Gas Chromatograph
detector
seal
valve
vial
oven
trap
Trap desorbed in Opposite direction
2020 © 2009 PerkinElmer
Analyzing Beer
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6.50 8.50 10.50 12.50 14.50 16.50 18.50 20.50 22.50 24.50 26.50 28.50 30.50 32.50 34.50Time0
100
%
Beer Flavors HS_GC_MS Scan EI+ TIC
2.95e1016.59;70
6.0446 12.02
4311.43
61
10.1543
22.3170
16.7655
18.1743 19.26
43
30.3388
26.018822.42
43
31.54104 33.77
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Pro
pyl a
ceta
teE
thyl
pro
pano
ate
Dim
ethy
l Sul
fide
Dia
cety
l
Eth
yl A
ceta
te
Isob
utan
ol
Isop
enta
nal
Isoamyl alcohol
2-M
ethy
l-1-b
utan
ol
Isob
utyl
ace
tate
Eth
yl b
utyr
ate
Isoa
myl
ace
tate
2-M
ethy
lbut
yl a
ceta
te
Eth
yl h
exan
oate
Lina
lyl a
nthr
anila
te
Eth
yl o
ctan
oate
á-P
hene
thyl
ace
tate
Met
hyl g
eran
ate
Eth
yl c
aprin
ate
Nice peak shapesGood peak separationRequired detection limitsRepeatable responseLinear response
Beer - Component Identification by Mass Spectrometry
Sample Size: 5 mLSample Temp: 70oCSample Load: 1 cycle Trap Load Temp: 25oCDry Purge: 6 minTrap high Temp: 300oCNeedle Temp: 160oCT Line Temp: 180oCColumn Flow: Pressure Pulse: 2mL/min for 0.4minAnalytical Flow Rate 1mL/min
Mass Range: 30 to 300 amu
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Sensitivity is AMAZING by HS Trap/GC/MS
DMS at 10 ppbSignal to Noise is 56940 to 1
Diacetyl at 10 ppbSignal to Noise is 1067 to 1
2323 © 2009 PerkinElmer
Hops EvaluationBeer Comparisons
Supplier InformationBrewing Investigation
Data of Beer Analysis from Long Trail Brewery
Thank you Long Trail and Bill Yawney
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Hop Volatile Comparison
1 2 3 4 5 6 7 8 9 10 11 120
2
4
6
8
10
12
Hop Volatile Compound ComparisonAverage Amounts - Sample 1 vs Sample 2
Sample 1Sample 2
Volatile Compounds
pp
m
Co
mp
ou
nd
#
Compound Name (Longer names have been
truncated)1 1R-à-Pinene2 β-Myrcene3 β-Pinene4 à-Phellandrene5 2,6-Dimethyl-1,3,5,7-octatetra6 Limonene7 1,4-Cyclohexadiene, 1-methyl-48 Furan, 3-(4-methyl-3-pentenyl)9 Copaene
10 1,6,10-Dodecatriene, 7,11-dime11 Bicyclo[3.1.1]hept-2-ene, 2,6-12 Caryophyllene
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Beer Volatile Comparison
Co
mp
ou
nd
N
ame
Ret
enti
on
T
ime
1-Propanol 8.022-Butanone, 4-hydroxy 9.721-Propanol, 2-methyl 10.451-Butanol, 3-methyl 12.541-Butanol, 2-methyl 12.80Propanoic acid ethyl ester 13.63n-Propyl acetate 13.74Mixture of methyl butanols 14.77Mixture of methyl butanols 14.93Acetic acid, 2-methylpropyl ester 16.28Butanoic acid, ethyl ester 17.371-Butanol, 3-methyl-, acetate 20.421-Butanol, 2-methyl-, acetate 20.55Hexanoic acid, ethyl ester 24.49Acetic acid hexyl ester 24.84Heptanoic acid, ethyl ester 26.93Acetic acid, heptyl ester 27.22Phenyl ethyl alcohol 27.45Octanoic acid 28.15Octanoic acid, ethyl ester 28.95Acetic acid, 2-phenylethyl ester 30.12Ethyl9-decanoate 32.11Decanoic acid ethyl ester 32.27Caryophyllene 33.93Alpha caryophyllene 34.55Decanoic acid ethyl ester 35.86
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Research and Development Project
Name RTDimethyl sulfide 8.3231- Propanol 9.3581- Propanol 9.591Acetic acid, anhydride with formic acid 10.1311-Propanol, 2-methyl 12.104Butanol, 3-methyl 13.004Formic acid 13.987Vinyl butyrate 14.4371-Butanol, 3-methyl 14.7001-Butanol, 2-methyl-, (S) 14.962Acetaldehyde, O-methyloxime 15.487Vinyl butyrate 17.528Butanoic acid, ethyl ester 19.178Butanoic acid, 3-methyl-,ethyl ester 21.406Cyclobutanone, 2,2,3-trimethyl- 21.744Cyclopentane 21.872Propanoic acid, 2-methyl-,2-methylpropyl ester 23.642Pentyl glycolate 25.938β-Myrcene 26.133Propanoic acid, 2-methyl-,2-methylbutyl ester 26.530Limonene 27.446Methyl 2-methyl hexanoate 28.286Butanoic acid, 2-methyl-,2-methylbutyl ester 28.631Decanoic acid, ethyl ester 28.924Octanoic acid, ethtyl ester 30.251Benzenebutanal 31.4522,6-Octadienoic acid, 3,7-dimethyl-, metyl ester, (E) 32.525Decanoic acid, ethyl ester 33.717
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Supplier Comparison
2828 © 2009 PerkinElmer
PerkinElmer Fermentation Experiment
Testing the process using HS/GC/MS
American Pale Ale
12mL sample taken approximately every eight hours starting from time zero (prior to adding the yeast) during the brewing process
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Grains Maris Otter Pale Malt Munich Malt Crystal Malt
Hops Chinook Centennial Amarillo Nelson Sauvin
Yeast SafAle American Ale 05 dry yeast, no starter
O.G. 1.058
IBU 45
Process Single infusion mash at 67°C Fermentation at 19-20°C
The ‘Profile’ Beer: American Pale Ale
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Results of Components Changing with Time
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Activity of Two Components over 111 Hours of Sampling
Dimethyl Sulfide (DMS) 2,3-Butanedione (Diacetyl)
Plot: Detector Response –vs- Time
Time Interval: Every Eight Hours
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Interesting Component … 3-Methyl-1-Butanol
0 20 40 60 80 100 1200
100000000
200000000
300000000
400000000
500000000
600000000
Plot: Detector Response –vs- Time
Time Interval: Every Eight Hours
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Concentration of Four Components through Fermentation
Time DMS Diacetyl 2,3-Pentanedione trans 2-Nonenal(hours) (PPB) (PPB) (PPB) (PPB)
Zero 16.5 81 15 4.77 18.8 1577 16 3.9
15 4.6 2779 37 3.823 3.0 2183 52 3.931 3.2 1658 59 3.839 3.0 862 43 3.947 4.0 715 85 4.053 3.6 422 93 3.9
66.5 3.3 249 41 3.970.75 3.7 341 129 4.0
79 3.4 109 72 3.987 3.4 95 91 3.9
95.5 3.6 74 65 3.9103 3.3 44 49 3.9111 3.2 48 64 4.0
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Specific Gravity Measurements over 95 hours
Profile Beer Specific Gravity
Time, hrs S.G Attn %0.0 1.058 0.07.0 1.058 0.0
15.0 1.054 6.923.0 1.052 10.331.0 1.050 13.839.0 1.042 27.647.0 1.035 39.755.0 1.030 48.364.5 1.025 56.970.8 1.023 60.379.0 1.020 65.587.5 1.016 72.495.0 1.014 75.9
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Quantifies defects
Characterizes beer by Providing component identity – WHAT IS IT? Qualitative information Providing relative component ratio information Providing concentration (quantitative) information – How much?
Enables the investigation of other beers – What makes my neighbors beer so very delicious?
Optimizes process control
Conclusion
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[email protected]@perkinelmer.com