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Hops–17
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HOP ESSENTIAL OILS BY CAPILLARY GAS
CHROMATOGRAPHY-FLAME IONIZATION DETECTION
This method identifies and measures selected
components of hop essential oil using a gas chro-
matograph equipped with a flame ionization detector.
Estimated concentrations of several components may be
determined using the optional internal standard.
Reagents
(a)
Pentane or hexane
(HPLC grade).
(b) 2-Octanol (reagent grade), a suggested internal
standard.
Apparatus
(a)
Autosampler vials,
glass nominal 1.8-mL volume.
(b)
Aluminum crimp seals
with Teflon lined septa for
autosampler vials.
(c)
Hand crimper
for autosampler vials.
(d)
Gas chromatograph
(GC) equipped with a flame
ionization detector (FID) and a data acquisition
system.
Operating Conditions for Gas Chromatograph
(a) GC column: 30 m × 0.25 mm, 25-µm film thickness
Supelcowax 10 column manufactured by Supelco,
(Bellefonte, PA) or equivalent column capable of
separating natural flavors and fragrances.
(b) Carrier gas: helium, ultra-high purity recommended.
Set head pressure to obtain a flow rate of 0.6
mL/min.
(c) Split ratio: 1:50.
(d) Injection port: 200°C.
(e) Flame ionization detector: 260°C.
(f) GC oven temperature program: 50°C, held for 1
min. Ramp from 50°C to 260°C at 3°C/min and
remain at 260°C for 15 min.
(g) Total run time: 86 min. Modifications can be made
to improve chromatography.
Sample Preparation
If no internal standard is used, prepare a 10% (v/v)
hop oil solution by adding 100 µL of hop oil and 900 µL
of the pentane or hexane in a 1.8-mL glass autosampler
vial.
If 2-octanol is used as an internal standard, prepare a
1% (v/v) 2-octanol stock solution in pentane or hexane.
Prepare a 10% (v/v) hop oil solution by adding 100 µL
of hop oil and 900
μL
of the 2-octanol stock solution in
a 1.8-mL glass autosampler vial.
Method
Condition the GC column per the manufacturer’s
recommendations prior to initial use. Adjust the FID gas
flows to the conditions recommended by the manu-
facturer; light the flame; and allow the detector to
equilibrate prior to sample injection.
Inject a 1-µL volume of the 10% hop oil solution.
Start integration after the solvent peak elutes, and
continue to integrate all essential oil components eluted
during the entire run time.
Calculation
Use the reference chromatogram shown in Fig. 1 to
aid in identifying selected components detected from the
hop oil solution injection. (Alternatively, retention times
for selected components may be determined by making
Fig. 1. Sample chromatogram, with selected components shown.
doi: 10.1094/ASBCMOA-Hops-17
Hops–17
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injections of diluted pure compounds using the same
chromatographic conditions as the 10% hop oil
analysis.) If an internal standard is not used, report the
area percent of each component identified, to the nearest
0.01%, as a portion of the total chromatographic area. If
an internal standard is used, calculate the estimated
concentration of each component based on internal
standard response.
Without an internal standard,
component integrator count
component
,
%
=
×
100
total integrator counts
With an internal standard,
wt of int std in sample
component, g/mL
=
int std integrator counts
component integrator counts
×
vol of essential oil in prepd sample
int std vol
wt of int std in prepd sample
=
total solvent vol
×
int std solution vol used
×
int std density
TABLE I
Repeatability and Reproducibility Coefficients of Variation
for Selected Hop Oil Components
a
Compound
Myrcene
2-Methyl-butyl isobutyrate
Caryophyllene
Farnesene
Humulene
Humulene:Caryophyllene
a
Repeatability %
(cv
r
)
0.6 – 3.4
1.8 – 11.2
0.8 – 2.4
1.4 – 2.8
0.6 – 3.3
0.2 – 7.4
Reproducibility %
(cv
R
)
4.2 – 12.3
8.8 – 20.4
5.3 – 10.7
4.2 – 10.2
5.1 – 14.8
2.2 – 10.4
Ranges from collaborative tests (Refs. 1–3).
Integrator count of standard = 438,520
Integrator count of component = 669,740
Component, g/mL
=
0.00737 669,740
×
438,520 0.1 mL
=
0.113 g/mL
Examples
Without an internal standard
Integrator count of component = 669,740
Total integrator counts = 6,293,580
Component =
669,740
×
100
=
10.64%
6,293,580
Notes
A sample chromatogram (Fig. 1) is shown to aid in
chromatogram peak identification.
In collaborative tests (Ref. 1,2,3), ranges of
repeatability and reproducibility coefficients of variation
for selected hop oil components were as shown in Table I.
References
1. American Society of Brewing Chemists. Report of Subcommittee
on Determination of Hop Essential Oils by Capillary Gas
Chromatography.
Journal
59:222, 2001.
2. American Society of Brewing Chemists. Report of Subcommittee
on Determination of Hop Essential Oils by Capillary Gas
Chromatography.
Journal
60:204, 2002.
3. American Society of Brewing Chemists. Report of Subcommittee
on Determination of Hop Essential Oils by Capillary Gas
Chromatography.
Journal
62:175, 2004.
4. Peacock, V. E., and McCarty, P. Varietal identification of hops and
hop pellets.
Tech. Q. Master Brew. Assoc. Am.
29:81, 1992.
2004
With an internal standard
Internal standard solution is 1 mL 2-octanol diluted
to 100 mL with hexane.
Prepared sample is 0.1-mL essential oil added to
0.9 mL internal standard solution.
Density of 2-octanol = 0.8193 g/mL
Weight of internal standard in prepared sample, g
1 mL
=
× 0.9 mL × 0.8193 g/mL
100 mL
= 0.00737 g

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