Hop bitterness analysis
ref : ASBC methods of Analysis, 8th Edition, 1992.
Transcribed by Dan McConnell (email@example.com)
Hop bitterness in beer (ASBC Method)
- Transfer 10.0 mL beer to a 50 mL centrifuge tube.
- add 50 uL octyl alcohol, 20 mL isooctane (HPLC grade) and 1 mL 3M HCl .
- shake vigorously for 15 minutes.
- centrifuge to separate the phases.
- read organic phase at 275 nm (1 cm cell) vs blank (20 mL isooctane, 50 uL
BU= Abs @275*50
Example: Abs =0.622 -- 0.622*50= 31.1 BU
- isooctane should have an Abs@275 <0.005.
- readings should be done ASAP due to decomposiotin by UV light
Alpha and Beta acids in hops: ASBC MoA Method
- place 5.000 +- .001 gr hops in an extraction bottle and add 100 mL toluene
- shake for 30 min with vigorous agitation
- let stand until clear or certrifuge (preferred)
- Dilution A:
dilute 5.0 ml of this extract to 100 mL with methanol.
- Dilution B:
dilute an aliquot of the dilution A with alkaline methanol (0.2 mL 6M NaOH per
100 mL MeOH) so that the Abs at 325 and 355 falls within the most accurate
range of the instrument.
- Immediatly read diluion B (1 cm) at 275, 325 and 355 vs a toluene
blank that was prepared and diluted in EXACTLY the same manner.
dilution factor, d= (volume dil A x volume dil B)/ (500 x aliq extract A x
aliq dil A)
% alpha acids= d x (-51.56 A355+ 73.79 A325-19.07 A275)
% beta acids= d x (55.57 A355-47.59 A325 + 5.10 A275)
- 5 gr hops extracted with 100 mL toluene
- 5 mL clear extract diluted to 100 mL with methanol=Dilution A
- 3 mL Dilution A diluted to 50 mL with alkaline methanol
- A325= 0.596
- d = (100 x 50) / (500 x 5 x 3) = 0.667
- alpha = 0.667 x [ -(51.56 x 0.615) + (73.79 x 0.596) - (19.07 x 0.132) = 6.5
- beta = 0.667 x [ (55.57 x 0.615) - (47.59 x 0.596) + (5.10 x 0.132) = 4.3
Alpha and Beta acids in hops by HPLC: ASBC MoA Method
- capable of 314 nm
- C18 (they recommend 250x4mm, 5 um ODS, RP18, Nucleosil-5)
- Mobile phase:
- MeOH: H2O: HPO4 (85%) /85:17:0.25 (v/v)
- 0.8 mL/min
- 10 uL
add 20 mL MeOH to 10.0 gr finely ground hops
add 100 mL diethyl ether- stopper and shake for 30 min.
add 40 mL 0.1M HCl
stopper and shake for 10 min.
allow to stand for 10 min to separate the phases
pipette 5.0 mL of the supernate to a 50 mL volumetric flask
bring to volume (50 mL) with methanol
filter before injecting (sample is stable 24 hours)
calculate based on a known calibration standard as follows
Response Factor, RF= [mass of calib extr (gr) x conc of component in calib
extr (%)] / area.
Component %= (2 x average sample peak area of component x RF) / mass of
Typical Retention times:
- 16 min
- n- + ad-humulone
- 19 min
- 27 min
- n- + adlupulone
- 34 min
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Spencer W. Thomas (firstname.lastname@example.org)