Alpha and Beta Acids in Hops by HPLC (reference: ASBC MoA. 8th edition, 1992) Method 1. Add 20 mL MeOH to 10.0 gr finely ground hops. 2. Add 100 mL diethyl ether. 3. Stopper and shake for 30 min. 4. Add 40 mL 0.1M HCl. 5. Stopper and shake for 10 min. 6. Allow to stand for 10 min to separate the phases. 7. Pipette 5.0 mL of the supernate to a 50 mL volumetric flask. 8. Bring to volume (50 mL) with methanol. 9. Filter before injecting (sample is stable 24 hours). 10. Calculate based on a known calibration standard as follows. Notes: * detector: capable of 314 nm * column: C18 (they recommend 250x4mm, 5 um ODS, RP18, Nucleosil-5) * Mobile phase: MeOH: H2O: HPO4 (85%) /85:17:0.25 (v/v) * Conditions: isocratic * Flow: 0.8 mL/min * Temp: ambiant * Sample: 10 uL * Typical Retention times: o cohumulone 16 min o n- + ad-humulone 19 min o colupulone 27 min o n- + adlupulone 34 min Calculations: 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 sample
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