高效液相色谱法同时检测啤酒中的葎草灵酮、希鲁酮和异α-酸
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摘要
为研究啤酒苦味物质组成,利用高效液相色谱法(high performance liquid chromatography,HPLC)建立了啤酒中的葎草灵酮、希鲁酮和异α-酸的检测新技术。以高甲酸颗粒酒花为原料,60℃高温空气氧化后,采用Alltima C18色谱柱分离,以体积分数为0. 05%的磷酸和乙腈为流动相梯度洗脱,结合色谱图和紫外光谱特性得到葎草灵酮和希鲁酮的参考标样;用C_(18)固相萃取柱(solid-phase extraction,SPE)进行样品前处理。该方法的相对标准偏差2. 18%～3. 59%;葎草灵酮、希鲁酮和异α-酸的最小检出限分别为0. 03、0. 03和0. 05 mg/L;回收率95%～115%。分析了市售不同品类啤酒中苦味物质组成,淡色Lager、皮尔森、小麦啤酒和黑啤酒中,葎草灵酮和希鲁酮含量均不高(<2 mg/L);而IPA啤酒中葎草灵酮、希鲁酮和异α-酸含量较高,且不同品牌苦味物质组成存在较大差异。该方法为后续开展葎草灵酮、希鲁酮的苦感研究及IPA啤酒苦味物质调控提供了有效的检测手段。
In order to study the composition of bitter substances in beer,high performance liquid chromatography was applied to quantitatively determine humuliones,hulupones,and iso-α-acid in beer. By oxidizing hop pellets at60 ℃,humuliones and hulupones as major products were separated using Alltima C18 column,and 0. 05% phosphoric acid and acetonitrile were used for mobile phase gradient elution. C18 solid-phase extraction( SPE) tubes were used for sample pretreatment. The relative standard deviations( RSDs) were 2. 18%-3. 59%. The minimum detection limits for humuliones,hulupones,and iso-α-acid were 0. 03,0. 03,and 0. 05 mg/L,respectively,and their recoveries were in a range of 95%-115%. It was found that most commercial beers,such as lager beer,pilsner,wheat beer and stout,contained less humuliones and hulupones( < 2 mg/L),however,India Pale Ale( IPA) had higher levels and their composition varied significantly among different brands. In conclusion,this method is effective for conducting further studies on bitter senses of humuliones and hulupones,and also for regulating bitter substances in IPA.
In order to study the composition of bitter substances in beer,high performance liquid chromatography was applied to quantitatively determine humuliones,hulupones,and iso-α-acid in beer. By oxidizing hop pellets at60 ℃,humuliones and hulupones as major products were separated using Alltima C18 column,and 0. 05% phosphoric acid and acetonitrile were used for mobile phase gradient elution. C18 solid-phase extraction( SPE) tubes were used for sample pretreatment. The relative standard deviations( RSDs) were 2. 18%-3. 59%. The minimum detection limits for humuliones,hulupones,and iso-α-acid were 0. 03,0. 03,and 0. 05 mg/L,respectively,and their recoveries were in a range of 95%-115%. It was found that most commercial beers,such as lager beer,pilsner,wheat beer and stout,contained less humuliones and hulupones( < 2 mg/L),however,India Pale Ale( IPA) had higher levels and their composition varied significantly among different brands. In conclusion,this method is effective for conducting further studies on bitter senses of humuliones and hulupones,and also for regulating bitter substances in IPA.
引文
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[8]杨朝霞,黄淑霞,田玉红,等.采用离子对试剂同时测定啤酒中异α-酸和还原异α-酸[J].分析实验室,2012,31(9):30-33.
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[10] TANIGUCHI Y,TANIGUCHI H,MATSUKURA Y,et al. Structural elucidation of humulone autoxidation products and analysis of their occurrence in stored hops[J]. J Nat Prod,2014,77(6):1 252-1 261.
[11] TANIGUCHI Y,YAMADA M,TANIGUCHI H,et al.Chemical characterization of beer aging products derived from hard resin components in hops(Humulus lupulus L.)[J]. Journal of Agriculture and Food Chemistry,2015,63(46):10 181-10 191.
[12] HASELEU G,INTELMANN D,HOFMANN T. Identification and RP-HPLC-ESI-MSMS quantitation of bitter-tastingβ-acid transformation products in beer[J]. Journal of Agriculture and Food Chemistry,2009,57(16):7 480-7 489.
[13] ALGAZZALI V,SHELLHAMMER T. Bitterness intensity of oxidized hop acids:Humulinones and hulupones[J].Journal of the American Society of Brewing Chemists,2016,74(1):36-43.
[14] OLADOKUN O,SMART K,COOK D. An improved HPLC method for single-run analysis of the spectrum of hop bittering compounds usually encountered in beers[J]. Journal of the Institute of Brewing,2016,122(1):11-20.
[15]林艳,单连菊,张沛,等.高效液相色谱法测定酒花、酒花颗粒及酒花浸膏中α-酸和β-酸[J].分析化学,2000,28(8):1 047.
[2] INTELMANN D,HASELEU G,DUNKEL A,et al. Comprehensive sensomics analysis of hop-derived bitter compounds during storage of beer[J]. J Agric Food Chem,2011,59(5):1 939-1 953.
[3] HASELEU G, LAGEMANN A, STEPHAN A, et al.Quantitative sensomics profiling of hop-derived bitter compounds throughout a full-scale beer manufacturing process[J]. Journal of Agriculture and Food Chemistry,2010,58(13):7 930-7 939.
[4]郝俊光,周月南,尹花,等.啤酒中源自酒花α-酸的苦味物质研究进展[J].食品与发酵工业,2017,43(12):232-240.
[5]郝俊光,王云岑,岳杰,等.源自酒花β-酸和硬脂酸的苦味物质研究进展[J].食品与发酵工业,2018,44(8):298-307.
[6]杨朝霞,李梅,武千钧.采用高效液相色谱法鉴别啤酒中异构化α-酸的种类[J].食品与发酵工业,2006,32(6):100-103.
[7]武千钧,杨朝霞,李梅,等.反相高效液相色谱法同时测定啤酒中的4种异构化α-酸[J].分析化学,2007,35(1):159.
[8]杨朝霞,黄淑霞,田玉红,等.采用离子对试剂同时测定啤酒中异α-酸和还原异α-酸[J].分析实验室,2012,31(9):30-33.
[9] YOSHIMASA T,YASUKO M,HIROMI O,et al. Identification and quantification of the oxidation products derived fromα-acids andβ-acids during storage of hops(Humulus lupulus L.)[J]. Journal of Agriculture and Food Chemistry,2013,61(12):3 121-3 130.
[10] TANIGUCHI Y,TANIGUCHI H,MATSUKURA Y,et al. Structural elucidation of humulone autoxidation products and analysis of their occurrence in stored hops[J]. J Nat Prod,2014,77(6):1 252-1 261.
[11] TANIGUCHI Y,YAMADA M,TANIGUCHI H,et al.Chemical characterization of beer aging products derived from hard resin components in hops(Humulus lupulus L.)[J]. Journal of Agriculture and Food Chemistry,2015,63(46):10 181-10 191.
[12] HASELEU G,INTELMANN D,HOFMANN T. Identification and RP-HPLC-ESI-MSMS quantitation of bitter-tastingβ-acid transformation products in beer[J]. Journal of Agriculture and Food Chemistry,2009,57(16):7 480-7 489.
[13] ALGAZZALI V,SHELLHAMMER T. Bitterness intensity of oxidized hop acids:Humulinones and hulupones[J].Journal of the American Society of Brewing Chemists,2016,74(1):36-43.
[14] OLADOKUN O,SMART K,COOK D. An improved HPLC method for single-run analysis of the spectrum of hop bittering compounds usually encountered in beers[J]. Journal of the Institute of Brewing,2016,122(1):11-20.
[15]林艳,单连菊,张沛,等.高效液相色谱法测定酒花、酒花颗粒及酒花浸膏中α-酸和β-酸[J].分析化学,2000,28(8):1 047.
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