中国传统豆制品中异黄酮的超高效液相色谱紫外检测器快速定量法

建立了超高效液相色谱-紫外检测器(UPLC-UV)测定豆制品中大豆苷、黄豆黄苷、染料木苷、大豆苷元、黄豆黄素、染料木素的检测方法。采用酸化提取条件,将豆制品中大豆异黄酮及其修饰物水解成3种葡萄糖苷和3种苷元。采用BEH C18(2.1 mm×50 mm,1.7 μm)色谱柱,以0.1%甲酸和乙腈为流动相进行梯度洗脱,梯度洗脱程序为:0~10 min,10%~45%乙腈;10~12 min,45%~100%乙腈。流速为0.3 mL/min,柱温45℃,在波长260 nm处进行紫外检测。6种大豆异黄酮组分在7.5 min内达到完全分离。建立了外标校正标准曲线(R2 ≥ 0.9998),检出限在0.05~0.1 mg·L-1之间,加标回收率在96.8%~102.0%之间。利用本方法对腐竹、豆干、素百叶、素鸡、嫩豆腐、老豆腐和内酯豆腐等7种传统豆制品中大豆异黄酮进行了定性定量分析,总大豆异黄酮含量在8.67~25.83 g/kg间。不同豆制品间6种大豆异黄酮中以大豆苷和染料木苷为主要成分,占88.0%~93.4%。结果表明,本研究建立的UPLC法可有效降低杂质干扰,色谱基线稳定且峰型良好,在10 min内实现对6种大豆异黄酮的快速定量检测,可良好应用于常见市售豆制品的营养评估与质量控制。     

超高效液相色谱法检测保健品中的大豆异黄酮各组分含量

目的建立超高效液相色谱法检测保健品中大豆异黄酮各组分含量的方法。方法利用超高效液相色谱仪,将供试品用80%甲醇溶解提取,用Waters XSELECT HSS T3色谱柱分离,以乙腈和1%磷酸溶液进行梯度洗脱,流速为0.5 mL/min,用紫外检测器在波长260 nm对大豆异黄酮组分进行检测。结果大豆异黄酮各组分的在各自范围内线性关系良好。大豆苷、大豆黄苷、染料木苷、大豆素、大豆黄素、染料木素的相关系数均大于0.9999,平均回收率为97.2%~103.3%,相对标准偏差为0.4%~4.8%。结论该方法运行时间短、结果的重复性好。超高效液相色谱法更为快捷简单,溶剂使用量更少,能够提高检验准确性,降低检验成本。     

HPLC-ESI-MSn法鉴定大豆中12种大豆异黄酮的结构

为了鉴定大豆异黄酮的结构,采用高效液相色谱-电喷雾离子源-离子阱多级质谱联用仪(HPLC-ESI-MSn)法分离鉴定大豆粉乙醇提取物,通过多级质谱提供的准分子离子峰和多级碎片离子信息,分析得到了12种大豆异黄酮的相对分子质量、保留时间并推断了它们异黄酮糖苷的组成结构、异黄酮糖苷中糖的类型等,这12种大豆异黄酮分别是大豆苷元、大豆黄素、染料木素、大豆苷、大豆黄苷、染料木苷、乙酰基大豆苷、乙酰基大豆黄苷、乙酰基染料木苷、丙二酰基大豆苷、丙二酰基大豆黄苷、丙二酰基染料木苷。     

HPLC法测定绿豆芽中四种大豆异黄酮的含量

建立同时测定绿豆芽中4种大豆异酮(大豆苷、染料木苷、大豆苷元和染料木素)含量的高效液相色谱方法,对从绿豆芽中提取4种大豆异黄酮的工艺进行优化,最佳工艺为:用60%的乙醇在55℃条件下超声提取60 min,料液比为1:12(g/mL),同时探究绿豆发芽不同天数4种大豆异黄酮的含量.采用日本岛津LC-20A高效液相色谱仪进行测定的色谱条件为:色谱柱为 phenomenex C18(150 mm×4.6 mm,5.0 μm),采用乙腈-0.2%甲酸为流动相进行梯度洗脱,流速为0.8 mL/min,检测波长为260 nm.大豆苷,染料木苷,大豆苷元,染料木素的线性范围分别是0.74 μg/mL～100.00 μg/mL,0.74 μg/mL～200.00 μg/mL,0.167 μg/mL～500.00 μg/mL,0.198 μg/mL～160.00 μg/mL(r>0.9996),线性关系良好;加样回收率(n=9)分别为100.02%,98.95%,99.28%,99.63%.     

霉菌型豆豉和纳豆中异黄酮含量的比较研究

目的:比较纳豆和豆豉中异黄酮含量的组成变化。方法:利用超声波提取,采用高效液相色谱法测定,色谱柱填料为Hypersil ODS2(4.6 mm×250 mm,5μm),流动相为甲醇(A)-水(B)进行梯度洗脱,检测波长为255 nm。结果:豆豉与纳豆相比较,豆豉中大豆苷、黄豆黄苷、染料木苷、大豆甙元、黄豆黄素、染料木素的含量分别为11,未检出,25,320,65,380μg/g;纳豆中中大豆苷、黄豆黄苷、染料木苷、大豆甙元、黄豆黄素、染料木素的含量分别为330,73,410,12,3.3,28μg/g。结论:豆豉中甙元的含量高,纳豆中糖苷的含量高。     

固相萃取-反相高效液相色谱测定食品中大豆异黄酮含量的方法研究

该研究建立了一种固相萃取-反相高效液相色谱同时测定大豆和大豆制品中黄豆黄素、黄豆黄苷、大豆苷、大豆苷元、染料木 素、染料木苷、鸢尾黄酮苷和鹰嘴豆芽素A共8种异黄酮含量的方法。 采用体积分数为90%甲醇水溶液提取异黄酮,提取液用C18固相 萃取柱净化,采用C18色谱柱分离,柱温40 ℃,以乙腈和磷酸水溶液为流动相,流速1.0 mL/min,紫外检测器254 nm进行检测,其重复测 定结果的相对标准偏差（RSD）均＜5%,加标回收率为79.3%～102.5%。所建立的反相高效液相色谱法是一种高灵敏度、高准确度的 测定方法,适用范围广,对食品中大豆异黄酮的质量控制和合理有效利用提供了参考依据,具有一定的理论意义和应用价值。     

蒸汽爆破对豆渣中大豆异黄酮的影响研究

采用高效液相色谱法研究蒸汽爆破处理对豆渣中大豆异黄酮组成和含量的影响。结果表明,豆渣经蒸汽爆破处理后,6种大豆异黄酮含量均显著增高,大豆苷、大豆苷元、黄豆黄苷、黄豆黄素、染料木苷、染料木素分别由汽爆前的3.69、11.22、0.84、2.76、2.13、27.45μg/g增至38.04、74.96、15.75、23.36、20.36、66.28μg/g。黄豆黄苷增幅最大,较汽爆前增高了17.75倍;染料木素增幅最小,较汽爆前增高了1.41倍。而且,大豆异黄酮的增幅随汽爆强度增加呈上升趋势,在汽爆压强2.0 MPa、维压时间30 s时达到最大值(黄豆黄苷为2.0 MPa、60 s)。研究表明,采用适宜的汽爆强度处理豆渣,能够显著提高大豆异黄酮含量,这为豆渣的开发利用提供了有益参考。     

超高效液相色谱-串联质谱法测定大豆中异黄酮素含量

目的 以染料木黄酮、大豆苷元和黄豆黄素的含量为指标,建立超高效液相色谱-串联质谱测定大豆中异黄酮素总量的方法。方法 样品用乙醇-水(3+1,V/V)提取,经过β-葡萄糖苷酶水解后,用Acquity UPLC?BEH C18柱(2.1 mm×100 mm, 1.7μm)分离,以甲醇和0.1%甲酸溶液(含5 mmol/L乙酸铵)作为流动相进行梯度洗脱,电喷雾负离子模式(ESI-)电离,多反应监测模式进行检测。结果 染料木黄酮、大豆苷元在5.0～500μg/L范围内线性关系良好(r>0.995),方法的检出限分别为2.7、4.0 mg/kg,在300.0、600.0、1 200 mg/kg添加水平的平均回收率为89.4%～102.2%,相对标准偏差(n=6)为1.5%～4.6%;黄豆黄素在0.5～50.0μg/L范围内线性关系良好(r>0.995),方法的检出限为0.6 mg/kg,在30.0、60.0、120.0 mg/kg添加水平的平均回收率为85.7%～104.0%,相对标准偏差(n=6)为1.3%～2.8%。结论 该方法简单、灵敏、准确可靠,适用于大豆中异黄酮素含量的测定...     

高效液相色谱法同时测定粮食中6种大豆异黄酮

目的建立高效液相色谱法(high performance liquid chromatography,HPLC)测定粮食中6种大豆异黄酮(大豆苷、大豆黄苷、染料木苷、大豆素、大豆黄素和染料木素)含量的分析方法。方法准确称取一定量粉碎混匀后的样品,经80%甲醇提取后取上清液,过0.22μm有机相滤膜上机。采用Thermo Syncronis C18柱(250 mm×4.6 mm,5μm),以0.5%甲酸水溶液(A)、乙腈(B)和甲醇(C)作为流动相进行梯度洗脱,流速0.8m L/min,柱温为35℃,于紫外检测器波长260 nm处检测,大豆异黄酮各组分含量以外标法进行定量。结果本方法在30 min内完成6种大豆异黄酮的分离分析,大豆异黄酮各组分浓度在0.2~50μg/mL范围内呈良好的线性关系(r0.999),平均加标回收率为96.9%~107.8%,相对标准偏差(relative standard deviation,RSD)为0.6%~5.0%,检出限为0.03~0.1μg/mL,定量限为0.1~0.3μg/mL。结论建立的方法具有较高的灵敏度和重复性,能满足不同粮食种类中大豆异黄酮含量测定。     

一种保健酒中5种大豆异黄酮及芝麻素的超高效液相色谱检测法

建立了一种保健酒中大豆异黄酮及芝麻素的超高效液相色谱检测法,以pH为3的磷酸水溶液和乙腈为流动相,检测器为PDA检测器,检测波长为287nm,本实验建立的色谱条件能使6种组分(大豆苷、大豆黄苷、染料木苷、大豆素、染木素、芝麻素)在10min内得到很好分离,该方法加标回收率及精密度较好,适用于保健酒中大豆异黄酮及芝麻素的分析检测。     

UPLC法测定大豆制品及相关制剂中大豆异黄酮含量

目的：建立同时测定大豆制品及相关制剂中4种大豆异黄酮类化合物大豆苷、染料木苷、大豆苷元、染料木素含量的超高效液相色谱法。方法：色谱柱：Acquity UPLC BEH C18柱(50mm×2.1mm,1.7μm),流动相：A为体积分数0.2%甲酸溶液,B为乙腈,梯度洗脱;流速：0.4mL/min;检测波长：254nm,柱温：30℃。结果：线性范围：大豆苷0.625～40.0mg/L(r=1.000),染料木苷0.625～40.0mg/L(r=0.9999),大豆苷元0.04～2.56mg/L(r=0.9997),染料木素0.04～2.56mg/L(r=0.9996)。大豆和Natrol? soy isoflavones中大豆苷、染料木苷、大豆苷元、染料木素平均回收率为98.5%～99.8%。结论：本方法简便、快速、灵敏、准确,适用于大豆制品及相关制剂中大豆异黄酮含量测定。     

Original article: Thermal dynamic properties of isoflavones during dry heating

Thermal stabilities of major soya isoflavones at different dry‐heating temperatures were determined in this study. The conversion of glucoside isoflavones to aglycone isoflavones was monitored as well. The thermal degradation of the isoflavones: glucoside form daidzin, glycitin, and genistin and aglycone form daidzein, glycitein, and genistein followed first‐order reaction kinetics at heating temperatures 100, 150, and 200 °C. The degradation rate constants of the six isoflavones were not significantly different at 100 °C. However, the constants increased with increasing heating temperature. The half‐life for the glucoside and aglycone isoflavones was from 144 to 169 min and 139 to 176 min at 100 °C, respectively. They decreased rapidly to 15.7–54.7 and 36.0–90.7 min when temperature increased to 150 °C. When heated at 200 °C, they further decreased to 5.8–6.0 and 15.7–21.2 min, respectively. The order of thermal stability from lowest to highest was glycitin < genistin < daidzin < glycitein < genistein < daidzein at temperature below 150 °C. However, their thermal stabilities were not different at 200 °C. The conjugated glucosides were cleaved from the isoflavones to produce their corresponding aglycones when heated at 150 °C or higher. The production of glycitein increased constantly and was the highest among the three aglycone isoflavones.     

Bioavailability of soy isoflavones in rats Part I: application of accurate methodology for studying the effects of gender and source of isoflavones

There are limited and controversial reports about the effects of gender and source of isoflavones on their bioavailability. Moreover, several previous studies have not used appropriate methodology to determine the bioavailability of soy isoflavones, which requires comparing the area under the plasma concentration-time curve after both oral and intravenous injection (IV) administration. Therefore, the present study was conducted to determine the bioavailability of isoflavones from different sources following both oral and IV administration in male and female rats. Three sources of isoflavones; Novasoy (a commercial supplement), a mixture of synthetic aglycones (daidzein, genistein and glycitein) and a mixture of synthetic glucosides (daidzin, genistin and glycitin) were tested. Following administration, blood samples were collected at several time points (0, 10, 30 min and 1, 2, 8, 24, 48 h post oral gavage and 0, 10, 30, 45 min and 1, 2, 3, 4, 8 h post-IV dosing) and plasma isoflavones were measured by LC/MS. Bioavailability values for daidzein, genistein and glycitein were significantly (p <0.05) higher (up to sevenfold) in Novasoy and the glucoside forms of isoflavones compared with those of the aglycone forms. Moreover, significant (p <0.05) gender differences in the bioavailability of 7-hydroxyl-3-(4'-hydroxyphenyl)-chroman (a metabolite of daidzein), glycitein and daidzein were observed for Novasoy, with higher values in male rats. In summary, the source of isoflavones and the sex of rats had significant effects on isoflavone bioavailability.     

Effects of extraction temperature,ionic strength and contact time on efficiency of bis(2‐ethylhexyl) sodium sulfosuccinate (AOT) reverse micellar backward extraction of soy protein and isoflavones from soy flour

BACKGROUND: Soy protein enriched with isoflavones has been linked to various disease‐preventing and health‐promoting activities owing to the antihypertensive, hypocholesterolaemic, antiobesity and antioxidative properties of isoflavones. The isoflavone profiles of soy‐based products are known to be highly dependent on the various chemical and physical treatments to which the products have been subjected. The aim of this research was to increase the efficiency of backward extraction of soy protein and isoflavones from bis(2‐ethylhexyl) sodium sulfosuccinate (AOT) reverse micelles by studying the effects of extraction temperature, ionic strength of the aqueous stripping solution and contact time on the amounts of soy protein and isoflavones backward extracted from an AOT/H₂O/isooctane reverse micellar system. RESULTS: By modifying the extraction temperature, ionic strength and contact time, 47.0–60.2% of protein, 43.3–68.4% of daidzin, 43.8–74.6% of genistin, 39.0–88.8% of glycitin, 20.8–92.6% of malonyl genistin, 20.2–52.0% of malonyl glycitin, 32.7–75.6% of acetyl genistin, 49.7–76.8% of daidzein and 19.6–38.1% of genistein present in the AOT reverse micellar solution were backward extracted into the aqueous stripping phase. Statistical analysis showed that there were significant linear and interactive effects of temperature and contact time on the backward extraction of daidzin, genistin, glycitin and daidzein. Significant linear and interactive effects of ionic strength and contact time were found in the backward extraction of daidzin and genistin. The backward extraction of genistein was only influenced by contact time and its interaction with temperature. CONCLUSION: This study showed the potential of reverse micelles as a protocol for extracting isoflavones from soy samples for analytical purposes. By modifying the extraction temperature, contact time and ionic strength, soy protein enriched with daidzin, genistin, daidzein and genistein could be produced from soy flour. The results represent an important contribution to current knowledge on utilising reverse micellar extraction in food technology. Copyright © 2007 Society of Chemical Industry     

不同品种大豆异黄酮组分及体外抗氧化活性分析

为研究不同品种大豆异黄酮组分和含量以及抗氧化活性,测定了50个特色大豆品种的百粒重、色泽（L*、a*、b*）,通过高效液相色谱法（HPLC）测定异黄酮组分,评估不同品种大豆甲醇提取物的DPPH、ABTS自由基清除能力,并用相关性分析、主成分分析及聚类分析法对样品进行分析。结果表明:大豆的百粒重范围为7.05~47.46 g。不同种皮色大豆L*、a*和b*呈现显著差异（P<0.05）。HPLC分析结果发现糖苷型异黄酮含量是大豆中主要的异黄酮组分,占总异黄酮含量的90%以上。其中染料木苷含量最高。安豆115品种的大豆异黄酮的总含量最高,为2500.78 μg/g;靖江丝瓜青品种的大豆异黄酮含量最低,为888.86 μg/g。安豆115品种的DPPH和ABTS自由基清除能力较强,分别为（16.11±0.25）和（8.12±0.04）μmol V_C/g。糖苷型异黄酮含量与DPPH和ABTS自由基清除能力有较高的线性相关性（R2分别为0.903和0.867）。主成分分析表明6项指标可用2个主成分来表示（累计贡献率达66.3%）。聚类分析将50个品种的大豆分为4类:第一类中黄豆黄素和黄豆黄苷含量较高;第二类中大豆苷、大豆苷元、染料木苷和染料木素含量较高;第三类和第四类中其大豆异黄酮含量基本处于中等水平和较低含量。综上所述,不同大豆品种的异黄酮组成、含量和抗氧化能力存在较大差异,可为其进一步综合开发利用提供一定理论基础。     

Daily intake of isoflavones based on the market basket method

Daily intake of isoflavones (daidzin, glycitin, genistin, daidzein, glycitein, and genistein) was determined quantitatively, based on the market basket method. Acid hydrolysis during extraction of foods was chosen to convert phytoestrogenes into the respective aglycons, facilitating HPLC analysis and allowing quantitation of total isoflavones as aglycones including both originally present glycosides and "free" aglycones. The isoflavones were extracted from samples with methanol and determined by reversed-phase HPLC analysis using a linear gradient of methanol-water as the eluent. From the results of hydrolysis, the daily intake of total isoflavon was 38.1 mg/adult Japanese. The values obtained by the market basket method and the National Nutrition Survey method were similar.     

腐乳发酵过程中大豆异黄酮变化的研究

以东北产大豆为原料,利用雅致放射毛霉(AS3.2278)菌株,研究了腐乳发酵过程中异黄酮总量和成分的变化。高效液相色谱检测结果表明:腐乳发酵过程中异黄酮糖甙含量降低,异黄酮甙原含量升高,后酵50 d 腐乳中总甙原含量约为白坯中的20倍。在6%食盐的条件下,发酵过程中染料木酮甙酶解速度高过黄豆甙,在后酵30d染料木酮和黄豆甙原含量分别达到局部峰值191.24μg/g(干物质)和106.65μg/g(干物质)。     

不同大豆种质种皮黄酮类色素的差异分析

为探究大豆种皮中黄酮类色素的含量和分布规律,选取167份大豆种质资源为实验材料,利用高效液相色谱(HPLC)法对大豆种皮黄酮类色素含量进行测定。结果表明：大豆种皮花色苷组分中矢车菊素-3-O-葡萄糖苷含量最多,异黄酮组分中大豆苷含量最高。花色苷组分在野生和半野生大豆种皮中高于栽培大豆。异黄酮组分中染料木苷在野生大豆中最高,黄豆黄苷在半野生大豆中最高,其他组分在栽培大豆中最高。栽培大豆黑色种皮花色苷组分、染料木苷和大豆苷元含量最高,青色种皮大豆苷和黄豆黄苷含量最高,双色种皮黄豆黄素含量最高。相关分析表明3类结合型糖苷内部、3类游离型苷元内部、3种花色苷组分内部两两相关极显著。大豆苷与游离型苷元、矢车菊素-3-O-葡萄糖苷相关显著。聚类分析将大豆材料划分为三大类群,第一类群除黄豆黄苷和飞燕草素-3-O-葡萄糖苷外,其他色素组分含量均最高,为黄酮类色素的研究和利用提供参考。