高压液相色谱法测定大豆异黄酮含量

大豆试样在酸性条件下水解后,采用岛津LC-10AtvpHPLC仪,紫外检测器(SPD-10Avvp),波长 254 nm,色谱柱:Inertsil ODS-3柱(5μm,4.6 mm×250 mm),流动相:甲醇:5％ 醋酸溶液30％～70％梯度洗脱,流速:1.0 ml/min,柱温:50℃条件下,测定水解提取物中大豆异黄酮含量。测定结果表明:不同品种、产地的大豆中异黄酮含量为 1.40 mg～3.30 mg/g,大豆胚芽中异黄酮含量最高,为13.18mg/g;其它豆类,如红豆、绿豆、青豆、豌豆等异黄酮含量为0.37mg～1.2mg/g,此测定法能为大豆的选育种、栽培及大豆异黄酮含量评价提供依据。     

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。     

Content and Profile of Isoflavones in Soy-Based Foods as a Function of the Production Process

Soy has been traditionally incorporated in diet as processed foods, such as soymilk, tofu, miso, tempeh, etc., and the consumption is commonly associated with a reduction of the development of chronic diseases due to their antioxidant, anti-inflammatory, and anti-allergic properties, among others. Many of the health benefits of soy have been attributed to isoflavones. They comprise a group of naturally occurring flavonoids consisting of heterocyclic phenols. Soy contains three types of isoflavones in four chemical forms: the aglycones daidzein, genistein, and glycitein; the β-glucosides daidzin, genistin, and glycitin; their 6″-O-malonyl-β-glucosides (6OMalGlc); and their 6″-O-acetyl-β-glucosides (6OAcGlc) conjugates. Industrial processing methods of soy-based food products commonly lead to the loss of isoflavones through removal of undesirable fractions. On the other hand, isoflavones can be transformed into different conjugates, which may have significant effects on the food texture and on the bioavailability and pharmacokinetics of the isoflavones. This article reviews the effect of a number of soybean processing treatments on the isoflavone content and profile. The preparation and manufacturing of different soy-based food and food ingredients, fermented and non-fermented, has been analyzed in terms of content and distribution of the three major isoflavone derivatives, daidzein, genistein, and glycitein, and their respective conjugates.     

Systematic evaluation of pre-HPLC sample processing methods on total and individual isoflavones in soybeans and soy products

There are mainly two protocols in isoflavone analysis, one that involves hydrolysis prior to HPLC analysis and the other direct HPLC analysis. In this study, three different hydrolysis methods were systematically re-evaluated, and compared with direct HPLC analysis. Acidic hydrolysis (1.2-3 M HCl in ethanol at 80 °C) showed a maximum conversion of ca. 92% from glucosides to aglycones in 2 h; however, longer reaction caused degradation of genistein. Alkaline hydrolysis using 2 M NaOH converted acetylglucosides and malonylglucosides to their respective glucosides within 10 min. Glucuronidase from H. pomatia effectively converted isoflavone glucosides and acetylglucosides to aglycones. Quantification of the total isoflavones in various soy food products showed no significant difference among direct injection and the three hydrolysis methods (P < 0.05). We conclude that direct analysis of isoflavones in crude extracts is a rapid and accurate method to obtain isoflavone profiles and compositions in soybean, soy foods and beverages.     

高效液相色谱电喷雾质谱联机检测黑豆异黄酮

采用C18色谱柱,以甲醇:水:甲酸30min内,0.8ml/min梯度洗脱(从20:80:0.1到80:20:0.1)黑豆70%乙醇提取液。紫外光谱和电喷雾质谱的数据表明:大豆苷、乙酰基染料木苷、染料木苷、丙二酰基大豆苷、丙二酰基黄豆苷和丙二酰基染料木苷是黑豆种子中最主要的异黄酮,总含量为0.373%,为普通大豆种子异黄酮含量的2～6倍;黑豆是异黄酮的丰富来源。     

高效液相色谱-电喷雾质谱联用法检测大豆异黄酮和皂苷

应用高效液相色谱 电喷雾质谱联用法 ,直接对大豆胚芽的 70 %乙醇室温提取液进行检测 .根据各色谱峰的质谱特征 ,在室温提取液中鉴定出 9种异黄酮 ,7种A组皂苷 ,4种B组皂苷和2种E组皂苷 .结果证明对于复杂天然产物的定性定量分析 ,高效液相色谱 电喷雾质谱 (HPLC/ESI MS)联用分析法是一种方便快捷有效的方法 .     

Microsatellite genotyping with HRM (High Resolution Melting) analysis for identification of the PGI common bean variety Plake Megalosperma Prespon

Legumes and common bean (Phaseolus vulgaris L.), in particular, are important crops worldwide, consumed either as dried seeds or fresh fruits. Correct identification of common bean varieties is important, in order to ensure food quality, safety and authenticity for consumers. Recently, DNA based methods, including molecular markers like microsatellites (SSR), have been developed for plant species or variety identification genotyping and for identification of their ingredients in the final food products. Here, we have applied High Resolution Melting (HRM) analysis coupled with four microsatellite markers to facilitate the identification of protected geographic indication (PGI) common bean variety ‘Plake Megalosperma Prespon’ (‘PMP’). The four microsatellite loci used were informative and were used to generate a unique melting curve profile of microsatellites for each variety tested. These microsatellite markers enabled the distinction and identification of the PGI (common bean variety ‘PMP’). Hence, this assay provided a flexible, cost-effective and closed-tube microsatellite genotyping method, well suited to varietal identification and authentication analysis in common beans.     

大豆豆脐中异黄酮的提取工艺优化

为了提高大豆豆脐在大豆加工行业的利用率以及更大程度地发挥大豆豆脐的营养价值,本文在国标大豆异黄酮检测方法的基础上,建立了一套适用于测定大豆豆脐中异黄酮含量的高效液相色谱方法。本实验以大豆豆脐为原料,采用乙醇-水溶液作为提取溶剂,通过单因素实验和正交实验确定超声波辅助提取大豆异黄酮的最佳工艺,结果表明:各因素对大豆异黄酮提取率影响大小的顺序为:提取温度(B)>提取时间(C)>料液比(D)>乙醇浓度(A);在乙醇浓度为80%、提取温度为80℃、提取时间为1.5h、料液比为1:35 g/mL时提取三次,大豆异黄酮的提取率可达10.88±0.120 mg/g。本方法准确、高效,能够提取出原料中90%以上的大豆异黄酮,该提取工艺稳定可行,可为大豆豆脐中异黄酮的提取提供理论依据。     

Evaluation of Different Sample Preparation Procedures Using Chemometrics: Comparison Among Photo-Fenton Reaction,Microwave Irradiation,and Direct Determination of Minerals in Fruit Juices

In this work, an exploratory study about the mineral characteristic of some fruit juice (passion fruit, pineapple, orange, and grape) applying chemometric tools was made. The juices analyzed were separated in different groups by principal components analysis and hierarchical components analysis in accordance with their minerals contents. Barium, C, Ca, K, Mg, Mn, Na, and Sr were determined by inductively coupled plasma optical emission spectrometry. Addition and recovery methods were used for confirmation of the results. The recovery range for microwave sample preparation was from 81% (for K) to 111% (for Mn), for photo-Fenton from 102% (for Ca e Mg) to 128% (for Na), and the direct analysis, without sample preparation step, from 96% (for K) to 139% (for Mn). A new method based on photo-Fenton reaction coupled with microwave radiation for juice decomposition was proposed; the obtained results were satisfactory and compatible with the conventional methods of analysis in accordance with Student’s statistic t test and chemometric tools.     

黑龙江省栽培大豆异黄酮含量的初步分析

大豆中含有大豆异黄酮，其主要活性成份为染料木素和大豆甙元。大豆异黄酮是引起大豆苦涩味的主要因子，同时也具有一定的生理活性。利用HPLC对黑龙江省46个主栽大豆品种（系）和6个特用豆品种的大豆异黄酮含量进行分析，筛选出3个高异黄酮含量的主栽品种和2个特用豆品系。并初步分析了遗传差异是影响大豆异黄酮含量的主要因素，同时栽培环境对大豆异黄酮含量也有影响。     

Identification and determination of isoflavones in germinated black soybean sprouts by UHPLC−Q-TOF-MS mass spectrometry and HPLC-DAD

Ultra-performance liquid chromatography coupled with electrospray ionization tandem quadrupole time-of-flight mass spectrometry (UHPLC?Q-TOF-MS) and HPLC-DAD with an ultrasonic extraction method was developed for qualitative and quantitative compositions and contents in black soybean sprouts. Operational conditions of ultrasonic extraction were optimized by single-factor experiment. Phytochemical compositions of black soybean sprouts were identified by UHPLC?Q-TOF-MS in positive ion mode. A total of 23 compounds were tentatively characterized and identified by means of accurate mass and characteristic fragment ions. Among of them, the six isoflavones in different germinated black soybean sprouts were further quantified by HPLC-DAD. The results indicated that the calibration curves showed good linearity (r² > 0.9994). The intra-day and inter-day precision variations were less than 1.65% and 1.73%, respectively. The recoveries were in the range of 94.95–106.45%. The results indicated that the maximum amounts of isoflavones were produced on the fourth to fifth day germinated black soybean sprouts.     

Comparative evaluation of flavonoid content in assessing quality of wild and cultivated vegetables for human consumption

BACKGROUND: Owing to the demand for ‘health foods’, commercial development of wild vegetables may find a sizable market niche if adequate agricultural methods are used to domesticate such species. Available techniques of cultivation (even in traditional farming) may provide many advantages, such as enhancement of the content of active principles in plants and improvement in the quality of the raw material to be processed on an industrial scale. In this context, the flavonoid composition and content of roots and leaves from five varieties of wild herbs (Cichorium intybus, Portulaca oleracea, Tragopogon porrifolius, Urtica dioica and Valerianella eriocarpa) and their cultivated relatives in central Italy were compared. The aims of the study were (i) to reveal the metabolic profile of particular bioactive metabolites present in some unusual food species by using a simple method of analysis, (ii) to quantify and compare the amount of polyphenolic metabolites in wild and cultivated plants and (iii) to evaluate the effect of growing conditions on polyphenolic variability. Data were subjected to statistical analysis. RESULTS: Each herb possessed a specific flavonoid fingerprint as indicated by high‐performance liquid chromatography (HPLC) data. The total phenolic content in cultivated leaves was greater than that in wild leaves for all herbs examined (P < 0.05), with the exception of P. oleracea. CONCLUSION: The results show that the polyphenolic content of the majority of the cultivated herbs (edible parts) is no lower than that of their wild relatives, thus indicating that these species are suitable for cultivation. Copyright © 2007 Society of Chemical Industry     

发酵处理对豆豉抗氧化活性影响的研究

发酵处理基本上不改变豆豉中异黄酮的总含量,而糖苷型大豆异黄酮在β-葡萄糖苷酶的作用下部分转化为游离型大豆异黄酮,使游离型大豆异黄酮的含量明显提高;在样品未添加任何抗氧化剂的前提下,通过试验发现豆豉的自身抗氧化性比原料熟黑豆强许多。     

Analysis of phenolic compounds and isoflavones in soybean seeds (Glycine max (L.) Merill) and sprouts grown under different conditions

Soybeans (Glycine max (L.) Merill) are popularly known as a healthy food in many Asian countries and are mostly consumed as soymilk, tofu, and fermented products such as miso, temph, and sufu. The objective of this study was to determine the variation and composition of phenolic compounds and isoflavone contents in soybean seeds [Glycine max (L.) Merill] and sprouts [Kongnamul] grown under dark conditions (producing yellow soybean sprouts) and in green and yellow boxes (producing green soybean sprouts). In seven soybean cultivars, the total phenolic content ranged from 6.67 μg⁻¹ in Pureunkong to 72.33 μg⁻¹ in Poongsannamulkong. The average total phenolic content in the green soybean sprouts (48.33 μg⁻¹) was higher than in the yellow soybean sprouts (29.75 μg⁻¹). The total phenolic content in the yellow soybean sprouts varied from 9.88 μg⁻¹ to 47.71 μg⁻¹, and the total phenolic content in the green soybean sprouts varied from 29.21 μg⁻¹ to 79.70 μg⁻¹. Only four phenolic compounds, p-hydroxybenzoic acid, salicylic acid, p-coumaric acid, and ferulic acid, were detected in all soybean cultivars. Syringic acid was not detected in yellow soybean sprouts, and myricetin was only detected in yellow soybean sprouts (4.65 μg⁻¹) from the Pureunkong cultivar grown under dark conditions. The total isoflavone content in soybean seeds ranged from 2.1 μg⁻¹ in Sowonkong to 33.0 μg⁻¹ in Pureunkong, and the mean total isoflavones was 10.61 μg⁻¹. Green soybean sprouts had higher average total isoflavones (1389.4 μg⁻¹) than yellow soybean sprouts (559.2 μg⁻¹), and the total isoflavone content was highest in the Pureunkong yellow soybean sprouts (756.3 μg⁻¹) and the Sowonkong green soybean sprouts (2791.6 μg⁻¹). In soybean sprouts, the higher the (malonyl)-daidzin or (malonyl)-genistein content, the higher the total isoflavone level. Our study suggests that producing soybean sprouts enriched in isoflavones under coloured-light sources is feasible.     

Chromatographic fingerprints and quantitative analysis of isoflavones in Tofu-type soybeans

Tofu-type soybeans can differ from conventional soy varieties in seed size, seed composition, flavour and nutrition. Using HPLC coupled with ESI-MS and PDA detection, a total of 19 isoflavones were detected and identified from Tofu-type soybean seeds, more than previously reported forms from many other soybeans. An HPLC/UV fingerprint study was performed for qualitative evaluation, which enabled the isoflavone profile of Tofu-type soybeans to be characterised and differentiated from other vegetative soybeans by similarity comparison. To meet the frequent quantitative application for isoflavone contents, a simple, precise and reliable method using HCl hydrolysis during sample extraction and LC/UV for the detection was developed and validated to quantitate total isoflavones in soybeans, and then applied to determine the total isoflavone contents of different Tofu-type soy varieties grown in different field locations over two growing seasons. A range of total isoflavone contents for Tofu-type soybeans was established to confirm the use of this analytical approach for quality control applications.     

HPLC法测定大豆提取物中大豆异黄酮含量的研究

主要建立了HPLC法测定大豆提取物中大豆异黄酮含量的方法。通过实验确定了HPLC法测定大豆提取物中大豆异黄酮的最佳条件是：采用Waters Symmetry5μmC18,250mm×4．6mm色谱柱;流速：1．0mL／min;波长260nm;柱温：35℃;流动相A为含0．1％（v/v）乙酸的乙腈溶液,流动相B为含0．1％（v／v）乙酸的超纯水溶液,流动相梯度：18％-35％N乙腈溶液。该方法灵敏度高、重现性好,回收率达98％,变异系数小于3％,为研究大豆异黄酮其它产品打下了良好的基础,操作方法简单,易于掌握。     

大豆中异黄酮含量的测定及其近红外分析

以100份中国核心大豆种质资源为材料,建立高效液相色谱法(HPLC)测定大豆中异黄酮5组分和总异黄酮。扫描大豆的近红外光谱,以傅里叶近红外光谱法(FT-NIRS)与HPLC技术相结合,采用偏最小二乘(PLS)回归和交叉验证法,探讨利用FT-NIRS技术预测异黄酮含量的可行性。总异黄酮近红外预测模型的内部交叉验证其校正决定系数(R2)和交叉检验均方误差(RMSECV)分别为0.8763和0.515,外部验证其校正决定系数(R2)和预测均方误差(RMSEP)则分别是0.9492和0.599。结果表明,利用FT-NIRS预测大豆中总异黄酮含量是可行的,但是各异黄酮组分的近红外模型不能达到准确预测要求。大豆异黄酮近红外模型的建立对今后大豆的异黄酮选育工作可以提供帮助。     

超声辅助离子液体提取黑豆异黄酮及其抑菌活性研究

以离子液体为提取剂,采用超声波辅助离子液体提取黑豆异黄酮,考察1-丁基-3-甲基咪唑氯盐的浓度、料液比、超声时间以及提取温度对黑豆异黄酮提取率的影响。采用响应面法对提取工艺进行优化,同时对异黄酮的抗菌活性进行研究。结果表明,黑豆异黄酮的最佳提取条件为:离子液体浓度0.65 mol/L,提取时间49 min,料液比1∶21(g/mL),提取温度60℃。此条件下异黄酮的提取率为0.688%,预测值和实际测定值接近,为黑豆中天然异黄酮的开发利用提供科学数据。