代表性大豆种质异黄酮主要组分含量鉴定

采用HPLC检测方法,对100份不同来源的代表性大豆种质进行异黄酮含量分析,结果显示大豆种质中主要含有6种异黄酮组分,分别为黄豆苷（D）、黄豆黄苷（GL）、染料木苷（G）、丙二酰基黄豆苷（MD）、丙二酰基黄豆黄苷（MGL）和丙二酰基染料木苷（MG）,且异黄酮含量在不同生态区间和品种间均存在显著差异,变异丰富;南方产区大豆品种的异黄酮总含量最高（2465.48ug/g）,黄淮大豆产区次之（2308.48ug/g）,北方大豆产区最低（1705.89ug/g）;6 种主要异黄酮组分含量的变异系数变化范围为33.44%～52.03%。相关分析表明异黄酮总含量与各主要组分含量之间均呈极显著正相关,与脂肪含量呈极显著负相关（r = -0.323**）。在此基础上,筛选出高异黄酮含量的大豆种质2份,分别为平顶黑豆（4459.91 ug/g）和PI-567479（4073.95 ug/g）,低异黄酮含量的大豆种质2份,分别为茶色豆（857.74 ug/g）和牡丰1号（922.82 ug/g）,可以用于大豆异黄酮育种或遗传研究。     

黑龙江省野生和栽培大豆异黄酮与其组分相关性分析

利用高效液相色谱法（HPLC）检测了黑龙江省556份不同生态区以及不同类型大豆的异黄酮、大豆苷和染料木苷含量,其中野生大豆243份,栽培大豆313份。结果表明：野生大豆异黄酮含量高于栽培大豆,同时筛选出高异黄酮含量种质3份,低异黄酮含量种质2份。异黄酮、大豆苷和染料木苷含量三者问的相关分析表明,大豆异黄酮含量与大豆苷含量及染料木苷含量、大豆苷含量与染料木苷含量均呈极显著正相关。     

豆粕营养成分对蝙蝠蛾拟青霉发酵产物的影响

为探索培养基中的豆粕以及制备豆粕的原材料大豆对蝙蝠蛾拟青霉发酵产物的影响,我们比较了不同品种大豆和用其制成的豆粕的主要营养成分含量,包括粗蛋白、粗脂肪、6种微量元素（钙、镁、铜、锌、铁、锰）和总异黄酮（大豆苷、黄豆黄苷、染料木苷、大豆苷元、黄豆黄素、染料木素）,以及豆粕作为培养基对蝙蝠蛾拟青霉发酵产物的生物量和有效成分含量包括腺苷、腺嘌呤、虫草素和麦角甾醇的影响,结果表明大豆中钙和大豆苷与发酵产物中腺苷含量正相关。豆粕中钙、大豆苷、大豆苷元、染料木素与发酵产物中腺苷含量正相关;豆粕中钙、镁、大豆苷、大豆苷元、染料木素、总异黄酮与发酵产物中腺嘌呤含量负相关;豆粕中铁与菌丝体干重值正相关。结果表明,培养基中的豆粕和原料大豆主要营养成分含量对蝙蝠蛾拟青霉发酵产物品质有显著影响。     

高效液相色谱(HPLC)技术鉴定中国南方大豆品种异黄酮主要组分

采用高效液相色谱(HPLC)技术检测中国南方六个省份的249份大豆品种异黄酮主要组分含量.结果显示大豆籽粒中可检测出6种主要的异黄酮组分,分别为大豆甙(Daidzin)、甲氧基黄豆甙原(Glycitin)、染料木甙(Genistin)、丙二酰基大豆甙(Malonyldaidzin)、丙二酰基黄豆甙原(Malonylglycitin)和丙二酰基染料木甙(Malonylgenistin).各组分中以丙二酰基(Malonyl)异黄酮组分含量最高(61.2%),且各组分间相关极显著.大豆品种间异黄酮含量变异较大,变异系数达49.6%.来自江苏省的品种海门红黄豆乙异黄酮含量最高(4932.3μg/g),品种宝应等西风含量最低(367.1μg/g).不同省份间异黄酮含量差异极显著,来自浙江省的大豆品种平均含量最高(2717.2μg/g),来自安徽省的平均含量最低(1181.8μg/g).异黄酮含量与生育期呈极显著正相关(r=0.319* * *),与百粒重呈显著正相关(r=0.132*),而与脂肪含量(r=-0.45* * *)和蛋白质含量(r=-0.136)呈负相关.     

大豆异黄酮的分离与纯化

天然大豆异黄酮作为健康食品在防治骨质疏松和癌症方面具有一定功效。由于化合物结构相似,异黄酮甙元特别是高纯度黄豆黄素（glycitein）的获得有一定难度,文献报道大多是通过盐酸水解异黄酮甙的方法获得,而这种方法对环境污染和工厂生产设备腐蚀较大。本文报道了用醇溶剂进行固相提取以及硅胶柱色谱方对含有三种大豆异黄酮甙元的混合物产品进行分离。通过低成本和无环境污染的固相提取方法得到纯度为97％的黄豆黄素和纯度超过95％的大豆黄素（daidzein）;95％纯度的另一种大豆甙元金雀异黄素（genistein）则通过硅胶柱色谱分离得到。应用硅胶柱色谱,一次性分离了一种含有两个异黄酮甙：大豆甙（daidzin）和黄豆甙（glycitin）的产品。     

兔肠道大豆异黄酮还原菌株的分离鉴定及其转化特性

【目的】从兔新鲜粪样中分离对大豆异黄酮黄豆苷原和染料木素具有转化作用的特定细菌菌株。【方法】在厌氧工作站内对獭兔新鲜粪样进行梯度稀释后涂板,挑取单菌落与底物黄豆苷原和染料木素分别厌氧混合培养,用高效液相色谱检测底物被转化情况。【结果】分离得到一株对大豆异黄酮黄豆苷原和染料木素均具有转化作用的革兰氏阳性严格厌氧细菌菌株AUH-JLR41(KJ188150)。根据产物的高效液相保留时间、紫外吸收图谱和质谱分析结果,将菌株AUH-JLR41代谢底物黄豆苷原和染料木素生成的产物分别鉴定为二氢黄豆苷原和二氢染料木素。经手性高效液相系统检测,产物二氢黄豆苷原和二氢染料木素均呈现两个等面积物质峰,表明这两个产物的对映体过量率均为0。通过转化动态研究发现,菌株AUH-JLR41分别在底物黄豆苷原和染料木素加入48 h和72 h后将底物全部转化为产物,该菌株能转化底物黄豆苷原和染料木素的最大浓度均为0.6 mmol/L。经BLAST比对,菌株AUH-JLR41的16S r RNA基因序列与斯奈克氏菌属菌株Slackia equolifaciens DZE(EU377663)的相似性高达99.6%。【结论】兔肠道分离的斯奈克氏菌属菌株Slackia sp.AUH-JLR41在厌氧条件下能将大豆异黄酮黄豆苷原和染料木素分别还原为二氢黄豆苷原和二氢染料木素。     

光照对大豆幼苗组织中异黄酮含量和分布的影响

利用高效液相色谱（ＨＰＬＣ）测定了不同光照处理的大豆（Ｇｌｙｃｉｎｅｍａｘ（Ｌ．）Ｍｅｒｒｉ．）幼苗不同组织的异黄酮类含量。子叶中最高,叶片和根部相对较少。子叶的异黄酮以大豆甙和染料木甙及其丙二酰基结合体为主,且在光照条件下,异黄酮含量随光照时间的增加而显著升高;相反,黑暗中的异黄酮含量随苗龄的增加呈下降趋势;当子叶由黑暗转为光照处理以后,异黄酮含量同样随光照时间的增加而升高。在叶片和根部异黄酮含量和种类也因光照条件的不同而有很大差异。光照条件下,叶片中以染料木甙及其丙二酰结合体和黄酮芦丁为主,且随时间增加呈上升趋势;黑暗中的黄化叶片,则以大豆甙和丙二酰结合体为主,但随时间的变化不明显。在幼苗根部,黑暗条件下几乎检测不出异黄酮的存在;光照条件下,则可检测到５种异黄酮,其中以大豆甙元及其衍生物占主要部分。实验证实了光照对大豆异黄酮的积累有明显的促进作用     

施氮水平对水稻生育后期地上部氨挥发的影响

采用温室盆栽模拟试验,研究了不同施氮水平下水稻开花后地上部氨挥发及其影响因素.结果表明：不同品种水稻开花后地上部日氨挥发量和开花至成熟期的氨挥发总量均随施氮量的增加而增加,且不同施氮水平间存在一定差异.花期和成熟期水稻地上部氨挥发量（y）与功能叶片谷氨酰胺合成酶（GS）活性（x₁）呈显著负相关,而与功能叶片质外体NH₄⁺浓度（x₂）呈显著正相关:y=-0.37846x₁+0.41821x₂+0.04925（R²=0.9471,n=16）.水稻氮素收获指数（x₁）和氮肥生理利用率（x₂）均与地上部氨挥发总量（y）呈显著负相关：y=-0.02117x₁+0.75186(R²=0.8426,n=8）和y=-1.10386x²+35.52676（R²=0.8489,n=8）,说明高氮水平下水稻氮肥利用率的下降与水稻地上部氨挥发量的增加有关.     

添加玉米和水稻秸秆对淹水土壤pH、二氧化碳及交换态铵的影响

通过温室土壤培养试验,研究了不同添加量玉米和水稻秸秆对淹水土壤pH、CO₂及交换态铵的影响.结果表明：在接近中性的土壤中加入秸秆可使土壤pH值降低,4 g·kg^-1玉米和水稻秸秆处理的土壤pH值均与对照差异显著（P＜0.05）,而1 g·kg^-1玉米和水稻秸秆处理与对照差异不显著（P＞0.05）.土壤溶液中的CO₂含量随秸秆添加量的增加而增大,1 g·kg^-1玉米和水稻秸秆处理的土壤溶液CO₂含量最大值分别为35.9％和31.9％（v/v）,与对照（25.8％）差异达显著水平（P＜0.05）,但两者间差异不显著（P＞0.05）;4 g·kg^-1玉米和水稻秸秆处理的土壤溶液CO₂含量最大值分别为54.2％和41.8％（v/v）,与对照差异极显著（P＜0.01）,两者间差异也达显著水平（P＜0.05）.在不施氮肥的情况下,添加秸秆可降低土壤铵态氮浓度,且铵态氮浓度随秸秆添加量的增加而降低,不同添加量处理间差异显著（P＜0.05）;在施入氮肥的情况下,1 g·kg^-1玉米和水稻秸秆处理提高了土壤铵态氮浓度,而4 g·kg^-1玉米和水稻秸秆处理降低了土壤铵态氮浓度.无论是否施入氮肥,玉米和水稻秸秆处理的土壤铵态氮浓度差异不显著（P＞0.05）.     

HPLC法测定甘薯叶片中的叶黄素

建立甘薯叶片中有效成分叶黄素含量的高效液相色谱测定方法,以寻求含量高的甘薯品种。采用Waters SunFire^TM C₁₈(150 mm×4.6 mm,5 μm )色谱柱;以甲醇—水为95:5(V/V)为流动相,流速为0.8 mL·min^-1;检测波长为445 nm;外标法定量。此色谱条件下,叶黄素含量在5~100 μg·mL^-1范围内时,与峰面积呈线性关系;样品平均回收率为99.5%;相对标准偏差 (RSD)为1.9%(n=5)。该方法灵敏、准确、专属性强,适用于甘薯叶片中叶黄素的测定;大部分供试品种间存在显著差异,其中以苏薯8号的含量最高。     

Coumestrin a coumestan derivative from soybean roots

A new coumestrol glycoside, coumestrin, has been isolated from soybean roots together with its aglycone, coumestrol, and the known isoflavones genistin, genistein, daidzin and daidzein. Their structures were determined by spectroscopic technique (¹H NMR, UV, IR, EIMS, CIMS and FDMS) and by some chemical transformations.     

Isoflavone glucoside stress metabolites of soybean leaves

The isoflavone glucosides daidzin, genistin and ononin, the isoflavones daidzein and formononetin, and glyceollins I-III accumulated in soybean leaves inoculated with phytopathogenic bacteria. Treatment of leaves with sodium iodoacetate or yeast extract also led to isoflavonoid accumulation. Various other stress-inducing treatments were not effective. Bacterially-induced accumulation of isoflavone glucosides and the occurrence of ononin and formononetin in soybean are reported for the first time.     

Physiological regulation of seed soaking with soybean isoflavones on drought tolerance of Glycine max and Glycine soja

In this study, the Glycine max Jackson cultivar (the relatively drought-sensitive) and the Glycine soja BB52 accession (the drought-tolerant one) were used as the experimental materials. Effects of seed soaking with exogenous soybean isoflavones (daidzin or genistin) on seed germination, and seedlings photosynthesis, relative electrolytic leakage, content of thiobarbituric acid reactive substances , and anti-oxidative activities were investigated under drought stress conditions. The results showed that, treatments of seed soaking with daidzin or genistin could improve seed germination, and alleviate cell damage, enhance anti-oxidative activities and photosynthesis in drought-stressed seedlings, therefore displayed mitigated effects on soybean drought injury, especially for the drought-sensitive G. max Jackson cultivar. Thus seed soaking with exogenous soybean isoflavones may be a usable approach to enhance drought tolerance of cultivated soybean in practice.     

Light Effect on the Tissue Contents and Distribution of Isoflavones in the Developing Seedling of Soybean

The contents of isoflavones in the developing soybean seedling tissues under different light conditions were examined by high-performance liquid chromatography (HPLC). The results indicated that the total contents of isoflavones in soybean seedling tissues were remarkably different, being the highest in the cotyledons and lower in the leaves and roots. In the cotyledons, the isoflavones like daidzin, genistin and their malonyl conjugates were the major metabolites and were increased markedly with the time of light exposure. In contrast, there was a decrease in the content of isoflavones in the dark. When the dark grown seedlings were tranferred to the light, an increase of the isoflavone content was observed. In the leaves and roots, there was also a marked difference in the contents and types of isoflavones due to various light conditions. Leaves contained mainly the isoflavones such as genistin, malonyl gensistin and flavones rutin, which were increased gradually under light condition, but in the dark, daidzin and malonyl daidzin were the major metabolites and did not change markedly with time. In the roots, no isoflavone was detected in the dark, while five isoflavones were found in the light-grown seedlings, and all root sections were predominated by daidzein and its conjugates. These results provided the evidence that light strongly stimulates the accumulation of isoflavones in soybean seedling tissues.     

Preparation of highly pure daidzin on oligo-β-cyclodextrin-Sepharose HP and investigation of chromatographic behavior of isoflavones by molecular docking

A novel method using column chromatography on oligo-β-cyclodextrin-Sepharose HP for the preparation of high purity daidzin from crude soybean samples was proposed in this work. The isoflavone of daidzin in sample A and B was purified under the optimum mobile phase composed of methanol/acetic acid/water=20.0/8.0/72.0 (v/v/v) at a flow-rate of 1.0 mL/min in one-step operation with a purity of 97.2% and 98.1%, a recovery of 95.3% and 96.3% respectively. The target products in isolated fraction were detected and characterized by HPLC analysis and ESI-MS spectrum. Preparative separation with sample-load of up to 2.42 mg/mL medium gave satisfactory results for daidzin with the purities over 97% and recoveries approximately 90%. Molecular docking simulations were utilized to help demonstrate the inclusion complexation between β-cyclodextrin and the isoflavones in samples through inclusion geometries and calculations of the binding energies. The prediction of the elution orders with AUTODOCK and SURFLEX-DOCK were validated by the chromatographic results.     

Isoflavonoids in root and hypocotyl of soybean seedlings (Glycine max,Fabaceae)

Isoflavonoids, some of which are highly fluorescent, are produced by soybean [Glycine max (L.) Merr.] and serve as chemical signals for certain aspects of nitrogen fixation and microbial resistance. This study was conducted to determine whether soybean mutants with nonfluorescent roots contained abnormal concentrations of isoflavonoids. Isoflavonoids were extracted from the root and hypocotyl of 4-d-old wild-type soybean seedlings (cv. Hark) having fluorescent roots and from four nonallelic mutant, near isogenic lines of Hark having nonfluorescent roots. In addition, isoflavonoids were extracted from the root and hypocotyl of 4-d-old seedlings of near isogenic lines of Hark harboring two pairs of the mutant alleles for nonfluorescent roots. Malonyl daidzin, daidzin, malonyl genistin, and genistein were the most abundant isoflavonoids extracted from either the root or hypocotyl of seedlings with either fluorescent or nonfluorescent roots. Extracted malonyl daidzin, malonyl genistin, and malonyl glycitin decomposed readily, yielding daidzin, genistin, and glycitin, respectively. The concentrations of malonyl genistin and genistein, two highly fluorescent compounds, were similar in both fluorescent and nonfluorescent roots. Thus, root fluorescence was not correlated with abundance per se of fluorescent isoflavonoids in roots. In addition, the abundance of isoflavonoids extractable from the hypocotyl did not correlate with root fluorescence.