蜜蜂蜂王不同于工蜂的关键因素-蜂王浆主蛋白1

蜂王浆是决定蜜蜂幼虫发育中级型分化,即成为蜂王还是工蜂的关键环境因素,而蜂王浆主蛋白(main royal jelly proteins,MRJPs)是反映蜂王浆新鲜度的重要指标。日本镰仓昌树以蜜蜂和果蝇为模型的最新研究表明,MRJP1是蜂王浆中决定蜜蜂级型分化的关键因子,该蛋白可通过激活虫体脂肪体中的表皮生长因子信号通路,引发个体增大、发育时间缩短和卵巢发育等蜂王特征的出现。因此,今后很有必要进一步开展MRJP1对人体的营养功能和作用机理研究,为MRJP1应用于功能食品提供科学依据。      

蜂王浆对免疫功能低下小鼠免疫功能的影响

研究蜂王浆对免疫低下小鼠免疫系统的调节作用。48只雄性昆明小鼠随机分为空白组、环磷酰胺模型组、酪蛋白组以及蜂王浆低、中、高剂量组,每组8只。除空白组外,其余5组每天腹腔注射80 mg/(kg bw)环磷酰胺,建立免疫低下小鼠模型。各组小鼠经不同样品连续灌胃30 d后测量小鼠体重、脾脏和胸腺质量及HE染色观察脾脏与肝脏的形态、白细胞计数、免疫球蛋白含量、细胞因子水平、脾淋巴细胞增殖能力、睾酮含量。造模后,脾脏和胸腺指数降低,小鼠脾脏和肝脏镜下形态学结构紊乱,白细胞计数、免疫球蛋白、IL-6水平显著降低,T/B淋巴细胞增殖能力显著下降;经蜂王浆干预后,相比于模型组,蜂王浆干预组小鼠上述指标有不同程度的改善,其中以高剂量组效果最为显著。蜂王浆可以增强环磷酰胺造成的免疫功能低下小鼠的免疫功能。     

Structures and properties of antioxidative peptides derived from royal jelly protein

We previously reported that royal jelly proteins (RJPs) hydrolyzed with protease N show the strong antioxidative activity against the peroxidation of linoleic acid. In this study, 29 antioxidative peptides were isolated from hydrolysate by membrane ultrafiltration, anion-exchange chromatography, gel filtration chromatography, and reverse-phase high performance liquid chromatography. We particularly focused on 12 small peptides with 2–4 amino acid residues: these structures were identified as Ala-Leu, Phe-Lys, Phe-Arg, Ile-Arg, Lys-Phe, Lys-Leu, Lys-Tyr, Arg-Tyr, Tyr-Asp, Tyr-Tyr, Leu-Asp-Arg, Lys-Asn-Tyr-Pro. Analysis of the antioxidative properties of these peptides revealed strong hydroxyl radical scavenging activity, but neither metal-chelating activity nor superoxide-anion radical scavenging activity differed significantly among these peptides. Moreover, three dipeptides (Lys-Tyr, Arg-Tyr, and Tyr-Tyr) containing Tyr residues at the C-terminal had strong hydroxyl-radical and hydrogen-peroxide scavenging activity. This suggests that the antioxidant properties of these peptides are due to a combination of these abilities to act as free-radical scavengers. Three tyrosyl dipeptides containing Tyr residues at their C-termini (Lys-Tyr, Arg-Tyr, and Tyr-Tyr) have phenolic hydroxyl groups, which scavenge the free radicals via the mechanism of donating a hydrogen atom from their hydroxyl group.     

Fast determination of royal jelly freshness by a chromogenic reaction

Royal jelly is one of the most important products of honeybees. Given its role in development of bee brood into fertile individuals of the royal caste it is also used in health products for human consumption. Royal jelly spoils and loses its health-promoting properties depending on storage duration and conditions. To ensure product quality before selling, it is therefore necessary to assess royal jelly freshness. Many indexes of freshness have been suggested, but they all lack reliability or require complex and time-consuming analyses. Here we describe a method to detect royal jelly freshness based on a chromogenic reaction between royal jelly and HCl. We demonstrate that analyses based on color parameters allow for the discrimination of royal jelly samples based on the duration of their storage. Color parameters of royal jelly stored at -18 and 4 °C for 28 d remained comparable to that of fresh samples, which supports the reliability of the method. The method of freshness determination described is practical, cheap, and fast and can thus be used in real-time when trading royal jelly. PRACTICAL APPLICATION: The method developed can be used to assess royal jelly freshness. It is practical, cheap, and fast and can thus be used in real-time when trading royal jelly.     

高效液相色谱法测定蜂王浆中6种糠醛类物质含量

目的建立同时测定蜂王浆中5-羟甲基-2-糠醛(5-HMF)、5-甲基糠醛(5-MF)、2-糠醛(F)、2-糠酸(2-OIC)、3-糠酸(3-OIC)、5-羟甲基-2-糠酸(5-HMFacid)等6种糠醛类物质含量的高效液相色谱分析方法。方法样品经去离子水提取,亚铁氰化钾和乙酸锌溶液沉淀蛋白后,用C18色谱柱分离,0.5%甲酸水溶液和甲醇梯度洗脱,在各化合物的最佳吸收波长处进行检测。结果在测试范围内,6种糠醛类物质的浓度与相应的峰面积呈线性关系,相关系数(R2)不小于0.9993。方法定量和定性检出限分别在0.07~0.26μg/m L和0.02~0.08μg/m L之间,加标回收率为86.2%~99.7%,相对标准偏差小于2.40%。结论 该方法操作简单,灵敏度高,适合蜂王浆中6种糠醛类化合物的测定。     

表没食子儿茶素没食子酸酯（EGCG）标准样品的研制

依据GB/T 15000.3-2008《标准样品工作导则（3）标准样品：定值的一般原则和统计方法》,开展表没食子儿茶素没食子酸酯（epigallocatechin gallate,EGCG）国家标准样品研制工作。以绿茶样品作为研究材料,采用溶剂提取、萃取、高速逆流色谱分离纯化,制备得到EGCG。通过紫外-可见吸收光谱、红外光谱、质谱和核磁共振波谱等方法对EGCG样品进行结构鉴定,并进行均匀性、稳定性检验。采用8 家实验室进行联合定值,并对定值结果进行数据分析。结果：EGCG标准样品均匀性和稳定性良好,定值结果为99.29%,置信度95%的扩展不确定度为0.16%。结论：所制备得到的EGCG样品满足GB/T 15000.3-2008的要求,可用于绿茶相关产品中EGCG的质量控制和方法验证。     

绿茶中表没食子儿茶素没食子酸酯生物活性研究进展

表没食子儿茶素没食子酸酯（EGCG）是从绿茶中提取的多酚物质含量最高的一种水溶性活性成分,在茶叶中约占到茶叶毛质量的7%～13%,也是茶多酚类物质中最具生理活性的物质。在现阶段植物和植物衍生物因其具有低副作用和在自然界中具有高可用性,作为发现和开发新抗菌和抗病毒药物的天然来源而备受关注。该综述总结了近几年绿茶中EGCG在抗病毒、抗菌、抗肿瘤、抗炎等方面的研究进展,为我国茶叶在健康产业中的应用提供理论支撑。     

水提酸沉法提取蜂王浆水溶性蛋白的工艺优化

以黑龙江密山蜂王浆作为原料,水溶性蛋白提取率作为评价指标,在单因素实验和PB实验的基础上,利用响应面法对水提酸沉法提取蜂王浆水溶性蛋白的工艺进行优化。实验结果显示,最佳工艺参数为离心转速10000 r/min、离心时间25 min、料水质量比1∶20、超声波提取时间41 min、沉淀时间43 min,此条件下蜂王浆水溶性蛋白提取率达到69.97%,是蜂王浆水溶性蛋白提取的较好方法。      

水提铵沉法提取蜂王浆水溶性蛋白工艺优化

选定黑龙江密山蜂王浆作为原料,以水溶性蛋白提取率作为评价指标。在单因素实验和PB实验的基础上,利用响应面优化法对水提铵沉法提取蜂王浆水溶性蛋白的工艺进行优化。实验结果显示,最佳工艺参数为料水质量比1∶20、超声波提取时间40 min、硫酸铵饱和度89%、pH5、沉淀时间50 min,此条件下蜂王浆水溶性蛋白提取率达到80.22%,是适用于蜂王浆水溶性蛋白提取的较好方法。      

EGCG乙酰化分子修饰取代度影响因素分析

为改善表没食子儿茶素没食子酸酯(epigallocatechin gallate,EGCG)的脂溶性和生物利用度,在乙酸乙酯体系中化学合成乙酰化EGCG。研究酰基供体乙酸酐用量、催化剂吡啶用量、溶剂乙酸乙酯用量、反应温度、反应时间对EGCG乙酰化分子修饰取代度的影响。结果表明,以0.19 g的EGCG为原料,乙酸酐用量0.04 m L、吡啶用量0.02 m L、乙酸乙酯用量20~60 m L、反应温度20~25℃、反应时间1~3 h有利于1~3取代度乙酰化EGCG生成;乙酸酐用量0.12~0.20 m L、吡啶用量0.06~0.20 m L、乙酸乙酯用量10~20 m L、反应温度17~25℃、反应时间5~9 h有利于4~6取代度乙酰化EGCG生成;乙酸酐用量0.40~0.80 m L、吡啶用量0.10~0.20 m L、乙酸乙酯用量5~10 m L、反应温度25~40℃、反应时间5~9 h有利于7~8取代度乙酰化EGCG生成。     

从蜂王浆中提取蜂王酸的研究

目的探讨从蜂王浆中提取蜂王酸（10-HAD）的工艺。方法二次有机溶剂提取，二次调pH，用高效液相色谱法对10-HDA进行定性分析。结果蜂王浆中10-HDA的最佳提取条件为：蜂王浆水溶液pH10，加入乙醚提取杂质，再调pH2加入乙醚提取10-HDA。结论此法可从蜂王浆中高效提取10-HDA。     

液相色谱-串联质谱法同时测定蜂王浆中15种高风险农药残留

摘 要：目的 建立液相色谱-串联质谱法(liquid chromatography-tandem mass spectrometry,LC-MS/MS)测定蜂王浆中苯菌灵、乙基硫菌灵、多菌灵等15种高风险农药残留的检测方法。方法 样品经提取净化后,以0.1%甲酸水溶液与甲醇为流动相进行梯度洗脱,采用电喷雾离子源正离子方式扫描,在多反应监测(multiple reaction monitoring,MRM)模式下检测。结果 实验验证15种农药的检测方法在一定的浓度范围内,具有良好线性关系,相关系数为0.9915~0.9999, 回收率范围为65.3%~113.5%,精密度为0.56%~9.11%,检出限为0.01 mg/kg,定量限为0.1 mg/kg。结论 该方法操作简单、准确可靠、灵敏度高,可同时完成蜂王浆中15种农药残留的检测。     

Antioxidant and Antimicrobial Activities of (‐)‐Epigallocatechin‐3‐gallate (EGCG) and its Potential to Preserve the Quality and Safety of Foods

Quality deterioration of fresh or processed foods is a major challenge for the food industry not only due to economic losses but also due to the risks associated with spoiled foods resulting, for example, from toxic compounds. On the other hand, there are increasing limitations on the application of synthetic preservatives such as antioxidants in foods because of their potential links to human health risks. With the new concept of functional ingredients and the development of the functional foods market, and the desire for a “clean” label, recent research has focused on finding safe additives with multifunctional effects to ensure food safety and quality. (‐)‐Epigallocatechin‐3‐gallate (EGCG), a biologically active compound in green tea, has received considerable attention in recent years and is considered a potential alternative to synthetic food additives. EGCG has been shown to prevent the growth of different Gram‐positive and Gram‐negative bacteria responsible for food spoilage while showing antioxidant activity in food systems. This review focuses on recent findings related to EGCG separation techniques, modification of its structure, mechanisms of antioxidant and antimicrobial activities, and applications in preserving the quality and safety of foods.     

基于荧光猝灭法的复合GA/EGCG与热变性乳清蛋白相互作用机制

目的:开发新型食品功能因子载体,有效调控载体在加工、运输和储存过程中功能因子的释放特性。方法:在不同温度(298.2,304.2,310.2 K)下,采用不同摩尔比(1∶0,3∶1,1∶1,1∶3,0∶1)复配的没食子酸(GA)和表没食子儿茶素没食子酸酯(EGCG)与热变性乳清蛋白(HWPI)之间相互作用,利用荧光猝灭法探究其相互作用机制。结果:与GA相比,EGCG与HWPI的亲和性更强,二者共存时,互相抑制彼此与HWPI的结合,结合常数均减小,亲和性降低;EGCG与HWPI的预先结合会促进低浓度、却抑制高浓度GA与HWPI的结合,而GA与HWPI的预先结合会抑制EGCG与HWPI的进一步结合。GA、EGCG与HWPI结合主要驱动力为离子作用力和疏水作用力。与单一多酚体系相比,GA/EGCG(3∶1)与HWPI反应后,体系内离子作用力和疏水作用力增强;GA/EGCG(1∶1)和GA/EGCG(1∶3)与HWPI的体系内主要以疏水相互作用为驱动力。结论:GA和EGCG与HWPI三者之间发生竞争关系;GA/EGCG与HWPI的主要相互作用力与多酚复配比例有关,与二元体系相比,GA/EGCG...     

Estimation and characterisation of major royal jelly proteins obtained from the honeybee Apis merifera

Royal jelly (RJ) contains many components, including proteins. We focused on major royal jelly proteins (MRJPs) under natural conditions, and attempted to determine the content ratios and molecular forms of MRJPs by size-exclusion HPLC, SDS–PAGE, 2-DE and MALDI TOF/TOF MS. Soluble RJ proteins were extracted by dialysis followed by several centrifugation techniques. Soluble RJ proteins were universally separated into five peaks (640 kDa, 280 kDa, 100 kDa, 72 kDa and 4.5 kDa) by size-exclusion HPLC on a Superose 12 column. Among these peaks, both the 280 kDa and 72 kDa peaks were major, but the intensity of the 280 kDa peak differed markedly among original RJ samples (n = 70). The main 280 kDa protein was separated into a 55 kDa band by reducing and non-reducing SDS–PAGE. This protein was also separated into multiple spots ranging from pH 4.2 to 6.5 by 2-DE. These spots were identified as MRJP 1 by MALDI TOF/TOF MS. From these results, MRJP 1 was thought to comprise an oligomer complex linked by non-covalent bonds under natural conditions. Another major protein, the 72 kDa peak on Superose 12 HPLC, was identified as MRJP 2.     

3,10-Dihydroxy-decanoic acid,isolated from royal jelly,stimulates Th1 polarising capability of human monocyte-derived dendritic cells

Different pharmacologically active components have been isolated from royal jelly. Some of them possess imunomodulatory activity, but the mechanisms of their effect on the immune system have not been elucidated yet. In this study we tested the effect of 3,10-dihydroxy-decanoic acid (3,10-DDA), a fatty acid isolated from royal jelly, on maturation and functions of human monocyte-derived dendritic cells (MoDCs). We showed that 3,10-DDA stimulated maturation of MoDCs by up-regulating the expression of CD40, CD54, CD86 and CD1a, and increased their allostimulatory potential in co-culture with allogeneic CD4⁺T cells. 3,10-DDA-treated MoDCs enhanced the production of IL-12 and IL-18, and stimulated the production of interferon-γ in co-culture with allogeneic CD4⁺T cells, compared to control MoDCs. In contrast, the production of IL-10 was down-regulated. In conclusion, our results suggest that 3,10-DDA stimulates maturation and Th1 polarising capability of human MoDCs in vitro, which could be beneficial for anti-tumour and anti-viral immune responses.     

Extraction of Epigallocatechin Gallate and Epicatechin Gallate from Tea Leaves Using β‐Cyclodextrin

Use of organic solvents to extract phenolic compounds from plants may result in environmental pollution and cause health problems in persons. Replacing organic extraction solvents by green extracting agents without affecting the extraction yield is one of the most pressing problems to be solved. The aim of this study is to evaluate the capacity of β‐cyclodextrin (β‐CD) to recover phenolic compounds from tea leaves. The extract obtained using the ethanol/water mixture presented the highest total phenolic content, followed by those obtained using β‐CD solution and water. HPLC analysis of the extracts showed that the addition of β‐CD to the extracting agent had a selective effect on the extraction of epigallocatechin gallate (EGCG) and epicatechin gallate (ECG). The extraction yield of EGCG and ECG using 15 g/L β‐CD were higher than that obtained using water and 50% ethanol. Molecular docking results indicated that the molecules of EGCG and ECG were more inclined to interact with β‐CD than epigallocatechin, epicatechin, and gallocatechin. The impact of β‐CD concentration, temperature, and time on EGCG and ECG extraction from tea leaves was investigated and the maximum amount of EGCG (118.7 mg/g) and ECG (54.6 mg/g) were achieved when extracted with 25 g/L aqueous β‐CD solution at 60 °C for 60 min. The present study indicates that aqueous β‐CD can be used as an alternative to organic solvents to recover EGCG and ECG from tea leaves.