注射用高纯度蛋黄卵磷脂制备新工艺的研究

本文探索建立了一条以新鲜蛋黄为原料,以乙醇为唯一提取溶剂,制备高纯度注射用蛋黄卵磷脂的新工艺。首先,用4倍重量的96%(v/v)乙醇作为沉淀剂将新鲜蛋黄中的蛋白质沉淀为滤饼,得到初级提取液;然后,再用4倍重量的96%(v/v)乙醇对滤饼抽提3次,提取残余卵磷脂,得到次级提取液。合并提取液,置9℃下静置分层,除去大部分蛋黄油。所得上清液上硅胶柱,用86%(v/v)的乙醇作为洗脱剂,纯化蛋黄卵磷脂,经高效液相色谱检测纯度达98%。并对提取过程进行了优化,得到最佳工艺条件:35℃的96%乙醇溶液,添加量为1∶4(w/w)提取蛋黄液,180 rpm转速下搅拌10 min。抽滤后,用60℃的96%乙醇提取滤饼3次,料液比为1∶4,静置15 min。最佳工艺条件下,蛋黄卵磷脂的提取率可达93.38%。     

卵磷脂促进小菜蛾颗粒体病毒感染的研究

采用有机溶剂从鸡蛋中提取卵磷脂,与小菜蛾颗粒体病毒混合添食感染2—3龄小菜蛾幼虫,所得数据经统计学方法处理,得到三组死亡率与时间的回归直线方程为y=18.1x-59.8(GV对照)、y=27.23x-99.85(GV+30mg卵磷脂/ml)、y=13.6x-18(GV+70mg卵磷脂/ml)。GV+70mg卵磷脂/ml与GV对照组比较,感染性提高了17％,LT_(50)提前了1.066天。结果表明,卵磷脂对小菜蛾颗粒体病毒感染具有促进作用。     

初级代谢产物和有性生殖促进剂对番茄红素发酵的影响

考察几种初级代谢中间产物对三孢布拉霉发酵生产番茄红素的影响,以及卵磷脂对三孢布拉霉正负菌接合孢子及发酵的影响。结果表明：发酵24h分别添加2．0％的柠檬酸和2．0％的三孢酸,番茄红素的产量分别达到0．99g／L和1．26g/L,比对照分别提高39．43％和32．63％;卵磷脂的添加能促进三孢布拉霉两性接合孢子的形成,进而促进番茄红素的合成,当大豆卵磷脂的添加量为0．3％时,番茄红素的产量为1．58g/L,比对照提高56．44％。     

大豆卵磷脂对小鼠的抗疲劳和抗氧化作用研究

目的:研究大豆卵磷脂的抗疲劳及抗氧化作用。方法:小鼠经口给予大豆卵磷脂30天后,采用负重游泳实验,观察记录小鼠游泳死亡时间;检测血清尿素氮、肝糖原;测定血清和肝匀浆超氧化物歧化酶(SOD)活性、谷胱甘肽过氧化物酶(GSH-Px)活力、丙二醛(MDA)含量。结果:给予大豆卵磷脂后,与对照组相比,实验组小鼠负重游泳时间明显延长,肝糖原消耗量减少,降低运动后血清尿素氮水平(P<0.05);升高小鼠血清和肝匀浆SOD活性及GSH-Px活力,降低MDA的含量(P<0.05)。结论:大豆卵磷脂具有抗疲劳和抗氧化作用。     

对盐和C_8-卵磷脂水溶液的准弹性激光散射研究

本文用自差拍光混频的准弹性激光散射技术,对生物组织重要组成部分的C_a-卵磷脂水溶液内微团的流体力学半径随卵磷脂浓度,温度,添加的单价盐离子类型和强度的变化情况,进行了系统性研究.并根据微团形成的梯级模型,推导出微团的大小和结构在微团形成过程中的变化,微团形成参数以及盐控制微团形成的规律.     

大豆卵磷脂对小鼠的抗疲劳和抗氧化作用研究

目的：研究大豆卵磷脂的抗疲劳及抗氧化作用。方法：小鼠经口给予大豆卵磷脂30天后,采用负重游泳实验,观察记录小鼠游泳死亡时间;检测血清尿素氮、肝糖原;测定血清和肝匀浆超氧化物歧化酶（SOD）活性、谷胱甘肽过氧化物酶（GSH—Px）活力、丙二醛（MDA）含量。结果：给予大豆卵磷脂后,与对照组相比,实验组小鼠负重游泳时间明显延长,肝糖原消耗量减少,降低运动后血清尿素氮水平（P〈0．05）;升高小鼠血清和肝匀浆SOD活性及GSH-Px活力,降低MDA的含量（P〈0．05）。结论：大豆卵磷脂具有抗疲劳和抗氧化作用。     

rUreB亚单位疫苗中试产品的纯度检测和肽图分析

分析重组幽门螺杆菌尿素酶B亚单位疫苗 (rUreB)中试产品的纯度 ,制备肽指纹图 ,证明rUreB三批中试产品在一级结构上的一致性和生产工艺稳定性。利用面积归一法检测产品的纯度 ,在非还原条件下用TPCK处理过的胰蛋白酶水解 3批中试产品 ,用反相HPLC制备分析肽图。结果显示 ,rUreB的 3批中试产品的纯度达到中试质量要求和肽图分析要求 ,3批中试产品的肽指纹图基本一致 ,重现性好。     

高纯度大豆卵磷脂对中国仓鼠肺细胞染色体畸变作用

目的研究高纯度大豆卵磷脂对中国仓鼠肺细胞（cHL）染色体畸变作用。方法测定高纯度大豆卵磷脂对CHL细胞的半数抑制浓度（IC50）,根据IC50设立不同剂量组,进行正式试验,分别观察高纯度大豆卵磷脂接触CHL6h,24h及加S9后6h染色体的畸变情况,根据标准进行结果判定。结果染毒6h,24h及加S9后染毒6h染色体畸变为阴性。结论高纯度大豆卵磷脂不能引起CHL细胞染色体产生畸变作用。     

真鲷仔鱼用微粒子饲料中肽酶与大豆卵磷脂添加效果的研究

本研究以酪蛋白分解物为蛋白源配制三种微粒子饲料MD-S、MD-T和MD-U对真鲷开口仔鱼进行饲养试验。以MD-S的配方为基准,MD-T采用粉状大豆卵磷脂和麸质代替液状大豆卵磷脂;MD-U则另外添加0.1%的肽酶。结果表明,微粒子饲料在水中浸泡15min后,MD-T的溶出率(35.5%)低于MD-S(46.8%)和MD-U(45.8%);实验结束时(20日龄),仔鱼的成活率以生物饵料(轮虫)组为最高(86.3%),其次是MD-T组为20.7%,显著高于(P<0.05)MD-S组(13.3%)和MD-U组(13.6%);生物饵料组的仔鱼全长(6.14±0.49mm)显著大于微粒子饲料组(4.23±0.30mm~4.46±0.30mm),各微粒子饲料组之间仔鱼的全长并不存在显著差异(P>0.05)。在孵化后第12d,微粒子饲料组的仔鱼肠上皮细胞发育良好,但至孵化后第18d,仔鱼肠上皮细胞大部分萎缩、并发生脱落。鱼体的蛋白质、DNA与RNA日间增长率微粒子饲料MD-T组高于MD-S和MD-U组,但都低于生物饵料组。由此可见,微粒子饲料中添加肽酶并无助真鲷仔鱼对其消化吸收;可是,使用粉状大豆卵磷脂与麸质代替液状卵磷脂能增强微粒子饲料的黏合性,可减少其营养成分的溶出率,从而提高微粒子饲料的饲育效果。     

31p磁共振波谱物质检测准确性的验证研究

目的:评价1.5T磁场下31PMRS磷化合物检测的准确性.方法:以卵磷脂为参照标准品,对照研究对同一位正常志愿者骨骼肌肉磷谱代谢特征,对其股骨下端先后行两次相同参数、相同条件状态下的31P波谱检测,第二次检测时在感共趣区内体外加入卵磷脂标准品,对比两次检测的结果变化.结果:对同一位正常志愿者先后两次进行渡谱检测,加入卵磷脂后各代谢物波谱变化为β-ATP:-24.46,α-ATP:+82.61,γ-ATP:-32.89,PCr:+64.32,Pi.+110.5,PDE:+462.57,PME:+15.71.加入卵磷脂后,PDE峰改变最明显.结论:1.5T磁场下31P MRS能准确检测体内化合物的存在及其定量分析,研究结果可信.     

Separation of human Fab fragments on negative mode Ni(II)-TREN-agarose chromatography

We evaluated the feasibility of using immobilized metal-ion affinity chromatography (IMAC) with nickel ion complexed with Tris(2-aminoethyl)amine (TREN) immobilized on agarose gel for purification of human Fab fragments by negative chromatography. Efficient purification of Fab fragments from digested human IgG (immunoglobulin G) (106.4% purity) was accomplished in Tris-HCl buffer at pH 7.5 without NaCl (based on total protein concentration and radial immunodiffusion of human Fab). A technological application of Ni(II)-TREN-agarose using non transgenic soybean protein extract spiked with human Fab fragments as feedstream was also studied. Experiments using Tris-HCl at pH 7.0 as loading buffer allowed the adsorption of almost all of the soybean proteins. Sixty-six percent of the loaded human Fab fragments were recovered in the flowthrough and washing fractions with about 90% purity. These results demonstrate that Ni(II)-TREN-agarose is a potential adsorbent for recombinant Fab fragment purification.     

Effect of lecithin nanoliposome or soybean lecithin supplemented by pomegranate extract on post-thaw flow cytometric,microscopic and oxidative parameters in ram semen

This investigation was carried out to study the effect of soybean lecithin 1.5% (wt/vol) (0, 2.5, 5 and 7.5 mg l⁻¹ pomegranate extract (PE)) or PE-loaded lecithin nanoliposome (0, 2.5, 5 and 7.5 mg l⁻¹) to Tris-based extender. Sperm motility (CASA), viability, membrane integrity (HOS test), abnormalities, mitochondrial activity, apoptosis status, lipid peroxidation, total antioxidant capacity (TAC)) and antioxidant activities (GPX, SOD) were investigated following freeze-thawing. No significant differences were detected in motility parameters, viability, membrane integrity, and mitochondria activity after thawing sperm between soybean lecithin and lecithin nanoliposomes. It was shown that PE5 significantly improved sperm total and progressive motility, membrane integrity, viability, mitochondria activity, TAC and reduced lipid peroxidation (malondialdehyde concentration). Moreover, the percentage of apoptotic sperm in PE5 extenders was significantly the lowest among other treatments. Sperm abnormalities, SOD and GPX were not affected by the antioxidant supplements. For apoptotic status, no differences were observed between soybean lecithin and lecithin nanoliposome. We showed that lecithin nanoliposome extender can be a beneficial alternative extender to protect ram sperm during cryopreservation without any adverse effects. It was also observed that regarding pomegranate concentration, PE5 can improve the quality of ram semen after thawing.     

Preparation and Purification of Xylitol-5-Phosphate from a Cell Extract of Lactobacillus casei Cl-16

A simple procedure which yields pure xylitol-5-phosphate is described. A cell extract of Lactobacillus casei Cl-16 from a 6-liter culture was used to synthesize up to 70 mg of xylitol-5-phosphate overnight from xylitol and phosphoenolpyruvate via a xylitol phosphoenolpyruvate:phosphotransferase system with a 53% yield. Centrifugation, filtration, precipitation as a barium salt, and ion-exchange batch chromatography permitted recovery of nearly 90% of the phosphorylated product synthesized. Thin-layer chromatography and enzymatic analysis indicated a purity level of more than 99%. The method was used to synthesize [U-¹⁴C]xylitol-5-phosphate, and it is suitable for the synthesis of many other nonmetabolizable sugar phosphates.     

4 种不同脂肪源对太平洋鲑生长和体组成的影响

在日粮中添加11.5%的4 种不同来源脂肪饲养180 尾初始重约为110g 的太平洋鲑(Oncorhynchus spp.)于水泥池中56d。实验分4 组,每组3个平行池,每池15尾鱼。研究日粮中4 种不同来源脂肪对淡水养殖太平洋鲑生长性能、体组成与品质的影响。4 组脂肪源分别为鱼油(实验1 组)、大豆油(实验2 组)、大豆磷脂(实验3 组)和玉米油(实验4 组)。实验表明:(1) 实验各组太平洋鲑存活率相似,但大豆磷脂组的特定生长率显著好于鱼油组、大豆油组和玉米油组(P0.05)。大豆磷脂组、大豆油组和玉米油组的饲料效益显著好于鱼油组(P< 0.05); (2) 大豆油组、大豆磷脂组和玉米油组太平洋鲑肠系膜脂肪与肝脏脂肪含量不同程度低于鱼油组,而肌肉中脂肪含量不同程度低于鱼油组; (3) 实验各组太平洋鲑肝脏脂肪、肌肉脂肪和肠脂中总多不饱和脂肪酸组成基本相似,但玉米油组、大豆磷脂组和大豆油组太平洋鲑总n-3 多不饱和脂肪酸比例较鱼油组显著下降,而总n-6 系多不饱和脂肪酸比例显著提高(P<0.05);(4) 玉米油组、大豆磷脂组和大豆油组太平洋鲑血浆中脂肪分解酶、甘油三酯和高密度脂蛋白指标较鱼油组不同程度上升;(5) 实验各组太平洋鲑解剖组织学检查未见异常病理变化。实验结果表明,淡水养殖条件下,太平洋鲑日粮中脂肪以添加大豆磷脂的生长性能最好,大豆油、玉米油和鱼油效果相似,添加玉米油、大豆磷脂和大豆油均不影响太平洋鲑健康状况和品质。       

Magnetometric studies on the reticulo-endothelial system of the liver after administration of different lipid emulsions.

A comparison of the burdening effects on the reticulo-endothelial system (RES) by soybean oil emulsions with two different emulsifiers [soybean lecithin (SOB) and egg lecithin (EGG)] or by a perfluorochemical emulsion (Fluosol-DAR, PFC) was performed using a magnetometric method. This method is based on measuring the relaxation of injected iron oxide (gamma-Fe2O3) above the liver after magnetization in a strong external magnetic field. Three different methods of evaluation of the data were chosen. First, according to the conventional monoexponential function, second, according to a monoexponential function with a constant, and third, using the ratio of the initial dynamic to total magnetic field strength. In contrast to SOB and EGG, the PFC emulsion which was used depressed the RES-capacity to less than 31% of the control values (p less than 0.001). Following the administration of SOB the phagosomal motion was significantly lowered after 6 h (p less than 0.01) and 1-2 days (p less than 0.05); thereafter no significant difference of the relaxation constants remained as compared to the control group. The fatty emulsion with egg lecithin showed no significant lowering of the RES-capacity during the entire observation period (p greater than 0.05). Our results indicate that the RES-capacity is diminished not only by a PFC emulsion, but also temporarily by a soybean lecithin emulsion, though not by an egg lecithin emulsion, when given in the same dosages.     

Lecithin essay in the diet of young Penaeus vannamei (Crustacea: Decapoda)

The effect of different lecithin sources and presentations on growth, food conversion ratio and survival of P. vannamei (290 mg +/- 0.02) was studied. The bioassay was designed in order to compare different dietary levels and different quality of lecithin. Squid lecithin, crude soybean (7%), deoiled soybean lecithin (3.48%) in combination with fish oil or squid neutral lipids, in a partially dilapidate formula. The isoenergetic diets were fed ad libitum to four replicate groups (tanks) of 15 shrimps each (5 x 4 x 15), during 28 days. The result of the bioassay with the partially dilapidate formulas was; the best growth rate (191%) and FCR (1.69 +/- 0.041) were obtained with the diet containing 7% of soybean crude lecithin as the unique lipid source. Followed by the diet countering 3.94% deoiled lecithin and 2.42% Menhaden oil (172% and 2.03 +/- 0.054 respectively). As expected, the worst results were obtained without the dietary lecithin 121% and 2.42 +/- 0.129). Crude soybean lecithin alone covered the phospholipid and neutral lipids requirements as well as the combination of deoiled soybean lecithin with fish or squid oil.     

In vitro comparison of egg yolk-based and soybean lecithin-based extenders for cryopreservation of ram semen

Substitution of egg yolk with soybean lecithin may reduce hygienic risks in extenders. Though a few studies have been performed on the effect of soybean lecithin in bull, to date evaluation of ram semen in vitro fertility after cryopreservation with use of soybean lecithin has not been studied. This study assessed the effect of 1% or 2% (wt/vol) soybean lecithin (L1 or L2) or 15% or 20% (vol/vol) egg yolk (E15 or E20) supplemented with 5% or 7% glycerol (G5 or G7) in a Tris-based medium for cryopreservation of ram (Oviss arries) semen. Although no significant difference was observed in pattern of capacitation, the best results in terms of sperm motility, viability postthaw, and cleavage rates were observed with L1G7 (51.9 ± 4.8%, 48.1 ± 3.5%, and 79.6 ± 3.9%, respectively) and E20G7 (51.8 ± 2.9%, 46.7 ± 4.0%, and 72.9 ± 6.4%, respectively). Our results also showed that 1% lecithin and 20% egg yolk was superior to 2% lecithin and 15% egg yolk. In terms of cleavage rate, 7% glycerol was superior to 5% glycerol. No significant difference was obtained between groups in terms of blastocysts rate per cleaved embryo. Therefore, we concluded that the optimal concentration of lecithin and egg yolk is 1% and 20%, respectively, along with 7% glycerol. In addition, our results suggest that lecithin can be used as a substitute for egg yolk.     

Peculiar behaviour of rabbit thymocytes in interaction with liposomes of different compositions shown by fluorescence polarization studies,lipid analysis,and uptake of vesicle-entrapped carboxyfluorescein

In order to obtain more information on membrane phenomena occurring at the cell surface of rabbit thymocytes we have performed experiments aimed at altering the lipid composition of the plasma membrane. Thymocytes were incubated at 37°C with phospholipid vesicles of different compositions. Vesicle-cell interaction was followed by measuring the degree of fluorescence polarization and the uptake of vesicle-entrapped carboxyfluorescein. Neutral and negatively charged liposomes prepared from egg phosphatidylcholine are currently used in investigations of vesicle-cell interaction. In this report we show that these liposomes do not interact with rabbit thymocytes as is evident from unaltered lipid fluidity measured in whole cells and in isolated plasma membranes. This was confirmed by experiments with vesicle-entrapped carboxyfluorescein showing hardly any uptake of the fluorophor from neutral and negatively charged egg phosphatidylcholine liposomes. Using both techniques substantial interaction was found with positively charged egg phosphatidylcholine liposomes and with liposomes prepared from soybean lecithin which is composed of a variety of phospholipids. The results of these experiments were supported by lipid analysis of cells treated with soybean lecithin liposomes. Increase in phosphatidylcholine contents of mixed phospholipid vesicles was further shown to result in decreased vesicle-cell interaction. From measurements of the quantity of carboxyfluorescein inside cells and the total amount of cell-associated carboxyfluorescein it is concluded that adsorption plays a prominent role in interaction between liposomes and rabbit lymphocytes. The grade of maturation of lymphocytes was also found to affect vesicle-cell interaction. The more mature thymocytes took up more vesicle-entrapped carboxyfluorescein from soybean liposomes than immature thymocytes. Mesenteric lymph node cells exhibited a still stronger interaction. The role of vesicle and cell surface charge and membrane fluidity of both vesicles and cells in interaction between liposomes and rabbit thymocytes is discussed.     

用膨化柱填料的方法从红藻中规模分离纯化R-藻红蛋白(英)

以Phenyl_sepharose作为柱填料 ,用streamline柱层析技术从红藻 (Palmariapalmata (Lannaeus)Kuntze)中规模分离捕光色素蛋白———R_藻红蛋白。由于不是传统的从层析柱上方进样 ,而是用泵将样品从streamline层析柱的下方加样 (从下至上 ) ,因而解决了用一般层析柱分离R_藻红蛋白时海藻抽提液中大量的粘性多糖堵塞层析柱的难题。用P .palmata粗提液上样后 ,分别用 0 .2mol/L、0 .1mol/L和 0 .0 5mol/L的 (NH4) 2 SO4溶液从相反的方向 (即从上到下 )洗脱层析柱 ,发现这些洗脱液中的藻红蛋白纯度已经较高。然后将洗脱液透析去盐 ,用阴离子交换柱层析 (Q_sepharose)进一步纯化。经过这两次柱层析后 ,R_藻红蛋白的纯度 (OD565/OD2 80 )超过 3.5 ,高于一般认可的R_藻红蛋白的纯度标准 3.2 ;产率为每克冷冻P .palmata可纯化 0 .12 2mg高纯度的R_藻红蛋白 ,比使用一般分离方法的产率要高 10倍。这些结果表明 ,使用本文报道的方法纯化藻红蛋白 ,将会使作为生化检测试剂的藻红蛋白市场价格大幅度下降。     

Tetramethylpyrazine production from glucose by a newly isolated Bacillus mutant

2,3,5,6-Tetramethylpyrazine (TTMP) was produced using a newly isolated Bacillus mutant. Culture medium optimization studies showed that soytone, an enzyme-hydrolysate of soybean meal, with the supplementation of vitamins, can fully replace yeast extract plus peptone in supporting TTMP production from glucose. In a 5-l fermenter, using the optimized medium which contained 20% glucose, 5% soytone, 3% (NH₄)₂HPO₄, and vitamin supplements, fermentations were carried out with stirring at 700 rpm, air flow at 1.0 vvm, controlled pH at 7.0, and temperature at 37 °C. TTMP reached 4.33 g l⁻¹ after 64.6 h cultivation. A product recovery method was described, which involved evaporation, crystallization, and lyophilization. The product purity was 99.88%, determined by GC with the normalization method. The main impurities were 2,3,5-trimethylpyrazine (0.09%) and 2-ethyl-3,5,6-trimethylpyrazine (0.02%), which were identified by GC/MS. ¹³C NMR determination also gave a consistent result. Natural and high purity of the product and the utilization of cheap green renewable materials make this process promising to compete with TTMP chemical synthetic methods.