Partial purification of a serum fraction from fasted pigs that inhibits proliferation of cultured myogenic cells

Sera from pigs fasted as little as 24 h appears to contain a factor(s) that inhibits proliferation of myogenic cells in culture. An inhibitor of myogenic cell proliferation has been partially purified from this sera by using a combination of gel filtration and immunoaffinity chromatography. The inhibitory activity elutes from a Sephacryl S-300 column at a Kav (elution minus void volume divided by total minus void column volume) between .41 and .59. Proteins banding at 76 and 67 kilodaltons appear to predominate on sodium dodecyl sulfate polyacrylamide gels of this fraction. Small quantities of each of these proteins were electrophoretically purified and used to elicit production of anti-76 and anti-67 immunoglobulin G in rabbits. These antibodies were used to prepare anti-76 and anti-67 column was particularly useful in isolating the inhibitor because it removed mitogens that made detection of the inhibitory activity difficult. The partially purified inhibitor inhibits proliferation of L6 myogenic cells in a concentration-dependent manner. On sodium dodecyl sulfate polyacrylamide gels, the predominant proteins in the inhibitor fraction band at approximately 63 and 61 kilodaltons. Inhibitors of myogenic cell proliferation may play an important role in balancing the effects of positive growth factors.     

胰岛素联合利奈唑胺抑制糖尿病细菌性肺炎的机制研究

试验旨在研究胰岛素联合利奈唑胺是否同时具有降糖、抗菌以及抑制α-溶血素活性，从而抑制糖尿病细菌性肺炎。本研究通过体外试验探讨了联合用药对高糖和金黄色葡萄球菌α-溶血素共同诱导的巨噬细胞炎症反应的作用，并探讨其机制。通过药物敏感试验测定最小抑菌浓度（MIC）；通过检测D_{600 nm}值绘制生长曲线来考察胰岛素、利奈唑胺单独或联合胰岛素的抗菌活性；通过溶血试验考察药物组合对α-溶血素活性的抑制效果；通过细胞毒性试验判定药物组合的体外安全性；借助Western blotting药物组合对炎症通路蛋白的作用。药物敏感性试验结果显示，胰岛素组未检测到金黄色葡萄球菌8325-4株和DU1090株的MIC值，利奈唑胺和胰岛素+利奈唑胺药物组合对这两种金黄色葡萄球菌的MIC均为0.5 μg/mL；由生长曲线结果可知，胰岛素不抑制正常组和高糖组金黄色葡萄球菌的生长，但是在利奈唑胺组和胰岛素+利奈唑胺药物组合组，0.25~4 μg/mL利奈唑胺可抑制金黄色葡萄球菌的生长；溶血试验结果显示，正常组和高糖组胰岛素均不抑制α-溶血素活性，0.25~4 μg/mL利奈唑胺均可抑制α-溶血素活性；细胞毒性试验显示，正常组和高糖组，细胞存活率均大于88.038%，50 nmol/L胰岛素、0.25 μg/mL利奈唑胺、50 nmol/L胰岛素+0.25 μg/mL利奈唑胺药物组合可提高高糖诱导的MH-S细胞存活率；Western blotting结果显示，50 nmol/L胰岛素单独使用对高糖和α-溶血素共同诱导小鼠肺泡巨噬细胞（MH-S细胞）中TLR2、MAPKs及NLRP3蛋白的表达水平没有抑制作用，而50 nmol/L胰岛素联合0.25 μg/mL利奈唑胺可降低高糖和α-溶血素共同诱导MH-S细胞中上述蛋白的表达水平，其抑制作用比单独使用利奈唑胺时显著。综上所述，胰岛素联合利奈唑胺可通过降糖、抗菌和抑制α-溶血素活性的方式抑制小鼠肺泡巨噬细胞炎症反应。     

白藜芦醇对体外培养的水牛卵丘细胞增殖活力及其相关基因mRNA表达的影响

【目的】研究白藜芦醇(resveratrol, RES)对水牛卵丘细胞体外培养过程中细胞增殖活力、激素分泌、卵丘扩展及抗凋亡和抗氧化能力的影响。【方法】用不同浓度(0(对照组)、1、10、20、30、40、50和60μmol/L)RES培养卵丘细胞,用CCK-8试剂盒测定细胞增殖活力,筛选最佳RES处理浓度及时间用于后续试验。用ELISA法测定培养液中卵丘细胞分泌的雌二醇(estradiol, E₂)和孕酮(progesterone, P₄)的含量,实时荧光定量PCR法测定卵丘细胞扩展、凋亡和抗氧化相关基因的相对表达量。【结果】与对照组相比,在体外培养24～36 h内,1、10和20μmol/L RES组水牛卵丘细胞的增殖活力均有升高趋势,10μmol/L RES处理36 h细胞活力最强,因此用于后续试验中细胞的处理。体外培养36 h时,10μmol/L RES组细胞增殖能力和E₂、P₄的分泌显著增加(P<0.05);10μmol/L RES组卵丘细胞扩展相关基因穿透素3(PTX3)、前列腺素...     

紫花苜蓿叶总黄酮提取及抗氧化性

研究紫花苜蓿（Medicago sativa L.）叶总黄酮的最优提取工艺及体外抗氧化能力。在单因素试验的基础上设计正交试验,通过方差分析和多重比较确定最优提取工艺。以1,1-二苯基-2-三硝基苯肼（2,2-diphenyl-1-picrylhydrazyl,DPPH）清除率测定法及还原力测定法评价苜蓿黄酮类化合物体外抗氧化活性。最佳提取工艺条件是：乙醇体积分数70%、提取温度60℃、提取时间30 min、超声功率180 w,总黄酮得率为6.43 mg·g^-1。苜蓿叶总黄酮在一定的质量浓度范围具有较明显的抗氧化活性,并随着质量浓度的增加活性增强。苜蓿叶总黄酮具有较强的还原力,清除DPPH自由基的半数抑制质量浓度（IC₅₀值）为0.608 mg·mL^-1。超声提取是一种高效的提取紫花苜蓿叶总黄酮方法。超声优化的苜蓿叶总黄酮提取工艺经济、稳定、合理可行。     

双氢睾酮对小鼠卵泡颗粒细胞增殖与抗苗勒管激素表达的影响

【目的】探讨双氢睾酮(DHT)对小鼠颗粒细胞增殖与抗苗勒管激素(AMH)表达的影响。【方法】给3周龄的昆明小鼠注射孕马血清促性腺激素(PMSG,10 IU/只)以获取颗粒细胞,颗粒细胞传代后48 h HE染色鉴定形态,绘制颗粒细胞的生长曲线,免疫荧光法鉴定促卵泡素受体(FSHR)的表达。当第2代颗粒细胞汇合度达到50%时,先用无血清的DMEM/F12培养基饥饿处理12 h,然后在培养基中添加不同浓度的DHT (0、10^-9、10^-8、10^-7、10^-6、10^-5 mol/L),培养48 h后检测颗粒细胞增殖情况,实时荧光定量PCR法及ELISA法分别测定AMH基因及蛋白的表达。在颗粒细胞培养液中分别添加10^-6 mol/L Flutamide (雄激素受体(AR)特异性抑制剂)、10^-7 mol/L DHT、10^-7 mol/L DHT+10^-6 mol/L Flutamide、10^-5 mol/L DHT、10^-5mol/L DHT+10^-8 mol/L 11-ketodihydrotestosterone (AR特异性激动剂),分别记为F、D7、DF、D5、DK组,以不添加药物为对照组。培养48 h后,检测各组颗粒细胞增殖及AMH蛋白含量。【结果】体外培养的小鼠颗粒细胞呈梭形或铺路石状,生长曲线呈S形,细胞普遍表达FSHR;与0 mol/L DHT组相比,10^-8、10^-7 mol/L DHT组细胞增殖分别显著和极显著增加(P＜0.05;P＜0.01),10^-5 mol/L DHT组细胞增殖显著降低(P＜0.05);10^-7 mol/L DHT组AMH基因的相对表达量和AMH蛋白含量均极显著增加(P＜0.01)。与D7组相比,DF组颗粒细胞增殖和AMH蛋白含量均极显著降低(P＜0.01);与D5组相比,DK组颗粒细胞增殖和AMH蛋白含量均极显著升高(P＜0.01)。【结论】10^-7 mol/L DHT能够显著促进颗粒细胞增殖和AMH表达,而10^-5 mol/L显著降低了颗粒细胞增殖以及AMH表达,且AR介导了DHT调控颗粒细胞增殖和AMH表达。     

雌激素和孕酮对犬子宫内膜基质细胞增殖和孕酮受体表达的影响

本试验应用不同消化分离途径获取犬子宫内膜基质细胞,调节培养液中雌激素(E₂)和孕酮(P₄)的浓度,采用MTT法测定E₂和P₄浓度水平对犬子宫内膜基质细胞体外增殖的影响,利用细胞免疫组织化学法鉴定细胞并测定细胞孕酮受体(PR)表达与激素浓度水平的相关性。结果表明,E₂浓度变化(15、30、100 pg/mL)对犬子宫内膜基质细胞的增殖和PR的表达均没有显著的调节作用(P>0.05);P₄(15、30 ng/mL)对犬子宫内膜基质细胞的增殖有显著促进作用(P<0.05),P₄(3、15、30 ng/mL)对犬子宫内膜基质细胞PR的表达具有显著的抑制作用(P<0.05),其影响程度与浓度和作用时间关系密切。     

Effects of Estrogen and Progesterone on Canine Endometrial Stromal Cells Proliferation and Progesterone Receptor Expression

In this study different digestion and isolation methods were applied to obtain canine endometrial stromal cells (ESCs), different concentration levels were set for estrogen (E₂) and progesterone (P₄), then MTT method was used to measure their effect on cell proliferation in vitro, also cell immunohistochemistry was used for cell identification and measurement of effect on progesterone receptors (PR) expression.The results indicated that E₂ (15, 30 and 100 pg/mL) showed no significant regulations on both cell proliferation and PR expression, P₄ (15 and 30 ng/mL) had significant promoting effect on proliferation of ESCs (P<0.05), P₄ (3, 15 and 30 ng/mL) showed significant inhibitory effect on PR expression (P<0.05), the regulation level was related to concentration and acting time.     

高铜对大鼠肾细胞炎性因子和细胞增殖的影响

旨在探讨铜中毒对大鼠肾组织炎性因子的表达和细胞增殖功能的影响。本研究选用32只20日龄健康大鼠随机分为4组,每组8只,其中,对照组日粮的铜（Cu）浓度为15 mg·kg^-1;高铜Ⅰ组日粮Cu浓度为30 mg·kg^-1;高铜Ⅱ组日粮Cu浓度为60 mg·kg^-1;高铜Ⅲ组日粮Cu浓度为120 mg· kg^-1。连续饲喂6个月,检测肾组织Ki-67、PCNA、IL-1β、IL-2、IL-6、IL-18、NF-κB、TNF-α的mRNA及其蛋白的表达。随着日粮中铜含量增高,肾组织中Ki-67、PCNA mRNA及其蛋白表达量显著降低（P<0.05）。炎性因子IL-1β、IL-2、IL-6、IL-18、NF-κB、TNF-α mRNA的表达水平显著升高（P<0.05）,IL-1β蛋白的表达水平呈剂量依赖性上升,IL-6和IL-18蛋白表达水平先上升,后下降。结果表明,日粮铜含量高于30 mg·kg^-1时,将不同程度地抑制大鼠肾组织的细胞增殖,促进炎性因子的表达,造成炎性损伤。     

Effects of High Copper on Inflammatory Factors and Cell Proliferation in Rat Kidney Cells

The purpose of this research was to investigate the effect of copper poisoning on the expression of inflammatory factors and cell proliferation in renal tissues of rat. In this study, 32 healthy rats aged 20 days were selected and randomly divided into 4 groups, 8 in each group. The rats were fed as follows: The copper concentration in the control group was 15 mg·kg^-1; high-copper group I: 30 mg·kg^-1; high-copper group Ⅱ: 60 mg·kg^-1; high-copper group Ⅲ: 120 mg·kg^-1. After continuously feeding for 6 months, the mRNA and their proteins expression of Ki-67, PCNA, IL-1β, IL-2, IL-6, IL-18, NF-κB, TNF-α in renal tissues were detected. The results demonstrated that the relative expressions of Ki-67,PCNA mRNA and proteins in renal tissues were significantly decreased with the increase of copper content in feed (P<0.05). Meanwhile, the mRNA expression levels of inflammatory factors including IL-1β, IL-2, IL-6, IL-18, NF-κB, TNF-α in renal tissues were significantly increased (P<0.05). Besides, the relative expression of IL-1β protein was up-regulated in a dose dependent manner, and the relative expression levels of IL-6 and IL-18 proteins increased first and then decreased. In summary, the finding suggested that high copper stimulation inhibited the cell proliferation of rat kidney tissue, promoted the expression of inflammatory factors, and caused inflammatory damage.     

Pre-exposure of cattle to drug-abbreviated Fasciola hepatica infections: the effect upon subsequent challenge infection and the early immune response

In this study we examined whether juvenile liver flukes are capable of stimulating protective immune responses in cattle. Four experimental groups of cattle were studied as follows: group A, a positive control, received a primary infection on day 0 and a secondary infection 28 days later; group B also received two infections but the primary infection was terminated by drug treatment on day 5; group C, received infections on days 0, 5 and 10 which were terminated by drug treatments on days 1, 6 and 11 and then a secondary infection on day 28; group D received an infection only on day 28. Juvenile flukes appear to induce protective responses because: (a) group B animals had significantly lower levels of gamma-GT (P<0.05) than group D; (b) both groups B and C exhibited lower parenchymal phase GLDH levels (P=0.006 and 0.041, respectively); and (c) both groups B and C had lower secondary phase eosinophilia (P=0.002 and 0.02, respectively) than those in group D. Sera taken from groups A-C contained antibodies reacting to a variety of proteins in adult fluke somatic antigen and excretory-secretory preparations, particularly to proteins of 52-60, 68-72 and 82-96 kDa. After secondary challenge the antibody responses of group A to these proteins declined while reactivity to proteins of 28-30 kDa increased. Antibody responses to the 28-30 kDa proteins were not detected in groups B-D until 3 weeks later than those observed in group A. Antibody responses to Fasciola hepatica cathepsin L proteases, which are known to induce protection, were monophasic, of the IgG1 isotype only and were not observed prior to secondary challenge in any of the four groups. In contrast, the response to another protective antigen fraction, a high molecular sized haem protein, was of a mixed IgG1/IgG2 nature and was detected within 14 days of primary infection. However, no significant difference in antibody titres to either protein preparation was observed after the secondary infection when groups B and C were compared to group D.     

Effect of weaning at different ages on serum insulin-like growth factor I (IGF-I), IGF binding proteins and serum in vitro mitogenic activity in swine.

We investigated the effects of weaning or fasting of 21- or 35-d-old swine by monitoring serum mitogenic activity, circulating insulin-like growth factor I (IGF-I) and its binding proteins using L6 myoblast bioassays, RIA and ligand blotting techniques. Serum samples were collected from 21- or 35-d-old animals just before and 36 h after weaning or fasting. Sera from 21- and 35-d-old weaned animals were not significantly altered in their ability to promote myoblast proliferation, whereas sera from 21- and 35-d-old fasted animals caused 29 and 21% decreases (P less than .05) compared with preweaning. The mitogenic activity of control serum was inhibited by serum from fasted animals but not by preweaned or weaned sera. Serum IGF-I levels were decreased 65 to 70% (P less than .05) with weaning or fasting at both ages. Unoccupied binding sites on circulating IGF binding proteins in the 155 kDa range decreased 18 to 19% with weaning at both ages and decreased 40% (P less than .05) with fasting at 21 d but only 17% at 35 d. Ligand blotting revealed that the 43 and 39 kDa IGF binding protein bands decreased with weaning and fasting at both ages, whereas the 29-kDa band increased with weaning and fasting. These data indicate that serum IGF-I and specific IGF binding protein bands decrease during weaning or fasting at 21 and 35 d of age. However, serum mitogenic activity did not always follow serum IGF-I levels.     

Expression and characterization of the recombinant swine interleukin-6

The swine interleukin-6 (SwIL-6) cDNA was cloned by RT-PCR and each expression system of recombinant SwIL-6 in Escherichia coli, insect cells, and mammalian cells was developed. Recombinant SwIL-6 produced in bacteria was applied for generation of the polyclonal antibodies. The rSwIL-6 was purified from supernatant of insect cells with a Q-sepharose or anti-SwIL-6 monoclonal antibody based immunoaffinity column. The antibodies showed that the molecular weight of rSwIL-6 was approximately 26kDa in E. coli, 25, 26, 30kDa in insect cells, and 26 and 30kDa in mammalian cells. These variations of molecular weight were probably due to the different modifications of glycosylation. All these recombinant proteins retained the antigenicity and biological activity on 7TD1 mouse cells.     

Growth-promoting effects of different fractions of extra-embryonic coelomic fluid on embryonic development

In the early stages of embryonic development, many growth-promoting molecules must be provided by the maternal system. These factors may be supplied locally to the embryo, by the decidua, the placenta, or the yolk sac. In this study the growth-promoting potential of extra-embryonic coelomic fluid (EECF) and its fractions was investigated. The embryonic requirement of growth-promoting molecules may be studied by reducing the growth-supporting capacity of serum. Thus, ultrafiltration of rat serum was carried out for 8 h using Millipore filters with a molecular weight exclusion of 30 kDa. Rat embryos at 9.5 days of age were cultured for 8 days for anembryonic yolk sacs, and then EECF was collected and divided into three different molecular weight fractions by ultrafiltration. Rat embryos were cultured for 48 h in whole rat serum and the serum retenate (which has low growth-supporting capacity) in the presence and absence of EECF, its fractions, or in EECF only. Embryos grown in retenate showed severe growth retardation, and the addition of EECF significantly improved embryonic growth. The fraction which contained the molecules with molecular weight between 10 and 30 kDa had significantly more effect on embryonic development than the other fractions. This fraction of EECF was analysed by gel electrophoresis. Three of the four protein bands observed in this fraction were identified by amino-terminal sequencing as alpha-fetoprotein precursor (22 kDa), apolipoprotein A1 precursor (24 kDa) and fetal haemoglobin Y2 chain (14 kDa), none of which are likely to be responsible for the growth-promoting activity. To further investigate growth-promoting proteins, EECF was Western-blotted to nitrocellulose membranes and probed with antisera against rat prolactin, epidermal growth factor, insulin-like growth factors I and II and human placental lactogen. No immunoreactive bands were detected in the EECF, suggesting that either these proteins are not present or are present at levels too low to be detected. Although the growth-promoting effect of the EECF was demonstrated in this study, the molecules responsible remain uncharacterized.     

NO自由基对猪圆环病毒2型复制的体外抑制作用研究

试验旨在探讨NO自由基(NO·)对猪圆环病毒2型(PCV2)复制的抑制作用。硝普钠(SNP)和抗坏血酸(VC)联合加入培养基中可产生NO·,而单独使用SNP产生NO的形式为NO^+。本研究采用MTT法测定药物的细胞毒性,通过Griess反应测定供体药物产生NO的水平。分别用SNP、VC、SNP+VC和对照药物乙酰青酶胺(NAP)处理PK-15细胞,6 h后接种1 MOI的PCV2病毒液,72 h后收获培养上清和细胞,通过间接免疫荧光试验(IFA)、Western blotting分析、病毒效价和实时荧光定量PCR试验检测病毒增殖水平。结果显示:SNP、VC和SNP+VC对PK-15细胞的最大安全浓度分别为500、62.5和31.25μmol/L;60μmol/L SNP和30μmol/L SNP+30μmol/L VC两种供体产生的NO量极显著高于非药物处理组和对照药物NAP处理组(P<0.01);与阳性对照组相比,60μmol/L SNP+30μmol/L VC处理组中的PCV2毒价和DNA拷贝数极显著下降(P<0.01),且PCV2 Cap蛋白的表达也受到明显抑制,而SNP和VC单独处理组上述指标无显著变化(P>0.05)。以上结果表明,SNP+VC产生的NO·对PCV2增殖有显著抑制作用,SNP产生的NO^+不影响PCV2增殖,NO体外抑制PCV2增殖主要是通过NO·实现。     

An AC-5 cathepsin B-like protease purified from Haemonchus contortus excretory secretory products shows protective antigen potential for lambs

The immunogenic properties of cysteine proteases obtained from excretory/secretory products (ES) of Haemonchus contortus were investigated with a fraction purified with a recombinant H. contortus cystatin affinity column. The enrichment of H. contortus ES for cysteine protease was confirmed with substrate SDS-PAGE gels since the cystatin-binding fraction activity was three times higher than total ES, despite representing only 3% of total ES. This activity was inhibited by a specific cysteine protease inhibitor (E64) and by recombinant cystatin. The one-dimensional profile of the cystatin-binding fraction displayed a single band with a molecular mass of 43 kDa. Mass spectrometry showed this to be AC-5, a cathepsin B-like cysteine protease which had not been identified in ES products of H. contortus before. The cystatin binding fraction was tested as an immunogen in lambs which were vaccinated three times (week 0, 2.5 and 5), challenged with 10 000 L3 H. contortus (week 6) before necropsy and compared to unvaccinated challenge controls and another group given total ES (n = 10 per group). The group vaccinated with cystatin-binding proteins showed 36% and 32% mean worm burden and eggs per gram of faeces (EPG) reductions, respectively, compared to the controls but total ES was almost without effect. After challenge the cystatin-binding proteins induced significantly higher local and systemic ES specific IgA and IgG responses.     

泛素激活酶抑制剂对猪精子获能状态、膜重构及体外受精的影响

【目的】评估泛素激活酶(E1)对猪精子获能、膜重构和体外受精的影响。【方法】选取5头长白猪采集精液,以硫醇酯法评估泛素激活酶(ubiquitin activating enzyme, E1)抑制剂(TAK-243)对精子中泛素(ubiquitin, Ub)与E1结合的影响;将精子分为鲜精组(Fresh)、DMSO组(DMSO)、获能组(Capacitated)及2.4、4.8和7.2 nmol/L TAK-243组,鲜精组用2 mL PBS重悬精子沉淀;获能组用2 mL获能液重悬精子沉淀;DMSO组及2.4、4.8和7.2 nmol/L TAK-243组分别用2 mL含0.07%DMSO及2.4、4.8和7.2 nmol/L TAK-243的获能液重悬精子沉淀,使用微生物动(静)态图像检测系统检测精子的动力学参数;以Western blotting法检测精子的酪氨酸磷酸化水平;Zn²⁺荧光探针检测精子的Zn²⁺含量;花生凝集素染色检测精子的顶体膜重构;免疫荧光法检测顶体表面精子黏附蛋白(AQN-1、AWN)的表达;体外受精法检测TAK-243...     

Identification and partial characterization of the hemolysin (HlyII) of Actinobacillus pleuropneumoniae serotype 2

The secreted hemolytic activity produced by Actinobacillus pleuropneumoniae serotype 2 reference strain is thermolabile, inactivated by proteinase K and requires Ca2+ as cofactor for its hemolytic activity. Purification of the hemolytic activity resulted in a fraction containing two proteins, one of 105 kDa and one of 125 kDa. These two proteins could be further separated by preparative SDS polyacrylamide gel electrophoresis. This purification step, resulted in loss of the hemolytic activity. Polyclonal antibodies were made against each of these proteins in rabbits. Neutralization experiments showed that antibodies made against the 105 kDa protein could neutralize the hemolytic activity produced by A. pleuropneumoniae serotype 2, while antibodies made against the 125 kDa protein were unable to neutralize the hemolytic activity. The 105 kDa protein therefore, is the hemolysin of A. pleuropneumoniae serotype 2, known as HlyII. This protein is closely related immunologically to the hemolysin I (HlyI) from A. pleuropneumoniae serotype 1. DNA::DNA hybridization experiments performed by the Southern blot method using the cloned structural gene of HlyI from A. pleuropneumoniae serotype 1 demonstrate that the structural genes of the two hemolysins (hlyIA and hlyIIA) are different and show at least 30% heterology. This confirms that HlyI and HlyII are two different proteins, although they have a very similar molecular weight and show strong immunological cross reactions.     

蚕蛹醇提物萃取组分对α-葡萄糖苷酶的抑制动力学研究

为了有效利用缫丝蚕蛹的功能性成分开发高附加值的降血糖药物,采用95%乙醇浸提蚕蛹活性物质,并依次选用石油醚、乙醚、乙酸乙酯、水饱和正丁醇对蚕蛹醇提物进行梯度极性溶剂萃取,获得不同萃取组分及水相组分。测定各相萃取组分对α-葡萄糖苷酶的体外抑制活性,结果显示:当石油醚、乙醚、乙酸乙酯、正丁醇萃取组分及水相组分的质量浓度为0.33mg/mL时,各相萃取组分对α-葡萄糖苷酶的体外抑制率分别为100%、58%、38.1%、36.93%、21.53%;各相萃取组分对α-葡萄糖苷酶的半数抑制浓度(IC50)分别为0.049、0.272、0.434、0.645、0.449 mg/mL。选择对α-葡萄糖苷酶抑制活性较强的石油醚相和乙醚相萃取组分,采用双倒数作图法检测其抑制作用类型均属于非竞争性抑制。     

Identification and partial purification of serologically defined Boophilus microplus larval antigens by natural ectoparasite exposure

In an effort to identify life-stage specific Boophilus microplus proteins that elicit a humoral response in cattle, soluble proteins were extracted from 10- to 14-day-old larvae and subsequently fractionated by size-exclusion chromatography and reverse-phase high pressure liquid chromatography. Several antigens were identified by Western blotting as potentially shared with other ixodid tick species since antibodies to these proteins were present in sera of calves not previously exposed to B. microplus. Six putative B. microplus-specific antigens were identified by antibodies in the sera of calves repeatedly exposed to B. microplus larvae. One of the antigens, a 19.1 kDa protein, was used in the development of a diagnostic kELISA for previous exposure to B. microplus. The 19.1 kDa protein did not have tryptic protease activity or inhibit bovine trypsin activity, but appeared to be allergenic in that a partially pure fraction elicited immediate-type hypersensitivity responses in calves previously exposed to B. microplus.     

模拟胃肠道消化过程中活性物质对牛乳的降脂作用

以牛乳为材料，研究模拟胃肠道消化过程中活性物质对牛乳中蛋白质体外消化率、抑制胰脂酶活性及胆固醇胶束溶解度的影响。结果表明：牛乳蛋白质体外消化率与活性物质添加量成反比，随表没食子儿茶素（epigallocatechin，EGC）添加量增加，蛋白质体外消化率从90.94%降至68.37%，随β-葡聚糖添加量的增加，蛋白质消化率从90.94%降至86.00%；当体系中加入乳蛋白时，随着乳蛋白添加量的增大，EGC对胰脂酶活性的抑制率有不同程度下降，β-葡聚糖及功能性成分混合液（β-葡聚糖2.5 g/L、表没食子儿茶素没食子酸酯 3.0 g/L、EGC 2.0 g/L）对胆固醇胶束溶解度的抑制率下降。活性物质对牛乳降脂作用的影响为功能成分混合液＞β-葡聚糖。