高产磷酯酶C菌株筛选及其抗血小板功能的研究

在湖北省、湖南省、江苏省、山东省、广东省156个点采土样1268份,利用卵黄琼脂的LB培养基初筛获得产生乳白色晕圈的菌株648株;再用卵黄琼脂滤纸圆片检测获得产生乳白色晕圈和透明圈菌株266株;这些菌株用卵黄琼脂杯蝶法筛选出产生大于13．0mm乳白色晕圈及少量透明圈共73株,经过菌生物量、卵黄杯碟法和抗兔血小板聚集率的测定,从73株菌中筛选出15株,菌号为：183,198,247,424,587,596,692,744,754—1,791—2,779,970,998,1107,1182;将此15菌株经菌生物量、卵黄琼脂杯碟法、NPPC法和抗人血小板聚集率的测定,从中选出754—1,779,970,1107四株菌进行分类鉴定和已知菌CW—W—90—3菌对照比较的研究。     

Influence of processing factors on the stability of model mayonnaise with whole egg during long-term storage

Mayonnaise-like oil-in-water emulsions with different stabilities—evaluated from the degree of macroscopic defects, e.g., syneresis—were prepared by different formulations and processing conditions (egg yolk weight, homogenizer speed, and vegetable oil temperature). Emulsions prepared with lower egg yolk content were destabilized for shorter periods. The long-term stability of emulsions was weakly related to initial properties, e.g., oil droplet distribution and protein coverage at the interface. Protein aggregation between oil droplets was observed and would be responsible for the instability of emulsions exhibited by the appearance defects. SDS-PAGE results for adsorbed and unadsorbed proteins at the O/W interface suggested that predominant constituents adsorbed onto the interface were egg white proteins as compared with egg yolk components when the amount of added egg yolk was low. In present condition, egg white proteins adsorbed at the O/W interface could be a bridge of neighboring oil droplets thereby causing flocculation in emulsions.     

The chicken egg yolk plasma and granule proteomes

Using 1-D SDS-PAGE, LC-MS/MS, and MS(3), we identified 119 proteins from chicken egg yolk, 86 of which were not identified in yolk previously. Proteins were roughly quantitated by calculating their exponentially modified protein abundance index (emPAI) to classify them as major or minor yolk components, and to estimate their distribution between yolk plasma and yolk granular fraction. The proteins with highest abundance were serum albumin, the vitellogenin cleavage products, apovitellenins, IgY, ovalbumin, and 12 kDa serum protein with cross-reactivity to beta2-microglobulin. In addition yolk contained many other serum and egg white proteins, the proteases nothepsin and thrombin, numerous protease inhibitors, and antioxidative enzymes, such as superoxide dismutase and glutathione peroxidase. Among the moderately abundant proteins were two alpha2-macroglobulin-like proteins different from egg white alpha2-macroglobulin, and the major biotin-binding protein of yolk. An unexpected identification was that of the eggshell matrix protein ovocleidin-116, which was previously thought to be eggshell-specific. The list of chicken egg yolk proteins provided in this report is by far the most comprehensive at present and may serve as a starting point for the characterization of less well-known yolk proteins.     

Is Aboriginal Food Less Allergenic? Comparing IgE-Reactivity of Eggs from Modern and Ancient Chicken Breeds in a Cohort of Allergic Children

Background

Hen''s egg allergy ranks among the most frequent primary food allergies in children. We aimed to investigate sensitization profiles of egg allergic patients and compare in vitro IgE reactivities of eggs from ancient chicken breeds (Araucana and Maran) with those from conventional laying hen hybrids.

Methodology

Egg allergic children (n = 25) were subjected to skin prick test, double blind placebo controlled food challenge, and sensitization profiles to Gal d 1–5 were determined by allergen microarray. IgE binding and biological activity of eggs from different chicken breeds were investigated by immunoblot, ELISA, and mediator release assays.

Principal Findings

We found that Gal d 1 and Gal d 2 are generally major egg allergens, whereas Gal d 3–5 displayed high sensitization prevalence only in patients reacting to both, egg white and yolk. It seems that the onset of egg allergy is mediated by egg white allergens expanding to yolk sensitization in later stages of disease. Of note, egg white/yolk weight ratios were reduced in eggs from Auraucana and Maran chicken. As determined in IgE immunoblots and mass analysis, eggs from ancient chicken breeds did not differ in their protein composition. Similar IgE-binding was observed for all egg white preparations, while an elevated allergenicity was detected in egg yolk from Araucana chicken.

Conclusion/Significance

Our results on allergenicity and biological activity do not confirm the common assumption that aboriginal food might be less allergenic. Comprehensive diagnosis of egg allergy should distinguish between reactivity to hen''s egg white and yolk fractions to avoid unnecessary dietary restrictions to improve life quality of the allergic child and its family.     

Simultaneous determination of nine fluoroquinolones in egg white and egg yolk by liquid chromatography with fluorescence detection

A liquid chromatographic (LC) method with fluorescence detection was developed for determination of nine fluoroquinolones (FQs) in egg white and yolk. Egg white samples were deproteinized with acidified ethanol (egg yolk samples with acetonitrile and acidified ethanol), followed by defatting with hexane once (white) or twice (yolk), and extracting FQs into acetonitrile. After acetonitrile was evaporated, the residue was dissolved in mobile phase, and FQs were detected in LC with a fluorescence detector. Recoveries for nine FQs from white and yolk were 74.7-85.6%, 79.1-91.2%, respectively, with excellent relative standard deviations. The limits of quantification were 5-20 ngg(-1).     

The egg white and yolk interactomes as gleaned from extensive proteomic data

Combinatorial peptide ligand libraries have recently allowed considerable advances in the mapping of chicken egg yolk and white proteomics. Data from literature have been regrouped and elaborated for network and pathway analyses in order to convey a unified view of these proteomes. Redundant proteins were excluded, while isoforms of the same proteins were maintained to reach a total of 260 distinct gene products for egg yolk and 148 for egg white having a match in the database. From these analyses, a role for proteins involved in cell development, proliferation and migration, cell-to-cell interaction and hematological system development emerged. Although it might turn out that, notwithstanding the extensive mapping, the currently available datasets might be still incomplete, a valuable insight could still be obtained about specific proteins playing a crucial role in antimicrobial responses, mainly histones, lysozyme and vitamin-binding proteins. In particular, SERPINB3 (ovalbumin Y, or Squamous Cell Carcinoma Antigen, SCCA1) was individuated in 8 out of 10 top score pathways in egg yolk and in 6 out 10 in egg white. SERPINB3 is a member of the ov-serpin family, participating in coagulation and inflammation responses. However, it is yet to be assessed how these observations could correlate with previous analyses about the role of egg yolk derived proteins in counteracting blood coagulation.     

Heat Resistance of Salmonella in Various Egg Products

The heat-resistance characteristics of Salmonella typhimurium Tm-1, a reference strain in the stationary phase of growth, were determined at several temperatures in the major types of products produced by the egg industry. The time required to kill 90% of the population (D value) at a given temperature in specific egg products was as follows: at 60 C (140 F), D = 0.27 min for whole egg; D = 0.60 min for whole egg plus 10% sucrose; D = 1.0 min for fortified whole egg; D = 0.20 min for egg white (pH 7.3), stabilized with aluminum; D = 0.40 min for egg yolk; D = 4.0 min for egg yolk plus 10% sucrose; D = 5.1 min for egg yolk plus 10% NaCl; D = 1.0 min for scrambled egg mix; at 55 C (131 F), D = 0.55 min for egg white (pH 9.2); D = 1.2 min for egg white (pH 9.2) plus 10% sucrose. The average Z value (number of degrees, either centigrade or fahrenheit, for a thermal destruction time curve to traverse one logarithmic cycle) was 4.6 C (8.3 F) with a range from 4.2 to 5.3 C. Supplementation with 10% sucrose appeared to have a severalfold greater effect on the heat stabilization of egg white proteins than on S. typhimurium Tm-1. This information should be of value in the formulation of heat treatments to insure that all egg products be free of viable salmonellae.     

Egg white hydrolysate inhibits oxidation in mayonnaise and a model system

The flavor deterioration of mayonnaise is induced by iron, which is released from egg yolk phosvitin under acidic conditions and promotes lipid oxidation. To prevent oxidative deterioration, natural components, rather than synthetic chemicals such as ethylenediaminetetraacetic acid have been required by consumers. In the present study, we evaluated the inhibitory effects of three egg white components with the same amino acid composition, namely egg white protein, hydrolysate, and the amino acid mixture, on lipid oxidation in mayonnaise and an acidic egg yolk solution as a model system. We found that the hydrolysate had the strongest inhibitory effect on lipid oxidation among the three components. The mechanism underlying the antioxidant effect was associated with Fe²⁺-chelating activity. Thus, egg white hydrolysate may have the potential as natural inhibitors of lipid oxidation in mayonnaise.     

Heterogeneity of serum, egg amylase and egg lysozyme in the interspecific hybrids of ducks

Polymorphism of serum and egg amylase by means of horizontal agarose gel electrophoresis and egg lysozyme by means of horizontal starch gel electrophoresis in Pekin, Muscovy ducks and their interspecific hybrids was studied. In the interspecific hybrids of ducks the codominant type of heredity of serum, egg yolk and egg white amylase isozymes, as well as egg white lysozyme, were found.     

Absorption, transportation and digestion of egg white in quail embryos

The present study was done to reveal how egg white is taken up by embryonic tissues, the pathway through which egg white is transported, and the location where it is digested during the development of the quail Coturnix japonica. Antiserum against quail ovalbumin was raised in rabbit and used as a probe. By immunoelectron microscopy, the uptake of ovalbumin on a small scale by receptor-mediated endocytosis was observed in the ectodermal cells of the yolk sac on days four to seven of incubation. The uptake of egg white on a large scale by fluid-phase endocytosis took place in the cells generally referred to collectively as the 'albumen sac'. The ovalbumin was transported through the albumen sac into the extraembryonic cavity during days eight to 10, and then into the amniotic cavity through the amnion approximately on day 10. Ovalbumin was present in the intestinal lumen on days 11 and 14, but it was not digested in the intestinal epithelial cells. The ovalbumin was detected in the yolk of embryos after day 10. Immunoblot testing, as well as a fluoroimmunoassay, revealed that the location where the amount of ovalbumin was highest changed chronologically from the extraembryonic cavity on day 10 to the amniotic cavity on day 11, the intestinal lumen on day 12 and then to the yolk on day 13. Several low molecular proteins which cross-reacted with the antiserum were observed in the extracts of the yolk. The reaction producing these proteins depended on low pH (approximately 3.0) and was inhibited by pepstatin A. The ovotransferrin was similarly digested. These results indicate that egg white is, for the most part, transported through the albumen sac to the yolk via the extraembryonic cavity, the amniotic cavity, and the intestinal lumen, and is digested in the yolk by aspartic proteinases.     

OSMOTIC RELATIONSHIPS IN THE HEN'S EGG,AS DETERMINED BY COLLIGATIVE PROPERTIES OF YOLK AND WHITE

The osmotic pressure of the yolk and white of the hen''s egg have been shown to be identical, by means of direct freezing point determinations, dialyses, and vapor pressure measurements. Dialysates of egg yolk slow the rate of ice formation compared with NaCl solutions. They also show a marked change of freezing rate as the freezing point is approached. The anomalous freezing behavior of this material may lead to errors in the determination of the true freezing point which would tend to make the value for the yolk erroneously low. The postulate of a vital activity at the yolk membrane maintaining an osmotic pressure difference is thus shown to be unnecessary, since a simple osmotic equilibrium exists between the yolk and the white.     

应用医疗CT扫描法观察恐龙蛋化石的内部结构

应用医疗ＣＴ扫描法观察恐龙蛋化石，可清晰地分辨出蛋形、蛋壳、卵蛋白、蛋黄及胚盘等结构，并可测出各部分具体数据，为恐龙蛋化石和其它生物化石的研究开辟了一个新的途径。     

Production of recombinant human erythropoietin/Fc fusion protein by genetically manipulated chickens

We previously reported the production of human erythropoietin (hEpo) using genetically manipulated (GM) chickens. The recombinant hEpo was produced in the serum and egg white of the GM chickens, and the oligosaccharide chain structures of the serum-derived hEpo were more favorable than those of the egg white-derived hEpo. In the present study, a retroviral vector encoding an expression cassette for a fusion protein of hEpo and the Fc region of human immunoglobulin G (hEpo/Fc) was injected into developing chicken embryos, with the aim of recovering the serum-derived hEpo from egg yolk through the yolk accumulation mechanism of maternal antibodies. The GM chickens that hatched stably produced the hEpo/Fc fusion protein not only in their serum and egg white, but also in the egg yolk as expected. Lectin blot analyses revealed that significant amounts of the oligosaccharide chains of hEpo/Fc produced in the serum and eggs of GM chickens terminated with galactose, and that the oligosaccharide chains of the serum- and yolk-derived hEpo/Fc incorporated sialic acid residues. Moreover, biological activity assessment using Epo-dependent cells revealed that the yolk-derived hEpo/Fc exhibited a comparable performance to the serum- and CHO-derived hEpo/Fc. These results indicate that transport of Fc fusion proteins from the blood circulation to the yolk in chickens represents an effective strategy for the production of pharmaceutical glycoproteins using transgenic chicken bioreactors.     

Availability of avidin-bound biotin to the chicken embryo.

Avidin, an exceptionally stable protein in egg white, binds the vitamin biotin with very high affinity and can induce biotin deficiency when fed to animals. To determine if biotin bound to avidin is available to the chicken embryo, the fate of [3H]biotin complexed to avidin was monitored during embryonic development. The majority (greater than 85%) of the [3H]biotin was extraembryonic until the day before hatching, when embryos swallow egg white and withdraw the yolk sac into their abdomen. Thus, biotin in the egg white of chicken eggs contributes little to the biotin status of the chick prior to hatching. After hatching, much of the [3H]biotin was assimilated. About 30% of the total was found in the liver and kidneys by 4 days of age. The biotin in liver was associated with large proteins and not with avidin. In a separate experiment, biotin injected into the egg white of biotin-deficient eggs failed to increase embryonic development or hatchability. Both experiments suggest that biotin in egg yolk is the primary and virtually sole source of biotin for the chicken embryo.     

Elevated plasma corticosterone decreases yolk testosterone and progesterone in chickens: linking maternal stress and hormone-mediated maternal effects

Despite considerable research on hormone-mediated maternal effects in birds, the underlying physiology remains poorly understood. This study investigated a potential regulation mechanism for differential accumulation of gonadal hormones in bird eggs. Across vertebrates, glucocorticoids can suppress reproduction by downregulating gonadal hormones. Using the chicken as a model species, we therefore tested whether elevated levels of plasma corticosterone in female birds influence the production of gonadal steroids by the ovarian follicles and thus the amount of reproductive hormones in the egg yolk. Adult laying hens of two different strains (ISA brown and white Leghorn) were implanted subcutaneously with corticosterone pellets that elevated plasma corticosterone concentrations over a period of nine days. Steroid hormones were subsequently quantified in plasma and yolk. Corticosterone-implanted hens of both strains had lower plasma progesterone and testosterone levels and their yolks contained less progesterone and testosterone. The treatment also reduced egg and yolk mass. Plasma estrogen concentrations decreased in white Leghorns only whereas in both strains yolk estrogens were unaffected. Our results demonstrate for the first time that maternal plasma corticosterone levels influence reproductive hormone concentrations in the yolk. Maternal corticosterone could therefore mediate environmentally induced changes in yolk gonadal hormone concentrations. In addition, stressful situations experienced by the bird mother might affect the offspring via reduced amounts of reproductive hormones present in the egg as well as available nutrients for the embryo.     

Carbohydrate compositional effects on tissue distribution of chicken riboflavin-binding protein

Riboflavin-binding proteins (RBP) purified from chicken egg white, yolk and the serum of laying hens differ in their carbohydrate compositions reflecting tissue-specific modifications of a single gene product. All three are complex glycoproteins having more than twice as many N-acetylglucosamine residues (>12) as mannose residues (approx. 6). Egg white RBP is distinctive in having only one sialic acid and two galactose residues. Serum RBP contains approx. five sialic acid and seven galactose residues. In addition there is one residue of fucose. The carbohydrate composition of yolk RBP indicates the hydrolysis, respectively, of one, one, two and 3 residues of sialic acid, fucose, galactose, and N-acetylglucosamine from its precursor, serum RBP. The effect of these differing levels of glycosylation on plasma clearance, ovarian uptake and tissue distribution of ¹²⁵I-labeled riboflavin-binding proteins in laying hens were compared. 2 h after intravenous injection, 19% of the egg white RBP, 29% of the yolk RBP, and 37% of the serum RBP remained in circulation. The kinetics of plasma clearance was distinctly biphasic for each of the radioiodinated proteins. The initial rapid-turnover component ($\text{t}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}\text{=13}\text{min}$) ranged from 27% of the serum RBP sample to 48% of the egg white RBP sample. The remaining slow-turnover components were cleared with half-lives fo 81 min (egg white RBP), 101 min (yokl RBP), and 121 min (serum RBP). 16 h after injection, only 4% of the egg white RBP was deposited in the yolk of developing oocytes while about 12% of the serum RBP and yolk RBP was deposited. This higly significant difference is apparently due to preferential, carbohydrate-dependent clearance of egg white RBP by the liver rather than preferential uptake of serum and yolk RBP by the ovarian follicle. We find no evidence for carbohydrate-directed uptake of riboflavin-binding protein by the ovarian follicle.     

Beneficial Effects of Simulated Gastro-Intestinal Digests of Fried Egg and Its Fractions on Blood Pressure,Plasma Lipids and Oxidative Stress in Spontaneously Hypertensive Rats

Background

We have previously characterized several antihypertensive peptides in simulated digests of cooked eggs and showed blood pressure lowering property of fried whole egg digest. However, the long-term effects of this hydrolysate and its fractions on blood pressure are not known. Therefore, the objectives of the study were to determine the effects of long term administration of fried whole egg hydrolysate and its fractions (i.e. egg white and egg yolk) on regulation of blood pressure and associated factors in cardiovascular disease such as plasma lipid profile and tissue oxidative stress.

Methods and Results

We used spontaneously hypertensive rats (SHR), an animal model of essential hypertension. Hydrolysates of fried egg and its fractions were prepared by simulated gastro-intestinal digestion with pepsin and pancreatin. 16–17 week old male SHRs were orally administered fried whole egg hydrolysate, non-hydrolyzed fried whole egg, egg white hydrolysate or egg yolk hydrolysates (either defatted, or not) daily for 18 days. Blood pressure (BP) and heart rate were monitored by telemetry. Animals were sacrificed at the end of the treatment for vascular function studies and evaluating plasma lipid profile and tissue oxidative stress. BP was reduced by feeding fried whole egg hydrolysate but not by the non-hydrolyzed product suggesting a critical role for in vitro digestion in releasing anti-hypertensive peptides. Egg white hydrolysate and defatted egg yolk hydrolysate (but not egg yolk hydrolysate) also had similar effects. Reduction in BP was accompanied by the restoration of nitric oxide (NO) dependent vasorelaxation and reduction of plasma angiotensin II. Fried whole egg hydrolysate also reduced plasma levels of triglyceride although it was increased by the non-hydrolyzed sample. Additionally the hydrolyzed preparations attenuated tissue oxidative stress.

Conclusion

Our results demonstrate that fried egg hydrolysates exert anti-hypertensive effects, improve plasma lipid profile and attenuate tissue oxidative stress in vivo.     

Effect of gravity on the interaction between the avian germ and neighbouring ooplasm in inverted egg yolk balls

The developmental capacities of an avian germ (from before symmetrization to the moment of laying) are strongly diminished after inversion of its egg yolk ball followed by culture in egg white. Our present experiments show that even when the avian germ is completely horizontally inverted (without an upper or lower border) below its egg yolk ball before symmetrization, symmetrization and gastrulation phenomena take place. The germ grows slower and becomes smaller than after normal incubation. After culture of inverted unincubated germs, localized on freshly laid eggs, the closure of the neural tube is impaired and it remains open over a long distance. Although a primitive streak (PS) develops, mesoderm migration (mainly from the lateral part of the area pellucida) is also impaired. On sections through the germinal disc one can see the abnormal upward migration into the depth of the ooplasm and yolk of cells from the germ wall and the development of large cellular extensions encircling the yolk globules. Most prominent is the loss of contact between the superficial cell layers and the deep layer elements (junctional endoblast and yolk endoblast in the area opaca). Large areas without deep layer elements (even visible on surface micrographs) develop in the area vasculosa and area vitellina interna. The margin of overgrowth grows and extends normally over the egg yolk ball. An autoradiographic study after labelling of the yolk layers in inverted egg yolks reveals that mainly compression of the peripheral subgerminal and perigerminal ooplasm takes place. This suggests that the compression by the neighbouring yolk and upwards growth of cells are at the origin of the impaired development. After return to the normal upward orientation of the germ on the topmost part of the egg yolk ball, a more or less pronounced restoration to normal development takes place (depending on the duration of the inversion period and the age of the germ).     

OSMOTIC RELATIONSHIPS IN THE HEN'S EGG

Data have been given to illustrate the difficulty of obtaining consistent freezing point data with a viscous fluid such as the yolk of the hen''s egg and a technique has been described for obtaining reproducible and accurate results consistently. Further freezing point data have been given which were obtained with both fertile and unfertile hen''s eggs by the use of a freezing point method previously described by the writer. These data show that there is a pronounced difference between the freezing points of the yolk and the white in contrast to data obtained by the use of the same method by Howard who found the freezing points of the yolk and the white to be the same. It was shown by freezing point determinations that even in a mixture of yolk and white osmotic equilibrium is slowly arrived at. This again emphasizes the fact established by Smith and Shepherd that since osmotic equilibrium between yolk and white is slowly arrived at, the postulation of a vital activity at the yolk membrane is unnecessary, since the steady state previously postulated need not be assumed to exist.     

Flow of egg white ovalbumin into the yolk sac during embryogenesis

Western blot analyses of yolk proteins of the White Leghorn hens showed that an ovalbumin-like molecule was included in day 16 eggs but not in the ovarian follicles, and very little in newly deposited eggs. Northern hybridization, as well as in vitro translation, of poly(A)+ RNAs prepared from the yolk sac membranes of developing embryos gave no signal for ovalbumin messages. These results imply that the present ovalbumin-like protein of yolk has its origin in egg white, not being a de novo synthesis product in the yolk sac membranes.