Review Yoshinori Mine and Jennifer Kovacs-Nolan

Journal of Poultry Science, .+ : +῎,3, ,**. ῌReview῍

Yoshinori Mine and Jennifer Kovacs-Nolan

Department of Food Science, University of Guelph Guelph, Ontario N+G ,W+, Canada

It is widely recognized that eggs are more than a source of dietary nutrients, and extensive studies identifying and characterizing the biologically active components of eggs have been carried out. Numerous biological activities have now been associated with egg components, including antibacterial and antiviral activity, immunomodulatory activity, and anti-cancer activity, indicating the importance of eggs and egg components in human health, and disease prevention and treatment. The potential of some of these biologically active components has already been realized, including egg white lysozyme and avidin, and yolk IgY and lecithin, which are currently produced on an industrial scale, and have been applied for the prevention and treatment of various medical conditions. The information presented here serves to demonstrate the signiﬁcant potential of biologically active egg components, for medical, nutraceutical, and food- fortiﬁcation applications.

Key words : Hen eggs, bio-active components, nutraceuticals, health and disease, functional foods

ῌῌῌῌῌῌῌῌῌῌῌῌῌῌῌῌῌῌῌῌῌῌῌ

Yoshinori Mine +321 : M. Sc. (Food Science) Shinshu University, Japan +33- : Ph. D. (Biochemistry) Tokyo University of Agriculture and Technology, Japan Current position Associate Professor and Chair in Egg Material Sciences Department of Food Science University of Guelph. Area of research interests : Egg Material Sciences, Bio-active protein/peptides, Molecular biology of egg allergens.

Jennifer Kovacs-Nolan ,**+ : M. Sc. (Food Science) University of Guelph , Ontario, Canada. Current position : Ph. D. student, Department of Food Science, University of Guelph, Ontario, Canada. Area of research interests : Food biochemistry/biotechnology. Bio-active components in eggs.

Received : October ,+, ,**-, Accepted : November +3, ,**- Corresponding author : Dr. Yoshinori Mine Department of Food Science, University of Guelph, Guelph, Ontario N+G ,W+, Canada Tel. : ῌ+῎/+3῎2,.῎.+,* ext. /,3*+ Fax : ῌ+῎/+3῎2,.῎00-+ E-mail : [email protected] 2 J. Poult. Sci., .+ (+)

Introduction

The egg is the largest biological cell originating from one cell division. It is an important source of nutrients, containing all of the proteins, lipids, vitamins, minerals and growth factors required by the developing embryo, as well as defense factors against bacterial and viral infection. The formation of an egg involves the conversion of the feed into egg constituents through a number of intricate and highly coordinated steps. The reproductive system of the hen, shown in Figure +, consists of the ovary and oviduct (Romano# and Romano#, +3.3). The ovary is the site of assembly of the primordial germ cells in the embryo (Burley and Vadehra, +323), which are later transformed into oocytes. Each oocyte becomes a follicle after being covered with the granulosa and theca layers. Yolk constituents are synthesized in the liver and are transported to the follicular walls via the blood. The follicle undergoes a rapid development during which most of the yolk is deposited prior to ovulation. When su$cient yolk has accumulated, the ovulated

Fig. +. The reproductive system of the hen (Romano# and Romano#, +3.3). Reproduced with permission from John Wiley & Sons, Inc., New York. Mine and Kovacs-Nolan : Biologically Active Components in Hen Eggs 3 ovum enters the oviduct (Burley and Vadehra, +323 ; Okubo et al., +331). In laying hens, the oviduct consists of ﬁve regions, infundibulum, magnum, isthmus, uterus and vagina. The ovulated ovum is held in the infundibulum, where the yolk probably acquires the outer layer of the vitelline membranes and the chalazal layer of the albumen (Burley and Vadehra, +323). In the albumen-secreting region (magnum), the egg albumen is secreted to cover the yolk, which is then immediately covered with the shell membranes (Burley and Vadehra, +323). Egg shell formation occurs in the uterus. The exact mechanism is unclear, but calcium from the blood is deposited to the shell, secreted by shell glands, and assembling as a crystalline-like calcium structure on the shell membranes (Burley and Vadehra, +323 ; Okubo et al., +331). The vagina is the last portion of the oviduct, and the end of the vagina connects with the cloaca. Eggs consist of about 3./ῌ eggshell (including shell membrane), 0-ῌ albumen, and ,1./ῌ yolk (Cotterill and Geiger, +311)(Table +). The main components are water (1/ῌ), proteins (+,ῌ), lipids (+,ῌ), as well as carbohydrates and minerals (Burley and Vadehra, +323 ; Sugino et al., +331). The proteins are distributed throughout the egg, with the majority found in the egg yolk and egg white, and a small proportion in the eggshell and shell membrane (Sugino et al., +331). The lipids are found almost exclusively in the egg yolk, mainly in the form of lipoproteins, and include phosphorous, nitrogen, and sugar-containing lipids (Burley and Vadehra, +323 ; Sugino et al., +331). Several minerals have also been found in eggs, most of them in the egg shell. Carbohydrates are a minor egg component, present throughout the egg, both as free and conjugated forms, attached to proteins and lipids (Sugino et al., +331). Diverse biological activities have nowbeen attributed to egg components, including antimicrobial and antiviral activity, protease inhibitory action, vitamin-binding properties, anti-cancer activity and immunomodulatory activity (Li-Chan et al., +33/). The role of these egg components in human health has been extensively studied, and will be discussed here.

Egg Shell and Membrane

Chicken eggshell is a mineralized structure consisting of approximately 3/ῌ calcium carbonate by weight, the rest made up of an organic matrix consisting mainly of glycoprotein and proteoglycans (Hincke, +33/ ; Hincke et al., +333). Generally considered a waste product, only recently has significant research been carried out to examine the potential biological activity of egg shell components (Suguro et al., ,*** ; Daengprok et al., ,**,). Eggshell calcium has been proposed for pharmaceutical applications for calcium deficiency therapies in humans, and in animals for bone mineralization and growth (Tsugawa et al., +33/). Eggshell powder was shown to have antirachitic e#ects in rats (Rovensky et al., +33.). In vitro, eggshell powder stimulated the growth of chick embryo cartilage cells (Schaafsma and Pakan, +333), and in humans the use of the eggshell powder resulted in an increase in bone mineral density (Schaafsma and Beelen, +333). In piglets, the apparent absorbability of calcium from eggshell powder was found to be at least as good as that from purified CaCO- (Schaafsma and Beelen, +333), and it has been suggested as a calcium supplement to 4 J. Poult. Sci., .+ (+)

Table +. Chemical composition of hen eggs (Modiﬁed from Mine, ,**,) ῌ Constituent Major components (relative ῌῌ w/w) ῍wῌv῎ EGG SHELL 3./ (including Inorganic salts (3+.21): Calcium carbonate (32..) shell membrane) Magnesium carbonate (*.2) Tricalcium phosphate (*.2) Proteins (0..) Water (+.1) Lipids (*.*-) EGG WHITE 0-.* Proteins (3.1ῌ+*.0): Ovalbumin (/.) Ovotransferrin (+,.*) Ovomucoid (++) Ovomucin (-./) Lysozyme (-..) G, Globulin (..*?) G- Globulin (..*?) Avidin (*.*/) Lipids (*.*-) Carbohydrates (*..ῌ*.3) Ash (*./ῌ*.0) EGG YOLK ,1./ Proteins (+/.1ῌ+0.0): Apovitellenin (I-VI) (-1.-) Lipovitellin apoproteins (.*.*) a-lipovitellin b-lipovitellin Livetins (3.-) a-livetin (serum albumin) b-livetin (a, glycoprotein) g-livetin (g globulin) Phosvitin (+-..) Biotin-binding protein (trace) Lipids (-,.*ῌ-/.*) Triglycerol (00) Phosphatidylcholine (PC) (,.) Phosphatidylethanolamine (PE) (,.2) Lysophosphatidylcholine (LPC) (*.0) Sphingomyelin (*.0) Cholesterol (/.*) Others (+.*) Carbohydrate (*.,ῌ+.*) Ash (+.+) Mine and Kovacs-Nolan : Biologically Active Components in Hen Eggs 5 increase bone mineral density in individuals with osteoporosis (Omi and Ezawa, +332 ; Schaafsma et al., ,***). A recent study demonstrated that a ,+ *** Da protein present in soluble eggshell matrix proteins may play an important role in increasingcalcium transport across intestinal epithelial cells in vitro (Daengprok et al., ,**-). Eggshell membrane is composed of collagen-like proteins, which are largely located in the inner membrane (Wong et al., +32.). Eggshell membrane-derived peptides, prepared by alkaline treatment, were shown to stimulate human skin ﬁbroblasts in vitro (Suguro et al., ,***). Eggshell membranes also contain antimicrobial substances, with lysozyme (Vadehra et al., +31,)andb-N-acetylglucosaminidase (Winn and Ball, +31/) activity reported.

Egg White

Ovalbumin Ovalbumin constitutes over half of the total egg white proteins (Ibrahim, +331). It is a monomeric phosphoglycoprotein, and has been used extensively as a model for studyingthe structure-function relationships of proteins (Li-Chan and Nakai, +323 ; Ibrahim, +331). Functionally, ovalbumin is important for the gelling, foaming, and emulsifyingproperties of eggwhite(Mine, ,**,), however, its biological role in the egg remains unknown. It has been suggested that ovalbumin may serve as a source of amino acids for the developingembryo (Ibrahim, +331). Ovalbumin may possess some immunomodulatory activity, as it was found to induce the release of tumor necrosis factor (TNF) alpha in a dose-dependant manner in vitro, when modified with dicarbonyl methylglyoxyl (Fan et al., ,**-), and im- munogenic ovalbumin peptides have been used to enhance immune responses for cancer immunotherapy (Vidovic et al., ,**, ; Goldberg et al., ,**- ;Heet al., ,**-). A vasorelaxingpeptide, ovokinin (OA -/2ῌ-0/), was isolated by the peptic digestion of ovalbumin (Fujita et al., +33/ a). Ovokinin (,ῌ1), a peptide produced by chymotrypsin digestion, and corresponding to OA -/3ῌ-0., was also found to possess vasorelaxingactivity (Matoba et al., +333). Both peptides were found to significantly lower the systolic blood pressure in spontaneously hypertensive rats, when administered orally (Fujita et al., +33/ b). The replacement of amino acids in the ovokinin (,ῌ1) peptide has resulted in enhanced anti-hypertensive activity, with the most potent derivative resultingin +**-fold more potent anti-hypertensive activity (Matoba et al., ,**+ ; Yamada et al., ,**,). Two angiotensin I converting enzyme (ACE)-inhibitory peptides were also identified in ovalbumin by peptic (OA +2-ῌ+2.) and tryptic (OA ,**ῌ,+2) digestions (Yoshikawa and Fujita, +33.). Finally, the phagocytic activity of macrophages was increased by the addition of OA 11ῌ2. and OA +,0ῌ+-. peptides, derived by peptic and chymotryptic digestions of ovalbumin, respectively (Tezuka and Yoshikawa, +33/). Lysozyme Lysozyme, which plays an important role in the natural defense mechanism (Kijowski et al., ,***), is a ubiquitous enzyme, present in almost all secreted body 6 J. Poult. Sci., .+ (+)

fluids and tissues of humans, as well as plants. The most plentiful source is hens’ egg white, containing around *.-ῌ*.. g of lysozyme per egg (Losso et al., ,***). Lysozyme acts as a mucopeptide N-acetylmuramyl hydrolase, exerting bacteriolytic activity by hydrolyzing the b(+ῌ.) linkage between N-acetylmuraminic acid and N-acetylglucosamine of peptidoglycan, the structural component of bacterial cell walls (Salton, +3/1). It has demonstrated antimicrobial activity against such organisms as Bacillus stearothermophilus, Clostridium tyrobutyricum, Clostridium thermosaccharolyticum, Clostridium sporogenes and Bacillus spp. (Losso et al., ,***), as well as Enterococcus faecalis, Weissella viridescens (Gill and Holley, ,**-), Brochothrix thermosphacta, Lactobaccilus sakei, Leuconostoc mesenteroides, Listeria monocytogenes (Gill and Holley, ,***), and Carnobacterium sp. 2./ (Nattress et al., ,**+) when used in conjunction with other compounds, such as nisin and EDTA. The enzymatic treatment of lysozyme, exposing amino acids 32ῌ++,, which have been found to exert antimicrobial action (Pellegrini et al., +331 ; Ibrahim et al., ,**+), as well as synthetic peptides corresponding to the C-terminal of lysozyme (During et al., +333), have been examined. Pellegrini et al. (,***) demonstrated that polypeptides derived from lysozyme were capable not only of damaging the outer membrane of Escherichia coli, but could also inhibit DNA and RNA synthesis. Along with numerous applications as a food antimicrobial agent, lysozyme has been added to oral health care products, such as toothpaste, mouthwash and chewing gum to protect against periodontis-causing bacteria and prevent infections in the oral mucosa (Sava, +330 ; Tenuovo, ,**,). Lysozyme has also been shown to exert antiviral activity, reportedly associated with its charge, rather than its lytic ability (Losso et al., ,***). Oral and topical applications of lysozyme were found to be e#ective in preventing and controlling several viral skin infections, including herpes simplex and chicken pox (Sava, +330), as well as acting as exerting anti-inflammatory action (Sava, +330). It has also been shown, when combined with immunotherapy, to be e#ective in improving chronic sinusitis (Asakura et al., +33*), and to normalize humoral and cellular responses in patients with chronic bronchitis (Sava, +330). Lee-Huang et al. (+333) found that chicken lysozyme also possessed activity against HIV type +. Finally, lysozyme has also been shown to act as an immune-modulating and immune-stimulating agent, enhancing immunoglobulin productivity, and regulating and restoring the immune responses in immune-depressed patients undergoing anti-cancer treatments (Sava, +330 ; Sugahara et al., ,***), and as an anti-cancer agent, inhibiting tumor growth in a number of experimental tumors (Sava, +323 ; Sava et al., +323, +33+ ;Daset al., +33, ; Pacor et al., +330, +333) and enhancing the e$cacy of chemotherapy treatments (Sava et al., +33/). Ovotransferrin Ovotransferrin is a monomeric glycoprotein, belonging to the transferrin family, a group of iron-binding proteins widely distributed in various biological fluids, which has the capacity to reversibly bind two iron ions per molecule (Ibrahim, ,***). Its suggested function is as an iron scavenger, to prevent its use by microorganisms, and as Mine and Kovacs-Nolan : Biologically Active Components in Hen Eggs 7 an iron delivery agent (Abdallah et al., +333). Ovotransferrin has demonstrated antibacterial activity against a wide spectrum of bacteria, including Pseudomonas spp., Escherichia coli, Streptococcus mutans (Valenti et al., +32-), Staphylococcus aureus, Bacillus cereus (Abdallah et al., +333)andSalmonella enteritidis (Baron et al., ,***). It has been suggested that ovotransferrin can exert antibacterial activity by permeating bacterial outer membranes, reaching the inner membrane and causing the selective permeation of ions and dissipation of electrical potential (Aguilera et al., ,**-). A 3,-amino acid ovotransferrin peptide, OTAP-3,, was also found to be capable killing Gram negative bacteria by crossing the bacterial outer membrane by self-promoted uptake, and damaging the cytoplasmic membrane (Ibrahim et al., ,***). It has also been shown that ovotransferrin possesses both antiviral activity, against Marek’s disease virus in chicken embryo fibroblasts (Giansanti et al., ,**,), as well as antifungal activity, against species of Candida (Valenti et al., +32/). Xie et al. (,**,) demonstrated that ovotransferrin can act as an im- munomodulator, modulating macrophage and heterophil functions in chickens. Fur- ther immunomodulating e#ects of ovotransferrin have also been shown, including the inhibition of proliferation of mouse spleen lymphocytes (Otani and Odashima, +331) and the enhanced phagocytic response of peripheral blood mononuclear cells and polymorphonuclear cells in dogs (Hirota et al., +33/). Ovotransferrin was also found to facilitate the recovery of chick eyes from induced myopia (Rada et al., ,**+). Finally, when administered with Syn ,+3*, an inhibitor of AmpC lactamases, lactoferrin increased the activity of various antibiotics against beta-lactamase- producing bacteria (Babini and Livermore, ,***). Ovomucoid Ovomucoid is a glycoprotein, arranged into three domains which are cross-linked by intradomain disulfide bonds (Kato et al., +321). It is relatively resistant to treatment with heat or digestive enzymes, and it is this stability that has resulted in its being one of the dominant egg white allergens (Cooke and Sampson, +331). Ovomucoid is also one of four egg white proteinase inhibitors, belonging to the group of serine proteinase inhibitors, namely inhibiting trypsin (Kato et al., +321 ; Saxena and Tayyab, +331). The incorporation of ovomucoid into polymeric microparticles, to overcome the degradation of protein drugs by proteolytic enzymes, has been examined. Agarwal et al. (,**+ b) found that when ovomucoid was used, the stability of insulin in poly- methacrylate based microparticulates was increased significantly. Inclusion of ovomucoid also resulted in targeting of drugs to the blood, by acting as a biospecific ligand to lectins on the walls of the gastrointestinal tract (Plate et al., ,**,). The presence of ovomucoid was found to enhance insulin flux across rat jejunum (Agarwal et al., ,**+ a), suggesting the use of ovomucoid to enhance the oral delivery of insulin. Using a rat model of experimental pancreatitis, intravenous ovomucoid was found to decrease the trypsin-like activity to the level of intact rats, and reduce primary pancreas destruction (Valueva et al., +33,). Synthetic ovomucoid peptides have also demonstrated immunomodulating activity, 8 J. Poult. Sci., .+ (+)

inducing T-cell secretion of cytokines interleukin- (IL) .,IL-+*,IL-+-, interferon- (IFN) gamma, and IL-0 (Holen et al., ,**+). Because of its allergenic nature, however, ovomucoid has limited biological and medical applications. Avidin Chicken avidin is a tetrameric glycoprotein, with an extremely high a$nity for the water soluble vitamin biotin (Green, +31/). The unique feature of this binding is the strength and specificity of the formation of the avidin-biotin complex, formed when avidin binds four molecules of biotin (Bayer and Wilchek, +33.). Avidin possesses antimicrobial activity, and has been found to inhibit the growth of biotin-requiring bacteria and yeasts (Green, +31/ ; Banks et al., +320). Avidin antimicrobial activity has also been attributed to its ability to bind to various Gram negative and Gram positive bacteria, including Escherichia coli K-+,, Klebsiella pneumoniae, Serratia marcescens, Pseudomonas aeruginosa, Staphylococcus aureus, and Staphylococ- cus epidermis (Korpela et al., +32.). Avidin has been used in cancer treatment, to localize and image cancer cells and to pre-target drugs to tumors. Because of its tight biotin binding and signal amplification due to the tetrameric structure of avidin, it leads to the accumulation of higher e#ective doses and increased persistence of biotinylated anti-cancer drugs, as compared to other immunotherapeutic procedures (Hytonen et al., ,**-). Tumor pre-targeting with avidin has also been found to be e#ective in increasing the uptake of tumor necrosis factor (TNF) alpha conjugated to biotin in vitro, improving the anti-tumor activity of TNF (Moro et al., +331 ; Corti et al., +332 ; Gasparri et al., +333). Yao et al. (+332) found that radiolabelled avidin also bound to lectins expressed on the surface of tumor cells, and localized highly and rapidly in various types of tumors in mice, thereby reducing radioactivity accumulation in other organs. It has also been found to be essential for the activity of adoptively transferred T-cells at tumor sites (Guttinger et al., ,***), and the utilization of avidin in drug delivery through the blood-brain barrier has been demonstrated, facilitating delivery of therapeutics to the brain (Bickel et al., ,**+). Ovomucin Egg white ovomucin is a macromolecular and heavily glycosylated glycoprotein, consisting of a peptide-rich a-subunit and a carbohydrate-rich b-subunit (Itoh et al., +321). Ovomucin serves physical functions within the egg, such as maintaining the structure and viscosity of the egg white albumen (Tsuge et al., +331 b), thus preventing the spread of microorganisms (Ibrahim, +331). It has also demonstrated several biological applications. Ovomucin has shown antiviral activity against Newcastle disease virus, bovine rotavirus, and human influenza virus in vitro (Tsuge et al., +330 a, b, +331 a, b ; Watanabe et al., +332 a). Ovomucin peptides, produced by treatment with the enzyme pronase, showed increased solubility, compared to its native form, while still retaining virus-binding activity (Tsuge et al., +331 a ; Watanabe et al., +332 a). Pronase-prepared glycopeptides of ovomucin have also demonstrated anti-tumor Mine and Kovacs-Nolan : Biologically Active Components in Hen Eggs 9 e#ects in a double grafted tumor system in mice (Watanabe et al., +332 b), suggested to be related to the anti-angiogenic activity of ovomucin, inhibiting tumor growth (Oguro et al., ,**+). Ovomucin peptides may also act as immunomodulators, showing macrophage- stimulating activity in vitro (Tanizaki et al., +331). Finally, ovomucin was found to inhibit cholesterol uptake in vitro by Caco-, cells, and reduce serum cholesterol in rats, displaying hypocholesterolemic action (Nagaoka et al., ,**,). Cystatin Egg white cystatin, part of a ῌsuperfamily” of cystatins, belongs to the Type , cystatins (Barrett, +320), and inhibits most cysteine proteinases, including ficin, papain, and cathepsins B, C, H and L (Li-Chan et al., +33/). It contains two disulphide bonds, but no carbohydrate (Barrett, +320). Low contents of cystatins in natural resources may limit their applications (Nakai, ,***), however, genetic modification and expression of cystatin has been examined, not only providing a source for increased quantities of cystatin, but also resulting in the production of recombinant cystatin with enhanced proteinase inhibitory activity (Ogawa et al., ,**,). Egg white cystatin has been shown to possess antibacterial activity, preventing the growth of group A streptococcus (Bjorck, +33*), Salmonella typhimurium (Nakai, ,***), and the periodontis-causing Porphyromonas gingivalis (Blankenvoorde et al., +330). Blankenvoorde et al. (+332) found that peptides derived from cystatin were also capable of inhibiting the growth of P. gingivalis. Several proteases, including cysteine proteases, are required for tumor progression and metastasis (Krol et al., ,**- a). Increased levels of cysteine proteases, and a decrease in cystatin, have been observed in various cancers (Konduri et al., ,**, ; Nagai et al., ,**-). Cystatin has been shown to inhibit tumor invasion in ras- transformed breast epithelial cells (Premzl et al., ,**+), and multifunctional inhibitors, composed of chicken cystatin in conjunction with other protease inhibitors, have been suggested for the therapy of solid tumors (Muehlenweg et al., ,*** ; Krol et al., ,**- a, b). Cystatins also produce less intensive side e#ects than other synthetic protease inhibitors currently used in medical treatments (Nakai, ,***). A relationship between cystatins, cytokines, and immune response has also been suggested. It has been observed that cystatin induced the synthesis of various cytokines (Kato et al., ,***), resulting in an up-regulation of nitric oxide release in vitro using mouse peritoneal macrophages (Verdot et al., +330, +333), as well as in vivo, greatly reducing parasite numbers in a mouse model of visceral leishmaniasis (Das et al., ,**+). Ovomacroglobulin (ovostatin) Ovomacroglobulin, also referred to as ovostatin, is a glycoprotein composed of four subunits joined in pairs by disulfide bonds (Kitamoto et al., +32,). It has demonstrated broad-spectrum inhibitory activity against various types of proteases, including serine proteases, cysteine proteases, thiol proteases, and metalloproteases (Kitamoto et al., +32, ; Molla et al., +321). The antimicrobial e#ects of ovomacroglobulin against Serratia marcescens and Pseudomonas aeruginosa, due to its proteinase inhibitory action, have been 10 J. Poult. Sci., .+ (+)

demonstrated both in vitro (Molla et al., +321 ; Miyagawa et al., +33+ b, c, +33.), and in vivo, where it was found to reduce corneal destruction in an experimental keratitis model in rabbits, as well as accelerate wound healing (Miyagawa et al., +33+ a ; Ijiri et al., +33- ; Miyagawa et al., +33.). Ovomacroglobulin was also found to enhance periodontal wound healing in rats, by accelerating fibroblast growth, collagen deposi- tion, and capillary formation in tissue (Ofuji et al. +33,). The proteinase inhibitory e#ects of ovomacroglobulin have also demonstrated several other biological e#ects, including the suppression of P. aeruginosa and Vibrio vulnificus septicemia due to the inhibition of kinin generating proteases (Maeda et al., +33- ; Maruo et al., +332), the in vitro inhibition of the inflammatory proteinase medullasin (Ikai et al., +323), and the suppression of metalloproteinases and vascular permeability in skin tissues, which play a role in tumor metastasis (Wu et al., ,**+). Ovoinhibitor Ovoinhibitor, like ovomucoid, is a serine proteinase inhibitor, inhibiting enzymes such as trypsin, chymotrypsin, and elastase, as well as various bacterial and fungal proteinases (Matsushima, +3/2 ; Vered et al., +32+ ; Tomimatsu et al., +300). It has been found to prevent the development of rotavirus gastroenteritis in a mouse model of rotavirus infection (Yolken et al., +321), and to inhibit the formation of active oxygen species by human polymorphonuclear leukocytes, which are associated with inflammatory diseases, mutagenicity and carcinogenicity (Frenkel et al., +321). It has also been used to study models of autoimmune arthritis in mice (Terato et al., +330). Proteinases are involved in the regulation of a number of biological processes, and have been implicated as contributors in several diseases, including viral diseases, such as HIV (Maliar et al., ,**,), and Alzheimer’s (Schimmoller et al., ,**,). Proteinase inhibitors, therefore, such as those from egg white, have significant potential for the treatment and prevention of proteinases-mediated diseases. The biological activities associated with egg white proteins are summarized in Table ,.

Egg Yolk

Immunoglobulin (Ig) Y Immunoglobulin (Ig) Y is the functional equivalent of IgG, the major serum antibody in mammals (Carlander et al., ,***). It is transferred to the developing embryo, to give acquired immunity to the chick (Carlander et al., +333 ;Simet al., ,***). Speciﬁc IgY can be produced by immunization of chickens with the desired protein, and then puriﬁed from the egg yolk (Gassmann et al., +33* ; Carlander et al., ,***). One of the primary applications of IgY is immunotherapy, for the passive immunization of immunocompromised individuals, and as an alternative to traditional antibiotic treatment. IgY has been produced against a number of bacteria and viruses, and has been shown to bind to and inhibit the infection and disease symptoms, in vitro and in vivo, of gastrointestinal pathogens such as human and bovine rotavirus (Yolken et al., Mine and Kovacs-Nolan : Biologically Active Components in Hen Eggs 11

Table ,. Biological properties of egg white proteins

Protein Properties Reference Ovalbumin Immunomodulating activity Tezuka and Yoshikawa, +33/ ; Vidovic et al., ,**, ;Fan et al., ,**- ; Goldberg et al., ,**- Anti-hypertensive activity of ovo- Fujita et al., +33/ a, b ; Matoba et kinin and ovokinin (,ῌ1) al., +333, ,**+ ; Yamada et al., ,**, ACE inhibitory activity of oval- Yoshikawa and Fujita, +33. bumin-derived peptides Lysozyme Antibacterial activity Losso et al., ,*** ; Gill and Holley, ,***, ,**- ; Nattress et al., ,**+ Antimicrobial action of lysozyme Pellegrini et al., +331, ,*** ; During peptides et al., +333 ; Ibrahim et al., ,**+ Antiviral activity Asakura et al., +33* ; Sava, +330 ; Losso et al., ,*** ; Lee-Huang et al., +333 Anti-inﬂammatory activity Sava, +330 Immunomodulating and immune- Asakura et al, +33* ; Sava, +330 ; stimulating activity Sugahara et al., ,*** Anti-tumor activity Sava, +323 ; Sava et al., +323, +33+ ; Pacor et al., +333 Protection against periodontis- Sava, +330 ; Tenuovo, ,**, causing bacteria Ovotrans- Antibacterial activity Valenti et al., +32- ; Abdallah et ferrin al., +333 ; Ibrahim, ,*** ; Baron et al., ,*** Antimicrobial activity of OTAP-3, Ibrahim et al., ,*** Antiviral activity Giansanti et al., ,**, Antifungal activity Valenti et al., +32/ Immunomodulating activity Hirota et al., +33/ ; Otani and Odashima, +331 ;Xieet al., ,**, Enhancement of antibiotic activity Babini and Livermore, ,*** Ovomucoid Serine proteinase inhibitor Kato et al., +321 ; Valueva et al., +33, ; Saxena and Tayyab, +331 Drug delivery to small intestine Agarwal et al., ,**+ a, b Biospeciﬁc ligand to gastrointes- Plate et al., ,**, tinal tract Immunomodulating activity Holen et al., ,**+ Avidin Antimicrobial activity Korpela et al., +32. ; Banks et al., +320 Pre-targeting and drug delivery Moro et al., +331 ; Corti et al., of anti-cancer drugs +332 ;Yao et al., +332 ; Gasparri et al., +333 ; Hytonen et al., ,**- 12 J. Poult. Sci., .+ (+)

Protein Properties Reference Application in immunotherapy Guttinger et al., ,*** Drug delivery to brain Bickel et al., ,**+ Ovomucin Antiviral activity Tsuge et al., +330 a, b, +331,a,b; Watanabe et al., +332 a Anti-tumor activity Watanabe et al., +332 b ; Oguro et al., ,**+ Immunomodulating activity Tanizaki et al., +331 Hypocholesterolemic activity Nagaoka et al., ,**, Cystatin Cysteine proteinase inhibitor Li-Chan et al., +33/ Antibacterial activity Bjorck, +33* ; Blankenvoorde et al., +330 ; Nakai, ,*** Inhibition of growth of P. gingi- Blankenvoorde et al, +332 valis by cystatin peptides Immunomodulating activity Verdot et al., +330, +333 ; Kato et al., ,*** ;Daset al., ,**+ Anti-tumor activity Muehlenweg et al., ,*** ; Premzl et al., ,**+ ; Krol et al., ,**- a, b Ovomacro- Serine, cysteine, thiol, and met- Kitamoto et al., +32, ; Molla et globulin allo proteinase inhibitor al., +321 ; Ikai et al., +323 ; Maeda et al., +33- ; Maruo et al., +332 ; Wu et al., ,**+ Antibacterial activity Molla et al., +321 ; Miyagawa et al., +33+ b, c, +33. Enhancement of wound healing Miyagawa et al., +33+ a ; Ofuji et al., +33, ; Ijiri et al., +33- Ovoinhibitor Serine proteinase inhibitor Matsushima, +3/2 ; Tomimatsu et al., +300 ; Vered et al., +32+ Antiviral activity Yolken et al., +321 Potential anti-inﬂammatory and Frenkel et al., +321 anti-mutagenic activity

+322 ; Hatta et al., +33- ; Kuroki et al., +33. ; Ebina et al., +330 ; Kovacs-Nolan et al., ,**+), bovine coronavirus (Ikemori et al., +331), Escherichia coli (Ikemori et al., +33, ; O’Farrely et al., +33, ; Yokoyama et al., +33, ; Imberechts et al., +331 ;Jinet al., +332 ; Marquardt et al., +333 ; Amaral et al., ,**,), Salmonella spp. (Peralta et al., +33. ; Yokoyama et al., +332 a, b ; Sugita-Konishi et al., ,*** ;Leeet al., ,**,), Yersinia ruckeri (Lee et al., ,***), Edwardsiella tarda (Hatta et al., +33.), Helicobacter pylori (Shin et al., ,**,), porcine epidemic diarrhea virus (Kweon et al., ,***), and infectious bursal disease virus (Eterradossi et al., +331), as well as Staphylococcus aureus, (Sugita-Konishi et al., +330 ; LeClaire et al., ,**,), Pseudomonas aeruginosa (Sugita-Konishi et al., +330 ; Carlander et al., ,**, ; Kollberg et al., ,**-). IgY against Streptococcus mutans have been shown to prevent adhesion of the bacteria in Mine and Kovacs-Nolan : Biologically Active Components in Hen Eggs 13 vitro and in vivo, and reduced dental caries development in an animal model (Otake et al., +33+ ; Hatta et al., +331 ; Chang et al., +333 ; Smith et al., ,**+). Chicken anti-venom IgY has been produced, for treatment of snake and spider bites, and was found to have a higher bioactivity than anti-venoms raised in horses (Thalley and Carroll, +33* ; Almeida et al., +332). Due to the structural di#erences between IgY and IgG (IgY does not fix mammalian complement components or bind human Fc receptors), IgY has been suggested for use to inhibit pig-to-human xenograft rejection (Fryer et al., +333 ; Leventhal et al., ,**+), and the use ofIgY has also been examined to replace the anti-inflammatory drugs used to treat Crohn’s disease and ulcerative colitis. Worledge et al. (,***) reported that anti-TNF antibodies produced in chickens were capable ofe #ectively treating acute and chronic phases ofcolitis in rats, and were also foundto neutralize human TNF in vitro, indicating its possible use for the treatment of inflammatory bowel disease in humans. Finally, IgY has been suggested for use in cancer treatment, to act as a carrier for anti-tumor drugs (Yang et al., +331), and has demonstrated anti-tumor activity, inhibiting the spreading and invasion ofglioma cells into adult rat optic nerve cells in vitro (Hensel et al., +332). Phosvitin Phosvitin is a highly phosphorylated protein, containing +*ῌ phosphorus and 0./ῌ carbohydrates (Taborsky and Mok, +301). Ninety-five percent ofthe iron in eggs is present in the yolk and is bound to phosvitin (Greengard et al., +30.), however its bioavailability is very low (Morris and Greene, +31,). This is due to the strong binding ofphosvitin, or its phosphopeptide derivatives, with Fe -ῌ, and the formation of phosvitin-iron complexes which promote the precipitation ofiron in the small intestine (Sato et al., +32., +32/, +321). However, Jiang and Mine (,***, ,**+) produced functional phosphopeptides, with a molecular weight of +***ῌ-*** Da, derived by tryptic hydrolysis following partial alkaline dephosphorylation. These peptides ex- hibited enhanced calcium-binding capacity and inhibited the formation of insoluble calcium phosphates. Phosvitin also demonstrated a capacity to inhibit iron catalysis ofphospholipid oxidations (Lu and Baker, +320). Thus, phosvitin could be useful in foods as a natural antioxidant. Lipoproteins (Low density lipoproteins-LDL) The low density lipoproteins (LDL) fraction of yolk plasma is made up of 23ῌ lipid and ++ῌ protein (Cook and Martin, +30,). The lipid content of LDL contains 1*ῌ triacylglycerol, .ῌ cholesterol, and ,0ῌ phospholipids (Martin et al., +30-). The immunomodulating activity of LDL has been demonstrated using human histocytic lymphoma cells (U-3-1), human monocytic leukemia cells (THP-+) and U- 3-1-derived macrophage-like cells (U-M) (Suzuki et al., +33.), promoting the growth ofall three cell types. LDL was also shown to enhance the production ofIgM in human-human hybridomas (Shinohara et al., +33-). 14 J. Poult. Sci., .+ (+)

Sialic acid Sialic acid is a term referring to derivatives of neuraminic acid, which have an acyl group on the amino nitrogen. The most widely distributed sialic acid in nature is N-acetylneuraminic acid (Neu/Ac) (Hartmann and Wilhelmson, ,**+). The use of egg yolk as a source of sialic acid has been examined, and was found to be an excellent source for the large scale preparation of Neu/Ac (Koketsu et al., +33, ; Juneja et al., +33.). Sialic acids possess many biological functions, including acting as receptors for microorganisms, toxins, and hormones, and masking receptors and immunological recognition sites of molecules and cells (Schauer, +321). Sialic acid may act as an anti-inflammatory agent. Cells carrying a carbohydrate ligand called sialyl-Lewis X were found to bind to endothelial leukocyte adhesion molecule-+ (ELAM-+), which mediates the adhesion of circulating leukocytes to the vascular endothelium during inflammation (Lowe et al., +33* ; Phillips et al., +33*). The anti-inflammatory e#ects of sialyl-Lewis X were also shown in vivo, reducing lung injury in rats (Mulligan et al., +33-) and reducing tissue injury in rabbit ears, caused by a temporary increase in blood supply (Han et al., +33/). Sialyloligosaccharides Several sialyloligosaccharides in chalaza, egg yolk membrane and delipided egg yolk have been isolated by acid hydrolysis or protease digestion (Juneja et al., +33+). These sialyloligosaccharides are likely to be naturally present as glycoproteins or glycopeptides (Koketsu et al., +33/ b). Sialylglycoconjugates, sialylgangliosides, sialyloligosaccharides and sialylglycoproteins have been reported to play various important roles in animal and human tissue cells. The oligosaccharide-enriched fraction of sialyloligosaccharide was found to significantly inhibit rotavirus infection both in vitro and in vivo, with the sialic acid moiety of the oligosaccharide playing an important role in viral inhibition (Koketsu et al., +33/ a). Sialyloligosaccharides have also been found to inhibit Salmonella infection by inhibiting the entry of bacteria through the gut (Sugita-Konishi et al., ,**,). Yolk lipids Dry egg yolk contains approximately 0*ῌ lipids (Juneja, +331), made up of around 0/ῌ is triglyceride, ,2ῌ is phospholipid, and /ῌ is cholesterol (Li-Chan et al., +33/). The fatty acid composition of the lipid fraction of egg yolk varies, and is influenced by the type of fat in the hen’s diet (Li-Chan et al., +33/). Egg yolk lipids have found numerous applications in the food industry, due to their surfactant (Juneja, +331) and antioxidant (Yamamoto et al., +33*, +33+ ; Juneja, +331) properties, and in the cosmetic industry, to replenish lipid deficiency in the skin (Juneja, +331). Egg yolk lipids have also demonstrated a number of medical applications. Rabinowich et al. (+321) noted that the administration of an egg yolk-derived lipid mixture to elderly individuals, formulated for in vivo rectification of rigidified cell membranes in an attempt to restore proper physiological function, resulted in an increase in lymphocyte responsiveness. As well, a common parenteral fat emulsion derived from egg yolk lipids is often used as a carrier of fat soluble drugs (Hartmann Mine and Kovacs-Nolan : Biologically Active Components in Hen Eggs 15 and Wilhelmson, ,**+). Cholesterol is an important component in cell membranes and is needed for the growth of infants. As well, it is a precursor of bile acids, sex hormones and cortex hormones (Juneja, +331). The supplementation of infant formulas with egg yolklipids has been suggested, to more closely resemble the mother’s milk, and it has been found that while providing essential nutrients, the yolklipids did not result in an increase in plasma cholesterol, indicating that it could safely be included in the infant diet (Makrides, ,**,). Finally, egg yolkcholesterol has also been suggested for use in the treatment of Smith-Lemli-Opitz syndrome, a condition in which the activity of 1-dehydrocholesterol delta(1)-reductase, the final enzyme in cholesterol biosynthesis, is reduced. Patients receiving supplemental dietary cholesterol from egg yolkexhibited a significant increase in plasma cholesterol levels, and a decrease in plasma levels of cholesterol precursors, which may be toxic (Linck et al., ,***). Phospholipids Phospholipids (PLs) are lipids which contain phosphate, and have a glycerol- phosphate backbone (Juneja, +331). About -+ῌ of the egg yolklipids are PLs ( ,-.), 1-ῌ of which is phosphatidylcholine (PC), followed by phosphatidylethanolamine (PE +/ῌ), lysophosphatidylcholine (LPC /.2ῌ), and sphingomyelin (SM ,./ῌ). The remaining -.1ῌ is made up of lysophosphatidylethanolamine (LPE ,.+ῌ), plasmalogen (*.3ῌ) and inositol phosholipid (IP) (Rhodes and Lea, +3/1). Phospholipids are amphiphilic molecules, having both polar and non-polar groups (Juneja, +331), making them ideal for studying membranes and preparing liposomes (aqueous compartments enclosed by a lipid bilayer (Hartmann and Wilhelmson, ,**+). Egg PC-containing liposomes have been shown to be e#ective for drug delivery to tumors (Chelvi and Ralhan, +33/ ; Avrilionis and Boggs, +330 ; Harasym et al., +331 ; Sharma et al., +332), and for drug delivery to the brain (Kobayashi et al., +330), reducing the toxicity and side e#ects, as well as prolonging the concentration compared to the free drug by itself. Feeding infants formula containing egg PLs was found to reduce the incidence of necrotizing enterocolitis, suggesting that one or more of the compounds of egg PLs may enhance the immature intestinal functions of infants (Carlson et al., +332). Egg yolkPLs were shown to decrease serum cholesterol levels in rats (Murata et al., +32,), with PC in particular reducing the intestinal absorption of cholesterol (Jiang et al., ,**+), and they have been found to improve memory retention and increase acetylcholine concentrations, a neurotransmitter which decreases in concentration in cases of Alzheimer’s disease (Masuda et al., +332 ; Favreliere et al., ,**-). A diet including egg PC was also found to enhance ῌmaze-learning” ability and brain functions in old mice (Lim and Suzuki, ,***). It has been suggested that egg PLs may exert a degree of antiviral activity by interfering with the viral envelope formation, and by protecting or restoring trans- membrane signaling functions in host cells (Liu and Watson, ,**,). Furthermore, bacterial or viral infections may also be reduced by an activation of immune cells by egg 16 J. Poult. Sci., .+ (+)

PLs, as immune cells grown in vitro require exogenous fatty acids, which could be provided by egg PLs (Liu and Watson, ,**,). More recently, research has focused on specific PL components, including ar- achidonic acid (AA), docosahexaenoic acid (DHA), and choline, whose metabolic products may play a role in membrane integrity, modulation of the membrane, and activation of immune cells (Liu and Watson, ,**,). Egg yolk PC is a significant source of choline, which is an important nutrient in brain development, liver function, and cancer prevention (Gutierrez et al., +331). DHA and AA are important in the maintenance of normal neural functions, and have been shown to promote e#ects that are hypolipidemic, anti-thrombic, vasodilatory, and anti-inflammatory (Sheppard, ,**,). The biological activities associated with egg yolk components are summarized in Table -.

Table -. Biological properties of egg yolk components

Component Properties Reference Immunoglob- Antibacterial activity O’Farrelly et al., +33, ; Yokoyama ulin Y et al., +33,, +332 a, b ; Hatta et al., +33. ; Sugita-Konishi et al., +330, ,*** ; Iberechts et al., +331 ;Jin et al., +332 ; Peralta et al., +332 ; Marquardt et al., +333 ;Leeet al., ,*** ; Amaral et al., ,**, ; LeClaire et al., ,**, ;Leeet al., ,**, ; Shin et al., ,**, ; Kollberg et al., ,**- Antiviral activity Hatta et al., +33- ; Kuroki et al., +33. ; Ebina et al., +330 ; Eterradossi et al., +331 ; Ikemori et al., +331 ; Kweon et al., ,*** ; Kovacs-Nolan et al., ,**+ Reduction of dental caries Otake et al., +33+ ; Hatta et al., +331 ; Chang et al., +333 ; Smith et al., ,**+ Anti-venom applications Thalley and Carroll, +33* ; Almeida et al., +332 Anti-inﬂammatory action Worledge et al., ,*** Cancer targeting and drug deliv- Yang et al., +331 ; Hensel et al., ery of anti-tumor drugs +332 Inhibition of xenograft rejection Fryer et al., +333 ; Leventhal et al., ,**+ Phosvitin Enhancement of calcium binding Jiang and Mine, ,***, ,**+ by phosvitin-derived peptides Antioxidant activity Lu and Baker, +320 Mine and Kovacs-Nolan : Biologically Active Components in Hen Eggs 17

Protein Properties Reference LDL Immunomodulating activity Shinohara et al., +33- ; Suzuki et al., +33. Sialyloigosac- Antiviral activity Koketsu et al., +33/ b charides Antibacterial activity Sugita-Konishi et al., ,**, Yolk lipids Antioxidant activity Juneja, +331 ; Hartmann and Wilhelmson, ,**+ Important for maintenance of Rabinowich et al., +321 ; Juneja, cell membranes +331 Drug carrier Hartmann and Wilhelmson, ,**+ Treatment of Smith-Lemli-Opitz Linck et al., ,*** symdrome Phospholipids Drug delivery to tumors and Chelvi and Ralhan, +33/ ; Avrilionis brain and Boggs, +330 ; Kobayashi et al., +330 ; Harasym et al., +331 ; Sharma et al., +332 Reduction of necrotizing enter- Carlson et al., +332 ocolitis in infants Reduction of serum cholesterol Murata et al., +32, ; Jiang et al., ,**+ Increase in acetylcholine concen- Masuda et al., +332 ; Lim and tration ; improvement in memory Suzuki, ,*** ; Favreliere et al., retention and brain function ,**- Antiviral activity Liu and Watson, ,**, Source of DHA and AA Sheppard, ,**,

Conclusion

In the last decade, extensive studies have been conducted to characterize the bio-physiological functions of egg components and to seek novel biologically active substances in hen eggs. By compiling these studies into a single focussed work, this review provides evidence that hen eggs contain various biologically active substances with specific benefits for human health, and significant potential for medical, nutraceutical and food-fortification applications.

References Abdallah FB and Chahine JM. Transferrins, the mechanism of iron release by ovotransferrin. European Journal of Biochemistry, ,0- : 3+,ῌ3,*. +333. Agarwal V, Nazzal S, Reddy IK and Khan MA. Transport studies of insulin across rat jejunum in the presence of chicken and duck ovomucoids. The Journal of Pharmacy and Pharmacol- ogy, /- : ++-+ῌ++-2. ,**+ a. Agarwal V, Reddy IK and Khan MA. Polymethylacrylate based microparticulates of insulin for oral delivery : preparation and in vitro dissolution stability in the presence of enzyme inhibitors. International Journal of Pharmaceutics, ,,/ : -+ῌ-3. ,**+ b. 18 J. Poult. Sci., .+ (+)

Aguilera O, Quiros LM and Fierro JF. Transferrins selectively cause ion e%ux through bacterial and artificial membranes. FEBS Letters, /.2 : /ῌ+*. ,**-. Almeida CM, Kanashiro MM, Rangel Filho FB, Mata MF and Kipnis TL. Development of snake antivenom antibodies in chickens and the purification from yolk. The Veterinary Record, +.- : /13ῌ/2.. +332. Amaral JA, Tino De Franco M, Carneiro-Sampaio MM and Carbonare SB. Anti- enteropathogenic Escherichia coli immunoglobulin Y isolated from eggs laid by immunized Leghorn chickens. Research in Veterinary Science, 1, : ,,3ῌ,-.. ,**,. Asakura K, Kojima T, Shirasaki H and Kataura A. Evaluation of the e#ects of antigen specific immunotherapy on chronic sinusitis in children with allergy. Auris Nasus Larynx, +1 : --ῌ -2. +33*. Avrilionis K and Boggs JM. Specific targeting of phototoxic haptenated liposomes to a hapten- specific B cell lymphoma. Cellular Immunology, +02 : +-ῌ,-. +330. Babini GS and Livermore DM. E#ect of conalbumin on the activity of Syn ,+3*,a+,/ dihydroxy- .-pyridon monobactam inhibitor of AmpC beta-lactamases. Journal of Antimicrobial Chem- otherapy, ./ : +*/ῌ+*3. ,***. Banks JG, Board RG and Sparks NHC. Natural antimicrobial systems and their potential in food preservation of the future. Biotechnology and Applied Biochemistry, 2 : +*-ῌ+.1. +320. Baron F, Fauvel S and Gautier M. Behaviour of Salmonella enteritidis in industrial egg white : egg naturally contains factors inhibitory to salmonella growth. In : Egg Nutrition and Biotechnology (Sim JS, Nakai N and Guenter W eds.). pp. .+1ῌ.-*. CAB International. Oxon. ,***. Barrett AJ. The cystatins : a diverse superfamily of cysteine peptidase inhibitors. Biomedica Biochimica Acta, ./ : +-0-ῌ+-1.. +320. Bayer EA and Wilchek M. Modified avidins for application in avidin-biotin technology : an improvement on nature. In : Egg Uses and Processing Technologies, New Developments (Sim JS and Nakai S eds.). pp. +/2ῌ+10. CAB International. Oxon. +33.. Bickel U, Yoshikawa T and Pardridge WM. Delivery of peptides and proteins through the blood-brain barrier. Advanced Drug Delivery Reviews, .0 : ,.1ῌ,13. ,**+. Bjorck L. Proteinase inhibition, immunoglobulin-binding proteins and a novel antimicrobial principle. Molecular Microbiology, . : +.-3ῌ+..,. +33*. Blankenvoorde MF, Henskens YM, van’t Hof W, Veerman EC, and Nieuw Amerongen AV. Inhibition of the growth and cysteine proteinase activity of Porphyromonas gingivalis by human salivary cystatin S and chicken cystatin. Biological Chemistry, -11 : 2.1ῌ2/*. +330. Blankenvoorde MF, van’t Hof W, Walgreen-Weterings E, van Steenbergen TJ, Brand HS, Veerman EC and Nieuw Amerongen AV. Cystatin and cystatin-derived peptides have antibacterial activity against the pathogen Porphyromonas gingivalis. Biological Chemistry, -13 : +-1+ῌ+-1/. +332. Burley RW and Vadehra DV. The Avian Egg, Chemistry and Biology. John Wiley & Sons, Inc., New York. +323. Carlander D, Stalberg J and Larsson A. Chicken antibodies : a clinical chemistry perspective. Upsala Journal of Medical Sciences, +*. : +13ῌ+3*. +333. Carlander D, Kollberg H, Wejaker P-Eand Larsson A. Peroral immunotherapy with yolk antibodies for the prevention and treatment of enteric infections. Immunologic Research, ,+ : +ῌ0. ,***. Carlander D, Kollberg H and Larsson A. Retention of specific yolk IgY in the human oral cavity. BioDrugs, +0 : .--ῌ.-1. ,**,. Carlson SE, Montalto MB, Ponder DL, Werkman SH, Korones SB. Lower incidence of necrotizing enterocolitis in infants fed a preterm formula with egg phospholipids. Pediatric Research, .. : .3+ῌ.32. +332. Chang HM, Ou-Yang RF, Chen YT and Chen CC. Productivity and some properties of immunoglobulin specific against Streptococcus mutans serotype c in chicken egg yolk (IgY). Journal of Agricultural and Food Chemistry, .1 : 0+ῌ00. +333. Chelvi TP and Ralhan R. Designing of thermosensitive liposomes from natural lipids for Mine and Kovacs-Nolan : Biologically Active Components in Hen Eggs 19

multimodality cancer therapy. International Journal of Hyperthermia, ++ : 02/ῌ03/. +33/. Cook WH and Martin WG. Egg lipoproteins. In : Structure and Functional Aspects of Lipopro- teins in Living Systems (Tria E and Scanu AM eds.). pp. /13ῌ0+/. Academic Press. New York. +30,. Cooke SK and Sampson HA. Allergenic properties of ovomucoid in man. Journal of Immunol- ogy, +/3 : ,*,0ῌ,*-,. +331. Corti A, Gasparri A, Sacchi A, Curnis F, Sangregorio R, Columbo B, Siccardi AG and Magni F. Tumor targeting with biotinylated tumor necrosis factor alpha : structure-activity relationships and mechanism of action on avidin pretargeted tumor cells. Cancer Research, /2 : -200ῌ-21,. +332. Cotterill OJ and Geiger GS. Egg product yield trends from shell eggs. Poultry Science, /0 : +*,1ῌ +*-+. +311. Daengprok W, Garnjanagoonchorn W and Mine Y. Fermented pork sausage fortified with commercial or hen eggshell calcium lactate. Meat Science, 0, : +33ῌ,*.. ,**,. Daengprok W, Garnjanagoonchorn W, Naivikul O, Pornsinlpatip P, Issigonis K and Mine Y. Chicken eggshell matrix proteins enhance calcium transport in the human intestinal epithelial cells, Caco-,. Journal of Agricultural and Food Chemistry, /+ : 0*/0ῌ0*0+. ,**-. Das S, Banerjee S and Gupta JD. Experimental evaluation of preventative and therapeutic potential of lysozyme. Chemotherapy, -2 : -/*ῌ-/1. +33,. Das L, Datta N, Bandyopadhyay S and Das PK. Successful therapy of lethal murine visceral leishmaniasis with cystatin involves up-regulation of nitric oxide and a favourable T cell response. Journal of Immunology, +00 : .*,*ῌ.*,2. ,**+. During K, Porsch P, Mahn A, Brinkmann O and Gie#ers W. The non-enzymatic microbicidal activity of lysozymes. FEBS Letters, ..3 : 3-ῌ+**. +333. Ebina T. Prophylaxis of rotavirus gastroenteritis using immunoglobulin. Archives of Virology Supplementum, +, : ,+1ῌ,,-. +330. Eterradossi N, Toquin D, Abbassi H, Rivallan G, Cotte JP and Guittet M. Passive protection of specific pathogen free chicks against infectious bursal disease by in-ovo injection of semi- purified egg-yolk antiviral immunoglobulins. Journal of Veterinary Medicine, B.. : -1+ῌ-2-. +331. Fan X, Subramaniam R, Weiss MF and Monnier VM. Methylglyoxal-bovine serum albumin stimulates tumor necrosis factor alpha secretion in RAW ,0..1 cells through activation of mitogen-activating protein kinase, nuclear factor kappaB and intracellular reactive oxygen species formation. Archhives of Biochemistry and Biophysics, .*3 : ,1.ῌ,20. ,**-. Favreliere S, Perault MC, Huguet F, De Javel D, Bertrand N, Piriou A and Durand G. DHA-enriched phospholipid diets modulate age-related alterations in rat hippocampus. Neurobiology of Aging, ,. : ,--ῌ,.-. ,**-. Frenkel K, Chrzan K, Ryan CA, Wiesner R and Troll W. Chymotrypsin-specific protease inhibitors decrease H,O, formation by activated human polymorphonuclear leukocytes. Carcinogenesis, 2 : +,*1ῌ+,+,. +321. Fryer J, Firca J, Leventhal J, Blondin B, Malcolm A, Ivancic D, Gandhi R, Shah A, Pao W, Abecassis M, Kaufman D, Stuart Fand Anderson B. IgY antiporcine endothelial cell antibodies e#ectively block human antiporcine xenoantibody binding. Xenotransplantation, 0 : 32ῌ+*3. +333. Fujita H, Sasaki R and Yoshikawa M. Potentiation of the antihypertensive activity of orally administered ovokinin, a vasorelaxing peptide derived from ovalbumin, by emulsification in egg phosphatidylcholine. Bioscience Biotechnology and Biochemistry, /3 : ,-..ῌ,-./. +33/ b. Fujita H, Usui H, Kurahashi K and Yoshikawa M. Isolation and characterization of ovokinin, a bradykinin B+ agonist peptide derived from ovalbumin. Peptides, +0 : 12/ῌ13*. +33/ a. Gasparri A, Moro M, Curnis F, Sacchi A, Pagano S, Veglia F, Casorati G, Siccardi AG, Dellabona P and Corti A. Tumor pretargeting with avidin improves the therapeutic index of biotinylated tumor necrosis factor alpha in mouse models. Cancer Researchearch, /3 : ,3+1ῌ ,3,-. +333. 20 J. Poult. Sci., .+ (+)

Gassmann M, Thommes P, Weiser T and Hubscher U. E$cient production of chicken egg yolk antibodies against a conserved mammalian protein. The FASEB Journal, . : ,/,2ῌ,/-,. +33*. Giansanti F, Rossi P, Massucci MT, Botti D, Antonini G, Valenti P and Seganti L. Antiviral activity of ovotransferrin discloses an evolutionary strategy for the defensive activities of lactoferrin. Biochemistry and Cell Biology, 2* : +,/ῌ+-*. ,**,. Gill AO and Holley RA. Surface application of lysozyme, nisin, and EDTA to inhibit spoilage and pathogenic bacterial on ham and bologna. Journal of Food Protection, 0- : +--2ῌ+.-0. ,***. Gill AO and Holley RA. Interactive inhibition of meat spoilage and pathogenic bacterial by lysozyme, nisin and EDTA in the presence of nitrite and sodium chloride at ,. degrees C. International Journal of Food Microbiology, 2* : ,/+ῌ,/3. ,**-. Goldberg J, Shrikant P and Mescher MF. In vivo augmentation of tumor-specific CTL responses by class I/peptide antigen complexes on microspheres (large multivalent immunogen). Journal of Immunology, +1* : ,,2ῌ,-/. ,**-. Green NM. Avidin. Advances in Protein Chemistry, ,3 : 2/ῌ+--. +31/. Greengard O, A Sentenac, N Mendelsohn. Phosvitin, the iron carrier of egg yolk. Biochim Biophys Acta 3* : .*0ῌ.*1, +30.. Gutierrez MA, Takahashi H and Juneja LR. Nutritive evaluation of hen eggs. In : Hen Eggs, Their Basic and Applied Science (Yamamoto T, Juneja, LR, Hatta H and Kim M eds.). pp. ,/ῌ-/. CRC Press. New York. +331. Guttinger M, Guidi F, Chinol M, Reali E, Veglia F, Viale G, Paganelli G, Corti A and Siccardi AG. Adoptive immunotherapy by avidin-driven cytotoxic T lymphocyte-tumor bridging. Cancer Research, 0* : .,++ῌ.,+/. ,***. Han KT, Sharar SR, Phillips ML, Harlan JM and Winn RK. Sialyl Lewis(x) oligosaccharide reduces ischemia-reperfusion injury in the rabbit ear. Journal of Immunology, +// : .*++ῌ .*+/. +33/. Harasym TO, Cullis PR and Bally MB. Intratumor distribution of doxorubicin following i.v. administration of drug encapsulated in egg phosphatidylcholine/cholesterol liposomes. Cancer Chemotherapy and Pharmacology, .* : -*3ῌ-+1. +331. Hartmann C and Wilhelmson M. The hen’s egg yolk : a source of biologically active substances. World’s Poultry Science Journal, /1 : +-ῌ,2. ,**+. Hatta H, Tsuda K, Akachi S, Kim M, Yamamoto T and Ebina T. Oral passive immunization e#ect of anti-human rotavirus IgY and its behavior against proteolytic enzymes. Bioscience, Biotechnology and Biochemistry, /1 : +*11ῌ+*2+. +33-. Hatta H, Mabe K, Kim M, Yamamoto T, Gutierrez MA and Miyazaki T. Prevention of fish disease using egg yolk antibody In : Egg Uses and Processing Technologies, New Develop- ments (Sim JS and Nakai S eds.). pp. ,.+ῌ,.3. CAB International. Oxon. +33.. Hatta H, Tsuda K, Ozeki M, Kim M, Yamamoto T, Otake S, Hirosawa M, Katz J, Childers NK and Michalek SM. Passive immunization against dental plaque formation in humans : E#ect of a mouth rinse containing egg yolk antibodies (IgY) specific to Streptococcus mutans. Caries Research, -+ : ,02ῌ,1.. +331. He X, Tsang TC, Luo P, Zhang T and Harris DT. Enhanced tumor immunogenicity through coupling cytokine expression with antigen presentation. Cancer Gene Therapy, +* : 003ῌ011. ,**-. Hensel T, Amberger VR and Schwab ME. A metalloprotease activity from C0 glioma cells inactivates the myelin-associated neurite growth inhibitors and can be neutralized by antibodies. British Journal of Cancer 12 : +/0.ῌ+/1,. +332. Hincke MT. Ovalbumin is a component of the chicken eggshell matrix. Connective Tissue Research, -+ : ,,1ῌ,--. +33/. Hincke MT, Gautron J, Tsang CPW, McKee MD and Nys Y. Molecular cloning and ultrastructural localization of the core protein of an eggshell matrix proteoglycan, Ovocleidinῌ ++0. Journal of Biological Chemistry, ,1. : -,3+/ῌ-,3,-. +333. Hirota Y, Yang MP, Araki S, Yoshihara K, Furusawa S, Yasuda M, Mohamed A, Matsumoto Mine and Kovacs-Nolan : Biologically Active Components in Hen Eggs 21

Y and Onodera T. Enhancing e#ects of chicken egg white derivatives on the phagocytic response in the dog. The Journal of Veterinary Medical Science, /1 : 2,/ῌ2,3. +33/. Holen E, Bolann B and Elsayed S. Novel B and T cell epitopes of chicken ovomucoid (Gal d +) induce T cell secretion of IL-0,IL-+-, and IFN-gamma. Clinical and Experimental Allergy, -+ : 3/,ῌ30.. ,**+. Hytonen VP, Laitinen OH, Grapputo A, Kettunen A, Savolainen J, Kalkkinen N, Martilla AT, Norlund HR, Nyholm TKM, Paganelli G and Kulomaa MK. Characterisation of poultry egg-white avidins and their potential as tools in pretargeting cancer treatment. Biochemical Journal, -1, : ,+3ῌ,,/. ,**-. Ibrahim HR. Insights into the structure-function relationships of ovalbumin, ovotransferrin, and lysozyme. In : Hen Eggs, Their Basic and Applied Science (Yamamoto T, Juneja LR, Hatta Hand Kim M eds.). pp. -1ῌ/0. CRC Press, Inc. New York. +331. Ibrahim HR. Ovotransferrin. In : Natural Food Antimicrobial Systems (Naidu AS ed.). pp. ,++ῌ ,,0. CRC Press, Inc. New York. ,***. Ibrahim HR, Sugimoto Y and Aoki T. Ovotransferrin antimicrobial peptide (OTAP-3,) kills bacteria through a membrane damage mechanism. Biochimica et Biophysica Acta, +/,- : +30ῌ,*/. ,***. Ibrahim HR, Thomas U and Pellegrini A. A helix-loop peptide at the upper lip of the active site cleft of lysozyme confers potent antimicrobial activity with membrane permeabilization action. The Journal of Biological Chemistry, ,10 : .-101ῌ.-11.. ,**+. Ijiri Y, Yamamoto T, Kamata R, Aoki H, Matsumoto K, Okamura R and Kambara T. The role of Pseudomonas aeruginosa elastase in corneal ring abscess formation in pseudomonal keratitis. Graefes Archive for Clinical and Experimental Ophthalmology, ,-+ : /,+ῌ/,2. +33-. Ikai A, Nakashima M and Y Aoki. Inhibition of inﬂammatory proteinases, medullasin, by alpha ,-macroglobulin and ovomacrogloulin. Biochemical and Biophysical Research Communica- tions, +/2 : 2-+ῌ2-0. +323. Ikemori Y, Kuroki M, Peralta RC, Yokoyama Hand Kodama Y. Protection of neonatal calves against fatal enteric colibacillosis by administration of egg yolk powder from hens immunized with K33-piliated enterotoxigenic Escherichia coli. American Journal of Veterinary Research, /- : ,**/ῌ,**2. +33,. Ikemori Y, Ohta M, Umeda K, Jr Icatlo FC, Kuroki M, Yokoyama Hand Kodama Y. Passive protection of neonatal calves against bovine coronavirus-induced diarrhea by administration of egg yolk or colostrum antibody powder. Veterinary Microbiology, /2 : +*/ῌ+++. +331. Imberechts H, Deprez P, Van Driessche E and Pohl P. Chicken egg yolk antibodies against F+2 ab ﬁmbriae of Escherichia coli inhibit shedding of F+2 positive E. coli by experimentally infected pigs. Veterinary Microbiology, /. : -,3ῌ-.+. +331. Itoh T, Miyazaki J, Sugawara Hand Adachi S. Studies on the characterization of ovomucin and chalaza of the hen’s egg. Journal of Food Science, /, : +/+2ῌ+/,+. +321. Jiang B and Mine Y. Preparation of novel functional oligophosphopeptides from hen egg yolk phosvitin. Journal of Agricultural and Food Chemistry, .2 : 33*ῌ33.. ,***. Jiang B and Mine Y. Phosphopeptides derived from hen egg yolk phosvitin : e#ect of molecular size on the calcium-binding properties. Bioscience, Biotechnology and Biochemistry, 0/ : ++21ῌ++3*. ,**+. Jiang Y, Noh SK and Koo SI. Egg phosphatidylcholine decreases the lymphatic absorption of cholesterol in rats. Journal of Nutrition, +-+ : ,-/2ῌ,-0-. ,**+. Jin LZ, Samuel K, Baidoo K, Marquardt RR and Frohlich AA. In vitro inhibition of adhesion of enterotoxigenic Escherichia coli K22 to piglet intestinal mucus by egg yolk antibodies. FEMS Immunology and Medical Microbiology, ,+ : -+-ῌ-,+. +332. Juneja LR, Koketsu M, Nishimoto K, Kim M, Yamamoto T and Itoh T. Large scale preparation of sialic acid from chalaza and egg yolk membrane. Carbohydrate Research, ,+. : +13ῌ+20. +33+. Juneja LR, Koketsu M, Kawanami H, Sasaki K, Kim M and Yamamoto T. Large-scale preparation of sialic acid and its derivatives from chalaza and delipidated yolk. In : Egg Uses 22 J. Poult. Sci., .+ (+)

and Processing Technologies, New Developments (Sim JS and Nakai S eds.). pp. +3,ῌ,*0. CAB International. Oxon. +33.. Juneja LR. Egg yolk lipids. In : Hen Eggs, Their Basic and Applied Science (Yamamoto T, Juneja LR, Hatta H and Kim M eds.). pp. 1-ῌ32. CRC Press, Inc. New York. +331. Kato I, Schrode J, Kohr WJ and Laskowski Jr M. Chicken ovomucoid : determination of its amino acid sequence, determination of the trypsin reactive site, and preparation of all three domains. Biochemistry, ,0 : +3-ῌ,*+. +321. Kato T, Imatani T, Miura T, Minaguchi K, Saitoh E and Okuda K. Cytokine-inducing activity of family , cystatins. Biological Chemistry, -2+ : ++.-ῌ++.1. ,***. Kijowski J, Lesnierowski G and Fabisz-Kijowska A. Lysozyme polymer formation and functionality of residuals after lysozyme extraction. In : Egg Nutrition and Biotechnology (Sim JS, Nakai S and Guenter W eds.). pp. ,03ῌ,2/. CAB International. Oxon. ,***. Kitamoto T, Nakashima M and Ikai A. Hen egg white ovomacroglobulin has a protease inhibitory activity. Journal of Biochemistry, 3, : +013ῌ+02,. +32,. Kobayashi K, Han M, Waterai S and Yasuda T. Availability of liposomes as drug carriers to the brain. Acta Medica Okayama, /* : 01ῌ1,. +330. Koketsu M, Juneja LR, Kawanami H, Kim M and Yamamoto T. Preparation of N- acetylneuraminic acid from delipidated egg yolk. Glycoconjugate Journal, 3 : 1*ῌ1.. +33,. Koketsu M, Nitoda T, Juneja LR, Kim M, Kashimura N and Yamamoto T. Sialyloligosaccharides from egg yolk as an inhibitor of rotaviral infection. Journal of Agricultural and Food Chemistry, .- : 2/2ῌ20+. +33/ a. Koketsu M, Seko A, Juneja LR, Kim M, Kashumura N, Yamamoto T. An e$cient preparation and strucutral chracterization of sialylglycopeptides from protease treated egg yolk. Journal of Carbohydrate Chemistry, +. : 2--ῌ2.+. +33/ b. Kollberg H, Carlander D, Olesen H, Wejaker PE, Johannesson M and Larsson A. Oral administration of specific yolk antibodies (IgY) may prevent Pseudomonas aeruginosa infections in patients with cystic fibrosis : A phase Ifeasibility study. Pediatr Pulmonol, -/ : .--ῌ..*. ,**-. Konduri SD, Yanamandra N, Siddique K, Joseph A, Dinh DH, Olivero WC, Gujrati M, Kouraklis G, Swaroop A, Kyritsis AP and Rao JS. Modulation of cystatin C expression impairs the invasive and tumorigenic potential of human glioblastoma cells. Oncogene, ,+ : 21*/ῌ21+,. ,**,. Korpela J, Salonen E-M, Kuusela P, Sarvas M and Vaheri A. Binding of avidin to bacteria and to the outer membrane porin of Escherichia coli. FEMS Microbiology Letters, ,, : -ῌ+*. +32.. Kovacs-Nolan J, Sasaki E, Yoo D and Mine Y. Cloning and expression of human rotavirus spike protein, VP2*, in Escherichia coli. Biochemical and Biophysical Research Communications, ,2, : ++2-ῌ++22. ,**+. Krol J, Kopitz C, Kirschenhofer A, Schmitt M, Magdolen U, Kruger A and Magdolen V. Inhibition of intraperitoneal tumor growth of human ovarian cancer cells by bi- and trifunctional inhibitors of tumor-associated proteolytic systems. Biological Chemistry, -2. : +*31ῌ++*,. ,**- a. Krol J, Sato S, Rettenberger P, Assfalg-Machleidt I, Schmitt M, Magdolen V and Magdolen U. Novel bi- and trifunctional inhibitors of tumor-associated proteolytic systems. Biological Chemistry, -2. : +*2/ῌ+*30. ,**- b. Kuroki M, Ohta Y, Ikemori Y, Peralta RC, Yokoyama H and Kodama Y. Passive protection against bovine rotavirus in calves by specific immunoglobulins from chicken egg yolk. Archives of Virology, +-2 : +.-ῌ+.2. +33.. Kweon C-H, Kwon B-J, Woo S-R, Kim J-M, Woo G-H, Son D-H, Hur W and Lee Y-S. Immunoprophylactic e#ect of chicken egg yolk immunoglobulin (IgY) against porcine epidemic diarrhea virus (PEDV) in piglets. The Journal of Veterinary Medical Science, 0, : 30+ῌ30.. ,***. LeClaire RD, Hunt RE and Bavari S. Protection against bacterial superantigen staphylococcal enterotoxin B by passive immunization. Infection and Immunity, 1* : ,,12ῌ,,2+. ,**,. Mine and Kovacs-Nolan : Biologically Active Components in Hen Eggs 23

Lee SB, Mine Y and Stevenson RMW. E#ects of hen egg yolk immunoglobulin in passive protection of rainbowtrout against Yersinia ruckeri. Journal of Agricultural and Food Chemistry, .2 : ++*ῌ++/. ,***. Lee EN, Sunwoo HH, Menninen K and Sim JS. In vitro studies of chicken egg yolk antibody (IgY) against Salmonella enteritidis and Salmonella typhimurium. Poultry Science, 2+ : 0-,ῌ0.+. ,**,. Lee-Huang S, Huang PL, Sun Y, Huang PL, Kung HF, Blithe DL and Chen HC. Lysozyme and RNases as anti-HIV components in beta-core preparations of human chorionic gonadotropin. Proceedings of the National Academy of Sciences U.S.A., 30 : ,012ῌ,02+. +333. Leventhal JR, Su A, Kaufman DB, Abecassis MI, Stuart FP, Anderson B and Fryer JP. Altered infectivity of porcine endogenous retrovirus by ῌprotective” avian antibodies : Implications for pig-to-human xenotransplantation. Transplant Proceedings, -- : 03*. ,**+. Li-Chan E and Nakai S. Biochemical basis for the properties of egg white. Critical Reviews in Poultry Biology, , : ,+ῌ/2. +323. Li-Chan ECY, Powrie WD and Nakai S. The Chemistry of eggs and egg products. In : Egg Science and Technology, .th Editon (Stadelman WJ and Cotterill OJ eds.). pp. +*/ῌ+1/. The Haworth Press, Inc.. New York. +33/. Lim SY and Suzuki H. Intakes of dietary docosahexaenoic acid ethyl ester and egg phosphatidylcholine improve maze-learing ability in young and old mice. Journal of Nutri- tion, +-* : +0,3ῌ+0-,. ,***. Linck LM, Lin DS, Flavell D, Connor WE and Steiner RD. Cholesterol supplementation with egg yolk increases plasma cholesterol and decreases plasma 1-dehydrocholesterol in Smith-Lemli- Opitz syndrome. American Journal of Medical Genetics, 3- : -0*ῌ-0/. ,***. Liu Y and Watson R. Reducing infection in infants with egg phospholipids. In : Eggs and Health Promotion (Watson RR ed.). pp. //ῌ0*. Iowa State Press. Ames. ,**,. Losso JN, Nakai S and Charter EA. Lysozyme. In : Natural Food Antimicrobial Systems (Naidu AS ed.). pp. +2/ῌ,+*. CRC Press, Inc. NewYork. ,***. Lowe JB, Stoolman LB, Nair RP, Larsen RD, Berhend TL and Marks RM. ELAM-+-dependent cell adhesion to vascular endothelium determined by a transfected human fucosyltransferase cDNA. Cell, 0- : .1/ῌ.2.. +33*. Lu CL and Baker R. Characteristics of egg yolk phosvitin as an antioxidant for inhibiting metal-catalyzed phospholipid oxidations. Poultry Science, 0/ : ,*0/ῌ,*1*. +320. Maeda H, Akaike T, Sakata Y and Maruo K. Role of bradykinin in microbial infection : enhancement of septicemia by microbial proteases and kinin. Agents Actions Supplementum, ., : +/3ῌ+0/. +33-. Makrides M, Hawkes JS, Neumann MA and Gibson RA. Nutritional e#ect of including egg yolk in the weaning diet of breast-fed and formula-fed infants : a randomized controlled trial. American Journal of Clinical Nutrition, 1/ : +*2.ῌ+*3,. ,**,. Maliar T, Balaz S, Tandlich R and Sturdik E. Viral proteinases-possible targets of antiviral drugs. Acta Virologica, .0 : +-+ῌ+.*. ,**,. Marquardt RR, Jin LZ, Kim JW, Fang L, Frohlich A and Baidoo SK. Passive protective e#ect of egg yolk antibodies against enterotoxigenic Escherichia coli K22῍ infection in neonatal and early weaned piglets. FEMS Immunology and Medical Microbiology, ,- : ,2-ῌ,22. +333. Martin WG, Tattrie NH and Cook WH. Lipid extraction and distribution studies of egg yolk lipoproteins. Canadian Journal of Biochemistry and Physiology, .+ : 0/1ῌ000. +30-. Maruo K, Akaike T, Ono T and Maeda H. Involvement of bradykinin generation in intravascular dissemination of Vibrio vulniﬁcus and prevention of invasion by a bradykinin antagonist. Infection and Immunity, 00 : 200ῌ203. +332. Masuda Y, Kokubu T, Yamashita M, Ikeda H and Inoue S. Egg phosphatidylcholine combined with vitamin B+, improved memory impairment following lesioning of nucleus basalis in rats. Life Sciences, 0, : 2+-ῌ2,,. +332. Matoba N, Usui H, Fujita H and Yoshikawa M. A novel anti-hypertensive peptide derived from 24 J. Poult. Sci., .+ (+)

ovalbumin induces nitric oxide-mediated vasorelaxation in an isolated SHR mesenteric artery. FEBS Letters, ./, : +2+ῌ+2.. +333. Matoba N, Yamada Y, Usui H, Nakagiri R and Yoshikawa M. Designing potent derivatives of ovokinin(,ῌ1), an anti-hypertensive peptide derived from ovalbumin. Bioscience Biotechnol- ogy and Biochemistry, 0/ : 1-0ῌ1-3. ,**+. Matsushima K. An undescribed trypsin inhibitor in egg white. Science, +,1 : ++12ῌ++13. +3/2. Mine Y. Recent advances in egg protein functionality in the food system. World’s Poultry Science Journal, /2 : -+ῌ-3. ,**,. Miyagawa S, Kamata R, Matsumoto K, Okamura R and Maeda H. Inhibitory e#ects of ovomacroglobulin on bacterial keratitis in rabbits. Graefe’s Archive for Clinical and Exper- imental Ophthalmology, ,,3 : ,2+ῌ,20. +33+ a. Miyagawa S, Matsumoto K, Kamata R, Okamura R and Maeda H. Spreading of Serratia marcescens in experimental keratitis and growth suppression by chicken egg white ovomacroglobulin. Japanese Journal of Ophthalmology, -/ : .*,ῌ.+*. +33+ b. Miyagawa S, Nishino N, Kamata R, Okamura R and Maeda H. E#ects of protease inhibitors on growth of Serratia marcescens and Pseudomonas aeruginosa. Microbial Pathogenesis, ++ : +-1ῌ+.+. +33+ c. Miyagawa S, Kamata R, Matsumoto K, Okamura R and Maeda H. Therapeutic intervention with chicken egg white ovomacroglobulin and a new quinolone on experimental Pseudomonas keratitis. Graefe’s Archive for Clinical and Experimental Ophthalmology, ,-, : .22ῌ.3-. +33.. Molla A, Matsumura Y, Yamamoto T, Okamura R and Maeda H. Pathogenic capacity of proteases from Serratia marcescens and Pseudomonas aeruginosa and their suppression by chicken egg white ovomacroglobulin. Infection and Immunity, // : ,/*3ῌ,/+1. +321. Moro M, Pelagi M, Fulci G, Paganelli G, Dellabona P, Casorati G, Siccardi AG and Corti A. Tumor cell pretargeting with antibody-avidin complexes and biotinylated tumor necrosis factor alpha. Cancer Research, /1 : +3,ῌ+3,2. +331. Morris ER and Greene FE. Utilization of the iron of egg yolk for hemoglobin formation by the growing rat. Journal of Nutrition, +*, : 3*+ῌ3*2. +31,. Muehlenweg B, Assfalg-Machleidt I, Parrado SG, Burgle M, Creutzburg S, Schmitt M, Auerswald EA, Machleidt W and Magdolen V. A novel type of bifunctional inhibitor directed against proteolytic activity and receptor/ligand interaction. Cystatin with a urokinase receptor binding site. The Journal of Biological Chemistry, ,1/ : --/0,ῌ--/00. ,***. Mulligan MS, Paulson JC, De Frees S, Zheng ZL, Lowe JB and Ward PA. Protective e#ects of oligosaccharides in P-selectin-dependent lung injury. Nature, -0. : +.3ῌ+/+. +33-. Murata M, Imaizumi K and Sugano M. E#ect of dietary phospholipids and their constituent bases on serum lipids and apolipoproteins in rats. Journal of Nutriton, ++, : +2*/ῌ+2*2. +32,. Nagaoka S, Masaoka M, Zhang Q, Hasegawa M and Watanabe K. Egg ovomucin attenuates hypercholesterolemia in rats and inhibits cholesterol absorption in Caco-, cells. Lipids, -1 : ,01ῌ,1,. ,**,. Nakai S. Molecular modiﬁcations of egg proteins for functional improvement. In : Egg Nutrition and Biotechnology (Sim JS, Nakai S and Guenter W eds.). pp. ,*/ῌ,+1. CAB International. Oxon. ,***. Nagai A, Terashima M, Harada T, Shimode K, Takeuchi H, Murakawa Y, Nagasaki M, Nakano A and Kobayashi S. Cathepsin B and H activities and cystatin C concentrations in cerebrospinal ﬂuid from patients with leptomeningeal metastasis. Clinica Chimica Acta, -,3 : /-ῌ0*. ,**-. Nattress FM, Yost CK and Baker LP. Evaluation of the ability of lysozyme and nisin to control meat spoilage bacteria. International Journal of Food Microbiology, 1* : +++ῌ++3. ,**+. O’Farrelly C, Branton D and Wanke CA. Oral ingestion of egg yolk immunoglobulin from hens immunized with an enterotoxigenic Escherichia coli strain prevents diarrhea in rabbits challenged with the same strain. Infection and Immunity, 0* : ,/3-ῌ,/31. +33,. Mine and Kovacs-Nolan : Biologically Active Components in Hen Eggs 25

Ofuji Y, Suzuki T, Yoshie H, Hara K and Adachi M. The e#ects of ovomacroglobulin on gingival wound healing in rats. Periodontal Clinical Investigations, +. : +-ῌ,,. +33,. Ogawa M, Nakamura S, Scaman CH, Jing H, Kitts DD, Dou J and Nakai S. Enhancement of proteinase inhibitory activity of recombinant human cystatin C using random-centroid optimization. Biochimica Biophysica Acta, +/33 : ++/ῌ+,.. ,**,. Oguro T, Ohaki Y, Asano G, Ebina T and Watanabe K. Ultrastructural and immunohistochem- ical characterization on the e#ect of ovomucin in tumor angiogenesis. Japanese Journal of Clinical and Electron Microscopy, -- : 23ῌ33. ,**+. Okubo T, Akachi S and Hatta H. Structure of hen eggs and physiology of egg laying. In : Hen Eggs, Their Basic and Applied Science (Yamamoto T, Juneja LR, Hatta H and Kim M eds.). pp. +ῌ+,. CRC Press, Inc. NewYork. +331. Omi N and Ezawa I. E#ect of eggshell Ca on preventing of bone loss after ovariectomy. Journal of Home Economics of Japan, .3 : ,11ῌ,2,. +332. Otake S, Nishihara Y, Makimura M, Hatta H, Kim M, Yamamoto T and Hirasawa M. Protection of rats against dental caries by passive immunization with hen egg yolk antibody (IgY). Journal of Dental Research, 1* : +0,ῌ+00. +33+. Otani H and Odashima M. Inhibition of proliferative responses of mouse spleen lymphocytes by lacto- and ovotransferrins. Food and Agricultural Immunology 3 : +3-ῌ,*,, +331. Pacor S, Giacomello E, Bergamo A, Clerici K, Zacchigna M, Boccu E and Sava G. Antimetastatic action and lymphocyte activation by the modified lysozyme mPEG-Lyso in mice with MCa mammary carcinoma. Anticancer Research, +0 : ,//3ῌ,/0.. +330. Pacor S, Gagliardi R, Di Daniel E, Vadori M and Sava G. In vitro down regulation of ICAM- + and E-cadherin and in vitro reduction of lung metastases of TS/A adenocarcinoma by a lysozyme derivative. International Journal of Molecular Medicine, . : -03ῌ-1/. +333. Pellegrini A, Thomas U, Bramaz N, Klauser S, Hunziker P and von Fellenberg R. Identification and isolation of a bactericidal domain in chicken egg white lysozyme. Journal of Applied Microbiology, 2, : -1,ῌ-12. +331. Pellegrini A, Thomas U, Wild P, Schraner E, and von Fellenberg R. E#ect of lysozyme or modified lysozyme fragments on DNA and RNA synthesis and membrane permeability of Escherichia coli. Microbiological Research, +// : 03ῌ11. ,***. Peralta RC, Yokoyama H, Ikemori Y, Kuroki M and Kodama Y. Passive immunization against experimental salmonellosis in mice by orally administered hen egg yolk antibodies specific for +.-kDa fimbriae of Salmonella enteritidis. Journal of Medical Microbiology, .+ : ,3ῌ-/. +33.. Phillips ML, Nudelman E, Gaeta FC, Periz M, Singhal AK, Hakomori S and Paulson JC. ELAM-+ mediates cell adhesion by recognition of a carbohydrate ligand, sialyl-Lex. Science, ,/* : ++-*ῌ++-,. +33*. Plate NA, Valuev IL, Sytov GA and Valuev LI. Mucoadhesive polymers with immobilized proteinase inhibitors for oral administration of protein drugs. Biomaterials, ,- : +01-ῌ+011. ,**,. Premzl A, Puizdar V, Zavasnik-Bergant V, Kopitar-Jerala N, Lah TT, Katunuma N, Sloane BF, Turk V and Kos J. Invasion of ras-transformed breast epithelial cells depends on the proteolytic activity of cysteine and aspartic proteinases. Biological Chemistry, -2, : 2/-ῌ 2/1. ,**+. Rabinowich H, Lyte M, Steiner Z, Klajman A and Shinitzky M. Augmentation of mitogen responsiveness in the aged by a special lipid diet AL 1,+. Mechanisms of Ageing Develop- ment, .* : +-+ῌ+-2. +321. Rada JA, Huang Y and Rada KG. Identification of choroidal ovotransferrin as a potential ocular growth regulator. Current. Eye Research, ,, : +,+ῌ+-,. ,**+. Rhodes DN and Lea CH. Phospholipids .. On the composition of hen’s egg phospholipids. Biochemical Journal, /. : /,0ῌ/--. +3/1. Romano# AL and Romano# AJ. The avian egg. Wiley, NewYork. +3.3. Rovensky JJ, Marek O, Schreiberova O and Stancikova M. Somatomadin-type activity of Biomin H. Casopis Lekaru Ceskych, +-- : ,+-ῌ,+.. +33.. 26 J. Poult. Sci., .+ (+)

Salton MJR. The properties of lysozyme and its action on microorganisms. Bacteriology Reviews, ,+ : 2,ῌ32. +3/1. Sato R, Lee YS, Noguchi T and Naito H. Iron solubility in the small intestine of rats fed egg yolk protein. Nutrition Reports International, -* : +-,3ῌ+-,0. +32.. Sato R, Noguchi T and Naito H. The formation and Iron binding property of phosphopeptides in the small intestinal contents of rats fed egg yolk diet. Nutrition Reports International, -+ : ,./ῌ,/,. +32/. Sato R, Noguchi T and Naito H. The e#ect of feeding demineralized egg yolk protein on the solubility of intra-intestinal Iron. Nutriton Reports International, -0 : /3-ῌ0*,. +321. Sava G. Reduction of B+0 melanoma metastases by oral administration of egg-white lysozyme. Cancer Chemotherapy and Pharmacology, ,/ : ,,+ῌ,,,. +323. Sava G, Benetti A, Ceschia V and Pacor S. Lysozyme and cancer : role of exogenous lysozyme as anticancer agent (review). Anticancer Research, 3 : /2-ῌ/3+. +323. Sava G, Ceschia V, Pacor S and Zabucchi G. Observations on the antimetastatic action of lysozyme in mice bearing Lewis lung carcinoma. Anticancer Research, ++ : ++*3ῌ+++-. +33+. Sava G, Pacor S, Dasic G and Bergamo A. Lysozyme stimulates lymphocyte response to ConA and IL-, and potentiates /-ﬂuorouracil action on advanced carcinomas. Anticancer Re- search, +/ : +22-ῌ+222. +33/. Sava G. Pharmacological aspects and therapeutic applications of lysozymes. EXS 1/ : .--ῌ..3. +330. Saxena I and Tayyab S. Protein proteinases inhibitors from avian egg whites. Cellular and Molecular Life Sciences, /- : +-ῌ,-. +331. Schaafsma A and Beelen GM. Eggshell powder, a comparable or better source of calcium than puriﬁed calcium carbonate : piglet studies. Journal of the Science of Food and Agriculture, 13 : +ῌ/. +333. Schaafsma A and Pakan I. E#ect of a chicken eggshell powder enriched dairy product on bone mineral density in person with osteoporosis or osteopenia. Nutrition, +/ : +/1. +333. Schaafsma A, Pakan I, Hofstede GJ, Muskiet, FA, Van Der Veer, E and De Vries PJ. Mineral, amino acid, and hormonal composition of chicken eggshell powder and the evaluation of its use in human nutrition. Poultry Science, 13 : +2--ῌ+2-2. ,***. Schauer R. Analysis of sialic acids. Methods in Enzymology, +-2 : +-,ῌ+0+. +321. Schimmoller F, Higaki JNand Cordell B. Amyloid forming proteases : therapeutic targets for Alzheimer’s disease. Current Pharmaceutical Design, 2 : ,/,+ῌ,/-+. ,**,. Sharma D, Chelvi TP, Kaur J and Ralhan R. Thermosensitive liposomal taxol formation : heat-mediated targeted drug delivery in murine melanoma. Melanoma Research, 2 : ,.*ῌ ,... +332. Sheppard EH. Specialty eggs : n-- fatty acid-enriched eggs. In : Eggs and Health Promotion Eggs and Health Promotion (Watson RR ed.). pp. -1ῌ.., Iowa State Press. Ames.,**,. Shin JH, Yang M, Nam SW, Kim JT, Myung NH, Bang WG and Roe IH. Use of egg yolk-derived immunoglobulin as an alternative to antibiotic treatment for control of Helicobacter pylori infection. Clinical and Diagnostic Laboratory Immunology, 3 : +*0+ῌ +*00. ,**,. Shinohara K, Kakiuchi T, Kobori M, Suzuki M and Fukushima T. Europium-liked immunoassay of IgM production by human-human hybridomas cultured with or without chicken egg yolk lipoprotein. Bioscience, Biotechnology and Biochemistry, /1 : +*+,ῌ+*+-, +33-. Sim JS, Sunwoo HH and Lee EN. Ovoglobulin IgY. In : Natural Food Antimicrobial Systems (Naidu AS ed.). pp. ,,1ῌ,/,. CRC Press. New York. ,***. Smith DJ, King WF and Godiska R. Passive transfer of immunoglobulin Y to Streptococcus mutans glucan binding protein B can confer protection against experimental dental caries. Infection and Immunity, 03 : -+-/ῌ-+.,. ,**+. Sugahara T, Murakami F, Yamada Y and Sasaki T. The mode of actions of lysozyme as an immunoglobulin production stimulating factor. Biochimica et Biophysica Acta, +.1/ : ,1ῌ -.. ,***. Mine and Kovacs-Nolan : Biologically Active Components in Hen Eggs 27

Sugino H, Nitoda T and Juneja LR. General chemical composition of hen eggs. In : Hen Eggs, Their Basic and Applied Science (Yamamoto T, Juneja LR, Hatta H and Kim M eds.). pp. +-ῌ,.. CRC Press, Inc. New York. +331. Sugita-Konishi Y, Shibata K, Yun SS, Yukiko HK, Yamaguchi K and Kumagai S. Immune functions of immunoglobulin Y isolated from egg yolk of hens immunized with various infectious bacteria. Bioscience, Biotechnology and Biochemistry, 0* : 220ῌ222. +330. Sugita-Konishi Y, Ogawa M, Arai S, Kumagai S, Igimi S, and Shimizu M. Blockade of Salmonella enteritidis passage across the basolateral barriers of human intestinal epithelial cells by speciﬁc antibody. Microbiology and Immunology, .. : .1-ῌ.13. ,***. Sugita-Konishi Y, Sakanaka S, Sasaki K, Juneja LR, Noda T and Amano F. Inhibition of bacterial adhesion and Salmonella infection in BALB/c mice by sialyloligosaccharides and their derivatives from chicken egg yolk. Journal of Agricultural and Food Chemistry, /* : -0*1ῌ-0+-. ,**,. Suguro N, Horiike S, Masuda Y, Kunou M and Kokubu T. Bioavailability and commercial use of eggshell calcium, membrane proteins and yolk lecithin products. In : Egg Nutrition and Biotechnology (Sim JS, Nakai N and Guenter W eds.). pp. ,+3ῌ,-,. CABInternational. Oxon. ,***. Suzuki M, Shinohara H, Yonekura M, Tsutsumi M and Shinohara K. Growth promoting e#ect of chicken egg yolk lipoproteins on human lymphocytic cell lines. Nippon Shokuhin Kogyo Gakkaishi, .+ : -1ῌ.,. +33.. Taborsky G and Mok C. Phosvitin : homogeneity of molecular weight. The Journal of Biological Chemistry, ,., : +.3/ῌ+/*+. +301. Tanizaki H, Tanaka H, Iwata H and Kato A. Activation of macrophages by sulfated glycopeptides in ovomucin, yolk membrane, and chalazae in chicken eggs. Bioscience, Biotechnology and Biochemistry, 0+ : +22-ῌ+223. +331. Tenuovo J. Clinical applications of antimicrobial host proteins lactoperoxidase, lysozyme and lactoferrin in xerostomia : e$cacy and safety. Oral Diseases, 2 : ,-ῌ,3. ,**,. Terato K, Ye XJ, Miyahara H, Cremer MA and Gri$ths MM. Induction by chronic autoimmune arthritis in DBA/+ mice by oral administration of type II collagen and Escherichia coli lipopolysaccharide. British Journal of Rheumatology, -/ : 2,2ῌ2-2. +330. Tezuka H and Yoshikawa M. Abstract of Annual Meeting of Japan Society for Bioscience, Biotechnology, and Agrochemistry, Tokyo, +33/,pp.+0-. Thalley BS and Carroll SB. Rattlesnakes and scorpion antivenoms from the egg yolk of immunized hens. Biotechnology, 2 : 3-.ῌ3-1. +33*. Tomimatsu Y, Clary JJ and Bartulovich JJ. Physical characterization of ovoinhibitor, a trypsin and chymotrypsin inhibitor from chicken egg white. Archives of Biochemistry and Biophys- ics, ++/ : /-0ῌ/... +300. Tsugawa N, Okano T, Higashino R, Kimura T, Oshio Y, Teraoka Y, Igarashi C, Ezawa I and Kobayashi T. Bioavailability of calcium carbonate, DL-calcium lactate, L-calcium lactate and powdered oyster shell calcium in vitamin D-deﬁcient or -replete rats. Biological and Pharmaceutical Bulletin, / : 011ῌ02,. +33/. Tsuge Y, Shimoyamada M and Watanabe K. Binding of egg white proteins to viruses. Bioscience, Biotechnology and Biochemistry, 0* : +/*-ῌ+/*.. +330 a. Tsuge Y, Shimoyamada M and Watanabe K. Di#erences in hemagglutination inhibition activity against bovine rotavirus and hen newcastle disease virus based on the subunits in hen egg white ovomucin. Bioscience, Biotechnology and Biochemistry, 0* : +/*/ῌ+/*0. +330 b. Tsuge Y, Shimoyamada M and Watanabe K. Structural features of newcastle disease virus- and anti-ovomucin antibody-binding glycopeptides from pronase-treated ovomucin. Journal of Agricultureal and Food Chemistry ./ : ,-3-ῌ,-32. +331 a. Tsuge Y, Shimoyamada M and Watanabe K. Bindings of ovomucin to newcastle disease virus and anti-ovomucin antibodies and its heat stability based on binding abilities. Journal of Agricultural and Food Chemistry, ./ : .0,3ῌ.0-.. +331 b. Vadehra DV, Baker RC and Naylor HB. Distribution of lysozyme activity in the exteriors of egg from Gallus gallus. Comparative Biochemistry and Physiology, .- : /*-ῌ/*2. +31,. 28 J. Poult. Sci., .+ (+)

Valenti P, Antonini G, Von Hunolstein C, Visca P, Orsi N and Antonini E. Studies of the antimicrobial activity of ovotransferrin. International Journal of Tissue Reactions, / : 31ῌ +*/. +32-. Valenti P, Visca P, Antonini G and Orsi N. Antifungal activity of ovotransferrin towards genus Candida. Mycopathologia, 23 : +03ῌ+1/. +32/. Valueva TA, Mateev NL, Mosolov VV and Penin VA. Protein proteinase inhibitor therapy in experimental pancreatitis : pharmacological characterization of the inhibitor. Agents Ac- tions Supplementum, -2 (Pt ,):,*-ῌ,+*. +33,. Verdot L, Lalmanach G, Vercruysse V, Hartmann S, Lucius R, Hoebeke J, Gauthier F and Vray B. Cystatins up-regulate nitric oxide release from interferon-gamma-activated mouse peritoneal macrophages. The Journal of Biological Chemistry, ,1+ : ,2*11ῌ,2*2+. +330. Verdot L, Lalmanach G, Vercruysse V, Hoebeke J, Gauthier F, and Vray B. Chicken cystatin stimulates nitric oxide release from interferon-gamma-activated mouse peritoneal macrophages via cytokine synthesis. European Journal of Biochemistry, ,00 : ++++ῌ+++1. +333. Vered M, Gertler A and Berstein Y. Inhibition of porcine elastase II by chicken ovoinhibitor. International Journal of Peptide and Protein Research, +2 : +03ῌ+13. +32+. Vidovic D, Graddis T, Chen F, Slagle P, Diegel M, Stepan L and Laus R. Antitumor vaccination with HER-,-derived recombinant antigens. International Journal of Cancer, +*, : 00*ῌ00.. ,**,. Watanabe K, Tsuge Y and Shimoyamada M. Binding activities of pronase-treated fragments from egg white ovomucin with anti-ovomucin antibodies and newcastle disease virus. Journal of Agricultural and Food Chemistry, .0 : ./*+ῌ./*0. +332 a. Watanabe K, Tsuge Y, Shimoyamada M, Ogama N and Ebina T. Antitumor e#ects of pronase- treated fragments, glycopeptides, from ovomucin in hen egg white in a double grafted tumor system. Journal of Agricultural and Food Chemistry, .0 : -*--ῌ-*-2. +332 b. Winn SE and Ball HR. b-N-acetylglucosaminidase activity of the albumen layers and membranes of the chicken’s egg. Poultry Science, /. : 133ῌ2*/. +31/. Wong M, Hendrix MJC, von der Mark K, Little C and Stern R. Collagen in the egg shell membranes of the hen. Developmental Biology, +*. : ,2ῌ-0. +32.. Worledge KL, Godiska R, Barrett TA and Kink JA. Oral administration of avian tumor necrosis factor antibodies e#ectively treats experimental colitis in rats. Digestive Diseases and Sciences, ./ : ,,32ῌ,-*/. ,***. Wu J, Akaike T, Hayashida K, Okamoto T, Okuyama A and Maeda H. Enhanced vascular permeability in solid tumor involving peroxynitrite and matrix metalloproteinases. Japanese Journal of Cancer Research, 3, : .-3ῌ./+. ,**+. Xie H, Hu# GR, Hu# WE, JM Balog and Rath NC. E#ects of ovotransferrin on chicken macrophages and heterophil-granulocytes. Developmental and Comparative Immunology, ,0 : 2*/ῌ2+/. ,**,. Yamada Y, Matoba N, Usui H, Onishi K and Yoshikawa M. Design of a highly potent anti-hypertensive peptide based on ovokinin (,ῌ1). Bioscience, Biotechnology and Biochem- istry, 00 : +,+-ῌ+,+1, ,**,. Yamamoto Y, Sogo N, Iwao R and Miyamoto T. Antioxidant e#ect of egg yolk on linoleate in emulsions. Agriculture and Biological Chemistry, /. : -*33ῌ-+*.. +33*. Yamamoto Y, Omori M and Miyamoto T. Antioxidant e#ects of egg yolk on linoleic acid and lard. Agriculture and Biological Chemistry, // : ,.*-ῌ,.*.. +33+. Yang J, Jin Z, Yu Q, Yang T, Wang H and Liu L. The selective recognition of IgY for digestive system cancers. Chinese Journal of Biotechnology, +- : 2/ῌ3*. +331. Yao Z, Zhang M, Sakahara H, Saga T, Arano Y and Konishi J. Avidin targeting of intraperitoneal tumor xenografts. Journal of the National Cancer Institute, 3* : ,/ῌ,3. +332. Yokoyama H, Peralta RC, Diaz R, Sendo S, Ikemori Y and Kodama Y. Passive protective e#ect of chicken egg yolk immunoglobulins against experimental enterotoxigenic Escherichia coli infection in neonatal piglets. Infection and Immunity, 0* : 332ῌ+**1. +33,. Yokoyama H, Peralta RC, Umeda K, Hashi T, Icatlo FC, Kuroki M, Ikemori Y and Kodama Y. Mine and Kovacs-Nolan :Biologically Active Components in Hen Eggs 29

Prevention of fatal salmonellosis in neonatal calves, using orally administered chicken egg yolk Salmonella-speciﬁc antibodies. American Journal of Veterinary Research, /3 : .+0ῌ.,*. +332. Yokoyama H, Umeda K, Peralta RC, Hashi T, Icatlo F, Kuroki M, Ikemori Y and Kodama Y. Oral passive immunization against experimental salmonellosis in mice using chicken egg yolk antibodies speciﬁc for Salmonella enteritidis and S. typhimurium. Vaccine, +0 : -22ῌ-3-. +332. Yolken RH, Willoughby R, Wee SB, Misku# R and Vonderfecht S. Sialic acid glycoproteins inhibit in vitro and in vivo replication of rotaviruses. Journal of Clinical Investigations, 13 : +.2ῌ+/.. +321. Yolken RH, Leister F, Wee SB, Misku# R and Vonderfecht S. Antibodies to rotavirus in chickens’ eggs :a potential source of antiviral immunoglobulins suitable for human con- sumption. Pediatrics, 2+ : ,3+ῌ,3/. +322. Yoshikawa M and Fujita H. Studies on the optimum conditions to utilize biologically active peptides derived from food proteins. In :Developments in Food Engineering (Yano, T, Matsuno R and Nakamura K eds.). pp. +*/-ῌ+*//. Blackie Academic and Professional. New York. +33..