Agarose Gel Electrophoretic Fractionation of Serum Proteins in Adult Cattle. II. A Study Of Cows with Different Diseases

In 283 dairy cows suffering from internal disorders the serum proteins were studied by agarose gel electrophoresis supplemented by total protein and albumin determinations. The clinical diagnoses could be grouped according to protein pattern. Group 1 (abomasal displacement and traumatic muscle injury) did not appreciably affect the serum protein concentrations and represented primarily non-inflammatory diseases or diseases of non-infectious origin. Group 2 (leukosis) occupied an exceptional position, with heavy lowering of albumin unaccompanied by a corresponding rise of total globulin. The γ-globulin concentration was significantly lowered. Group 3 (acute traumatic peritonitis) represented an acute inflammatory process with increase chiefly of the α-globulins, while the γ-globulin concentration was normal. Group 4 (chronic traumatic peritonitis, summer mastitis, chronic subclinical mastitis, chronic laminitis and polyarthritis, urinary tract infection, abscess, and sub-acute-chronic pneumonia) comprised diseases chiefly of chronic inflammatory character and with infectious origin. Especially characteristic was the heavy rise of globulins in general and of γ-globulin in particular. In most cases there was a large increase also of the α- and p-globulin fractions.     

Studies of Electrophoresis on Cellulose Acetate Membrane of Serum Proteins from Normal Horses,Sheep and Pigs

A method for the rapid electrophoresis on a cellulose acetate membrane of serum proteins from horses, sheep and pigs is discussed. The various main globulin fractions in the serum of these animals were experimentally identified. Normal values for the percentage composition of serum from normal horses, sheep and pigs were calculated. In the horse there was great individual variation in the shape of the β-fraction, assumed to be due to different transferrin types. The mean value for β-globulin of 19.5 % in the horse was higher than for the other two species. The albumin percentage was highest in the sheep and lowest in the pig, 48.5 % and 43.2 % respectively. The sheep had the highest γ-globulin percentage, 22.8 %, while the horse had the lowest with 19.0 %. Finally the values were compared with corresponding figures reported by other authors and the results discussed.     

Some Effects of Protein Deficiency in Young Growing Pigs. III. Ferrokinetic Investigations

Ferrokinetic investigations have been carried out in pigs which had undergone protein starvation and rehabilitation. After long term protein deprivation haematocrit levels, blood haemoglobin levels, serum iron concentrations, serum total iron binding capacity and blood volume per kg body weight were all reduced in the deficient animals. The values for the rehabilitated animals were in the normal range. The erythrocytes of the protein starved pigs showed an increased rate of uptake of radioactive iron compared with the control pig. However, the turnover of iron as a whole was reduced greatly in the protein deprived group compared to the rehabilitated and control pigs. Accumulation of iron occurred in the livers, spleens and kidneys of the protein starved animals. The rehabilitated pigs had lower liver stores of iron than the control pig. On the basis of the data presented it has been concluded that these effects are mainly due to a shortage of amino acids for protein synthesis.     

Geographic variation in blood plasma protein concentrations of young herring gulls (Larus argentatus) and Caspian terns (Sterna caspia) from the Great Lakes and Lake Winnipeg

Relative and total amounts of plasma protein fractions are affected by infections, inflammation, and nutritional and physiological status, and are therefore important health indicators in free-living animals. Our objectives were: (1) to examine intercolony differences in plasma protein fractions in prefledgling gulls and terns; (2) to investigate relationships between plasma proteins and other physiological measures such as weight loss, growth, and immune function; and (3) to examine potential associations between organochlorine exposure and plasma proteins. During 1992, blood was collected from 3-week-old herring gull (Larus argentatus) chicks from six sites on Lakes Superior, Huron, Michigan, Erie, and Winnipeg and from 3-week-old Caspian tern (Sterna caspia) chicks from five sites on Lakes Huron, Michigan, and Ontario. These sites provided a wide gradient of organochlorine contamination. Plasma proteins were separated by high-resolution agarose gel electrophoresis and stained with Coomassie brilliant blue dye. Six major fractions were quantified: prealbumin, albumin, α-globulins, β₁-globulins, β₂-globulins, and γ-globulins. Total protein, prealbumin, albumin, and γ-globulin concentrations and the albumin/globulin ratio did not differ among sites. Total protein, albumin, and the albumin/globulin ratio were not decreased in birds experiencing food stress or weight loss. Intersite differences were found in α- and β-globulins. In gulls, β₂-globulins were positively associated with polychlorinated biphenyls (PCBs) and 1,1-dichloro-2,2-bis(p-chlorophenyl)ether (DDE). In terns, PCBs were negatively associated with α-globulins and positively associated with β₁-globulins. Additional research is needed to identify individual proteins and elucidate causal relationships between the particular protein concentrations and factors such as contaminants, growth, and condition.     

Effect of fetal calf serum and serum protein fractions on the uptake of liposomal phosphatidylcholine by rat hepatocytes in primary monolayer culture

We studied the effect of fetal calf serum and serum protein fractions on the interaction of phospholipid vesicles consisting of phosphatidylcholine, cholesterol and dicetylphosphate (molar ratio 7 : 2 : 1), with rat liver parenchymal cells in a primary monolayer culture. During incubation of such vesicles with fetal calf serum part of the labeled phosphatidylcholine is transferred to a lipoprotein particle similar to the one we identified previously as a derivative of high density lipoprotein (Scherphof, G., Roerdink, F.H., Waite, M. and Parks, J. (1978) Biochim. Biophys. Acta 542, 296–307). When the particle thus formed is incubated with the cells a transfer of the phospholipid label to the cells is observed. When vesicles are incubated with the cells in presence of serum such lipoprotein-mediated lipid transfer may conceivably contribute to the total lipid uptake observed. However, we found that the presence of fetal calf serum in the culture medium greatly diminished rather than increased the total transfer of liposomal lipid to the cells. Also bovine serum albumin and bovine β-globulins reduced this transfer, although to a lesser extent than whole serum. α-Globulins, on the other hand, were as effective as complete serum in reducing the uptake of liposomal phospholipid. A γ-globulin fraction failed to exhibit any effect on the uptake of [¹⁴C]phosphatidylcholine by the cells.All protein fractions which were able to inhibit cellular uptake of liposomal phospholipid were shown to bind to the phospholipid vesicles. Furthermore, lipid vesicles preincubated with fetal calf serum and then separated from it showed reduced transfer of labeled phosphatidylcholine to parenchymal cells.These observations were taken to suggest that the diminished uptake of liposomal lipid may be caused by a modification of the liposomal surface membrane as a result of the binding of certain serum proteins. On the other     

Tryptophanyl fluorescence of sodium dodecyl sulfate treated and 2-mercaptoethanol reduced proteins: a simple assay for tryptophan

Relative tryptophanyl fluorescence intensities of eleven different proteins (bovine liver glutamate dehydrogenase, bovine pancreas trypsin and α-chymotrypsinogen, egg white lysozyme, ovalbumin, bovine serum albumin and γ-globulin, bovine heart and rabbit muscle lactate dehydrogenases, rabbit muscle glyceraldehyde-3-phosphate dehydrogenase, and yeast alcohol dehydrogenase) were evaluated as a function of the physical state of the protein, i.e., native, denatured with intact disulfide bonds, and denatured with reduced disulfide bonds.     

The enhancement of muscle trichinosis in rabbits after immunization with heterologous serum proteins

The enhancement of muscle trichinosis in rabbits after immunization with heterologous serum proteins. International Journal for Parasitology4, 193–196. Rabbits were immunized with electrophoretically-purified rat serum albumin, β-globulin or γ-globulin in Freund's Complete Adjuvant and then inoculated with Trichinella spiralis larvae. Muscle parasitism was greatly enhanced in all the treated animals; this, plus the high levels of anti-rat and low levels of antiparasite antibody, indicates that antigenic competition can increase parasite numbers in trichinosis. Antigenic competition may play a role in the establishment of natural infections by T. spiralis by deflecting a protective immunological response by the host.     

Fractionation of Protein Mixtures Through Differential Adsorption on a Gradient of Substituted Agaroses of Increasing Hydrophobicity

Data are presented which indicate the feasibility of protein fractionation at high salt concentrations (≤ 3 M NaCl) through differential hydrophobic (non-ionic) adsorption on a series of columns of agaroses substituted with ligands of increasing hydrophobicity. By means of such a hydrophobicity gradient of connected columns, separation of mixtures of γ-globulin and serum albumin, as well as group separation of proteins in dialyzed human blood serum, has been achieved.     

Antidinitrophenyl antibodies in pigs and bulls

Using antigens fixed to cellulose (immunosorbents) the formation of antibodies at an average concentration of 1 mg per ml. of serum in pigs and bulls was shown after repeated immunization by dinitrophenylated γ-globulin in adjuvant. The amount of antibodies to the protein bearing the dinitrophenyl group was only exceptionally higher than 5% of the total amount of antibodies to the immunizing antigen. The dinitrophenyl group fixed directly to cellulose, binds only about one third of antibodies to the immunizing antigen. The antibodies were purified by precipitation with the immunizing antigen, followed by decomposition of the precipitate by a solution of dinitrophenol and separation of the mixture of the antibody, hapten and antigen on DEAE-Sephadex. The yield of antibodies was 60 to 70%. The antibodies were characterized as pure γ-globlin with a slightly lower electrophoretic heterogeneity than that of non-specific γ-globulin.     

Serum proteins in the leopard seal, Hydrurga leptonyx, in Prydz Bay, Eastern Antarctica and the coast of NSW, Australia

Blood protein analysis including total serum protein and albumin by chemical methods, fibrinogen estimation and serum protein electrophoresis (SPE) was performed on the leopard seal, Hydrurga leptonyx. The most commonly observed SPE pattern was eight fractions designated albumin, alpha(1a), alpha(1b), alpha(2a), alpha(2b), beta(1), beta(2) and gamma-globulin. Significantly higher total serum protein and albumin concentrations, as determined by chemical methods, and significantly higher alpha(2)-globulin concentrations, determined by SPE, were seen in free-ranging male seals compared to females, whilst significantly higher beta-globulin concentrations were seen in female seals. Season of sampling influenced fibrinogen and beta(2)-globulin concentrations, whereas there were no significant differences in any protein concentrations with moult status. Qualitative comparison of SPE traces of leopard seals in Antarctica with "sick" individuals in NSW, Australia revealed obvious differences, as did quantitative comparison of protein concentrations where differences in alpha(1), alpha(2), beta(1), beta(2), and gamma-globulin concentrations were seen. These findings suggest that SPE is a useful tool for investigating serum proteins in the leopard seal, with applications for the investigation of "sick" individuals and the assessment of variation in homeostasis. This technique could also be used to identify the presence of environmental stressors, subclinical disease and physiological variation within specific seal populations.     

Response of goats to partial replacement of dietary protein by a leaf meal mixture containing Leucaena leucocephala,Morus alba and Tectona grandis

This study investigated the effects of Leucaena leucocephala–Morus alba–Tectona grandis (2:1:1) based leaf meal mixture as a protein supplement to partially (50%) replace conventional supplements on the performance of female goats. Local female goats (12) were divided into three groups in a completely randomized design to receive either a leaf meal mixture (LMTM) based supplement, or groundnut cake (GNC) or soybean meal (SBM) incorporated concentrate, and wheat straw as basal diet for a period of about 4 months. Blood enzymes and immunological profile were monitored in all goats at 0, 30, 60 and 90 days of feeding. While dry matter intake (DMI) (g kg^−0.75 BW) of goats given LMTM was higher (P<0.05) than SBM, it was comparable to their counterparts fed GNC. Digestibility of nutrients, except for depressed crude protein digestibility (P<0.05) in SBM group, was comparable among dietary treatments. Nitrogen intake and fecal excretions were similar irrespective of supplements, the urinary excretion of N in goats given GNC was significantly higher than comparable excretion in goats on LMTM and SBM. Balance of N (g per day) remained positive in all goats, however it was significantly higher in animals on LMTM as compared to GNC. The DCP, TDN and ME values of the composite diets were comparable among dietary treatments. Intake of DCP, TDN and ME was close to stipulated NRC requirements. Blood enzymes (alkaline phosphatase (ALP), alanine aminotransferase (ALT) and aspartate aminotransferase (AST)) and serum protein electrophoresis pattern (albumin, α-globulin, β-globulin and γ-globulin) did not differ significantly due to the replacement of 50% dietary protein by LMTM throughout the experiment. It may be concluded that leaf meal mixture could be used inexpensively as a vegetable protein supplement to wheat straw based diets for goats without any adverse effect on voluntary intake, nutrient utilization, serum enzymes and immune status.     

Carbohydrate, protein and glycoprotein metabolism by the guinea-pig liver in chronic metribuzin poisoning

Male guinea-pigs weighing 400-600 g, 8 months old, were given metribuzin directly into the gastric lumen over a period of 30 days (20 animals) or 90 days (20 animals) 6 times a week. In the liver of the poisoned animals, the glycogen level and the AspAT and AlAT activities, while in the serum the total protein and the fractions albumin, alpha 1-globulin and gamma-globulin significantly decreased; serum glucose and the serum fractions alpha 2-globulin and beta-globulin, each showed an increase. The glycogen level in the liver, total protein, glucose as well as the alpha 1 and alpha 2 globulin fractions in the serum showed not appreciable difference between 30 and 90 days of intoxication. After 90 days of metribuzin treatment AspAT and AlAT dropped in the liver and rose in the serum, in comparison to the 30-day values. As to the parameters of glycoprotein metabolism, the intoxicated animals showed a significant decrease and increase in concentration of hexosamines and sialic acids in the liver and serum, respectively. Metribuzin intoxication also cause a significant decrease in activity of glucosamine phosphate isomerase and significant increase in activity of glycosidases in the liver. The results suggest that metribuzin disturbs the metabolism of carbohydrates, proteins and glycoproteins in the guinea-pig liver.     

Purification of human alpha-2-macroglobulin by chromatography on Cibacron Blue Sepharose.

A simple and efficient procedure has been devised for the isolation of α-2-macroglobulin from human plasma (type 1-1 haptoglobulin). The primary step is gel filtration and affinity chromatography on Cibacron Blue Sepharose, which selectively removes albumin and retards lipoproteins and γ-globulin, while effecting the molecular sieving of the remainder of the plasma proteins. This results in the separation of about 40% of the α-2-macroglobulin as a homogeneous component. A second step, gel filtration on Ultrogel AcA 22, may be utilized to separate α-2-macroglobulin in contaminated fractions obtained after Cibacron Blue Sepharose chromatography.     

An extracellular 5'-nucleotidase with both monoesterase and diesterase activity from Micrococcus sodonensis.

Bovine γ-globulin was separated into four fractions by chromatography on cellulose phosphate. The chromatographic distribution was similar to that reported for human and dog γ-globulin. More than 80% of a nonspecific phagocytosis stimulating factor (leucokinin) present in the serum was isolated in γ-globulin fraction IV. Bocine red blood cells and polymorphonuclear leukocytes bind γ-globulin without appreciable selectivity for any of the four chromatographic fractions, but they do selectively bind the phagocytosis stimulating factor. Splenectomy caused no observable change in either the chromatographic distribution or phagocytosis stimulating activity of bovine serum γ-globulin. The tetrapeptide tuftsin stimulates phagocytosis by bovine neutrophiles, but on a molar basis the activity of tuftsin was only 10% that of the phagocytosis stimulating factor. If the factor exerts its effect, as has been proposed, by having a phagocytosis stimulating peptide cleaved from it by an enzyme on the leukocyte membrane, that peptide must differ in structure from tuftsin. This conclusion is supported by the inability of trypsin to liberate an active peptide from bovine serum.     

Existence of γ-helix in natural proteins

Serum albumin is known to act as a carrier for a variety of molecules and metal ions. This property of the protein could be due to the presence of different types of secondary structures in its molecules. The most commonly known are the α-helix, β-sheets, β-turns and random coil forms. A rigorous analysis of human serum albumin has been carried out by using four different approaches. Comparative studies have revealed that the segment Asp 107 to Val 122 of this protein assumes a γ-helical structure. Under favourable circumstances, two prolines at the i and (i + 5)th positions can initiate a γ-helix. Further requirements for the formation and stability of γ-helix are discussed.     

The Effects of Sugar Alcohols on Metabolism of Growing Pigs

In two experiments of 3 × 3 Latin square with growing pigs, the effect was investigated of supplementation of 5 % or 10 % (2.5 % vs. 5 % DM) polyol mixture and 2.5 % or 5 % of xylitol on digestibility of diet, N-balance, blood clinical-chemical parameters and insulin level in serum. Apparent digestibility of crude protein was lower for the diet with 10 % polyol mixture compared to the control. Sugar alcohols were not found in faeces. Arabinitol, mannitol and rhamnitol were excreted in the urine 25–67 %. Little sorbitol and xylitol were found in urine on diets with polyol mixture 5–10 %. On xylitol diets the pigs did not excrete xylitol in urine. Plasma glucose rose in pigs fed xylitol. Blood total protein and albumin decreased in pigs fed polyol mixture. ALAT-activities were higher for xylitol diets than for the controls. Serum insulin tended to increase in pigs fed polyol mixture 10 % one hour after feeding, and in xylitol feeding two hours after feeding; these values were higher with increasing xylitol inclusion in diet.     

Changes in serum lipoproteins during metamorphosis of the bullfrog,Rana catesbeiana

Serum lipoproteins of the bullfrog, Rana catesbeiana, were studied during metamorphosis. Adult bullfrog has essentially one lipoprotein, designated β-lipoprotein. This β-lipoprotein migrates during electrophoresis to β-globulin region and it has a low hydrated density such that it exhibits floatation in a solvent of density 1.063. On the other hand, tadpole serum has one more lipoprotein, designated as α-lipoprotein, in addition to the β-lipoprotein. The α-lipoprotein migrates to the α-globulin region in zone electrophoresis and corresponds to the so called high density lipoprotein judging from ultracentrifugal behavior. Serum α-lipoprotein disappears and β-lipoprotein content decreases during metamorphosis.     

The proteins and protein-bound carbohydrates of the serum of the developing pig

1. The concentrations of total nitrogen, hexosamine and protein-bound hexose and the amounts of these constituents precipitated by trichloroacetic acid were determined in the serum of pigs at various ages from 57 days after copulation to 42 days after birth. The concentrations of the same constituents were also determined in the serum of mature pigs. 2. A rise in the concentrations of total nitrogen, hexosamine and protein-bound hexose between 90 days' gestation and term was entirely due to material soluble in trichloroacetic acid. This material disappeared from the serum by 7 days after birth. 3. Electrophoresis of the serum proteins showed that the concentration of alpha(1)-globulin fell steadily during gestation, being lower at term than at 57 days' gestation. No alpha(1)-globulin was detected at 7 days of age. 4. It was concluded that high values for non-protein nitrogen in the serum of newborn piglets, determined after precipitation of the proteins with trichloroacetic acid, is largely due to the presence of one or more mucoproteins and not to alpha(1)-globulin. 5. The protein migrating in the alpha(1)-globulin position is possibly a foetal protein of the fetuin type.     

Effects of dietary incorporation of different antioxidant extracts and free-range rearing on fatty acid composition and lipid oxidation of Iberian pig meat

This investigation was designed to evaluate the effects of feeding either free range or in confinement using concentrated diets with the same ingredients and oil source (5.5% of olive oleins) but with different antioxidant supplementation [control diet with a basal level of α-tocopheryl acetate (control); 200 mg/kg synthetic all-rac-α-tocopheryl acetate (Eall-rac); 200 mg/kg natural RRR-α-tocopheryl-acetate (ERRR-); flavonoid extract-enriched diet (AFlav); and phenolic compound-enriched extract (APhen)] on the fatty acid composition and lipid oxidation of Iberian pig muscle longissimus dorsi. The α-tocopherol concentration was significantly higher in muscles from free-range and ERRR- pigs than in muscles from Eall-rac pigs, and γ-tocopherol was only detected in muscles from free-range pigs. Longissimus dorsi muscles from free-range pigs had a significantly lower content of saturated fatty acids and higher content of polyunsaturated fatty acids than muscles from the other five groups of pigs fed in confinement; however, no significant effect on monounsaturated fatty acids was observed. No effect of dietary antioxidant supplementation (synthetic or natural α-tocopherol, flavonoid extract, or phenol extract) on the fatty acid composition of muscles was observed. A significant influence of dietary treatment on lipid oxidation was observed after 3 (P < 0.01), and 7 and 10 (P < 0.001) days of refrigerated storage, respectively. The lowest thiobarbituric acid-reactive substances (TBARS) values were found in pork chops from the free-range and ERRR- groups, intermediate values from the Eall-rac group, followed by AFlav and APhen, while the highest TBARS values corresponded to muscles from pigs fed the control concentrate. The source of α-tocopherol had a significant effect on lipid oxidation (P < 0.05), whereas the AFlav and APhen groups had similar TBARS values.