Equation for osmotic pressure of serum protein (fractions).

The colloid or protein osmotic pressure (Pi) is a function of protein molarity (linear) and of Donnan and other effects. Albumin is the major osmotic protein, but also globulins influence Pi. Equations based on concentrations of albumin and nonalbumin (globulin concentration + fibrinogen concentration) protein approximate Pi better than albumin alone. Globulins have a wide range of molecular weights, and a 1956 diagram indicated that Pi of globulin fractions decreased in the order alpha1-, alpha2-, beta-, and gamma-globulin. The molecular weight of the serum protein fractions had been extrapolated, so van't Hoff's law and nonlinear regression analysis of the curves permitted expression of the diagram as an equation: product Pi(s,Ott,2 degrees C,cmH2O)=x(alb)(0.338C(tot)+0.00339C(tot)(2))+x(alpha1)(0.518C(tot)+0.0107C(tot)(2))+x(alpha2)(0.203C(tot)+0.00155C(tot)(2))+x(beta)(0.187C(tot)+0.000577C(tot)(2))+x(gamma)(0.161C(tot)+0.000223C(tot)(2)), where Pi(s,Ott,2 degrees C,cmH2O) is Pi of serum at 2 degrees C (in cmH2O) computed from the 1956 diagram, C(tot) is the concentration (g/l) of total protein in serum, and x(alb), x(alpha1), x(alpha2), x(beta), and x(gamma) are the fractions of albumin, alpha1-, alpha2-, beta-, and gamma-globulin, respectively. At one and the same concentration of fractions, Pi("Ott") decreases in the order alpha1-globulin, albumin, alpha2-globulin, beta-globulin, and gamma-globulin.     

Continuous measurement of protein osmotic pressure in blood and lymph of sheep

We have continuously measured protein osmotic pressure of blood and lymph in sheep to compare two kinds of needle osmometers (rigid and flexible) with a membrane osmometer (Wescor). We also compared the averaged values of the continuous measurement with osmotic pressure calculated from total protein and albumin fraction, using the Yamada equation. The rigid-needle and membrane osmometers showed excellent correlation (y = 1.00x + 0.06; r greater than 0.99). The flexible-needle osmometer tended to overestimate osmotic pressure (avg 16%). We used the rigid-needle osmometer for continuous measurements of protein osmotic pressure of blood and lymph in anesthetized or unanesthetized sheep to observe changes in protein osmotic pressure of blood and lymph through the three different interventions. The relationship between the theoretical values (x) and the continuous measurements (y) of osmotic pressure was good (y = 0.99x + 0.16, r = 0.97), but after various interventions, the continuously measured protein osmotic pressure tended to exceed the calculated measurements. The continuous measurement should be monitored with spot samples measured in a stationary osmometer or by calculation of osmotic pressure from total protein concentration and albumin fraction.     

Effects of hypoproteinemia on lung microvascular protein sieving and lung lymph flow

Experiments were conducted on five chronically instrumented unanesthetized sheep to determine the effects of sustained hypoproteinemia on lung fluid balance. Plasma total protein concentration was decreased from a control value of 6.17 +/- 0.019 to 3.97 +/- 0.17 g/dl (mean +/- SE) by acute plasmapheresis and maintained at this level by chronic thoracic lymph duct drainage. We measured pulmonary arterial pressure, left atrial pressure, aortic pressure, central venous pressure, cardiac output, oncotic pressures of both plasma and lung lymph, lung lymph flow rate, and lung lymph-to-plasma ratio of total proteins and six protein fractions for both control base-line conditions and hypoproteinemia base-line conditions. Moreover, we estimated the average osmotic reflection coefficient for total proteins and the solvent drag reflection coefficients for the six protein fractions during hypoproteinemia. Hypoproteinemia caused significant decreases in lung lymph total protein concentration, lung lymph-to-plasma total protein concentration ratio, and oncotic pressures of plasma and lung lymph. There were no significant alterations in the vascular pressures, lung lymph flow rate, cardiac output, or oncotic pressure gradient. The osmotic reflection coefficient for total proteins was found to be 0.900 +/- 0.004 for hypoproteinemia conditions, which is equal to that found in a previous investigation for sheep with a normal plasma protein concentration. Our results suggest that hypoproteinemia does not alter the lung filtration coefficient nor the reflection coefficients for plasma proteins. Possible explanations for the reported increase in the lung filtration coefficient during hypoproteinemia by other investigators are also made.     

Acid-base effects of altering plasma protein concentration in human blood in vitro

We altered the concentration of plasma proteins in human blood in vitro by adding solutions with [Na+], [K+], and [Cl-] resembling those in normal blood plasma, either protein-free or with a high concentration of human albumin. After equilibrating the samples with a gas containing 5% CO2-12% O2-83% N2 at 37 degrees C, we measured pH, PCO2, and PO2; in separated plasma, we determined the concentrations of total plasma proteins and albumin and of the completely dissociated electrolytes (strong cations Na+, K+, Mg2+ and anions Cl-, citrate3-). With PCO2 nearly constant (mean = 35.5 Torr; coefficient of variation = 0.02), lowering plasma protein concentration produced a metabolic alkalosis, whereas increasing plasma albumin concentration gave rise to a metabolic acidosis. These acid-base disturbances occurred independently of a minor variation in the balance between the sums of strong cations and anions. We quantified the dependence of several acid-base variables in plasma on albumin (or total protein) concentration. Normal plasma proteins are weak nonvolatile acids. Although their concentration is not regulated as part of acid-base homeostasis, hypoproteinemia and hyperalbuminemia per se produce alkalosis and acidosis, respectively.     

Osmotic reflection coefficient for total plasma protein in lung microvessels

The osmotic reflection coefficient (sigma) for total plasma proteins was estimated in 11 isolated blood-perfused canine lungs. Sigma's were determined by first measuring the capillary filtration coefficient (Kf,C in ml X min-1 X 100g-1 X cmH2O-1) using increased hydrostatic pressures and time 0 extrapolation of the slope of the weight gain curve. Kf,C averaged 0.19 +/- 0.05 (mean +/- SD) for 14 separate determinations in the 11 lungs. Following a Kf,C determination, the isogravimetric capillary pressure (Pc,i) was determined and averaged 9.9 +/- 0.5 cmH2O for all controls reported in this study. Then the blood colloids in the perfusate were either diluted or concentrated. The lung either gained or lost weight, respectively, and an initial slope of the weight gain curve (delta W/delta t)0 was estimated. The change in plasma protein colloid osmotic pressure (delta IIP) was measured using a membrane osmometer. The measured delta IIP was related to the effective colloid osmotic pressure (delta IIM) by delta IIM = (delta W/delta t)0/Kf,C = sigma delta IIP. Using this relationship, sigma averaged 0.65 +/- 0.06, and the least-squares linear regression equation relating Pc,i and the measured IIP was Pc,i = -3.1 + 0.67 IIP. The mean estimate of sigma (0.65) for total plasma proteins is similar to that reported for dog lung using lymphatic protein flux analyses, although lower than estimates made in skeletal muscle using the present methods (approximately 0.95).     

Plasma volume, albumin and globulin concentrations and their intravascular masses. A comparative study in endurance athletes and sedentary subjects.

Plasma volume, hematocrit, intravascular protein concentration, colloid osmotic pressure and the intravascular mass of proteins were measured in 49 sedentary subjects and 40 endurance athletes (long-, middle distance runners, cyclists). The plasma volume in sedentary subjects was 42.7(35.8-51.7) ml/kg body weight (BW) as compared to 54.6(46.7-65.9) ml/kg BW in athletes. The protein concentrations were 71.0 (66.5-77.1) g/l in sedentary subjects and 69.0 (64.8-75.2) g/l in athletes. The respective numbers for the hematocrit were 44.6 (40.1-49.25)% and 42.8 (38.2-49.6)%, for the colloid osmotic pressure 38.0 (36.0-40.5) cm H2O (n=35) and 30.0 (25.0-34.4) cm H2O (n=31), for the intravascular mass of proteins 3.09 (2.45-4.01) g/kg BW and 3.75 (3.31-4.67) g/kg BW. All differences were statistically significant at least on the 5% level. The physiological consequences for athletes of having a lower hematocrit and lower protein concentration but a higher intravascular mass of proteins (+22%) for their waterbalance as well as for their dietary protein intake are discussed. Endurance exercise stimulates mainly the synthesis of albumin and globulins produced by the liver resulting in an expansion of the PV. The protein synthesis of the RES does not seem to respond to exercise stimulus.     

Role of apolipoprotein D in the transport of bilirubin in plasma

Apolipoprotein D (apo D) is a 30-kDa glycoprotein of unknown function that is associated with high-density lipoproteins (HDL). Because unconjugated bilirubin has been shown to bind apo D with a 0. 8:1 stoichiometry, we examined the contribution of this protein to transport of bilirubin in human plasma. Density gradient centrifugation analysis using physiological concentrations of [(14)C]bilirubin reveals that 9% of unconjugated bilirubin is associated with HDL, with the remaining pigment bound primarily to serum proteins (i.e., albumin). The percentage of total plasma bilirubin bound to HDL was found to increase proportionally with bilirubin concentration. Affinity of human apo D for bilirubin was determined by steady-state fluorescence quenching, with Scatchard analysis demonstrating a single binding site for unconjugated bilirubin with an affinity constant (K(a)) of approximately 3 x 10(7) M(-1). Incorporation of apo D into phosphatidylcholine vesicles had no effect on K(a), suggesting that a lipid environment does not alter the affinity of the protein for bilirubin. Using stopped-flow techniques, the first-order rate constant for bilirubin dissociation from apo D was measured at 5.4 s(-1) (half-time = 129 ms). Our findings indicate that HDL is the principal nonalbumin carrier of bilirubin in human plasma and further support the proposition that the affinity of HDL for bilirubin is primarily the result of binding to apo D.     

Experimental determination of net protein charge, [A]tot, and Ka of nonvolatile buffers in bird plasma.

The quantitative mechanistic acid-base approach to clinical assessment of acid-base status requires species-specific values for [A]tot (the total concentration of nonvolatile buffers in plasma) and Ka (the effective dissociation constant for weak acids in plasma). The aim of this study was to determine [A]tot and Ka values for plasma in domestic pigeons. Plasma from 12 healthy commercial domestic pigeons was tonometered with 20% CO2 at 37 degrees C. Plasma pH, Pco2, and plasma concentrations of strong cations (Na, K, Ca), strong anions (Cl, L-lactate), and nonvolatile buffer ions (total protein, albumin, phosphate) were measured over a pH range of 6.8-7.7. Strong ion difference (SID) (SID5=Na+K+Ca-Cl-lactate) was used to calculate [A]tot and Ka from the measured pH and Pco2 and SID5. Mean (+/-SD) values for bird plasma were as follows: [A]tot=7.76+/-2.15 mmol/l (equivalent to 0.32 mmol/g of total protein, 0.51 mmol/g of albumin, 0.23 mmol/g of total solids); Ka=2.15+/-1.15x10(-7); and pKa=6.67. The net protein charge at normal pH (7.43) was estimated to be 6 meq/l; this value indicates that pigeon plasma has a much lower anion gap value than mammals after adjusting for high mean L-lactate concentrations induced by restraint during blood sampling. This finding indicates that plasma proteins in pigeons have a much lower net anion charge than mammalian plasma protein. An incidental finding was that total protein concentration measured by a multianalyzer system was consistently lower than the value for total solids measured by refractometer.     

Paraoxon hydrolysis in human serum mediated by a genetically variable arylesterase and albumin.

Gel filtration chromatography resolves human serum paraoxonase into two fractions: (1) a high molecular weight fraction that is completely inhibited by EDTA and coelutes with arylesterase (E.C.3.1.1.2); and (2) a second fraction that is closely associated with albumin, is only partially inhibited by EDTA, and has relatively little arylesterase activity under the assay conditions used. The activity of the high molecular weight fraction is stimulated by NaCl, whereas the albumin associated activity is partially inhibited by NaCl and is not present in serum derived from an analbuminemic individual. Our data suggest that albumin itself, rather than a protein bound to or cofractionating with albumin, mediates paraoxonase activity. The variation in levels of the activity of the nonalbumin, high molecular weight enzyme is responsible for the observed polymorphism of paraoxonase activity in human serum or plasma. An optimal assay of polymorphic paraoxonase activity should be based on activity measurements of the nonalbumin fraction. It is considered likely that only the nonalbumin fraction is responsible for in vivo hydrolysis of paraoxon.     

Energetics of a hexagonal-lamellar-hexagonal-phase transition sequence in dioleoylphosphatidylethanolamine membranes.

The phase diagram of DOPE/water dispersions was investigated by NMR and X-ray diffraction in the water concentration range from 2 to 20 water molecules per lipid and in the temperature range from -5 to +50 degrees C. At temperatures above 22 degrees C, the dispersions form an inverse (HII) phase at all water concentrations. Below 25 degrees C, an HII phase occurs at high water concentrations, an L alpha phase is formed at intermediate water concentrations, and finally the system switches back to an HII phase at low water concentrations. The enthalpy of the L alpha-HII-phase transition is +0.3 kcal/mol as measured by differential scanning calorimetry. Using 31P and 2H NMR and X-ray diffraction, we measured the trapped water volumes in HII and L alpha phases as a function of osmotic pressure. The change of the HII-phase free energy as a function of hydration was calculated by integrating the osmotic pressure vs trapped water volume curve. The phase diagram calculated on the basis of the known enthalpy of transition and the osmotic pressure vs water volume curves is in good agreement with the measured one. The HII-L alpha-HII double-phase transition at temperatures below 22 degrees C can be shown to be a consequence of (i) the greater degree of hydration of the HII phase in excess water and (ii) the relative sensitivities with which the lamellar and hexagonal phases dehydrate with increasing osmotic pressure. These results demonstrate the usefulness of osmotic stress measurements to understand lipid-phase diagrams.     

Protein osmotic pressure and the state of water in frog myoplasm

We measured the osmotic pressure of diffusible myoplasmic proteins in frog (Rana temporaria) skeletal muscle fibers by using single Sephadex beads as osmometers and dialysis membranes as protein filters. The state of the myoplasmic water was probed by determining the osmotic coefficient of parvalbumin, a small, abundant diffusible protein distributed throughout the fluid myoplasm. Tiny sections of membrane (3.5- and 12-14-kDa cutoffs) were juxtaposed between the Sephadex beads and skinned semitendinosus muscle fibers under oil. After equilibration, the beads were removed and calibrated by comparing the diameter of each bead to its diameter measured in solutions containing 3-12% Dextran T500 (a long-chain polymer). The method was validated using 4% agarose cylinders loaded with bovine serum albumin (BSA) or parvalbumin. The measured osmotic pressures for 1.5 and 3.0 mM BSA were similar to those calculated by others. The mean osmotic pressure produced by the myoplasmic proteins was 9.7 mOsm (4 degrees C). The osmotic pressure attributable to parvalbumin was estimated to be 3.4 mOsm. The osmotic coefficient of the parvalbumin in fibers is approximately 3.7 mOsm mM(-1), i.e., roughly the same as obtained from parvalbumin-loaded agarose cylinders under comparable conditions, suggesting that the fluid interior of muscle resembles a simple salt solution as in a 4% agarose gel.     

Decreased Coomassie brilliant blue colour yield with glycated proteins.

Collagen, myosin and albumin were incubated for 7 days at 20 degrees C with fructose, ribose or glyceraldehyde. For thus-formed glycated proteins, quantities were determined by the Conway microdiffusion technique and by the colorimetric method based on Coomassie brilliant blue G-250 colour yield. It was found that when albumin was glycated with increasing amounts of glyceraldehyde, the colour yield was decreased by 7-33%. In collagen, myosin and albumin incubated with 0.5 mol/l fructose, 0.5 mol/l ribose or 0.1 mol/l glyceraldehyde, protein concentration was not changed, as proved by the Conway microdiffusion technique; the Coomassie brilliant blue G-250 colour yield was up to 50% lower, depending on the protein used, and was decreased much less when proteins were incubated with less sugar.     

Virial coefficients of protein solutions

An osmometer capable of measuring protein osmotic pressures up to 100 cms. of mercury pressure has been described. The principle of the osmometer is to set a given pressure and to permit the protein concentration to equilibrate with the pressure. The higher virial osmotic coefficients of egg albumin in various electrolytes and in 1 m urea as well as a function of NaCl concentration are reported. The virial coefficients of bovine serum albumin and of bovine methemoglobin in 1 m NaCl are also given. It appears that the primary cause for the departure of the osmotic pressure from ideality is due to the covolumes of the proteins.     

Oestriol binding to plasma proteins

Simple diffusion experiments indicated that oestriol was retained by human pregnancy plasma more effectively than by albumin solutions of a corresponding concentration. Oestriol bound (Ka = 6 X 10(6) l/mol at 4 degrees C) to a glycoprotein which had been isolated from plasma by adsorption to Concanavalin A. The free energy of binding at 37 degrees C was -38 kJ/mol. Competition experiments indicated that the oestriol binding glycoprotein had properties expected of sex hormone binding globulin. The distribution of oestriol among the protein fractions of human pregnancy plasma--glycoprotein bound 7.8%, albumin bound 78.6%, unbound 13.6%--suggests that this glycoprotein plays little part in the transport of oestriol.     

A model to explain the osmotic pressure behavior of hemoglobin and serum albumin

Previously published osmotic pressure data on hemoglobin and bovine serum albumin were used to determine the osmotically unresponsive solvent volume per unit dry mass of protein. A model is presented that accounts for the osmotic pressure of globular proteins based on a surface-associated osmotically unresponsive solvent volume. The model also accounts for changes in the osmotically unresponsive solvent volume owing to changes in pH, cosolute salt concentration, protein conformation, and protein aggregation.     

Early fluid and protein shifts in men during water immersion

High precision blood and plasma densitometry was used to measure transvascular fluid shifts during water immersion to the neck. Six men (28-49 years) undertook 30 min of standing immersion in water at 35.0 +/- 0.2 degrees C; immersion was preceded by 30 min control standing in air at 28 +/- 1 degrees C. Blood was sampled from an antecubital catheter for determination of blood density (BD), plasma density (PD), haematocrit (Ht), total plasma protein concentration (PPC), and plasma albumin concentration (PAC). Compared to control, significant decreases (p less than 0.01) in all these measures were observed after 20 min immersion. At 30 min, plasma volume had increased by 11.0 +/- 2.8%; the average density of the fluid shifted from extravascular fluid into the vascular compartment was 1006.3 g.l-1; albumin moved with the fluid and its albumin concentration was about one-third of the plasma protein concentration during early immersion. These calculations are based on the assumption that the F-cell ratio remained unchanged. No changes in erythrocyte water content during immersion were found. Thus, immersion-induced haemodilution is probably accompanied by protein (mainly albumin) augmentation which accompanies the intravascular fluid shift.     

Method of high-precision microsample blood and plasma mass densitometry

The reliability of the mechanical oscillator technique (MOT) for blood and plasma mass density measurements on small samples is quantified in this paper. Sources of measurement errors that can reduce both the accuracy and precision of density determinations include storage of plasma samples, inhomogeneity of blood samples, and density reading before adequate temperature equilibration. Measurements on fractions from identical samples and repeated samplings from test subjects under steady-state conditions revealed a 10(-2) g/l reproducibility of density readings. The mean plasma density (PD) readings did not change significantly after up to 1-wk storage at +4 degrees C or up to 2 mo storage at -20 degrees C. The variability of the PD findings increased with storage time and were generally higher with storage at -20 degrees C, compared with +4 degrees C. Densitometers of different sizes were used to evaluate rheological influences on blood density (BD) readings. Linear correlations between PD and plasma protein concentration, between BD and blood hemoglobin concentration, and between erythrocyte density and mean corpuscular hemoglobin concentration were significant (P less than 0.001). Rapid density measurements with up to 10(-2) g/l reliability on small (less than 0.1 ml) volumes of biological fluids and continuous blood densitometry can be performed with use of the MOT.     

Effects of haemorrhage on colloid osmotic pressure in the pigeon

1. Colloid osmotic pressure, plasma osmolality and plasma protein concentration, percentage composition and A/G ratio were measured before and after haemorrhage in the pigeon. 2. Colloid osmotic pressure, total protein, albumin and beta-globulin concentrations decreased significantly immediately post-haemorrhage, but were significantly elevated after one week. 3. Osmolality and A/G ratio values were significantly increased post-haemorrhage. 4. The results are discussed in relation to fluid exchange across capillaries.     

Theoretical and Practical Aspects of Albumin Esterase Activity

Russian Journal of Bioorganic Chemistry - Unlike many other plasma proteins, albumin is barely glycosylated and plays an important role in maintaining the colloidal osmotic blood pressure; it can...     

Effect of hypoproteinemia on lung fluid balance in awake newborn lambs

To study the influence of plasma protein concentration on fluid balance in the newborn lung, we measured pulmonary arterial and left atrial pressures, lung lymph flow, and concentrations of protein in lymph and plasma of eight lambs, 2-3 wk old, before and after we reduced their plasma protein concentration from 5.8 +/- 0.3 to 3.6 +/- 0.6 g/dl. Each lamb underwent two studies, interrupted by a 3-day period in which we drained protein-rich systemic lymph through a thoracic duct fistula and replaced fluid losses with feedings of a protein-free solution of electrolytes and glucose. Each study consisted of a 2-h control period followed by 4 h of increased lung microvascular pressure produced by inflation of a balloon in the left atrium. Body weight and vascular pressures did not differ significantly during the two studies, but lung lymph flow increased from 2.6 +/- 0.1 ml/h during normoproteinemia to 4.1 +/- 0.1 ml/h during hypoproteinemia. During development of hypoproteinemia, the average difference in protein osmotic pressure between plasma and lymph decreased by 1.6 +/- 2 Torr at normal left atrial pressure and by 4.9 +/- 2.2 Torr at elevated left atrial pressure. When applied to the Starling equation governing microvascular fluid balance, these changes in liquid driving pressure were sufficient to account for the observed increases in lung fluid filtration; reduction of plasma protein concentration did not cause a statistically significant change in calculated filtration coefficient. Protein loss did not influence net protein clearance from the lungs nor did it accentuate the increase in lymph flow associated with left atrial pressure elevation.(ABSTRACT TRUNCATED AT 250 WORDS)