1156
Langmuir 1993,9, 1156-1157
Notes A New Role of Serum Albumin and a New Interpretation for the High Sedimentation Rate of Erythrocytes in Analbuminemia Takayoshi Matawnoto,',+ Hiroshi Inoue,' and Masakazu Takahashit
BSA-20mM Phosphate buffer
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Department of Polymer Chemistry, Kyoto University, Kyoto 606, Japan, and Department of Pathology, Sasaki Institute, Tokyo 101, Japan Received July 13, 1992. In Final Form: September 28, 1992
Introduction The biological and physiological function of serum albumin has not been substantiallyclarified. Physiological or clinical disease in analbuminemia (a genetic mutant with albumin deficient blood) is very slight, and also the rates of growth and reproduction of the analbuminemia were essentially no different from the normal cas8.l On the basis of this fact, some doubt has been entertained about the biochemical or physiological role of serum albumin as a carrier by unspecific adsorption of various hormones, nutrients, and drugs. In this paper, we discuss a new role of serum albumin in terms of blood flow which is a completely different approach from the biochemical or physiological viewpoint, using the aqueous colloids of the serum albumin, and the serums of the normal rat and the analbuminemic rat. Experimental Section The bovine serum albumin (BSA, A0281) employed was purchased from Sigma Ltd. This sample is essentially globulinfree and the fatty acid content is less than 0.005%. The weight average molecular weight M, of the dispersing particle was measured by means of a low angle laser light scattering photometer (A = 633 nm, Chromatix KMX6) and the specific refractive index increment dnldc was measured with a differential refractometer (A = 633 nm, Chromatix KMX16). In Figure 1, Kc/Re is plotted against the BSA concentration c in a medium of 20 mM sodium phosphate buffer solution at pH 7.0. Here K is the optical constant and R, is the excess Rayleigh ratio of the system. The value of M, of the dispersing particle can be calculated as a reciprocal value of the intercept of the ordinate using dn/dc = 0.183 mL g-l and estimated at 11.7 X 104. That is, the BSAmolecules disperse in an association state of N (=Mw/ Mo)= 1.77 in a concentration range at least from 0.02 to 0.1%. Here, MOis the molecular weight of a BSA molecule, which is calculated as 66 267 from the amino acid ~equence.~ The serums of the rats employed were extracted from the analbuminemicrat (Nagase Anulbuminemia Rat, NAR, 20 weeks old, male) and the normal rat (SDR, 20 weeks old, male).
Results and Discussion Figure 2 showsthe logarithmicflow curves of shear stress
* To whom correspondence should be addressed.
Kyoto University. Sasaki Institute. ( 1 ) Aoki, K.,Takaai, T.,Terada. H.,Ed. Serum Albumin: Kodansha: Tokyo, 1989. (2) Nagase, S.; Shimamura, K.; Sumiya, S.Science 1979,205, 590. (3) Reed, R. G.;Putnam, F. W.;Peters,T., Jr. Biochem. J. ISSO, 19, t
887.
n 0
Mw=1.17x105
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6 8 10 12 '10% I g ml Figure 1. KclRe plotted against the BSA concentration c. 4
BSA-20mM buffer b 0.001%
0
0.1%
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2
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log ( y / s-' ) Figure 2. Logarithmic flow curves of isotropic colloids of native bovine serum albumin in phosphate buffer solution over a wide concentration range from0 to 5 $7, (opensymbols), and logarithmic flow curves of the analbuminemic rat (NAR) and normal rat (SDR) (closed symbols). u vs shear rate +,which have been measured with a coneplate Weissenberg rheogoniometer, for the native BSA colloids in a 20 mM sodium phosphate buffer solution (pH 7.0) over a wide concentration range from 0 to 5 wt 7% (open symbols). In this figure, the flow curves of the serums of NAR and SDR are also shown by the closed symbols. The buffer solution shows Newtonian flow, and the aqueous colloids of the higher concentrations beyond 0.01 7% apparently show a plateau region which approximately correspondsto the yield stress. The BSA colloids also show relatively large values of dynamic modulus in the same concentration range. The yield stress and modulus are a measure of the strength of a solidlike structure in the c~lloid.~ There are two notable points in these flow curves of the BSA aqueous colloids. One is that the system shows relatively clear yield stress even at very low concentrations such as 0.01 7% ,and the second is that the value of yield stress remains almost constant over an extremely wide range of the BSA concentration. Considering that the association number of the diapersing particle is 1.77 in the concentration range from 0.02 to 0.1 % ,the yield stress in the BSA colloid seemsto originate (4) Mataumoto, T.; Yao, S.; Onogi,
S.J. Rheol. 1986, 29, 177.
0743-7463/93/2409-1156$04.~/0 0 1993 American Chemical Society
Langmuir, Vol. 9, No.4, 1993 1157
Notes
PS TF
BSA O Q Y
G’ b 0
SDR
e
log( c / gqg-’) Figure 3. Shear stress u, (open circles) at
= 0.1 s-l in the plateau region and dynamic modulus C’ (closed circles) at w = 0.1 8-1 plotted against the BSA concentration c. The yield stress or the modulus is also plotted for the usual colloidsof polystyrene beads (PS)and titanate fibers (TF).
from a certain ordered arrangement (the albumin lattice) of the BSA molecules. The formation of the albumin lattice is assumed to be extremely accelerated by shear flow.s It turns out that the serum of SDR apparently belongs to the group in which the albuminlattice is formed, but the serum of NAR is in the group which cannot form the albumin lattice. Figure 3 shows the shear stress (ay,open circles) at i. = 0.1 s-l in the plateau region and dynamic modulus G’ at o = 0.1 s-l plotted against the BSA concentration c. For the BSA colloid, ay increases in proportion to c1 at very low concentrations and remains almost constant (or increases very slightly) at higher concentrations. The dynamic modulus is so small that it cannot be detected at concentrations lower than 0.01% for the BSA colloid. In Figure 3,the concentration dependence of the yield streas or modulus is also shown for the usual colloids of sphericalpolystyrene beads (PS, diameter = ca. 100nm)6J and rodlike titanate fibers (TF).* The effective charge (5) Mataumoto, T.; Chiba, J. J. SOC.Rheol., Jpn. 1991,19, 147. (6) Lindsay,H.; Chaikin, P. H. J . Chem. Phys. 1982, 76,3774.
number of polystyrene beads is ca. 300,and the rigidity of the system diesappearsat the electrolyte concentration of 10-200 p M in the medium, depending on the particle content. The behavior of the BSA colloid is very different from that of the usual colloids. In a comparison of the BSA colloid with the usual colloids, the BSA colloid flows relatively easily at high concentrations but does not flow easily at low concentrations. This means that the BSA colloid retains an almost constant viscosity at a low level over a wide concentration range and that the system can support the stress of 0.142 Pa. Considering that the difference in specificgravity between the erythrocytesand the serum is ca. 0.07, this stress is large enough to prevent the sedimentation of erythrocytes in the blood.9 The BSA concentration in serum is estimated at ca. 4% in a normal case, but in analbuminemia? the concentration decreases to 1 / 2 ~ - 1 / 4 ~of the normal case as shown by arrows on the abscissa in Figure 3. The stress levels for SDR and NAR are also shown on the ordinate. The BSA concentration in the serum of analbuminemia is in the region where the yield stress decreases sharply. That is, the sedimentation rate of erythrocytes,which is measured in a stationary state, increases noticeably in analbuminemia. However, consideringthat the shear stress to which blood is submitted in the arterioles is estimated to be in the range of ca. 1.3-3 Pa,1othe yield stress of 0.1-0.2 Pa can be easily overcome in the flowing state of the blood. That is, the deficit of serum albumin does not exert a serious influence on the blood flow in the vessels, though the existence of the albumin is more favorable for transportation of the erythrocytes. In previous papers,6J1using rheological and SAXS data, the authors reported that ovalbumin colloids also show behavior similar to that of BSA colloids and that the behavior is retained at various ionic strengths and pH values. It may be a common characteristic for various albuminsthat they form the albumin lattice in the systems to prevent the sedimentation of certain objecta, but the albumins still retain the systems under conditions of easy flow. (7) Okubo, T. J . Chem. SOC.,Faraday Trans. 1 1989,85,455. (8)Mikami, Y.; Onogi, S. J. SOC.Rheol., Jpn. 1977,5, 110. (9) Phillips, R. A.; et al. J. Biol. Chem. 1950,83, 305. (10) Koyama, T. Biorheology 1985,22, 379. (11) Mataumoto, T.; Inoue, H. J . Chem. SOC.,Faraday Trans. 1991, 87, 3385; Colloid Polym. Sci. 1992, 270, 687.
