Study of interaction between drug enantiomers and serum albumin by capillary electrophoresis

The interaction between drugs and human serum albumin (HSA) was investigated by capillary electrophoresis (CE). It involves stereoselectivity, drug displacement and synergism effects. Under protein-drug binding equilibrium, the unbound concentrations of drug enantiomers were measured by frontal analysis (FA). The stereoselectivity of verapamil (VER) binding to HSA was proved by the different free fractions of two enantiomers. In physiological pH (7.4, ionic strength 0.17 phosphate buffer) when 300 mu M (+/-) VER were equilibrated with 500 mu M HSA, the concentration of unbound S-VER was about 1.7 times its antipode. The binding constants of two enantiomers, KR-VER and KS-VER, were 2670 and 850 M-1, respectively. However, no obvious stereoselective binding of propranolol (PRO) to HSA was observed. Trimethyl-beta-cyclodextrin (45 mM) was used as a chiral selector in pH 2.5 phosphate buffer. Several drug systems were studied by the method. When ibuprofen (IBU) was added into VER-HSA solution. R-VER was partially displaced while S-VER was not displaced at all. A binding synergism effect between bupivacaine (BUP) and verapamil was observed and further study suggested that verapamil and bupivacaine occupy different binding site of HSA (site II and site III, respectively).

Competitive adsorption between bovine serum albumin and collagen observed by atomic force microscope

Atomic force microscopy (AFM) was used to study the competitive adsorption between bovine serum albumin (BSA) and type I collagen on hydrophilic and hydrophobic silicon wafers. BSA showed a grain shape and the type I collagen displayed fibril-like molecules with relatively homogeneous height and width, characterized with clear twisting (helical formation). These AFM images illustrated that quite a lot of type I collagen appeared in the adsorption layer on hydrophilic surface in a competitive adsorption state, but the adsorption of BSA was more preponderant than that of type I collagen on hydrophobic silicon wafer surface. The experiments showed that the influence of BSA on type I collagen adsorption on hydrophilic surface was less than that on hydrophobic surface.

Adsorption of human serum albumin onto gold: a combined electrochemical and ellipsometric study

Human serum albumin adsorption onto gold surfaces was investigated by electrochemical and ellipsometric methods. Albumin adsorption onto gold was confirmed by the change of the open circuit potential of gold and by the ellipsometric parameter variation during albumin immobilization. In both experiments the parameters reached stable values within 10-15 min. The albumin adsorption layer thickness measured with the ellipsometer was about 1.5 nm. The adsorption of albumin Under applied potential was also investigated and it was found that both positive and negative applied potential promote albumin adsorption. Changes in the optical parameters of bare gold and albumin adsorbed onto gold surface under applied potential were investigated with in Situ ellipsometry. The similarity and reversibility of the optical changes showed that adsorbed albumin was stable on the gold surface Under the applied potential range (-200-600 mV). The cyclic voltammograms of K3Fe(CN)(6) on the modified gold surface showed that albumin Could partly block the oxidation and reduction reaction. (C) 2004 Elsevier Inc. All rights reserved.

Competitive adsorption of collagen and bovine serum albumin - effect of the surface wettability

The competitive adsorption of collagen and bovine serum albumin (BSA) on surfaces with varied wettability was investigated with imaging ellipsometry, and ellipsometry. Silane modified silicon surfaces were used as substrates. The results showed that surface wettability had an important effect on protein competitive adsorption. With the decrease of surface wettability, the adsorption of collagen from the mixture solution of collagen and BSA decreased, while the adsorption of BSA increased. (C) 2003 Elsevier B.V. All rights reserved.

Frog albumin is expressed in skin and characterized as a novel potent trypsin inhibitor

A novel potent trypsin inhibitor was purified and characterized from frog Bombina maxima skin. A full-length cDNA encoding the protein was obtained from a cDNA library constructed from the skin. Sequence analysis established that the protein actually comprises three conserved albumin domains. B. maxima serum albumin was subsequently purified, and its coding cDNA was further obtained by PCR-based cloning from the frog liver. Only two amino acid variations were found in the albumin sequences from the skin and the serum. However, the skin protein is distinct from the serum protein by binding of a haem b (0.95 mol/mol protein). Different from bovine serum albumin, B. maxima albumin potently inhibited trypsin. It bound tightly with trypsin in a 1: 1 molar ratio. The equilibrium dissociation constants (K-D) obtained for the skin and the serum proteins were 1.92 x 10(-9) M and 1.55 x 10(-9) M, respectively. B. maxima albumin formed a noncovalent complex with trypsin through an exposed loop formed by a disulfide bond (Cys(53)-Cys(62)), which comprises the scissile bond Arg(58)(P-1)-His(59)(P-1'). No inhibitory effects on thrombin, chymotrypsin, elastase, and subtilisin were observed under the assay conditions. Immunohistochemical study showed that B. maxima albumin is widely distributed around the membranes of epithelial layer cells and within the stratum spongiosum of dermis in the skin, suggesting that it plays important roles in skin physiological functions, such as water economy, metabolite exchange, and osmoregulation.

Apoptotic activity of frog Bombina maxima skin albumin

Albumin, the most abundant protein components of blood plasma, is synthesized and secreted by liver cells in vertebrates. Recently, it was demonstrated that frog Bombina maxima albumin is also expressed in skin. Both B. maxima albumins from skin and serum

Phosphorus fractions and alkaline phosphatase activity in sediments of a large eutrophic Chinese lake (Lake Taihu)

Spatial, vertical, and seasonal variations in phosphorus fractions and in alkaline phosphatase activity (APA) were investigated in sediments in a large-shallow eutrophic Chinese lake (Lake Taihu) in 2003-2004. The phosphorus content was highest in the most seriously polluted lake area. Iron-bound phosphorus (Fe(OOH)-P) dominated (47% on average) among the phosphorus fractions determined according to Golterman (Hydrobiologia 335:87-95, 1996). Notably, organically-bound P comprised a further significant additional portion (acid-soluble + hot NaOH-extractable organic P = 25%), which was highest at the most polluted sites. The Fe(OOH)-P content was the lowest in spring (April, 2004), suggesting that degradation of organic matter led to the release of iron-bound phosphates. Sediment APA showed a significant positive relationship with both organically-bound P and Fe(OOH)-P. Consequently, organically-bound P is an important portion of the sediment phosphorus in Lake Taihu. It is mainly derived from freshly-settled autochthonous particles and from external discharges. Organically-bound P induces APA and may lead to the release of bioavailable phosphates from the organic sediments, thereby accelerating lake eutrophication.

X-Ray diffraction determination of the fractions of hexagonal and twinned phases in cubic GaN layers grown on (001)GaAs substrate

Being an established qualitative method for investigating presence of additional phases in single crystal materials, X-ray diffraction has been used widely to characterize their structural qualities and to improve the preparation techniques. Here quantitative X-ray diffraction analysis is described which takes into account diffraction geometry and multiplicity factors. Using double-crystal X-ray four-circle diffractometer, pole figures of cubic (002), {111} and hexagonal {10 (1) over bar0} and reciprocal space mapping were measured to investigate the structural characters of mixed phases and to obtain their diffraction geometry and multiplicity factors. The fractions of cubic twins and hexagonal inclusions were calculated by the integrated intensities of rocking curves of cubic (002), cubic twin {111}, hexagonal {10 (1) over bar0} and hexagonal {10 (1) over bar1}. Without multiplicity factors, the calculated results are portions of mixed phases in only one {111} plane of cubic GaN. Diffraction geometry factor can eliminate the effects of omega and X angles on the irradiated surface areas for different scattered planes. (C) 2001 Elsevier Science B.V. All rights reserved.

Apoptotic activity of frog Bombina maxima skin albumin

Albumin, the most abundant protein components of blood plasma, is synthesized and secreted by liver cells in vertebrates. Recently, it was demonstrated that frog Bombina maxima albumin is also expressed in skin. Both B. maxima albumins from skin and serum (BmA-skin and BmAserum) have similar biochemical characteristics except that the former contains haem b. Present studies showed that BmA-skin exhibited cytotoxic activity on H9 and C8166 cells. Pretreated with hemin to induce erythroid differentiation, K562 cells lost their resistance to cytotoxicity of BmAskin. After treating cells with BmA-skin for 48 h, 50 percentage cytotoxic concentrations (CC50) of BmA-skin on H9, C8166 and hemin-treated K562 cells were 1.31±0.09, 1.59±0.08 and 2.28±0.06 μM, respectively. The cell death induced by BmA-skin was mediated by apoptosis of the tested cell lines, as demonstrated by nuclear morphological changes, DNA fragmentation and DNA hypodiploidy of apoptosis cells. At BmA-skin concentration of 2 μM, 27.3%, 19.7% and 17.8% of H9, C8166 and hemin-treated K562 cells were found to be apoptotic. In contrast, BmA-serum possessed no cytotoxic and apoptosis-inducing activity on all the cell lines tested, even with concentration used up to 15 μM. These results indicated that bound haem b in BmA-skin contributed significantly to its cytotoxic and apoptosis-inducing activity on the cell lines assayed.

Investigation on Fe, Mn, Zn, Cu, Pb and Cd fractions in the natural surface coatings sampled and surficial sediments collected in the Songhua River, China

Natural surface coatings sampled (NSCSs) from the surface of shingles and surficial sediments (SSs) in the Songhua River, China were employed to investigate the similarities and difference in fractions of heavy metals (Fe, Mn, Zn, Cu, Pb, and Cd) between NSCSs and SSs using the modified sequential extraction procedure (MSEP). The results show that the differences between NSCSs and SSs in Fe fractions were insignificant and Fe was dominantly present as residual phase (76.22% for NSCSs and 80.88% for SSs) and Fe-oxides phase (20.33% for NSCSs and 16.15% for SSs). Significant variation of Mn distribution patterns between NSCSs and SSs was observed with Mn in NSCSs mainly present in Mn-oxides phase (48.27%) and that in SSs present as residual phase (45.44%). Zn, Cu, Pb and Cd were found dominantly in residual fractions (>48%), and next in solid oxides/hydroxides for Zn, Pb and Cd and in easily oxidizable solids/compounds form for Cu, respectively. The heavy metal distribution pattern implied that Fe/Mn oxides both in NSCSs and SSs were more important sinks for binding and adsorption of Zn, Pb and Cd than organic matter (OM), and inversely, higher affinity of Cu to OM than Fe/Mn oxides in NSCSs and SSs was obtained. Meanwhile, it was found that the distributions of heavy metals in NSCSs and SSs were similar to each other and the pseudo-total concentrations of Zn, Cu, Pb and Cd in NSCSs were greater than those in SSs, highlighting the more importance for NSCSs than SSs in controlling behaviours of heavy metals in aquatic environments.