Dissimilar pH-dependent adsorption features of bovine serum albumin and α-chymotrypsin on mica probed by AFM

We studied bovine serum albumin (BSA) and α-chymotrypsin adsorption onto mica surfaces over a large pH range by atomic force microscopy (AFM) measurements in liquid. Data analyses (height, roughness and roughness factor) brought new insights on the conformation of proteins in soil environments, with mica as a model of soil phyllosilicates and non-hydrophobic surfaces. Validation of AFM approach was performed on BSA, whose behavior was previously described by nuclear magnetic resonance and infra-red spectroscopic methods. Maximum adsorption was observed near the isoelectric point (IEP). A stronger interaction and a lower amount of adsorbed proteins were observed below the IEP, which contrasted with the progressive decrease of adsorption above the IEP. We then studied the adsorption of α-chymotrypsin, a proteolytic enzyme commonly found in soils. AFM pictures demonstrated a complete coverage of the mica surface at the IEP in contrast to the BSA case. Comparison of the AFM data with other indirect methods broadened the understanding of α-chymotrypsin adsorption process through the direct display of the protein adsorption patterns as a function of pH.     

Adsorption of bovine serum albumin at the air/water interface and its effect on the formation of DPPC surface film

The adsorption of bovine serum albumin (BSA) at the air/water interface and its effect on the transport of dipalmitoylphosphatidylcholine (DPPC) to form a surface film were studied with tensiometry, infrared reflection absorption spectroscopy (IRRAS), and ellipsometry. For 1, 10, 100, and 1000 ppm BSA solutions, the steady-state tension ranges from 55 to 50 mN m⁻¹. At pulsating area (at 20 cycles min⁻¹), both the minimum and maximum tensions decrease with increasing bulk concentration. Even though the steady-state tension is similar for 100 and 1000 ppm BSA, IRRAS and ellipsometry results indicate that the adsorbed density is higher for 1000 ppm BSA. For 1000 ppm/1000 ppm BSA/DPPC mixture, the tension behavior was found to be similar to that of 1000 ppm BSA when alone. Results from IRRAS and ellipsometry also demonstrate that BSA is the dominant adsorbed component at the air/water interface. Thus, at 1000 ppm, by adsorbing fast and possibly irreversibly, BSA interferes with the transport and adsorption of DPPC and inhibits its ability to lower the surface tension. However, when DPPC is introduced via a spread monolayer mechanism, DPPC expels partly or completely the adsorbed BSA monolayer and then controls the tension behavior with little or no inhibition by BSA. Thus, the competitive adsorption of DPPC and BSA depends strongly on the path or mechanism of introducing DPPC to the surface and involves path-dependent nonequilibrium adsorption phenomena.     

牛血清蛋白对二硬脂酰基磷脂酰乙醇胺单层膜结构的影响

利用Langmuir-Blodgett(LB)技术结合原子力显微镜(AFM),研究了牛血清蛋白(BSA)在气/液界面上对二硬脂酰基磷脂酰乙醇胺(DSPE)单层膜结构的影响.通过改变亚相的pH值和BSA浓度,获得了不同条件下DSPE单层膜的等温线、吸附曲线和压缩循环曲线等.实验结果表明,亚相中BSA的存在对DSPE单层膜的压缩性、稳定性以及相变行为产生了较大的影响.吸附动力学结果表明,DSPE单层膜对BSA分子的吸附量存在一定的阈值,且该阈值的大小与亚相pH值相关.通过分析实验数据可知,当亚相pH=3时,BSA的疏水残基几乎全部暴露在外面,2种分子之间的相互作用最强;而pH=7时,BSA仅有少量的疏水残基暴露在外面,2种分子之间的相互作用最弱.原子力显微镜观测到的单层膜形态变化特点与曲线分析结果一致.该研究为了解牛血清蛋白与磷脂分子之间的相互作用机理提供了重要的实验基础和理论依据.     

Restructuring of hydrophobic surfaces created by surfactant adsorption to mica surfaces

Hydrophobic surfaces created by the adsorption of a monolayer of surfactants, such as CTAB or DODAB, to mica display long-range mutual attraction when placed in water. Initially, this attraction was considered to be due to hydrophobic interaction, but more careful measurements using AFM showed that the surfactant monolayer undergoes rearrangements to produce charged patches on the surface; therefore, the nature of the long-range interaction is due to the electrostatic interaction between patches. The monolayer rearrangement depends on the nature of the surfactant and its counterion. To study possible monolayer rearrangements in molecular detail, we performed detailed molecular dynamics computer simulations on systems containing a monolayer of surfactants RN(CH(3))(3)(+)Cl(-) (R indicates a saturated hydrocarbon chain) adsorbed on a mica surface and immersed in water. We observe that when chain R is 18 carbons long the monolayer rearranges into a micelle but it remains a monolayer when the chain contains 24 carbons.     

Vibrational Spectroscopy As a Probe of Surface Heterogeneity: CO on NaCl(100)

The fundamental (Δv = 1 ← 0) and the first overtone (Δv = 2 ← 0) transitions of monolayer physisorbed CO on NaCl(100) single crystal surfaces at 5 K have been investigated using polarized Fourier transform infrared spectroscopy. Comparisons of the transition frequencies, bandwidths and absorbances for these two transitions together with those of dilute isotopes allow us to isolate the contribution of dynamic and static coupling effects of molecules within the monolayer. Homogeneous and heterogeneous effects can also be distinguished from band profiles of these various transitions. We conclude that the residual line broadening of 0.1 cm^?1 at 5 K for the isolated CO isotopes arises from heterogeneous surface effects. We set an upper limit of 0.07 cm^?1 for the linewidth of single CO molecules on NaCl(lOO) due to surface heterogeneity. A lower limit of × 10^?8 cm^?1 is provided by the vibrational lifetime of adsorbed CO molecules. Sharpness of the vibrational features shows that infrared spectroscopy of adsorbate is a sensitive method for probing surface and adlayer irregularities of adsorbed molecules on a single crystal.     

Dynamic Behaviors and Morphology Change of Anionic Phospholipid DPPG Monolayer Caused by Bovine Serum Albumin at Air-Water Interface

The interaction between bovine serum albumin (BSA) and the anionic 1.2-dipalmitoyl-snglycero- 3-(phospho-rac-(1-glycerol)) (sodium salt) (DPPG) phospholipid at different subphase pH values was investigated at air-water interface through surface pressure measurements and atomic force microscopy (AFM) observation. By analyzing surface pressure-mean molecular area (π-A) isotherms, the limiting molecular area in the closed packing state-the concentration of BSA (A_lim-[BSA]) curves, the compressibility coefficient-surface pressure (C_S^-1-π) curves and the difference value of mean molecular area-the concentration of BSA (ΔA-[BSA]) curves, we obtained that the mean molecular area of DPPG monolayer became much larger when the concentration of BSA in the subphase increased at pH=3 and 5. But the isotherms had no significant change at different amount of BSA at pH=10. In addition, the amount of BSA molecules adsorbed onto the lipid monolayer reached a threshold value when [BSA]>5×10^-8 mol/L for all pHs. From the surface pressure-time (π-t) data, we obtained that desorption and adsorption processes occurred at pH=3, however, there was only desorption process occurring at pH=5 and 10. These results showed that the interaction mechanism between DPPG and BSA molecules was affected by the pH of subphase. BSA molecules were adsorbed onto the DPPG monolayers mainly through the hydrophobic interaction at pH=3 and 5, and the strength of hydrophobic interaction at pH=3 was stronger than the case of pH=5. At pH=10, a weaker hydrophobic interaction and a stronger electrostatic repulsion existed between DPPG and BSA molecules. AFM images revealed that the pH of subphase and [BSA] could affect the morphology features of the monolayers, which was consistent with these curves. The study provides an important experimental basis and theoretical support to understand the interaction between lipid and BSA at the air-water interface.     

Probing protein adsorption onto mercaptoundecanoic acid stabilized gold nanoparticles and surfaces by quartz crystal microbalance and zeta-potential measurements

The adsorption characteristics of three proteins [bovine serum albumin (BSA), myoglobin (Mb), and cytochrome c (CytC)] onto self-assembled monolayers of mercaptoundecanoic acid (MUA) on both gold nanoparticles (AuNP) and gold surfaces (Au) are described. The combination of quartz crystal microbalance measurements with dissipation (QCM-D) and pH titrations of the zeta-potential provide information on layer structure, surface coverage, and potential. All three proteins formed adsorption layers consisting of an irreversibly adsorbed fraction and a reversibly adsorbed fraction. BSA showed the highest affinity for the MUA/Au, forming an irreversibly adsorbed rigid monolayer with a side-down orientation and packing close to that expected in the jamming limit. In addition, BSA showed a large change in the adsorbed mass due to reversibly bound protein. The data indicate that the irreversibly adsorbed fraction of CytC is a monolayer structure, whereas the irreversibly adsorbed Mb is present in form of a bilayer. The observation of stable BSA complexes on MUA/AuNPs at the isoelectric point by zeta-potential measurements demonstrates that BSA can sterically stabilize MUA/AuNP. On the other hand, MUA/AuNP coated with either Mb or CytC formed a reversible flocculated state at the isoelectric point. The colloidal stability differences may be correlated with weaker binding in the reversibly bound overlayer in the case of Mb and CytC as compared to BSA.     

氢原子在Be(0001)表面吸附的密度泛函理论研究

采用第一性原理的密度泛函理论研究单个氢原子和多个氢原子在Be(0001)表面吸附性质.给出了氢吸附Be(0001)薄膜表面的原子结构、吸附能、饱和度、功函数、偶极修正等特性参数.同时也讨论了相关吸附性质与氢原子覆盖度(0.06-1.33ML)的关系.计算结果表明:氢原子的吸附位置与覆盖度之间有强烈的依赖关系,覆盖度低于0.67ML时,氢原子能量上易于占据fcc或hcp的中空位置;覆盖度为0.78ML时,中空位与桥位为氢原子的最佳吸附位;覆盖度在0.89到1.00ML时,桥位是氢原子吸附能量最有利的位置;以上覆盖度中Be(0001)表面最外层铍原子的结构均没有发生明显变化.当覆盖度为1.11-1.33ML,高覆盖度下Be(0001)表面的最外层铍原子部分发生膨胀,近邻氢原子渗入到铍表面次层,氢原子易于占据在hcp和桥位.吸附结构中的氢原子比氢分子中的原子稳定.当覆盖度大1.33ML时,计算结果没有发现相对于氢分子更稳定的吸氢结构.同时从分析偶极修正和氢原子吸附垂直高度随覆盖度的变化关系判断氢覆盖度为1.33ML时,在Be(0001)表面吸附达到饱和.     

Interactions of adsorbed albumin with underpotentially deposited copper on gold piezoelectrodes

The adsorption of a model protein, bovine serum albumin (BSA), on Au electrodes was investigated using the Cu adatom probe method and Electrochemical Quartz Crystal Nanobalance (EQCN) technique. The adsorption of BSA was confirmed by AFM imaging and has been found to be controlled by kinetics. Using the Cu adatom probe method, we were able to reconstruct the entire BSA adsorption transient Theta(BSA) vs. t. The adsorption rate constant k(1), determined from this transient is k(1)=2.45x10(5) L mol(-1) s(-1). We have found that the bulk Cu(0) deposition process is blocked by BSA adsorption and it decays exponentially with time during BSA adsorption. It ceases completely when a full monolayer of BSA is formed. In contrast to that, the mass associated with Cu-u.p.d. decreases only to ca. 50% of that in the absence of BSA, indicating that Cu adatoms can penetrate (wedge) into the space between the surface Au atoms and the adsorbed BSA molecules. In addition to that, we have found that the degree of penetration of Cu adatoms can be controlled by the applied deposition potential. By selecting a sufficiently cathodic potential, we were able to deposit a full Cu-u.p.d. monolayer, independent of the BSA surface coverage extending from Theta(BSA)=0 to Theta(BSA) approximately 1. The positive shift of Cu(ad) desorption peak potential E(p), observed in the presence of adsorbed BSA, has been interpreted in terms of Frumkin exchange interaction forces between Cu(ad) and BSA(ad), on the basis of our earlier theoretical model, expanded here to include adsorbed species in two monolayers. This expansion is possible owing to the fast rate of Cu adatom penetration in the interfacial region. From the plots of E(p) vs. Theta(BSA), the presence of strong attractive interactions between Cu(ad) and BSA(ad) was deduced. These interactions result in a super-shift of the Cu-u.p.d. desorption peak potential, corresponding to the exchange interaction coefficient g(M,X)<-4, indicating on a possibility of the formation of a stable interface complex.     

牛血清白蛋白在气-液界面上的吸附行为及其与含芘手性探针分子的相互作用研究

利用表面压力-时间曲线对牛血清白蛋白(BSA)在气液界面上的吸附行为和对手性探针分子D/L-[4-(1-芘基)]丁酰基-苯丙氨酸(PPs)的界面手性识别, 以及由此引起的气液界面上BSA的构象变化进行了研究. 结果表明, 界面上形成的稳定单层膜经历了漫长的构象调整过程; BSA的表面压力的变化说明其对亚相中的探针分子很强的浓度依赖性和对手性分子的区分能力. 在较高的PLP和PDP探针分子浓度下, BSA的成膜性均受到了很大抑制, 但较低的PLP和PDP探针分子浓度却转而对BSA成膜有利; 与PLP相比, PDP能更有效地与BSA在界面结合, 其复合膜的稳定性更好.     

Bovine serum albumin adsorption onto colloidal Al2O3 particles: a new model based on zeta potential and UV-vis measurements

We investigated the adsorption of bovine serum albumin (BSA) on colloidal Al2O3 particles in an aqueous environment. Changes in the zeta potential of the Al2O3 particles upon the adsorption of BSA were measured using an electro-acoustic technique. The mass of protein adsorbed was determined by using UV-vis spectroscopy. The change of the isoelectric point of the Al2O3 powder-protein suspension was found to be a function of adsorbed protein mass. It was shown that approximately one monolayer of BSA was needed to fully mask the surface and to compromise the charge of Al2O3. From titration experiments it follows that about 30-36% of the negatively charged groups of the protein form bonds with the protonated and charged Al2O3 surface. On the basis of our observations we introduced a new adsorption model for BSA on Al2O3 particles.     

Controlled rearrangement of adsorbed undecanol films on mica surfaces induced by an atomic force microscopy tip

An undecanol film adsorbed on a mica surface was found to rearrange and spread in a position-controlled way induced by a tapping mode atomic force microscopy (AFM) probe. AFM images of varying scanning times showed that before forming an ordered monolayer the undecanol molecules were adsorbed on the mica surface in the disordered and disorganized status. With the proceeding of scanning, these undecanol molecules gradually formed an ordered and flat film. Such behavior was caused by the formation of a stable film and had never been reported for other alcohols.     

Determination of carcinoembryonic antigen using a novel amperometric enzyme-electrode based on layer-by-layer assembly of gold nanoparticles and thionine

Electrochemical sensing of carcinoembryonic antigen(CEA)on a gold electrode modified by the se- quential incorporation of the mediator,thionine(Thi),and gold nanoparticles(nano-Au),through co- valent linkage and electrostatic interactions onto a self-assembled monolayer configuration is de- scribed in this paper.The enzyme,horseradish peroxidase(HRP),was employed to block the possible remaining active sites of the nano-Au monolayer,avoid the non-specific adsorption,instead of bovine serum albumin(BSA),and amplify the response of the antigen-antibody reaction.Electrochemical ex- periments indicated highly efficient electron transfer by the imbedded Thi mediator and adsorbed nano-Au.The HRP kept its activity after immobilization,and the studied electrode showed sensitive response to CEA and high stability during a long period of storage.The working range for the system was 2.5 to 80.0 ng/mL with a detection limit of 0.90 ng/mL.The model membrane system in this work is a potential biosensor for mimicking the other immunosensor and enzyme sensor.     

Synthesis and characterization of poly(ethylene glycol) methyl ether endcapped poly(ethylene terephthalate)

Linear and branched poly(ethylene terephthalate) (PET) copolymers with polyethylene glycol) (PEG) methyl ether (700 or 2000 g/mol) end groups were synthesized using conventional melt polymerization. DSC analysis demonstrated that low levels of PEG end groups accelerated PET crystallization. The incorporated PEG end groups also decreased the crystallization temperature of PET dramatically, and copolymers with a high content of PEG (>17.6 wt%) were able to crystallize at room temperature. Rheological analysis demonstrated that the presence of PEG end groups effectively decreased the melt viscosities and facilitated melt processing. XPS and ATR-FTIR revealed that the PEG end groups tended to aggregate on the surface, and the surface of compression molded films containing 34.0 wt% PEG were PEG rich (85 wt% PEG). PEG end-capped PET (34.0 wt% PEG) and PET films were immersed into a fibrinogen solution (0.7 mg/mL BSA) for 72 h to investigate the propensity for protein adhesion. XPS demonstrated that the concentration of nitrogen (1.05%) on the surface of PEG endcapped PET film was statistically lower than PET (7.67%). SEM analysis was consistent with XPS results, and revealed the presence of adsorbed protein on the surface of PET films.     

Measuring hydrogen–deuterium exchange in protein monolayers

A Fourier transform infrared (FTIR) spectroscopy assay to measure hydrogen–deuterium exchange (HDX) in surface‐adsorbed protein monolayers is developed to provide information on protein tertiary structure, because the typical secondary structural analysis of our surface and solution protein samples proved to be very similar. Adsorbed protein HDX is quantified by exposing the protein to a 50% deuterated NaPO₄ buffer solution and then measuring the normalized intensity change of the amide II band in the FTIR reflection spectrum. When collected as a function of exchange time, this intensity follows the kinetics of the exposure of the protein amides to solvent. HDX kinetics have been obtained for bovine serum albumin (BSA) in solution and adsorbed to gold surfaces. Using experiments designed to allow comparisons between protein in solution and on surfaces, the extent of HDX was found to increase over that observed for BSA in solution, consistent with an increase in the exposure of albumin amide groups and protein unfolding upon adsorption. We also show that BSA adsorbs to the surface of gold in multilayers and that the increase in amide exposure is present only in the first adsorbed monolayer. Published in 2009 by John Wiley & Sons, Ltd.     

Catalytic currents of albumin at the hanging mercury drop electrode

Bovine serum albumin (BSA), as well as completely reduced BSA denoted by P (SH)₃₅, are adsorbed on the hanging mercury drop electrode (HMDE) from alkaline buffer solutions. When time is allowed, a monolayer is adsorbed from very dilute (10^?9M) BSA solutions in ammoniacal and borate buffers. With a monolayer of adsorbed protein the voltammograms at the HMDE are then identical in a given ammoniacal or borax buffer containing cobalt(III) or (II) and different BSA concentrations. Voltammograms of P (SH)₃₅ are virtually identical with those of native BSA. At the HMDE the second Brdi?ka current is proportional to concentration of cobalt(III) or (II) and the first current nearly so. Incompletely or completely adsorbed BSA or P (SH)₃₅ is not desorbed on keeping the HMDE for one hour in ammonia buffers. An incomplete layer of adsorbed BSA or P (SH)₃₅ is relatively rapidly desorbed at ?1.6 V (vs. SCE) and a complete film at ?1.65 V, some desorption occurring at ?1.6 V. Upon desorption, the second Brdi?ka current decreases faster than the first one; this is particularly striking in 1 M ammonia buffer. The rate of desorption is increased by calcium chloride, but the rate of adsorption is not, or only slightly, increased in the presence of calcium. Incomplete adsorption occurs at ?1.60 V (vs. SCE) and no adsorption at ?1.65 V. Indications are obtained that “presodium currents” yield a slight plateau at ?1.67 to ?1.70 V, the plateau currents being attributed to adsorbed BSA, while unadsorbed BSA yields catalytic currents without a plateau, the currents merging with the residual one of the buffer. Calcium chloride greatly increases the presodium currents. From many kinetic data obtained at the dropping mercury electrode (DME) and from results at the HMDE it is concluded that, depending on the BSA concentration, Brdi?ka currents at the DME are partly of a kinetic and partly of a surface adsorption nature and partly diffusion-controlled. Adsorption equilibrium is not attained at the DME at 25° at concentrations of BSA smaller than 10^?6M.     

INTERACTIONS BETWEEN POLY(ETHYLENE OXIDE) COATED SURFACES AND BETWEEN SUCH SURFACES AND PROTEINS

Surfaces coated with poly(ethylene oxide) containing nonionic polymers are of interest in medical applications due to, among other things, the low adsorption of proteins on such surfaces. In this paper we have studied the interfacial properties of surfaces coated with PEO by measuring the forces acting between two such surfaces in water and across a protein solution as well as between one such surface and a surface carrying adsorbed proteins. One type of surface coating was a graft copolymer of poly(ethylene imine) and poly(ethylene oxide) where the cationic poly(ethylene imine) group anchored the polymer to negatively charged mica surfaces. Three different ways to prepare this coating was used and compared. It was found that this coating was not stable in the presence of lysozyme, a small positively charged protein, when the PEO graft density was low. The other type of coating was obtained by adsorbing ethyl(hydroxyethyl)-cellulose onto hydrophobised mica surfaces. The driving force for adsorption is in this case the hydrophobic interaction between nonpolar segments of the polymer and the surface. The EHEC coating was stable in the presence of lysozyme and the interactions between adsorbed layers of lysozyme and EHEC coated surfaces are purely repulsive due to long-range steric forces.     

Generation of nanostructures of mica supported lysozyme molecules in aqueous solution by atomic force microscopy

Nanostructures of lysozyme molecules adsorbed to mica were generated by the tip of an atomic force microscope in contact, tapping, and force-distance mode in aqueous solution. In contact mode at high ionic strength and adjusted lysozyme concentration a monolayer of defined pattern and orientation could be formed by the scan process of the tip. A lysozyme monolayer with minimal pattern size of about 60 nm was achieved by line scan. At larger loading forces besides a monolayer also 3D-aggregates of lysozyme molecules could be generated. In force-distance mode the volume of 3D-aggregates grows with increasing generation time, lysozyme concentration in the bulk phase, loading force, and frequency of up- and down-movement of the substrate toward the fixed cantilever. In tapping mode 3D-aggregates could be generated as well. It is postulated that reduction of electrostatic interaction between the oppositely charged lysozyme molecules and mica surface by sufficient high ionic strength is essential for monolayer formation. It is discussed that for the underlying mechanism of monolayer generation in contact mode lysozyme molecules of the bulk phase adsorb to the tip, become pulled off and attach to the mica surface by the scan process of the tip.     

Fe(CO)5 thin films adsorbed on Au(111) and on self-assembled organic monolayers: II. Thermal transformations

The thermal transformations of as-deposited Fe(CO)(5) films adsorbed on Au(111)/mica and C(4), C(8), C(12), and C(16) self-assembled methyl-terminated monolayer organic surfaces have been studied using infrared spectroscopy to probe how the physical restructuring influences the sensitivity of these systems to low-energy electron beams. A companion publication shows that the as-deposited monolayers are composed of molecules physisorbed with one axial and two equatorial carbonyl groups directed toward the substrate; subsequent layers are preferentially oriented with the C(3) molecular axis aligned perpendicular to the substrate (i.e., one axial carbonyl group directed toward the substrate). In this work, we show that the as-deposited films are structurally unstable above 125 K on Au(111)/mica surfaces and above 100 K on the organic self-assembled monolayers. Above these thresholds, the layered structures transform into three-dimensional aggregates, implying strongly nonwetting behavior for Fe(CO)(5) on each of these substrates; molecular desorption from this aggregate structure takes place between 140 and 160 K. The irreversibility of this temperature-induced transformation demonstrates that the as-deposited layered films do not represent a thermodynamically well-defined phase; this key feature of the as-deposited films is believed to be the cause of the discrepancies in previous attempts to understand Fe(CO)(5)/surface structures based on infrared results. Moreover, the thermally induced transformation to 3D aggregate structures is shown to decrease the apparent sensitivity of the adsorbed Fe(CO)(5) to low-energy electron-induced decarbonylation (0-10 eV) by over 3 orders of magnitude.     

Structural and topographical characteristics of adsorbed WPI and monoglyceride mixed monolayers at the air-water interface

In this work we have analyzed the structural and topographical characteristics of mixed monolayers formed by an adsorbed whey protein isolate (WPI) and a spread monoglyceride monolayer (monopalmitin or monoolein) on the previously adsorbed protein film. Measurements of the surface pressure (pi)-area (A) isotherm were obtained at 20 degrees C and at pH 7 for protein-adsorbed films from water in a Wilhelmy-type film balance. Since the surface concentration (1/A) is actually unknown for the adsorbed monolayer, the values were derived by assuming that the A values for adsorbed and spread monolayers were equal at the collapse point of the mixed film. The pi-A isotherm deduced for adsorbed WPI monolayer in this work is practically the same as that obtained directly by spreading. For WPI-monoglyceride mixed films, the pi-A isotherms for adsorbed and spread monolayers at pi higher than the equilibrium surface pressure of WPI are practically coincident, a phenomenon which may be attributed to the protein displacement by the monoglyceride from the interface. At lower surface pressures, WPI and monoglyceride coexist at the interface and the adsorbed and spread pi-A isotherms (i.e., the monolayer structure of the mixed films) are different. Monopalmitin has a higher capacity than monoolein for the displacement of protein from the air-water interface. However, some degree of interactions exists between proteins and monoglycerides and these interactions are higher for adsorbed than for spread films. The topography of the monolayer corroborates these conclusions.