biomimetics
Article Surfactant Interactions with Protein-Coated Surfaces: Comparison between Colloidal and Macroscopically Flat Surfaces
Helena Mateos 1,* , Alessandra Valentini 2, Francesco Lopez 3 and Gerardo Palazzo 4
1 CSGI (Center for Colloid and Surface Science), via Orabona 4, 70125 Bari, Italy 2 School of Chemical Engineering, University of Birmingham, Birmingham B15 2TT, UK; [email protected] 3 Department of Agricultural, Environmental and Food Sciences (DiAAA) and CSGI, University of Molise, Via De Sanctis, 86100 Campobasso, Italy; [email protected] 4 Department of Chemistry, University of Bari, via Orabona 4, 70125 Bari, Italy; [email protected] * Correspondence: [email protected]
Received: 16 May 2020; Accepted: 29 June 2020; Published: 1 July 2020
Abstract: Surface interactions with polymers or proteins are extensively studied in a range of industrial and biomedical applications to control surface modification, cleaning, or biofilm formation. In this study we compare surfactant interactions with protein-coated silica surfaces differing in the degree of curvature (macroscopically flat and colloidal nanometric spheres). The interaction with a flat surface was probed by means of surface plasmon resonance (SPR) while dynamic light scattering (DLS) was used to study the interaction with colloidal SiO2 (radius 15 nm). First, the adsorption of bovine serum albumin (BSA) with both SiO2 surfaces to create a monolayer of coating protein was studied. Subsequently, the interaction of these BSA-coated surfaces with a non-ionic surfactant (a decanol ethoxylated with an average number of eight ethoxy groups) was investigated. A fair comparison between the results obtained by these two techniques on different geometries required the correction of SPR data for bound water and DLS results for particle curvature. Thus, the treated data have excellent quantitative agreement independently of the geometry of the surface suggesting the formation of multilayers of C10PEG over the protein coating. The results also show a marked different affinity of the surfactant towards BSA when the protein is deposited on a flat surface or individually dissolved in solution.
Keywords: protein-coated surfaces; surfactant; DLS; ζ potential; BSA; colloids
1. Introduction Protein/surfactant interactions find important applications in industry and science. Their behavior at the air/liquid or liquid/liquid interface has been widely investigated due to their importance in the formation and stabilization of foams and emulsions [1–7]. Protein adsorption to solid surfaces is also related to dirt cleaning, or ultimately, it can be associated with biofilm formation, in which case, it is important to minimize or remove proteins adsorbed to the solid surface [8–10]. Studies of adsorption and protein/surfactant interaction at the liquid/solid interface are, however, not common. Furthermore, predicting protein/surfactant interactions at a solid surface is often difficult and not intuitive, especially when dealing with soft and flexible proteins that might change conformation when in contact with a surface. The subject of this paper is to investigate the potential use of colloidal surfaces instead of flat macromolecular ones for the research of interactions at the liquid/solid interface. The use of colloidal surfaces in combination with dynamic light scattering (DLS) could present many potential advantages since it can be applied to a wide range of materials including those that are unsuited for the external
Biomimetics 2020, 5, 31; doi:10.3390/biomimetics5030031 www.mdpi.com/journal/biomimetics Biomimetics 2020, 5, x FOR PEER REVIEW 2 of 13 Biomimetics 2020, 5, 31 2 of 13 (DLS) could present many potential advantages since it can be applied to a wide range of materials including those that are unsuited for the external coatings. Furthermore, working with colloidal solutionscoatings. increases Furthermore, substantially working the with analyzed colloidal surface solutions area, increases which could substantially lead to more the analyzedrepresentative surface results.area, which could lead to more representative results. WeWe have have studied studied thethe interactioninteraction between between bovine bovine serum serum albumin albumin (BSA) (BSA) and the and non-ionic the non surfactant‐ionic C10PEG, at the glass/water interface in flat and colloidal forms (Scheme1). Using glass is an interesting surfactant C10PEG, at the glass/water interface in flat and colloidal forms (Scheme 1). Using glass is ancase interesting scenario case since scenario it is a very since common it is a very household common surface. household On the surface. other On hand, the BSA’s other adsorptionhand, BSA’s at a variety of interfaces has been extensively studied previously [10–15] and C10PEG is a common and adsorption at a variety of interfaces has been extensively studied previously [10–15] and C10PEG is a commonrelevant and surfactant relevant widely surfactant used widely in industry. used Inin thisindustry. paper, In we this compare paper, thewe resultscompare of the interactionsresults of thebetween interactions both between solid-surface both conformationssolid‐surface conformations using surface using plasmon surface resonance plasmon (SPR), resonance dynamic (SPR), light dynamicscattering light (DLS), scattering and laser (DLS), Doppler and laser electrophoresis Doppler electrophoresis (LDE) measurements. (LDE) measurements.
SchemeScheme 1. Graphical 1. Graphical representation representation of ofthe the interaction interaction between between bovine bovine serum serum albumin albumin (BSA) (BSA)-coated‐coated
SiOSiO2 and2 and surfactant surfactant on onboth both flat flat (A) ( Aand) and colloidal colloidal surfaces surfaces (B). (B ).
2. 2.Materials Materials and and Methods Methods BovineBovine serum serum albumin albumin (BSA) (BSA) protein protein (purity (purity > 98%)> 98%) and and Ludox Ludox AM AM colloidal colloidal silica silica system system were were purchasedpurchased from from Sigma Sigma-Aldrich‐Aldrich (St. (St. Louis, Louis, MO, MO, USA) USA) and and used used without without any any further further purification. purification. Na HPO and NaH PO were purchased from Fluka (Buchs, Switzerland) and J. T. Baker (Phillipsburg, Na2HPO2 4 and4 NaH22PO44 were purchased from Fluka (Buchs, Switzerland) and J. T. Baker (Phillipsburg,NJ, USA), respectively. NJ, USA), respectively. The water was The purified water was by means purified of aby Milli-Q means system of a Milli (Millipore‐Q system Corp., (Millipore Bedford, Corp.,MA, Bedford, USA) and MA, hereafter USA) is and denoted hereafter as MQ is denoted water. Aqueous as MQ water. phosphate Aqueous buffer phosphate was prepared buffer from was MQ preparedwater. The from surfactant MQ water. (decanol The surfactant ethoxylated (decanol with an averageethoxylated number with of an eight average ethoxy number (EO) groups) of eight was ethoxysupplied (EO) by groups) Sasol with was asupplied purity grade by Sasol of 95%. with a purity grade of 95%. WeWe used used surface surface plasmon plasmon resonance resonance (SPR) (SPR) to to evaluate evaluate the the interaction interaction of of C10 CPEG10PEG on on a previously a previously depositeddeposited BSA BSA layer layer on on a a flat, flat, macroscopic silica-coatedsilica‐coated sensor sensor surface surface (the (the thickness thickness of of the the silica silica layer layerwas was 10 nm). 10 nm). A dual-wavelength A dual‐wavelength (670 (670 and and 785 785 nm), nm), multiparametric multiparametric SPR SPR instrument instrument (MP–SPR (MP–SPR Navi Navi200, 200, BioNavis BioNavis Ltd., Tampere,Ltd., Tampere, Finland) Finland) was used was to recordused to the record SPR spectra. the SPR Contrary spectra. to traditionalContrary SPR,to traditionalMP SPR allowsSPR, MP the SPR measurement allows the measurement up to 5 µm above up to the 5 μ solidm above surface. the solid Spectra surface. were acquiredSpectra were in the acquiredrange of in 50 the◦ torange 77.5 ◦of. The50° to fluidic 77.5°. system The fluidic was system cleaned was before cleaned and afterbefore each and experiment after each experiment using a glass usingslide a inglass the slide place in of the the place sensor. of A the flow sensor. of 200 Aµ flowL/min of of 200 purified μL/min MQ of waterpurified was MQ set water for 10 minwas followedset for 10by min a 1%followed SDS solution by a 1% in SDS water solution for 10 in min water and for the 10 final min flow and of the MQ final water flow for of other MQ water 10 min. for BSA other and 10surfactant min. BSA and samples surfactant were preparedsamples were in phosphate-bu prepared inff phosphateered saline‐buffered (PBS) (pH saline 7.2) (PBS) and injected (pH 7.2) (250 andµ L injectedinjection (250 loops) μL injection on a silica-coated loops) on a sensorsilica‐coated in kinetic sensor titration in kinetic mode titration at a constant mode at flow a constant of 20 µ flowL/min. of All20 μ theL/min. experiments All the experiments were done in were triplicate. done in The triplicate. affinity The constants affinity were constants estimated were using estimated Winspall using 3.02 Winspall(Max Planck 3.02 Institute(Max Planck for Polymer Institute Research, for Polymer Mainz, Germany)Research, andMainz, OriginPro Germany) (OriginLab and Corporation,OriginPro (OriginLabNorthampton, Corporation, MA, USA) Northampton, software. MA, USA) software. TheThe interaction interaction between between colloidal colloidal particles particles and and BSA BSA was was studied studied by by adding adding increasing increasing BSA BSA concentrationsconcentrations ranging ranging from from 0.08 0.08 to to8 mg/mL 8 mg/mL to toa 2.5 a 2.5 mg/mL mg/mL Ludox Ludox solution solution in in10 10 mM mM phosphate phosphate bufferbuff erat atpH pH = 7.4= 7.4. . The The resulting resulting samples samples were were analyzed analyzed by by following following the the changes changes in ζ‐ inpotentialζ-potential (laser (laser DopplerDoppler electrophoresis, electrophoresis, LDE) andand size size distribution distribution (dynamic (dynamic light light scattering, scattering, DLS) usingDLS) a using Nanosizer a NanosizerZS (Malvern ZS (Malvern instruments, instruments, Malvern, Malvern, UK). Instrumental UK). Instrumental conditions conditions are explained are explained elsewhere elsewhere (see [16] for (seeLDE [16] and for [17 ]LDE for DLS.and The[17]ζ -potentialfor DLS. wasThe subsequently ‐potential evaluatedwas subsequently from the electrophoreticevaluated from mobility the electrophoreticaccording to themobility Smoluchowski according approximation. to the Smoluchowski approximation.
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ToTo studystudy the the interactions interactions between between BSA-coated BSA‐coated Ludox Ludox particles particles and and surfactant surfactant C10 CPEG10PEG, we, we chose chose a solutiona solution of of 2.5 2.5 mg mg/mL/mL Ludox Ludox and and 8 mg 8 mg/mL/mL BSA BSA concentration. concentration. Then, Then, surfactant surfactant was was added added in ain 0.5 a 0.5 to 10to wt%10 wt% concentration concentration range range keeping keeping the the bu ffbufferer concentration concentration constant. constant. 1 1H didiffusionffusion NMR andand fluorescencefluorescence measurementsmeasurements were performedperformed usingusing thethe samesame set-upsset‐ups describeddescribed elsewhereelsewhere [[17].17].
3.3. Results and Discussion TheThe experimentalexperimental strategystrategy used inin thisthis investigationinvestigation isis asas follows:follows: from SPRSPR experimentsexperiments it isis possible to obtain information on the deposition of the analytes on a flat SiO surface. Then, DLS and possible to obtain information on the deposition of the analytes on a flat SiO22 surface. Then, DLS and EDL can be used to obtain the same information on a nano-sized SiO surface. The comparison of both EDL can be used to obtain the same information on a nano‐sized 2SiO2 surface. The comparison of setsboth of sets experiments of experiments allows allows us to compare us to compare the use ofthe flat use versus of flat nanosized versus nanosized surfaces from surfaces the interaction from the of BSA and further interaction between BSA and a non-ionic C PEG surfactant. interaction of BSA and further interaction between BSA and a 10non‐ionic C10PEG surfactant. 3.1. Interaction of BSA on a Silica Surface 3.1. Interaction of BSA on a Silica Surface A first analysis of the interaction between BSA and SiO2 is necessary to understand how to obtain A first analysis of the interaction between BSA and SiO2 is necessary to understand how to obtain a monolayer with a good surface coverage of the protein on the sensor chip’s surface. Moreover, a monolayer with a good surface coverage of the protein on the sensor chip’s surface. Moreover, understanding the deposition of BSA on the SiO2 surface is crucial to further investigate its interaction understanding the deposition of BSA on the SiO2 surface is crucial to further investigate its interaction with surfactants. Therefore, we injected increasing concentrations of the protein to extract information with surfactants. Therefore, we injected increasing concentrations of the protein to extract about the affinity and needed concentration to create a good coverage on the chip. The experiment information about the affinity and needed concentration to create a good coverage on the chip. The shown in Figure1 is performed by sequential injections (also called kinetic titration) of the analyte experiment shown in Figure 1 is performed by sequential injections (also called kinetic titration) of (BSA) in increasing concentrations. After the first injection is finished, and only the buffer is being the analyte (BSA) in increasing concentrations. After the first injection is finished, and only the buffer flushed, the SPR intensity signal reaches a stable plateau, meaning that the analyte is not being desorbed is being flushed, the SPR intensity signal reaches a stable plateau, meaning that the analyte is not from the surface. Then, the second analyte concentration is injected, and so on. The change in the being desorbed from the surface. Then, the second analyte concentration is injected, and so on. The minimum angle with time can be related to either, an increase in the thickness of the deposited layer, change in the minimum angle with time can be related to either, an increase in the thickness of the or an increase in surface coverage. deposited layer, or an increase in surface coverage.
FigureFigure 1.1. Surface plasmon resonance (SPR) sensogram of a kinetic titration of BSA on aa macroscopicmacroscopic silica-coatedsilica‐coated sensorsensor (the(the thicknessthickness ofof thethe silicasilica layerlayer waswas 1010 nm).nm). The surface coverage (ng/cm2) is usually calculated using the De Feiter equation(Equation (1)) [18], The surface coverage (ng/cm2) is usually calculated using the De Feiter equation(Equation (1)) which predicts the surface coverage to be the optical thickness (D0 in Equation (2)) divided by the [18], which predicts the surface coverage to be the optical thickness (D0 in Equation (2)) divided by refractive index increment with concentration (dn/dC Equation (3)): the refractive index increment with concentration (dn/dC Equation (3)):
D 0 ΓΓ = (1)(1) dn / /dC