Langmuir Technique Research Articles

Creation of giant two-dimensional crystal of zinc oxide nanodisk by method of single-particle layer of organo-modified inorganic fine particles

In this study, the formation and structure of a single-particle layer of organo-zinc oxide are investigated using surface-pressure-area (π-A) isotherms, out-of-plane X-ray diffraction (XRD) analysis, and atomic force microscopy (AFM). Further, techniques for achieving the solubilization of inorganic fine particles in general solvents have been proposed, and a single-particle layer has been formed using such an inorganic solution as a “spreading solution” for an interfacial film. Surface modification of ZnO is performed using a long-chain carboxylic acid. Accordingly, a regular arrangement of ZnO can be easily achieved in order to overcome the relatively weak van der Walls interactions between inorganic materials. A condensed Langmuir monolayer of these particles is also formed. A multiparticle layered structure is constructed by the Langmuir–Blodgett (LB) technique. Out-of-plane XRD measurement results for a single-particle layer of organo-ZnO clearly show a sharp peak at 42Å. This peak is attributed to the distance between ZnO layers. The AFM image of this single-particle layer of organo-ZnO shows a particle assembly with a uniform height of 60nm. These aggregated particles form large two-dimensional crystals. In other words, a regular periodic structure along the c-axis and a condensed single-particle layer had been fabricated using Langmuir and LB techniques.

Structure and Electrical Properties of the Selected Crown Ether-Derivatized Bis(2,2'-bithienyl)Methanes in Langmuir-Blodgett Films

Physical properties of thin films of thiophene derivatives and their polymers are greatly affected by the structure and orientation of their molecules, as well as the local environment.1 In order to obtain thin films of such compounds for practical applications, from electronic devices to chemo- and biosensors.2-3, it is important to control orientation and order of the molecules in the formed functional materials. Interesting and efficient way to allow for such a control is Langmuir and Langmuir-Blodgett technique. In the presented contribution formation and properties of ultrathin films of the series of bis(2,2’-bithienyl)methane derivatives appended with crown ethers of varying size are presented. Uniform Langmuir films of liquid-expanded type were obtained at water-air interface for all of the used compounds. The influence of metal ions on the orientation of the molecules in the films was relatively small. Obtained Langmuir films were subsequently transferred onto the solid substrates by using Langmuir-Blodgett technique forming mono- and multilayers. In order to further elucidate structure of the formed layers AFM and STM techniques have been applied. The morphology of the multilayer films depended both on the size and orientation of the crown ether moiety in the molecules. However, in all cases formed multilayers were porous (Fig. 1). In case of monolayers, the STM studies helped to ascertain organization of the molecules in the Langmuir-Blodgett film. On the other hand the scanning tunnelling spectroscopy (STS) allowed acquisition of information about the electrical properties of the studied thiophene derivatives. Bibliography 1. D. Hu, J. Yu, K. Wong, B. Bagchi, P. J. Rossky, P. F. Barbara, Nature, 405, (2000), 1030. 2. A. G. MacDiarmid, Angew. Chem., Int. Ed., 40, (2001),2581. 3. R. Singhal, A. Chaubey, K. Kaneto, W. Takashima, B. D. Malhotra, Biotechnol. Bioeng., 85 (2004), 277. Figure 1

Creation of high-density and low-defect single-layer film of magnetic nanoparticles by the method of interfacial molecular films.

A technique to solubilize fine magnetic inorganic particles in general organic solvents is proposed via surfaces modification by long-chain carboxylic acids. This organic modification should overcome the relatively weak van der Waals interactions between the nanoparticles, allowing the formation of ordered arrangements of the modified Fe3O4 and CoFe2O4 materials. Using nanodispersions of these organo-modified magnetic nanoparticles as "spreading solutions", Langmuir monolayers of these particles were formed. Multiparticle layered structures were constructed by the Langmuir-Blodgett (LB) technique. The fabrication of single- and multiparticle layers of organo-modified magnetic nanoparticles was investigated using surface pressure-area (π-A) isotherms, out-of-plane X-ray diffraction (XRD), in-plane XRD, and atomic force microscopy (AFM). The out-of-plane XRD profile of a single-particle layer of organo-modified Fe3O4 clearly showed a sharp peak which was attributed to the distance between Fe3O4 layers along the c-axis. The AFM image of single-particle layer of organo-modified CoFe2O4 revealed integrated particle organization with a uniform height; these aggregated particles formed large two-dimensional crystals. For both nanoparticle species, regular periodic structures along the c-axis and high-density single-particle layers were produced via the Langmuir and LB techniques.

Electrical and electrochemical measurements in nanostructured films of polythiophene derivatives

Thin films from polythiophene derivatives can be fabricated by some experimental methods, including the Langmuir techniques, which are particularly suitable if some degree of molecular control is required. In this study, electrical and electrochemical measurements in Langmuir-Schaefer and spin-coated films of polythiophene derivatives were carried out in order to compare the influence of the deposition technique in these properties. Through cyclic voltammetry and UV-visible absorption measurements it was possible to estimate the energy diagrams for all the polythiophene films. Besides, electrochromic effect in the polythiophene thin films was observed.

Supramolecular arrangements of an organometallic forming nanostructured films

Organometallic materials have become subject of intensive research on distinct technological applications. The Langmuir-Blodgett (LB) and Langmuir-Schaefer (LS) techniques have proven to be suitable to address challenges inherent to organic devices, in which the film properties can be tuned at molecular level. Here, we report on the supramolecular arrangement of zinc(II)-protoporphyrin(IX) dimethyl ester (ZnPPIX-DME) using the Langmuir, LB and LS techniques, leading to nanostructured films. The π-A isotherms showed that π-π stacking interaction among ZnPPIX-DME molecules takes place at the air/water interface, favoring the formation of Langmuir films closely packed. The controlled growth of the LB and LS films was monitored via UV-Vis absorption spectroscopy, with the thickness per monolayer within 1.3 and 1.7 nm. The homogeneous topography found at microscale is no longer preserved at nanoscale, which is found rougher according to AFM data. The FTIR indicated that the ZnPPIX-DME is isotropically arranged on both LB and LS films.

Comparison of cholesterol and 25-hydroxycholesterol in phase-separated langmuir monolayers at the air-water interface.

Langmuir monolayer studies combined with fluorescence microscopy provide powerful insights into the phase behavior of cholesterol and cholesterol analogue/phospholipid monolayer systems at the air-water interface. These studies have established the ability of cholesterol and similar molecules to condense the average molecular area of the monolayer as well as to laterally organize the monolayer into sterol-rich and sterol-poor regions. Oxysterols are one class of molecules that deserve particular attention due to their metabolic and physical effects on the membrane and the functioning of mammalian cells. We systematically explore the miscibility of 25-hydroxycholesterol (25OH) with 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) in lipid monolayers. Like cholesterol, the 25OH/DMPC system exhibits phase separation; however, due to the difunctional nature of 25OH, there are significant differences. Using fluorescence microscopy and traditional Langmuir techniques, we investigate the average molecular packing and micron scale phase separation within a 25OH/phospholipid monolayer. We present evidence of the nucleation and growth of domain formation, the expansion of the monolayer induced by 25OH, and a model to describe our results. We conclude that 25OH and other similar hydroxysterols provide a useful and independent test of cholesterol's behavior within monolayer leaflets.

Formation and structure of fine multi-particle layered organo-modified zirconium dioxides fabricated by Langmuir–Blodgett technique

Formation and structure of multilayered organization of organo-modified zirconium dioxide are investigated by surface pressure–area (π–A) isotherms, out-of-plane and in-plane X-ray diffraction (XRD) analysis, and atomic force microscopy (AFM). In this study, solubilization technology of inorganic fine particles into general solvent has proposed and formation technology of highly ordered single-particle layer has also established by using that inorganic solution as “spreading solution” of the interfacial film. The surface modification of zirconia particles is performed using several long-chain carboxylic acids of different lengths. Accordingly, it is easily achieve a regular arrangement of ZrO2 particles to overcome the relatively weak van der Waals interactions between the inorganic materials. A Langmuir monolayer of these particles is extremely condensed. A multi-particle layered structure is constructed by the Langmuir–Blodgett (LB) technique. The out-of-plane XRD measurement of multilayered organization of stearate-modified ZrO2 particles confirms sharp peaks at 59Å. AFM images on a mesoscopic scale of this single-particle layer of stearate-modified ZrO2 show the aggregate of particles with 50nm diameters. However, fine particles with ∼5nm diameter are confirmed from high-resolution AFM observations in the case of single-particle layers transferred at low surface pressure. That is to say, a regular periodic structure along the c-axis and a hierarchical aggregated particle form are fabricated by Langmuir and LB techniques.

Monolayer Behavior of Binary Systems of Lactonic and Acidic Forms of Sophorolipids: Thermodynamic Analyses of Langmuir Monolayers and AFM Study of Langmuir^|^ndash;Blodgett Monolayers

The synergic effect and miscibility of the lactonic and acidic forms of sophorolipids (SLs) produced by Starmerella bombicola NBRC 10243 were first evaluated through atomic force microscopy (AFM), together with the Langmuir monolayer technique. The π-A isotherm of a pure lactonic sophorolipid (LS) monolayer mostly exhibited a liquid expanded monolayer, while that of an acidic sophorolipid (AS) monolayer showed a liquid condensed monolayer, suggesting that the lactonization of SLs makes the molecules more bulky and prevents them from adopting a close-packed arrangement. Plots of the mean area per molecule of mixed LS/AS monolayers gave positive deviations from the ideal curves, implying that the LS and AS molecules are miscible. Interestingly, the positive deviation of excess area (Aex) from ideality was most significant at a mole fraction (XAS) of 0.3, which closely resembles the composition of the LS/AS mixture secreted by S. bombicola in culture. The AFM images of mixed LS/AS monolayers transferred at 20 mN/m revealed no phase-separated microdomain structures, but rather showed small protruding objects for all compositions, indicating that LS and AS are partially miscible, as predicted by the positive deviations from the ideal curves. Cross-section analysis of the AFM images indicated that the observed protruding objects are AS-rich monolayers formed on the LS/AS monolayer. Our results clearly demonstrate that AFM combined with the Langmuir technique is useful for the exploration of the miscibility and synergic effects of microbial products.

Improved Mechanical Stability of Acetoxypropyl Cellulose upon Blending with Ultranarrow PbS Nanowires in Langmuir Monolayer Matrix

Cellulose and cellulose derivatives have long been used as membrane fabrication. Langmuir monolayer behavior, which naturally mimics membranes, of acetoxypropyl cellulose (APC) and lead sulfide (PbS) nanowire mixtures at different volume ratios is reported. Surface pressure (π)-area (A) isotherms of APC and PbS nanowires mixtures at different volume ratios show a gradual decrease in the monolayer area with increasing volume fraction of PbS nanowires. Change of surface potential with monolayer area at different volume ratios also reveals a gradual increase in the surface potential indicating incorporation of PbS nanowires within APC matrix. The compressibility and elastic constants measurements reveal an enhancement of the elasticity upon incorporation of PbS nanowires up to certain volume fractions. An enhancement in stability of the blend is observed upon PbS nanowire incorporation to the APC matrix. Rheological measurements also support the robustness of the mixture of APC and PbS nanowires in 3D bulk phase. Such robust ultrathin films of cellulose based-nanowire blend obtained by means of the Langmuir technique may lead to novel routes for designing cellulosic-based thin films and membranes.

Thermodynamic analysis and AFM study of the interaction of palmitic acid with DPPE in Langmuir monolayers

Palmitic acid (PA), which could induce the apoptosis of endothelial cells, is related to kinds of diseases. The influence of PA on 1, 2-dipalmitoyl-sn-glycero-3-phosphoethanolamine (DPPE) monolayer was studied in this paper, which is essential to understand those physiological and pathological processes. The behaviors and microstructures of monolayer of PA/DPPE binary systems on subphase of pH 6.2 and 3 were investigated by using a combination of Langmuir technique and Atomic force microscopy (AFM) in this paper. The two kinds of molecular arrangement situations were provided at different molar ratios. The thermodynamic properties of PA/DPPE mixed monolayer have been analyzed quantitatively based on the regular solution theory. The results have clearly manifested that excess molecular area (Aex), excess Gibbs energy (ΔGex) and interaction parameter (ω) of PA/DPPE mixed monolayers obtained positive deviation for XPA=0.1, 0.3, 0.5 and 0.7, suggesting an intermolecular repulsive interaction existed, the stability of the system decreased and the binary systems were metastable. The negative deviation was found at XPA=0.9, suggesting an attractive force existed between molecules and was beneficial to the pack of molecules and increasing the stability of the system as well. The value of activity coefficient f1 and f2 of mixed systems presented opposite changing trend at subphase of pH 6.2. However, the trend was similar at subphase of pH 3. These results provided some experiment evidences and theory references to cure atherosclerosis and acute coronary syndrome caused by excess free fatty acid.

Influence of the 524-VAAEIL-529 sequence of annexins A6 in their interfacial behavior and interaction with lipid monolayers

Annexin A6 (AnxA6), a calcium- and membrane-binding protein, is expressed in mammalian cells in two isoforms: AnxA6-1 and AnxA6-2, the latter lacking the 524-VAAEIL-529 sequence at the start of repeat 7. The different intracellular localization of these two isoforms suggests distinct function in membrane dynamics. The aim of this work was to analyze the behavior of AnxA6 isoforms at the air/water interface alone and in the presence of membrane mimicking lipid monolayers. Using Langmuir technique showed that AnxA6-2 was less adsorbed to the neat air–water interface than AnxA6-1 at acidic pH and minor differences in their PM-IRRAS spectra were observed. Both isoforms exhibited similar behavior towards cholesterol monolayer. However, the interactions of AnxA6-2 with cholesterol ester monolayer were most favorable compared to AnxA6-1. Our experimental data are discussed in relation with the different intracellular localization of the two isoforms and with our constructed model of AnxA6-2 with the known crystal structure of AnxA6-1 showing the persistence of the 516–529 α-helix in AnxA6-2 despite the absence of the 524-VAAEIL-529 sequence.

Miscibility and phase separation in mixed erucylphosphocholine–DPPC monolayers

Binary Langmuir monolayers composed of 1,2-dipalmitoyl-sn-phosphatidylcholine (DPPC) and erucylphosphocholine (ErPC) – a new generation anti-cancer drug of phospholipids-like structure, have been studied with classical Langmuir technique complemented with Brewster angle microscopy (BAM) and Grazing Incidence X-ray Diffraction (GIXD). In the course of surface pressure (π)–area (A) isotherms for film containing mole fraction of ErPC (XErPC) equal to 0.1, 0.2 and 0.3, two collapses appear – the value of the first one is close to the collapse pressure observed for pure ErPC, while the second one occurs at the pressure similar to that for pure DPPC. Such a behavior could imply that the investigated system is immiscible – the expulsion of a component collapsing at a lower pressure (ErPC) from mixed monolayer occurs at the first transition while DPPC (collapsing at a higher pressure value) is expelled at the final collapse. To confirm this hypothesis further analysis has been performed. Namely, thermodynamic analysis based on the calculation of the excess free enthalpy of mixing (ΔGMexc) evidenced for mutual miscibility of DPPC and ErPC in the region of surface pressures below the first collapse, which has been additionally confirmed with BAM images. On the other hand, at high surface pressures, above the first collapse (50mN/m) GIXD experiments complemented with BAM images proved phase separation of the investigated lipids and 2D crystalline domains formation. The interpretation of the X-ray scattering results enabled us to propose a possible model of molecular packing, according to which only condensed domains of the 1:1 lipids ratio are periodically ordered; whereas at the other proportions the lipids are located randomly within the domains.

Cholesterol accelerates the binding of Alzheimer's β-amyloid peptide to ganglioside GM1 through a universal hydrogen-bond-dependent sterol tuning of glycolipid conformation

Age-related alterations of membrane lipids in brain cell membranes together with high blood cholesterol are considered as major risk factors for Alzheimer's disease. Yet the molecular mechanisms by which these factors increase Alzheimer's risk are mostly unknown. In lipid raft domains of the plasma membrane, neurotoxic Alzheimer's beta-amyloid (Abeta) peptides interact with both cholesterol and ganglioside GM1. Recent data also suggested that cholesterol could stimulate the binding of Abeta to GM1 through conformational modulation of the ganglioside headgroup. Here we used a combination of physicochemical and molecular modeling approaches to decipher the mechanisms of cholesterol-assisted binding of Abeta to GM1. With the aim of decoupling the effect of cholesterol on GM1 from direct Abeta-cholesterol interactions, we designed a minimal peptide (Abeta5-16) containing the GM1-binding domain but lacking the amino acid residues involved in cholesterol recognition. Using the Langmuir technique, we showed that cholesterol (but not phosphatidylcholine or sphingomyelin) significantly accelerates the interaction of Abeta5-16 with GM1. Molecular dynamics simulations suggested that Abeta5-16 interacts with a cholesterol-stabilized dimer of GM1. The main structural effect of cholesterol is to establish a hydrogen-bond between its own OH group and the glycosidic-bond linking ceramide to the glycone part of GM1, thereby inducing a tilt in the glycolipid headgroup. This fine conformational tuning stabilizes the active conformation of the GM1 dimer whose headgroups, oriented in two opposite directions, form a chalice-shaped receptacle for Abeta. These data give new mechanistic insights into the stimulatory effect of cholesterol on Abeta/GM1 interactions. They also support the emerging concept that cholesterol is a universal modulator of protein-glycolipid interactions in the broader context of membrane recognition processes.

Simultaneously propagating voltage and pressure pulses in lipid monolayers of pork brain and synthetic lipids

Hydrated interfaces are ubiquitous in biology and appear on all length scales from ions and individual molecules to membranes and cellular networks. In vivo, they comprise a high degree of self-organization and complex entanglement, which limits their experimental accessibility by smearing out the individual phenomenology. The Langmuir technique, however, allows the examination of defined interfaces, the controllable thermodynamic state of which enables one to explore the proper state diagrams. Here we demonstrate that voltage and pressure pulses simultaneously propagate along monolayers comprised of either native pork brain or synthetic lipids. The excitation of pulses is conducted by the application of small droplets of acetic acid and monitored subsequently employing time-resolved Wilhelmy plate and Kelvin probe measurements. The isothermal state diagrams of the monolayers for both lateral pressure and surface potential are experimentally recorded, enabling us to predict dynamic voltage pulse amplitudes of 0.1-3 mV based on the assumption of static mechanoelectrical coupling. We show that the underlying physics for such propagating pulses is the same for synthetic and natural extracted (pork brain) lipids and that the measured propagation velocities and pulse amplitudes depend on the compressibility of the interface. Given the ubiquitous presence of hydrated interfaces in biology, our experimental findings seem to support a fundamentally new mechanism for the propagation of signals and communication pathways in biology (signaling), which is based neither on protein-protein or receptor-ligand interaction nor diffusion.

Interaction of resveratrol with membrane glycerophospholipids in model system in vitro

Resveratrol is a polyphenol that among other sources occurs in grapes and for this reason, red wines also contain considerable amounts of this compound. Interactions of resveratrol with pure molecular species of phosphatidylcholine (PC), phosphatidylethanoloamine (PE) and phosphatidylserine (PS) were studied with the Langmuir technique on monolayers and with differential scanning calorimetry on unilamellar liposomes. Resveratrol caused a modest increase in the mean molecular area (MMA) of dipalmitoyl-PC (DPCC) and PE (DPPE) monolayers, but profoundly increased the MMA of dipalmitoyl-PS (DPPS). The resveratrol-induced increase in MMA was enlarged in PS species containing stearolyl and oleoyl acyls suggesting that increase in the acyl chain length and unsaturation enhanced the resveratrol–phospholipid interaction. In liposomes resveratrol lowered Tm (main transition temperature) and increased TC1/2 (transition interval) in DPPE, DPPS and 1-palmitoyl-2-oleoyl-PS (POPS), suggesting that resveratrol causes an increase in the anisotropy of these liposomes. In DPPE and POPS liposomes resveratrol caused a lowering of ΔH, further substantiating a pure enhancement of anisotropy for these molecular species. However, resveratrol caused a marked increase of ΔH in DPPS liposomes, indicating that, in addition to increase anisotropy, DPPS and resveratrol attracted each other specifically. This study has clearly shown interactions between resveratrol and glycerophospholipids on a molecular level and that these interactions are influenced by the acyl chain length, degree of unsaturation and head group of the lipids.

AFM Studies on Curcumin Based Zn(II) Complex Molecules for Applications as Anticancer Agents

We present our recent atomic force microscopy studies on a curcumin based Zn(II) complex anticancer agent. We investigated the morphology of the molecule on a nanoscale level to test the effect of different substrates and solvents on the shape and the aggregation properties of the molecule. Self assembled films were produced by Langmuir technique at different surface pressures and by spin coating. Measurements showed small molecular aggregates with a height of 0,5 nm. The wetting properties of the substrate play a role in the aggregation properties and the film formation. The obtained results will help in understanding the interaction of the studied molecule with DNA or chromonic systems.

Investigation of the interactions of lupane type pentacyclic triterpenes with outer leaflet membrane phospholipids – Langmuir monolayer and synchrotron X-ray scattering study

Lupane type pentacyclic triterpenes (LTs) are pharmacologically active natural products isolated from different plants. They have broad spectrum of therapeutic action ranging from anticancer via anti-HIV, antibiotic to anti-inflammatory and anti-protozoal activity. Many scientific papers underline that the key stage in the LT mechanism of action is their incorporation into cellular membrane and the interaction with the structural lipids. In our research we apply Langmuir monolayers as a versatile platform for the investigation of these phenomena, since till now important aspects concerning this issue are incomprehensible. We focus our attention on the interactions of lupeol and betulinic acid with choline-headgroup structural lipids: a representative of saturated glycerophosphatidylcholines (DPPCs), and octadecyl-sphingomyelin, a representative of membrane sphingolipids. Application of complementary physicochemical techniques such as the Langmuir technique, Brewster angle microscopy, and grazing incidence X-ray diffraction supported by thermodynamic analysis enabled us to investigate the intermolecular interactions in such binary model systems. Our results corroborate that LT is miscible with the outer leaflet membrane phospholipids, both DPPC and SM in the whole range of mole ratios. Moreover, the introduction of LT into the phospholipid film, even in small proportion, leads to the loss of periodical ordering of the phospholipid molecules and the disappearance of the diffraction signal as observed by GIXD. Our results also proved that LT does not form any surface complexes of fixed stoichiometry resembling the well characterized lipid rafts.

Interactions between bovine serum albumin and Langmuir films composed of charged and uncharged poly(N-isopropylacrylamide) block copolymers

The thermoresponsive poly(N-isopropylacrylamide) (PNIPAAM) and NIPAAM block copolymer derivatives are attractive for drug delivery applications as they contract reversibly at lower critical solution temperatures (LCST) close to physiological conditions. In order to investigate biomaterial-protein compatibility, we have studied the interaction between PNIPAAM copolymer films spread at the air–water surface and bovine serum albumin (BSA) injected below the precompressed polymer films, using the Langmuir technique coupled with Brewster angle microscopy (BAM). A PNIPAAM homopolymer was applied together with a number of PNIPAAM-based di- and triblock copolymers, to assess effects of e.g., charge and hydrophobicity on protein–polymer interactions. The nature and strength of protein–polymer interaction was found to be tunable, ranging from complex formation (PNIPAAM homopolymer) to mixed monolayers and electrostatic cross-linking, according to the nature of the co-monomer. Results show that intercalation versus adsorption can be controlled through polymer composition.

Molecular Rearrangement in a Zinc Stearate Langmuir Film Dependent on a Film Preparation Method Studied Using Polarization-Modulation Infrared Reflection Absorption Spectroscopy and X-ray Absorption Fine Structure

Molecular dynamic rearrangement in a Langmuir (L) film of zinc stearate in a cross section image has been analyzed by employing polarization modulation infrared reflection spectrometry (PM-IRRAS) and X-ray absorption fine structure (XAFS). The number of coordination on a zinc cation is evaluated by XAFS, and the coordination structure of the carboxylic group is revealed by the IR analysis. An L film prepared at a fixed area without using film-compression bar exhibits time-dependent change of the coordination structure about a zinc cation, and it attains a highly ordered coordination structure in the carboxylic group region. However, an L film prepared by the Langmuir technique using a compression bar exhibited ignorable spectral changes in both IR and X-ray analyses. The hydrocarbon chains in the compressed L film are better uniformed than the uncompressed film, but the uniformed molecular arrangement of the carboxylic group is restricted to be rearranged, and it does not attain a stable structure in terms of coordination on the zinc cation.

Lupane-Type Pentacyclic Triterpenes in Langmuir Monolayers: A Synchrotron Radiation Scattering Study

Lupane-type pentacyclic triterpenes (lupeol, betulin, and betulinic acid) are natural products isolated from various plant sources. The terpenes exhibit a vast spectrum of biological activity and are applied in therapies for different diseases, among which the anticancer, anti-HIV, antihypercholesteremic, and antiinflammatory are the most promising. These chemicals possess amphiphilic structure and were proved to interact strongly with biomembranes, which can be the key stage in their mechanism of action. In our studies, we applied Langmuir monolayers as versatile models of biomembranes. It turned out that the three investigated terpenes are capable of stable monolayer formation; however, these monolayers differ profoundly regarding their physicochemical characteristics. In our research, we applied the Langmuir technique (surface pressure-mean molecular area (π-A) isotherm registration) coupled with Brewster angle microscopy (BAM), but the main focus was on the synchrotron radiation scattering method, grazing incidence X-ray diffraction (GIXD), which provides information on the amphiphilic molecule ordering in the angström scale. It was proved that all the investigated terpenes form crystalline phases in their monolayers. In the case of lupeol, only the closely packed upright phase was observed, whereas for betulin and betulinic acid, the phase situation was more complex. Betulinic acid molecules can be organized in an upright phase, which is crystalline, and in a tilted phase, which is amorphous. The betulin film is a conglomerate of an upright crystalline monolayer phase, tilted amorphous monolayer phase, and a crystalline tilted bilayer. In our paper, we discuss the factors leading to the formation of the observed phases and the implications of our results to the therapeutic applications of the native lupane-type triterpenes.