Interfacial structurization between triolein and water from pH and buffer ions.

HYPOTHESIS: Understanding and manipulating the oil/water interface is important across various industries, including food, pharmaceuticals, cosmetics, and detergents. Many of these processes occur under elevated pH conditions in buffer systems, where base-catalyzed hydrolysis of triglyceride ester bonds leads to amphiphilic reaction products such as fatty acids. EXPERIMENTS: Here, pH-triggered alterations of the triolein/water interface are analyzed in the presence of phosphate (PB) and tris(hydroxymethyl)aminomethane (TRIS). Ellipsometry at the liquid/liquid interface, tensiometry, and scanning small angle X-ray scattering are used to study the formation of structures at the oil/water interface. Confocal Raman microscopy, nuclear magnetic resonance spectroscopy, and in silico modeling analyze compositional changes in the interfacial region. FINDINGS: pH and buffer ions were discovered to significantly modify the triglyceride/water interface, contrary to the decane/water control. Decreasing interfacial tensions from 32.4 to 2.2 mN/m upon pH increase from 6.5 to 9.5 is seen with multilamellar interfacial layers forming at pH around 9.0 in the presence of TRIS. Oleic acid from triolein hydrolysis and its further interaction with TRIS is held responsible for this. The new understanding can guide the design of pH- and ion-responsive functional materials and optimize industrial processes involving triglyceride/water interfaces.

Biocompatible Rhamnolipid Self-Assemblies with pH-Responsive Antimicrobial Activity.

There is an urgent need for alternative antimicrobial materials due to the growing challenge of bacteria becoming resistant to conventional antibiotics. This study demonstrates the creation of a biocompatible pH-switchable antimicrobial material by combining bacteria-derived rhamnolipids (RL) and food-grade glycerol monooleate (GMO). The integration of RL into dispersed GMO particles, with an inverse-type liquid crystalline cubic structure in the core, leads to colloidally stable supramolecular materials. The composition and pH-triggered structural transformations are studied with small-angle X-ray scattering, cryogenic transmission electron microscopy, and dynamic light scattering. The composition-structure-activity relationship is analyzed and optimized to target bacteria at acidic pH values of acute wounds. The new RL/GMO dispersions reduce Staphylococcus aureus (S. aureus) populations by 7-log after 24 h of treatment with 64 µg mL-1 of RL and prevent biofilm formation at pH = 5.0, but have no activity at pH = 7.0. Additionally, the system is active against methicillin-resistant S. aureus (MRSA) with minimum inhibitory concentration of 128 µg mL-1 at pH 5.0. No activity is found against several Gram-negative bacteria at pH 5.0 and 7.0. The results provide a fundamental understanding of lipid self-assembly and the design of lipid-based biomaterials, which can further guide the development of alternative bio-based solutions to combat bacteria.

Human antimicrobial peptide inactivation mechanism of enveloped viruses.

HYPOTHESIS: Enveloped viruses are pivotal in causing various illnesses, including influenza and COVID-19. The antimicrobial peptide LL-37, a critical part of the human innate immune system, exhibits potential as an antiviral agent capable of thwarting these viral threats. Its mode of action involves versatile and non-specific interactions that culminate in dismantling the viral envelope, ultimately rendering the viruses inert. However, the exact mechanism of action is not yet understood. EXPERIMENTS: Here, the mechanism of LL-37 triggered changes in the structure and function of an enveloped virus is investigated. The bacteriophage "Phi6" is used as a surrogate for pathogenic enveloped viruses. Small angle X-ray and neutron scattering combined with light scattering techniques demonstrate that LL-37 actively integrates into the virus's lipid envelope. FINDINGS: LL-37 addition to Phi6 leads to curvature modification in the lipid bilayer, ultimately separating the envelope from the nucleocapsid. Additional biological assays confirm the loss of virus infectivity in the presence of LL-37, which coincides with the structural transformations. The results provide a fundamental understanding of the structure-activity relationship related to enveloped viruses. The knowledge of peptide-virus interactions can guide the design of future peptide-based antiviral drugs and therapies.

Solvent Dependent Folding of an Amphiphilic Polyaramid.

A series of synthetic alternating and amphiphilic aromatic amide polymers were synthesized by a step growth polymerization. Alternating meta- and para-linkages were introduced to force the polymer chain into a helical shape in the highly polar solvent water. The polymers were analyzed by 1H NMR spectroscopy and SEC in polar aprotic solvents such as DMSO and DMF. However, the polymers also showed good solubility in water. 1H NMR spectroscopy, small-angle X-ray scattering, and dynamic light scattering provided clear evidence of polymer folding in water but not DMF. We employed parallel tempering metadynamics in the well-tempered ensemble (PTMetaD-WTE) to simulate the free energy surfaces of an analogous model polymer in DMF and water. The simulations gave a molecular model of an unfolded structure in DMF and a helically folded tubular structure in water.

Single particle investigation of triolein digestion using optical manipulation, polarized video microscopy, and SAXS.

HYPOTHESIS: Understanding how soft colloids, such as food emulsion droplets, transform based on their environment is critical for various applications, including drug and nutrient delivery and biotechnology. However, the mechanisms behind colloidal transformations within individual oil droplets still need to be better understood. EXPERIMENTS: This study employs optical micromanipulation with microfluidics and polarized optical video microscopy to investigate the pancreatic lipase- and pH-triggered colloidal transformations in a single triolein droplet. Small-angle X-ray scattering (SAXS) provides complementary statistical insights and allows for detailed structural assignment. FINDINGS: Optical video microscopy recorded the transformation of individual triolein emulsion droplets, with the smooth surface of these spherical particles becoming rough and the entire volume eventually being affected. The polarized microscopy revealed the coexistence of at least two distinct structures in a single particle during digestion, with their ratio and distribution altered by pH. The SAXS analysis assigned the optical anisotropy to emulsified inverse hexagonal- and multilamellar phases, coexisting with isotropic structures such as the micellar cubic phase. These results can help understand the phase transformations inside an emulsion droplet during triglyceride digestion and guide the design of advanced food emulsions.

Protein Cage-Stabilized Emulsions: Formulation and Characterization.

The formulation of Pickering emulsions using protein cages is gaining interest for applications in molecular delivery. Despite the growing interest, methods to investigate the at the liquid-liquid interface are limited. This chapter describes standard methods to formulate and protocols to characterize protein cage-stabilized emulsions. The characterization methods are dynamic light scattering (DLS), intrinsic fluorescence spectroscopy (TF), circular dichroism (CD), and small angle X-ray scattering (SAXS). Combining these methods allows understanding of the protein cage nanostructure at the oil/water interface.

Computer-Aided Drug Design and Synthesis of Rhenium Clotrimazole Antimicrobial Agents.

In the context of the global health issue caused by the growing occurrence of antimicrobial resistance (AMR), the need for novel antimicrobial agents is becoming alarming. Inorganic and organometallic complexes represent a relatively untapped source of antibiotics. Here, we report a computer-aided drug design (CADD) based on a 'scaffold-hopping' approach for the synthesis and antibacterial evaluation of fac-Re(I) tricarbonyl complexes bearing clotrimazole (ctz) as a monodentate ligand. The prepared molecules were selected following a pre-screening in silico analysis according to modification of the 2,2'-bipyridine (bpy) ligand in the coordination sphere of the complexes. CADD pointed to chiral 4,5-pinene and 5,6-pinene bipyridine derivatives as the most promising candidates. The corresponding complexes were synthesized, tested toward methicillin-sensitive and -resistant S. aureus strains, and the obtained results evaluated with regard to their binding affinity with a homology model of the S. aureus MurG enzyme. Overall, the title species revealed very similar minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) values as those of the reference compound used as the scaffold in our approach. The obtained docking scores advocate the viability of 'scaffold-hopping' for de novo design, a potential strategy for more cost- and time-efficient discovery of new antibiotics.

Supramolecular design of CO2-responsive lipid nanomaterials.

HYPOTHESIS: Stimuli-responsive materials can innovate in various fields, including food and pharmaceutical sciences. Their response to a specific stimulus can be utilized to release loaded bioactive molecules or sense their presence. The biocompatibility and abundance of CO2 in the environment make it an exciting stimulus for such applications. We hypothesize the formation of CO2-responsive self-assemblies of oleyl-amidine in water. Their integration into glycerol-monooleate-based (GMO) dispersions is further thought to form CO2-switchable liquid crystalline nanoparticles. The switch from an non-charged acetamidine surfactant to its cationic amidinium form triggers curvature changes that ultimately induces phase transitions. EXPERIMENTS: The CO2-switchable lipid (E)-N,N-dimethyl-N-((Z)-octadec-9-en1-yl)acetimidamide (OAm) is synthesized and formulated into emulsions and dispersed liquid crystals with GMO. The supramolecular structure and its response to CO2 are characterized using small angle X-ray scattering, dynamic light scattering, ζ-potential measurements and cryogenic transmission electron microscopy. FINDINGS: Depending on the composition, OAm is discovered to self-assemble into a variety of CO2-responsive lyotropic liquid crystalline structures that can be dispersed in excess water. CO2-triggered colloidal transformations from unstructured OAm-in-water emulsions to direct micelles; dispersed inverse hexagonal phase to direct rod-like micelles, and sponge phase to vesicles are discovered. These structural changes are driven by the reaction of OAm's amidine headgroup with CO2. The results provide a fundamental understanding of CO2-triggered functional nanomaterials and may guide their future design into delivery platforms and biosensors.

Mesenchymal Stem Cells Sense the Toughness of Nanomaterials and Interfaces.

Stem cells are known to sense and respond to the mechanical properties of biomaterials. In turn, cells exert forces on their environment that can lead to striking changes in shape, size and contraction of associated tissues, and may result in mechanical disruption and functional failure. However, no study has so far correlated stem cell phenotype and biomaterials toughness. Indeed, disentangling toughness-mediated cell response from other mechanosensing processes has remained elusive as it is particularly challenging to uncouple Youngs' or shear moduli from toughness, within a range relevant to cell-generated forces. In this report, it is shown how the design of the macromolecular architecture of polymer nanosheets regulates interfacial toughness, independently of interfacial shear storage modulus, and how this controls the expansion of mesenchymal stem cells at liquid interfaces. The viscoelasticity and toughness of poly(l-lysine) nanosheets assembled at liquid-liquid interfaces is characterised via interfacial shear rheology. The local (microscale) mechanics of nanosheets are characterised via magnetic tweezer-assisted interfacial microrheology and the thickness of these assemblies is determined from in situ ellipsometry. Finally, the response of mesenchymal stem cells to adhesion and culture at corresponding interfaces is investigated via immunostaining and confocal microscopy.

PEGylation of Phosphatidylglycerol/Docosahexaenoic Acid Hexosomes with d-α-Tocopheryl Succinate Poly(ethylene glycol)2000 Induces Morphological Transformation into Vesicles with Prolonged Circulation Times.

Considering the broad therapeutic potential of omega-3 polyunsaturated fatty acids such as docosahexaenoic acid (DHA), here we study the effect of PEGylation of DHA-incorporated hexosomes on their physicochemical characteristics and biodistribution following intravenous injection into mice. Hexosomes were formed from phosphatidylglycerol and DHA with a weight ratio of 3:2. PEGylation was achieved through the incorporation of either d-α-tocopheryl succinate poly(ethylene glycol)2000 (TPGS-mPEG2000) or 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-methoxy-poly(ethylene glycol)2000 (DSPE-mPEG2000) at a concentration of 1.5 wt %. Nanoparticle tracking analysis, synchrotron small-angle scattering, and cryo-transmission electron microscopy were employed to characterize the nanodispersions. The results show that PEGylated lipids induce a structural transition from an inverse hexagonal (H2) phase inside the nanoparticles (hexosomes) to a lamellar (Lα) phase (vesicles). We also followed the effect of mouse plasma on the nanodispersion size distribution, number, and morphology because changes brought by plasma constituents could regulate the in vivo performance of intravenously injected nanodispersions. For comparative biodistribution studies, fluorescently labeled nanodispersions of equivalent quantum yields were injected intravenously into healthy mice. TPGS-mPEG2000-induced vesicles were most effective in avoiding hepatosplenic clearance at early time points. In an orthotopic xenograft murine model of glioblastoma, TPGS-mPEG2000-induced vesicles also showed improved localization to the brain compared with native hexosomes. We discuss these observations and their implications for the future design of injectable lyotropic nonlamellar liquid crystalline drug delivery nanosystems for therapeutic interventions of brain and liver diseases.

Optical tweezer platform for the characterization of pH-triggered colloidal transformations in the oleic acid/water system.

HYPOTHESIS: Soft colloidal particles that respond to their environment have innovative potential for many fields ranging from food and health to biotechnology and oil recovery. The in situ characterisation of colloidal transformations that triggers the functional response remain a challenge. EXPERIMENTS: This study demonstrates the combination of an optical micromanipulation platform, polarized optical video microscopy and microfluidics in a comprehensive approach for the analysis of pH-driven structural transformations in emulsions. The new platform, together with synchrotron small angle X-ray scattering, was then applied to research the food-relevant, pH-responsive, oleic acid in water system. FINDINGS: The experiments demonstrate structural transformations in individual oleic acid particles from micron-sized onion-type multilamellar oleic acid vesicles at pH 8.6, to nanostructured emulsions at pH < 8.0, and eventually oil droplets at pH < 6.5. The smooth particle-water interface of the onion-type vesicles at pH 8.6 was transformed into a rough particle surface at pH below 7.5. The pH-triggered changes of the interfacial tension at the droplet-water interface together with mass transport owing to structural transformations induced a self-propelled motion of the particle. The results of this study contribute to the fundamental understanding of the structure-property relationship in pH-responsive emulsions for nutrient and drug delivery applications.

Donor-Acceptor Stenhouse Adduct-Polydimethylsiloxane-Conjugates for Enhanced Photoswitching in Bulk Polymers.

Donor-acceptor Stenhouse adducts (DASAs) are a rapidly emerging class of visible light-activated photochromes and DASA-functionalized polymers hold great promise as biocompatible photoresponsive materials. However, the photoswitching performance of DASAs in solid polymer matrices is often low, particularly in materials below their glass transition temperature. To overcome this limitation, DASAs are conjugated to polydimethylsiloxanes which have a glass transition temperature far below room temperature and which can create a mobile molecular environment around the DASAs for achieving more solution-like photoswitching kinetics in bulk polymers. The dispersion of DASAs conjugated to such flexible oligomers into solid polymer matrices allows for more effective and tunable DASA photoswitching in stiff polymers, such as poly(methyl methacrylate), without requiring modifications of the matrix. The photoswitching of conjugates with varying polymer molecular weight, linker type, and architecture is characterized via time-dependent UV-vis spectroscopy in organic solvents and blended into polymethacrylate films. In addition, DASA-functionalized polydimethylsiloxane networks, accessible via the same synthetic route, provide an alternative solution for achieving fast and efficient DASA photoswitching in the bulk owing to their intrinsic softness and flexibility. These findings may contribute to the development of DASA-functionalized materials with better tunable, more effective, and more reversible modulation of their optical properties.

Improved Purification of Human Granzyme A/B and Granulysin Using a Mammalian Expression System.

Cytotoxic lymphocytes release proteins contained within the cytoplasmic cytolytic granules after recognition of infected or tumor target cells. These cytotoxic granular proteins (namely granzymes, granulysin, and perforin) are key immunological mediators within human cellular immunity. The availability of highly purified cytotoxic proteins has been fundamental for understanding their function in immunity and mechanistic involvement in sepsis and autoimmunity. Methods for recovery of native cytotoxic proteins can be problematic leading to: 1) the co-purification of additional proteins, confounding interpretation of function, and 2) low yields of highly purified proteins. Recombinant protein expression of individual cytolytic components can overcome these challenges. The use of mammalian expression systems is preferred for optimal post-translational modifications and avoidance of endotoxin contamination. Some of these proteins have been proposed for host directed human therapies (e.g. - granzyme A), or treatment of systemic infections or tumors as in granulysin. We report here a novel expression system using HEK293T cells for cost-effective purification of high yields of human granzymes (granzyme A and granzyme B) and granulysin with enhanced biological activity than previous reports. The resulting proteins are free of native contaminants, fold correctly, and remain enzymatically active. Importantly, these improvements have also led to the first purification of biologically active recombinant human granulysin in high yields from a mammalian system. This method can be used as a template for purification of many other secreted cellular proteins and may lead to advances for human medicine.

(Anti)viral Material Design Guided by Scattering Methods.

Viruses are nature's own nanoparticles that are highly symmetric and monodisperse in size and shape with well-defined surface chemistry. They have evolved for optimal cell interactions, genetic information delivery and replication by the host cell over millions of years. These features render them into very efficient pathogens that place a severe burden onto the health of our society. At the same time, they are highly interesting objects for colloidal studies and building blocks for advanced bio-inspired materials for health applications. Their characterisation requires sophisticated experimental techniques such as scattering of X-rays, neutrons, and light to probe structures and interactions from the nanometre to the micrometre length-scale in solution. This contribution summarizes the recent progress in the field of virus self-assembly and virus-based biopolymer composites for advanced material design. It discusses the advances and highlights some of the challenges in the characterization of structure and dynamics in these materials with a focus on scattering techniques. It further demonstrates selected applications in the field of food and water purification.

Cell medium-dependent dynamic modulation of size and structural transformations of binary phospholipid/ω-3 fatty acid liquid crystalline nano-self-assemblies: Implications in interpretation of cell uptake studies.

Lyotropic non-lamellar liquid crystalline (LLC) nanoparticles, with their tunable structural features and capability of loading a wide range of drugs and reporter probes, are emerging as versatile injectable nanopharmaceuticals. Secondary emulsifiers, such as Pluronic block copolymers, are commonly used for colloidal stabilization of LLC nanoparticles, but their inclusion often compromises the biological safety (e.g., poor hemocompatibility and enhanced cytotoxicity) of the formulation. Here, we introduce a library of colloidally stable, structurally tunable, and pH-responsive lamellar and non-lamellar liquid crystalline nanoparticles from binary mixtures of a phospholipid (phosphatidylglycerol) and three types of omega-3 fatty acids (ω-3 PUFAs), prepared in the absence of a secondary emulsifier and organic solvents. We study formulation size distribution, morphological heterogeneity, and the arrangement of their internal self-assembled architectures by nanoparticle tracking analysis, synchrotron small-angle X-ray scattering, and cryo-transmission electron microscopy. The results show the influence of type and concentration of ω-3 PUFAs in nanoparticle structural transitions spanning from a lamellar (Lα) phase to inverse discontinuous (micellar) cubic Fd3m and hexagonal phase (H2) phases, respectively. We further report on cell-culture medium-dependent dynamic fluctuations in nanoparticle size, number and morphology, and simultaneously monitor uptake kinetics in two human cell lines. We discuss the role of these multiparametric biophysical transformations on nanoparticle-cell interaction kinetics and internalization mechanisms. Collectively, our findings contribute to the understanding of fundamental steps that are imperative for improved engineering of LLC nanoparticles with necessary attributes for pharmaceutical development.

Human Antimicrobial Peptide Triggered Colloidal Transformations in Bacteria Membrane Lipopolysaccharides.

Growing concerns of bacterial resistance against conventional antibiotics shifts the research focus toward antimicrobial peptide (AMP)-based materials. Most AMPs kill gram-negative bacteria by destroying their inner membrane, but have to first pass the outer membrane covered with lipopolysaccharides (LPS). Their interplay with the LPS is crucial for bactericidal activity, but is yet to be elucidated in detail. In this study, self-assemblies of Escherichia coli LPS with the human cathelicidin AMP LL-37, free and encapsulated into glyceryl monooleate (GMO) lipid nanoparticles, are analyzed using synchrotron small angle X-ray scattering, dynamic light scattering, and cryogenic transmission electron microscopy. Circular dichroism spectroscopy is used to study modifications in LL-37's secondary structure. LPS is found to form elongated micelles and the addition of LL-37 induces their transformation to multilamellar structures. LPS' addition to GMO cubosomes triggers the swelling of the internal cubic structure, while in multilamellar GMO/LL-37 nanocarriers it causes transitions into unstructured particles. The insights on the interactions among LPS and LL-37, in its free form or encapsulated in GMO dispersions, may guide the design of LPS-responsive antimicrobial nanocarriers. The findings may further assist the formulation of antimicrobial nanomaterials with enhanced penetration of LPS layers for improved destruction of bacterial membranes.

Multifunctional Anthracene-Based Ni-MOF with Encapsulated Fullerenes: Polarized Fluorescence Emission and Selective Separation of C70 from C60.

We report an anthracene-based Ni-MOF [Ni(II) metal-organic framework, {[Ni(µ2-L)2Cl2]·x(C6H6)·y(MeOH)}n (1), L = anthracene-9,10-diylbis(methylene)diisonicotinate] whose crystal structure reveals the presence of hexagonal channels with a pore size of 1.4 nm that can accommodate guests such as C60 and C70. Both confocal fluorescence and Raman microscopy results are in agreement with a homogeneous distribution of fullerenes throughout the single crystals of 1. Efficient energy transfer from 1 to the fullerenes was observed, which emitted partially polarized fluorescence emission. Stronger binding between 1 and C70 versus C60 was confirmed from HPLC analysis of the dissolved material and provides a basis for the selective retention of C70 in liquid chromatography columns packed with 1.

Ethanol Inactivation of Enveloped Viruses: Structural and Surface Chemistry Insights into Phi6.

Lipid-enveloped viruses, such as Ebola, influenza, or coronaviruses, are a major threat to human health. Ethanol is an efficient disinfectant that is widely used to inactivate these viruses and prevent their transmission. However, the interactions between ethanol and enveloped viruses leading to their inactivation are not yet fully understood. This study demonstrates the link between ethanol-induced viral inactivation and the nanostructural and chemical transformations of the model virus Phi6, an 85 nm diameter lipid-enveloped bacterial virus that is commonly used as surrogate for human pathogenic viruses. The virus morphology was investigated using small-angle X-ray scattering and dynamic light scattering and was related to its infectivity. The Phi6's surface chemistry was characterized by cryogenic X-ray photoelectron spectroscopy, and the modifications in protein structure were assessed by circular dichroism and fluorescence spectroscopy. Ethanol-triggered structural modifications were found in the lipid envelope, detaching from the protein capsid and forming coexisting nanostructures.

pH-responsive aminolipid nanocarriers for antimicrobial peptide delivery.

HYPOTHESIS: pH-responsive aminolipid self-assemblies are promising platforms for the targeted delivery of antimicrobial peptides (AMPs), with the potential to improve their therapeutic efficiency and physico-chemical stability. EXPERIMENTS: pH-sensitive nanocarriers based on dispersed self-assemblies of 1,2-dioleoyl-3-dimethylammonium-propane (DODAP) with the human cathelicidin LL-37 in excess water were characterized at different pH values using small-angle X-ray scattering, cryogenic transmission electron microscopy, and dynamic light scattering. Fluorescence and electrophoretic mobility measurements were used to probe the encapsulation efficiency of LL-37 and the nanocarriers' surface potential. FINDINGS: Upon decreasing pH in the DODAP/water systems, normal oil-in-water emulsions at pH ≥ 5.0 transitioned to emulsions encapsulating inverse hexagonal and cubic structures at pH between 4.5 and 4.0, and mostly positively-charged vesicles at pH < 4.0. These colloidal transformations are driven by the protonation of DODAP upon pH decrease. The larger lipid-water interfacial area provided by the DODAP self-assemblies at pH ≤ 4.5 allowed for an adequate encapsulation efficiency of LL-37, favouring the formation of vesicles in a concentration-dependent manner. Contrary, LL-37 was found to dissociate from the emulsion droplets at pH 6.0. The knowledge on the pH-triggered self-assembly of LL-37 and DODAP, combined with the results on peptide release from the structures contribute to the fundamental understanding of lipid/peptide self-assembly. The results can guide the rational design of future pH-responsive AMP delivery systems.

Applying Cryo-X-ray Photoelectron Spectroscopy to Study the Surface Chemical Composition of Fungi and Viruses.

Interaction between microorganisms and their surroundings are generally mediated via the cell wall or cell envelope. An understanding of the overall chemical composition of these surface layers may give clues on how these interactions occur and suggest mechanisms to manipulate them. This knowledge is key, for instance, in research aiming to reduce colonization of medical devices and device-related infections from different types of microorganisms. In this context, X-ray photoelectron spectroscopy (XPS) is a powerful technique as its analysis depth below 10 nm enables studies of the outermost surface structures of microorganism. Of specific interest for the study of biological systems is cryogenic XPS (cryo-XPS). This technique allows studies of intact fast-frozen hydrated samples without the need for pre-treatment procedures that may cause the cell structure to collapse or change due to the loss of water. Previously, cryo-XPS has been applied to study bacterial and algal surfaces with respect to their composition of lipids, polysaccharides and peptide (protein and/or peptidoglycan). This contribution focuses onto two other groups of microorganisms with widely different architecture and modes of life, namely fungi and viruses. It evaluates to what extent existing models for data treatment of XPS spectra can be applied to understand the chemical composition of their very different surface layers. XPS data from model organisms as well as reference substances representing specific building blocks of their surface were collected and are presented. These results aims to guide future analysis of the surface chemical composition of biological systems.