Biomolecules In Aqueous Solution Research Articles

Viral gene detection based on multifunctional microspheres and AgNPs coated substrate interface area SERS scheme

Surface-enhanced Raman spectroscopy (SERS) has emerged as an effective method for detecting viral genes in aqueous solutions. However, SERS has a low efficiency and poor reproducibility in multi-component aqueous solutions. Thus, developing a SERS scheme with stable hotspots generation, significant enhancement effects, and efficient viral gene enrichment remains a challenge. This study sought to proposes an enhanced hotspot generation scheme combining multifunctional microspheres and SERS substrates, which can achieve stable enhancement in the interface area. Multifunctional microspheres can be used to selectively enrich target biomolecules in aqueous solutions. Furthermore, simulation results show that these microspheres can synergistically form a stable enhancement area with SERS substrates. Finally, this scheme was used for immunocapture and Raman detection of hemagglutinin 7 (H7) and neuraminidase 9 (N9) specific gene in an aqueous solution containing multiple types of influenza viral genes. The relationship between the characteristic peaks and concentrations was analysed using two-dimensional correlation analysis. Spectral data analysis revealed an exceedingly high linear correlation between the selected characteristic peak intensity and concentrations. The calculation results show that the SERS enhancement factor of the proposed scheme for the H7 and N9 specific gene are 9.23×107 and 1.59×108, respectively. Notably, the detection limit of this scheme was 1 pM. Therefore, this scheme holds the promise to realize the detection of viral genes in heterogeneous, complex, and dynamic aqueous biological samples.

Graphene exfoliation using multidomain peptides.

Liquid-phase exfoliation using biomolecules in aqueous solution is a promising approach to obtain high quality 2D nanosheets. For example, the well-studied graphene-binding peptide, P1 (sequence HSSYWYAFNNKT), has been previously investigated and shown to have a good ability to exfoliate graphene sheets in aqueous conditions under sonication, maintaining colloidal stability. Building on this, the biomolecular exfoliant and assembly motif (BEAM) peptide, that features a graphene-binding domain at one end and a hexagonal boron nitride (h-BN) binding domain at the other, separated by a 10-carbon fatty acid chain in the centre, is shown to exfoliate graphene sheets from bulk graphite in aqueous media. An in-depth examination of the ability of the BEAM to both facilitate sheet exfoliation under sonication conditions and also maintain colliodal stability is provided through molecular dynamics simulations. These findings open new possibilities for designing multi-functional molecules that can both exfoliate and organise 2D materials into heterostructures under ambient conditions in aqueous media.

Flexible Specific Determination of Glucose in Solution, Blood Serum, and Sweat Using a Terahertz Hydrogel‐Functionalized Metamaterial

Terahertz metamaterials have shown promises in biomolecules detection, however, aqueous solvents such as blood serum strongly absorb the terahertz waves, interfering with the detection. To avoid such interference, an ultrathin, flexible, biogel‐based metamaterial that can specifically detect the transmission of terahertz waves through biomolecules in aqueous solution is proposed and demonstrated. The glucose in water can be detected with a sensitivity of 0.0446 dL mg−1 and a detection limit of 1.64 mg dL−1, as well as glucose in human serum. The metamaterial that detects glucose in human sweat is part of a wearable device. The biosensor functions well for more than 100 detection cycles. The results suggest a strategy for real‐time detection of glucose and potentially other biomolecules.

Thermodynamic study of calcium d-pantothenate drug in the presence of aqueous solutions including some amino acids: Volumetric and compressibility properties

The impressive and remarkable efforts have been made over the past years to improve the production, formulation and purification of active ingredients pharmaceutical which leads to the discovery of novel drug preparations and delivery systems. The optimal design of such processes requires a molecular-level understanding of the interactions between drug molecules and biological membranes. The thermodynamic investigation provides deep and complete knowledge of interactions and the choice of appropriate and suitable compounds in pharmaceutical fields. Particularly, the analysis of drugs + biomolecules in aqueous solutions is the central issue in many types of research because they exert their impact by interacting with biological membranes. This work is aimed to measure the density and speed of sound for the calcium d-pantothenate, vitamin B5 calcium salt (CDP) in aqueous solutions of amino acids (glycine, l-alanine, and l-leucine) at temperature ranges (293.15 to 308.15) K. By correlating of the measured physical quantities in with standard relations, apparent molar volumes, Vϕ,m, and apparent molar isentropic compression, κs,ϕ,m, are calculated. In addition, apparent molar isobaric expansion, Eϕ,m0, and Hepler’s constant are computed. For fitting the experimental Vϕ,m and κϕ the Redlich-Meyer equation was employed that the important quantities; standard partial molar volume, Vϕ0 , and partial molar isentropic compression, κϕ0, were obtained.

The Ion-Dipole Correction of the 3DRISM Solvation Model to Accurately Compute Water Distributions around Negatively Charged Biomolecules.

The 3D reference interaction site model (3DRISM) provides an efficient grid-based solvation model to compute the structural and thermodynamic properties of biomolecules in aqueous solutions. However, it remains challenging for existing 3DRISM methods to correctly predict water distributions around negatively charged solute molecules. In this paper, we first show that this challenge is mainly due to the orientation of water molecules in the first solvation shell of the negatively charged solute molecules. To properly consider this orientational preference, position-dependent two-body intramolecular correlations of solvent need to be included in the 3DRISM theory, but direct evaluations of these position-dependent two-body intramolecular correlations remain numerically intractable. To address this challenge, we introduce the Ion-Dipole Correction (IDC) to the 3DRISM theory, in which we incorporate the orientation preference of water molecules via an additional solute-solvent interaction term (i.e., the ion-dipole interaction) while keeping the formulism of the 3DRISM equation unchanged. We prove that this newly introduced IDC term is equivalent to an effective direct correlation function which can effectively consider the orientation effect that arises from position dependent two-body correlations. We first quantitatively validate our 3DRISM-IDC theory combined with the PSE3 closure on Cl-, [ClO]- (a two-site anion), and [NO2]- (a three-site anion). For all three anions, we show that our 3DRISM-IDC theory significantly outperforms the 3DRISM theory in accurately predicting the solvation structures in comparison to MD simulations, including RDFs and 3D water distributions. Furthermore, we have also demonstrated that the 3DRISM-IDC can improve the accuracy of hydration free-energy calculation for Cl-. We further demonstrate that our 3DRISM-IDC theory yields significant improvements over the 3DRISM theory when applied to compute the solvation structures for various negatively charged solute molecules, including adenosine triphosphate (ATP), a short peptide containing 19 residues, a DNA hairpin containing 24 nucleotides, and a riboswitch RNA molecule with 77 nucleotides. We expect that our 3DRISM-IDC-PSE3 solvation model holds great promise to be widely applied to study solvation properties for nucleic acids and other biomolecules containing negatively charged functional groups.

Study on the interactions of tetradecylpyridinium dl-mandelic acid and cetylpyridinium dl-mandelic acid with some small biomolecules in aqueous solution

Interactions of two active pharmaceutical ingredient ionic liquids (API-ILs) tetradecylpyridinium dl-mandelic acid ([TetPy][MAN]) and cetylpyridinium dl-mandelic acid ([CetPy][MAN]) with some small biomolecules have been studied by using conductometric, density and UV–vis spectroscopic techniques. The chosen small biomolecules are glycine, l-alanine, l-valine, l-leucine and glycylglycine (AAGG). The critical micelle concentration (cmc) of [TetPy][MAN] and [CetPy][MAN] in aqueous AAGG solution is calculated from the measured conductance values, thus yielding some thermodynamic parameters of the micellization, such as the standard Gibbs energy change (ΔGm0), the standard enthalpy change (ΔHm0) and the standard entropy change (ΔSm0). On the basis of the experimental densities of ternary solutions of (AAGG + API-ILs + water), apparent molar volume at infinite dilution (V2,φ0), apparent molar expansibility at infinite dilution (Eφ0), transfer partial molar volume (ΔtV0) and hydration number (nH) of AAGG in aqueous API-ILs solution were calculated. The dominant hydrophobic-hydrophilic and hydrophobic-hydrophobic interactions were involved in the investigated systems. The binding constant (Kb) between AAGG and API-ILs has been calculated from UV–vis spectroscopic data. Effect of some factors including concentration, temperature, molecular structure, etc. on the aforementioned parameters and the existing solvent–solute interaction has been discussed.

Terahertz transmission characteristics of xanthan gum colloids in a microfluidic chip

Terahertz time-domain spectroscopy (THz-TDS) technology has great application prospects for the detection of biological macromolecules, whereas microfluidic technology uses micro-channel networks to manipulate liquids, which can reduce the influence of hydrogen bonds in liquids on THz waves. The combination of these two technologies makes the detection of THz waves in aqueous solutions of biomolecules more accurate and efficient. In this study, microfluidic technology and THz-TDS technology were applied to study the THz transmission characteristics of xanthan gum, providing three feasible methods for the detection of xanthan gum colloids.

Ultrasensitive Mid-Infrared Biosensing in Aqueous Solutions with Graphene Plasmons.

Identifying nanoscale biomolecules in aqueous solutions by Fourier transform infrared spectroscopy (FTIR) provides an in situ and noninvasive method for exploring the structure, reactions, and transport of biologically active molecules. However, this remains a challenge due to the strong and broad IR absorption of water which overwhelms the respective vibrational fingerprints of the biomolecules. In this work, a tunable IR transparent microfluidic system with graphene plasmons is exploited to identify ≈2nm-thick proteins in physiological conditions. The acquired in situ tunability makes it possible to eliminate the IR absorption of water outside the graphene plasmonic hotspots by background subtraction. Most importantly, the ultrahigh confinement of graphene plasmons (confined to ≈15nm) permits the implementation of nanoscale sensitivity. Then, the deuterium effects on monolayer proteins are characterized within an aqueous solution. The tunable graphene-plasmon-enhanced FTIR technology provides a novel platform for studying biological processes in an aqueous solution at the nanoscale.

Aggregation Properties of Albumin in Interacting with Magnetic Fluids.

In this study, interactions of Fe3O4 magnetic nanoparticles with serum albumin biomolecules in aqueous solutions were considered. The studies were conducted with the laser correlation spectroscopy and optical analysis of dehydrated films. It was shown that the addition of magnetite to an albumin solution at low concentrations of up to 10−6 g/L led to the formation of aggregates with sizes of up to 300 nm in the liquid phase and an increase in the number of spiral structures in the dehydrated films, which indicated an increase in their stability. With a further increase in the magnetite concentration in the solution (from 10−4 g/L), the magnetic particles stuck together and to albumin, thus forming aggregates with sizes larger than 1000 nm. At the same time, the formation of morphological structures in molecular films was disturbed, and a characteristic decrease in their stability occurred. Most stable films were formed at low concentrations of magnetic nanoparticles (less than 10−4 g/L) when small albumin–magnetic nanoparticle aggregates were formed. These results are important for characterizing the interaction processes of biomolecules with magnetic nanoparticles and can be useful for predicting the stability of biomolecular films with the inclusion of magnetite particles.

QSPR model for estimation of photodegradation average rate of the porphyrin-TiO2 complexes and prediction of their biodegradation activity and toxicity: Engineering of two annihilators for water/waste contaminants

4-Nitrophenol (4-NP) and 4-chlorophenol (4-CP) are classified as emerging contaminants and essential environmentally harmful waste. Due to the water solubility of the mentioned hazardous pollutants, experimental researches are widely discussed. A novel quantitative structure-property relationships (QSPR) model is presented to predict the photodegradation average rate of 4-NP and 4-CP (r͞4-NP/4-CP) by biomolecules in aqueous solutions. This QSPR model can predict the r͞4-NP/4-CP values for porphyrin-TiO2 complexes only from their molecular structures by calculation of metal interaction with ligand and fragment effects. For deriving the new model, the measured values of r͞4-NP/4-CP of 70 porphyrin complexes were collected from various references. The values of the regression coefficient (R2) for both training (56 compounds) and test (14 compounds) sets are 0.999. The calculated values of the mean squared error (MSE) for training and test sets are 0.157 and 0.053, respectively. The biodegradation activities of porphyrins are anticipated through the quantitative structure-activity relationships (QSAR) method to reveal their relevance with simple structural descriptors. All of the photocatalysts were investigated for designing novel complexes with higher r͞4-NP/4-CP and the lower ecotoxicological effect that was satisfying.

Stretching the resolution limit of atomic force microscopy.

Atomic force microscopy (AFM) is unique in visualizing functional biomolecules in aqueous solution at ~1 nm resolution. By borrowing localization methods from fluorescence microscopy, AFM has been shown to discern structural domains that may be separated by only a few Ångströms.

Investigations of solute–solvent interactions in aqueous solutions of amino acids ionic liquids having the common nitrate as anion at different temperatures

Amino acids ionic liquids (AAILs) have huge potential applications in the green chemistry, as these are synthesized from the naturally occurring 20 amino acids and having strong hydrogen bonding ability, hydrophobic tuning, biodegradability, low-toxicity. These specialities are useful for the solubilization and stabilization of biomolecules in aqueous solutions and biofluids. We realize that the buffering capacity of such ionic salts in biofluids may cause some–sort of control in biological processes. It is expected that similar to carbonate–bicarbonate equilibria, the interaction of amino acids may be contributing to ionic interactions. In the present work, we report the synthesis of amino acids–based nitrate protic ionic liquids involving an infrequently used cations amino acids such as glycine ([HOOC–CH2–NH3]+) and ւ–alanine ([HOOC–(CH3)CH–NH3]+) with common inorganic planner anion as nitrate ([NO3]−). To understand the ionic and molecular interactions between the new class of solvents so called “bio–ionic liquids” such as glycine nitrate ([HOOC–CH2–NH3]+ [NO3]−) and ւ–alanine nitrate ([HOOC–(CH3)CH–NH3]+ [NO3]−) in aqueous solutions, the density property has been measured in the concentration range of 0.01974–0.52080 mol·kg−1 at different temperatures (288.15–308.15 K in steps of 5 K) and at 0.1 MPa. Further, experimental density data were used to calculate the apparent molar volume (Vϕ) of solute, partial molar volume of solute (V¯2) and partial molar volume of solvent (V¯1) as a function of concentration at 288.15–308.15 K. Using the appropriate thermodynamic equations incorporating the corrections due to Debye–Hückel limiting law, the apparent molar volume of solute at infinite dilution (Vϕ0) at all studied temperatures has been calculated along with the deviation parameters. The isobaric expansivity coefficient (α) and apparent molar expansivity (Eϕ) of solute at finite concentrations as well as limiting apparent molar expansivity (Eϕ0) of solute are also obtained at 293.15, 298.15 and 303.15 K. The results have been discussed to obtain information about the ion–solvent and ion–ion interactions along with influence of hydrophobic group on the volumetric properties.

The cation effect on the solubility of glycylglycine and N-acetylglycine in aqueous solution: Experimental and molecular dynamics studies

The specific interactions of ions with biomolecules in aqueous solutions play a very important role in the life sciences and biotechnology. This work aims to study the effect of NaCl, KCl, NH4Cl, CaCl2 or MgCl2 on the solubility of two glycine derivatives, glycylglycine (a.k.a. diglycine) and N-acetylglycine, and to understand the nature of the interactions present on these systems. Experimentally, upon increasing the concentration of the salts, the solubility of N-acetylglycine decreased while the solubility of diglycine increased, with divalent cations inducing a greater salting-in on the solubility of diglycine than monovalent cations. For diglycine, the results from molecular dynamics simulations correlate well with the salting-in effect with interactions involving the cation and the carboxylate group, while for neutral N-acetylglycine the interactions between the chloride anion and the hydrogen atom of the carboxylic acid group, and between the carbonyl group of the peptide bond and the cation, can be exploited to describe the generalized salting-out effect.

Sulfonated Pyrene as a Photoregulator for Single-Stranded DNA Looping.

Controlling the conformation and function of biomolecules through photoregulators holds great promise as a spatiotemporally controllable tool for disease control. In addition, introducing photoregulators into biomolecules has also found applications in constructing smart nanomaterials. In spite of the astonishing advances that have been made in the past few years, realizing highly controllable and efficient regulation over the conformation and function of biomolecules under physiological conditions is still challenging. Herein, sulfonated pyrene SPy was synthesized and used as a photoregulator to control the looping of single-stranded DNAs (ssDNAs) in aqueous solution. Due to its water solubility, SPy merits use in the study of biomolecules in aqueous solution. The looping of the doubly SPy-modified ssDNAs is stimulated by irradiation and regulated by SPy. Photoionization generates the radical cation of SPy (SPy.+ ). The association of SPy.+ with its neutral counterpart, SPy, gives rise to the dimer radical cation of SPy (SPy2 .+ ). During the association process, the stabilization energy released to form SPy2 .+ provides a driving force for the looping of ssDNAs. Conversely, the formed loop conformations were trapped by the formation of SPy2 .+ , and this allowed the looping dynamics to be investigated. The results reported herein suggest potential of SPy as a photoregulator for controlling the conformation and function of biomolecules under physiological conditions.

Study on Chemical Modifications of Glutathione by Cold Atmospheric Pressure Plasma (Cap) Operated in Air in the Presence of Fe(II) and Fe(III) Complexes

Cold atmospheric pressure plasma is an attractive new research area in clinical trials to treat skin diseases. However, the principles of plasma modification of biomolecules in aqueous solutions remain elusive. It is intriguing how reactive oxygen and nitrogen species (RONS) produced by plasma interact on a molecular level in a biological environment. Previously, we identified the chemical effects of dielectric barrier discharges (DBD) on the glutathione (GSH) and glutathione disulphide (GSSG) molecules as the most important redox pair in organisms responsible for detoxification of intracellular reactive species. However, in the human body there are also present redox-active metals such as iron, which is the most abundant transition metal in healthy humans. In the present study, the time-dependent chemical modifications on GSH and GSSG in the presence of iron(II) and iron(III) complexes caused by a dielectric barrier discharge (DBD) under ambient conditions were investigated by IR spectroscopy, mass spectrometry and High Performance Liquid Chromatography (HPLC). HPLC chromatograms revealed one clean peak after treatment of both GSH and GSSH with the dielectric barrier discharge (DBD) plasma, which corresponded to glutathione sulfonic acid GSO3H. The ESI-MS measurements confirmed the presence of glutathione sulfonic acid. In our experiments, involving either iron(II) or iron(III) complexes, glutathione sulfonic acid GSO3H appeared as the main oxidation product. This is in sharp contrast to GSH/GSSG treatment with DBD plasma in the absence of metal ions, which gave a wild mixture of products. Also interesting, no nitrosylation of GSH/GSSG was oberved in the presence of iron complexes, which seems to indicate a preferential oxygen activation chemistry by this transition metal ion.

Strong and Weak Hydrogen Bonds in Protein–Ligand Recognition

The hydrogen bond has justifiably been termed the ‘master key of molecular recognition’. It is an interaction that is weaker than the covalent bond and stronger than the van der Waals interaction. The ubiquity and flexibility of hydrogen bonds make them the most important physical interaction in systems of biomolecules in aqueous solution. Hydrogen bonding plays a significant role in many chemical and biological processes, including ligand binding and enzyme catalysis. In biological processes, both specificity and reversibility are important. Weaker interactions can be made and broken more easily than stronger interactions. In this context, it is of interest to assess the relative significance of strong and weak interactions in the macromolecular recognition processes. Is protein–ligand binding governed by conventional, that is, electrostatic N–H…O and O–H…O hydrogen bonds, or do weaker interactions with a greater dispersive component such as C–H…O hydrogen bonds also play a role? If so, to what extent are they significant? Most proteins, involving as they do, main chains, side chains, and differently bound forms of water, do not really have a static fixed structure, but rather have a dynamic, breathing nature. This tendency may to some extent be lessened by the ligands which are small molecules, but in the end, it is reasonable to expect that the strong and weak hydrogen bonds inside the protein and also at the protein–ligand interface will also have dynamic character; arguably, the weaker the hydrogen bond, the greater its dynamic character. These are often central to the much debated mechanisms of binding such as conformational selection and induced fit. All protein–ligand interactions must compete with interactions with water; both the protein and the ligand are solvated before complexation and lose their solvation shell on complex formation. Conversely, the entropic cost of trapping highly mobile water molecules in the binding site is large. However, in favorable cases, these losses are suitably compensated by the enthalpic gain resulting from water-mediated hydrogen bonds. In effect, the enthalpy–entropy balance is a fine one, and for a water molecule to be able to contribute to binding affinity, it has to be in a binding site that provides the maximum number of hydrogen-bond partners at the optimum distance and orientation. In summary, hydrogen bonds are crucial to the recognition of ligands by proteins. Integration of knowledge gained from more high-quality protein–ligand structures into theoretical and computational molecular models will be an exciting challenge in the coming years.

Ultrafast structural rearrangement dynamics induced by the photodetachment of phenoxide in aqueous solution

The elementary processes that accompany the interaction of ionizing radiation with biologically relevant molecules are of fundamental importance. However, the ultrafast structural rearrangement dynamics induced by the ionization of biomolecules in aqueous solution remain hitherto unknown. Here, we employ femtosecond optical pump-probe spectroscopy to elucidate the vibrational wave packet dynamics that follow the photodetachment of phenoxide, a structural mimic of tyrosine, in aqueous solution. Photodetachment of phenoxide leads to wave packet dynamics of the phenoxyl radical along 12 different vibrational modes. Eight of the modes are totally symmetric and support structural rearrangement upon electron ejection. Comparison to a previous photodetachment study of phenoxide in the gas phase reveals the important role played by the solvent environment in driving ultrafast structural reorganization induced by ionizing radiation. This work provides insight into the ultrafast molecular dynamics that follow the interaction of ionizing radiation with molecules in aqueous solution.

Interaction Between an Active Pharmaceutical Ingredient Ionic Liquid Benzalkonium Salicylate and Small Biomolecules in Aqueous Solution: UV Absorption, Conductivity, and Volumetric Study

UV absorption spectroscopy, electrical conductivity and density experiments have been used to investigate the interactions of some small biomolecules (amino acids/dipeptides) with an active pharmaceutical ingredient in ionic liquid form (API-IL), benzalkonium salicylate (BaSal), in aqueous solution. A number of useful parameters, such as critical micellar concentration (cmc), aggregation number (Nagg) and limiting molar conductivity (Λ0) of BaSal, standard partial molar volumes (\(V_{2,\phi }^{ \circ }\)), corresponding volumes of transfer from water to aqueous BaSal solutions (ΔtrVo), standard partial molar expansibilities (\(E_{\phi }^{ \circ }\)), hydration number (nH) of small biomolecules, as well as the binding constants (Kb) for small biomolecule–BaSal complexes have been evaluated. The dependence of the properties on concentration, temperature and alkyl chain length of amino acids/dipeptides is examined. The results are used to identify the solute–solvent physicochemical interactions occurring in the studied systems.

Cold atmospheric Plasma Jet-Generated Oxidized Derivatives of Tryptophan and Their Selective Effects on Murine Melanoma and Fibroblast Cells

Cold atmospheric pressure plasma is widely used in research for biomedical applications and is a promising therapy to selectively eradicate cancer cells. However, fundamental information related to the plasma modification of biomolecules in aqueous solution remains elusive. In this work, we studied the chemical basis of tryptophan (Trp) oxidation using an Ar plasma jet and investigated the effects of plasma-treated tryptophan (PTT) on cell viability. The results show that the main product of the Trp of plasma induced oxidation is a mixture of hydroxyl derivatives and hydroperoxides in aqueous form. The products result primarily from the hydroxyl radical (·OH) attacking the Trp, which can be explained by the interaction with both the aromatic and the pyrrole rings. We observed that the PTT has a different proliferation effect between the growth of the B16 melanoma cells and the L929 fibroblast cells. The experimental results indicated that the effect of the PTT is dose-dependent in the B16 cells, ranging from cell proliferation to cytotoxic damage with apoptosis. Furthermore, we examined the intracellular changes in hydrogen peroxide (H2O2) and superoxide radicals ( $${\text{O}}_{2}^{{\bar{ \cdot }}}$$ ) following the Trp-derived treatment using the fluorescence probe method. Rigorous identification of the reactive oxygen species (ROS) produced by PTT in cells as $${\text{O}}_{2}^{{\bar{ \cdot }}}$$ and H2O2 helped establish the cellular source of ROS. It is shown that the intracellular H2O2 might originate from the activation of NADPH. These results suggested that the amino acid and protein hydroperoxides may play a crucial role in the action of plasma on the biologic target.

There and back again: from the origin of life to single molecules

What is life? There is hardly a more fundamental question raised by aspiring researchers, and one less prone to ever be answered in a scientifically satisfying way. In the long, productive and highly influential period of research following his Nobel-recognised work on relaxation kinetics, Manfred Eigen made seminal contributions towards a quantifiable definition of life, with a strong focus on its evolutionary character. In the last years of his time as an active researcher, however, he devoted himself to another, purely experimental topic: the detection and analysis of single biomolecules in aqueous solution. In this short review, I will give an overview of the groundbreaking contributions to the field of single molecule research made by Eigen and coworkers, and show that both, in its intrinsic motivation, and in its consequences, single molecule research strongly relates to the question of the physical–chemical essence of life. In fact, research on living systems with single molecule sensitivity will always refer the researcher to the question of the simplest possible representation, and thus the origin, of any biological phenomenon.