2D IR analyses of rate processes in lipid–antibiotic monomolecular films
Introduction
Two dimensional infrared correlation spectroscopy (2D IR) has proven to be a valuable tool due to its ability to enhance spectral resolution and identify overlapped spectral features [1]. This has proven especially valuable in infrared spectroscopic studies of biomolecules, as this enables one, for example, to identify discrete and unique protein secondary structure conformations as well as interconversion of one form to another as a result of changes in external environment [2], [3], [4], [5], [6], [7]. Two-dimensional IR correlation analysis has also been used to analyze structure in monomolecular films. The phase behavior of phospholipid monolayers have been studied using 2D IR and it was shown how these methods could distinguish bands due to co-existing phases in a disorder-order phase transition in the monolayer [8], [9].
Standard 2D IR methods have been most successfully employed in simplifying complex spectra and facilitating band assignments through resolution enhancement [10]. In addition to these uses, 2D IR can also be used to determine the temporal order of events that occur in a set of dynamically varying spectra upon sample perturbation. The basis for this determination is the relative signs of the synchronous and asynchronous cross-peak at coordinate (ν1, ν2) in the 2D correlation plots [11].
While it is certainly possible to determine the relative sequence of molecular events based on standard 2D IR methods, this procedure tends to be difficult to implement for highly overlapped spectra and may lead to uncertainties. In order to more quantitatively describe the degree of coherence between spectral intensity changes and the sequence of molecular events in a set of dynamic spectra, we have recently developed a modified 2D IR correlation method called βν correlation analysis [12]. This method is a variation of asynchronous cross-correlation, in which dynamically varying spectra are correlated against a mathematical function with a varying phase angle. We recently applied βν correlation analysis to surface pressure-induced changes in the IRRAS spectra of phospholipid monolayers at the A/W interface, and showed how the relative rates of acyl chain and methyl group reorientation could be quantitatively determined [13] and have also applied this analysis in the study of conformational changes and relative reorientation rates of hydrophobic surfactant proteins SP-B and SP-C at the A/W interface [14].
Our current objective is to use polarization-modulation infrared reflection absorption spectroscopy (PM-IRRAS) to study the interactions of the antibiotic tetracycline with a phospholipid monolayer at the air–water interface. It has been shown that tetracycline induces significant changes on phospholipid monolayers and the strongest interactions are observed for the DPP A system due to specific dipole–dipole interactions [15], [16]. The aim of the current study is to unambiguously identifying the specific regions of interaction between the two molecules at the air–water interface.
To accomplish this aim, we use both conventional 2D IR and βν correlation analysis to analyze the PM-IRRAS spectra obtained from the lipid–antibiotic interactions. In addition, we introduce a new model-dependent 2D correlation method, kν correlation analysis. Our results are able to clearly identify the functional groups involved in this lipid–antibiotic interaction and the order in which their respective functional groups reorient upon increasing monolayer surface pressure.
Section snippets
Materials
Tetracycline hydrochloride (TC) was obtained from Sigma (St. Louis, MO, purity > 99%) while 1,2-dipalmitoyl-sn-glycero-3-phosphatidic acid (DPPA) (>99%) was obtained from Avanti Polar Lipids (Alabaster, AL). The chemical structures of tetracycline hydrochloride and DPPA are shown in Fig. 1. HPLC grade chloroform (J.T. Baker, Phillipsburg, NJ) was used as the spreading solvent and typical DPPA concentrations of 1 mg/mL were used for making the spreading solutions. Ultrapure H2O obtained from a
Monolaver IR spectroscopv
Previous research using monomolecular films of DPPA on a tetracycline-containing subphase has indicated that specific inter-molecular interactions may occur between the polar head groups of the phospholipid in the condensed phase and the tetracycline molecules dissolved in the subphase [15], [16]. In order to test this hypothesis, we employed polarization-modulation infrared reflectance–absorption spectroscopy (PM-IRRAS) and obtained IRRAS spectra at the air–water (A/W) interface for monolayer
Conclusions
Polarization modulation infrared reflection spectra of a dipalmitoyl phosphatidic acid monolayer on a subphase containing 5 mM tetracycline hydrochloride were collected under varying surface pressures. Two-dimensional IR, βν and kν correlation analyses were performed on these PM-IRRAS spectra to gain a better understanding of the surface pressure-induced effects on the interaction between the phospholipid and antibiotic.
The synchronous 2D IR correlation map reveal strong correlation behavior
Acknowledgements
The work described here was supported by the US Public Health Service through National Institutes of Health grant EB001956 (R.A.D.).
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Cited by (13)
Effect of tetracycline antibiotic on the monolayers of phosphatidylcholines at the air-water interface
2015, Colloids and Surfaces A: Physicochemical and Engineering AspectsCitation Excerpt :Apart from the plateau region, the DPPC isotherm under the influence of TC in the subphase rises more steeply, and therefore we are inclined to think that this is due to the hydrophobic interactions of long alkyl chains facilitating the organization and packing of the hydrophobic part of lipid molecules forming the monolayers and promoting better ordering, even in the presence of unsaturated bond (DOPC). From the above data it seems unlikely that TC (in its cationic form at this concentration and pH) can penetrate the hydrophobic moiety of the monolayer, and this observation is supported in the literature [14,18]. Having said that, let us now focus on the CPD results that were acquired simultaneously with pressure-area isotherms and compressibility modulus data.
Orientation of solid supported lipid bilayers examined by βν-correlation analysis
2012, Vibrational SpectroscopyCitation Excerpt :Using the βν correlation analysis the relative rates of conformational ordering in the individual components of binary monolayer mixture spread at the air–water (A/W) interface was determined [10]. Also the phase transitions in Langmuir–Blodgett polymer films and in phospholipid monolayers as well as secondary structure of protein in lipid mixture, the lipid-antibiotic interactions and changes in micelle structure triggered by a thermally induced helix-to-sheet protein folding transition were studied by this approach [11–13]. To the best of our knowledge, until now only Dluhy's group put into practice the βν-correlation method.
PM-IRRAS at liquid interfaces
2011, Biointerface Characterization by Advanced IR SpectroscopyProjection two-dimensional correlation analysis
2010, Journal of Molecular StructureAdsorption and intercalation of tetracycline by swelling clay minerals
2009, Applied Clay ScienceProgress in two-dimensional (2D) correlation spectroscopy
2006, Journal of Molecular Structure