The Pearl is our solution to making transmission IR spectroscopy as easy as ATR-FTIR for analysing viscous liquids and greases. The same ease-of-use benefits are apparent in other situations also, however. Below we outline some appearances of the Pearl accessory in the scientific literature.
Investigating bacterial resistance to antimicrobial agents
Schmid et al. investigated ion lipid pairs as a proxy for the chemistry involved in the development of antimicrobial resistance in bacteria [1]. By studying simple chemical analogues for more complex biological systems insights can be gained and used to support the development of novel antimicrobial agents. For the pathogen Staphlyococcus aureus increased formation of a cationic lipid, even at relatively low concentrations, is known limit the effect of cationic antimicrobial peptides on the bacteria. This resistance arises from an ionic interaction between a negatively charged phosphate functional group in the bacteria’s membrane and a positively charged amine group in the lipid. Schmid and co-workers therefore chose to investigate the interaction between an organophospholipid (dipalmitoylglcero-3-phosphoglycerol, DPPG) and a quaternary ammonium compound (dihexadecyldimethylammonium, DHDAB) in an aqueous solution. With the former acting as a proxy for the bacterial membrane and the latter as a proxy for the cationic lipid. DHDAB was carefully selected by the authors to avoid functional groups that might obscure DPPG’s PO peaks in the FTIR spectrum.
After subtraction of the water solvent peaks, clear peaks were observed at 1221 and 1201 cm-1, assigned to the PO functional groups. The ratio of these two peaks was measured against concentration, with the 1221 cm-1 peak increasing relative to the 1201 cm-1 peak as the molar ratio of DPPG:DHDAB was decreased. This change in peak ratio provides direct evidence for the formation of the ion pair.
Further changes in the spectrum were also observed in the C-H stretching region (3000-2800 cm-1) as the molar ratio was adjusted. Initially as the relative concentration of DHDAB was increased, the CH2 peaks red shifted with increasing concentration of DHDAB, indicating an increase in the order and tighter packing of the alkyl chains. At relative molar ratios >0.5 DHDAB, the trend reversed, and the peaks began to blue shift, indicating a decrease in order and increase in the gauche conformer. These changes were assigned to Van der Walls intermolecular forces between the alkyl chains of the two ions. This implies that the interaction in the antimicrobial resistant bacteria may not be solely due to ionic bonding between the membrane phosphate and amine, which helps to explain why a small relative increase in amine concentration in the bacteria can have such a hugely negative effect on the binding between the antimicrobial agent and bacteria. This insight will hopefully prove invaluable in the fight against this bacterium through the development of novel therapeutics targeting this specific mode of resistance.
Pearl’s horizontal Oyster cells prove a benefit
In the study a Pearl fitted with a 25 μm CaF2 Oyster cell provided a unique benefit: at concentrations approaching a 50:50 mixture of anion and cation the ion pairs coagulate and form a significant amount of sedimentation. This could result in erroneous data in a traditional style cell where the sediment would sink to the bottom, however owing to the innovative design of the cell, whereby the cell is rotated 90 degrees and held horizontally within the IR beam, any sedimentation was retained within the IR beam and analysed as part of their measurement.
The effect of pH on peptide nanotubes
Castelletto et al. have studied a self-assembled peptide nanotube, the surfactant like arginine3–leucine12 (R3L12) [2]. This type of nanotube has interesting applications including biocatalysis and confinement of pharmaceutical molecules for targeted release of drugs, amongst others. These peptide nanotubes form a wide array of structures including coils, and β-sheets [3,4]. In their previous work [5] the authors reported that under acidic conditions R3L12 formed α-helix structures with cross linkers between the nanotubes. This structure is unique in the scientific literature to date. The nanotubes self-assembled into a structure where the arginine moiety was present on the walls of the tubes (both inner and outer faces) with the leucine fragment embedded within the nanotube.
For their study involving the Pearl, the authors were interested in probing the effect of switching the environment to a more basic one, with the pH varied from 9 to 13, at concentrations from 0.04-0.07 wt% of peptide. To probe the effect on the nanotubes the authors used a variety of techniques including transmission electron microscopy (TEM), circular dichosim (CD) spectroscopy, small angle X-ray scattering (SAXS) and FTIR spectroscopy. The CD spectroscopy demonstrated that there was a loss α-helix structure as the pH was raised, with nanotubes detected by SAXS measurements only at pH 9. The TEM and SAXS measurements demonstrated that, at higher pH’s, the nanotubes reformed into globular, micelle like structures. Whilst TEM, CD and SAXS were able to demonstrate that a change had taken place, only the FTIR spectroscopy could be used to probe those changes at a molecular level. The use of the Pearl fitted with a CaF2 oyster cell was therefore critical in understanding why the nanotubes fell apart at higher pH’s.
The FTIR spectra confirmed the loss of the α-helix structure, with the peak at 1665 cm-1, assigned to the amide I band consistent with an α-helix structure present at pH 9, before declining in the spectra recorded at pH 12 and 13. The amide III band remained consisted across different pH levels. In the CH region peaks at 2871 and 2959 cm-1 were assigned to symmetric and asymmetric mode of CH3 terminal groups and at 2930 cm-1 to CH/CH2 stretching modes. These peaks are assigned to the 2-Methylpropyl side chain on the leucine peptide. These peaks were observed to decline in intensity when the pH was raised which indicated the formation of a highly disordered molecular state, potentially with the formation of R3L12 dimers.
References
[1] Castelletto, V., Seitsonen, J., Ruokolainen, J. & Hamley, I.W., Soft Matter., 17, (2021), 3096-3104. DOI: 10.1039/D0SM02095H
[2] Schmid, M., Wölk, C., Giselbrecht, J., Chan, K.L.A. & Harvey, R.D., Colloids Surf. B, 169, (2018), 298-304. DOI:10.1016/j.colsurfb.2018.05.031
[3] Morris, K.L., Zibaee, S., Chen, L., Goedert, M., Sikorski P. & Serpell, L.C., Angew. Chem., Int. Ed., 52, (2013), 2279 —2283. DOI: 10.1002/anie.201207699
[4] Pandya, M.J., Spooner, G.M., Sunde, M., Thorpe, J. R., Rodger, A. & Woolfson, D. N., Biochemistry, 39, (2000), 8728 —8734. DOI: 10.1021/bi000246g.
[5] Castelletto, V., Seitsonen, J. , Ruokolainen, J., Piras, C., Cramer, R., Edwards-Gayle C.J.C, & Hamley I.W.,, Chem. Commun., 56, (2020), 11977–11980. DOI: 10.1039/D0CC04299D
Related Posts
GET IN TOUCH