Search Menu
Photonics Media Photonics Buyers' Guide Photonics EDU Photonics Spectra BioPhotonics EuroPhotonics Industrial Photonics Photonics Showcase Photonics ProdSpec Photonics Handbook
More News
Email Facebook Twitter Google+ LinkedIn Comments

  • Confocal Raman Microscopy Studies Drug-Membrane Interactions

Photonics Spectra
Aug 2006
Lauren I. Rugani

Investigating drug-membrane interactions, such as a drug’s ability to pass through a membrane or to bind to proteins within it, helps researchers understand the effects of drugs on membrane functions. Christopher B. Fox, Robert A. Horton and Joel M. Harris of the University of Utah in Salt Lake City have applied optical-trapping confocal Raman microscopy to study the accumulation of salicylate and ibuprofen within phospholipid vesicles and to determine the disordering effects of the drugs on the lipid membranes.

Micro Update 3_1 of 2_Confocal_raman_microscope.jpg

The optical-trapping confocal Raman microscope setup includes a Kr+ laser (left), an inverted fluorescence microscope (center) and a monochromator and CCD detector (bottom right).

Other techniques used to perform similar studies face obstacles, such as the need for a spectroscopic label or high concentrations of lipid, which can deplete drug concentration. Optical-trapping confocal Raman microscopy does not require labeling for the assessment of drug-membrane interactions, and spectroscopic analysis can be conducted with a low drug concentration.

“This technique is uniquely suited to measuring the local composition of a lipid vesicle, so that the accumulation of drug molecules in the lipid membrane can be monitored,” explained Harris, a chemistry professor at the university.

In the optical trapping experiments, a beam from a Coherent Inc. Kr+ laser, operating at 647.1 nm, is passed through the Raman microscope setup centered on a Nikon inverted fluorescence microscope, and is focused on a 0.6-μm-diameter spot inside a flow cell. The focused beam traps individual 1,2-dimyristoyl-sn-glycero-3-phosphocholine vesicles in solution containing varying drug concentrations for three minutes, while a CCD camera from Andor Technology collects spectra over a three-pixel, or 66-μm, vertical dimension.

Micro Update 3_2of2_Image of trapped vesicle.jpg
A 0.6-μm 1,2-dimyristoyl-sn-glycero-3-phosphocholine lipid vesicle is optically trapped by a focused laser beam.
Raman spectra of the vesicles were compared, revealing that both ibuprofen and salicylate exhibit disordering effects on the membrane. Drug concentration in the vesicles and changes in membrane order were evaluated, respectively, by distinct Raman scattering peaks (at 813 cm–1 for salicylate and at 1611 cm–1 for ibuprofen) and by peak intensity ratios (1086:1065 cm–1 and 2925:2881
cm–1). Both peak intensities vary with drug concentration, confirming the disordering effect. Raman scattering intensity from ibuprofen indicates that it accumulates in the membrane, while salicylate shows no significant partitioning. Because the volume of a lipid membrane is small relative to the trapped vesicle, Harris noted, it is difficult to determine quantitatively the accumulation of drugs unless the partitioning is large.

These results are supported by spatial profiling analysis of surface-adhered vesicles. Using confocal Raman microscopy, spectra from ∼5-μm-diameter 1,2-dipalmitoyl-sn-glycero-3-phosphocholine vesicles — adhered to the surface of the glass cover slip — are gathered in 0.5-μm increments to create a profile of drug concentration in the lower membrane, interior and upper membrane. Ibuprofen was observed to partition preferentially into the membrane, whereas salicylate showed no local increase in concentration.

The team is developing a high thermal conductivity sample holder and temperature controller for use in temperature-dependent studies. “We plan to study the phase-transition behavior of individual lipid vesicles and the effects of drug molecules on these transitions,” Harris said.

Analytical Chemistry, online May 28, 2006, doi:10.1021/ac0605290.

Terms & Conditions Privacy Policy About Us Contact Us
back to top

Facebook Twitter Instagram LinkedIn YouTube RSS
©2016 Photonics Media
x Subscribe to Photonics Spectra magazine - FREE!