Spatially focused deposition of capillary electrophoresis effluent onto surface-enhanced Raman-active substrates for off-column spectroscopy

Abstract

Surface-enhanced Raman scattering (SERS) is employed to obtain distinctive spectra of compounds that are efficiently separated by capillary electrophoresis (CE) and deposited onto planar SERS-active substrates. A simple method is described that explains how to prepare SERS-active substrates by depositing a silver-colloid solution onto frosted-glass microscope slides, using a high-efficiency nebulizer. Scanning electron micrographs reveal a layered coating of fairly uniform-sized, 100-nm silver nanoparticles with interstitial spaces ranging from a few to tens of nanometers. The on-column separation is monitored by laser-induced fluorescence, while electrofilament depositing the CE effluent onto a moving SERS-substrate. Subsequently, the SERS spectra and off-column electropherograms are obtained with a simple confocal Raman spectrometer. The test compounds used to demonstrate this technique include compounds of biological significance: benzyloxyresorufin, riboflavin, and resorufin. CE and Raman conditions are evaluated to determine their affects on the SERS signals. An average off-column efficiency of 100,000 plates/m and a signal reproducibility of 11% relative standard deviation were achieved. Characteristic spectra with major Raman bands exhibiting signal-to-noise ratios of greater than 3 were obtained for a 3.2-nL injection of 10(-6) M (706 fg) resorufin. Forming a self-assembled monolayer (SAM) on the substrate increases the sensitivity of the SERS technique and decreases the on-substrate broadening. Calibration plots for both plain- and SAM-SERS substrates are demonstrated.

Achievements in resonance Raman spectroscopy

“…However, in CE it is essential that during deposition the electrical current be maintained. Appropriate devices have been developed by Devault and Sepaniak [105] and He et al [106]. In the latter case the capillary outlet is metallized to maintain electrical contact and the effluent is deposited by capillary forces.…”

Recent strategies toward microfluidic‐based surface‐enhanced Raman spectroscopy

“…Collecting separated fractions into microvials or on a moving substrate are common approaches toward off‐column SERS analysis. On‐line coupling is challenging for four main reasons: (1) accumulation of SERS signal is limited by the flow within the column, (2) laser‐induced photodeposition of NPs influences intensity of after‐going analyses, (3) in‐solution nanostructures can influence separation process, and (4) slow adsorption/desorption process results in memory effects.…”

SERS and Separation Science

“…As highlighted by the authors, because of the difference in surface affinities and cross sections for different analytes, calibration steps will be necessary for any quantification, but CE-SERS is an effective separation and identification method for multiple analytes even when CE separation may be incomplete. DeVault and Sepaniak [7] used a modified version of electrospray to spatially focus the effluent from a CE capillary onto an SERS-active substrate, in this case silver colloid solutions sprayed onto roughened glass slides, for off-line CE-SERS detection. Their experiments highlighted the importance of careful buffer selection for the CE separation as, while there are a large range of buffers available for separating charged analytes, not all of them will be compatible with an SERS surface.…”