Journal of Chromatography A, 1226 (2012) 43–47 Contents lists available at ScienceDirect Journal of Chromatography A jou rn al h om epage: www.elsevier.com/locat e/chroma Photodeposited silver nanoparticles for on-column surface-enhanced Raman spectrometry detection in capillary electrophoresis Jan Pˇ rikryl, Karel Klepárník ∗ , Frantiˇ sek Foret Institute of Analytical Chemistry of the Academy of Sciences of the Czech Republic, Brno, Czech Republic a r t i c l e i n f o Article history: Available online 22 July 2011 Keywords: SERS Capillary electrophoresis Photodeposition Metal nanoparticle a b s t r a c t A new, simple photo-deposition method of silver nanoparticles induced by laser inside a fused-silica capil- lary is described and tested. Silver nanoparticles are immobilized using Ar-ion laser beam of a wavelength of 488 nm and power of 3.6 mW for 60 min. The photodeposited compact spot of a size of ∼10 m is tem- porary and spatially stable and resistant to a hydrodynamic ﬂow. The deposit has very good properties for surface-enhanced Raman scattering and serves well for detection in capillary electrophoresis. The advantage of this approach is that neither the silver nanoparticles nor the chemicals for their preparation are components of the background electrolyte during the electrophoretic separation. Thus, the substrate formation and separation of analytes are two independent processes and can be performed under their optimum conditions. The zone broadening due to the sorption of analytes on the immobilized nanopar- ticles can be signiﬁcantly reduced by an addition of 20% solution of methanol. The efﬁciency of capillary electrophoresis and detection selectivity of surface-enhanced Raman scattering induced by He–Ne laser at 632.8 nm is demonstrated by the 3D electropherograms of rhodamines 123 and B as model samples. The limits of detection of about 49 and 150 fmol (1 and 2 M) have been reached for rhodamine B and 123, respectively. © 2011 Elsevier B.V. All rights reserved. 1. Introduction The discovery of surface-enhanced Raman scattering (SERS) has revealed its great potential not only in analytical chemistry but also in biology, pharmacy, environmental chemistry, biomedicine, etc. Although the ﬁrst SERS experiment with pyridine adsorbed on an electrochemically roughened silver electrode was performed by Fleischmann et al. as early as in 1974 [1], SERS mechanism has not been completely explained yet. The method is based on coupled optical responses of nanostructured metals and analyte molecules in a close proximity [2]. According to the electromagnetic theory, the laser induced plasmon enhances electric ﬁeld strength at metal surfaces inducing structure-speciﬁc Raman scattering on molecules in the proximity to such a metal surface (e.g., adsorbed due to physical forces). However, many experiments proved also chemical mechanism of the enhancement [3,4]. The theory of the chemical mechanism, which does not involve the surface plasmon, is based on the charge transfer within the complex between a metal particle and chemisorbed molecule. Currently, SERS has the status of a well established analytical spectrometric method providing highly sensitive quantitative [5–8] and qualitative analyses [9]. ∗ Corresponding author. Tel.: +420 532 290 122; fax: +420 541 212 113. E-mail address: kleparnik@iach.cz (K. Klepárník). The vibration frequencies of an analyte affect the spectrum of the scattered light, which is used for the analyte identiﬁcation. This is based on the detection of characteristic bands of functional groups or the ﬁngerprint region of the vibration spectra. Thus, a high speciﬁcity and sensitivity of this method makes SERS a highly applicable analytical method. Even single molecule (SM) sensitivity of SERS was proposed by Nie and Kneipp [10,11] theoretically and, several years later, SM sensitivity of SERS has been experimentally proved by various techniques [12–19]. Many publications describe various SERS applications such as protein detection [4,20], SERS imaging of living cells [4,21], cancer diagnostics [22], glucose sensing or SERS immunoassays [4]. In most of the applications mentioned above, SERS was used as a stand-alone method. However, relatively little attention was focused to coupling of SERS with separation methods such as liquid chromatography [23–26] or capillary electrophoresis (CE). In 2000, Nirode et al. published a simple way to integrate the on-column SERS detection in CE by usage of SERS-active silver nanoparticles (Ag NPs) suspended in the CE background electrolyte (BGE) [27]. They reported limits of detection (LOD) of 1 M and low-nanomolar for riboﬂavin and rhodamine 6G (Rh6G), respectively. However, the SERS spectra degraded with time, probably due to the sorption of nanoparticles on the capillary wall. Moreover, the inﬂuence of NPs on the separation efﬁciency was not discussed in detail. Recently, the research in the ﬁeld of coupling CE and SERS was focused on 0021-9673/$ – see front matter © 2011 Elsevier B.V. All rights reserved. doi:10.1016/j.chroma.2011.07.045