164 Research Article Received: 17 March 2008 Accepted: 18 July 2008 Published online in Wiley Interscience: 19 September 2008 (www.interscience.wiley.com) DOI 10.1002/jrs.2099 Surface-enhanced Raman scattering study of L-tryptophan A. E. Aliaga, a I. Osorio-Rom ´ an, a P. Leyton, b C. Garrido, a J. C ´ arcamo, a C. Caniulef, a F. C ´ elis, c G. Díaz F., c E. Clavijo, a J. S. G ´ omez-Jeria a and M. M. Campos-Vallette a* Surface-enhanced Raman scattering (SERS) spectra of tryptophan (Trp) were obtained. A unique SERS spectrum of Trp, corresponding to the most stable conformation and orientation on the metal surface, is observed after a stabilization period. The Trp molecules interact with the surface through both the carboxylate and amino groups; the aliphatic moiety is close to the surface. The pyrrole ring of the indole moiety is farther from the surface than the benzene fragment. The observed spectra vary depending on both the preparation of the silver colloid and the aggregation time. The interpretation of the experimental results is supported by theoretical treatment of the molecule on the silver surface. Copyright c  2008 John Wiley & Sons, Ltd. Keywords: tryptophan; Raman spectroscopy; SERS; metal surface Introduction Amino acids are the molecular basis of proteins and enzymes. There are a varying number of amino acidic residues required to make up different proteins. Vibrational studies on such biological materials, in particular those related to surface-enhanced Raman scattering (SERS), have been intensively pursued in the last 10 years. SERS substrates provide an innovative and powerful approach to protein and amino acid studies. [1] The resultant SERS spectrum of a protein represents the point of interaction with metal, influenced by all the amino acids. To interpret this composite spectrum, it is necessary to have a fundamental understanding of the interactions of individual amino acids and peptides with metal substrates. Important advantages offered by SERS are the use of a small quantity of sample, detection of very low concentration, and the use of low laser power. The most relevant characteristic inherent to Raman spectroscopy is the insignificant Raman cross section of water, which allows study of biological materials in their natural media. SERS studies on tryptophan-related molecular systems such as tryptamine, [2] indole, [3] and proteins [4] have been published. SERS of other molecular systems involving L-tryptophan (Trp) has been reported. [5 – 12] Vidugiris et al. [6] studied the orientation of biological systems including Trp deposited on a silver electrode. Podstawka et al. [7] have proposed that the imino group of Trp in an S–S containing protein may bind to a colloidal silver surface. It was found [8] that Trp adsorbs on a silver surface via both the carboxylate (COO − ) and the amino (NH 2 ) groups; the nitrogen atom of the indole ring binds to the surface only when Trp is the C-terminal residue of a peptide. Stewart and Fredericks [9] showed that the adsorption of 19 L-amino acids was via the ionized COO − group, and that the side chain of most of the molecules was also in close proximity to the surface; the spectra also indicated that, in contrast, the amine terminal was protonated and relatively far from the surface. Reyes-Goddard et al. [10] published a review of the photodiagnosis using Raman and SERS of body fluids, including a section on amino-acid-based solutions; an SERS spectrum of Trp acquired on a Tollen reaction substrate using a near infrared (NIR) laser was displayed. Recently, Zhang et al. [11] developed a sensor, a hollow core photonic crystal fiber, for the detection of SERS from molecules in solution with silver nanoparticles; Trp was detected in the range 10 −4 to 10 −5 M. From this work, one can deduce that the SERS spectrum of Trp depends on several factors such as the laser line, the physical chemistry characteristics and the nature of the metal substrate, pH, ionic strength and concentration of the solution, aggregation time, and the eventual chemical reaction between the amino acid and the metal surface. The SERS spectrum reflects the mechanisms involved in the spectral enhancement, electromagnetic (EM) and charge-transfer (CT), [13] and the organization and orientation of the analyte on the metal surface. The present contribution deals with the SERS spectrum of Trp obtained under specific experimental conditions, particularly the time accommodation of the analyte in a silver colloidal solution at neutral pH. We also perform calculations on a simplified molecular model in which Trp interacts as zwitterionic species with a large silver surface. To our knowledge no calculations of this type have been performed. Experimental Materials Tryptophan of analytical grade was purchased from GIBCO and used as received. Stock solutions of tryptophan in water were ∗ Correspondence to: M. M. Campos-Vallette, Laboratorio de Espectroscopia Vibracional, Universidad de Chile, Las Palmeras 3425, Santiago, Chile. E-mail: facien05@uchile.cl a Facultad de Ciencias, Universidad de Chile, Santiago, Chile b Facultad de Ciencias B´ asicas y Matem´ aticas, P. Universidad Cat´ olica de Valparaíso, Valpo, Chile c Facultad de Ciencias, Universidad de Playa Ancha, Valpo, Chile J. Raman Spectrosc. 2009, 40, 164–169 Copyright c  2008 John Wiley & Sons, Ltd.