DROP COATING DEPOSITION RAMAN – A NEW METHOD IN RAMAN SPECTROSCOPY FOR BIOMOLECULAR SAMPLES OF LOW CONCENTRATIONS

V. Kopecký Jr., V. Baumruk, J. Kapitán, J. Štěpánek

Institute of Physics, Faculty of Mathematics and Physics, Charles University, Ke Karlovu 5, 121 16 Prague 2, Czech Republic, e-mail: kopecky@karlov.mff.cuni.cz

Raman spectroscopy can be a powerful probe of structure and function of biomolecules. However, its usage, particularly in biochemistry, is often limited by amount and concentration of samples. Recently reported new technique of non-enhanced Raman spectroscopy – a drop coating deposition Raman (DCDR) method [1] – based on a coffee ring effect [2] enables nondestructive measurements of solutions with concentration of biomolecules down to 1 µM (ca. 0,01 mg/mL). The method is based on a deposition of a small amount of sample solution (typically units of mL) on a Teflon-coated stainless steel surface with almost no Raman signal. The hydrophobic surface enables drying of a sample. It has been demonstrated that even after drying its solution structure is preserved and Raman spectra taken from the dried deposit and (concentrated) solution are virtually identical [1].

Increasing number of papers demonstrates usefulness of DCDR method. It has been shown that DCDR can serve as an analytical tool in identification of insulin variants [3] or as a detection method of peptide tyrosine phosphorylation [4]. DCDR is useful also in chemometric analysis as was demonstrated by oligosaccharide identification and quantification in mixtures [5]. Possible coupling of DCDR with HPLC analysis and extension to proteomic analysis generally can bring new quality to scientific research.

Despite of significant increase of information about the practical usage of DCDR method, the basic questions remain unanswered: (i) how is it possible that the samples are almost identical in solution and in a drop deposit? (ii) are they really identical or little specific differences do exist? (iii) is it possible to estimate humidity of drop deposited samples? (iv) what samples remain intact after deposition and what samples are not suitable for DCDR method?

The aim of this work is to answer these questions as well as to give an overview of DCDR method and our first experience with it.

The authors thank M. Trchová (Institute of Macromolecular Chemistry, AS CR) for her kind assistance with Raman microscopy and K. Hofbauerová and K. Bezouška (Department of Biochemistry, Charles University) for a generous gift of protein samples. Ministry of Education of the Czech Republic and the Grant Agency of the Czech Republic are gratefully acknowledged for support (No. MSM 0021620835, No. 202/05/0628, respectively).

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[2] R. D. Deegan, O. Bakajin, T. F. Dupont, G. Huber, S. R. Nagel, T. A. Witten, Nature 389 (1997) 827–829.

[3] C. Ortiz, D. Zhang, Y. Xie, V. J. Davisson, D. Ben-Amotz, Anal. Biochem 322 (2004) 245–252.

[4] Y. Xie, D. Zhang, G. K. Jarori, V. J. Davisson, D. Ben-Amptz, Anal. Biochem. 332 (2004) 116–121.

[5] M. F. Mrozek, D. Zhang, D. Ben-Amotz, Carbohydr. Res. 339 (2004) 141–145.