**Calculation of the Absorption Spectrum from an ATR Infrared
Experiment **

**Barbara Grobelnik and Jože Grdadolnik* **

*National Institute of Chemistry, Hajdrihova 19, Ljubljana, Slovenia
* Corresponding author: E-mail:
joze.grdadolnik@ki.si *

**Abstract
**The most efficient and precise method for separation of absorption and
reflection contributions in the ATR (attenuated total reflection) spectrum is
the calculation of pure absorption spectrum from optical constants. However, the
precision of calculated optical constants depends on the alignment of the
optical component of an ATR attachment. The arrangement of the ATR attachment
always produces imperfections in optical path alignment. Therefore, we
calculated the expected error analytically. The calculated error of Rs and Rp is
in the range between 3%–8% per deviation of the incidence angle of 1°. To reduce
the error of calculated reflectivities, a new procedure for recording the ATR
spectrum is proposed. It is based on the calibration of the number of reflection
by probing the spectrum of pure liquid water. The proposed procedure
significantly reduces the error (< 2%) due to a divergence of the incidence
angle. This approach is included in the program for routine calculation of the
optical constants and absorption spectra of anisotropic solutions. A structural
analysis of three dipeptides in water solutions is shown as an example of the
application of an ATR attachment and the calculation of the absorption spectrum.
The calculated spectra of dipeptides and bulk water provide ideal conditions for
reliable subtraction. The OH stretching region in the difference spectrum of the
leucine dipeptide shows characteristic patterns of rearranged water molecules in
the vicinity of solutes. From the positions of negative and positive bands we
hypothesize that the presence of leucine molecules creates a denser structure of
water molecules near the solutes. The modelling of the amide III band shape in
the spectra of the methionine and asparagine dipeptide in water reveals the
conformation of the dipeptide backbone. The methionine dipeptide possesses
mainly the

**Keywords:** Infrared spectroscopy, ATR experiment, calculation of
optical constants, anisotropic solutions, preferential dipeptide conformations