330
T. Roliński et al.
solution spectra are rather broad and noisy, the information content is definitively
larger for the spectra of solids, which are much sharper, more intense, and noisefree. It is well known that in solution the amino acids can exist in different conformational forms, which makes the Raman spectra very difficult to interpret. Similarly
to the situation in solution, amino acids in the solid phase are present as zwitterionic
structures. Now, however, only one conformer is present, of which the structure is
usually known from X-ray studies. thus, by measuring the Raman spectra in solids,
one avoids the problem of disentangling the mixture of conformers. Actually, different polymorphs are possible, which can be interconverted, e.g., by applying high
pressure [4]. however, this complication should be considered an advantage rather
than a drawback, since interconversion between different forms, if performed in a
controlled fashion, can provide additional information on the vibrational structure.
In this work, we report the spectra of twenty basic amino acids, recorded for powder samples on the same instrument using four different laser wavelengths, ranging
from uv (325 nm), through visible (514.5 and 632.8 nm) to the NIR (785 nm) region. We critically compare the spectra of the amino acids, using similarity analysis
techniques. the connection with the possibility to identify particular amino acids
in multi-component mixtures is underlined. Finally, examples of identification of
components in mixtures containing up to eight amino acids are presented.
12.2 Materials and Methods
Samples the solid amino acids used for recording the Raman spectra were purchased from Aldrich and merck. the samples from Aldrich included glycine,
410225; L-Alanine, A7627; L-Isoleucine, I7403; L-Asparagine anhydrous, 11149;
L-Cysteine, 168149; L-Phenylalanine, P212; L-tyrosine, T8566; L-tryptophan,
T8941; L-Serine, S4500; L-glutamine, G8540. the compounds from merck included L-Lysine monohydrate, 1.12233.0100; L-valine, 1.08495.0025; L-Leucine,
1.05360.0025; L-Serine, 1.07769.0010; L-Arginine, 1.01542.0100; L-Aspartic acid,
1.00126.0100; L-Proline, 1.07434.0010; L-glutamic acid, 1.00291.0250; L-methionine, 1.05707.0100; L-histidine, 1.04351.0025; L-threonine, 1.08411.0010.
Measurements the spectra were recorded with Renishaw Invia Raman microscope, using four laser lines for each amino acid sample: 785 nm (Renishaw
hPNIR785), 632.8 nm (heNe, Renishaw RL633), 514.5 nm (Ar
+
, Stellar Pro
modu-Laser, LLC) and 325 nm (heCd, Kimmon). the spectrometer was equipped
with 1200, 1800 and 2400 grooves/mm gratings, cutoff optical filters, 1024 × 256
pixels Peltier-cooled RenCam CCd detector, and an automatic translation stage.
the 20 × objective was used for visible and the 15 × objective for uv excitation.
the laser power on the sample was 50 mW or less. the spectral resolution was
5–6 cm
−1
in the visible region and 10 cm
−1
in the uv range, with the wavenumber accuracy of 2 cm
−1
. the Raman shift was calibrated using both the Rayleigh
line and the 520.6 cm
−1
line of crystalline silicon. the wavelength dependence of
T. Roliński et al.
solution spectra are rather broad and noisy, the information content is definitively
larger for the spectra of solids, which are much sharper, more intense, and noisefree. It is well known that in solution the amino acids can exist in different conformational forms, which makes the Raman spectra very difficult to interpret. Similarly
to the situation in solution, amino acids in the solid phase are present as zwitterionic
structures. Now, however, only one conformer is present, of which the structure is
usually known from X-ray studies. thus, by measuring the Raman spectra in solids,
one avoids the problem of disentangling the mixture of conformers. Actually, different polymorphs are possible, which can be interconverted, e.g., by applying high
pressure [4]. however, this complication should be considered an advantage rather
than a drawback, since interconversion between different forms, if performed in a
controlled fashion, can provide additional information on the vibrational structure.
In this work, we report the spectra of twenty basic amino acids, recorded for powder samples on the same instrument using four different laser wavelengths, ranging
from uv (325 nm), through visible (514.5 and 632.8 nm) to the NIR (785 nm) region. We critically compare the spectra of the amino acids, using similarity analysis
techniques. the connection with the possibility to identify particular amino acids
in multi-component mixtures is underlined. Finally, examples of identification of
components in mixtures containing up to eight amino acids are presented.
12.2 Materials and Methods
Samples the solid amino acids used for recording the Raman spectra were purchased from Aldrich and merck. the samples from Aldrich included glycine,
410225; L-Alanine, A7627; L-Isoleucine, I7403; L-Asparagine anhydrous, 11149;
L-Cysteine, 168149; L-Phenylalanine, P212; L-tyrosine, T8566; L-tryptophan,
T8941; L-Serine, S4500; L-glutamine, G8540. the compounds from merck included L-Lysine monohydrate, 1.12233.0100; L-valine, 1.08495.0025; L-Leucine,
1.05360.0025; L-Serine, 1.07769.0010; L-Arginine, 1.01542.0100; L-Aspartic acid,
1.00126.0100; L-Proline, 1.07434.0010; L-glutamic acid, 1.00291.0250; L-methionine, 1.05707.0100; L-histidine, 1.04351.0025; L-threonine, 1.08411.0010.
Measurements the spectra were recorded with Renishaw Invia Raman microscope, using four laser lines for each amino acid sample: 785 nm (Renishaw
hPNIR785), 632.8 nm (heNe, Renishaw RL633), 514.5 nm (Ar
+
, Stellar Pro
modu-Laser, LLC) and 325 nm (heCd, Kimmon). the spectrometer was equipped
with 1200, 1800 and 2400 grooves/mm gratings, cutoff optical filters, 1024 × 256
pixels Peltier-cooled RenCam CCd detector, and an automatic translation stage.
the 20 × objective was used for visible and the 15 × objective for uv excitation.
the laser power on the sample was 50 mW or less. the spectral resolution was
5–6 cm
−1
in the visible region and 10 cm
−1
in the uv range, with the wavenumber accuracy of 2 cm
−1
. the Raman shift was calibrated using both the Rayleigh
line and the 520.6 cm
−1
line of crystalline silicon. the wavelength dependence of