Characterization of Pollen by Vibrational Spectroscopy

Abstract

Classification, discrimination, and biochemical assignment of vibrational spectra of pollen samples belonging to 43 different species of the order Pinales has been made using three different vibrational techniques. The comparative study of transmission (KBr pellet) and attenuated total reflection (ATR) Fourier transform infrared (FT-IR) and FT-Raman spectroscopies was based on substantial variability of pollen grain size, shape, and relative biochemical composition. Depending on the penetration depth of the probe light, vibrational techniques acquire predominant information either on pollen grain walls (FT-Raman and ATR-FT-IR) or intracellular material (transmission FT-IR). Compared with the other two methods, transmission FT-IR obtains more comprehensive information and as a result achieves superior spectral identification and discrimination of pollen. The results strongly indicate that biochemical similarities of pollen grains belonging to the same plant genus or family lead to similar features in corresponding vibrational spectra. The exploitation of that property in aerobiological monitoring was demonstrated by simple and rapid pollen identification based on relatively small spectral libraries, with the same (or better) taxonomic resolution as that provided by optical microscopy. Therefore, the clear correlation between vibrational spectra and pollen grain morphology, biochemistry, and taxonomy is obtained, while successful pollen identification illustrates the practicability of such an approach in environmental studies.

Vibrational microspectroscopy enables chemical characterization of single pollen grains as well as comparative analysis of plant species based on pollen ultrastructure

“…On the other hand, the spectrum of the grain that has distal polar orientation has strong signals of sporopollenins. The sporopollenin bands at 1605, 1515, 1171 and 833 cm -1 can be associated with the vibrations of aromatic rings (Zimmermann 2010;Schulte et al 2008;Zimmermann and Kohler 2014). The reason for this discrepancy in spectral signatures of pollen for the two orientations is the spatial arrangement of the main biochemicals in Pinaceae pollen grains.…”

“…Lipids are characterized by the strong vibrational band at 1745 cm -1 (C=O stretch), as well as by weaker bands at 1462 cm -1 (CH2 deformation) (Gottardini et al 2007;Pappas et al 2004;Zimmermann 2010;Zimmermann and Kohler 2014). Proteins are characterized by two strong and broad bands at 1650 cm -1 (amide I: C=O stretch) and 1550 cm -1 (amide II: NH deformation and C-N stretch), while carbohydrates have strong absorption in 1200-900 cm -1 region (C-O-C and C-OH stretch) (Gottardini et al 2007;Pappas et al 2004;Schulte et al 2008;Zimmermann 2010;Zimmermann and Kohler 2014). On the other hand, the spectrum of the grain that has distal polar orientation has strong signals of sporopollenins.…”

“…Pollen grains of various species can vary quite a lot in grain shape and size, number, type and position of apertures, and structure and sculpture of the grain wall. In the last decade, vibrational spectroscopy of pollen has been performed in macroscopic mode on bulk samples, resulting in the clear identification of main chemical constituents (Zimmermann and Kohler 2014;Zimmermann 2010;Parodi et al 2013;Pappas et al 2003;Gottardini et al 2007;Schulte et al 2010;Schulte et al 2009;Schulte et al 2008;Ivleva et al 2005).…”

“…However, the process not only destructs the sample partially, it also significantly increases the spectral acquisition time. An alternative solution is the application of FT-Raman spectroscopy employing a near infrared (NIR, 1064 nm) laser excitation (Zimmermann 2010). Unfortunately, the use of a near infrared laser excitation in FT-Raman spectroscopy results in a lower spatial resolution and sensitivity compared to dispersive Raman where lasers with wavelengths in the visible region are employed.…”

On the Taxonomic Resolution of Pollen and Spore Records of Earth’s Vegetation

“…In the case of Late Silurian (∼425 Ma) cryptospores from Sweden, Fourier transform infrared spectroscopy has been used to demonstrate that the wall of these fossils contains sporopollenin and is of similar composition to the wall of fossilized trilete spores but is chemically distinct from the wall of chitinozoans and leiospheres (Steemans et al 2010). Such spectroscopic techniques have also found application in the monitoring of airborne allergenic pollen (Dell'Anna et al 2009;Zimmerman 2010). In a similar but geologically younger example, the carbon isotopic composition of fossil grass pollen has been used to show that grasses using C 4 photosynthesis were present on the Earth in the Early Oligocene (∼33 Ma; Urban et al 2010).…”