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Matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS) is a well-implemented analytical technique for the investigation of complex biological samples. In MS, the sample preparation strategy is decisive for the success of the measurements. Here, sample preparation processes and target materials for the investigation of different pollen grains are compared. A reduced and optimized sample preparation process prior to MALDI-TOF measurement is presented using conductive carbon tape as target. The application of conductive tape yields in enhanced absolute signal intensities and mass spectral pattern information, which leads to a clear separation in subsequent pattern analysis. The results will be used to improve the taxonomic differentiation and identification, and might be useful for the development of a simple routine method to identify pollen based on mass spectrometry.
Matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS) is a well-implemented analytical technique for the investigation of complex biological samples. In MS, the sample preparation strategy is decisive for the success of the measurements. Here, sample preparation processes and target materials for the investigation of different pollen grains are compared. A reduced and optimized sample preparation process prior to MALDI-TOF measurement is presented using conductive carbon tape as target. The application of conductive tape yields in enhanced absolute signal intensities and mass spectral pattern information, which leads to a clear separation in subsequent pattern analysis. The results will be used to improve the taxonomic differentiation and identification, and might be useful for the development of a simple routine method to identify pollen based on mass spectrometry.
RATIONALE
A fast and reliable online identification of pollen is not yet available. The identification of pollen is based mainly on the evaluation of morphological data obtained by microscopic methods.
METHODS
Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-TOF MS) was applied to the analysis of extracts and milled pollen samples. The obtained MALDI data were explored for characteristic peak patterns which could be subjected to a multivariate statistical analysis.
RESULTS
Two sample preparation methods are presented, which require only minimal or no chemical extraction of the pollen. MALDI pollen spectra could be recorded showing various peak patterns. A multivariate statistics approach allowed the classification of pollen into clusters indicating similarities and differences between various species.
CONCLUSIONS
These results demonstrate the potential and the reliability of MALDI-TOF MS for the identification and, in combination with multivariate statistics, also for the classification of pollen.
Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) has been suggested as a promising tool for the investigation of pollen, but the usefulness of this approach for classification and identification of pollen species has to be proven by an application to samples of varying taxonomic relations.
MALDI time-of-flight mass spectrometry (MALDI-TOF MS) has become a widely used tool for the classification of biological samples. The complex chemical composition of pollen grains leads to highly specific, fingerprint-like mass spectra, with respect to the pollen species. Beyond the species-specific composition, the variances in pollen chemistry can be hierarchically structured, including the level of different populations, of environmental conditions or different genotypes. We demonstrate here the sensitivity of MALDI-TOF MS regarding the adaption of the chemical composition of three Poaceae (grass) pollen for different populations of parent plants by analyzing the mass spectra with partial least squares discriminant analysis (PLS-DA) and principal component analysis (PCA). Thereby, variances in species, population and specific growth conditions of the plants were observed simultaneously. In particular, the chemical pattern revealed by the MALDI spectra enabled discrimination of the different populations of one species. Specifically, the role of environmental changes and their effect on the pollen chemistry of three different grass species is discussed. Analysis of the Group formation within the respective populations showed a varying influence of plant genotype on the classification, depending on the species, and permits conclusions regarding the respective rigidity or plasticity towards environmental changes.