Mass spectroscopy, anthocyanins

Giusti, M.M. et al.. Electrospray and tandem mass spectroscopy as tools for anthocyanin characterization, J. Agric. Food Chem., Al, 4657, 1999. 

Many techniques for the analysis of anthocyanins have been used for almost a century and are still of importance, along with considerable advances in technologies such as mass spectroscopy (MS) and nuclear magnetic resonance (NMR). This section summarizes the analytical procedures for quantitative and qualitative analyses of anthocyanins, including classical and modem techniques. 

Many methods are currently available for the qualitative analysis of anthocyanins including hydrolysis procedures," evaluation of spectral characteristics, mass spectroscopy (MS), " nuclear magnetic resonance (NMR), and Fourier transform infrared (FTIR) spectroscopy. - Frequently a multi-step procedure will be used for... 

Ethyl acetate is used on the C18-adsorbed anthocyanins to remove polyphenolic compounds. More efficient polyphenolic recovery will be accomplished if residual water is removed from the cartridge with a nitrogen gas stream for 2 to 3 min before application of ethyl acetate. After washing the column with ethyl acetate, water should not be added because some anthocyanins could be eluted and lost. Ethyl acetate removal of polyphenolics is particularly recommended if the anthocyanin fraction is to be subsequently analyzed by electrospray mass spectroscopy. 

Considerable advances in methods for the detection, identification and quantification of anthocyanins have been made recently, particularly with the advent of UPLC and improved mass spectroscopy methods. Additional work is needed, particularly with quantitation using MS. However, the biggest limitation in the quantitation of anthocyanins is the commercial availability of anthocyanin standards, which are nonexistent for most anthocyanins, and those that are available are fairly expensive. Some laboratories have been able to isolate and purify the particular anthocyanins of interest (Ling et ah, 2009 Nakamura et al., 2010), but most laboratories do not have the equipment, time, and/or expertise to fully validate the anthocyanin standards. 

Anthocyanins occur as salts within the cell. Extraction under very mild conditions yields genuine anthocyanins that have been studied electrophoretically. NMR spectroscopy and mass specfrometry (especially fast atom bombardment or FAB) have been extremely useful for the determination of stmctures of relatively intact anthocyanins (Brouillard, 1988). 

Due to the increasing demands of anthocyanin extracts together with the market and prize competition, blending and adulteration of berry products appears to become attractive. Consequently, detailed analysis of anthocyanins is necessary to prove the content, composition and source of anthocyanins. Traditional methods for the determination of the "total anthocyanin" content by UV/VIS spectroscopy are nowadays augmented by HPLC with UV-detection and mass spectrometry. Only the latter techniques fulfill the current need for reliable results in terms of content and identity of individual anthocyanins. Based on this information, the real versus the labeled amount of anthocyanins, the identity of the... 

Quantification of anthocyanins/anthocyanidins is usually performed either by UVA/ IS-spectroscopy or by HPLC with UV-detection. For qualification and structural elucidation HPLC coupled to mass-spectrometry is used. Other techniques include thin layer chromatography, gas chromatography and capillary zone electrophoresis (Figure 17). 

UVA/IS spectroscopy HPLC/UV (mass spectrometry) Figure 17. Steps in the analysis of anthocyanins. 

Tamura, H., Hayashi, Y., Sugisawa, H., Kondo, T., Stmcture determination of acylated anthocyanins in Muscat Bailey A grapes by homonuclear Hartmann-Hahn (HOHAHA) spectroscopy and liquid chromatography-mass spectrometry, Phytochem. Anal., 1994, 5, 190-196. 

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