Pollen is the unique source of all the nutrients that bees require for healthy growth. They use it to feed themselves as well as the larvae in the hive and without it, the hive would perish. Bees collect pollen from flowers and mix it with a sticky secretion from their stomachs, so that it can be stored in pollen baskets on their legs for transport to the hive.
Bee pollen is a mixture of plant pollen, bee saliva and nectar and is rich in proteins, carbohydrates, amino acids, pigments, vitamins and minerals. The actual composition and proportions of the various components of a particular pollen, like honey, are related to the distribution of the source plants. These components fix the composition so that quality control standards can be established and subsequent adulteration of the products can be detected.
This is an important factor because bee pollen, just like honey, has been used for millennia as a dietary supplement by humans. Many declarations have been made regarding the health benefits but, tellingly, the US FDA does not allow pollen marketers in the United States to make health claims, as there is currently no scientific basis for these.
The pigments present in bee pollen, consisting primarily of flavonoids and anthocyanins, account for the variation in colour of pollen and honey. However, other non-coloured flavonoids can also be present. and their presence in one particular bee pollen has attracted the attention of Iberian researchers.
Federico Ferreres from the University of Murcia at Espinardo in Spain and David Pereira, Patricia Valentao and Paula Andrade from Porto University, Portugal, became interested in bee pollen from the plant Echium plantagineum, also known as Paterson's curse. It is an invasive weed found in southern Europe and its bee pollen has been extensively studied. However, no work has been carried out on the phenolic compounds, including flavonoids, which might be present.
The researchers analysed bee pollen from central Portugal which had been confirmed to originate from E. plantagineum. An acidic methanol extract was subjected to HPLC with two detectors, a photodiode array detector and a mass spectrometer. HPLC is an important technique for separating the complex mixtures of flavonoids present in food extracts, especially when limited supplies of the sample prevent ready isolation and purification of the components.
The extract was separated on a reversed-phase octadecylsilica column operating at room temperature, with a mobile phase gradient of methanol in aqueous acetic acid. Twelve flavonoids eluting within 30 minutes were detected at a wavelength of 350 nm. Since the visible range covers 390-750 nm, these flavonoids were colourless.
The compounds were glycosides of quercetin, isorhamnetin and kaempferol but HPLC alone was not sufficient to differentiate between them. The HPLC eluent was directed to an ion trap mass spectrometer operating in negative-ion electrospray ionisation mode for MS/MS and MS/MS/MS experiments. This was essential to identify the various isomers having the same flavonoid core but different sugar chains.
The mass spectral fragmentation patterns characterised the linkages between the glycoside groups to allow their identification. In this way, isomeric kaempferol 3-O-neohesperidoside and 3-O-rutinoside were distinguished, as were several other isomeric pairs.
The research team declared that this is the first time that non-coloured flavonoids have been reported from this pollen and the value of HPLC for isomer separation and mass spectrometry for isomer identification has been illustrated.
The results expand the phytochemical knowledge of bee pollen from E. plantagineum. They will be useful for the quality control of commercial dietary supplements prepared from this bee pollen and for detecting their adulteration with other substances, which might be cheaper pollens from other sources or even non-pollen matter.
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* Rapid Communications in Mass Spectrometry 2010, 24, 801-806: "First report of non-coloured flavonoids in Echium plantagineum bee pollen: differentiation of isomers by liquid chromatography/ion trap mass spectrometry"
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