“Bee pollen” is pollen collected from flowers by honey bees. It is used by the bees to nourish themselves, mainly by providing royal jelly and brood food, but it is also used for human nutrition. For the latter purpose, it is collected at the hive entrance as pellets that the bees bring to the hive. Bee pollen has diverse bioactivities, and thus has been used as a health food, and even as medication in some countries. In this paper, we provide standard methods for carrying out research on bee pollen. First, we introduce a method for the production and storage of bee pollen which assures quality of the product. Routine methods are then provided for the identification of the pollen’s floral sources, and determination of the more important quality criteria such as water content and content of proteins, carbohydrates, fatty acids, vitamins, alkaloids, phenolic and polyphenolic compounds. Finally, methods are described for the determination of some important bioactivities of bee pollen such as its antioxidant, anti-inflammatory, antimicrobial and antimutagenic properties. Métodos estándar Para la investigación del polen El "polen de abeja" es el polen recogido de las flores por las abejas melíferas. El polen de abeja es utilizado para nutrir a las propias abejas, principalmente para proporcionar jalea real y alimento para las crías, pero también se utiliza para la nutrición humana. Para este último fin, se recoge en la entrada de la colmena en forma de gránulos que las abejas llevan a la colmena. El polen de abeja tiene diversas bioactividades, por lo que se hautilizado como alimento para la salud, e incluso como medicamento en algunos países. En este artículo, proporcionamos métodos estándar para llevar a cabo investigaciones sobre el polen de abeja. En primer lugar, presentamos un método de producción y almacenamiento de polen de abeja que garantiza la calidad del producto. A continuación, se ofrecen métodos de rutina para la identificación de las fuentes florales del polen y la determinación de los criterios de calidad más importantes, como el contenido de agua y de proteínas, carbohidratos, ácidos grasos, vitaminas, alcaloides y compuestos fenólicos y polifenólicos. Por último, se describen métodos para la determinación de algunas bioactividades importantes del polen de abeja, como sus propiedades antioxidantes, antiinflamatorias, antimicrobianas y antimutagénicas.

Propolis is a “natural” remedy with prominent biological activity, which is used as dietary supplement. In the absence of clinical studies that would substantiate these claims, information on the biological activity of propolis is valuable. This study comprises chromatographic, image processing and chemometric approach for phenolic profiling of Serbian, Croatian and Slovenian propolis test solutions. Modern thin-layer chromatography equipment in combination with software for image processing was applied for fingerprinting and data acquisition, whereas the principal component analysis was used as pattern recognition method. Characterization of phenolic profile was performed along with the determination of the botanical and geographical origin of propolis. High-performance thin-layer chromatograms reveal that Central and Southeastern European propolis samples are rich in flavonoids. In addition, phenolic compounds proved to be suitable markers for the determination of European propolis authenticity.

Information on compositional, nutritional and functional properties of bee-pollen, as a health-promoting food, is essential for defining its quality. Concerning the nutritional importance of phenolic compounds, the aim of this study was to determine the phenolic profile and antioxidant activity of twenty-four bee-pollen samples collected from different regions of Serbia. High-performance thin-layer chromatographic (HPTLC) fingerprinting was used for profiling of bee-pollen samples according to the botanical type. HPTLC hyphenated with image analysis and a pattern recognition technique confirmed the grouping of samples caused by the specific phenolic composition of pollens of different botanical origin. Flavonoid glycosides in bee-pollen samples were identified by applying ultra-high-performance liquid chromatography (UHPLC) coupled with linear ion trap-Orbitrap mass spectrometry (LTQ Orbitrap MS). Eight out of twenty-seven flavonol glycosides were identified in bee-pollen samples for the first time. All analyzed bee-pollen samples showed a high number of phenolic compounds which may have therapeutic potential.

To replace common organic solvents that present inherent toxicity and have high volatility and to improve the extraction efficiency, a range of natural deep eutectic solvents (NADESs) were evaluated for the extraction of phenolic compounds from Agrimonia eupatoria. Screening of NADES efficiency was carried out based on the total phenolic and flavonoid content and radical-scavenging activity, determined by spectrophotometry, as well as phenolic compounds quantified, obtained using ultra-high-performance liquid chromatography with a diode array detector and a triple-quadrupole mass spectrometer. Increased extraction efficiency when compared with organic solvent was achieved using NADES mixtures choline chloride (ChCl):urea 1:2 and choline chloride:glycerol 1:1. Flavonol glycosides were the most abundant compounds in all extracts. The COSMO-RS model provided insights into the most important intermolecular interactions that drive the extraction process. Moreover, it could explain the extraction efficiency of flavonol glycosides using ChCl:glycerol NADES. The current article offers experimental evidence and mechanistic insights for the selection of optimal NADES to extract bioactive components from Agrimonia eupatoria.