Phenolic quantification and botanical origin of Portuguese propolis

Highlights

• The phenolic quantification of Portuguese propolis was achieved by HPLC.

• The phenolic composition of Populus x Canadensis Moench buds and Cistus ladanifer L. resin was evaluated by LC–MS/MS.

• Populus spp. and Cistus ladanifer were establish as two main floral sources for Portuguese propolis.

• Kaempferol-3,7-dimethyl-ether is a possible marker compound for discrimination of for Cistus propolis.

Abstract

The production of propolis by honeybees results from a selective collection of exudates from leaf buds and plants present in the hive neighborhood leading to a resin with many potentialities in the pharmaceutical industry. This study aims to quantify the phenolic content in propolis from different Portuguese regions and in the potential floral sources, Populus x Canadensis Moench buds and Cistus ladanifer L., in order to establish links with geographical and botanical origin.

The Portuguese propolis revealed a phenolic profile with marked differences in concentrations: the richness in flavonoids is common in all regions, but more evident in propolis from central interior, south and Madeira. The composition of poplar type propolis common in temperate zones was observed in the north, central coast and Azores, while the central interior and south samples, with a composition rich in kaempferol derivatives, resemble the C. ladanifer exudates, a spontaneous bush widespread in the Mediterranean. The compound kaempferol-3,7-dimethyl-ether, absent in the poplar type propolis, can be regard as a possible marker for the discrimination of these two types of propolis.

Introduction

The honeybee exploit nature as a store to fulfill its nutritional needs, collecting nectar and pollen, but also as a source for substances with other goals such as construction material or to maintain the antiseptic environment in the hive (Burdock, 1998, Bogdanov and Bankova, 2011). With the time evolution, bees were able to find in the surroundings of their nest the best source of materials for the desired proposes. Thus, it is not surprising the potentialities exhibit by propolis, a complex natural product gathered by the honeybees from resinous exudates of buds, leaves, branches and barks present in the vicinity of the beehive. Also named as the “bee glue” it plays an important role to guarantee the bee colony health.

Since ancient times, propolis is used in traditional medicine and now is gaining popularity in health foods as well in cosmetic products (Bogdanov and Bankova, 2011). A great number of research studies focused on the pharmacological and biological properties present by propolis, including antihepatotoxic, antitumor, antioxidative, antimicrobial, anti-inflammatory and immunomodulatory among others (Banskota et al., 2001, Bankova et al., 2000). These bioactivities are closely linked with the chemical composition, particularly with the richness in phenolic compounds, which accounts for approximately half of the resin content, while beeswax, volatiles and pollen represents the other 30%, 10% and 5%, respectively, of it. (Bogdanov and Bankova, 2011). The propolis chemical composition varies greatly with the plant origin of the resin and thus with the geographic and climatic characteristics of the site (Bankova, 2005). The specificity of local flora is very important, not all plants are resin providers and bees have a marked preference for one or a few sticky resin sources, which are, at the same time sources of biologically active phytochemicals (Salatino et al., 2011). Based on this knowledge, propolis was typified according to their plant origin and its main chemical constituents (Bankova, 2005). In temperate zones of the world, poplar buds (Populus spp.) are the main sources of the bee glue with flavones, flavanones, fenolic acids and their esters as major compounds. Exceptions can be found, for example, the birch propolis type found in Russia, which has its origin in species like Betula verrucosa, where the main compounds are flavones and flavonols different from those found in poplar propolis (Bankova, 2005). Also a Mediterranean propolis type was found in Sicily, Crete and Malta, whose main compounds are diterpenes most probably originated in coniferous plant of the genus Cupressaceae (Popova et al., 2009). Tropical propolis has a totally different compositional pattern: the green propolis type, found in Brazil, has its main plant source on the leaves of Baccharis spp. and mainly contains prenylated phenylpropanoids (Bankova et al., 2005). In Venezuela and Cuba, the main plant sources are the flower exudates of Clusia species, originating a propolis rich in prenylated benzophenones (Bankova et al., 2005). C-prenylflavonoids (or propolins) have been described in propolis from Pacific islands, where the resin sources are the fruit exudates of the tree Macaranga tanarius (Chen et al., 2003). The propolis typification on the basis of plant sources knowledge is a useful tool for its chemical standardization and thus for ensuring the quality and safety necessary for its commercialization (Bankova, 2005, Salatino et al., 2011).

Recently, the phenolic profile of Portuguese propolis was characterized by liquid chromatography with diode-array detection coupled to electrospray ionization tandem mass spectrometry (LC/DAD/ESI-MSⁿ) (Falcão et al., 2013). Forty samples from different continental regions and islands were analyzed allowing the detection of seventy six polyphenols and the establishment of two different propolis groups: the common temperate propolis, which contained the typical poplar phenolic compounds such as flavonoids and their methylated/esterified forms, phenylpropanoid acids and their esters and an uncommon propolis type with an unusual composition in quercetin and kaempferol glycosides, some of them never described in propolis. The data suggest a diversified botanical origin for the Portuguese propolis besides poplar buds (Falcão et al., 2013).

Following those finds, we now present the phenolic quantification of propolis from the different Portuguese continental regions and islands. The results assort the compounds with major contribution to the propolis composition and allow the establishment of links with the geographical origin of this beehive product, a key factor for propolis commercial valorization. The inclusion of two potential floral sources of Portuguese propolis in this study, the buds exudates and surface material present on the leaves and stems of Populus x Canadensis, male and female specimens and Cistus ladanifer L. enable the correlation between the phenolic profile and the plant source of the resin, and the proposal of kaempferol-3,7-dimethyl-ether as a marker compound for C. ladanifer propolis.