Laura Bortolotti

Detection of Honey Adulteration by Sugar Syrups Using One-Dimensional and Two-Dimensional High-Resolution Nuclear Magnetic Resonance

The importance of honey adulteration detection has recently increased owing to the limited production levels in recent years and the relative high price of honey; therefore, this illegal practice has become more and more attractive to producers. Hence, the need has arisen for more effective analytical methods aiming at detecting honey adulteration. The present research presents an effective method to detect adulteration in honey falsified by intentional addition of different concentrations of commercial sugar syrups, using one-dimensional (1D) and two-dimensional (2D) nuclear magnetic resonance (NMR) coupled with multivariate statistical analysis. Sixty-three authentic and 63 adulterated honey samples were analyzed. To prepare adulterated honeys, seven different sugar syrups normally used for nutrition of bees were used. The best discriminant model was obtained by 1D spectra, and leave-one-out cross-validation showed a predictive capacity of 95.2%. 2D NMR also furnished acceptable results (cross-validation correct classification 90.5%), although the (1)H NMR sequence is preferable because it is the simplest and fastest NMR technique.

Nectar Flavonol Rhamnosides Are Floral Markers of Acacia (Robinia pseudacacia) Honey

With the objective of finding floral markers for the determination of the botanical origin of acacia (robinia) honey, the phytochemicals present in nectar collected from Robinia pseudacacia flowers were analyzed by high-performance liquid chromatography-tandem mass spectrometry. Eight flavonoid glycosides were detected and characterized as kaempferol combinations with rhamnose and hexose. Acacia honey produced in the same location where the nectar was collected contained nectar-derived kaempferol rhamnosides. This is the first time that flavonoid glycosides have been found as honey constituents. Differences in the stability of nectar flavonoids during honey elaboration and ripening in the hive were shown to be due to hydrolytic enzymatic activity and to oxidation probably related to hydrogen peroxide (glucose-oxidase) activity. Acacia honeys contained propolis-derived flavonoid aglycones (468-4348 microg/100 g) and hydroxycinnamic acid derivatives (281-3249 microg/100 g). In addition, nectar-derived kaempferol glycosides were detected in all of the acacia honey samples analyzed (100-800 microg/100 g). These flavonoids were not detected in any of the different honey samples analyzed previously from different floral origins other than acacia. Finding flavonoid glycosides in honey related to floral origin is particularly relevant as it considerably enlarges the number of possible suitable markers to be used for the determination of the floral origin of honeys.

Influence of brood rearing temperature on honey bee development and susceptibility to poisoning by pesticides

Summary Adult honey bees (Apis mellifera) usually maintain colony brood rearing temperature between 34–35°C by thermoregulation. The brood may, however, also be subjected to suboptimal temperature. Here we investigated whether a decrease of brood rearing temperature may have effects on larval mortality, adult emergence, longevity, morphology and susceptibility to poisoning by pesticides (dimethoate). Using the in vitro rearing protocol of Aupinel (2005), we were able for the first time to control the brood temperature not only during the pupal stage but also during the larval stage. Honey bee larvae were reared in vitro at 35°C (optimal) and 33°C (suboptimal) from 12 h after hatching for 15 days. Dimethoate was tested by ingestion either on 4-day old larvae or on 7-day old adults. Our results showed that lower rearing temperature had no significant effects on larval mortality and adult emergence, but adult bee mortality was strongly affected. Moreover, adult workers emerging at 33°C were significantly more susceptible to dimethoate. Larval LD50 (48 h) was, however, 28 times higher at 33°C than at 35°C. The striking differences between larvae and adults may be explained by differential larval metabolism at 33°C and resulting slower active ingredient absorption. We conclude that adult honey bees reared at even slightly suboptimal brood temperature may be more susceptible to pesticide poisoning and be characterised by reduced longevity. Thus, low temperature brood rearing could be another stress factor for colonies.

Chemical and Functional Characterization of Italian Propolis Obtained by Different Harvesting Methods

The composition and antioxidant activity of Italian poplar propolis obtained using three harvesting methods and extracted with different solvents were evaluated. Waxes, balsams, and resins contents were determined. Flavones and flavonols, flavanones and dihydroflavonols, and total phenolics were also analyzed. To characterize the phenolic composition, the presence of 15 compounds was verified through HPLC-MS/MS. The antioxidant activity was evaluated through 1,1-diphenyl-2-picrylhydrazyl radical and reducing power assays. The ability of propolis to inhibit lipid oxidation was monitored by analyzing hydroperoxide and TBARS formation in lipids incorporated into an oil-in-water (O/W) emulsion. Acetone shows the highest extraction capacity. Wedge propolis has the highest concentration of active phenolic compounds (TP = 359.1 ± 16.3 GAEs/g; TFF = 5.83 ± 0.42%; TFD = 7.34 ± 1.8%) and seems to be the most promising for obtaining high-value propolis more suitable to prepare high-quality dietary supplements (TBARS = 0.012 ± 0.009 mmol std/g; RP = 0.77 ± 0.07 TEs/g).

Use of Quinoline Alkaloids as Markers of the Floral Origin of Chestnut Honey

To identify potential floral markers of chestnut honey, the phytochemicals present in chestnut floral nectar collected by bees were analyzed. Two nitrogen-containing compounds were detected, isolated, and identified as 4-hydroxyquinaldic acid (kynurenic acid) and 4-quinolone-2-carboxylic acid by (1)H NMR and (13)C NMR. In addition, chestnut nectar contained the monoterpene 4-(1-hydroxy-1-methylethyl)cyclohexa-1,3-diene-1-carboxylic acid, its gentiobioside ester, and the flavonol quercetin 3-pentosylhexoside. These nectar markers were found in different chestnut unifloral honey samples, although the flavonol was not detected in all samples analyzed. The terpenoid derivatives had previously been found in linden and tilia honeys. These results show that quinoline alkaloids are potentially good markers of chestnut honey, as they were not detected in any other unifloral honey analyzed so far. They are present at concentrations ranging from 34 to 65 mg/100 g of honey in the samples analyzed. In addition, the terpenoid and flavonoid derivatives present in nectar, although not exclusively characteristic of this floral origin, are good complementary markers for the determination of the floral origin of chestnut honey.

The Status of Honey Bee Health in Italy: Results from the Nationwide Bee Monitoring Network

In Italy a nation-wide monitoring network was established in 2009 in response to significant honey bee colony mortality reported during 2008. The network comprised of approximately 100 apiaries located across Italy. Colonies were sampled four times per year, in order to assess the health status and to collect samples for pathogen, chemical and pollen analyses. The prevalence of Nosema ceranae ranged, on average, from 47–69% in 2009 and from 30–60% in 2010, with strong seasonal variation. Virus prevalence was higher in 2010 than in 2009. The most widespread viruses were BQCV, DWV and SBV. The most frequent pesticides in all hive contents were organophosphates and pyrethroids such as coumaphos and tau-fluvalinate. Beeswax was the most frequently contaminated hive product, with 40% of samples positive and 13% having multiple residues, while 27% of bee-bread and 12% of honey bee samples were contaminated. Colony losses in 2009/10 were on average 19%, with no major differences between regions of Italy. In 2009, the presence of DWV in autumn was positively correlated with colony losses. Similarly, hive mortality was higher in BQCV infected colonies in the first and second visits of the year. In 2010, colony losses were significantly related to the presence of pesticides in honey bees during the second sampling period. Honey bee exposure to poisons in spring could have a negative impact at the colony level, contributing to increase colony mortality during the beekeeping season. In both 2009 and 2010, colony mortality rates were positively related to the percentage of agricultural land surrounding apiaries, supporting the importance of land use for honey bee health.