Bee Behavior
Numerous honeybees were seen on alecrim plants fragmenting vegetative apices (bud, leaf primordia and young leaves). The fragments in the mandibles have sticky aspect from the beginning of the collecting process, indicating liberation of resinous substances from trichomes and ducts. The bees manipulate the fragments, a mass of resinous material being the final product. Using the first pair of legs, the bees move the resin mass to the median legs and then to the opposite corbicula (Fig. 1). The resin masses may be transferred to the corbicula of the same side. Very rarely the material is transferred directly from the first pair of legs to the corbiculae.
The time spent from the beginning of the collecting process to the deposit of the resin mass in the corbiculae was, on average, 7 min. The frequency of visits varied, depending on several factors, among which was the hive's demand for propolis.
Histology of Resin Masses and Propolis
Figure 2 evidences the high degree of fragmentation of the material in the resin mass. Some details are relevant as diagnostic of the resin plant source. Resiniferous ducts are conspicuous in histological sections of leaves of Baccharis. Several ducts are apparent in Fig. 2, which shows a fragment of a young alecrim leaf from a resin mass. Another leaf fragment from a resin mass with a resiniferous duct in cross section is shown in Fig. 3; glandular and non-glandular trichomes are also visible. In Fig. 4, several leaf fragments in the residue of the propolis sample are seen, most with conspicuous ducts. Glandular and non-glandular trichomes of alecrim leaves were commonly seen. Viewed with polarized light, alecrim glandular trichomes show brilliant contents, which correspond to starch grains (Fig. 5) and were detected in histological preparations of green propolis (Fig. 6). Figures 2–5 evidence that the fragments correspond to young plant leaves, because the chlorenchyma in all cases is still undifferentiated, no palisade and spongy parenchyma being apparent.
Slides prepared from the propolis samples showed no fragments of plants other than alecrim. Hence the strategy of choosing a time of intense vegetative alecrim growth, the use of hive boxes with slits to promote propolis production and the restriction of such production to a short time was successful to attain a product from a single plant source.
Chemical Analyses
Tables 1–4 list the substances identified in shoot apices of female and male alecrim plants, in the resin masses and in the pooled propolis sample. Table 1 refers to phenolic compounds, Table 2 to terpenoids (sesqui, di and pentacyclic triterpenoids), Table 3 to waxy substances, and Table 4 to compounds from other classes. A total of 64 substances were identified, but other compounds (probably unknown) were also detected. Prenylated (compounds 10–17) and non-prenylated (compounds 1–9) cinnamic acid derivatives were often detected.
Among the identified substances, 42 were detected in male and 33 in female apices, 29 in the resin masses and 34 in the propolis sample. No exact match is observed in the distribution of compounds among the four resin sources. A higher score of coincidences is observed comparing resin masses and propolis (23 matches), most substances corresponding to phenolics, some of them common in propolis, such as derivatives of cinnamic acids. There are 17 matches comparing resin masses either with male, female or both apices, 13 comparing propolis/apices and 12 comparing all three sources.
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