Tetragonisca angustula honey was fractioned in a SiO 2 column to furnish three fractions (A-C) in which four hydroxycinnamic acid-Spermidine amides (HCAAs), known as N′, N″, N‴-tris-p-coumaroyl spermidine, N′, N″-dicaffeoyl, N‴-coumaroyl spermidine, N′, N″, N‴-tris-caffeoyl spermidine and N′, N″dicaffeoyl and N‴-feruloyl spermidine were identified in the fractions B and C by electrospray ionization tandem mass spectrometry. A primary culture model previously infected with Neospora caninum (72 h) was used to evaluate the honey fractions (A-C) for two-time intervals: 24 and 72 h. Parasitic reduction ranged from 38% on fraction C (12.5 µg/ml), after 24 h, to 54% and 41% with fractions B and C (25 µg/ml) after 72 h of treatment, respectively. Additionally, HCAAs did not show any cell toxicity for 24 and 72 h. For infected cultures (72 h), the active fractions B (12.5 µg/ml) and C (25 µg/ml) decreased their NO content. In silico studies suggest that HCAAs may affect the parasite's redox pathway and improve the oxidative effect of NO released from infected cells. Here, we presented for the first time, that HCAAs from T. angustula honey have the potential to inhibit the growth of N. caninum protozoa. K E Y W O R D S hydroxycinnamic acid-spermidine amides, immune response, Neospora caninum, nitric oxide, nitrogenous compounds, Tetragonisca angustula honey | 1105 LIMA et al.

The antioxidant activity of Tetragonisca angustula honey (TAH) and its ethanolic extract (TAEE) were investigated. The total levels of phenolic (TPC) and flavonoids (TFC) were also evaluated. The results for TPC were 19.91 ± 0.38 and 29.37 ± 1.82 mg GAE g-1 and for TFC 0.20 ± 0.02 and 0.14 ± 0.01 mg QE g-1 of TAH and TAEE, respectively. Antioxidant activities were 73.29 ± 0.49% and 93.36 ± 0.27% in the DPPH● assay and 71.73 ± 4.07% and 97.86 ± 0.35% in ABTS●+ for TAH and TAEE, respectively. The total reducing activity was determined by the method of reducing power (PR) and iron ion (Fe III) and the results varied in PR from 151.7 ± 25.7 and 230.7 ± 25.2 mg GAE L-1, for TAH and TAEE respectively and for (Fe III) in EC50 0.284 in TAEE and 0.687 in TAH. Chemical analysis by HPLC-DAD of the ethanolic extract (TAEE) revealed the presence of ferulic acid as majority phenolic component in the extract. The 1H NMR analysis confirmed this structure and showed the also presence of glucose, citric acid, succinic acid, proline and hydrocarbon derivatives. In addition, the botanical origin was also investigated and showed a multifloral characteristic, having found 19 pollen types with a botanical predominance of the Anacardiaceae family, with Tapirira pollen occurring as predominant (42.6%) and Schinus as secondary (25.7%). The results showed that T. angustula honey is an interesting source of antioxidant phenolic compounds due to its floral origin and can act as a protector of human health when consumed.

Neosporosis is a disease caused by Neospora caninum sp (JP Dubey), an intracellular parasite that affects many animal species, fostering abortions and neurological disorders. Recent studies on Neospora caninum have shown that glial cells have been considered a model of in vitro infections of these protozoa. Honey has been used since ancient times for its anti-inflammatory and antimicrobial properties. The present study aimed to evaluate the in vitro reactivity of glial cells (astrocytes and microglia) infected with N. caninum treated with Jataí honey at 1% (Tetragonisca angustula). After 72 hours from infection, mitochondrial metabolism, dehydrogenase lactate activity (LDH), nitric oxide production (NO) and the number of parasites were checked. Cultures treated with honey at 1% have shown mitochondrial metabolism and cell permeability increase. There was an increase in nitrite production, possible indicating a microglial activation. Nitric Oxide values found in this study may indicate an immune response against Neospora caninum tachyzoites, since such increase may contribute to reduce the number of tachyzoites. So, our results suggest that honey had a protective effect over astrocytes and microglia cultures infected with Neospora caninum.