Determination of Dicyclohexylamine in Beeswax by Aqueous Normal Phase Liquid Chromatography Coupled With Tandem Mass Spectrometry.
Dicyclohexylamine (DCH) is an excipient present in commercial formulations of fumagillin (Fumagilin-B® and Fumidil-B®), an antibiotic which is employed in apiculture for the control of nosema disease. DCH has been demonstrated to be stable in harvested honey; however, its fate in the beehive has not been investigated. In this study, DCH residues were determined in beeswax samples collected from brood chambers and honey supers from various apiaries throughout Alberta. The determination of DCH was performed using a simple extraction procedure followed by low-temperature cleanup and finally analysis with aqueous normal phase liquid chromatography-electrospray ionization tandem mass spectrometry. The method was found to yield high accuracy and precision (94.9 ± 1.7% to 110.8 ± 6.6%) for replicate beeswax samples spiked with 4, 80, 360 and 2,000 μg kg-1 of DCH. With a limit of quantification of 1 μg kg-1, 100% of the 61 beeswax samples analyzed were found to contain residues of DCH ranging from 15 to 6,410 μg kg-1. The results indicate that DCH can accumulate in beeswax when Fumagilin-B® or Fumidil-B® is applied in apiculture and that these concentrations can vary significantly among beekeeping operations.
In Vivo Inhibitory Assessment of Potential Antifungal Agents on Nosema ceranae Proliferation in Honey Bees
Nosema ceranae Fries, 1996, causes contagious fungal nosemosis disease in managed honey bees, Apis mellifera L. It is associated around the world with winter losses and colony collapse disorder. We used a laboratory in vivo screening assay to test curcumin, fenbendazole, nitrofurazone and ornidazole against N. ceranae in honey bees to identify novel compounds with anti-nosemosis activity compared to the commercially available medication Fumagilin-B®. Over a 20-day period, Nosema-inoculated bees in Plexiglas cages were orally treated with subsequent dilutions of candidate compounds, or Fumagilin-B® at the recommended dose, with three replicates per treatment. Outcomes indicated that fenbendazole suppressed Nosema spore proliferation, resulting in lower spore abundance in live bees (0.36 ± 1.18 million spores per bee) and dead bees (0.03 ± 0.25 million spores per bee), in comparison to Fumagilin-B®-treated live bees (3.21 ± 2.19 million spores per bee) and dead bees (3.5 ± 0.6 million spores per bee). Our findings suggest that Fumagilin-B® at the recommended dose suppressed Nosema. However, it was also likely responsible for killing Nosema-infected bees (24% mortality). Bees treated with fenbendazole experienced a greater survival probability (71%), followed by ornidazole (69%), compared to Nosema-infected non-treated control bees (20%). This research revealed that among screened compounds, fenbendazole, along with ornidazole, has potential effective antifungal activities against N. ceranae in a controlled laboratory environment.
Synthesis and antiproliferative activity of ligerin and new fumagillin analogs against osteosarcoma.
Ligerin (1) is a natural chlorinated merosesquiterpenoid related to fumagillin (2) exhibiting a selective antiproliferative activity against osteosarcoma cell lines and an in vivo antitumor activity in a murine model. Semisynthesis of ligerin analogs was performed in order to study the effects of the C3-spiroepoxide substitution by a halogenated moiety together with the modulation of the C6 chain. Results showed that all derivatives exhibited an in vitro antiproliferative activity against osteosarcoma cell lines and that chlorohydrin compounds were equally or more active than their spiroepoxy analogs. Among semisynthetic analogs, the parent compound 1 was the best candidate for further studies since it exhibited higher or equivalent activity compared to TNP470 (3) against SaOS2 and MG63 human osteosarcoma cells with a four times weaker toxicity against HFF2 human fibroblasts. Quantitative videomicroscopy analysis was conducted and allowed a better understanding of the mechanism of its antiproliferative activity.