Anti-angiogenic activity of Gracilaria coronopifolia J.G. Agardh extract by lowering the levels of trace metals (iron, zinc and copper) in duck chorioallantoic membrane and in vitro activation of AMP-kinase.

AMP-activated protein kinase (AMPK) is an intracellular energy sensor important in metabolic regulation, cell growth, and survival. However, the specific role of AMPK signaling pathway in the inhibition of angiogenesis remains unclear. The study highlights the activity on AMP activated protein kinase signaling pathways of a marine algae, Gracilaria coronopifolia, and its effects on angiogenesis. It was found that the most potent extract, GCD, inhibited angiogenesis significantly in the duck chorioallantoic membrane assay and also activated the enzyme AMP-kinase, in vitro. The dichloromethane extract was found most active in inhibiting angiogenesis in the duck chorioallantoic membrane (IC50 = 1.21 µg/mL) followed by GCH (IC50 = 3.08 µg/mL) (p = 0.479) and GCM (IC50 = 8.93 µg/mL) (p = 0.042). Benferroni post hoc analysis revealed that there was no significant difference between the percent inhibitions of GCH and GCM extracts (p = 0.479). Consequently, angiogenic inhibition caused lowering of iron, zinc, and copper levels in the duck CAM. Thin layer chromatography and gas chromatography-mass spectrometry revealed the components of each extracts. Notably, this is the first report on the kinase activity of a red algae G. coronopifolia extracts and a colorimetric-based quantification of angiogenesis based on metal content of CAM. Our data also suggest a novel therapeutic approach for inhibiting angiogenesis through the AMPK pathway.

Optimization using central composite design for antihistamines separation by nonaqueous capillary electrophoresis with electrochemical and electrochemiluminescence detections.

The first detailed examination of non-aqueous capillary electrophoresis with electrochemical and electrochemiluminescence detections (NACE-EC/ECL), separation parameters and their interactions via central composite design was presented. This concept was demonstrated by examining the optimization separation conditions of seven antihistamines (chlorpheniramine, cyproheptadine, diphenhydramine, doxylamine, methapyrilene, terfenadine, and triprolidine) by NACE-EC/ECL. To evaluate the NACE separation quality, the chromatography resolution statistic function (CRS(-1) function) with regard to the resolution and migration time was established as the response variable. The influences of three experimental variables (buffer apparent pH value (pH*), buffer (TBAP) concentration, and separation voltage) on the response were investigated. A set of optimal conditions was obtained from central composite design: 9.2 mM TBAP in ACN (pH* 4.0) and voltage (12.7 kV), and under these optimum conditions, the seven antihistamines could be well separated in less than 10 min. The obtained electropherograms indicated that the dual EC/ECL detection system was indispensable since the six antihistamines (except for triprolidine) displayed both EC and ECL response, whereas triprolidine only displayed the EC response. This work is instructive for investigators in simplifying the NACE-EC/ECL development procedures for multi-component analysis.