New α-glucosidase inhibiting anthracenone from the barks of Harungana madagascariensis Lam

Abstract Two new 10-hydroxy-9(10H)-anthracenone, madagascenone A (1) and B (2), were isolated from the barks of Harungana madagascariensis Lam. The structures of the compounds were determined using 1D- and 2D-NMR and mass spectroscopic techniques. Both of the compounds showed an in vitro α-glucosidase inhibition with IC50 = 69.9 ± 4.21 and 122.3 ± 1.13 μM, respectively, more potent than the standard acarbose (IC50 = 840 ± 1.23 μM).

α-Glucosidase, an enzyme secreted from the epithelium of intestine, catalyses the hydrolysis of polysaccharides to monosaccharide. Inhibition of α-glucosidase interferes in the digestion of carbohydrate and ultimately decreases the risk of postprandial hyperglycaemia in diabetic patients. 1-Deoxynojirimycin, acarbose and miglitol have been developed as α-glucosidase inhibitors for clinical use. However, use of acarbose cause gastrointestinal tract disturbance and there is a need to develop the safe and effective compounds to manage hyperglycaemia of the diabetes (Choudhary et al. 2011). Many natural products have been reported as α-glucosidase inhibitors from marine sponges (Ramanjooloo et al. 2015), sesquiterpenes and flavonoids from plant Zingiber zerumbet (Ajish et al. 2015), from guggul and resin (El-Mekkawy et al. 2013), and many other secondary metabolites showed excellent α-glucosidase inhibition activity (Salah El Dine et al. 2014;Sun et al. 2014).
In continuation of our search of new bioactive compounds from different plant source of Nigeria, we isolated two new α-glucosidase inhibiting anthranols. The spectroscopic data and biological activities of aforementioned compounds are presented here.
Compounds 1 and 2 were tested against α-glucosidase enzyme and found to be potently active of 97.1 and 97.9% of inhibition, respectively, with reference to standard acarbose (59.1%) ( Table 1). Both of the compounds are more potent than standard and this research verified the use of H. madagascariensis by local people in Nigeria to treat diabetes. Compounds 1 and 2 may be lead compounds to develop medicine for decreasing the risk of postprandial hyperglycaemia in diabetic patients (Figure 1).

General experimental conditions
JASCO DIP-360 digital polarimeter was used for the measurement of optical rotations. The infrared (IR) spectra were recorded on JASCO A-302 IR spectrophotometer. For ultraviolet (UV) spectra, Thermo Evolution-300 spectrophotometer was used. 1D-and 2D-NMR spectra were recorded on a 500 MHz on Bruker Avance-500 nuclear magnetic resonance spectrometer. The low-resolution EI was performed on Finnigan MAT 311 mass spectrometer. The HRFAB-MS was recorded on the Finnigan MAT 95 XP mass spectrometer. The column chromatography was performed on silica gel (230-400 mesh, E. Merck), while thin-layer chromatography (TLC) was carried out on pre-coated preparative silica gel plates, GF-254 (20 × 20 cm, 0.5 mm thick, E-Merck).

Plant material
The stem back of H. madagascariensis (Haronga) was collected from Idi-Ayunre in Ibadan, Oyo State, Nigeria in 2010. The plant was identified by Mr D.S.P. Esimekhuai, Botanist, Herbarium of the Department of Botany, University of Ibadan, Ibadan, Nigeria. A voucher specimen

α-Glucosidase inhibition assay
The α-glucosidase inhibition assay was performed according to the slightly modified method of Oki et al. (1999). α-Glucosidase (E.C.3.2.1.20) from Saccharomyces sp. was purchased from Wako Pure Chemical Industries Ltd. (Wako 076-02841), while other chemicals were purchased from various commercial sources. The inhibition was measured spectrophotometrically at pH 6.9 and at 37 °C using 0.5 íM p-nitrophenyl α-d-glucopyranoside (PNP-G) as a substrate and 250 m units/mL of enzyme, in 50 mM sodium phosphate buffer containing 100 mM NaCl. Acarbose (0.78 mM) was used as positive controls. The increment in absorption at 400 nm, due to the hydrolysis of PNP-G by α-glucosidase, was monitored continuously with a spectrophotometer (Spectra Max, Molecular Devices CA, USA) (Choudhary et al. 2011).

Conclusion
This research work concluded the finding of the two new potent α-glucosidase enzyme inhibitors, which may be the lead compounds to develop medicine for decreasing the risk of postprandial hyperglycaemia in diabetic patients

Supplementary material
NMR table, HMBC figure, NOESY figure, and mass and NMR spectra of compounds 1 and 2 are available online.