In Vitro Antioxidant Activities of Annona squamosa Leaf and Identification of Bioactives by LC-ESI-MS-Qtof

Annona squamosa Linn. (Family; Annonaceae) commonly known as custard apple. A bark and root  is used to prevent diarrhea and dysentery. A decoction of the leaves is used for cold and also recognized for several medicinal properties. The present study reports the antioxidant potential of various fractions isolated from leaves of custard apple and their  chemical constituents were identified by liquid chromatography with high resolution mass spectral analysis. Dried leaf powder was successively extracted with various solvents including hexane, ethyl acetate, acetone, methanol, and water by increasing polarity at 60 to 80 °C. All lyophilized extracts exhibited wide range of free radical scavenging (DPPH and ABTS) activity and antioxidant capacity by phosphomolybdenum method. Acetone extract showed the highest  antioxidant activity (73% to 88% at 35 ppm) in DPPH and ABTS methods among tested fractions. The same trend was observed in phospomolybedum method.  The degree of radical scavenging activity was comparable to total phenolics present in each extract. Further, the acetone fraction was separated on C18 column with gradient mobile phase and analyzed for their active constituents by rapid and high sensitive liquid chromatography-electrospray ionization-quadrupole-time of flight-mass spectrometry (LC-ESI-Q-TOF-MS). We have identified two compounds using accurate mass spectral data as methy armepavine and xylopine. The results of the present study provide the use of best solvent to isolate antioxidant rich fractions which might provide potential biological properties.  Further studies are needed to establish effectiveness of the health benefits in in vivo models. This project is based upon work supported by the USDA–NIFA # 2010-34402-20875 "Designing Foods for Health" through the Vegetable & Fruit Improvement Center.