Research On The Mechanism Of Ammonium Sulfate Precipitating Limonin In Pomelo Juice And The Water Extract Of Pomelo Peel

Limonin is a compound of highly oxygenated tetracyclic triterpenoids present in plants of Rutaceae and Meliaceae,which possesses a variety of biological activity such as antioxidant,antibacterial,anticancer and larvicidal,and has good application value.In our previous study,it has been observed ammonium sulfate could precipitate limonin in pomelo juice.On the basis of this phenomenon,limonin precipitation was prepared from pomelo juice and water extract of pomelo peel,purified and identified to research the mechanism of limonin existing in aqueous solution and precipitated by ammonium sulfate.Main results are as follows:(1)The optimal condition of preparing limonin precipitation from pomelo juice was:pH 5,10%saturation of ammonium sulfate,reaction temperature 90 ~oC.The purity of precipitation was80.2%after washed by hot water of 45 ~oC.Using acetonitrile and water as solvent system,limonin precipitation was crystallized at room temperature,and the crystal of 98.2%purity was obtained.(2)The condition to extract limonin by hot water from pomelo peel powder was optimized as follows:liquid-to-solid ratio of 20 mL/g,extraction temperature 80oC and time for 30 min.The best condition to precipitate limonin from water extract of pomelo peel was:reaction temperature 50 ~oC,reaction time for 90 min,and 90%saturation of ammonium sulfate.After purification by macroporous adsorption resin D101,extraction and crystallization,limonin crystal of 96.4%purity was gained.(3)Acid and pectin promote the dissolution of limonin in aqueous solution.The acid can make limonin partially hydrolyze to the form of soluble hydroxyl acid;pectin can combine with limonin molecule,or limonin wrap in reticular structure of pectin,which promote limonin stability existing in pomelo juice and the water extract of pomelo peel.(4)The limonin crystal from pomelo juice and water extract of pomelo peel was analysed by nuclear magnetic resonance and crystal diffraction.The results showed that crystal did not contain ammonium and sulphate ions,which indicated ammonium sulfate did not combine with limonin molecule.In addition,limonin precipitation was prepared by limonin standard reacting with ammonium sulfate aqueous solution;after analysed with infrared spectroscopy,it was confirmed that ammonium sulphate did not combine with limonin molecule.(5)The ability of hydrogen,sulphate and ammonium ions to precipitate limonin was researched,and it was found that sulphate ion played a major role in the process of ammonium sulfate precipitating limonin.The mechanism of ammonium sulfate precipitating limonin is sulphate ion destroys the combination of hydroxy acid limonin and water,the combination of limonin with pectin,and the combination of pectin with water,which cause the precipitation of limonin.This paper found that ammonium sulfate could precipitate limonin in pomelo juice and the water extract of pomelo peel,then preliminary illustrated the mechanism of limonin existing in juice and the water extract.The research provides some references to further process and utilize citrus fruits.