Study On Functional Lipids In Walnut Oil And Their Efficacy

Walnut belongs to the genus of Juglans regia,cultivated with a long history and distributed widely in China.It is an important economic crop in Shandong,Hebei and Shanxi,and has a great production.The lipids content of walnut reaches to 60%-70%.Walnut oil is rich in unsaturated fatty acids,phytosterols,vitamin E and other pharmacological efficacy ingredients,which contains the pharmacological activities of improving memory,anti-oxidation,anti-tumor and so on.Unsaturated fatty acids play an important role in walnut oil.To study whether the method of preparation have an effect on the content and efficacy,in this paper,walnut oil,which was prepared by cold pressing,aqueous enzymatic extraction,soxhlet extraction,respectively,was selected as the object,and the research was carried as follows:First,the method of HPLC-ELSD for the determination of fatty acids in walnut oil was established.After being optimized,the chromatographic conditions were determined as:with Eclipse XDB-C18(4.6×150,5?m)column,mobile phase:methanol/water(containing 0.1%acetic acid)(83:17,v/v),column temperature:30 ?,flow rate:1.0mL.min-1,temperature of ELSD:70 ?,N2 flow rate:2.0mL.min-1.Method validation was carried out,and results showed that the linearity,precision,stability,reproducibility and recovery of the method were good.And the molecular structure of each fatty acid was identified by HPLC-MS,then the composition of fatty acid was determined.Based on the above method,the content of fatty acids in walnut oil was determined quantitatively.The saponification conditions of walnut oil samples were investigated through single factor,and the content of total fatty acids was selected as index.The optimum saponification conditions were as follows:the concentration of KOH-ethanol solution was 1.0mol.L-1,and the time of saponification was 40min at 70 ?.The content of fatty acids and total fatty acids in the walnut oil which was prepared by different methods were compared under the optimum saponification conditions,and the results showed that both of which in aqueous enzyme-extracted walnut oil were higher than other two methods.We explored the nuclear magnetic resonance(NMR)spectroscopy for the determination of the content of fatty acids in walnut oil,and compared the content with HPLC.The result showed that there was no significant difference between them,and confirmed the reliability of NMR.And then,we established a method for the determination of triglycerides with UV detector.The chromatographic conditions were determined as follows:with Agilent XDB-C18(4.6×250mm,5?m)column,mobile phase:acetonitrile/isopropanol/n-hexane(66:20:14,v/v/v),flow rate:0.8mL.min-1,temperature of column:20 ?,wavelength of UVD:210nm.Under these conditions,components were all well separated.After that,three walnut oils were characterized by infrared spectroscopy(IR).According to the characteristic absorption peaks of each group in the IR spectrum,the infrared spectrum for quality control of walnut oil was preliminarily established.Finally,Morris-water maze and shuttle box method were used to compare the effect of walnut oils prepared by different methods on the ability of learning and memory of mice.ICR mice were selected as the test object,fed with different dosages of enzyme-extracted walnut oil(low,medium,high),walnut oil-?-cyclodextrin microcapsules(medium)and cold-pressed walnut oil(medium)successively.After 4 weeks,the tests were carried out.The result of Morris-water maze showed that the escape latency in the low and high dosage of enzyme-extracted walnut oil groups was significantly lower(P<0.05)than that in normal control group,and was lower than that in positive control group,too.And the number of collapse and residence time in the target area of low and high dosage of enzyme-extracted walnut oil groups were higher than control group.The result of shuttle box experiment showed that the rate of active avoidance in the high dose of enzyme-extracted walnut oil was higher than normal control group and other groups(P<0.05),and the time of passive avoidance was shortened significantly(P<0.05).It was indicated that the improvement of learning and memory ability of enzyme-extracted walnut oil was better than cold-pressed walnut oil.