| China is the largest donkey breeding country in the world and has a donkey breeding history of nearly4000years. The donkey fat is one of the common edible animal fats in North China. But until now, the basic research and functional study of donkey fat is still in the blank all over the world.This research focuses on the composition of donkey fat. With gas chromatograph and thin layer chromatograph, donkey fat’s total fatty acid composition, sn-2fatty acid composition and trans fatty acid composition were analyzed. Also tocopherol and cholesterol were analyzed with the high performance liquid chromatograph. The efraction index and solid fat content in different temperatures were determinated with the nuclear magnetic vibration instrument and abbe refractometer instrument. Donkey fats fatty acid value, iodine value, saponification value, peroxide value, relative density were determinated, too. The conclusions are followings: the oleic acid content（36.28%） is the highest among all the unsaturated fatty acids（59.30%） in total fatty acids in donkey fat，and essentidal fatty acids content in the sn-2is higher than that in total fatty acids in donkey fat. The trans fatty acids content（0.206%） in donkey fat is far below the national standard of infantal food. Donkey fat contains all four kinds of tocopherols（85.9mg/kg）and the alpha tocopherol content is the highest in the four types of tocopherol. There is very litter cholesterol in donkey fat（0.03187mg/kg). With the nuclear magnetic vibration method the completely melt temperature of donkey fat is determinated as39-40℃.In order to research the differences of donkey fat and some other animal fats, the comparision of properties, composition and trace element content with donkey, pig, cattle, sheep, chicken, duck and ostrich fats is needed. The conclusions are followings. Firstly, in all the studied livestock animal fats, donkey fat has the highest unsaturated fatty acid content in total fatty acids. Secondly, the total polyunsaturated acid content in donkey fat is higher than that in all the other studied animal fats and close to the poultry fats.The sn-2unsaturated acid content in donkey fat is also higher than that in all the studied livestock animal fats, mainly higher in essential fatty acids. Thirdly, the trans fatty acid content of donkey fat is the lowest among all the studied animal fats. The donkey fat has almost no cholesterol. In terms of tocopherol content, donkey fat is also the highest and its tocopherol types are most complete among all the studied animal fats. On the basis of the donkey oil research, this topic focuses on the preparation method of functional fats with donkey fat as basic material. In this thesis, a research model about synchronous directional manufacture of functional fat in the same system with lipase working on decomposition and synthesis of the substrate in anhydrous solvent free environment is established.With three dimensional motion constant speed and pressure method, the ester exchange rate of donkey fat and conjugated linoleic acid (CLA) is enhanced. The experimental finding and screening of specific lipase is in the first place and then single factor reaction conditions will be confirmed. Reaction condition of enzymatic directional ester exchange has been optimized with response surface analysis. Ultimately, the best enzyme substrate reaction conditions are determined like followings: lipase novozymes435as a biological catalyst, solvent-free environment, simultaneous decomposition and synthesis reaction, ester exchange reaction temperature to be60℃, the substrate ratio to be1:3(donkey oil: conjugated linoleic acid w/w), the enzyme added amount to be7%(donkey oil mass ratio), reaction time is55hours and the optimal forecasted exchange rate value to be47%.The discussion focuses on the reaction mechanism of enzymatic directional interesterification, the middle transition state composite structure of acidolysis reaction, acid-ester exchange and ester-ester exchange, and reaction mechanism within molecule of acidolysis reaction. Moreover, synergy factors of active centre and reaction mechanism of lipase are also involved in this disccusion. The reaction mechanism of acidolysis, acid ester exchange at the same time, inhibition of intermolecular ester-ester exchange are also narrated in this researched. Because several reactions complete at the same time in solvent-free conditions, the production of the intermediate is avoided. The conditions for separation and purification of a mixture of triacylglycerol and fatty acid composition are also created because of this.The functional triacylglycerol content after enzymatic directional ester exchange is purified and seperated using secondary molecular distillation with the rotor film molecular distiller. The optimum conditions for molecular distillation method are followings. For the first level of molecular distillation, the parameters are2.0Pa system pressure, distillation temperature230℃, the feed rate2.0mL/min and blown film speed150rpm; for the second level, the parameters are0.2Pa system pressure, distillation temperature310℃, the feed rate of1.5mL/min and blown film speed350rpm.In order to verify the product safety, an evaluation of food safety toxicology for conjugated linoleic acid of donkey fat triacylglycerol is needed. According to the national standards <Toxicology Evaluation Program for Food Safety>, in this research, mouse and rats were put in acute experiment through the mouth, genetic toxicology experiment (including the Ames test, bone marrow micro nuclear test and sperm malformation test) and30days feeding test. After testing, it is seen that all indicators of the triacylglycerol products in this research meet the national standards, so it could be concluded that CLA function oil is a kind of non-toxic, safe oil without genetic mutations and could be used in food and pharmaceutical industries.In order to verify the functionality of the product, a preliminary study of antitumous effect about recombined CLA donkey triacylglycerol is also necessary. Human prostate lymph node cancer cells LNCaP is needed to test anti-proliferative effect of recombined CLA donkey triacylglycerol. Using anti-cancer functional CLA standard sample (99.95%purity) as a positive comparison and normal donkey fat as a negative comparison, a comparative test about antitumous effect to human prostate lymph node cancer cells between both of them and functional fat from experiment with47%CLA content is done. The MTT method is used every24hours to see proliferation of tested cells in96h. We fourd in the first4determinations in72h, donkey fat, ethanol solution of0.025μmol/L density and RPMI-1640cell culture fluid could not inhibit cancer cell growth while CLA sample from Sigma Ltd., but raw material with80%content CLA in molecular rearrangement experiment and the anticancer functional oil with47%content CLA showed ability of inhibiting cancer cell growth. In summary, donkey fat does not have a capacity of cancer cell proliferation.99.95%CLA sample could inhibit human prostate lymph node cancer cells growth while recombined CLA donkey triacylglycerol also showed the potential, similar capacity like CLA sample. |
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