| Objective:In this paper,the submicron-emulsion was prepared by elaeagnus mollis oil which was used as raw material,and its preparation process was optimized.The quality of submicron-emulsion was evaluated and stability of the preparation were investigated.Futhermore,its anti-fatigue effect and mechanism were discussed.Method:(1)Formulation and preparation study of elaeagnus mollis:(1)The appropriate HLB value of elaeagnus mollis oil was investigated;(2)The formulation of elaeagnus mollis oil submicron-emulsion include the emulsifier,the co-emulsifier,preservatives,and sweeteners were subjected to single factor screening;(3)The optimal composition ratio of each component in the formulation was determined;(4)The four key factors such as temperature,mixing time,speed and emulsification methods were investigated by orthogonal-design.On the basis of single factor,orthogonal design was used to optimize the preparation process,and the elaeagnus mollis oil was prepared as O/W submicron-emulsion;(5)the homogeneous pressure and the number of homogenization were explored to prepare submicron-emulsion.(2)Quality evaluation of elaeagnus moliis oil submicron-emulsion:According to the provisions of the Chinese Pharmacopoeia(2015),“Health food management approach”and“Health food stability testing guidelines”,the assay methods of character identification,particle diameter,density,Zeta electric potential,centrifugal test,and centrifugal stability coefficient(K_e)were studied.Gas chromatography(Gas chromatography,GC)and High performance liquid chromatography(HPLC)were used to determine the content of linoleic acid in the submicron-emulsion of elaeagnus mollis oil.The two methods were investigated.The formulation stability test was conducted to examine the storage conditions and time.Study on relieving exercise-induced fatigue of elaeagnus mollis oil submicron-emulsion:one hundread female mice were randomly divided into control group,low dose group(1.25 ml/kg),middle dose group(2.5 mL/kg),high dose group(5 mL/kg)and submicron-emulsion group(23.7 mL/kg),twenty in each group.The mice in each experimental group were continuously gavaged for 30 days,and the swimming training was performed once every other day for 20 minutes to 30 minutes.Ten mice in each group were subjected to a weight-bearing swimming test and their exhausted swimming time was measured.In the other 10 mice,the elisa kit was used to measure the changes in blood biochemical parameters such as Blood lactic,Hepatic glycogen and Blood urea nitrogen before and after exercise.In order to study the effect of elaeagnus mollis oil and submicron-emulsion on exercise-induced fatigue,and further explore its anti-fatigue mechanism.Results(1)The optimum HLB value of the elaeagnus mollis oil was 9 after screening,and the Surfactant(SF)of the elaeagnus mollis oil submicron-emulsion was a mixed surfactant consisting of monoglyceride,polyglycerol fatty acid ester and lecithin.Cosurfactant(CoSF)is a food grade glycerine.The mass ratio of SF/CoSF is Km=1:1,SF:CoSF:oil=4:4:5,in a word,the best formulation is composed of 4%of surfactants,4%of glycerin,5%of butternut oil,76%of distilled water,1%of potassium sorbate,and 10%of sucrose.The preparation process was as follows:The mixed emulsifier was dissolved in elaeagnus mollis oil and heated in a water bath at 70?as oil phase,glycerol was dissolved in distilled water and heated in a 70?water bath to serve as water phase.Water phase was slowly added to the oil phase,then the mixture were heated at 70?water bath,while heating continued stirring 30-60min.Under the condition of 70?,the stirring speed of 1500~2000 r/min,the uniform and stable milky white emulsion were formed.Finally,the high-pressure homogenizer was used to homogenize the emulsion under high pressure,and the homogenization condition was homogenized under 600 bar homogenization pressure for 6 times to obtain the final submicron-emulsion formulation.(2)The appearance of the winged milk is uniform milky white liquid,the particle size distribution is between 100~1000nm,the average density is 0.9702~0.9735g/cm~3,pH is 5.3~5.5,?potential of-40~-50mv;There is no delamination after centrifugation at 4000r/min for 15 min.The content of linoleic acid in the elaeagnus mollis oil sub-microemulsion with Gas chromatography method:the linear range of linoleic acid was 0.0506~1.012mg/ml,and quantitative detection limit and minimum detection limit were 79.2?g/m L(S/N=10)and 23.7?g/mL(S/N=3)respectively.the intra-day precision RSD was 0.47%~1.74%,the intraday precision was 0.33%~1.23%.The RSD value of the stability test was 1.32%,the recovery rates were between 98.4%and 102.1,the average recovery was 100.1%and the RSD was 1.07%.The content of linoleic acid in homemade wing milk is 9.87~10.18mg/mL,the RSD value of each batch is0.96%~1.62%.The content of linoleic acid in the elaeagnus mollis oil sub-microemulsion with High performance liquid chromatography.The linear range of linoleic acid was0.0506~1.012mg/mL,and quantitative detection limit and minimum detection limit were 21.08?g/mL(S/N=10)and 6.325?g/mL(S/N=3)respectively.The intra-day precision RSD was 0.38%~0.6%,the intraday precision was 0.98%~1.54%,the RSD value of the repeatability test was 1.01%,the recovery was between 98.2%and102.4%,the average recovery was 100.41%and the RSD was 1.43%.The content of linoleic acid in homemade wing milk is 9.91~10.21mg/mL,the RSD value of each batch is 1.15%~1.34%.The stability test results show that at the high temperature condition(60°C),the elaeagnus mollis oil submicron-emulsion was stratified by centrifugation test on the10th day,and there was no significant change in other indicators except K_e value;under the condition of low temperature and light,each component including appearance,the quality evaluation index and linoleic acid content were no significant change.Accelerated test results show that submicron-emulsion was stratified by centrifugation test at 40?,2 months,and the K_e value is large;the indicators have not changed significantly at 30?.long-term stability test results show that all the indicators no significant change.(3)There was no significant difference in the loading swim time between the high/low-dose group and the control group(P>0.05),while the middle dose group was significantly longer than that of the control group(P<0.05).Compared with the control group and high/low-dose group,the submicron-emulsion group was not statistically significant,but the loading swim time was slightly longer.The biochemical indicators in the middle dose group of elaeagnus mollis oil,including the hepatic glycogen storage and recovery rate,BLA,BUN and clearance rate,were remarkably changed when compared with that of the control group(P<0.05).There was no significant difference between the high-dose group and the low-dose group(P>0.05).All indicators of the submicron-emulsion group were superior to the control group and high/low-dose group.Conclusion:The study was assessed from some aspects,such as prescription screening,preparation process inspection,quality evaluation,functional evaluation of anti-fatigue and other aspects,the results of the preparation of elaeagnus mollis oil submicron-emulsion.The elaeagnus mollis oil submicron-emulsion was prepared successfully.The preparation has the advantages of masking the sweet smelling and taste of the oily phase,improving the bioavailability and promoting the absorption of the human.All indicators are in line with relevant operations under"National Food Safety Standard for Food Additives"(GB2760-2014),"Chinese Pharmacopoeia"(2015)fourth,"stability test of raw materials and pharmaceutical preparations"and"health food inspection and evaluation of technical specifications"(2003).The preparation is a novel oral preparation with good biocompatibility,high stability,good safety,and relieves exercise fatigue.Having a certain research value and market application prospects. |
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