| Objective: The medicinal use of Prinsepia utilis Royle oil has a long history.It is the first time in the world that Mosuo people extracted Prinsepia utilis Royle oil from fruit and it was used for health care and food.Mosuo people called it "Qingnamanan" or "Qingnayou" because they thought that longterm consumption of Prinsepia utilis Royle oil can bring them health,longevity and beauty.It can be seen that Prinsepia utilis Royle oil has a high health value.Though Prinsepia utilis Royle has been planted in large quantities in Yunnan province,there are few researches and formulation studies on its special medicinal and healthy functions.Therefore,it has great development potential in the field of health care.In this paper,the basic researchs on the pharmacological activity and formulation of Prinsepia utilis Royle oil were carried out,which provides the research basis for the industrialization.This paper carried out resrearches on the following aspects:(1)to detect the physical and chemical indexes and analyze the components of Prinsepia utilis Royle oil and establish GC internal standard method to determine the content of 4 fatty acids in Prinsepia utilis Royle oil;(2)to investigate the effect of Prinsepia utilis Royle oil on inhibiting the proliferation of various cancer cells in vitro and to conduct a developmental toxicity test based on zebrafish model;(3)the microcapsules were prepared by spray drying and the preparation process was optimized by single factor and orthogonal test;(4)the stability of microcapsules of Prinsepia utilis Royle oil was preliminarily investigated.Method:(1)Prinsepia utilis Royle oil was extracted from the fruit of Prinsepia utilis Royle by supercritical CO2 fluid.The physicochemical indexes such as acid value,iodine value,saponification value,peroxide value and refractive index of Prinsepia utilis Royle oil were detected by the methods of general principles 0713 and 0622 of 2015 edition of the Chinese Pharmacopoeia.GC/MS was used to analyze the fatty acids and sterols in Prinsepia utilis Royle oil.On this basis,GC internal standard method was established to determine the contents of oleic acid,linoleic acid,stearic acid and palmitic acid in Prinsepia utilis Royle oil at different extraction periods through supercritical CO2 fluid extraction.(2)The inhibitory effects of Prinsepia utilis Royle oil on colon cancer cells(SW480),breast cancer cells(MDA-MB-231,MCF-7),glioma cells(C6),hepatocellular carcinoma cells(Hep G2)and prostate cancer cells(PC-3)were investigated by classical MTT method.Finally,the developmental toxicity test in vivo was carried out based on zebrafish model.(3)In order to improve the stability of Prinsepia utilis Royle oil,maltodextrin and acacia gum were used as wall materials,the Prinsepia utilis Royle oil was prepared into stable and uniform emulsion by high speed shear disperser and high pressure homogenizer,and finally Prinsepia utilis Royle oil microcapsules were prepared by a spray dryer.Single factor and orthogonal test were used to optimize the process of spray drying to prepare microcapsules.Sensory evaluation,infrared spectrum scanning analysis,optical microscope and scanning electron microscope observation were carried out on the microcapsules.The tapped density,angle of repose,moisture content and particle size were measured to evaluate the quality of the optimized microcapsules.(4)Finally,influencing factors test,accelerated stability test and longterm stability test of the microcapsules were preliminarily investigated.Results:(1)The CO2 supercritical fluid extraction method was used to extract Prinsepia utilis Royle oil and its extraction rate was 31.13 %.The following physical and chemical properties were dected in order to evaluate the quality of the oil,acid value 0.86 mg KOH/g,iodine value 101.7 g I/100 g,saponification value 190.3 mg KOH/g,refractive index 1.4630,peroxide value 2.20 g/100 g.The fatty acid and phytosterols composition of Prinsepia utilis Royle oil were both analyzed by GC/MS.Palmitic acid,palmitoleic acid,stearic acid,cis-oleic acid,cis-linoleic acid,arachidic acid and cis-linolenic acid of fatty acid in Prinsepia utilis Royle oil were respectively 16.24 %,0.30 %,8.60 %,38.34 %,34.73 %,0.63 % and 1.16 % while the trans-fatty acid was less than 0.10 %.It was shown that ?-sitosterol,fucoidol,cycloheximide,2,4-methylenecycloartenol,stigmasterol,campesterol and squalene of unsaponifiable matters in Prinsepia utilis Royle oil were respectively 53.19 %,14.43 %,10.46 %,8.63 %,8.03 %,3.71 % and 1.55 %.The oil extracted by CO2 supercritical fluid extraction method has good quality and high unsaturated fatty acid content.It took short extraction time but had a high yield of extraction oil and green environment friendly.(2)The content of palmitic acid,stearic acid,oleic acid and linoleic acid of Prinsepia utilis Royle oil was determined by GC internal standard method with phenyl benzoate as the internal standard.The method was simple,convenient and accurate.The different content of fatty acid of Prinsepia utilis Royle oil extracted by supercritical CO2 fluid extraction during different period was investigated.The highest content of linoleic acid of the oil collected from 1.0 to 1.5 hours was 348.05 mg·g-1.The highest content of the oil collected from 1.5 to 2.0 hours was 341.79 mg·g-1.The content of oleic acid and linoleic acid of the oil collected from 2.0 to 2.5 hours was lower than other periods.(3)The inhibitory effect of Prinsepia utilis Royle oil on SW480 cells was evaluated by MTT experiment testing.The half-inhibitory concentration was 238.7 ?g/m L,which may be related to the inhibition of cell growth induced by Prinsepia utilis Royle oil.Then the zebrafish model was used to evaluate the in vivo developmental toxicity.It was shown that Prinsepia utilis Royle oil has low teratogenicity,low lethality and good hatching rate on zebrafish embryos when the concentration was less than 600 ?g/m L.(4)The preparation process of microcapsules of Prinsepia utilis Royle oil was obtained through the orthogonal test optimization.They were shown as below: the ratio of composite wall material/core was 2/1,the peristaltic pump injection speed was 4 g/min,the inlet air temperature was 170 ?,the atomization pressure was 5×103 KPa,the solids content was 5 %,and the shear rate was 6000 rpm.The microcapsule encapsulation rate of Prinsepia utilis Royle oil was 90.85 ± 1.42 %.The microcapsules were light yellow solid powders which had no special odor.The bulk density and tap density of the microcapsules were 0.283 g/m L and 0.417 g/m L,respectively,while the angle of repose was 40.2°.The particle size of microcapsules under the optical microscope was 2 ? m44 ? m.The microcapsules under the scanning electron microscope had a spherical structure which were full and its surface was smooth,no clear cracks and holes,but the wall material had a small amount of adhering agglomerates.The microcapsules under the laser particle size analysis shown that the average particle size,D90 and were respectivele 0.949 ?m,2.063 ?m and 0.274.Finally,Fourier transform infrared spectroscopy was used to determine the formation of microcapsules of Prinsepia utilis Royle oil. (5)After 10 days of high temperature test,the peroxide value of Prinsepia utilis Royle oil increased to 1.734 g/100 g,which is about 62 times as much as that of 0 day.However,the microcapsules increased to 0.613 g/100 g after 10 days of the high temperature test./100 g,about 13 times as much as the microcapsules before the high temperature test.After 10 days of intense light test,the peroxide value of the oil increased to 0.304 g/100 g,which was about 11 times as much as the oil before intense light test.However,the microcapsules increased to 0.103 g/100 g after 10 days of intense light test,about twice as much as the microcapsules before intense light test.The peroxide value of the oil in the high-humidity environment had no obvious changed after 10 days in the high-humidity environment.However,the microcapsules absorbed moisture and its weight gained 6.50% after 10 days in the high-humidity environment.It was condensed into a block,which turns from pale yellow to dark yellow and its peroxide value was about 3 times as much as the microcapsules before the high humidity test.After 3 months of accelerated stability test,there was no significant change in the appearance of the microcapsules.The maximum value of peroxide was 0.288 g/100 g,which increased slightly compared with the original sample.After 6 months of long-term stability testing,there was no significant change in the appearance of microcapsules.The maximum value of peroxide was 0.138 g/100 g,which was about 2.8 times as much as the microcapsules before the test.Conclusion: The physical and chemical properties,compositions of fatty acids and the unsaponifiable matters of Prinsepia utilis Royle oil extracted by supercritical CO2 fluid was tested in this study.GC internal standard method was established for simultaneous determination of 4 main fatty acids of Prinsepia utilis Royle oil and the methodological study showed the method was stable and accurate,which provided a more perfect quality control method for the control of Prinsepia utilis Royle oil.Antitumor activity in vitro and the developmental toxicity test in the zebrafish of Prinsepia utilis Royle oil was also carried out.Finally,the microcapsules optimized by orthogonal test were prepared by spray drying,which can improve the stability during storage and transportation,and broaden the product development route of Prinsepia utilis Royle oil. |
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