Study On The Preparation Of Neem Oil Microemulsion And Its Acanicidal Activity Of Neem Oil In Vitro

Gamasoidosis is a kind of common verminosis of chronicity and contact,which caused by the acarus in the branches of itch-mite and scab-mite parasitized on the body surface of livestock and birds.The gamasoidosis has a bad effect on the throughput of animals,even becomes the cause of death in them.It also affects the development of stockbreeding by increasing the cost of raising and treating,which makes a big loss to stockbreeding and some concerning fields.At,present,chemicals are used to prevent the gamasoidosis home and abroad,and prevention in immunity and biography is under exploring.However,chemicals give rise to some trouble in quality safety of livestock product because of its high toxicity,long residual period and pollution of the environment. Therefore,it bears significant meaning to filter acaricide of high effeciency,low toxicity and environmental protection from natural chemicals of plant source,and it also benefit the sustainable development in stockbreeding and medical prevention in clinical gamasoidosis.Mordern research shows the neem oil distilled from neem seeds can effectively control verminosis such as ticks,acarus and cooties parasitized on the body surface of human being and animals.Neem oil mainly includes EC and EW,in which the EC will cause environmental pollution and EW is prone to delamination,doing harm to the eco-enviroment and product quality;even worse,it needs a great deal of dosage.In order to solve the problems mentioned above,the reseach will stress on the study for developmet of new neem oil microemulsion according to its own characteristics.The neem oil in the research will be used not only as chemicals but as oil phase,in order to optimize the preparation of neem oil.Then the neem oil microemulsion will be put to test its acaricidal activity in vitro and the skin stimulation on rabbits.The results for the research is mainly as follows:1,The process of the experiment:the materials to prepare the microemulsion includes neem oil,TW60,TW80,SDS,SDBS,methanol,ethanol,isopropanol,n-butylalcohol, n-pentanol,n-hexyl alcohol,n-heptanol,n-octyl alcohol,propanediol,propanetriol, Polyethylene glyco1400,distilled water and so on.Based on the direction of microemulsion,titration can be helped draw pseudo-ternary phase diagrams,and confirm the distribution of microemulsifing area,then prepare the neem oil microemulsion by optimizing the directions and choosing the best one to make it.TEM(transmission election microscope) is used to examine the morphological structure of neem oil microemulsion. The stability of them can be examined by dilution,centrifugalization,low temperature, heat storage and long-stem storage.The result of the experiment shows that if TW80 and SDBS are equally mixed in mass ratio of 4:1 to make the compound surfactant,and the compound surfactant and n-hexyl alcohol are mixed in mass ratio of 4;1 to make the emulsifier system,then mixed neem oil,emulsifier system and water in mass ratio of 1:3.5:5.5,altogether put it into water bath of 40℃,stir in 800 rounds per minute for 15 minutes,and finally it will appear inaurate clear transparent neem oil microemulsion as the best technical direction.The neem oil microemulsion made under the best technical direction is of O/W type.Observed throngh TEM,the liquid drops of it shaped in balls, which equally distributed in size.After 100 times dilution centrifugalization at low or high speed,no changes can be seen by naked eyes.Put the neem oil microemulsion in the storage of-20℃,4℃,and 54℃for 15 days,it only freezed under the condition of-20℃, no changes in other situations.And when the temperature increased to room temperature,it resumes a transparent liquid.The long-term stability of neem oil microemulsion indicates that it can be stored in natural environment for 6 months and it can keep stable in appearance.2,Different dosage forms and their different blank preparation of neem oil is used on rabbits to compare the acaricidal activity on itch-mites of neem oil microemulsion,neem oil EW and atolein solution.The result shows the three are of comparatively strong acaricidal activity that the time for complete killing the larvas is respectively 192.50,212.50,337.50 minutes,in which the time of neem oil microemulsion is obviously shorter than that of atolein solution(P＜0.05),while there is no obvious difference between the time for neem oil microemulsion and that of neem oil EW(P＞0.05).The time of three blank preparations for complete killing the larvas is longer than their respective preparations as:231.25,437.50,2 495.00 minutes.The Median lethal time(LT₅₀) of the Sarcoptes scabiei var.cuniculi of neem oil microemulsion,neem oil EW and atolein solution is shorter than their respective blank preparations as:81.746 3,95.551 5,156.679 9 minutes.The toxicity regression equations for them are Y=-6.026 9+3.151 4X,Y=-9.978 4+5.039 0X,Y=-11.406 8+5.196 7X.The LT₅₀ of three blank preparations for larva itch-mites on rabbits are 89.066 6,165.401 0,1576.194 6 minutes.From the above data,it makes clear that the absorbency by skin ofneem oil microemulsion is stronger than that of neem oil EW and atolein solution.And microemulsion works better and has more stability than neem oil EW,so that it can be infinitely diluted with water and is more convienient for use.3,The thrill experiment on rabbit skin is used to examine the thrill of animal skin by microemulsion and neem oil microemulsion.The result shows when putting the neem oil microemulsion on full skin for only once,during 72 hours,the stimulus response(RS) scores for blank microemulsion and neem oil microemulsion of low and high concentration are 0.When putting on worn skin,from the beginning of 72 hours,the RS scores for both are 0.33,less than 0.4.When putting the neem oil microemulsion on full skin for several times,the RS scores for blank microemulsion and neem oil microemulsion of low and high concentration are 0.When putting on worn skin,on the exact day of removing the chemicals,the RS scores for both blank microemulsion and neem oil microemulsion of low and high concentration are 0.33,less than 0.4.After 24 hours of removing the chemicals, the RS scores is up to the maximum 0.66.After 48 hours and 72 hours,the scores decreas to 0.33.From the above data,it can be seen that the stimulus response of neem oil microemulsion is of light stimulation,so it can be used in skin because of its subtle stimulation.