| We prepared a sub-microemulsion preparation of scutellaria effective fraction phospholipid complex in nasal systems which depended on the theory and pratice of "nasal-brain pathway" and effective parts of scutellaria as raw materials, and safe, stable, effective and controllbale as guiding principle for preventing and treating cerebral ischemic injury through nasal delivery. In order to lay the foundation for the traditional Chinese medicine preparation which prevent and treat erebral ischemic injury and investigate the feasibility of Chinese medicine preparation which deliver drug through nasal into brain, we took a system study obout the process of extraction and purification, preparation, quality control, the evaluation of safety and effectiveness, and absorption mechanism, and so on.Firstly, we prepared effective parts that dfferent contents of baicalin from scutellaria by using water extraction and acid precipitation method, and screening specification of optimum effective parts by using antioxidation in vitro and pharmacodynamics test, and determining the route of administration and effective dose. The preparation technologies for effective parts are as the follows:scutellaria was decocted for three times in water (after boiling), combined extracts and concentrated0.8g/ml, added HCl to adjust to pHl.0-2.0, stood for1h under70℃and stirred precipitate which added proper water, adjusted its pH to7.0by20%NaOH, added specified amount of ethanol (the content of alcohol were10%and40%), stood, filtrated, adjusted filtrate’s pH to1.0-2.0by HCl, stood for1h under80℃and cooled to room temperature, filtrated,washed precipitate’s pH to5.0by ethanol and dryed by decompression drying to get sample which purity of62%and85%, in the same time, bought98%scutellaria extracts as. We researched different content of extracts which antioxidation in vitro and pharmacological test by intravascular suture blocking method to establish a focal cerebral ischemia., results showed85%extracts had the better effect of antioxidation and efficacy; explored the route of administration and effective dose, determined the better route was nasal administration and initially to determinn the effective dose.In view of the water and lipid-soluble of scutellaria extracts are all poor and difficult through the biofilm to affect the bioavailability, we studied the modification of extracts by the technology of phosphlipid complex, results showed that significantly improved oil-water partition coefficient and bioavailability of extracts in phosplipid complex. The preparation technologies for phosplipid complex are as the follows:used absolute ethylalcohol as complex solvent, the ratio of phosplipid and extract was2:1, the extract concentration was3.0mg/ml, the reaction temperature was50℃.Combined the propertise of phospholipid complex and characteristics of intranasal administration, we studied the screening of formulation by investigating swine nasal mucosa permeability and animal nasal mucosa irritation for oil solution, aqueous solution, microemulsion, sub-microemulsion and insitu gel. Sub-microemulsion had the better permeability in the test of swine nasal mucosa permeability, the apparent permeability coefficient is37.09±1.75×10-3(μg·cm-2·s-1), significantly greater than other formulations; investigated the toad palate mucomembranous ciliotoxicity of formulation and its stimulation on the rat nasal mucosa, results showed sub-microemulsion is the best form for scutellaria extract phosplipid complex in nasal systems.We studied the formulation factors and preparation processes of scutellaria extract phosplipid complex sub-microemulsion, and determined the prescription of preparation by using orthogonal design method and central composite design-response surface optimization. The preparation technologies are as the follows:weighed scutellaria extracts, sobean lecithin as above the prescription, placed at a reactor, added to absolute ethanol, refluxed in a kettle at60℃that reaction time was2h, fully composite, recovered solvent by vacuum concentration to get the remining amount of ethanol was about30%, then added the medium-chain triglycerides, soybean oil, emulsified phosphoilpids, oleic acid, vitamine E, and continued recovered to without ethanol taste, which was the oil phase. Glycerin, poloxamer, sodium EDTA were dissolved in appropiate amount of warm-up distilled water, which was the aqueous phase. Two-phase was heated to60℃respectively, added the aqueous phase to the oil phase, then mixed. Colostrum was sheared for9min by19000rev/min at60℃, and cooled to room temperature, conducted homogenization in high pressure homogenizer for6times that high pressure homogenization pressure was600bar, for3times that low pressure homogenization pressure was60bar, adjusted its pH to5.0by0.1mol/l NaOH, and filtrated, which was scutellaria extract phosplipid complex sub-microemulsion. Influence factors test results showed the appropriate storage condition of the formulation is low temperature(4-8℃) keeping away from direct sunlight; In the research of preparation,because phospholipids complex is hard to dry and easy to absorb water, at the same time, complex in oil phase solubility smaller, addition mount of durg can’t completely dissolved and uniform dispersion in oil phase, finally, we used the patent technology solute phase transfer technology successfully solved the problem of dissolved and scattered, got the high drug loading of the submic ro emulsion, meanwhile, avoid the phospholipids complex dry process, and explored t he mechanism of phase transfer technology.According to the pharmacopoeia of the People’s Republic of China on the2010edition, we made further study about the quality standards of sub-microemulsion, including qualitative identification, general quality inspection, and content determination. At the end we established scientific and feasible quality control methods. Results that physical and chemical properties research showed preparation process was rational and feasible that encapsulation efficiency of preparation was89.49%; nasal absorption accorded with first order kinetic and belonged to a passive absorption.We researched the effcet of scutellaria extract phosplipid complex sub-microemulsion on cerebral ischemia in rats by used bilateral carotid artery blocking method to establish a complete cerebral ischemia and intravascular suture blocking method to establish a focal cerebral ischemia. Results showed that compared with the model group, Brain indexes of the drug-treated group were all lower than that of the model group (P<0.01), the content of brain water was significantly decreased (P<0.01, P<0.05), brain tissue pathomorphology and nerve function were markedly improved (P<0.01), neurological deficits scores were significantly decreased(P<0.01); the content of SOD and ATPase were increased, while the content of MDA was significantly deceased in drug-treated group (P<0.01) for complete cerebral ischemia in rats; the content of SOD and Ca2+-ATP were highly increased (P <0.05) for focal cerebral ischemia in rats. Results of toxicity and irritation tests showed that sub-microemulsion has no obvious toxicity.With baicalin as a evaluation index, we studied the brain dynamics and absorption mechanism of sub-microemulsion. Results showed that plasma bioavailability of the nasal adminstration was72%; brain targeting index was7.88which comparaed with intravenous administration; the main way into the brain was olfactory pathway that drug molecules were absorbed through the olfactory mucosa, the uptake of which by neuron axon terminals, arrived at olfactory bulb through axoplasm, further to reach the olfactory, then enter the cerebellum, the brain and other parts.We used effective parts of scutellaria as research object to explore the feasibility of traditional Chinese medicine by intranasal administration for treat diseases of the brain, practice insoluble active ingredients of traditional Chinese medicine phospholipid complex modifid technology, innnovatively invented the formulations molding patented technology(phase conversion technique) that high drug loading sub-microemulsion. This paper provides a basis and reference for the development of traditional Chinese medicine insoluble components and the prevention and treatment for cerebrovascular diseases by nasal drug delivery. |
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