Preliminary Study On Preparation, Pharmacology And Toxicology Of Modern Five-Kind Traditional Chinese Medicine Based On Sulforaphene

Radish is a kind of common cruciferous plants, radish roots and sprouts are vegetables which are daily consumed. Radish seed can be used as a traditional Chinese medicine and its name is Laifuzi. Radish, as a kind of cruciferous plant, contains abundant of myrosinase and glucoraphenin, and myrosinase can rapidly hydrolyze gluocoraphenin and convert it to sulforaphene when myrosinase contacts with glucoraphenin after the radish tissues are broken. As a kind of isothiocyanates, sulforaphene possesses a strong activity of inducing phase II enzymes to improve the ability of scavenging carcinogen from human body, and sulforaphene can effectively inhibit the growth and proliferation of cancer cells and kill cancer cells. This thesis mainly study the separation and preparation technology of crude sulforaphene extract, the acute toxicity of crude sulforaphene extract and the anticancer activities of crude sulforaphene extract in vivo and in vitro.The efficient separation and preparation process of crude sulforaphene extract was established by selecting the radish seed as a raw material which contained abundant of glucoraphenin and myrosinase. Study showed that the hot pressing pretreatment processing can eliminate hydrogen sulfide from the hydrolysis reaction system, effectively avoiding the rapid degradation reaction of newly generated sulforaphene, and then the effective accumulation of sulforaphene was realized. Vitamin C can effectively restore the thiol group of myrosinase to efficiently activate myrosinase and then the formation rate of sulforaphene was improved. The formation rate of sulforaphene was significantly increased with the decreasing of temperature, and low temperature was advantageous to the enzyme hydrolysis of sulforaphene. The optimum pH value was 5.0 for glucoraphenin hydrolysis reaction catalyzed by myrosinase, and this pH value was conducive to the efficient accumulation of sulforaphene in the hydrolysis reaction system. SP-850 resin can rapidly adsorb and enrich sulforaphene from hydrolysis water solution,80% ethanol water solution can effectively elute sulforaphene from SP-850 resin, the concentrated sulforaphene solution was extracted with dichloromethane, and then the crude sulforaphene extract was obtained with a high content of sulforaphene. Under the optimal separation condition, the production yield of crude sulforaphene extract was 9.25 g·kg-1, the content of crude sulforaphene extract was 81.5% and the total yield of the separation and preparation process was 88.3%.Study on the anticancer activity in vitro showed that crude sulforaphene extract could dose dependently inhibit the growth and proliferation of NCI-H460, A549, SK-OV-3, Hela, HepG2, MDA453, MCF-7, A375, BxPC-3 and LNCaP cancer cells. The half inhibitory concentration were 3.22 μg·mL-1,7.38 μg·mL-1,7.16 μg·mL-1,3.92 Hg·mL-1,4.92μg·mL-1,6.33 μg·mL-1,5.84μg·mL-1,0.964 μg·mL-1,3.65 μg·mL-1 and 6.71 μg·mL-1. A375 cancer cells was the most sensitive for crude sulforaphene extract, and the half inhibition concentration for A375 cancer cells was lower than other cancer cells about generally 3-8 times. The result of acute toxicity experiment showed that the half lethal dose of crude sulforaphene extract for experimental mice was 186.8mg·kg1, and confirmed the crude sulforaphene extract had moderate toxicity on experimental mice. The main cause of mice death could be strong excitatory effect of crude sulforaphene extract on the central nervous system of experimental mice, and the certain injury effect of crude sulforaphene extract on the gastrointestinal tissue of experimental mice. The anticancer activity of crude sulforaphene extract in vivo was studied, and experimental result showed that crude sulforaphene extract could not significantly inhibit the growth of A375 cancer cells xenograft tumor under a BID dose between 50 mg·kg-1 and 250 mg·kg-1. Experimental mice could had a rapid metabolism processing for crude sulforaphene extract in vivo, the crude sulforaphene extract administrated intragastrically was metabolically eliminated rapidly, and the toxicity of crude sulforaphene extract on experimental mice was significantly reduced, but at the same time the anticancer activity of crude sulforaphene extract in vivo was also significantly reduced.