Studies On Lyophilization Injection Of Lamiophlomis Rotata (Benth.) Kudo (LRK)

Objective: Lamiophlomis rotata (Benth.) Kudo is a kind of medical material which is common used in Tibet,with the quality of promoting blood flow, alleviating pain, eliminating wind and drying jaundice.Recently, people pay close attention to it.Because of its good effect and little side effect,it is generally used.It is reported that total flavonoids of Lamiophlomis rotata (Benth.) Kudo(LRK) is the effect position to alleviate pain,which can be used to cure pain caused by bone fracture, rheumatalgia,etc.Some reporte indicates that it can be used to alleviate pain at the fist stage of cancer,which can avoid side effect caused by indomethacin.At present,only the dosage form for oral use is going on sale.This article intend to study on lyophilization injection of LRK, by intravenous injection administration to deepen its development,widen its clinical application, convenient patients who can not take it orally.This thesis is to investigate the best technology of extracting totle flavonoids of LRK,enrich it and get the intermediate;To investigate the formula of lyophilization injection of LRK and technique of preparation,to control its quality comprehensively,to make the quality standard of lyophilization injection of LRK;To study the bio-security of lyophilization injection of LRK including irritability,hemolysis and vas stimulation;To develop a RP-HPLC method for the preliminary study of fingerprint on raw material ,intermediate and lyophilization injection of LRK,which can provide evidence for its quality controllability.Method: Chose the best technology of extracting totle flavonoids of LRK by orthogonal design;Separate and purify totle flavonoids of LRK with AB-8 macroporous adsorptive resins;Use modified gelatin method to get rid of tannin in the extract,evaporate alcohol and then get through AB-8 macroporous adsorptive resins,firstly use warter to flush,then use 10% alcohol, 20% alcohol, 30% alcohol and 50% alcohol in order;With mannitol ,glucose, lactose,etc. as skeleton excipients to prepare lyophilization injection of LRK; The content of galuteolin in lyophilization injection of LRK was detected by RP-HPLC method(Methanol:0.4%H3PO4 Water=50:50 as mobile phase and uv detection at 347nm, with DiamonsiLTM ODS C18 250×4.6mm, 5μm as chromato bar;Flow rate was 1.0ml/min and 20μl was injected every time);The freeze-drying curve was obtained when the lyophilization injection of LRK was produced in GMP workshop;the physical and chemical properties of the injection were evaluated.Guinea pigs were irritated then attacked,level of anaphylaxis was observed comparing with negative and positive control groups;the lyophilization was also investigated to show the effect to 2% red cell sedimention solution and stimulation to the vain of rabbit ear,the local reaction of inject-spots were observed by naked eyes and vein tissues were sliced up to show the pathological change in microscope.RP-HPLC was applied to the preliminary study of fingerprint and analysed with the traditional Chinese medicine fingerprint software.Results: The optimum extraction technology was: with 20% alcohol 8 times weight of raw material as extractant,heat and recirculate three times and 1h pertime;cobine 30% alcohol and 50% alcohol extraction, evaporation to dryness to get intermediate; the optimum formula include total flavonoids of LRK intermediate 60mg and mannitol 240mg,prepared for the solution of 6ml per ampoule to be lyophilized; The solution of lyophilization injection of LRK showed negative in sensitivity test to guinea pig and had no coagulative and hemolysis effect to red cell.The vein of rabbit ear had an intact endodermis and showed no dilation,denaturalization putrescence.In the fingerprint,24 common peaks of lyophilization injection of LRK were confirmed.Coclutions: The extraction technology is reproducible and the extraction achieve the level of crude drug for injection. The formular of lyophilization injection of LRK is rational,the preparation method can be applied in lab and also provide gist for workshop production.The quality standard is reliable,which can control quality of lyophilization injection of LRK.Bio-security tests showed lyophilization injection of LRK has no irritability,no effect of coagulation and hemolysis and no significant stimulative influence to vein.The RP-HPLC method is simple and accurate with a good reproducibility and specificity and the preparatory study of fingerprint can be used scientifically as a quality control method for lyophilization injection of LRK and its raw material.