| Centella asiatica (Centella asiatica (L.) Urb.) is the whole dry plant of the Umbelliferae Centella asiatica-Centella asiatica, and was used as a traditional medicine for thousands of years in many countries and regions, in Chinese medicine it also has been used for oral and external for 2000 years,it is still used as one of the main raw material of Herbal tea in Guangdong too. Currently, Centella asiatica are used in clinical to treat burns, inhibite scar proliferation and antidepressant. Although it was reported that Centella asiatica could be used to cure gastric ulcer, there were no Centella asiatica for the treatment of gastric ulcer drug on domestic market yet.Centella asiatica were included in Chinese Pharmacopoeia 2005 edition for the first time, and its active ingredients Asiaticosides added in the Chinese Pharmacopoeia 2010 edition as a new item.In this thesis use modern pharmaceutical technology to develop the effective parts of Centella asiatica herbs Asiaticosides to a clear treatment of gastric ulcer drug, and the the purpose of this thesis is to explore the best preparation process, quality standards and preliminary observe its anti-ulcer effects of Asiaticosides sustained-release pellets.1. TLC qualitative distinction, UV and HPLC quantitative analysis methods were established for the target constituents measurement of Asiaticosides, those methods mentioned above are accurate and reliable, simple and rapid, and provides an effective analytical method of the studying Asiaticosides sustained-release pellets. In the TLC chromatograph research, after a comparative study, chloroform-methanol-water (13:7:2) were selected as developing systems,10% acid solution as developer sulfuric, and experiments proved its effective. A compared experiment founded that ultrasound with extraction solvent methyl alcohol is a best extraction method. High performance liquid chromatography with diode array detector and wavelength scanning function, and 205nm were used as detection wave length, which consistent with the highly effective liquid phase chromatography's examination wave length in Asiaticosides item in "Chinese Pharmacopoeia" 2010 Edition. The chromatograph conditions are:chromatographic column: KromasilC18 column (4.6×250mm,5μm); Flowing:Methyl cyanide:Peroxide solution= 26:74; Speed of flow:1ml/min; Examination wave length:205nm; The column is warm: 25℃.2.In the refining research of Asiaticosides, the content of Asiaticosides extraction rate and yield of dry extract were used as monitoring indicators, to optimize the extraction and purification process. And observed the single factor effect of extraction solvent on Asiaticosides content. Orthogonal design was used to optimized the four factors related to extraction process:ethanol concentration, solvent volume, extraction time and number of process.The optimized extraction process are:Asiaticosides herbs were extract 3 times with eight times the volume of 50% ethanol boils backset. AB-8 macroporous resin was selected in purification process from AB-8, D-101, HPD200, HPD100, for its maximum cleaning speed, the highest transfer rate of the index components. The rate of extracting and the content of Asiaticosides in the dry extract is 328.81%,56.21%. respectively.Centella asiatica extract purification process described as follows:the concentrated solution (the grass of Centella asiatica 0.0625g in every 1mL solution), according to the ratio of raw herb and resin 1:1, was added in AB-8 macroporous resin column at first, 1ml/min,10% ethanol solution eluted, then lml/min,2BV 70% ethanol elution, the eluent was collected in the following rotary evaporator vacuum for the recovery of ethanol at 70℃, at last, when the relative density of the eluent reached 1.15g/ml (70℃), the eluent was dried in Vacuum drier at the degrade of 70℃, and the dry paste was smashed and reserved.3.The main physical and chemical propertie of the Asiaticosides's were tested. The results indicated that the purified water and dilute hydrochloric acid can be used as Asiaticosides solvent in the dissolution test. The porosity of Asiaticosides extract powder is relatively large, the average bulk density was 0.652 g/ml. The angle of repose of Asiaticosides extract powder is more than 40°, the fluidity is not good, and the relative humidity is small in 54% medicinal powder's water-absorptivity, but in 54% above the medicinal powder water absorbability increases obviously. Asiaticosides shows stability in light, heat, moisture conditions. The interaction between Asiaticosides extract and some accessories to be adopted,by TLC, and the results show that no changes occurred in Asiaticosides combined with HPMC, EC, carbomer 934P, lactose, stearic acid. In artificial gastric juice and intestinal fluid, Kabo (carbomer) could provide the controlled release of drug effect, in a lower density than other ingredient, as a result, Carbomer 934P was selected as the main accessories.4. Based on the references studying, both the appropriate suitable pill preparative technology and the best prescription were designed, by inspecting each factors related to pill formation craft, combined with the results of the stomach mucous membrane surface experiment in vivo and release test in vitro. The prescription determination of the micro pill is as follows:Asiaticosides(50% of the total amount), MCC, (25.5%, of the total amount), carbomer 934P(13% of the total amount), stearic acid (6.5% of the total amount), ethyl cellulose(5% of the total amount),, wetting agent are the water。The cat's foot total glucoside 50%, MCC25.5%, Kabo 934P13%, stearic acid 6.5, ethyl cellulose 5%,.This Asiaticoside extended-release pellets show better pill roundness, higher yield,and the extended-release satisfied the anticipated demand, under selected pill preparative technology total glucoside releases slowly The preparation craft of Asiaticosides sustained-release pellets are as follows:Asiaticosides extracts and accessories were filtrated by 80 mesh filter, and mixed with 13%(w) Carbopol 934P and stearic acid 6.5%(w) at a high-speed stiring, refrigeraeted 2h immediately after melted in a constant temperature water bath at 60℃, then were put in a dryer to room temperature, crushed and the grinding through 80 mesh sieve at last; More Asiaticosides mixed with MCC, (25.5%, of the total amount), carbomer 934P(13% of the total amount), stearic acid (6.5% of the total amount), ethyl cellulose(5% of the total amount), and water(the wetting agent),were prepared the micro pellets through the Extrusion spheronization system, the prepared micro pellets first put in a desiccator drying 2h,at the 40℃, then heated 80℃for 2h, the purpose micro pellets were Picked out form a 20～40 mesh sieve.5. The purpose micro pellets were coated with the Release coating fluid:Killcoat SR30D, by using the multifunctional Coating Machine Granulator. Prescription and process factors were investigated, and the Prescription of Release coating fluid were optimizated with the method of central composite design-response surface. The final coating process of Asiaticosides sustained-release pellets are:the polymer contains 43.86% Kollicoat SR30D, and the coating weight add by 20%.. Prescription are:43.86% Kollicoat SR30D, 2% propylene glycol,0.5% lactose,46.14%water,0.5% titanium dioxide,2% talc,5% water. Parameters of:the coating conditions:60℃inlet air temperature,0.08Mpa spray pressure,0.08Mpa protective air pressure,0.5-0.8ml/min constant pump flow rate,10% added coating weight, and the coating curing for curing 12h at the temperature of 40℃.6. Pharmacodynamic studies Confirmed that:To some extent Asiaticosides could scavenge oxygen free radicals, improve antioxidant capacity, stimulate the growth of granulation tissue mucosa, and promote inflammation damages and ulcer, all of these prove that the anti-gastric ulcer effects of Asiaticosides.The Animal experiments showed that compared with modle group, the ulcer indexies of rabeprazole sodium acetic acid and the asiaticosides high and middle dose group were significantly lower (P<0.05), the ulcer indexies of asiaticosides high and middle groups were significantly lower than Hericium group (P<0.05) and was not significant different from rabeprazole group(P> 0.05), all of these result reveals Asiaticosides show the anti-gastric ulcer effects in a dose-dependent manner, and could be used to treat the experimental ulcer in rats induced by acetic acid. SOD activity and MDA content of asiaticosides high and middle dose was stronger than rabeprazole sodium in Gastric tissue from Acetic acid induced gastric ulcer animals. Asiaticosides high dose histological examination showed that the inflammatory cell infiltration mainly infiltrated in mucous layer, a small amount of fibrous scar tissue and inflammatory granulation tissue around the mucosal were in visible. |
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