Research And Development Of A Compound Preparation For Treating Osteoarthritis

Objective:To compare the effect of S-adenosy-L-methionine combined with meloxicam on knee osteoarthritis induced by mono-iodoacetate in rats,and explore the therapeutic effect of induced osteoarthritis in rats and its mechanism.To compare the effect of ademetionine, meloxicam and combination of multiple pain, and explore the mechanism. Study of ademetionine hygroscopicity and meloxicam by solvent selection. Compound particle movement, shape, particle size distribution as the index to determine the preparation process of compound enteric coated capsule in the treatment of osteoarthritis of the prescription composition and preparation of the compound enteric coated capsules were prepared. Through the determination of the critical relative humidity of the prescription, the reference is provided for the control of the environmental humidity in the production process of the preparation. The feasibility of the preparation technology was verified by the three batch amplification experiments.The quality standard for the treatment of osteoarthritis of compound enteric capsules was established.Methods:(1)the function of the compatibility of ademetionine with meloxicam on knee osteoarthritis:KM mice were randomly divided into 7 groups according to body weight matched after the adaptive feeding of 1w. Through the hot plate method and writhing test, to study the effect of ademetionine, meloxicam and compatibility between the two on the pain reaction in mice.Wistar rats were randomly divided into 7 groups after the normal feeding of 1w. After intragastric administration for 21 days, the rats were 3% sodium pentobarbital intraperitoneal injection of superficial anesthesia, and OA model was induced by sodium mono-iodoacetate in unilateral knee joint of rats. The mean arthritis index was evaluated at different time, and the histological change of histopathology on knee joint cartilage and synovial was observed by light microscopy.Cartilage oligomeric matrixprotein(COMP), matrix metalloproteinase-3(MMP-3),nitric oxide(NO),cytokine interleukin-1β(IL-1β) and tumor necrosis factor-α(TNF-α)in serum or synovial fluid were detected by ELISA.(2) The preparation technology research of compound capsules in the treatment of osteoarthritis : Microcrystalline cellulose, lactose, silica powder, β-cyclodextrin and starch as excipients, with hygroscopic index screening ademetionine anti hygroscopic materials kinds and dosage. Poloxamer 188(F68) and polyvinylpyrrolidone k-30(pvp-k30) as accessories,with the dissolution degree index screening of meloxicam increase the type and amount of solvent. Screening of the best compound preparation by the index of the mobility of capsule contents, particle size distribution, forming.Finally, to determine the composition of treating osteoarthritis of compound prescription of enteric capsules.According to the preparation process, the production of 3 batches of samples study on the enlarging experiment. In accordance with the 2010 edition of the "Chinese pharmacopoeia " method, the 3 batch of samples for routine inspection.(3)Establishing the quality standard of compound capsules in the treatment of osteoarthritis : To identify compound ademetionine and meloxicam by chemical method, Ultraviolet spectrophotometric method and High performance liquid chromatography. Determination and related substances of compound ademetionine and meloxicam was examined by HPLC method. To carry out methodology investigation.In vitro release of the drug was investigated at pH l.2 or 6.8 of the dissolution medium by paddle method.Finally, the quality standard of compound capsules in the treatment of osteoarthritis was established.Results:(1)the function of the compatibility of ademetionine with meloxicam on knee osteoarthritis : The different proportion of ademetionine with meloxicam can extend the mouse hot plate reaction and twisting body response latency.Among, the high dose group can significantly inhibit the thermal and chemical stimulation induced pain.ademetionine with meloxicam and the compound could obviously reduce the degree of rat knee joint inflammation symptoms, improve the knee articular cartilage and synovium lesions, reduce the activity of COMP and MMP-3 in serum, and reduce the level of IL-1β,TNF-α and NO in serum and synovial fluid.(2) The preparation technologyresearch of compound capsules in the treatment of osteoarthritis : Microcrystalline cellulose and lactose were selected as anti moisture absorption auxiliary material of ademetionine, SAMe,MCC and lactose at the dosage ratio of 14∶5∶2.Poloxamer188 was selected as the increasing solvent of meloxicam, MLX and F68 at the dosage ratio of 5∶3.The preparation process: F68 dissolved in an appropriate amount of ethanol, is placed in the corresponding MLX of solid powder, stir, 40℃water bath boiling dry ethanol, vacuum drying, grinding, through a 100 mesh sieve spare, to MCC and lactose, same raw material powder and 100 mesh, according to the proportion of weighing, mixing, with 98% ethanol made of soft material, 24 orders granulation, vacuum drying, 24 orders whole grain, add 0.15% silica powder can flow,filling in the enteric coated capsule shell.The preparation of the drug particles with good flow and formability, particle size distribution, the critical relative humidity of78%, the difference and the content uniformity of the test are in line with the requirements.( 3) Establishing the quality standard of compound capsules in the treatment of osteoarthritis : Determination and related substances of compound ademetionine and meloxicam was examined by HPLC method.Method validation results show that the method is stable, accurate and reproducible. In vitro release test showed, three batches of samples ademetionine and meloxicam are no dissolution in pH as dissolution medium of 1.2 at 2 hours internal, but the release were in accordance with the relevant provisions in pH as dissolution medium of 6.8 at 45 min internal.Conclusion:S-ademetionine compatibility of meloxicam caused by heat and chemical stimulus pain effectively. the ademetionine and meloxicam can slow the pathological process in OA rat model. The compatibility can improve the curative effect of different level.The mechanism may be associated with the disorder of COMP and MMP-3 in serum, and the reduction of inflammation factors in serum and synovial fluid. Treatment of osteoarthritis of compound enteric coated microcapsules preparation process is stable and feasible, the quality of stable and controllable,and in vitro dissolution trials have shown good enteric properties, suitable for in the later stages of industrialization production. The established HPLC method have accurate,stable and good repeatability,and can be used as a method for determiningthe content of compound ademetionine with meloxicam.