Investigation Of Collagen Double Layer Minipellet Of Erythropoietin

Erythropoietin (EPO) , a glycoprotein hormone which has a molecular weight of 30,400Da, is the main regulator of human erythropoiesis. EPO is clinically used for therapy of renal anemia and related conditions. Recombination human erythropoietin (rh-EPO) is widely used in clinic because EPO produced in kidney is very little. There are many disadvantages when EPO is injected for therapy of renal anemia and some malignant tumors, such as repeated administration of large amounts of drug, long course of administration. We have developed a matrix system for the slow delivery of EPO using collagen as a biodegradable drug carrier, the double layer minipellet (DLMP). Collagen has suitable characteristics for medical applications, which bring DLMP many advantages including slow delivery, large amount of loading drugs. Therefore EPO double layer minipellet (EPO-DLMP) is expected to improve the stability of EPO in formulations and reduce the frequency of administration.In order to control the quality of EPO-DLMP, the method for determination of EPO quantity by HPLC was established, which could measure the content of EPO in EPO-DLMP accurately.According to the behavior of EPO-DLMP in vitro release, various factors were studied. The main variables were the collagen/EPO ratios (A), various contents of CS (B), various diameters of extrusion nozzles(C) and EPO-DLMP with various lengths (D). The orthogonal experimental design was chosen to optimize the condition of formula of EPO-DLMP. The analysis showed (A) 1000:1, (B) 1%, (C) 1.4mm and (D) 15mm were the optimal formulation. Under the optimal formulation, the numeric value of deviation was 7.59.The quantity of EPO in EPO-DLMP was measured by ELISA and the stability of EPO-DLMP was valuated with the indexes of content changes of EPO in EPO-DLMP in 4 ℃ and relative humidity 25% for 30 days. The results showed that the EPO-DLMP stored in the conditions mentioned above would keep EPO stable.The phannacokinetic parameters of market EPO injection (EPO-I) and EPO-DLMP were studied after subcutaneous administration EPO-I or EPO-DLMP to rabbits. EPO concentration in serum was determined by ELISA. Comparing with EPO-I, it showed longer elimination half life, larger area under the curve values and lower clearance, longer Tmax of EPO-DLMP (P < 0.05). The results of phannacokinetics showed that EPO-DLMP had better slow-release effect than EPO-I.The pharmacodynamics of the two formulations were evaluated by the method of reticulocyte count after they were subcutaneously administrated. The results showed that EPO-DLMP could increase the numbers of reticulocyte persistently and effectively which is potential to treat anemia.