| Objective:To study the critical influence factors in the preparation tablets of fresh antler by lyophilized method and to optimize the preparation technology. In order to provide a reference for the packaging, storage conditions and stability of quality, the author decided to measure the stability and critical relative humidity (CRH) of the lyophilized tablets of fresh antler. In addition, the author also did the preliminary evaluation experiments to demonstrate the lyophilized molding technology has effects on preserving fresh antler peptides activity effectively and provide theoretical support and technical support on the others biopharmaceuticals with large number of peptide and protein by the lyophilized molding technology.Methods:Fresh antler as the model drug, the lyophilized molding technology was used to prepare the tablets of fresh antler and preparation were obtained by means of single factor test with promoting cell proliferation activity, moisture content, disintegration time, and appearance as evaluation criterion. The changes in appearance, moisture, disintegration time and the biological activity of fresh antler tablets under high humidity, high temperature and strong light, accelerated test and long term test were investigated and got its CRH by the reducing heavy law. The preliminary evaluation both quantitative and pharmacodynamics of fresh antler, antler fresh freeze-dried preparation and processing velvet were evaluated by comparative evaluation method. While diabetic wound was considered as the animal model in pharmacodynamics study, and the wound healing rate, histological changes, tumor necrosis factor(TNF-a), vascular endothelial growth factor(VEGF) and hydroxyproline(HOP) content as indicator to evaluate the effect on wound healing and provide new processing method on velvet activity preservation.Results:(1) The optimal preparation conditions of the lyophilized fresh antler tablets were as follows:The frozen storage temperature of fresh antler was-40℃, acetate buffer was used as diluent to mix with the fresh antler powder according to ratio of 1’.2, then to homogenize for 10 min under liquid nitrogen protection with 10% trehalose as lyoprotectant, and the homogenate was lyophilized in vacuum after poured into the mold to prepare the lyophilized fresh antler tablets. The tablets could disintegrate within 30s completely and its moisture content was less than 7%. In addition, there was no difference in promoting cell proliferation compared with fresh antler.(2) The stability results were as follow:under influence factors indicate the tablets were stable under strong light, while the appearance, moisture, disintegration time under high humidity, and the biological activity under high temperature have changed obviously(p<0.01). To compare with 0 month, the appearance, moisture, disintegration time have changed under accelerated test and long term test, the changes were not fitted the requirements of orally disintegrating tablets. Its CRH is 72.59%.(3) The preliminary evaluation results were as follow:a. The soluble protein contents of fresh antler, lyophilized fresh antler tablets and processing antler were 4.90±0.11,3.77±0.12 and 0.31±0.03 mg/g. To compared with processed products the soluble protein content, there was a significant difference (p<0.01) in fresh antler and lyophilized fresh antler tablets.b. The NIH/3T3 cell proliferation activity of fresh antler, lyophilized fresh antler tablets and processing antler were 41.36±11.28,35.36±9.64 and 19.74±3.63%. To compared with processed group, there was a significant difference on cell proliferation activity (p<0.01) in fresh antler, lyophilized fresh antler tablets.c. The effects on chronic wound healing in diabetic of fresh antler, lyophilized fresh antler tablets and processing antler have showed, to compared with the lyophilized fresh antler group (80.47±6.79), the wound healing rate of processing antler(64.9±16.63) and control group(63.07±13) have an obviously difference (p<0.05 orp<0.01); in addition, there were obvious changes in inflammatory cells infiltration, telangiectasia and proliferation, TNF-a, VEGF levels of fresh antler, lyophilized fresh antler tablets were significantly higher than the control group and the processing antler group in the early wound healing. Compared with fresh group, TNF-a, VEGF level of lyophilized group showed no significant difference (p>0.05) at all time points. Therefore the author inferred the lyophilized molding technology could save the activity of velvet proteins effectively.Conclusion:The lyophilized fresh antler tablets prepared by optimal technology has highly biological activity, the other indexes were fit for the standardization and was easy to take. However it is very sensitive to humidity and temperature, they are the major influence factors on its quality. So it is necessary to package with moistureproof, store sealed in a cool dry place with the relative humidity is under 72.59%. In addition, to compare with the processing antler, the protein content and biological activity in lyophilized fresh antler tablets has significantly improved. It may was announced velvet protein content and biological activity could be saved effectively by lyophilized molding technology, and the lyophilized molding technology could apply to this aspects widely, which include numerous protein to improve its availability.
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