Extraction Process of Total Alkaloids of Uncaria Rhynchophylla, and Its Acute Toxicity and Protective Effect on HUVEC

OBJECTIVE To optimize the optimal extraction process of Uncaria rhynchophylla by orthogonal experiment, to study the acute toxicity of Uncaria rhynchophylla total alkaloids to mice and protective effect on H₂O₂-induced injury of human umbilical vein endothelial cells(HUVEC). METHODS The L₉(3⁴) orthogonal test method was used, ethanol concentration, extraction times, extraction time, and material ratio were selected as the factors to be investigated, the contents of isorhynchophylline, rhynchophylline, hirsuteine and hirsutine were determined by HPLC, the transfer ratios were calculated, and the comprehensive score of principal component analysis was taken as the index to optimize the extraction process of Uncaria rhynchophylla total alkaloids. KM mice were selected. After intragastric administration of total alkaloids solution of Uncaria rhynchophylla for twice, maximum tolerated dose(MTD) was calculated, the changes of mice within 14 d were observed. The eyeballs were removed to collect blood. The contents of urea nitrogen(BUN), glutamic-pyruvic transaminase(GPT) and glutamic- oxaloacetic transaminase(GOT) in the serum of mice were determined. HUVECs were selected and damaged by 1.0 mmol·L^-1 H₂O₂ to observe the protective effects of total alkaloids of Uncaria rhynchophylla. The survival rate of HUVECs and the contents of lactate dehydrogenase(LDH), endothelin-1(ET-1), nitric oxide(NO) in the supernatant were measured. RESULTS The optimum paratemeters of Uncaria rhynchophylla total alkaloids extraction process were as following:Uncaria rhynchophylla was extracted by 70% ethanol for twice, each time for 2 h, and the material ratio was 1:8. Within 14 d of the acute toxicity test, the mice's activity was normal, no obvious adverse reactions were observed, and no death after administration; the MTD was 21.0 g·kg^-1·d^-1, which was equivalent to the daily dose of clinical adults 1 377 times. Compared with blank group, GPT in Uncaria rhynchophylla total alkaloids group was significantly decreased(P<0.01). Compared with the blank group, the survival rate of HUVEC and the NO content in the supernatant of the model group were significantly decreased(P<0.01 or P<0.05), while the LDH and ET-1 contents were significantly increased(P<0.01); compared with the model group, the cell viability of Uncaria rhynchophylla total alkaloids(5 μg·mL^-1) was significantly increased(P<0.01), and the content of LDH and ET-1 in the supernatant was significantly decreased(P<0.01 or P<0.05). CONCLUSION The optimized extraction process of total alkaloids of Uncaria rhynchophylla is simple and feasible, total alkaloids of Uncaria rhynchophylla has less acute toxicity in mice, and has a protective effect on HUVECs damage.