基于经皮给药途径研究白芍对制川乌6个酯型生物碱组织分布的影响

Research on the effects of Paeoniae Radix Alba on the tissue distribution of six aconite alkaloids in Aconiti Radix Cocta based on transdermal administration

目的:探讨制川乌配伍白芍经皮给药对制川乌中6个酯型生物碱组织分布的影响。方法:SD大鼠随机分为2组,分别经皮给予制川乌-白芍凝胶及制川乌凝胶,连续5 d,末次给药后4、8、20 h采集大鼠皮肤、脑、心、肺、肝、脾、肾等组织,以溶剂萃取法对样品进行提取,应用LC-MS/MS方法对组织中6个酯型生物碱乌头碱(AT)、次乌头碱(HT)、新乌头碱(MT)、苯甲酰乌头原碱(BA)、苯甲酰次乌头原碱(BH)、苯甲酰新乌头原碱(BM)的含量进行测定。采用spss统计软件对各组织中药量进行t检验,比较配伍前后6个酯型生物碱在各组织中分布的变化情况。应用主成分分析法(principal component analysis,PCA)、偏最小二乘判别法(partial least quares discriminant analysis,PLS-DA)分析配伍对6个酯型生物碱在皮肤及毒靶器官心脏、脑组织中分布的影响。结果:大鼠多次经皮给予制川乌凝胶后,6个酯型生物碱分布较广,主要分布在皮肤,其次为肝、脾、肺、肾,而在脑、心脏组织中的含量较低;配伍后单酯型生物碱苯甲酰新乌头原碱(BM)、苯甲酰乌头原碱(BA)在皮肤组织中的分布显著增加,而3个单酯型生物碱苯甲酰新乌头原碱(BM)、苯甲酰乌头原碱(BA)、苯甲酰次乌头原碱(BH)在心脏和脑组织的分布明显减少;同时双酯型生物碱新乌头碱(MT)、次乌头碱(HT)、乌头碱(AT)在各组织中的分布均呈下降趋势;基于皮肤、心脏、脑组织中6个酯型生物碱的分布规律,利用PCA统计方法可将实验组分为单用组与配伍组;PLS-DA法分析显示,白芍对制川乌中6种酯型生物碱在皮肤、心脏、脑组织中分布的影响由大到小的顺序分别为HT、AT、MT、BM、BH、BA,MT、AT、BA、BH、BM、HT,AT、BH、BA、HT、MT、BM。结论:制川乌配伍白芍经皮给药,可促进部分单酯型生物碱在皮肤中的分布与吸收,同时减少6个酯型生物碱在心脏、脑等毒靶器官的分布,从而发挥"增效减毒"的作用。

Objective： To investigate the effects of Paeoniae Radix Alba on the tissue distribution of six aconite alkaloids in herb couple of Aconiti Radix Cocta-Paeoniae Radix Alba after transdermal administration in rats. Methods： The Aconiti Radix Cocta gel and Aconiti Radix Coeta combined with Paeoniae Radix Alba gel were continuously administered to SD rats of SPF grade for five days, respectively. Tissues of skin, brain, heart, lung, liver, spleen and kidney were collected at 4, 8 and 20 h after transdermal administration. Samples were extracted by ethyl acetate and six aconite alkaloids including aconitine （ AT ）, hypaeonitine （ HT ） , meaconitine （ MT ）, benzoylaconine （ BA ）, benzoylhypaconine （ BH ）, and benzoylmesaconine （ BM ） in each tissue were determined by LC-MS/MS method. Data were analyzed using SPSS software to compare tissue distribution of six aconite alkaloids based on t-test. Principal component analysis （ PCA ） and partial least squares diseriminant analysis （ PLS-DA ） were used to identify the changes of six aconite alkaloids content in tissues of skin, brain and heart. Results： In Aconiti Radix Cocta group, the six aconite alkaloids were mainly distributed in skin, followed with liver, spleen, lung and kidney, while low in brain and heart. The distribution of BM and BA in the skin tissue was significantly increased after compatibility, but in the heart and brain tissue, the distribution of 3 kinds of mono ester alkaloids i. e. BM, BA and BH were significantly reduced. At the same time, the contents of the diester-type alkaloids of MT, HT and AT showed a downward trend in every tissue when Aconiti Radix Cocta compatibility with Paeoniae Radix Alba. Based on the distribution of 6 alkaloids in skin, heart and brain, PCA could directly classify the Aconiti Radix Cocta group and compatibility group. The PLS-DA indicated that Paeoniae Radix Alba could influence the distribution of 6 alkaloids in skin, heart and brain, and the effect sequences of 6 alkaloids from large to small were as follows： HT, AT, MT, BM, BH, BA; MT, AT, BA, BH, BM, HT; AT, BH, BA, HT, MT, BM, respectively. Conclusion： The compatibility of Paeoniae Radix Alba could decrease the toxicity of Aconiti Radix Cocta, and promote the absorption and distribution of partial mono-ester aconite alkaloids at the administration site such as skin, meanwhile, reduce the distribution of the six aconite alkaloids in brain and heart, which was beneficial to lower the toxicity of the Aconiti Radix Cocta in central nervous system and cardiovascular system.