独活凝胶贴膏穴位贴敷治疗类风湿关节炎模型家兔的药动学与药效学研究

Pharmacokinetics and Pharmacodynamics Study of Acupoint Application with Duhuo Gel Plaster in the Treatment of Rheumatoid Arthritis in Model Rabbits

目的研究独活凝胶贴膏穴位贴敷治疗类风湿关节炎(RA)模型家兔的药物代谢动力学(PK)与药物效应动力学(PD)。方法取10只健康新西兰雌性家兔,2只作为正常对照组;其余8只家兔肩胛区注射卵清蛋白佐剂构建RA模型,随机分为模型组(空白凝胶贴膏)和穴位给药组(独活凝胶贴膏),各4只。采用微透析取样技术对模型家兔的关节腔液取样,采用超高效液相色谱-串联质谱(UPLC-MS/MS)法测定关节腔液中独活有效成分二氢欧山芹醇当归酸酯和蛇床子素的含量,并建立PK参数;采用酶联免疫吸附(ELISA)法测定血清中白细胞介素1β(IL-1β)、肿瘤坏死因子-α(TNF-α)的水平,并计算抑制率,作为PD指标;采用WinNonlin ExamplesGuide 5.2.1软件拟合PK-PD模型。结果穴位给药组家兔关节腔液中蛇床子素一级吸收速率常数(k_(01))为0.22 h^(-1),一级消除速率常数(k_(10))为0.10 h^(-1),0~14 h的血药浓度-时间曲线下面积(AUC_(0~14 h))为6584.18(ng•h)/mL,吸收相半衰期(k_(01)_LH)为2.54 h,消除相半衰期(k_(10)_LH)为5.89 h,达峰时间(t_(max))为5.70 h,达峰浓度(C_(max))为454.31 ng/mL;关节腔液中二氢欧山芹醇当归酸酯k_(01)为0.20 h^(-1),k_(10)为0.12 h^(-1),AUC_(0~14 h)为2185.67(ng•h)/mL,k_(01)_HL为2.69 h,k_(10)_HL为5.94 h,t_(max)为6.52 h,C_(max)为150.53 ng/mL。穴位给药组家兔血清中IL-1β和TNF-α水平建立的抑制百分数(E_(IR))与药效中效应室浓度(Ce)的PK-PD模型方程分别为E_(IR)=91.72Ce/598.35+Ce,E_(IR)=95.72 Ce/568.08+Ce。结论蛇床子素和二氢欧山芹醇当归酸酯在RA模型家兔体内的PK行为与药效紧密联系,独活凝胶贴膏经穴位给药可达到预期疗效。

Objective To study the pharmacokinetics(PK)and pharmacodynamics(PD)of acupoint application with Duhuo Gel Plaster in the treatment of rheumatoid arthritis(RA)in model rabbits.Methods Ten healthy New Zealand female rabbits were selected,two were taken as the normal control group,the other eight rabbits were injected with ovalbumin adjuvant into the scapular area to construct RA model and they were randomly divided into the model group(blank gel patch)and acupoint administration group(Duhuo Gel Plaster),with four rabbits in each group.Microdialysis sampling technology was used to sample the articular cavity fluid of model rabbits.Ultra-high performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS)method was used to determine the content of active ingredients columbianadin and osthole in Angelicae Pubescentis Radix in the articular cavity fluid,and PK parameters were established.Enzyme-linked immunosorbent assay(ELISA)was used to determine the levels of serum interleukin 1β(IL-1β)and tumor necrosis factor-α(TNF-α),and the inhibition rate was calculated as a PD index.WinNonlin Examples Guide 5.2.1 software was used to fit the PK-PD model.Results The first-order absorption rate constant(k_(01))of osthole in the articular cavity fluid of rabbits in the acupoint administration group was 0.22 h^(-1),and the first-order elimination rate constant(k_(10))was 0.10 h^(-1).The area under the plasma concentration-time curve from 0 to 14 h(AUC_(0-14 h))was 6584.18(ng·h)/mL,the half-life of the absorption phase(k_(01)_LH)was 2.54 h,the half-life of the elimination phase(k_(10)_LH)was 5.89 h,the peak time(t_(max))was 5.70 h,and the peak concentration(C_(max))was 454.31 ng/mL.Columbianadin k_(01) in articular cavity fluid was 0.20 h^(-1),k_(10)was 0.12 h^(-1),AUC0-14 h was 2185.67(ng·h)/mL,k_(01)_HL was 2.69 h,k_(10)_HL was 5.94 h,t_(max) was 6.52 h,and C_(max) was 150.53 ng/mL.The PK-PD model equations of the inhibitory percentage(E_(IR))established by the levels of serum IL-1βand TNF-αin the acupoint administration group and the concentration-effect(Ce)relationship in PD were E_(IR)=91.72Ce/598.35+Ce,E_(IR)=95.72Ce/568.08+Ce,respectively.Conclusion The PK behaviors of osthole and columbianadin in RA model rabbits are closely related to the PD.Duhuo Gel Plaster can achieve the expected efficacy by acupoint administration.