[關(guān)鍵詞]
[摘要]
目的 超濾法結(jié)合超高效液相色譜(UPLC)法同時檢測抗601合劑中2種指標性成分——黃芩苷、綠原酸體外血漿蛋白結(jié)合率。方法 建立UPLC法同時測定牛血清白蛋白(BSA)、大鼠空白血漿和正常人空白血漿超濾液中的黃芩苷、綠原酸含量,進行專屬性、精密度、準確度、穩(wěn)定性、超濾膜回收率等方法學考察;配制黃芩苷質(zhì)量濃度分別為28.2、141.0、282.0 mg/L,綠原酸質(zhì)量濃度為6.40、32.0、64.0 mg/L的抗601合劑溶液,分別與BSA溶液、大鼠空白血漿、人空白血漿混勻后,置于37℃水浴溫育4 h,于超濾離心管中10 000 r/min離心20 min,取超濾液10 μL注入UPLC分析測定,計算超濾液中黃芩苷和綠原酸濃度及血漿蛋白結(jié)合率。結(jié)果 建立的UPLC法專屬性、精密度、準確度、穩(wěn)定性、超濾膜回收率均良好,能夠滿足定量分析測試要求;黃芩苷與BSA、大鼠空白血漿和正常人空白血漿的蛋白結(jié)合率分別為(73.70±1.65)%、(82.72±1.64)%、(87.33±1.84)%,綠原酸與3種血漿的蛋白結(jié)合率分別為(66.78±1.91)%、(74.18±2.01)%、(78.54±2.97)%,無濃度相關(guān)性,兩成分與BSA溶液蛋白結(jié)合率顯著低于與大鼠空白血漿、人空白血漿結(jié)合率(P<0.05)。結(jié)論 抗601合劑中的黃芩苷、綠原酸與不同種屬血漿均有較高的蛋白結(jié)合率,在考察范圍內(nèi)無濃度相關(guān)性,歸類于中速處置類藥物,體內(nèi)消除情況與其血漿蛋白結(jié)合率可能密切相關(guān)。
[Key word]
[Abstract]
Objective Ultrafiltration combined with ultra high performance liquid chromatography (UPLC) method were established for the simultaneous determination of plasma protein binding rate of two index components baicalin and chlorogenic acid in Anti-601 Mixture. Methods The UPLC method was established for the simultaneous determination of baicalin and chlorogenic acid in bovine serum albumin (BSA), blank plasma and normal human blank plasma ultrafiltration, and the methods of specificity, precision, accuracy, stability and ultrafiltration recovery rate were investigated. The Anti-601 Mixture of 3 concentrations was prepared with the concentration of baicalin was 28.2, 141 and 282 mg/L, and the concentration of chlorogenic acid was 6.40, 32.0 and 64.0 mg/L. After mixing the Anti-601 Mixture with BSA solution, rat blank plasma and human blank plasma respectively, the solutions were incubated at 37 C in water bath for 4 h, and placed in the ultrafiltration centrifuge tube for 10000 r/min centrifugation 20 min, and 10 μL ultrafiltration was injected into UPLC to determine the concentration of baicalin and chlorogenic acid in the ultrafiltration and the binding rate of plasma protein. Results The method had high sensitivity, good specificity and reproduction, with simple management in fulfilling the requirements. The plasma protein binding rate of baicalin with BSA, rat plasma and human plasma were (73.70±1.65)%, (82.72±1.64)%, (87.33±1.84)% respectively, and the plasma protein binding rate of chlorogenic acid with BSA, rat plasma and human plasma were (66.78±1.91)%, (74.18±2.01)%, (78.54±2.97)% respectively. No concentration correlation was observed. The binding rate of two components to BSA solution protein was significantly lower than that of rat blank plasma and human blank plasma (P<0.05). Conclusion The two major compounds in Anti-601 Mixture showed a high binding power to plasma protein of different species in vitro, and it was independent of the investigated concentrations. It is classified as medium speed treatment drugs, elimination in vivo may be closely related to plasma protein binding rate.
[中圖分類號]
[基金項目]
十三五南京市衛(wèi)生青年人才培養(yǎng)工程第三層次(QRX17173);南京藥學會—常州四藥醫(yī)院藥學科研基金(2016YX010);南京醫(yī)科大學科技發(fā)展基金面上項目(2015NJMU068)