Effects of Prenylated Compounds Derived from Psoralea Corylifolia against Human CYP1A1 and Corresponding Molecular Docking Validation
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摘要: 目的 考察补骨脂富含的异戊烯基成分对CYP1A1活性的影响, 并采用分子对接技术进行验证。方法 以7-乙氧基试卤灵为CYP1A1的探针底物, 采用超高效液相串联三重四级杆质谱(UHPLC-TQD-MS)对代谢产物试卤灵的质谱条件进行优化; 应用CYP1A1体外孵育体系, 评价补骨脂富含的补骨脂二氢黄酮、补骨脂定、异补骨脂查尔酮、异补骨脂二氢黄酮和补骨脂二氢黄酮甲醚在不同浓度(1、10、100 μmol·L-1)对CYP1A1活性的影响; 采用不同的抑制动力学模型计算相关的抑制常数(Ki); 应用Autodock 4.2软件, 将活性成分与CYP1A1进行分子对接。结果 初筛结果显示, 补骨脂二氢黄酮、补骨脂定、异补骨脂查尔酮、异补骨脂二氢黄酮、补骨脂二氢黄酮甲醚对CYP1A1均表现出抑制活性, IC50值分别为0.28、0.49、0.36、2.24、4.07 μmol·L-1; 根据最小赤池信息量准则(AIC)和施瓦茨信息准则值(SC)即为最佳模型的原则, 补骨脂二氢黄酮和异补骨脂查尔酮对CYP1A1的抑制为竞争性抑制, Ki常数分别为0.12、0.23 μmol·L-1; 补骨脂定对CYP1A1表现出非竞争性抑制, 抑制常数Ki为0.59 μmol·L-1; 分子对接结果显示, 补骨脂二氢黄酮和异补骨脂查尔酮与CYP1A1蛋白之间均能产生氢键、π-π键以及疏水作用, 结合自由能分别为-10.145、-8.286 kcal·mol-1(1 kcal=4.2 kJ), 这可能是它们与CYP1A1亲和力较强进而产生强抑制活性的原因。结论 补骨脂富含的异戊烯基成分是CYP1A1强抑制剂(IC50 < 5 μmol·L-1); 异戊烯基成分C环1位和2位脱水开环会增加对CYP1A1的抑制活性; A环C-6位异戊烯基取代的化合物比C-8位取代的化合物表现出更强的CYP1A1抑制活性; A环7-OH甲基化会减弱CYP1A1的抑制活性; 分子对接结果也证实了补骨脂二氢黄酮和异补骨脂查尔酮与CYP1A1有较强的亲和力。Abstract: OBJECTIVE To investigate the effects of prenylated compounds derived from Psoralea corylifolia against human CYP1A1, and validate the obtained results by molecular docking.METHODS 7-Ethoxyresorufin was used as the probe substrate for human CYP1A1. In addition, the ultra-high performance liquid chromatography tandem triple quadrupole mass spectrometry (UHPLC-TQD-MS) conditions for resorufin (the specific metabolite of 7-ethoxyresorufin by CYP1A1) were optimized. Based on the optimized condition, 7-ethoxyresorufin was incubated with CYP1A1 in the absence (control) and presence of bavachin, psoralidin, isobavachalcone, isobavachin and bavachinin at different concentration (1, 10, 100 μmol·L-1), respectively. Three kinetic models were applied to calculate the Kivalues by nonlinear regression for competitive inhibition, noncompetitive inhibition, and mixed-type inhibition, respectively. The molecular docking between prenylated compounds and CYP1A1 was performed by Autodock 4.2.RESULTS These findings demonstrated that bavachin, psoralidin, isobavachalcone, isobavachin and bavachinin exhibited potent inhibitory effects against CYP1A1, and the corresponding IC50values were 0.28, 0.49, 0.36, 2.24, 4.07 μmol·L-1, respectively. Based on the smallest Akaike information criterion (AIC) and Schwarz criterion (SC) principles, bavachin and isobavachalcone exerted competitive inhibition against CYP1A1 with Ki values of 0.12 and 0.23 μmol·L-1, respectively, while psoralidin displayed noncompetitive inhibitory effects against CYP1A1(Ki=0.59 μmol·L-1). Molecular docking results indicated that hydrogen bond, π-π bond and hydrophobic interaction could be formed between bavachin (or isobavachalcone) and CYP1A1 protein crystals, and their binding free energies were -10.145 and -8.286 kcal·mol-1(1 kcal=4.2 kJ), respectively. This might be the reason why these compounds have strong affinity with CYP1A1, and thus produce strong inhibitory activity.CONCLUSION Prenylated compounds in Psoralea corylifolia are identified as potent inhibitors for CYP1A1 (IC50 < 5 μmol·L-1). When dehydration reaction occurrs in bonds 1 and 2, the inhibitory effects of herbal compounds against CYP1A1 increases. Herbal compounds with isopentene group at C-6 position exhibits stronger inhibitory effects against CYP1A1 than those with isopentene group at C-8 position. Herbal compounds with C-7-OCH3 displays weaker inhibitory effects against CYP1A1 than those with C-7-OH. The molecular docking results also validates the presence of strong affinity between bavachin (or isobavachalcone) and CYP1A1 protein.
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表 1 补骨脂异戊烯基成分对CYP1A1抑制作用的动力学参数(x±s, n=3)
化合物 IC50/(μmol·L-1) 类型 Ki/(μmol·L-1) α R2 AIC SC 补骨脂提取物 14.97±2.17# — — — — — — 补骨脂二氢黄酮 0.28±0.05 竞争性 0.12±0.02 — 0.988 8 -100.21 -97.22 非竞争性 0.26±0.02 — 0.983 5 -92.32 -89.33 混合型 0.14±0.03 8.97±11.26 0.989 4 -99.23 -95.25 补骨脂定 0.49±0.25 竞争性 0.28±0.05 — 0.965 8 -82.00 -79.01 非竞争性 0.59±0.04 — 0.984 7 -98.03 -95.05 混合型 0.87±0.29 0.48±0.27 0.986 0 -97.89 -93.91 异补骨脂查尔酮 0.36±0.02 竞争性 0.23±0.03 — 0.983 5 -110.90 -107.91 非竞争性 0.60±0.06 — 0.970 6 -99.42 -96.44 混合型 0.24±0.05 36.49±123.24 0.983 5 -109.01 -105.03 异补骨脂二氢黄酮 2.24±0.70 — — — — — — 补骨脂二氢黄酮甲醚 4.07±0.85 — — — — — — 注: #代表IC50值的单位是mg·mL-1。 
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