目的 研究采用靶向代谢组学方法，观察苍术麸炒前后对脾虚大鼠血清氨基酸水平的变化，探讨苍术炮制机制。方法 72只大鼠随机分成正常组、模型组、低剂量生苍术组(生低组)、中剂量生苍术组(生中组)、高剂量生苍术组(生高组)、低剂量麸炒苍术组(麸炒低组)、中剂量麸炒苍术组(麸炒中组)和高剂量麸炒苍术组(麸炒高组)，共8组，每组9只。除正常组外，模型组通过过度疲劳、苦寒泻下和饮食不节法造模。对各组大鼠采用酶联免疫吸附测定(Enzyme Linked Immunosorbent Assay，ELISA)方法测定颌下腺中水通道蛋白 3(Aquaporin-3，AQP-3)、水通道蛋白 4(AQP-4)、水通道蛋白5(AQP-5)、水通道蛋白8(AQP-8)的含量，结肠中水通道蛋白1(AQP-1)、水通道蛋白2(AQP-2)、水通道蛋白9(AQP-9)、紧密连接蛋白1(zonula occludes protein-1，ZO-1)的含量。对生中组和麸炒中组采用超高效液相色谱-三重四极杆质谱(Ultra high performance liquid chromatography-triple quadrupole tandem mass spectrometry，UH-PLC-MS/MS)检测血清中24种氨基酸水平的变化。结果 与正常组比较，模型组中颌下腺AQP-3、AQP-4、AQP-5、AQP-8的含量显著降低(P＜0。05);模型组中结肠AQP-1和AQP-2的含量显著升高(P＜0。05)，AQP-9和ZO-1的含量显著降低(P＜0。05)。生苍术和麸炒苍术对上述指标均有调节作用。经进一步同等剂量的生苍术和麸炒苍术相比，麸炒中组使上述指标含量升高更明显。与正常组比较，模型组血清中L-缬氨酸等4种物质的水平显著升高(P＜0。05)，L-酪氨酸等11种物质的水平均显著降低(P＜0。05)。经进一步比较，麸炒苍术对上述指标调节效果优于生苍术。靶向代谢通路分析表明，这些差异代谢物主要与精氨酸和脯氨酸代谢、氨酰tRNA生物合成、甘氨酸、丝氨酸和苏氨酸代谢、乙醛酸和二羧酸代谢通路有关。结论 苍术炮制后的增效机制可能与对脾虚大鼠血清中氨基酸调节有关。
Targeted Metabolomic Analysis of Amino Acids with Spleen Deficiency Syndrome Rats after Administering Raw and Bran-Fried Cangzhu(Atractylodis Rhizoma)
Objective This study used targeted metabonomics to observe the effects of raw and bran-fried crude Cangzhu(At-ractylodis Rhizoma)on the changes of serum amino acid levels in rats with spleen deficiency syndrome rats and explore the pro-cessing mechanism of Cangzhu(Atractylodis Rhizoma).Methods Seventy-two rats were randomly assigned to control group(CG),model group(MD),raw Cangzhu(Atractylodis Rhizoma)low-dose group(LR),raw Cangzhu(Atractylodis Rhizoma)medium-dose group(MR),raw Cangzhu(Atractylodis Rhizoma)high dose group(HR),bran-fried Cangzhu(Atractylodis Rhi-zoma)low-dose group(LB),bran-fried Cangzhu(Atractylodis Rhizoma)medium dose group(MB)and bran-fried Cangzhu(Atractylodis Rhizoma)high dose group(HB),eight groups with 9 in each group.In addition to the control group,the model group adopted the complex method of improper diet,excessive fatigue and bitter cold diarrhea to create the spleen deficiency mod-el of rats.The contents of aquaporin(AQP)-3,AQP-4,AQP-5 and AQP-8 in submandibular gland,the contents of AQP-1,AQP-2,AQP-9 and zonula occludes protein-1(ZO-1)in intestinal were determined by enzyme-linked immunosorbent assay(ELISA).The changes of 24 kinds of amino acids in serum of MR and MB groups were detected by Ultra High Performance Liquid Chromatography-Triple Quadrupole Tandem Mass Spectrometry(UHPLC-MS/MS).Results Comparing with those of the CG group,there were lower levels of AQP-3,AQP-4,AQP-5 and AQP-8 in submandibular gland of MG group(P＜0.05),higher levels of AQP-1 and AQP in intestinal of MG group(P＜0.05),lower levels of AQP-9 and ZO-1 in intestinal of MG group(P＜0.05).Both raw and bran-fried crude CangZhu(Atractylodis Rhizoma)can regulate the above indexes.Af-her further comparison with the same dose of raw and bran-fried crude CangZhu(Atractylodis Rhizoma),the content of the a-bove indexes in the MB group was higher than in the HR group.Compared with those of the CG group,the levels of L-Valine and 4 kinds of substances in the serum increased significantly(P＜0.05),while the levels of L-Tyrosine and 11 kinds of sub-stances significantly decreased(P＜0.05).After further comparison,the adjustment effect of bran-fried crude CangZhu(At-ractylodis Rhizoma)was better than raw crude CangZhu(Atractylodis Rhizoma).Targeted metabolomic pathway analysis showed that these differential metabolites were mainly related to the arginine and proline metabolism,aminoacyl-tRNA biosynthesis,gly-cine,serine and threonine metabolism,glyoxylate and dicarboxylate metabolism.Conclusion The synergistic mechanism of bran-fried Cangzhu(Atractylodis Rhizoma)may be related to the regulation of amino acids in serum of rats with spleen deficiency.
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