小叶黄杨TLC鉴别及不同部位中紫丁香苷和香草酸HPLC测定

目的：建立小叶黄杨中紫丁香苷和香草酸的TLC鉴别和HPLC含量测定方法，研究药材中不同部位紫丁香苷和香草酸的含量差异。方法：TLC鉴别法，使用硅胶GF₂₅₄薄层板，以三氯甲烷-甲醇-甲酸（8：1.2：0.5）作为展开剂；HPLC法，使用Xbridge C₁₈色谱柱（4.6 mm×250 mm，5 μm），以甲醇-0.2%磷酸水溶液（17：83）为流动相，流速1 mL·min^-1，检测波长264 nm，柱温35℃。结果：TLC法操作简便，准确，斑点清晰，专属性好。紫丁香苷、香草酸质量浓度分别在3.36~302.4和0.55~695.5 μg·mL^-1（r=0.999 5）范围内线性关系良好，3种浓度水品平均回收率分别在99.8%~103.0%和98.4%~101.1%范围内。小叶黄杨不同部位中紫丁香苷和香草酸的含量存在差异，紫丁香苷在粗茎、细枝、叶和地上部分中的含量分别为0.030~0.324、0.002~0.518、0.013~0.907和0.010~0.667 mg·g^-1，香草酸在粗茎、细枝、叶和地上部分中的含量分别为0.025~0.077、0.021~0.087、0.012~0.132和0.020~0.123 mg·g^-1。结论：该方法能鉴别小叶黄杨，准确测定紫丁香苷和香草酸的含量，为开发利用小叶黄杨资源提供了科学依据。

Objective: To establish the methods of TLC identification and HPLC determination of the contents of syringin and vanillic acid in Buxus sinica, and determine different contents of syringin and vanillic acid in different fractions of Buxus sinica.Methods: The thin-layer plate silica of gel GF₂₅₄ and organic solvent chloroformmethanol-formic acid(8:1.2:0.5, v/v/v)were used in TLC identification. The HPLC method was performed on a Xbridge C₁₈ chromatographic column(4.6 mm×250 mm, 5 μm)with the mobile phase consisting of methanol -0.2% phosphoric acid solution(17:83)at the flow rate of 1 mL·min^-1.The detection wavelength was set at 264 nm, and the column temperature was set at 35℃.Results: TLC method was simple and accurate, and the spots were clear with good separation. The linear range of syringin and vanillic acid were 3.36-302.4 and 0.55-695.5 μg·mL^-1(r=0.999 5), and the recoveries on three concentration levels were 99.8%-103.0% and 98.4%-101.1% respectively. The contents of syringin and vanillic acid were obviously different in different parts of Buxus sinica. The contents of syringin in main stem, withe, leaves and aerial part were 0.030-0.324 mg·g^-1, 0.002-0.518 mg·g^-1, 0.013-0.907 mg·g^-1 and 0.010-0.667 mg·g^-1, respectively. The contents of vanillic acid in main stem, withe, leaves and aerial part were 0.025-0.077 mg·g^-1, 0.021-0.087 mg·g^-1, 0.012-0.132 mg·g^-1 and 0.020-0.123 mg·g^-1, respectively.Conclusion: The method can be used to identify Buxus sinica effectively, and to determine the contents of syringin and vanillic acid in different parts accurately, which provides a scientific basis for development and utilization Buxus sinica resources.

[1] 林云良,邱明华,李忠荣.黄杨中的非生物碱化学成分[J].云南植物研究, 2006, 28(4):429 LIN YL, QIU MH, LI ZR. Non-alkaloid constituents of Buxus sinica (Buxaceae)[J]. Acta Bot Yunnan, 2006, 28(4):429
[2] 吕霞,胡明通,于盱.黄杨属植物化学成分的研究进展[J].安徽农业科学, 2012, 40(24):12028 LÜ X, HU MT, YU X. Research advances in the chemical constituents of Buxus[J]. J Anhui Agric Sci,2012,40(24):12028
[3] 张越浩,关焕玉,王永林,等.小叶黄杨非生物碱类成分[J].中国实验方剂学杂志, 2014, 20(20):114 ZHANG YH, GUAN HY, WANG YL, et al. Non-alkaloid constituents from Buxus sinica [J]. Chin J Exp Tradit Med Form, 2014,20(20):114
[4] 梁感荣,毛庭枝,梁成钦.小叶黄杨化学成分的研究[J].广西植物, 2009, 29(5):703 LIANG GR,MAO TZ,LIANG CQ. Studies of chemical constituents of Buxus mycrophylla[J]. Guihaia,2009, 29(5):703
[5] 宋媛媛,李媛,张洪泉.丁香苷抗炎镇痛作用及部分机制研究[J].中国野生植物资源, 2010,29(4):27. SONG YY, LI Y, ZHANG HQ. Anti inflammatory and analgesic effects of clove glycosides and its mechanism[J]. Chin Wild Plant Resour, 2010,29(4):27
[6] 汪琢,姜守刚,祖元刚.刺五加中紫丁香苷的提取分离及抗肿瘤作用研究[J].时珍国医国药, 2010, 21(3):752 WANG Z, JIANG SG, ZU YG. Study on the extraction and isolation and anti tumor effect of the syringin from Acanthopanax[J]. Lishizhen Med Mater Med Res, 2010, 21(3):752
[7] 姜月红,王燕颖.紫丁香苷对大鼠肝缺血再灌注损伤保护作用的初步研究[J].中国实验诊断学杂志, 2016, 20(3):359 JIANG YH, WANG YY. Preliminary research on the protective effects of syringin against hepatic ischemia repefusion injury in rats[J]. Chin J Lab Diagn,2016, 20(3):359
[8] 党晓伟,李清.紫丁香苷在大鼠体内的药动学研究[J].中成药, 2012,34(7):1246 DANG XW,LI Q. Pharmacokinetic study of syringin in rat plasma[J]. Chin Tradit Pat Med, 2012, 34(7):1246
[9] 杜传荣,林发全,逯东伟,等.香草酸酰胺衍生物的制备及其止血活性的动力学法研究[J].发光学报,2016, 34(4):478 DU CR, LIN FQ, LU DW, et al. Synthesis of vanillylamide derivatives and their hemostatic activity in vitro by kinetics methods[J]. Chin J Lumin, 2016,34(4):478
[10] MOTIRAM KR, TEJASWINI G, PATIL MA, et al. Vanillic acid ameliorates cationic bovine serum albumin induced immune complex glomerulonephritis in BALB/c mice[J]. Durg Dev Res, 2016, 77(4):171
[11] 仁爽,李铮,王京辉,等.中药女贞子饮片中8个成分的含量比较研究[J].药物分析杂志,2015, 35(8):1393 REN S, LI Z, WANG JH, et al. Comparative study of the content of eight components in Ligustri Lucidi Fructus pieces[J]. Chin J Pharm Anal,2015, 35(8):1393
[12] 吴发明,张思荻,曾俊,等. HPLC-ELSD法测定不同产地麦冬中4种代表性成分的含量[J].药物分析杂志, 2016, 36(8):1370 WU FM, ZHANG SD, ZENG J, et al. Determination of the 4 representative components in Maidong from different producing areas by HPLC-ELSD method[J]. Chin J Pharm Anal, 2016, 36(8):1370
[13] 杨娟艳,王芳,郭婷,等.黔产小叶黄杨中环维黄杨星D的鉴别及不同部位含量测定[J].生物技术进展,2016, 6(5):341 YANG JY, WANG F, GUO T, et al. The TLC identification and the cyclovirobuxine D content determination in different parts of Buxus sinica from Guizhou[J]. Curr Biotechnol, 2016, 6(5):341
[14] 郭婷,董莉,魏秀丽,等.小叶黄杨中环维黄杨星D含量测定的两种高效液相色谱法评价[J].中国临床药理学杂志, 2016, 32(19):1797 GUO T, DONG L, WEI XL, et al. Evaluation on the two kinds of HPLC methods for determination of cyclovirobuxium D in Buxus sinica [J]. Chin J Clin Pharmacol, 2016,32(19):1797
[15] 谢昀,于国治,徐新盛,等.反向离子对色谱法测定黄杨木中环维黄杨星D[J].中草药, 2005,36(11):1726 XIE Y, YU GZ, XU XS, et al. Determination in the content of cyclovirobuxine D from Buxus mycrophylla by reverse ion pair chromatography[J]. Chin Tradit Herb Drugs, 2005, 36(11):1726