复方苦参肠炎康片HPLC指纹图谱及多成分含量测定

Fingerprint of Fufang Kushen Changyankang Tablets and the Content Determination of Its Multi-components by HPLC

关键词:  复方苦参肠炎康片  指纹图谱  高效液相色谱法  多组分测定  质量控制

Key words:  Fufang Kushen Changyankang tablets  fingerprint  HPLC  multi-component determination  quality control

摘要:

目的 采用HPLC建立复方苦参肠炎康片指纹图谱定性和8个成分定量分析的测定方法，为复方苦参肠炎康片的质量控制提供更为可靠的方法和依据。方法 采用HPLC同时测定复方苦参肠炎康片中京尼平苷酸、苦参碱、绿原酸、氧化苦参碱、芍药苷、黄芩苷、盐酸小檗碱、甘草酸铵的含量，采用Agilent TC-C₁₈柱（250 mm×4.6 mm，5 μm），流动相为乙腈（A）-0.05%磷酸溶液（B），梯度洗脱；检测波长为220 nm，柱温为30℃，流速为1.0 mL·min^-1，进样量为10 μL。选取黄芩苷为参照峰，并使用中药色谱指纹图谱相似度评价系统对指纹图谱进行相似性评价，确定共有峰。结果 京尼平苷酸、绿原酸、苦参碱、氧化苦参碱、芍药苷、黄芩苷、盐酸小檗碱、甘草酸铵质量浓度的线性范围分别为1.080~9.723（r=0.999 8），5.213~46.92（r=0.999 6），18.95~170.5（r=0.999 4），31.79~286.1（r=0.999 7），49.53~445.8（r=0.999 4），113.5~1022（r=0.999 5），70.77~636.9（r=0.999 6），10.69~96.21（r=0.999 5）μg·mL^-1；定量限分别为0.867 0，1.051，1.123，1.021，0.903 1，0.896 1，1.063，1.216 μg·mL^-1；平均加样回收率分别为98.6%，99.0%，98.4%，98.7%，98.8%，99.1%，98.9%，98.8%（RSD均<2.0%，n=6）；仪器精密度、重复性、稳定性（48 h）试验的RSD均<2.0%（n=6）。14批次复方苦参肠炎康片的指纹图谱与其指纹图谱共有模式相比较相似度均>0.9，标定了共有峰22个，并对其中8个共有峰进行了指认和归属。结论 所建立的HPLC指纹图谱分离效果良好，特征性强，方法稳定简单，多指标成分含量测定同时结合指纹图谱分析能更为全面地对复方苦参肠炎康片的质量进行控制。

Abstract:

OBJECTIVE To establish a method for the qualitative analysis of fingerprints and quantitative analysis of 8 components of Fufang Kushen Changyankang tablets by HPLC, and to provide a more reliable method and basis for the quality control of Fufang Kushen Changyankang tablets. METHODS The determination was performed on AgilentTC-C₁₈(250 mm×4.6 mm, 5 μm) column with mobile phase consisted of acetonitrile(A)-0.05% phosphoric acid(gradient elution) at the flow rate of 1.0 mL·min^-1. The detection wavelength was set at 220 nm. The column temperature was set at 30℃ and the injection volume was 10 μL. Baicalin was selected as the reference peak. The HPLC fingerprints were analyzed and the similarity was evaluated with Similarity Evaluation System for Chromatographic Fingerprint of Traditional Chinese Medicine. RESULTS The linear range was 1.080-9.723 μg·mL^-1 (r=0.999 8)for geniposidic acid, 5.213-46.92 μg·mL^-1 (r=0.999 6) for chlorogenic acid, 18.95-170.5 μg·mL^-1(r=0.999 4) for matrine, 31.79-286.1 μg·mL^-1(r=0.999 7) for oxymatrine, 49.53-445.8 μg·mL^-1(r=0.999 4) for paeoniflorin, 113.5-102 2 μg·mL^-1(r=0.999 5) for baicalin, 70.77-636.9 μg·mL^-1(r=0.999 6)for berberine hydrochloride, 10.69-96.21 μg·mL^-1(r=0.999 5) for ammonium glycyrrhizinate, limits of quantitation were 0.867 0, 1.051, 1.123, 1.021, 0.903 1, 0.896 1, 1.063, 1.216 μg·mL^-1; average recoveries were 98.6%, 99.0%, 98.4%, 98.7%, 98.8%, 99.1%, 98.9%, 98.8% (RSD<2.0%, n=6), respectively; RSDs of precision, reproducibility and stability tests (48 h) were <2.0%(n=6). Compared with the common pattern of fingerprints, the similarities of 14 batches of samples were >0.9 with 22 common peaks, 8 of which were identified and assigned. CONCLUSION The HPLC fingerprints established have a good separation effect with strong characteristics. The method is stable and simple. The better quality control of Fufang Kushen Changyankang tablets can be achieved by multi-index composition determination combined with fingerprint analysis.