CAJ | 학술논문

目的：建立LC-MS/MS法测定人血浆中羟基红花黄色素A( QA)的浓度。方法血浆样品中加入等体积0．2 mol ·L-1的乙酸铵后,采用固相萃取技术进行提取QA,洗脱液直接进行 LC-MS/MS分析。 QA和内标异鼠李素-3-O-新橙皮苷(SLS)通过Agilent ZORBAX SB C18(3．0 mm ×100 mm,3．5μm)色谱柱进行等度洗脱分离,流动相组成为0．2 mmol ·L-1乙酸铵水溶液/甲醇=30/70。质谱检测采用负离子模式,扫描方式为多反应检测,QA的目标离子对为m/z 611.131/490．900,SLS 的目标离子对为 m/z 623．032/298．800。结果 QA和SLS的保留时间分别2．7 min和3．9 min左右,空白血浆无干扰影响含量测定；血浆中 QA 的线性范围为8．57~4185μg·L-1( r为0．9949~0．9992),定量下限为8．570μg·L-1,日内日间精密度 RSD 均小于7%；低、中、高3个浓度下的平均介质效应分别为116．62%、119．06%和115．72%( RSD 分别为3．27%、3．42%和4．93%),平均提取回收率分别为77．75%、80．90%和80．76%( RSD分别为1．70%、1．78%和4．15%)；血浆样本于室温放置4 h、-70℃反复冻融3次及-70℃冰冻保存31 d的情况下均稳定；浓度超出定量上限的QA血浆样本经空白血浆稀释6．25倍后可准确测定。结论建立的测定血浆样本中QA浓度的LC-MS/MS分析方法简便、快速、准确灵敏,适用于QA人体药代动力学的系统研究。
목적：건립LC-MS/MS법측정인혈장중간기홍화황색소A( QA)적농도。방법혈장양품중가입등체적0．2 mol ·L-1적을산안후,채용고상췌취기술진행제취QA,세탈액직접진행 LC-MS/MS분석。 QA화내표이서리소-3-O-신등피감(SLS)통과Agilent ZORBAX SB C18(3．0 mm ×100 mm,3．5μm)색보주진행등도세탈분리,류동상조성위0．2 mmol ·L-1을산안수용액/갑순=30/70。질보검측채용부리자모식,소묘방식위다반응검측,QA적목표리자대위m/z 611.131/490．900,SLS 적목표리자대위 m/z 623．032/298．800。결과 QA화SLS적보류시간분별2．7 min화3．9 min좌우,공백혈장무간우영향함량측정；혈장중 QA 적선성범위위8．57~4185μg·L-1( r위0．9949~0．9992),정량하한위8．570μg·L-1,일내일간정밀도 RSD 균소우7%；저、중、고3개농도하적평균개질효응분별위116．62%、119．06%화115．72%( RSD 분별위3．27%、3．42%화4．93%),평균제취회수솔분별위77．75%、80．90%화80．76%( RSD분별위1．70%、1．78%화4．15%)；혈장양본우실온방치4 h、-70℃반복동융3차급-70℃빙동보존31 d적정황하균은정；농도초출정량상한적QA혈장양본경공백혈장희석6．25배후가준학측정。결론건립적측정혈장양본중QA농도적LC-MS/MS분석방법간편、쾌속、준학령민,괄용우QA인체약대동역학적계통연구。
Aim To establish a LC-MS/MS method for the determination of hydroxysafflor yellow A ( QA ) in human plasma. Methods After being added into 0. 2M ammonium acetate (1∶1,V/V), QA was extrac-ted using solid-phase extraction technique, and the eluent was directly injected into LC-MS/MS systems. Agilent ZORBAX SB C18 (3. 0 × 100 mm, 3. 5 μm) column and isocratic elution system composing of meth-anol and 0. 2 mM ammonium acetate (70 ∶ 30, V/V) provided chromatographic separation of QA and internal standard isorhamnetin-3-O-neohespeidoside ( SLS) fol-lowed by detection with mass spectrometry. The mass transition ion-pair was followed as m/z 611 . 131→490. 900 for QA and m/z 623. 032→298. 800 for SLS. Results The retention time of QA and SLS was 2. 7 min and 3. 9 min respectively, with no interference in human blank plasma. The proposed method showed good linearity over the concentration range of 8. 57 ~4185 μg·L-1 for QA with a correlation coefficient≥ 0 . 9949 . The lower limit of quantitation was 8. 570 μg ·L-1 . The intra-batch and inter-batch precision and accuracy were within 7%. The average matrix effect ranged from 115. 72% to 119. 06% with RSD less than 5%. The average extraction recovery ranged from 77. 75% to 80. 76% with RSD less than 5%. Stability of human samples after 4 h at room temperature, after the three freeze-thaw cycles and after 31 days at -70℃, and post-preparative stability of the processed sam-ples after 24 h was acceptable. Plasma samples with the concentration beyond the upper quantitation limit could be accurately determined after being diluted using 6. 25 times ( V/V ) of human blank plasma. Conclusion Our current LC-MS/MS method is sensitive, accurate and convenient, and is proved to be suitable for the sys-tematic study on clinical pharmacokinetics of QA.