北京大学学报(医学版) ›› 2020, Vol. 52 ›› Issue (3): 591-596. doi: 10.19723/j.issn.1671-167X.2020.03.030

• 技术方法 • 上一篇    下一篇

高效液相色谱法测定小鼠血浆中苯并三唑类紫外线吸收剂UV-327和UV-328

朱梅青,崔蓉()   

  1. 北京大学公共卫生学院劳动卫生与环境卫生学系,北京 100191
  • 收稿日期:2020-01-09 出版日期:2020-06-18 发布日期:2020-06-30
  • 通讯作者: 崔蓉 E-mail:cuirong19@sohu.com

Determination of UV-327 and UV-328 in mouse plasma by high performance liquid chromatography

Mei-qing ZHU,Rong CUI()   

  1. Department of Occupational and Environmental Health Science, Peking University School of Public Health, Beijing 100191, China
  • Received:2020-01-09 Online:2020-06-18 Published:2020-06-30
  • Contact: Rong CUI E-mail:cuirong19@sohu.com

摘要:

目的 建立小鼠血浆中苯并三唑类紫外线(ultraviolet, UV)吸收剂UV-327和UV-328的高效液相色谱测定方法。方法 在小鼠血浆中加入正己烷-丙酮溶液(体积比1 ∶1)涡旋混匀提取,上清液于50 ℃下氮气吹干,残渣用甲醇溶解后离心过滤,采用高效液相色谱-紫外检测器测定小鼠血浆中UV-327和UV-328的浓度。分析柱为Waters Symmetry?C18柱 (250 mm×4.6 mm,5 μm),流动相为100%甲醇,等度洗脱,流速1.0 mL/min,紫外检测波长为340 nm,以UV-320为内标,保留时间定性,内标法定量。 结果 UV-327、UV-328在0.05~10.0 mg/L范围内呈现良好的线性关系(r=0.999 7),检出限为0.01 mg/L,定量限为0.03 mg/L。小鼠血浆中,低、中、高3种浓度(0.50、1.00、2.00 mg/L)UV-327和UV-328的平均回收率分别为91.7%~101.0%和97.5%~103.9%,日内精密度(n=6)分别为2.9%~6.6%和2.7%~7.4%,日间精密度(n=3)分别为6.0%~9.3%和6.6%~8.6%,提取回收率分别为98.8%~103.8%和99.8%~100.9%。血浆样品室温下放置6 h和-40 ℃放置15 d,UV-327测定值的相对偏差分别为0.9%~3.5%和7.4%~15.0%,UV-328测定值的相对偏差分别为2.0%~4.3%和2.1%~13.8%。3种加标浓度的小鼠血浆样品室温下至少可以放置6 h,-40 ℃冰箱中可以保存15 d。结论 高效液相色谱法简便、快速,准确度、精密度和灵敏度较高,可应用于小鼠血浆中UV-327和UV-328的分析测定。

关键词: 高效液相色谱, UV-327, UV-328, 小鼠, 血浆

Abstract:

Objective: To establish a high performance liquid chromatography (HPLC) method for the determination of ultraviolet (UV) absorbers UV-327 and UV-328 in mouse plasma.Methods: N-hexane-acetone (volume ratio 1 ∶1) was added to a mouse plasma sample as the extraction solvent for vortex extraction, and the supernatant was dried at 50 ℃ with nitrogen. Thereafter the residue was redissolved with methanol, centrifuged and filtered. The separation was performed on a Waters Symmetry?C18 column (250 mm×4.6 mm, 5 μm), and the concentrations of UV-327 and UV-328 in the mouse plasma were determined by HPLC with an UV detector. The elution was isocratic at a flow rate of 1.0 mL/min with a mobile phase composed of 100% methanol, and the UV detection wavelength was 340 nm. The retention time was used for qualitative analysis, and the internal standard method was used for quantitative analysis using UV-320 as the internal standard. Results: The calibration curves of UV-327 and UV-328 were linear with correlation coefficients of 0.999 7 over the concentration range of 0.05 to 10.0 mg/L. The limit of detection was 0.01 mg/L, and the limit of quantitation was 0.03 mg/L. The average recoveries at low, medium and high three concentrations (0.50, 1.00, 2.00 mg/L) in the mouse plasma were 91.7%-101.0% for UV-327, and 97.5%-103.9% for UV-328. The intra-day precisions (n=6) of UV-327 were 2.9%-6.6%, and 2.7%-7.4% for UV-328. The inter-day precisions (n=3) of UV-327 were 6.0%-9.3%, and 6.6%-8.6% for UV-328. The extraction recoveries of UV-327 were 98.8%-103.8%, and 99.8%-100.9% for UV-328. The measured relative deviations of UV-327 in the mouse plasma samples placed at room temperature for 6 hours and -40 ℃ for 15 days were 0.9%-3.5% and 7.4%-15.0%, and the measured relative deviations of UV-328 were 2.0%-4.3% and 2.1%-13.8%, respectively. The mouse plasma samples could be stored at room temperature for 6 hours at least and -40 ℃ for 15 days at three spiked concentration levels.Conclusion: The method was simple and fast with high accuracy, precision and sensitivity, and could be applied to the determination of UV-327 and UV-328 in mouse plasma.

Key words: High performance liquid chromatography, UV-327, UV-328, Mouse, Plasma

中图分类号: 

  • R194.2

图1

小鼠血浆中UV-320、UV-327和UV-328测定的高效液相色谱图"

表1

小鼠血浆中UV-327和UV-328准确度与精密度实验结果"

Analyte Spiked
concentration/(mg/L)
Measured
concentration/(mg/L), x?±s
Average recovery Intra-day precision
(RSD, n=6)
Inter-day precision
(RSD, n=3)
UV-327 0.50 0.46±0.04 91.7% 6.6% 9.3%
1.00 1.01±0.08 101.0% 4.3% 7.7%
2.00 2.00±0.12 99.9% 2.9% 6.0%
UV-328 0.50 0.49±0.04 97.5% 7.4% 8.6%
1.00 1.04±0.07 103.9% 3.6% 6.6%
2.00 2.01±0.14 100.8% 2.7% 6.8%

表2

小鼠血浆中UV-327和UV-328提取回收率结果(n=6)"

Analyte Spiked
concentration/(mg/L)
Peak area ratio Extraction
recovery
A B
UV-327 0.50 3.00 2.89 103.8%
1.00 5.77 5.84 98.8%
2.00 11.38 11.25 101.2%
UV-328 0.50 2.26 2.24 100.9%
1.00 4.68 4.69 99.8%
2.00 9.28 9.29 99.9%

表3

小鼠血浆中UV-327和UV-328稳定性实验结果(n=6)"

Analyte Spiked
concentration/
(mg/L)
Instant Room temperature for 6 h -40 ℃ for 15 d
Measured
concentration/
(mg/L), x?±s
Measured
concentration/
(mg/L), x?±s
Relative deviation Measured
concentration/
(mg/L), x?±s
Relative deviation
UV-327 0.50 0.45±0.01 0.45±0.01 0.9% 0.51±0.04 13.3%
1.00 0.95±0.04 0.94±0.01 1.1% 0.88±0.02 7.4%
2.00 2.00±0.04 1.93±0.17 3.5% 1.70±0.04 15.0%
UV-328 0.50 0.47±0.01 0.49±0.02 4.3% 0.46±0.01 2.1%
1.00 1.02±0.02 1.04±0.04 2.0% 0.92±0.02 9.8%
2.00 2.10±0.05 2.03±0.13 3.3% 1.81±0.05 13.8%

图2

实际样品测定色谱图"

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