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Journal of Peking University (Health Sciences) ›› 2020, Vol. 52 ›› Issue (3): 591-596. doi: 10.19723/j.issn.1671-167X.2020.03.030

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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

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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

CLC Number: 

  • R194.2

Figure 1

Chromatographic diagram of UV-320, UV-327 and UV-328 in mice plasma A, blank plasma; B, blank plasma spiked with UV-320 (0.20 mg/L); C, blank plasma spiked with UV-327 (0.50 mg/L) and UV-320 (0.20 mg/L); D, blank plasma spiked with UV-328 (0.50 mg/L) and UV-320 (0.20 mg/L); E, blank plasma spiked with UV-327 (0.50 mg/L), UV-328 (0.50 mg/L) and UV-320 (0.20 mg/L)."

Table 1

Accuracy and precision of UV-327 and UV-328 in mice plasma"

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%

Table 2

Extraction recovery of UV-327 and UV-328 in mice plasma (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%

Table 3

Stability test results of UV-327 and UV-328 in mice plasma (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%

Figure 2

Chromatographic diagram of actual sample determination A, chromatogram diagram of UV-327 after 2 h gavage in mice plasma; B, chromatogram diagram of UV-328 after 2 h gavage in mice plasma."

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