Abstract
Objective To establish the HPLC method for detecting neonatal serum FB. Methods The chromatographic column was ODS-C18 column (150.0 mm × 4.6 mm × 5.0 μm). The mobile phase of methanol-ammonium acetate ice acetic acid solution was 96:4, flow rate 0.9 ml/min, column temperature 30℃, detection wavelength 458 nm, and injection volume 20 μl. The precision, accuracy, recovery rate, and stability of the method were examined. The differences between different groups were calculated by SPSS software package. Results The retention time of FB was 9.583 min. The lower limit of quantification was 0.625 μmol/L. The linear range of serum FB was 0.625-160.000 μmol/L. The standard curve: y=73 403x – 22 975. The r2 is 0.999 5. The slope deviation was 726.649 686 4. The intercept deviation was 30 119.924 55. The average intra-day and inter-day precision of each gradient concentration were less than 5%. The relative recovery was 95%-105%. Different gradient concentration extraction recovery range was 68%-78%. The repeated freeze-thaw stability average RSD was 3.020%. The average RSD value of stability under different storage conditions was 0.565%. The concentration of serum FB in 13 jaundiced newborns we measured were (548.20±107.19) nmol/L. Conclusion HPLC detecting serum FB is simple in operation, and has high specificity, accuracy, precision and stability. Key words: Serum free bilirubin (FB); Neonates; HPLC; Detection
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