RESUMEN
Objective@#To establish a high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method for the detection of serum oleic acid (OA), and preliminarily evaluate the role of OA in insulin resistance (IR) of type 2 diabetes (T2DM). @*Methods@#OA-［ 13 C 5 ］ was used as isotope-labeled internal standard, and the ion pairs of OA and OA-［ 13 C 5 ］ were 281.3/281.3 and 286.3/286.3, respectively. The ultrapure water was used as mobile phase A and methanol: acetonitrile (1∶1, v/v) as mobile phase B in a ZORBAX SB-Aq C18 reversed phase column. Meanwhile, the gradient elution system with a flow rate of 0.3 mL/min was used. According to the CLSI guidelines (EP15-A3), the reliability of the established method was evaluated by detecting the performance indicators such as precision, trueness, linear range, stability and carrying contamination rate. Serum OA levels were detected by the established HPLC-MS/MS method in 109 patients with clinically diagnosed T2DM and 100 healthy controls. The insulin resistance index (HOMA-IR) was calculated to evaluate IR, and the relationship between OA and IR was further analyzed. @*Results@#The established HPLC-MS/MS method for the detection of serum OA had good specificity and linearity in the range of 10-1 000 μmol/L (y=0.007 55x+0.004 83,r=0.997 7), and the low limit of quantification (LLOQ) was 10 μmol/L. It also had good precision, and the within-run coefficient of variation (CV) and total CV were not more than 1.62% and 1.73%, respectively, indicating that the method was suitable for the detection of serum OA. The serum OA levels in T2DM patients ［(425.58 ± 220.17) μmol/L］ were significantly higher than that in the healthy controls ［(113.20±58.00) μmol/L］, and serum OA levels were significantly correlated with HOMA-IR in T2DM patients and healthy controls. The area under the receiver operating characteristic (ROC) curve (AUC) of OA for the diagnosis of IR was 0.689. When the cut-off value identified by Youden index was 235.8 μmol/L, the sensitivity and specificity were 70.4% and 63%, respectively. When OA combined with fasting blood glucose (FBG) to diagnose IR, the AUC increased to 0.806, which was significantly higher than that of OA (P＜0.05). @*Conclusion@#A scientific and efficient HPLC-MS/MS method for the quantitative detection of serum OA is established successfully, which provides a reliable method for the dynamic monitoring of the changes of OA levels in the patients with metabolic diseases.
RESUMEN
Objective To establish an ultra high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method for detecting α-hydroxybutyrate (α-HB) in serum.Methods Electrospray ionization negative ion and multiple reaction monitoring mode were used to detect serum α-HB.The linearity,low limits of quantification,precision,recovery and interference of UHPLC-MS/MS were evaluated.The reference interval of this method was established in 130 serum samples (62 males and 68 females) from Shanghai East Hospital.Dixon method was used to judge the outliers and K-S test was used to analyze the data normality.The standard curve was scored by linear regression analysis.Results The total run time was 4 min of UHPLC-MS/MS method for the determination of α-HB.It has a good linear relationship in the range of 0.5-40.0 mg/L(r=0.999 4);the low limit of quantification was 0.5 mg/L;the in-batch and inter-batch coefficient of variation precision were less than 4.1％ and 6.3％,respectively;the recovery ranged between 95.8％ and 103.8％.Hemolytic samples (about 5 g/L hemoglobin),lipemic samples (about 12 mmol/L triglyceride),icteric samples (about 150 μmol/L total bilirubin) had no significant interference to the detection.The reference range of the apparent healthy population was 1.46-6.48 mg/L.Conclusions A method for the determination of serum α-HB by UHPLC-MS/MS was established.The method was simple,rapid,and could be used for the detection of clinical samples.