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Objective:To establish a candidate reference method for serum progesterone using isotope dilution liquid chromatography tandem mass spectrometry (ID-LC/MS/MS) in our laboratory, validate the analytic performance of five clinical routine detection systems to explore the comparability of serum progesterone detection by different detection systems.Methods:A candidate reference method for serum progesterone using ID-LC/MS/MS method was established. The sample was pretreated by liquid-liquid extraction method, and the reversed phase liquid phase separation in positive ion mass spectrometry mode was used to detect progesterone in human serum, and the detection time of a single sample was controlled within 5 minutes by gradient elution. In order to improve the accuracy of the method, the bracketing calibration method (BCM) was used to establish the standard curve. The sensitivity, accuracy, precision and specificity of BCM and classical calibration curve method were evaluated according to CLSI C62-A, EP15-A2, EP6-A2 and EP9-A3, and the analytical performance and comparability of five clinical routine progesterone detection systems were evaluated,compared with ID-LC/MS/MS method, the bias at medical decision level 2 and 25 ng/ml was evaluated to see if they were <1/2TEa (12.5%).Results:The limit of detection (LOD) of ID-LC/MS/MS was 0.005 ng/ml. The recoveries of BCM method and classical calibration curve method are 97.95%-101.58% and 96.88%-110.70%, respectively. The measurement results of BCM method for certified reference materials are within its declared uncertainty range. The intra-and inter-assay coefficient of variation ( CV) of BCM method was less than 3.0%, which was better than that of classical calibration curve method ( CV: 2.48%-9.33%). The precision and linear range of the five clinical routine detection systems can meet the detection requirements. The measurement bias of detection system 1, 3 and 5 at 25 ng/ml of medical decision level was less than 1/2TEa, and the measurement bias at 2 ng/ml of medical decision level was more than 1/2TEa. The measurement bias of detection system 2 and 4 at two medical decision levels was less than 1/2TEa. Conclusion:The candidate reference method for serum progesterone ID-LC/MS/MS established in our laboratory meets the requirements of the reference method. BCM has better detection performance than classical calibration curve method. The precision and linearity of the five progesterone clinical detection systems are satisfactory. The five clinical detection systems could meet the clinical requirements at the medical determination level of 25 ng/ml, however, only two of the five clinical detection systems meet the clinical requirements at the medical determination level of 2 ng/ml.
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Objectives:To establish a candidate reference method of isotope dilution liquid chromatography tandem mass spectrometry (ID-LC/MS/MS) for the determination of human plasma normetanephrine, and to evaluate the performance of the method. The method was used to quantify the samples of the external quality assessment program, and to initially evaluate the detection status of plasma normetanephrine.Methods:The isotope standard solution of normetanephrine was selected as the internal standard, the gravimetric method was used for sampling, and the standard curve method was used for quantification. Protein precipitation combined with weak cation solid phase extraction was used for pretreatment, and ultra-high liquid chromatography-coupled triple quadrupole mass spectrometry was used for LC/MS analysis. According to the relevant EP documents, the specificity, matrix effect, detection limit, quantification limit, precision, accuracy, and uncertainty of the method were estimated. This method is used to quantify the samples of the 2020 National Center for Clinical Laboratories external quality assessment program of normetanephrine. Taking the average value of this method as the target value, the optimal allowable total error standard of biological variation as the evaluation limit, the quality of the laboratory testing was evaluated.Results:The method had good specificity, and the interferences and matrix effects did not affect the detection results. The detection limit and quantification limit of plasma normetanephrine were 1.08 pg/g and 3.54 pg/g, respectively. The intra-batch coefficient of variation ( CV) and total CV were 0.43%-1.10% and 0.61%-1.42%, respectively. The relative recovery rates were 98.5%～101.9%. The relative expansion uncertainty of the four plasma samples were 3.10%, 2.34%, 2.16%, and 1.73%, respectively. The results of the external quality assessment program showed that the pass rates of the 202013 and 202014 samples were 80% and 85%, respectively. Conclusions:The study established a candidate reference method of ID-LC/MS/MS for the measurement of plasma normetanephrine. The method is accurate, precise and simple, and is expected to be used as a reference method for the determination of plasma normetanephrine, and can be applied to quantify the samples of the external quality assessment program.
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Objective To develop a method for the simultaneous determination of 15mycotoxins in peanuts by ultra high performance liquid chromatography-tandem mass spectrometry with QuEChERS EMR-Lipid approach and stable isotope dilution. Methods The samples were extracted by 2% formic acid acetonitrile-water (50 : 50, V/V) and then purified with QuEChERS EMR-Lipid approach.The mycotoxins were fully separated on a pentafluorophenyl column under a gradient elution with methonal-0.01%formic acid aqueous solution.The mycotoxins were analyzed by UPLC-MS/MS with multiple reaction monitoring (MRM) mode and quantified by isotope internal standard method. Results Fifteen mycotoxins had good linear relationship in the certain correlation ranges with the correlation coefficients all above 0.995 and the detection limits were 0.1－10 μg/kg.The mean recoveries ranged from 81.2% to 115.3% with RSD (n=6) varying from 2.1% to 10.7%. Conclusion The method is simple, highly sensitive, practical, and proves to be suitable for quantitative analysis of 15 mycotoxins in peanuts.
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A fast, simple and cost-effective UPLC-MS / MS method was established for determination of 16 kinds of mycotoxins in vegetable oils with stable isotope dilution technique. Samples were extracted by acetonitrile-water-acetic acid 84: 15: 1(V/ V) and then diluted using water without any further clean-up steps. The mycotoxins were fully separated on a pentafluorophenyl column. Matrix effects were efficiently compensated by the [ 13 C]-labelled internal standards. The mean recoveries at three different concentration levels ranged from 74. 2％ to 105. 6％ , with RSD varied from 0. 3％ to 13. 9％ . Finally, the method was applied to analyze several kinds of vegetable oil samples. The method was simple, rapid, high sensitive and suitable for the determination of mycotoxins in vegetable oils.
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Objective To develop an isotope dilution liquid chromatography tandem mass spectrometry (ID-LC-MS/MS) method for determining the concentration of 5-methyltertrahydrofolate (5MT) in human serum.Methods After qualitative analysis of 5MT reference,its purity was identified by two methods with different principle,i.e.,mass balance and quantitative nuclear magnetic resonance.The definite value and uncertainty of 5MT concentrations in 6 serum samples containing same level of 5MT were assessed by ultra performance liquid chromatography-triple quadrupole tandem mass spectrometer equipped with electrospray ionization in monitoring mode of multiple reactions,in which 13C5-5MT was used as the internal standard.The precision,accuracy,limit of detection (LOD) and limit of quantification (LOQ) of the method were also evaluated.Results After qualitative analysis for confirming the principal component in the sample,the purity of 5MT reference was measured as 76.65％ by mass balance and quantitative nuclear magnetic resonance method.The definite value of 5MT in the serum sample was 8.22 ng/mL,the extended uncertainty was 0.54 ng/mL,LOD was 0.05 ng/mL,LOQ was 0.50 ng/mL and the repeatability and inter-batch precision CV were within 5.0％.The accuracy of measured results was verified by using NIST SRM 1955 since all the values of 3 levels were within the range of certified value with the extended uncertainty.Conclusion A accurately quantitative determination for the concentration of 5 MT in human serum with high specificity and sensitivity was established by using ID-LC-MS/MS method.
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Stable isotope dilution mass spectrometry(ID-MS)is a quantitative method for determining the content of unlabeled compounds with stable isotope labeled compounds as internal standard. Stable isotope dilution liquid chromatography-mass spectrometry(ID-LC-MS)has the advantages of high sensitivity, no quantitative separation of the analytes, and effective elimination of the biological matrices. The method has been widely used in quantitative analysis of biomarkers, drugs and their metabolites. In this paper, the mechanism and characteristics of ID-LC-MS are briefly introduced, and its medical applications are summarized.
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Stable isotope dilution mass spectrometry(ID-MS)is a quantitative method for determining the content of unla?beled compounds with stable isotope labeled compounds as internal standard. Stable isotope dilution liquid chromatography-mass spec?trometry(ID-LC-MS)has the advantages of high sensitivity,no quantitative separation of the analytes,and effective elimination of the biological matrices. The method has been widely used in quantitative analysis of biomarkers,drugs and their metabolites. In this pa?per,the mechanism and characteristics of ID-LC-MS are briefly introduced,and its medical applications are summarized.
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A high throughput screening method based on QuEChERS purification and stable isotope dilution-liquid chromatography coupled to high resolution time-of-flight mass spectrometry was developed for the simultaneous rapid determination of 86 kinds of glucocorticoids (GCs) in cosmetics.The analytes were extracted by acetonitrile, and then the extracts were purified using an improved QuEChERS method.The chromatographic separation was performed on a novel multiple chromatographic retention mechanisms column of Poroshell 120 PFP (100 mm × 2.1 mm, 2.7 μm) with gradient elution using 0.2% (V/V) acetic acid and acetonitrile as mobile phase.The accurate mass database of parent ions and mass spectra library of fragment ions of 86 GCs were established under positive ionization mode with electrospray ionization source.Based on the method described above, the qualitative identifications of the 86 GCs were accomplished without the contrast of standard substances.The results demonstrated that the linear range of this method was from 2 μg/L to 200 μg/L with good correlation coefficients of R2>0.99.The average recoveries of the 86 GCs ranged from 66.2% to 112.8%, and the relative standard deviation (RSDs) was 4.6%-13.9% at three different spiked levels.The limit of detection (LOD) and quantification (LOQ) were 0.006-0.015 mg/kg and 0.02-0.05 mg/kg, respectively.The method is simple, efficient, reliable and accurate, and is suitable for high throughput screening of 86 GCs added illegally in cosmetics.
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Objective To summarize and review the comparative results of the isotope dilution mass spectrometry in Cholesterol Reference Method Laboratory Network (CRMLN)project of the Centers for Disease Control and Prevention (CDC)of the United States in order to provide quality controls for determination of blood lipid.Methods The isotope dilution liquid chromatography tandem mass spectrometry (ID-LC/MS/MS)methods for determination of serum total cholesterol (TC)and triglycerides (TG)were developed to participate in the comparison of CRMLN.The survey was conducted every three months before 2016 and every half year from 2016.Four kinds of reference materials with two parallel tubes for each material and each tube in duplicate were determined in every survey.At least two certified reference materials used as quality control samples were analyzed simultaneously in each determination.Results In the 15 comparisons the CV of the TC determination method in our laboratory was 0.43％ while the CV of all the participated laboratories in CRMLN was 0.42％.The bias from the overall mean value in our laboratory was 0.22％ while the bias from the CDC target values was 0.58％.The CV of the TG determination method in 15 tests of our laboratory was 0.62％ and the bias was-0.98％ from the overall mean value and-0.80％ from the target values of CDC.Among the 60 results for comparison,98％ (59/60)of CV in the TC determination met with CDC requirement for precision (CV≤ 1％),and 70％ of bias (42/60) of the results met with CDC requirement for accuracy (bias ≤ 1％).For the 60 results in comparison of TG determination,92％ (55/60)of bias of the results met with the accuracy requirement of CDC (bias ≤2.55％).Conclusion In CRMLN comparison the results of TC and TG determined by ID-LC/MS/MS method were consistent with the values which were certified by CDC and determined by other network laboratories.The comparative surveys may play an important role in the standardization of lipid determination,and should be expected to provide experiences and technical supports in the comparative plan for reference measurement in our country.
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A method was established for the simultaneous determination of the total fatty acid esters of chloropropanols in edible oils by gas chromatography-mass spectrometry combined with isotope dilution technology. The samples were hydrolyzed with sodium methylate-methanol, and then purified by diatomite cartridge. After being derivatized with heptafluorobutyrylimidazole ( HFBI ), the target analytes were determined by GC-MS with the deuteriumchloropropanols esters as the internal standards. An excellent linear correlation in the range of 0. 050-2. 000 mg / L was acquired for 3-monochloropropane-1,2-diol (3-MCPD) esters, 2-MCPD esters, dichloropropan-2-ol (1,3-DCP) esters and 2,3-dichloropropan-1-ol (2,3-DCP) esters, with all the correlation coefficients (r) higher than 0. 9995. The limits of detection (LODs) for 3-MCPD esters, 2-MCPD esters, 1,3-DCP esters and 2,3-DCP esters were 0. 015, 0. 015, 0. 030, and 0. 030 mg / kg, respectively, and the limits of quantitation (LOQ) were 0. 050, 0. 050, 0. 100, and 0. 100 mg / kg, respectively. The average spike recoveries of the four kinds of chloropropanols esters in blank extra virgin olive oil matrix were typically in a range of 87. 0% -110. 5% with the relative standard deviations (RSDs) less than 10. 1% . The detection rates of 3-MCPD esters, 2-MCPD esters, 1,3-DCP esters and 2,3-DCP esters in 74 edible oil samples were 94. 6% , 63. 5% , 5. 4% , and 0% , respectively. The contamination levels of 3-MCPD esters, 2-MCPD esters and 1,3-DCP esters were in the range of not detected (ND) to 10. 646 mg / kg, ND to 3. 617 mg / kg and ND to 0. 089 mg / kg, respectively. This method is accurate and rugged for the simultaneous determination of total fatty acid esters of chloropropanols in edible vegetable oils.
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An enantioselective method was developed for the separation and determination of three chiral hexabromocyclododecanes ( HBCDs ) including α-HBCD, β-HBCD, γ-HBCD in soil and earthworm by HPLC-ID-MS/MS. d18-HBCDs used as internal standards were added to the samples before extraction. HBCDs enantiomers were extracted from soil by accelerated solvent extraction ( ASE ) with n-hexane/DCM (1:1,V/V) at 100℃ and 10 MPa for 5 min, and further cleaned up using silica column. HBCDs enantiomers were extracted from earthworm by vortex turbulence with ethyl acetate. The extracts were orderly sulphonated by sulfuric acid, and purified by silica column. For all HBCDs enantiomers, good linearities were obtained in the concentration range of 0. 25-50 ng/mL. Limits of detection ( LOD) and limits of quantification ( LOQ) were 0. 00544-0. 00766 ng/g and 0. 0173-0. 0244 ng/g, respectively in soil. The recoveries of spiked samples at 0. 05 and 2. 5 ng/g levels were 80. 0%-95. 9% with relative standard deviations ( RSD, %) of 5. 7%-11. 9% in soil. Limits of detection (LOD) and limits of quantification (LOQ) were 0. 0103-0. 0148 ng/g and 0. 0328-0. 0471 ng/g, respectively in earthworm. The recoveries of spiked samples at 0. 1 and 5 ng/g levels were 78. 0% -94. 4% with relative standard deviations ( RSD, %) of 6. 1% -12. 2% in earthworm. This method can meet the requirements of determination of trace HBCDs in soil and earthworm.
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A method was developed for the quantification of human growth hormone ( hGH ) by protein purification and isotope dilution-high performance liquid chromatography mass spectrometry ( HPLC-IDMS ) . The hGH was purified and fractionated by fast protein liquid chromatography ( FPLC ) , then hGH molecular weight was accurately determined by Fourier transform ion cyclotron resonance mass spectrometer ( FTICR-MS). The purified hGH was hydrolyzed and the separation was performed on an KINETEX C18 column (150 mmí2 mm I. D. , 2. 6 μm) with water ( containing 0. 1% TFA) and acetonitrile isocratic solution as the mobile phase at a flow rate of 0 . 2 mL/min and 40℃. The electrospray source was operated in the positive ion mode, and monitored in the multiple reaction monitoring ( MRM) mode. The measured hGH molecular weight by FTICR-MS was only 0. 31 Da difference from theoretical value. Three amino ( proline, valine and phenylalanine) were clearly separated by isocratic elution within 5 min. Under the optimized conditions, the content of hGH was 186 . 80 μg/g with a RSD of 0 . 5%. The detection results of hGH in international comparison by this method were consistent with the reference value, which validated the feasibility of the established method. The developed method is simple, practical, accurate, reliable and reproducible, and can be used for the hGH quantitation of pure hGH CRM to provide reference for the routine detection of hGH.
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A method for the determination of methylmercury ( MeHg ) and ethylmercury ( EtHg ) in aquatic products was developed using gas chromatography-mass spectrometry with stable isotope-labelled internal standard. After ultrasonication assisted hydrochloric acid extraction, MeHg and EtHg in samples were extracted into toluene under the presence of sodium chloride and then back-extracted into cysteine aqueous solution. The MeHg and EtHg were released from their complexes with cysteine by adding cupric ions, and then derived with sodium tetraphenylborate. Under the optimal chromatographic conditions, MeHgPh and EtHgPh, the resulting derivatives, were separated completely on a DB-5MS capillary column and detected by electron impact ionization mass spectrometry in the selective ion monitoring ( SIM) mode, and quantified by a stable isotope dilution method using the d3-methylmercury as internal standard. The calibration curves were linear in the range of 1-500 μg/L of MeHg and EtHg. Concentration of 0. 828 mg Hg/kg with relative standard deviation ( RSD ) of 3 . 2% ( n=6 ) was obtained for MeHg in GBW 10029 . This was in good agreement with the certified values of (0. 84±0. 03) mg Hg/kg. The average recoveries were 94%-101% and 81%-104% for MeHg and EtHg spiked in aquatic samples, with RSDs of 1. 9%-4. 7% and 3. 1%-8. 2%(n=6), respectively. The limits of detection (S/N=3) of the two targets were 0. 1-0. 3μg/kg. This method was sensitive, accurate and could meet the demand of the determination of methylmercury and ethylmercury in aquatic products.
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Objective To establish a method for measuring serum cotinine by isotope dilution liquid chromatography tandem mass spectrometry (ID-LC/MS/MS) and provide an assay that can be applied to theevaluation of the level of smoke exposure and to the risk analysis of smoking related diseases.Methods Blood samples were collected from 94 apparently healthy subjects from October to December in 2010 and centrifuged,and the sera were separated.Serum samples were mixed with [ D3 ] -cotinine ( as the internal standard) and treated with acetonitriles to precipitate protein.After centrifugation,the supernatants were transferred and evaporated under a stream of nitrogen until dryness and reconstituted with mobile phase.Then the residuals were analyzed by LC/MS/MS system with multiple reaction monitor model; the concentration of cotinine were quantified by the isotope internal standard method and the stand curve was employed with a series of calibration.To estimate the precision of the method,five frozen serum pools were repeatedly analyzed in five runs,and every pool was analyzed in triplicate.In addition,the recovery rates were analyzed with the serum sample added with different levels of standard.The stability of cotinine in serum preserved at room temperature,4 ℃ and - 80 ℃,respectively.Finally,the levels of cotinine of 94 healthy subjects were measured to evaluate the distribution of cotinine with different smoke statuses.Results Serum cotinine measured by ID-LC/MS/MS was separated well with few interferences.The correlation coefficients between the peak area ratios and cotinine concentrations were higher than 0.9993.The values of within-run coefficients of variation (CV) of five frozen serum pools (0.68,48.42,94.34,250.95 and 287.04 μg/L) were 2.19％,0.78％,0.75％,0.65％ and 0.67％,respectively.The values of total CV were 4.71％,1.40％,1.98％,1.10％ and 1.03％,respectively.The limit of detection (LOD) and limit of quantitation ( LOQ ) were 0.013 and 0.050 μg/L,respectively.The analytical recoveries ranged from 99.22％ to 102.67％.The samples could maintain stability within 2 d at room temperature,7 d at 4 ℃ and 3 months at -80 ℃ resulting the accuracy of measurements from 99.28％ to 100.87％ and the CV＜5％.The levels of cotinine of 94 healthy subjects were measured and shown skewed and leptokurtic distribution.The concentrations of twenty smokers,fourteen former smokers and sixty non-smokers were 116.40 (63.17 -241.12),0.67 (0.15 - 0.95 ) and 0.22 (0.15 - 0.42 ) μg/L,respectively.Furthermore,the level of cotinine of former smokers (Z =-2.12,P ＜0.05) and smokers (Z =-6.67,P ＜0.001) were statistically higher than non-smokers.Conclusions An ID-LC/MS/MS method for serum cotinine detection has been established.It is hoped that the method will be applied to the assessment of smoke exposure and its association with the risks of smoking related diseases since it is simple,specific,precise,sensitive and accurate.
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Objective To develop a candidate reference method for the measurement of serum glucose based on isotope dilution liquid chromatography tandem mass spectrometry(ID-LC/MS/MS)Methods An internal standard [~(13)C_6]glucose was added to serum samples and equilibrated with endogenous glucose.Serum proteins were removed by a precipitation with anhydrous ethanol.Serum glucose and the internal standard were then reacted with 1-phenyl-3-methyl-5-pyrazolone and the formed derivatives were analyzed by liquid chromatography tandem mass spectrometry with multiple reaction monitoring(MRM).The method was calibrated with bracketing calibrators and serum glucose concentrations were calculated by comparing the peak area ratios of samples with that of the calibrators.Results The within-run,between-run and total coefficients of variation averaged 0.36%,0.47%and 0.61%,respectively.The analytical recoveries ranged from 99.0% to 100.9%.Results of analyzing the certified reference material SRM 965a showed an average biases of-0.20%.Conclusions An ID-LC/MS/MS method for measuring serum glucose has been developed.The method is highly precise and accurate and may be used as a candidate reference method.
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A method was developed for determining residual dexamethasone and betamethasone in swine liver by HPLC-MS/MS with isotope dilution. The samples were digested by β-glucuronidase/aryl sulfatase, and extracted) with acetonitrile. Further cleanup was performed on C_(18)) cartridge and liquid-liquid extraction with sodium) carbonate solution. Then the supernatant was dried under nitrogen and residues were dissolved in mobile) phase. The samples were analyzed by HPLC-MS/MS on a Hypercarb C_(18)) column with a mixture of acetonitrile-water-formic acid as the mobile phase. The samples were quantified with the internal standard calibration curve method using isotope dilution. The limit of detection for dexamethasone and betamethasone in swine liver was 0.12 μg/kg and 0.14 μg/kg, respectively. The limits of quantification were 0.42 μg/kg and0.47 μg/kg), respectively. The average recoveries were 97.3%-111.0%, the intra-assay relative standard deviations were 1.85%-5.65% and the inter-assay relative standard deviations were 2.8%-8.0% at spiked levels of 0.75-2.00 μg/kg. There was a good linear correlation (the correlation coefficient is above 0.9997) between the ratio of peak areas of quantitative ion-pair to internal standard and concentration of analyte) in the range of 10-500 μg/L.
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A comprehensive analytical method based on solid phase extraction-isotope dilution-gas chromatography tandem mass spectrometry has been developed for the determination of musk xylene in cosmetics. Various cosmetic samples, including cream, lotion, powder, shampoo and lipstick, were extracted under ultrasonication. The extract was centrifuged, and the upper solution was concentrated by rotary evaporator. The reconstructed solution was then cleaned up by Sep-Pak Silica solid phase extraction cartridge. Qualitative and quantitative analysis was carried out under the MRM mode after the chromatographic separation on DB-5 MS(30 m×0.25 mm, 0.25 μm) capillary column. The limit of quantitation(LOQ) for musk xylene was 5 μg/kg. The mean recoveries at the three spiked levels of 5-50 μg/kg were 81.1%-86.9%, with the intra-day precision less than 10% and the inter-day precision less than 12%. The method is accurate, rapid, sensitive and adapt to the inspection of musk xylene in cosmetics.
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Characteristics and measurement principles of reference methods in clinical biochemistry were described.Implementation of reference systems is one of the most effective approaches to improve the accuracy and comparability of clinical laboratory test results.Reference methods are the key components of reference systems.Reference methods should have measurement uncertainties that meet the requirements of the intended use,and thus should be based on reliable measurement principles.For the well-defined biochemistry analytes,reference methods have been almost all based on instrumental analysis.Isotope dilution mass spectrometry (ID/MS) is considered most reliable and has been the major analytical principle of the reference methods.ID/MS analysis is accurate but expensive.Use of other validated instrumental analyses as reference measurement principles would be justified.
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Objective To develop a method for the determination of total cholesterol in serum by isotope dilution liquid chromatography tandem mass spectrometry (ID-LC/MS/MS).Methods Serum samples were supplemented by addition of [3,4-~(13)C_2]-cholesterol,hydrolyzed with alcoholic sodium hydroxide and oxidized into cholest-4-ene-3,6-dione by chromic acid.The oxidation products were analyzed by LC/MS/MS using atmospheric pressure chemical ionization (APCI) source and detection modes of multiple reaction monitoring (MRM) and single ion recording (SIR).Signals (peak areas) of the internal standard were corrected for the contributions of cholesterol and the signal ratios of cholesterol to internal standard for the calibrations were linearly regressed against cholesterol concentrations.The resulted regression equation was used for the calculation of serum cholesterol concentrations.Results The correlation coefficients between the peak area ratios and cholesterol concentrations were 0.999 9 and higher.Under MRM mode,the average within-run CV of the results obtained on 3 serum samples was 0.95% (ranged from 0.92% to 0.99%) and the total CVwas 0.86% (0.82% to 0.89%),and under SIR mode,the within-run CV was 0.64% (from 0.54% to 0.77%) and the total CVwas O.69% (0.62% to 0.81%),respectively. Results on certified reference materials (SRM 1951 a Level Ⅰ and Level Ⅱ;GBW 09145 and GBW 09147) showed an average bias of 0.23% (0.14% to 1.00%) under MRM mode,and 0.24% (0.07% to 1.27%) under SIR mode.Conclusions An ID-LC/MS/MS method for serum cholesterol has been developed.It is specific and precise and may be used as a candidate reference method.
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Objective To develop a candidate reference method for the measurement of progesterone in human serum.Methods The serum sample is mixed with the internal standard [3,4-~(13)C_2] progesterone.After extraction with n-hexane and purified by a aqueous solution of 2-Hydroxypropyl-?- cyclodextrin (HP-?-CD),the serum progesterone and labeled progesterone are converted to the 3-enol heptafluorobutyrate and analyzed by gas chromatography mass spectrometry (GC/MS) with selected ion monitoring.The concentration of serum progesterone is calculated by bracketing method.Results The results gave coefficients of variation (CVs) of 0.69% to 2.12%.The analytical recoveries ranged from 98.3% to 100.1%.The results of measuring certified reference materials of serum progesterone are agree with the target value.Conclusion The procedure for measuring progesterone in serum is a highly accurate and precise method and may be used as a candidate reference method for serum progesterone assays.