Background:Accurate quantification of 17-hydroxyprogesterone(17-OHP)in serum is vital for clinical and research applications.However,inter-laboratory variability in test results exists owing to the lack of a standardi...Background:Accurate quantification of 17-hydroxyprogesterone(17-OHP)in serum is vital for clinical and research applications.However,inter-laboratory variability in test results exists owing to the lack of a standardized reference measurement procedure(RMP).Therefore,in this study,we developed a highly accurate,cost-effective,and user-friendly candidate RMP(cRMP)for analyzing 17-OHP in serum.Methods:We quantified 17-OHP in serum using a one-step liquid–liquid extraction method with the addition of 17-OHP-^(13)C_(3),followed by liquid chromatography–tandem mass spectrometry.The ability of these methods to suppress interference was evaluated by chromatographic analysis.We assessed accuracy,specificity,the lower limit of quantitation,linearity,and matrix effects by following the standards specified by the Clinical and Laboratory Standards Institute.The consistency between the developed cRMP and the chemiluminescence method was evaluated through experiments with 120 clinical samples.Results:The developed cRMP required only 8 min for accurate quantification of serum 17-OHP without bias from matrix effects or interference from 19 metabolites added as potential interferents.The method exhibited favorable measurement performance,with a quantitation limit of 0.086 ng/mL,linear range of 0.1–400 ng/mL,a total imprecision of≤2.90%,spike recovery of 100.1%–100.6%,and relative deviations from assigned target values(RfB Institution)of−2.91%to 1.10%.The cRMP demonstrated good consistency with the conventional assay(chemiluminescence method),with a correlation coefficient R of 0.96977.Conclusion:A cRMP with high accuracy,cost-effectiveness,and convenient operation was developed for quantifying 17-OHP in serum.Its simplicity and robust performance make it an invaluable addition to the arsenal of analytical tools available for laboratories.This method can enhance the accuracy and reliability of 17-OHP measurements across various laboratories.展开更多
Using guinea pig andrenal microsomes, we studied the inhibitory effects of 20α-hydroxyprogesterone on steroid hydroxylation reactions catalyzed by cytochromes P-450. When 17α-hydroxyprogesterone was used as a substr...Using guinea pig andrenal microsomes, we studied the inhibitory effects of 20α-hydroxyprogesterone on steroid hydroxylation reactions catalyzed by cytochromes P-450. When 17α-hydroxyprogesterone was used as a substrate, 20α-hydroxyprogesterone functioned as a competitive inhibitor on the C_(17)—C_(20) bond cleavage reaction of P-450_(17α lyase). The inhibition constant, K_i was 1.37 μmol/L. 20α-hydroxyprogesterone also competitively inhibited the convertion of 17α-bydroxyprogesterone to 11-deoxycortisol by the action of P-450_(c21). The value of K_i was 1.73μmol/L. When progesterone was used as a substrate, 20α-hydroxyprogesterone inhibited neither the 21-hydroxylation of P-450_(c21). the C_(17)—C_(20) bond cleavage, nor 17α-hydroxylation of P-450_(17α lyase). Based on the seresults, we can deduce that the production of androstenedione from progesterone by the action of P-450_(17α lyase) proceeds through a successive monooxygenase reaction.展开更多
基金Science and Technology Project of Dongguan,Grant/Award Number:20211800905072Graduate Education Innovation Program Project of Guangdong Province,Grant/Award Number:2022KCXTD010+6 种基金Discipline Construction Project of Guangdong Medical University,Grant/Award Numbers:4SG22098G,4SG22259G,4SG23030G,4SG23143GNational Natural Science Foundation of China,Grant/Award Numbers:81870016,82270013Guangdong Basic and Applied Basic Research Foundation,Grant/Award Number:2022A1515010525Guangdong Medical UniversityShenzhen YHLO Biotech Co.,Ltd.Huazhong University of Science and TechnologyShenzhen Maternal and Child Health Care Hospital。
文摘Background:Accurate quantification of 17-hydroxyprogesterone(17-OHP)in serum is vital for clinical and research applications.However,inter-laboratory variability in test results exists owing to the lack of a standardized reference measurement procedure(RMP).Therefore,in this study,we developed a highly accurate,cost-effective,and user-friendly candidate RMP(cRMP)for analyzing 17-OHP in serum.Methods:We quantified 17-OHP in serum using a one-step liquid–liquid extraction method with the addition of 17-OHP-^(13)C_(3),followed by liquid chromatography–tandem mass spectrometry.The ability of these methods to suppress interference was evaluated by chromatographic analysis.We assessed accuracy,specificity,the lower limit of quantitation,linearity,and matrix effects by following the standards specified by the Clinical and Laboratory Standards Institute.The consistency between the developed cRMP and the chemiluminescence method was evaluated through experiments with 120 clinical samples.Results:The developed cRMP required only 8 min for accurate quantification of serum 17-OHP without bias from matrix effects or interference from 19 metabolites added as potential interferents.The method exhibited favorable measurement performance,with a quantitation limit of 0.086 ng/mL,linear range of 0.1–400 ng/mL,a total imprecision of≤2.90%,spike recovery of 100.1%–100.6%,and relative deviations from assigned target values(RfB Institution)of−2.91%to 1.10%.The cRMP demonstrated good consistency with the conventional assay(chemiluminescence method),with a correlation coefficient R of 0.96977.Conclusion:A cRMP with high accuracy,cost-effectiveness,and convenient operation was developed for quantifying 17-OHP in serum.Its simplicity and robust performance make it an invaluable addition to the arsenal of analytical tools available for laboratories.This method can enhance the accuracy and reliability of 17-OHP measurements across various laboratories.
文摘Using guinea pig andrenal microsomes, we studied the inhibitory effects of 20α-hydroxyprogesterone on steroid hydroxylation reactions catalyzed by cytochromes P-450. When 17α-hydroxyprogesterone was used as a substrate, 20α-hydroxyprogesterone functioned as a competitive inhibitor on the C_(17)—C_(20) bond cleavage reaction of P-450_(17α lyase). The inhibition constant, K_i was 1.37 μmol/L. 20α-hydroxyprogesterone also competitively inhibited the convertion of 17α-bydroxyprogesterone to 11-deoxycortisol by the action of P-450_(c21). The value of K_i was 1.73μmol/L. When progesterone was used as a substrate, 20α-hydroxyprogesterone inhibited neither the 21-hydroxylation of P-450_(c21). the C_(17)—C_(20) bond cleavage, nor 17α-hydroxylation of P-450_(17α lyase). Based on the seresults, we can deduce that the production of androstenedione from progesterone by the action of P-450_(17α lyase) proceeds through a successive monooxygenase reaction.
