Research article Received: 3 December 2013,
Revised: 31 March 2014,
Accepted: 3 April 2014
Published online in Wiley Online Library: 2 June 2014
(wileyonlinelibrary.com) DOI 10.1002/bmc.3232
Rapid determination of corticosterone in mouse plasma by ultra fast liquid chromatography-tandem mass spectrometry Huan Lia,b†, Xiao Liua,c†, Yanhong Pohb, Li Wua, Qi-Gang Zhoud,e* and Bao-Chang Caia* ABSTRACT: Major depressive disorder is a severe, life-threatening and highly prevalent psychiatric disorder. A high percentage of people suffering from depression are characterized by hyperactivity of the hypothalamic–pituitary–adrenal axis, resulting in plasma glucocorticoid (cortisol in human and corticosterone in rodent) elevations. Glucocorticoid is a critical molecule in the onset of pathology of depression. A simple, highly sensitive and specific method based on ultra-fast liquid chromatography–tandem mass spectrometry method has been developed for the quantitation of corticosterone in mouse plasma for the first time, which provides technical support for the high-throughput measurement for clinical determination of corticosterone in biological samples. Samples were spiked with methanol to precipitate the protein, and then chromatographed on an Agilent Zorbax Eclipse Plus C18 (100 × 2.1 mm,1.8 μm) column by linear gradient elution with methanol and 0.1% formic acid as the mobile phase within 5 min. The detection of corticosterone was performed on ultra-fast liquid chromatography–triple quadrupole tandem mass spectrometry in the positive ion. The ions [M + H]+ m/z 347.2 → m/z 311.1 for corticosterone and [M + H]+ m/z 363.2 → m/z 327.2 for hydrocortisone (internal standard) were used for quantitative determination. The lower quantification limit for corticosterone was 1 ng/mL. The validated method was successfully applied to the quantitation of corticosterone in mouse plasma. Copyright © 2014 John Wiley & Sons, Ltd. Keywords: corticosterone; MDD; plasma; quantitative determination; UFLC-MS/MS
Introduction
1860
Major depressive disorder (MDD) is a severe, life-threatening, and highly prevalent psychiatric disorder, affecting 10–30% of women and 7–15% of men (Bartolomucci and Leopardi, 2009; Zhou et al., 2007). It is established that a high percentage of people suffering from depression are characterized by hyperactivity of the hypothalamic–pituitary–adrenal (HPA) axis, resulting in plasma glucocorticoid (cortisol in human and corticosterone in rodent) elevations (Young et al., 1991). A prominent mechanism by which the brain reacts to acute and chronic stress is activation of the HPA axis (Airan et al., 2007). Long-term activation of the HPA axis induces a persistent increase in cortisol/corticosterone level in the plasma of human/rodent, which causes damage and molecular pathway dysfunction in the brain limbic system (Yu et al., 2008). Glucocorticoid is a critical molecule in the onset of pathology of depression. However, the existing methods, including highperformance liquid chromatography (HPLC), gas chromatography–mass spectrometry (GC-MS), fluorescence spectrometry and ELISA for monitoring corticosterone level in the plasma, are not precise, stable and convenient enough for studying depression. The detection time for corticosterone was 25 min using HPLC. Sample preparation for fluorescence spectrometry or GC-MS analysis is tedious, requiring more than 30 min static reaction and complicated sample derivatization procedure for analysis. ELISA employs a colorimetric enzyme reaction which can be affected by many factors. The method requires the avoidance of contact with light and metal, strict control of the
Biomed. Chromatogr. 2014; 28: 1860–1863
amount of acid added in sample analysis, and a long detection time, leading to a lack of proper validation testing (Little et al., 2008; Zhou et al., 2011; Shu et al., 2003; Hay and Mormède, 1997; Zhong and Suo, 1998; Sink et al., 2008). All of the methods listed above cannot achieve high-throughput screening, which is a clinical requirement for disease diagnosis. The timely advent of the technique of liquid chromatography coupled with tandem mass spectrometry has greatly enabled bioanalysts to * Correspondence to: Qi-Gang Zhou, Lerner Research Institute, 9500 Euclid Avenue Cleveland, Ohio 44195, US. E-mail: [email protected] Bao-Chang Cai, College of Pharmacy, Nanjing University of Chinese Medicine, No.138 Xianlin Road. Nanjing 210023, Jiangsu Province, China. E-mail: [email protected] †
The first two authors contribute equally to this work.
a
College of Pharmacy, Nanjing University of Chinese Medicine, Nanjing, China
b
School of Applied Science, Temasek Polytechnic, Singapore
c
College of Science, Cleveland State University, Cleveland, USA
d
College of Pharmacy, Nanjing Medical University, Nanjing, China
e
Lerner Research Institute, Cleveland Clinic, Cleveland, USA Abbreviations used: CMS, chronic mild stress; ESI, electrospray ionization; HPA, hypothalamic–pituitary–adrenal; MDD, major depressive disorder; MRM, multiple reaction monitor; UFLC-MS/MS, ultra-fast liquid chromatography–tandem mass spectrometry.
Copyright © 2014 John Wiley & Sons, Ltd.
Rapid determination of corticosterone in mouse plasma rise to the challenge of short timelines in biosample quantitation. It has been made possible to drastically reduce the chromatographic run time owing to the utilization of multiple reaction monitoring (MRM) mode. MRM contains two stages: a first stage selecting the mass of the intact analyte (parent ion) and, after fragmentation of the parent by collision with gas atoms, a second stage selecting a specific fragment of the parent, collectively generating selected reaction monitoring. The two mass filters produce a very short chromatographic run time with very specific and sensitive response for the selected analyte that can be used to detect and integrate a peak in a simple one-dimensional chromatographic separation of the biosample, which has in turn met the need for highthroughput approaches to biological sample analysis. Also, the analytes are routinely measured using this approach at high-throughput with a relatively high accuracy (coefficient of variation
Revised: 31 March 2014,
Accepted: 3 April 2014
Published online in Wiley Online Library: 2 June 2014
(wileyonlinelibrary.com) DOI 10.1002/bmc.3232
Rapid determination of corticosterone in mouse plasma by ultra fast liquid chromatography-tandem mass spectrometry Huan Lia,b†, Xiao Liua,c†, Yanhong Pohb, Li Wua, Qi-Gang Zhoud,e* and Bao-Chang Caia* ABSTRACT: Major depressive disorder is a severe, life-threatening and highly prevalent psychiatric disorder. A high percentage of people suffering from depression are characterized by hyperactivity of the hypothalamic–pituitary–adrenal axis, resulting in plasma glucocorticoid (cortisol in human and corticosterone in rodent) elevations. Glucocorticoid is a critical molecule in the onset of pathology of depression. A simple, highly sensitive and specific method based on ultra-fast liquid chromatography–tandem mass spectrometry method has been developed for the quantitation of corticosterone in mouse plasma for the first time, which provides technical support for the high-throughput measurement for clinical determination of corticosterone in biological samples. Samples were spiked with methanol to precipitate the protein, and then chromatographed on an Agilent Zorbax Eclipse Plus C18 (100 × 2.1 mm,1.8 μm) column by linear gradient elution with methanol and 0.1% formic acid as the mobile phase within 5 min. The detection of corticosterone was performed on ultra-fast liquid chromatography–triple quadrupole tandem mass spectrometry in the positive ion. The ions [M + H]+ m/z 347.2 → m/z 311.1 for corticosterone and [M + H]+ m/z 363.2 → m/z 327.2 for hydrocortisone (internal standard) were used for quantitative determination. The lower quantification limit for corticosterone was 1 ng/mL. The validated method was successfully applied to the quantitation of corticosterone in mouse plasma. Copyright © 2014 John Wiley & Sons, Ltd. Keywords: corticosterone; MDD; plasma; quantitative determination; UFLC-MS/MS
Introduction
1860
Major depressive disorder (MDD) is a severe, life-threatening, and highly prevalent psychiatric disorder, affecting 10–30% of women and 7–15% of men (Bartolomucci and Leopardi, 2009; Zhou et al., 2007). It is established that a high percentage of people suffering from depression are characterized by hyperactivity of the hypothalamic–pituitary–adrenal (HPA) axis, resulting in plasma glucocorticoid (cortisol in human and corticosterone in rodent) elevations (Young et al., 1991). A prominent mechanism by which the brain reacts to acute and chronic stress is activation of the HPA axis (Airan et al., 2007). Long-term activation of the HPA axis induces a persistent increase in cortisol/corticosterone level in the plasma of human/rodent, which causes damage and molecular pathway dysfunction in the brain limbic system (Yu et al., 2008). Glucocorticoid is a critical molecule in the onset of pathology of depression. However, the existing methods, including highperformance liquid chromatography (HPLC), gas chromatography–mass spectrometry (GC-MS), fluorescence spectrometry and ELISA for monitoring corticosterone level in the plasma, are not precise, stable and convenient enough for studying depression. The detection time for corticosterone was 25 min using HPLC. Sample preparation for fluorescence spectrometry or GC-MS analysis is tedious, requiring more than 30 min static reaction and complicated sample derivatization procedure for analysis. ELISA employs a colorimetric enzyme reaction which can be affected by many factors. The method requires the avoidance of contact with light and metal, strict control of the
Biomed. Chromatogr. 2014; 28: 1860–1863
amount of acid added in sample analysis, and a long detection time, leading to a lack of proper validation testing (Little et al., 2008; Zhou et al., 2011; Shu et al., 2003; Hay and Mormède, 1997; Zhong and Suo, 1998; Sink et al., 2008). All of the methods listed above cannot achieve high-throughput screening, which is a clinical requirement for disease diagnosis. The timely advent of the technique of liquid chromatography coupled with tandem mass spectrometry has greatly enabled bioanalysts to * Correspondence to: Qi-Gang Zhou, Lerner Research Institute, 9500 Euclid Avenue Cleveland, Ohio 44195, US. E-mail: [email protected] Bao-Chang Cai, College of Pharmacy, Nanjing University of Chinese Medicine, No.138 Xianlin Road. Nanjing 210023, Jiangsu Province, China. E-mail: [email protected] †
The first two authors contribute equally to this work.
a
College of Pharmacy, Nanjing University of Chinese Medicine, Nanjing, China
b
School of Applied Science, Temasek Polytechnic, Singapore
c
College of Science, Cleveland State University, Cleveland, USA
d
College of Pharmacy, Nanjing Medical University, Nanjing, China
e
Lerner Research Institute, Cleveland Clinic, Cleveland, USA Abbreviations used: CMS, chronic mild stress; ESI, electrospray ionization; HPA, hypothalamic–pituitary–adrenal; MDD, major depressive disorder; MRM, multiple reaction monitor; UFLC-MS/MS, ultra-fast liquid chromatography–tandem mass spectrometry.
Copyright © 2014 John Wiley & Sons, Ltd.
Rapid determination of corticosterone in mouse plasma rise to the challenge of short timelines in biosample quantitation. It has been made possible to drastically reduce the chromatographic run time owing to the utilization of multiple reaction monitoring (MRM) mode. MRM contains two stages: a first stage selecting the mass of the intact analyte (parent ion) and, after fragmentation of the parent by collision with gas atoms, a second stage selecting a specific fragment of the parent, collectively generating selected reaction monitoring. The two mass filters produce a very short chromatographic run time with very specific and sensitive response for the selected analyte that can be used to detect and integrate a peak in a simple one-dimensional chromatographic separation of the biosample, which has in turn met the need for highthroughput approaches to biological sample analysis. Also, the analytes are routinely measured using this approach at high-throughput with a relatively high accuracy (coefficient of variation
