Abstract
5-Azacytidine (5AC), a nucleoside analogue and hypomethylating agent, has anticancer properties and has been utilized in the treatment of various malignancies. 5AC is unstable and rapidly hydrolyzed to several by-products, including 5-azacytosine and 5-azauracil. A sensitive, reliable method was developed to quantitate 5AC using LC/MS/MS to perform pharmacokinetic and pharmacodynamic studies on 5AC combination therapy trials. Blood samples were collected in a heparinized tube and immediately processed for storage. To increase the stability of 5AC in plasma, 25 ng/mL tetrahydrouridine was added to the plasma and snap frozen. Plasma samples were extracted using acetonitrile then cleaned up by Oasis MCX ion exchange solid-phase extraction cartridges. 5AC was separated on an YMC Jsphr M80™ C 18 column with gradient elution of ammonium acetate (2 mM) with 0.1% formic acid and methanol mobile phase. 5AC elutes at 5.0 ± 0.2 min with a total run time of 30 min. Identification was through positive-ion mode and multiple reaction monitoring mode at m/z+ 244.9 → 113.0 for 5AC and m/z+ 242.0 → 126.0 for 5-methyl-2′-deoxycytidine, the internal standard. The lower limit of quantitation of 5AC was 5 ng/mL in human plasma, and linearity was observed from 5 to 500 ng/mL fitted by linear regression with 1/x weight. This method is 50 times more sensitive than previously published assays and successfully allows studies to characterize the pharmacokinetics and pharmacodynamics of 5AC.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 81-88 |
| Number of pages | 8 |
| Journal | Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences |
| Volume | 813 |
| Issue number | 1-2 |
| DOIs | |
| State | Published - Dec 25 2004 |
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Keywords
- 5-Azacytidine
- LC/MS/MS
- Pharmacokinetics
ASJC Scopus subject areas
- Biochemistry
Cite this
Quantification of 5-azacytidine in plasma by electrospray tandem mass spectrometry coupled with high-performance liquid chromatography. / Zhao, Ming; Rudek-Renaut, Michelle; He, Ping; Hartke, Carol; Gore, Steve; Carducci, Michael A; Baker, Sharyn D.
In: Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences, Vol. 813, No. 1-2, 25.12.2004, p. 81-88.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Quantification of 5-azacytidine in plasma by electrospray tandem mass spectrometry coupled with high-performance liquid chromatography
AU - Zhao, Ming
AU - Rudek-Renaut, Michelle
AU - He, Ping
AU - Hartke, Carol
AU - Gore, Steve
AU - Carducci, Michael A
AU - Baker, Sharyn D.
PY - 2004/12/25
Y1 - 2004/12/25
N2 - 5-Azacytidine (5AC), a nucleoside analogue and hypomethylating agent, has anticancer properties and has been utilized in the treatment of various malignancies. 5AC is unstable and rapidly hydrolyzed to several by-products, including 5-azacytosine and 5-azauracil. A sensitive, reliable method was developed to quantitate 5AC using LC/MS/MS to perform pharmacokinetic and pharmacodynamic studies on 5AC combination therapy trials. Blood samples were collected in a heparinized tube and immediately processed for storage. To increase the stability of 5AC in plasma, 25 ng/mL tetrahydrouridine was added to the plasma and snap frozen. Plasma samples were extracted using acetonitrile then cleaned up by Oasis MCX ion exchange solid-phase extraction cartridges. 5AC was separated on an YMC Jsphr M80™ C 18 column with gradient elution of ammonium acetate (2 mM) with 0.1% formic acid and methanol mobile phase. 5AC elutes at 5.0 ± 0.2 min with a total run time of 30 min. Identification was through positive-ion mode and multiple reaction monitoring mode at m/z+ 244.9 → 113.0 for 5AC and m/z+ 242.0 → 126.0 for 5-methyl-2′-deoxycytidine, the internal standard. The lower limit of quantitation of 5AC was 5 ng/mL in human plasma, and linearity was observed from 5 to 500 ng/mL fitted by linear regression with 1/x weight. This method is 50 times more sensitive than previously published assays and successfully allows studies to characterize the pharmacokinetics and pharmacodynamics of 5AC.
AB - 5-Azacytidine (5AC), a nucleoside analogue and hypomethylating agent, has anticancer properties and has been utilized in the treatment of various malignancies. 5AC is unstable and rapidly hydrolyzed to several by-products, including 5-azacytosine and 5-azauracil. A sensitive, reliable method was developed to quantitate 5AC using LC/MS/MS to perform pharmacokinetic and pharmacodynamic studies on 5AC combination therapy trials. Blood samples were collected in a heparinized tube and immediately processed for storage. To increase the stability of 5AC in plasma, 25 ng/mL tetrahydrouridine was added to the plasma and snap frozen. Plasma samples were extracted using acetonitrile then cleaned up by Oasis MCX ion exchange solid-phase extraction cartridges. 5AC was separated on an YMC Jsphr M80™ C 18 column with gradient elution of ammonium acetate (2 mM) with 0.1% formic acid and methanol mobile phase. 5AC elutes at 5.0 ± 0.2 min with a total run time of 30 min. Identification was through positive-ion mode and multiple reaction monitoring mode at m/z+ 244.9 → 113.0 for 5AC and m/z+ 242.0 → 126.0 for 5-methyl-2′-deoxycytidine, the internal standard. The lower limit of quantitation of 5AC was 5 ng/mL in human plasma, and linearity was observed from 5 to 500 ng/mL fitted by linear regression with 1/x weight. This method is 50 times more sensitive than previously published assays and successfully allows studies to characterize the pharmacokinetics and pharmacodynamics of 5AC.
KW - 5-Azacytidine
KW - LC/MS/MS
KW - Pharmacokinetics
UR - http://www.scopus.com/inward/record.url?scp=8844285234&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=8844285234&partnerID=8YFLogxK
U2 - 10.1016/j.jchromb.2004.09.012
DO - 10.1016/j.jchromb.2004.09.012
M3 - Article
C2 - 15556519
AN - SCOPUS:8844285234
VL - 813
SP - 81
EP - 88
JO - Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences
JF - Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences
SN - 1570-0232
IS - 1-2
ER -