Abstract
A novel analytical approach for the targeted profiling of bile acids (BAs) in human serum/plasma based on liquid chromatography quadrupole time-of-flight mass spectrometry (LC-QTOF-MS) is presented. Reversed-phase chromatography enabled the baseline separation of 15 human BA species which could be readily detected by accurate mass analysis in negative ion mode. Blood proteins were removed by methanol precipitation in the presence of deuterium-labeled internal standards which allowed BA quantification in 50 μl plasma/serum. The assay was validated according to FDA guidance achieving quantification limits from 7.8 to 156 nM. Calibration curves prepared in charcoal-stripped serum/plasma showed excellent regression coefficients (R 2 > 0.997) and covered quantities from 7.8 to 10,000 nM depending on the analyzed species. Intra- and inter-day accuracy and precision were below 15 % for all analytes. Apparent extraction recoveries were above 97 %, and ion suppression rates were between 4 and 53 %. Mean BA level in serum/plasma from healthy volunteers ranged from 11 ± 4 nM (tauroursodeoxycholic acid) to 1321 ± 1442 nM (glycochenodeoxycholic acid). As a proof of concept, the assay was applied to plasma samples derived from a clinical phase I study of myrcludex B, a novel first-in-class virus entry inhibitor for the treatment of chronic hepatitis B and D. The results demonstrate that myrcludex-induced inhibition of the hepatic BA transporter Na+-taurocholate cotransporting polypeptide (NTCP) significantly affects plasma BA level. These observations provide novel insights into drug-induced metabolic responses and will be indispensable for the assessment of side effects and dose-finding processes during future clinical trials.
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de Aguiar V, Tarling EJ, Edwards PA (2013) Cell Metab 17:657–669
Stieger B, Meier Y, Meier PJ (2007) Pflugers Arch 453:611–620
Dawson PA, Lan T, Rao A (2009) J Lipid Res 50:2340–2357
Barnes S, Gallo GA, Trash DB, Morris JS (1975) J Clin Pathol 28:506–509
Tribe RM, Dann AT, Kenyon AP, Seed P, Shennan AH, Mallet A (2010) Am J Gastroenterol 105:585–595
Patterson AD, Maurhofer O, Beyoglu D, Lanz C, Krausz KW, Pabst T, Gonzalez FJ, Dufour JF, Idle JR (2011) Cancer Res 71:6590–6600
Galeazzi R, Lorenzini I, Orlandi F (1980) Dig Dis Sci 25:108–112
Lutjohann D, Hahn C, Prange W, Sudhop T, Axelson M, Sauerbruch T, von Bergmann K, Reichel C (2004) Int J Clin Pharmacol Ther 42:307–313
Lepri L, Heimler D, Desideri PG (1984) J Chromatogr 288:461–468
Fujihira E, Takahashi N, Minato A, Uenoyama K, Ogiso T (1972) Chem Pharm Bull (Tokyo) 20:2719–2721
Sian MS, Rains AJ (1979) Clin Chim Acta 98:243–252
Shaw R, Elliott WH (1976) Anal Biochem 74:273–281
Elliott WH (1980) Lipids 15:764–769
Eneroth P, Gordon B, Ryhage R, Sjovall J (1966) J Lipid Res 7:511–523
Murphy GM, Billing BH, Baron DN (1970) J Clin Pathol 23:594–598
Roda A, Roda E, Aldini R, Capelli M, Festi D, Sama C, Mazzella G, Morselli AM, Barbara L (1980) Clin Chem 26:1677–1682
Steiner C, von Eckardstein A, Rentsch KM (2010) J Chromatogr B Anal Technol Biomed Life Sci 878:2870–2880
Burkard I, von Eckardstein A, Rentsch KM (2005) J Chromatogr B Anal Technol Biomed Life Sci 826:147–159
Tagliacozzi D, Mozzi AF, Casetta B, Bertucci P, Bernardini S, Di IC, Urbani A, Federici G (2003) Clin Chem Lab Med 41:1633–1641
Scherer M, Gnewuch C, Schmitz G, Liebisch G (2009) J Chromatogr B Anal Technol Biomed Life Sci 877:3920–3925
Garcia-Canaveras JC, Donato MT, Castell JV, Lahoz A (2012) J Lipid Res 53:2231–2241
Xiang X, Han Y, Neuvonen M, Laitila J, Neuvonen PJ, Niemi M (2010) J Chromatogr B Anal Technol Biomed Life Sci 878:51–60
Wang H, Yeh CY, Li K, Chung-Davidson YW, Li W (2015) J Chromatogr B Anal Technol Biomed Life Sci 980:72–78
John C, Werner P, Worthmann A, Wegner K, Todter K, Scheja L, Rohn S, Heeren J, Fischer M (2014) J Chromatogr A 1371:184–195
Han J, Liu Y, Wang R, Yang J, Ling V, Borchers CH (2015) Anal Chem 87:1127–1136
Jantti SE, Kivilompolo M, Ohrnberg L, Pietilainen KH, Nygren H, Oresic M, Hyotylainen T (2014) Anal Bioanal Chem 406:7799–7815
Wang Y, Griffiths WJ (2007) Curr Anal Chem 3:103–126
Paul M, Ippisch J, Herrmann C, Guber S, Schultis W (2014) Anal Bioanal Chem 406:4425–4441
Bobeldijk I, Hekman M, de Vries-van der W, Coulier L, Ramaker R, Kleemann R, Kooistra T, Rubingh C, Freidig A, Verheij E (2008) J Chromatogr B Anal Technol Biomed Life Sci 871:306–313
Ding J, Lund ET, Zulkoski J, Lindsay JP, McKenzie DL (2013) Bioanalysis 5:2481–2494
Want EJ, Coen M, Masson P, Keun HC, Pearce JT, Reily MD, Robertson DG, Rohde CM, Holmes E, Lindon JC, Plumb RS, Nicholson JK (2010) Anal Chem 82:5282–5289
Voelker T, Wang H, Irish M, Wang J, Harrison S, Reuschel S, Bennett P, Meng M (2013) Bioanalysis 5:1229–1248
Urban S, Bartenschlager R, Kubitz R, Zoulim F (2014) Gastroenterology 147:48–64
Haefeli WE, Blank A, Mikus G, Mier W, Alexandrov A, Urban S (2012) Hepatology 56:372a
Momose T, Tsubaki T, Iida T, Nambara T (1997) Lipids 32:775–778
Guidance for industry: bioanalytical method validation (2001) U.S. Department of Health and Human Services, Food and Drug Administration, Center for Drug Evaluation and Research (CDER), Center for Veterinary Medicine (CVM), Rockville, MD, USA. http://www.fda.gov/downloads/Drugs/Guidances/ucm070107.pdf. Accessed on 15 Jan 2015
Dalsgaard PW (2013) J Forensic Sci Criminol 1:1–6
Ni Y, Lempp FA, Mehrle S, Nkongolo S, Kaufman C, Falth M, Stindt J, Koniger C, Nassal M, Kubitz R, Sultmann H, Urban S (2014) Gastroenterology 146:1070–1083
Nkongolo S, Ni Y, Lempp FA, Kaufman C, Lindner T, Esser-Nobis K, Lohmann V, Mier W, Mehrle S, Urban S (2014) J Hepatol 60:723–731
Oehler N, Volz T, Bhadra OD, Kah J, Allweiss L, Giersch K, Bierwolf J, Riecken K, Pollok JM, Lohse AW, Fehse B, Petersen J, Urban S, Lutgehetmann M, Heeren J, Dandri M (2014) Hepatology 60:1483–1493
Kubitz R, Droge C, Kluge S, Stindt J, Haussinger D (2014) Drug Discov Today Technol 12:e55–e67
Gong IY, Kim RB (2013) Drug Metab Pharmacokinet 28:4–18
Vaz FM, Paulusma CC, Huidekoper H, de RM, Lim C, Koster J, Ho-Mok K, Bootsma AH, Groen AK, Schaap FG, Oude Elferink RP, Waterham HR, Wanders RJ (2014) Hepatology 61:260–267
Hofmann AF, Hagey LR (2014) J Lipid Res 55:1553–1595
Lepist EI, Gillies H, Smith W, Hao J, Hubert C, St CR III, Brouwer KR, Ray AS (2014) PLoS One 9, e87548
MacFarlane GD, Scheller DG, Ersfeld DL, Shaw LM, Venkatarmanan R, Sarkozi L, Mullins R, Fox BR (1999) Clin Chem 45:1449–1458
Mashige F, Tanaka N, Maki A, Kamei S, Yamanaka M (1981) Clin Chem 27:1352–1356
Mashige F, Imai K, Osuga T (1976) Clin Chim Acta 70:79–86
Acknowledgments
We thank Markus König for his excellent technical assistance. This work was supported by grants from the Hans Lutz Merkle Foundation, the Robert Bosch Foundation (Stuttgart, Germany), the Medical Faculty of the University of Heidelberg, and the German Center for Infection Research (DZIF).
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Haag, M., Hofmann, U., Mürdter, T.E. et al. Quantitative bile acid profiling by liquid chromatography quadrupole time-of-flight mass spectrometry: monitoring hepatitis B therapy by a novel Na+-taurocholate cotransporting polypeptide inhibitor. Anal Bioanal Chem 407, 6815–6825 (2015). https://doi.org/10.1007/s00216-015-8853-5
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DOI: https://doi.org/10.1007/s00216-015-8853-5