An LC-MS/MS Method for the Determination of Cycloserine in Human Plasma: Application to Pharmacokinetic Study


  • Phongpan Mokmued Department of Medical Sciences, Ministry of Public Health, Thailand
  • Triporn Wattananat Department of Medical Sciences, Ministry of Public Health
  • Surakameth Mahasirimongkol Department of Medical Sciences, Ministry of Public Health, Thailand
  • Hideki Yanai Department of Clinical Laboratory, Fukujuji Hospital, Japan
  • Taisei Mushiroda Laboratory for Pharmacogenomics, RIKEN Center for Integrative Medical Sciences, Japan
  • Katsushi Tokunaga Department of Human Genetics, The University of Tokyo, Japan


Cycloserine, human plasma, LC-MS/MS, pharmacokinetics


A simple and rapid liquid chromatography-tandem mass spectrometric (LC-MS/MS) method was developed and validated for the determination of cycloserine in human plasma. Carbamazepine was used as an internal standard. The method utilized protein precipitation with acetonitrile followed by dilution with 50% methanol. The separation was performed on a C18 column using methanol-0.1% formic acid (70:30, v/v) as the mobile phase. Detection was performed on a quadrupole mass spectrometer using an electrospray ionization interface in multiple reaction monitoring mode. The method was proven to be specific, sensitive, accurate, precise, and linear over the concentration range of 0.3-70 µg/mL with correlation coefficients greater than 0.995. The intra- and interday precisions and accuracies were within 10.7%. The method was successfully applied to a pharmacokinetic study of cycloserine following the oral administration of 250 and 750 mg cycloserine capsules in 11 healthy Thai volunteers under fasting conditions.


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Batson, S., Chiara, C. D., Majce, V., Lloyd, A. J., Gobec, S., Rea, D., … Roper, D. I. (2017). Inhibition of D-Ala:D-Ala ligase through a phosphorylated form of the antibiotic D-cycloserine. Nature Communications, 8: 1939. doi: 10.1038/s41467-017-02118-7

Brennan, P.J., Young, D.B., & Robertson, B.D. (2008). Handbook of antituberculosis agents; Tuberculosis, 88(2): 100-101.

David, V., Ionescu, M., & Dumitrescu, V. (2001). Determination of cycloserine in human plasma by high-performance liquid chromatography with fluorescence detection, using derivatization with p-benzoquinone. Journal of Chromatography B: Biomedical Sciences and Applications, 761(1), 27-33. doi: 10.1016/s0378-4347(01)00293-6

Han, M., Jun, S. H., Lee, J. H., Park, K. U., Song, J., & Song, S. H. (2013). Method for simultaneous analysis of nine second-line anti-tuberculosis drugs using UPLC-MS/MS. Journal of Antimicrobial Chemotherapy, 68(9), 2066-2073. doi: 10.1093/jac/dkt154

Kumar, A. H., Polisetty, A. K., Sudha, V., Vijayakumar, A., & Ramachandran, G. (2018). A selective and sensitive high performance liquid chromatography assay for the determination of cycloserine in human plasma. Indian Journal of Tuberculosis, 65(2), 118-123. doi: 10.1016/j.ijtb.2017.08.034

Kim, Y., Lee, Y., Lee, E., Lee, S., Kwon, J., Shim, W. K. C.-K., & Lee, M. (2000). Bioequivalence assessment of closerin capsule to Dura seromycin capsule of cycloserine after a single oral dose administration to healthy male volunteers. Int. Journal of Clinical Pharmacology and Therapeutics, 38(10), 461-466. doi: 10.5414/cpp38461

Mao, Z., Wang, X., Li, B., Jin, J., Xu, M., Liu, Y., & Di, X. (2017). A simplified LC-MS/MS method for rapid determination of cycloserine in small-volume human plasma using protein precipitation coupled with dilution techniques to overcome matrix effects and its application to a pharmacokinetic study. Analytical and Bioanalytical Chemistry, 409(11), 3025-3032. doi: 10.1007/s00216-017-0249-2

Mulubwa, M., & Mugabo, P. (2019). Sensitive Ultra-performance Liquid Chromatography Tandem Mass Spectrometry Method for Determination of Cycloserine in Plasma for a Pharmacokinetics Study. Journal of Chromatographic Science, 57(6), 560–564. doi: 10.1093/chromsci/bmz028

Park, S.-I., Oh, J., Jang, K., Yoon, J., Moon, S. J., Park, J. S., … Chung, J.-Y. (2015). Pharmacokinetics of Second-Line Antituberculosis Drugs after Multiple Administrations in Healthy Volunteers. Antimicrobial Agents and Chemotherapy, 59(8), 4429-4435. doi: 10.1128/aac.00354-15

Patel, D. S., Sharma, N., Patel, M. C., Patel, B. N., Shrivastav, P. S., & Sanyal, M. (2011). Development and validation of a selective and sensitive LC-MS/MS method for determination of cycloserine in human plasma: Application to bioequivalence study. Journal of Chromatography B, 879(23), 2265-2273. doi: 10.1016/j.jchromb.2011.06.011

Pattar, V. P., & Nandibewoor, S. T. (2016). Electrochemical studies for the determination of an antibiotic drug, d-cycloserine, in pharmaceutical and human biological samples. Journal of Taibah University for Science, 10(1), 92-99. doi: 10.1016/j.jtusci.2015.05.003

Polagani, S. R., Pilli, N. R., Maddela, R., Gajula, R., & Gandu, V. (2013). A rapid and sensitive liquid chromatography-tandem mass spectrometric assay for cycloserine in 50μL of human plasma: Its pharmacokinetic application. Journal of Pharmaceutical and Biomedical Analysis, 76, 21-27. doi: 10.1016/j.jpba.2012.11.036

Stepanova, E. S., Ovcharov, M. V., Barsegyan, S. S., & Chistyakov, V. V. (2016). Determination of Cycloserine in Blood Plasma by HPLC/MS: Application to Bioequivalence Studies. Pharmaceutical Chemistry Journal, 50(3), 195-199. doi: 10.1007/s11094-016-1422-1

Yaroshenko, D. V., Grigoriev, A. V., & Sidorova, A. A. (2014). Development and validation of a LC-MS/MS method for d-cycloserine determination in human plasma for bioequivalence study. Analytical and Bioanalytical Chemistry, 406(3), 923-927. doi: 10.1007/s00216-013-7521-x

Zhou, H., Wu, G., Hu, X., Zhu, M., Zhai, Y., Liu, J., … Wu, L. (2015). Pharmacokinetic Properties and Tolerability of Cycloserine Following Oral Administration in Healthy Chinese Volunteers: A Randomized, Open-Label, Single- and Multiple-Dose 3-Way Crossover Study. Clinical Therapeutics, 37(6), 1292-1300. doi: 10.1016/j.clinthera.2015.03.015

Zhang, W., Wan, S., Chen, L., Wang, X., Wang, Z., & Huang, Y. (2018). Determination of cycloserine in microdialysis samples using liquid chromatography-tandem mass spectrometry with benzoyl chloride derivatization. Biomedical Chromatography, 32(6). doi: 10.1002/bmc.4187

Zhu, M., Nix, D. E., Adam, R. D., Childs, J. M., & Peloquin, C. A. (2001). Pharmacokinetics of Cycloserine under Fasting Conditions and with High-Fat Meal, Orange Juice, and Antacids. Pharmacotherapy, 21(8), 891-897. doi: 10.1592/phco.21.11.891.34524

US Department of Health and Human Services, Food and Drug Administration, Center for Drug Evaluation and Research, Center for Veterinary Medicine. (2018). Bioanalytical Method Validation Guidance for Industry. Retrieved from

WHO treatment guidelines for drug-resistant tuberculosis. (n.d.). Retrieved October 11, 2018, from;jsessionid=3FEEDBD4BB16F729EC82D51FD68B0FE2?sequence=1






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