Drug Delivery and Translational Research

, Volume 9, Issue 6, pp 1057–1066 | Cite as

Total drug quantification in prodrugs using an automated elemental analyzer

  • Yingwen Hu
  • David M. Stevens
  • Sonny Man
  • Rachael M. Crist
  • Jeffrey D. ClogstonEmail author
Original Article


Polymeric prodrugs have become an increasingly popular strategy for improving the pharmacokinetic properties of active pharmaceutical ingredients (API). Therefore, identifying a robust method for quantification of the API in these prodrug products is a key part of the drug development process. Current drug quantification methods include hydrolysis followed by reversed phase high-performance liquid chromatography (RP-HPLC), size exclusion chromatography (SEC)-based molecular weight determination, and mass spectrometry. These methods tend to be time-consuming and often require challenging method development. Here, we present a comparative study highlighting the automated elemental analyzer as a facile approach to drug quantification in this up-and-coming class of therapeutics. A polymeric prodrug using poly(l-lysine succinylated) (PLS) and the drug lamivudine (LAM) was prepared and analyzed using the elemental analyzer in comparison to the traditional approaches of hydrolysis followed by RP-HPLC and SEC using multi-angle light scattering (MALS) detection. The elemental analysis approach showed excellent agreement with the conventional methods but proved much less laborious, highlighting this as a rapid and sensitive analytical method for the quantitative determination of drug loading in polymeric prodrug products.


Elemental analysis Drug loading Polymer–drug conjugates Nanoparticle Nanomedicine Prodrug 


Funding information

This project has been funded in whole or in part with federal funds from the National Cancer Institute, National Institutes of Health, under Contract No. HHSN261200800001E.

Compliance with ethical standards


The content of this publication does not necessarily reflect the views or policies of the Department of Health and Human Services nor does mention of trade names, commercial products, or organizations imply endorsement by the U.S. Government.

Conflict of interest

The authors declare that they have no competing interests.

Supplementary material

13346_2019_649_MOESM1_ESM.docx (54 kb)
ESM 1 (DOCX 54 kb)


  1. 1.
    Koynova R, Tenchov B. Recent Progress in liposome production, relevance to drug delivery and nanomedicine. Recent Pat Nanotechnol. 2015;9(2):86–93.CrossRefGoogle Scholar
  2. 2.
    Bae Y, Kataoka K. Intelligent polymeric micelles from functional poly(ethylene glycol)-poly(amino acid) block copolymers. Adv Drug Deliv Rev. 2009;61(10):768–84. Scholar
  3. 3.
    Abd Ellah NH, Abouelmagd SA. Surface functionalization of polymeric nanoparticles for tumor drug delivery: approaches and challenges. Expert Opin Drug Deliv. 2017;14(2):201–14. Scholar
  4. 4.
    Wadhwa S, Mumper RJ. Polymer-drug conjugates for anticancer drug delivery. Crit Rev Ther Drug Carrier Syst. 2015;32(3):215–45.CrossRefGoogle Scholar
  5. 5.
    Li C, Wallace S. Polymer-drug conjugates: recent development in clinical oncology. Adv Drug Deliv Rev. 2008;60(8):886–98. Scholar
  6. 6.
    Dragojevic S, Ryu JS, Raucher D. Polymer-based prodrugs: improving tumor targeting and the solubility of small molecule drugs in cancer therapy. Molecules. 2015;20(12):21750–69. Scholar
  7. 7.
    Pang X, Jiang Y, Xiao Q, Leung AW, Hua H, Xu C. pH-responsive polymer-drug conjugates: design and progress. J Control Release. 2016;222:116–29. Scholar
  8. 8.
    Senanayake TH, Lu Y, Bohling A, Raja S, Band H, Vinogradov SV. Encapsulation of poorly soluble drugs in polymer-drug conjugates: effect of dual-drug nanoformulations on cancer therapy. Pharm Res. 2014;31(6):1605–15. Scholar
  9. 9.
    Chang M, Zhang F, Wei T, Zuo T, Guan Y, Lin G, et al. Smart linkers in polymer-drug conjugates for tumor-targeted delivery. J Drug Target. 2016;24(6):475–91. Scholar
  10. 10.
    Zou J, Yu Y, Li Y, Ji W, Chen CK, Law WC, et al. Well-defined diblock brush polymer-drug conjugates for sustained delivery of paclitaxel. Biomater Sci. 2015;3(7):1078–84. Scholar
  11. 11.
    Liu Z, Wang Y, Zhang N. Micelle-like nanoassemblies based on polymer-drug conjugates as an emerging platform for drug delivery. Expert Opin Drug Deliv. 2012;9(7):805–22. Scholar
  12. 12.
    Etrych T, Kovar L, Strohalm J, Chytil P, Rihova B, Ulbrich K. Biodegradable star HPMA polymer-drug conjugates: biodegradability, distribution and anti-tumor efficacy. J Control Release. 2011;154(3):241–8. Scholar
  13. 13.
    Sousa-Herves A, Wurfel P, Wegner N, Khandare J, Licha K, Haag R, et al. Dendritic polyglycerol sulfate as a novel platform for paclitaxel delivery: pitfalls of ester linkage. Nanoscale. 2015;7(9):3923–32. Scholar
  14. 14.
    Penugonda S, Kumar A, Agarwal HK, Parang K, Mehvar R. Synthesis and in vitro characterization of novel dextran-methylprednisolone conjugates with peptide linkers: effects of linker length on hydrolytic and enzymatic release of methylprednisolone and its peptidyl intermediates. J Pharm Sci. 2008;97(7):2649–64. Scholar
  15. 15.
    Abu-Fayyad A, Nazzal S. Synthesis, characterization, and in-vitro antitumor activity of the polyethylene glycol (350 and 1000) succinate derivatives of the tocopherol and tocotrienol isomers of vitamin E. Int J Pharm. 2017;519(1–2):145–56. Scholar
  16. 16.
    Badawy SI, Williams RC, Gilbert DL. Chemical stability of an ester prodrug of a glycoprotein IIb/IIIa receptor antagonist in solid dosage forms. J Pharm Sci. 1999;88(4):428–33. Scholar
  17. 17.
    Yoshikawa M, Endo H, Hoshino K, Sugawara Y, Takaiti O, Kanda S, et al. A new method for the high performance liquid chromatographic determination of TA-870, a dopamine prodrug (catechol ester compound). Biomed Chromatogr. 1990;4(5):181–7. Scholar
  18. 18.
    Zhang H, Wang J, Mao W, Huang J, Wu X, Shen Y, et al. Novel SN38 conjugate-forming nanoparticles as anticancer prodrug: in vitro and in vivo studies. J Control Release. 2013;166(2):147–58. Scholar
  19. 19.
    Yang JY, Zhang R, Pan HZ, Li YL, Fang YX, Zhang LB, et al. Backbone degradable N-(2-hydroxypropyl)methacrylamide copolymer conjugates with gemcitabine and paclitaxel: impact of molecular weight on activity toward human ovarian carcinoma xenografts. Mol Pharm. 2017;14(5):1384–94. Scholar
  20. 20.
    Veronese FM, Schiavon O, Pasut G, Mendichi R, Andersson L, Tsirk A, et al. PEG-doxorubicin conjugates: influence of polymer structure on drug release, in vitro cytotoxicity, biodistribution, and antitumor activity. Bioconjug Chem. 2005;16(4):775–84. Scholar
  21. 21.
    Hussain MA, Abbas K, Amin M, Lodhi BA, Iqbal S, Tahir MN, et al. Novel high-loaded, nanoparticulate and thermally stable macromolecular prodrug design of NSAIDs based on hydroxypropylcellulose. Cellulose. 2015;22(1):461–71. Scholar
  22. 22.
    Culmo RF. The elemental analysis of various classes of chemical compounds using CHN: PerkinElmer Application Note.Google Scholar
  23. 23.
  24. 24.
    Fadeeva VP, Tikhova VD, Nikulicheva ON. Elemental analysis of organic compounds with the use of automated CHNS analyzers. J Anal Chem. 2008;63(11):1094–106. Scholar
  25. 25.
    Inukai Y, Tanaka Y, Matsuda T, Mihara N, Yamada K, Nambu N, et al. Removal of boron(III) by N-methylglucamine-type cellulose derivatives with higher adsorption rate. Anal Chim Acta. 2004;511(2):261–5. Scholar
  26. 26.
    Karnitz O, Gurgel LVA, de Freitas RP, Gil LF. Adsorption of Cu(II), Cd(II), and Pb(II) from aqueous single metal solutions by mercerized cellulose and mercerized sugarcane bagasse chemically modified with EDTA dianhydride (EDTAD). Carbohydr Polym. 2009;77(3):643–50. Scholar
  27. 27.
    Shah KJ, Mishra MK, Shukla AD, Imae T, Shah DO. Controlling wettability and hydrophobicity of organoclays modified with quaternary ammonium surfactants. J Colloid Interface Sci. 2013;407:493–9. Scholar
  28. 28.
    Hermanova S, Zarevucka M, Bousa D, Pumera M, Sofer Z. Graphene oxide immobilized enzymes show high thermal and solvent stability. Nanoscale. 2015;7(13):5852–8. Scholar
  29. 29.
    Matejovic I. Determination of carbon, hydrogen, and nitrogen in soils by automated elemental analysis (dry combustion method). Commun Soil Sci Plant Anal. 1993;24(17–18):2213–22. Scholar
  30. 30.
    Huang YF, Kuan WH, Lo SL, Lin CF. Total recovery of resources and energy from rice straw using microwave-induced pyrolysis. Bioresour Technol. 2008;99(17):8252–8. Scholar
  31. 31.
    Sheykhan M, Ma'mani L, Ebrahimi A, Heydari A. Sulfamic acid heterogenized on hydroxyapatite-encapsulated gamma-Fe2O3 nanoparticles as a magnetic green interphase catalyst. J Mol Catal A Chem. 2011;335(1–2):253–61. Scholar
  32. 32.
    Li PH, Li BL, Hu HC, Zhao XN, Zhang ZH. Ionic liquid supported on magnetic nanoparticles as highly efficient and recyclable catalyst for the synthesis of beta-keto enol ethers. Catal Commun. 2014;46:118–22. Scholar
  33. 33.
    U.S. Provisional Application No. 62/572,733, filed October 16, 2017.Google Scholar
  34. 34.
    PCT/US2018/55794, filed October 15, 2018.Google Scholar
  35. 35.
    Hamid MHM, Elsaman T. A stability-indicating RP-HPLC-UV method for determination and chemical hydrolysis study of a novel naproxen prodrug. J Chem. 2017;5285671:10. Scholar
  36. 36.
    Bennett DB, Adams NW, Li X, Feijen J, Kim SW. Drug-coupled poly(amino acids) as polymeric prodrugs. J Bioact Compat Polym. 1988;3(1):44–52. Scholar
  37. 37.
    Li C, Wildes F, Winnard P, Artemov D, Penet MF, Bhujwalla ZM. Conjugation of poly-L-lysine to bacterial cytosine deaminase improves the efficacy of enzyme/prodrug cancer therapy. J Med Chem. 2008;51(12):3572–82. Scholar
  38. 38.
    Wang C, Tang F, Wang XY, Li LD. Synthesis and application of biocompatible gold core-poly-(L-lysine) shell nanoparticles. Colloid Surface A. 2016;506:425–30. Scholar
  39. 39.
    Sriram D, Yogeeswari P, Gopal G. Synthesis, anti-HIV and antitubercular activities of lamivudine prodrugs. Eur J Med Chem. 2005;40(12):1373–6. Scholar
  40. 40.
    Kim KH, Kim ND, Seong BL. Discovery and development of anti-HBV agents and their resistance. Molecules. 2010;15(9):5878–908. Scholar
  41. 41.
    Hollander I, Kunz A, Hamann PR. Selection of reaction additives used in the preparation of monomeric antibody-calicheamicin conjugates. Bioconjug Chem. 2008;19(1):358–61. Scholar
  42. 42.
    Quiles S, Raisch KP, Sanford LL, Bonner JA, Safavy A. Synthesis and preliminary biological evaluation of high-drug-load paclitaxel-antibody conjugates for tumor-targeted chemotherapy. J Med Chem. 2010;53(2):586–94. Scholar
  43. 43.
    Ludlam PR, King JG. Size exclusion chromatography of urea formaldehyde resins in dimethylformamide containing lithium-chloride. J Appl Polym Sci. 1984;29(12):3863–72. Scholar

Copyright information

© Controlled Release Society 2019

Authors and Affiliations

  1. 1.Nanotechnology Characterization Laboratory, Cancer Research Technology Program, Leidos Biomedical Research, Inc.Frederick National Laboratory for Cancer ResearchFrederickUSA

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