J. Sep. Sci. 2014, 37, 295–303 295 Veronika ˇ Sol´ ınov ´ a Martin Maxmili ´ an Kaiser Milo ˇ s Luk ´ aˇ c Zlatko Janeba V´ aclav Ka ˇ si ˇ cka Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, Prague, Czech Republic Received October 3, 2013 Revised November 8, 2013 Accepted November 10, 2013 Research Article Enantiopurity analysis of new types of acyclic nucleoside phosphonates by capillary electrophoresis with cyclodextrins as chiral selectors CE methods have been developed for the chiral analysis of new types of six acyclic nucleoside phosphonates, nucleotide analogs bearing [(3-hydroxypropan-2-yl)- 1H-1,2,3-triazol-4-yl]phosphonic acid, 2-[(diisopropoxyphosphonyl)methoxy]propanoic acid, or 2–(phosphonomethoxy)propanoic acid moieties attached to adenine, guanine, 2,6- diaminopurine, uracil, and 5-bromouracil nucleobases, using neutral and cationic cyclodex- trins as chiral selectors. With the exception of the 5-bromouracil-derived acyclic nucleoside phosphonate with a 2-(phosphonomethoxy)propanoic acid side chain, the R and S enan- tiomers of the other five acyclic nucleoside phosphonates were successfully separated with sufficient resolutions, 1.51–2.94, within a reasonable time, 13–28 min, by CE in alkaline BGEs (50 mM sodium tetraborate adjusted with NaOH to pH 9.60, 9.85, and 10.30, respec- tively) containing 20 mg/mL -cyclodextrin as the chiral selector. A baseline separation of the R and S enantiomers of the 5-bromouracil-derived acyclic nucleoside phosphonate with 2-(phosphonomethoxy)propanoic acid side chain was achieved within a short time of 7 min by CE in an acidic BGE (20:40 mM Tris/phosphate, pH 2.20) using 60 mg/mL quaternary ammonium -cyclodextrin chiral selector. The developed methods were applied for the assessment of the enantiomeric purity of the above acyclic nucleoside phosphonates. The preparations of all these compounds were found to be synthesized in pure enantiomeric forms. Using UV absorption detection at 206 nm, their concentration detection limits were in the low micromolar range. Keywords: Acyclic nucleoside phosphonates / CE / Chiral analysis / Cyclodextrins / Nucleotide analogs DOI 10.1002/jssc.201301092 1 Introduction Acyclic nucleoside phosphonates (ANPs) represent a dis- tinguished class of antiviral agents and drugs [1–4]. Three ANPs have been approved worldwide for clinical use [1, 2]: (S)-1-[3-hydroxy-2-(phosphonomethoxy)propyl]cytosine (cid- ofovir, Vistide R  ) for the treatment of cytomegalovirus re- tinitis in AIDS (acquired immunodeficiency syndrome) pa- tients, 9-[2-(phosphonomethoxy)ethyl]adenine (as its oral prodrug adefovir dipivoxil, Hepsera R  ) for the treatment of chronic hepatitis B virus infections [5], and (R)-9-[2- (phosphonomethoxy)propyl]adenine, PMPA, (as its oral pro- Correspondence: Dr. V´ aclav Kaˇ siˇ cka, Institute of Organic Chem- istry and Biochemistry, v.v.i., Academy of Sciences of the Czech Republic, Flemingovo nam. 2, 166 10 Prague 6, Czech Republic E-mail: kasicka@uochb.cas.cz Fax: +420-220-183-592 Abbreviations: AIDS, acquired immunodeficiency syndrome; ANP, acyclic nucleoside phosphonate; CD, cyclodextrin; HIV, human immunodeficiency virus; PMPA, (R)-9-[2- (phosphonomethoxy)propyl]adenine; QA--CD, quaternary- ammonium-–cyclodextrin drug tenofovir disoproxil fumarate, Viread R  ) for the treat- ment of HIV (human immunodeficiency virus) infection (itself or in combination with other antivirals) and chronic hepatitis B virus infections [6]. Their mechanism of ac- tion is based on interfering with DNA polymerase or re- verse transcriptase resulting in viral DNA chain termina- tion. Many other ANPs were found to exhibit important antiviral properties, e.g. derivatives of 2,6-diaminopurine and “open ring” derivatives of 2,4-diaminopyrimidine. (S)-9- [2-Hydroxy-3-(phosphonomethoxy)propyl]adenine possesses a broad spectrum activity against DNA viruses (including herpes-, adeno-, pox-, and iridoviruses), as well as against retroviruses [4]. Acyclic nucleoside analogs of adenosine, like (S)-9-(2,3-dihydroxypropyl)adenine produced in Czechoslo- vakia as Duvira gel R  , and structurally related 3-(adenin- 9-yl)-2-hydroxypropanoic acid constitute another class of broad-spectrum antivirals interfering with (S)-adenosyl-L- homocysteine hydrolase, thus inhibiting viral RNA matura- tion. ANPs exhibit not only antiviral but also cytostatic [7], antiparasitic [8–10], and immunomodulatory activities [11]. Dedicated to Prof. Frantiˇ sek ˇ Svec on the occasion of his 70th birthday. C  2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim www.jss-journal.com