Beta-blockers are chiral compounds with enantiomers that have different bioactivity, which means that while one is active, the other can be inactive or even harmful. Due to their high consumption and incomplete degradation in waste water, they may reach surface waters and affect aquatic organisms. To address this issue we developed a chromatographic method suitable for determining beta-blocker enantiomers in surface waters. It was tested on five beta-blockers (acebutolol, atenolol, bisoprolol, labetalol and metoprolol) and validated on bisoprolol enantiomers. Good enantioseparation of all analysed beta-blockers was achieved on the Chirobiotic V column with the mobile phase composed of methanol/acetic acid/triethylamine (100/0.20/0.15 v/v/v) at a flow rate of 0.5 mL/min and column temperature of 45 °C. Method proved to be linear in the concentration range from 0.075 µg/mL to 5 µg/mL, and showed good recovery. The limits of bisoprolol enantiomer detection were 0.025 µg/mL and 0.026 µg/mL and of quantification 0.075 µg/mL and 0.075 µg/mL. Despite its limitations, it seems to be a promising method for bisoprolol enantiomer analysis in surface water samples. Further research could focus on waste water analysis, where enantiomer concentrations may be high. Furthermore, transferring the method to a more sensitive one such as liquid chromatography coupled with tandem mass spectrometry and using ammonium acetate as the mobile phase additive instead of acetic acid and triethylamine would perhaps yield much lower limits of detection and quantification.

Beta-blokatori su kiralni spojevi s enantiomerima različite bioaktivnosti, dakle, dok je jedan enantiomer aktivan, drugi može biti neaktivan ili čak štetan za organizam. Zbog njihove visoke potrošnje i nepotpune razgradnje u pogonima za preradu otpadnih voda, postoji mogućnost da se pojave u prirodnim vodama i negativno utječu na vodene organizme. Stoga je u ovom radu razvijena kromatografska metoda za određivanje enantiomera beta-blokatora u prirodnim vodama. Metoda je testirana na pet beta-blokatora (acebutolol, atenolol, bisoprolol, labetalol i metoprolol) te validirana za enantiomere bisoprolola. Dobra enantioseparacija svih analiziranih beta-blokatora postignuta je na koloni Chirobiotic V sastava mobilne faze metanol/octena kiselina/trietilamin (100:0,2:0,15 v/v/v) pri protoku od 0,5 mL/min i temperaturi od 45 °C. Metodom je postignuta dobra linearnost u području od 0,075 μg/mL do 5 μg/mL s dobrim analitičkim povratom. Granice detekcije pojedinih enantiomera bisoprolola bile su 0,025 μg/mL i 0,026 μg/mL, a granice kvantifikacije 0,075 μg/ mL za oba enantiomera. Unatoč ograničenjima metode, pokazala se kao obećavajuća metoda analize enantiomera bisoprolola u površinskim vodama. Daljnja istraživanja mogla bi se izvoditi na otpadnim vodama, gdje bi koncentracije enantiomera mogle biti više. Također, korištenjem osjetljivije metode, primjerice vezanoga sustava tekućinske kromatografije i tandemne spektrometrije masa, te korištenjem amonijeva acetata kao aditiva mobilnoj fazi umjesto octene kiseline i trietilamina, mogle bi se postići znatno niže granice detekcije i kvantifikacije.

Department of Chemistry Faculty of Science University of Zagreb Zagreb Croatia

Department of Physical and Macromolecular Chemistry Faculty of Science Charles University Prague Czech Republic

Zobrazit více v PubMed

Poirier L, Tobe SW. Contemporary use of β-blockers: clinical relevance of subclassification. Can J Cardiol. 2014;30(Suppl 5):S9–15. doi: 10.1016/j.cjca.2013.12.001. PubMed DOI

Nguyen LA, He H, Pham-Huy C. Chiral drugs: an overview. Int J Biomed Sci. 2006;2:85–100. PMID: 23674971. PubMed PMC

Hernando MD, Gómez MJ, Agüera A, Fernández-Alba AR. LC-MS analysis of basic pharmaceuticals (beta-blockers and anti-ulcer agents) in wastewater and surface water. Trend Anal Chem. 2007;26:581–94. doi: 10.1016/j.trac.2007.03.005. DOI

Massarsky A, Trudeau VL, Moon TW. β-Blockers as endocrine disruptors: the potential effects of human β-blockers on aquatic organisms. J Exp Zool. 2011;315:251–65. doi: 10.1002/jez.672. PubMed DOI

Stanley JK, Ramirez AJ, Mottaleb M, Chambliss CK, Brooks BW. Enantiospecific toxicity of the β-blocker propranolol to Daphnia magna and Pimephales promelas. Environ Toxicol Chem. 2006;25:1780–6. doi: 10.1897/05-298r1.1. PubMed DOI

de Andrés F, Castañeda G, Ríos Á. Use of toxicity assays for enantiomeric discrimination of pharmaceutical substances. Chirality. 2009;21:751–9. doi: 10.1002/chir.20675. PubMed DOI

Sun L, Xin L, Peng Z, Jin R, Jin Y, Qian H, Fu Z. Toxicity and enantiospecific diffrences of two β-blockers, propranolol and metoprolol, in the embryos and larvae of Zebrafish Danio rerio. Environ Toxicol. 2014;29:1367–78. doi: 10.1002/tox.21867. PubMed DOI

Stringham RW, Ye YK. Chiral separation of amines by high-performance liquid chromatography using polysaccharide stationary phases and acidic additives. J Chromatogr A. 2006;1101:86–93. doi: 10.1016/j.chroma.2005.09.065. PubMed DOI

Ali I, Gaitonde VD, Aboul-Enein HY, Hussain A. Chiral separation of β-adrenergic blockers on CelluCoat column by HPLC. Talanta. 2009;78:458–63. doi: 10.1016/j.talanta.2008.11.043. PubMed DOI

Santoro MIRM, Cho HS, Kedor-Hackmann ERM. Enantiomeric separation and quantitative determination of atenolol in tablets by chiral high-performance liquid chromatography. Drug Dev Ind Pharm. 2000;26:1107–10. doi: 10.1081/DDC-100100275. PubMed DOI

Morante-Zarcero S, Sierra I. Comparative HPLC methods for β-blockers separation using different types of chiral stationary phases in normal phase and polar organic phase elution modes. Analysis of propranolol enantiomers in natural waters. J Pharm Biomed Anal. 2012;62:33–41. doi: 10.1016/j.jpba.2011.12.029. PubMed DOI

Armstrong DW, Chen S, Chang C, Chang S. A new approach for the direct resolution of racemic beta adrenergic blocking agents by HPLC. J Liq Chromatogr. 1992;15:545–56. doi: 10.1080/10826079208017191. DOI

Bosáková Z, Cuřínová E, Tesařová E. Comparison of vancomycin-based stationary phases with different chiral selector coverage for enantioselective separation of selected drugs in high-performance liquid chromatography. J Chromatogr A. 2005;1088:94–103. doi: 10.1016/j.chroma.2005.01.017. PubMed DOI

Mistry B, Leslie JL, Eddington ND. Enantiomeric separation of metoprolol and α-hydroxymetoprolol by liquid chromatography and fluorescence detection using a chiral stationary phase. J Chromatogr B. 2001;758:153–161. doi: 10.1016/S0378-4347(01)00166-9. PubMed DOI

Evans SE, Davies P, Lubben A, Kasprzyk-Hordern B. Determination of chiral pharmaceuticals and illicit drugs in wastewater and sludge using microwave assisted extraction, solid-phase extraction and chiral liquid chromatography coupled with tandem mass spectrometry. Anal Chim Acta. 2015;882:112–26. doi: 10.1016/j.aca.2015.03.039. PubMed DOI

Hefnawy MM, Sultan MA, Al-Shehri MM. Development of an HPLC method for the quantitation of bisoprolol enantiomers in pharmaceutical products using a teicoplanin chiral stationary phase and fluorescence detection. J Liq Chromatogr RT 2006; 29:2901–14. DOI

Bagnall JP, Evans SE, Wort MT, Luben AT, Kasprzyk-Hordern B. Using chiral liquid chromatography quadrupole time-of-flight mass spectrometry for the analysis of pharmaceuticals and illicit drugs in surface and wastewater at the enantiomeric level. J Chromatogr A. 2012;1249:115–29. doi: 10.1016/j.chroma.2012.06.012. PubMed DOI

Barclay VKH, Tyrefors NL, Johansson IM, Pettersson CE. Chiral analysis of metoprolol and two of its metabolites, α-hydroxymetoprolol and deaminated metoprolol, in wastewater using liquid chromatography-tandem mass spectrometry. J Chromatogr A. 2012;1269:208–17. doi: 10.1016/j.chroma.2012.09.090. PubMed DOI

Agency for Medicinal Products and Medical Devices of Croatia (HALMED) http://www.halmed.hr Annual report on drug utilization for 2017 [displayed 17 October 2018]. Available at.

State Institute for Drug Control. http://www.sukl.cz Annual summary on drug deliveries for 2017 [displayed 18 December 2018]. Available at.

Meierjohann A, Brozinski JM, Kronberg L. Seasonal variation of pharmaceutical concentrations in a river/lake system in Eastern Finland. Enivoron Sci Processes Impacts. 2016;18:342–9. doi: 10.1039/C5EM00505A. PubMed DOI

Lee HB, Sarafin K, Peart TE. Determination of β-blockers and β2-agonists in sewage by solid-phase extraction and liquid chromatography-tandem mass spectrometry. J Chromatogr A. 2007;1148:158–67. doi: 10.1016/j.chroma.2007.03.024. PubMed DOI

Miège C, Favier M, Brosse C, Canler JP, Coquery M. Occurrence of betablockers in effluents of wastewater treatment plants from the Lyon area (France) and risk assessment for the downstream rivers. Talanta. 2006;70:739. doi: 10.1016/j.talanta.2006.07.002. –. PubMed DOI

International Conference on Harmonization of Technical Requirements for Registration of Pharmaceuticals for Human Use. ICH Harmonized Tripartite Guideline: Validation of Analytical Procedures: Text and Methodology Q2(R1) [displayed 27 November. 2018. https://database.ich.org/sites/default/files/Q2_R1__Guideline.pdf Available at.

Beesley TE, Lee JT. Method development strategy and applications update for CHIROBIOTIC chiral stationary phases. J Liq Chromatogr R T. 2009;32:1733–67. doi: 10.1080/10826070902959489. DOI