{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,7]],"date-time":"2026-07-07T14:38:16Z","timestamp":1783435096530,"version":"3.54.6"},"reference-count":39,"publisher":"MDPI AG","issue":"19","license":[{"start":{"date-parts":[[2025,9,26]],"date-time":"2025-09-26T00:00:00Z","timestamp":1758844800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"European Union\u2019s Horizon Europe research and innovation program under the Marie Sklodowska-Curie","award":["101130235"],"award-info":[{"award-number":["101130235"]}]},{"name":"National Research, Development and Innovation Office, Hungary","award":["NKFIH FK 146930"],"award-info":[{"award-number":["NKFIH FK 146930"]}]}],"content-domain":{"domain":["www.mdpi.com"],"crossmark-restriction":true},"short-container-title":["IJMS"],"abstract":"<jats:p>Tetrahydroprotoberberine alkaloids (THPBs) are bioactive natural products bearing stereogenic centers that frequently exhibit enantiomer-specific pharmacological effects. Xylopinine (XPN), a representative THPB, shows cytotoxic, antimicrobial, and antimalarial activity in vitro, and displays pronounced stereoselectivity in vivo, with the naturally occurring (S)-enantiomer emphasizing the need for reliable enantioselective analysis. In this study, we present the synthesis of racemic XPN from norlaudanosine, and its first comprehensive cyclodextrin-assisted capillary electrophoresis screening dedicated to the enantioseparation of XPN. Sulfated- and sulfobutyl-ether-\u03b2-cyclodextrin (S-\u03b2-CyD, SBE-\u03b2-CyD) provided efficient resolution (Rs &gt; 3), while heptakis-(6-deoxy-6-(2-carboxyethyl)thio)-\u03b2-CyD (subetadex, SBX) yielded outstanding separation (Rs &gt; 9). The enantiomer migration order was consistently R,S, except when using SBE-\u03b2-CyD, which showed the inverse sequence. Chiral HPLC using a Chiralpak AD column in polar organic mode with methanol modified with 0.1% diethylamine as mobile phase enabled the semi-preparative isolation of XPN enantiomers, with the (S)-enantiomer exceeding 95% purity. The absolute configuration was confirmed by circular dichroism spectroscopy. 1H NMR titration and 2D rotating-frame nuclear Overhauser effect correlation spectroscopy (ROESY) consistently revealed multi-site recognition of XPN by SBX, supporting the inclusion of both aromatic rings (A and D).<\/jats:p>","DOI":"10.3390\/ijms26199405","type":"journal-article","created":{"date-parts":[[2025,9,26]],"date-time":"2025-09-26T08:31:28Z","timestamp":1758875488000},"page":"9405","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["Enantioselective Complexation of Xylopinine: A Cyclodextrin-Assisted CE and NMR Study"],"prefix":"10.3390","volume":"26","author":[{"ORCID":"https:\/\/orcid.org\/0009-0004-8348-4545","authenticated-orcid":false,"given":"Erzs\u00e9bet","family":"V\u00e1rnagy","sequence":"first","affiliation":[{"name":"Department of Pharmacognosy, Semmelweis University, \u00dcll\u0151i \u00fat 26, H-1085 Budapest, Hungary"},{"name":"Center for Pharmacology and Drug Research & Development, Semmelweis University, \u00dcll\u0151i \u00fat 26, H-1085 Budapest, Hungary"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5341-319X","authenticated-orcid":false,"given":"Gerg\u0151","family":"T\u00f3th","sequence":"additional","affiliation":[{"name":"Center for Pharmacology and Drug Research & Development, Semmelweis University, \u00dcll\u0151i \u00fat 26, H-1085 Budapest, Hungary"},{"name":"Department of Pharmaceutical Chemistry, Semmelweis University, H\u0151gyes Endre u. 9, H-1092 Budapest, Hungary"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"S\u00e1ndor","family":"Hosztafi","sequence":"additional","affiliation":[{"name":"Center for Pharmacology and Drug Research & Development, Semmelweis University, \u00dcll\u0151i \u00fat 26, H-1085 Budapest, Hungary"},{"name":"Department of Pharmaceutical Chemistry, Semmelweis University, H\u0151gyes Endre u. 9, H-1092 Budapest, Hungary"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Milo","family":"Malanga","sequence":"additional","affiliation":[{"name":"CarboHyde Ltd., Berlini u. 47-49, H-1045 Budapest, Hungary"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ida","family":"Fej\u0151s","sequence":"additional","affiliation":[{"name":"Department of Pharmacognosy, Semmelweis University, \u00dcll\u0151i \u00fat 26, H-1085 Budapest, Hungary"},{"name":"Center for Pharmacology and Drug Research & Development, Semmelweis University, \u00dcll\u0151i \u00fat 26, H-1085 Budapest, Hungary"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7056-6825","authenticated-orcid":false,"given":"Szabolcs","family":"B\u00e9ni","sequence":"additional","affiliation":[{"name":"Integrative Health and Environmental Analysis Research Laboratory, Department of Analytical Chemistry, Institute of Chemistry, ELTE E\u00f6tv\u00f6s Lor\u00e1nd University, P\u00e1zm\u00e1ny P\u00e9ter S\u00e9t\u00e1ny 1\/A, H-1117 Budapest, Hungary"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2025,9,26]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"8","DOI":"10.1093\/pcp\/pcl041","article-title":"Identification of a WRKY Protein as a Transcriptional Regulator of Benzylisoquinoline Alkaloid Biosynthesis in Coptis Japonica","volume":"48","author":"Kato","year":"2007","journal-title":"Plant Cell Physiol."},{"key":"ref_2","doi-asserted-by":"crossref","unstructured":"O\u2019Connor, S.E. (2010). ChemInform Abstract: Alkaloid Biosynthesis. ChemInform, 41.","DOI":"10.1002\/chin.201012266"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"1113","DOI":"10.1517\/14712598.2012.704014","article-title":"Berberine on Metabolic and Cardiovascular Risk Factors: An Analysis from Preclinical Evidences to Clinical Trials","volume":"12","author":"Derosa","year":"2012","journal-title":"Expert Opin. Biol. Ther."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"504","DOI":"10.1002\/ptr.6252","article-title":"Berberine and Barberry (Berberis vulgaris): A Clinical Review","volume":"33","author":"Imenshahidi","year":"2019","journal-title":"Phyther. Res."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"150","DOI":"10.1016\/j.phytochem.2006.10.004","article-title":"Quaternary Protoberberine Alkaloids","volume":"68","author":"Marek","year":"2007","journal-title":"Phytochemistry"},{"key":"ref_6","doi-asserted-by":"crossref","unstructured":"Menezes, L.R.A., D\u2019Sousa Costa, C.O., Rodrigues, A.C.B.D.C., Santo, F.R.D.E., Nepel, A., Dutra, L.M., Silva, F.M.A., Soares, M.B.P., Barison, A., and Costa, E.V. (2016). Cytotoxic Alkaloids from the Stem of Xylopia laevigata. Molecules, 21.","DOI":"10.3390\/molecules21070890"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"278","DOI":"10.1002\/ptr.4710","article-title":"HPLC Analysis of Stephania rotunda Extracts and Correlation with Antiplasmodial Activity","volume":"27","author":"Bory","year":"2013","journal-title":"Phyther. Res."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"537","DOI":"10.1016\/j.jep.2014.04.024","article-title":"Ethnobotany, Phytochemistry and Pharmacology of Stephania rotunda Lour","volume":"154","author":"Desgrouas","year":"2014","journal-title":"J. Ethnopharmacol."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"132","DOI":"10.1016\/j.jep.2007.02.005","article-title":"Antimalarial Activity of Alkaloids Isolated from Stephania rotunda","volume":"112","author":"Chea","year":"2007","journal-title":"J. Ethnopharmacol."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"677","DOI":"10.1055\/s-2003-41126","article-title":"Aporphine Alkaloids from Guatteria spp. with Leishmanicidal Activity","volume":"69","author":"Montenegro","year":"2003","journal-title":"Planta Med."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"381","DOI":"10.1016\/j.jep.2012.10.052","article-title":"New Antiplasmodial Alkaloids from Stephania rotunda","volume":"145","author":"Baghdikian","year":"2013","journal-title":"J. Ethnopharmacol."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"10","DOI":"10.1254\/jjp.16.10","article-title":"Comparison of the Optical Isomers of Xylopinine","volume":"16","author":"Nakanishi","year":"1966","journal-title":"Jpn. J. Pharmacol."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"447","DOI":"10.1007\/s10822-019-00194-z","article-title":"Significantly Different Effects of Tetrahydroberberrubine Enantiomers on Dopamine D1\/D2 Receptors Revealed by Experimental Study and Integrated in Silico Simulation","volume":"33","author":"Ge","year":"2019","journal-title":"J. Comput. Aided. Mol. Des."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"108","DOI":"10.1016\/j.jchromb.2005.08.012","article-title":"Chiral Liquid Chromatography Resolution and Stereoselective Pharmacokinetic Study of Tetrahydropalmatine Enantiomers in Dogs","volume":"826","author":"Hong","year":"2005","journal-title":"J. Chromatogr. B Anal. Technol. Biomed. Life Sci."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"293","DOI":"10.1002\/chir.20161","article-title":"Stereoselective Pharmacokinetics of Tetrahydropalmatine after Oral Administration of (-)-Enantiomer and the Racemate","volume":"17","author":"Hong","year":"2005","journal-title":"Chirality"},{"key":"ref_16","first-page":"34","article-title":"Chiral HPLC Determination and Stereoselective Pharmacokinetics of Tetrahydroberberine Enantiomers in Rats","volume":"43","author":"Zhang","year":"2011","journal-title":"Chirality"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"2716","DOI":"10.3762\/bjoc.17.183","article-title":"The Ethoxycarbonyl Group as Both Activating and Protective Group in N-Acyl-Pictet-Spengler Reactions Using Methoxystyrenes. A Short Approach to Racemic 1-Benzyltetrahydroisoquinoline Alkaloids","volume":"17","author":"Keller","year":"2021","journal-title":"Beilstein J. Org. Chem."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"939","DOI":"10.1007\/s00216-005-0238-8","article-title":"Chiral High-Performance Liquid Chromatographic Separation of the Enantiomers of Tetrahydropalmatine and Tetrahydroberberine, Isolated from Corydalis Yanhusuo","volume":"384","author":"Zhai","year":"2006","journal-title":"Anal. Bioanal. Chem."},{"key":"ref_19","doi-asserted-by":"crossref","unstructured":"V\u00e1rnai, B., Zsila, F., Szak\u00e1cs, Z., Gar\u00e1di, Z., Malanga, M., and B\u00e9ni, S. (2022). Sulfobutylation of Beta-Cyclodextrin Enhances the Complex Formation with Mitragynine: An NMR and Chiroptical Study. Int. J. Mol. Sci., 23.","DOI":"10.3390\/ijms23073844"},{"key":"ref_20","doi-asserted-by":"crossref","unstructured":"Doh\u00e1rszky, A., V\u00e1gi, E.M., K\u00f6ncz\u00f6l, \u00c1., Simon, A., V\u00e1rnagy, E., Muratov, M., Steiger, K.I., V\u00e1rnai, B., B\u00e9ni, S., and Riethm\u00fcller, E. (2024). Kratom Alkaloids: A Blood\u2013Brain Barrier Specific Membrane Permeability Assay-Guided Isolation and Cyclodextrin Complexation Study. Molecules, 29.","DOI":"10.3390\/molecules29225302"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"461375","DOI":"10.1016\/j.chroma.2020.461375","article-title":"Single Isomer Cyclodextrins as Chiral Selectors in Capillary Electrophoresis","volume":"1627","author":"Kalydi","year":"2020","journal-title":"J. Chromatogr. A"},{"key":"ref_22","first-page":"364","article-title":"Enantiomeric Separation by Cyclodextrin Modified Capillary Zone Electrophoresis (CD-CZE) of Quaternary Tetrahydroprotoberberine Alkaloids","volume":"57","author":"Koyama","year":"2000","journal-title":"Chem. Pharm. Bull."},{"key":"ref_23","doi-asserted-by":"crossref","unstructured":"Yan, B., Huang, Z.A., Yahaya, N., and Chen, D.D.Y. (2020). Enantioselective Analysis in Complex Matrices Using Capillary Electrophoresis-Mass Spectrometry: A Case Study of the Botanical Drug Corydalis Rhizoma. J. Chromatogr. B Anal. Technol. Biomed. Life Sci., 1152.","DOI":"10.1016\/j.jchromb.2020.122216"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"6721","DOI":"10.1021\/acs.joc.6b01419","article-title":"Synthesis of Tetrahydroisoquinoline Alkaloids and Related Compounds through the Alkylation of Anodically Prepared \u03b1-Amino Nitriles","volume":"81","author":"Benmekhbi","year":"2016","journal-title":"J. Org. Chem."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"3297","DOI":"10.6023\/cjoc202304032","article-title":"Asymmetric Synthesis of (S)-(-)-Xylopinine and (S)-(+)-Laudanosine","volume":"43","author":"Zhang","year":"2023","journal-title":"Chinese J. Org. Chem."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"2689","DOI":"10.1002\/ejoc.200500095","article-title":"Enantioselective Synthesis of (+)-(S)-Laudanosine and (\u2013)-(S)-Xylopinine","volume":"2005","author":"Mujahidin","year":"2005","journal-title":"Eur. J. Org. Chem."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"e202200878","DOI":"10.1002\/asia.202200878","article-title":"Visible Light Driven Metal-Free Photoredox Catalyzed \u03b1-Benzylation and \u03b1-Oxygenation of N-Substituted Tetrahydroisoquinolines: Applications to Synthesis of Natural Products","volume":"17","author":"Elavarasan","year":"2022","journal-title":"Chem. An Asian J."},{"key":"ref_28","doi-asserted-by":"crossref","unstructured":"V\u00e1rnagy, E., T\u00f3th, G., Hosztafi, S., Dob\u00f3, M., Fej\u0151s, I., and B\u00e9ni, S. (2025). Chiral Recognition Mechanism of Benzyltetrahydroisoquinoline Alkaloids: Cyclodextrin-Mediated Capillary Electrophoresis, Chiral HPLC, and NMR Spectroscopy Study. Molecules, 30.","DOI":"10.3390\/molecules30051125"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"2783","DOI":"10.1021\/ja01188a044","article-title":"Curariform Activity and Chemical Structure. II. Synthesis in the Benzyltetrahydroisoquinoline Series1","volume":"70","author":"Craig","year":"1948","journal-title":"J. Am. Chem. Soc."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"753","DOI":"10.1002\/jhet.5570050603","article-title":"A Modified Synthesis of Codamine under Eschweiler-Clarke Conditions","volume":"5","author":"Kametani","year":"1968","journal-title":"J. Heterocycl. Chem."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"95","DOI":"10.1002\/chir.22772","article-title":"Enantioseparation of Racecadotril Using Polysaccharide-Type Chiral Stationary Phases in Polar Organic Mode","volume":"30","author":"Foroughbakhshfasaei","year":"2018","journal-title":"Chirality"},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"1818","DOI":"10.1002\/elps.202100076","article-title":"Enantioseparation of Solriamfetol and Its Major Impurity Phenylalaninol by Capillary Electrophoresis Using Sulfated Gamma Cyclodextrin","volume":"42","author":"Malanga","year":"2021","journal-title":"Electrophoresis"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"311","DOI":"10.1007\/s10847-013-0301-6","article-title":"Studies on \u03b1-, \u03b2-, and \u03b3-Cyclodextrin Inclusion Complexes of Isoquinoline Alkaloids Berberine, Palmatine and Coralyne","volume":"78","author":"Hazra","year":"2014","journal-title":"J. Incl. Phenom. Macrocycl. Chem."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"264","DOI":"10.1002\/hlca.200490013","article-title":"Berberine\/\u03b3-Cyclodextrin Inclusion Structure Studied by 1H-NMR Spectroscopy and Molecular-Dynamics Calculations","volume":"87","author":"Kamigauchi","year":"2004","journal-title":"Helv. Chim. Acta"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"5297","DOI":"10.2147\/IJN.S464994","article-title":"Supersaturated Drug Delivery System of Oxyberberine Based on Cyclodextrin Nanoaggregates: Preparation, Characterization, and In Vivo Application","volume":"19","author":"Huang","year":"2024","journal-title":"Int. J. Nanomedicine"},{"key":"ref_36","doi-asserted-by":"crossref","unstructured":"Miskolczy, Z., Megyesi, M., and Bicz\u00f3k, L. (2022). Entropy-Driven Inclusion of Natural Protoberberine Alkaloids in Sulfobutylether-\u03b2-Cyclodextrin. Molecules, 27.","DOI":"10.3390\/molecules27217514"},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"158","DOI":"10.1016\/j.chroma.2016.04.004","article-title":"CEval: All-in-one software for data processing and statistical evalua-tions in affinity capillary electrophoresis","volume":"1445","year":"2016","journal-title":"J. Chromatogr. A"},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"2764","DOI":"10.1002\/jssc.201200502","article-title":"Affinity capillary electrophoresis method for investigation of bile salts complexation with sulfobutyl ether-\u03b2-cyclodextrin","volume":"35","author":"Jensen","year":"2012","journal-title":"J. Sep. Sci."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"1032","DOI":"10.1002\/elps.201100489","article-title":"Determination of stability constants of complexes of neutral analytes with charged cyclodextrins by affinity capillary electrophoresis","volume":"33","year":"2012","journal-title":"Electrophoresis"}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/26\/19\/9405\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,9,26]],"date-time":"2025-09-26T08:38:11Z","timestamp":1758875891000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/26\/19\/9405"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,9,26]]},"references-count":39,"journal-issue":{"issue":"19","published-online":{"date-parts":[[2025,10]]}},"alternative-id":["ijms26199405"],"URL":"https:\/\/doi.org\/10.3390\/ijms26199405","relation":{},"ISSN":["1422-0067"],"issn-type":[{"value":"1422-0067","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,9,26]]}}}