Cyclodextrins: Applications in chemical analyses

Transcription

1 Cyclodextrins: Applications in chemical analyses Tamás Sohajda May 7th, 2015 CYCLONHIT Workshop Research Center of Natural Sciences of Hungarian Academy of Sciences, Budapest 1

2 The ultimate purpose Analytical purposes Physical isolation of incompatible compounds Control volatility and sublimation Influence chemical stability Catalysis Aqueous solubility enhancement Guest specific interactions Increase bioavailability Reduce or eliminate sideeffects and irritations 2

3 Fields of CD enabled chemical analysis Everything comes down to supramolecular complex formation Chiral separation Chiral columns Single molecule sensors Sample preparation Non-chiral separation Selective recognition of analytes Sensitivity improvement Determination of complex stability Complex stoichiometry Complex structure 3

4 Fields of CD enabled chemical analysis Everything comes down to supramolecular complex formation Determination of complex stability Complex stoichiometry Complex structure 4

5 Determination of complex stability Capillary electrophoresis Short analysis, easy to optimize, minute material consumption - costs Great performance, highly variable parameters, simple sample preparation Automatization, aqueous/non-aqueous UV active analytes are preferred, dependent on protonation state Poor robustness of the methods + A + A + - 5

6 Determination of apparent stability constants vincadifformine RAMEB c CD 75 mm binding and enantioseparation (stoichiometry) (S) (R) R S = mm 25 mm 10 mm R S =2.09 R S =1.70 R S = mm R S =0.6 0 mm t (min) T. Sohajda et al, J Pharm Biomed Anal, 53, ,

7 Screening results (Dapoxetine) CD derivative RAMEB-CD Binding constant (1/M), Resulation Resolution value K = 510 (1) Apparent average binding constants RAMEG-CD CM-β-CD DS~3 CM-γ-CD CE-β-CD SP-α-CD DS~2 K S = 360 (2) K R = 590 (8) R S = 3,32 K < 5 K S = 54 (5) K R = 61 (3) R S = 1,38 K < 5 K < 5 Preferred cavity size/ds/susbtituent for complexation Enantioselectivity SP-β-CD DS~4 SP-γ-CD DS~2 SHP-γ-CD DS~3 K S = 280 (6) K R = 310 (8) R S = 1,01 K < 5 K < 5 Enantiomer Migration Order SB-α-CD DS~4 K = 28 (1) SB-β-CD DS~4 K S = 610 (1) K R = 690 (2) R S = 0,62 G. Neumajer et al, J Pharm Biomed Anal, 62, 42-47,

8 Determination of apparent stability constants vincadifformine HPGCD H 11 H 12 H 10 H 9 NH O c CD /c Vindiff H N O CH H 3 C 22 H 9 (+) H 9 (-) H 11 (+) H 11 (-) binding (enantioseparation) H 9 (+/-) H 11 (+/-) H 10, 12 (+/-) 8 T. Sohajda et al, J Pharm Biomed Anal, 53, , 2010

9 Δδ x 0,018 Complex stoichiometry Job s plot 1H NMR stoichiometry Tosylated pregabalin - BCD Tos-preg β-cd 0,013 0,008 0,003-0,002 0,0 0,1 0,2 0,3 0,4 0,5 0,6 0,7 0,8 0,9 1,0 x -0,007-0,012 Sz. Beni et al, J Pharm Biomed Anal, 51, ,

10 Complex structure 2D ROESY Tosylated pregabalin - BCD H 3 C CH 3 O - structure (OH) 7 O HN O S O CH 3 (OH) 7 (OH) 7 CH 3 O O S O NH O - (OH) 7 (OH) 7 H 3 C CH 3 Sz. Beni et al, J Pharm Biomed Anal, 51, , 2010 (OH) 7 10

11 Comparison Method Costs Time Material need Analytes Information quality Capillary electrophoresis NMR spectroscopy Phase solubility 11

12 Fields of CD enabled chemical analysis Everything comes down to supramolecular complex formation Chiral separation Sensitivity improvement Single molecule sensors Sample preparation Non-chiral separation Selective recognition of analytes Chiral columns 12

13 Non-chiral separations Dye trace analysis textile industry Using the right CD at the right concentration the selectivity can be enhanced! Fluorescein Brilliant Blue Tartrazine Allura Red Curcumine 10 mm Hydroxypropyl BCD 10 mm Random methyl BCD 10 mm BCD without CD time / min 13

14 Non-chiral separations Using the right CD at the right concentration the selectivity can be enhanced! 14

15 Non-chiral separations Pharmacopeial example Iloprost monograph, polar related substances: Mobile phase: mix 330 m of acetonitrile R1 and 670 ml of 12 g/l BCD adjusted to ph 2.0 with phosphroic acid R1. 15

16 Non-chiral separations Pros: Selector type, quality and concentration is flexible Cheap columns and additives Cons: The selector may affect detection (UV, MS) High material need Reliable selector quality? 16

17 Chiral separations H 3 C O O H N aspartame (Asm) (Nutrasweet ) N O O CH 3 O O NH 3 + O - CH 3 100% 90% 80% 70% 60% 50% 40% 30% 20% 10% 0% 43% 36% 34% 41% 34% 25% 39% 42% 41% 58% 32% 25% 24% 18% 8% Racemic Racemát Tiszta Enantiopure enantiomer Nem Achiral királis F F F F N N N N NH 2 * H 3 C CH 3 OH O pregabalin (Preg) (Lyrica ) H 3 C N CH 3 O N H F NH 2 O vinpocetin F sitagliptin (Sgli) (Januvia ) dapoxetine (Dpx) (Priligy ) 17

18 Chiral separations Alogliptin + SBECD EOF 5 mm CD Rs=4.67 S R 4 mm CD EOF S R Rs=3.54 Requirements At least for one enantiomer: μ free μ complex For the complexes formed: μ S,cplx μ R,cplx (K S K R ) 3 mm CD 2 mm CD 1 mm CD 0.5 mm CD 0 mm CD S R EOF S S R R EOF EOF EOF EOF Rs=1.69 Rs=1.95 Rs=1.56 Rs=0.41 Rs= t / min + R + S + - I. Fejos et al, Electrophoresis, 35, ,

19 Application fields for chiral separations Different biological activity Chiral analysis of natural compounds Follow-up analytics of enantioselective synthetic routes Control of Active Pharmaceutical Ingredients (APIs) Agrochemistry (herbicides) 19

20 Application fields for chiral separations Cosmetics (fingerprint) Food industry (infections) Environment (biodegradation) Archeology (dating) Space industry (MOD I-II-II) Murchison meteorite 20

21 CD-bound chiral columns Permethrinic acid Naproxen Clopidogrel 0.5% (R) impurity 21

22 CD-bound chiral columns Pros: The selector does not affect detection Possibility for semi-preparative separations Cons: Expensive and very specific columns 22

23 Sensitivity improvement In the CD cavity the analyte microenvironment suffers a significant and temporary change. Enhanced spectral response, fluorescence or phosphorescence by lipophilic environment Protection of the fluorescing/phosphorescing state by the CD cavity from quenchers L Szente and J Szeman, Anal Chem, 85, ,

24 Spectroscopic methods Taguchi K, J Am Chem Soc, 108, ,

25 Sample preparation Aim: to concentrate the compontent of interest to an adequate level How: Selective trapping and removal of the analyte or the interfering matrix using CDs CD enabled cartridges or Selective precipitation L Szente and J Szeman, Anal Chem, 85, , 2013 Improved accuracy and reliability, decreased/eliminated matrix effects 25

26 Sample introduction Sample preparation Analyte enrichment (complexation) Elution (decomplexation) L Szente and J Szeman, Anal Chem, 85, ,

27 Fluorescence detection CD enabled sensors Chromophore or fluorophore tagged CDs The presence of a competitive guest changes the fluorescence spectra Selective to compounds with higher affinity towards the CD cavity than the fluorophore L Szente and J Szeman, Anal Chem, 85, ,

28 Future? applications Diagnostic applications Early diagnosis of Alzheimer disease, even prognosis Determination of the amount of ethane and butane in the expired breath breath biomarkers A network of carbon nanotubes coated with CDs and CD derivatives Enhanced sensitivity for volatile biomarkers in breath Q Ashton Acton, Parkinson s Disease: New Insights for the Healthcare Professionals,

29 Future? applications Diagnostic applications Cyclodextrin based supramolecular biosensors - cancer BCD with linker Adamantylbiotin linker Streptavidin Gold Rhodamine labelled IgG fragment Biotinylated protein Build-up of a bionanostructure Ludden et al, J Am Chem Soc, 130, ,

30 Future? applications Diagnostic applications Cyclodextrin based supramolecular biosensors - cancer Ludden et al, J Am Chem Soc, 130, ,

31 Future? applications DNA sequencing Pore in the Hole Identifying each DNA base by changes in the ion current flowing across the pore Individual human genome analysis Continuous DNA sequencer/reader Analyte recognizing sensor Protein nanopore with covalently tagged cyclodextrin adaptor 31

32 Future? applications DNA sequencing DNA array chip prototype with amino-β-cyclodextrin 550 nucleotide analyzed in 1 minute! 32

33 Future applications 33

34 Thanks for your attention 34