Federico Marini , Remo Bucci, Ilaria Ginevro & Antonio Magrì

1. Coupling of spectroscopic, thermoanalytic and chemometric techniques for the resolution of enantiomeric mixtures Federico Marini, Remo Bucci, Ilaria Ginevro & Antonio Magrì Dipartimento di Chimica, Università di Roma “La Sapienza”

2. ENANTIOMERS

3. DRUG PAIN ENANTIOMERIC MIXTURE DIFFERENT PHARMACOKINETIC BEHAVIOUR: study of the absorption process, distribution, metabolism and excretion DIFFERENT PHARMACODYNAMICAL BEHAVIOUR: study of the mechanisms of action between the active principle and the receptor

4. ADVANTAGES IN THE USE OF THE ACTIVE ENANTIOMER ONLY Half quantity of active principle Reduction of the metabolic burden on the liver Reduction of the metabolic burden on the kidneys Elimination of the undesirable effects of the inactive form Elimination of the intermediate products of the bio-inversion R→S

5. SINGLE ENANTIOMER DRUGS d-ibuprofen (S)-citalopram

6. Non-steroidic anti-inflammatory Analgesic Antipyretic KETOPROFEN

7. RACEMIC KETOPROFEN

8. d-KETOPROFEN

9. KETOPROFEN (2) • (S) is the active enantiomer (cyclooxigenase, which is the target enzyme is stereo-selective) • (S) is not gastrolesive while (R) is • (S) doesn’t lead to the formation of ulcerae, while (R) is ulcerogenic at the level of both intestines. More pharmaceutical industries are moving towards the use of single enantiomers and therefore it is necessary to device a suitable way of determining enantiomeric excess

10. SCOPE OF THE WORK Study of the possibility of devicing a rapid and cheap method to determine enantiomeric excess in pharmaceutical preparation DSC Tf (racemate) = 95°C Tf (enantiomer) = 78°C IR 3500-2500 cm-1 1700-1300 cm-1

11. DSC Analysis • Particularly suited for the determination of purity of compounds (Van’t Hoff law) • There can be deviations from linearity that need to be corrected. • Some enantiomeric mixtures melt with decomposition so it’s difficult to estimate the melting temperatures (that are needed for the computation)

12. EXPERIMENTAL DESIGN • Check repeatability, reproducibility and effect of some experimental factors: • Way of preparing lab mixtures (directly in mortar or by dissolution and riprecipitation) • DSC scan rate • Open/sealed capsula • Identity of mixtures prepared starting from the single enantiomer plus the racemate and of those prepared mixing the two enantiomers Preliminary study using mandelic acid

13. MANDELIC ACID IR 3500-2500 cm-1 1700-1300 cm-1 DSC Tf (racemate) = 118°C Tf (enantiomer) = 132°C

14. PHASE DIAGRAM Prigogine-Dufay Schroeder-Van Laar

15. SOME CONSIDERATIONS • No difference in preparing the mixtures by directly weighing and mixing in mortar or by dissolution and riprecipitation. • No detectable effect of scan rate • No difference in mixing R+S or Rac+S • Sealed capsula provided better reproducibility • Presence of one eutectic • It’s difficult to measure Tfclose to the composition of the eutectic

16. IR SPECTROSCOPY

17. PLS CALIBRATION • 2 significant components (about 94% Y-variance explained)

18. MCR Better results with three component, probably due to background effects

19. MCR (2)

20. KETOPROFEN - PLS

21. KETOPROFEN – MCR

22. MCR(2)

23. CONCLUSIONS • Coupling of IR spectroscopy and chemometrics seems to provide a rapid and cheap tool to determine enantiomeric excess to an acceptable accuracy • Possibility of improving these results by variable selection (GA, iPLS_GA) • Next step is the validation on real formulates

24. СПАСИБО!