Temperature-Induced Inversion of the Elution Order of Enantiomers in Gas Chromatography:  N-Ethoxycarbonyl Propylamides and N-Trifluoroacetyl Ethyl Esters of α-Amino Acids on Chirasil-Val-C₁₁ and Chirasil-Dex Stationary Phases

Abstract

Inversion of the elution order of enantiomers caused by enthalpy-entropy compensation at the isoenantioselective temperature (Tiso) was experimentally observed by gas chromatography on the diamide-type chiral stationary phase (CSP), Chirasil-L-Val-C11, with N-ethoxycarbonyl propylamide (ECPA) derivatives of a number of alpha-amino acids. For the first time, a clear visual representation of the increase of the apparent enantioseparation factor alpha app from 1.00 to 1.08 as the temperature is raised from 120 to 170 degrees C is presented. The increase of alpha app is accompanied by a concomitant reduction of the retention factors of the enantiomers. The Tiso values were in the range from 110 to 130 degrees C depending on the nature of the alpha-amino acid. On the contrary, the Tiso values of the N(O)-trifluoroacetyl ethyl ester derivatives (TFA-Et) of the same alpha-amino acids were approximately 80 degrees higher than that of ECPA derivatives. The comprehensive thermodynamic investigation of the enantioseparation of ECPA and TFA-Et derivatives of valine and alanine using the retention increment method showed that the Delta L,D(DeltaH) difference between the diastereomeric selector-selectand associates was almost the same for ECPA and TFA-Et derivatives despite a much stronger bonded selector-selectand association taking place for the ECPA derivatives. On the other hand, the Delta L,D(DeltaS) values were found to be more negative in the case of ECPA derivatives, resulting in the unusually low values of Tiso. A temperature-dependent inversion of the elution order of enantiomers was also observed on the cyclodextrin-type CSP, Chirasil-Dex, with TFA-Et derivatives of several alpha-amino acids. The Tiso values were in the range from 20 to 170 degrees C depending on the nature of the alpha-amino acid. The results obtained demonstrate the necessity to conduct temperature-dependent studies in order to optimize the enantiomeric separation of single racemates isothermally or of mixtures of racemates in temperature-programmed runs using enantioselective GC. It is also shown that consideration of the elution order of enantiomers and the value of the apparent enantioseparation factor alpha app alone, without performing temperature-dependent measurements, can easily lead to wrong conclusions regarding the enantiorecognition mechanism.

Salient Features of Enantioselective Gas Chromatography: The Enantiomeric Differentiation of Chiral Inhalation Anesthetics as a Representative Methodological Case in Point

“…Below T iso enantioseparation is governed by the predominant enthalpic contribution to enantiorecognition, whereas above T iso it is governed by the predominant entropic contribution (ΔS D < ΔS L ) to enantiorecognition with the stronger bonded enantiomer D (ÀΔH D > ÀΔH L ) bizzarrely eluted as the first peak. Whereas the sign of the enantioselectivity changes at T iso , the association constants K The existence of an isoenantioselective temperature T iso in enantioselective GC has been observed independently in hydrogen-bonding equilibria [111,112] and in metal complexation equilibria [113,114] whereby the intriguing peak reversal for the enantiomers below and above T iso was clearly evident in the gas-chromatograms. A report of a peak reversal for the enantiomers of methyl lactate on a modified cyclodextrin selector (Lipodex E) [115] could not be ascertained [112] while a temperature-induced reversal of the elution order has been observed for the enantiomers of N-trifluoroacetyl-α-amino acid ethyl esters on the selector Chirasil-Dex [112].…”

“…Whereas the sign of the enantioselectivity changes at T iso , the association constants K The existence of an isoenantioselective temperature T iso in enantioselective GC has been observed independently in hydrogen-bonding equilibria [111,112] and in metal complexation equilibria [113,114] whereby the intriguing peak reversal for the enantiomers below and above T iso was clearly evident in the gas-chromatograms. A report of a peak reversal for the enantiomers of methyl lactate on a modified cyclodextrin selector (Lipodex E) [115] could not be ascertained [112] while a temperature-induced reversal of the elution order has been observed for the enantiomers of N-trifluoroacetyl-α-amino acid ethyl esters on the selector Chirasil-Dex [112]. Thus the derivatives of valine and leucine showed a peak reversal on Chirasil-Dex below and above T iso ¼ 70 and only a single peak was observed at the coalescence temperature.…”

“…Thus the derivatives of valine and leucine showed a peak reversal on Chirasil-Dex below and above T iso ¼ 70 and only a single peak was observed at the coalescence temperature. For the isoleucine derivative the isoenantioselective temperature was as low as T iso ¼ 30 [112]. A different enthalpy/entropy compensation approach based on the plot of ln R 0 vs ΔH for the second eluted enantiomer has also been described [105].…”

“…The use of (9) were used according to (10) in which the adjusted retention time t 0 R (¼ total retention time t R minus unretained-peak holdup time t M ) of the enantiomers was related to an inert reference standard. For α-amino-acid selectors and metalcomplex selectors, n-alkanes have been used as reference standards as they undergo neither hydrogen-bonding nor complexation with transition metal ions [112,114]. However, in the case of cyclodextrin selectors, minor molecular association with n-alkanes has been detected [24].…”

“…A theoretical treatment of the retention-increment R 0 approach has been put forward for n-alkane reference standards which undergo a definite, albeit negligible, interaction with modified cyclodextrins [24]. Indeed, n-alkanes could previously not be used as reliable reference standards in a thermodynamic study of derivatized amino acids and modified cyclodextrins [112]. However, due to the very large retention factors k involved for "compound B" and Lipodex E, the use of n-alkanes as references standards was straightforward [134].…”