CHIRALITY 4174-177 (1992)
Chromatographic Resolution of the Chiral Isomers of Several P-Blockers Over Cellulose Tris(3,5dimethylphenylcarbamate) Chiral Stationary Phase C.B. CHING, B.G. LIM, E.J.D. LEE, AND S.C. NG Department of Chemical Engineering (C.B.C., B.GL.), Department of Pharmacology (EJBL.), and Department of Chemistty (S.CN.), National Universib of Singapore, Singapore
ABSTRACT The optical resolution of seven 0-blockers which have in common the Nisopropyl-3-aryloxy-2-hydroxypropylamine moiety was carried out by HPLC using the cellulose tris(3,5-dimethylphenylcarbamate)chiral stationary phase to quantitatively characterize the enantioselectivity of these compounds. The capacity factors and separation factors at different column temperature were determined with some qualitative trends derived. A compensation effect was observed for these compounds where there exists an approximately linear relationship between the enantiomeric differences in enthalpic and entropic energies. o 1992 Wiley-Liss, Inc.
KEY WORDS: chiral resolution, 0-blockers, HPLC, cellulose tris(3,5-dimethylphenylcarbamate) chiral stationary phase INTRODUCTION 0-Adrenergicblocking agents @blockers) have considerable utility in the management of various cardiovascular disorders, such as hypertension, angina pectoris, and cardiac arrhythmias. Despite the fact that the P-blocking properties of these drugs are found only in the levorotatory enantiomers, most of the P-blockers are used chemically as the racemate.2 This is mainly due to the technical difficulties involved in preparing pure enantiomers of racemic compounds. Recently, the enantiomers of five P-blockers were successfully resolved using cellulose tris(3,5-dimethylphenylcarbamate) absorbed on macroporous silica gel as the chiral stationary phase (CSP).3 This CSP had also been used for the HPLC assays of P-blockers in human plasma and ~ r i n e .However, ~?~ to date, there appears to have been no detailed study of the separation mechanism of the P-blockers on this derivatized cellulose liquid chromatographic column. In our present study, seven 0-blockers (acebutolol, alprenolol, atenolol, metoprolol, oxprenolol, pindolol, and propranolol) which have in common the N-isopropyl-3-aryloxy-2-hydroxypropylamine moiety, were used in an attempt to quantitatively characterize the enantioselective resolution of these compounds. The structures of these compounds are shown in Table 1. THEORY Capacity Factor and Separation Factor For chromatographic separation of two enantiomeric compounds, the separation factor (u),which is a measure of relative peak separation can be expressed as the ratio of the capacity factors (k) of the enantiomers. If the capacity factors of each enantiomer are denoted by kl and ri, for the first and second eluting isomers respectively, = @
1992 Wiley-Liss, Inc.
b/k,
(1).
The capacity factor can be calculated from the following expression:
k = - - r - TO - r / u - ro/u
(2)
To I U
TO
where TO is the mean retention time for an unretained compound (i.e., a compound which penetrates the macropore space but is not actually adsorbed) and u is the superficial velocity for the elution. The mean retention time (T or first moment) for each enantiomericpeak (1and 2) and the unretained compound can be determined by integrating the peak numerically, ft'
JO
ct dt
f"
ct dt
Jt,
f"
ct dt
JO
where tl is any time after the emergence of the first peak and before the emergence of the second peak. For compounds with no baseline separation, r1 and r2 can be estimated from the time for the peaks to reach their highest values. The Gibbs free energy may be related to the separation factor, AGO = - R T l n k
(4). Consequently, the free energy difference between the two eluting peaks will be 6(AG") = AGj - AG = -RT In (b/kl) = -RT In (a) (5). Received for publication August 13, 1991;accepted October 25, 1991. Address reprint requests to C.B. Ching, Department of Chemical Engineering, National University of Singapore, 10 Kent Ridge Crescent, Singapore 0511.
175
CHROMATOGRAPHIC RESOLUTION OF P-BLOCKERS
TABLE 1. Structures of the seven P-blockers General structure: R-CLC%-CH(OH~14-NH-CH(CHJ2 Compound Structure of R CO-CH3
Acebutolol H3C-CH2-CH2-CO-NH-
Atenolol Metoprolol
H2N-CO-CH3-
0-
H3C-O-CH2-CH2-
0 0
-
Instrumentation The solvent delivery system was a high-pressure liquid chromatographic pump (Shimadzu Model LC-9A) and sample injection was performed using a Rheodyne Model 7125 syringe loading valve fitted with a 10 pl sample loop. The analytical Chiralcel O D 0 column (250 x 4.6mm i.d.) was a CSP containing cellulose tris(3,5-dimethylphenylcarbamate)polymer absorbed on 10-pm macroporous silica (Daicel Chemical Industries, Ltd., Japan) (see Fig. 1).The mobile phase used for the optical resolution was a mixture of hexane, isopropanol, and diethylamine (80:20:0.1). The temperature of the column was kept constant by using a column water jacket with water circulated through the jacket from an external thermostatic bath. The eluting enantiomers were monitored by a spectrofluorometer (Varian Model 2070) with appropriate excitation and emission wavelengths selected on the basis of the maximum absorbance in the presence of the mobile phase. The milivolt signal from the spectrofluorometerwas digitized with the aid of a data acquisition/controlunit which was interfaced with a microcomputer for data storage and processing. A polarimeter (ACS Chiralmonitor Model 750/25), which was in series with the spectrofluorometer, was used to indicate the eluting sequence of the enantiomers. For propranolol, the sequence was further confirmed by chromatography of its enantiomers.
RESULTS AND DISCUSSION The capacity factors, k, and k- , for the ( + ) - and ( - )-isomers, respectively,of the seven P-blockers,and their separation Propranolol factors, CL, were calculated based on Eqs. (1)and (2), and are shown in Table 2. The capacity factors and separation factors for some of these compounds,reported previously by Okamoto et al. (Table 2), are in general consistent with the present data, with the exception of the data for pindolol for which the separation factor obtained in the present study is much larger. Cellulose tris(3,5-dimethylphenylcarbamate) exists as polyAn expansion of Eqs. (4)and (5) to involve the enthalpy (H) meric chains of derivatized D-( +)-glucose residues in p-1,4linkand entropy (S)terms yields ages. These chains lie side by side in bundles and are twisted together to form rope like helical structures into which the (6) enantiomer of interest partitions and interacts stereospecificalIn(k) = - W / R T + AS"/R or ly. It has been postulated8 that these chiral cavities or ravines of the stationary phase have a high affinity for aromatic (7). In(a) = - G ( W ) / R T+ G(AS")/R groups, and for compounds containing such groups, it is that The enthalpic and entropic energies for the binding of each part of the molecule that will enter the cavity. For separation enantiomer of the seven P-blockers to the chiral stationary phase can be obtained by solving Eq. (6).Similarly, the enthalpic and entropic energies difference between the two enantiomers can be obtained by solving Eq. (7). RO
EXPERIMENTAL Reagents and Chemicals Racemic atenolol and propranolol were gifts from ICI (UK), and metoprolol from Hassle (Sweden). Racemic alprenolol, acebutolol, oxprenolol, and pindolol were purchased from Sigma Chemical Co. (USA).Propranolol enantiomers were gifts from ICI (UK). 1,3,5-Tri-tert-butyl-benzene was purchased from Aldrich Chemical Co. (USA). Hexane and isopropanol (HF'LC grade) were purchased from Fisher Scientific (USA) and diethylamine (AR grade) was purchased from BDH ChemicalsLtd. (UK).
Fig. 1. Structure of cellulose tris(3,5-dimethylphenylcarbamate)CSP (Chiralcel OD).
176
CHING ET AL.
TABLE 2. Capacity factors and separation factors of the seven p-blockersat different temperatures 283K
290K
297 K
304K
310 K
1.54 1.76 1.14
1.46 1.63 1.11
1.33 1.49 1.12
1.22 1.38 1.13
1.20 1.35 1.13
0.35 1.67 4.79
0.38 1.69 4.39
0.33 1.25 3.83
0.31 1.10 3.50
0.30 0.94 3.16
3.87b/2.27c
4.34 8.45 1.94
3.72 7.09 1.91
3.69 6.42 1.74
3.14 5.75 1.83
2.97 5.31 1.79
1.58
0.56 2.11 3.77
0.55 1.97 3.57
0.54 1.60 2.95
0.50 1.43 2.86
0.49 1.36 2.75
1.40 12.90 9.19
1.21 9.540 7.89
0.98 6.16 6.30
0.86 4.93 5.72
0.77 3.88 5.03
3.75 54.29 14.49
3.52 46.53 13.23
2.95 35.87 12.17
3.03 28.14 9.29
2.74 23.21 8.46
3.17
1.69 4.75 2.81
1.54 3.70 2.40
1.57 3.37 2.15
1.35 2.76 2.05
1.38 2.81 2.04
1.43
298 Ka
Acebutolol k+
kU
Alprenolol k+ kU
Atenolol k+ kU
Metoprolol k+
Q U
Oxprenolol k+ kU
6.03
Pindolol k+ kU
Propranolol k+ kU
5.07
2.29
“Reported capacity factors and separation factors by Okamoto et al? The values were obtained using Daicel ODTHcolumn (250 x 0.46 an i.d.) and the mobile phase was hexane-isopropanol-diethylamine (80:20:0.1).Column temperature was 25’C for all cases. *Mobile phase was hexane-isopropanol (9O:lO). ‘Column was 50 x 2 an i.d.
to OcCuT, the “fit” of the aromatic moiety in the cavity needs to be reasonably tight but also at least one of the substituents on the chiral center needs to be able to interact with the steric environment just outside the cavity. According to Okamoto et al., the hydrogen bonding between the hydroxyl group of the P-blockers and the carbonyl group of the CSP seems to play the most important role for effective chiral recognition. It was not possible, from the values of K and a of the seven p-blockersobtained in the present study, to define a clear structure-resolvability relationship for these compounds. However, the following qualitative trends were observed: 1. The presence of polar amido-functionality in the case of atenolol increases the retention of both enantiomers, although this does not necessarily enhance the separation factor since atenolol has one of the lowest separation factor of the various compounds studied. 2. The presence of relatively nonpolar side chain substituted in either an ortho or para position within the phenylene moiety seems to enhance the separation of enantiomers. Moreover, this enhancement seemed particularly marked when there is an ether linkage within this side chain, as in the case of oxprenolol. 3. The presence of the indoline moiety in the case of pindolol
evidently gives rise to a marked enhancement in enantiomeric separation in contrast to propranolol which has a naphthalene moiety. 4. The presence of a bulky ketonic functionalityin acebutolol may make it difficult for the enantiomers to fit into the cavity of the CSP accounting for the poor enantiomeric resolvability. 5. The separation factors for compounds with substituents at the ortho position are larger compared with compounds that have substituents at the para position. This could be due to steric hindrance introduced by the long chain attached at the para position. There are large differences in the enantioselectivity of the columns for different compounds but in all cases the (-)-isomer is the more strongly retained species and has the larger heat of adsorption. There is an approximately linear relationship between the 6 ( M )and S(ASo)values for all compounds, as may be seen from Figure 2. Such behavior is not uncommon in both adsorption and reaction systems and is referred to as a “compensation effect.”g This may be understood from general thermodynamic considerations. In the adsorbed state the more strongly held species (larger - AH) will in general have less freedom of movement, and therefore a lower entropy than the less strongly bound species. That AHo and A.S” [and therefore 6 ( W ) and S(ASo)] should be directly
177
CHROMATOGRAPHIC RESOLUTION OF P-BLOCKERS 50
40
Oxpranolol
/
n
Y I
0
30
C
E
\
7
20 n 0
v)
a
m
NOTATION solute concentration (g m -3) capacity factor k universal constant (8.314 J mol-’ K - *) R T temperature (K) elution time (sec) t superficial velocity = volumetric flow rate/column U cross-sectional area (ms AG” free energy change (J mol A€P- enthalpy change (J mol -I) A 9 entropy change 0 mol - K - ’) U separation factor mean retention time (sec) ‘5
Alprenolol /in:otoi
x
w
“lock and key” relationship between the isomers and the chiral stationary phase, a more extensive study is required to provide comparative data, not only between different sorbates on a single stationary phase but also between the behavior of different structurally related stationary phases, has to be undertaken.
to
I
0
LITERATURE CITED
kebutolol
-10
8
4
- b (AHo)
12
16
(kJ/kmol)
Fig. 2. Compensation plot showing approximately linear relationship between 8 ( A W ) and 6(AS’) for all compounds.
correlated is therefore not surprising, but the commonly observed linearity of such correlations is not explained by this simple argument. CONCLUSION For j3-blockers, pindolol and oxprenolol, the selectivity between the (+)- and (-)-isomers is remarkably high and it appears that a large scale chromatographic separation of these compounds over the cellulose tris(3,5-dimethylphenylcarbamate) chiral stationary phase should be practically feasible, although other factors such as the saturation capacity would have to be considered. The selectivity for acebutolol is substantially lower, probably not sufficient for large scale practical resolution and the potential of other alternative stationary phases would have to be explored. In order to understand the
1. Princhard. B.N.C., Owens, C.W.I., Tuckman, J., eds. Clinical features of adrenergic agonists and antagonists. In: Adrenergic Activators and Inhibitors-Handbook of Experimental Pharmacology, Vol 54, Pt 11. Berlin: Springer-Verlag, 1980: 559497. 2. Lee, E.J.D., William, W.H. Chirality: Clinical pharmacokinetics and pharmaccdynamic considerations. Clin. Phannacokinet. 18(5):33%345, 19%. 3. Okamoto, Y., Kawashima, M., Aburatani, R., Hatada, K.. Nishiyama, T., Masuda, M. Optical resolution of P-blockers by HPLC on cellulose triphenylcarbamate derivatives. Chem. Lett. 1237-1240,1986. 4. Hartmann, C., Krauss, D., Spahn, H., Mutschler, E. Simultaneous determination of (R)-and (S)-celiprolol in human plasma and urine: High-performance liquid chromatographic assay on a chiral stationary phase with fluorimetric detection. J. Chromatogr. 496.387-396, 1989. 5. Rutledge, D.R., Garrick, C. Rapid high-performance liquid chromatographic method for the measurement of the enantiomers of metoprolol in serum using a chiral stationary phase. J. Chromatogr. 497181-190, 1989. 6. Koller, H., Rimb&k, K.H., Mannschreck, A. High pressure liquid chromatography on triacetylcellulose:Characterization of a sorbent for the separation of enantiomers. J. Chromatogr. 28289-94, 1983. 7. Hesse, G., Hagel, R. Complete separation of a racemic mixture by elution chromatography on cellulose triacetate. Chromatographia 6(6):277-280, 1973. 8. Francotte, E., Romain, M.. Lohmann, D., Muller. R. Chromatographic resolution of racemates on chiral stationary phases: I. Influence of the s u pramolecular structure of cellulose triacetate. J. Chromatogr. 347:2537, 1985. 9. Cremer, E. The compensation effect in heterogeneous catalysis. Adv. Catal. 775-91, 1955.
Chromatographic Resolution of the Chiral Isomers of Several P-Blockers Over Cellulose Tris(3,5dimethylphenylcarbamate) Chiral Stationary Phase C.B. CHING, B.G. LIM, E.J.D. LEE, AND S.C. NG Department of Chemical Engineering (C.B.C., B.GL.), Department of Pharmacology (EJBL.), and Department of Chemistty (S.CN.), National Universib of Singapore, Singapore
ABSTRACT The optical resolution of seven 0-blockers which have in common the Nisopropyl-3-aryloxy-2-hydroxypropylamine moiety was carried out by HPLC using the cellulose tris(3,5-dimethylphenylcarbamate)chiral stationary phase to quantitatively characterize the enantioselectivity of these compounds. The capacity factors and separation factors at different column temperature were determined with some qualitative trends derived. A compensation effect was observed for these compounds where there exists an approximately linear relationship between the enantiomeric differences in enthalpic and entropic energies. o 1992 Wiley-Liss, Inc.
KEY WORDS: chiral resolution, 0-blockers, HPLC, cellulose tris(3,5-dimethylphenylcarbamate) chiral stationary phase INTRODUCTION 0-Adrenergicblocking agents @blockers) have considerable utility in the management of various cardiovascular disorders, such as hypertension, angina pectoris, and cardiac arrhythmias. Despite the fact that the P-blocking properties of these drugs are found only in the levorotatory enantiomers, most of the P-blockers are used chemically as the racemate.2 This is mainly due to the technical difficulties involved in preparing pure enantiomers of racemic compounds. Recently, the enantiomers of five P-blockers were successfully resolved using cellulose tris(3,5-dimethylphenylcarbamate) absorbed on macroporous silica gel as the chiral stationary phase (CSP).3 This CSP had also been used for the HPLC assays of P-blockers in human plasma and ~ r i n e .However, ~?~ to date, there appears to have been no detailed study of the separation mechanism of the P-blockers on this derivatized cellulose liquid chromatographic column. In our present study, seven 0-blockers (acebutolol, alprenolol, atenolol, metoprolol, oxprenolol, pindolol, and propranolol) which have in common the N-isopropyl-3-aryloxy-2-hydroxypropylamine moiety, were used in an attempt to quantitatively characterize the enantioselective resolution of these compounds. The structures of these compounds are shown in Table 1. THEORY Capacity Factor and Separation Factor For chromatographic separation of two enantiomeric compounds, the separation factor (u),which is a measure of relative peak separation can be expressed as the ratio of the capacity factors (k) of the enantiomers. If the capacity factors of each enantiomer are denoted by kl and ri, for the first and second eluting isomers respectively, = @
1992 Wiley-Liss, Inc.
b/k,
(1).
The capacity factor can be calculated from the following expression:
k = - - r - TO - r / u - ro/u
(2)
To I U
TO
where TO is the mean retention time for an unretained compound (i.e., a compound which penetrates the macropore space but is not actually adsorbed) and u is the superficial velocity for the elution. The mean retention time (T or first moment) for each enantiomericpeak (1and 2) and the unretained compound can be determined by integrating the peak numerically, ft'
JO
ct dt
f"
ct dt
Jt,
f"
ct dt
JO
where tl is any time after the emergence of the first peak and before the emergence of the second peak. For compounds with no baseline separation, r1 and r2 can be estimated from the time for the peaks to reach their highest values. The Gibbs free energy may be related to the separation factor, AGO = - R T l n k
(4). Consequently, the free energy difference between the two eluting peaks will be 6(AG") = AGj - AG = -RT In (b/kl) = -RT In (a) (5). Received for publication August 13, 1991;accepted October 25, 1991. Address reprint requests to C.B. Ching, Department of Chemical Engineering, National University of Singapore, 10 Kent Ridge Crescent, Singapore 0511.
175
CHROMATOGRAPHIC RESOLUTION OF P-BLOCKERS
TABLE 1. Structures of the seven P-blockers General structure: R-CLC%-CH(OH~14-NH-CH(CHJ2 Compound Structure of R CO-CH3
Acebutolol H3C-CH2-CH2-CO-NH-
Atenolol Metoprolol
H2N-CO-CH3-
0-
H3C-O-CH2-CH2-
0 0
-
Instrumentation The solvent delivery system was a high-pressure liquid chromatographic pump (Shimadzu Model LC-9A) and sample injection was performed using a Rheodyne Model 7125 syringe loading valve fitted with a 10 pl sample loop. The analytical Chiralcel O D 0 column (250 x 4.6mm i.d.) was a CSP containing cellulose tris(3,5-dimethylphenylcarbamate)polymer absorbed on 10-pm macroporous silica (Daicel Chemical Industries, Ltd., Japan) (see Fig. 1).The mobile phase used for the optical resolution was a mixture of hexane, isopropanol, and diethylamine (80:20:0.1). The temperature of the column was kept constant by using a column water jacket with water circulated through the jacket from an external thermostatic bath. The eluting enantiomers were monitored by a spectrofluorometer (Varian Model 2070) with appropriate excitation and emission wavelengths selected on the basis of the maximum absorbance in the presence of the mobile phase. The milivolt signal from the spectrofluorometerwas digitized with the aid of a data acquisition/controlunit which was interfaced with a microcomputer for data storage and processing. A polarimeter (ACS Chiralmonitor Model 750/25), which was in series with the spectrofluorometer, was used to indicate the eluting sequence of the enantiomers. For propranolol, the sequence was further confirmed by chromatography of its enantiomers.
RESULTS AND DISCUSSION The capacity factors, k, and k- , for the ( + ) - and ( - )-isomers, respectively,of the seven P-blockers,and their separation Propranolol factors, CL, were calculated based on Eqs. (1)and (2), and are shown in Table 2. The capacity factors and separation factors for some of these compounds,reported previously by Okamoto et al. (Table 2), are in general consistent with the present data, with the exception of the data for pindolol for which the separation factor obtained in the present study is much larger. Cellulose tris(3,5-dimethylphenylcarbamate) exists as polyAn expansion of Eqs. (4)and (5) to involve the enthalpy (H) meric chains of derivatized D-( +)-glucose residues in p-1,4linkand entropy (S)terms yields ages. These chains lie side by side in bundles and are twisted together to form rope like helical structures into which the (6) enantiomer of interest partitions and interacts stereospecificalIn(k) = - W / R T + AS"/R or ly. It has been postulated8 that these chiral cavities or ravines of the stationary phase have a high affinity for aromatic (7). In(a) = - G ( W ) / R T+ G(AS")/R groups, and for compounds containing such groups, it is that The enthalpic and entropic energies for the binding of each part of the molecule that will enter the cavity. For separation enantiomer of the seven P-blockers to the chiral stationary phase can be obtained by solving Eq. (6).Similarly, the enthalpic and entropic energies difference between the two enantiomers can be obtained by solving Eq. (7). RO
EXPERIMENTAL Reagents and Chemicals Racemic atenolol and propranolol were gifts from ICI (UK), and metoprolol from Hassle (Sweden). Racemic alprenolol, acebutolol, oxprenolol, and pindolol were purchased from Sigma Chemical Co. (USA).Propranolol enantiomers were gifts from ICI (UK). 1,3,5-Tri-tert-butyl-benzene was purchased from Aldrich Chemical Co. (USA). Hexane and isopropanol (HF'LC grade) were purchased from Fisher Scientific (USA) and diethylamine (AR grade) was purchased from BDH ChemicalsLtd. (UK).
Fig. 1. Structure of cellulose tris(3,5-dimethylphenylcarbamate)CSP (Chiralcel OD).
176
CHING ET AL.
TABLE 2. Capacity factors and separation factors of the seven p-blockersat different temperatures 283K
290K
297 K
304K
310 K
1.54 1.76 1.14
1.46 1.63 1.11
1.33 1.49 1.12
1.22 1.38 1.13
1.20 1.35 1.13
0.35 1.67 4.79
0.38 1.69 4.39
0.33 1.25 3.83
0.31 1.10 3.50
0.30 0.94 3.16
3.87b/2.27c
4.34 8.45 1.94
3.72 7.09 1.91
3.69 6.42 1.74
3.14 5.75 1.83
2.97 5.31 1.79
1.58
0.56 2.11 3.77
0.55 1.97 3.57
0.54 1.60 2.95
0.50 1.43 2.86
0.49 1.36 2.75
1.40 12.90 9.19
1.21 9.540 7.89
0.98 6.16 6.30
0.86 4.93 5.72
0.77 3.88 5.03
3.75 54.29 14.49
3.52 46.53 13.23
2.95 35.87 12.17
3.03 28.14 9.29
2.74 23.21 8.46
3.17
1.69 4.75 2.81
1.54 3.70 2.40
1.57 3.37 2.15
1.35 2.76 2.05
1.38 2.81 2.04
1.43
298 Ka
Acebutolol k+
kU
Alprenolol k+ kU
Atenolol k+ kU
Metoprolol k+
Q U
Oxprenolol k+ kU
6.03
Pindolol k+ kU
Propranolol k+ kU
5.07
2.29
“Reported capacity factors and separation factors by Okamoto et al? The values were obtained using Daicel ODTHcolumn (250 x 0.46 an i.d.) and the mobile phase was hexane-isopropanol-diethylamine (80:20:0.1).Column temperature was 25’C for all cases. *Mobile phase was hexane-isopropanol (9O:lO). ‘Column was 50 x 2 an i.d.
to OcCuT, the “fit” of the aromatic moiety in the cavity needs to be reasonably tight but also at least one of the substituents on the chiral center needs to be able to interact with the steric environment just outside the cavity. According to Okamoto et al., the hydrogen bonding between the hydroxyl group of the P-blockers and the carbonyl group of the CSP seems to play the most important role for effective chiral recognition. It was not possible, from the values of K and a of the seven p-blockersobtained in the present study, to define a clear structure-resolvability relationship for these compounds. However, the following qualitative trends were observed: 1. The presence of polar amido-functionality in the case of atenolol increases the retention of both enantiomers, although this does not necessarily enhance the separation factor since atenolol has one of the lowest separation factor of the various compounds studied. 2. The presence of relatively nonpolar side chain substituted in either an ortho or para position within the phenylene moiety seems to enhance the separation of enantiomers. Moreover, this enhancement seemed particularly marked when there is an ether linkage within this side chain, as in the case of oxprenolol. 3. The presence of the indoline moiety in the case of pindolol
evidently gives rise to a marked enhancement in enantiomeric separation in contrast to propranolol which has a naphthalene moiety. 4. The presence of a bulky ketonic functionalityin acebutolol may make it difficult for the enantiomers to fit into the cavity of the CSP accounting for the poor enantiomeric resolvability. 5. The separation factors for compounds with substituents at the ortho position are larger compared with compounds that have substituents at the para position. This could be due to steric hindrance introduced by the long chain attached at the para position. There are large differences in the enantioselectivity of the columns for different compounds but in all cases the (-)-isomer is the more strongly retained species and has the larger heat of adsorption. There is an approximately linear relationship between the 6 ( M )and S(ASo)values for all compounds, as may be seen from Figure 2. Such behavior is not uncommon in both adsorption and reaction systems and is referred to as a “compensation effect.”g This may be understood from general thermodynamic considerations. In the adsorbed state the more strongly held species (larger - AH) will in general have less freedom of movement, and therefore a lower entropy than the less strongly bound species. That AHo and A.S” [and therefore 6 ( W ) and S(ASo)] should be directly
177
CHROMATOGRAPHIC RESOLUTION OF P-BLOCKERS 50
40
Oxpranolol
/
n
Y I
0
30
C
E
\
7
20 n 0
v)
a
m
NOTATION solute concentration (g m -3) capacity factor k universal constant (8.314 J mol-’ K - *) R T temperature (K) elution time (sec) t superficial velocity = volumetric flow rate/column U cross-sectional area (ms AG” free energy change (J mol A€P- enthalpy change (J mol -I) A 9 entropy change 0 mol - K - ’) U separation factor mean retention time (sec) ‘5
Alprenolol /in:otoi
x
w
“lock and key” relationship between the isomers and the chiral stationary phase, a more extensive study is required to provide comparative data, not only between different sorbates on a single stationary phase but also between the behavior of different structurally related stationary phases, has to be undertaken.
to
I
0
LITERATURE CITED
kebutolol
-10
8
4
- b (AHo)
12
16
(kJ/kmol)
Fig. 2. Compensation plot showing approximately linear relationship between 8 ( A W ) and 6(AS’) for all compounds.
correlated is therefore not surprising, but the commonly observed linearity of such correlations is not explained by this simple argument. CONCLUSION For j3-blockers, pindolol and oxprenolol, the selectivity between the (+)- and (-)-isomers is remarkably high and it appears that a large scale chromatographic separation of these compounds over the cellulose tris(3,5-dimethylphenylcarbamate) chiral stationary phase should be practically feasible, although other factors such as the saturation capacity would have to be considered. The selectivity for acebutolol is substantially lower, probably not sufficient for large scale practical resolution and the potential of other alternative stationary phases would have to be explored. In order to understand the
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