extending the scope of enantiomer separation by capillary supercritical fluid chromatography on...

Extending the Scope of Enantiomer Separation by Capillary Supercritical Fluid Chromatography on Immobilized Poly- siloxane-Anchored Permethyl-p-cyclodextrin (Chirasil-Dex) Martin Jung and Volker Schurig* lnstitut fur Organische Chemie der Universitat, Auf der Morgenstelle 18, W-7400 Tubingen, FRG

Key Words: Enantioselective capillary (open-tubular) SFC Polysiloxane-anchored cyclodextrin stationary phase Immobilized Chirasil-Dex Separation of cyclodextrin derivatives by SFC Non-steroidal anti-inflammatory drugs (NSAIDs)

Summary Immobilized polysiloxane-anchored permethyl-fl-cyclodextrin (Chirasil-Dex) with a cyclodextrin content of approximately30 YO by weight, previously employed as a versatile chiral stationary phase for the separation of enantiorners by GC, has been used for the separation of enantiomers by capillary supercritical fluid chromatography (SFC). A considerable number of racemates could be resolved, e.g. aromatic alcohols, amino alcohols (TFA derivatives), and underivatized acids. Many pharmaceutical compounds were among those analyzed, including several NSAlDs (e.g. ibuprofen and ketoprofen), a steroidal drug (norgestrel), a barbiturate (hexobarbital), and others.

Among the racemates resolved were many which cannot be analyzed by GC owing to low volatility or decomposition at elevated temperatures. For two racemates, analysis temperature and mobile phase density were systematically varied to give constant analysis times or capacity factors k. Low temperatures (ca SOOC) yielded the best separation in term of separation factor, a, or resolution, R,, even though higher densities had to be used. In comparison with GC, capillary SFC was able to furnish higher separation factors and similar resolution. The applicability of capillary SFC for the analysis of mixtures of cyclodextrin derivatives, e.g. those used in the synthesis of Chirasil-Dex, was, furthermore, demonstrated.

1 Introduction It is only in recent years that supercritical fluid chromatography (SFC) on chiral stationary phases has been introduced as a tool in the growing field of enantiomer analysis [ 11. Several reviews have been published [2-51. Comparisons of enantiomer separations by LC and subcritical fluid chromatography (SubFC) under otherwise similar conditions have clearly demonstrated the considerably higher speed of analysis achievable by the latter [6, 71; this is attributed to higher diffusion coefficients. Other advantages over LC include compatibility with most common detectors (FID, MS) and higher efficiency, especially if open-tubular columns are used. Compared with GC, SFC offers low analysis temperatures; these are known to enhance enantioselectivity and prevent thermal decomposition and racemization. SFC is, therefore, a technique well-suited to the separation of the enantiomers of non-volatile or thermally labile solutes which cannot easily be analyzed by GC.

There have been only few reports on enantiomer separations by capillary SFC on chiral stationary phases. All describe the use of polysiloxanes containing chiral side chains [ 8-14] or block

copolymers with alternating chiral units and polysiloxane units [ 15, 161. The flexible polysiloxane moiety provides favorable mass transfer and coating properties.

Cyclodextrin derivatives are widely used as chiral selectors for HPLC [17, 181 and GC [19, 201. Macaudiere et al. 16, 211 used commercial HPLC columns packed with silica-bonded P-cyclodextrin (Cyclobond@) in SubFC. In preliminary accounts 122, 231, we recently reported our first results of enantiomer separations by capillary SFC on polysiloxane-anchored permethyl-P-cyclodextrin (Chirasil-Dex), which has proved to be a versatile chiral stationary phase for GC 124, 251. Meanwhile, Markides e t al. [26) have suggested a cyclodextrin-containing block copolymer for the resolution of some racemates by capillary SFC. In this study, the scope of our previous work has been considerably extended.

2 Experimental 2.1 Synthesis

The preparation of permethylated 6-monokis-(oct-7-enyl)-P- cyclodextrin and of Chirasil-Dex by hexachloroplatinic acid (H2PtC16)-catalyzed hydrosilylation has been described in detail elsewhere (stationary phase 4 in reference 27).

2.2 Instrumentation

SFE was performed with a commercial Carlo-Erba SFC 3000 system (Fisons/Carlo Erba, Milan, Italy), equipped with an SFC 300 syringe pump, pneumatic Valco valve injector (0.2 pl sample loop), and FID (operated at 360 "C). The split ratio was approximately 1 : 50. Carbon dioxide (99.9995 %, Messer Griesheim, Dusseldorf, FRG) was used a s mobile phase at an average linear velocity of approximately 2.0-2.7 cm s-l(60 "C, 15 MPa). The flow was controlled with a deactivated frit restrictor (Dionex/Lee Scientific, Salt Lake City, USA). Pump and oven were controlled by means of the Fisons Instruments SFC 300 software package, whereas data acquisition was performed with the "Chromatogra- phy Workstation Baseline 810" (Millipore, Milford, Mass., USA). Gas chromatography was performed as described elsewhere [ 271.

2.3 Column Preparation

Fused silica tubing (50 pm i.d., Chrompack, Middelburg, The Netherlands) was heated at 260 "C for 2 h in a slow stream of hydrogen. The capillary was statically coated at 30 "C, without further deactivation, with a carefully filtered solution (20 mg ml-'1 of Chirasil-Dex in diethyl ether, yielding a film thickness of ca 0.25 pm. Immobilization was accomplished thermally at 190 "C for 20 h

0 1993 Dr. Alfred Huethig Publishers Journal of High Resolution Chromatography 21 5

Extending the Scope of Enantiomer Separation by Capillary SFC on Chirasil-Dex

with a very low flow of hydrogen, a s previously described 127-291. The column was finally rinsed with methanol, then with mixtures of methanol and dichloromethane, and then with pure dichloromethane. In order to prevent clogging, the proportion of dichloromethane was increased only gradually.

Immobilization was monitored by GC (inlet pressure 0.5 MPa hydrogen or 0.8 MPa helium) by measuring the capacity factors of n-dodecane and n-tridecane and the separation factors of 1- phenylethanol and y-nonalactone before and after immobilization and rinsing; the degree of immobilization was ca 85 %. The column efficiency measured by GC was ca 9000-10000 effective plates per meter for n-tridecane and 1-phenylethanol.

2.4 Solutes

All solutes were commercial products (Sigma or Aldrich), the pharmaceutical compounds were kindly provided by Professor Dr G Blaschke, Munster, FRG, and Prof W H Pirkle, Urbana, Ill , USA Methanol and dichloromethane were used as solvents Amines were derivatized with trifluoroacetic acid anhydride for 30 min at ambient temperature

3 Results and Discussion

3.1 Scope

Several different chiral solutes were tested by capillary SFC on immobilized Chirasil-Dex (containing approx. 30 % permethyl- 0-cyclodextrin by weight). The chromatographic conditions and resolution values, R,, are listed in Table 1. (The separation factors, a , are not given under programmed conditions.) Typical chromatograms are shown in Figure 1. Significant improvements in resolution and baseline stability were observed for those solutes which had already been investigated in previous work ( e . g . ibuprofen, cis-permethrinic acid, 1-(1-naphthy1)ethanol) [22]. Two thousand effective plates per meter were measured for 1-phenylethanol (60 "C, 0.2 g ml-l, k = 5.3). No loss in enantioselectivity or column performance was observed even after using the column for SFC for several months.

Some of the chiral solutes described here (e.g. all of the aromatic alcohols) can also be separated into their enantiomers by GC on the same stationary phase in a shorter analysis time (c.f. 125, 271). Our interest was, therefore, focused on solutes which either

Table 1

Chromatographic data for enantiorner separations by open-tubular SFC on a 5 m x 50 pm fused silica column coated with a 0.25 pm film of immobilized Chirasil-Dex (mobile phase, carbon dioxide).

Racemic solute Temp Density program Analysis R,a)

["CI is ml-'l [g ml-l min-l] Iminl min gradient time

l-Phenylethanolb) l-Phenylpropanolb) l-(2-Naphthyl)ethanolb) 1-( 1-Naphthyl)ethanolb) 1,2,3,4-Tetrahydro-l -naphthol ( a-tetralol)b) p-( 1 -Hydroxyethyl)biphenyP' 1 -( 9-Anthryl)-2,2,2-trifluoroethanol Diethyl tartrate Benzyl mandelateb) 10,2-Camphorsultam Ephedrine (TFA) Norephedrine (TFA) Synephrine (TFA) Hexobarbital Ethosuximid Warfarin Norgestrel cis-Permethrinic acid trans-Permethrinic acid Ibuprofen Ketoprofen Cicloprofen Flurbiprofen Carprofen Pirprofen Naproxen Tiaprofenic acid Tropic acid

60 60 60 60 60 60 60 60 60 50 60 60 60 90 60 90 60 50 50 60 60 45 45 90 60 60 60 60

0.35 (10 min) 0.35 (10 min) 0.35 (10 min) 0.35 (10 min) 0.20 (10 min) 0.40 (10 min) 0.40 (10 min) 0.24 ( 8 min) 0.22 (10 min) 0.30 (10 min) 0.32 ( 5 min) 0.32 ( 5 min) 0.32 ( 5 min) 0.31 ( 3 min) 0.21 ( 5 min) 0.32 ( 5 min) 0.28 ( 5 min) 0.24 (10 min) 0.24 (10 min) 0.25 ( 2 min) 0.38 ( 5 min) 0.44 (10 min) 0.38 (10 min) 0.52 (10 min) 0.32 (10 min) 0.30 (10 min) 0.35 (10 min) 0.26 (10 min)

0.015 0.015 0.015 0.015 0.008 0.015 0.006 0.005 0.006 0.008 0.006 0.006 0.008 0.006 0.003 0.005 0.006 0.008 0.008 0.0035 0.004 0.004 0.004 0.008c) 0.005 0.005 0.008 0.010

15 16 23 25 37 20 56 21 48 35 17 19 22 12 30 34 39 45 39 59 63 83 84 46 71 75 61 42

3 3 2 5 3 4 1 8 2 6 3 6 1 9 4 2 1 3 4 7 4 1 3 6 3 7 4 6 1 8 1 6 1 4 2 7 2 7 3 7 1 2 3 7 2 6 3 4 1 4 1 6 1 3 1 1

given by R, = 1 177 ( t R ( 2 ) - t R ( 1 ) ) 1 (WO ~(2) + WG 5 ( l ) ) where tR is retention time and WG 5 the peak width at half height b)6 5 m x 50 pm 1 d column C)final density 0 82 g ml-'

21 6 VOL. 16, APRIL 1993 Journal of High Resolution Chromatography


0

0.35 0:4 0 :5 t I I I

0 10 20 30 min

a) aromatic alcohols

L L I I

0.4 0.5 0:6 g/ml I I I

0.4 0.5 0.6 g/ml I I I

0 10

c) p-( 1-hydoxyethy1)biphenyl

20 min

1 0.3

I 0.4

I 0.5 g/ml

I I I

0 10 20 30 min

(not rac.)

..I L 0.2 013 0.4 g/ml

I I I I 1 0 10 20 30 40 min

b) a-tetralol

CF, - CH - OH

&

t I I I

I I I I

0.4 0.5 0.6 0.7 g/ml

0 20 40 60 min

d) 1 -(9-anthryl)-2,2,2-trifluoroethanol

HoXCooEt no COOEt

0.24 r I 0 10 3

e) 10,2-carnphor sultam 9 diethyl tartrate

li

- 1 0.35 g/ml

0 I I 30 min

Journal of High Resolution Chromatography VOL. 16, APRIL 1993 21 7


I

0.24 0.3 0.4 0.5 g/ml 0.21 0.25 0.29 g/ml

' I 6 ' I0 20 30 40 min I b Ib 2b 30 min

h) ethosuximid g) cis-permethrinic acid

0.28 0.3 0.4 0.5 g/ml r I 0 10 2-0 30 40 min

i) norgestrel

I I 0.25 0.3 0.4 g/ml

6 i0 40 80 min

I ) ibuprofen

y 3 co I

L - 0.32 0.35 0.4 0.45 g/ml

I I I I

0 10 20 30 min

k) warfarin

I I 1 0.7 g/ml 0.38 0.4 0.5 0.6

I I I I I

0 20 40 60 80 min

m) flurbiprofen

218 VOL. 16, APRIL 1993 Journal of High Resolution Chromatography


COOH

I ' I ' I 1 . I 0.35 0.4 0.5 0.6 0.7 0.8 g/ml

6 2b 4b 6b min

n) tiaprofenic acid

0.32 0.35 0.4 0.45 g/ml I 1 I 0 10 20 m i n

p) synephrine (TFA) Figure 1

I I I .

0.52 0.6 0.7 0.8 0.82 g/ml I I I 1

0 10 20 30 40 50 min

0) carprofen

I

0.32 0.35 0.4 g/ml I r r 0 10 20 min

q) ephedrine (TFA) and norephedrine (TFA)

Separations of enantiomers by open-tubular SFC on a 5 m x 50 pm id. fused silica column coated with a 0.25 pm film of immobilized Chirasil-Dex (chromatographic conditions listed in Table 1).

constant capacity factors

Constant analysis times (using the same restrictor)

cH3NTNH hexobarbital

I

Constant capacity factors

COOH

Constant analysis

cicloprofen times (using the same restrictor)

0 10 20 M 40 mm o 10 20 mm

Figure 2

Separations of the enantiomers of hexobarbital and cicloprofen by SFC under different chromatographic conditions which yield the capacity factors (left) or analysis times (right) (cf. Table 2). (Note that cicloprofen decomposes at higher temperatures.)

same

Journal of High Resolution Chromatography VOL. 16, APRIL 1993 21 9

Extendinq the Scope of Enantiomer Separation by Capillary SFC on Chirasil-Dex

cannot be analyzed by GC or require a rather high analysis temperature. Many NSAIDs, such as ibuprofen, ketoprofen, cicloprofen, flurbiprofen, and other chiral drugs could be separated. Many polar analytes such as diethyl tartrate or underivatized acids could be chromatographed with very little peak tailing. Only a few solutes such as underivatized carprofen, which is both an acid and a secondary amine, exhibited some peak tailing. Most amines, e.g. synephrine or ephedrine, had to be derivatized Little or no enantioselectivity was observed for, e.g , many P-blockers, 1,l'-binaphthyl and 2,2'-derivatives thereof, oxazepam, lorazepam, fenoprofen, etololac, thahdomide, testo- sterone, promethazin, and the chiral coordination compound iron tetracarbonyl dimethyl fumarate. Many of the solutes used in this work can also be resolved by capillary electrophoresis on capil- laries coated with Chirasil-Dex [30] (and vice versa); although resolution is often found to be somewhat lower, analysis is faster in CE.

For those solutes (1-phenylethanol, diethyl tartrate, ibuprofen) which were also investigated by Markides et al. 1261 on a cyclodextrin-containing copolymeric stationary phase, the values for the resolution, Rs, measured here under similar chromatographic conditions, are somewhat better, even though their copolymeric phase had a much higher cyclodextrin content (ca 67 % permethyl-p-cyclodextrin by weight). This finding is in agree- ment with our GC results for Chirasil-Dex 1271 which indicated that little extra enantioselectivity was obtained by increasing the cyclodextrin beyond ca 30 %. (In that study the immobilization properties and the long-term stability of Chirasil-Dex were also investigated.) It should be mentioned that the copolymeric phase of Markides e t al. [26] is only suitable for SFC where it is swollen by

the supercritical mobile phase. Also, because cross-linking has not yet been achieved, only volatile compounds could be resolved by SFC on that phase [26]

3.2 Optimization of Chromatographic Conditions and

In a previous systematic study of SFC on Chirasil-Dex (231 we found that chiral separation factors, a , decrease with increasing temperature, a s is common in enantiomer analysis. Separation factors were, on the other hand, found to decrease with increasing mobile phase density. At constant temperature, a is considerably lower in SFC than in GC. Similar results were observed for Chirasil-Val 1311, but not for Chirasil-Metal 1321. It was assumed that this behavior could, in part, be attributed to competition between solute and mobile phase molecules for the selector sites. We further found that the resolution, R,, at constant pressure increases a s the temperature is reduced, but goes through a maximum and finally decreases as the temperature drops below ca 60 "C.

Since low mobile phase density and low analysis temperature both enhance enantioselectivity, but also increase retention, one question which is very important in practice remains to be answered: what is the best way to utilize a given analysis time or capacity factor in order to obtain the best enantiomer separation? In order to find an answer, we are now considering different combinations of temperature and density which yield the same capacity factor, k, or the same analysis time. Two different examples are shown (Figure 2, Table 2): hexobarbital, which can also be analyzed by GC (Figure 3), and cicloprofen, which

Comparison with GC

Table 2

Enantiomer separations of hexobarbital and cicloprofen by SFC under different chromatographic conditions which yield the same capacity factors (left) or analysis times (right) (chromatograms are shown in Figure 2).

T da) Pb) a RS

["CI d P to kc' a R S

Hexobarbital

50 70 90

110 130 150

Cicloprofen

50 70

90 110 130

Constant capacity factors (= 6)

0.300 9.0 1.595 9.1 0.330 11.6 1.476 6.5 0.310 13.2 1.370 5.2

0.275 13.9 1.288 4.9 0.238 13.9 1.196 3.5 0.192 12.7 1.149 3.1

Constant capacity factors (= 8)

0.560 11.9 1.223 1.9 0.500 15.0 1.197 2 .1

0.450 17 3 1166 1.7 0.400 18.7 (1.137) ( l . O ) f )

Constant analysis times ( ~ 1 1 . 6 min)

0.460 10.7 346 1.0 0.390 12.8 196 2.6 0.350 14.4 150 3.5 density programd) 0.285 14.3 110 5.2 0.238 13.9 97 6.0 0.187 12.4 94 6.5

Constant analysis time ( ~ 2 5 min)

0.650 13.5 261 4.7 0.500 15.0 176 7.7 density programe) 0.410 16.1 123 11.1 0.360 17.1 99 14.0 0.308 17.1 82 15.1

1.498 1.437 1.346

1.270 1.200 1.147

1.221 1.197

1.168 (1.139) (-1

2.4 3.9 5.3 4.6 4.4 3.5 2.9

1.6 2.1 2.2 1.8 (1 .3)f) Nf)

a) density [g ml I ]

b, pressure [MPa] c, capacity factor for the second eluted enantiomer dl 0.313 g ml-I for 3 min, then programmed at 0.006 g ml-I min-I

0 46 g ml-I for 5 min, then at 0.006 g ml-I min-' bad peak shape because of solute decomposition during chromatography



Rs=3.4 CL = 1.16

data from Table 2 are displayed in Figure 4.

Figure 3

Separation of the enantiomers of hexobarbital by GC on long and

I

*) constant capacity factors a

short capillary columns coated with Chirasil-Dex: inlet pressure,

l S 6 O l 1.40

In a. 0.4

0.2 t

,*/ / /'

1 >* /'

1'

hexobarbital /* S C / * : i +/-+

/ + 4 c lopr ofen : GC ,,* /

4 '.+ '+.+ cicloprofen

1.20

t '\* hexobarbital ',SC

*\* GC :

:

1.00 50 100 150 TCOCl

0 .o 1 . . 20 25 30 1/

CK-'] c) constant analysis times

(using the same restrictor)

GC

A+"+ cicloprofe>+

I 50 100 150 T[OC]

Figure 4

Plots of the data listed in Table 2.

These results show that low analysis temperatures are favorable in SFC even though higher densities have to be used in order to keep the capacity factors or analysis times constant In other words, the enantioselectivity is more drastically influenced by temperature than by density Somewhat decreasing efficiencies however, render very low temperatures less favorable in terms of the resolution, R, If both capacity factors and analysis times could be kept constant, the maximum resolution for hexobarbital in Figure 4c should be expected around rather than around 90 "C (since the unfavorable, very small capacity factors k a t low temperatures would be avoided) As a result temperatures around 60 "C seem to be optimum in most circumstances

In comparison with GC, capillary SFC furnishes considerably higher enantioselectivity, a , in a given analysis time under typical

Journal of High Resolution Chromatography VOL. 16, APRIL 1993 221


4-

experimental conditions, because of the lower efficiency of SFC, however, the resolution of hexobarbital by SFC (Table 2 , Figure 4) is still somewhat lower than by GC (Figure 3)

3.3 Analysis of Mixtures of Cyclodextrin Derivatives by SFC

Mixtures of cyclodextrin derivatives such as those encountered in the synthesis of Chirasil Dex were analyzed by capillary SFC on Chirasil-Dex (Figures 5 and 6) It is not known whether self-recognition of the cyclodextrins plays a role in this technique Previous analytical methods have included LC (e g [331), high temperature GC at 300-400 "C (experimentally difficult) [33], GC after reductive cleavage of the glycosidic bonds [34], and mass spectrometric techniques (e g (351)

0.49 0.5 I 1 I 5 1 0 5 10 15 min

Figure 5

Separation of permethylated a- , p-, and y-cyclodextrins by SFC on a 5 m x 50 pm i.d. capillary column coated with immobilized Chirasil- Dex: mobile phase, carbon dioxide; temperature, 100 "C; density, 10 min at 0.49 g ml-' then programmed at 0.01 g ml-I min-'.

1 O A octenyl groups

4 Conclusion Although the enantioselectivities observed in this work are rather low in comparison with those commonly observed in chiral HPLC

[36], the above-mentioned advantages of capillary SFC, particu- larly the much higher efficiency (enabling the analysis of complex mixtures) and the high compatibility with FID or MS as detection techniques, render the described method an important alterna tive Future developments might further improve enantioselectivity by adopting elements common in enantiomer separation by HPLC on silica-bonded cyclodextrins, e g the use of cyclodextrin derivatives which bear some underivatized hydroxyl or other polar groups or the use of more polar mobile phases, i e addition of modifiers

Acknowledgment

Support of this work by the Deutsche Forschungsgemeinschaft and Fonds der Chemischen Industrie is gratefully acknowledged

References

111 P.A. Mourier, E. Eliot, M H Caude, and R H Rosset. Anal. Chem. 57 (1985) 2819.

121 P Macaudiere, M. Caude, R . Rosset. and A. Tambute, J Chromatogr. Sci. 27 (1989) 383.

131 M.L. Lee and K.E. Markides (Eds). Analytical Supercritical Fluid Chromatography and Extraction, Chromatography Conferences Inc.. Provo. Utah, USA (1990) 543-553

141 Z. Juvancz and K E Markides, LC-GC Intl 5 (1992) 44

151 M. Schfejrner and V. Schung, in. B. Wenclawiak (Ed.), Analysis with Supercritical Fluids: Extraction and Chromatography, Springer, Ber- lin, FRG (1992) 134-150

161 P. Macaudiere. M Caude. R . Rosset, and A Tambute, J. Chromatogr 405 (1987) 135

I71 P. Macaudiere, M. Caude. R Rosset. and A. Tambute, J. Chromatogr Sci 27 (1989) 583

[81 W Roder, F J Ruffing, G Schomburg, and W.H. Pirkle, HRC & CC 10 (1987) 665.

191 F. J. Ruffing, J A Lux, W. Roder. and G Schomburg, Chromatographia 26 (1988) 19.

[ l o ] J. S. Bradshaw, S. K. Aggarwal, C. J. Rouse, B.J. Tarbet, K. E Markides, and M.L. Lee, J. Chromatogr 405 (1987) 169

1111 G. Lai, G. Nicholson, U. Muhleck, and E. Bayer, J. Chromatogr. 540 (1991) 217.

[12l V. Schurig, D Schmalzing. and M. Schleimer, Angew. Chem. 103 (1991) 994; Angew. Chem. Int. Ed Engl 30 (1991) 987.

[13l R. Brugger, P Krahenbuhi, A Marti, R. Straub. and H. Arm, J. Chromatogr. 557 (1991) 163.

[14] A. Marti, P. Krahenbuhl, R Brugger, and H. Arm, Chimia 45 (1991) 13.

1151 B.E. Rossiter, P. Petersson. D F Johnson, M. Eguchi, J.S. Bradshaw,

1161 P Petersson. K E Markides, D F Johnson, B E. Rossiter. J.S.

1171 S H Han and D W Armstrong, Chapter 10 in reference 36

1181 D. W. Armstrong, Anal Chem 59 (1987) 84A

[19l V. Schurigand H. -P. Nowotny, Angew. Chem. 102 (1990) 969; Angew

[20l W. A. Konig, Enantioselective Gas Chromatography with Modified

[21l P. Macaudiere, M. Caude, R. Rosset. a n d A Tambute, J. Chromatogr.

K.E. Markides, and M.L Lee, Tetrahedron Letters 32 (1991) 3609

Bradshaw, and M L. Lee, J Microcol. Sep. 4 (1992) 155

Chem. Int Ed Engl 29 (1990) 939

Cyclodextrins, Hiithig, Heidelberg, FRG (1992).

450 (1988) 255



V. Schurig, Z Juvancz, G Nicholson, and D Schmalzing, HRC 14 (1991) 14.

D. Schrnalzing, G. J. Nicholson, M. Jung, and V Schurig, J. Microcol. Sep. 4 (1992) 23.

V. Schurig, D. Schmaizing, U. Muhleck, M. Jung, M. Schleimer, P. Mussche, C. Duvekot, and J. C. Buyten, HRC 13 (1990) 713.

D Schmalzing, M. Jung. S Mayer, J. Rickert, and V. Schurig, HRC 15 (1992) 723.

a) P. Petersson, K.E Markides, G. Yi, S.R Reese, B.E. Rossiter. J.S. Bradshaw, and M L. Lee, in. P. Sandra and M.L. Lee (Eds), Proc. 14th Int. Symp Capillary Chromatogr., Baltimore, USA 25-29 May, 1992, Huthig, Heidelberg, FRG (1992) 750; b) J.S. Bradshaw, G. Yi. B.E. Rossiter, S.L. Reese. P. Petersson, K.E Markides, and M.L. Lee, Tetrahedron Lett. 34 (1993) 79.

M Jung and V Schurig, J Microcol. Sep 5 (1993). 11.

1281 D Schrnalzing, Doctoral Thesis, University of Tubingen, FRG (1991)

[291 G. Lai, U. Miihleck, G.J . Nicholson, J. Schmid, and E. Bayer,

1301 S. Mayer and V. Schurig, J Liq Chromatogr 16 (1993) 915.

1311 G. Lai, G. Nicholson. U. Muhleck, and E. Bayer, J Chromatogr 540

I321 M Schleimer and V. Schurig, J Chromatogr 1993, in press

1331 G. Schornburg, A Deege, H Hinrichs, E. Hdbinger, and H Husmann,

[341 G. Wenz. P. Mischnick, R . Krebber. M. Richters, and W.A. Konig, HRC

1351 J. W. Metzger, M. Jung, D Schmalzing. E Bayer, and V Schurig.

1361 A.M. Krstulovic (Ed.) Chiral Separations by HPLC, Ellis Horwood.

Ms received January 29, 1993 Accepted: March 4, 1993

Chromatographia 32 (1991) 241

(1991) 217

HRC 15 (1992) 579

13 (1990) 724.

Carbohydr Res. 222 (1991) 23, and literature cited therein

Chichester, UK (1989)

Journal of High Resolution Chromatography VOL. 16, APRIL 1993 223

extending the scope of enantiomer separation by capillary supercritical fluid chromatography on...

Documents