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Paper electrophoretic study of ion-pair formation
Journal of Chromatography,
403 (1987) 388-391
Elsevier Science Publishers B.V., Amsterdam -
Printed in The Netherlands
CHROM. 19 684
Note
Paper electrophoretic
study
XVI*. Resolution of optical amino acid ligands
of ion-pair isomers
formation
of some cobalt(lll)
complexes
with
S. FANALI*, P. MASIA and L. OSSICINI Istituto di Cromatograjia del C.N.R., Rome (Italy)
Area della Ricerca di Roma, C.P. IO, 00016 Monterotondo
Scala,
(Received May 7th, 1987)
Ion-pair or outer-sphere complex formation is widely used in chromatography to achieve good separations of various compounds. Many years ago in our laboratory we studied ion-pair formation by electrophoretic techniques and succeeded in developing a good method for the separation of several metal complexes and racemic mixture&’ In this paper we report the electrophoretic study of some racemic cobalt(III) complexes with mixed ligands such as [Co(en),Aa12+, where Aa is an amino acid, using as counter ions aqueous solutions of sodium (+)- or (-)-tartrate, antimony1 ( + )-tartrate and arsenyl ( + )-tartrate. EXPERIMENTAL
A high-voltage paper electrophoresis apparatus (HVE Camag system) and Whatman No. 1 paper were used. The time required for the experiments varied from 15 to 35 min and the voltage from 2000 to 3500 V; the electric field applied was generally 50 V/cm. The cooling plate was kept at 68°C. Evaporation from the paper strips during the run was negligible. The electroosmotic flow in the HVE was measured by observing the movement of hydrogen peroxide; the results given are distances already corrected for electroosmotic flow. The samples were prepared as described below. Racemic mixtures were placed side by side with the two resolved isomers in the electropherograms. For the detection of the spots aqueous ammonium sulphide or Dragendorff reagent was used. A Camag TLC-HPTLC scanner was used for adsorption measurements directly on the electropherograms. Preparation of samples
Literature methods were used for the synthesis of racemic cobalt(II1) complexes*. Resolution of the optical isomers was achieved using a cationic-exchange column (Sephadex SP, C24; sodium (+ )-tartrate (0.18 M) was used as the eluent for * For Part XV, see J. Chromatogr., 318 (1985) 440. 0021-9673/87/$03.50
0
1987 Elsevier Science Publishers B.V.
3x9
NOTES (a)
o”o 00
0
0
-- 1
0+
.-
.- 3 2
-- 4
I-,
00
t
1
complexes, where Aa = (1) D-alanine, (2) LFig. 1. High-voltage electropherograms of [Co(en),Aa]” alanine, (3) L-valine and (4) D-valine. Voltage, 2500 V; paper, Whatman No. 1; time, 35 min. (a) 0.18 M solution of sodium (+)-tartrate @H 5); (b) 0.18 M solution of sodium (-)-tartrate.
the resolution of [Co(en)+alanine12 + and [Co(en),L-valine]’ + and antimony1 ( + )tartrate for complexes containing D-alanine and D-valine. As eluates containing optical isomers were very dilute and contained large amounts of the chiral counter ions, it was necessary to treat the samples in the following way. The samples were placed on a chromatographic column filled with a cationic exchanger [Chelex 100 (Bio-Rad (0)
0
0
1
0+
(b)
00
0 -
00 0
0+
00 Fig. 2. Resolution of (+)- and (-)-forms of [Co(en)zAa]2+ complexes (Aa as in Fig. 1) on Whatman No. 1 paper at 3500 V for 35 min. (a) 0.2 A4 solution of potassium antimony1 (+)-tartrate; (b) 0.2 M solution of sodium arsenyl ( + )-tartrate.
390
NOTES
(b)
(a)
Fig. 3. Resolution of (+)- and ( -)-[Co(en),~-alanine12+ on Whatman No. 1 paper at 3000 V for 35 min in a 0.18 M sodium (+)-tartrate solution. (a) Electropherogram; (b) scan of the separate isomers in (a). Wavelength, 565 nm; slit length, 4 mm; slit width, 0.1 mm; VP (plate travel) = V, (recorder chart speed) = 1 mm/s.
Labs.), 100-200 mesh] and eluted with water. The complexes remained at the top of the column and the anionic chiral counter ion was eluted. Subsequently, the complexes were eluted from the column with 0.1 M hydrochloric acid. After neutralization, the samples were dried and the optical rotations were measured with a PerkinElmer 141 polarimeter. It was thus possible to obtain pure resolved complexes without interfering anions. RESULTS
Figs. l-3 show the electrophoretic separations of some cobalt(II1) complexes with mixed ligands. Fig. la and b show the electropherograms of the four complexes studied when the counter ions were L( +)-tartrate or D( ---tartrate. It can be seen that with L( + )-tartrate solution we were able to resolve the ( + )- and (-)-forms of [Co(en)2r_.-Ala]2+ and [Co(en)+Val]’ + and with D( -)-tartrate we resolved the complexes containing D-alanine and D-valine. When we used antimony1 ( + )-tartrate as the electrolyte (Fig. 2a) all the racemic mixtures were resolved into their (+)- and (-)-isomers. Fig. 3 shows as an example the scan of one resolution obtained with this electrophoretic method. Hence we were able to set up a chromatographic method to obtain pure resolved complexes for standard purpose without interfering anions, and to devise an electrophoretic separation method for all the complexes studied. As previously reported6, in all the experiments antimony1 (+)-tartrate was found to be the best separating agent of the chiral anions used. The resolutions obtained were very good, as has been shown previously, when hydrogen bonding is involved. Although the electrophoretic method is mainly for analytical purposes, the use of a scanning apparatus allows the separations obtained to be monitored and the relative amounts of the separated isomers to be determined.
NOTES
391
CONCLUSIONS
The advantages of the above method are as follows: the time required for the separations is rather short (within 3.5 min, whereas for high-performance ion-pair chromatography an analysis time of 100 min is necessaryg); it is possible to study solution interactions, as the support does not interfere with the separation processes; and there is complete separation of racemic components from their mixtures and not only enrichment of one with respect to the other. REFERENCES L. Ossicini and C. Celli, J. Chromatogr., 115 (1975) 655. V. Cardaci, L. Ossicini and T. Prosperi, Ann. Chim. (Rome), 68 (1978) 713. V. Cardaci and L. Ossicini, J. Chromatogr., 198 (1980) 76. S. Fanali and L. Ossicini, J. Chromatogr., 212 (1981) 374. S. Fanali, V. Cardaci and L. Ossicini, J. Chromatogr., 265 (1983) 131. S. Fanali, L. Ossicini and T. Prosperi, J. Chromatogr., 318 (1985) 440. L. Ossicini and S. Fanali, Electrophoresis ‘86 -Proceedings of the Fifth Meeting of the International Electrophoresis Society, London, 9-12 September, 1986. 8 C. T. Liu and B. E. Douglas, Inorg. Chem., 3 (1964) 1356. 9 D. X. Buckingham, J. Chromatogr., 313 (1984) 93.
1 2 3 4 5 6 7
403 (1987) 388-391
Elsevier Science Publishers B.V., Amsterdam -
Printed in The Netherlands
CHROM. 19 684
Note
Paper electrophoretic
study
XVI*. Resolution of optical amino acid ligands
of ion-pair isomers
formation
of some cobalt(lll)
complexes
with
S. FANALI*, P. MASIA and L. OSSICINI Istituto di Cromatograjia del C.N.R., Rome (Italy)
Area della Ricerca di Roma, C.P. IO, 00016 Monterotondo
Scala,
(Received May 7th, 1987)
Ion-pair or outer-sphere complex formation is widely used in chromatography to achieve good separations of various compounds. Many years ago in our laboratory we studied ion-pair formation by electrophoretic techniques and succeeded in developing a good method for the separation of several metal complexes and racemic mixture&’ In this paper we report the electrophoretic study of some racemic cobalt(III) complexes with mixed ligands such as [Co(en),Aa12+, where Aa is an amino acid, using as counter ions aqueous solutions of sodium (+)- or (-)-tartrate, antimony1 ( + )-tartrate and arsenyl ( + )-tartrate. EXPERIMENTAL
A high-voltage paper electrophoresis apparatus (HVE Camag system) and Whatman No. 1 paper were used. The time required for the experiments varied from 15 to 35 min and the voltage from 2000 to 3500 V; the electric field applied was generally 50 V/cm. The cooling plate was kept at 68°C. Evaporation from the paper strips during the run was negligible. The electroosmotic flow in the HVE was measured by observing the movement of hydrogen peroxide; the results given are distances already corrected for electroosmotic flow. The samples were prepared as described below. Racemic mixtures were placed side by side with the two resolved isomers in the electropherograms. For the detection of the spots aqueous ammonium sulphide or Dragendorff reagent was used. A Camag TLC-HPTLC scanner was used for adsorption measurements directly on the electropherograms. Preparation of samples
Literature methods were used for the synthesis of racemic cobalt(II1) complexes*. Resolution of the optical isomers was achieved using a cationic-exchange column (Sephadex SP, C24; sodium (+ )-tartrate (0.18 M) was used as the eluent for * For Part XV, see J. Chromatogr., 318 (1985) 440. 0021-9673/87/$03.50
0
1987 Elsevier Science Publishers B.V.
3x9
NOTES (a)
o”o 00
0
0
-- 1
0+
.-
.- 3 2
-- 4
I-,
00
t
1
complexes, where Aa = (1) D-alanine, (2) LFig. 1. High-voltage electropherograms of [Co(en),Aa]” alanine, (3) L-valine and (4) D-valine. Voltage, 2500 V; paper, Whatman No. 1; time, 35 min. (a) 0.18 M solution of sodium (+)-tartrate @H 5); (b) 0.18 M solution of sodium (-)-tartrate.
the resolution of [Co(en)+alanine12 + and [Co(en),L-valine]’ + and antimony1 ( + )tartrate for complexes containing D-alanine and D-valine. As eluates containing optical isomers were very dilute and contained large amounts of the chiral counter ions, it was necessary to treat the samples in the following way. The samples were placed on a chromatographic column filled with a cationic exchanger [Chelex 100 (Bio-Rad (0)
0
0
1
0+
(b)
00
0 -
00 0
0+
00 Fig. 2. Resolution of (+)- and (-)-forms of [Co(en)zAa]2+ complexes (Aa as in Fig. 1) on Whatman No. 1 paper at 3500 V for 35 min. (a) 0.2 A4 solution of potassium antimony1 (+)-tartrate; (b) 0.2 M solution of sodium arsenyl ( + )-tartrate.
390
NOTES
(b)
(a)
Fig. 3. Resolution of (+)- and ( -)-[Co(en),~-alanine12+ on Whatman No. 1 paper at 3000 V for 35 min in a 0.18 M sodium (+)-tartrate solution. (a) Electropherogram; (b) scan of the separate isomers in (a). Wavelength, 565 nm; slit length, 4 mm; slit width, 0.1 mm; VP (plate travel) = V, (recorder chart speed) = 1 mm/s.
Labs.), 100-200 mesh] and eluted with water. The complexes remained at the top of the column and the anionic chiral counter ion was eluted. Subsequently, the complexes were eluted from the column with 0.1 M hydrochloric acid. After neutralization, the samples were dried and the optical rotations were measured with a PerkinElmer 141 polarimeter. It was thus possible to obtain pure resolved complexes without interfering anions. RESULTS
Figs. l-3 show the electrophoretic separations of some cobalt(II1) complexes with mixed ligands. Fig. la and b show the electropherograms of the four complexes studied when the counter ions were L( +)-tartrate or D( ---tartrate. It can be seen that with L( + )-tartrate solution we were able to resolve the ( + )- and (-)-forms of [Co(en)2r_.-Ala]2+ and [Co(en)+Val]’ + and with D( -)-tartrate we resolved the complexes containing D-alanine and D-valine. When we used antimony1 ( + )-tartrate as the electrolyte (Fig. 2a) all the racemic mixtures were resolved into their (+)- and (-)-isomers. Fig. 3 shows as an example the scan of one resolution obtained with this electrophoretic method. Hence we were able to set up a chromatographic method to obtain pure resolved complexes for standard purpose without interfering anions, and to devise an electrophoretic separation method for all the complexes studied. As previously reported6, in all the experiments antimony1 (+)-tartrate was found to be the best separating agent of the chiral anions used. The resolutions obtained were very good, as has been shown previously, when hydrogen bonding is involved. Although the electrophoretic method is mainly for analytical purposes, the use of a scanning apparatus allows the separations obtained to be monitored and the relative amounts of the separated isomers to be determined.
NOTES
391
CONCLUSIONS
The advantages of the above method are as follows: the time required for the separations is rather short (within 3.5 min, whereas for high-performance ion-pair chromatography an analysis time of 100 min is necessaryg); it is possible to study solution interactions, as the support does not interfere with the separation processes; and there is complete separation of racemic components from their mixtures and not only enrichment of one with respect to the other. REFERENCES L. Ossicini and C. Celli, J. Chromatogr., 115 (1975) 655. V. Cardaci, L. Ossicini and T. Prosperi, Ann. Chim. (Rome), 68 (1978) 713. V. Cardaci and L. Ossicini, J. Chromatogr., 198 (1980) 76. S. Fanali and L. Ossicini, J. Chromatogr., 212 (1981) 374. S. Fanali, V. Cardaci and L. Ossicini, J. Chromatogr., 265 (1983) 131. S. Fanali, L. Ossicini and T. Prosperi, J. Chromatogr., 318 (1985) 440. L. Ossicini and S. Fanali, Electrophoresis ‘86 -Proceedings of the Fifth Meeting of the International Electrophoresis Society, London, 9-12 September, 1986. 8 C. T. Liu and B. E. Douglas, Inorg. Chem., 3 (1964) 1356. 9 D. X. Buckingham, J. Chromatogr., 313 (1984) 93.
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