1788 Irena Malinowska1 Marek Studzin´ski1 Henryk Malinowski2 1

Department of Planar Chromatography, Chair of Physical Chemistry, Faculty of Chemistry, Maria CurieSk"odowska University, Lublin, Poland 2 Vexler and Baldin Laboratory of High Energy Physics, Join Institute of Nuclear Research, Dubna, Russia

Received March 17, 2011 Revised May 16, 2011 Accepted May 16, 2011

J. Sep. Sci. 2011, 34, 1788–1795

Research Article

Some aspects of TLC in homogenous magnetic fields This article consists of two parts. First part is a short review about the role of magnetic phenomena in natural environment, human surroundings, and his activities such as science, engineering, and medicine. The second part of the article presents a set of experiments, their results, and data obtained in a static homogenous magnetic field, generated by a pair of permanent magnets and outside it. Adsorption chromatographic systems were investigated: as chromatographed substances – polyaromatic hydrocarbon (PAH), as stationary phase – silica gel 60, as monocomponent mobile phases – n-hexane, n-heptane, n-octane, and benzene were used and binary mobile phases n-hydrocarbons – benzene. Magnetic field influences retention and efficiency of investigated chromatographic systems. Experimental data analysis (RF, N) allows us to propose some explanations of the differences between experiment results performed in induced magnetic field and outside it, and in consequence on the changes in the interfacial phenomena induced by field presence. Keywords: Magnetochromatography / Polyaromatic hydrocarbons / TLC DOI 10.1002/jssc.201100249

1 Introduction It is widely known that magnetic fields may influence a great number of processes in animated and unanimated nature. Combustion of liquid hydrocarbons, contained in gasoline–air mixture after magnetization in special device (magnetizer), is more efficient than without it. Passing the hydrogen through magnetic field, we obtain molecules of para-hydrogen (both nuclear spins are directed in the same way). Magnetic water treatment in heating devices and heat exchangers prevents effective installations against scale deposition. It was also observed that the presence of magnetic field influences living organisms. Various treatments consisting of magnetic field application can effectively accelerate healing of many different injuries and burns. Magnetic field also seems to mitigate rheumatic pain and brings relief in other diseases. Drugs subjected to magnetic field change their pharmacological activity and in effect their therapeutical dose is changed. Plants growing in magnetic field whose intensity is different from natural earth’s synthesize different amounts of nutrition components in their roots and plants. On the basis of examples presented above, it becomes necessary to state a question about the mechanism of Correspondence: Professor Irena Malinowska, Plac Marii CurieSk"odowskiej 3/216, 20-031 Lublin, Poland E-mail: [email protected] Fax: 148815333348

Abbreviation: PAH, polyaromatic hydrocarbon

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magnetic field influence on natural processes. Are the reported changes concerning only molecules/particles of investigated systems, or might they influence interfacial phenomena of this systems? Presented problem is extremely important specially for biological systems, where many processes take place in the interfacial area, and that is the reason why we decided to investigate whether there is any influence of magnetic field on interfacial phenomena using liquid chromatography. In the last few years, a number of articles about using the magnetic field for the separation of molecules/particles in liquid phase were presented. In those publications, the new word ‘‘magnetochromatography’’ was used [1–7] although it is not precisely defined yet. Nomizu et al. and Kim et al. used it for the description of magnetic particles’ separation from liquid phase (in this case, ‘‘magnetochromatography’’ does not seem to be a good word). The BIONANO Company uses it for static separation systems Dynabeadss, where in every particle of sorbent a ferromagnetic core is placed. Finally, Barrado and his team used the magnetic field as additional separation factor in chromatographic systems containing paramagnetic immobile phase. According to the studies of authors presented above, it seems to be interesting to examine the influence of magnetic field on the retention of compounds. Analyzing the differences of retention between experiments obtained in magnetic field, and outside it, will allow us to determine the influence of magnetic fields on detailed parts of chromatographic system (stationary phase, mobile phase, and chromatographed substances). In our previous study [8], differences in retention values of inactive polyaromatic www.jss-journal.com

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hydrocarbon (PAH) in monocomponent and binary mobile phases on silica gel were presented. Comparing retention of PAH on TLC and HPTLC plates it was observed that the influence of magnetic field on retention depends on the distance of chromatogram development. The explanation of that phenomenon may bring some information on the influence of magnetic field on chromatographic systems, and that is the reason why, in our present article, we decided to focus on that problem. As an investigation method, TLC was chosen. Its main advantages are simplicity, the possibility of chromatographic system rapid modification, and low operation costs. Besides, the equipment used for planar chromatography does not (or we would rather say should not) interfere with the external magnetic field used in the experiment. Moreover, this method, thanks to high level of instrumentation in present time, is very precise and reliable.

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Figure 2. Distance between walls of magnets versus inductivity of obtained magnetic field, and the spacing during the experiment.

Table 1. Chromatographed substances

2 Materials and methods 2.1 Magnets The measurements of magnetic field influence were carried out using the TLC technique. For this purpose, special modification of the chromatographic instruments was performed. ‘‘Sandwich’’-type chromatographic chamber was placed between a pair of neodymium magnets (after dismounting all of its ferro- and paramagnetic parts) (Fig. 1). The permanent magnets used in the experiment are described by the following parameters: (i) (ii) (iii) (iv) (v)

dimensions: 20 mm  50 mm  100 mm ¨ e) bHc coertion – min. 8992 A/m (11.3 kO ¨ e) jHc coercion – min. 955 A/m (12 kO energy density: 286–303 kJ/m3 max. working temperature: 353.15 K

Obtained inductivity of field between magnets was in the range of 0.44–0.48 T. In our experiments, 0.44 T inductivity was used (Fig. 2).

Id

Name

1W

Biphenyl

2W

Phenanthrene

3W

Pyrene

4W

Fluoranthene

5W

Chrysene

6W

Benzo-(a)-pyrene

Structure

2.2 Chromatographic conditions

Figure 1. Scheme of chromatographic system in magnetic field.

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TLC was carried out using stock, glass-backed, silica gel plates distributed by Merck (Darmstadt, Germany). The dimension of the used plates was 5 cm  10 cm. Chromatographic chamber – ‘‘sandwich’’ type – was manufactured by Chromdes (Lublin, Poland). The temperature during the experiment was 2170.21C/294.1570.2 K. As chromatographed substances, some PAHs were used (Table 1). www.jss-journal.com

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Figure 3. Images of chromatographic plates. (A) In the magnetic field. (B) Without magnetic field. Stationary phase: TLC SiO2 – 60. Mobile phase: benzene. Chromatographed substances – PAH (Table 1).

Substances were applicated onto the plates as 2.5 mm wide bands using CAMAG Linomat 5 applicator. Detection was performed using CAMAG Reprostar 3 and Baumer Optronics camera. Data were collected using CAMAG DigiStore2 software and evaluated using CAMAG VideoScan v. 1.02.00. Investigations were carried out using monocomponent and binary mobile phases. As monocomponent mobile phases, n-hexane, n-heptane, n-octane, benzene, and toluene were used. As binary phases, mixtures of n-hexane/benzene in 9:1, 7:3, and 1:1 volume fractions were used. Every measurement was repeated three times or more. All blunders were rejected.

3 Results and discussion The fact that confirms the influence of magnetic field on chromatographic process is the change in retention and band shapes of chromatographed substances. Figure 3 shows the images of chromatographic plates developed using exactly identical mobile phases in exactly the same conditions obtained in the presence of magnetic field and without it. As shown in the figure, the retention and bandwidth for the corresponding spots are different. Thus, we can assume that the change in retention is a result of the influence of magnetic field on chromatographic system.

3.1 Influence of magnetic field on the retention of chromatographed substances Presented research is just beginning, it was carried out on the simplest possible chromatographic systems with silica & 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

Figure 4. Pyrene peaks obtained in magnetic field and without it in n-hexane for different development distances.

gel as stationary phase and apolar chromatographed substances and mobile phases. Influence of magnetic field on retention may be described by comparing the corresponding substances’ RF values in magnetic field and outside it in different mobile phases. www.jss-journal.com

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3.1.1 Monocomponent mobile phases For the sake of properties of investigated chromatographic system, only apolar solvents were allowed to use, because in polar mobile phases almost all investigated substances were migrating on the solvent front, or they were outside the analytic range (Fig. 3). In case of apolar monocomponent mobile phases, without p-bonds in the molecule, it was observed that there is practically no influence of the magnetic field on retention [8]. In spite of it, in the case of n-hexane, n-heptane, and n-octane, it was

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noticed that differences in RF values of chromatographed substances depend on the distance of chromatogram development. The biggest differences between retention of chromatographed substances in magnetic field and outside it were observed on the distance of development which equals 4.5 cm (Fig. 4). Aside from the cases presented above, different RF values of investigated substances on different distances of the development were observed, regardless of whether chromatograms were developed in the field or outside the field.

Figure 5. RF values and their standard deviations of investigated PAH obtained without and in magnetic field in following mobile phases: n-hexane, n-heptane, and n-octane phases: (A) n-Hexane in magnetic field, (B) n-hexane outside the field, (C) n-heptane in magnetic field, (D) n-heptane outside the field, (E) n-octane in magnetic field, (F) n-octane outside the field.

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Changes in chromatographed substance retention on different development distances in the case of monocomponent mobile phases are explained by: (i) Incomplete moistening of adsorbent, by mobile phase (creation of a front) – but in the case of chromatography with the flow generated by capillary forces, this phenomenon practically does not take place. (ii) Evaporation of solvent from chromatographic plate surface to internal chamber space. In the case of ‘‘Sandwich’’-type chamber (used in experiment), internal chamber space volume is small, and it saturates quite fast and hence evaporation in the presented process proceeds only in the beginning of chromatogram development. Evaporation of solvent from the plate generates evaporative flow. Described phenomenon gives the fact that bigger volume of solvent flows through the sorbent bed than in case when evaporative flow does not take place. As a result, we can observe elongation distance of migration for chromatographed substances, in spite of the fact that there is no change in solvent front migration distance, what finally causes increase in chromatographed substances’ RF values. Considering retention data, it is noticeable that differences between corresponding RF values on the distance 4.5 cm and longer (6.5 and 8.5 cm) (DRF) are higher in magnetic field than DRF values obtained during the experiment without magnetic field (Figs. 5 and 6). Intensity

Figure 7. Computer simulation of magnetic field induced by pair of permanent magnets used in experiment, with field lines, local values of flux density, and local direction of B vector represented by arrows.

of magnetic field is practically constant on whole distance of development (8.5 cm) (Fig. 7); retention changes caused by the presence of field on short distance (4.5 cm) may be explained as a result of differences in the rate of evaporation of solvent from the surface of chromatographic plate to internal chamber space. The fact which proves the existence of evaporation flow are small differences between retention in the magnetic field and outside it obtained for n-octane and carbon tetrachloride on different distances of development (Fig. 6A and B). Saturated vapor pressures of those eluents are significantly lower than n-hexane and n-heptane.

3.1.2 Binary mobile phases

Figure 6. (A) Differences of RF values (DRF) for 4.5 and 6.5 cm distances of development obtained for chromatographed PAHs. (B) Comparison of differences between DRF values obtained for 4.5 and 6.5 cm distances of development obtained in magnetic field (DRFmag) and outside the field (DRFnmag) for n-hexane, n-heptane, and n-octane used as mobile phases.

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Differences in RF values versus migration distance were also present when binary mobile phases were used (n-hexane/ benzene). Similarly as in monocomponent mobile phases, rate of migration of chromatographed PAHs is higher in magnetic field than outside it for 4.5 and 8.5 cm distances of development. However, bigger differences for 4.5 cm are observed. It proves that, in this case, evaporative flow is also generated. As it was discussed above, differences in distances of migration of chromatographed substances on shorter development distances (4.5 cm) are the results of solvent evaporation from chromatographic plate. Similar situation is observed in binary mobile phase, but the differences between retention of investigated solutes on distance 4.5 cm in binary mobile phases are bigger than in monocomponent mobile phases (n-hexane and benzene). This phenomenon can be explained by evaporation of www.jss-journal.com

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mobile-phase components from the surface of chromatographic plate and, as a consequence, generation of evaporative flow. But, in the case of binary mobile phase, evaporation of the mobile phase to internal chamber space causes a change of composition of the mobile phase on the surface of the plate, because the saturated vapor pressures of the binary mobile-phase components are different. In the described investigation, n-hexane has a higher vapor pressure than benzene, and mobile-phase vapors are enriched in n-hexane in relation to bulk phase (in investigated system, the deviation of the Rault’s law is very small and hence it can be omitted). In consequence, mobile phase on the sorbent layer is enriched in less-volatile component, in the present case – benzene, which is simultaneously more active component. Thus, the elution strength of the mobile phase during the development of chromatogram increases, and the effect of evaporation is bigger than we observed for monocomponent mobile phases. The additional effect that can influence retention of chromatographed substances is demixion of binary mobile phase. All factors mentioned above can be used as explanation of bigger differences of RF values in binary mobile phases observed in this experiment. Similar dependencies were also observed for n-hexane/toluene mobile phase. In examined binary mobile phases, the presence of magnetic field increases obtained RF values, and differences between retention of PAHs in magnetic field and outside the field are bigger for shorter distances. The differences also depend on the composition of mobile phase (Fig. 8). As shown in Fig. 4, shape of the peaks is different when the chromatogram was developed in magnetic field and

outside it. The differences are also observable when the presence of magnetic field does not affect retention of chromatographed solute (Fig. 4C). Change in retention and bandwidth influences the efficiency of chromatographic systems. Figure 9 shows the differences in the number of theoretical plates (N) obtained in magnetic field and outside it on distances 4.5 and 8.5 cm. As presented above, the efficiency of separation on distance 4.5 cm is higher for experiments carried out in magnetic field, for n-hexane and n-heptane and comparable for n-octane. For longer distances of development, effect of magnetic field for n-hexane and n-heptane may be neglected,

Figure 8. RF values obtained in magnetic field (m-designated) and outside the field for examined PAHs on different distances of development. Mobile phase: n-hexane/benzene, (A) 9:1 v/v and (B) 7:3 v/v.

Figure 9. Number of theoretical plates obtained for examinated PAHs in n-hexane (A), n-heptane (B), and n-octane (C) as mobile phase obtained for different distances of development in magnetic field (m-designated) and outside it.

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and for n-octane efficiency of chromatographic system decreases in the presence of investigated magnetic field. Analyzing the peak shapes developed in and outside the magnetic field, one can see that they differ not only in their width but also in their surface area. The surface area of the peaks is used for quantitative evaluations and is repeatable for TLC method, especially when used with full instrumentation. When it comes to the specification of the quantitative evaluations in TLC, only the part of the substance that is on the surface layer of the sorbent is detected. The peak shape image of the chromatographic band that is obtained is just the top of the upper part of the substance in the mobile phase. However, considering the fact

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that vertical distribution of substance is repeatable enough to be used for quantitative evaluation in planar chromatography, and it can be connected with surface area of the chromatographic peak obtained using VideoScan software. The surface area of the peaks in the magnetic field and outside it differs. The differences in the surface areas are present independently of whether there was a retention change or not. Figure 10 shows the example of the calibration curves in and outside the magnetic field. The data show that the peak surface changes in the magnetic field together with the amount of the substance in the spot are more regular than when outside the field (higher R2 coefficient). Magnetic field can change the relationship between the analytic signal and the amount of the substance – for benzo-(a)-pyrene and chrysene; the signal from videodensitometric evaluation of spots obtained in magnetic field is higher than outside the field – steeper curve of the peak surface area versus amount of the examined substance was obtained. In such cases, the quantitative evaluations carried out in the field are more accurate in most of the cases. In consequence of the inversely proportional dependence between limit of detection (LOD) and slope of calibration curve (assuming similar values of standard deviation), the presence of magnetic field lowers the LOD for substances shown in Fig. 10A and C (calculated LODs for biphenyl and benzo-(a)-pyrene are 0.9853 and 0.3233 mg in the magnetic field and 1.5331 and 0.9281 mg outside the magnetic field, respectively). However, the magnetic field does not always cause the increase in the peak area or the signal amplification – the example of that phenomenon may be the presented calibration curves obtained for chrysene – but the R2 values are still higher in the field than outside it (Fig. 10B). The width change of the peaks tells us about the diffusion processes taking place on the surface of the chromatographic plate, whereas the height (area) modification of the peak tells us about the concentration of the substance in the surface layer of the sorbent. Different areas of the peaks represent different substance distributions in the sorbent layer in and outside the magnetic field.

4 Concluding remarks

Figure 10. Quantitative dependencies obtained in magnetic field and outside it for (A) benzo-a-pyrene, (B) chrysene, and (C) biphenyl.

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The presence of the magnetic field modifies the solvent evaporation rate from the sorbent layer – the phenomenon that generates changes in the evaporative flow. It is clearly observable for short distances of chromatogram development. The experiments carried out for monocomponent mobile phases with high vapour pressure show that the evaporative flow is stronger in the magnetic field than outside it. This observation proves the increment of solvent evaporation rate from chromatographic plate in magnetic field. In case of two-component mobile phases, evaporative flow is also generated, but it is not responsible for the change in retention from the distance of development dependence. As a result of ingredients evaporation of mobile phase from chromatographic plate, the composition www.jss-journal.com

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of mobile phase in the sorbent layer changes. The mobile phase on the surface of the chromatographic plate is enriched in less volatile component. In case of mobile phases presented in this article, the less volatile ingredient is also more active and hence the elution strength of the solvent on the plate surface increases. Magnetic field also influences the diffusion processes taking place in stationary phase (it is proved by differences in band width for the experiments carried out in and outside magnetic field) and vertical distribution of chromatographed substance in sorbent layer (which was proved in our experiment as a difference in peak areas obtained for two identical chromatographic systems in the magnetic field and outside it). Considering that the presence of magnetic field modifies retention and spot width of chromatographed substances, it also influences the efficiency of the chromatographic system. In many cases, the efficiency of separation is increased, what is represented as decrement of height equivalent to a theoretical place value.

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5 References [1] Nomizu, T., Yamamoto, M., Wanatabe, K., Anal. Sci. 1996, 12, 829. [2] Nomizu, T., Yamamoto, M., Wanatabe, K., Anal. Sci. 2001, 17, 1177–1190. [3] Kim, S. B., Nakada, C., Murase, S., Okada, H., Oharab, T., Physica C 2007, 463– 464, 1306–1310. [4] Fukui, S., Shoji, Y., Ogawa, J., Oka, T., Yamaguchi, M., Sato, T., Ooizumi, M., Imaizumi, H., Ohara, T., Sci. Technol. Adv. Mater. 2009, 10, 014610. [5] Barrado, E., Rodrı´ques, J. A., J. Chromatogr. A 2006, 1128, 189–193. [6] Barrado, E., Rodriguez, J. A., Castrilejo, Y., Anal. Bioanal. Chem. 2006, 385, 1233–1240. [7] Barrado, E., Rodriguez, J. A., Castrilejo, Y., Talanta 2009, 78, 672–675. [8] Malinowska, I., Studzin´ski, M., Malinowski, H., JPC-J. Planar. Chromatogr. 2008, 21, 379–385.

The authors have declared no conflict of interest.

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Some aspects of TLC in homogenous magnetic fields.

This article consists of two parts. First part is a short review about the role of magnetic phenomena in natural environment, human surroundings, and ...
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