JPH05322849A - Kinetic chromatography method - Google Patents

Abstract

PURPOSE:To separate substances within an organic solvent effectively by adding Star Burst Dendrimmer(SBD) into a cataphoresis buffer liquid with a high organic solvent concentration. CONSTITUTION:An SBD which is a single-dispersion gigantic molecule with highly regular and symmetrical structure is applied as an ion interface activator in a kinetic chromatography under a high organic solvent concentration. Methanol is used as a reaction solvent, michael is added to acryl acid methyl of amine component, an excessive amount of etylenediamine is reacted with generated ester, and amide connection is generated, thus obtaining SBD at the end of amino group. A gigantic SBD is formed by repeating this sort of two-stage reaction. When the SBD is used as a carrier 50 of kinetic chromatography, it is stable even under presence of an organic solvent, thus forming micell (cataphoresis) properly and establishing conductive chromatography even under presence of a high-concentration organic solvent.

Description

Detailed Description of the Invention

[0001]

FIELD OF THE INVENTION The present invention relates to an electrokinetic chromatography method, and more particularly to an improvement of the electrokinetic chromatography method used for separating substances in an organic solvent.

[0002]

2. Description of the Related Art Capillary electrophoresis has attracted attention as an electrophoretic method that can be used as an instrument. Since the principle of separation is different from that of ordinary high performance liquid chromatography, the separation selectivity is different and the sample has high performance. It has the advantages that the amount is small and that the absolute detection sensitivity is high, and that it has the advantages that the data processing similar to chromatography can be performed by adopting the online detection method. This capillary electrophoresis method is a general term for electrophoresis performed in a capillary having an inner diameter of about 0.1 mm or less, and generally, an electric field of several hundred V per cm is applied for separation. As the separation mode of capillary electrophoresis, capillary zone electrophoresis performed in free solution is the most common, but isoelectric focusing method performed in free solution and capillary gel electrophoresis method using gel-filled capillaries are also used. Has been done.

However, since these are all electrophoretic, they are applied to the analysis of charged substances, that is, ions, and are not effective for the separation of neutral substances. Therefore, an electrokinetic chromatography method has been attracting attention as a capillary electrophoresis method effective for separating such neutral substances.
The most commonly used electrokinetic chromatography method is the micellar electrokinetic chromatography method, in which an ionic surfactant micelle is added to a buffer for electrophoresis to facilitate incorporation into the micelle. By utilizing the difference between the two, neutral substances that cannot be separated by electrophoresis are separated.

[0004]

However, in the above-mentioned conventional electrokinetic chromatography method, the micelles of the surfactant are significantly less likely to be formed when the ratio of the organic solvent in the electrophoresis buffer is increased, and the separation state is It may change or may become impossible to separate. For this reason, in particular, the separation of the fat-soluble component has been extremely difficult because it is necessary to use an organic solvent due to its solubility. The present invention has been made in view of the above problems of the prior art, and an object thereof is to provide an electrokinetic chromatography method capable of efficiently performing capillary electrophoresis even when the concentration of an organic solvent is high.

[0005]

In order to achieve the above-mentioned object, the electrokinetic chromatography method according to claim 1 of the present application comprises adding a starburst dendrimer to an electrophoresis buffer having a high organic solvent concentration to form a capillary. It is characterized by performing electrophoresis. Further, the electrokinetic chromatography method according to claim 2 is characterized in that the organic solvent concentration is 50% by volume or more.

[0006]

The electrokinetic chromatography method according to the present invention comprises:
Since it is carried out at a high organic solvent concentration as described above, it is possible to easily dissolve the fat-soluble components even when the components are separated. Under such a high organic solvent concentration, it is difficult to form micelles with an ordinary ionic surfactant, but according to the present invention, since the starburst dendrimer is used, good micelles are formed and high. Capillary electrophoresis in electrophoresis buffer with organic solvent concentration is possible.

[0007]

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT A preferred embodiment of the present invention will be described below with reference to the drawings. FIG. 1 shows a conceptual diagram of a capillary electrophoresis method to which the present invention is applied. In the figure, both ends of the fused silica tube 10 which constitutes a capillary are electrically connected to platinum electrodes 16 and 18 through electrolytic solutions 12 and 14. The platinum electrodes 16 and 18 have a high-voltage power supply 2
Higher voltage than 0 (eg maximum output 30kV, 100μA)
Is applied, and the current value can be monitored by the ammeter 22. Further, a detector 24 is provided in the middle of the fused silica tube 10. Therefore, the substance separated in the fused silica tube 10 is detected by the detector 24.

Here, the separation in the fused silica tube 10 is performed as shown in FIG. In the figure, the inner surface of the fused silica tube 10 has a negative charge. Therefore, the liquid in the fused silica tube has an equal amount of positive charge to neutralize the negative charge on this surface. Furthermore, many of the positive charges are attracted to the negative charges on the surface, forming an electric double layer. Therefore, when an electric field is applied to both ends of the silica tube 10, the internal sample liquid 26 having a positive charge is pulled toward the negative electrode, and the whole sample liquid moves integrally. This is an electroosmotic flow, and in capillary electrophoresis, movement occurs due to this electroosmotic flow simultaneously with normal electrophoresis.

However, in this capillary electrophoresis, it is necessary that the substance to be separated has an electric charge. Therefore, in the present invention, the micelle electrokinetic chromatography method (MEKC) is used to measure neutral substances.
To adopt. In this electrokinetic chromatography method, as shown in FIG. 3, a neutral substance 30 is taken into a micelle 34 formed of, for example, an anionic surfactant 32, and the micelle 34 is
Since a negative charge appears on the surface of the, the neutral substance 30 is moved / separated together with the micelle 34.

However, as shown in FIG. 4, when methanol was added as an organic solvent to the electrophoresis buffer, there was a delay in the migration time of each solute as the concentration of methanol increased. It is considered that this is because the addition of methanol reduced the ionic strength of the separated solution, and consequently the electroosmotic flow velocity. Especially when the methanol content exceeds 50%, the separation time is remarkably increased, and the separation efficiency is also sharply reduced. Therefore, it is understood that the addition of a large amount of the organic solvent to the separation solution increases the separation time, which is not preferable in terms of separation efficiency.
The separation selectivity in sodium dodecyl sulfate-micelle electrokinetic chromatography (SDS-MEKC) is SD.
Since the alkyl chain length of S is shorter than that of cetyltrimethylammonium chloride (CTAC), it is completely lost at 50% methanol concentration due to its poorer stability to organic solvents. As described above, the addition of an organic solvent to the separation solution in MEKC reduces the Elution range and also the separation efficiency. Therefore, it is considered that MEKC in the presence of 50% or more of the organic solvent is not practical. Was there.

Therefore, the inventors of the present invention aimed to apply the starburst dendrimer to electrokinetic chromatography under high organic solvent concentration. That is, starburst dendrimers (SBDs) are monodisperse macromolecules with highly ordered and symmetrical structures. FIG. 5 shows a synthetic scheme of SBD. In the figure, using methanol as a reaction solvent, a two-step reaction of (A) amine component to methyl acrylate Michael addition and (B) ester amidation was repeated and synthesized. In the reaction (A) in the scheme, a branched structure of SBD is formed, and three bonds are formed for the ammonia molecule and two bonds are formed for the amino group, and the terminal is methoxy derived from methyl acrylate. Converted to carbonyl. When the produced ester is reacted with a large excess of ethylenediamine in the reaction (B), an amide bond is produced and an amino group-terminated SBD is obtained.

By repeating the above two-step reaction, a huge SBD is formed as shown in FIG. Next, a specific method of forming the SBD will be described. (A) Synthesis of Generation 0.5 In a 200 ml flask, 28% ammonia water 0.5 g
(0.082 mol) is dissolved in 100 ml of methanol,
23.4 g (0.27 mol, Mw = 86.09) of methyl acrylate (MA), which was 3.3 times equivalent to that of ammonia, was added, and the mixture was stirred at room temperature for 48 hours. After the reaction, the solvent was distilled off at 45 ° C. and further distilled under high vacuum under reduced pressure to remove unreacted methyl acrylate. As a result, a slightly viscous colorless transparent liquid was obtained. (B) Synthesis of Generation 1.0 10 g of generation 0.5 triester (0.036 mo
L, Mw = 275) dissolved in 50 ml of methanol
180 g of ethylenediamine (EDA) 82 times equivalent to 3.0 of eneration (3.00 mol, Mw = 60.
The mixture was gradually added dropwise to 500 ml of a methanol solution containing 10) while stirring, and then reacted at room temperature for 55 hours. After the completion of the reaction, most of the excess ethylenediamine was distilled off with an evaporator, and the pressure was reduced under high vacuum at 50 ° C. for 12 hours to remove the excess ethylenediamine. As a result,
A pale yellow viscous liquid was obtained. (C) Synthesis of Full Generation SBD According to the method of (b) above, Half Genaration as a raw material
Full Generation SBD was synthesized by reacting a large excess of EDA with SBD. Half Genera
Table 1 below shows the equivalent amount (n), required amount (g), and reaction time of EDA necessary for synthesizing Full Generation SBD from 5.0 g of tionSBD. Also, Generation 2.0
In the subsequent SBD synthesis, the solvent was distilled off and then S
BD was dissolved in a small amount of methanol, 1-butanol (Mw = 74.12 bp = 117.7 ° C.) was added, the solvent was further distilled off under reduced pressure, and ethylenediamine was removed by azeotropic distillation.

[0013]

[Table 1] ──────────────────────────────────── Start G 0.5 1.5 2. 5 3.5 4.5 5.5 5.5 6.5 G G 1.0 2.0 3.0 3.0 4.0 5.0 6.0 7.0 ─────────────── ────────────────────── n 82 330 980 2600 7000 17000 40000 EDA (g) 90 113 113 141 174 226 271 316 Reaction time (days) 2 4 6 8 12 14 18 ──────────────────────────────────── (d) Synthesis of Half Generation SBD (a) ).
Half Generation SBD was synthesized by reacting 10% excess MA with respect to all the end groups of SBD. After completion of the reaction, the solvent was distilled off, and the solvent and unreacted MA were distilled off under high vacuum. If the number of moles of Full Generation SBD is a and the number of end groups is Z, the required amount of MA (g)
Is calculated as a × 2 × Z × 1.1 × 86.09.
Table 2 below shows Hal from Full Generation SBD 5.0g
The amount of MA and the reaction time required to synthesize f Generation SBD are shown.

[0014]

[Table 2] ──────────────────────────────────── Start G 1.0 2.0 3. 0 4.0 5.0 5.0 Production G 1.5 2.5 3.5 3.5 4.5 5.5 6.5 ───────────────────── ──────────────── MA (g) 7.9 5.4 4.7 4.7 4.4 4.3 4.2 Reaction time (days) 2 4 7 10 10 10 ─ ─────────────────────────────────── (e) Synthesis of carboxyl group-terminated SBD Half Generation SBD (G = 2.5) 5.0 g to 50
After dissolving in ml of methanol and adding 1.2 g (0.03 mol) of sodium hydroxide in an amount equal to that of the terminal group, the mixture was stirred at room temperature for 6 hours. After the reaction was completed, the solvent was distilled off, and the solution dissolved in about 5 ml of methanol was dropped into ether to give SB.
D was precipitated. The ether was filtered off, washed with a small amount of ether, and dried under reduced pressure. As a result, white fine powder was obtained. (F) Alkylation of Full Generation SBD 2.0 g of SBD (G = 3.0) was dissolved in 30 ml of methanol, and a 2-fold equivalent amount of 1,2-Epox to all terminal amino groups was dissolved.
1.60 g (20 mmol, Mw = 72.11) of ybutane was added, and the mixture was stirred at room temperature for 24 hours. After the reaction, the solvent is distilled off,
It was dissolved in a small amount of methanol and precipitated in water-acetone. Dissolve the precipitate in methanol and use high vacuum at 50 ° C for 1
It was dried under reduced pressure for 2 hours (this alkylated SBD was treated with G = 3.
To as 0-C _4: C ₄ indicates the number of carbon atoms in the alkyl group). Similarly for 1,2-Epoxydecane, SBD (G =
3.0) 2.0 g to 3.11 g (20 mmol, Mw =
156.27) were reacted _{(G = 3.0-C 10)} . (G) Alkylation of Half Generation SBD 2.0 g of SBD (G = 3.5) was dissolved in 50 ml of methanol, and 1.2 times equivalent amount of Octylamine 1.
66 g (12.9 mmol, Mw = 129.24) was added and reacted at room temperature for 5 days. After completion of the reaction, the solvent was distilled off with an evaporator, and the pressure was reduced under high vacuum at 50 ° C. for 12 hours (G = 3.5-C ₈ ). Similarly for Propylamine,
For SBD (G = 3.5) 2.0 g, 0.76 g (1
2.9 mmol, Mw = 59.11) was reacted (G = 3.
5-C _3).

The SBD formed as described above was used as a carrier 5 for electrokinetic chromatography as shown in FIG.
When used as 0, the SBD carrier was stable even in the presence of an organic solvent, so that it was revealed that the electrokinetic chromatography system was established even in the presence of a high concentration of organic solvent. Further, since the separation solution containing SBD has a high ionic strength, the current value was higher than that of a general micelle system, but in the presence of an organic solvent, the current value decreased to a range suitable for separation.

FIG. 8 shows the relationship between the current value and the electroosmotic flow rate when methanol was added to the separation solution containing SBD. As is clear from the figure, a reverse electroosmotic flow was observed when Full Generation SBD was added to the separation solution even in the presence of an organic solvent. When the voltage was kept constant, the current value and the electroosmotic flow mobility decreased due to the decrease in ionic strength due to the increase in the organic solvent concentration. However, in SBD-electrokinetic chromatography, by applying a high voltage, an electroosmotic flow velocity similar to that in water was obtained.

Furthermore, the presence of SBD in the separation solution gives a sufficiently large electroosmotic flow velocity even in methanol, and MEKC can be applied even in a non-aqueous solvent if SBD is added to the separation solution. It became clear that Generally, in an HPCE system in which a positive direction electroosmotic flow flows, it is known that the electroosmotic flow velocity increases but the electroosmotic flow mobility decreases when the methanol concentration of the separation solution is increased with a constant current. There is. A similar tendency was observed in a system in which reverse electroosmotic flow was obtained in the presence of SBD. Next, the effect of adding an organic solvent in SBD-electrokinetic chromatography was examined.
Table 3 below shows Full Generation SB
The results of adding methanol to a separation solution containing D and having a pH of 5 to separate aromatic hydrocarbons having high hydrophobicity are shown.

[0018]

[Table 3] Effect of addition of organic solvent ──────────────────────────────────── MeOH (%) C GE I (mM) (V / Cm) (μA) ───────────────────────────────────── 50 5 3.0 400 20.0 10 3.0 300 24.0 60 20 3.0 3.0 300 28.0 70 10 3.0 400 400 24.0 80 10 3.0 400 16.0 100 10 3.0 400 13.6 20 3.0 400 20.8 ───────────────────────────────────────── ───────────────────────────────── MeOH (%) t ₀ / t _p ¹⁾ 10 ⁴ X μ _ep・ ( P) ²⁾ α ( _P / _N ) ³⁾ ─────────────── ───────────────────── 50 0.83 0.221 1.30 0.84 0.295 1.24 60 0.80 0.283 1.20 70 0.87 0.221 1.13 80 0.85 0.248 1.10 100 0.88 0.210 1.19 0.81 0.272 1.25 ──────────── ───────────────────────── C: SBD concentration t _p: travel time pyrene t _O: travel time 2 solutes having no charge): Electro-osmotic mobility of pyrene 3): Relative retention time of pyrene and naphthalene

In Table 3 above, in order to compare the magnitude of retention of the solute by SBD, t ₀ with respect to the traveling time of pyrene is compared.
Ratio (t ₀ / t _p ) and the effective electrophoretic mobility of pyrene (μ _ep ·
(P)). As is clear from Table 3, t ₀ / t _p
The values of and (μ _ep · (P) showed almost constant values without being affected by the addition of the organic solvent.
In the SBD-kinetic chromatography system using BD, even if the concentration of methanol in the separation solution is reduced, SB
No increase in solute retention due to D was observed. Further, the separation coefficient α of naphthalene and pyrene decreased as the organic solvent concentration increased, but the peak shape was improved by the addition of the organic solvent, so the actual separation was improved. This result is in contrast to the trend in reverse phase chromatography where solute retention decreased with increasing organic solvent content in the mobile phase. The hydrophobicity of SBD is small, but Full Generation
When methanol is added to the pH = 5 separation solution containing SBD, it is considered that the contribution of the hydrophobic interaction to the retention of elution is small.

As a result, it is suggested that if SBD is added to the separation solution, electrokinetic chromatography can be applied even in a non-aqueous solvent. Even if only SBD is added to methanol, almost no current flows, but about 0.0
An electric current was passed by adding 5% acetic acid and an appropriate electroosmotic flow was obtained. The separation selectivity of the carboxyl group-terminated SBD changed depending on the pH of the separation solution, which is considered to be due to the change of the hydrophobic environment inside or outside the SBD molecule depending on the pH. Therefore, the same effect is expected in the presence of an organic solvent,
For the purpose of increasing the hydrophobicity of the SBD end, an SBD having an alkyl chain bound thereto was also examined. Next, Table 4 shows the separation results of aromatic hydrocarbons in methanol.

[0021]

[Table 4] Separation of polycyclic aromatics in methanol ──────────────────────────────────── Carrier E (v / cm) I ( μA) t 0 / t p 1) ───────────────────────────────── --G = 3.0 400 13.6 0.88 G = 3.0-C ₄ 500 7.2 0.89 G = 3.0-C ₁₀ 500 12.4 0.80 G = 3.5 -C ₃ 500 16.0 0.92 G = 3.5-C ₈ 500 30.4 0.92 ────────────────────────── ─────────────────────────────────────────────── Carrier 10 ⁴ Xμ _ep・ (P) ²⁾ α (P / N) ³⁾ α (T / 0) ⁴⁾ ────────── ────────────────────────── G = 3.0 0.180 1.19 1.24 G = 3.0-C ₄₀ . 186 1.25 1.32 G = 3.0-C ₁₀ 0.273 1.25 1.39 G = 3.5-C ₃ 0.113 1.28 1.41 G = 3.5-C ₈₀ 0.098 1.44 1.43 ──────────────────────────────────── 4) 0-terphenyl Relative retention time of triphenylene As is clear from Table 4, Full Generation SBD
The terminal amino groups and epoxy degree of G = 3.0-C ₄ bound to an alkyl chain by the reaction and, G = the 3.0-C ₁₀ carriers, appear the effect of terminal alkyl chain of, mu _ep・ The value of (P) was increased. But G
= 3.5−C ₃ , G = 3.5−C ₈ has a small μ _ep
(P) values were given and no effect of alkyl chains on solute retention was observed. It is considered that such a difference in the amount of retention with respect to the solute mainly affects the separation characteristics of the SBD before binding the alkyl chain. That is,
Amino group terminal SBD in aqueous solution of pH = 5.0
(G = 3.0) holds the carboxyl group terminal S
It is considered that this is due to the fact that it is larger than BD (G = 3.5).

In the reverse phase chromatography, the separation coefficient (α (T / T /
O)) is an index for evaluating the density of the alkyl chains in the stationary phase, because the two compounds have almost the same degree of hydrophobicity and a different three-dimensional shape. Also in SBD-electrokinetic chromatography, it is suggested that the separation coefficient (α (T / O)) of these compounds serves as an index for evaluating the stereoselectivity of the carrier. Incidentally, FIG.
Chromatography of triphenylene and o-terphenyl in methanol is shown in. In electrokinetic chromatography using SBD with an alkyl chain attached, no significant increase in solute retention was observed, but the separation of triphenylene and o-terphenyl was improved and the α (T / O) of Table 4 The value increased.

The above results show that the alkyl chains bound to the outer surface of SBD improve the stereoselectivity of SBD for elution, but do not increase the retention for elution.
Next, improvement of separation by SBD-electrokinetic chromatography in methanol / buffer solution was investigated. That is, in a separation solution containing a high concentration of organic solvent, Full
The retention due to the hydrophobic interaction of Generation SBD was small. Therefore, in order to increase the interaction between SBD and the sample, separation in 50% ethanol was examined, and the results are shown in Table 5 below.

[0024]

[Table 5] Separation of polycyclic aromatics loaded in 50% methanol ─────────────────────────────────── -Carrier C (mM) pH E (v / cm) 10 ⁴ Xμ _ep · (P) ───────────────────────────── ──────── G = 3.0 5.0 5.0 400 400 0.221 G = 7.0 0.6 5.0 400 400 0.311 G = 2.5 5.0 11.0 500 -0.250 G = 3.5 5.0 5.0 5.0 500 ... G = 3.5 5.0 9.0 400 -0.284 G = 3.5 5.0 11.0 400-0. 381 * G = 3.5 5.0 11.0 400 -0.295 G = 6.5 1.0 11.0 400 -0.390 2.0 11.0 400 -0.623 ───── ─────── ──────────────────────── ─────────────────────────── ── t ₀ / t _p α (P / N) α (T / 0) α (F / D) ────────────────────────── ─── 0.83 1.30 1.46 1.14 0.79 1.41 1.79 1.21 0.83 1.12 1.20 …… 0.97 1.00 …… 1.00 .81 1.25 1.38 1.13 0.61 1.25 1.46 1.15 0.80 1.06 ...... 0.78 1.28 1.47 1.09 0.67 1. 26 1.42 1.15 ──────────────────────────── As is clear from Table 5, SBD having any end group
Also, higher generation SBDs have larger μ _ep · (P)
Gave a value. As shown in FIG. 10, Full Generation S
When BD was used, the increase in solute retention was small, and the peak tended to show tailing. On the other hand, in the alkaline separation solution, the SBD at the carboxyl group end
The retention of solutes was increased with the use of and the separation was improved even with the previous SBD. In FIG. 11, G =
The separated state of aromatic hydrocarbons at 3.5 (pH = 11) is shown. Further, when the methanol content of the separated solution of G = 3.5 (pH = 11) is 50% to 70%, μ in Table 5
The absolute value of _ep · (P) decreased to 0.381 to 0.295.

As described above, in the carboxyl group-terminated SBD system, the contribution of the hydrophobic interaction was observed in the retention of elution. Even when using an organic solvent as a separation solution, Fu
In the SBD-electrokinetic chromatography system using ll Generation SBD for separation, reverse electroosmotic flow was observed. Therefore, it is considered that the SBD exists on the inner wall of the capillary, and the SBD adsorbed on the inner wall interacts with the sample molecules, resulting in a decrease in separation efficiency.
Since the magnitude of the interaction with the solute depends on the molecular weight of SBD, it is considered that addition of high generation Full Generation SBD is disadvantageous in terms of separation efficiency.

On the other hand, Half Gener under alkaline conditions
In the ation SBD, a positive electroosmotic flow from the positive electrode to the negative electrode was observed, suggesting that the SBD existing on the inner wall is small. In the SBD-kinetic chromatography system using the Half Generation SBD, the adsorption of SBD is effective due to the repulsion between the anionic charge of the terminal carboxyl group and the silanol group on the inner surface of the capillary, as in the case of protein separation in the alkaline region. It is suppressed. Further, the carboxyl group-terminated SBD imparts different selectivity depending on the pH of the separation solution.

FIG. 12 shows the results of studies on the influence of the change in the pH of the separation solution on the separation. Naphthalene and pyrene are separated under alkaline conditions but not in the acidic region. It is believed that the high pH separation solution increases the hydrophobicity inside the carboxyl-terminated SBD molecule, favoring the alignment of solute molecules on the SBD. On the contrary, it is considered that the cationic charge existing inside the carboxyl group-terminated SBD reduces the hydrophobicity in the separation solution having a low pH, which makes the retention of the solute disadvantageous. From the chromatography of Figure 12, the pH
= 5.0, pyrene and naphthalene were not separated, but at pH = 3.0, these compounds were separated. That is, it can be considered that at pH = 3.0, the effect of the hydrophobicity of the SBD terminal, which is a neutral type, appeared.

As described above, the carboxyl group-terminated SBD
Changed the hydrophobic environment inside and outside the molecule depending on the pH of the separation solution, and gave different separation selectivity to aromatic compounds. The above results indicate that the interaction between the carboxyl group-terminated SBD and the solute depends on the hydrophobicity inside the SBD molecule, and the increase in the hydrophobicity inside the SBD molecule increases the separation ability. Can be done.

[0029]

As described above, according to the electrokinetic chromatography method of the present invention, since the starburst dendrimer is used as the carrier, the substance can be efficiently separated especially in the organic solvent.

[Brief description of drawings]

[Figure 1]

FIG. 2 is an explanatory diagram of the concept of capillary electrophoresis.

FIG. 3 is an explanatory diagram of a separation concept of micellar electrokinetic chromatography.

FIG. 4 is an explanatory view of the influence of addition of an organic solvent on the micelle electrokinetic chromatography shown in FIG.

FIG. 5 is an explanatory diagram of a synthetic scheme of starburst dendrimer.

FIG. 6 is an explanatory diagram of the structure of a starburst dendrimer.

FIG. 7 is an explanatory diagram of a separation concept of electrokinetic chromatography using a starburst dendrimer.

FIG. 8 is an explanatory diagram of a relationship between an organic solvent concentration and electroosmotic flow in electrokinetic chromatography using a starburst dendrimer.

FIG. 9 is an explanatory diagram of the effect of an alkyl chain at the end of a starburst dendrimer in methanol.

FIG. 10 is an explanatory view of a separated state of polycyclic aromatics by Full Generation SBD (G = 7.0).

FIG. 11 is an explanatory diagram of a separated state of polycyclic aromatics by Half Generation SBD (G = 3.5).

FIG. 12 is an explanatory diagram of changes in separation selectivity depending on pH.

Claims (2)

[Claims] 1. An electrokinetic chromatography method, wherein a starburst dendrimer is added to an electrophoresis buffer solution having a high organic solvent concentration, and capillary electrophoresis is performed. 2. The electrokinetic chromatography method according to claim 1, wherein the concentration of the organic solvent is 50% by volume or more.