Original

Chromatography, Vol.28 No.1 (2007)

Original

Specific separation of polar analytes on a medium polar stationary phaseby gas chromatography with carrier gas containing vaporous modifiers
Hiroshi Ito1,2*, Takashi Sugiura3, Hidenori Nagai3, Atsushi Murase1,Shuji Kodama4, Yoshinori Inoue5, Atsushi Yamamoto3 , Kazuichi Hayakawa2
1
2

Toyota Central R&D Labs., Inc., 41−1, Nagakute, Nagakute−cho, Aichi 480−1192, Japan

Graduate School of Natural Science and Technology, Kanazawa University, Kakuma−machi, Kanazawa−shi, Ishikawa 920−1192, Japan3

College of Bioscience and Biotechnology, Chubu University, 1200, Matsumoto−cho, Kasugai−shi, Aichi 487−8501, Japan
4Toyama Institute of Health, 17−1 Nakataikoyama, Imizu−shi, Toyama 939−0363, Japan

5

ChromaNik Technologies Inc., 6−3−1, Namiyoke, Minato−ku, Osaka 552−0001, Japan
Received for review December 12, 2006. Accepted December 28, 2006.

ABSTRACT
We have developed a novel generator that constantly introduces a vaporous modifier into the inert carrier gas. The effect of different modifiers on the retention times of analytes using a medium polar stationary phase and the generator was studied. When using formic acid as amodifier, the line of the van’t Hoff plot of acetic acid was convexly curved. The inflection point was in the range of 100 to 120 °C, which corresponds to the boiling point of formic acid under pressurized conditions. The inflection point was found to be related to the association offormic acid.
Key words: vaporous modifier, introducer, polar analytes, medium polar stationary phase

instruments were commercially available which gave the effective

Introduction
One of the problems of GC is the use of inert gases such as ni-results on the analyses of phenols produced in the manufacturing

trogen and helium as a carrier gas. The separation in GC is only de-process of the coal liquefaction,8 and surfactants and additives inpendent on an affinity between the stationary phase and an analytesoap.9 But GC with steam modifier was deficient in flexibility andbecause the affinity of the inert gas may be disregarded. Conse-

simplicity of the hardware and was left behind in the field of sepa-

quently, the use of GC column with different polarity has been ef-

ration science that was making rapid progress. Unfortunately, afective in improving the separation of the overlapped peaks.

clear explanation for the separation mechanism in GC with theIn the 1970 s, many researchers have examined several or-

modifier has not been made. Clarifying the interaction between

ganic and inorganic vapors in GC as the carrier gases aiming at di-analytes, modifier and stationary phase during the separation proc-

versity of manners in separation and extension of the GC applica-

ess would be of concerned in the revival of GC with modifier.

tions.1−7 The practical use of GC with modifier seemed to be not

Therefore, we developed a novel generator of modifier which con-

far ahead by various investigations in which the method of intro-sists of a simple structure and seams to be applicable for capillary

ducing the modifiers and the hyphenation to many detectors werecolumn GC.
In this paper we basically evaluate the prototype generator andincluded. In regard to the steam modifier, specially designed GC

Correspondence: Hiroshi Ito, Organic Analysis Lab., Toyota Central R&D Labs., Inc., 41−1 Nagakute, Nagakute−cho, Aichi 480−1192, Japan
Fax: +81−561−63−6448
E−mail: hiroshi-ito@mosk.tytlabs.co.jp―２５―

Chromatography, Vol.28 No.1 (2007)

investigate the elution behaviors of analytes by using various vola-[vinylpyrolidone−divinylbenzene], 50/80 mesh, Waters, Milford,

tile liquids as modifiers. Moreover, we report the fundamental in-MA, USA). It was washed with tetrahydrofuran and methanol, andvestigation on the behavior of formic acid modifier in the column

then was heated to dryness in a vacuum prior to packing them into

which indicates the largest change in retention of organic acid ana-

the stainless tubes (1/8 in. o.d. × 2.17 mm i.d. × 1 m).

lytes.
Determination of the modifier concentration in carrier gas
The concentration of formic acid modifier in the carrier gas

Experimental

was determined by ion chromatography (761 Compact IC,Generator of modifier
A schematic diagram of newly developed generator of modi-

Metrhom Shibata, Tokyo, Japan) equipped with photometric detec-

fier is shown in Fig.1. It has a direct ventilation structure and thetor (870−UV, Jasco, Hachiouji, Japan). The absorbed solution usedcarrier gas enters into the humidification chamber from gas line

here was 3 mM KOH in which the carrier gas was bubbled for 3

saturated with modifier vapor. Quartz or acrylic fiber was used asmin.

the absorbent of the modifier and was covered by the punching
Reagents

metal plate. A Teflon porous membrane film with a pore size of
1.2 µm set at the exit side prevented the contamination of theStraight-chained (C2−C7) fatty acids, their methyl esters, andmodifier particle. The generator has a duplex-tube structure and thetheir 1-amino derivatives were purchased from Wako Pure Chemi-

aqueous solution of 20% ethylene glycol was circulated throughcal (Osaka, Japan). Volumes of 2 µL of working solutions withthe overcoat tube to keep the humidification chamber temperature

concentration of 1 mg/mL of each compound were injected into theconstant. The amount of vaporous modifier generated was con-GC column. Other reagents were purchased from Wako.

trolled by the dilution rate of the vapor with the carrier gas or by
Results and discussion

the temperature of the humidification chamber.

Evaluation of the generator of modifier
There are many methods for generating the vaporous modifiers such as natural or heating vaporization, bubbling, supersonic atomization, nebulization and so on. In this work we chose themethod in which the vaporous modifier was vaporized in contact
with carrier gas. The generator can be directly placed in the carrier
gas line near the injector because its mechanical structure is simple.
Full contact between two phases is important for the constant
supply of the vaporous modifier. The acrylic fiber is well−ventilated and exhibits an excellent property for the absorption anddesorption of water. To prevent the leak of water drops from Teflon porous membrane film, a spatial layer between the absorbent
Figure 1. Schematic of the generator

and the membrane film in the introducer was necessary. In regard
to water supply, we have confirmed that the relative humidity inthe carrier gas was constant more than 8 hr after introduction.10

Gas chromatography
The GC instrument used was a GL Sciences (Tokyo, Japan)

Next, we monitored the concentration of formic acid in theGC 353 B equipped with a flame ionization detector (FID), whichcarrier gas. When using organic compounds as a modifier, quartz

included the prototype generator of modifier. To prevent the con-fiber which is chemically and physically stable was substituted as

densation of modifier vapor, the carrier gas line between the injec-

an absorbent instead of for the acrylic fiber. The column packed

tor and the generator was heated by a ribbon heater. Operating con-with Porapak R was preconditioned at 120 °C. The generator of

ditions were as follows: injector temperature, 280°C; detector tem-modifier containing 0.3 g of formic acid was kept at 0 °C with theperature, 320°C; column temperature, isothermally 80−200°C;

carrier gas flow of 50 mL He/min. The concentration of formic

carrier gas flow rate, 55.8 mL/min. The calculation of the retention

acid in 10 mL of absorbed solution was 450 ppm after 30 min, butfactor, k, was performed by using the column dead volume, V0,decreased slowly and became constant at 360 ppm 1 hr later. After-

which was obtained from the signal of a thermal conductivity de-wards, it was approximately constant of about 360 ppm until 6 hr.

tector on injecting air.

After the introduction of modifier, the baseline of FID was very
stable after 1 hr of introduction until 6 hr and the repeatability ofThe column packing material used here was Porapak R (poly―２６―

Chromatography, Vol.28 No.1 (2007)

Hiroshi Ito, Takashi Sugiura, Hidenori Nagai, Atsushi Murase,
Shuji Kodama, Yoshinori Inoue, Atsushi Yamamoto, Kazuichi Hayakawa

to the analysis of high boiling point fatty acids. Generally, fatty ac-the GC analysis was satisfactory.

ids are prone to give large tailing peaks in the medium polar colEffect of the modifier on the retention of analytes on Porapak R

umn. If formic acid introduced successively as a modifier was dis-column

tributed in the medium polar stationary phase, the modifier can

We investigated the effect of steam modifier on the retention

produce in the polar column and the retentions and peak shapes ofof analytes. The non-polar packing material, Gaskropak 54 [polyhigh boiling point fatty acids change steeply. Figure 4. shows themodifier.10

We

chromatograms of such compounds with and without formic acidchose the medium polar packing material, Porapak R, as the sta-modifier at column temperature of 200 °C. As for the n−valerictionary phase of this study in considering of the application to ana-acid (b.p.186 °C), the retention time became shorter (4.10 min)lytes with wide range of polarities and boiling points.

with the modifier. Moreover, it showed a clear peak. n−Caproic(styrene/divinylbenzene)], was unstable to steam

The retention behavior of acetic acid on the Porapak R col-acid (b.p. 205 °C) and n-heptanoic (b.p. 174 °C) acids, which wereumn was examined by introducing water, formic acid, lactic acidstrongly adsorbed on the polar stationary phase and not eluted

and propionic acid as modifiers. Figure 2 is van’t Hoff plot when

without modifier, could be eluted at 8.08 min and 16.00 min.

the temperature of generator was set at 40 °C except for formic acidof 20 °
C. The retention behavior of acetate was not affected bywater and was changed slightly by lactate and propionate modifiers. The change in retention time of acetate, however, became
larger when using formic acid modifier as the column temperature
decreased. In formic acid modifier, van’t Hoff plot of acetic acidshowed convex curve. The boiling point of formic acid calculated
from the pressure at the column entrance (760 + 678 mmHg) isabout 124 °C, and thus that in the column is in the region 100 to124 °C. Curved lines of van’t Hoff plot seem to be inflective in thisregion. It was estimated that formic acid modifier was promoted toself-associate below its boiling point and was adsorbed on a methacrylic acid site of the medium polar stationary phase, and as a result, the decrease of the stationary phase polarity caused the decrease in retention of the analyte.
Figure 2. van’t Hoff plot of acetic acid with various modifiers
Symbols: ● ; Without modifier,With modifier: □ ; Lactic acid, ▲ ; Propionic acid,■ ; Formic acid, ＊ ; H2O

To elucidate the change of retention behavior by formic acidmodifier, the thermal dependence for the retention of formic acidon the Porapak R was examined. A van’t Hoff plot of formic acidwas curved, and the curved line seemed to be composed by twokinds of straight lines (Fig. 3). The temperature at the intersectionof two regression lines was 107 °C, and it agreed with the boilingpoint of formic acid under pressurized state in the column. This result suggests that the distribution coefficient in the stationary phase
was changed at the boiling point of formic acid. Larger the slope ofthe regression line, higher the content of modifier in the stationary
phase depending on the hydrophilic interaction of formic acid. If
the polar site of the stationary phase was masked by polar interaction with the modifier, the polarity of the stationary phase might bedecreased, as a result, the retention of polar analytes also will rapidly decrease. This hypothesis explains the extreme decrease in retention of acetic acid observed in Fig. 2. On the other hand, thecontent of formic acid in the stationary phase above boiling point
was smaller as shown in Fig 3., and the association characteristics
of formic acid will be lost in this range.11

Figure 3. van’t Hoff plot for formic acid modifier

We applied the separation technique with formic acid modifier―２７―Chromatography, Vol.28 No.1 (2007)

a stationary phase on GC analysis. However, it was suggested thatthe presented approach could produce new stationary phases of GC
column which were created by using various vaporous modifiers
added to the inert carrier gas. Thus, any samples with various polarities might be analyzed by the unique column.
References[1]

Tsuda, T.; Tokoro N.; Ishii D.; J. Chromatogr., 1970, 46,[2]

Tsuda, T.; Yanagihara H.; Ishii D.; J. Chromatogr., 1974,241.
101, 95.
[3]

Tsuda, T.; Ichiba T.; Muramatsu H.; Ishii D.; J. Chroma-[4]

Wagaman K. L.; Smith T. G..; J. Chromatogr. Sci., 1971, 9,togr., 1977, 130, 87.
Figure 4. Separation of the high boiling point ingredient bymodifier of formic acid

241.

Conclusion[5]

Nonaka A.; Anal.Chem., 1972, 44 , 271.

[6]

Nonaka A.; Anal.Chem., 1973, 45, 483.

[7]

We developed a generator of vaporous modifier into the car-Guillemin C.L.; Martinz F.; Thiault S.; J. Chromatogr. Sci.,
1979, 17 , 677.

rier gas of GC. This equipment was evaluated by applying some
[8]modifier compounds to the polar packed column. It was recognizedNarita H.; Patent abstracts of Japan, Publication number S
60−161482.

that the elution of volatile acids was fastened by introducing the[9]modifiers. Formic acid modifier changed the elution behavior ofHatayama S.; Patent abstracts of Japan, Publication number
S 61−47559

analytes in the neighborhood of the boiling point of the modifier. It[10] H. Ito, Patent abstracts of Japan, Publication number H 18−

was presumed that the phase of modifier adsorbed on the packing

162097material changed at this temperature. Further examination of the ef-[11] Miyamoto S.; Nakamura Y.; Iwai Y.; Arai Y.; J. Chem. Eng.

fect of a modifier to the mobile phase on the retention mechanism

Data, 1999, 44, 48.

will be necessary. However, it is suggested that the present approach might be a significant technique in changing the property of―２８―