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442 Anal. Chem.

1982, 5 4 , 442-447

Separation of Organic Amine Compounds on Silica Gel with


Reversed-Phase Eluents

Brian A. Bldiingmeyer,* John K. Del Rios, and Jack Korpi


Waters Associates, 34 Maple Street, Mliford, Massachusetts 0 1757

Instead of trylng to eliminate the contrlbutlon to retention from surface silanols (17).However, it has been shown that if a
surface sllanol groups in bonded phase packlngs, It Is posslble “complete” coverage is obtained with the primary reaction,
to expiolt silanoi groups in a posltlve manner by uslng bare no additional bonding from a secondary reaction occurs and
silica gel as the adsorbent using aqueous eluents. Many this end capping does not reduce the concentration of surface
separatlons of llpophllic amlnes which are very dlfflcult on silanols (18). Nevertheless, amine-containingsample molecules
bonded reversed-phase packlngs are easily accomplished have badly tailed peaks when separated on most CI8 re-
wlth good peak symmetry on slllca. Retentlon of organlc versed-phase packings when using aqueous/organic eluents
amines on slllca uslng reversed-phase eluents appears to be containing an inorganic salt.
dependent upon electrostatic and adsorption forces. By use
Instead of vigorously trying to eliminate the surface silanol
contribution to retention of bonded columns, this paper
of lnorganlc or organlc amlnes as retentlon modifiers, sepa-
demonstrates that a better approach is to use bare silica gel
ratlons can be adjusted. AddHonally, lhls paper conflrms that
and exploit silanol interaction. Many sepaations of lipophilic
surface sllanols play a key role In determlning retention on amines which are unsatisfactory (excessive retention and
bonded phases probably through an electrostatlc contrlbutlon tailing) on CISreversed-phase packings can be satisfactorily
as could be expected from an lonlzed hydroxyl group existing, accomplished on silica using a routine reversed-phase eluent
In essence, as an ion-exchange slte. Excessive talling of of acetonitrile/aqueous salt solution. Silica packings behave
organlc amlnes on the reversed-phase packlng Is not due to quite nicely in this mode for the separation of organic amines.
the presence of sllanols but due to their Inaccessibility. This paper also confirms that surface silanols are a major
contributor to retention of organic amine compounds on
bonded CIS packings. However, contrary to popular belief,
The use of bare (unbonded) silica gel with aqueous eluents it is not just the presence of silanols that is important but the
for the separation of basic lipophilic amines may be more accessibility of them. Under the conditions used in this study,
appropriate than using the popular technique of reversed- the contribution of the silanols to retention of an organic
phase LC on bonded-phase packings. This is quite significant amine sample is mainly due to electrostatic attraction.
since the popular belief is that silica should not be used in
aqueous eluents. However, many separations of organic EXPERIMENTAL SECTION
amines which are very difficult and have poor peak symmetry Apparatus. The chromatographic system was a Waters As-
on bonded reversed-phase packings are easily accomplished sociates, Inc. (Milford, MA), Model 244 ALC which included a
with good peak symmetry on silica. Retention of the basic Model 6000A solvent delivery system, Model U6K injector, Model
amines on silica using reversed-phase eluents appears to be 440 absorbance detector, and a Model R401 refractometer. The
dependent upon electrostatic and adsorption forces. Sepa- columns were Radial-PAK C18and Silica (8 mm X 10 cm) car-
rations can be adjusted by using inorganic salts or organic tridges which were maintained under radial compression in the
amines as retention modifiers. radial compression separation system (Waters). The analog
outputs of the absorbance (W)and refractive index (RI) detectors
When using bonded-phase columns, many approaches to were recorded with a Model 730 data module (printer, plotter,
achieving a separation of ionic compounds have been dem- integrator) (Waters). The time equivalent (to)of the void volume
onstrated. Paired-ion chromatography has been used to obtain (Vo) was determined by injecting 5 PL of acetonitrile and
retention with hydrophilic amines (1-3). Ion suppression measuring the time from injection to the first deviation from the
which has worked well for separation of organic acids has had base line. This time was also confirmed by using a marker of
only limited application in separating lipophilic ammonium uracil. Times were converted to retention volumes using the flow
compounds (I). Generally, the separations which have been rate. The k’values were calculated automaticallyby programming
accomplished with ion suppression have been reported on the Model 730. Samples were injected automatically using the
PBondapak ClS (4).Other CISbonded-phase columns require Model 710B Waters intelligent sample processor (WISP). Au-
tomatic control of flow rate, injection, and data documentation
the addition of an organic amine modifier a t acidic pH to were accomplished by using the Model 720 system controller.
obtain good retention and peak symmetry with lipophilic Frontal analysis has been used to determine the ion-exchange
amine samples (1, 3-13). Another approach to obtain good capacity of bonded packings (19). This same approach is used
peak symmetry in the bonded-phase mode is to add an organic to determine the exchange capacity of silica. The column is first
ammonium compound as a mobile phase modifier at pH equilibrated with dilute hydrochloricacid and purged with water
values as high as 11(14,15). However, the basic pH will affect until no detectable acid elutes. Dilute sodium hydroxide is then
column life (14) as columns develop voids and channels in the introduced into the column. The breakthrough elution of the salt
bed structure due to dissolution of the packing, even those front is determined with a refractometer and confirmed by atomic
with good bonding and end capping (16). absorption analysis for sodium.
Since it is believed that the excessive retention and tailing Reagents and Mobile Phases. The salts used in this study
were purchased from Fisher Scientific (Fairlawn,NJ). Ammonium
of organic amines on bonded phases is due to interaction of hydroxide,sodium hydroxide, hydrochloric acid, and phosphoric
residual silanols on the packing, much effort has been directed acid were obtained from Baker Chemical Go. (Phillipsburg,NJ).
toward the chemistry of preparing a bonded phase where Purified water, obtained from a Milli-Q System (Millipore,
residual silanols are eliminated. Secondary bonding reactions, Bedford, MA), and a chromatographic grade acetonitrile and
after the primary reaction, have been claimed to reduce the methanol (Waters) were used for eluent preparation.

0003-2700/82/0354-0442$0 1.2510 0 1982 American Chemical Society


ANALYTICAL CHEMISTRY, VOL. 54, NO. 3, MARCH 1982 443

A. Name pKa Structure A.

Benzocaine 2.8 0COOC2H5

LidOEslne

Etidocaine

TstraCme

B. Name pKa Structure

HC- coon
Msieic Acid H
H c- coon

uuuL-
(I I* II >.

Figure 2. Effect of column surface upon retention or organic amines:


(A) separation of Radiai-PAK C,*; (B) separation on a column with
C. 50% coating level of Cia; (C) separation of Radial-PAK Silica. Mobile
Name pKa Structure phase is acetonitrile/water (60:40)with 4 mM dibasic ammonium
phosphate (pH 7.8). Sample components are (1) benzocaine, (2)
Ildocaine, (3) etiiocaine, and (4) tetracaine.

Use of halogen acids and salts is not recommended by equip-


ment manufacturers. However, NaCl was used to maintain
constant ionic strength during pH adjustment in part of this study
to avoid confounding the results. After each set of experiments,
the system was purged with water and at no time was NaCl
allowed to stagnate in the LC system. Similar precautions were
taken when using dilute HCl in the experiments to determine the
ion-exchange capacity of silica gel.
There were four probe mixtures used in this study. Mixture
A contained the anestheticsbenzocaine, lidocaine, etidocaine, and
tetracaine. The structures and their basic pK, values are shown
in Figure 1A. Mixture B contained maleic acid, chlorphenirmine,
D. and propranolol. These structures are shown in Figure 1B.
Name Name pKa
Mixture C contained phenylpropanolamine, quinidine, and pro-
Amllriplyllm 9.4 Dsaipramlnr 10.2 methazine. These structures are shown in Figure 1C. Mixture
D consists of the tricyclic antidepressants, amitriptyline, imipr-
amine, nortriptyline, desipramine, and protriptyline. These are
shown in Figure 1D. The antidepressant compounds were used
as probe molecules when evaluating organic amine modifiers. All
samples were pharmaceutical grade and from a variety of sources.
Imlpramlns 9.5 PlOtriplyiine 10
Samples were prepared at a concentration of 1 mg/mL and
dissolved in the mobile phase.
RESULTS AND DISCUSSION
Separation of Organic Amines on Silica Gel. Organic
amines have been traditionally separated on bonded phase
Nonriplyilne 9.7
columns using an inorganic salt in the eluent in order to reduce
the tailing which would occur if the salt were not present. An
example of such a separation is shown in Figure 2A for the
mixture A on a Radial-PAK C18 cartridge with dibasic am-
monium phosphate in the eluent. Tetracaine which is the
Flgure 1. Structures and basic pK, of compounds: (A) mixture A, (B) most basic amine was retained the longest with poor peak
mixture B, (C) mixture C, (D) rnixture D. symmetry. To understand the role of the surface silanols and
their contribution to retention, we analyzed these compounds
Mobile phases were filtered through either a 0.45-pm cellulose on a column which was purposely coated to half of the CI8level
acetate filter (HAWP 04700) or a 0.5-pm PTFE filter (FHUP of the original Radial-PAK Cl& This partially coated column
04700) (Millipore)and degassed before use in an ultrasonic bath had approximately 50% silanol content and the effect upon
with the solution under vaouum. retention and tailing would be revealing. If peak tailing is
444 ANALYTICAL CHEMISTRY, VOL. 54, NO. 3, MARCH l982

due solely to the silanol interaction, the peak symmetry should


be worse than on the fully coated column.
Figure 2B shows the separation on the column with
coating level of half of the column of Figure 2A. There was
no decrease in peak symmetry. In fact, retention of the
tetracaine (peak 4) increased slightly while the peak symmetry I/
2 3
improved. Thus, it appears that silanols do contribute to
retention but the amount of silanols is not the main cause of
peak asymmetry. The logical extension of this approach is
to separate the compounds on unbonded silica gel as shown
in Figure 2C. In this situation, peak symmetry is quite good
and retention is decreased.
6 18 30 42 mls.
The chromatograms in Figure 2 indicate that the surface
silanols by themselves are not deleterious to the retention of
organic amines when using organic, aqueous eluents containiig
an inorganic salt. In fact, silica gel itself appears to be the
preferred adsorbent for the separation of these bases. The
key to good peak symmetry is not the presence or absence of
residual silanols but more probably the accessibility of the
surface groups. If surface silanols are freely accessible, there
is good symmetry. The cause of tailing on the bonded Cla
packing may be due to the situation such that after the re-
action of the alkyl silane, the amount of active silanols as
compared to reactive silanols is increased (20). This could
result in an inhomogeneous distribution of active silanols on
the surface and produce discrete regions exhibiting strong 18 30 42 mls.
electrostatic interactions which could lead to peak tailing. Flgure 3. Retention of organic amines on Radial-PAK Silica: (A)
Additionally, it is feasible that the remaining silanols may be separation of mixture B, (1) maleic acid, (2) chlorpheniramine, and (3)
in micropores which are blocked by the presence of the thicker propranolol; (E) separatlon of mixture C, (1) promethazine, (2) phe-
bonded phase. Either or both of the described possibilities nylpropanolamine,and (3) quinidine. Mobile phase is acetonitriWwater
would affect mass transfer due to reduced accessibility of the (80:40) with 4 mM dibasic ammonium phosphate (pH 7.8).
surface silanols and result in the type of skewed peak which
is shown in Figure 2A. Regardless of why tailing occurs for a pH of 7.8 reduces the retention of all the amine samples.
Retention of benzocaine and maleic acid is not influenced by
organic amines on the CISbonded phases, lipophilic amines
exhibit minimal tailing on bare silica when an aqueous/or- ionic strength as these compounds are not positively charged
at the pH of the eluent. The interaction appears to be
ganic, inorganic salt eluent is used.
electrostatic as indicated by the large decrease in retention
The organic amines, quinidine and promethazine in mixture a'rising when the salt concentration is increased. This behavior
B, and chlorpheniramine and propranolol in mixture C are is typical of that reported in aqueous, ion-exchange systems
more lipophilic and basic (high pKa) than those compounds where retention of an ionic compound is reduced as the double
in mixture A and also have excessive retention and poor peak layer is reduced due to the increased ionic strength.
symmetry on the fully coated Radial-PAK CIS column in an Effect of pH. If the retention is dependent upon ion
eluent of acetonitrileaqueous dibasic ammonium phosphate exchange, pH should also contribute to retention. Since there
(60140). With these more lipophilic bases, addition of higher is a large contribution to retention from ionic strength, it is
salt concentration only slightly improved the separation on necessary to maintain a constant ionic strength when inves-
the bonded phase CIS column. Retention volume was of the tigating the effect of pH. This means that salts which are
order of several hundred milliliters. The best separation for buffers such as dibasic ammonium phosphate cannot be used
mixtures B and C was also on the unbonded silica column. and, therefore, sodium chloride was used with appropriate
The separations are shown in Figure 3. precautions. Figure 5 shows the effect of pH upon the organic
It is interesting to note that the order of retention on the amine samples. For all the ionic samples, as pH is increased,
silica column is the same as that on the Radial-PAK CIS retention of the organic amines is increased to a maximum
column for most of these samples. Furthermore, lidocaine and value after which retention of the sample decreases. Those
etidocaine, compounds with the same p K , elute with differing samples which are not ionic over the pH region studied are
retention. Promethazine and phenylpropanolamine have the not influenced by pH.
same pKa and also elute with differing retention. Quinidine, An explanation of this behavior can be made in terms of
whose pKa is less than promethazine and phenylpropanol- the effect of pH upon the ionization of surface silanols and
amine, elutes as a longer retention volume than either of them. the effect upon the ionic character of the sample. The pK,
Therefore, the retention of the amines must be due to an of silica is reported to be about 6.5 (21),so as pH goes from
electrostatic attraction and an adsorptive interaction (affinity 3 to 7.5 the ionic nature of the surface increases. For samples
to the stationary phase). Furthermore, the silica seems to be with pKas of 8-8.5, the compounds are ionic at pH 6 and
behaving in a "reversed-phase" mode when using aqueous/ nonionic at pH 10. This combination of the surface becoming
organic mobile phases. ionic at pH 3-7.5 and the samples decreasing in ionic nature
Influence of Ionic Strength. The retention of organic from pH 6 to 10 explains the behavior seen in Figure 4A of
amines using reversed-phase solvents on silica columns results increased retention until pH 6 and declining retention after
in good peak symmetry. The retention could be due to two that. For the samples with pKa values around 9-10, the same
contributions, ion exchange and/or adsorption. To elucidate two effects are occurring. As pH is increased from 3 to 8 the
the role of an ion-exchange interaction, one needs to define surface hydroxyls which are ionized (21) participate in ionic
the influence of salt concentration upon retention at concltant attraction toward the samples and the retention of the ionic
pH. Figure 4 shows that increasing the salt concentration at amines is increased. From pH 8 to 12, there is a decrease in
ANALYTICAL CHEMISTRY, VOL. 54, NO. 3, MARCH 1982 445

A. 150 r A.

3 4 5 6 7 8 9 1 0
PH

3 4 5 6 7 8 9 10
PH
C.

3 4 5 6 7 8 9 1 0
PH

Flgure 5. Effect of pH upon retention: (A) compounds are (1) ben-


zocaine, (2) lidocaine, (3) etidocaine, and (4) tetracaine; (B) compounds
are (1) maleic acid, (2) chlorpheniramine, and (3) propranolol; (C)
compounds are (1) promethezine, (2) phenylpropanolamine, and (3)
quinidine. Mobile phase is acetonitriWwater (60:40) with 4 mM sodium
0.1 1.o 10.0 100.0 chloride. The pH is the value of the aqueous stock solution containing
(NHq)2HPOq3@!
sodium chloride before the mobile phase is prepared. Column is
Flgure 4. Effect of ionic strength upon retention: (A) compounds are Radial-PAK Silica.
(1) benzocaine, (2) lidocaine, (3) etidocaine, and (4) tetracaine: (B)
compounds are (1) maleic acid, (2) chlorphenlramine, and (3) propra- surements below 30% acetonitrile were too difficult to in-
nolol; (C) compounds are (1) promethazine, (2) chlorpheniramine, and terpret because of excessively retained and broad peaks. The
(3) quinidine. Mobile phase is acetonitrile/water (pH 7.8) (60:40). relationship of sample retention with solvent composition
Column is Radial-PAK Silica. which occurs on bare silica is typical of that observed when
the ionic (positive) character of the samples and this reduces using bonded phases in the same type of eluent. Because of
the ionic interaction and decreases retention. This is what this similarity it can be concluded that the silica is a nonpolar
is indicated in Figure 4H,C. surface with respect to the aqueous/organic eluent. In the
Ion-Exchange Capacity. Silica is known to exhibit ion- terms of traditional chromatographicnomenclature, the silica
exchange capacity (22). Additionally, it has been reported gel is retaining organic amines in a reversed-phase mode. This
that bonded CISreversed-phase packings have ion-exchange seemingly anomalous behavior may in fact not be, since it is
capacity (19). Measurement of the ion-exchange capacity of known that siloxane groups on the silica are hydrophobic (21,
the silica column used in this study using frontal analysis 22). Thus, the situation can be viewed as a sample interacting
resulted in a value of 3 mequiv/column which is consistent with a surface consisting of silanols which are ionic and hy-
with the effect of pH and ionic strength upon retention. drophilic and of siloxane groups which are hydrophobic. In
Effect of Solvent Strength upon Retention. Evidence essence, the surface is more nonpolar than the eluent, so, with
is strong that the retention of organic amines on silica depends constant pH and ionic strength, typical reversed-phase be-
upon electrostatic and adsorption forces. To investigate the havior is exhibited. The mechanism of retention may be
adsorption contribution, we varied the acetonitrile/water ratio similar to the ion-interaction model for reversed-phase ion-pair
while maintaining constant pH and ionic concentration. The liquid chromatography where retention is also proposed to
effect of eluent concentmtion is shown in Figure 6. As the be due to electrostatic and adsorptive forces (1, 3). In the
aqueous portion of the eluent increases, the retention of the ion-interaction model, charge is dynamic, while on the silica
organic amines increases. The observed linear relationship surface, charge is static.
of a plot of log k' vs. solvent composition is typical for re- Reversed-phase behavior of silica has been shown for ni-
versed-phase systems. The concentrations of acetonitrile trobenzene, resorcinol, and crown ethers (23,24). However,
above 70% were not investigated because the concentration data for crown ethers had a minimum in retention at 0.4
of buffer salt was not soluble in the eluent. Retention mea- volume fraction of water in methanol. This is contrasted to
446 ANALYTICAL CHEMISTRY, VOL. 54, NO. 3, MARCH 1982
A. 1
100.0

k 10.0

1.o
0.3 0.4 0.5 0.6 0.7
Volume Fraclion of Water

u , z
6 12 I 8 mls.
Flgure 7. Separation of tricyclic antidepressantson Radial-PAK Silica.

-
Compounds are (1) amitryptiiine, (2) imipramine, (3) nortryptiiine, (4)
desiprimine, and (5) protryptliine. Mobile Dhase is methanol/water
(80:20) with 5 mM butylamine.
t
1.0
0.3 0.4 0.5 0.6 0.7
volume Fraction of Water

,"
k C

1.0 -0.3 0.4 0.5 0.1) 0.1


Volume Fraction 01 Water
Flgure 8. Effect of solvent strength upon retention with constant pH
6 7 8
PH
9 OI 11

Flgure 8. Effect of pH upon retention of tricyclic antidepressants.


Compounds are (1) amitryptiiine, (2)imipramine, (3) nortryptiiine, (4)
and ionic strength: (A) compounds are lidocaine (2), etidocaine (3),
tetracaine (4). and benzocaine (l),always eluted near V , and is not desiprimine, and (5)protryptiiine. Column is Radial-PAK Silica. Mobile
shown: (B) compounds are chiorpheniramine (2), propranolol (3),and phase is methanol/water (80:20)with 5 mM butylamine. The pH which
maleic acid (l),always eluted near V,, and is not shown: (C) com- is plotted is the value of the aqueous stock solution containing bu-
pounds are promethazine (l),chlorpheniramlne (2),and quinidine (3). tylamine before the mobile phase is prepared.
Mobile phase Is acetonitrile/water with 4 mM dibasic ammonium
phosphate at pH of 7.8. Column Is Radial-PAK Silica. lationship of log k 'vs. butylamine concentrationwas observed
over a concentration range of 5 to 20 mM in a methanol/water
the trend observed for the organic amines where the behavior (80:20) eluent. This is similar to the effect of ionic strength
is more typical of reversed phase. upon retention shown in Figure 4 which resulted from adding
Analysis of Tricyclic Antidepressants. Tricyclic an- dibasic ammonium phosphate to the eluent used to separate
tidepressants are a class of organic amines used for the the other amines.
therapeutic management of depression. Various methods for The effect of pH upon the retention of the tricyclics is
the measurement of these compounds have been proposed. shown in Figure 8 and is different than that observed using
Liquid chromatography has advantages in accuracy and the inorganic salt. At a pH of 10, the samples should be
precision, specificity, and convenience (25). Chromatography partially ionized so that as pH is decreased, there should be
has been on bonded phases in the reversed-phase mode (5, an increase in retention due to increased electrostatic at-
15,26,27) and on silica in the normal-phase mode (28,29). traction of the sample to the silica surface. This is what is
One paper utilized methanol with 0.2% propylamine as the observed with pH values 11to 8 and is similar to that observed
eluent on a silica column (30). Substitution of butylamine when using an inorganic salt. However, when going from pH
for the propyl analogue in this system resulted in a good 8 to 3 using the butylamine, a reversal in the elution order
separation (31). However, neither author reported how the occurred for three members of this group.
separation would behave as a reversed-phase system. As was demonstrated earlier (Figure 5 ) when using the
The similarity of the butylamine/methanol eluent to the inorganic salt as a modifier, retention of organic amines of
organic-buffer salt eluent used in this work indicated that the high pK, decreased when going from pH 8 to 3 due to reduced
tricyclic separation might be enhanced using a reversed-phase ionic attraction caused by protonization of the surface silanol;
eluent on silica. Indeed, the separation was improved with this trend was not observed in this situation. Because of this
the best peak symmetry and retention times resulting from reversal in retention of these amines, it is probable that the
an eluent of methanol/water (8020) with 5 mM butylamine butylamine is actually modifying the silica column and con-
ds shown in Figure 7. tributing to retention in a way other than simply competing
As the water concentrationratio was increased (maintaining for the ionic sites on the surface.
the concentration of butylamine at 5 mM) the retention ex- Three additional amines, urea, dicyclohexylurea,and am-
pressed by log k' increased linearly. This is the same type monia (ammonium hydroxide), were used as modifiers at a
of behavior which was observed previously in Figure 6. concentration of 5 mM in the methanol/water (80:20) eluent
The effect of increasing the concentration of butylamine at a pH of 11. The retention of the tricyclics varied depending
was to decrease the retention of the tricyclics. A linear re- upon which amine modifier was used. The effect of the
Anal. Chem. 1902, 5 4 , 447-450 447

modifier as expressed by the retention of the tricyclic anti- (10) Baker, J. K. Anal. Chem. 1979, 51, 1693.
depressants was in the following order: ammonia (longest (11) Knox, J. H.; Jurand, J. J . Chromatogr. 1977, 142, 651.
(12) Kikta, E. J.; Grushka, E. Anal. Chem. 1978, 48, 1098.
retention) > urea > butylamine > dicyclohexylurea. The type (13) Kirkland, J. J. Chromatographla 1975, 8, 661.
of amine made a large difference in retention which was more (14) Wehrll, A.; Hildenbrand, J. C.; Keller, H. P.; Stampfli, R. J . Chroma-
fogr. 1978, 149, 199.
than would be expected from only a reduction in ionic at- (15) Kraak, J. C.; Bljster, P. J . Chromatogr. 1977, 143, 499.
traction to the ion-exchange site. This is additional evidence (16) Rabel, F. M. J . Chromatogr. Scl. 1980, 18, 394.
that the organic amine is adsorbed and changes the surface (17) Cooke, N. H.; Olsen, K. J . Chromatogr. Sci. 1981, 18, 512.
(18) Berendsen, 0. E.; Pikaart, K. A,; DeGalan, L. J . Liq. Chromatogr.
character. 1980, 3 , 1437.
(19) Thurman, E. M. J . Chromatogr. 1979, 185, 625.
ACKNOWLEDGMENT (20) Snyder, L. R. “Prlnciples of Adsorption Chromatography”; Marcel Dek-
ker: New York, 1968; pp 155-182.
The authors thank I,. Jaros and J. Balderson for help in (21) Iler, R. K. “The Chemlstry of Sllica”; Wiley: New York, 1978; p 660.
the preparation of this manuscript, B. Murphy for technical (22) Unger, K. “Porous Silica”; Elsevier: Amsterdam, 1979; pp 130-141.
assistance, and U. N e w and P. McDonald for helpful dis- (23) Nahum, A.; Horvath, C. J . Chromatogr. 1981, 203, 53.
(24) BIj, K. E.; Horvath, C.; Melander, W. R.; Nahum, A. J . Chromatogr.
cussions. 1981, 203, 85.
(25) Scogglns, B. A.; Maguire, K. P.; Norman, T. R.; Burrows, G. D. Clin.
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(1) Bldllngmeyer, B. A. J . Chromatogr. Scl. 1980. 18, 525. 143, 535.
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(5) Twitchett, P. S.; Moffat, A. C. ./. Chromatogr. 1975, 111, 149. ston-Salem, N.C.) 1978, 24, 87.
(6) Wahlund, K. G.; Sokolowski, A. J . Chromafogr. 1978, 151, 299. (30) Kuss, Von H. J.; Nathman, M. Drug Res. 1978, 28, 1301.
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. , Proelas. H. F.: Lohmann. H. J.: Miles. D. G. Clln. Chem. (Winston-&-
18)
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man, E. J . Chromafogr. 1977, 142, 715. 1981.

Liquid-Phase Photoionization Detector for Liquid


Chromatography
David C. Locke,” Bhim S. Dhingra, and A. D. Baker
Department of Chemlstty, City University of New York, Queens College, Flushing, New York 11367

A direct liquid-phase photoionlratlon detector with picogram molecules with a-electron systems typically have ionization
sensitivity to polycyclic aromatic hydrocarbons eluted with potentials (IP)of the order of 9 eV and show a sharp rise in
hexaneI2-propanol from slllca columns Is described. Llght photocurrent for photon energies slightly larger than the IP.
from a mlcrowave-excited continuum xenon source Irradiates In condensed phases, however, a steep threshold for pho-
the HPLC effluent In a 10-pL flow cell. The amplified photo- toionization does not usually exist (3))and because of sta-
currents are llnear with concentratlon over at least 6 orders bilization of the positive ion and photoelectron formed, by
of magnltude. No signal is observed for phenols or chlorinated solvation, r-electron photocurrents are observed for incident
compounds, although nanogram quantltles of various 0- and photon energies typically 2-4 eV less than the vacuum IP value
N-containing compounds are detectable. (4).Molecules with electrons only, such as alkanes, have
vacuum IP values 1-2 eV higher than those for a-electrons
and a smaller lowering by solvation of the liquid-phase ion-
ization threshold.
Although a multitude of detectors has been developed for Typical HPLC eluents have vacuum IP values reported (2)
HPLC (I),there is yet to be invented a universal and sensitive as follows: tetrahydrofuran, 9.45 eV; 2-propanol, 10.09 eV;
detector. In attempting t o develop such a detector, we have ethyl acetate, 10.15 eV; n-hexane, 10.18 eV; methanol, 10.85
investigated techniques based on photoionization. In a pre- eV; chloroform, 11.42 eV; acetonitrile, 12.20 eV; and water,
vious publication (2))a photoionization detector (PID) was 12.59 eV. Resonance lines from low pressure (0.1 torr) rare
described in which resonance radiation from xenon or krypton gas discharge lamps are characteristic of the filling gas: Xe
discharges, or Lyman a radiation from a hydrogen/helium emits a resonance line of wavelength 1469.6 A, equivalent to
lamp, was used to irradiate the flash-evaporatedeffluent from photon energy of 8.44 eV; Kr, 1235.8 %.,10.03 eV; Ar, 1048.2
a HPLC column. To circumvent the problem of effluent A, 11.83 eV; Ne, 735.9 A, 16.85 eV; and He, 584.3 A, 21.22 eV
evaporation, we now report on a PID based on the direct (5, 6). Thus in the liquid phase, to avoid eluent ionization,
irradiation of the liquid e€fluent stream. Measurement of the resonance lines cannot generally be used as the source of
resulting solute photocurrent allows detection of picogram ionizing radiation. At higher gas pressures, however, micro-
quantities of certain polycyclic aromatic hydrocarbons and wave- and RF-excited discharges in rare gases emit continua,
various other substances eluted from normal-phase (silica) which for example in the case of xenon at 200 torr has a fairly
HPLC columns with hexane/2-propanol mobile phases. sharp low wavelength cutoff at 1600 A, corresponding to 7.81
Photoionization in liquids is a more complex phenomenon eV (7). This is insufficient to ionize liquid n-hexane,for which
than that in gases. In the gas phase at low pressures, organic the photoionization threshold is observed at 9.59 eV (8))but
0003-2700/82/0354-0447$01.25/0 0 1982 American Chemical Soclety