Gas Chromatographic Separation of Some Unsaturated Hydrocarbons Using Thallium(1) Tetraphenylborate as Stationary Phase George E. Baiulescu Department of Analytical Chemistry, University of Bucharest, Romania
Vasile A. llie Institute of Chemical Research, Bucharest, Romania
Several stationary phases are available for the gas chromatographic separation of olefins, particularly those of the terpene series (1, 2); but the best and most commonly used, silver nitrate in diethylene glycol or ethylene glycol (3, 4 ) has considerable drawbacks. Silver ion is rapidly reduced above 65 "C to the detriment of column performance; even a t 40 "C, an operating life of only a few days is often obtained, and the column behavior is irreproducible. A t 25 OC, the column is stable for long periods and gives reproducible separation (3, 4 ) but the resulting long retention times for terpenes and other high boiling olefins are inconvenient; attempts t o improve the situation by reducing the proportion of silver nitrate in the stationary phase give columns with low capacity that are unsuitable for preparative-scale operation. Several attempts were made to separate some unsaturated hydrocarbons on lithium caproate ( 5 ) as well as on transition-metal complexes (6). The capacity of Tl(1) in thallium nitrate to interact selectively as the silver ion in silver nitrate with unsaturated hydrocarbons is reported in ( 7 ,8 ) . The present paper studies the behavior as stationary phases of the thallium tetraphenylborate compared with that of sodium in the separation of a series of olefins and aromatic hydrocarbons. The use of sodium, potassium, rubidium, cesium, and thallium tetraphenylborates as stationary phases in gas chromatography was previously mentioned (9). EXPERIMENTAL The preparation of thallium(1) tetraphenylborate was made by precipitation from a concentrated solution of thallium(1) sulfate (Riedel de Haen A.G.) with a diluted solution of sodium tetraphenylborate (Merck), followed by filtration and washing of the precipitate with distilled water until the thallium ions did not react. The precipitate was dried in the room atmosphere. The thermograms of sodium and thallium tetraphenylborates (9) show a good thermal stability, the loss of substance being observed up to 200 O C . Measurements were carried out on a gas chromatograph Fractovap Model C (Carlo-Erba, Milano), with a flame ionization detector. Nitrogen was used as carrier gas. The solution peaks were recorded using a Speedoniax G 0-1 mV (Leeds & Northrup). The different components were introduced in the chromatographic column using a Terumo microsyringe of 1 ~ 1 Each . component was (1) (2) (3) (4) (5) (6)
R. A.
Bernhard. Anal. Chem., 34, 1576 (1962).
M. H. Klouwen and R. J. Heide, J. Chromatogr.,7 , 297 (1962). J. Heriing, J. Shabtai, and E. Gil-Av. J. Chromatogr.,6 , 349 (1962). E . Smith and R . Ohlson, Acta. Chem. Scand., 16, 351 (1962). M. Muhlstadt. Chem. Ber., 93, 2638 (1960). G. P. Cartoni et a/., "Gas Chromatography 1960." R. P. W. Scott, Ed..
Butterworths, London, 1960, p 273-93. (7) B. T. Guran and L. B. Rogers, J. Gas. Cbromafogr.,3, 269 (1965). (8) D. V. Banthorpe, C. Gatford, and B. R. Hollebone, J. Gas. Chromatogr., 6, 61 (1968). (9) G. E. Baiulescu and V. A. Hie. Anal. Chem.,44, 1490 (1972)
separately introduced (0.1 PI) in the chromatographic column. All the chemical compounds used as standards were of chromatographic purity. Phenylacetylene, phenyl tert- butylacetylene and benzo-tert- butylcyclobutene were synthetized and identified by means of elemental analysis, IR spectra, and NMR measurements. The olefin retention was studied using chromatographic columns of aluminum with internal diameter of 4 mm and a length of 6 m. The columns were filled with thallium and sodium tetraphenylborates deposited in the proportion of 25% on hexamethyl-silanized Chromosorb W 30-60 mesh. The retention data for aromatic hydrocarbons were determined on thermostable glass columns with internal diameter of 3.6 mm and a length of 1.4 m. The stationary phases were solved in acetone and deposited in proportion of 15% on hexamethyl-silanized Chromosorb W 60-80 mesh. The chromatographic columns were filled by tapping and conditioned in a carrier gas flow a t 180 "C for 12 hr.
RESULTS AND DISCUSSIONS
Table I gives the relative retentions, calculated against n-pentane a t 58 "C, of several olefins on columns with sodium and thallium tetraphenylborates. Table I also gives the relative retentions of some aliphatic hydrocarbons having boiling points (IO)close to those of the olefins. The similar relative retentions of olefins and saturated hydrocarbons having close boiling points, obtained with sodium tetraphenylborate show the lack of noticeable interaction between the stationary phase and the components. The olefins and especially diene hydrocarbons are strongly retained on thallium(1) tetraphenylborate. This interaction is due to thallium(1) ions having low-lying dorbitals which can form partial covalent bonds with the r-orbitals of the adsorbed molecules. Cyclopentadiene does not elute from the column of thallium tetraphenylborate under our conditions (column temperature 58 "C, carrier gas flow 20.1 ml/min). Table I1 gives the relative retentions of some aromatic hydrocarbons and their derivatives, calculated referred to n-decane a t 129 "C. The substances elute generally on the column of sodium tetraphenylborate in the increasing order of their boiling points. Higher differences in the relative retentions of the aromatic hydrocarbons on the column of thallium tetraphenylborate comparative to that of sodium, should be due to the thallium(1) ions which are able to interact with the a-orbitals of the aromatic hydrocarbons. The presence of an ethinyl radical in the phenylacetylene causes the lowering of the aromatic character of the molecule which explains its relative retention, practically the same on both chromatographic columns. The introduction of a tert-butyl radical in the molecule of phenylacetylene causes differences between the relative retentions on the chromatographic columns. (10)
"Dictionary of Organic Compounds," I. Heilbron and H. M. Bunbury, Ed., Russian ed., Izd. I. L.. Moscow, 1949.
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Table I. Relative Retentions (Referred to n-Pentane at 58 "C) of Some Olefins on Columns with NaB (CeHs) and TIB (CsH5)4 Compound
Boiling point, OC
1,3-Butadiene n-Butane 2-MethylbutadieneL3 n-Pentane cis-2-Pentene 2-Methyl-2-butene Cyclopentadiene 4-Methyl-truns-2pentene 2,3-Dimethyl-1,3butadiene n-Hexane
NaB(CoH6)r
-2.6 -0.5 35 .O
0.35 0.29 1.10
36.0 37.0 38.6 41-42 53.9
1.oo
1.14 1.20 1.36 2.18
2.35
69-70
3.29
6.00
68.9
3.15
3 .OO
Benzene Cyclohexane Toluene E thylbenzene o-X ylene m-Xylene p-Xylene Styrene Phen ylacetylene Isopropyl benzene Mesitylene tert-Butylbenzene l-Methyl-4isopropylbenzene n-Decane Naphthalene Phenyl-tertbut ylacetylene Benzo-tert-butylcyclobutene p-Di-tert-butylbenzene n-Tridecane 1-Methylnaphthalene 2-Methylnaphthalene Hexamethylbenzene Diphenyl n-Pentaclecane
-
Boiling point, OC
80.08 81 .O
110.6 135.5 144.1 139.3 138.0 145.0 142-144 152 .O 164.8 168.0 177.0
Compound
l-Methyl-4isopropyl benzene Naphthalene Phenyl-tertbutylacetylene Benzo-tertbutylcyclobutene p-Di-tert-butylbenzene 1-Methylnaphthalene n-Decane n-Pentadecane
0.65 0.31 1.76 1.oo
1.24 1.38 -
a
NaB(C,H,),
TlB(C,H,),
0.06 0.06 0.24 0.43 0.47 0.46 0.46 0.47
0.18 0.06 0.41 0.70 0.94 0.83 0.76 0.75 0.60 1.01 1.57 1.59 1.71
0.56
0.57 0.86 0.91 1.11 1.oo
1.oo
174.0 218 .O 72'12 mm.
3.74 2.56
5.98 8.59
57'/1 mm.
1.90
3.77
236.5
6.69
17.23
234 .O 241 .O
5.90 5.65
7.66 10.30
240-242
6.74
10.76
264.0
9.04
18.69
14.93 20.81
17.38 32.20
254-255 270.5
AS
TLB(CEHS) 1
Table 11. Relative Retentions (Referred to n-Decane at 129 "C) on Columns with NaB(CsH5)4 and TlB (CgHj) Compound
Table IV. Entropy Changes for Selected Compounds, Calculated Using log K Values at 129 "Ca AS/AH
NaB(C6Hd4
TlB(CaHa)r
-27.2
-38.5
2.65 2.45
-29.9 -29.9
-33.3 -42.8
2.54 2.37 2.56 2.38
-27.2
-38.0
2.60 2.41
-35.3 -32.1 -29.2 -41.8
-45.8 -38.1 -32.3 -43.9
2.49 2.51 2.64 2.43
-
Na
T1
2.36 2.36 2.48 2.33
The entropy changes are in units of cal/deg. mole.
Table 111 gives the logarithms of the capacity ratios and the adsorption heats of some compounds on sodium and thallium tetraphenylborates. The adsorption heats were calculated from the slope of the plotted straight lines log K us. 1/T X104. The adsorption heat of n-pentadecane on thallium tetraphenylborate was estimated for log K a t 156 "C = 0.900. The straight line slopes were calculated by means of the least squares method. The standard error of the slope has been estimated using the regression analysis. The adsorption heat values of various components on thallium tetraphenylborate were higher than those of the same components on a column of sodium tetraphenylborate. The differences were significant especially in the case of the aromatic hydrocarbons and their derivatives, which may be explained by the interaction ability of Tl(1) ion with these components. Relative changes in entropies of adsorption were calculated assuming that the capacity ratios were directly proportional to the thermodynamic adsorption constant Ka (11) in order to make relative, but not absolute comparisons (Table IV). The relative entropy changes for the selected compounds are higher on the column of thallium tetraphenylborate compared to that of sodium, the highest differences being observed with the aromatic compounds. The ratios A S / A H are lower on thallium tetraphenylborate than on the sodium tetraphenylborate and decrease for the superior homologs implying that heats of adsorption increased more rapidly than the corresponding values for entropies. (1 1) J. E. Heveran and L. B. Rogers, J. Chromafogr.,25, 213 (1966).
Table 111. Log Capacity Ratios for Some Compounds Eluted from NaB(CsH5)(,TIB(C&)4 Columns and Their Heats of Adsorption, Kcal/mole TlB (C6%)
NaB (CsHa)r log K ,
Compound
106'C
1-Methyl-4-isopropylbenzene
-0.435 0.121
Naphthalene Phenyl-tert-butylacetylene benzo-tert-butylcyclobutene
p-Di-tert-butylbenzene 1-Methylnaphthalene n-Decane n-Pentadecane 1848
log K , 129OC
-0.814 -0,288 -0.453 -0.050 -0.219 -0,584 0.432 -0.036 0.362 -0,109 -0.469 -0.861 0.457 1.061
log K , 146OC
-0.996 -0,527 -0.690 -0.794 -0.348 -0.338 - 1,075 0.119
-AH
10.28 i 0.87 11.79 f 0.41 11.65 i 0.37 10.46 f 0.41 14.17 i. 0.14 12.80 f 1.05 11.06 i. 0.59 17.18 i. 0.82
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log K ,
4
logK, 106OC
log K , 146'C
129'C
0.763 1.192 1.492 1.083 1.883 1.578 0.435 ...
0.256 -0,104 0.409 0.799 0.493 0.956 0.207 0.599 0.809 1.259 0.689 1.035 0,022 -0.287 1.530 1.087
-AH
15.68 zkO.37 14.03 f 1.55 17.96 f 1.32 15.77 + 0.87 19.42 =t0.46 16.13 f 0.18 13.02 f 0.46 18.87 f 1.07
CONCLUSIONS The present paper shows that the olefins and aromatic hydrocarbons are selectively retained on columns of thallium tetraphenylborate compared t o those of sodium tetraphenylborate. The stronger retention of these compounds on the column of thallium tetraphenylborate should be due
to the possible interaction between thallium d-orbitals and the ir-orbitals of unsaturated hydrocarbons. Therefore, the thallium tetraphenylborate may be used as selective stationary phase for the separation of hydrocarbons. RECEIVEDfor review January 28, 1974. Accepted June 4, 1974.
Determination of Long Chain Fatty Acids as 2-Naphthacyl Esters by High Pressure Liquid Chromatography and Mass Spectrometry M. J. Cooper and M. W. Anders Department of Pharmacology, University of Minnesota, Minneapolis, Minn. 55455
Long chain polyunsaturated fatty acids have assumed increased importance in recent years; in particular, some CZOacids are known precursors of the biologically import a n t prostaglandins. A number of analytical methods have been employed for the determination of fatty acids. For example, microdiffusion and gas chromatography have been used for the analysis of short and medium chain fatty acids (1) and Umeh (2) described the gas chromatographic analysis of C ~ - C ~acids O as their p - bromophenacyl and p - phenylphenacyl esters. Ehrsson ( 3 ) reported the gas chromatographic detection of C4-C10 acids as their pentafluorobenzyl derivatives. Recently, the gas chromatography-mass spectrometry of C1-C'o acids as their benzyl esters was reported ( 4 ) . Mike5 et al. ( 5 ) used high pressure liquid chromatography (HPLC) with complex-forming stationary phases for CIS fatty acid methyl esters. Grunert and Bassler (6) reported the gas chromatographic analysis of fatty acid methyl esters. Derivatives have been widely employed in gas chromatographic analysis to improve chromatographic properties and to increase the sensitivity of detection. I t is apparent, however, that derivatives should be useful in liquid chromatographic analysis for the same purposes. Henry et al. ( 7 ) and Papa and Turner ( 8 ) have employed 2,4-dinitrophenylhydrazone derivatives for the HPLC analysis of various carbonyl compounds and Fitzpatrick and Siggia (9) described the preparation and analysis of benzoate derivatives of hydroxy steroids. Fluorigenic labeling has also been employed in HPLC analysis ( I O , 11).Dunham and Anders (12) reported the base-catalyzed conversion of prostaglandin E to B to enhance the sensitivity of detection for HPLC analysis.
(4) (5) (6) (7) (8) (9) (10) ( 11) (12)
J. W. A. Meijer and L. Hessing-Brand, Clin. Chim. Act. 43, 215 (1973). E. 0. Umeh, J. Chromatogr., 56, 29 (1971). H. Ehrsson, Acta Pharrn. Suecica, 8, 113 (1971). U. Hintze. H. Roper, and G. Gercken, J. Chromafogr., 87, 481 (1973). F. Mike;, V . Schurig, and E. Gil-Av, J. Chromatogr., 83, 91 (1973). A. Grunert and K. H. Bassler, Fresenius' Z. Anal. Chem., 267, 342 (1973). R. A. Henry, J. A. Schmit, and J. F. Dieckman, J. Chromafogr. Sci., 9, 513 11971). L. J. Papa and L. P . Turner, J. Chromatogr. Sci., 10, 747 (1972). F. A. Fitzpatrick and S . Siggia, Anal, Chem., 45, 2310 (1973). R. W. Frei and J. F. Lawrence, J. Chromatogr., 83, 321 (1973). R. W. Frei, J. F. Lawrence, J. Hope, and R. M. Cassidy. J. Chromatogr. Sci., 12, 40 (1974). E. W. Dunham and M. W. Anders. Prostaglandins, 4, 85 (1973).
We report here the HPLC analysis of unsaturated CIS and CZOfatty acids. Since the methylene interrupted polyunsaturated acids show no specific ultraviolet absorption, the 2-naphthacyl esters were prepared. The structures of the derivatives were confirmed by both electron impact (EI) and chemical ionization (CI) mass spectrometry (MS).
EXPERIMENTAL Reagents. Reagent grade solvents were used without further purification. 8-Naphthacyl bromide (cu-bromo-2'-acetonaphthone) and N,N- diisopropylethylamine were obtained from Aldrich Chemical Company. Dihomo-y-linolenic acid was obtained from the Hormel Institute, and oleic acid from Calbiochem; all other fatty acids were purchased from Sigma Chemical Company. Derivatization Procedure. The acid (10 pmoles), 2-naphthacyl bromide (20 pmoles), and N,N-diisopropylethylamine (40 pmoles) were dissolved in 1 ml of dimethylformamide. The reaction mixture was heated a t 60' for 10 minutes, a t which time the reaction was complete. An aliquot of the reaction mixture was injected into the liquid chromatograph. For the MS analyses, the esters were prepared on a larger scale. Acid (100 pmoles), 8-naphthacyl bromide (92 pmoles), and N,Ndiisopropylethylamine (184 pmoles) were dissolved in 10 ml of dimethylformamide and stirred for 20 hr a t room temperature. The solvent was removed in uacuuo at 50°, the residue was dissolved in ethyl acetate and washed with sodium bicarbonate solution followed by water and dried over sodium sulfate. The ethyl acetate solution was evaporated to dryness leaving the derivatives as oils a t room temperature; only 8-naphthacyl oleate was a crystalline compound. The derivatives were recrystallized from ether a t -Zoo. Since the methylene interrupted polyunsaturated acids are very prone to oxidation, the reaction and work-up were conducted under nitrogen and all vessels were protected from light. High Speed Liquid Chromatography. Analyses were conducted using a chromatograph fabricated in this laboratory. A Varian constant pressure, gas driven reservoir/pump was employed with nitrogen as the pressurizing gas. A Pharmacia 1205 UV monitor measured absorbance a t 854 nm. The column was a 3-ft X 0.07-in. i.d. stainless steel tube packed with Corasil-Cls (Waters Associates). Methano1:water (8515) served as the eluent and a flow rate of 12 ml/hr was obtained a t a pressure of 300 psig. The solvents were degassed by boiling just before use. Samples were injected directly onto the column. Mass Spectrometry. A Finnigan 1015 quadrupole mass spectrometer was used. The ionizing current and voltage were 50 pA and 50 eV, respectively, when E1 spectra were recorded. CI spectra were recorded using methane as the reactant gas; the source parameters were 500 FA and 100 eV. The scan time was 10 sec. All samples were introduced through the solid probe and the spectra were recorded a t probe and ion source temperatures of 200'.
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