Infrared Spectra of Steroids New Solvents f o r Steroids Dificultly Soluble in Carbon DisulJide
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WILLI.4M T A R P L E Y AND CECELIA V I T I E L L O Chemical Research Department, Schering Corp., Bloom$eld, N. J . In studies of the infrared spectra of certain highly purified steroids from 2 to 15 microns (5000 to 666 cm. -l), difficulty is frequently encountered inobtaining spectra suitable for comparison purposes. These steroids are often only slightly soluble in the usual nonpolar solvents. Two approaches have resulted in some success in overcoming this difficulty; 0.2% solutions of As-pregnene-3(P)-ol-20-one in carbon disulfide yielded suitable fingerprint region spectra in a cell of long path length (8.5 mm.). Stabilized polar liquids with heavy atoms in their molecular structure were investigated in a n effort to find solvents t h a t would produce little or no interference in
the spectra. The spectra of purified bromoform and methylene bromide in 0.2- and 1-mm. cells are shown together with 17-ethinylestradiol in bromoform and 16,17 (a)-oxido-As-pregnene-3(P)-ol-SO-one in methylene bromide solutions. Purified bromoform and methylene bromide w-ere found to be good solvents for these steroids and yielded fingerprint spectra far superior to chloroform and carbon tetrachloride. By supplementing spectra in bromoform with those in methylene bromide only narrow regions of the spectrum remain unavailable. Both solvents produce no interference in the double bond region where carbon disulfide is not usable.
A
AIOSG the difficulties ill the infrared spectrometry of solids may be included the problems of preparing sufficiently thin films ( 9 ) , loss of energy from the beam through scattering by crystalline samples ( 2 , 4 , 11, 19, 2 2 ) ) and spectral variations due to partial sample orientation and possible polymorphism (1), Although mulling with mineral oil or perfluorokerosene has been used to reduce scattering losses, it has been the experience of this laboratory that interpretations of solid spectra, particularll- those of steroids, are frequently questionable because of the above effects, the impossibility of reproducing exactly the amount of sample in the beam, and concurrent resolution differences. The presence of absorption band differences between several samples of the same compound in niineral oil mull spectra, which vere absent when spectra of the same samples n-ere determined in solution, has been attributed to polymorphism (12, 1 7 ) . Solution spectra, although more reliable in interpretation, are complicated by the existence of relatively fen solvents which do not absorb throughout large intervals of the 2 to 15 micron (5000 to 666 cm.-lj region (20). The use of solvent-filled compensation cells in the reference beam of double-beam infrared
spectrometers can reduce interferences from less intense solvent absorption bands, but is of no value when absorption is so great as to render the instrument inoperative (21). A suitable solvent for infrared spectrometry should have a spectrum relatively free from absorption bands and a fair degree of stability, should not react with the sample or the materials of the absorption cell, and must dissolve enough solute to give absorption bands of satisfactor?- intensity in the fingerprint region of the spectrum. For steroid samples, approximately 1 to 3% solutions in sodium chloride cells of 1-mm. path length are required ( 7 ) . Carbon disulfide, carbon tetrachloride, and chloroform (3, 13, 15, 20) are the solvents most frequently used.' Carbon disulfide has by far the fewest absorption bands, but many steroids, particularly those with several hydroxyl groups, cannot be dissolved to a sufficiently great estent to yield satisfactory spectra (16). Similarly, carbon tetrachloride is a poor solvent for many steroids and in addit,ion absorbs so strongly from 7.8 to 15 microns (1280 to 666 c m - l ) a s to preclude obtaining satisfactory spectra in the fingerprint region. Chloroform has been used frequently in steroid studies ( 1 4 ) and will usually dissolve sufficient sample. Unfortunately, the
WAVE LENGTH, MICRONS
Figure 1. A5-Pregnene-3(p)ol-20-one in Carbon Disulfide 3 mg. per ml.
315
ANALYTICAL CHEMISTRY
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Figure 2.
Bromoform in 0.2-3Im. and l - l l m . Path Lengths
multiplicity of absorption bands permits only scattered observation in the fingerprint region ( 6 , 16) and in addition masks portions of the functional group region. During the past year, two approaches in these laboratories have found some success in overcoming the above difficulties. -4n absorption cell of 8.5-mm. path length was prepared by soldering (10)sodium chloride windows to a brass spacer. It is thus possible to introduce 8.5 times as much sample into the infrared beam as with conventional 1-mm. cells. With relatively insoluble samples, such as highly purified A3-pregnene-3(@)-ol-20-one,suitable spectra in the fingerprint region were obtained (Figure l), although a concentration of only 0.2% in carbon disulfide could be prepared. There is some broadening of the major carbon disulfide absorption bands, as v-ell as increased interference from the smaller bands. 3Iinor variations in purity of carbon disulfide are more apparent a t this path length and mahe it ndvisable to determine a solvent reference spectrum on each ne\\shipment. Theoretical considerations ( 2 , 19), as well as published spectra (6, 8, 18), suggest that the replacement of chlorine in halogenated methanes by heavier elements (leading to displacement of the carbon-halogen absorption bands to longer nave length) might
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result in good solvents with fewer interferences in the 2 to 15 micron (5000 to 666 cm.-I) region. In preliminary studies, methylene iodide was considered but soon proved to be unstable. Freshly distilled methylene iodide discolored rapidly and gave many absorption bands. Both redistilled bromoform (Figure 2) and methylene bromide (Figure 3) were found to give suitable spectra in cells of 0.2- and 1-mm. path length. I n general, the solubility of steroids in these solvents parallels that in the corresponding chlorinated solvents, a tremendous increase over that in carbon disulfide and carbon tetrachloride. Bromoform is the preferred solvent, as it causes less interference than methylene bromide in the functional group region, and there is no interference in the region around 8 microns, frequently needed for con-
II firmation of the presence of -C-0-Cof acetate groups. By recourse to spectra in methylene bromide as well as in bromoform (0.2-mm. path length), only the narrow regions of the spectrum2.40 to 2.45 microns (4170 to 4070 em.-'), 3.25 to 3.40 microns (3070 to 2940 em-'), 8.40 to 8.70 microns (1190 to 1149 em.+), and 14.8 to 15.0 microns (676 to 666 cm.-')-remain unavailable. The spectra of 17-ethinylestradiol and 16,17(a)-oxido-Aj-pregnene-3( p)ol-20-one (Figures 4 and 5) are typical results. Correla-
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Figure 3.
Methylene Bromide in 0.2-Mm. and 1-Mm. Path Lengths
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17-Ethinylestradiol in Bron
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30 mg. per ml.
tions of absorption band TT-ave length with structureusingchloroform as a solvent appertrh apply hithoUt shift h bromoform and methylene bromide. Similarly, resolution of adjacent bands seems not to have been affected. EXPERIMENTAL
Determination of Infrared Spectra. A Perbin-Elmer Model 12C single-beam spectrometer with sodium chloride prism N&E used in the early phases of this work. More recently a PerkinElmer Model 21, doublebeam spectrometer was used. No attempt was made t o compensate for solvent absorption bands, but the absorption bands of carbon dioxide, and water vapor in the air, are automatically compensated in the latter instrument. All spectra were run under conditions usual in this laboratory for survey studies. Solute concentration and p?th length of the cell are indi cated on the mectra. About 25 mlnutes were taken to record e balancu maticd ;form. In order t o dbtain bromoform of sufficient spec,ity, ethyl alcohol and other impurities must he removed. orm (Merok or Baker U.S.P.) was fractionated under 1 (25 mm. of mercury) in a 50-cm. r;ll-glassSnyder colomn ~
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and the forerun ot aU% Was ulsearaeu. I n e main lractlon, buyo (boiling paint 49-50' C., 25 mm. of mercury), %-ascollected while the colored residue amounting to 10% was discarded. Tho usable main fraction w u stored in small brown glass bottles, 60 that oply %?ugh bromofprm for 3 or 4 da&use,need exposed to the air. pive grams of mercury per IUU mi. 01 Dromotmm were added as a stabilizer. When stored under laboratory conditions, out of direct sunlinht. and tiebtlv closed when not, in n ~ e no .
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Methylene bromide (Dow Technical) was distilled at atmospheric pressure, and the forerun 10% was discarded. The main fraction, 80% (bailing poiFt 97.5-98.5" C.], TVZ%? stored brown Elass bottles, Bind tne realclue was umarueu. I'he lnfrared ~~
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were expended in itdfurther $ f t c a t i o n ." Thh m a t e n d wa8 stable for a t least 3 months, as judged by spectrometrio criteria. Methylene bromide solutions can also be warmed for short p s i o d s on the steam bath without decomposition. lnao
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16)7 L*) 0 N ~ O - -PREGNENE-3p)h-ZO-U b~ IN CHEBI~.O.ZMM mlH LENGTH
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16,17(a)-0~ido-As-6-p~egnene-3(~)ol-20-one in Methylene Bromide 25 mg. per ml.
ANALYTICAL CHEMISTRY
318 Possible Reaction of Steroid Samples with Bromoform or Methylene Bromide. Over 125 spectra have been run in bromoform and methylene bromide, with no evidence of reaction between solute and solvent. Occasionally, solutions of steroids in bromoform have developed a noticeable yellow color after standing about 3 hours. The infrared spectra of such colored solutions even after standing for 3 days proved to be identical with those obtained immediately after dissolving. Furthermore, on shaking with mercury, the color was completely removed from the solutions with no resultant change in the infrared spectrum. ACKNOWLEDGVENT
Thanks are due to Betty Blasko, who assisted in obtaining the spectra. LITERATURE CITED
R. B., Gore, R. C., Williams. E. F., Linsley, S. G., and Petersen. E. M.. ANAL.CHEY..19. 620-7 (1947). (2) Barnes, R. B., Liddel, U.. and Williams, V. Z:, IND.EKG.CHEY., ANAL.ED.,15, 659-709 (1943). (3) Blout, E. R., Fields, bl., and Karplus, R., J . A m . Chem. Soc., (1) Barnes,
70, 194 (1948). (4) Bonham, L. C., Hunt, J. I f . and Wisherd, M. P., AN.~L. CHEY., 22, 1478-97 (1950). (5) Decius, J. C., J . Chem. Phys., 16, 214-22 (1948). (6) Dobriner, K., Perkin-Klmfr1nstriLment N e w , 1, Yo. 1, 3 (1950).
(7) Dobriner, K., Liberman, S., Rhoads, C. P., Jones, R. Ii., Williams, V. Z., and Barnes, R. B., J . Bid. Chem., 172, 297 (1948). (8) Emschwiller, G., and Lecomte, J., J . phys. radium, 8 , 130-44 (1937). (9) Furchgott, R. F’., Rosenkrantz, H., and Shorr, E., J . Biol. Chem., 163, 375-86 (1946). (10) Gore, R. C., private communication. (11) Henry, R. I,.,J . Optical SOC.Am., 38, 7 7 5 8 9 (1948). (12) Jones, R. N., Chemistry in Canada, 3 (June 1950). and Dobriner, K., Vitamins and Hormones, 7, 293 (13) Jones, R. Ii., (1949). (14) Jones, R. N., Humphries, P., Packard, E., and Dobriner, K., J . Am. Chem. Soc., 72, 86-91 (1950). (15) Marion, L., Ramsay, D. h.,and Jones, R. N., Ibid., 73, 305 (1951). (16) Nelson, D. H., Reich, H., and Samuels, L. T., Science, 111, 578 (1950). (17) Pheasant, R., J . Am. Chem. Soc., 72, 4303 (1950). (18) Plyler, E. K., Smith. TT. H., and Acquista, N., J. Researrh Natl. Bur. Sfandnrds, 44, 503 (1950). (19) Randall, H. M., E’uson-X., Fowler, R. G., and Dangl, J. R.
Infrared Determination of Organic Structure, New York, D. Van Nostrand Go., 1949. (20) Torkington, P., and Thompson, H. W., Trans. Faraday Soc., 41, 184-6 (1945). (21) White, J. E., and Liston, hl. D., J . Optical Soc. Am., 40, 93-101 (1950). (22) Williams. V. Z., Rea. Sei. Inslruments, 19, 135-78 (1948).
RECEIVED May 5 , 1951. Presented before the Meeting-in-Miniature, North Jersey Section, AMERICAS C H E w c A L SOCIETY. January 28, 1952.
Correlation of Infrared Spectra Parafins, OleJins, and Aromatics with Structural Groups H. L. MCMURRY
AND VERNON THORNTON
Research Division, Phillips Petroleum Co., Bartlesville, Okla. This work was undertaken to improve the reliability and convenience of structural anal) ses of hydrocarbons br infrared spectra, and to provide factual data which might eventually aid in understanding the vibrational origins of certain bands. Correlations between the spectra and structural properties of paraffins, olefins, and aromatics are tabulated and charted in a manner convenient to the needs of the practicing spectroscopist. They should improve the reliability of structural analyses of hydrocarbons because (1) wave-length ranges for the correlation
A
CORRELATIOS band is any band which empirical observation has demonstrated to be associated with a specific atom grouping (hereafter termed “structure”). Its presence in the spectrum of a material indicates that the structure may be present; its absence is certain evidence that the structure is missing. Any correlation band will vary in position, depending on the environment of the associated structure. Most tables and charts of correlations assign wave-length ranges to the bands (1, 3, 6, 10, 1 2 ) . Many correlations which have been deduced from the spectra of molecules involving nonhydrocarbon as well as hydrocarbon substituents are found to have much narrower ranges when they are deduced from hydrocarbon spectra only. Figure 8 illustrates this for the correlations for benzene ring substitutions. I n making structural analyses of petroleum chemicals by infrared, it is useful to use correlation tables and charts giving the narrower wave-length ranges valid for the hydrocarbons.
bands have been deduced from only hydrocarbon spectra. Often the ranges thus determined are considerably narrower than shown in data published heretofore. This makes the bands more definitive. (2) Intensity data are given. These are sometimes useful in identifying the structures giving rise to bands. In some cases they make possible estimates of the concentrations of groups. (3) Critical evaluations of the correlations are given. This makes the limitations involved in using any specific correlation band more evident.
This report presents correlation tables and charts for paraffinic, olefinic, and aromatic structures. The paraffin correlations were deduced from a study of paraffin spectra only; the olefins are from the spectra of unconjugated alkyl substituted compounds. The aromatic correlations were obtained partly from the spectra of alkyl aromatics and, where the data on hydrocarbons were limited, from a comparison of the spectra of alkyl aromatics with published correlation charts. The problem of determining how the correlations given here for paraffinic structures are modified when these are adjacent t o olefinic or aromatic structures, and of deteimining the effect on the olefinic and aromatic correlations when other hydrocarbon structures than alkyl groups are substituted, cannot be solved empirically without more data than are n o x available. The sprctral data on hydrocarbons are from the American Petroleum Institute catalog of infrared spectra of hydrocarbons. The aromatic correlations were partl? deduced with the aid of
