Analysis of High Molecular Weight Alcohols by the Mass Spectrometer The Wax Alcohols of Human Hair Fat R. A. BROWN and W. S. YOUNG The Atlantic Refining Co., Philadelphia, f a . NICHOLAS NICOLAIDES Section of Dermatology, Department of Medicine, University of Chicago, Chicago,
High molecular weight alcohols separated from hair oil have been analyzed by high temperature mass spectrometry. Spectra of to Cn straight-chain alcohols were identified. Alcohols having an even m~mberof carbon atoms are present in significantly greater quantity than the adjacent odd homolog. pure compounds in theo l ' to range were used as standards.
'"
D
URING the analysis of human hair fat, a fraction was isolated Iyhich was apparently a complex mixture of straight-chain primary alcohols as determined by urea adduct formation, infrared spectra, and recrystallization properties. The recent success of high temperature mass spectrometry as applied to the analysis of mixture of hydrocarbons of molecular weight as high as 700 (1, 4 ) suggested that the method could easily be adapted to this mixture of alcohols whose average molecular weight was 296, and indeed, such has been the case. Two homologous series were distinguished, an aliphatic one ranging from cI6to c2,,and the mono-olefinic series ranging from c18 to c*,. EXPERIMENTAL
Preparation of Sample. Hair fat was obtained by two methods: from exhaustive batchwise ether extraction of pooled cut hair secured from haircuts of inmates of state institutions and from daily wiping of the scalp of a medical student with fat-free cotton swabs soaked in ether, The experimental subjects either had no access to hair preparations (tonics, pomades, etc.) or agreed not to use them.
100 80 60 d
40
Y
d
20
'
0 100 80
"
-2-
5
4 '*
1
111.
The removal of free fatty acids, saponification, and chromat OgraPhY of the UnsaPOnifiables were Performed as Previously described (3). The benzene eluate of the chromatogram was next subjected t o a urea separation (6, 6) in which the fat was boiled with urea and 95% ethyl alcohol (in the proportion 1 gram of fat to 5.5 grams of urea and 7.5 ml. of alcohol), and cooled, then petroleum ether was added and the mixture was filtered. On evaporation of the filtrate a second crop of crystals was obt,ained. Both crops of crystals were then combined, washed repeatedly with petroleum ether, and decomposed with water, and the straight-chain alcohols were recovered by ether extraction. The mixture of alcohols represented about 8% of the hair fat of adults and about 4% of that of boys. Mass Spectrometry. A Consolidated 21-103-4 mass spectrometer was used which had been modified t o have operating characteristics similar to those discussed in the literature (1, 4). Spectra were obtained of wax alcohol mixtures derived from several samples of hair fat from men, R70men, and boys 6 to 12 years of age. To interpret t'hese results calibration spectra were taken of cia, ciz, ci4~Ci6, Cia, cz4, Czs, c30, c31, c 3 3 , and C3a normal alcohols as Well as of Oleyl alcohol. -1Cza isoalcohol (containing a terminal isopropyl group) and tlvo alcohols, CZ3 and cZ5, of the ante-iso series (containing a terminal secondary butyl group) were also examined: since these alcohols have been found t o occur in wool wax (2). The Cia to Cis alcohols were obtained from commercial suppliers. The normal saturated alcohols differed from each other in an orderly manner. For this reason data for compounds not available as calibration standards could be estimated by interpolation. Complete spectra of the Clo to C18 alcohols are being published by the API Project 44. RESULTS AND DISCUSSION
The alcohol spectra proved to be interesting, and numerous points of characteristic behavior were observed. Parent ions are either absent or of low abundance. The intensity of mass-31 ions, a -CH20H fragment, varies inversely w i t h m o l e c u l a r weight, whereas mass44 ions =CHCHzOH are relatively constant. Ions at the high mass end of the spectrum occur a t intervals of 14 mass units which are alternativelv high ---.1-Octacosanol a n d low i n a b u n d a n c e . I-Nonacasanol Furthermore, the mass of .-.- 1-Triacontanal abundant ions are defined by the equations:
60
Mass = Jf
- (18 + n28)
Mass = Af
- (20
a
40
20
+ n28)
vihere M is the molecular xeight of the alcohol and n may have the value 0 or 1.
0
I80 194 208
222
236 250
264
278 292
306
320 3 3 4
348
362 376
390
Mass of Ion
Figure 1. Mass Spectra of Aliphatic Alcohols
1653
404 418
432 446
460
The spectrum of l-octadecenol indicated that monoolefinic alcohols behaved like aliphatic ones.
1654
ANALYTICAL CHEMISTRY
The C Zis0 ~ alcohol spectrum was similar to that of the normal
Czr alcohol. Ante-iso alcohols differed from is0 and normal compounds in that a relatively high peak occurred a t M-31 and a t mass 70. Occurrence of ions according to the formula M - (20 n28) is illustrated in Figure 1. Shown here are relative ion intensities a t masses 180 to 446 in intervals of 14 for 1-octadecanol, l-tetracosanol, 1-octacosanol, 1-nonacosanol, 1-triacontanol, l-hentriacontanol, and 1-dotriacontanol as well as a mixture. Since the molecular weight of 1-octadecanol is 270, relatively intense ions should occur a t masses 250 and 222 with weak ones a t 236 and 208. l-Tetracosanolj of molecule weight 354, has intense ions a t 334 and 306 with weak ones a t 320 and 292. [These spectra were obtained a t the time when a stainless steel tube was located beta-een the leak and ion source. Subsequently, this tube was replaced with one of glass. After the modification alcon28) ions, hol spectra showed a marked decrease in ,M - (20 indicating that these ions are due to thermal and not to electronic decomposition. In this case the heated metal surface apparently catalyzed decomposition. Alternation of the basic nature of the spectra casts no uncertainty on the analysis reported, as they were based on reproducible spectral data. Analysis based on these changed spectra would, however, be calculated using a different combination of peaks from that described.] The 11-48 peak, however, diminishes in comparative size with increasing molecular weight although a reversal occurs between C31 and c32. The ratio of the 127/226 ions of n-hexadecane are shown as a reference to indicate the manner in which relative peak heights varied during calibration of the mass spectrometer.
+
+
homolog spread for two different samples within the same group check within the accuracy of the method and the very similar analysis of samples obtained by two different methods (extraction of pooled cut hair us. scalp wiping) indicates that no gross contamination was involved. Of particular interest is the appearance of alcohols containing odd as well as even numbers of carbon atoms of both the saturated and unsaturated variety. As shown by analysis, however, alcohols with an even number of carbon atoms are present in greater amounts. Visual inspection of the mixture spectrum in Figure 1 shows this fact. Urea separation is not discriminating for small terminal branchings of long chain material such as 2-methyl (iso) and 3-methyl (ante-iso) compounds ( 2 , 6). For this reason, the possibility of detecting such compounds by mass spectrometry was briefly investigated. From the 69 and 70 peaks it was estimated that at the maximum ante-iso alcohols were present to less than 10%. The spectrum of the is0 alcohol, however, was too similar to that of the normal alcohols to discriminate between them. The method of mass spectrometry is particularly suited for the analysis of naturally occurring straight-chain alcohols (and fatty acids after first reducing them to alcohols) because of the fact that these substances usually occur as complex mixtures of closely related homologs. Furthermore, as little as 30 mg. of material is required for a complete analysis. ACKNOWLEDGMENT
The authors wish to thank George Martin of The Atlantic Refining Co., who made extensive calculations required in this work.
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Table I. Human source Samplea T y p e of alcohol, mole, 70 Cl6 c 1 7
cis
ClS c 2 0 C21 c 2 2
c 2 5 C24
C25 C26 C2i
Totals a
b C
Mass Spectrometric Analysis of Straight-Chain Alcohols from Human Hair Fat Men VI1
A1iph.b 2 1 15
Women
w
I JV 0lef.C 0 0 4
4
2
17 7 8 6 8 3 0.5 0 71.5
11
2 5 1 2
1 0.5 0 28.5
Aliph. 2
1 18 4
18 6 8 5 8 3 1 0.5 74.5
AK Olef. 0
0 3 2 10
2 4 1 2 0.5 0.5 0.5 25.5
Aliph. 1 0 19 4 22 9 10
VFW Olef. 0 0 5
0.5
2 12 2 3 1 1 0.5
73.5
26.5
4
4 0 0
0 0
Aliph. 6 1 16 4
18 6 9
5 6 2 0.5 0.5 74
VI F W Olef 0 0
4 2 10 2 4 1
2 0.5 0.5 0
26
Aliph. 5 2 17 4 14 '6 7 4 6
2 1 0.5 68.5
Olef. 0 0 5 2 13 2 5 1 2 0.5 0.5 0.5 31.5
Boys 6 t o 12, I B Aliph. Olef. 2 0 0 0 12 4 4 3 20 10 2 10 4 10 1 6 7 1 0.5 2 0.5 1 0 0 74 26
Sample AK was from scalp wipings; t h e remaining samples were from pooled c u t hair extracts Saturated alcohols. Mono-olefinic alcohols.
To determine the composition of the mixtures, lengthy but straightforward calculations were carried out. First, calibration spectra of the C16 to Cao alcohols were obtained by experiment or interpolation of observed data. Unicomponent mixture peaks were then calculated for mono-olefinic alcohols using the mass series 220, 231. 218, 262, etc. This TTas done by starting a t the heaviest mass and successively removing peak contributions from one molecular ITeight species to another using calibration data to calculate these contributions. Aliphatic peaks were calculated from the mass series 222, 236, 250, 264, etc., following the same procedure as for mono-olefins. In this case, however, contributions of mono-olefinic to aliphatic alcohols must be taken into consideration. Finally, peaks were converted to partial pressures which in turn were changed to volume units so as to express composition in volume per cent. The homolog distribution of straight-chain alcohols obtained from the hair fat of men, women, and boys is listed in Table I. This distribution for both saturated and mono-olefinic alcohols is essentially the same for all three groups of human beings. The
The Clo to C18 alcohols were kindly provided by Reuben Jones of the Eli Lilly Co., Indianapolis, Ind., and the C Z normal, ~ C2( iso, C23, and C25 ante-iso alcohols, by K. E. Murray of the Commonwealth Scientific and Industrial Organizat,ion, Melbourne, Australia. LITERATURE CITED
(1) Bronrn, R. A , AIelpolder, F. W., and Young, W. S.,Petroleum P T O C C S 7,204 S ~ ~ Q(1952). , (2) Murray, K. E., and Schoenfeld, R., -J. Am. Oil Chemists' SOC., 29,416 (1952). (.3.) Nicolaides, S . , and Rothman, S., J . Invest. Dermatol., 19, 389 (1952). (4) O'Neal, 11. J., Jr., and Wier, T. P., Jr., ASAL. CHEM.,23, 830 (1951). (5) Rudloff, E. V., Chemistry & I n d u s t r y , 1951,338. (6) Zimmerschied, W. J., Dinerstein, R. A,, Weitkamp, A. W., and RIarschner, R. F., I n d . Eng. Chem., 42, 1300 (1950). RECEIVED f o r review March 18, 1954. Accepted July 1, 1954. Investigation supported in part by t h e Medical Research and Development Division, Office of the Surgeon General, Department of t h e h r m y , under Contract S o . D.4-49-007-3ID-411.
