DEPARTMENT O F AGRICULTURE, AGRICULTURAL RESEARCH ADMIXISTRATION, BCREAU OF ENTOMOLOGY AND PLANT QUARANTINE ]
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CONSTITUENTS OF PYRETHRUM FLOWERS. XVII. T H E ISOLATION O F FIVE PYRETHROLONE SEMICARBAZOWES F. B. LAFORGE AND W. F. BARTHEL Received October id, 19.64
A new conception of the chemical nature of the pyrethrins has become necessary since their alcoholic-ketonic component, “pyrethrolone,” formerly considered to be a homogeneous compound, has been shown to be a mixture of two or more constituents (1). This conclusion was reached following the observation that fractions were obtained on distillation which showed a progressive increase in index of refraction and a progressive decrease in terminal-methyl content with increase in distillation temperature. Certain observations of discordant phenomena, especially relating to refraction and absorption (2), when referred to chemical behavior, may now be readily reconciled if it is accepted that pyrethrolone and all its derivatives consist of mixtures. Although the constituents with the higher terminal-methyl content tended to accumulate in the earlier fractions, only incomplete separations could be accomplished owing to the small differences in boiling range. The pronounced tendency of pyrethrolone to polymerize on protracted heating was another dificulty. Nevertheless, a fraction, n:’ 1.5467, could be isolated, although only in small quantity, that contains but one equivalent of terminal methyl. This fraction consists almost entirely of that form of pyrethrolone corresponding to formula I, and may be present to the extent of 80% or more in preparations obtained from pyrethrin semicarbazone that has been repeatedly recrystallized with loss of the more soluble constituents.
CH,
I HzH H H /X-C-C=C-C=CH2
:p0 0 H
I It is responsible for the observed double chromophoric effect,’ by reason of its conjugated pentadienyl side chain, the grouping also responsible for the production of formaldehyde on ozonization. A satisfactory concentration of the constituents of low boiling range, characterized thus far only by their high terminal-methyl content and low refractive index, could not be accomplished by distillation of pyrethrolone. Fractionation 1 From a spectrographic study, Gillam and West (2 b) demonstrated the presence in pyrethrolone of two separate chromophoric systems, each containing more than one double linkage, the a,#?-unsaturated ketone feature, and a conjugated diene system. 106
CONSTITUENTS OF PYRETHRUM FLOWERS
107
mas practical, however, with the lower-boiling and more stable acetyl derivatives. Thc: refractive index and the terminal-methyl content of the various fractions were found to be the most reliable criteria of quality, the distillation temperature having only approximate significance. Yo attempt was made to apply any special refinements in this distillation, because it was known that more or less polymerization always occurred, but fortunately this property seems to be especially, perhaps exclusively, characteristic of the major constituent of pyrethrolone of formula I, while the rare constituents appear to be the more stable ones. The distillate obtained by redistillation of the lower fractions of acetyl pyrethrolone furnished a semicarbazone mixture from which two members were isolated. One of these, m.p. 150-151”, strongly predominated. By its ready solubilit,y in benzene it could be easily separated from its companion, m.p. 151-152”, which was almost insoluble and was present in much smaller quantity. The combined higher-boiling fractions likewise yielded two semicarbazones, which could also be separated from each other by their pronounced difference in solubility in benzene. The more soluble one melted a t 133”,its difficultly soluble companion a t 173-175”. It appears from evidence to be presented that these four compounds probably consist of two pairs of stereoisomers. This seems still more likely when the corresponding “pyrethrolone semicarbazones,” obtained from the acetyl derivatives by saponification, are compared among themselves and with the semicarbazone of the conjugated form of pyrethrolone of formula I, designated from now on as “pyrethrolone C”. Of the four new pyrethrolones, the members of the first pair will be designated as “A-1” and “A-2”, those of the second pair as “B-1” and “B-2”. All five of the corresponding semicarbazones resemble one another by their difficult solubility in all the usual solvents and by their similar melting points, all located between 200” and 218”. All five melt with decomposition and gas evolution. Their similar solubilities account for failure t o observe any fractionation on recrystallization of mixtures of them. Semicarbazone of pyrethrolone A-1. This semicarbazone is the derivative of the pyrethrolone that, except for the compound of formula I, predominates in all ordinary “pyrethrolone” preparations. Its empirical formula, CllH17N302, differs from that of the semicarbazone of pyrethrolone C (and the accepted formula for “pyrethrolone semicarbazone”) by possessing one less carbon atom. Its acetyl derivative was isolated in quantity about equal to the combined yields of all the other minor constituents. On carefully controlled hydrogenation 2.35 moles of hydrogen was absorbed, corresponding to slightly more than two unsaturated linkages. The excess hydrogen absorption, together with the fact that values for carbon and hydrogen deviate somewhat from the theory, indicates that the material may contain a small amount of some other compound, probably semicarbazone of pyrethrolone B-1 Samicarbazone of pyrethrolone A-2. This companion t o the semicarbazone of pyrethrolone A-1 was obtained from its acetyl derivative (m.p. 152”), which remained undissolved after extraction of the main product with benzene. Except
108
F. B. LAFORGE AND W. F. BARTHEL
for the marked difference in solubility of its acetyl derivative, this semicarbazone exhibits the same chemical characteristics as the semicarbazone of A-1 with which it is associated. It is represented by the same empirical formula and contains two unsaturated linkages. Semicarbazone of pyrethrolone B-I. The acetyl derivative of this semicarbazone, m.p. 133”, was isolated from the mixture obtained from the higher-boiling fractions of acetyl pyrethrolone. It was separated from the accompanying isomer by its high solubility in benzene. The corresponding pyrethrolone semicarbazone obtained by saponification of the acetyl compound is the most insoluble of all the five semicarbazones. However, it is represented by formula C12H17N302,and hence is isomeric with the semicarbazone of pyrethrolone C. Like the latter, it contains three unsaturated linkages and, on hydrogenation, two of these are saturated a t a rate about ten times that of the third one. It may be inferred from these facts that this compound, like semicarbazone of pyrethrolone C, contains a doubly unsaturated side chain. Semicarbazone of pyrethrolone B-2. The corresponding acetyl derivative, m.p. 173-175”, being very dBicultly soluble in benzene, remains undissolved after the more soluble component is extracted from the mixture by this solvent. The semicarbazone itself is represented by the same empirical formula, CI2H1,N302, as is that of B-1. It melts several degrees lower and, like its companion, contains three unsaturated linkages, two of which are hydrogenated a t about ten times the rate of the remaining one. The mixture of the acetyl derivatives of this isomeric pair contains both members in about equal amounts. But their combined yield was equal to only about 50430% of the distillate employed. The remaining part consisted of viscous material quite soluble in ether, which hardened to a solid resin on heating. We surmise that it consisted of partially polymerized or altered pyrethrolone derivatives. Similar material had remained as the last still residue after each distillation. It would, therefore, seem that the isolation of the more rare, but probably more stable, constituents of “pyrethrolone” has been facilitated by the greater instability of the predominating one. Semicarbazone of pyrethrolone C. This derivative was prepared in a state approaching punty from the last fraction obtained on refractionation of the higher-boiling constituents of “pyrethrolone.” It melts a t 218” and resembles the other pyrethrolone semicarbazones in solubility. On hydrogenation it shows behavior identical with that of the semicarbazones of B-1 and B-2. It contains one terminal methyl group. Since these three semicarbazones are isomeric with one another, it seems likely that the boiling points of the corresponding pyrethrolones are near together, which would account for the difficulty in separating pyrethrolone C from its isomers by distillation. Those constituents with a lower molecular weight would tend to accumulate in the lower-boiling fractions . In what proportions the different pyrethrolones are combined with the chrysanthemum acids is unknown. The terms “pyrethrin I” and “pyrethrin 11,”
CONSTITUENTS O F PYRETHRUM FLOWERS
109
however, must henceforth be regarded as defining not compounds, but groups characterized only according to the acid component, which in each case is esterified with more than one and probably with several pyrethrolones. EXPERIMENTAL
The starting material was 325 g. of a concentrate prepared from a commercial pyrethrum extract by the selective-solvent process described elsewhere (3). It contained 94% of total pyrethrins as determined by the mercury-reduction method and 100% by hydrogenation. This material was used for the preparation of the semicarbazones by the usual procedure. The proportions of reagents and solvents per 100 gm. of concentrate were as follows: Semicarbazide hydrochloride 60 g., pyridine 65 ml., water 80 ml., and ethanol 350 ml. After 48 hours at room temperate the crystallization was completed by cooling in an ice-salt bath, tho separated material was washed with cold 50% ethanol and then with water, and dried in the air. The yield was 220 g. The mother liquor and washings were concentrated in vacuum, and the separated semicrystalline material was extracted with ether. The solution was washed with dilute acid and dried, and the solvent was removed, leaving a residue that partly crystallized on standing. From this residue another 40 g. of clean crystalline pyrethrin semicarbazone was isolated by dissolving the syrupy by-products i n cold methanol. The non-crystalline material still contained pyrethrin semicarbazone, for on saponification i t yielded some pyrethrolone semicarbazone. For the most part i t consisted of a sticky syrup, which we believe to be partly polymerized pyrethrins. Fractionation of acetyl pyrethrolone. About 65 g. of "pyrethrolone," prepared via its sernicnrbazonc from the crude mixture of pyrethrin semicarbazones, was distilled in the usual manner from a Claisen flask with a vacuum of about 1 mm. The first 31 g. of composite distilled VI-as collected over the temperkture rangc 135-140", nz 1.5350; the second fraction, 29 g., over the range 140-145", n?,? 1.5450. The first distillate furnished 32 g. of the acetyl derivative [Staudinger and Ruzicka (2a)j which showed a refractive index n: 1.5158. This was subjected to a fractionation with a 25-1" insulated column, and six unequal fractions were collected over the range 120-130", p = 1.0 mm. The index of refraction (n?,?) of each ww measured and observed t o increase regularly from 1.4961 to 1.5140. The major portion, 16 g., of the combined fractions 1 to 4 was redistilled in the same apparatus but under a vacuum of 0.2-0.1 mm.; i t yielded 10 g. of distillate, b.p. 103-112", ng 1.4963, acetyl 21.5%. The residue in the flask and 4.4 g. of a seventh fraction, n?,? 1.5140, acetyl 21.5%, were separately distilled without the column, yielding 2.6 g. .4cetyl derivative of semicarbazone of pyrethrolone A - i . Eight and a half grams of the distillate boiling at 103-112" was dissolved i n a solution composed of 6 g. of semicarbazide hydrochloride in 8 ml. of water, 8 ml. of pyridine, and 60 ml. of ethanol. After about 48 hours partial evaporation had occurred, and complete precipitation of the crystalline reaction products was induced by addition of water. The washed and dried crude material was extracted with warm benzene, in which most of i t dissolved. The cold solution was allowed to stand for a short time and then filtered from the insoluble constituent. The benzene solution was concentrated and, on addition of warm petroleum ether, the semicarbazone separated in well-defined crystals. After i t had been washed with ethyl ether, the dried compound melted at 140-145". It was recrystallized by boiling with 40-50 parts of :bbsolute ether, filtering from a small residue, and concentratingthe solution to about one-fifth of i t s volume. It then melted a t 150-151", [a]: f50.0" (c = 1.45). The yield was 6.8 g . The insoluble material melted a t 125-135" and may be a mixture of semicarbasones of pyrethrolone B-1 and B-2. After complete evaporation of the ethereal solution, a very soluble, low-melting semicarbazone mixture remained. Anal. Calc'd for C I Z H I O N ~mol. O ~ : wt. 265; N , 15.84; CHaCO, 16.22; 3 CHa, 17.0. Found: N , 15.73, 15.83; CH,CO, 16.40,16.67; CHa, 15.25,14.30,14.75. Semicurbalone of pyrethrolone A - i . One part of the acetyl derivative was suspended
110
F. B. LAFORGE AND W. F. BARTHEL
a t 0' in about 5 parts of methanol i n which 0.1 part of sodium had been dissolved. After 12 hours in the ice-box and a few hours standing at room temperature, the semicarbazone was completely precipitated by addition of water, washed, and dried. It was recrystallized by dissolving in a boiling mixture of methanol and ethyl acetate and concentrating the solution. It melted at 200-201", with decomposition, after sintering a few degrees lower. The yield was almost quantitative. Hydrogenation reveals the presence of two unsaturated linkages. Xmax. 2630, c = 22,000. A n a l . Calc'd for CI1Hl,N30~: mol. wt. 223; C, 59.2; H, 7.60; 2 CH3,13.4. Found: C, 60.06, 60.15; H , 7.34, 7.46; CH3, 10.45, 10.75. Acetyl derivative of semicarbazone of pyrethrolone A-2. The residue from the acetyl semicarbazone mixture that was not dissolved by benzene was separated and recrystallized from ethyl acetate. It melted then at 151-152". The yield was only 1.0 g. Anal. Calc'd for C1,HI9N3O3:mol. wt. 265; N, 15.84; CH3C0, 16.22; 3 CH3, 17.0. Found: N, 15.95; 15.73; CH3C0, 16.80; CH3, 13.65, 13.4, 14.5. Semicarbazone of pyrethrolone A-8. This semicarbazone was obtained in quantitative yield from the acetyl compound i n the same manner as was that of pyrethrolone A-1. It was also recrystallized from the same solvents and melted a t 199-200" with decomposition. It contains two double bonds. hmax. 2650, e = 21,000. Anal. Calc'd for CllH1~N302: mol. wt. 223; C, 59.2; H, 7.68; 2CH3, 13.4. Found: C, 59.64, 59.77; H, 7.58, 7.46; CH3, 9.8, 9.7. If the fractionation of the acetyl compounds has been incomplete, the semicarbazone of acetyl pyrethrolone A-2 may contain some of the acetyl semicarbazone of pyrethrolone B-2. I n this case separation of the two by crystallization is not possible. On saponification, however, only the semicarbazone of B-2 separates from the methanol solution, while the semicarbazone of A-2 is soluble and separates in crystalline form on addition of several volumes of water. Acetyl derivative of semicarbazone of pyrethrolone B-1. The major portion, 12.8 g., of the combined fractions 9, 6, and 7 was dissolved i n a solution of 10 g. of semicarbazide hydrochloride, 13 ml. of water, 10 ml. of pyridine, and 80 ml. of ethanol. The crystalline product, which had separated on 48 hours standing and upon addition of water, was contaminated with a sticky material which was also dispersed in the turbid aqueous-alcoholic filtrate. This substance could easily be removed from the crystalline semicarbazone mixture b y washing with ether. It probably consisted of polymerized pyrethrolone C. The crystalline mixture was separated into a soluble and an insoluble constituent by extraction with benzene. On concentration of the benzene solution and addition of petroleum ether, the soluble compound was precipitated, probably with solvent of crystallization. I n this condition i t readily dissolved in ethyl ether but soon separated in the form of crystalline needles, which were then almost insoluble in this solvent. It melted a t 130-133". It is also obtained solvent-free and melting at the same temperature by evaporation of the benzene solution with a stream of air a t room temperature. [a]: +49.0° (in methanol c = 2.5). A n a l . Calc'd for C141&9Nd&: mol. wt. 277; X, 15.16; CHsCO, 15.52; 2 CH3, 10.83. Found: N, 15.12, 15.15; CHsCO, 16.2, 17.1; CH,, 10.55, 10.60. Semicarbazone of pyrethrolone B-1. The acetyl compound was saponified in the manner described above and the semicarbazone isolated and recrystallized from methanol-ethyl acetate mixture, in which i t is much more difficultly soluble than the other analogs, 100 parts of the boiling solvent mixture being required for solution of 1 part of substance. It melted a t 214", sintering a few degrees lower. The yield was about quantitative. The compound contains three double bonds, two of which are hydrogenated a t ten times the rate of the third. Xmax. 2650, E = 20,500; Xmax. 2280, E = 25,000. A n a l . Calc'd for ClPH1,NaOt: mol. wt. 235; C, 61.26; H, 7.28; 1 CHa, 6.4. Found: C, 60.78, 60.62; H, 7.32, 7.33; CHs, 8.25, 8.25. Acetyl derivative of semicarbazone of pyrethrolone B-B. This compound constitutes the fraction difficultly soluble in cold benzene which was left after extraction of the more
111
CONSTITUENTS OF PYRETHRUM FLOWERS
soluble component. It was recrystallized by dissolving in the boiling solvent and concentrating the solution. It melted a t 173-175". It is isomeric with the acetyl semicarbazone of pyrethrolone B-1. The yield was 2.9 g. Anal. Calc'd for Cl&BNSOS: mol. wt. 277; N, 15.16; CHaCO, 15.52; 2 CHa, 10.83. Found: N , 15.19, 15.09; CHsCO, 15.7, 16.1; CHa, 11.25, 11.00. Semicarbazone of pyrethrolone B-2. The semicarbazone prepared by saponification of the acetyl derivative, after recrystallization from methanol-ethyl acetate as previously described, melted a t 207-208". It contains three unsaturated linkages, two of which are hydrogenated about ten times as rapidly as the third one. Xmax. 2650, B = 20,500; Xmax. 2280, e = 25,000. Anal. Calc'd for C&17S;802: mol. wt. 235; C, 61.26; H, 7.28; 1 CH3, 6.4. Found: C, 61.29; 61.11; H, 7.29, 7.17; CHa, 8.8, 9.3. Semicarbazone of pyrethrolone C . This is the derivative of the predominating alcoholicketonic component of the pyrethrins. It was prepared in small quantity in a state approaching purity from the pyrethrolone fraction, ng 1.5467, obtained by distillation of the second pyrethrolone fraction. After recrystallization in the usual manner it melted at 217-218". Three unsaturated linkages were shown to be present in its structure, two of
TABLE I HYDROGENATION OF THE PYRETHROLONE SEMICARBAZONES SEMICAPBAZONE
YOLECULAP WEIGHT
WEIGHT, 0 .
TIME, MIN.
COP. VOLUME OF HI ML.
[OLES OF Hz/XOLE
A-1
223
0.1049
A-2
223
.0994
B-1
235
.lo49
B -2
235
.lo49
C
235
.0930
5 30 5 40 5 40 5 35 5 44
15.9 24.9 14.9 21.1 19.9 28.5 20.2 29.2 17.3 25.6
1.51 2.36 1.50 2.11 2.00 2.86 2.02 2.92 1.99 2.88
OF SUBSTANCE
which were hydrogenated at ten times the rate of the third. [For spectrographic data see Gillam and West (2 b).] Anal. Calc'd for C12H17Na02;mol. wt. 235; N, 17.87; CH3, 6.4. Found: N, 18.12, 18.17; CHa, 6.8, 6.7. It is possible that pyrethrolone C is a mixture of pyrethrolone B-1 and B-2. Hydrogenations. All the five pyrethrolone semicarbazones were hydrogenated i n a n apparatus with all-glass connections described by Joshel (4). The gas volume was measured over mercury and corrected with reference t o the vapor pressure of the 50-50 mixture of methanol and ethyl acetate employed as the solvent. The catalyst was palladium on calcium carbonate. The volume changes were observed over short intervals i n all instances but are indicated in Table I in abbreviated form. Figure 1 is the graphic presentation of the complete data for the same examples. The preparation of these pyrethrolone semicarbazones haa been carried out -on four samples of starting material without change in the procedure and with identical results except i n one instance. In the exceptional case, apparently the same compounds were isolated, but they showed terminal-methyl contents different from those reported in the experimental part, although agreeing with them with respect t o all the other analytical data. The semicarbazones of pyrethrolones A-1 and A-2 showed a n apparent terminalmethyl content corresponding to 3 moles. This W&B confirmed i n the case of the corre-
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F. B. LAFORGE AND W. F. BARTHEL
sponding acetyl derivatives which showed a content of 4 moles. The semicarbazones of B-1 and B-2 likewise showed a n apparent methyl content of 3 moles and their acetyl derivatives 4 moles. These terminal-methyl values are difficult to account for and must be the result of secondary reactions. Spectrographic data with respect to compounds B-1 and B-2, which indicate the presence of a conjugated diene system, exclude the presence of more than two carbon-linked methyl groups. It is noteworthy that pyrethrolone C always
FIGURE I
-
R A T E S OF H Y D R O G E N A T I O N OF
PYRETHROLONE S E M I C A R B A Z O N E S
shows a n appreciable excess above the theory for 1 mole of terminal methyl. This indicates that it exists in a state of equilibrium between two forms.
We wish t o express our appreciation to R. E. Davis and Harry Bastrom, of the Bureau of Animal Industry, U. S. Department of Agriculture, for supplying the spectrographic data. SUMMARY
Pyrethrolone as prepared by acid hydrolysis of its semicarbazone is a mixture of at least five related compounds. The major part is the one represented by
CONSTITUENTS OF PYRETHRUM FLOWERS
113
formula I, and has been temporarily designated as “pyrethrolone C.” Two isomers of this form have been isolated as their semicarbasones. These three compounds resemble one another with respect to behavior on hydrogenation, and spectrographic data indicate that they all contain a conjugated side chain. Compounds B-1 and B-2 are probably stereoisomers. 13esides these two isomers of pyrethrolone C, a second pair of analogous compounds, differing from them by containing one less carbon atom and one less unsaturated linkage, have been isolated and characterized. They contain two terminal-methyl groups. The process by which these constituents of “pyrethrolone” were obtained is described. I ~ L T S V I L L EMD. , REFERENCES (1) L.4FORGE AND BARTHEL, J. erg. Chem., 9, 242 (1944). (2) (a) STAUDINGER AND RUZICKA, Helv. Chim. Acta, 7, 212 (1924). (b) GILLAM AND WEST, J . Chem. Soc., 671 (1942). (c) LAFORGE AND HALLER, J. Org. Chem., 2,546 (1938). (3) BARTHEL, HALLER, AND LAFORGE, Soap and Sanit. Chem., 20, No. 7,121 (1944). (4) JOSHEL,Ind. Eng. Chem., Anal. Ed., 15,590 (1943).
