- Email: [email protected]
Separation of polythiadioxopiperazine antibiotics by thin-layer chromatography
NOTES
592 CHROMz 5013 Separation
of polythiadioxopiperazine
antibiotics
by thin-layer
chromatography In recent years the number of 2,5-epipolythia-3,6-dioxopiperazine antibiotics has increased significantly due to isolation of new members of the series and by chemical transformations of the naturally-occurring compounds. By extensive chemical and/or X-ray crystallographic investigations the structure of gliotoxinl (I), sporides (III, X = z) was determined and min293 (II, X = 2, R = OH) and dehydrogliotoxin4 other closely related compounds have since been isolated or synthesized3~~. More recently several compounds related to acetyl aranotinO-8 (IV) have been described
CH,OH (I)
CH,OH
-
(VI
_n
cl_
OH
OH
I
”
(i.e. WD) and the synthesis of r,4-dimethyl-S+piperazinedione (VI) has also been reported. Chetomin, another sulfur-containing natural product is also believed to contain the epipolythiadioxopiperazine moiety lo. Most of the above compounds po+ sess biological activity 10-12.This communication deals with the thin-layer chrotnatol
Issueclas NRCC No.. 114g6
J. Citrorrtatog.,53
(1970)
592-594
593
NOTES
graphic behavior of fifteen of these sulfur-containing compounds. The separation of polysulfides (II, X = I, 2, 3, 4, R = OH) and (III, X = I, 2, 3, 4) differing only in the number of sulfur atoms in the 2,s bridge is particularly noted.
Gliotoxin (I) was isolated from the fungus P. tevlikowslzii4 and converted into dehydrogliotoxin (III, X = z) as described 4. Treatment of dehydrogliotoxin (III, X = 2) with triphenylpl~osphine13 gave (III, X = I) and with hydrogen polysulfide gave (III, X = 3) and (III, X = 4). Sporidesmin (II, X = 2, R = OH) was isolated from P. chavtamvlz and reaction with triphenylphosphine gave (II, X = I, R = 0H)a. Sporidesmin El4 (II, X = 3, R = OH), B (II, X = 2, R = H)i6 and D (VII)10 were also isolated from the above fungus and the tetrasulfide (,II, X = 4, R = OH) has also been isolated and prepared by treatment of (II, X = 2, R = OH) with hydrogen Bisdethiodi(methylthio)-acetylaranotin (V) (BDA) and acetylaranotin polysulfidei’. (IV) were gifts of Eli Lilly Laboratories and the 1,4-dimethyl-2,5-piperazinedione (VI) was prepared by the method of TROWN~*. Chetomin was obtained from Chaetowzi~nn cochZiodesll. CJtronantogva$Jtic jwocerlacres The solvent systems used and the RF values for the compounds (I-VII) are given in,Table I. Commercially available Merck Silica Gel 1T,,4 plates (20 x 20 cm, thickness 0.25 mm) were used and 5 ,ul of an 0.0002 y0 solution (in chloroform) was applied to the plate. Detectiou All compounds TABLE I RF VALUES
FOR
IT, s -= I, r< = 01-I II, S = 2, R = OH III, III, III, III, IV V VI VII
s S S
s s s s
= 3, R = OH = 4, R = OH = 3-, R = H
= = = -
I 2 3 4
under short wave UV light. When
EPIPOLY1’HIA~IOXOPIPrsRAZINE
I
II, II, II,
were visible
ANTIBIOTICS
0.15
0.50 0.53 0.57 0.54 0.49 0.59 OS-l7
0.2G
0.58
0025
0.55 OS49
0.15
o.oG
0.20
0.09
0.32 0.38 0.36
o.oG
0.31 0.39 0.39 o-25 0.41 0.20 0.44 0.43 0.23 0.32 0.23 0.22 0.24 0.09
O.I’i
0.21
0.09 0.07 0.03
0.39
0.11
0.25 0.40 o-35 0.19 0.30 0.20 0.16
0. I I
0.19
0.05
the chromato-
0.22 0.15 0.05 0.15 0.11
0.13 0.04 0.01
*
0.17 0.24 0.24
0.23
0.17 0.1
I
0.G5
0.09
O.G5
0.07 0.01
0.5G
0.01
0.51
.r. Ch’omafo.g.,
0*.55
53 (1970)
592-594
NOTES
594
grams were sprayed with neutral aqueous 5 Y0 silver nitrate solution spots of black. (or brown) silver sulfide were obtained where the sulfur compounds were located. ’ The results obtained (Table I) show that by judicious choice of solvent systems separation of most of these sulfur-containing compounds can be obtained. In the dehydrogliotoxin series (III, X = 1-4) all four polysulfides as well as gliotoxin (I) are clearly separated with the solvent system benzene-ethyl acetate (4 : I). The monoand tetrasulfides of sporidesmin (II, X = I and 4, R = OH) have different Rp in benzene-ethyl acetate (4: I) (Rp 0.32 and 0.21) and in benzene-ether-acetic acid (70: 30: I) and (RF 0.31 and o.z~), from the chromatographically similar di- and trisulfides (II, X = z and 3, R = OH). Separation of the latter two compounds is readily effected in hexane-tert.-butanol (9: I). The above solvent systems have all been used for the separation of large quantities of material (100-300mg) by preparative thin-layer chromatography. Sporidesmin B (II, X = 2, R = H) is generally less polar and sporidesmin D (VII) more polar than the other members of the series in most of the solvent systems used. Chetomin and (VI) are the two most polar compounds and both BDA (V) acetylaranotin (VI) are readily separable in all five solvent systems. With chloroform as solvent the values are low for a single elution however with repeated elution (four times) excellent separations of most of the structurallysimilar polysulfides is obtained. Atlaatic Regional Laboratory, NatioptaL Halifax, Nova Scotia (Canada)
Research
Council
of Canada,
R.
RAHMAN
s. SAFE A. TAYLOR
i AND A. M. MAT~~IESON,A&Z Crysl., 23 (1gG7) 439. I FRIDRICHSONS 18 (1965) 1043. 2 J._~?IZIDRICHSONS AND A. M. MATHIESON,AC~~C~~~~., citccl. 3 S. SAFE AND A. TAYLOR, J. Chsm. Sec. (C), (x970) 432 and rcfcrcnces AND L.C.VINING, J. Chcm. Sot. (C), (1966) 1799. 4 G. LOWE, A.TAYLOR I<. C. MURDOCK AND R. 13.ANGIER, Chern. Cownun, (x970) 55. N. R-NELSON AND J.H. VAN DEN HENDE, J.A t~z.C/~rtt.Soc.,go (1968) 65x9.. 2 D, B.~ONSULICH, I?. A. MILLER, P. W.TROWN, W. FUL~~OR, J. KARLINEI~ AND G. MORTON, Biochm. Biophys.. 7 Rcs. Commun., 33 (1968) 219. L.L. HUCKSTEP, D. H. LIVELY, D.C.DELONG, M. M. MAIE+I AND N. NEUSS, s R. NAGARAJAN, ,J. Am. Chent., Sot., go (rg6S) zgSo. go (IgGS) 65x7. 9 J. W. MONCRIEFF, J, Am. ,Chem. Sot., AND G. N. WOGAN (Eclitors), J3iochevrtistvy of SOIIIC FoodDo~m IO A. TAYLOR, in R. I. MATELES Micvobial Toxins, MIT press, 1967, p. Gg and refercnccs citccl. AND A, TAYLOIZ, Can. J. Micvobiol., 13 (1967) 1577 and rcfcrcnccs cited. II D. BREWER 12 N.N~~~~,L.D.UOECK,D.R.BRANNON,J.C.CLINE,D.C.I~ELONG,NI.GORMAN,L.L.I-IUCKSTEP, D. H. LIVELY, J. MABE, M. M. IMAIXSFI,13. B. MULLOY, R. NAGARAJAN, J. 1). NELSON AND W. M. STARK, in Anti~~&robiaZ A&vrl and CItemotlwnpy, American Society for Microbiology, 1968, p. 213. S. SAFE AND A. *rAyLO& Chem. Common. (rgGS) 1571. 13 D. BI~EWER, R. RAHMAN, S. SAFE AND A. TAYLOII,,{. Clbettt. Sot. (C), (1969) 1665. I4 R. RAHMAN, E. P. WHITE AND R. J. RBI~AIIABI,J. Chem. Soc.,(IgG3) 3172. I.5 J. W. RONALDSON,A.TAYLOR, J. CJwn. Sot. (C), (1909) 1564. IG W. D. JA&IIESON, R. RAI-IMAN AND A. TAYLOR, to be published. 17 S. SAFE AND A\.TAYLOR, Res. Commun., 33 (1968) 402. IS 1’. W. l’t~ow,u, Bsochem. Biophys.
Received
August 28th, 1970
J. Clwomntog.,
53 (1970)
592-594
592 CHROMz 5013 Separation
of polythiadioxopiperazine
antibiotics
by thin-layer
chromatography In recent years the number of 2,5-epipolythia-3,6-dioxopiperazine antibiotics has increased significantly due to isolation of new members of the series and by chemical transformations of the naturally-occurring compounds. By extensive chemical and/or X-ray crystallographic investigations the structure of gliotoxinl (I), sporides (III, X = z) was determined and min293 (II, X = 2, R = OH) and dehydrogliotoxin4 other closely related compounds have since been isolated or synthesized3~~. More recently several compounds related to acetyl aranotinO-8 (IV) have been described
CH,OH (I)
CH,OH
-
(VI
_n
cl_
OH
OH
I
”
(i.e. WD) and the synthesis of r,4-dimethyl-S+piperazinedione (VI) has also been reported. Chetomin, another sulfur-containing natural product is also believed to contain the epipolythiadioxopiperazine moiety lo. Most of the above compounds po+ sess biological activity 10-12.This communication deals with the thin-layer chrotnatol
Issueclas NRCC No.. 114g6
J. Citrorrtatog.,53
(1970)
592-594
593
NOTES
graphic behavior of fifteen of these sulfur-containing compounds. The separation of polysulfides (II, X = I, 2, 3, 4, R = OH) and (III, X = I, 2, 3, 4) differing only in the number of sulfur atoms in the 2,s bridge is particularly noted.
Gliotoxin (I) was isolated from the fungus P. tevlikowslzii4 and converted into dehydrogliotoxin (III, X = z) as described 4. Treatment of dehydrogliotoxin (III, X = 2) with triphenylpl~osphine13 gave (III, X = I) and with hydrogen polysulfide gave (III, X = 3) and (III, X = 4). Sporidesmin (II, X = 2, R = OH) was isolated from P. chavtamvlz and reaction with triphenylphosphine gave (II, X = I, R = 0H)a. Sporidesmin El4 (II, X = 3, R = OH), B (II, X = 2, R = H)i6 and D (VII)10 were also isolated from the above fungus and the tetrasulfide (,II, X = 4, R = OH) has also been isolated and prepared by treatment of (II, X = 2, R = OH) with hydrogen Bisdethiodi(methylthio)-acetylaranotin (V) (BDA) and acetylaranotin polysulfidei’. (IV) were gifts of Eli Lilly Laboratories and the 1,4-dimethyl-2,5-piperazinedione (VI) was prepared by the method of TROWN~*. Chetomin was obtained from Chaetowzi~nn cochZiodesll. CJtronantogva$Jtic jwocerlacres The solvent systems used and the RF values for the compounds (I-VII) are given in,Table I. Commercially available Merck Silica Gel 1T,,4 plates (20 x 20 cm, thickness 0.25 mm) were used and 5 ,ul of an 0.0002 y0 solution (in chloroform) was applied to the plate. Detectiou All compounds TABLE I RF VALUES
FOR
IT, s -= I, r< = 01-I II, S = 2, R = OH III, III, III, III, IV V VI VII
s S S
s s s s
= 3, R = OH = 4, R = OH = 3-, R = H
= = = -
I 2 3 4
under short wave UV light. When
EPIPOLY1’HIA~IOXOPIPrsRAZINE
I
II, II, II,
were visible
ANTIBIOTICS
0.15
0.50 0.53 0.57 0.54 0.49 0.59 OS-l7
0.2G
0.58
0025
0.55 OS49
0.15
o.oG
0.20
0.09
0.32 0.38 0.36
o.oG
0.31 0.39 0.39 o-25 0.41 0.20 0.44 0.43 0.23 0.32 0.23 0.22 0.24 0.09
O.I’i
0.21
0.09 0.07 0.03
0.39
0.11
0.25 0.40 o-35 0.19 0.30 0.20 0.16
0. I I
0.19
0.05
the chromato-
0.22 0.15 0.05 0.15 0.11
0.13 0.04 0.01
*
0.17 0.24 0.24
0.23
0.17 0.1
I
0.G5
0.09
O.G5
0.07 0.01
0.5G
0.01
0.51
.r. Ch’omafo.g.,
0*.55
53 (1970)
592-594
NOTES
594
grams were sprayed with neutral aqueous 5 Y0 silver nitrate solution spots of black. (or brown) silver sulfide were obtained where the sulfur compounds were located. ’ The results obtained (Table I) show that by judicious choice of solvent systems separation of most of these sulfur-containing compounds can be obtained. In the dehydrogliotoxin series (III, X = 1-4) all four polysulfides as well as gliotoxin (I) are clearly separated with the solvent system benzene-ethyl acetate (4 : I). The monoand tetrasulfides of sporidesmin (II, X = I and 4, R = OH) have different Rp in benzene-ethyl acetate (4: I) (Rp 0.32 and 0.21) and in benzene-ether-acetic acid (70: 30: I) and (RF 0.31 and o.z~), from the chromatographically similar di- and trisulfides (II, X = z and 3, R = OH). Separation of the latter two compounds is readily effected in hexane-tert.-butanol (9: I). The above solvent systems have all been used for the separation of large quantities of material (100-300mg) by preparative thin-layer chromatography. Sporidesmin B (II, X = 2, R = H) is generally less polar and sporidesmin D (VII) more polar than the other members of the series in most of the solvent systems used. Chetomin and (VI) are the two most polar compounds and both BDA (V) acetylaranotin (VI) are readily separable in all five solvent systems. With chloroform as solvent the values are low for a single elution however with repeated elution (four times) excellent separations of most of the structurallysimilar polysulfides is obtained. Atlaatic Regional Laboratory, NatioptaL Halifax, Nova Scotia (Canada)
Research
Council
of Canada,
R.
RAHMAN
s. SAFE A. TAYLOR
i AND A. M. MAT~~IESON,A&Z Crysl., 23 (1gG7) 439. I FRIDRICHSONS 18 (1965) 1043. 2 J._~?IZIDRICHSONS AND A. M. MATHIESON,AC~~C~~~~., citccl. 3 S. SAFE AND A. TAYLOR, J. Chsm. Sec. (C), (x970) 432 and rcfcrcnces AND L.C.VINING, J. Chcm. Sot. (C), (1966) 1799. 4 G. LOWE, A.TAYLOR I<. C. MURDOCK AND R. 13.ANGIER, Chern. Cownun, (x970) 55. N. R-NELSON AND J.H. VAN DEN HENDE, J.A t~z.C/~rtt.Soc.,go (1968) 65x9.. 2 D, B.~ONSULICH, I?. A. MILLER, P. W.TROWN, W. FUL~~OR, J. KARLINEI~ AND G. MORTON, Biochm. Biophys.. 7 Rcs. Commun., 33 (1968) 219. L.L. HUCKSTEP, D. H. LIVELY, D.C.DELONG, M. M. MAIE+I AND N. NEUSS, s R. NAGARAJAN, ,J. Am. Chent., Sot., go (rg6S) zgSo. go (IgGS) 65x7. 9 J. W. MONCRIEFF, J, Am. ,Chem. Sot., AND G. N. WOGAN (Eclitors), J3iochevrtistvy of SOIIIC FoodDo~m IO A. TAYLOR, in R. I. MATELES Micvobial Toxins, MIT press, 1967, p. Gg and refercnccs citccl. AND A, TAYLOIZ, Can. J. Micvobiol., 13 (1967) 1577 and rcfcrcnccs cited. II D. BREWER 12 N.N~~~~,L.D.UOECK,D.R.BRANNON,J.C.CLINE,D.C.I~ELONG,NI.GORMAN,L.L.I-IUCKSTEP, D. H. LIVELY, J. MABE, M. M. IMAIXSFI,13. B. MULLOY, R. NAGARAJAN, J. 1). NELSON AND W. M. STARK, in Anti~~&robiaZ A&vrl and CItemotlwnpy, American Society for Microbiology, 1968, p. 213. S. SAFE AND A. *rAyLO& Chem. Common. (rgGS) 1571. 13 D. BI~EWER, R. RAHMAN, S. SAFE AND A. TAYLOII,,{. Clbettt. Sot. (C), (1969) 1665. I4 R. RAHMAN, E. P. WHITE AND R. J. RBI~AIIABI,J. Chem. Soc.,(IgG3) 3172. I.5 J. W. RONALDSON,A.TAYLOR, J. CJwn. Sot. (C), (1909) 1564. IG W. D. JA&IIESON, R. RAI-IMAN AND A. TAYLOR, to be published. 17 S. SAFE AND A\.TAYLOR, Res. Commun., 33 (1968) 402. IS 1’. W. l’t~ow,u, Bsochem. Biophys.
Received
August 28th, 1970
J. Clwomntog.,
53 (1970)
592-594