EPA-650/2-74-040
July 1974
Environmental Protection Technology Series
I
55
a
» . . t » « » » . f . » « r * » i » » « » . . » * » . • I
•.'.•.•.*.*.'.».•.'.*.*.*.*.*••••.'.'.' .*.»•* «».1 *.*.•**.• •*•".•'.•
-------
RESEARCH REPORTING SERIES
Research reports of the Office of Research and Development, U.S. Environ-
mental Protection Agency, have been grouped into series. These broad
categories were established to facilitate further development and applica-
tion of environmental technology. Elimination of traditional grouping was
consciously planned to foster technology transfer and maximum interface
in related fields. These series are:
1. ENVIRONMENTAL HEALTH EFFECTS RESEARCH
2 . ENVIRONMENTAL PROTECTION TECHNOLOGY
3. ECOLOGICAL RESEARCH
4. ENVIRONMENTAL MONITORING
5. SOCIOECONOMIQ ENVIRONMENTAL STUDIES
6. SCIENTIFIC AND TECHNICAL ASSESSMENT REPORTS
9. MISCELLANEOUS
This report has been assigned to the ENVIRONMENTAL PROTECTION
TECHNOLOGY series. This series describes research performed to
develop and demonstrate instrumentation, equipment arid methodology
to repair or prevent environmental degradation from point and non-
point sources of pollution. This work provides the new or improved
technology required for the control and treatment of pollution sources
to meet environmental quality standards.
-------
EPA-650/2-74-040
SYNTHESIS AND PURIFICATION
OF CARCINOGENIC POLYNUCLEAR
AROMATIC HYDROCARBON STANDARDS
by
Dr. Robert Fuhrmann, Allen Tunick,
and Louis Palmer
Allied Chemical Corporation
Corporate Research and Development
Morristown, New Jersey
Contract No. 68-02-0545
Project No 26 AEK, 42
Program Element No. 1AA010
EPA Project Officer: James E. Meeker
Chemistry and Physics Laboratory
National Environmental Research Center
Research Triangle Park, North Carolina 27711
Prepared for
OFFICE OF RESEARCH AND DEVELOPMENT
U.S. ENVIRONMENTAL PROTECTION AGENCY
WASHINGTON, D. C. 20460
-------
EPA REVIEW NOTICE
This report has been reviewed by the National Environmental Research
Center - Research Triangle Park, Office of Research and Development,
EPA, and approved for publication. Approval does not signify that the
contents necessarily reflect the views and policies of the Environmental
Protection Agency. nor does mention of trade names or commercial
products constitute endorsement or recommendation for use.
This document is available to the public for sale through the National
Technical Information Service, Springfield, Virginia 22161.
-------
TABLE OF CONTENTS
Page
L
ABSTRACT v
1.0 INTRODUCTION 1
2.0 SUMMARY 1
2.1 Benzo(a)pyrene 1
2.2 Benzo(a)anthracene 1
2.3 Benzo(e)pyrene 1
2.4 Chrysene 1
2.5 Benzo(b)fluoranthene 2
2.6 Benzo(k)fluoranthene 2
3.0 PROPOSED SYNTHESES OF COMPOUNDS IN PHASE I
(from T. P. No. DU-72-8312) 2
3.1 Benzo(a)pyrene 2
3.2 Benz(a)anthracene 5
3.3 Benzo(e)pyrene 6
3.4 Chrysene 7
3.5 Benzo(b)fluoranthene 7
3.6 Benzo(k)fluoranthene 9
4.0 DISCUSSION OF EXPERIMENTAL RESULTS 10
4.1 Benzo(a)pyrene 10
4.2 Benzo(a)anthracene 11
4.3 Benzo(e)pyrene 11
4.4 Chrysene 12
4.5 Benzo(b)fluoranthene 12
4.6 Benzo(k)fluoranthene 13
ill
-------
TABLE OF CONTENTS (Cont'd)
5.0 EXPERIMENTAL 13
5.1 Purification of Pyrene 13
5.2 Pyrene-1-carboxaldehyde 14
5.3 Purification of Hexahydropyrene 14
5.4 3- [l,2, 3, 6, 7 ,8-Hexahydro-4-pyrenoyl] -
propionic acid (HPPA) 15
5.5 4- £l,2, 3, 6, 7, 8 -Hexahydropyr eny l] -
butyric acid (HPBA) 15
5.6 1,2,3,6,7 ,8,9,10,11,12-decahydrobenzo (e)-
pyrene-9-one (DBPO) 16
5.7 1,2,3,6,7,8,9,10,11,12-decahydrobenzo (e)-
pyrene 17
5.8 Chrysene 18
5.9 9- 2-chlorobenzylidene -fluorene (CBF) 18
5.10 Benzo (b) fluoranthene 18
5.11 Purification of Acenaphthenequinone 19
5.12 7 ,12-dicyanobenzo (k) fluoranthene
(DCBF) (F. Goetz) 19
5.13 7 ,12-benzo (k) f luoranthenedicarboxamide
(BFDC) 19
5.14 Benzo (k) fluoranthene 20
6.0 ANALYSIS AND PURIFICATION OF FINAL PRODUCTS 21
6.1 Introduction 21
6.2 Purification and Analysis of Chrysene 26
6.3 Analysis and Purification of Benzo (b) -
fluoranthene 27
6.4 Analysis and Purification of Benzo (k) -
fluoranthene 29
iv
-------
ABSTRACT
This report details the experimental work performed under
Phase I of Contract 68-02-0545 which called for the prepar-
ation of samples of the following six condensed poly-
cyclic aromatic hydrocarbon compounds at a purity exceeding
99.9+%:
(1) Benzo(a)pyrene - (20 g.)
(2) Benzo(a)anthracene - (10 g.)
(3) Benzo(e)pyrene - (10 g.)
(4) Chrysene - (10 g.)
(5) Benzo(b)fluoranthene - (10 g.)
(6) Benzo(k)fluoranthene - (10 g.)
Due to termination of the contract before completion, only
the chrysene compound was completed to the required purity
and quantity. Four of the other compounds were completed
only partially through the steps in their syntheses. The
efforts of a subcontract on the sixth compound, Benzo (a)-
anthracene, were unsuccessful.
Experimental details as well as analytical methods and
safety procedures developed during the course of this work
are described herein.
-------
1.0 INTRODUCTION
This report details the experimental work performed under
Phase I of the contract #68-02-0545, based on Technical
Proposal No. DU-72-8312 to the Environmental Protection
Agency. Phase I called for the preparation, in purity
exceeding 99.9%, of samples of the following six con-
densed polycyclic aromatic hydrocarbons:
(1) Benzo(a)pyrene - (20 g.)
(2) Benzo(a)anthracene - (10 g.)
(3) Benzo(e)pyrene - (10 g.)
(4) Chrysene - (10 g.)
(5) Benzo(b)fluoranthene - (10 g.)
(6) Benzo(k)fluoranthene - (10 g.)
Due to termination of the contract before completion of
Phase I, not all of the above compounds are available for
shipment under the terms of the contract. The available
compounds and intermediates will be shipped in bulk,
according to the latest instructions from the contracting
authority.
2.0 SUMMARY
2.1 Benzo(a)pyrene
The second step of the eight step synthetic sequence
was completed and pyrene-1-carboxaldehyde is ready
for shipment. However, difficulties encountered in
step 3 prompted us to propose an alternate approach,
discussed in more detail in section 4.0 of this
report.
2.2 Benzo(a)anthracene
Initial plans called for the obtainment of commer-
cial material and development of purification and
analytical methods. Materials ordered from Princeton
Organic and Aldrich Chemical were not delivered at
termination of the contract by the E.P.A.
2.3 Benzo(e)pyrene
Four out of five synthetic steps have been completed.
Sufficient quantities of the immediate precursor for
compound (3) have been prepared and this compound
will be shipped.
2.4 Chrysene
Sufficient quantities have been prepared in adequate
purity and the compound will be shipped.
- 1 -
-------
2 . 5 Benzo(b)fluoranthene
Sufficient quantity has been prepared and in a purity
which may be sufficient, but has not been clearly
demonstrated.
2 .6 Benzo(k)fluoranthene
The synthesis of this compound has been completed.
Since only one run of the last synthetic step has
been completed, the amount of material prepared is
only 9 g. However, sufficient synthetic precursor
is available to prepare additional amounts of com-
pound 6. Purification procedures have also been
developed. The available amount of compound 6 in
impure state, plus its immediate precursor, will be
shipped.
3.0 PROPOSED SYNTHESES OF COMPOUNDS IN PHASE I
(from T.P. No. DU-72-8312)
Phase I will consist of the synthesis of the following
compounds:
3.1 Benzo(a)pyrene
Pyrene (I), the starting material, will be of the
purest commercial grade (zone refined). Prior to
use, the purity of the starting material is checked
by a combination of chromatographic and fluorometric
techniques (and further purification if necessary).
Previously published1-3 Friedel-Crafts synthesis of
benzo(a)pyrene from pyrene will not be used because
of the lack of selectivity of these conditions for
1-substitution on pyrene. Instead, the Vilsmeier-
Haack synthesis'*^ of pyrene-1-carboxaldehyde (II)
will be effected. The aldehyde (II) is freed from
pyrene as much as possible by distillation and
1 J. W. Cook et.al., J. Chem. Soc. , 1933, 398.
2 W. Winterstein et.al., Ber., 68, 1082 (1935).
3 H. Vollmann et. al., Ann. 531, 48 (1937).
4 M. deClercq and R. H. Martin, Bull. Soc. Chim. Beiges,
64_, 367 (1955) .
5 N. P. Buu-Hoi et.al., J. Chem. Soc., 1958, 48.
- 2 -
-------
crystallization (Pyrene will be completely removed
later before it can be involved in side reactions) .
If impurities beside traces of pyrene are present,
these are removed by chromatographic absorption.
2-(2,4,10-Trioxa-3-adamantyl)-ethyl bromide (A) is
prepared by the procedure of Stetter6 >7 from
2-bromopropionitrile and the phosphorane (B) , prepared
by the reaction of the phosphonium salt from A and
triphenylphosphine and n-butyl lithium in tetrohydro-
f uran , is allowed to react with aldehyde II. The pro-
duct, 1- (l-pyrenyl)-3-(2,4 ,10-trioxa-3-adamantyl) -
propene-1 (III) is
2) BuLi,THF
reduced to 1- (1-pyrenyl) -3- (2 ,4 ,10-trioxa-3-adamantyl) -
propane (IV) with platinum catalyst in a hydrogen
atmosphere (slight hydrogenation of the polycycle
at this stage will be inconsequential, but will be
prevented by careful control of the amount of hydrogen
absorbed) . The ortho ester IV is hydrolyzed with
acid to 4- (1-pyrenyl) -butyric acid (V) which can be
freed of any traces of pyrene by continuous extraction
of an aqueous solution of the sodium salt. The acid
is cyclized to 7 ,8 , 9 ,10-tetrahydrobenzo (a) pyrene (VII)
which is carefully dehydrogenated with palladium
on alumina (charcoal contains hydrocarbon impurities)
in specially purified mesitylene as solvent in a
nitrogen atmosphere to give the desired benzo (a) pyrene
(VIII). If dehydrogenation is not absolutely complete,
the dehydrogenation process is repeated and, if neces-
sary, chromatographic absorption is applied.
6 H. Stetter and K. H. Steinacker, Ber. 85, 451 (1952)
7 H. Stetter and K. H. Steinacker, Ber. 87, 205 (1954)
— 3 —
-------
CH3
POCl-
B
0 (page 3)
II
PtO
H30
III
IV
V
HF
press
W.K.
VII
pa,Ai.2.o3
mesitylene
VIII
* Wolff-Kishner reduction
- 4 -
-------
3.2 Benz(a)anthracene
The starting material, 1-bromonaphthalene (I), will
be of the highest purity available and will be fur-
ther purified by fractional freezing and low temper-
ature crystallization after a careful distillation.
The Grignard reagent from I in tetrahydrofuran is
reacted with high purity phthalic anhydride (partic-
ularly free of quinones and of benzoic acid) to give
the keto acid II which is cyclized with benzoyl
chloride containing a trace of sulfuric acid to
7,12-dihydrobenz(a)-anthracene-7,12-dione (III) which
is purified by distillation and, if necessary, chrom-
atographically purified to give purest material. The
quinone is reduced to the hydrocarbon benz(a)anthracene
(IV) by zinc dust melt and subsequently chromatograph-
ically purified1 -** .
1) Mg,THF
2)
0COC1
cat.
An,ZnC.l2 /rrr
=\ heat
0
III
IV
i K. Elbs, Ber. 19_, 2209 (.1886).
2 S. Gabriel and J. Coleman, Ber. 33, 446 (1900)
3 G. Heller and K. Schulke, Ber. 41_, 3627 (1908)
4 C. Graebe, Ann. 340, 254 (1905).
- 5 -
-------
3.3 Benzo(e)pyrene
Reduction of pyrene1"3 results in the formation of
mostly 1,2,3,6,7,8-hexahydropyrene (I) under con-
trolled conditions. The commercially available
material must be freed of other isomers by recrys-
tall^zation followed by zone refinement and/or
chromatography. The synthesis I through VI is then
applied during which the ketone IV is chromatograph-
ically purified. (Steps III - VI are substantially
the same as V - VIII in 3.1.)
A1C13
HF
OH
press,
III
W.K.*
0
Pd,Al203
mesitylene
* Wolff-Kishner reduction
1 J. W. Cook and C. L. Hewett, J. Chem. Soc., 1933, 401
2 I. Kagehira, Bull. Chem. Soc. Japan, 6_, 241 (1931).
3 W. Treibs and G. Meuer, Ber. 91, 1910 (1958).
- 6 -
-------
3.4 Chrysene
Purest commercial chrysene contains benzo (b)carbazole
as the main impurity. This can be removed by KOH
fusion, followed by potassium fusion and distillation
from potassium (repeated twice). Final purification
is done by column chromatography- If anthracene or
higher annulated analogues are suspected impurities,
they will be removed by maleic anhydride fusion and
caustic extraction prior to final purification.
3.5 Benzo(b)fluoranthene
Fluorene (I) (highest commercial purity--e.g., zone-
refined) is converted to methyl 9-fluorene carboxy-
late (II) by literature procedure, then cyanoethylated
(III) with acrylonitrile and base catalyst. Compound 3
is saponified and dehydrated to the anhydride (IV)
with acetic anhydride. This compound is isomerized
to the keytone V by heating with A1C13 (nitrobenzene)
and decarboxylated to VI by heat (Cu catalyst).
Synthesis to this stage is analogous to literature
procedure. (A. Campbell and S. H. Tucker, J. Chem. Soc.
1949, 2623) The synthesis of the final product VI - X
follows the general procedure given in 3.1, Steps
II - VIII.
OCH-
III
1) BuLi
2) C0.2
>
3) CH3OH
cat. H30~
+
2.) H30
3) Ac20
OCH-:
base cat.
- 7 -
-------
heat
Cu cat.
1) B (page
2) H2PtO
V
VI
Pd,Al203
=f
raesitylene
1) H30
>
2) HF
press.
VII
VIII
1) W.K.*
2) Pd,Al303
mesitylene
IX
* Wolff-Kishner Reduction
- 8 -
-------
3.6 Benzo(k)fluoranthene
The synthesis of benzo(k)fluoranthene is effected
via literature2 procedures (I - V). The starting
material, acenaphthene (I), is highest purity,
zone-refined material.
K2Cr207
HOAc
II
base cat.
,1) OH ,H20
2) HoO+
heat
Cu cat.
Ill
IV
V
Literature references are as follows:
i Steps I - II, Org. Synth. Coll., Vol. Ill, p. 1.
2 Steps II - V, H. Maurev et.al., C. R. Acad. Sci. Paris,
223, 951 (1946) .
-------
The following sections detail the work performed and final
status of each of the six compounds listed above. Experi-
mental details as well as analytical procedures and results
are given in Sections 5 and 6 of this Report.
4.0 DISCUSSION OF EXPERIMENTAL RESULTS
4.1 Benzo(a)pyrene
The synthesis originally proposed by F. Goetz (see
below) called for condensation of pyrene-1-carboxalde-
hyde with a Wittig reagent, (B), prepared by a separate
6-stage synthetic sequence, as follows:
EtOH
HC1
JJH2C1
EtOH
OH
n-BuLi,
THF
BrCH2CH2CN
/
BrCH2CH2C-0
0
0_P=CHCH0C-0
BrCH2CH2C-OEt
BrCH0CH0C-OEt
2 2t
OEt
03P
Br
G
03PCH2CH2C-
= B
* cis-phloroglucitol
from hydrogenation
of phloroglucinol
Pyrene was purified and formylated by two different
procedures, only one of which, the Villsmayer-Haack
synthesis using N-methylformanilide and POCl^,
afforded sufficiently pure pyrene-1-carboxaldehyde.
The preparation of the previously unreported reagent
(B) was found to involve unexpected difficulties, and
the synthesis was discontinued. Before termination
of the contract, a different approach, based on direct
Friedel-Crafts acylation with succinic anhydride under
conditions which afford high selectivity toward
1-substitution, was planned as outlined below:
0H
AlClo
H-,NNH-
KOH
Diethylene
Glycol
- 10 -
-------
The rest of the synthesis would be as outlined in the
original proposal. The commercial availability of
compound VI, Aldrich 18,061-0, presents another alter-
native if it can be adequately purified.
Alternatively, pyrene-1-carboxaldehyde might be
utilized to synthesize compound V in the following
way:
ArCHO — » ArCHCH2CH2CHO
CIMgCH2CH2CH(OEt)
THF
then H+, H20 OH
> ArCH2CH2CH2C02H
VI
The pyrene-1-carboxaldehyde was found to contain no
impurities detectable by thin layer chromatography
and is available for shipment.
4.2 Benzo(a)anthracene
Benzo(a)anthracene, with guaranteed purity of 99.9%,
was ordered from Princeton Organics, Inc. Shipment
of this material was not made by the time that work
on the present contract was terminated, so the order
was cancelled. This compound, in 99% purity, was
also ordered from Aldrich Chemical Company, but also
not received before contract termination.
4.3 Benzo (e)pyrene
1,2,3,6,7,8-Hexahydropyrene was purified by recrystal-
lization from abs. ethanol and the resulting material
was found to contain no TLC-detectable impurities.
Friedel-Crafts acylation with succinic anhydride and
aluminum chloride in purified nitrobenzene by a modi-
fied procedure based on the method of Cook and Hewett
(J. Chem. Soc. 398(1958)) produced a good yield of
1,2,3,6,7,8-hexahydro-4-pyrenoylpropionic acid which
was recrystallized from benzene. Wolff-Kishner reduc-
tion (Huang-Minion modification) gave the corresponding
hexahydropyrenylbutyric acid in very good yield. The
product was purified via its sodium salt, then by re-
crystallization (twice) from methanol. Friedel-Crafts
cyclization with anhydrous hydrofluoric acid at 25°
- 11 -
-------
gave a very good yield of 1,2 ,3 ,6 ,7 ,8,9,10,11,12-deca-
hydrobenzo(e)pyrene-9-one which was purified by recrys-
tallization first from ethanol and methanol, then from
ethanol alone. Further purification by column chroma-
tography on silica gel (benzene elution) followed by
recrystallization from ethanol gave material free of
impurities detectable by thin-layer chromatography.
Reduction to the corresponding decahydrobenzo (e)pyrene
was accomplished by the Huang-Minion modification of
the Wolff-Kishner reduction, which afforded a very
good yield of product purified by recrystallization
from benzene. Further purification was necessary to
remove a colored impurity, and a satisfactory chroma-
tographic scheme was developed. However, contract
termination prevented further work at this point. The
impure 1,2,3,6,7,8,9,10,11,12-decahydrobenzo(e)pyrene
is available for shipment.
4.4 Chrysene
Chrysene with guaranteed minimum purity of 99.9% was
purchased from Princeton Organics, Inc., and analytical
methods based on thin-layer, high-pressure liquid, and
gas-liquid partition chromatographies were developed
for purity determination. Three volatile impurities,
totaling 0.2% by peak area on GLPC (flame ionization
detection), were observed, and the material was re-
turned to the supplier. A sample of Chrysene obtained
from Aldrich Chemical Company and found to be ~ 96%
pure by GLPC was sent to Prof. A. R. McGhie at the
University of Pennsylvania for zone-purification. The
purified material was analyzed by the same methods and
found to be of > 99.9% purity. This material is avail-
able in sufficient quantity and is ready for shipment.
4.5 Benzo(b)fluoranthene
The synthesis originally outlined in the contract pro-
posal was abandoned when a simpler alternative synthesis
was found to be satisfactory. The condensation of
fluorene with 2-chlorobenzaldehyde was performed by the
Method of Hammer, Stauner and Chardonnens (Helv. 49,
1723 (1966)) in methanolic sodium methoxide. The re-
sulting 9-/2-chlorobenzylidene7-fluorene was obtained
in good yield and recrystallized from ethanol. Although
the product displayed a lower melting point than the
literature value, it was found to afford benzo(b)fluor-
anthene in a form readily purified to the level required,
The dehydrohalogenative ring closure was based on
British Patent 459,108 (1/1/37), example 1, and the
- 12 -
-------
method of Badger and Spotswood (J. Chem. Soc. 1959, 1635).
Benzo(b)fluoranthene was obtained in high yield and
recrystallized from benzene/ethanol. Chromatography on
silica gel afforded material free of any TLC-detectable
impurities. Further purification by zone-refining
(Prof. McGhie) produced material with one TLC-detectable
impurity. The determination of the absolute amount of
this^impurity was not undertaken due to contract
termination. This material is available for shipment,
as is a large quantity of relatively crude benzo(b)-
fluoranthene which was recrystallized from ethanol/ben-
zene, but neither chromatographed nor zone-refined.
4.6 Benzo(k)fluoranthene
This synthesis followed the general outlines of that
originally proposed. Acenaphthenene quinone (Aldrich)
was purified by recrystallization from glacial acetic
acid and condensed with o-phenylenediacetonitrile
(Aldrich) in piperidine solution as described by
Orchin and Reggel JAGS, 73, 436 (1951)). The resulting
7,12-dicyanobenzo(k)fluoranthene was purified by re-
crystallization from N,N-dimethylacetamide and then
hydrolyzed to the corresponding dicarboxamide with
potassium hydroxide in 2-ethoxyethanol containing a
small amount of water. The benzo(k)fluoranthene-7,12-
dicarboxamide was purified by recrystallization from
N,N-dimethylformamide. Conversion to benzo(k)fluo-
ranthene by means of heating with 100% phosphoric acid
at 200° was complicated by foaming, and a low yield of
crude product was obtained. Sufficient precursor
dicarboxamide remains to produce enough crude material
for final purification to obtain the required amount
for the contract by an improved procedure (see experi-
mental) . An apparently satisfactory purification
procedure has also been developed for the crude benzo (k)-
fluoranthene. Contract termination precluded further
work in this area.
5.0 EXPERIMENTAL
5.1 Purification of Pyrene
The sample of Aldrich pyrene had a light yellow color
and contained a number of components detected by thin
layer chromatography (TLC). Analysis by gas chroma-
tograph (GC) indicated 99.0% purity by peak area. A
method was developed by column Chromatography to purify
the pyrene.
- 13 -
-------
A column 4.0 cm x 26.0 cm was packed with basic aluminum
oxide, and carbon tetrachloride as a solvent. A concen-
trated solution of 200 g pyrene in carbon tetrachloride
was passed down the column and washed with additional
carbon tetrachloride. The flow fate was not controlled
as the process was similar to a filtration. The impur-
ities were adsorbed at the top of the column where a
dark brown area formed. A brown cast developed over
the column as the pyrene moved down, the effluent how-
ever was colorless. A greenish-yellow color appeared
in the eluent after short exposure to air and light.
The solvent was removed by vacuum distillation and the
pyrene obtained was white. The recovery was nearly
quantitative.
GC analysis indicated 99.9% pyrene and only trace im-
purities present. TLC analysis showed no impurities
present.
This technique was repeated twice using a fresh column
each time and over 600 gm of pyrene was purified.
5.2 Pyrene-1-carboxaldehyde
The method of Vollman, Becker, Corell and Streeck
(Ann., 531; 107 (1937)) was used. To a solution of
266 g. Eastman N-methylformanilide in 200 ml. dry
o-dichlorobenzene kept under dry Nitrogen, was added
dropwise, over two hours with stirring and external
cooling (to maintain T < 25°C), 266 g. of B & A phos-
phorous oxychloride. To the resulting clear, dark red
solution was added 197 g. of purified pyrene (see above)
and the mixture was heated and stirred at 90 - 95°C for
2 hours. Upon cooling, a precipitate formed, and it
was collected by suction filtration, washed well with
benzene and then added to distilled water to afford the
crude aldehyde which was co-llected by filtration and
recrystallized from ethanol. A first crop of 101.0 g.,
M.P. 126.8 - 127.6° (lit. 126°), a second crop of
8.2 g. M.P. 126.5 - 127-0°, and a third crop of 4.3 g.,
M.P. 123 - 124.5° were obtained. Total yield was 517.
Crops I and II were free of impurities detectable by
thin layer chromatography.
5.3 Purification of Hexahydropyrene
The sample of hexahydropyrene from Aldrich Chemical Com-
pany was analyzed by thin layer chromatography (TLC) which
revealed the presence of two impurities. The product
has the largest Rf (Chromatogram XXIV, analytical sec-
tion) . Due to the large quantity of reagent required, a
chromatographic purification procedure was not developed.
- 14 -
-------
It was found that the impurities could be removed by
a slow recrystallization of the sample from absolute
ethanol. A 500 g. sample of hexahydropyrene was dis-
solved in one liter of absolute ethanol at 70°C. The
solution was then allowed to cool slowly over a 12-hour
period in an insulated container. TLC analysis of
material recrystallized in the above manner indicated
that both impurities had been removed .
5.4 3-(J,2 /3 ,6 ,1 , 8-Hexahydro-4-pyrenoy^| -propionic acid (HPPA)
The procedure of Cook and Hewett (J. Chem. Soc . , 398
(1933)) was employed with a modified work-up as follows:
A solution of 43 g. Eastman succinic anhydride in 415 ml.
Aldrich nitrobenzene was slowly treated with 120 g.
B & A anhydrous aluminum chloride with stirring and cool-
ing. Then 90 g. of purified (see above) S-hexahydro-
pyrene was added gradually with stirring, and the mixture
was stirred overnight at room temperature. The mixture
was then poured into ice and water and concentrated
hydrochloric acid was added to dissolve the aluminum
salt precipitates. The mixture was then extracted three
times with a total of 2 liters chloroform, and the com-
bined extracts were washed with water and evaporated to
leave a thick slurry of the crude product in nitroben-
zene. This was thinned with benzene, cooled, suction-
filtered and the residue washed well with cold benzene.
Recrystallization from benzene afforded, after drying in
vacuo, 88.2 g. (66%), M.P. 176.5 - 178°C (lit. 173.5°).
Another crop of 12.0 g., M.P. 176.0 - 178°, was obtained
(total 75% yield) from the mother liquors. No TLC
analysis was deemed feasible at this stage.
5.5 4-0,2 ,3 ,6 ,7 ,8-HexahydropyrenyI} -butyric acid (HPBA)
A mixture of 77 g. (0.25 mole) of HPPA, 33 g. B & A
85% KOH, 16.9 g. Eastman 95% hydrazine and 250 ml.
Eastman diethylene glycol was heated gently until the
KOH went into solution and then refluxed for 1 hour
(T = 155°C) . Water was then distilled off until the
temperature reached 205°C, and refluxing was maintained
for 4.5 hours during which time the temperature was
218 - 224°C. The mixture was cooled, poured into water,
acidified to pH2 with HCl , filtered, and the residue
washed well with water. After drying in vacuo, 75.2 g.
crude material was obtained, wich afforded, after two
recrystallizations from methanol , 32.9 g. M.P. 130-133°.
The mother liquors yielded 37 g. which was converted to
the sodium salt with 5% sodium bicarbonate solution.
- 15 -
-------
The salt was filtered, washed with water, benzene, water
again, and then acidified with HC1 and extracted into
benzene. The benzene solution was extracted twice with
water, dried over anhydrous magnesium sulfate, and
evaporated to yield 34.8 g. Recrystallization from
200 ml. methanol afforded 27.7 g. M.P. 132 - 134°C.
This was combined with the other crop to afford 60.6 g.
(82.4%). One more recrystallization from —500 ml.
methanol afforded 58.2 g., M.P. 130 - 133°C. Again,
TLC analysis was not deemed feasible and the material
was carried through the next step of the synthesis.
5.6 1,2,3,6,7,8,9,10,11,12-decahydrobenzo(e)pyrene-9-one (DBPO)
To 58.0 g. (197 mmol) HPBA in a 1 liter polyethylene
bottle, was added 340 g. Matheson anhydrous liquid
hydrogen fluoride, and the resulting mixture was stirred
magnetically until the starting material was completely
dissolved. The purple solution was left standing at
room temperature for 70 hours and the HF was then re-
moved by purging with N2 into a solution of potassium
hydroxide in water. The residue was treated with aqueous
potassium carbonate and then taken to pH 11 with potas-
sium hydroxide. The resulting mixture was extracted
three times with a total of 1500 ml. diethyl ether, and
the combined extracts were washed twice with water, dried
over MgSC>4, and evaporated to leave 50.8 g. (93%) of
the crude ketone as a dark orange solid. This was re-
crystallized from 700 ml. ethanol-methanol (2:1) to
afford 44.8 g. dark orange crystals, M.P. 147 - 148.5°C.
Another recrystallization as above gave 40.7 g.,
M.P. 147.5 - 148.5° lighter orange crystals.
Thin layer chromatographic (TLC) results indicated the
material contained several components with lower R£
values than the product. Since the sample was very sol-
uable in benzene and had good resolution from the impur-
ities on silica gel, a method of purification was
developed by column chromatography.
A column 8 cm x 40 cm was packed with silica gel and
benzene. A solution of 39.2 g. sample in 150 ml benzene
was placed on the column and washed with additional
benzene. The flow rate was maintained at 5 cc per min-
ute. As the sample moved down the column, an orange
colored band moved ahead of the main component. This
material had not been detected by TLC as its concentra-
tion in the sample was very low. (This material was
isolated.) The other impurities remained near the top
- 16 -
-------
of the column. Movement of the components on the column
was checked periodically by examining the column with
long-wave ultraviolet light.
The product was isolated as it eluted from the column
and the solvent was removed by vacuum distillation. The
recovery was 99%. After recrystallization from absolute
ethanol, TLC analysis indicated the absence of any
impurities.
5.7 1,2,3,6,7,8, 9 ,10 ,11,12-decahydrobenzo(e)pyrene
A mixture of 35.4 g. (128 mmoles) purified (DBPO), 27.4 g.
B & A 85% potassium hydroxide, 13.5 g. Eastman 95% hydro-
zine and 200 ml. Eastman diethylene glycol were heated
gently to dissolve the KOH, then refluxed for one hour
(T = 150°C). Water was distilled off until the temper-
ature reached 210°C, and reflux was maintained for four
hours, during which time some condensation of product
accumulated on the condenser. The mixture was cooled,
poured into water and extracted with benzene. Some solid
remained undissolved at this stage. The benzene solution
was washed with water, dried over MgS04, and evaporated
to leave 11.9 g. yellow solid. The undissolved solid was
rinsed out of the separatory funnel, washed with methanol
and dried to afford 22 g. This was combined with^the
11.9 g. and recrystallized from benzene to afford 27.1 g.
(81%) of large yellow prisms, M.P. 190 - 195° (lit 196 -
197 ). A second crop of 1.8 g., M.P- 187 - 193° was
obtained by recrystallization of 5.1 g. material left
upon evaporation of the mother liquors. Sublimation of
the second crop was not effective for purification. The
27.1 g. first crop was set aside while a purification
scheme was worked out as follows:
Analysis of the material by thin layer chromatography (TLC)
revealed the presence of three components. The product
was separated from two impurities by large Rf differences
in the system used (Chromatogram XXVIII, analysis section).
A method to purify this material was developed by column
chromatography- A column 2.0 cm x 20.0 cm was packed
using Mallinckrodt CC-7 silica gel in hexane, toluene
(1:1). To this column 100 mg of the sample was applied
in the above solvent system, and the flow rate set at
0.2 ml per minute. The movement of the bands was moni-
tored by briefly exposing the column to long wave ultra-
violet light (360 my), the bands appearing as fluores-
cent areas. The product was eluted from the column and
the solvent distilled off under vacuum. The solid
obtained was colorless, and TLC analysis revealed no
impurities.
- 17 -
-------
Plans were made to purify the entire sample using a
10 cm x 50 cm column, however, the contract was termin-
ated before this could be completed.
5.8 Chrysene
See Section 6.
5.9 9-£2-chlorobenzylidene7-fluorene (CBF)
In two separate preparations a total of 93 g. (0.66 mole)
Aldrich 2-chlorobenzaldehyde, 33.2 g. (0.60 mole)
fluorene, 250 ml. MC & B 4.35 M (1.11 mol) sodium
methoxide in methanol, and 1425 ml. B & A anhydrous
methanol were refluxed for eight hours. The yellow
oil which precipitated was separated, and the methanol
layer was reduced about two-thirds on the rotary evap-
orator. The newly precipated yellow oil was added to
the previous quantity and the total was triturated with
dilute aqueous HC1, then with water to afford 155.1 g.
(91%) of a clear yellow oil which resisted attempts at
crystallization but crystallized spontaneously after
standing at -10°C for several days. Several recrystal-
lizations from anhydrous ethanol failed to give material
with the reported melting point of 71°C. A main crop
of 96.9 g. M.P. 57 - 61°C was obtained along with
several smaller crops. A small sample of this material
was carried through the next step of the synthesis (see
below) and was found to afford benzo(b)fluoranthene in
good yield and in a form readily purified by column
chromatography, so no further purification or analysis
was undertaken on the impure CBF.
5.10 Benzo(b)fluoranthene
A96.5g. (0.33 mole) sample of CBF, 400 g. B & A 85%
KOH, and 1000 ml. Aldrich quinoline were refluxed
(T = 230°C) with good stirring for 3 hours. The mix-
ture was cooled to 130°, poured into a mixture of
1800 ml. cone. HC1 and 20 kg. ice in a large glass
carboy. The resulting mixture was filtered, and the
residue was washed with water, dissolved in ~1300 ml.
warm benzene, quickly extracted with water and the
benzene solution evaporated. The residue was dried
in vacuo overnight, then recrystallized from 2.5 liters
benzene-ethanol (1:2) to afford 51.0 g. (61%) of fine,
shiny, grey-brown needles, M.P. 167.5 - 168.0°C
(lit. 167°, Badger and Spotswood).
Details of final purification will be found in Section 6.
- 18 -
-------
5%11 Purification of Acenaphthenequinone
Thin layer chromatographic analysis of an Aldrich
Chemical sample of acenaphthenequinone revealed the
presences of several impurities. The material was pur-
ified by recrystallization from glacial acetic acid.
Long brown needles were obtained, M.P. 260 - 261°C.
TLC analysis of this material revealed one black spot
on a purple background in the presence of ultraviolet
light, 254 my (Chromatogram XXIX, analytical section).
5.12 7,12-dicyanobenzo (k)fluoranthene (DCBF) (F. Goetz)
At 10°C, separate solutions of 60 g. (0.33 mole) purified
1,2-acenaphthenequinone in 250 ml. freshly distilled
piperidine and of 70 g. (0.45 mole) purified o_-phenylene-
diaectonitrile in 350 ml. of the same solvent~were pre-
pared. The acenaphthenequinone solution was added over
20 min. with stirring at 10 - 15°C to the solution of
the dinitrile. Then 150 ml. more piperidine was added,
and the temperature was allowed to rise to 40°C with
stirring. At this point precipitation of the product
made stirring difficult, and the mixture was left stand-
ing at room temperature for two days. The mixture was
then diluted with 1000 ml. "Spectrograde" DMF and fil-
tered. The residue was carefully washed with four
1000 ml. portions of DMF, then with methanol, and fin-
ally dried to yield 90.2 g. (89%) of a light yellow
solid (see below).
Due to the limited solubility and high melting point of
this compound, thin layer and gas chromatographic analy-
sis were not attempted. From the crude material, a
100 g. sample was recrystallized from eight liters of
N,N-dimethylacetamide, then washed with N,N-dimethyl-
formanide, methanol, and diethyl ether. 88 g. of small
yellow needles were obtained which decomposed at
358 - 360°C.
5.13 7,12-benzo(k)fluoranthenedicarboxamide (BFDC)
An 87 g. sample of DCBF was added to a solution of 1.8 kg
B & A 85% KOH in 9 1. Aldrich 2-ethoxyethanol and 720 ml.
water at 100°C and the resulting mixture was refluxed
with stirring for 48 hours. The mixture was cooled to
100°C diluted with 6 1. water and allowed to, cool to
30°C with constant stirring. The off-white product was
filtered, washed first with a mixture of 4 1. H20 and
2 1. 2-ethoxyethanol then with 6 1. methanol, and air
dried.
- 19 -
-------
Limited solubility and decomposition at higher temper-
ature hindered the analysis of this intermediate.
Purification was accomplished by recrystallization
from N,N-dimethylformamide. (Glacial acetic acid was
not used due to insufficient solubility.) Three crops
of crystals were obtained:
Crop 1, 35.1 g., M.P. 419 - 421°C
Crop 2, 28.1 g., M.P. 418°C
Crop 3, 2.3 g., M.P. 413°C
5.14 Benzo(k)fluoranthene
To 1300 ml. (2.8 kg) 100% H3P04, (prepared from 1190 ml.
(2006 g.) 85% H3P04 and 791 g. P2°s) in a two liter r.b.
flask was added 35.0 g. BFDC (batch 1). The mixture was
stirred magnetically and heated. When the temperature
reached 160° much foaming occurred. (Suggestion for
better results: Add dicarboxamide gradually to well
stirred (mech.) H3?04 at -—160° then heat to 200° for
8 hours.) The foam that overflowed was collected, washed
well with water, 5% sodium bicarbonate, water again,
and dried to leave 7.4 g. solid. The remainder of the
main reaction mixture was heated to maintain 220° for
4 hours with frequent swirling (since magnetic stirring
was inefficient). The mixture was quenched by pouring
onto excess ice and the aqueous suspension extracted
with a total of 2.5 1. benzene. The benzene solution
was washed with water, 5% NaHCC>3' an<^ then dried over
MgSC>4 and flashed to leave 17.1 g. solid. This was
recrystallized from benzene (hot filtration to remove
insoluble material) to afford 5.9 g. M.P. 214.5 - 215°C.
The 7.4 g. material obtained from overflow above was
recrystallized in the same way to afford 1.4 g.
M.P. 215.5 - 215.5°C. The benzene mother liquors from
both crops were combined and concentrated to afford a
third crop of 2.2 g. M.P. 214.5 - 215°C.
See Section 6 for purification scheme.
- 20 -
-------
6<0 ANALYSIS AND PURIFICATION OF FINAL PRODUCTS
6.1 Introduction
Analysis of the aromatic hydrocarbons and their inter-
mediates listed in this report was accomplished by
three techniques, liquid (L.C.), gas (G.C.) and thin
layer chromatography (T.L.C.). T.L.C. was used as the
main screening technique in detecting impurities.
When a final product or intermediate was found to con-
tain an impurity, methods of purification were developed
by recrystallization, column chromatography or zone
refining.
The purified final products in each synthesis were ana-
lyzed by all the T.L.C. systems listed in Table I and
by the liquid and gas chromatographic techniques devel-
oped to analyze the crude materials. The application of
these techniques insures that all the detected impuri-
ties have been removed by the purification procedure
and the material has a minimum purity of 99.9%.
Table I lists the adsorbents and solvent systems used
in the screening program. All of the chromatographic
adsorbents with the exception of magnesium hydroxide1
and Porapakz T were obtained from Brinkmann Instruments,
Inc.3. Eastman4 chromagram silica gel sheets were also
used.
Layers of magnesium hydroxide, Porapak T, acetylated
cellulose and acetylated polyamide were prepared in the
laboratory using Chromaflex T.L.C. plates5. This tech-
nique gave the best and most reproducible layers in
the shortest period of time. The other adsorbents used
were obtained in a prepared state from the suppliers.
The charge transfer systems were prepared by soaking
the plate for three minutes in an acetone solution of
the complexing agent; the plate was then air dried for
12 hours and kept in a dark area. It was found that
prolonged exposure to light, and possibly air, cause
the layer to darken, decreasing the sensitivity of the
analysis.
All T.L.C. plates were developed in Chromaflex develop-
ing tanks5, then air dried until no odor of solvent
could be detected. The plates were then analyzed in
the presence of long and short wave ultraviolet light
(360, 254 mp) and with an iodine vapor chamber. A small
21 ~
-------
walk-in dark room was constructed on the lab bench, and
long and short wave ultraviolet lamps were mounted 45°
to the bench surface. A black cloak of polyethylene
was worn to prevent the reflection of fluorescent light
onto the plate. This light is created by the inter-
action of the ultraviolet light and the normal laboratory
clothing; its presence hinders the analysis.
Liquid chromatographic analysis was carried out using
an 830 DuPont liquid chromatograph7 equipped with an
ultraviolet detector (254 my). Many problems were
noted with this equipment, and excessively long equi-
libration times were required before reproducible
data could be obtained.
Gas chromatographic analysis was done on a Packard
7400 gas chromatograph8 equipped with glass columns,
on-column injection, and a flame ionization detector.
All zone refining of the crude final products were
done by Professor Andrew McGhie at the University of
Pennsylvania.
Table IV lists the order in which copies of the
chromatograms appear.
- 22 -
-------
TABLE
T.L.C. Screening Systems
Adsorbents:
Alumina
Silica Gel.
Magnesium Hydroxide
Micro-crystalline
Cellulose
Porapak T
Cellulose 10%, 20%,
50%, 40%
Acetylated
Polyamide 11, 6, 6.6
Polyamide 11, 6, 6.6
Acetylated
Solvent Systems;
Carbon Tetrachloride
Benzene, Heptane, Chloroform (1:1:1)
Benzene
Methylene Chloride, Benzene (1:1)
Toluene
Toluene
Methylene Chloride
Tetrahydrofuran
Methyl Ethyl Ketone
DimethyIformamide, Water (1:1)
Methanol
Acetone
Ether, Methanol, Water (4:4:1)
Toluene, Ethanol, Water (4:17:4)
Tetrahydrofuran, DimethyIformamide,
Water (1:1:1)
Tetrahydrofuran, Diethyl ether,
Hexane (2:7:4) (1:2:2)
Toluene, Ethanol, Water (4:17:4)
Charge Transfer System
Benzene, Heptane (1:1)
Silica Gel
0.2% Caffeine
0.3% 2,4,7,-Trinitro-9-fluorenone
0.3% 1,3,5-Trinitrobenzene
- 23 _
-------
TABLE II
Suppliers of Equipment and Chromatographic Materials
Fisher Scientific Co.
52 Tadem Road
Springfield, New Jersey 07801
Waters Associates, Inc.
61 Fountain Street
Framingham, Massachusetts 01701
Brinkmann Instruments, Inc
Canteague Road
Westbury, New York 11590
Eastman Kodak Company
Rochester, New York 14650
Kontes
Vineland, New Jersey 08360
Ultra-Violet Products, Inc.
San Gabriel, California
E. I. DuPont Company, Inc.
Wilmington, Delaware 19898
8 Packard Instruments Company, Inc
2200 Warrenville Road
Downers Grove, Illinois 60515
-24 -
-------
TABLE III
Conditions for Liquid and Gas Chromatographic Analysis
of_
Aromatic Hydrocarbons
Liquid Chromatography;
830 DuPont Liquid Chromatograph
Column - One Meter ODS
Solvent System - Methanol, Water (6:4)
Flow Rate - One milliliter per minute
Pressure - 1000 P.S.I.
Temperature - 50°C
Detector - Ultraviolet 254 my
Attenuator - 4x, 0.04 OD full scale
Chart Speed - 0.2 inch per minute
Gas Chromatography;
Packard 7400 Gas Chromatograph F.I.D.
Column - 6 ft. Chromasorb G Coated with 1.0% OV-17
Carrier Gas - Helium 60 cc per minute
Detector Temperature - 210°C
Injector Port. Temperature - 210°C
Chromatogram IV - Isothermal 235°C, Chart Speed 0.5 inch
per minute
Chromatogram V - Program 10 minutes at 205°C then 2° per
minute to 235°C then held 10 minutes
- 25 -
-------
6 .2 Purification and Analysis of Chrysene
Two samples of commercial grade chrysene from Aldrich
Chemical Company and Eastman Organic Chemicals were
obtained and analyzed by T.L.C., L.C., and G.C. A
specially purified sample from Princeton Organics was
also included in the analysis. The chromatograms of
these analyses are presented in Chromatograms I thru V.
The results of this analysis show the Eastman sample
to be the poorest of the group, containing two impur-
ities . The Aldrich sample contains the same two
impurities, but one is present in only trace amounts.
The Princeton sample was by far the cleanest. How-
ever, it did not meet the purity specifications stated
in its purchase contract, and the sample was returned.
Attempts to effect a preliminary purification by column
chromatography on Mallinkrodt CC-7 silica gel with
toluene as a solvent showed only a minor improvement
(see Chromatogram VI for T.L.C. analysis of the three
fractions collected). The only impurity that was re-
moved from the bulk of the sample was the one produc-
ing the white colored spot with the lowest Rf.
A fresh sample of Aldrich Chemical Company chrysene
was submitted to Professor Andrew McGhie for zone re-
fining. The sample was successfully purified by the
following procedure:
(1) Approximately 50 gms of the chrysene supplied
(Aldrich Chemical Co.) was sublimed and charged
into a 15 mm O.D. zone melting tube. The sample
was then given 25 zone passes on a Sloan-McGowan
zone melter. After zoning, the tube was cut up
and seven samples taken for L.C. and G.C. analyses.
A green band was observed at the top of the Z.R.
tube and a red band at the bottom, indicating
impurities with K<1 and K>1.
(2) A center section of this zone melting tube was
recast and rezone melted in a 12 mm O.D. tube
for 70 zone passes. This tube was sectioned and
analyzed by L.C. A persistent impurity was ob-
served throughout the tube which was removed
only slowly by zone melting.
(3) Approximately 16.5 gms of the purest material
from this tube was then continuously chromato-
graphed under Argon through a 50 mm O.D. tube of
Woelm neutral alumina using benzene (A.C.S.
certified, Fisher) as eluant.
- 26 -
-------
After 21 hours, the chrysene had been eluted completely
leaving a 4 cm band of blue violet fluorescent impurity
at the top of a 50 cm Column. Chromatography was
carried out in the dark, using DuPont Kapton sheet as
light shield. The apparatus used was a modified
Sangster and Irvine unit. After this treatment, the
chrysene was recrystallized from the benzene eluant.
The purified sample received from Professor McGhie was
analyzed by all the T.L.C. systems listed in Table I
and the L.C. and G.C. procedures listed in Table III.
No impurities have been detected by these techniques,
and, thus, all of the detectable impurities in the
crude material have been removed. These analytical
results indicate this material to have purity of 99.9%.
(Refer to Chromatograms VII thru XVI).
6.3 Analysis and Purification of Benzo(b)fluoranthene
The synthesized sample of benzo(b)fluoranthene was
analyzed by a number of T.L.C. systems. The most
effective systems appear in Chromatogram XVII. This
chromatogram reveals the presence of at least three
impurities in the crude material.
A purification method based on the best T.L.C. system
was developed for column chromatography. A column
4.0 cm x 25.0 cm was packed with Brinkmann3 neutral
silica gel and toluene. The solvent was purified by
passing it through a precolumn of neutral alumina.
A 1.0 g. sample of crude benzo(b)fluoranthene was
applied to the column as a solution in toluene. It
was then eluted with toluene and the movement of the
bands was monitored by exposing the column to long-
wave ultraviolet light. The exposure time was kept
to a minimum to prevent photochemical reactions from
taking place. After the product was eluted from the
column, the solvent was distilled off under vacuum
and the material analyzed by T.L.C., L.C. and G.C.
techniques. T.L.C. analysis (Chromatogram XVIII)
revealed the presence of one spot for the product; no
impurity spots were detected. The L.C. and G. C.
techniques developed were not capable of resolving
the impurities in the crude material and were not
thought to be of value here. This purification was
then scaled-up as described below.
A column 7.5 cm x 33.0 cm was packed in the same man-
ner as above; 25.9 g. of benzo(b)fluoranthene was
dissolved in 750 cc of toluene and applied to the
- 27 -
-------
column. The toluene had been purified by passing it
through a precolumn of neutral alumina. A total of
25.2 g. of the applied sample was collected over
five fractions, a 97.3% recovery. The solvent was
distilled off under vacuum and the fractions analyzed
by T.L.C.
T.L.C. analysis (Chromatograms XIX, XX) showed frac-
tions one and two to contain one spot for the product.
The combined weight for these fractions is 5.8 g. The
chromatograms of fractions three and four show that
they might contain a trace impurity with a lower Rf
than the product. It was difficult to determine if
any impurity was present in this area due to the tail-
ing of the product. For this reason the fractions
were combined (18.9 g.) and submitted to Professor
McGhie for zone refining. The chromatogram of frac-
tion five showed that it contained the impurities of
the crude material (0.5 g.). This was expected as
all the impurities in the crude material have lower
Rf 's than the product in this system. (Fractions
3 and 4)
The column purified benzo(b)fluoranthene was zone re-
fined in the following manner:
Approximately 20 g. of benzo(b)fluoranthene was
pumped extensively for 2 days on a high vacuum
line to remove the occluded chromatographic sol-
vent (alcohol). The sample was then melted under
0.5 atmos. Argon into a 10 mm O.D. Pyrex zone
melting tube to give an ingot - 35 cm long. The
tube was sealed under - 0.75 atmos. Argon and
given 60 zone passes on a Sloan-McGowan zone
melter (zoning speed l"/hr., £/L = 1:20). The
upper two-thirds of the ingot was taken as pure
product.
Analysis of the zone refined benzo(b)fluoranthene by
T.L.C. (Chromatograms XXI, XXII) revealed the presence
of an impurity spot just under the product spot. Zone
refining of the sample did not improve its purity.
The nature and concentration of this impurity have
not been determined as no quantitative techniques
have been developed that are capable of sufficiently
resolving the impurity. Further work on the analysis
and possible purification of this material was stopped
due to the termination of the contract.
- 28 -
-------
6.4 Analysis and Purification of Benzo(k)fluoranthene
(see Section 5.14)
Three recrystallization fractions of benzo(k)fluor-
anthene were submitted for analysis and purification.
The fractions were analyzed by T.L.C. (Chromato-
gram XXIII) and found to contain several components.
A method of purification based on the T.L.C. separa-
tion was developed for column chromatography:
A column 7.5 cm x 33.0 cm was packed with Mallinckrodt
CC-7 silica gel and hexane-toluene (3:1). All sol-
vents were distilled before use. The three fractions
were to be combined totaling 11.5 g., dissolved in
the solvent system and applied to the column. The
product would have been eluted from the column and
the solvent distilled off under vacuum.
The above purification method was not applied to the
sample due to the termination of the contract.
- 29 -
-------
TABLE IV
List of Chromatograms of Analysis Section
Chromatogram
I thru V
VI
VII thru XVI
XVII
XVIII thru XX
XXI and XXII
XXIII
XXIV
XXV
XXVI
XXVII
XXVIII
XXIX
Title
Crude Chrysene
Column Purified Chrysene
Pure Chrysene
Crude Benzo(b)fluoranthene
Column Purified Benzo(b)fluoranthene
Zone Refined Benzo(b)fluoranthene
Crude Benzo(k)fluoranthene
Pyrene
Pyrene-1-Carboxyaldehyde
Hexahydropyrene
Decahydrobenzo(e)pyrenone
Decahydrobenzo(e)pyrene
Acenaphthenequinone
- 30 -
-------
Chromatogram I
Crude Chrysene
Solvent front
Solvent System;
x
Toluene
Y
Pentane
Diethyl Ether
(19:1)
Adsorbent;
t
Silica Gel
Eastman
Detection:
U.V. 254 mu
Iodine Vapor
Quantity;
4-y as 0.4% solution
Methylene Chloride
Code;
A - Princeton Organic
B - Aldrich Chemical
C - Eastman Organic
P - Purple
L,B - Light Blue
W - White
Dotted line indicates
trace amounts
Starting Spots
J/.B
i .'•••»,
B
B
31
-------
Chrornatogram II
Crude Chrysene
Solvent Front
Solvent System;
Tetrahydrofuran
Diethyl Ether
Hexane (1:1:1)
Adsorbent;
Polyamide 11
Brinkmann
Detection;
U.V. 254 my
Quantity;
8y as 0.4% Solution
Methylene Chloride
Code;
A - Princeton Organic
B - Aldrich Chemical
C - Eastman Organic
P - Purple
L,Y - Light Yellow
W - White
Starting Spots
32
-------
Chromatogram III
Crude Chrysene
Solvent Front
Solvent System:
Toluene
Ethanol
Water (4:17:4)
Adsorbent;
Cellulose 40%
Acetylated
Brinkmann
Detection:
U.V. (360,254 my)
Quantity:
16y as 0.4% Solution
Methylene Chloride
Code:
A - Princeton Organic
B - Aldrich Chemical
C - Eastman Organic
P - Purple
W - White
Dotted line indicates
trace amount and tailing.
Starting
at)
rX
v \J
to'vK
'" ';••'•;
'
''&> :M
("•
0
W
B
33
-------
i_J r~_r "rr1
! 1'- T
64X — ;
S-:: -:::
: :._ _ r r :
-------
Chromatogram V
L.C. Analysis of Aldrich Chemical Chrysene
i-U
I 1 I '" '
;i!';j';-l-
-n-4, '4-U-
; t i
J-^U t -I
> ! M
II
;•>!'!
-t I !
i ! £ ^ i- M i i . f i i ; :
rltf-rlT;1'-
':#;;!'I h
iH-i
....: o
0) •
UU-l-.-H.hL4- .l".:~:uu
iilTf ^r, i -••!'!
-------
Chromatogram VI
Column Purification of Chrysene
X
Solvent Front
Solvent System:
X
Toluene
Tetrahydrofuran
Diethyl Ether
Hexane (1:2:2)
Adsorbents:
X
Silica Gel
Eastman
Polyamide 11
Brinkman
Quantity:
4y as 0.4% Solution
Methylene Chloride
Code;
I,II,III -
Fraction from column
chromatography
purification of
Aldrich chrysene
P - Purple
Starting Spots
L,B
0 O O
i
36
II
IV
O_
III
r\
J
A
L,B
n in
-------
Chromatogram VI
Column Purification of Chrysene
X
Solvent Front
Solvent System:
X
Toluene
Tetrahydrofuran
Diethyl Ether
Hexane (1:2:2)
Adsorbents:
X
Silica Gel
Eastman
Polyamide 11
Brinkman
Quantity:
4y as 0.4% Solution
Methylene Chloride
Code:
I,II,III -
Fraction from column
chromatography
purification of
Aldrich chrysene
P - Purple
Starting Spots
L.B
0 o O
i
36
II
W
o_
III
A
\J
n in
-------
Chromatogram VII
Pure Chrysene
Solvent Front
Solvent System;
Toluene
Adsorbent:
Silica Gel.
Eastman
Detection;
U.V. (360,254 my)
Iodine Vapor
Quantity:
Numbers refer to j/spot.
as 0.4% Methylene
Chloride Solution
Color;
Purple
Starting Spots
8 12 16 20 24
37
-------
Pure Chrysene
Solvent Front
Solvent System;
Tetrahydrofuran
Diethyl Ether
Hexane (1:2:2)
Adsorbent;
Polyamide II
Brinkmann
Detection;
U.V. (360, 254 my)
Iodine vapor
Quantity;
Numbers refer to y/spot
as 0.4% Methylene
Chloride Solution
Color;
Purple
Starting Spots
\J
8 12 16
20 24
38
-------
Chromatogram IX
Pure Chrysene
Solvent Front
Solvent System;
X
Acetone
Methanol
Adsorbent:
PQrapak T
Waters Assoc.
Direction;
U.V. (360,254 my)
Iodine vapor.
Quantity:
4y as 0.4% Solution
Methylene Chloride
Color:
Purple
Starting Spots
BBC*.
39
-------
Chromatogram X
Pure Chrysene
A B
Solvent Front — —
Solvent System;
Benzene
Heptane (1:1)
Adsorbent;
Charge Transfer
Silica Gel
Brinkmann
A - 0.3% Caffeine
B - 0.3% T.N.F.
C - 0.3% T.N.B.
Detection:
Visible Light
Quantity:
Numbers refer to
y/spot as 0.4%
Methylene Chloride
Solution
Color;
Brown on tan
background
Code;
Dotted line indi-
cates solvent front
Starting Spots
! Br
^ 0
1 \J
. 4 8
i
Br
0 0
4 8
!
•
<
Br
o 0
!
r~
4 8
40
-------
Chromatogram XI
Pure Chrysene
\J
A
Solvent System;
Diethyl Ether
Methanol, Water
(4:4:1)
Quantity;
A - 8y
All other 4y
•per spot 0.4%
Methylene Chloride
Solution
Adsorbent:
Cellulose
W - 10% Acetylated
X - 20%
Y - 30%
Z - 40%
Brinkmann
Detection:
U.V. (360, 254 my)
Iodine Vapor
Color: Purple
Code; Solid line indicates
starting spots.
Dotted line indicates
solvent front.
41
-------
Chromatogram XII
Pure Chrysene
r\
r\
\J
L
JU-
4 8
Solvent System:
Diethyl Ether,
Methanol, Water
(4:4:1)
Adsorbent;
Cellulose 20%
Acetylated
12
16
2Q
24
28
32
Detection;
U.V. C360,254
Iodine Vapor
Quantity;
Numbers refer to y/spot
as 0.41 .Solution
Methylene Chloride
Color: Purple
Code:
Solid line indicates
starting spots.
Dotted line indicate!
solvent front.
42
-------
Chromatogram XIII
Pure Chrysene
r\
\J
12
16
20
24
28
32
Solvent System;
Toluene, Ethanol
Water (4:17:4)
Adsorbent;
Cellulose 30%
Acetylated
Detection;
U.V. C360,254 my)
Iodine Vapor
Quantity;
Numbers refer to y/spot
as 0.4% solution
Methylene Chloride
Color; Purple
Code;
Solid line indicates
starting spots.
Dotted line indicates
solvent front.
43
-------
Chromatogram XIV
L.C. Analysis of Pure Chrysene
TTTtjTTl—rr-
-------
Chromatogram XV
Analysis of Pure Chrysene
i i
•T: t
1 x 10
-10
-------
Chromatogram
60 ' .11.55 ! '..-HO' '. '.J.-..30 "-
70 .. . 6.0-.-,- !-, . 50,~ : , , .46
-------
Chromatogram XVII
Crude Benzo[b]Fluoranthene
Solvent front
Solvent System:
Toluene
Adsorbent;
Alumina
Brinkmann
Detection;
U.V. (360,254 my)
Quantity:
4y per spot as 0.4% Solution
Methylene Chloride
Color:
L,B - Light Blue
W - White
L,0 - Light Orange
Dotted line indicates slight
tailing.
Starting spot.
o
47
-------
Chromatogram XVIII
Purified Benzo[b]Fluoranthene - Small Column
Solvent front "- ^~~
Solvent System:
Toluene
Adsorbent;
Alumina
Brinkmann
Detection:
U.V. (360,254 my)
Quantity;
4y per spot as 0.4% Solution
Methylene Chloride
Color;
L,B - Light Blue
Starting Spot
48
-------
Chromatogram XIX
Purified Benzo[b]Fluoranthene - Large Column
Solvent front
Solvent System;
Toluene
Adsorbent;
Alumina
Brinkmann
Detection;
U.V. (360,254 my)
Iodine Vapor
Quantity;
4y per spot as 0.4%
Solution
Methylene Chloride
Color:
L,B - Light Blue
W - White
L,0 - Light Orange
Dotted lines indicate
tailing
Starting Spots
0
0
« i
'
4,
49
-------
Chromatogram XX
Purified Benzo [b] f luoranthene - Large^ Column
Solvent Front
Solvent System:
Toluene
Adsorbent:
Alumina
Brinkmann
Detection:
U.V. (360,254 my)
Iodine Vapor
Quantity;
4y per spot as 0.4%
Solution
Methylene Chloride
Color:
L,B - Light Blue
W - White
L,O - Light Orange
Dotted lines indicate
tailing
1 Starting Spots
L.B
50
-------
Chromatogram XXI
Zone Refined Benzo[b]fluoranthene
Solvent Front
Solvent System:
Toluene, Ethanol,
Water (4:17:4)
Adsorbent;
X
Polyamide 6.6 Acetylated
Y
Polyamide II Acetylated
Brinkmann
Detection:
U.V. (360,254 my)
Iodine Vapor
Quantity;
4y per spot as 0.4%
solution
Methylene Chloride
Color:
L,B - Light Blue
Starting Spot
L.B
51
-------
Chromatogram XXII
Zone Refined Benzo[b]fluoranthene
Solvent Front
Solvent System:
Benzene
Heptane
(1:1)
Adsorbent:
Charge Transfer
0.3% Caffeine Coated
on Silica Gel.
Brinkmann
Detection:
U.V. (360,254 my)
Iodine Vapor
Quantity:
4y per spot as 0.4% Solution
Methylene Chloride
Color:
L,B - Light Blue
On dark purple background
L,B
Starting Spot
52
-------
Chromatogram XXIII
Crude Benzo[k]fluoranthene
Solvent Front
Solvent System;
Hexane, toluene
(3:1)
Adsorbent;
Silica Gel.
Brinkmann
Detection;
U.V. (360,254 my)
Quantity;
4y as 0.4% solution
Methylene Chloride
Color;
L,B - Light Blue
W - White
L,Y - Light Yellow
Starting Spots
Y
B
53
-------
Chromatogram XXIV
Pyrene
Solvent Front
Solvent System;
Carbon Tetrachloride
Adsorbent;
Alumina
Brinkmann
Detection;
U.V- 254,360 my
Quantity:
10y as 0.5% solution
Methylene Chloride
Code:
A - Crude
B - Purified
Dotted area indicates
trace amount
Color:
L,G - Light Green
W - White
Starting Spots
54
O-v.
A
B
-------
Chromatogram XXV
Pyrene-1-Carboxyaldehyde
Solvent System:
Benzene
Heptane
Chloroform
(1:1:1)
Adsorbent;
Alumina
Brinkmann
Detection:
U.V. 360 mu
Quantity:
10y as 0.5% Solution
Methylene Chloride
Code:
A - Crude
B - Purified
Color:
R,0 •
L,B •
O
W
L,Y •
Red Orange
Light Blue
Orange
White
Light Yellow
Solvent front
Starting Spots
R.O
o
A
B
55
-------
Chromatogram XXVI
Hexahydropyrene
Solvent System:
Tetrahydrofuran
Diethyl Ether
Hexane
(10:35:20)
Adsorbent;
Polyamide 11
Brinkmann
Detection:
U.V 254 my
Quantity:
10y as 1.0% solution
Methylene Chloride
Code;
A -• Crude
B - Purified
Color:
W - White
D,B - Dark Blue
Solvent Front
Starting Spots
— W —
r\
-D,B
B
56
-------
DecahydrobenzoIe]pyrenone
Solvent front
Solvent System:
Benzene
Adsorbent:
Silica Gel
Eastmann
Detection:
U.V. 254, 360 my
Color;
Yellow - Green
Quantity;
5y as 0.1% solution
Methylene Chloride
Starting spot
57
-------
Chromatogram XXVIII
Decahydrobenzo[e]pyrene
Solvent Front
Solvent System:
Hexane
Toluene
(3:1)
Adsorbent;
Silica Gel
Eastmann
Detection;
U.V. 254 mu
Quantity:
10y as 0.5% solution
Methylene Chloride
Color:
Y,G - Yellow-Green
W - White
Starting spot
W ,
-^-
58
-------
Chromatogram XXIX
Acenaphthenequinone
Solvent front.
Solvent System:
Methylene Chloride
Benzene (1:1)
Adsorbent:
Silica Gel.
Eastmann
Detection:
U.V. 254 my
Color;
Black
Quantity:
5y as 0.1% solution
Starting spot,
59
-------
TECHNICAL REPORT DATA
(Please read Instructions on the reverse before completing)
1. REPORT NO.
EPA-65Q/2-74-Q40
4. TITLE AND SUBTITLE
2,
3. RECIPIENT'S ACCESSIQWNO.
SYNTHESIS AND PURIFICATION OF CARCINOGENIC
POLYNUCLEAR AROMATIC HYDROCARBON STANDARDS
7. AUTHOR(S)
Dr. Robert Fuhrmann, Allen Tunick and Louis Palmer
8. PERFORMING ORGANIZATION REPORT NO.
6. REPORT DATE
iltllY 1974
i. PERFORMING <
ORGANIZATION CODE
9. PERFORMING ORGANIZATION NAME AND ADDRESS
Allied Chemical Corporation
P. 0. Box 1021R
Morristown, New Jersey 07960
10. PROGRAM ELEMENT NO.
1AA010
11. CONTRACT/GRANT NO.
68-02-0545
12. SPONSORING AGENCY NAME AND ADDRESS
Office of Research and Development
U.S. Environmental Protection Agency
Washington, D. C. 20460
13. TYPE OF REPORT AND PERIOD COVERED
Final
14. SPONSORING AGENCY CODE
15. SUPPLEMENTARY NOTES
16. ABSTRACT
Experimental work was performed with the objective of preparing six condensed
polynuclear hydrocarbons at a purity equal tovor exceeding 99.9+^. The amounts
to be prepared were 20 g. of benzo(a)pyrene, and 10 g.each of benz(a)anthracene,
benzo(e)pyrene, chrysene, benzo(b)fluoranthene, and ber»zo(k)fluoranthene.
At the time the contract was terminated., chrysene was the only compound whose
quantity and purity had been shown to meet the specifications. Four other compounds
benzo(a)pyrene, benzo(e)pyrene, benzo(b)fluoranthene, and benzo(k)fluoranthene were
in the process of purification and synthesis. No progress was obtained in the
purification efforts on. benz(a)anthracene.
Experimental details as well as analytical methods and safety procedures
developed during the course of the work are described.
17.
KEY WORDS AND DOCUMENT ANALYSIS
DESCRIPTORS
b.lDENTIFIERS/OPEN ENDED TERMS
c. COSATI Field/Group
Aromatic Polycyclic Compounds
Synthesis Crystallization
Purification
Gas Chromatography
Zone melting
Chromatographic analysis
Primary Standards
18. DISTRIBUTION STATEMENT
Unlimited
19. SECURITY CLASS (This Report)'
Unclassified
21. NO. OF PAGES
63
20. SECURITY CLASS (Thispage)
Unclassified
22. PRICE
EPA Form 2220-1 (9-73)
60 -
------- |