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Current Progress in the Determination of the Polychlorinated Biphenyls
* Hk W^HiM.norcnt, P. Rkiciik, and II. S. Oixorr
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The vit products of a global technological society
*.In*vitably become componenta of Cha environment. Many of
.
thcaa waste eaterlala have the capacity to inflict peraanant
changes upon the ecosystem. Among the synthetic pollutants
which are accumulated by wildlife the chlorinated hydrocarbons
. of biocide origin appear to be tha most abundant. Residue
. levels in marina fish and birds now frequently surpass those
- 1 recorded in terrestrial and fresh-water species (1,2.3). la
i1
..
California, the concentrations of chlorinated hydrocarbons
in petrel, pelagic blrda which do not approach land axeept .
'.to breed on off-shore islands, are many times higher than
' those recorded in species inhabiting agricultural and urban
. areas (1). Ocher classes of environmental pollutants which
'.. might be similarly accumulated by organisms and similarly
. ' dispersed around Che world could be expected to share soma ` ;"* or all of the following properties of tha biocide derivatives* `
Which have contributed to their unexpected distribution. in;:' ' '
V -.the global ecosystem. \
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* `I . ' . .
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1) High world production. U. S. production of DDT in 1965
1M
hnni of EA.irwaawai.1 CoM.nua.lfv* A Tii*Wy. Vat i. Na. *. iwa. yyl.UArd hjr Sp.iaprr.VrfU. Urm Yarfc lar.
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vat 140 Billion pounds (4). Vorld yearly production la thorefore In tha order of 10** grant, only five orders of magnitude leas
than tha anount of carbon fixed by plants into organic natter (J).
Other pollutants which are present' In ecologically significant
amounts In the global ecosystem suet, like p,p'-DDE, therefore
derive fro* primary materials that are produced In comparable
omounte.
'
2) Chemical stability. p,p'-DDE, which may bs tho mpst abundant
of the eynthatic pollutants In the environment, le comparatively
rcelatant to degradation by the usual detoxification mechanlame
of vertebrataa, to microbial action, and to non-blologieal *. breakdown In the environment. Zte half-life, therefore, would
eppeer to ba groetor then ten yoere. .
1
f
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5) Insolubility In water. Water-soluble waste products will bo
diluted to a greater extent in,the see end may or may not bo
subsequently accumulated by organlame. tha non-poler nature of
p,p'--DDE and tho other chlorinated hydrocarbons explains why
they ere concentrated in fat tissue end why they are present la
* - *
i .
greatest amounts In- the terminal carnivores of food" chains.
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4) Mobility and ncrlnl dispersal. Even though vapor pressures
of potential pollutants might ba vary low, the amounts entering "
.tha atmosphere could become ecologically significant over periods
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of time, especially when rates of vaporisation era greatly . ` ' '
increased by codlstilletlon with water (6) Incineration of ' ' '
vaote mar veporlxac can be fu fallout.
Amor. . the arocl Their use
resins dc end resla
Uald
have long and blrdu polychlor evidence. 1) Haag si compounds with a cor chlorlne-M as bloddi 2) RctcntJ of peaks 1 cannot ba have rater, era ldanti
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...yeet* material* would also significantly increase the rates of
j^va^orlsacloa of some compound*. Tha global distribution of DOT
-etn bi fully explained only by aarlal dlaparaal vlch subsequent
^fallout, In conjunction with tranaport by water (2).
.Among Industrial product* which meat thcao critarla are
|>alberoclora, which era.mixture* of chlorinated polyphenyla.
CilJheir uaafulnpaa in tha manufacture ..of many plaocica, paints and .
.rpaslop dative* to a largo part from their chemical atablllty
.
4)and..raalatanca. to degradation*.
...
of ..Unidentified peak* produced by halogen-containing compound*
have.long been evident in gas chromatograaa of extracts of fish
o.end birds... The Identification of cost of these compounds as . 1
.^polychlorinated biphenyls (PCS) is now based upon the following /
- evidence.
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1) Kiss spectromctrlc analysts In Sweden. Kaas i.pectra of
e
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compounds present In extracco of fish and.se* birds obtained
,
with a combined gas chromatograph-maas spectrometer showed that '
chlorine-containing compounds which could not be identified 1 ?
.....a* blocideo were polychlorinated biphenyls (7).
2) Retention times In ens chronntogrnphlc nnalvsli. The majority .'
. of peaks in chromatogram* of extract* of fish and birds which
' ' " cannot be identified aa biocides that amargs after p.p'-DOE , ; .
have ratantlon times on both polar and non-polar columns which
* ... *
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*
' are identical with those of the principal PCB compound*. Tha
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^ major peaks produce ' characteristic pattern on each kind of
column. The same pattorn# of peak* aro evident in extracts of
wildlife fron around the world (1,2,8,9).
- '*
3) Chlorine content. - Mlcrocoulometrlc analysis shows that the
compounds-identified as PCS contain halogens. . '
4) Saponification and nitration. Unlike the peaks of toxaphene ;
. (10); p.p'-DDT, p,p'-DDT, and p,p'-DDD, tho PCS peeks ere not
' removed or displaced by dehydrohalogenatlon with alcoholic KOH.
Vlgorduanltratlon carried out at room temperature with fuming
nitric acid (11) removes the chromatographic peaks of both PCB
end the DOT compounds, but does not dostroy the toxaphene peaks.
At 100* the DDT compounds arc tetranltrated by this procedure v-'
- (12). The nitrated derivatives would not pass through the
coenonly used CI.C coluana. A milder nitration process, carried '
^
out at 0* with nitric acid rather, chan fuming nitric acid,
..
"destroys tho-DDT, but not the PCB peaks. . . ............
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3) Distribution in the global environment. Concentrations of
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PCS in fish and birds are highest in such industrial outfalls
'
as San Francisco Bay and San Diego Bay and concentration gradients
. apparently exist from these areas to regions more remote (1,2). ' 1
A conslatency in tho ratios of p,p'-DDE to PCB among tha birds
'
. of a given region suggested tha existence of regional fallout
.* *
patterns (1). PCB has been found in all aaa birds from tho
' \
Pacific Ocean so far analysed, but they could not bd detected *
' * -
in pcngul
/
compounds The
chromatog with thos DC-200:
QF-li
The colura Chromosort of 195*. underline^ on DC-200. peak emar On QF-1 cc with o,p'with tha c N interfarar shoulder c
J' As a
of tha val .recant lit
\ . reported i
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J in penguin eggs Iron the'Antarctic which did contain DDT
. compound# (1). .
.*' The retention tinea relative to p.p'-DDE, of peak# In . .
chrosatograas of extract* of fleh and blrda which are identical .
with thoae of PCS compound* in Cho coamorcial preparations ere!
OC-MO* 1.25, 1.48. 1.75. 2.05, 2.41, 2.50, 2.90. 3.41, 3.88,
. 5.53; dleldrini 1.00; p.p'-DDDi 1.27; p.p'-DDTi 1.68
` * QF-li 1.10, 1.33. 1.40. 1.65, 1.72. 2.14, 2.59, 3.23, 3.88, .
' 4.84; dleldrlnt 1.49; p.p'-DDDt 1.75; p.p'-DDTi 1.91
* The coluano employed were 10Z DC-200 and 3Z QF-1, both on
,
. Chroaoeorb U, 80-100 aeah and were aaintained at a temperature '
of 195*. The retention tinea of the three principal peak* are ..
. . . \/
.*
: ` underlined. The first of these, with a retention,tla* of 1.25 '
. *. on DC-200 columns emergen with p.p'-DDD. Another aajor PCB
.
; ' '' peak emerges slightly later than p.p'-DDT on DC-200 coluans.
t i,
. . On QF-1 coluans, cho compound with retention time 1.33 emergos
. with o,p'-DDT and cho compound with retention tlaa 1.75 interferes
. ^wlth the determination of p.p'-DDD. On this column there is no
. interference with p.p'-DDT. Dieldrln appears as n trailing ..
4.
' shoulder on the peak with retention time 1.40.
As a result of this interference, it appears that many , >
of the values of p.p'-DDD, p.p'-DDT and o,p'-DDT reported in
'
\ . j. t .
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. .recent literature are erroneoua. Tlu DDD velueo originally
... . , . #
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. reported in Pacific sea birds in a paper from chi* laboratory .
T '.y
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(13) were coo high because of PCS Interference and have
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subsequently been corrected (1).' p,p*-DDE is.(he moat abundsnt
i.i ; s-;::.- > r.
of the DOT compounds in the environment and there is no
significant PCS interference in the detcralnatlon of p,p'-DDE.
Consequently the total DDT residues reported in the past, before
Che extent of PCB interference was known, would not be greatly
changed after correction for this Interference. o,p'-DDT, which
has been shown to be an active estrogen (14,13) is present in
very low aaounts in the global environment, although o,p'-DDS
Is comparatively more abundant (1,3).
' When both DDT and PCB peaks are present in chronatograaa,
the clue to the relative amount of PCB cones fron the holght -
of those PCB peaks which do not Interfere with any of the DDT
coepounds. With the exception noted below, the three major
i peaks usually have approximately' the sane height on chromacogr
obtained with an electron capture detector. If the peak with
retention time 1.48 on DC-200 columns is approximately as high
as the "ODD" and "DDT" peaks, no or vary small amounts of
0 p,p*-DDD and p,p'-DDT are present. When one or both of those
peaks Is relatively higher than the PCB peak with retention tine
1.48, the amount of p,p'-DDD end p,p*-DDT can be estimated from
I
*
the changes in peak height with saponification. The sapooifi-
.\ * t
cation procedure is rapid, and elnce only-a relative change in
peek heights is determined, It naed not be quantitative. An
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aliquot of in ethyl si aqueous NsC the hexane have been t contributlo can thereby
PCS hn hydroxylase
/' partially r observed An accumulate that /the PC of oomparsb ' Since individual future, the (10), also present be . used (1,2) mlcrocoulom' PCB compoun> fish and bi by the coaa>
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aliquot of tho extract la refluxed for approximately 5 .hlnutcs
1I
In ethyl alcohol with 5Z KOil, to which hexane and concentrated
. ............................................ -.............................................................
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aqueous NaCl solution aro added in turn* Chromatography I
_thJiexanalayer shows chat the p.p'-DDD and p.o'-DDT peaka
have been displaced, but the PCS paaka are unchanged* The
. #'
contribution of PCS to tho peaka of the original chromatograms
'
can thereby ba estimated, provided the detector response la linear* .0
(l!;) ,_PCB^haa; been shown to ba a powerful inducor of steroid -
..^hydroxylases in birds .(1) and, with p,p'-DDE,' nay therefore bo '
^'.partially responsible for the aberrant calcium physiology
^observed in species of raptorial and fish-eating birds which
^accumulate chlorinated hydrocarbons .(16,17) *..'Xt. is probable '
... that the PCD In human food supplies would induce the .synthesis
' .of compsrAbla enzymes in men*....
_ ft...s#>-%. ...............................
...............
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" . . .. Since It 1s highly unlikely that primary standards of
-
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individual PCS compounds will be svaliable in tho foreseeable ' .
^ future,t the quantification procedures, like those for toxaphene
r'
(10), also, a mixture of polychlorinated coap.oundo, must for the
present ba approximate.* The quantification procedure we have *'
LmV C *1..' ** ..............` ............
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.t
^uaed (1,2) is based primarily, upon the. determination with s
Csm** l*c*.r.o..c..o...u..l.o..m. - otrlc dotheto.1r...o..f.....t.h..e' l ' tot*al ' ch lorine content of the . pPCB compounds emerging after p,p'-DDE. TJjo. chromatograms of .
fish and^blrdextracts most cloely_.resseabls those.produced -
vb^.tha^ ^osmerclal^eroclor .yhich^ Jia* an. average, .chlorine content '
as the HD3D" and "GOV' punks,, no or very'-a.-.'ll
'*
* -
p,"p -ODD and p.n'-DDT. jVo present. ^<11 ..no or both
.ai -
,
*
peaks 1* relatively hlgkoV^ char. V<?3 pe.-.k rfich roc*-.-.;.-. i. . .
1.43. the amount of p,^`-r/JO '..-.d , r,1-3&7 cun 'cu i?.;-:*;:. . . . .%
the rhnr.noi ir. *>cnk h--i
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-\ ..iof S4X." PCB concent rations In fish and birds were therefore
derived freathe chlorine valuta by aaeuaing chat the chlorine
, Vontenc vas also1 54Z. ` This procedure would oecAolcnal!/ include
..other chlorinated pollutants with the PCB. It aay, however,
be toore meaningful to measure total organic chlori--
,.of non-blocldc origin in tissues of wildlife chan parte per
'.pillion concentrations of individual compounds. This might
be e better index of physiological effecta such as enzyme
induction.
, Since we do not yet have in our laboratory the perriancnt
. use of a mlcrocouloaecric detector, we have had to depend upon
.electron-capture for routine quantification of PCB. Each PCB
compound was initially assumed to produce tho same peak height,
with the electron capture detector, as an equivalent amount of ,
) - p,p'-DDE. Amounts of sample injected into tho
r nogrepb
were adjusted in order to fall within cha linear response
range of the electron capturo detector. The amount of total
PCB considered as p,p'-DOE was then multiplied by a factor . (
which would yield a value cquaj. to that obtained with a micro- .
coulomctrlc detoctor. This procedure wae periodically checked
by electron capture determination of standard PC&
. Although this method yields approximate results, it does
. \
permit accurate determinations of relative concentrations. It
\<
. has shown.that 'some species of birds, notably certain raptorial
species, of PCB (1 end conci. eleo ahov concentrn
The and 1.33 chromato? Bay and 1 is preaen tho Gulf aerial fa Californl teated th selective in the et evaporate h> diehea co / 'then irra goralclda of peaks asaentlal height of was irrod
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species, ohcerwatcrs and pctrola, accumulate high concentratIona
of PCS (1). Terminal carnivores of food chain* nay accumulate
and concentrate the PCS present in prey epeclee (1). It has
also shown that regional fallout patterns exist (1), with highest
,Ui .
......................
...
...
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,
concentrations la Industrial regions.
....................................................
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. The PCB compound with retention time 1.25 on DC-200 columns'
and 1.33 on QP^l columns produces one of the prominent peeks in
chromatograms of extracts of fish and birds from San Francisco
Bay end In the commercial mixtures, yec It Is not proaent, or
Is present In relatively low amounts, In cxtracto of birds from
*!.:
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*
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the Cult of California. PCB la this area must derive from
.
' aerial fallout, slnco thore are few Industrial areas in Baja
.............................
California or along the coast of western Mexico. Wo thersfore
.
wi 1... '
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...
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tested the possibility that this compound might have been
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selectively degraded by ultraviolet light during Its transport
in the atmosphere. Standard PCB solutions In hexane vara '
evaporated to dryness in Petri dishes, or were applied to Petri
dishe-a...c..o..n...t.a..i.n..i.n...g....e...i.t.h...e..r. water of 3X NoC l solution, * they.wars
then irradiated at a distance of. 4 Inches with a 15-vatC
...........i..
.............. .... ........................................
.
germicidal lamp emitting at 2537 X. In each case the pattern
" -
....... .............. '
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of. pe'a.*k.s: *o'n - twh.ae chr.oI ,mat1 o*grams *of....C..h..e....i.r..r.a...d..i.a. te' d *'PCB w. asj*'-`
'
essentially unchanged, except for a relative, reduction in the
1 '. * .'lieu y 4 * * 1 , *
-
*.............
*
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*
height of this-peak. The affect, was. more pronounced when PCB
i. a* * t CV..I*
*
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1
yas irradiated on the. aqueous surfaces*. .. .^ V
: . > , V . . - .-/ * : *.
Alchcu.-.h
.\ . ' . . ` '
permit jccu.atc dote.Ai.*..`.tin-.*, a of r.-lACivt-
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haA shown
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Acknowlodgcmonta
Vo thank tho Monsanto Chemical Company for supplying samples , . of tho Aroclora. Tho National Sclonco Foundation (GD6362) provldod
0 financial oupport. .
Roforoncos
1. R. V. RISEBROUCH, D. B. PEAKALL, S. C. KERMAN, H. N. KIRVQi
and P. R1ECHE, Nature 220, 1098 (1966).
.
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2. R. V. RISEBROUCH, Chemical Fallout, Flrot Rochoator Conferonco
on Toxicity, In tho prooa.
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3. R. V. RICEERCUGH, D. B. MENZEL, D. J. MARTIN and K. S. OLCOIT, in preparation.
'0
U. Tho Pootlcldo Review, Unltod Statoe Dopartmont of Agrloultur*
.... '.;<w66).
. . ..
$. E. S. NIELSON, Ann. Rev. Plant Phyalol. 11, 3Ul (I960).
, 6. F. AGREE, Jr., M. BEROZA, and H. C. BOWMAN, Agrlc. Food Chen.
11, 270 (1963).
" . .....................
/
7. G. WIDXARK, J. Aas. Off. Anal. Chon. 50, 1069 (1967).
,
0. D. C. H0LE2S, J. H. SIttONS, and J.O'G. TATTON, Naturo 216,
227(1967).
" .
9. A. V. HOLDEN and.K, MARSDEN, Nature 216, 127U (1967).
1
(
. 10. T. E. ARCHER and D. 0. CROSBY, Pull. Environ. Cont. Toxicol.
1 * 1, 70 (1966).
.
:
11. F. ERRO, A. BEVDRJK, and H. BECKMAN, Bull. Environ. Cont.
Toxicol. 2, 372 (1967).
.
.
.
12. M. S. SCHECTER and K. L? HALLER, J. Am. Chen. Soo. 66, 2129 .. (19UL).
13. R. E. RISEBROUCH. D. B. MENZEL, D. J. MARTIN, and H. S. (JLCOTT, Nature 216, 509 (1567).
Hi. R. M. WELCH, W. LEVIN, and A. H. CONNEY, Toxicol.' Appl. . Pharmacol., In tho preeo.
15- J. BITMU. H. C. CECIL, S. J. HARRIS, and C. F. FRIES, Science
162, 371 (I960).
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16. D. A. RATCLIFFE, Nature 215, 200 (1967).
17. J. J. HICKEY and D. V. ANDERSON, Science 162, 271 (I960).
18. F. A. GUNTHER and J. H. BARKLEY) Bull. Environ. Cont. TCxlool.
1, 39 (1966).
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Expai for Ori
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In 1 utility o compounds to signal, for facil. fractions 0.17 yg. visually < used at t; | minimum d< j
Ballrlin a( Eatim Vat. 4. Ha. 4. 19W.'
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