Document RJ3YwnM51brzE4zMKJjLdzjpE

%rutila, Belgium February 22, 1967 m Dr. R. Vxme.t Kelly 8t. Louie J.K. Buchanan, Ut teuii ^ s_ . J.K. Filer, Brussels ^';''5CCp7^XP,V, Berdy, MCI. ' Kugene Wilde, tt Louie Dear Dr. Kelly, 1 should like to refer to your letter dated the lOth of February and our telephone discussion of yesterday's date. 1 enclose a copy of SOren Jensen's original paper which in fact was presented in English - Dr, Jensen apologises for the typing etc. but this copy is taken frost his original paper which has not yet been re-typed and tidied up. 1 asked our agent in Sweden to dig out the various pieces of information that you required and I attach a copy of his reply dated February 17th which i think fully covers the point raised in your letter. If there axeany snail points outstanding, please let me know and X will try to get you further information. X as. sanding the only copy of Mr. Jensen's paper to you and therefore if any of the other recipients of this letter needs to have access, they can take a copy from yourself. 1 would however emphasise the post script on page 1 of Ola Palm's latter in which he states Mr. Jensen has consented to send us the full copy of his paper on the understanding that this is used only internally within the Monsanto organisation. TOWOLDMON0003200 WATER_PCB-00000222 RISING & STRAND 0P/30 aktiebolao TELErOM; 3401 8S TflEBHAM; HENHIS TELEX; H24 roiTHmg smj eankoird- 73.0441 SSVtEoIcVAkChCoHlm47v February 17, 19o7 Mr. David Wood Monsanto Europe BRUSSELS J Belgium Dear David, re: AROCLORS Reference is made to our telephone conversation ar.ci I have tried to dig out the information you asked for. Before going into any detail, however, I would like to refer to your letter to Mr. Soren Jensen , dated February 8. Mr. Jensen would be interested in samples of as many isomers as you can put at his disposal. The quantities of course are very small indeed and he would need only about 10 mg of each. In theory you could put the lot behind a stamp and mail it to Mr. Sdren Jensen but in actual practice it is of course not as easy as that. Anyway, Mr. Jensen would be interested in both the lowchlorinated and the high-chlorinated types since this would help him to get a better picture of the metabolism. Now to the various points you raised. Enclosed you will find the original articles as published in "Dagens Nyheter" and "Svenska Dagbladet" on November 29, 1960. The people who organised the meeting wnere Mr. Jensen's paper was presented was a Committee - 190A Ars Naturv&rdskcmmitt& - operating under Statens Naturveti-hoKapI.. Forskningsr&d (The Swedish Natural Science Research Council). The Committee (for Conservation of Natural Resources of 1964) is werking under the chairmanship of a Dr. B. Lundholm. .. The meeting was organised under the auspices of Naturv&rdskommittn and was called apparently only in order to give Mr. Sbren Jensen an opportunity to publish his findings. Tha timing was made so as to coincide with a visit of Dr. A. Holden from Scotland. TOWOLDMON0003201 WATER_PCB-00000223 RISING 8. STRAND Mr. D. Wood - MCL Dr. Alan Holden - address Fresh Water Fisheries Laborai. eiy, Pitlochry Pearcher, Scotland - is the coordinator l'or the twelve OECD countries, as far as the influence of biocides on Nature goes. Dr. Holden apparently is a specialist on such questions and particularly on fish. There is no intention that Dr. Holden, or the group of twelve OECD countries as such, should carry on any further investigations on the particular problem unearthed by Mr. Jensen. Any continuation will be carried on in Stockholm by Mr. Jensen or cooperating Swedish labora tories. The meeting of November 2J offered only one paper, i.e. that of Mr. Jensen. No decisions were made on any further action. The work clone by Mr. Jensen was paid for by the abovementioned Committee for Conservation of Natural Resources. The further studies planned for on toxicology have not been started on yet since there is no money available. It is possible, however, that the same Committee of 1964 will be asked to subsidise even that work. If toxicological studies are to be made, these probably will be carried out at Karolinska Institutet, Division of Toxicology under Professor Bo Holmstedt. The full address of this institute is Kgl. Karolinska Institutet Avd. Toxikologi Solnavagen 1 Stockholm 60. Since no decision has been made on toxicological invest igations, obviously no information can be given as to the scope of the planned investigations. The LKB press release of January 10, 19uY mentionsc tne research of twelve OECD countries. As was mentioned above, there is no question of any central investigations to be made under the auspices of the twelve countries. There is only the question of coordinated efforts by way of interchange of information. Enclosed you will find photo copies of the original paper of Mr, Sdren Jensen. We have taken this copy here and Mr. Jensen apologizes for the state of the paper which is his own typing. He has not yet had an oppor tunity to have it properly retyped for publication. I nope the above answers your questions. Yours oii'.C'ireiy , h .-.A' MONS 090517 TOWOLDMON0003202 WATER_PCB-00000224 rising . strand Mr. D. Wood - MCL P.S. David For the sake of good order, I would like to mention that Mr. Sdren Jensen has sent us these papers with the understanding that they are only for internal use within the Monsanto organisation. Ola. MONS 090518 TOWOLDMON0003203 WATER_PCB-00000225 > Mr Chairman. ladles and gen*tolo,Cmon, In honor to our Brittlah goet I will try to hold this lecture in Bnglioh. As tho tltlo or this loeture states, I tun today going to tell about tbo dloeovory of some hithorto unoboervod chlorinated hydrocarbons having up to eight chlorine in tho nolooule and round'in reoiduo cnalyuio. Tha chonioftl none of polychlorinotod birenyle ( Xn tho following oollod FOB). To get Xcnillar with PCS I will start with tho ohemietry and tozioologi. i Choraletry Vit The nain-oharaoterietio of FCB le 1. Their very high stability* As an example they eon be boiled with nitrin add without being destroyed. 2. They are t)0dly metabolised in living organism* 3* if more than 4 chlorine areipret^eent they are non lnflomable. It is door that these thxeo characteristics doss it easy to undorst&nd that when they havo entorod the jllvlng organism the will have a low persistence . But it is difficult "to explain how they find their way into the living organism* Ono thing seems to be dear, they don't ooaa from agriculture! use, but from a technical one and most probable it Comes to the nature via wastes that are tried to be burnt up, because then wo havo them, at.onco in the air, beoauee of their non inflomability. . - . .Toxlcologi . MUN 049048 i Pho PCB woro introduced in 1929 and as early as 1936 Jones and Alien reported that 23 out of 24 non employed In manufacturing of PCB suffered from an acno fora eruption of the akin. Acne did not appear until 6 to 8 months after the material was first usod. In 1937 Drinker reported that rats exposed to chlorinated biphenyls In concentration of appkoxlmately 1 ug/s? for 16 hours a day for 6 weeks chovrdd damage of tho livor. After that time tho allonod concentration of PCB in air is 0,5 ng/a5. (Por DDt the some value Is 0*5 * 1 mg/a^). The soao authors finished tholr sxperiaents in 1938, and related that those compounds have an lnjouriouo effect, manifested solely in the liver. Chlorinated bipohnylo appeared to.be the moot lnjouriouo chlorinated compounds of all tested. !' l TOWOLDMON0003204 WATER_PCB-00000226 Gireenburg, Mayer and Smith 1939 reported that PCB and polychlorinated naphtalcaee are blamed for the death of three young workers. and that5* pregnant women and pereone who have at any time hud any livor diseases aro particularly euspeetible, t Wedol, Hnllor and Benton gave 1942 animals PCB including administration by inhalation, ingootion and akin abeorbtion. histological examination of tho visoera showed important toxic effect only in the skin and liyor, end tho dogonoratlon effects in the liver are essentially the sane whnt- ovor was'tho asthod for the administration. Paribok (l955/ found as on occupational poison in the eleotrioal Industry, mixed totra and po&tu chlorobiphenyl cauoos folliculitis, comedo, pyodermia and other okin affections, and that its principal toxic effect is fatty degeneration of the liver. ' Miller (1944) injected 69 mg PCB (4 and 5 ohlorine) subcontnneouoly in 32 guinoa pigs, Bight to ten days after injection, fat dropleta wero noted in the liver cells, and after 16 days they were present in modcrato or very large numbers. Babbits and rats were also tested in this lnvootl- gatlon, as well ae the PCB wae admlnietated both continously, cubcuntia- ously or ingested In the food. In the feeding experiment Q guinea pigs received 2 doses Of 69 mg of the chlorinated biphenyl 1 week apart. Death occurred in 11 to 29 days. Finally Me laughlin 1964 reported a method to test the chemical toxicity and teratogenic effect by injection Into the yolk Bac of fertile eggs prior to inoubation, PCB was found between the eight compounds among 100 tested having the highest order of toxicity. Mo hatch wae found at a level of ?3 mg pc egg. At a level of 10 mg per egg, one chick hatched out of 20 injected eggs, but died 2 days later. Some embryos which were examined after they died, showed weak deformities (often a short upper brek) and growth retardation. Lead acetate resulted as an example in no hatch at a level of 1 mg per egg. Autopsy of the dead eobyos have showed exten slve brain damage. Mercurio chloride showed no hatch even at a level of , 0,5 mg per egg. MGNS 049049 . As the analytical chemistry is a pronounced service science I have been in contact with many scientists from other fields during the work with residue analysis, and I have always found this contact very stimulating for my own work. This co-operation often demands that we are talking the same scientific language. Because of this need 1 will today try to give a lecture in low level analytical chemistry for biologists, illustrated by the residue analysis of polychlorinated biphenyls. The lecture will be divided in the following throe sub-divisions: . TOWOLDMON0003205 WATER_PCB-00000227 1. Chemistry of ?CB and their tocioology. 2. Analytical mathodo for rooidue analysis and proof of structures, 5. -Behaviour of PCB In nature, dlfferenoiee In metabolising xato of the PCB components, potenaation in an ecological oerie, con centration levels and examples of samples which have boon provod to con tain PCB* A rooidue analyste con be divided int ' 1, Extraction of tbs pesticides from the biological material, * followed by a careful eleaning-up to take away interferring substancee, moat often fate* a 2* Identification analysis by mean of gas chromatography. Thin- ^ layer ohromAtography and maae spectrometry. 3, Quantitative analysis. . At an ecological laboratory in Biksaueiet in Stockholm 1-2 g of a samplo e is cut out of tha biological material and transferred into a weighed and oarefully cleaned test tubs, and stored at -20 until analysis. Smaller yw samples have been used, min. 5 mg of body fat, and with dry materials such as hair, feathers, pins needles 100 mg are sufficient to reach the deolred tO ng/g level In residue analysis. In cases of water proofs 1 1. is used for reaching the 10 pg/g. level* B*l(homog) In order to facilitate complete extraction of the fatty materials from the biological sample, the doulbe amount of finely powdered anhydrous ' magnesium sulphate la added to the sampling tube, and the whole ie'homogenleed with an intertable homogonizer, The resulting powder is transferred into a special Soxhlet extraotor. After 4 hours of extraction the solvent is evaporated, leaving the fat In a email weighed teat tube at the bottom 'flox.-tube) of the extractor* This fat la dissolved in methylene chloride In such a way that 100 til (o,1 ml) contain 20 mg of fat. The 100 ul solution is now transferred to a little object glaae, 3 x 7 cm, covered with a sillcagel layer 1 mm thick, in order to fora a line 0,7 cm from one end of the slide* Inserting this thin-layer plate into a vessel the bottom of which is covered by a few mm of methylene chloride, the rv solvent will be sucked up In the dry layer of sillcagel, and at least reach the upper end of the plate. The fact io that the fat has a greater affinity to_ the powder on the plate than the chlorinated hydrocarbon have. -- and we get a separation. The fat being more polar than the chlorinated hydrocarbons will never go longer than 2 cm before the MONS 049050 TOWOLDMON0003206 WATER_PCB-00000228 solvent roacbos the upper part of the Blase. Tho front of the fat eppoaro qulto visible against a lamp, and with tho aid of a razor binda tho zone abovo tho fat is transferred to tho olutlon tubo and the chlorinated biooidoo absorbed on the powder can now be oluted by one ml of ether. The concentration is sufficient for detection of tho chlorinated hydrocarbons down to tho 10-12 g levol. Tho noxt stop In tho analytical procedure concerns the separation of tho difforont chlorinated hydrooarbone that the sample may contain. Ad matter of fact, this is a broublosome tank. It is easy to estimate what is not prosont, but more difficult to say exactly one la present. Vfe ouffor from tho negative demonstration, as will be shorn later. . At flrot a fow words about the separation of the components preoont in the sample and their visualization. The separation is aooompliehod by mean of a gas chromatograph fitted to a dotoctor that transfers its impulse to a recorder. The system is shortly .described: A spirally formed glass tube with an inner diameter of 2 mm end about 2 m in length is filled up by a support, covered with an thin layer of an oil. The tube is hoated in the chromatograph to about 200. through the tubo a stream of nitrogen continously follows. When about 10 ul (1/100 of 1 ml) of the purified sample la injected Into the tube, the components of the sample will be evaporised and go forward through the column with the gas stream. As the constituents have different affinity to the column filling they will pass ths column with different spood and it will taka different time for then to roach tho dotoctor at the other end of tho glass tubs. It the temperature and the nitrogen flow are held constant this time, tho retention time, has a spoclflevalue for a certain compound. This is true, but unfortunately it is also a fact that two components oaa have tho-same retention time. This is one of the bigger problems in gaa chromatographic analysis of unknown samples, as will soon be obvious. To make it possible to estimato the retention time it la necessary to visualise the chlorinated hydrocarbons, ?or that purpose more or loss specific detoators are used. The detector most often used in postlcldo analysis is the so called sloctron capture detector, which can detect down to one pieogram ( 10 g of lindan). Unfortunately this detector is not specific for chlorine, bu gives answer also for oxygencontaining compounds. The response hers is mucj: lower but can be counterbalanced if the concen tration of the oxygon containing is much higher, M0N$ 049051 Tho principle for tho electron capture detector is shortly: At the end of the gas chromatographic tube is placed a little tube con taining a foil mado of tltnniinn tritido. Thie 1r an-rodlant. The o- TOWOLDMON0003207 WATER_PCB-00000229 TJ OO particles ors reacting with the nitrogen molecules coining from the column. Then we get + N2 -- e" + Mg. Over the detector we have a tcnoion of SO volt and by mean of the electrons wc trill get a constant electrical curront over the detector. This standing current is transferred to a one-mV rocordor as a constant baseline. When now a chlorinated hydrocarbon lonvoo tho column thlo compound has a high affinity to tfeo eleotrono and this moans thnt tho amount of elootrons will diminish, and they will diminish proportionally to the amount of chlorine. The electrical curront will also diminish and this ie noted as a peak on the recorder. The area of tho peak will be proportional to the amount of substance in the sample. By moan of a standard injection it is now possible to compare the reten tion tlmo and the aroa of an unknown component with the retention time and area of the known standard. As said before thiB detector is not speci fic for chlrlne but anyhow very useful, because of its high sensitivity. The ayetom described has, ae we have seen, two disadvantages: 1. Two difforont compounds can have the name retention time and be detected ae ono peak. 2. A regietrated peak does not need to be chlorinated , because the detector is not specific. If the sample is injected in two different columns with different chemical properties we have inoreaeed' the chance for a good eeparatlon. If two compounds have the same retention time on one column they may not have it on another. When a result seems doubtful, - if the compound being rssponsible for a certain peak contains chlorine or not - it 1b possible to concentrate the sample and analyse,it on a less aenoitlve detector suoh as the mlcrocloumetrlc one, which le specific for chlorine. The oompound is burned in a furnace and teh generated chlorine titrated directly. MONS 049052 As ie aeen from the two last mentioned possibilities it is anyhow possible to get a rather high degree of certainty in residue analysis, but it is _ a rather tiae-sonsumeing work.When using this method just described, we very ofton found that many chromatograms from residue analysis of most carefully purified samples still contain a large number of peaks. Uapy of these have retention times that do not agree with any known chlorinated pesticides, or their metabolites. This chromatogram con serve as an example. It was obtained by residue analysis of a sea-eagle found deed in the archipelago of Stockholm. In the range of the known peaks, there are no many unidentified that there also muct be an obvious risk of the . known penko to bo covered by unknown ones. If thl R Fftmark In fonnrt +T1P. fhii tvbotO + o n*f wnnw TOWOLDMON0003208 WATER_PCB-00000230 titobivo anal,, A e oust be brought into quebe\\ 'ii\nn.. In the present investi gation it is shown that moot of the unknown peak of chromatograms at rooiduo analysis of chlorinated pesticides are due to polychlorinated biphenyls. I will show a chromatogram of human fat analysed on a ao ca lled S? 96 column, the most often used type in pesticide analyses* Early retention tinea were in agreemont with DEB, EDIop and DDTpp, Next slide shows the same sample analysed on a QB-1 column. Now the former 2 EEC peake have divided into 4 peaks,and two of them are still in agreement with DD2pp and op., the two new wore unknown. ' Logically, those unknown components were at first thought to be metabolites of the insecticides. Against that spoke that neigther treatment nor conoontratod sulfuric acid in other. This treatment made it rather euse that the compounds did not contain oxygen. In Swodon residues of organic mer oury have boen investigated rather Intensively in the Swedish fauna. Ae those compoundo give very high responses to the electron capture doteo- tor it was also investigated if the unknown peaks could have a mercuric origin. It was found that the water-ecological aeries had high residues of both mercury (Weotermaxk,Johnels) and the unknown ones, when ths same indivi duals were analysed. Anyhow, the pheasant suffering most from mercury poisoning only contained low levels of electron capturing compounds and those belonged ' to the normal insectisldes. Therefore the unknown could hardly be mercu rials or metabolites of them. HONS 049053 As the eagle sample giving the chromatogram shown in fig. to, could be estimated to contain SDT and LEE up to 13 g/kg in extractable fat, the amount of unknown compounds also were suggested to be in tho same range, and tehn sufficiently high to do a run on the combined gas chromatograph - mass spectrometer. If this could be done successfully it would be possible' to get very important informations about the chemical nature of the unknown, for ex. the mclekular weight numbers of chlorine etc. This method is up to now the method giving the highect degree of certainty in the low level analytical chemistry, amounts of 100 ng substance being enough. As this method for identification of totally unknown reeidues surely will be very Important in the future (when f.ex. a biologist has found that fieheo in a river die) it may^loasible by mean of this method to find out exactly what compoundo are responsible for the death. For this reason, I will go into come details with this method. In the actual ease we took the extract from 20 mg eagle and concentrated TOWOLDMON0003209 WATER_PCB-00000231 ' ' (Y O it as much as possible and made an injection on the gaa chromatograph o combined with the mass apectrometer. The result was the chroamtogram shown on the next slido. Every time the recorder showed that u compound is leaving the column, the effluent ie led to the naas epeetromotor. Now juet a few words about the maae epee, 1* The moleculee leering the eolumn are bonded with electrons at K. Vo havo how got the molocule positive charged, but with the same maoo no before. This )i* is accelerated in a vacuum and will then get a kinetic energi. whero is the speed. Next comes the magnetic field that . trios to bend the direction of the molecule. This will be big for & small molecule and less for o If we have a sieve in the other end we can dlreotly read the molecular weight. Added to this parent molecule M+ we will also gat addition in formations, because of the fact that H+ may not bo sthblo, a port of them will be broken down before they reach the aleve in the othor end. ?.ex. M DD5? U ,, cci? Maes spectrograms from the different unknown peaks in the eaglo sample ae shown, Tho mass numbers equal to the molecular weights of the unknowns could be read to 426,392, 358, 324. Astonishingly, the molecular diffe rences were constantly 34 mass units. This difference shows a familarlty in origin of the unknown. Sow the fact is that chlorine ezsioto ae a mixture of two isotopes with atom weights 35 and 37 in proportion 75s25. If the molecule has one chlorine, this will give two molecule peuke, one for Cl,. and one for Cl--. If there are two chlorine wo have the pooolbility 35 37 of one with only Cl^, one with both and 37 and one with 2 C1^ and therofore MONS 049054 TOWOLDMON0003210 WATER_PCB-00000232 She relation of the peaks found on the different mass epee woret Molecular weight ; 324 358 392 426 Ohlorino content 56 78 An explanation 0f the familiarity of the compounds can be given if one eubetanoe is built from the former by substituting a hydrogen with chlorine m 01, SCI HC1 M++ 34 Thon it is possib) io calculate the molecular weight of the parent hydrocarbon VHC. PKC " K " s MC1 + x "h * wh#r* M ie the molecular weight of the component having x chlorine atoms. P.ex. for m - 426 and 8 Cl we will get . 426 - 280 + 9a 154 and equal with the other molekylo. Iho moot probable formula with carbon and hydrogen giving this moleoular weight 1b C^ 2 and 'this can only be satisfied when the parent-hydro carbon is biphenyl, and the unknown being polychlorinated biphenyls. This explanation was later fully verified by injection of a synthetic PDC on the mess spec. Furthermore extensive gaa chromatographic investigations proved that the FBC standard gave peaks with the same retention time as the unknown * peaks from the sea eagle. With the method just described 1 suppose that we have a new possibility to study the residues in the air because the pine needles can allways be . We have had great difficult; in Quantifying the ?CB, but when getting a little more time it will be possible. We have done a few calculations on a few species, and I suppose they are right within a factor 2. We have found the residue to be from It has been my statement here to-day to present thie method for studies of dofiling of the nature, and with thie method a new typo of dofilia* agents has been found to be present In nature, and a few experiment have shown where they may be found. How this method is going to be used in the first hadn to estimate how the situation is Ln nature *b a whole, and in the ether hand to find the leaks throug which they find its way to nature. Soem maybe are present here today to get news about the leaks, and to then 1 want to say cose back in a year. MUNS 049055 TOWOLDMON0003211 WATER_PCB-00000233 / 9 So much I think I can coy again that the PCB hardly con coma from agriculture. A a euppoft for this suggestion I can say that we have found PCB in eagle feathers from Rikamuseot from 1944, where hardly any chlorinated pesticides were used in agricultrue. One more thing that I find important to say is that in contrast to the mercury problem this does not scorn to bo a pure Swedish problem, 1 have just studied chromatograms takon from London air, and they doarly contain PCB, and dr. Holdon han told me that ho also find them in his fishsamples. But finally in waiting at more results Xnshould like to point)$e morething. It is proved that PCI eomos to naturo, wo dent know now where they are usod, but thoy are very persistant to chemicals and to fire* X think the poison jury should try to state that a oontsnt of PCB shall always be found in an open declara tion. HONS 049056 ! TOWOLDMON0003212 WATER_PCB-00000234 TOWOLDMON0003213 WATER_PCB-00000235 8506^0 SNOW O o cP x*l Si NQ, t o* 3 s. a mm, 5 53 9 .1 5 *) MV *S 55 5? 5 5: .0 lo SN V> N. N C\ TOWOLDMON0003214 WATER_PCB-00000236 TOWOLDMON0003215 WATER_PCB-00000237 ( 0906*0 SNOW o*>3 9. **) 9. 0. 9 H 5: Curve ( * **\ Curve 2 * 5! Cur vo 3 CUrvp 4 *S to * *> $ . s ts * b S s V * * \ o TOWOLDMON0003216 WATER_PCB-00000238 TOWOLDMON0003217 WATER_PCB-00000239  ok .Q v o N > ^s Q -C-uAt^ CL $ s <K*>. *SS. ~Grttm&? 5'Cvtr*e. o 6-utice <D o o. fc * *M *> k Ot \Q MONS 049062 TOWOLDMON0003218 WATER_PCB-00000240 TOWOLDMON0003219 WATER_PCB-00000241 TOWOLDMON0003220 WATER_PCB-00000242 TOWOLDMON0003221 WATER_PCB-00000243 TOWOLDMON0003222 WATER_PCB-00000244 TOWOLDMON0003223 WATER_PCB-00000245 8906*0 SNOW I C t ~r~ TOWOLDMON0003224 WATER PCB-00000246 6906*0 SNOW % V ssr c> TOWOLDMON0003225 WATER PCB-00000247 TOWOLDMON0003226 WATER_PCB-00000248 y o s TOWOLDMON0003227 WATER_PCB-00000249 ?06*0 SNOW TOWOLDMON0003228 WATER_PCB-00000250 /9 ifo TOWOLDMON0003229 WATER_PCB-00000251