Determination of Fluoride in Vegetation R. J . ROWLEY, J . G. GRIER, AND R. L. PARSONS Aluminum Research Laboratories, .4luminicm Co. of Imerira, Vew Kensington. P u . The methods generall? used for determining fluoride in vegetation gile l o w results with samples high in sirica. 4nalysis of synthetic mixtures showed that large quantities of silica inhibit recovery of fluoride. During ashing, there seems to be formed a refractory compound from which fluoride is not evolved by double distillation. * i n alkaline fusion completely liberates this combined fluoride. Collection of 500 ml. of distillate further increases the accuracy of the determination. 4lthough not all vegetation requires an alkaline fusion or collection of 500 ml. of distillate, the method has been retised to include both. Appreciable savings in time, as w e l l as increased accuracy, result from these modifications, which hale been tested on numerous samples and known mixtures.
T
HE generally accepted methods for determining fluoride vegetation are based on separation of fluoride using the ~~eIl-1~no~?-ri JVillard and Winter distillation method (41, following a preliminary ashing treatment with lime or other alkaline fixative :ddrd to prevent loss through volatilization. lleasurement of the fluoride isolated by distillation is usually based on the reaction between fluoride and thorium nitrate in the presence of sodium alizarinsulfonate indicator. For milligram amounts of fluorid?, it is possible to perform a tlirect titration. Colorimetric methods based on various fading re:ic.tions involving fluoride ion are also in use, particularly for smdl amounts of fluoride. li-here only a few micrograms are p r w r n t in the entire sample, a back-titration procedure is frequent 1y carried out in Sessler tubes. Preliminary ashing, followed by double distillation arid back-titration, has been adopted by t,he ilssociation of Official .4gricult,ural Chemists ( 1 ) . The purpose of the distillation step is to separate the fluoride froni constituents which either retain the fluoride in the form of iiisoluble compounds or interfere with the final measurement. There has been a general belief that fluoride can be satisfactorily recovered froni ashed vegetation by double distillation, first with sulfuric acid, and then with perchloric acid. Recent work i n this and other laboratories (3)has shown that ashing of vegetation rontaining appreciable amounts of silica may result in unrewtive combinations of fluoride, from which fluoride cannot be sep:iixted quantitatively under conditions of the Willard and l\.inter distillation. Tivo modifications for eliminating lo\?-recoveries result,ing from fixation associated wit,h the ashing treatment are: alkali fusion of the ash and collection of 500 ml. of distillate. A single distillation with perchloric arid has been shown to be adequate for isol:iting the fluoride in n form suitable for measurement. 4 p i)rer.iahle savings in time result from elimination of the double c1iqtill:ition and evaporation formerly believed necessary. This 15-ork supplements and confirms conclusions of Remmert and 1':irks (3).who first called attention to the low results obtained usiiig the procedure of the Association of Official Agricultural Chemists 011 certain types of vegetation high in silica. ('oiisiderable detail is included in the procedure given here. hecanuse of the diffic.ult>-experienced with previous methods for cleterinining fluorine. Estreme care must be exercised in order to avoid high blniiks and t o obtain significant results in t h s determiii:ition of mii.rogram quantities of fluorine. ill
being made alkaline, was concentrated by evaporation, transferred to the distillation flask, treated with perchloric acid, and distilled a t 135" C. Ordinarily, 250 ml. of distillate were collected. The other technique involved fusing 1 gram of the ash with sodium hydroxide, transferring the melt t o the distilling flask, adding perchloric acid, and distilling at a temperature of 135" C., collecting 500 ml. of distillate. The data in Table I indicate that higher results mav brobtained h~ the fusion method
Table J . Comparison of Double Distillation and Fusion-Single Distillation Methods Fluoride, P.P.LI. Fusion-sin& Douhle distillation diptillat ion
Sample
Grass hay Grass h a s fluoride
+ added
Red r l o r e r hay Red clover hav added fluoride
-
Strait
1.4 1.4 1 4 18 23 18 17 18
3.6 3 0
2.0 48 47 47 22 40 18
18
8: 83 81 3.4
Moat methods for determination of fluoride which involve distillation recommend collection of 250 ml. of distillate. This volume is suitable for samples of low silica content, but with highly siliceous mat,erials, the fluoride is not readily released. To prove this point, a number of samples were run hy thc fusionsingle distillation method, the distillate being collected in three portions of 230 ml. each. and the fluoride being determined ill each portion. Table I1 shows the effect of t,he volume distilled on the recovery of fluoride. These results indic.ate that for R mrietj. of vegetation samples it is neclessarj- to collect 500 ml.
Table 11.
Effect of Volume of Distillate on Recover:of Fluoride Flrioride in 2.50 MI.. P.P.31. 1st 2nd 3rd ~~~
40 41 36 37 3 3 34 39
EXPERIMENTAL WORK
One of the first experiments involved the determination of the fluoride content of several samples by the two different methods. In the double dist,illation method. a portion of the ash was distilled in the presence of sulfuric acid at a temperature of 163' C. :inti 400 ml. of distillate were Follec*te.tl. This solution. after,
h:ind R raniples are duplicates
1061
2 7 6 5 : 2 1
8
8 9
3 0 0 0 0 8 5
2 8 0 4 1 4 6
~,,~~~o;;f~~d Frar t ions Yq I
100 100 100 100 100 98 99
o
0
0
0
0 8 1
1062
ANALYTICAL CHEMISTRY
distillate. In no case examined would a large error be introduced if the t h k d 250-ml. portion were disregarded. In order to establish the depressing effect of silica on recovery of fluoride, the authors prepared a number of mixtures containing silica and measured amounts of fluoride. The specified amount of silica in the form of sodium silicate was added to the distilling Ha8k. An aqueous solution of sodium fluoride was then added directly to the flask in the desired amount. Perchloric acid (70 to 72%) was added and the distillation was made a t 135' C., the distillate being collected in three portions of 250 ml. each and andyaed separately. Table 111shows that when %largeamount of fluoride is present, the efficiency of recovery dmxasea with an increase in the amount of silica present. When the silioa content is high, recovery of fluoride may not be oomplete IT-ith callerition of as much as 750 ml. of distillate. The data listed here do not represeint a test of the pro:,osed .... J Aw ...LL.. fusion method, as the silica. and Humins wen: uuu/ /LUMU ~muer before distillation. The range of both silica and fluoride isgreater however, than is generally encountered in vegetation. In order to establish the accuracy of the proposed method, samples of known fluoride content u~ouldbe required. , As there were no vegetation samples of known composition available, the recovery of measured additions of Huoride was studied in detail. Weighed portions of vegetation, obtained and prepared according to the standard procedure, were treated with varying quantities of an aqueous solution of sodium fluoride. These synthetics were ashed, fused. and distilled, and the fluoride was determined by back-titration (Table IV).
These percentage recoveries are considered setisfactory for elements in this range of concentrations. Results are ordinarily rounded off to the nearest whole number. PROCEDURE
The following method for the determination of fluoride in vegetation embodying the proposed modifications is now employed by all laboratorics of the Aluminum Co. of America.
,
Apparatus. Wiley cutting mill. Porcelain casseroles, Coors No.6,1200-ml. capacity, for ashing. Inconel dishes (5.5 inches in diameter and 3 inches high) are also satisfactory (obtainable from Precision Metal Spinning Co.. 9825 Dixie Highway, Clarkston 2, Mich.) Sampling jam, Mason jars wit,h tight fitting covers and rings. Wide-mouthed Erlenmeyer Raskn with rubber stoppers. ~
-
Table 111. Effect of Silica on Remover? of Fluoride
_. ..
SiOn. ME.
.
rruarlne I" "-".I. ZDV 1 1 1 . . yo1st 2nd 3rd
Total F Recovered. 3'%
With 200 Miorogrsma of F and Varying Amounts o i Si02 00 100 0 0 100 50 93 3 0 101 ion 94 4 0 98 300 85 7 3 94 5nu 76 14 3 93
A.
Figure 1. Apparatus Used for Distillation of Fluoride 1. 9
3: 4.
Stearngenerator Distilling flask Liebig oondenser Steam releaae tube
6. 6. 7.
Clamp 0" inlet tube to 3. Outlet water maniiold 0 llllminllm distilling LR o~l l~ . . . ..l.l.ll.l"lota r_""" Thermometer 16: Fisher burnem Inlet water manifold 11. Fletcher burners 1R. Safety tube
12. ,?
16.
Wire triangle
dil..min..m . . . I ......I.I
port Receiver$
..,". ___." "_ L.nOJa
V O L U M E 25, NO. 1. J U L Y 1 9 5 3
1063
Recovery of Added Fluoride
T a b l e IV. No.
F Added, P.P.M.
1 2 3 4
0.0
F Found, P.PM.
F Reoovered, $b
Grass Hay 15.2 25.0 45.6 76.3
10.0 30.0 fin. 0
.., 99.2
100.9 101.5
No. 35 Gram Hay 1 2 3
10.0 30.0
4
60.0
1
0.0 10.0
4.1 13.4 31.8 59.2
0.0
...
95.0 91.4 92.4
straw 2 3 4
30.0 GO. 0
1
n. 0
4.9 14.3 32.6 59.2
...
96.0 93.4 91.2
No. 51 Red Clover Hay 12.1 22.3 39.5 69.4
10.0
2 3 4
30.0
60.0
1ba:g 93.8 96.3
Timothy 0.0 10.0 30.0 60.0
1
2 3 4
3.4 11.8 29.9
60.7
...
88.1 89.5 95.7
A ~ p l eLeaves 0.0
1 2
14.0
3
30.0
35.1.36.1 51.2:" " 64.5. 94.8.
... .-.
Nickel beakers, 250-ml. Multiple still (Figure 1). This is a variation of a model described by Churchill ( 8 ) . Steam generator, 2000-ml. Florence flask made of borosilicate glass. Each flask is fitted with a No. 8 stopper having five holes for inserting borosilicate glass tubing 6 mm. in outside diameter. Three of the glass tubes are bent a t right.augles for introducing steam to three distilling flasks, The fourth tube serves as an outlet for controlline. the steam D E S S U ~ ~ . The fifth is a safetv
i s ~ b e i downward t to attach to an upright condenser. The side tube is fitted with 8. No. 3 one-hole stopper, and the main neck is fitted with a No. 3 two-hole stopper through which pt~ssa therrnornet,er and a hornailieste elms inlet tube. 6 mm. in outside ~~~.~~~~~~~ diameter, for admitting the s7eam. s7eeam. Both the thermometer and inlet tube are immersed in the acid solution containing the sample. Borosilicate glass Liebig condenser, 300-mm. jacket. Steam release tube. This tube is clamped when the sample is heine distilled. CGmp on inlet tube to the distilling flask. Thermometer, range 100" to 200" C.,,of sufficient length for 130" C. mark to extend beyond etopper wlth bulb immersed. Inlet water manifold. Outlet water manifold. ~~~~
~.-.~..
~
~~
~
..
~~~
~
~~
C&talog No. 11-312), for use in distilling flask to prevent superheating and to SUDDIV .. . silica for formation of fluosilicic mid durine distillaion. Pumice, granular. Buret stand. Attach a double buret clamp to a rad that fastens a sheet of aluminum to the base. This sheet is bent a t right angles to give a background and is painted with ~ white hitr ensrnd i*oPPimlTP21 i .____.-___,_ r~*Pim,r~II Fluorescent lamp, to provide illumination for titrating. Microburets, having &ml. capacity, 0.01-ml. divisions, and reservoir holding about 50 ml. During titrations, read to 0.005 I
~
rnl ...
Nessler tubes, matched set of 50-ml. high-form tubes TT-ith shadowless bo**---YYYIIID. Nessler tub es, matched set of 100-ml. hish-fi - x m tubes a i t h shadowless b ottoms. Nessler tubt3 rackoreomparator. Reagents. Calcium oxide, speoially prepare:d, fluorinofree ( F i s h e r Scientific Co., N o . C-117). wide slurry, 15 g r a ~ r ~per a liter. S o d i u m rtliaarindfouate indicator, for direct titration, 0.400 gram per 500 ml. of water. S o d i u m alizsrinsulfonate indicator, for backtitration, 0.050 gram per 500 ml. of water. Sodium hydroxide solution, 2 grams per liter (about 0.05 N ) . Hydrochloric acid, 1 to 250 (about 0.05 N ) . Chloroseetate buffer solution. Dissolve 9.45 grams ofmonochloroscetie acid and 2.0 grams of sodium hydroxide in 100 ml. of water. Silver perchlorate solution, 50% (50 grame in 50 ml. of water). Standard thorium nitrate solution, 0.1 iv (equivalent to 1.9 mg. of fluorideperml.). Dissolve 13.8 grams of thorium nitrate tetrahydratein water and dilute to 1 liter. T o standardize, weigh 0.100 gram of 100% natural cryolite (selected, handnicked crvstalsi or of iOO% s o d i u m f l u o r i d e , tran'&er to a distilling Figure 2. Stand for Titrating flask, distillwithperehloric Fluorides acid, and collect 250 mi. of distillate. Titrate 50ml. aliquots (0.02 gram of cryolite or sodium fluoride) with 0.1 N thorium nitrate. Carry B blank through the mme procedure. Deduct t h r millifor the liters of thorium nitrate used for the blank from:hose standards to obtain the net milliliters of thorium nitrate.
~
in Figure 1. The 2-inch holes and the di&lling flasks m u i have a close fit, so that the flame heats only the portion of the flask conbaining the liquid. Excessive heating volatilizes the perchloric acid. Fisher burners, utility model, one for each steam generator. Fletcher burners, one for each distilling flask. The burners are taDDed from manifolds so that the barrels reach to within about 1.5hehes below theplate. Wire triangle, to allow the flame to heat the sides of the steam generator. Sunnart made of duminum angle. GiLmanifold. Receivers, 500-ml. volumetric flasks. Safety tube, one for each stemn generator. Borosilicate glass tubing. 2 feet lone and 6 mm. in outside diameter, extending to 1 mtural rubber for h s k connections. Soft-glass beads, 3 mm. in diameter (Fisher Scientific Co..
Mg. of F per ml. =
mg. af cryolite titrated >< 0.543 net ml. of thorium nitraE ~
OP
mg. of sodium fluoride titrated X 0.4524 net ml. of thorium nitrate Standard thorium nitrate mluwon, 0.04 N (equivalent t? 0.76 mg. of fluorine per ml.). Dilute 400 ml. of 0.1 N thorium rutrate to 1 liter and mix. Standardize a6 follows: Dilute 50 ml. of distillate obtained in the standardization of 0.1 N thorium nitrate to 500 ml. Titrate 50-, 25-, and 10-ml. diquats (containing 0.002,0.001, and 0.0004 gram of cryolite or sodium fluoride) 1%-ith 0.04 N thorium nitrate. Deduct a determined blank and make calculationsasfor0.1 N thoriumnitrate. Sodium fluoride stock solution, 1mg. of fluorine per ml. Dissolve 2,210 grams of 100% sodium fluoride in 1 liter of water. Pipet a 25-ml. aliquot into a 250-ml. Claisen flask, add 35 ml. of perchloric acid, steam-distill a t 135" C., and collect 250 ml. of
:. of F per ml.
=
.
1064
A N A L Y T I C A L CHEMISTRY
distillate. T o a 100-ml. aliquot of the distillate, add 1 ml. of sodium alizarineulfonate solution and neutralize using sodium hydroxide and hydrochloric acid solutions. Add 1 ml. of chloroacetate buffer and titrate with 0.1 S thorium nitrate. Deduct a determined blank.
a Fisher buiner. Allow to cool, wash doxn uith water, and heat to disintegrate the melt. Distillation. Fill a steam generatoi about t n o thirds full of water. .2dd a pellet of sodium hydroxide and a few drops of phenolphthalein to ensure that the n ater remains alkaline at all tinier .idd a gianule of pumice and heat the Aater to boiling. CILCULATIOS (FOR STOCK SOLUTIOX). Place a 500-ml volumetric flask under the condenser to receive distillate. MI. of Th(liO,), X mg. of F per ml. Th(SO1)l theTranqfer the contents of the nickel beaker to a distilling flask l r g . of F per ml. = ml. of stock solution (cleaned with brush and distilled water) containing five t o six glass beads. Insert in the main neck of the flask a rubber stopper Standard sodium fluoride, equivalent to 0.01 ing. of fluorine through which pass a thermometer and a glass inlet tube. Set per nil. Dilute 10 ml. of stock solution to I liter. the flask in the 2-inch-diameter hole in the metal plate and connect t o a condenser. Rinse the sides of the beaker with 50 ml. Sampling. Representative samples should be selected, taking of perchloric acid (70 to 72%) and add 1 ml. of silver perchlorate cognizance that contamination may arise fiom a variety of solution. Transfer to the distilling flask by means of a small sourws such as fertilizers, road dust, etc. Evidence of such funnel attached to the glass inlet tube. Rinse the beaker and contamination should be noted, as uell as other pertinent inadd the rinsings to the flask. Mix the contentcl of the flask and attach to the steam generator. formation, including a complete description of the sample, locaHeat the distilling flask to 135" C. Open clamp 5 on the steam tion of sampling, and weather conditions prior to sampling. inlet tube and close steam release tube 4 uith a clamp. Steam This last ie of importance chiefly for survey norA. Roots and distill a t 135" C. Swirl the contents of the distilling flask fresoil should be avoided by using hand shears in sampling green quently to minimize deposition of any siliceous residues which might retain fluoride. After collecting 500 ml. of distillate, disforage. Tn-igs are ordinarily excluded in the sampling of leaves. connect the rubber tubing from the steam inlet tube, open the 1.iine is added to green vegetation for maintaining sufficient steam clamps, and turn off the burners. alkalinity to fix the fluoride during storage, although this treatFollowing distillation, the determination mal be complete1 ment is not necessary with dried samples. Other minor differences in the preliminary treatment of green and dried vegetation by titration with 0.04 S thorium nitrate. by back-titration. ale specified below. or by titration uith a photometric titrator (to be descrihed in a subsequent paper). A. GREENVEGET.4TIOX-GRASSj LEAVES,ETC. Place alterTitration with 0.04 N Thorium Nitrate. The 0.04 S titration nate cuttings of the vegetation in sampling jars, for the determethod is normally used for determining 0.02 to 1 mg. of fluoride mination of fluoride, and in weighed 250-ml. Erlenmeyer flasks for determination of moisture. Stopper the flasks tightly. Two per 100-ml. volume titrated. An aliquot of the distillate is ways of adding the lime and determining the sample weight are: buffered to pH 2.9 to 3.1 and titrated with standard thorium 1. Weigh jar in laboratory. Collect 100 to 200 grams of vegenitrate to the first definite color change produced by lake fortation and add 3 grams of previously weighed calcium oxide in mation of excess titrant with sodium alizarinsulfonate indicator. the field. Weigh jar and contents in laboratory. 2. Weigh jar and calcium oxide slurry (200 ml.) in laboratory. During titiation, thorium fluoride forms. A blank made bv Collc~tvegetation in field. Weigh jar and contents in labousing the same quantities of reagents and the same technique ratory. as for the sample is carried through the preliminary treatment. If the samples are to be analyzed nithin a few hours after distillation, and titration. collection, the calcium oxide may be added immediately on rePipet an aliquot of distillate into a 400-ml. beaker and dilute turn to the laboratory. I n any case) storage of green vegetation to 100 mi. Add 1 ml. of sodium alizarinsulfonate indicator and a t rooni temperature should be kept a t an absolute minimum, sodium hydroxide solution (10 grams per liter) dropwise until a pink color is obtained. Discharge the pink color by adding dropbecause fermentation occurs with subsequent generation of preswise 0.05 S hydrochloric acid. Add 1 ml. of chloroacetate buffer w r i~ n the containers and possible loss of fluoride. solution. Place the beaker on the titrating stand. Titrate with 0.04 After calculating the ueight of the sample, transfer the conthorium nitrate while stirring with a glass rod until a faint permatents of the sampling jar into a porcelain casserole and rinse nent pink color is observed. the jar with water, adding the rinsings to the sample. Cover the Cartv a blank through the same piocedure and deduct the ve,getntion with water, add a few drops of phenolphthalein, and blank titration from the sample titration for the net milliliters of nil\; R. DRIED VEGETATIOK-CURED H 4y, DRIED LEAYES. thorium nitrate C~LCULXTIO\ STRIVS,ETC. Diqintegrate the sample in a Wiley cutting mill and mix. Place a 10- t o 25-gram sample in a weighed 250-ml. P.p in. of F (dry basis) = net ml. of std. Th(SO9)r X mg. of Erlenmeyer flask, stopper tightly, and reserve for determination of moisture. g. of total ash ml. of distillate Weigh a 50-gram sample into a porcelain casserole. Add 3 F'nil' nil. of aliquot g. of ash fused X grams of calcium oxide, cover with water. add a few dram of ~ _1000 _ phenolphthalein, and mix. Moisture Determination. Weigh the 250-nil. Erlenmeyet, g. of sample X 100 flash containing 10 to 25 grams of unlimed sample, dt,y 21 hour$ a t 105' C., cool in a desiccator, and weigh. C.4LCUL.tTIOS. Back-Titration, The back-titration method is normally used for determining less than 0.02 mg of fluoride per 50-ml. volume g. of loss x 100 % moisture = - g. of sample titrated. An aliquot of distillate is added to a Kessler tube. adjusted to definite acidity with hydrochloric acid, and titrated Ashing and Fusion. Digest the prepared sample in the caswith standard thorium nitrate to the first definite pink color. serole or Inconel dish on a hot plate and add calcium oxide as reh blank aliquot of water is tieated with the same volume of quired to keep the water alkaline. Stir occasionally and evaporate thorium nitrate and titrated with standard sodium fluoride until to dryness. .leh the sample on the hot plate as completely as possible in order t.o prevent the dry material from hursting into the lahe color matches that of the sample. flanie when it is placed in the muffle. Transfer to a muffle fur1. DETER\IIUATIOS O F ACIDITY I N SXMPLE DISTILL~TE. nace a t 600" C. and ignite until ashing is complete, as indicated by a white or light gray ash. Transfer 50 ml. of distillate into a 50-ml. Kessler tube and add After cooling, partially pulverize the ash with a pestle and mix 1 ml. of sodium alizarinsulfonate and 0.05 S sodium hydrovide to a pink color. Kote the milliliters required for neutralization thoroughly. Scrape to remove any adhering material and weigh and discard the clolution. If more than 5 ml. of sodium hydroxide the entire c0ntent.s of the casserole. Transfer to a stoppered bottle. is required repeat the distillation to ohtain a sample distillate Weigh 1 gram of ash and transfer to a nickcl beaker. Add 5 that k more nearly neutral grams of sodium hydroxide pellets and fuse for a few minutes over 2. TITRITIO\O F St\IPLE D I S T I L L ~TII E ITH T H O R I L r \ I S I A\-
V O L U M E 25, NO. 7, J U L Y 1 9 5 3 Transfer jo ml. of distillate into a 5o-nil. xessler tube (sample tube) and add 1 nil. of sodium alizarinsulfonat,e. Adjust acidity with 0.05 S hydrochloric acid until the equivalent of 2 ml. is present-that is, 2 ml. minus milliliters of sodium hydroxide required under Section 1. Add thorium nitrate solution (0.26 gram of tetrahydrate per liter) from a microburet until a faint pink color appears. When 50 ml. of distillate require from 2 t o 5 nil. of sodium hydroxide for neutralization, omit the hydrochloric acid addition to samplc distillate, hut add t o the standard tube the same quantity of acid as was found in the sample. 3. TITRATIOS OF FLUORIDE STANDARD.Pour 50 nil. of water into a 60-ml. Sessler tube (st,andard tube), add 1 ml. of sodium itlizarineulfonate, add 2 ml. of 0.05 .V hydrochloric acid to match the sample (see Section 2 ) , and add standard sodium fluoride solution equivalent t o about 80% of the fluoride present in the sample aliquot as indicated by the thorium nitrate required under Section 2. Mix thoroughly, add the same volume of thorium nitrate required for titration of the sample under Section 2, and again mix thoroughly. The standard t,ube will be more highly colored than the sample tube. From a microburet add sodium fluoride to the standard until the color matches that of the sample. Equalize the volumes in the two tubes and mix each solution thoroughly by inverting, allowing all air bubbles to escape before making color comparisons. Check thcx end point by adding 1 or 2 drops of standard sodium fluoride t o the standard tube. If the colors were matched, a d i d n c t failing of the color in the standard tube should occur. TR.iTE.
1065
4. B L ~ \ E D I E T E R XITIOX. II~ Deduct :I blank obtained b v carrying the qame amounts of all reagents used through the entire process of afihing, fusion, distillation, and titration. With proper attention t o detail., total blanks of the order of 5 mic~ograms are obtainable. CALCULiTION. P.P.M. of F (dry baiis) = ml. of didillate net ml. of std. S a F X mp. of F / m I X ml,of aliouot X 1000 g. of total ash - % moisture) g. of ash fused g. of m n p l e X 100
(
LITERATURE CITED
of Official Agricultural Chemists, “hIethods of .Inalysis,” p. 389-96. 1950. 12) Churchill. H. V..ISD.ENG.CHEJI...INAL. ED..17. 720 (1945). ( 3 ) Remmert, L. F.,’P;t~ks. T. D., Lawrence. 1.LI., and ;\lcBurney, E. H.. . ~ N . L I . . ( ’ H E M , . 25, 450 11953). (4) Willard, H. H., and \T-inter, 0. B., ISD. ESG. C‘HEX., SAL. En., 5, 7 (1933). (1) Association
RECEIVEDfor reviex Janiiary 1 7 . 1953. Accepted A p r i l 11, 1953. P r c sented before t h e Pittsburgh r o n f r r e n c e on Analytical C‘heiiiiatry and A P plied Spwtro.scopy. Pitt?hiirgli, Pa., March 2 t o 6, 1933.
Determination of DDT and Related Substances in Human Fat 4RNOLD 31. MATTSON, J4SET T. SPILLiNE, CURTIS B 4 K E K , A Y D GEOKGE W. PEARCE Coiiiniccnicable Diseuse Center, Public Health Serrire, K‘.S. Department of Heulth, Education, and F’elfare, Box 769, Savannah, Ga. Preliminary e\idence of the occurrence of DDE as well as DOT i n human fat made it necessary to stud) the Schechter-Haller method critically as applied to fat containing DDT and the degradation product, DDE. Through empirical standardization it was possible to make differential determinations of DDT and DDE totaling 5 micrograms in unknown fat samples, with rough estimations down to 2 micrograms. 4 modified Davidow column was used in a rapid method €or isolating DDT and DDE. Among 50 samples all but 2 contained substantial qiiantities of DDE. The total DDT plus DDE ranged from 0 to 80 p.p.m., with DDE representing from 39 to 86% of the total. Chromatographic and spectrophotometric data provided e\ idence that the degradation prodnct i s TIDE.
I
S THIC c o u ~ s cof u study on the occurrence of DDT in human fat it became clear that a detailed study of both the method of isolating DDT from fat and the final estimation by t,he Schechter-Haller procedure was necessary. This became more essential \\.hen evidence was obtained that a substantial quantity of DDK [ l.l-dichloro-2,2-bis(p-chlorophen~-l)ethylene] as well as D D T \\.as present in the samples. .4 preliminary report on this observation has been given hy Pearcr et a!.(8). The present paper reports the results of the study on the methods eniployed and .solile additional evidence as to the presence of DE)],; in human fat. SEP4RATION O F FAT
The isolation of DDT and related compounds from biological materials invariably has presented difficulties, espec’ially in dealing with fats or oils because of the pronounced solubility of chlorinated hydrocarbons in fat, solvents. The mo.st conimonly used methods for separating DDT from fat are bayed on a sulfuric acid treatment which partially sulfonates the fat and greatly reduces its affinity for DDT and thus permits rxtraction of the D D T in an inert solvent such as chloroform or c a h o n tetra-
chloride. I n the method ot &.herhter et ai. (f01.the opemtion i.. carried out in separator) funnels. Daviclow ( 3 ) has adapted the sulfuric acid treatment and solvent estrac.tion to a chroniatographic column. \Tith modifications, the Dsvidow rolunin has been found to be niorp rapid and consideralll>~ leps laborious well as in that in the writers’ experience. I11 the present work already report,ed, the modified Davidox column has been employed. T h e fat ~ a m p l e sv w e extracted in preparation for chromatographing as IoIIo\vR: Samples of known weight n-rye ground in a mortar with xnhydrous sodium sulfat,e and then transferred to 500-ml. gl. SQstoppered bottles uqing C.P. carbon tetrachloride :is solvent and transfer agent (50 to 70 nil. of carbon tetrachloride per 2 grams of fat). The bottles were shaken mechanically for a t least 2 hours. The contents xvere then filtered and the filtrate was chroiii:ttographed through previously prepared Davidow columns ( 2 ) . SCHECHTER-IIALLER A N L Y S I S
The Schcchter-Halier ( 1 1 ) method for determining D D T in micro quantities is now generally accepted as the best available. Various modifications have been introducecl from time to time and it is. doubtful whether any t v o laboratories follow exactly
