Radiochemical Determination of Lead-210 in Products and Biological Materials
Mill
CLAUDE W. SILL and CONRAD P. WlLLlS
Health and Safefy Division, U. S. Atomic Energy Commission, Idaho Falls, Idaho
b The determination of lead-210 through its bismuth-2 10 daughter has been re-examined critically for application particularly to complex inorganic systems in addition to biological materials. Special emphasis has been placed on complete recovery of both lead and bismuth, so that either element can be used in the determination. In the large number of cases in which secular equilibrium i s known to exist, the bismuth fraction can be counted and results obtained within a few hours, eliminating the lengthy delays common to many other procedures. A combination of potassium fluoride and pyrosulfate fusions ensures complete decomposition of siliceous solids. Extraction with a solution of diethylammonium diethyldithiocarbamate in chloroform separates lead and bismuth quantitatively from large quantities of other metals in strongly acid solution. Dithizone then eliminates the remaining interferences and separates the two elements from each other prior to beta counting. High activities of gamma-emitting lead2 1 2 and bismuth-207 tracers were used to detect and identify any source of error larger than about 0.05%. Recovery of both elements through the entire procedure i s over 99% from 500 ml. of river water or liquid mill effluents, 1 gram of ores or mill tailings, 100 grams of blood, tissue, feces, or grain, 1500 ml. of urine, or large samples of air dusts. Overall recovery i s still about 98% even from 100 grams of soil or bone ash. Separation of lead and bismuth from each other i s better than 99.9%. Several serious sources of error are discussed.
F
regulations require that the concentration of soluble lead-210 in liquid effluents be less than 1 x 10-7 pc. per ml. for release to uncontrolled environments (5). In fact, only radium-226 and -228 have lower maximum permissible concentrations (MPC), To determine compliance with the regulations by mills engaged in processing uranium ores, analytical procedures capable of determining concentrations a t least as small as 0.1 MPC are reEDERAL
quired. At 20% counting efficiency, 0.1 NPC amounts to only 2 c.p.m. on a 500-ml. sample. Also, because of the severe biological consequences resulting from this nuclide, very sensitive procedures are required for its determination in biological materials and environmental samples. Holtzman ( 7 ) reports that the radiation dose to the human skeleton from lead-210 and its bismuth and polonium daughters is more than twice that from radium-226 and its daughters and about equal to that from radium-226 and radium-228 combined. Lead-210 has a half life of 22 years and emits beta particles with a maximum energy of only 0.018 m.e.v., making direct determination of this nuclide very difficult. The procedure generally employed involves determination of either the 5.01-day bismuth-210 or the 138-day polonium-210 daughters in equilibrium with the lead310 parent. Much of the work on determination of lead-210 has been directed a t determination of the natural levels occurring in water, air, and biological materials, where sensitivity is of prime importance; relatively little has been done on more complicated inorganic systems. Consequently, the polonium procedure seems to have been preferred because of its relatively simple chemistry and the higher inherent sensitivity obtainable with alpha counters because of their higher counting efficiencies and lower backgrounds. However, its long half life is a distinct disadvantage when time is important. Few samples of either biological or mineralogical types could safely be assumed to contain such a long-lived daughter in exact equilibrium with its parent. It is generally necessary to separate the lead210 and then allow sufficient time for ingrowth of fresh polonium-210 to a significant fraction of its equilibrium value. It is not uncommon to permit delays of 1 to 3 months in the analysis ( 7 ) to obtain sufficient activity bearing a known relationship to its parent. Such lengthy delays are obviously costly and undesirable, but are necessary if the inherent advantage of sensitivity is not to be lost. Sensitivity is not of paramount importance in analysis of mill products
and many other mineralogical types of samples. Most ores, tailings, liquid effluents, etc., from mills processing uranium ores generally contain concentrations of lead-210 high enough to be determined easily on moderately sized samples in reasonable counting times, in comparison to the natural levels present in the environment and in biological materials. Even 0.1 MPC for release to uncontrolled environments can be reached without much difficulty. The bismuth-210 daughter of lead-210 has a very energetic 1.17m.e.v. beta emission that is easily detected with good efficiency. Furthermore, the daughter’s half life is so short that secular equilibrium with its longlived parent can be assured on many types of biological and mineralogical samples without significant delay. Any sample known to have been captive or chemically unaltered for a few weeks can be analyzed almost immediately. The longest delay will be required of samples such as barren raffinates or certain biological tissue in which bismuth might have concentrated significantly in comparison to lead. However, even when regrowth of a daughter in the freshly purified lead-210 parent is required to ensure an unequivocal relationship between them, a maximum delay of only 5 days is required when counting bismuth-210 to come within a factor of 2 of the maximum sensitivity obtainable. The usual procedure is to hold the specimen for at least 35 days before counting to allow any bismuth210 not associated with lead-210 to decay to less than 1% of its original activity ( 3 ) . Aronson and Hammond (3) reduce the waiting time by separating lead from bismuth with dithizone a t p H 3.8 and then holding the lead fraction for 11 days before counting freshly ingrown bismuth-210. Special consideration is given in the present procedure toward keeping the total elapsed time of the analysis as small as possible. Particularly, all separations are designed to give equally complete recovery of lead and bismuth, so that either fraction can be used, depending on whether or not secular equilibrium exists. Determination of lead-2 10 through its bismuth-210 daughter is complicated VOL. 37, NO. 13, DECEMBER 1965
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by the possible presence of other lead, bismuth, and thallium nuclides from all the naturally occurring radioactive series. Ordinarily, a 5-hour delay between separation and counting allows 99.9% or more decay of 26.8-minute lead-214 or 19.7-minute bismuth-214 from the uranium-238 chain or of 36.1minute lead-211, 2.16-minute bismuth21 1, or 4.79-minute thallium-207 from the uranium-235 chain. The bismuth210 can then be counted without separation from the lead-210, since the soft beta from the latter nuclide is completely absorbed if the window of the counting chamber is thicker than about 1 mg. per sq. cm. However, many low-grade uranium ores and mill products contain enough natural thorium to contribute activities of 10.6hour lead-212 that are much larger than that of the lead-210. Consequently, counting of a lead fraction requires a delay of several days to ensure adequate decay of any possible lead-212. On the other hand, if bismuth-210 is separated from the lead, a few hours produces sufficient decay of the 60.5-minute bismuth-212 and its 3.1-minute thallium-208 daughter to eliminate all interference from the natural thorium chain with a considerable saving in time. Generally, it is convenient to allow the separated bismuth fraction to decay overnight, which will give complete elimination of interferences from all naturally occurring lead and bismuth isotopes while causing only about 10% decrease in sensitivity due to decay of the bismuth-210. The separated lead210 fraction can also be used, since the &day delay necessary for 50% ingrowth of the bismuth-210 daughter will also be adequate for complete decay of any lead-212 that could reasonably be expected. Dithizone has always been a reagent of choice for the spectrophotometric determination of lead and bismuth. Both elements can be separated simply and completely from practically all others except polonium, monovalent thallium, and divalent tin by extraction from an ammoniacal citrate solution containing cyanide. Lead can then be stripped from the dithizone extract with a pH 3.4 buffer, leaving bismuth quantitatively in the organic phase (4). The same separations should be equally adaptable to the radiochemical determination of lead-210, using either the lead or bismuth fractions as desired. These separations have been used so widely for the spectrophotometric determination of lead and bismuth with such excellent results that it is surprising that the same chemistry has not been used more extensively for their radiochemical determination. In a recent review (6), only two of 15 radiochemical procedures cited for lead employed dithizone, one of which (2) was not developed for 1662 *
ANALYTICAL CHEMISTRY
radiochemical use. More recently, Aronson and Hammond (3) and Jaworowski (8) have applied similar systems to the determination of lead-210 in biological material and Talvitie and Garcia (14) used dithizone for the determination in various environmental samples. Jaworowski used dithizone for the initial separation of lead-210 and bismuth-210 from the sample but changed to a cupferron extraction for separating the elements from each other. He injects an unfortunate note of confusion into the dithizone chemistry of lead and bismuth by stating that the two elements cannot be separated from each other with dithizone a t low pH because “the bulk of lead goes into the aqueous phase together with bismuth,” a conclusion reported to have been confirmed by two other groups of investigators. The present investigation conclusively reaffirms the longestablished fact that excellent separations of the two elements from each other are easily obtained when proper consideration is given to the effect of dithizone concentration on the acidity required. The present procedure has been developed particularly to provide for certain problems encountered in the analysis of mill products and other mineralogical samples that are uniquely different from those encountered in analysis of biological materials. The procedure is designed to be used in a sulfate system both to accommodate the high concentrations of sulfates resulting from leaching of ores with sulfuric acid and to provide for dissolution of lead present as the insoluble sulfate. Concentrated sulfuric acid and alkali sulfates can then be used to facilitate the wet decomposition of organic materials and potassium fluoride and/or pyrosulfate fusions can be used to obtain complete decomposition of refractory silicates with simultaneous volatilization of silica (10). The separations and high concentrations of complexing agents and extractants used are designed t o accommodate the large quantities of other elements, including stable lead, that are usually present in large samples of ores and raffinates. EXPERIMENTAL
Instrumentation. A low-background beta counter must be used to obtain maximum sensitivity with the shortest counting time. I n the present work, a background counting rate of about 0.7 c.p.m. and a counting efficiency for bismuth-210 of about 20% were obtained initially with a 1-inch counting chamber with the Lowbeta counter manufactured by the Sharp Laboratories of Beckman, Inc. However, about 25 to 50% of the polonium-210 present in the sample accompanies bismuth-210 in the present
procedure and special precautions are necessary to prevent interference from counting the alpha emission. The upper discriminator could be set so that hard betas would count efficiently while the polonium-210 alphas are counted with an efficiency of only about 1%. However, if any absorbing materials are present in the sample being counted, many of the alphas are degraded in energy below the discriminator setting and their counting rate increases significantly. Consequently, the safest practice is to use an absorber sufficiently thick to stop all alphas, which is particularly necessary with instruments not equipped with discriminators. An aluminum absorber approximately 6 mg. per sq. cm. thick will block nearly all of the polonium-210 alphas while reducing the counting rate of bismuth-210 by only about 20%. An absorber of 5.5 mg. per sq. cm. thickness was easily constructed from a piece of one of the heavier aluminum foils generally available for wrapping purposes. The absorber was made in the form of a cap that fits over the top of the planchet holder to serve the additional function of lessening contamination of the counting chamber. Standardization. The counting efficiency was determined by counting a n aliquot of a solution of bismuth-210 that had been standardized by 4-T counting. The bismuth-210 was obtained by activation of natural bismuth with thermal neutrons. Unless very high specific activities are known to be present, aliquots of different sizes should be 4 - r counted and the results extrapolated t o zero volume to eliminate the effect of self-absorption on the true disintegration rate. h long-lived standard was then prepared for continuing use from an aliquot of a commercially available lead-210 solution containing bismuth-210 in secular equilibrium, which was standardized by counting under the recommended conditions. Tracer Studies. 411 separations and methods of sample treatment were checked with both lead and bismuth tracers, so that either element could be employed in the analysis as desired. Approximately 1.4 x 105 c.p.m. of bismuth-207 or 4 x 105 c.p.m. of lead-212 (12) were used for each experiment. Each fraction was gamma-counted for 1 minute in a 3-inch thallium-activated sodium iodide well crystal in aqueous solutions of 75-ml. volume, observing the necessary precautions to obtain precision (12). Any loss as large as 0.05% was detectable and each tracer could be accounted for to within 100.0 f 0.3% in nearly all cases. Apparatus. If the inside bottoms of both the 250-ml. Erlenmeyer flasks and particularly the 150-ml. beakers used for evaporation of D D T C and dithizone extracts, respectively, are pitted extensively with an engraving tool, the chloroform will evaporate smoothly without a boiling chip. Otherwise, violent bumping and con-
Dissolve 8.0 grams of phthalic acid and 3 grams of sodium hydroxide in 200 ml. of water by warming gently until solution is complete. Cool and dilute to 900 ml. Mix the sodium perchlorate and sodium acid phthalate solutions and dilute to 1 liter. Adjust the pH to 2.7 by careful dropwise addition of 72% perchloric acid using a pH meter. LEAD-BISMUTH CARRIER. Prepare a solution of lead and bismuth perchlorates in 1% perchloric acid containing 100 fig. per ml. of each element. Tap and River Waters. Place 500 ml. of the neutral water sample to be analyzed and 1 ml. of lead-bismuth carrier in a 1-liter beaker. Add 40 ml. of concentrated hydrochloric acid and a boiling chip and boil vigorously for 5 minutes to dissolve lead sulfate, or hydroxides, phosphates, or sulfides of lead or bismuth that might be present in small quantities. If much insoluble material is present, filter the solution to prevent interference with phase separation during the subsequent extraction, using a little paper pulp as a filter aid. If any lead activity that might be associated with the siliceous material is desired, treat the inReagents. DIAMVONIUM CITRATE, solubles as described for ores and mill tailings. Cool the solution to room 40%. Dissolve 400 grams of ditemperature, add about 100 mg. of ammonium citrate, (XHJzHCsHs0,, in solid ascorbic acid, and transfer to a 825 ml. of distilled water to obtain 1 1-liter separatory funnel. Add 20 ml. liter of solution. of a 1% solution of diethylammonium SODIUMCYANIDE, 40y0 (sulfide-free). diethyldithiocarbamate (DDTC) in Dissolve 400 grams of sodium cyanide chloroform, shake vigorously for 1 in 775 ml. of water to obtain 1 liter of minute, and draw the extract off into a solution. Add 10 mg. of lead acetate 250-ml. Erlenmeyer flask. Repeat the dissolved in a few milliliters of water extraction with an additional 10 ml. of and heat the solution gently until any DDTC. Add a few milliliters of chlorolead sulfide produced flocculates well. form to recover the last extract Filter the solution while still warm completely and to rinse the stem of the through a double glass-fiber filter paper funnel. Discard the aqueous solution. in a Biichner funnel, using very gentle Add 2 ml. of 50% sodium hydrogen suction. If more than a light brown sulfate (NaHS04) and 2 ml. of conturbidity was produced, repeat the centrated nitric acid to the flask contreatment with lead acetate and filtertaining the combined extracts and heat ing. Store the solution in a polygently on a hot plate until the chloroethylene bottle. form has been expelled. Place the flask HYDROXYLAMINE, 20%. Add 2 on a high-temperature hot plate covered grams of hydroxylamine hydrochloride with a thin piece of asbestos cloth and and 2 drops of 0.1% phenolsulfoneheat strongly until nitric acid has been phthalein indicator to 10 ml. of water. expelled and a black cake of charred Add concentrated ammonium hydroxide organic matter is obtained. As soon solution with continuous swirling and as the charred mass is completely dry cooling until the salt has dissolved and immobile and no further charring and the red alkaline color of the inis evident, cool the flask, add 3 ml. of a dicator is produced. Prepare fresh 1 to 1 mixture of concentrated nitric each day in the quantity required. and 72% perchloric acids, and boil until DITHIZONE, 0.006 and 0.03%. Shake the black suspension becomes colorless 1 liter of chloroform vigorously with and most of the excess perchloric acid 100 ml. of water containing about 0.5 has evaporated. If the bisulfate cake gram of hydroxylamine hydrochloride has a yellow color while hot and is made alkaline to phenolsulfonephthalein colorless when cold, some iron is present indicator with ammonium hydroxide. but oxidation of organic matter is Filter the chloroform through a dry probably complete. However, if addouble acid-washed filter paper into a ditional charring is produced or yellow glass-stoppered borosilicate glass bottle noniron colors are still present on heatcontaining 60 or 300 mg. of dithizone ing to near dryness, repeat the evaporaas required. Add 5 ml. of ethanol as a tion with a few drops of mixed oxidizing preservative and store in a darkened acids to complete the elimination of refrigerator when not in use. organic matter. BUFFER,p H 2.7. Dissolve 20 grams Cool the Erlenmeyer flask and add 50 of anhydrous sodium perchlorate in ml. of water, 2 ml. of concentrated about 50 ml. of water and filter if hydrochloric acid, and a boiling chip. necessary. Add 2 or 3 drops of 7201, perchloric acid followed by a few drops Boil the solution vigorously for a few of %yosodium metabisulfite sufficient minutes until lead sulfate and any to produce a slight odor of sulfur dioxide. anhydrous ferric sulfate that might be sequent loss of sample occur frequently unless great care is taken. Separatory funnels having Teflon stopcocks should be used if possible to eliminate grease and facilitate cleaning. Grease dissolved from conventional stopcocks during chloroform extraction is not destroyed completely during wet ashing and makes the beaker very greasy and difficult to wash. Failure to transfer the bismuth or lead activities from the beaker to counting planchet quantitatively is one of the most likely sources of error in the entire procedure. Similarly, activity dissolved from greasy stopcocks used with high-level samples is one of the most common sources of contamination of blanks and low-level samples. If conventional stopcocks must be used, they should be cleaned frequently and greased lightly with Vaseline petroleum jelly or other nonsilicone lubricant. The funnel should then be washed thoroughly with chloroform to remove excess grease from the stopcock bore and funnel stem before using.
present have dissolved and a clear solution is obtained. Remove the flask from the hot plate and add 4 drops of 0.1% phenolsulfonephthalein indicator, 5 ml. of 4ooj, diammonium citrate solution, and enough concentrated ammonium hydroxide added dropwise with continuous swirling to produce the red alkaline color of the indicator plus an excess of 1 ml. ildd 5 ml. of 40% sulfide-free sodium cyanide and 1 ml. of 20% hydroxylamine solution. Allow the solution to stand hot without further heating for about 2 minutes, or more if much iron is present, then cool thoroughly in a bath of cold running water. The solution will have a pH of about 9.5. The pH should be above 9.0 where the extraction of lead and bismuth is most efficient but below 10.2 where there is still sufficient dithizone left in the chloroform to indicate the end of extraction. Transfer the solution to a 250-ml. separatory funnel with sufficient water to make a total volume of about 80 ml. Add 10 ml. of 0 . 0 0 6 ~ 0dithizone and shake vigorously for 1 minute to extract lead, bismuth, and thallium nuclides. Do not use the same graduated cylinder to measure the dithizone as was used with the dithiocarbamate solution or serious error will frequently occur in the lead-bismuth separation due to residual DDTC. Draw off the chloroform extract into a second 250-nil. separatory funnel containing 25 nil. of distilled water, repeat the extraction twice with consecutive 10-ml. portions of dithizone, and combine both extracts with the first. The first extract will change almost completely to a red color on shaking for a few seconds because of the color of the dithizonates of the lead and bismuth carriers added. The second and third extracts will retain the bright green color of unchanged dithizone with most water samples. If the first extract changes abruptly to a bright red color with the first two or three shakes because of the presence of significant quantities of lead or bismuth in the sample, add 5-ml. portions of 0.03% dithizone and shake vigorously after each addition until the new portions stop changing to red before drawing off the extract. Resume the extraction with 0.006% reagent and make one extraction after the dithizone remains the bright green color of unchanged dithizone. Each time the dithizone extract is drawn off, including all later steps, add 1 or 2 ml. of chloroform to the aqueous layer and draw off without shaking to recover the previous extract completely and to rinse the stem of the separatory funnel for quantitative recovery. Discard the aqueous solution and rinse the separatory funnel thoroughly with water. Shake the combined dithizone extracts vigorously for 1 minute with 25 ml. of distilled water and draw the extract into the original cleaned 250-ml. separatory funnel containing 35 ml. of pH 2.7 buffer. Rinse the wash solution and separatory funnel tvith 2 or 3 ml. of chloroform but do not shake, or an emulsion generally results. Discard the aqueous solution and rinse the separatory funnel thorVOL. 37,
NO. 13,
DECEMBER 1965
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oughly with water. Shake the dithizone extract with the p H 2.7 buffer solution vigorously for 1 minute to strip the lead out of t'he organic layer. Note the tinie of this first strip to correct for subsequent decay of bismuth-210. Draw the chloroform layer into the second 250-ml. separatory funnel containing another 35-ml. portion of pH 2.7 buffer and repeat the strip. If over 30 ml. of 0.037, dit'hizone lvas used, make a third strip to ensure complete separation of lead. Draw off the dithizone extract into a 150-ml. beaker. Add the strip solution in the second separatory funnel to that' in the first one and extract with one 10-nil. portion of fresh dithizone to recover a small quantity (ca. 2%) of bismuth that will have been stripped froin the previous dithizone extracts. AlIo\v the extract to settle thoroughly and separate it cleanly from the aqueous solution to prevent Contamination of the bismuth fraction n-ith some of its long-lived parent and interference with proper decay corrections. I d d t,he extract to the beaker containing the main dithizone extract. If the determination is also to be made from the lead fraction, reserve the combined strip solutions for further treatment. .Add 5 nil. of concentrated nitric acid to the 150-nil. beaker containing the combined dithizoiie extracts. Cover the beaker with a watch glass and heat carefully until the chloroform and all but about 0.2 nil. of nitric acid have evaporated, but do not allovi the beaker to bake. Cool) add 5 nil. of 309.', hydrogen peroxide! and repeat the evaporation to about 0.2-nil. volume. *ldd2 nil. of concentrated nitric acid and heat until all excess peroxide has been decomposed and frothing has ceased. Add concentrated nitric acid if necessary to adjust the volume to about 2 ml. Transfer the solution to a 1-inch stainless steel ringed counting planchet that has been cleaned by boiling for a few minutes in concentrated nitric acid, and begin evaporating the solution under a n infrared lamp. .Add 2 nil. of concentrated nitric acid to the empty
Table 1. Hours
beaker, cover with the same watch glass, and boil for a few seconds to reflux the cover and sides. Without evaporating more than about 0.5 ml. of acid, transfer the rinse while still hot to the planchet under the infrared lamp. With high-counting samples, allow a few minutes for some of the solution to evaporate froin the planchet and repeat the nitric acid rinse of the beaker to obtain quantitative transfer. Evaporate the solution to dryness. When completely dry, heat the planchet on a high temperature hot plate until residual organic matter is \vel1 charred and evolution of moisture and organic fumes has ceased. If more than 20 ml. of 0.03y0 dithizone was used in the extraction, or if considerable grease is present, heat the planchet intermittently over a very small flame to burn off the charred material and fui ther reduce self-absorption. Do not heat the planchet hot enough to produce more than a barely perceptible red glow for even a few seconds, or a detectable quantity of lead or bismuth will be volatilized. Store the planchet in a covered Petri dish to prevent contamination or accumulation of dust and allow to stand overnight to permit decay of bismuth212. Count in a low-background counter for a n appropriate length of time. Generally, 30 minutes viill suffice for water samples as far as federal regulations for disposal are concerned. However, counting times required can range from as little as 1 minute with some ores, tailings, and raffinates to several hours to determine the natural levels in air, m-ater, biological materials, etc. (1, 7 ) . Subtract a blank that has been carried through the same procedure at the same time as the samples using identical quantities of the same batch of reagents. Correct the observed counting rate of the bismuth-210 back to the time of the lead-bismuth - 0 693t -~
separation by dividing by e is where t is the elapsed time and the half life of bismuth-210. Corrections for decay can be made most con-
Decay Factors" for Bismuth-210 as a Function of Time Days
2 3 4 5 0.5007 0.6603 0,5750 0.7583 0.4950 0 6527 0.5684 0,7496 0.5619 0.4893 0.6453 0.8509 0.7410 0,5555 0,4837 0,7325 0.6379 0.8412 0,4782 0.6306 0.5491 0.7241 0,8315 0,4727 0.6233 0.5428 0.7158 0.8220 0.4673 0.6162 0,5366 0.7076 12 0.8126 0.5304 0.4619 0.6091 0.6995 14 0,8033 0.5243 0,4566 0.6915 0.6021 0.7941 16 0.5183 0.4514 0,5952 0.6835 0.7850 18 0.5124 0.4462 0,5884 0.6757 0.7760 20 0.5065 0.4411 0,5817 0.6680 22 0,7671 a Factors given are values of e-x' for bismuth-210 using a half life of, 5.01 days and cover through the fifth day recommended for ingrowth of bismuth-210 in the lead-210 fractions. Factors for longer times can be obtained by multiplying the factor for the time in excess of the largest even multiple of 5 days from Table I by (1/2)", where n is the number of 5-day half lives involved. For example, the fraction of the initial activity remaining after 17 days and 14 hours is 0.6995 X (l/2Y or 0.0874. 0 1.0000 0.9885 0.9772 0.9660 0,9549 0.9439 0.9331 0.9224 0.9119 0.9014 0.8911 0.8809
0 2 4 6 8 10
1664
e
1 0.8708 0.8608
ANALYTICAL CHEMISTRY
veniently by using the factors given in 2-hour increments in Table I. If secular equilibrium between the bismuth-210 and its parent is not likely to have been achieved] or if a confirmatory check on this condition is desired, a completely unambiguous analysis can be obtained from the fraction containing the lead-210, although longer time is required. To the strip solution in the separatory funnel add 1 ml. of concentrated hydrochloric acid, 2 ml. of 40% ammonium citrate, 4 drops of 0.1% phenolsulfonephthalein indicator, and enough concentrated ammonium hydroxide added dropwise with continuous swirling to produce the red alkaline color of the indicator plus a n excess of 1 ml. Add 1 ml. of 40% sodium cyanide and 0.5 ml. of 20% hydroxylamine and make consecutive 1-minute extractions each with 10 ml. of dithizone until two extracts are obtained that retain the bright green color of unchanged dithizone, as occurred with the original extractions. Generally, three extractions will suffice for water samples. Combine the extracts in a 150-ml. beaker. Rinse the separatory funnel with 2 or 3 nil. of chloroform and evaporate the combined extract to about one-half its volume to remove ammonia. Add 5 ml. of concentrated nitric acid and evaporate the remainder of the chloroform. Oxidize the organic matter with hydrogen peroxide, transfer the final nitric acid solution to a counting planchet, and dry as a i t h the bismuth fraction. However, add 0.5 ml. of water to each nitric acid rinse to ensure complete dissolution of lead nitrate. Store the planchet in a covered Petri dish for approximately 5 days for ingronth of bismuth-210 before counting. Calculate the equilibrium activity of bismuth-210, and therefoie of lead-210, by dividing the observed activity by the fraction of the iiiaxinium equilibrium activity that has grown in since the leadbisnuth separation was made. The growth factors are given by the ex-0.6931
pression 1 - e t ' l 2 , where the t's have the meaning given above and can also be obtained conveniently from Table I by subtracting the decay factor for the time in question from unity. Other Water Samples. Certain modifications are necessary in preparing the sample when other than fairly clean tap and river waters are analyzed. Some liquid mill effluents such as the barren raffinates and other process wastes generally contain large quantities of heavy metals such as iron, copper, and vanadium. To prevent loss of lead in the DDTC extraction due to too high a n acidity, neutralize any strong acid present with concentrated ammonium hydroxide until a permanent precipitate begins to form in the presence of heavy metals or to a phenolsulfonephthalein end point in their absence. Reacidify the solution with 8 ml. of concentrated hydrochloric acid for each 100 ml. of
sample taken and boil for 5 minutes to dissolve any lead sulfate or bismuth phosphate that might be present. Cool the solution and add solid ascorbic acid in approximately 0.5-gram portions until the solution is decolorized or until no further color changes result, to ensure complete reduction of iron. Complete the analysis as described for tap and river waters, beginning with the DDTC extraction. If large quantities of copper or other elements extracting into DDTC from strong acid solution are present, use a 5% solution of DDTC in chloroform and repeat the 1O-ml. extractions as necessary to obtain complete extraction. Generally, one extraction after the highly colored extracts have been removed will quffice. If a precipitate forms during the first extraction, add more DDTC solution and re-extract before drawing off the extract. After charring the extracts, use 10 ml. of 1 to 1 nitric-perchloric acid initially and repeat with 5-ml. portions as necessary to complete the oxidation of the large quantities of organic matter present. Waters containing organic material or even small quantities of sulfur compounds, particularly those giving rise to sulfides, must be evaporated and wetashed with nitric and perchloric acids. Evaporate most of the excess perchloric acid and dissolve the remaining salts by boiling with 50 nil. of water and 6 nil. of concentrated hydrochloric acid. Filter the solution if necessary, cool, add ascorbic acid, and transfer to a separatory funnel with about 25 ml. of water. Extract with three consecutive 1O-nd. portions of 1% DDTC in chloroform. Finish as described. Ores and Mill Tailings. Place 1 gram of sample in a 250-ml. Erlenmeyer flask and add 1 ml. of leadbismuth carrier and 5 ml. each of concentrated nitric and 7270 perchloric acids. Heat the flask on a hot plate until fumes of perchloric acid are evolved and all organic matter has been oxidized. If necessary, add a few drops of a 1 t o 1 mixture of nitric and perchloric acids to complete the oxidation. Boil the solution until only about 1 ml. of perchloric acid remains. Cool the flask, add 50 ml. of water, 6 ml. of concentrated hydrochloric acid, and a boiling chip, and boil gently for about 5 minutes to dissolve bismuth phosphate and metallic sulfates, particularly those of lead and calcium. Cool and transfer the solution and insoluble material to a 100ml. Lusteroid centrifuge tube with enough water to make the total volume about 80 ml. Mix and centrifuge a t 2000 r.p.m. for 5 minutes. If the supernate is not completely clear, centrifuge for additional time. Decant the supernate into a 250-ml. separatory funnel and reserve for later treatment. Transfer the insoluble material from the centrifuge tube to a 50-ml. platinum dish with 5 consecutive 1-ml. portions of 48y0 hydrofluoric acid, add 1 ml. of lead-bismuth carrier, and evaporate carefully to dryness on a hot plate. Sprinkle 3 grams of anhydrous potassium fluoride uniformly over the resi-
due, fuse, and transpose with 6 ml. of concentrated sulfuric acid and 2 grams of anhydrous sodium sulfate to a pyrosulfate fusion (10). Clean the outside of the platinum dish and place it upright in a 400-ml. beaker. Add 5 ml. of concentrated hydrochloric acid and 50 ml. of water t o the platinum dish and swirl some of the solution into the beaker, Cover the beaker with a watch glass and heat gently to boiling with occasional rapid swirling until the cake becomes detached from the sides of the dish and all small isolated pieces of the cake around the sides have dissolved. Remove the dish and rinse it with about 20 ml. of water. Continue boiling the solution gently until the cake has distintegrated completely and cool to room temperature. If the solution is completely clear, transfer it to a second 250-ml. separatory funnel. Add sufficient ascorbic acid to the solutions from the pyrosulfate fusion and from the original perchloric acid treatment to reduce all iron and leave an excess of about 0.5 gram. Extract each solution with three consecutive 10-ml. portionsof 1% DDTC. Combine all six extracts in a 250-ml. Erlenmeyer flask, add sodium acid sulfate and nitric acid, and finish as described for tap and river waters. Do not combine the two solutions before the extraction or precipitation of potassium perchlorate and/or calcium, or barium sulfates will cause loss of lead and interfere with phase separation. If relatively large quantities of lead and other elements might be present, use either a 5% solution of DDTC and/or a 0.03% solution of dithizone as required to obtain complete recovery in a reasonable number of extractions when using samples as large as 1 gram, particularly on uranium ores. Make one extraction with 5% DDTC after the original dark extracts have become light brown or yellow to colorless, and one with the o.o3YO dithizone that retains the brilliant green color of the unchanged reagent. If a detectable turbidity still remains after cooling the hydrochloric acid solution of the pyrosulfate fusion, transfer the solution to a 90-ml. borosilicate centrifuge tube and centrifuge a t 2000 r.p.m. for 5 minutes before decanting the supernate carefully into the second 250-ml. separatory funnel. Drain the tube as thoroughly as possible and extract the solution with DDTC as described. Add 15 ml. of 72y0 perchloric acid to the tube and heat over a blast burner with continuous swirling until the barium and/or calcium sulfate has dissolved. Cool the perchloric acid to room temperature and transfer it to a third 250-ml. separatory funnel containing 150 ml. of water and 5 ml. of concentrated hydrochloric acid with continuous gentle swirling. Rinse the tube with 1 ml. of perchloric acid and a few milliliters of water, adding each in turn to the main solution. Add 10 ml. of 1% DDTC and shake the separatory funnel vigorously for 1 minute as soon as possible. Repeat the extraction with one additional 10-ml. portion of 1% DDTC and combine it with the
main extracts in the Erlenmeyer flask. Larger samples can be analyzed if necessary by the procedure given for soils. Soil. Although samples up to about 1 gram can be analyzed by the procedure given for ores and mill tailings, much larger samples are required to determine the natural levels of lead-210 present in soil without excessive counting times. The following changes in the above procedure will permit 100 grams of soil to be decomposed completely and made ready for analysis in about 6 hours. Smaller samples can be analyzed with a proportionate reduction in reagents and apparatus. A 100-gram sample will permit detection of lead-210 activities larger than approximately 0.01 pc. per gram with a 30-minute count or 0.002 pc. per gram with an 18-hour count. Add 100 ml. of 72% perchloric acid, 10 nil. of concentrated nitric acid, and 1 ml. of lead-bismuth carrier to 100 grams of soil in a 1-liter Erlenmeyer flask and evaporate until most of the excess perchloric acid has been volatilized. Boil the residue gently for 5 minutes with 250 ml. of water and SO ml. of concentrated hydrochloric acid. Cool the solution thoroughly in a cold water bath to precipitate as much potassium perchlorate as possible before centrifuging. Transfer both solution and insoluble material into five 100-ml. Lusteroid centrifuge tubes with about 50 ml. of water. Centrifuge for a t least 10 minutes, decant the supernates back into the Erlenmeyer flask, add 150 ml. of water, and reserve for later use (Solution 1). Add 2 mi. of 48% hvdrofluoric acid to the insoluble matter in each tube, swirl vigorously, and pour the suspension into a 500-ml. platinum dish. Use an additional 10 ml. of 48% hydrofluoric acid for each tube to complete the transfer of insolubles to the platinum dish, Do not add more than 2 ml. of hydrofluoric acid initially while the insoluble material is still in the tube, or the violent reaction that frequently results will expel the boiling acid and serious burns may result . Add 1 ml. of lead-bismuth carrier and evaporate the solution to dryness. Repeat the evaporation with one 50-ml. and two 25-ml. portions of hydrofluoric acid. Vse 85 grams of anhydrous potassium fluoride in the fluoride fusion and transpose to a pyrosulfate fusion with 95 ml. of concentrated sulfuric acid and 50 grams of anhydrous sodium sulfate. The fluoride fusion can be made on such large samples most conveniently by placing the dish in a muffle furnace for 10 minutes a t 1000° C. After cooling, place the platinum dish in a 2-liter beaker and decompose the cake by boiling with 300 ml. of water. Transfer the solution and precipitate of barium and calcium sulfates to six 100ml. Lusteroid centrifuge tubes and centrifuge a t 2000 r.p.m. for 5 minutes. VOL. 37, NO. 13, DECEMBER 1965
1665
Decant the supernates back into the 2liter beaker, add 20 ml. of concentrated hydrochloric acid, and boil for 5 minutes. Dilute to 500 ml., cool, and reserve for later use (Solution 2). Transfer the precipitates from the centrifuge tubes to a 600-ml. beaker with 300 ml. of 1M hydrochloric acid and heat the solution to boiling. If the coarsely crystalline precipitate of calcium sulfate dissolves, leaving only a fine suspension of barium sulfate, cool and transfer the solution to four 90-ml. round-bottomed borosilicate centrifuge tubes. Centrifuge a t 2000 r.p.m. for 5 minutes. If the calcium sulfate does not dissolve completely, add 1M hydrochloric acid until it does and centrifuge in a proportionately larger number of tubes. Combine all supernates in a 500-ml. separatory funnel (Solution 3). Distribute 15 ml. of 72% perchloric acid evenly among the glass centrifuge tubes used and heat each tube until the barium sulfate dissolves. Ignore a slight turbidity of original sample that escaped complete decomposition. Cool and pour the perchloric acid solutions into a 250-ml. separatory funnel containing 150 ml. of water and 5 ml. of concentrated hydrochloric acid while swirling the funnel continuously. Rinse all the tubes consecutively with one 2-ml. portion of 72% perchloric acid and then with a few milliliters of water, adding each rinse in turn to the main solution (Solution 4). Extract as quickly as possible with 20 ml. of 1% DDTC as described. Add 0.5 gram of ascorbic acid to Solution 3 and extract with 20 ml. of 1% DDTC. Add sufficient ascorbic acid to Solutions 1 and 2 to reduce all iron and leave an excess of a t least 0.5 gram and transfer the solutions into 1-liter separatory funnels. Extract each solution with 20 ml. of 1% DDTC. Combine the extracts from all four solutions in a 250-ml. separatory funnel containing 50 ml. of water and 5 ml. of 72% perchloric acid. Re-extract each of the four solutions with an additional 10 ml. of 1% DDTC and combine with the main extracts. Calcium sulfate will begin to precipitate from Solutions 2 and 3 and barium sulfate from Solution 4 shortly after the solutions become saturated with chloroform, and the extraction should be completed without delay. Shake the combined extracts with the wash solution vigorously for 30 seconds and draw the extract into a 250-ml. Erlenmeyer flask. Add sodium acid sulfate and nitric acid and finish as for tap and river waters. Air Dusts. Collect an appropriate sample of air dust, using suitable high-volume sampling equipment.
A volume of about 20 cu. meters of air will allow detection of one tenth of the maximum level (4 X 10-12 IC. per ml.) of soluble lead-210 permitted for release to uncontrolled environments in a 30-minute count with the counter described. At least 2000 cu. meters will be required for determination of the natural levels present in air. The filtering media employed must be 1666
ANALYTICAL CHEMISTRY
analytically clean, very retentive, and capable of handling large volumes of air. Glass fiber papers permit complete decomposition of both filter and siliceous dust in a single container (10) but contain a suprisingly large quantity of barium which requires special treatment. Filters made of polystyrene or cellulose fibers are easily wet-ashed but require separate treatment of the insoluble residue from the sample. The following directions are for a 4inch circle of each type paper but can be scaled up to handle larger sizes. To decompose polystyrene filters such as Microsorban, place the filter in a 500-ml. Erlenmeyer flask and add 10 ml. of concentrated sulfuric acid, 5 ml. of concentrated nitric acid, 1 gram of anhydrous sodium sulfate, and 1 ml. of lead-bismuth carrier. Heat the solution strongly on a high-temperature hot plate until the polystyrene has been charred completely to a black immobile mass. Cool the flask, add 10 ml. each of concentrated nitric and 72% perchloric acids, and boil until all organic matter has been oxidized and about 1 ml. of colorless sulfuric acid remains. Add a few drops of a 1 to 1 mixture of concentrated nitric and perchloric acids if necessary to facilitate oxidation of the last of the organic matter. Cool the solution, add 50 ml. of water and 6 ml. of concentrated hydrochloric acid, and finish as described for ores and mill tailings. For cellulose papers such as MSA 2133, add 3 ml. of concentrated sulfuric acid, 10 ml. of concentrated nitric acid, 1 ml. of lead-bismuth carrier, and 1 gram of anhydrous sodium sulfate to the paper in a 500-ml. Erlenmeyer flask and boil. Each time the solution turns black because of charring of the cellulose, add 1 or 2 ml. of Concentrated nitric acid to clear the solution. When the solution can be evaporated nearly to fumes of sulfuric acid without producing colors darker than a light black or red, add 2 or 3 ml. of a 1 to 1 mixture of nitric and perchloric acids and evaporate to fumes of sulfuric acid to oxidize the last of the organic matter without further attention. Add water and hydrochloric acid and finish as with the polystyrene papers. For glass fiber papers, fold the paper into quarters with the sample toward the inside of the paper. Place the paper in a 50-ml. platinum dish and moisten the folded edge with 1 ml. of water while holding the other edge of the paper in place. Moisten the rest of the paper with 1 ml. of lead-bismuth carrier and 6 drops each of concentrated nitric and 72% perchloric acids. Add 5 ml. of 48% hydrofluoric acid a few drops at a time where the paper is the wettest to moderate the vigorous reaction that occurs with the glass fibers. Evaporate the solution to dryness, adding a few drops of a 1 to 1 mixture of nitric and perchloric acids if necessary to complete oxidation of the organic matter. Starting with the potassium fluoride fusion, complete the determination as described for the in-
soluble fraction under ores and mill tailings, including treatment of the barium sulfate that is frequently present. Bone. Place 100 grams of bone ash and 1 ml. of lead-bismuth carrier in a 1-liter beaker and add 170 ml. of 72% perchloric acid. Cover the beaker with a watch glass and heat the solution to boiling. After most of the bone ash has dissolved, place the beaker on the hottest part of an uncovered high-temperature hot plate and heat strongly until the black suspension of carbon that is generally present has been oxidized and a yellow solution due to decomposition products of perchloric acid is obtained. Remove the cover glass and continue heating with occasional swirling until the yellow color has nearly disappeared and the thick solution solidifies on the sides when the beaker is swirled. Do not prolong the heating until the solution itself becomes more turbid. Cool the solution and add 350 ml. of water. Stir the cake with a glass rod to dissipate the heat generated on dissolution of the cake in water and prevent the violent bumping that might otherwise occur. Add 10 ml. of concentrated hydrochloric acid, cover the beaker with a watch glass, and boil the solution vigorously for 5 minutes to dissolve bismuth phosphate and lead and/or calcium sulfates. Cool to room temperature and transfer to a 1-liter separatory funnel with about 100 ml. of water, making a final volume for extraction of about 600 ml. Shake vigorously for 1 minute with 20 ml. of 1% DDTC followed by two consecutive extractions with 10-ml. portions. Draw each extract off into a 250-ml. separatory funnel containing 5 ml. of 72% perchloric acid and 50 ml. of water. Shake the separatory funnel vigorously for 30 seconds and draw the extract into a 250-ml. Erlenmeyer flask. Extract the wash solution with 5 ml. of chloroform and add the extract to the Erlenmeyer flask. Add sodium acid sulfate and nitric acid to the combined extracts and finish as described for tap and river waters. For smaller samples, reduce the scale of both apparatus and reagents. Use 1.7 ml. of 72% perchloric acid per gram of bone ash but not less than a total of 5 ml. for dissolution of the sample and a final volume of 6 ml. per gram of sample with a minimum of 75 ml. for the extraction. In the presence of such high concentration of salts and phosphoric acid, use not more than 1.2 ml. of concentrated hydrochloric acid per 75-ml. final volume, or extraction of lead will be seriously incomplete. If hydrochloric acid is not present, turbid solutions frequently result and some bismuth tends to precipitate as the very insoluble phosphate. Urine. Transfer a 24-hour sample (ca. 1500 ml.) to a 2-liter beaker. Rinse the original container with 5 ml. of concentrated nitric acid and add the acid to the main solution. Add 10 ml. of 85% phosphoric acid and a boiling chip, cover the beaker with a watch glass, and heat the solution to
boiling. Add 1 ml. of 0.1% phenolsulfonephthalein indicator and sufficient 8-14 sodium hydroxide while stirring the solution vigorously to precipitate all calcium phosphate and convert the indicator to its red alkaline form. If necessary, add a few drops of indicator down the side of the beaker and observe the color locally around the drops. An excess of 1 or 2 ml. of the sodium hydroxide will do no harm, but larger excesses should be avoided. Boil the solution vigorously for 10 minutes, leaving the cover glass slightly askew to prevent frothing. Decant the supernate, filter, and wet-ash the precipitate and paper as described previously ( 1 1 ) . After evaporating the 72% perchloric acid to about 2 ml. as described, add 50 ml. of water and 5 ml. of concentrated hydrochloric acid, and boil for 2 minutes. Cool, dilute to 100 ml., and extract with 3 consecutive 10-ml. portions of 1% DDTC. Add sodium acid sulfate and nitric acid to the extract and finish as described for tap and river waters with one important exception. Discard the bismuth dithizone extract and count only the lead fraction after 5 days as described. Recovery of bismuth from urine on calcium phosphate is very incomplete because of the competition from the chloride present. In any case the results would be very difficult to interpret in terms of lead-210, because of the uncertainty about the equilibrium conditions obtaining on a sample so recently altered chemically. Feces, Tissue, Blood, and Grain. Directions have been given (11) for decomposition of 50-gram samples using combinations of nitric, perchloric, and sulfuric acids. Larger samples can be accommodated by direct scale-up of the procedures given. Add 1 ml. of lead-bismuth carrier before beginning the decomposition. When oxidation of organic matter is complete, evaporate excess perchloric or sulfuric acid until less than 2 ml. remains, but do not allow the solutions to dry and bake even in local spots. Add 100 ml. of water and 5 ml. of concentrated hydrochloric acid and boil for about 5 minutes to dissolve bismuth phosphate and lead, calcium, and iron sulfates. Cool, filter if necessary, and dilute to a 100-ml. volume. Reduce with ascorbic acid and extract with three consecutive 10-ml. portions of 1% DDTC. Finish as described for tap and river waters. RESULTS
After all developmental work had been completed, each of the procedures described was repeated in the presence of lead-212 and bismuth-207 tracers in separate runs to determine the recovery of each element actually obtained under the final conditions recommended. The results obtained
Table II.
Distribution of Lead and Bismuth in Recommended Procedurefor 500 MI. of River Water
Element Dresent. 41, .. dithizone" 0.03Yc dithizoneb Bismuth Lead Bismuth I
0.006';:
Lead Fraction First 20-ml. extract of 1% DDTC (96.5) 3.2 Second 10-ml. extract of 1% DDTC Third 10-ml. extract of 1% DDTC 0.2 Aqueous after 3 DDTC extractions 0.0 Insoluble material and/or filter paper 0.1 First Dz extract (94.7)c 1.1 Second Dz extract Third Dz extract 0.1 Fourth Dz extract ... Aqueous after last Dz extraction 0.4 Beaker containingoriginal alkaline Dz 0.2 solution Aqueous wash of Dz extracts 0.0 Second pH 2.7 buffer strip 0.3 Main bismuth Dz fraction 0.0 0.1 Dz extract of first pH 2.7 strip Main lead Dz fraction (first 2 Dz extracts) 93.7d Third Dz extract of final lead extraction 0.1 Fourth Dz extract of final lead extrac... tion 0.2 Aqueous after final lead Dz extraction Material balance 99.7 Total recovery of lead and bismuth in combined fractions 98.8 a 100 pg. each of lead and bismuth present. b 2.1 mg. of lead and 100 fig. of bismuth present. Red extract; no excess dithizone. Purple extract; excess dithizone present.
with 500-ml. water samples are shown in Table 11. The first set of data was obtained under conditions described for tap and river waters, in which 100 pg. each of lead and bismuth carriers was extracted from a typical sample of river water with 1% DDTC and 0.006% dithizone. The second set of data was obtained from river water to which 2 mg. of natural lead was added in addition to the regular lead-bismuth carrier and extracted with 1% DDTC and 0.03% dithizone. Each fraction was kept separate to show the actual distribution of both lead and bismuth under the conditions employed. Except for the runs with 2 mg. of added lead, in which the first three dithizone extracts were combined, only the fraction containing the major part of the tracer was carried on through the remaining steps of the procedure. The minor fractions were counted and subtracted from 100% to obtain the quantity in the main fraction. These values are shown in parentheses for comparison and should not be included in the material balance, since they are accounted for in subsequent fractions. All values not enclosed in parentheses were determined by actual counting of each final fraction obtained. The material balances show the excellent accountability of all tracer obtained. The bottom line in Table I1 gives the total recovery of both lead and bismuth that results when the fractions are
(96.9) 2.8 0.1 0.0 0.2 (64.7)" 31.9 0.2
...
0.0
0.1 0.0 0.8 63.2 0.9
(93.9) 5.4 0.6 0.1
(95.9) 3.9 0.2 0.0
cii : o ) c
(0:3)c (7.6)" (87.7)d 0.3 0.0
(33.2)c (19.6)d 0.1 0.0
... 0.0 2.1 0.0 0.1
... 0.0 0.1 95.2 0.1
O.Od
71. le
0.0"
0.0
20. 6 d
O.Od
...
0.0 100.2
100.2
0.0 0.0 99.8
99.6
99.2
99.8
0.1 0.0
combined as recommended in the procedure. The data of Table I1 show particularly the excellent efficiency of both the DDTC and dithizone extractions and the lead-bismuth separation a t pH 2.7 in the presence of up to 2 mg. of lead carrier. KO lead could be detected in the final bismuth fraction nor bismuth in the final lead fraction. Each of the other procedures described was checked out in final form in a similar manner with similar results. With ores, mill tailings, air dusts, feces, tissue, blood, and grain, the efficiency of the DDTC extraction is even greater than that shown in Table I1 because of the smaller aqueous volume to be extracted. Over 98% of both bismuth and lead can be recovered consistently in the DDTC extractions from 100 grams of either bone ash or soil. After the DDTC extraction, the procedure is essentially independent of the type of sample being analyzed. Although 99.1% of the lead was recovered as the phosphate from 1500 ml. of urine, less than 15% of the bismuth could be recovered under the same conditions. Since no difficulty is encountered in precipitating over 99% of both lead and bismuth with calcium phosphate from 10 grams of bone ash, the chloride present in urine apparently forms a complex with bismuth in alkaline solution of competitive stability with bismuth phosphate, which is not true of lead. In the DDTC extraction from VOL 37, NO. 13, DECEMBER 1965
1667
acid solution, the reverse is true and much higher concentrations of hydrochloric acid are permissible for extraction of bismuth than for lead. As a final proof of the efficiency of the separations employed and the equivalence of the determination from either the lead or bismuth fractions when in secular equilibrium, a 1-gram sample of a pitchblende ore was analyzed. The growth and decay curves of the lead-210 and bismuth310 fractions, respectively, are shown in Figure 1. The solid curves are not best-fit curves drawn to the experimental data points but are theoretical curves for a 5.0-day half life with data points superimposed. Curve 1 was obtained by drawing a straight line with a 5.0-day slope through the point on the ordinate obtained by extrapolation of the three highest data points back to zero time. This maximum activity of the bismuth fraction should also be the final activity of the lead fraction after regrowth of its bismuth-210 daughter to equilibrium. Curve 2 was then calculated from this equilibrium value, again using a 5.0-day half life. The excellent fit of both sets of data to the respective theoretical curves proves conclusively that both fractions are radiochemically pure and equivalent, since one curve was calculated from the other. Both curves were followed for 6 half lives without finding any evidence of radiochemical impurity. In view of the high and reproducible recovery of over 99% obtained consistently in the tracer studies for both lead and bismuth and the radiochemical purity obtained, it is evident that the precision and accuracy obtainable in actual analysis will be dependent almost entirely on the counting statistics obtained and the accuracy with which the counting efficiency can be determined and reproduced. To verify this point, duplicate analyses mere made on uranium ores using relatively highgrade material to obtain sufficient activity to keep the statistical uncertainty on a 30-minute count as small as the chemical and mechanical errors being checked. Since most errors encountered in analytical work are not likely to be proportional to sample size, samples of different size were used to emphasize such errors and increase confidence in the accuracy of the results when agreement between duplicates is obtained. With samples giving counting rates of 5 x 103 to 20 X lo3c.p.m. and differing by a factor of 2 in size, the larger sample in every case resulted in a lower activity per gram by as much as 15%. Furthermore, when the bismuth fraction was recounted at later times to verify the correct rate of decay of bismuth210, the values corrected back to the time of separation increased with time 1668
ANALYTICAL CHEMISTRY
'-
I
\ 10
103~
3
ELAPSED TIME, days
Figure 1 . Separation of bismuth-2 10 from lead-2 10 1. 2.
Decoy of bismuth-21 0 fraction Growth of bismuth-210 in lead-210 fraction
since separation, with the larger sample showing the greater increase. Both effects were shown to be due primarily to increasingly larger coincidence losses a t the higher counting rate which are reversed as the counting rates decrease because of decay. The low-background anticoincidence counter employed has a resolving time of 300 microseconds, which results in coincidence losses of 1% a t 1.8 X 103 c.p.m. and about 15% a t 25 X lo3 c.p.m. If the higher counting rates are desired, either to obtain higher statistical precision in shorter counting times or to accommodate more active samples without using inconveniently small samples, instruments with much faster resolution must be used instead of the slower lowbackground counters. One instrument used in this laboratory employs methane as the counting gas in a specially designed tube with a collimated window and a 1-microsecond scaler and can be used a t counting rates a t least as high as 4 x 105 c.p.m. without detectable coincidence loss. Obviously, similar considerations apply to the 4-i7 counter used in standardizing the solution used in determining the counting efficiency. Absorption errors resulting from impurities in reagents used in constant quantities in the analysis or from incomplete decomposition of organic reagents such as dithizone will produce a similar effect when samples of different size are used. Since the spectrum of bismuth-210 contains an abnormally large proportion of low-energy betas, absorption losses are higher than usual with this nuclide, particularly if the activity is not distributed uniformly over the counting planchet. When sufficiently small samples and pure reagents are used to avoid both coincidence and absorption losses, duplicate values obtained from differentsized samples invariably agree more closely than the statistical uncertainty
between them, which is about 1.5% a t the 95% confidence level for a 30-minute count a t the level employed. For example, the pitchblende sample mentioned above gave values of 5.99 X lo3 and 6.00 X lo3 d.p.m. per gram from the bismuth fraction and 5.97 X lo3and 5.89 X l o 3 d.p.m. per gram from the lead fraction from 0.5- and 1-gram samples, respectively. This particular material was selected hopefully as being a primary unaltered mineral to permit a direct evaluation of the accuracy of the procedure by comparing the lead-210 activity obtained with that calculated from the uranium content. The ore contains 0.973% U30a, which gives 6.05 x lo3 d.p.m. per gram if the lead210 is actually in secular equilibrium with its uranium-238 parent. The values obtained amount to 98.8y0 of the uranium equivalent, which is not statistically different from loo%, and adds considerable confidence that the accuracy is as good as is inferred from the tracer data and the precision obtained on actual samples of different size. An independent determination of the polonium-210 daughter, which has to be in secular equilibrium with its lead-210 parent, gave a value of 6.00 f 0.18 X l o 3 d.p.m. per gram for seven determinations on samples varying in size from 0.1 to 1.5 grams, The polonium analysis is 99.1% of the uranium equivalent in excellent agreement with the lead values. DISCUSSION
Effect of Alkaline Earths. I n a previous investigation (11), submilligram quantities of several elements, including bismuth, were found to be precipitated quantitptively under certain conditions by relatively small quantities of barium or strontium sulfates and less efficiently by calcium sulfate. Lead is also carried in the
barium sulfate lattice even under conditions where lead sulfate itself does not precipitate. Soils contain an average of approximately 0.035% strontium, 0.05% barium, and 1.5% calcium. Mill tailings have been found frequently to contain appreciable quantities of barium, perhaps concentrated many times over the concentration present originally in the ore or, in some cases, added deliberately during processing of the ore to increase decontamination of the liquid effluents from radium. As expected from the previous work, even a few milligrams of barium or strontium sulfates and large quantities of calcium sulfate will contain a significant fraction of the lead and bismuth activities present despite the large quantity of carriers employed. If the pyrosulfate fusion is clear, which is generally true if a potassium fluoride fusion was used to dissolve the original sample, both original insolubles and silica will be known to have been eliminated completely. Consequently, any material that does not dissolve in the hot acid solution of the pyrosulfate cake must be presumed to be an alkaline earth sulfate and must be treated to recover the bismuth and lead that it contains. Under the conditions employed, the barium sulfate from 1 mg. of barium produces a noticeable turbidity containing about 2.5% of the bismuth. Five milligrams of barium produces distinct turbidity and 15% loss of bismuth while 0.2 mg. can be seen only by the Tyndall beam from a flashlight and causes only 0.3% loss. Consequently, quantities of barium sulfate not apparent to the naked eye will not cause errors larger than about 1%. Fortunately, the quantity of barium sulfate usually encountered even in 100 grams of soil is easily soluble in boiling 7297, perchloric acid and can be kept in solution long enough to permit a DDTC extraction if the perchloric acid is poured into a large volume of water in the absence of significant quantities of sulfate. Dilute hydrochloric acid is used to reduce coprecipitation of lead sulfate. Also, high concentrations of sulfates are required for bismuth to be carried efficiently by barium sulfate, so that little bismuth will be lost if some barium sulfate is reprecipitated. The effect of alkaline earths can be demonstrated by the distribution of lead and bismuth obtained by the present procedure on a 100-gram sample of soil. Approximately 5097, of the lead and bismuth in the sample was removed in the original acid leach (Solution I). Of that remaining in the insoluble portion, only 3.0% of the bismuth and 9.3% of the lead were present in the entire 500 ml. of solution of the pyrosulfate cake (Solution 2). I n contrast, 90.4% of the bismuth and
56.8% of the lead were present in the hydrochloric acid solution of the calcium sulfate (Solution 3) and 6.6% of the bismuth and 33.9% of the lead were in the barium sulfate (Solution 4). Effect of Iron. Statements continue to appear in the literature that “the direct extraction of traces of lead with dithizone is not applicable t o solutions of highly alloyed steels as rapid oxidation of t h e reagent by iron(II1) occurs in the initially reduced and complexed alkaline cyanide solutions used for extracting lead.” That such statements are untrue can be readily demonstrated by the fact that a t least 10 grams of either potassium ferrocyanide or ferricyanide in 100 ml. of solution does not interfere with dithizone extraction in presence of excess hydroxylamine. The trouble usually encountered is due to the slowness of the complexing reaction between iron and cyanide in alkaline solution. When cyanide is added to an ammoniacal citrate solution of ferric iron, the yellow color of the ferric citrate complex changes to a bright red color suggestive of an iron oxide sol. If the quantity of iron present is small, the color quickly disappears on addition of hydroxylamine because of rapid formation and reduction of the ferricyanide complex. When large quantities of iron are present, complexation by cyanide in alkaline solution is relatively slow, perhaps because of precipitation of colloidal ferric oxide by the strongly alkaline cyanide. If all iron is not complexed and the resulting ferricyanide is reduced before the extraction is begun, the dithizone is oxidized rapidly by the ferricyanide as the latter is formed, and extraction is impossible or severely retarded. If excess cyanide and hydroxylamine are added to a boiling ammoniacal citrate solution of iron and the solution is allowed to cool spontaneously to room temperature, the deep red color produced initially from as much as 1 gram of iron will have lightened to the light yellow color of the ferrocyanide by the time the solution has cooled and no trouble will be experienced during subsequent extraction. The solution must not be boiled after addition of cyanide, since alkaline solutions of ammonium cyanide produce a deep red color on boiling that eventually produces a reddish brown precipitate. The quantities of iron extracting into DDTC will ordinarily not require more than 1 or 2 minutes in hot solution to eliminate all interference from this source. Effect of Silica. Maynes and McBryde (9) observed a loss of about 5% of their lead to the glass walls of the separatory funnel containing the alkaline citrate-cyanide solution a t pH 9.5 and found it necessary to rinse the glassware with dilute acid and re-
extract with dithizone to obtain complete recovery of lead. Loss of lead to glass surfaces from alkaline solution has been confirmed with radioactive tracers in the present work. In radiochemical work, the loss can be reduced to less than 0.3% (Table 11) a t pH 9.5 by using 100 pg. of lead carrier, but increases again to 1 or 2% a t pH 12. This loss of lead is very similar to the losses observed with beryllium in alkaline solution ( I S ) and is probably due to anion exchange on the glass. However, a much more serious loss occurs from silica in the sample. A 15% loss of lead was produced when 10 mg. of silica was added to the alkaline solution either as hydrated silicic acid or as the precipitate produced when soluble sodium metasilicate was added to the alkaline solution and the solutions were filtered. Similar results were obtained with bismuth, but the losses were only one-third as large as those with lead. Either hydrated silicic acid or dehydrated silica can be filtered off while the solution is acidic with virtually no loss of either lead or bismuth. If lead and bismuth are extracted with DDTC from acid solution, water samples need not be evaporated to dryness to dehydrate silica to avoid this serious error. Dithizone Extraction. From an 80ml. volume containing 5 ml. each of 40% diammonium citrate and 40% potassium cyanide and 100 pg. each of lead and bismuth, recovery of lead is greater than 95% in a single extraction with 10 ml. of 0.006% dithizone in chloroform from a p H of 9 to 11. Two consecutive extractions give better than 99% recovery from a pH of 8 to a t least 12. Bismuth appears to be estracted somewhat less efficiently than lead in the presence of citrate, recovery being only 80 to 85% in the first extraction in the pH range from 8 to 10 when 100 pg. of each element is present. Since 10 ml. of 0.006% dithizone contains only enough reagent to react with about 200 pg. of lead or bismuth, recovery of bismuth in the first extract will decrease as the lead content of the sample increases (Table 11). However, after most of the lead carrier has been removed in the first extraction, recovery of bismuth is much more efficient in the second extraction where there is a larger excess of dithizone and over 9997, recovery is still obtained in two extractions from pH 8 to 10 (Table 11). If the extraction is continued until two extracts having the bright green color of unchanged dithizone are always obtained with the 0.006% reagent, or one green extract with the 0.03% reagent, complete recovery of both lead and bismuth is ensured. Lead dithizonate is easily separated from bismuth dithizonate by stripping VOL. 37, NO. 13, DECEMBER 1965
1669
the former into an aqueous solution at p H 3.4 when approximately 0.005% dithizone and equal volumes of aqueous and organic phases are employed as in the usual spectrophotometric determination of lead. However, when higher concentrations or larger volumes of dithizone are used to accommodate larger quantities of lead and bismuth for radiochemical separations, lead is stripped incompletely unless the acidity is also increased. Obviously, the converse is equally true. If the dithizone concentration is reduced, a n increasing quantity of bismuth will accompany the lead unless the acidity is also reduced. This fact probably explains Jaworowski’s inability to keep part of the bismuth from following the lead, since he used the equivalent of approximately 0.003% dithizone to permit simultaneous determination of stable lead as well as lead-210. As shown by the data of Table 11, a n equal volume of a p H 2.7 buffer will strip approximately 98% of the lead from at least 30 ml. of 0.03% dithizone without losing more than about 0.8% of the bismuth per strip from as little as 30 ml. of 0.006% dithizone. Even this small quantity of bismuth is easily recovered by extracting the strip solution with a small portion of fresh dithizone. The higher concentration of dithizone will easily accommodate 2 mg. of lead, which is more than most samples to be analyzed will contain. I n contrast, an equal volume of pH 3.4 buffer will strip only 65 to 70% of the lead from the larger quantity of dithizone. Consequently, about 10% of the total lead will be left in the bismuth fraction after two consecutive strips when a pH 3.4 buffer is used with the larger quantity of dithizone. If the bismuth fraction is counted after 18 hours, the results will be too high by only 1%, because of correction for slightly more decay than actually occurred. On the other hand, results from the lead fraction will be low by the full 10%. Diethyldithiocarbamate Extraction. Although not very selective, extraction with D D T C is a particularly useful separation for application t o inorganic systems, since both lead and bismuth can be extracted very efficiently from strongly acid solutions and separated from large quantities of iron, aluminum, calcium, magnesium, phosphates, etc., that are frequently encountered. For example, liquid effluents from mills processing uranium ores frequently contain such high concentrations of metallic salts that the solution cannot be made ammoniacal for dithizone extraction when large samples are used. Even if sufficient citrate could be used to prevent precipitation of metal phosphates or hydroxides, a copious precipitate of 1670
ANALYTICAL CHEMISTRY
manganese ferrocyanide forms frequently on addition of hydroxylamine to the ammoniacal citrate-cyanide solution that makes any extraction impossible. Extraction with DDTC is undoubtedly the simplest and most convenient method for separating lead and bismuth from samples containing high concentrations of calcium phosphates (2, 6) such as bone, urine, feces, and soil. Extraction from acid solution also avoids the serious error that results from even small unsuspected quantities of silica during dithizone extraction from alkaline solution. Lead is extracted to better than 99% from 100 ml. of 1M hydrochloric acid by a single 10-ml. portion of 1% DDTC in chloroform. At higher concentrations of hydrochloric acid, extraction of lead drops sharply until less than 5% is removed from 3M acid. The reduced extractability is undoubtedly due to formation of a competing chloride complex rather than a n acid effect, because recovery is again better than 99% in 3M perchloric acid. I n contrast, bismuth is extracted in excess of 99% from a t least 6.11 solutions of either hydrochloric or perchloric acid. Since hydrochloric acid dissolves both lead sulfate and bismuth phosphate readily, a 1M solution is used for extraction. The extraction is not affected significantly by moderate concentration of sulfates or phosphates. However, with high concentrations of phosphates, as with large samples of bone ash, the hydrochloric acid concentration must be kept small to prevent serious loss of lead. Extraction with DDTC is much more efficient than other systems applicable to acid solutions, and volumes of solution as large as 1 liter can be extracted directly with small volumes of extractant. Direct extraction not only saves the time that would otherwise be required to evaporate large volumes of solution but avoids other associated problems as well. Most tap and river waters contain relatively large quantities of calcium and some contain surprisingly high concentrations of soluble silicate. Evaporation of such solutions generally leads to precipitation of calcium sulfate, calcium silicate, or silicic acid which must be filtered off and retreated, particularly when large samples are employed. In many cases, both calcium phosphate and calcium silicate are copiously precipitated when the p H is raised for dithizone extraction despite a high concentration of citrate. Many investigators recommend the addition of an aqueous solution of the sodium salt of diethyldithiocarbamic acid to the acidic sample followed by extraction of the precipitates produced into chloroform. Under these conditions, iron will extract virtually completely even from the reduced state
in the presence of excess reducing agent apparently because of the rapid air oxidation of ferrous iron that occurs in the presence of high concentrations of diethyldithiocarbamic acid. If the extraction is made with a solution of the diethylammonium salt in thloroform, very little iron is extracted in the presence of reducing agents because of the very low concentration of diethyldithiocarbamic acid in the aqueous phase and the short period of contact with that in the chloroform phase in the presence of air. Copper increases the extractability of iron slightly, probably because of its catalytic effect on the air oxidation of ferrous iron. Ascorbic acid as a reducing agent for iron in acid solution is markedly superior to the sulfite-iodide combination generally recommended (2, 6, 8). Although cobalt, nickel, quinquevalent vanadium, and several other elements are coextracted with lead and bismuth from strong acid solution, only copper has been encountered in sufficient quantity to be a problem. If a 5% solution of DDTC in chloroform is used, all the copper from even 500-ml. samples of mill effluents can usually be extracted with less than 50 ml. of reagent. Since copper is not extracted by dithizone in the presence of cyanide, no further problem is encountered. However, in the presence of excess copper, a precipitate only slightly soluble in chloroform forms in the first extraction and interferes seriously with phase separation. If excess DDTC is added before drawing off the previous extract, the precipitate redissolves rapidly and subsequent phase separation and extraction are perfectly normal. Probably, the initial precipitate formed in the presence of excess copper is a less soluble 1 to 1 complex, while a 1 to 2 copper-DDTC complex easily soluble in chloroform is formed in the presence of excess DDTC. Surprisingly, DDTC cannot be destroyed completely by any combination of nitric, sulfuric, and perchloric acids. However, the extract must not be evaporated to dryness with perchloric acid alone or the organic residue will ignite in a sudden violent burst of white-hot fire and most of the lead will be volatilized. If DDTC is evaporated to fumes with concentrated nitric and perchloric acids and water is added, a clear and colorless solution is obtained that becomes turbid and develops a strong odor of hydrogen sulfide on standing. The organic matter remaining still has a definite ability to form strong chloroform-soluble complexes with lead and bismuth from acid solution that will prevent complete stripping of lead dithizonate in the subsequent dithizone extraction. After oxidation with nitric acid, the residual organic matter is not charred signifi-
cantly even by boiling concentrated sulfuric acid but is easily charred thermally on heating to dryness. The resulting char can then be oxidized completely and safely with a mixture of nitric and perchloric acids. Sodium acid sulfate is added as an acid buffer to prevent hydrolysis and thermal decomposition of lead and bismuth sulfates during heating to dryness and to prevent precipitation of very insoluble anhydrous sulfates of iron and other metals that might be extracted. Reduction and Volatilization of Lead. While checking the applicability of the potassium fluoridepyrosulfate fusion in platinum (10) to the direct decomposition of solid samples prior to the determination of lead, several per cent of the lead-212 tracer being used was always unaccounted for, presumably lost by volatilization. In addition, the platinum dish itself always contained several per cent of the tracer after the fluoride fusion, only part of which could be removed in the subsequent pyrosulfate fusion. The remainder could not be removed by any
means. Apparently, some of the lead is reduced to the metal in the slightly alkaline fluoride fusion by organic matter in the sample or by reducing gases in the burner used to make the fusion, some of which is then volatilized a t the high temperature employed and some is permanently alloyed with the platinum dish. This residual 10.6-hour lead-212 tracer decays off in a few days and the dish can be put back into service but it is obvious that any contamination of expensive platinum dishes by 22year lead-210 is intolerable. Therefore, samples should be wet-ashed vigorously in glassware to oxidize all organic matter completely and to leach out as much of the lead activity as possible before fusing the residue in platinum, particularly when relatively radioactive samples such as ores, mill tailings, etc., are being analyzed. The preliminary acid treatment has the additional distinct advantage of leaching out other easily reducible oxides and phosphates, both of which are particularly corrosive to platinum ware.
LITERATURE CITED
(1) Adams, J. A. S., .Lewder, W. M.,
“The Natural Radiation Environment,” University of Chicago Press, Chicago,
1964. (2) Analytical Methods Committee, Analyst 84, 127 (1959). (3) Aronson, A. L., Hammond, P. B., Nucleonics 22, No. 2, 90 (1964). (4) Bambach, K., Burkey, R. E., IND. ENG.Cnm.. ANAL.ED. 14. 904 (1942).
(5) Code of Federal Regulations,‘ Titie 10, Part 20, Federal Register, Nov. 17, 1win
(6)-Gibson, W. M., “The Radiochemistry of Lead,” National Academy of Sciences, Nuclear Science Series, NAS-NS 3040 (1961). (7) Holtzman, R. B., Health Phys. 9, 385 (19fIR\. \----,.
(8) Jaworowski, Z., Nukleonika 8, No. 5, 333 (1963). (9) Maynes, A. D., McBryde, W. A. E., ANAL.CHEM.29, 1259 (1957). (10) Sill, C. W., Ibid., 33, 1684 (1961). (11) Sill, C. W., Willis, C. P., Ibid., 36, 622 (1964). (12) Ibid., 37, 1176 (1965). (13) Sill, C. W., Willis, C. P., Flygare, J. K., Jr., Ibid., 33, 1671 (1961). (14) Talvitie, N. A., Garcia, W. J., Ibid., 37, 851 (1965). RECEIVEDfor review July 19, 1965. Accepted October 7, 1965.
An X-Ray Fluorescence Method for the Determi nutoi n of Ca Icium, Potassiu m, ChI ori ne, Sulfur, and Phosphorus in Biological Tissues GEORGE V. ALEXANDER laboratory of Nuclear Medicine and Radiation Biology, University o f California, 10s Angeles, Calif.
b An x-ray fluorescence method for the determination of calcium, potassium, chlorine, sulfur, and phosphorus in dried biological tissues, fluids, and organic extracts from tissues has been developed. The procedure has been developed and substantiated by the use of synthetic standards representing the extremes of concentration found in biological systems. Beyond the determination of a sensitivity constant for each element the method requires only the application of an absorption correction. This correction is determined in part by measuring the attenuation of Ti Ka x-rays by the sample and in part by the determined concentrations. Details of the method, including examples, are presented.
T
GENERAL LACK of suitable procedures for determining sulfur in a broad range of organic-based materials such as is found in biological tissues, fluids, and extracts prompted HE
the development of this x-ray fluorescence procedure. A number of x-ray procedures have been developed for sulfur but in each case have been applied to such a limited range of samples-e.g., gasoline ( 5 ) , oil (4), oil additives (a), or blood serum (7)-that the possible extension to other organic materials without resorting to an infinitely complicated set of standards has been doubtful. Sulfur, chlorine, and potassium are lost totally or in part during the thermal ashing of biological tissues at temperatures above 400’ C. This is also true for sulfur when tissues are ashed by low temperature-activated oxygen techniques. To avoid these losses it is necessary to carry out the analysis on fresh or dry tissue. The purpose of this paper is to present the details of an x-ray fluorescence procedure suitable for determining sulfur, calcium, potassium, chlorine, and phosphorus in biological tissues and organic components separated from these tissues.
90024 EXPERIMENTAL
Apparatus. The x-ray fluorescence equipment used for this work is commercially available from the North American Phillips Co. The x-ray tube is a tungsten target FA-60 which is mounted to irradiate the sample from above. The lead aperture in the conventional tube was replaced with a small trapezoidal mask which limits the irradiated area a t the sample surface to 0.5 X 0.5 inch. The x-ray supply voltage and current are regulated to within *0.25% and *0.10%, respectively. The spectrometer is equipped with a 2-inch, 0.125-inch spacing entrance soller slit, a flat ethylenediamine dextrotartrate (EDDT) crystal (2d = 8.808A.), a 4-inch1 0.007-inch spacing exit soller, and a P-10 gas flow proportional counter. The standard entrance soller slit is modified with apertures so that only the 0.5- X 0.5-inch sample area can be viewed by the spectrometer. The entire system is enclosed in a helium atmosphere mainVOL. 37, NO. 13, DECEMBER 1965
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