Document Moj2R2GMq3N093y7DqN1LwgYa

DETEHLlIMiLTIOH OF LEAD . - . Iho-dification of ,tiie b'ischer - Leopoldi Uetnod' u.itkin she Xas'c feu years ditkisone (diphenylthioeurb'azone). H-s round k-xteusive use as a .reagent in'determining minute daounrs of lead, particularly .in biological material.' lie f foliating4Uetnod bias', beor developed in our laboratory as a , :3u h ;oil obtaining -rapid as usll as .ad curate.; analyses for ': sr: l3att. ,0 or/nore, and, incorporates ideas from >ds .01 u-uscrler eopoldi (1), .'ilkins, fllloughby, -a . vw / ..inter et: -1. (5). eocodure ' ' - ;s. - lyrex plussnore;should be. used tnrougk- oguiob type', separatory funnels of capacity --ju .a . --a .p'uliX ,ced :w'i_- ,U.- adjustable '_\.LC . Joi -1'-J ' uO uo' reus of .,i:. s..co, placeo..directly, one 'k bbcr, vi ra llbX* .1 pkassaure -oust be' lead free ikon. put into Ukou. flat -nitric.facia of about o volume per cent is use a for rinsing, followed by die-rilled'viater. a : ; .loer'ck;l;s:iblueV labal/reagents ire- .'preferred:. fiigii grade ;of 'Pheridcal 'insures diiou reagent blank, dke special reagents re ..ireo. .,,re as rcalo on: . -- ' : a. - .. - g , i ob 1e-;di-suikied .ter Oredueeu ar nrnsteal fi y -rasseili 0.' f .< reagent ,aiai-onia. g . - f; f =wv irasselii. ;-0'.;kf.i-reagent- nitridfacid. go. 'gtp puff ..o id)-' Jrasaelli.; g.go, reagent i.gdrcckloric acid. :ip.I,fSpecial aeidusolution, lead, free,pcontaining- 48 o vo3ru.u .or cons of uLtric acid,; '4fvolume''-per' .gritko'i "&ydro.T " P caloric ,,cid m d one balance- as tauter. - - ,.,,,u - v- b- ,,ck uS uco, for Pits o o i w k o n Bustgbe; obtained by' file g |feaistiliation:bcffreagontsf (3} and '(4.) tnrougii a' silica still. .Uiie:..gv,ityr;;used, must' be:..reagent (1.) redistilled iron a rvrex still.f> ,f. :g'; g ' .. . . ' o k :; f: a : f; pd) Ucaiinn.hydroxide solution - 1 5 ,,per -cent by . ci,Pur. _ is solutibniis poade by dissolving 10 ibs-.kof kferck-'s' vreagent :.nadk,; {pbiiGtof orn}: in -13 .65-l i t e r s 1of rater. :'After .staiidgiiggboga/clearfisoin^ dram:' off ,b y .ibeans. of ,a u ... \ > - si., on. . -. o ' = f.; , 17) Girric acid solution,-. Dissolve 400 gratis' off f -citiuepkeidh-k.gP:;';\(g:tue'koonbnyarate o) in about -500 l.-of . ' ' .nistliied a::ator'v:-neutralise gltk aiuaonia to "pH' S.5-4.G usirigf '- U..bigger coat bronpkenol blue as an indicator, add a kittle : . ' copper salt, and-pa so inikgb. ;Let stand over nigiit and filter, m i l off every trace :ofpHab ana dilute to one liter 'uitii ' distilled rater, f ffis 'stock -'solution nay be diluted ten tines' ..'itk arstiiled 'Uater ucii tkat 1 ml. - 0.04 gra. of citric, acid. r.ad:;:a -Siiiaii' m o u n t pfb cnlorofoxn as a preservative. A ^ 0003SS4 a v: (S ) ; p e d a l aomonia for"neutralization. - Redistil .'. reagent (E }, passing.'the evolved gas into ice-cold radio- , m p tilled. r a t e r . : m-pP ' - . . ' ' , . (Cj Potassium cyanide solution - 10 per cent b/Y. : {10} Tar e k *s Reagent Chloroform. . p, . T (11) ' DIthizcne purified. To 50 nl. of chloroform ui.-m. vJ-- . _a.d--- of commercial ditiiizone. such as produced by.the...; mustunr kodak Company. 'hake.out rith three .consecutive. 100 ' ' ul.% portions of .oiie. volume par cent numonia using redistilled, load'Treed aiuionia. -l-cidify- the ammonia extracts with dilute uO.yoIune par cent redistilled',- lead free hydrochloric acid. - 'sdsdblva the precipitated-bithizone with chloroform using: m threevpr :;four 0 nl.-portions. Cash- t h e 'chloroform sntracts '''' tuo or three tines with redistilled v/ater. Rvaporate the chloroform d-ith gentle heat and remove the last traces at COCO, .fa vacuo. ; . ' (Id) Lithizoiie solution no. 1. This solution ' contains--;hS up. of commercial dithizone (approniuately do p puiPity] pep liter ofmchlorofo.ru. tore in croon glass. h JOu u _l0 . v.no _,__i.. ^ mj^1 l Xu --._U x'Q.* -- . :m(15}. D ithizohe solution ho. 'Sw- This solution con-' s T T s -TO)ipms.h ofPreagent )(11j -per Titer, of chloroform.`'m-Ttore T n .hrovumflass bcttlemundor; a' layer -of udner cent: sulphur cus p acid.mmuse aTv = approximately'PuO gamma lb. ' ' ' d. ' \ P c l T T p a c i a l Pi'uiimmne Tnfraction solution.m'- Place lupa: ImPgo .separatory, funnel', .one. liter of aU'-auieoUspoolution J . ' O V m - x .. om .m:-jmv.e-- if ^m j a-aoi hO --L u-X rux.mmlvC \S j. , .aid'ID :_T. of '.reagent (Id) \.ad shake vigorously for several mThut es-.releasing pressure .formed at. the start. If the . - --chlerpfdrip:,layer;jin- ue.Tmuid'the; mater layer colorless',. 'draws off . o c Ij i c .Oj.;. l u or _u ce~eat. the 'b^er&Mbnmdntii the;; culoroforu- layer'.is1--eishor. colorless or) slightly green..;. h o :- _c m . . . . . , usm.... c.e msaaeu snail as necessary ms mms-ove . -xa.1 -of. lead., une, Viator .layer should have cnly-u sligkt- H M 'O O " H & m-a . . 0 i_. a Suii'. C j j --,.. ;.......S U ~ .-v v S ; ....... u Si. g x 1C pc*~~ i-i. u X O iij --i.. a o ,,. S;....I a ecbuefcolOrlbss^vtfter-.Ttasidingpseveral hours in'she .sun- . ight* sv,fhe solution ;ism then rea d y for use. ' p . . ; m 1; (:lo} -Ttanaprd lead- solution. - Dissolve'0.15S8. gram f lead nitrate' crystals i n .p u o volume per centnitric acid o ethat,-pho final-yolune .is", one liter. Dilute'this''stock .. . elusion con :T,es ,;ith. vone per cent nitric'acid sucir. that - zTL - lo ^m u .To. The cue cr cent --'nitric acid used for -his o d u t i c n .mist C v U c T n rev.istilled: nitric acid and P rec.iatilled '..mtor. ;. - m .: ! (15 ) Tnt.icators. '0.04) per cent -brouphenol'blue,,ph .) C .oi or cent phenol-',fed maid 0.04 pier cent met a ere sol- " V purple,...prnen nmis up.-.Tccording -to Clark and Luos. Tetliyl orange, .avpieous ,miade up as a saturated solution. ' - - `(IV) '-/c;;.g-er solution, axssolve I grams of electro lytic cophor'. in minimum LuGo-and ualce-vvo tc 500 nl. - u-! P h 0003385 transfer the.complete sample to a beaker of suitable ' Pise. 'hull nitric'acid in the proportion of 100 mi./of acid, per -iter of urine. Fh:'h evaporates bn- the-.hot 'plate to a low.,volume , suck that: pransfeh nap 'bevnuade gto a 50G ml.1glass dish, rinsing the' beaker with a shall unaount, of, hot rater. ihmporate to ' aryness on the hot-plate, char -gradually at. the'mouth of an open electric muffle -furnace and finally ignite in a closed ruffle 'It -450--5.00-R i. to a white ash. Benove -'from the muffle and. alio;; to cool. gwfhisablve ash by treatment, with 50- volume .per cent-nitric .acihufsihg' for final solution 15 ml. of nitric" acid par liter of original-urine1. - -Stir '-with -heat'u n t i l .a l l 'soluble salts ':h._LO-l.1/Ci ^ g: h i filter into" a 500;mi. beaker, through a 12.5 .cm. .ihatiiian ' ho. 10 filter, p a p e r r i n s i n g the .glass dish and filter paper,- with .co ..no;. hi-lute--- the' filtrate to- approximately 400 w a i f : _w.- ..3 !-tr,,i; ,:.e ,iw,, sodium hydronide-:-solution to -p H :'B.d-a.o- asing ;a ten drops of aqueous methyl orange as indicator.... should--oho end point'be overstepped, a lev; drops of 10.-volumeg --nr ,,ujciv. 1c* 1c - id n a y o h added'; to the faintest: -pink : eou^rdolcn. . d ;p.-- i ,,o .it-- - UJ-- -,,if ,,lion no .stand over n i a h t - F i l t e r - on a -' . '-~S nnbnpbh .dphatiiah'h a . d-0-filler "paper, washing the', pro cipit at e ' _ ,,u fr jo.1 ___ .u ,,pa-ro.-mi sulphide- .waterVcbntaining:v;0..1.;; :pbrweent'wo'f its:-volume 'of'-hydrochloric acid,' aiscard ;the V; filtrate, 'and dissolve' t h e ..precipitated.sulpli-idest-from.the:'--. h i Iter paper buck no'the original: beaker using hot; {lead' free) .special ucid solution, '-followed by an.suing with' hot water.. :-,,lfeihateV:this process, until- the sulphides-are completely : hxssoiyed.' .:Concentrate-, the ,final- solution on '-the hot, plate to .about -hOggl,. thereby..removing traces' of ;hydrogen, sulphide- and - dissolving free onl ;hur. :should a clear solution not '.be ; obt-ii-ca lu is best to repeat -the- gassing, operation. bknnsfer g o u 50 ;._i. _lass stoppered -cylinder, .-.and-.ailutev .--O1u U -- x a G- jj O -^o.-.- -ah. . Faeces a '"hf 'brassier, the -saiiple-to a weighed 50-0 ni.; vitreoo.il dish, rinsing .'the. container'with.-diot water, svaporate to dryixoss on yno hoc place. bweigh and, rcord 'tWsdry weight hdien constancy i-sao-otuined. . f w w --. ; Place: in jthselectric raffle a t 4 D G O - 5 0 0 oov3rnight ana pxheu cool neigh again , recording the' ash weight p :;f - , ' -heat -tliepash aitlr 10 rxl. of 50 volume per cent nitric acid and evaporate to,: dryness, on the hot plate. Place-in a l e y a f f l e mid ignite -'to 'd white ash. a uenove and -men cool treat the ash v/Ith 50 volume' per cent nitric acid using . , ,'tis.r enough to dissolve-soluble :salts. the ash- iron -faeces ; verge of ten .contains none carbon uiiiciris very difficult to --* ve Uu':.e p a oioxl.: ..-v."- a- . . - - filter into;a yOQfixl. boaaer using the same- procedure : - ,,s:.pith urine. _-ot/ever, due to;-the presence of large "p amounts of -triCalciuni' phosphate present -in solution, the sample must .be. neutralized very carefully to prevent she.precipitation of abnormal aricants of this salt prior to passing. y o dium hydro:-;!de rust be added slcviypxith rapid' sxirring u n t i l 'the opalescence persists, the pH is sou 5.4- ;a avie .-sor barely acid to reliiyl orange - .'Udo evif 1 . -;aS: ti-tll -IvO,.,,^ain neutrelite the final filtrate, after:' pilution,:,Pith soinrs'hyarouide solution, this tine, using i o,.a.,j. e -^.iu.xCuivCl ..aa a,-,a-.a a n u^i.d oO ja-3 final filtrate tc 5 r:l. in a-glass stoppered cylinder. . ': i v.\'U,.-\, 0-, 'issuGS.y .-.aa- ,,.-a q . ` - .. f . . f. of - If : (1) Large- lie sues; . .- _ _ r o _ : r ^ a "i^sue a _ y ,, f u r cf ,.ib_bie sine, ynyblhgauhe ;cGhtainer;ys'iasaal nith -uater. treat 'uithgaif pf;. ." u c '- -: 3" - --..-;g- s1 a .o "-or -cent nitric .acid..,.- ffa.llovr an -u nie/brdpeh; -oeaner -uve un ivjil- -c a a l __u a s i g >ia a X 1/ o t a r ; . l a t e v. a t 1 1 ie ,.- 1 i s s u e 1.".i. kO' p a r t i a i l : t sax r\ rH fd-p..OC: p h i : : f e e y " - m a y s . ; "to--". a - 5 0 C :i l l .f v i t r e o s vu i o US ; o : fLii, ,:,o n e x x u f f i e a t , 4 5 0 ^ - 5 0 0 G O OciJ- 0ingf'thd' t i o n . y.'l; 1in.. ' fu'C ii o r .ya-O b u f f i ` 1i i . t h ' uXiO . .c -a - .0 a t e " i : o ..." '. \ s a. <J. Z J _ O O - n o a i a i n i n g i-_ t n e 1' Oa ir s .... .... a..'VO t h e v i i i X`0 O-s i l - d i . 0 li c d h t . a i n i : jxiQ ..o.otasnea rxa - - - U'JOU a i n .cautiously iunir a --Lxi e . -^,3 :'U.3 >i ^ '.J J '.3b, v- -- 0Si-jb,XXS X----0OS S_. XSSpOS aid aaul._ be treated u h a _ suitable: .auouht.'of p O ; voluixo:;: ;parsceht:shitric,;;h U p o mevahoiatidh to dryness. final - xitiori- ixx:;the';:suffie,' rhe ash obtained- is- ''generally'unite, ;coi iiihg crily traces of carbon. . ... i .-Ju.u of. laJ a n a l ash: nith 50 volume y ceiru uirric -.Cib ubSsji..n--g - f,;u:_Ss \j OxlC h to aissolve all soluble;, salts .f hiltr,. ureutraiine andigas, folioeiug .the-, sane procedure 887 -o- aa for faeces if;the '.sample contains large 'amounts of ;y calciimi Ond; phosphtrous, otherwise follow the sane procedure as used for ; u r i h e . .: :; : ,. : ' f :: . . - ~ ' -/ H..:all Tissues. . , ' y y /. - , f Transfer the tissue to a, clean vitreosll dish;, o f ' . a Ire...rinsing the container usual wit La- hot - ... .1 hdd nitric :nci:'redistilled'-from a vitreosii still, j.hnro:cinately: 1 'hi. per'grauiof tissue.; Heat on the hot'relite..;'until dlyostlbn Is partially comolate and encess Place in mouth of open electric muffle. furnace until ' !s .complete then ignite .in the closed uuffls at m b ' '-5 00-O.-; Hemcve:, from furnace' ana alien cool moisten the .ash with a pop. reai atilled,; nitric acid." .mpuin.cautioi m.,0UCKlluuii-lL' s-Uj `hnanauiile and- rei unite '"to .ii.rue uni. my-urcaj as'nomi i,ha-a: v e f r e a l. . w .c. j. j.e , a j . l o r i e ,:c r d h r e a l s t i l l e d : x'r ;_* ___lKO.Lt KJ "...i n g r a s s a o l e t o m l . .1 a s s st o w n e r e d cy i t o ' t h e ; w.,;-,-,l-a- ,h,i4io. tv-wfvf * iar v i m ' - U i -3 a.d aX.icu a.j. h o ad- hue U 0.131i..IK-Ui-id - * .m ;' W 3 sent )isfrom, ;d ;ca n r 1,L :O' 0.-1.lxO'O'uodd by hi l> C aiO .C.1 0ix3 'OKKiI'CIQ, n : cenar mprunea v w .j'.ith -hit his,on3'.:30 at ion" f.d 1. ,..-1 ;a:r.cOer ilo. vsiirlrP mitdhpie '..C ..`-J U ` '0a h e m 1 0 ;c 0 LfC-. 2l ' `' "i immure, ad...rangincb.bb twe en. py ancii100^ m a m a suit;able'.' ..ulipuct nayne' itiooxeea. oy letnod error . ' m3 '.h_Ln.c'.O . Uy n_o d. tiy.wO"V"OQ. U ilean-' government Otto and y s 'transferred to. the ;separatory funnel, nun', a, f .r e a g e n t { 7 )yand. Z drops of pile110 1 Hed indicator. .3 - d rodist iilsd; arfloniai drop, by drop, swirling '-the -funnel-. for, ii i h n ,funtil 'dibtihet; yellow- coloration develops.0- i d fn;hfowi;'add y 'ml., olwraageht; \ 9Op.;follow by ..the-,.',audit ion of ionia,.; uswbef or.. until hi pod color;.develops,'indicating 'a oro: -v-.rTor rr . y o j _ .j - / *. ;xion uCa oj, . iithizb'ne 3 olu'cion ho.l.e.-ihe stoppered funnel is ohahen vigorously, rbloaoiny 'Che ;yressure .devoloped. at the start.' Depending;. '' upon, the- amount ,ofe ieadyyresent, the color of the chloroform olidbe yyiil n m : s ; c..r.d.a.,.b'rilliant red-orange', with appreciable .ts of.,lead,ytlireugh' purple vo 'the unchanged green color ox y h i t h i r o n e .solution. Hntrction of. the aoueous solution /<ff f)003S88 -6- is repeated with successive 5 ml * portions of the dithizone solution, delivering each after extraction, to another separatory funnel, until the color of a newly added portion of dithizone remains unchanged. The aqueous phase is extracted once more, as a precautionary measure, and is then discarded. The total volume of dithizone solution used is noted as it gives a preliminary indication of the amount of lead present, and is therefore of value as a guide in the final lead estimation. - :V In special cases such, as the analysis of small tissues which does not entail a preliminary separation of lead as the sulphide, the chloroform solution of the lead-dithizone complex is first washed with 25 ml. of water containing one drop of reagent (9), quantita tively separated, and transferred to another separatory funnel.;! Contamination of either phase by the other.is avoided by running out the chloroform layer until about 0.2 ml. remains. Small portions of chloroform are successively added and withdrawn, maintaining a chloro form trap at all times-, until the separation of all ; ehlorofoi'm-soluble components is complete. The lead is removed from the organic complex by ;isbaking the clllerofqrm solution with 20 ml. of one, 'volume per cent nitric acid; the dithizone: remains in ehe chloroform phase, whereas the lead, in the form of -the;nitrate,hpasses:into the, aqueous phase. , w .v After separations of the two, phases, the aqueous ::h.v phase is washed repeatedly with small portions of chloroform until a freshly added portion remains color less. Draw <off the .:chloroform phase -conpletely('dis- , ; carding, and adjust the aqueous phase to pH;7 * 5 A adding .special, ammonium, hydroxide drop by drop and using -one: drop of Phenol Hed as an indicator. The sample is now jeady for the final analysis. Pinal Analysis for L e a d .,: A , '. A: standard is .prepared .by ^introducing into a separatory funnel, 3 ml. of standard lead solution, reagent (13), and adding 17 ml. of one volume per cent nitric acid. Adjust to pH 7*5 in the usual manner. Now placed in the supporting rack are the sample and the standard. To each, add 2 ml. of reagent (9). Both solutions are now distinctly alkaline. K F 0(j3cbO Add dithizone solution No. 2 from a burette in 1 ml. portions, shaking the-.separatory funnel after each addition, and allowing the chloroform phase to separate until a slight purplish tinge is produced. At this point dilute the chloroform phase to 20 ml. by the addition of chloro form, shake well and allow a final separation.' Withdraw . the chloroform phase into another separators^, funnel contain ing 20 ml. of reagent (1*1-} . Shake well and repeat this operation four times, until the red chloroform soluble dlthizoneAcomplexrremains. . ' '' A '. .' Treat with 10 ml. of 10 volume per cent hydrochloric acid and shake to liberate the lead in the aqueous phase as lead, chloride, leaving the dithizone as chloroform ' soluble -to be measured by colorimetry. The operation just mentioned must be carried out in the dark room. A Duboscq colorimeter is used, equipped; with a suitable blue glass filter and electrical; illumination. Remove moisture f r o m A the stem of the separatory funnel by means of a cotton swab..'--Draw--off the bluish-green dithizone solution into a . colorimeter cup. This applies to the standard as well as r the unknown. Set the unknown at 31 and use the standard : for making readings. Each division corresponds to one :gamma of lead. ,A . __ Removal of Bismuth Interference. . -mm '-Bismuth, vif ^present;, seriously interferes, inasmuch A p-,; as it reacts withr^ the -dithizone' reagent under-the- 'same fpAfv ;A conditions as specified for lead* This-- elements may. be-- -.v. present as a result of previous; .medi;oatio:h.;P;\ -Ap.;A `'A'^AvA;'^ Therefore it is imperative, to test,' for "theApresMicel A/ :.;: o f bismuth. To; the nitric acid, extract containing lead ; and bismuth (if present) as the corresponding nitrates is vadded 'two' drops of met a eresol purple. The solution is ' Adjusted to pH 2*0'vby the-addition of special ammonia A diluted to 10 volume per cent. Five'ml. of dithizone: solution No. 1 is added. If upon shaking the stoppered separator^ .there is- no change in color, bismuth is not present. The-chloroform dithizone containing phase is v. ., removed in the manner as described in the initial separation of lead and the final analysis for lead is continued after readjusting to pH 7 5 , A 7 A.: A -;.AA If bismuth, is present in large amounts the concentration of dithizone solution No.l may be increased, to facilitate separation. However, the solution must not be so concen trated as to mask the color changes during extraction. A solution containing 0.20 gram of dithizone per liter will : -- be found suitable. This method of separation has been described by Willoughby et al {^}. Discussion . , ..' Assuming Beer's Law to hold, a separate blank deter mination was made consisting of all reagents used in the final determination of lead, that is, following the initial separation. This blank was found to be 1 gamma. Therefore the colorimeter must be set at 31 instead of 30 Like wise a blank run upon all reagents used in the analysis, including the initial separation and final determination showed an analysis of 3 gamma. ,, : Therefore in calculating the final analysis it is necessar;- to subtract 3 gamma from the colorimeter reading obtained, multiply this result by the factor of aliquotion and, subtract a reagent blank for the amount of nitric acid ,and;-'S.odi'uiii-`;hydro2:i'de5splution used in preparation of the, sample. Nitric acid may be analyzed spectrographically. The analysis of nitric acid.is expressed as gamma Pb per liter and the analysis of sodium hydroxide as gamma Pb per 100 ml. The amount of Pb contained in all other reagents used for preparation is so small that it can be neglected, considering the sensitivity of this method of analysis. . Constant care must be exercised throughout the entire procedure to avoid lead contamination. Separatory funnel - stopcocks may be lubricated with pure white vaseline. /Glass stoppers may be .anchored "to t h e .tops 'of the separatory funnels with, stopper ties made of rubber,,which/ permit of ;feqs; removal ofAt hd bstdppers.,,. but" do not' al1ow them to hang in contact with the sides of the funnels. Dithizone . Solution No.l and .2 as 'well as- chloroform m a y b e .drawn1 from - graduated dispensing burettes, the solutions themselves being stored in brown glass bottles. Pyrex glass is not necessary for these particular containers. No lubrication of stopcocks is necessary in this instance, as the liquid furnishes its own lubrication* ' A dark room has been found necessary for the final colorimetric reading, since day light, particularly when the sun is shining, often causes the dithizone- to partially oxidize. Even a trace of the yellow oxidized dithizone leads to imperfect color matching, as well as incorrect colorimeter readings. 7/hen using a dark room it is possible to make eleven consecutive analyses using one standard. K ^ ooossdi Literature Cited (1) Fischer, H . , and Leopold!, G., Wiss. Veroffenlich, Siemens - Konzern, 12, Heft 1, (19335 (2) Wilkins, E. 3., Fr., Willoughby, C. E., Kraemer, E. 0., and Smith, F. L., 2nd, J,Ind. 'Eng. Chem., Anal. Ed., 33 (1935) (3) Winter, 0. B., Robinson, Helen M . , Lamb, Francis W . , and Miller, E. F., Ind. Eng. Chem., Anal. Ed., 7, 265 (1935) (M-) Willoughby, C. E., Wilkins, E. S ., -Jr., and Kraemer, E, 0., Ind. Eng. Chem., Anal. Ed. 7, 285 (1935)