Analyfaca Chunrca Ada Clsevier Putlish~ng Co:npany, Alnstcrdaln Print4 in I’hc h’cthrrlenti

175

THE NEPHELOMETRIC DETERMINATION OF SMALL AMOUNTS OF SULI’HATE ION

J hl MARTIN AND W I S1 EPI-EN

Dcfxulnrort of Clreuzlstry, Tlrc Ujtlvrrsaly, I’ 0 Uor 363, Btrnrtqlrarn (~?Inglrcrrd)

(liccclvcd Alarch rqth, 1967)

The accurate determmation of small amounts of sulphatc is a matter of con- siderable importance m many branches of pure and applied chemistry and in many biochemical studies Direct calorimetric procedures arc few and not entirely satis- factory, reduction to hydrogen sulphidc and its conversion to mcthylene blue is a sensitive process, but the many papers pubhshed on it testify to its basic unrehabihty. Methods based on tllc precipitation of barium sulphate (e g. from bat-mm chloramlatc) are subject to the many mtcrfcrcnces associated with this precipitation Nevertheless, the barium ion remams the most widely used means for the determmation of the sul- phate ion, and many rngcnious methods have been proposed, particularly for titrrme- tric and calorimetric purposes

The rather low solublhty of barium sulphate, particularly m mixed solvent systems of water and polyhydric alcohols has an obvious apphcation m turbidimctry and nephelometry, and numerous such procedures are to be found in the literature. The basic method of DENIS A&D REEDI which deals with I-IO p p m of sulphate ion m a 25-ml sample has been modified by later workers; the method of TOENNIE~ AND

BAKAYP is the most recent and thorough cxammation of this process. Up to the present, little or no attention has been given to the rcplaccment of

barium by other prcclpltants for the sulphate ion m thcsc nephclomctrrc procedures. BenLidme 1s unfortunately less sensitive a precipitant for the sulphate Ion than barium chloride, and substltutcd benzidmes are, m general, even less sensitive, so that no advantage can accrue from the apphcation of these reagents m turbidimetrrc or nephelometric procedures However, the same cannot be said for one other precipitant for the sulphate ion, 4-ammo-4’-chlorodiphenyl (trivrally named CAD) which has found considerable apphcatron as a reagent for small quantities of sulphate by titrl- metric” and spectrophotornetrlc 4 methods BELCHER AND STEPHENS have indicated the superior sensitivity of this reagent to barium chloride, and state that it can detect by a precrpitation reaction as little as o I p p.m. of sulphate ion m a volume of 50 ml. This sensitivity IS about IO times as great as that of the basic nephelomctric method with barium chloride. The precipitate of CAD sulphate 1s extremely fmely drvlded, and suspensions of this salt should be Ideal for turbidrmetric and nephelometric ap- plications. HASLAM, HAMILTON ANU SQUIRRELL~~ m an account of the detection of certain clemcnts m plastic materials by the flask combustion method, recommend a turbidimetric CAD hydrochloride method as an alternative to the standard barium chloride method for the detection and semiquantitative determination of sulphur m the test material The suspension of CAD sulphate 1s stabihsed by the presence of gum ghatti and peptone in the reagent solutions, and the optical density of the sus-

Anal. Clram, Acta, 39 (x967) 175-180

pension 15 measured at 700 nm after 0 5 h Only scmiquantltat~vc applications arc

pr(qXJ5ed for NTIOUnt~ Of $Ul]>llUr (Jf tile Or&r I--2’%, in the test material ‘I’hesc! wrkers

d0, hcJWe\Tr, emphaslse the sensitivity of tllc CAD hydrocl~lor~dc reagent and rcc-

ommcntl Its use kvhen only trace amounts of sulphur are to Ix dctccted

Nepliclometric proccdurcs arc gcncrally more scnsltlvc than Lomparable turbi-

dimctric procedures III wl~~cll tllc ccmccntration of a suspcnslon 15 mcasurccl by climi- JlutlcJn of Incid(*nt light In nel)lielomctty, tlic liglit scattcrccl by the particles of the

suspcn5ion 15 measured dircc,tly at an angle (usually r)o”) to tlic Incident beam Various type5 of nephcl’nnctcl arc’ av:ulablc c.ommcrc.i4ly in w11i~h almost all then scattcrccl

light from JU5t dhJVC cJ” t0 ()O” from tlic incitlc*nt beam 15 collcctetl and measured

Sucli instruments enable less stringent ctJnclitions to Ix pl~~cccl on requirements of

parti& size, which for p.qXJW~ of maximal scattering should be about o 2-o 4 1.4

Smaller particles scatter less 1ntcnsclv, but cvcnly in all clircctrons, larger particks ;il~J scatter less intensely IJut m,iinly within IO0 from tllc inc.ltlcnt beam Altllou~h the

Intensity of ltglit scclttercd at a given angle 15 dcp~~ndcnt cJn the slzc and shape of the

suspcnclccl particles, on tlic wavclcngtli of tlic inciclcnt light and on the ratio of tlic

refractive mcliccs of the particle and the medium in wliich it is suspended, ah well as

on Its concentration, particle 51%~ 1% the most significant variable in any analytical

pTJWdUW involving ncplielomctt ic mea5urcmcnt57, the rcproductble formation of

similarly si~ctl p4 ticlcs 15 the prcrcquisite for successful nqdiclomctr~c dctcrminations In tlic prcsscnt invcstigntion, conditions liavc been cstablislicd for tlic detcr-

mlnatlon of o 025-0 25 mg of sulphate in Io-ml volumes of test so~utloll (z c Conccn-

trxtions of 2 5-45 p 1) fn of 4ulplix.te) by ncldiclometr~c meaburc’nient of CAD sul-

phate suspen5ion5, stnbiliscd with gum ghatti

A study of the effect of other anionlc spcc ic\ on tlic dctcrrnination has shown

that cliloridc and nitrate ions prcscnt in concentrations 10 times th‘it of the maximal

4pllatc concuntlatlon ate without effect Fluorlclc and l~llosl~llatc ions arc knowr> to

lntcrfcrc wltli the rccommcndcd titrrmetric mr*thod for sulphate using CAD liydro-

chlorlclc and tlicsc Ions would bc expected to cau5c a sirxiil,Lr mtcrfercrice in tlic nc-

plielomctric mctliod A x0-fold conccntratlon of fluoride ion (2 e 2 5 mg F-/r0 ml) gives

9 curdy gelatinous prccipitatc with the CAD reagent, which dis,~ppears when the

concentration is reduced to about I 8 mg of fluoride However, in the presence of this

amount of fluoricle ion, the apparent sulplintc content 15 consiclcrably higher than that

actually pxcscnt, ancl only wlicn the fluorlclc concentration 15 reduced to 0 25 mg does

this lntcrfcrcncc tllsappcar With phosphate, the CAD rcqcnt forms prcclpltntes with

conccnttations below 0 25 nig of I-1#0.1- in ro-ml volumes and it 15 necessary to

remove phosphate ions from the test sample by trcatmcnt wit11 magnesia mixture

A proccdurc which clcals lrltll 1Jhosphntc m amounts up to 2 5 mg of H#O.I- in IO ml of test sample has been clcvclolxxl t(J ovcrcomc tllis mtcrfclencc No other anionic

spcc~cs has been cxarnlned as a posslblc mtcl fcrcncc, but it 15 known that bromide and

~ocl~2c ions, and the organic acid\, acctlc, cltllc and tartnrlc acids, arc without effect

on the precipitntlon of sulphntc ions by CAD llydrochloricle~ Catlomc mterfercnces

have not been cxammcd 111 the lxcscnt lnvcstlgntlon Of the common cations, only

alummlum 1% known to fotrn a sparingly soluble complex salt with the reagent m

neutral solutions cont,unmg sulphate ions, zinc, copper and mngncsnxn form soluble

complexes with the lcagcnt, but should not intcrfcrc when prcscnt m Io-fold amounts

If hgh concentrations of cations (other than the alkali metals) were present m the test

28nnl. Clrrrtr A tla. 39 (1967) I 75-180

SEPHELOMETRIC DETERhlIXATION OF s04*- 177

solution, these could be convenlcntly removed by an Ion-exchange resin, and the rcsult- mg acid test solution neutrahscd before apphcatlon of the ncphelometrlc procedure

EXPERIMESTAL

Nephelometrlc measurements were made with the EEL Nephelometer Head m conJunction with the EEL Cintgalvo Type 20 Test tubes (2” dlam , volume cn. 25 ml) provided with tile instrument were used to cont‘un the suspensions

CAI_l hydrochlonde solrrtlon (Sol&on A) DIWJ~W 2 o g of 4-ammo-4’-chlolo- cllphenyl hyclrochlorrcle~ III Yoo ml of hot (70”) o 05 N hydrochloric acid Cool the solution and flltcr through a Whatman No 40 paper If the solution 15 to be stored for prolonged periods, dark bottles should be used

CA I> lrydvoclrlortde-~c~lollc soluttorl (Sohtgon 13) D~ssolvc o 025 g of blologlcal reagent grade peptonc m 50 ml of solution A by warmIng to 7o” Cool and filter through 2 Whatman No 40 papers Prepare tins solution datl\

GIrlIt gltnttt sollltlntl rh~0ivc 0 5 g of finely grcjuncl q.nn ghatti in 200 ml of o 05 N l~yclrocl~lor~c acid by warming to 70~ Cool the solut ton and filter off undlssolvcd material on double Whatman No 40 papers Store the solutron at o-5” anti discard at the first signs of the form,Ltion of \mnll p,Miclc+ 111 tllc solution

Cornpovtc /wectfittatrott veugent (Solrrtlon C) MIX IO ml of solution 13 and 90 ml of solution A shortly before use

I~otaswirnt ertlfhitc solittzon (0 025 mg sulfiliatc per ml) Ihsolvc 0 0454 g of freshly clrrcd A I< potassium sulphatc m I htrc of water.

Mfig:r~~n mtdrrve I>is~olvc 20 g of A I< ammomum chloride and I0 g of A R. magnesnun chloride hcxahydrSltc In xoo ml of water

Proced ttve Neutral test solrttaom (jhoqjllatc nbsctrt) To x0 nil of the neutral test solution,

contamcd m a 5o-ml stoppered flask (volumctrlc flasks arc convenient, but not cssentral), add 12 ml of the composite reagent, solutron C After 15 see, stopper the flask, swirl the contents for a further 15 XX and Invert the flask once per second f4jr 30 WC Leave for 45 mm and then add with gentle swirling, 3 ml of gum ghattl solu- tton After a furtl.\er 15 mm, transfer the whole of the suspenston to one of the nephclo- meter tubes (Note), takmg care to avoid formatlon of au- bubbles m the \uspcnsmn, measure It. cffcctlvc llgllt scattcrmg power m the nephelomcter, previously adlusted to gvc maximal galvanomcter deflection (x00) with the most concentrated standard suspcnslon (o 25 mg of sulphatc per IO ml) and zero deflection on dlstllled water Prepare a cahbratlon graph for amounts of sulphate ion in the range o 025-0 25 mg per I0 ml of ctanclard solution and detcrmmc the sulphatc content of the test solutions from the standard graph A blank dctcrmmatron should be included m each series of cahbratlons

Note l’hc ncphelomcter tubes sl~oulcl he thoroughly cleaned perrodlcally in chromic-sulphurlc acid solution to mlnlmlsc the risk of air bubbles forming on the walls of tubes when the suspensions arc poured into them

178 J >I MARTIS, W. I. STEPHEN

Aczd tesl solzctzonc To IO ml of the test solution contained in a qj-ml beaker, add o 3 ml of ammonls solution (d o 88), and carefully cvaporatd the 5olutlon to a volume of 3-5 ml Transfer the solution to a ro-ml mcasurmg cylmdcr and rmse the beaker with small amounts of water to give a volume of 7 ml m the measurmgcyhndcr Transfer this volume of solution to the so-ml preclpltatlon vessel and rinse the beaker and tllc cylmdcr with z ml of water, made up to 3 ml m the cylmclcr , add this 3 ml to the 7 ml in the prcclpltatlon vessel. Prcclpltatc the sulphate as tlcscrlbcd above Treat the standards m the same way to compensate for the traces of sulphatc present m the additional reagents

Pitosjlratc +wnt To 15 ml of test solution, containing 2 s-25 p p m. of sulphatc, add o I ml of 4 N llydrochlorlc acid, o G ml of magnc\la mixture and o 3 ml of ammoma solution (cl o 88) MIX well ant1 store ovcrnlght at o” Ccntrlfugc the prc- clpltatcd magncslum ammomum phosphatc~ and plpettc* IO ml of the clear superna- tant l~lu~cl rnto a zs-ml beaker Remove cxccs~ of ammonia by bolhng and make up the volume to IO ml a~ descrlbcd under Aczd test colrltzom Prepare stand‘trds from IO-ml allcluots of sulphatc solution, treated with ldcntlcnl amounts of the phosphatc- precipitating x-cage&s, and adjust the volumes to IO ml as tlcscrlbed above Dctcrmmc the amount of sulphate m the test solutmn from the cahbratlon graph and multiply the result by the volume corrcctlon factor, I oG7

I~ESlJJ.TS AND 1)ISCUSSION

The analytical procedure 15 designed to deal with lo-ml test samples contaln- mg sulphate ion at conccntratlons in the range z 5 to 25 p p m It can be extended to slightly higher conccntrntlons (30 p.p m ) without impairment of accuracy The results in Table I arc typical, and show an avcragc error of +2 80/, Comparison of the new

‘lhJ3LI~ 1

RI’SULTS I3Y TIIE RI:COMMI~ NDISD I’l~OCI~IJURl:

(Volun~c of test sntnplc IO ml) _____ - -_-_------_- _-..___ ._- -_-

SlIlpllUtC Slllpl~tlk Crror Slrlplln~c Srllpllulc LWOY

fwcsrnl folr~lfl (‘%,) pYCSCNt fo1rm~ PXI

(P P “1 ) (P p “1 I CP P “1 I (P p “1 ) -_A ---- _--

30 0 30 5 3-17 15 0 15 5 i-33 24 0 24 5 i-21 I2 0 12 0 00

22 5 22 0 - 2.3 IO 0 98 - 20

Ia 0 IG a --G7 85 a0 - 5.9 16 5 165

+Z

50 49 - 20

15 8 16.2 --- -- --_

method with the standard barium sulphate method8 mcllcatcs that the CAD method 1s consldcrably more scnsltlvc and capable of dealing with a smaller total amount of sulphate in the test solution

No difficulty 1s cxpcrlenced in obtaining lmear cahbratlon graphs by the recommended procedure The lmc need not pass through the ongm, its intercept on the RXIS of the galvanometer rcaclmgs 15 gcncrally small (bctwecn z and 5%) and l\ obtamed from the measurcmcnt of the blank solutions The small variation m the

SCPHELOMETRIC DETERMISATIOS OF s042- 179

blank values depends largely on the age of the reagents, in particular, that of the gum ghattl solution The properties of this solution change slightly from day to day, de- spltc storage of the solution m a refrigerator, SO that no two standard graphs are exactly the same, unless constructed on the same day It is, therefore, cssentlal that tests and standards arc prepared as nearly as possible under identical condltlons, certainly with the same reagents, and on the same day Alternative forms of the gum ghattl solution, and mdccd, altcrnatlvc stablhsers such as gum acacia, gave rise to very inferior results. zind the procedure descrlbcd above was established only after consldcrablc variation of the conclltlons for the prcparatlon of stable suspensions

When sulpllatc-pliosphr~te solutions wcrc exammcd, the results indicated that a constant increment of 2 p 1) m of sulphatc was present in each sample Expen- ments with solutions contarnmg standard amounts of sulphate, and no phosphate, but with the addltton of the rcqulsltc amounts of rcagcnts necessary to remove the phosphate ion also gave consistently high results, Indicating that the reagents con- tained appreclablc amounts of sulphate ion a3 lmpurlty (Fig. I) It is, therefore, cs- sentlnl to treat standard samples m exactly the same way a~ the test solutions, under these conclltlons, It 15 posslblc to cletcrmme sulphatc m the prcscncc of up to 25 p p m.

of H2PO4- without any 1059 of prcci5ion

005 010 015 020 0 25 mg S0,2- in lOem sample

Fig I Typical callbratlon graphs (A) Samples from which o 25 mg HaPOd- was prccqdatctl; (U) standard sulphatc solutions

The new method should fmd applicntlon for the detcrmmatlo,n of traces of sulphate ion m water samples, or of sulphur m organic and blologlcal ma.terials after conversion to sulphate ion

Because of the known carcmogcnlclty of bcnaldme and 4-ammodlphcnyl, doubts have been cast on 4-ammo-4’-chlorodlphenyl as a possible carcmogen. Physlol- oglcal studies with rats have +own that this compound has no sgniflcant propertic as a carcinogen, compared with the parent amme and the 4-fluoro-derlvatlv@, Its use as an analytical reagent should not bc restrlcted on this account.

WC arc grateful to Professor R. BELCHER for 111s mterest in this work, Dr I;. L ROSE (1.C I Pharmaceuticals Division) for his comments on the carcmogcmclty of the reagent and the University of Birmingham for financial support to J .M.M

AIWZ Chrm Ada, 39 (1967) 175-180