AOAC Official Method 980
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A3A5550456124ECE007720FE38435C65 |
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0.13 |
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2 |
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日期： |
2016-2-28 |
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46.2.12,AOAC Of fi cial Method 980.24,Sulfanilic Acid, Schaeffer’s Salt,4,4￠-(Diazoamino)-dibenzene-sulfonic Acid,and 6,6￠-Oxybis(2-Naphthalene Sulfonic Acid),in FD&C Yel low No. 6,Liquid Chromatographic Method,First Ac tion 1980,Fi nal Ac tion 1981,A. Apparatus,(a) Liq uid chromatograph.—With gra di ent elu tion ca pa bil ity,and au to matic sam pler, e.g., DuPont 830 with Model 838,pro gram ma ble gra di ent and Model 834 au to matic sam pler with 5 mL,loop (or equiv a lent). Op er ating con di tions: eluant flow rate,0.6 mL/min; tem per a ture, am bi ent; gra di ent pro file, pro gram IX 70,Fig ure 980.24A, is scribed on pro gram charts pro vided with,Model 838 pro gram ma ble gra di ent and run at time set ting of 40 min;,equil i bra tion time, 10 min be tween each gra di ent run.,(b) Detectors.—(1) DuPont 254 nm de tec tor (low pres sure Hg,source) with at ten u a tion set at 0.08 AUFS (or equiv a lent). (2) DuPont,385 nm multiwave de tec tor (me dium pres sure Hg source and,325–385 nm fil ter) with at ten u a tion set at 0.04 AUFS (or equiv a lent).,(c) Col umn.—DuPont Zipax SAX col umn (Cat. No. 852950005),(or equiv a lent). Con di tion new col umn by heat ing 50 h at 50°C with,0.01M Na2B4O7 flow ing at 1000 psi and then let ting it rest un used,2 weeks.,(d) Spectrophotometer.—For use in vis i ble and UV range.,B. Re agents,(a) Eluants.—(1) Pri mary eluant.—0.01M Na2B4O7. (2),Sec ond ary eluant.—0.2M NaClO4 in 0.01M Na2B4O7.,( b ) St a n d a r d s o l u t i o n s .—(1) S u l f a n i l i c a c i d,(SA).—Ap prox i mately 0.25 mg/mL H2O. (2) Schaeffer’s salt,(SS).—Ap prox i mately 0.35 mg Na salt of 2-naphthol-6-sulfonic,acid/mL H2O. (3) 6,6￠-Oxybis(2-naphthalene sulfonic acid),(DONS) . — A p p r o x i m a t e l y 1 . 2 mg/mL H2 O . ( 4 ),4 , 4 ￠ - ( D i a z o a m i n o ) - d i b e n z e n e - s u l f o n i c acid,(DAADBSA).—Ap prox i mately 0.15 mg/mL 0.2% NaOH.,De ter mine ex act con cen tra tion of stan dard so lu tions (1)–(3) from,UV spec tra of di luted aliquots in H2O; that of stan dard so lu tion (4),from vis i ble spec tra of di luted aliquots in 1% NaOH. Ap prox i mate a,(mg/mL, 1 cm) at spec i fied nm are: (1) 81.6, 247; (2) 292, 231; (3),179, 238; and (4) 86.5, 410.,C. Suit abil ity Test,Pre pare test so lu tion con tain ing 1% FD&C Yel low No. 6 and,rel a tive to the FD&C Yel low No. 6, 0.2% SA, 0.3% SS, 0.1%,DAADBSA, and 0.5% DONS. Set pa ram e ters as in A(a) and run,blank gra di ent (no in jec tion) and test so lu tion. Com pare,chromatogram to that of Fig ure 980.24B. If sim i lar res o lu tion is not,at tained, ad just op er at ing pa ram e ters to those needed to re solve,com pounds.,D. Cal i bra tion,For each cal i brat ing so lu tion, dis solve 0.500 g FD&C Yel low,No. 6 (free of SA, SS, DAADBSA, and DONS) in ca 25 mL H2O,Figure 980.24A. Gra di ent pro file pro gram for,sep a rat ing in ter me di ates and re ac tion by-products from,FD&C Yel low No. 6 by LC.,Fig ure 980.24B. Chromatogram of FD&C Yel low No. 6,(peak 8). 1 = In jec tion; 2 = sulfanilic acid; 3 = gra di ent,front peak; 4 = Schaeffer’s salt; 5 = DAADBSA;,6 = un known; 7 = higher sulfonated sub sid iary;,9 = un known; 10 = DONS. Top trac ing from 254 nm,de tec tor; bot tom trac ing from 325–385 nm de tec tor.,2006 AOAC INTERNATIONAL,add 5 mL 0.01M Na2B4O7 and aliquots in di cated in Ta ble 980.24,and di lute to 50 mL with H2O.,Cal cu late con cen tra tions of com pounds be ing de ter mined as,weight % of FD&C Yel low No. 6. Cal cu late % con cen tra tions in,cal i bra tion so lu tions as:,C = V ′ C￠ ′ 0.2,where V = vol ume stan dard so lu tion taken; C￠ = con cen tra tion,stan dard so lu tion, B(b), de ter mined spec tro pho to met ri cally, in,mg/mL; and 0.2 = 100 (%)/500 (mg).,Mea sure ar eas of peaks from SA, SS, and DONS, us ing 254 nm,de tec tor, and of DAADBSA, us ing 360 nm de tec tor. Areas may be,ob tained from in te gra tor or by mul ti ply ing peak height by peak,width at half peak height. Plot % con cen tra tion of each com pound,against peak ar eas. Use cal i bra tion so lu tions 1–5 to de ter mine,re ten tion times of the 4 com pounds.,E. De ter mi na tion,Weigh 0.500 g test sample, dis solve in ca 25 mL H2O, add 5 mL,0.1M Na2B4O7, and di lute to 50 mL with H2O. Chromatograph test,solutions in ter spersed with cal i bra tion so lu tions. Com pare,chromatograms of test so lu tions with those of cal i bra tion,so lu tions. Iden tify by re ten tion times and mea sure ar eas of peaks,cor re spond ing to the 4 com pounds. Use cal i bra tion plots to,de ter mine % SA, SS, DAADBSA, and DONS.,Ref er ence: JAOAC 63, 61(1980).,CAS-135-76-2 (Schaeffer’s salt),CAS-121-57-3 (sulfanilic ……
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