S ID N E Y G O T T L IE B a n d P A U L B. M A R S H
Bureau of Plant Industry, Soils, and A g ricu ltu ral E ngineering, U . S. D ep artm ent of A g ric u ltu re , B eltsville, M d .
The co lor reaction of 4-am inoa ntipyrine with the te x tile m ildew preventive 2 ,2 '-m e th y le n e b is [4 -c h lo ro p h e n o l] in the presence of potassium ferricyanide and d ilu te sodium carbonate has been found adaptable to quantitative analysis for this ph en olic material and has been used for its determ ination in fabric. A b s o rp tio n curves and standard calibration curves are given for several other com m ercially im portant phenols, with the suggestion that this co lor reaction may find application in the quantitative determ ination of many p h en olic fungicides, germ icides, and other materials.
R
E C E N T L Y certain fungicidal m aterials h ave come in to extensive use as mildew preventives for fabrics destined to be used in tro p ical areas (4). One of th e com pounds w hich has been m ost widely used for th is purpose is 2,2'-m ethylcnebis[4-chloro- phenol], know n to th e textile trad e as C om pound G-4. T he m ethod m ost comm only used in th e p a st for th e determ ination of th is com pound on fabric is a m odification of th e halogen m icro
procedure of W illard an d T hom pson (5). T h is m ethod, how
ever, has been found in practice to be tim e-consum ing an d the resu lts to be uncertain in som e cases because of th e presence of inorganic or organically b ound chlorine in o th er form s on the fabric. B o th these difficulties are avoided b y th e m ethod here described. P relim inary d a ta on several additional phenolic com
pounds suggest th a t th e new procedure m ay be useful in th e d eterm in atio n of phenolic fungicides, germicides, and other m aterials.
T h e reaction used is based on th e observation m ade by E m er
son th a t 4 -am inoantipyrine condenses w ith arom atic qmines (1) in th e presence of acid oxidizing agents an d w ith phenols (2,3) in th e presence of alkaline oxidizing agents to yield a series of an- tip y rin e dyes. O n th e basis of th is reaction E m erson proposed a color te s t for phenols (2). T h e stru c tu re suggested for th e dyes produced in th e reactio n is show n in th e following exam ple for phenol :
C ,H 5 N / \ C IL — N C = 0
C H 3— O
K 3F e (C N )«
A lkali -N IL
C6I-I5
E m erson concluded t h a t su b stitu tio n to o k place in th e position p a ra to th e phenolic OH group a n d th a t th e s tru c tu ra l require
m en ts for th e reaction were as follows:
January, 1946
GAMMAS OF PHENOLIC COMPOUND
Figure 1 . Standard Calibration Curves for D yes Produced by Reaction of 4 -A m in o a n tip y rin e with o-Phenylphenol, 2 ,2 '-
M e th y le n e b is [4 -c h lo ro p h e n o l], and Salicylanilide
’Filters used for data in Figures 1, 2 , and 3 are those suggested in text where reactions of various compounds with 4-aminoantipyrine are described
1. A t least one free phenolic group m ust be present.
2. T h e position p a ra to the phenolic OH m u st either be un- su b stitu te d , or su b stitu te d by halogen, carboxyl, sulfonic acid, hydroxyl, or m ethoxyl, w hich groups are expelled in th e reaction.
■ S u b stitu tio n in th e p a ra position by alkyl, aryl, nitro, benzoyl, jiitro so , or aldehyde groups blocks th e reaction.
U n d er certain conditions the reaction of 4-ainlnoantipyrine w ith phenols is readily am enable to q u an titativ e treatm en t in th e d eterm in atio n of phenolic fungicides. A m ethod (o f the de
term in atio n of 2,2'-m ethylenebis[4-chIorophenol] on fabric is p resented below and absorption and stan d ard calibration curved are given for o-phenylphenol, salicylanilide, 2,2'-m ethylenebis- [3,4,6-trichlorophenol], 4,4'-isopropylidenediphenol, 4,4'-isopro- pylidenebis|2-chlorophenol], 2,4-dichlorophenol, pentachloro- phenol, and tetrabrom o-o-cresol.
D E T E R M IN A T IO N O F 2 ,2 '-M E T H V L E N E B IS [4 -C H L O R O P H E N O L ]
T h is com pound reacts w ith 4-am inoantipyrine in an alkaline
•oxidizing m edium to form a red dye. U nder the conditions de
s c r ib e d below th e dye develops alm ost im mediately and is rela
tively stab le for several hours. T he stan d ard calibration curve, m ade w ith an Aminco T y p e F photom eter, balanced against dis
tilled w ater, an d th e absorption curve, taken w ith a Coleman M odel 10S spectrophotom eter, are shown in Figures 1 and 4, respectively. T h e procedure for determ ination of this compound
in fabric is p resented below.
So l u t i o n s. 4-A m inoantipyrine, 2 % . Two grams o f 4-amino-
;aritipyrine (m .p. 108-109°) dissolved in 100 g c . of distilled w ater.
'T h is com pound is m ade from an tip y rin e by the m ethod described by E m erson (I ). I t h as been k ep t in solution in stoppered dark b o ttle s in th e lab o rato ry for several m onths w ithout noticeable
■deterioration.
Po tassiu m ferricyanide, 8% . E ig h t gram s of c.p. potassium ferricyanide dissolved in 100 cc. of distilled w ater.
Sodium carbonate, 0.025% . c.p. anhydrous sodium earbon-
•ate (4.5 gram s) dissolved in 18 liters of distilled w ater. T he p H of th is solution should be betw een 10.4 and 10.6.
St a n d a r d Cu r v e. D issolve 200 mg. of 2,2'-m ethylenebis- [4-chlorophenol ] in 100 cc. of acetone in a volum etric nask place 1 cc. of th is solution in a second 100-cc. volum etric flask, and
•dilute to th e m ark w ith sodium carbonate solution. T his solu
tio n , containing 2 0 .micrograms of G-4 per cc. is used to obtain
suitable aliquots covering th e range of 20 to 100 m icrogram s.
Place each aliq u o t in a 25-cc. volum etric flask an d ad d 0.5 cc.
of am inoantipyrine reagent. D ilu te to th e m a rk w ith th e so
dium carb o n ate solution, a d d 0.25 cc. of 8 % potassium ferri
cyanide, an d shake vigorously. A fter 5 m inutes, pour th e solu
tion into a colorim eter tu b e an d m easure th e color, using a su it
able filter (green, a b o u t 500 m illim icrons). T h e sam e grade of p ro d u ct should be used in m ailing up th e sta n d a rd curve as was applied to th e cloth sam ple in question.
Fa b r i c An a l y s i s. F o r sam ples of cloth containing up to 2%
of com pound G-4, weigh a 1-gram sam ple of th e very finely c u t m aterial to th e n earest m illigram , an d place th e sam ple in a 200- cc. beaker. A dd 50 cc. of 0.25% sodium carbonate solution and h e a t to gentle boiling. Boil gently for 5 m inutes. P o u r th e hot solution off th e fabric into a 200-cc. volum etric flask (Pyrex).
R ep eat th e extraction twice w ith 50 cc. of th e sodium carbonate solution, each tim e boiling for 5 m inutes an d com bining th e ex
tra c ts w ith th e first one in th e volum etric flask. D ilute th e solu
tion to near th e m ark w ith th e sodium carbonate solution, using th e d iluent to w ash th e textile m aterial twice. Cool the flask to room tem p eratu re an d dilute exactly to th e m ark . F ilte r ab o u t 20 cc. through a d ry filter an d use the proper aliq u o t of th is solu
tion (usually 1 to 2 cc.) to g et a reading on th e sta n d a rd curve.
Place th e aliq u o t in a 25-cc. volum etric flask an d ad d 0.5 cc. of 2% 4-am inoantipyrine solution. D ilu te to th e m ark w ith 0.025%
sodium carbonate solution, ad d 0.25 cc. of 8 % potassium ferri
cyanide solution, shake vigorously, an d a fte r 5 m inutes p o u r in to a colorim eter tu b e an d m easure th e color. A fter mixing th e
re-Table I. Comparative D eterm ination of 2 ,2 '-M e th y le n e b is [4 - ch lorop hen ol] (C o m p oun d G - 4 ) on Fabric
(B y th e W illard an d T h o m p so n halogen m icro p ro ced u re a n d th e 4-am in o a n tip y rin e m e th o d . T h e rp p licatcs in th e a m in o a n tip y rin e m e th o d a re
d ifferen t in d iv id u a l sam p les c u t from a single piece of c lo th .) Sam ple
N o.
P e r C e n t o n F a b ric
A m in o a n tip y rin e W illa rd -T h o m p so n m eth o d
0 .8 9 0 .9 1 0.880 .8 7 1.7 6 1 .7 3 1 .7 6 1.7 4 1 . 1 0 1.0 5 1 .0 7 1.0 6 1.68 1.6 7
m eth o d 0 .9 9
1 .7 5
1 .0 7
1 .6 9
GAMMAS OF PHENOLIC COMPOUND
Figure 2 . Standard C alibration Carves for Dyes Produced b y R e
action of 4 -A m in o a n tip y rin e with 2 ,2 '-M e th y le n e b is f3 ,4 ,6 -tr i- c h lo ro p h en o l), 4 ,4 '-ls o p ro p y lid e n e d ip h e n o l, 4 ,4 '-ls o p ro p y lid e n e -
b is[2-ch lo ro p h en o l], and 2 ,4 -d ich lo ro p h en o l
18 I N D U S T R I A L A N D E N G I N E E R I N G C H E M I S T R Y Vol. 18, No. 1
GAMMAS OF PHENOLIC COMPOUND
Figure 3. Standard C alibration Curves for D yes Produced by Reaction of 4 -A m in o a n tip y rln e with P entachlorophenol and
Tetrabrom o-o-cresol
agents, th e tu b e s should n o t be exposed to d irect su n lig h t or stro n g artificial light, as th is som etim es causes fading of th e colors.
Figure 4 . A b s o rp tio n Curves for D yes Produced b y Reaction of 4 -A m in o a n tip y rin e with 2 ,2 '-M e th y ie n e b is [4 -c h lo ro p h e n o l], 2 ,4 - D ich lo ro p h en o l, S alic ylan ilid e , and 4 ,4 '-ls o p ro p y lid e n e b is [2 -
chlorophenol]
1 0 0 micrograms of phenolic compound used to develop color
A com parison of results o b tain ed on analyzing four sam ples of fabric by th e W illard and T hom pson m ethod (chlorine d eterm in a
tion) and th e 4-am inoantipyrine m ethod is given in T ab le I.
I t w as found t h a t th e colorim etric d eterm inations could be m ade on 16 sam ples of fabric in less th a n 2 hours, including w eighing of sam ples, extraction, developm ent, an d m easurem ent of color.
D A T A O N O T H E R P H E N O L S
A bsorption curves and sta n d a rd calibration curves were m ade for a group of o th er phenols according to th e m ethods described above for C om pound G-4. T h e several sta n d a rd calibration curves are shown in Figures 1, 2, an d 3, an d th e absorption curves in Figures 4 an d 5.
Salicylanilide, a fabric preserv ativ e know n com m ercially as S hirlan, produces a red dye w hich fades very g rad u ally ; calibra
tion an d absorption curves are show n in Figures 1 an d 4.
o-Phenylphenol, a n in d u stria l preserv ativ e for casein paints, cosm etics, leath er finishes, an d sizing m aterials w hich is know n com m ercially as Dowicide 1, produces a stab le red dye w ith a cali
b ratio n curve as show n in Figure 1. 2,2'-M ethylenebis[3,4,6- trichlorophenol], a germ icide for use in soaps, form s a stab le red color w ith calibration an d absorption curves as show n in Figures 2 an d 5. T h e large a m o u n t of inorganic chloride p resen t is said to m ake th e determ ination of th is germ icide in soap very difficult by th e W illard an d Thom pson procedure. 4,4'-Isopropylidene- diphenol, a m aterial currently in th e early stages of comm ercial developm ent, form s a stab le an tip y rin c dye, whose stan d ard calibration a n d ab sorption curves are show n in Figures 2 an d 5, respectively. 4,4'-Isopropylidenebis[2-chlorophenol], a chlo
rin ated analog of th e m aterial previously m entioned, is claimed to be a p o te n t fungicide an d has been suggested as a fabric pre
servative. S ta n d a rd calibration a n d absorption curves are shown in Figures 2 an d 4.
2,4-D ichlorophenol, a n in term ed iate in th e m an u factu re of th e weed-killer, 2,4-dichlorophenoxyacetic acid, form s a stab le red color w ith ,stan d ard calibration an d absorption curves as shown in F igures 2 an d 4.
T etrabrom o-o-cresol, used on a lim ited scale as a fabric pre
servative, reacts w ith 4-am inoantipyrine to produce a green color w ith a n ab so rp tio n m axim um a t 540 m y, which slowly changes to a red color. A fter 1 h o u r th e red color is fully developed and is stab le for a t least 12 hours th ereafter. T h e sta n d a rd calibra
tio n curve an d ab sorption curve for th e red color are given in Figures 3 a n d 5, respectively. M easurem ents were ta k e n for bo th curves exactly 60 m inutes a fte r com bination of th e reagents.
P entachlorophenol, a wood an d fabric preservative know n u n d er th e tra d e nam e of D ow icide 7, reacts w ith 4-am inoantipy
rine to give a green color w hich g radually fades to colorless over a period of 2 h ours. A satisfacto ry calibration curve could be
ob-WAVE .LENGTH IN M ILLIM IC R O N S
»
Figure 5 . A b s o rp tio n Curves for D yes Produced b y Reaction of 4 -A m ln o a n tip y rin e with 4 ,4 '-ls o p ro p y lid e n e d ip h e n o l, 2 ,2 '- M e t h -
y le n e b is [3 ,4 ,6 -tric h lo ro p h e n o l], and Tetrabrom o-o-cresol For first two compounds 1 0 0 micrograms of each compound were used to develop color, as described in section on standard curves. For tetrabromo-o-cresol 4 0 0
micrograms of phenolic compound were used.
tained by m easuring th e color a t 30 m inutes a fte r developm ent of color, using a 640 filter (Figure 3). In th is reaction i t was found th a t 0.5 cc. of the potassium ferricyanide solution and 0.75 cc. of th e am inoantipyrine solution gave th e b est develop
m en t of color.
D IS C U S S IO N
I t can be seen from th e d a ta p resented t h a t 4-am inoantipyriue prom ises to be a very useful reag en t for th e q u a n tita tiv e estim a
tion of phenols. T h e only elem ent appearing to require special care in th e analysis is th e p H of th e reaction m edium . D evia
tions of 0.5 of a pH u n it in eith er direction from th e lim its given caused changes in b o th th e in ten sity a n d sta b ility of th e color produced. A tte m p ts to utilize buffer m ixtures w ere n o t success
ful, since th e ions used in th e buffer m ixtures interfered w ith th e developm ent of th e color. T h u s boric acid-sodium hydroxide,
A N A L Y T I C A L E D I T I O N 19 glycine-sodium hydroxide, an d sodium te trab o rate-so d iu m car
bonate could n o t bo used.
I t will be noticed th a t th e reactions of 4,4'-isopropylidenedi- phenol and 4,4'-isopropylidenebisl2-chlorophenol] represent an anomaly to th e stru c tu ra l requirem ents for the reaction as re
ported by E m erson (2). B o th of these compounds have an ary l alkyl s u b s titu e n t p a ra to th e phenolic OH group which sup
posedly would block th e reaction since they could n o t be expelled by the ferricyanido oxidizing agent. T he readiness w ith which these com pounds re a c t w ith 4-am inoantipyrine suggests th a t som ething o th er th a n a p a ra su b stitu tio n tak es place and brings out th e possibility t h a t in cases where th e p ara position is blocked ortho-quinoid stru c tu re s m ay be form ed. M ore work is needed to elucidate th e exact n a tu re of the reaction between phenols and 4- am inoantipyrine.
A C K N O W L E D G M E N T
T he au th o rs wish to express th eir g ra titu d e to R . E. H orsey, W . G um p, an d E. N ikaw itz of G ivaudan-D elaw anna, Inc., for cooperation in several phases of th is investigation an d to R . K . Zuck for assistance in th e experim ental work. T h a n k s are also due to th e Dow C hem ical C om pany for furnishing sam ples of phenolic m aterials.
LITE R A TU R E CITED
(1) E isen sta ed t (E m erson), E . I ., J . Org. Chem ., 3, 153 (1938).
(2) E m erson, E . I ., Ib id ., 8, 417 (1943).
(3) Em erson, E . I., U . S. P a te n t 2,194,201 (1940).
(4) M arsh, P . B „ el al., U . S. D e p t. A gr., Tech. B u ll. 8 9 2 ,1 - 2 2 (1945).
(5) W illard, II. H ., and T h om p son , J. J., J . A m . Chem . Soc., 52, 1893 (1930).