MICROCHEMICAL JOURNAL 15, 585-589 (1970)

Spectrophotometric Determination of Iron

with Orthophenanthroline

G. S. R. KRISHNA MURTI, A. V. MOHARIR, AND V. A. K. SARMA

Dil.isicm of Agricult~cml Piz~.~ics, I~~tliatr Agriculrrtr-al Rrsenrci~ Institute,
Nell% Dcllri, I&in

Orthophenanthroline forms with ferrous iron in aqueous solution a

soluble orange-red complex, stable between pH 2 and 9 (IO). Meas-
urement of the color intensity at 5 1S-540 m/l, has been widely used for
the determination of traces of iron in reagent chemicals (I I ), biological
materials (S), plant extracts (/, .?). and citric acid extracts from soils
(7). Several reagents have been suggested for the reduction of ferric
iron prior to formation of the complex. Of these, hydroxylamine
hydrochloride has been considered to bc the most efficient (2). How-
ever, for systems containing citrate, hydroxylamine hydrochloride has
been variously reported to bc unsuitable (I) and quite suitable (7).
The accuracy of the calorimetric determination of iron with ortho-
phenanthroline depends on (a) completeness of reduction of Fe”+ to
Fe’-: (b) pH of iron solution bcforc complex formation, (c) stability
of the colored complex, and (d) interference of other ions in the for-
mation of the color.
Several reagents have been tried as reductants. Sodium dithionite,
though reducing iron completely. introduces turbidity in the test solu-
tion within a few minutes (5). Sodium sulfite and formaldehyde com-
plex with Fe’: and thus cause considerable error in the determination
(2). Sodium and potassium formates lead to error due to formic acid
complexing with Fc:l+ (13).
The ferrous iron-orthophenanthrolinc complex, once formed, is
stable even up to 15 days. and has constant color intensity between pH
2 and 9 (IO, 9). It has been found that the intensity of the color
increases with time if the pH of the iron solution is adjusted to a value
>4 before reduction at room temperature with hydroxylamine hydro-
chloride, indicating incompleteness of reduction ( 7). Complete reduc-
tion of the iron can, however, be achieved in such solutions by heating
the solution with the reductant and a pH 3.5 acetate buffer for 1 hour at
65°C (7).
It has been reported that most anions and cations, up to a concentra-
585
586 KRISHNA MURTI, MOHARIR, AND SARMA

tion 250 times that of iron, do not interfere with iron determination by
orthophenanthroline (2). No common soil ions except orthophosphate
are considered to interfere in the determination (6).
The methods recommended by earlier workers are time consuming
and difficult to apply when the number of samples is large. It was,
therefore, considered necessary to develop a rapid and reliable modifi-
cation of the method. Thioglycolic acid has been successfully used to
reduce iron to eliminate interference in the calorimetric determination
of zirconium (4). It was thought worthwhile to investigate the suitabil-
ity of thioglycolic acid as reductant in the spectrophotometric determi-
nation of iron with orthophenanthroline. The procedure so developed is
presented below.
MATERIALS AND METHODS

Iron standards were obtained by dilution of 100 ppm iron stock solu-
tion prepared from spectroscopically pure ferric oxide. A Lumetron
Model 402-E photoelectric calorimeter with a narrow band 515 rnp
monochromatic filter was used for measurement of transmittance of the
solutions after development of color.
Proposed method. A suitable aliquot (< 150 ,Lg of Fe) of the solu-
tion is taken in a 50-ml volumetric flask and 10 ml of (pH 3.5) M
sodium citrate-citric acid buffer, followed by 1 ml of 4% thioglycolic
acid, is added. The contents of the flask are mixed well, 2 ml of 0.4%
orthophenanthroline are added, and the volume is made up to 50 ml.
Transmittance of the solution is measured at 515 rnp after 5 minutes.
A standard curve in the range O-150 pg of Fe is prepared in the same
way using aliquots of standard iron solution, and the iron content of the
test solution is read from the curve.
RESULTS AND DISCUSSION

The optical densities obtained with different amounts of iron are pre-
sented in Table 1 and show that Beer’s law is obeyed in the range O-3
ppm Fe. Development of color was instantaneous, and the color of the
complex was stable for 72 hours.

EfJect of pH
Aliquots of standard iron solution representing 62.5 pg of Fe were
buffered with 15 ml of citrate-citric acid buffers of different pH values,
and the amount of iron was determined by the proposed method. Table
2 gives the amount of iron recovered from each solution and the final
pH of the solution. Complete recovery in all cases showed that reduc-
tion by thioglycolic acid was complete at all the pH values studied. In
 DETERMINATION OF IRON 5x7

TAfSf.f- I

OPWAL Dmstrtts OF %AVDAIW ftwN sOt.UTIONS ,zr 515 tnP Umt;

I ttt. Pruwost II MLTIIOD

I-inal iroti
C‘OiIC
No. ( ,‘,,I11 1

I 0.25
7 0.50
3 I .oo
1 I io
5 2.00
6 2.50
1 i .oo

the proposed method, however, addition of pH 3.5 buffer is rccom-

mended to prevent the possible precipitation of phosphates from test
solutions.
Aliquots of standard iron solution representing 50 /kg of Fc were
adjusted to different pH values between 2 and 9 by predetermined ali-
quots of 0.1 N NaOH and 0.1 N HCI. The iron in the solutions was
then determined by the present method and the methods of Olson (9)
and Krishna Murti et al. (7). The results, given in Table 3, show that
complete recovery of iron was obtained using the method of Krishna
Murti et al. (7) and the present method. Olson’s (9) method gave
extremely low values for iron when the pH of the solution was greater
than 2. In a solution of pH higher than 4, the iron forms hydroxylated
complexes which are difficult to reduce (7). Hydroxylamine hydro-
chloride is capable of reducing the iron only with heating at 65°C for 1
hour (7). Thioglycolic acid, however, reduced the iron instantly at
room temperature at any pH.
588 KRISHNA MURTI, MOHARIR, AND SARMA

Interference of Other Ions

Suitable aliquots representing various concentrations of several other
ions in the final solution were taken with 50 /.lg of iron and the amount
of iron was determined by the proposed method. No interference of Ca
(10,000 ppm), Mg (1200 ppm), Al (150 ppm), Ti (80 ppm), Zr (50
ppm>, PO4 (30 ppm), Mn (20 ppm), Ba (20 ppm), Zn (20 ppm),
Cu (2 ppm), and Ni (0.6 ppm) was found in the determination of iron
with the proposed method.
Free Iron Oxides Extracted from Soil Clay
The free iron oxide content in the clay fraction (<2 p of two soils
from Madhya Pradesh, India, were extracted by the dithionite-
-citrate-bicarbonate method of Mehra and Jackson (8). Suitable ali-
quots of the extract were taken and the iron was determined by the
present method and the method of Krishna Murti et al. (7). The
amounts of iron obtained are presented in Table 4. The closenessof the
values show that the proposed method is reliable and accurate for such
determinations.

TABLE 3

EFFECT OF INITIAL pH OF IRON SOLUTION ON THE DETERMINATION OF IRON

BY VARIOUS METHODS

Amount of iron found (,~g)

Amount of Initial
iron taken pH of iron Olson Krishna Murti Proposed
No. h! solution a (1965) et al. (1966) method
1 50.0 2.0 50.0 50.0 50.0
2 50.0 4.0 20.5 49.5 50.0
3 50.0 6.0 15.0 50.0 50.0
4 50.0 9.0 0 49.5 50.0
a pH adjusted by 0.1 N HCl or 0.1 N NaOH

TABLE 4
DETERMINATION OF IRON IN EXTRACIX OF FREE IRON OXIDES FROM CLAY
FRACTIONS OF Sons
Amount of iron found (pg)

No. Clay sample Proposed method Krishna Murti ef al.

1 Umaria 97.5 96.5
2 Raogarh 64.5 64.5
 DETERMINATION OF IRON 589

SUMMARY
A modified procedure is presented for the spectrophotometric determination of
iron by orthophenanthroline. Reduction of iron prior to development of color is
accomplished by thioglycolic acid, and is instantaneous at room temperature. The
system is buffered at pH 3.5 with sodium citrate-citric acid to avoid precipitation
of hydroxides and phosphates from the test solution. Moderate amounts of Ca,
Mg, Al, Ti, Zr. PO,, Mn, Ba. Zn are permissible but more than 2 ppm of Cu
and 0.6 ppm of Ni interfere.

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