Mikrochim.

Acta [Wien] 1989, I, 329--336

Mikrochimica
Acta
9 by Springer-Verlag1989

Complex Compounds of Tetracycline

with WO42- and MoO42- Ions. Investigation
by pH-Metric and Conductometric Methods

Dragan Veselinovi61, * and Milena Jeliki6-Stankov 2

1 Institute of Physical Chemistry, Faculty of Science, University of Belgrade,
Studentski Trg 12-16, YU-11000 Beograd, Yugoslavia
2 Institute of Analytical Chemistry, Faculty of Pharmacy, University of Belgrade,
Dr. Suboticfi 8, YU-11000 Beograd, Yugoslavia

Abstract. By using pH-metric and conductometric methods it has been

found that tetracycline (H3TC) forms with WO]- and MOO]- ions the
following complex compounds: [WO3HTC] 2-, [WO3(H2TC)2]2- and
[MoO3(H2TC)d 2-. Stability constants log flf = 7.86 and logfl~ = 7.80 for
[WO3HTC] 2- and [MoO3HTC] 2-, respectively, have been calculated
from pH-metric measurements.
Key words: tetracycline, WO]- complex, MOO]- complex.

Investigation of antibiotics and tetracycline (H3TC) complexes with several

bivalent and trivalent metal ions begins with the works of Albert [1, 2].
Considerable decrease of H3TC concentration in the organism was found
due to complex formation during simultaneous therapeutic application of
H3TC, ferropreparations and antacide drugs [3, 4]. Complex compounds of
H3TC with different metal ions enable determination of H3TC contents in
pharmaceutical preparations by both colorimetric and spectrophotometric
methods [5--8].
In our work we investigated composition and stability constants of
H3TC complexes with WO42- and MOO]-. Available literature data refer to
the use of tetracycline and (NH4)eMoO4 [9, 10] or Na2WO4 [11], colored reac-
tions which lead to decomposition of the H3TC molecule for colorimetric
and spectrophotometric determination of this antibiotic in pharmaceutical
preparation. No data have been found in the literature on the conditions of
complexation with WO42- and MoO 2- ions.

* To whom correspondence should be addressed

330 D. Veselinovi6 and M. Jeliki6-Stankov

Experimental
Reagents
The reagents used were: H3TC - HC1 standard (Institute for Drug Control of the Socialist
Republic of Croatia), Na2MoO4" 2H20 "Merck" and Na2WO4" 2H20 "Reanal". All p.a.
H3TC solutions were prepared by dissolving exact quantities of H3TC standard in water
while the initial Na2WO4 and Na2MoO4 solutions were standardized gravimetrically [12].
Solutions were prepared with fresh bidistilled water with the conductivity of 3.33 -10 -6 S
and pH > 6.0.

Equipment
A pH meter Iskra Model MA 5705, a conductometer Iskra Model MA 5951 and a thermostat
VEB Prgfgerfite, precision + 0.020 C, were used. The pH-meter was calibrated using buffer
solutions radiometer pH = 7.02, and pH = 4.00, at 20 ~C. The cell for conductivity measure-
ments was calibrated by using 0.01 M KC1 solution ( C = 7.09 9102 S, at 25~ Conductivity
measurements were performed with an error of 1% while the pH was measured with the
precision of _+0.01 pH units.

Results and Discussion

Composition of Complexes
Composition of complex compounds of H3TC with WO]- or M0024- was
determined by the Job's method [13, 14] using pH-metric and conducto-
metric measurements in a series of three solutions, as follows:
(a) x ml of H3TC and (10 - x) ml of Na2WO4 o r N a 2 M o O 4 solutions,
(b) x ml of H3TC and ( 1 0 - x ) ml of water, and
(c) x ml of N a 2 W O 4 o r N a 2 M o O 4 solution with x ml of water.
Corresponding series of solutions (a), (b), (c) were prepared for each x value
between 1 and 9 (Tables 1 and 2). The concentration used was always
5.10 -4M. Measurement of pH of solutions was performed after thermo-
stating at 20 +_0.02~ for 10 rain. Their specific conductivity was measured

Table 1. pH values of solution mixtures of 5 91 0 - 4 M Na2WO 4 o r N a 2 M o O 4 and H3TC

Na2WO4 or Na2WO~ or Na2WO] or H3TC 1, H3TC2+ H20

Na2MoO4 Na2MoO 2+ H 2 0 Na2MoO24+ H3TC
ml pH 1 pH 2 pH 1 pH 2 pH 1 pH 2

1.00 5.41 5.99 3.20 3.33 2.97 3.21

2.00 5.48 6.00 3.26 3.48 3.03 3.23
3.30 5.57 6.12 3.56 3.87 3.04 3.28
3.50 5.58 6.15 3.61 3.94 3.06 3.30
4.50 5.59 6.16 4.06 4.21 3.08 3.33
5.00 5.60 6.18 4.30 4.36 3.09 3.36
6.00 5.62 6.20 4.73 4.56 3.13 3.48
8.00 5.63 6.22 5.14 4.90 3.29 3.92
Complex Compounds of Tetracycline with WO 2- and MOO]- Ions 331

Table 2. Specific conductivity values of solution mixtures of 5 91 0 - 4 M Na2WO4 or Na2MoO4

and HaTC

Na2WO4 or Na2WO] or Na2WO41 or H3TC 1, H3TC 2 q- H20

Na2MoO4 Na2MoO ] + H20 NaRMoO42 + H3TC
C[S cm -11 C[S cm -1] C[S cm -~1
] 2 1 2 1 2
ml .10 .5 "10 .5 .10 .5 .10 -5 .10 .5 .10 .5

1.00 1.879 2.580 12.40 11.92 12.92 ]2.92

2.00 3.298 3.340 ]0.69 8.857 11.48 11.07
3.30 4.768 4.690 8.159 7.046 10.00 9.688
3.50 4.920 4.920 7.750 6.888 9.539 9.393
4.50 6.457 5.960 6.597 7.046 8.379 7.948
5.00 6.888 6.739 6.597 7.21] 7.471 7.38]
6.00 8.159 8.156 7.561 8.379 6.200 6.079
6.70 -- 8.857 -- 9.118 -- 5.081
8.00 11.07 10.33 10.00 10.68 3.264 3.100

at room temperature which was kept constant. The obtained results are
presented in Tables 1 and 2.
It is evident that H3TC and Na2WO4 or Na2MoO4 solution mixtures have
higher pH values than H3TC solution. The decrease of H + ions concen-
tration is due to the following possible reactions,
2 H 3 T C + N a 2 W O 4 ~- N a 2 [ W O 3 ( H 2 T C ) 2 ] -+- H 2 0 , (1)
2H3TC + Na2MoO4 Na2[MoO3(H2TC)2] + H20, (2)
H3TC + Na2WO4 Na2[WO3HTC] + H 2 0 , (3)
H 3 T C q- N a 2 M o O 4 Na2[MoOBHTC] + H20, (4)
resulting in the complex formation and also followed by decrease of the
total ions concentration. In the other case, i.e. if there was no complexation
according to reactions (1)--(4), H + ions concentration in a solution
containing both Na2WO4 and H3TC [solution (a)] would be equal to the sum
of H + concentrations in each solution separately [solution (b) plus solution
(c)]. Due to the complex formation the concentration of H + ions, actually of
all ions present in solution (a), decreases as a function of the quantity of the
formed complex, thus leading to an increase of the pH value and a decrease
of specific conductivity of the solution. Concentration decrease of H + ions
has been calculated by using the following equation,
A[H +] = [H+]a - [[H+]b -ff [H ~]r (5)
where [H+]a is H + ions concentration in solution (a), while [H+]b and [H+]:
denote concentrations in solutions (a), (b) and (c), respectively.
Decrease of specific conductivity C has been calculated from the
equation
A C = Ca - [CB + Cd, (6)
332 D. V e s e l i n o v i 6 a n d M. J e l i k i 6 - S t a n k o v

where Ca is specific conductivity of solution (a) while Cb and Co are, respec-

tively, specific conductivities of solutions (b) and (c).
The results obtained by pH and conductivity measurements are
presented in Tables 1 and 2. Low pH values of solutions (a) and (b) are not
affected by CO2 dissolved in water. Change of the pH of solution (c) due
to dissolution of CO2 is not considerable since it only insignificantly
contributed to the A[H +] value.
Dependence of A[H +] on WO 2- or MOO]-, and o f A C o n WO 2- of MoO z-
molar fraction *n (*n = mol of metal salt/(mol of metal salt + tool of H3TC))
is shown in Figs. I and 2, respectively. For calculation of H + from the pH
value, activity factors of H + ions were taken to be equal to unity.

A [H+] 10-4

-4

-3

-2

-1

0.5 1.0 n MoO 2-,wO 2-

Mol.e fraction of metat satt
Fig. 1. Job's curve obtained by pH-metric measurements; 1" CH3TC = 5 ' 1 0 - 4 M~
CNa2WO4 = 5" 10 -4 M ; 2: CH3TC = 5" 1 0 - 4 M , CNa2MoO4 = 5 " 1 0 - 4 M

AC 1G5

-8

-6

-4

-2

i i

o n MoOh WOe-
Mote fraction of meta[ salt
Fig. 2. J o b ' s c u r v e o b t a i n e d b y c o n d u c t o m e t r i c m e a s u r e m e n t s ; 1: CH3TC = 5 " 10 -4 M,
CNa2WO4 = 5 9"10- 4 M ; 2: CH3TC= 5 " 10 - 4 M, CNa2MoO4 = 5 ' 10 -4 M
Complex Compounds of Tetracyclinewith WO2- and MoO42- Ions 333

The maxima of the curves appear at WO42- or MoO ]- molar fractions of

n = 0.5 and n = 0 . 3 3 , pointing to the formation of the complexes
H 3 T C ' W O ] - or H 3 T C ' M o O ] - = 1"1 and 2"1 according to reactions
(1)--(4).
The literature data indicate the possibility of formation of colloidal
tungstic acid [15] due to the presence of H + ions according to the reaction
NazWO4 + 2H + # H2WO4 + 2Na +,
leading to the increase of the pH of solution. H + ions can also cause poly-
merization [16], whereby hexatungstic polyanion is formed according to the
reaction
6WO 2- + 7HA @ [ H W 6 0 2 d 5 - 4- 7A- + 3H20,
which is followed by the increase of the pH of the solution, too. Products of
this reaction are colorless. Complex compounds formed with H3TC are
colored. The absorption spectra of 4 . 1 0 - S M H3TC in 4 "10-4M Na2WO4
and 5 . 1 0 - 5 M H3TC in 5 . 1 0 - 4 M Na2MoO4, recorded with respect to the
H3TC solution at pH = 6.00, have the maxima at 394 and 390 nm, respec-
tively (Fig. 3).
These results have proven the observed change of pH and specific
conductivity to be due to the formation of colored complex compounds, not
of colloidal acids or polyanions, which is in agreement with the literature
data [17--19].

Calculation of Stability Constants

Relative stability constants of the complexes [WO3HTC] 2- and
[MoO3HTC] 2- were calculated on the basis of pH-metric measurements

0.8 2

0.6

0.4

Fig. 3. A b s o r p t i o n spectra of the

H 3 T C - M o O 2- c o m p l e x a n d H3TC-
0.2 Na2WO42- c o m p l e x ;
1: CH3TC= 5 " 1 0 - 5 M ,
CMoO2- = 5 910 -4 M;
2: CII3TC= 4 910-5 M,
370 390 410 430 ~,nrn Cwo42- = 4 -10 -4 M
334 D. Veselinovi6 and M. Jeliki6-Stankov

using the data obtained for the solutions with excessive metal salts as
compared to H 3 T C . The stability constant of the complex can be calculated
from the following equations, established for the equilibrium of WO]- ions
in the solution,

[H3TCI~ = [H3TCI + [H2TC-] + [ HTC 2-1 + [ W O B H T C ] 2-, (7)

[WO 2- ] = [WO 2-1 + [WO3HTC] 2-, (8)
[H +1 = [H2TC-] + 2 [ H T C 2 - ] , (9)

Kdl = [H+][HeTC-]
[H3TCI ' (10)

[H +1[HTC2-]
G2 = [H2TC-] (11)

The dissociation constants of H 3 T C [20] are Kal = 5.01 910 -4,

Kd2= 2.09.10 .8 and gd3 ~-2.04.10 -4~ Stoichiometric concentrations of
H 3 T C and WO]- or MOO]- are marked by having an index "o". H 3 T C
partial dissociation constants are Kdl and Kd2 whereas dissociation of the
third H § ion, occurring in the alkaline medium which contains the complex,
has not been taken into account due to a very low Kd3 value. Approximate
WO]- , H2TC-, HTC 2- and [ W O 3 H T C ] 2- ion concentrations in the solution
were calculated from Eqs. (7)--(11) assuming the activity coefficient of H +
ions to be equal to unity, i.e. that pH -- - l o g c.+. The ionic strength of the
solution and the H § ion activity factor were calculated by using the
obtained approximate ion concentration. The correct H § ion concentration
was calculated from the pH and H + activity factor. This H + ion concen-
tration value was further used to calculate concentrations of HTC 2-, WO ]-
and [ W O 3 H T C ] 2- ions as well as the relative stability constant of the
complex, by introducing the obtained concentration values into

[WO3 A]
Kr- [WO]-I[AI" (12)

The results are presented in Table 3. Because the difference in the values
obtained for the constants is within the measurement error, the calculation
procedure has not been repeated.

Table 3. Stability constants of [ W O 3 H T C ] 2- and

[MoO3HTC] 2 complexes

Complex pH # log ]3~

[WO3HTC] 2- 4.73 6.18 " 1 0 - 4 7.86

5.14 8.06 ' 10 4 7.16
[MoO3HTC] 2- 4.56 6.28 910 -4 7.80
4.90 8.13.10 -4 7.13
Complex Compounds of Tetracycline with WO 2- and MoO 2- Ions 335

The complexation process between H3TC and WO]- or MOO]- ions

having WO3 or MOO3, respectively, as the complex components, can be
presented by the reactions
WO]- + 2H + ~ WO3 + H20, (13)
WO3 + A ~ WO3A, (14)
whereby 02- is substituted from WO]- by the coordinatively bonded ligand A.
The stability constant obtained for relation (14) can be expressed as
[WO3A] (15)
K = [WOg][AI'
but cannot be positively determined due to the concentration of WO3 mole-
cules in the solution. Since the numerical value of the reaction constant
expressed by Eq. (13) has not been calculated so far it is also impossible to
calculate the stability constant for (15) even if the constant for (12) is
known. From the equilibrium constant of reaction (13), expressed by
[WO3][H20] (16)
K e =- [WO2_l[H+]2,

and from the stability and relative stability constants of the complex (Eqs.
(15) and (12), respectively), the expression
log K, = a - 2pH (17)
is obtained after taking the logarithm. Here a = log Ke - log [H20] + log K.
It follows from Eq. (17) that the difference between log K, values deter-
mined at different pH, log K~, is equal to twice the difference between pH
values, ApH. The same holds for molybdenum complexes formed in the
reaction of the same type as for tungsten. The results log AK~ = 0.70 and
ApH = 0.41 as well as A log K~= 0.67 and ApH = 0.34 obtained for
tungsten and molybdenum complexes, respectively (see Table 3), support
the above conclusion.

References
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336 Complex Compounds of Tetracycline with WO]- and MOO]- Ions

[16] P. J. Bettinger, S. Y. Tyree, J. Am. Chem. Soc. 1957, 79, 3355.

[17] M. Sugi6, D. Veselinovi6, D. Su2njevi6, Bull. Soc. Chim. [Beograd] 1964, 29, 121.
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Received November 10, 1987. Revision May 9, 1988.