Ugye et al / Chemistry Journal (2013), Vol. 03, Issue 05, pp.

128-132 ISSN 2049-954X

Research Paper

Temperature Effects on the Rate of Reaction of Plasma

Albumin with Formaldehyde in Water Solution and
Ethanol-Water-Mixtures
J.T. Ugye1*, A. Uzairu2, S.O. Idris2 and H.O. Kwanashie3
1
Department of Chemical Sciences, Federal University, Wukari, P.M.B 1020, Wukari
2
Department of Chemistry, Ahmadu Bello University, Zaria
3
Department of Pharmacology and Therapeutics, Ahmadu Bello University, Zaria
*Phone: +2348069519202
*E-Mail: torugye2007@yahoo.co.uk

Abstract
The effects of temperature on the rate of reaction of plasma albumin and formaldehyde in water solution and in ethanol
water mixtures (1-25%) were investigated under seven temperatures from 20-45 oC. The effects of the temperature
variations on the reaction of plasma albumin with formaldehyde were then established from a plot of ln(At-A∞) against
time at each of the varied temperatures. Also the Erying plots, logk2/T vs. 1/T as a function of different temperatures were
plotted and the thermodynamic parameters were calculated from the relation ln(k2 / T) = - ΔH҂/RT + ΔS҂/ R + 23.8 while
the activation energy was calculated by using the expression Ea = ΔH҂ + RT and the plot of lnk against 1/T according to the
Arrhenius equation. The result of the temperature dependent study in water solution showed a mean second order rate
constant k2=, 2.47 (±0.1) mol-1s-1, while that in ethanol-water-mixtures showed a mean value of (2.47 (±0.4). The rate
constants of the reaction in both media were observed to have decreased steadily with increase in temperatures, it could
then be asserted that the reaction was not favoured at high temperatures in both media.

Keywords: Temperature Effects, Water Solution, Ethanol-Water Mixtures, Reaction Rate, Plasma Albumin, Formaldehyde

1. Introduction
Formaldehyde is a known toxin to humans. Its residue in
food for human consumption is proscribed because of its
possible carcinogen nature. The main sources for the form-
aldehyde pollution in air are reported to include painting,
coating material and cigarette smoking (USEPA, 2012).
However, formaldehyde is known to be toxic to humans
and other animals at high concentrations and particularly
toxic to humans at levels above 0.08% w/v of formaldehy-
de in human blood and irritating to the respiratory tract,
eye and skin (Chun, et al, 2007). Inhalation of a large amo-
unt of formaldehyde is reported to cause severe nasal tum-
ours and irritation of the upper respiratory tract in both
rats, mice and humans. Data from human exposures indic-
ated that high concentrations of formaldehyde may lead to
pulmonary oedema (USDL, 2009). Recent studies have sh-
own that neuro- degeneration is closely related to misfoldi-
ng and aggregation of neuronal tau. Neuronal tau is an
important protein in promoting and stabilizing the microtu-
bule system involved in cellular transport and neuronal
morphogenesis. Formaldehyde is reported to penetrate thr-
ough the blood-brain barrier and could cause some lesions
to central nervous system, especially our visual system
(Shcherbakova et al, 1986). It has also been established
that high levels of formaldehyde reacts with proteins and
nitrogen atoms in the environment to form reversible and
irreversible adducts in vitro, in vivo and clinically, the
lethal dose of formaldehyde for human beings is found to
be about 0.08% w/v in the circulation (Erkrath et al,
1981). Long-term exposure to Formaldehyde has been fou-
nd to cause irreversible neurotoxicity and is related to cent-

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 Ugye et al / Chemistry Journal (2013), Vol. 03, Issue 05, pp. 128-132
ral nervous system cancer (brain astrocytoma) (Stroup et
al, 1986 and Kilburn et al, 1994). In addition, inhaled for-
maldehyde has been shown to cause behavioural and mem-
ory disorders in rats and has been classified as ‘probably
neurotoxic (Pitten, et al, 2000). It has also been reported
that inhaled formaldehyde gas has negative effects on the
central nervous system, and these effects may appear acut-
ely in the form of headaches, malaise, sleeping disorders,
fatigue, anorexia and dizziness. Formaldehyde has been
reported to have the potential of significantly mis-folding
DNA and native soluble proteins into attendant amyloid
fibrillation and insoluble fibrils comprising of cross- ß-
sheets (Chun, et al, 2005 and 2007). It also has been found
to have the potential of oxidizing protein-adducts with the
attendant increase in acid levels in the blood even if it
remains in the human body for only a short time Lazo et al
(2008). There is, however, paucity of information on the
effect of temperature on the reaction of formaldehyde with
plasma albumin. This study therefore aims to explore the
effects of temperature on the reaction of low concentrate-
ons of formaldehyde on plasma albumin at temperature ra-
nge of between 20-45 oC in water solution and ethanol wat-
er solutions (1-25 %) for better insight in this all important
human and food toxin.
Rajbar (1968) reported that human blood proteins like he-
moglobin and serum albumin undergoes a slow non-enzy-
matic glycation with oxidizing agents such as formaldehy-
de, mainly by forming Schiff bases between є- amino gro-
ups of lysine and some times arginine and glucose molecu-
les in the blood. Elevated glycol albumin has been report-
ed in diabetes mellitus (Iberg and Fluckiger, 1986). Glyca-
tion is found to result in the formation of advance Glycos-
ylation End-Products (AGE), which results in abnormal
biological effects. AGEs are reported to be antigenic and
represent many of the important neo antigens found in coo-
ked and stored foods and also found to be channel blockers
for calcium, magnesium etc. (Vazquez et al, 2008). Becau-
se of the toxicities of formaldehyde to humans, quest for
information about how this substance causes specific dise-
ase conditions through its reactions with bio molecules in
the human body has become a necessity. To date, there are
a few scientific studies reporting the detailed mechanisms
of the reaction of formaldehyde with bio-molecules, espec-
ially proteins. In fact Chun et al (2007) were the only ones
to report the kinetics and mechanisms of the reaction of
formaldehyde with a human protein -protein tau using a
sodium dodecyl sulphate-polyacrylamide gel electrophore-
sis (SDS-PAGE) method. Their study found that tau prote-
in was cross-linked and aggregated by formaldehyde via a
second order mechanism thus indicating formaldehyde is
neurotoxic. Formaldehyde in the human body is distributed
by blood and its possible reaction with plasma albumin is
worth exploring at body temperature range.
2. Materials and Methods
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ISSN 2049-954X
The stock solutions of the plasma albumin used in the
study were prepared by dissolving 50 gdm-3, (0.75x10-3
mol.dm-3) in distilled water. The dilute aqueous solution
0.51x10-4 mol dm-3 was prepared by diluting 50 g of the
plasma albumin in a volumetric flask followed by addition
of 10 cm3 of (0.1 mol.dm-3 HCl) which was further diluted
to mark in a 1000 cm3 volumetric flask with distilled wat-
er. Other lower albumin working solutions of the plasma
albumin (40 g.dm-3) was also prepared by the dilution of
1.6 cm3 of the 0.75x10-2 mol.dm-3 stock solution into a 10
cm3 volumetric flask to obtain the 0.6x10-3 mol.dm-3 work-
ing albumin solution.
Also the hemiacetals were prepared readily by method of
Tong (2003) by dissolving the various formaldehyde conc-
entrations in the (1-25%) ethanol-water mixtures in 1:1
ratio and the stock solution left to stand for two hours in
some beakers. Fresh preparations were made for each runs.
The rate of disappearance of plasma albumin in the format-
ion of the adduct from the reaction of formaldehyde with
plasma albumin in aqueous solution as well as in the
ethanol-water mixtures was followed in situ at its maxim-
um wavelength (235nm) in a compartment of a UV spectr-
ophotometer. The kinetic measurements were carried out
at constant conditions with the concentrations of the form-
aldehyde in excess of 0.27x10-1 mol.dm-3 while that of
plasma albumin was kept constant at 0.51 x10-4 mol.dm-3.
The reaction was carried out by mixing 1-2 cm3 of the
plasma albumin with 2-3 cm3 of 0.27x10-1 mol.dm-3 HCHO
at ionic strength of [0.5 NaCl] mol.dm-3. The effects of
temperature on the rate constants in the reaction of plasma
albumin and formaldehyde under the given constant condi-
tions were investigated under seven temperatures from 20-
45 oC. The temperature dependence of the reactions was
carried out at constant reactants concentration, Ionic stren-
gth, µ= 0.5 [NaCl) mol.dm-3 and pH 7.0 ±3 in (1-25%)
ethanol-water mixtures as well as in water solution. The
effect of the temperature variations on the reaction of plas-
ma albumin with formaldehyde was then established from
a plot of ln(A∞-At) against time at each of the varied temp-
eratures. Triplicate values were taken in each case to obta-
in the pseudo-first and second order rate kinetics.
The energy of activation was determined from the plot of
lnk against 1/T according to the Arrhenius equation
lnk = Ea/RT+lnA
Where:
k=The rate constant
Ea= activation energy
T= temperature in Kelvin
R= gas constant and
A= frequency factor
Also the Erying plots, that is logk2/T vs. 1/T as a function
of different temperature constant were plotted and the ther-
129
 Ugye et al / Chemistry Journal (2013), Vol. 03, Issue 05, pp. 128-132 ISSN 2049-954X

modynamic parameters were calculated from the relation temperatures in water. Since the thermodynamic properties
ln(k2 / T)= - ΔH҂/RT + ΔS҂/ R + 23.8 while the activation of the activated complex are usually taken as a measure of
energy was calculated by using the expression Ea = ΔH҂ + the solution effects (Asadi et al, 2010).
RT (Asadi et al, 2010). The values of the thermodynamic
parameters energy were determined and the results were The typical linear Erying plot of ln(k2 /T) vs. 1/T at the
recorded. seven different temperature range of (20-45 oC) for plasma
albumin and excess 2.7 x 10-2 mol.dm-3 of formaldehyde in
3. Results and Discussion water was also calculated as given in Figure 1. The plot
gave a positive slope and an approximate correlation value
3.1. Effects of Temperature on the Rate of the Reaction of 0.9 (r2 = 0.892). The large negative ΔS҂ (-220.795 J mol-
1
in Water Solution K-1) parameter as shown in (Table 2) indicated random
distribution of the solvent molecules in activated state i.e.
The results of the temperature dependence on the reaction the reaction passed through an ordered transition state (El-
rate in water solution were as presented in Tables 1-3 and Mallah et al, 2010). The small positive values of Ea (1.687
Figure 1. kJ Mol-1) and negative value of ΔH҂ (-1.715 kJmol-1) and a
Table 1. Experimental Results of Pseudo-First Order and Second Order Rate Constants at 2.7 x 10-1 mol/dm3 HCHO of Formaldehyde With
Plasma Albumin Reaction at Different Temperature in Water System

K kobs k2 1/T logk2 k2/T log (k2/T)

293 0.071 2.63 0.00341 0.420 0.00897 -2.047
298 0.068 2.52 0.00336 0.401 0.00847 -2.072
303 0.068 2.52 0.00329 0.401 0.00847 -2.072
308 0.067 2.48 0.00325 0.394 0.00806 -2.094
310 0.067 2.48 0.00323 0.394 0.00806 -2.094
313 0.065 2.41 0.00319 0.382 0.00778 -2.109
318 0.061 2.26 0.00314 0.354 0.00710 -2.149

Table 2. Experimental Pseudo-First Order and Second Order Rate large positive ΔG҂ (65.643 kJmol-1) clearly showed that the
Constants Values for the Reaction of Plasma Albumin and reaction was not spontaneous in aqueous solution over the
formaldehyde in Water At Varying Plasma Albumin Con-
centrations and Constant Conditions at Human Body Te- considered temperature range. Activation energy values
mperature T= 37.0 oC, Formaldehyde 0.27 x10-5], NaCl calculated from Erying plots in this study were found
[0.5] mol dm-3] pH= 6.5., λmax = 235nm negative for this reaction. However it has been reported
that it is possible to have a negative activation energy valu-
PABNH2 es when rates of reaction decreases with increase in tempe-
S/No kobs sec-1 k2 mol-1s-1
[moldm-3] rature (Jensen, 2010). Reactions exhibiting these negative
1 2.1 x10-2 0.066 3.14 activation energies are reported typically to be barrierless
2 3.5 x10-2 0.078 2.23 reactions, in which the reaction proceeding relies on the
capture of the molecules in a potential well. Increasing the
3 5.1 x10-2 0.084 1.65
temperature leads to a reduced probability of the colliding
4 7.5 x 10-2 0.083 1.11 molecules capturing one another (with more glancing colli-
4.6 10-2 (±0.02) 0.078(±0.008) 02.03 (±0.87) sions not leading to reaction as the higher momentum carr-
ies the colliding particles out of the potential well), expres-
The results showed a second order rate constant but unlike sed as a reaction cross section that decreases with increase-
in most reactions in which the rate increases with tempera- ng temperature. Such a situation no longer leads itself to
ture, the increase was not sharp. This could be due to the direct interpretations as the height of a potential barrier
denaturation of the plasma albumin substrates which are (Jensen, 2010).
responsible for the reaction, hence, decrease in the concen-
tration of the plasma albumin. The mean second order rate Suffice to say that if positive activation energy keeps a rea-
constant of 2.47 (±0.1) mol.-1s-1 at the varying temperatu- ction from occurring until that amount of energy is provid-
res (20-45 oC) is greater than the average second order rate ed from the environment, then negative activation energy
constant value of 1.32 (±0.58) mol-1s-1 at 37 oC as shown in would imply you could not stop the reaction from occurri-
Table 3. As both reactions were carried out in water solute- ng if is tried. The reactants already have plenty of energy
on, it indicated that the reaction was not favoured at high for the reaction so the only way to prevent the reaction

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 Ugye et al / Chemistry Journal (2013), Vol. 03, Issue 05, pp. 128-132 ISSN 2049-954X

would be to keep the reactants apart. If this energy is nega- and ΔH҂ (-1.715 kJ.mol-1) and large ΔG҂ (61.180 kJ.mol-1)
tive then it can be said that this is a barrierless reaction and clearly showed that the reaction was not spontaneous in the
the reaction rate is proportional to the collision frequency. ethanol-water mixtures over the considered temperature ra-
nge and at the formaldehyde concentration. According to
Table 3. Thermodynamics and Other Parameters for Reaction in Asadi et al (2010) the negative ΔS҂ value indicates that the
Various Ethanol-Water Media. Temp. = 20-37 oC, ionic
Strength = 0.5 mol.dm-3 reaction is compatible with the associative mechanism in
this case a cross-linking / binding reaction as earlier report-
ed in the literature (Bedino, 2003).
Solvent Ea ∆G≠ ∆H≠ ∆S≠
Comp. (%) (kJ mol ) (kJmol ) (kJmol ) (Jmol-1K-1)
-1 -1 -1

1 1.290 66.880 -0.951 -225.351

5 0.891 59.232 -1.612 -202.139
15 0.355 59.301 -2.148 -202.802
20 0.355 59.301 -2.148 -202.802
Mean Values 0.723 61.180 -1.715 -208.270

Figure 2. Combined Erying Plots of 1/T vs. logk2/T in 1%, 5%, 15%b
and 20% Ethanol-Water Mixtures

Figure 1. Erying Plot of ln(k2/T) Versus 1/T in Water System
3.2. Effects of Temperature on the Rate of the Reaction
in (1-25%) Ethanol-Water) Mixtures
Tables 2 and Figures 2 showed second order rate constant
results for the reaction in the ethanol-water mixtures (1-
25%). The rate constants were observed to have decreased
steadily with increase in temperatures just as in water med-
ium and it could be due to solvent effect and the denaturat-
ion of the plasma albumin substrates which were response-
ble for the reactions. The mean second order rate constant
of 2.47 (±0.4) mol-1s-1 at the varying temperatures (20-45
o
C) was also observed.
The typical linear Erying plots of ln(k2 /T) vs. 1/T at the se-
ven different temperature range of (20-45 oC ) in Ethanol-
water mixture (1-25%) were also calculated as presented
in (Figures 2). The thermodynamic parameters were calcu-
lated as in water solution based on the method of Asadi et
al (2010) and the mean values are as given in Table 3. Si-
milar to the result obtained in water solution, the negate-ve
ΔS҂ parameter (-208.27 J mol-1K-1) indicated random distr-
ibution of the solvent molecules in activated state i.e. the
reaction passed through an ordered transition state (Asadi
et al, 2010). Also the small values of Ea (0.723 kJ.mol-1)
4. Conclusion
The results of the temperature dependent study of this rea-
ction in water solution and in ethanol-water mixtures sho-
wed a second order rate constant. The rate constants of the
reaction in both ethanol-water mixtures and water solutio-
ns decreased steadily with increase in temperatures. It cou-
ld be asserted that the reaction was not favoured at high
temperatures in water. The results of the thermodynamic
parameters in both media showed small negative values of
Ea and ΔH and a large positive ΔG҂ which clearly showed
that the reaction was not spontaneous in both ethanol-
water mixtures and water solution over the considered te-
mperature range. Reactions exhibiting these negative acti-
vation energies are reported typically to be barrierless rea-
ctions, in which the reaction proceeding relies on the capt-
ure of the molecules in a potential well. Other than these
reasons it could be asserted that this reaction is a barrierle-
ss reaction both in water solution and ethanol-water mixt-
ures.
Acknowledgements
This work was not supported by any donor agent, however,
we wish to appreciate the contributions of Prof. Kehinde
Okonjo of the Department of Chemistry, Convenant Univ-
ersity, Ota-Nigeria, Dr. S.G. Yiase and Mr S.O. Adejo as
well as the technical staff of ABU Zaria and the Benue
State University, Makurdi for their assistance and in facili-
tating the conduct of the bench Chemistry of this study in
their various laboratories.

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