Download Experiment For Measuring The Partition Coefficient

Chemistry 1
Running Head: PARTITION COEFFICIENT
Experiment For Measuring The Partition Coefficient
Chemistry 2
Introduction
The ratio of concentration of a solution in two different phases is known as the
partition coefficient. For the calculation of partition coefficient, a chemical substance
is used which is known as solute. Solute is a soluble component in solution of two
different type of liquids which can not be mixed with each other. This substance is
mixed in the solution for having a constant at an equilibrium. Such constant is termed
as partition coefficient.
Formula of partition coefficient:
The partition coefficient can be Claudette by dividing concentration of solute
in solvent A by concentration of solute in solvent B. This can be presented as
mathematical equation presented as below:
P = CA / C B
where CA = concentration of solute in solvent A
CB = concentration of solute in solvent B
(Hodgson, 2004)
Experiment for the having partition coefficient in a chemical reaction
For the experiment, water and oil were taken in to account as two different
non-miscible solvent and Salicylic acid was used as solute because it is soluble in
both solvents. For having the partition coefficient between such type of non-aqueous
and an aqueous phase, following formula will be used :
P = Coil / C water
with help of partition coefficient, the absorption capacity of a particular drug
in a particular solution can be predicted easily. Logarithmic form of such type of
coefficient also enable the researcher to calculate the hydrophocity and lipophilicity of
the solute used in the experiment. As the body membrane of human is permissible for
Chemistry 3
only some specific outside ionic species, only some specific drugs are allowed to
enter in to human body through membrane in unionized form. In this way, the
importance of a particular drug is directly dependent upon the lipophilicity, state and
PH level of the drug. Different parts of the human body possess different PH level.
Because of this the drugs for different body part must have different ionized states.
Further, determination of acid dissociation constant is the other important
usage of partition coefficient. Henderson-Hasselbalch equation can be used for the
calculation of such acid dissociation constant (Ka). According to the equation :
Ka = [H+][A-] / [HA]
where [H+] = concentration of hydrogen ions
[A-]= concentration of ionized acid
[HA]= concentration of unionized acid
Objective of the experiment
The experiment was conducted to determine the overall impact of different PH levels
and and degree of ionization over coefficient of salicylic acid. With the help of the
partition coefficient of a weak dissociating system, the determination for salicylic acid
(pKa) can be obtain by following formulas:
P = CO / [HA]
Ka = [H+][A-] / [HA]
P’ = CO / ([HA] + [A-]) = CO/CW
where p' = “apparent” partition coefficient which is obtained from experiment
with the help of above three equation a combined equation can be derived as:
1/P’ = 1/P + Ka/[H+]P
where P = partition coefficient
Ka = dissociation constant
CO = total concentration in oil phase
CW = total concentration in aqueous phase
with the help of different concentrations of Salicylic acid, different values of
Chemistry 4
absorbance of the acid can be derived. Such values are very useful in constructing a
calibration curve and a graph demonstrating the concentration of the acid in aqueous
phase. In the graph which is shows the values of 1/P' against the [1/H+], a line can be
yielded which is having the 1/P as an intercept and Ka/P as a slop (Hodgson, 2004).
Methods
The experiment was conducted in to two parts :
Part I determination of concentration of salicylate ions for producing the curve of
calibration.
For reveling the level of absorbance of salicylate ion in the solution at
different concentration, the experiment was done. With the help of results of the
experiments a calibration curve was drawn.
In the experiment five test tubes were used. 6 ml water was taken in in one test
tube which was used as a blank solution. In other test tubes, the solution of 2 ml ferric
nitrate in 5ml water was taken. Further, 1 ml salicylic acid was added to the test tubes
at different concentrations for example 0.00125M, 0.0025M, 0.00375M, and 0.005M.
For determining the absorbance of salicylic acid in each test tube, a calorimeter was
set to the wavelength of 624 nm. After this each sample was inserted in to calorimeter
with the help of cuvettes and absorbencies were read.
The values retrieved from such experiment ware plotted in a graph. The equation of
the line in the grpah will be used in determining the concentration of salisylate in the
next step of experiment.
Part B: measurement of Salicylate ion concentration in the solution of four different
PHs.
The experiment is conducted into pairs in which two 100 ml volumetric flask
were used. In each of flasks, 10 ml solution of 0.2 M sodium salicyalte was pipetted.
Chemistry 5
In these flasks
four different buffers with pH level of 2.6, 3.0, 3.4, and 3.8 was used in the
experiment. Two of the four different buffer solutions were added in two flasks. Such
mixture of solutions in each of flasks were shaken and mixed properly. Further, about
25 ml solution from each of flasks was taken out in a separate funnels. In each funnel,
about 25ml of Tesco sunflower oil was added. After this, the funnel was left for
settling the mixture so that two different levels of solution can be sighted easily. This
process was repeated with each funnel. The level of pH level of each solution was
measured. After having such measurement, 10 ml solution was mixed with water in a
50 ml volumetric flask. 1 ml of this dilute solution was pipetted out in the 2mL of
ferric nitrate solution and 5mL of water. then absorbance of the solution was
measured.
After this measurement, two flasks were washed out with detergent and than
the whole process was repeated by using remaining two buffer solutions.
Results
Table 1: Absorbance of Salicylate at Different Concentrations
Concentration of Salicylate
0.00125M
0.00250M
0.00375M
0.00500M
Absorbance (624nm)
0.125
0.253
0.378
0.502
Graph A
Table 2: pH and Absorbance Readings of 0.2M Sodium Salicylate with Different
Buffers
Buffer pH
pH of Solution after
Separation
Absorbance
2.6
3.0
3.4
3.8
Chemistry 6
0.258
0.318
0.328
0.353
3.34
3.56
3.96
4.07
Table 3: Apparent Partition Coefficient at Different Hydrogen Ion Concentrations
1/[H+]
1/P’
2187.7616
1.7972
3630.7805
3.8193
9120.1089
4.5937
11748.9756
7.3333
Graph B
Calculations
With the help of the slope equation from the graph the concentration of
salicylate ion in the water can be determined as below:
y = 100.4x + 0.000
0.113 = 100.4x
x1 = 0.00257M
x2 = 0.00317M
x3 = 0.00327M
x4 = 0.00352M
The initial concentration of salicylate after incorporating the dilution factor of 10:
0.2M/10 = 0.02M
The concentration of salicylate in oil can be determined by:
Co = Cw - Ca
Where: Co is the concentration of salicylate in oil
Cw = initial concentration of salicylate ion
Ca = Concentration of salicylate in aqueous phase.
Chemistry 7
0.02 = Cw + Co
Co1 = 0.02 – (x1 * 5)
Co1 = 0.02 – (0.00257 * 5)
Co1 = 0.00715M
C02 = 0.00415M
Co3 = 0.00356M
Co4 = 0.00240M
the apparent partition coefficient P’ = Co/Ca
thus
1/P’1 = Ca1/Co1
1/P’1 = 0.01285/0.00715
1/P’1 = 1.79720
1/P’2 = 3.81928
1/P’3 = 4.59370
1/P’4 = 7.33333
with the help of equation pH = -log10[H+
the inverse concentration of proton in the solution can be calculated as:
1/[H+]1 = 1E-pH
1/[H+]1 = 1E-3.34
1/[H+]1 = 2187.7616
1/[H+]2 = 3630.7805
1/[H+]3 = 9120.1089
1/[H+]4 = 11748.9756
(E = 1 x 10x)
form the equation y = 0.0005x + 1.2419, p can be determined as below:
1/P = 1.2419
P = 0.8052
Using P, Ka can now be calculated:
m = Ka/P
Ka = m x P
Ka = 0.0005 x 0.8052
Ka = 4.0261E-04
Chemistry 8
Consequently,
pKa = -log10Ka
pKa = -log10(4.0261E-04)
pKa = 3.40
% Error from literature for Ka = 0.001000 mol/dm3 and pKa = 3.00
% Error of Ka = |[(Obtained – Actual)/ Actual]| x 100%
% Error of Ka = |[(4.0261E-04 – 1.0000E-03)/1.000E-03]| x 100%
% Error of Ka = 59.74%
% Error of pKa = [(Obtained – Actual)/ Actual] x 100%
% Error of pKa = [(3.40 – 3.00)/3.00] x 100%
% Error of pKa = 13.33%
Discussion:
After having a thorough analysis of results, it can be discussed that the
solubility of a drug in the aqueous phase has a inverse relationship with the solubility
of that drug in a non aqueous solution such as oil. With the help of different graphs a
linear relationship of concentration of salicylate ions and its absorbencies in different
solutions can be proved. Such fact is also backed by Beer Lambert law. According to
the law, there is an empirical relationship is found between the absorbance of light and
the properties of the material in which the light is traveling. In the lights of results of
experiments it can be implied that the pH level of the aqueous layer in the mixture
increases with the increase in the pH in the solution. Further, the results of the
experiments also show that the value of apparent partition coefficient decreases with
the increases in the degree of ionization of salicylate ion. This results was confirming
the theory of higher lipophillcity which states that an unionized drug can passes
through the biological membrane, more easily in compared to ionized drug. After
being ionized, the overall partition coefficient of the drug decreases and consequently
its volatility through the biological membrane gets affected adversely.
Chemistry 9
In the experiment when the pH is increased from 2.6 to 3.8, the concentration
of the solution was also increased from 0.00257M to 0.00352M. This is showing that
higher the PH of the buffer solution, higher will be the concentration of the salicyalte
ion in the solution. Furthermore, the values of partition coefficient were 0.556, 0.262,
0.218, and 0.136 at different pH level i.e. 2.6, 3.0, 3.4, and 3.8 which showing an
inverse relationship between in these two entities. The experiment also backs the fact
that in comparison to the solution having higher pH values, an unionized drug ionized
better in the basic conditions.
Discussion of errors in the experiments
there are two major sources of errors in an experiments namely human and
non-human experimental or instrumental errors. In the experiment the most possible
human error can be incurred because of the inaccurate separation of two different
layers of solution. Such type of inaccuracy may occur due to not giving enough time
to solution because of which the proper separation does not take place. This improper
separation can cast an adverse impact over the ionization process of salicylate ion in
the non-aqueous phase of solution.
Further the next source of error is instrumental errors which can be arose due
to lack of cleanliness of the cuvettes and separating funnels used in the experiment.
The uncleanliness of such apparatuses can present difficulties in the reading of
absorbencies of the solution. Moreover, the consistency of the temperature of the
room in which the experiment is conducted is also an important source of error. This
can affect the value of partition coefficient during the experiment. At last, the
blanking of calorimeter is also an important reason which can affect readings of
absorbence of the solutions.
The theoretical values of ka and pH of Salicylic acid are 2.75E-05 and 3.00
Chemistry 10
respectively (Reynolds, 2005). in comparison to this the value of Ka and pH of
salicylic acid retrieved form the experiment are 4.0261E-04 and 3.40 which is
showing the errors of 56.74% and 13.33% respectively. This observation implies that
there is an experimental error in the collection of data.
Importance of partition coefficient
This partition coefficient is used for determining the lipophilicity and
hydrophobicity of a particular drug in the human body. The lipophilicity and
partitioning of a particular drug is very helpful for identifying the areas of human
body from where it can pass through. The partition coefficient is very important in the
pharmaceutical science as it is used in determining and identifying the proper
formulation of the drug for a particular diseases. Such fact can be seen in the drugs
which is taken orally, and are absorbed in the small intestine. Literatures are
acknowledging that in the stomach the pH level is very low. Because of this reason,
the drug cannot be ionized. Thus it can be implied that the basic drugs are absorbed by
intestine more easily in comparison to the acidic drugs (Borchardt, 2004).
Alternative methods for measuring the partition coefficient
there are different other methods such as automatic titration, chromatography
with electrometrics, aqueous titration and Sirius Potentiometric method available for
the measurement of the partition coefficient. Furthermore a filter probe device can
also be used for measurement of pKa and Log P (Earll, 1999).
for measuring log p of the solution, '1-octanol/water' system is used because it
is very useful in the measurement of water absorption. 1-octonol water solvent
provides a platform where salicyate ions can be easily ionized and can move from one
phase to another (Sangster, 1997). The methods of taking any drug such as rectal,
nasal, topical applications, and parenteral are heavily dependent over the partitioning
Chemistry 11
properties of a particular drug in the body. The selection of the method of drug
delivery is dependable upon the compatibility of the drug with the mucus layer as well
as lipid bi-layer of human body. In this way, the partition coefficient and lipophillicty
of a drug plays an vital role in pharmaceutical science.
For optimizing the log p of the drug there are a number of factors such as
absorbence level of drug, pH level of the medium, partition properties of the drug,
which are needed to be considered in the formulation of a particular drug. For
enhancing the effectiveness of a drug it is necessary to have a proper understanding of
different aspects of drug such as where it will be absorbed in human body, where in
the human body the process of partitioning will take place, what will be the chemical
structure of the drug and what are level of lipophilicity and hydrophobicity of that
particular drug (Flanagan, Taylor and Watson, 2007).
Conclusion
After having an intense data analysis, the partition coefficient of the used
solution was found 0.8052. The pKa , and the Ka of that solution were 3.40, and
4.0261E-04 respectively. Form the analysis it can be concluded that in-spite of having
high degree of errors in the measurement of Ka and the pKa i.e. 9.74% and 13.33%
respectively, the methods used for the experiment is effective.
Chemistry 12
References
Borchardt, R.T. (2004).Pharmaceutical profiling in drug discovery for lead selection.
Springer.
Earll M. (1999) A guide to Log P and pKa measurements and their use. Retrieved
February 22, 2011, from
http://www.raell.demon.co.uk/chem/logp/logppka.htm .
Flanagan, R.J., Taylor, A, and Watson, I.D. (2007). Fundamentals of analytical
toxicology. Wiley-Interscience.
Hodgson, E. (2004). A Textbook of Modern Toxicology. (4th ed.). John Wiley and
Sons.
Reynolds J. Martindale: The Complete Drug Reference (34th edi.). London:
Pharmaceutical Press.
Sangster, J. (1997). Octanol-water partition coefficients: fundamentals and physical
chemistry. John Wiley and Sons.