Elements of the Partition Coefficient - Research Paper Example

 Introduction Partition coefficient is the ratio of two-phased solute concentration. As a constant, it is defined as the concentration of solute in solvent A (CA) divided by the concentration of solute in solvent B (CB) expressed in this formula: P = CA / CB . A solute is a soluble component mix with immiscible liquids to achieve equilibrium of constant temperature. In an experiment, researchers used oil and water as solvents and salicylic acid was used as solute. The latter determined the partition coefficients between a non-aqueous phase (oil) and an aqueous phase (water). Thru partition, coefficient was expressed with the concentration in the non-aqueous or lipophilic phase as the numerator. This was expressed in a formula “P = Coil / Cwater.” In that context, the drug’s absorption can be predicted using the partition coefficient. In logarithmic partition, coefficient values are calculated to determine the solute’s hydrophocity, lipophilicity and it’s anent absorbability. Since body membranes are usually impermeable to foreign ionic species, only un-ionized drug molecules can partition into the membrane. Hence, the lipophilicity correlates with the state of the drug including its acidity or pH level. Note that pH levels vary in all part of the body which implies that drugs partitioned in a specific part will also have diverse un-ionized states. “Partition coefficient’ also determines the acid dissociation constant or Ka. This can be calculated using the equation of Henderson-Hasselbalch where the “product of hydrogen ion ([H+]) and ionized acid ([A-]) concentrations” is divided by the “concentration of unionized acid ([HA]). This formula is expressed with the following: Ka = [H+][A-] / [HA]  This experiment aims to determine the different pHs and degree of ionization affected the partition coefficient of salicylic acid. Inspired by “partition coefficient” for weakly dissociating systems, the pKa determination of salicylic acid can be resolved using the following equations:  P = CO / [HA]  Ka = [H+][A-] / [HA]  P’ = CO / ([HA] + [A-]) = CO/CW  In this formula, P’ is the “apparent” partition coefficient obtained experimentally. The combination such three equations would result to  1/P’ = 1/P + Ka/[H+]P  of which P is partition coefficient  Ka is dissociation constant  CO is total concentration in oil phase; and CW is total concentration in aqueous phase. Researchers will also obtain the “absorbance values” using different concentration of salicylic acid because this is relevant in setting a calibration curve and a graph to reflect the concentration in the aqueous phase. Moreover, a graph plotting 1/P’ against the reciprocal of hydrogen ion concentration (1/[H+]) yielded a line with an intercept of 1/P and a slope of Ka/P. Using the partition coefficient and the dissociation constant, the pKa was calculated.  Methodology This experiment comprised of two parts: (a) determination of salicylate concentration to produce a calibration curve and (b) salicylate concentration measured in solutions of four different pHs. a. Salicylate Concentration Determination to Produce a Calibration Curve  It aims to resolve the absorbance of salicylate solution at different concentrations to produce a calibration curve. As part of the method, researcher will use five test tubes. Test Tube 1 will contain 6mL of water which will be used as the blank solution to keep the calorimeter at zero. Other test tubes will contain 5mL of water which is added to 2mL of ferric nitrate solution. Further, 1mL of different concentrations of salicylate was added to test tube 1; 0.00125M for test tube 2; 0.0025M for test tube 3; 0.00375M for test tube 4; and 0.005M for test tube 5. With calorimeter set at a wavelength of 624nm, the absorbance of each test tube of salicylate solution was determined by pouring solution in cuvettes and inserting them into the calorimeter. Data will be figured in a graph. Meanwhile, the equation of the line was used to know the concentrations of salicylate seen in part B.  b. Salicylate Concentration Measured in Solutions of Four Different pHs  In duo, 10mL of 0.2M sodium salicylate solution was pipetted into each of the two 100mL volumetric flasks and with two of the four buffer solutions supplied (pH of buffers used was 2.6, 3.0, 3.4, and 3.8 respectively). Contents of the flask were mixed and 25mL of each solution was pipetted into separating funnel. Tesco Sunflower Oil will be added in this funnel and be mixed again for 10 minutes. The funnel was clamped until the contents settled and two different layers were visible.  The lower aqueous layer was run into a beaker. This was repeated for each funnel. The solution’s pH are thereafter measured followed by the dilution of a five-fold by pipetting 10mL of the solution into a 50mL volumetric flask of water. 1mL of this diluted solution was mixed with 2mL of ferric nitrate solution and 5mL of water. Thereafter, the absorbance was measured.  After the aqueous layers of the first two buffers are assayed, both flasks were thoroughly washed with water and detergent. This experiment was repeated for the remaining two buffer solutions.  Results Table 1: Absorbance of Salicylate at Different Concentrations  Concentration of Salicylate Absorbance (624nm)  0.00125M 0.00250M 0.00375M 0.00500M 0.125  0.253  0.378  0.502 Graph A  Table 2. pH and Absorbance pH and Absorbance Readings of 0.2M Sodium Salicylate with Different Buffers Buffer pH pH of Solution after Separation Absorbance 2.6 3.34 0.258 3.0 3.56 0.318 3.4 3.96 0.328 3.8 4.07 0.353  Table 3: Apparent Partition Coefficient at Different Hydrogen Ion Concentrations  1/[H+] 1/P’  2187 3630 9120 11748 .7616 .7805 .1089 .9756 1.7972 3.8193 4.5937 7.3333 Graph B  Calculations  Using the slope equation of line of best fit from the calibration curve, concentration of salicylate in water can be determined thru this formula: 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 being mixed has the dilution factor of 10 is 0.2M/10 = 0.02M . This is multiplied by 5 as a five-fold dilution. The salicylate’s concentration in oil can be determined by the formula:  Co = Cw - Ca where Co is the salicylate’s concentration in oil; Cw is the initial concentration of salicylate; and Ca is the concentration of salicylate in aqueous phase. This can be figured as follows: 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  Then the apparent partition coefficient (P’) can be found by: P’ = C¬o/Ca and further the inverse can be found by calculating 1/P’. 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  Using the equation pH = -log10[H+] and the pH of the solution after separation, the inverse proton concentration ([H+]) is calculated this way, 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)  From the equation y = 0.0005x + 1.2419, P can be determined using the y-intercept:  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  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  Following the result of this experiment, it can be inferred that the drug solubility in the aqueous phase is inversely proportional to the drug solubility in the non-aqueous/lipophilic phase. It also bared the linear relationship with the concentration of salicylate and its absorbance. This affirmed the Beer Lambert Law which pointed the empirical relationship of the absorbance of light to the properties of the material through which the light is traveling. Focusing on the relation of ‘pH of the solution’ at different salicylate/acidic concentrations, it also bared that as the pH of the solution increases, so does the pH in the aqueous layer. It was also noted that as the pH of the solution at the degree of ionization of salicylate increases, there is decrease in the value of the apparent partition coefficient. This is attributed to partitioning of the drug, and since it was unionized, it crossed from one phase to the other like passing through a biological lipid membrane. It can therefore be concluded that higher lipophilicity allow unionized drugs to cross easily at biological membranes compared to ionized drugs. This is showed when the pH increased from 2.6 to pH 3.8, concentrations increased from 0.00257M to 0.00352M respectively. Further, it also indicates that as the pH increases, more ionization of the salicylate occurs, hence the increase of salicylate’s concentration as the buffer solutions increased. Moreover, the apparent partition coefficient values showed pHs at 2.6, 3.0, 3.4, and 3.8 were 0.556, 0.262, 0.218, and 0.136 respectively. It bared the increase in pH that is directly proportional to the increase in dissociation of the unionized salicylate. Also, lipophilic activity decreased as the pH increased due to unionized drug ionizing. The decrease of the apparent partition coefficient occurred relative to the increase of pH. Records state that the Ka and the pH of salicylic acid are 2.75E-05 and 3.00 respectively [1]. The values that were calculated through this experiment were a Ka of 4.0261E-04 and a pH of 3.40. These values had a percent error of 56.74% and 13.33%, respectively, and may have figuratively resulted to experimental errors. Although the pH is relatively close to that of the literature, caution could have impeded percent error.  Partition coefficient is not only used to determine the partitioning of a particular drug but also the lipophilicity and hydrophobicity of particular drug. In reference to the biology, partitioning and lipophilicity of the drug signify where it will separate in the body and when it can cross biological membranes which are directly proportional to solubility of the drug. This shows that partition coefficient plays a vital role in pharmaceutical sciences to determine the relevance of drug. This is exemplified by the fact that medicines taken are digested into the body for absorption. Data revealed once drug reaches the stomach, its acidity hinder its ionization. Therefore since the stomach easily ionizes acidic drug than basic drugs, the lipophilicity of the drug would have to be more basic. Hence, slightly basic drug can be greatly absorbed in the intestine than in the stomach.  There are other methods than the shake-flask in measuring partition coefficients. These are automatic titration and chromatography and the usage of a Sirius Potentiometric method [2]. Another method is the use of a filter probe device where a continuous draw off of a particular phase is achieved.  Conclusion In conclusion, the experimental methods pointed that the partition coefficient, the pKa , and the Ka were found at 0.8052, 3.40, and 4.0261E-04 respectively. Although errors were observed in the Ka and the pKa which is relatively high at 59.74% and 13.33% respectively, but still the experiment was generally successful.     References  1. Reynolds JEF. Martindale: The Complete Drug Reference. 34th Edition. 2005. London: Pharmaceutical Press.  2. Earll M. A guide to Log P and pKa measurements and their use. CChem MRSC. 1999. Available through: http://www.raell.demon.co.uk/chem/logp/logppka.htm. Read More