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P ka and log p presentation

This document discusses pKa (acid dissociation constant) and log P (partition coefficient). It defines pKa as the negative base 10 logarithm of Ka, which is a quantitative measurement of acid strength. Common methods to determine pKa include potentiometric titration and spectrophotometric methods. Log P is the ratio of concentrations of a compound between octanol and water, and is a measure of lipophilicity. Common methods to determine log P include the shake flask method and HPLC. Both pKa and log P values are important parameters in drug design and development as they influence absorption, distribution, and other pharmacokinetic properties.

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‘pKa and log P determination’ Karnataka College Of Pharmacy Bangalore-64 By- Pramod Sarraf 1
pKa (Logarithmic acid dissociation constant) Introduction: • Understanding the concept of ‘Ka’ before ‘pKa’ • Ka or acid dissociation constant is a quantitative measurement of the strength of an acid in solution. • Let us consider the dissociation of the compound ‘HA’ HA ⇌ A- + H+ The Ka for this reaction will be given by: Ka = [A−][H+] [HA] • Expressing acidity in terms of Ka can be inconvenient for practical purposes, therefore, pKa is used. • pKa can be defined as ‘the negative base-10 logarithm of acid dissociation constant (Ka) of a solution’. pKa = -log10Ka • Example:  The Ka constant for acetic acid is 0.0000158, but the pKa constant is 4.8, which is a simpler expression.  The smaller the pKa value, the stronger the acid.  The pKa value of lactic acid is about 3.8, so that means lactic acid is stronger than acetic acid. 2
3 • A weak acid has a pKa value in the approximate range of -2 to 12 in water. • Acids with a pKa value of less than about -2 are said to be strong acids. Methods of pKa determination: 1. Potentiometric titration- • In potentiometric titration, a sample is titrated with acid or base using a pH electrode to monitor the course of titration. • The pKa value is calculated from the change in shape of the titration curve compared with that of blank titration without a sample present. • Relationship between pH and pKa: pH = pKa+log10 [A−] [HA] Fig.1. Titration curve of pH vs volume of titrant.
4 2. Spectrophotometric methods- • Main advantage is higher sensitivity. • The compound must contain a UV-active chromophore close enough to the site of the acid–base function in the molecule. • Spectral data are recorded continuously during the course of titration by a diode-array spectrometer. • The absorption spectra of the sample changes during the course of the titration to reflect the concentration of neutral and ionized species present. • The largest change in absorbance occurs at the pH corresponding to a pKa value. • The determination of pKa values by UV–VIS assumes that the solute of interest is pure or that its impurities do not absorb in the UV–VIS range, since the spectra of impurities can overlap with those corresponding to the solutes of interest. 3. NMR titration- • The protonation of a basic site leads to electronic deshielding effects on the adjacent NMR-active nuclei. • The average chemical shifts of all the measurable NMR-active nuclei, as a function of pH, are expected to reflect the fractional protonation of each basic group of a molecule. • NMR titrations have not yet been applied to the pKa determination of APIs.
5 3. Liquid chromatography (LC)- • LC is used as a powerful technique for the determination of dissociation constants, as it requires only a small quantity of compounds. • Studied samples do not need to be pure and poor water solubility is not a serious drawback. • This method does not include measuring solute or titrant concentrations, just retention times. • Also, calculation is straightforward and independent of solute purity. 4. Computational methods- • Theoretical pKa values can be calculated by computational methods (e.g., SPARC and and ACD/Lab). • ARChem’s SPARC software is an on-line calculator that estimates the macroscopic and microscopic pKa of any organic compound solely from its chemical structure. • ACD/Lab pKa is a software program that calculates accurate acid-base ionization constants (pKa values) under 25°C and zero ionic strength in aqueous solutions for almost any organic structure.
6 Applications of pKa: • The pKa values of proteins and amino acid side chains are of major importance for the activity of enzymes and the stability of proteins. • Buffer solutions are used extensively to provide solutions at or near the physiological pH for the study of biochemical reactions; the design of these solutions depends on a knowledge of the pKa values of their components. • In pharmacology, ionization of a compound alters its physical behaviour and macro properties. This is exploited in drug development to increase the concentration of a compound in the blood by adjusting the pKa of an ionizable group. • Knowledge of pKa values is important for the understanding of coordination complexes, which are formed by the interaction of a metal ion with a ligand. • In chemical oceanography in order to quantify the solubility of iron(III) in seawater at various salinities, the pKa values for the formation of the iron(III) hydrolysis products Fe(OH)2+, Fe(OH)+ 2 and Fe(OH)3 are determined.
7 pKa determination for aspirin: Requirements- Aspirin, ethanol, sodium hydroxide, pH meter. Principle- Aspirin is a weak acid and partially ionizes in water. HA + H2O ↔ H3O+ + A- It’s acid dissociation constant, Ka is given by: Ka = [A−][H3O+] [HA] Aspirin and sodium hydroxide react in a 1:1 mole ratio: CH3OCOC6H4COOH + NaOH → CH3OCOC6H4COONa + H2O Method- • A burette is filled with 0.1M sodium hydroxide solution. • 0.36g of aspirin is weighed in 250ml beaker and 10ml of 95% ethanol is added and volume is made up with deionized water. • 2ml portions of sodium hydroxide solution is added from burette to the beaker, stirring well between each additions and recording the pH using a pH meter. • The pH begins to rise rapidly near the end-point. • After adding 18ml of sodium hydroxide solution, addition is continued in 0.5ml portions. • After adding about 22ml, additions in 2ml portions is started again. • The addition is continued until total of 36ml has been added.
8 Processing data- • A graph of pH against volume of 0.1M sodium hydroxide is plotted. • The end-point is calculated from the graph of titration. • This is checked against the expected value by calculating the number of moles of aspirin used (relative molecular mass of aspirin = 180) and therefore the volume of 0.1M sodium hydroxide solution needed to react with it in a 1:1 mole ratio. • From the graph, the pH is estimated at half-way point of the titration. This gives a value for the pKa of aspirin. Volume of 0.1M sodium hydroxide Fig.2. Titration graph for aspirin against 0.1M sodium hydroxide solution.
9 Log P (Logarithm of partition-coefficient) Introduction: • Partition coefficient, abbreviated as P, is defined as a particular ratio of the concentrations of a solute between two immiscible solvents at equilibrium. • The logarithm of this ratio is thus ‘log P’. log Pow = log [unionized solute]o [unionized solute]w • This ratio is therefore a measure of the difference in solubility of the compound in these two phases. • The partition-coefficient generally refers to the concentration ratio of unionized species of the compound. • When one of the solvents is water and the other is a non-polar solvent, then the log P value is a measure of lipophilicity or hydrophobicity. Choice of solvents: • The choice of partition solvent has been subject to debate in recent years. • The most commonly used solvent has been octan-1-ol or octanol. • Octanol was chosen as a simple model of a phospholipid membrane.
10 • Hydrophobic drugs with high octanol/water partition coefficients are mainly distributed to hydrophobic areas such as lipid bilayers of cells. • Hydrophilic drugs with low octanol/water partition coefficients are found primarily in aqueous regions such as blood serum. • If one of the solvents is a gas and the other a liquid, a gas/liquid partition coefficient can be determined e.g. the blood/gas partition coefficient of a general anesthetic measures how easily the anesthetic passes from gas to blood. Component log Pow(octanol/water) T (°C) Acetamide -1.16 25 Methanol -0.81 19 Formic acid -0.41 25 Diethyl ether 0.83 20 p-Dichlorobenzene 3.37 25 Hexamethylbenzene 4.61 25 Table 2. Partition coefficient data of some compounds.
11 Measurement of log P: 1. log P by Shake Flask- • The shake flask method is the oldest and most tedious way of measuring log P values. • Equimolar quantities of drug are added into equal volumes each of n-octanol and water. • It is very important to pre-saturate the solvents in prolonged shake-flask experiments. • The experiment must be performed over 3 days or more to ensure equilibrium is reached. • The phases are separated and absorbance is measured and log P is calculated from formula for P. 2. log P by HPLC- • Compounds with known Log P's are injected onto a C18 reverse phase HPLC column and their capacity factors used to create a calibration curve. • Unknown compounds are then injected and their capacity factors used to predict log P. 3. pH metric- • This technique determine lipophilicity pH profiles directly from a single acid-base titration in a two phase water-organic solvent system. • Hence, a single experiment can be used to measure the log P.
12 4. Prediction- • QSAR (Quantitative Structure Activity Relationship) can be used to derive partition coefficients. • Estimates of partition coefficients can be made using a variety of methods including fragment based, atom based and knowledge based that rely soley on knowledge of the structure of the compound. Optimum log P values for certain classes of drugs: • Optimum CNS penetration - log P = 2±0.7 • Optimum oral absorption - log P = 1.8 • Optimum intestinal absorption - log P =1.35 • Optimum colonic absorption - log P = 1.32 • Optimum sub lingual absorption - log P = 5.5 • Optimum percutaneous absorption - log P = 2.6  For reasonable absorption log P should be below 5
13 Applications of log P: • In pharmacology, log P strongly affects how easily the drug can reach its intended target in the body, how strong an effect it will have once it reaches its target and how long it will remain in the body in an active form. Hence, It is an essential criteria in pre-clinical drug discovery for assessment of new drug candidates. • In pharmacokinetics, log P has a strong influence on ADME properties of a drug. • In environmental science partition coefficient can be used to predict the mobility of radioactive compounds in groundwater and hence study water pollution and its toxicity to animals and aquatic life. • In metallurgy, partition coefficient is an important factor in determining how different impurities are distributed between molten and solidified metal. • In FMCG, partition coefficient is taken into account e.g.in the formulation of topical ointments, dyes, hair colors, etc.
14 Log P determination of Levofloxacin: Determined by shake flask method in n-octanol:water system 10mg of drug is added into 50ml each of n-octanol and water The mixture is shaken for 24 hours until equilibrium is reached Phases are separated using a separating funnel, filtered and suitably diluted Absorbance is measured at 258nm using UV spectrophotometer Partition coefficient (P) is calculated using: P = Co Cw and hence log P can be calculated. NOTE: Using similar techniques for log P determination of levofloxacin, log P can be found out for salicylic acid, various steroids, antibiotics, etc.
15 Log P determination of Permethrin: Method- Reverse phase HPLC Preparation of diluent- Methanol and water is taken in the ratio 95:10 v/v. Preparation of reference standard solution- Equimolar solutions of thiourea, triphenylamine, phenanthrene, naphthalene, phenol and methyl benzoate are prepared into different 10ml volumetric flasks, using the above diluent solution. Preparation of permethrin sample solution- The solution is made using 2.95mg of permethrin in 10ml volumetric flask, using the diluent solution. Determination of log P- The capacity factor, k´ for each reference standard and test item can be calculated for the HPLC system from their respective retention times. k´ = (𝑡𝑅−t0) t0 Where, k´= capacity factor 𝑡𝑅 = retention time (min) t0 = dead time (min) • A correlation graph is generated of ‘log k´’ vs ‘log Pow’ for the reference standards. • The log Pow for permethrin is the interpolated from the correlation graph. NOTE: • Capacity factor, k´ - It is an indication of how long a compound can be retained by the stationary phase. • Retention time, 𝑡𝑅 - It is the measure of time taken for a solute to pass through a chromatography column. • Dead time, t0 - It is the elution time of an unretained component
16 Fig.3. Calibration curve of known reference standard log k´ vs log Pow.
17 REFERENCES:  Svehla G. Vogel’s Textbook of Macro and Semimicro Qualitative Inorganic Analysis. Longman Group Limited, London. 1975 (5). 28-31:130-131.  Babić S, Horvat A. J. M, Pavlović D. M and Kaštelan-Macan M. Determination of pKa Values of Active Pharmaceutical Ingredients. Trends in Analytical Chemistry, 2007, 26 (11). 1043-1060.  Wikipedia contributors. (2018, August 31). Acid dissociation constant. In Wikipedia, The Free Encyclopedia. Retrieved 13:24, September 1, 2018, from https://en.wikipedia.org/w/index.php?title=Acid_dissociation_constant&oldid=857405027 .  Wikipedia contributors. (2018, June 14). Partition coefficient. In Wikipedia, The Free Encyclopedia. Retrieved 19:51, September 1, 2018, from https://en.wikipedia.org/w/index.php?title=Partition_coefficient&oldid=845795904.  Mark E. A Guide to log P and pKa Measurements and Their Use. Mark’s Analytical Chemistry, 2006.  Shahwal V. K, Dubey B. K and Bhoumick M. Preformulation Study of Levofloxacin. International Journal of Advances in Pharmaceutics, 2012, 1(1). 1-8.  Rao T. N, Srinivasarao T and Botsa P. A Simple Reverse Phase HPLC Method for Determination of Partition Coefficient of Permethrin Pesticide. Chromatography, 2016, 1 (1). 1-5.  Jeffery G. H, Bassett J, Mendham J and Denny R. C. Vogel’s Textbook of Quantitative Chemical Analysis. Longman Scientific & Technical, England. 1989 (5). 31-33:162-163.
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