Non aqueous Titration theory and principles ppt

1. Principles of Non-Aqueous Titration & Application in Pharma Industries Dr. A. Amsavel M.Sc., B.Ed., Ph.D.

2. An Overview Introduction Non-Aqueous Titration Acid - Base Theory Solvents for NAT Indicators & Potentiometric End Point Acid & Base Titration Pharmacopeia USP <541> Potentiometric Titration Precaution To Be Taken Estimation Of Errors And Elimination Of Errors

3. Introduction Titrimetry / Volumetric Analysis: Volumetric analysis is performed for Quantitative determination of assay / content. It is simple and commonly used technique in Chemical Industries. Analysis conducted in Aqueous and non-aqueous medium. ◦ Simple and easy ◦ Fast and can be done on site ◦ Less expensive ◦ Estimation of content or Assay of chemical ◦ Precise and Accurate - depends on method & Specificity

4. Limitation of Aqueous Titration Titration in Water medium has limitations: To determine the substances which are insoluble in water. To titrate of week acids or weak bases To titrate separately for a mixture of acids with nearer dissociation constants. Similarly a mixture of bases. The substances, which are either to weakly acidic or too weakly basics to give sharp end point in water The above can be overcome by non-aqueous titration (NAT). It is easy to perform and get accurate result.

5. Non-Aqueous Titration (NAT) Non Aqueous Titration: Titration performed in solvent medium which does not contain water. Substance is dissolved in a solvent and titrated using acid or base as titrant. Principle & theory are almost same as Aqueous titration Reaction is carry out in non-aqueous medium Extensively used for organic acids and bases, nitrogen containing compunds Principle is based on Brønsted-Lowry Theory

6. Where to Use NAT Advantage of NAT: ◦ Titration of week acids or weak bases, by improving reactivity of low or poor reactive substances To titrate separately for a mix of acids or bases with near dissociation constants. To determine the substances which are insoluble in water. Titration of chemicals which shows poor or not giving sharp end point in aqueous titrations (weakly acidic or basic substances)

7. Advantages of NAT Organic acids and bases that are insoluble in water can be tested using appropriate solvents Very week acid and bases can be analysed in NAT Principle of NAT is same as aqueous titration. Mixture of week acids can be tested in single or mixture of solvents. Biological ingredients (acidic or basic) can be selectively titrated using proper solvents. Eg. Nitrogen containing compounds Non aqueous titrations are simple and accurate. Pharmacopeias recommend non-aqueous titration; Eg. Ephedrine , codeine phosphate, tetracycline, piprazine Anti-histamines etc

8. What are Acids & Bases Acids: ➢Arrhenius acid: Any substance that, when dissolved in water, increases the concentration of hydronium ion (H3O+) ➢Bronsted-Lowry acid: A proton donor; conjugate base ➢Lewis acid: An electron acceptor Bases: ➢Arrhenius base: Any substance that, when dissolved in water, increases the concentration of hydroxide ion (OH-) ➢Bronsted-Lowery base: A proton acceptor ; conjugate acid ➢Lewis acid: An electron donor

9. Brønsted-Lowry Theory The conjugate acid of a base is the base plus the attached proton and the conjugate base of an acid is the acid minus the proton + A - HB+ B + HA Base proton Acid Acid Receiver Proton source Proton source Receiver Base Proton

10. Equilibrium Constant Kaand Kb The Equilibrium Constant for a Brønsted acid is represented by Ka, and base is represented by Kb. CH3COOH(aq) + H2O (l) H3O+(aq) + CH3COO–(aq) [H3O+][CH3COO–] Ka= ––––––––––––––––– [CH3COOH] Notice that H2O is not included in either equilibrium expression. NH3(aq) + H2O(l) NH4+(aq) + OH–(aq) [NH4+] [OH–] Kb= ––––––––––––– [NH3] pH of 1M ACOH =2.4

11. Solvent Selection for NAT Solvents, which are used is non-aqueous solvent, and plays major role in NAT analysis. Classified as four types: 1. Aproticsolvents:Chemically neutral 2. Protogenicsolvents:Acidic nature readily donate protons. 3. Amphiprotic solvent:Which are slightly ionize and donate and accept protons. 4. Protophilcsolvents: Posses high affinity for protons.

12. Aprotic Solvents Aprotic Solvents: ◦ Solvents are Chemically Neutral, ◦ Un-reactive under the titration conditions; do not undergo reactions with acids and bases ◦ They possess low dielectric constants. ◦ Do not cause ionization in solutes and ◦ Aprotic solvents are frequently used to dilute the reaction mixture and enhance the end point. Eg. Toluene, carbon tetrachloride , acetonitrile, benzene, and chlorinated hydrocarbons

13. Protogenic Solvents ProtogenicSolvents: ( Protogenic – Proton producing) Solvents which are more acidic than water. It is Acidic in nature and readily donate protons. Also called as Acidic solvents. Strength / ability to donate protons enhance the strength of weak bases. Since more acidic character of solvents , tend to have a leveling effect on the bases when used. Eg . Anhydrous Hydrogen fluoride , Sulphuricacid, Formic acid , acetic acid etc.

14. Protophilic Solvents Protophilic Solvents: (Philic- affinity) Solvents which are more basic than water. Solvent react with an acidic solute with the formation of a solvated proton. It shows the conjugate base of the acid. It possess a high affinity for protons. Eg. Liq ammonia, Amine (Ethylenediamine), Ketones, ethers, dioxane, amines-hydrazine and etc…. Leveling effect: Weak acids are normally used in the presence of strongly protophilic solvents as their acidic strengths are then enhanced and then it behave like strong acids; this is known as the leveling effect.

15. Amphiprotic Solvents Solvents posses both the properties of protophilic as well as protogenic. Similar to water, possesses both acidic and basic properties (donating and accepting of protons). Which are slightly ionize and donate or accept protons. ◦ Eg Alcohols (Methanol, Ethanol, etc.) , weak organic acids ( Acetic acid) ◦ Acetic acid makes weak acid into storing base

16. Acetic Acid Acetic acid slightly ionise and combine both protogenic and protophilic properties and able to donate and to accept protons Acetic acid is slightly ionize and dissociate to produce protons CH3COOH CH3COO – + H+ But in the presence of perchloric acid, a far stronger acid, it will accept a proton: CH3COOH2++ ClO4– CH3COOH + HClO4 The CH3COOH2+ion can very readily give up its proton to react with a base, so basic properties of a base is enhanced, so titrations between weak base and perchloric acid can often be accurately carried out using Acidic acid .

17. Leveling Solvents In general, strongly protophilic solvents are important to force equilibrium equation to the right. CH3COOH2++ ClO4– CH3COOH + HClO4 This effect is so powerful that, in strongly protophillic solvents, all acids act as of similar strength. HB B - + H + Similarly protogenic solvents, which cause all bases ( week also) to act as they were of similar strength. Solvents, which act as above (week to strong) , are known as Levelling Solvents.

18. Properties of the Solvents The choice and Properties of a solvent for the particular titration is very critical. ▪ The solvent should dissolve the substance to be titrated. ▪ Should not introduce interfering side reactions with either the substance to be titrated or the titrant. ▪ Should permit a large change in the solvated proton concentration near the equivalence point. ▪ Should be easily purified or available in pure and ▪ Preferably should be less expensive. If the solvent is to be used for a differentiating titration it should be neither strongly acidic nor strongly basic to avoid "LEVELING" effects.

19. Titration Of Bases The titrant should be a very strong acid. Eg Perchloric acid in Dioxane The solvent should not be basic properties Aprotic solvents, such as benzene, chloroform, carbon tetrachloride, chlorobenzene, either alone or mixed with glacial acetic acid may sometimes be used for titration with acetous perchloric acid To determine primary , secondary , tertiary amines, heterocyclic amines

20. Titration Of Acids The titrant should be a solution of a strong base Solutions of quaternary ammonium hydroxides in organic solvents, e.g. tetra-butylammonium hydroxide in benzene - methanol or IPA or triethyl-n-butylammonium hydroxide in benzene – methanol. Solution of Sodium or Potassium methoxide in Toluene - methanol Solvent (s): A mixture of benzene and methanol very weak acids (e.g., many phenols) usually require a more strongly basic solvent, such as DMF, anhydrous ethylenediamine or butylamine To determine week organic acids. Precaution: Amine may absorb carbon dioxide from the atmosphere

21. Selection of Solvents for NAT Acetic acid used for titration of: Weak bases, Nitrogen containing compounds Acetonitrile and or with Acetic Acid: Metal ethanoates Alcohols (IPA, nBA) : Soaps and salts of organic acids etc DMF: Benzoic acid, amides etc

22. Titrants for NAT Perchloric acid in acetic acid ◦ Amines, amine salts, amino acids, salts of acids Potassium Methoxide in Toluene-Methanol ◦ Week organic acid Quaternary ammonium hydroxide in acetonitrile-pyridine ◦ Acids, enols, imides & sulphonamides

23. Endpoint Detection End point detection is critical for titration, it is to know the completion of reaction and accurate determination. 1) Visual indicators: • Observe a colour change or precipitation at the endpoint. – Reaction progress checked by addition of external or self indicator 2) Electrochemistry: • Potentiometry - measure voltage change ( pH electrode) • Amperometry - measure change in current between electrodes in solution • Conductance – measure conductivity changes of solution Later two used for coloured, turbid & accurate end point

24. Improvement of End Point The end points may often be improved by the addition of aprotic solvents in order to depress the solvolysis of the neutralisation product. Potentiometric titrations are used for coloured solutions and also for compounds which remain feebly acidic or basic notwithstanding the levelling effect of the solvent. Visual indicators may be employed for compounds which behave as sufficiently strong acids or bases in appropriate non-aqueous solvents. The suitability of a visible indicator for a particular titration must be determined by performing a potentiometric titration and observing the colour change of the indicator simultaneously. Temperature of standardization and test shall be same since coefficients of expansion and dissociation constant may vary

25. Indicators for NAT 1. Crystal violet is used as a 0.5% (w/v) solution in glacial acetic acid. Its colour change is from Violet through blue, followed by green, then to greenish- yellow, 2. Methyl red is used as a 0.2% (w/v) solution in dioxane with a yellow to red colour change. 3. I-Naphthol benzein 0.2% (w/v) solution in acetic acid gives a yellow to green colour. It gives sharp end points in Nitromethane containing acetic anhydride for titrations of weak bases against perchloric acid.

26. Indicators for NAT 4. Oracet blue B is used as a 0.5 % (w/v) solution in acetic acid and it is superior to crystal violet for titrations of bases in acetic acid with standard perchloric acid. The end point : from blue to pink. 5. Thymol blue 0.5 % (w/v) in methanol is used for titrations of substances acting as acids in dimethylformamide solution. The end point: change from yellow to blue 6. Methyl violet 0.2% (w/v) in chlorobenzene, violet to blue.

27. USP Titrimetry <541> Non-Aqueous Titration Non-Aqueous titrimetry performed for , ◦ Acids, acid halides, acid anhydrides, carboxylic acids, amino acids, enols such as barbiturates and xanthines, imides, phenols, pyrroles, and sulfonamides. ◦ Bases, amines, nitrogen-containing heterocyclic compounds, oxazolines, quaternary ammonium compounds, alkali salts of organic acids, alkali salts of weak inorganic acids, and some salts of amines.

28. USP Titrimetry <541> Choice of Titrants: Basic compound: A volumetric solution of perchloric acid in glacial acetic acid is preferred, or perchloric acid in dioxane is used as required. Acidic compound: A volumetric solution of sodium methoxide or Lithium methoxide in a mixture of methanol and toluene tetra-n-butylammonium hydroxide and trimethyl hexadecyla mmonium hydroxide (in benzene-methanol or isopropyl alcohol),

29. USP Titrimetry <541> Systems for Nonaqueous Titrations Typeof Solvent Acidic (for titration of bases and their salts) Relatively Neutral (for differential titration of bases) Acetonitrile Alcohols Chloroform Benzene Basic (for titration of acids) Relatively Neutral (for differential titration of acids) Acetone Acetonitrile Methyl Ethyl Ketone Methyl Isobutyl Ketone Solvent # Glacial Acetic Acid Acetic Anhydride Formic Acid Propionic Acid Dimethylformamide n-Butylamine Pyridine Ethylenediamine Sulfuryl Chloride Toluene Chlorobenzene Ethyl Acetate Dioxane Methyl Red Methyl Orange p-Naphthol benzein Morpholine tert-Butyl Alcohol Indicator Crystal Violet Quinaldine Red p-Naphtholbenzein Thymol Blue Thymolphthalein Azo Violet Azo Violet Bromothylmol Blue p-Hydroxyazobenzene Alphezurine 2-G Malachite Green o-Nitroaniline p-Hydroxyazobenzene Antimony–calomel Antimony–glass Antimony–antimony # Platinum–calomel Glass–calomel Thymol Blue Electrodes Glass–calomel Glass–silver–silver chloride Mercury–mercuric acetate Glass–calomel Calomel–silver–silver chloride Antimony–calomel Glass–calomel Glass–platinum# # Relatively neutral solvents of low dielectric constant such as benzene, toluene, chloroform, or dioxane may be used in conjunction with any acidic or basic solvent in order to increase the sensitivity of the titration end-points. ## In titrant. reproducedFrom USP

30. Potentiometric Titration Electrode Systems Titration Indicating Electrode Glass Equation (1) Reference Electrode Calomel or silver–silver chloride Calomel (with potassium nitrate salt bridge) Applicability (2) Acid-base E = k + 0.0591 pH Titration of acids and bases E = E° + 0.0591 log [Ag+] Precipiti metric (silver) Silver Titration with or of silver involving halides or thiocyanate E = E° + 0.0296(log k′ − pM) E = E° + (0.0591/n) × log [ox]/[red] Complexo metric Mercury– mercury (II) Platinum Calomel Titration of various metals (M), e.g., Mg+2, Ca+2Al+3, Bi+3, with EDTA Oxidation- Reduction Calomel or silver–silver chloride Titrations with arsenite, bromine, cerate, dichromate, nitrite, exacyonoferrate(III), iodate, permanganate, thiosulfate (1) Appropriate form of Nernst equation describing the indicating electrode system: k = glass electrode constant; k′ = constant derived from Hg–Hg(II)–EDTA equilibrium; M = any metal undergoing EDTA titration; [ox] and [red] from the equation,ox+ ne ⇄ ⇄ red. (2)Listing is representative but not exhaustive. Reproduced From USP

31. Pontentiomertic Titration Potentiometric titration : The end-point of the titration is determined by variation of the potential difference between 2 electrodes in the solution to be examined as a function of the quantity of titrant added. Determine the volume consumed between the 2 points of inflexion. Electrode : Selection of appropriate Electrode is important to determine the accurate determination Preferred electrode for acid-base titrations is glass- calomel or glass-silver-silver chloride electrode , unless otherwise specified in Pharmacopeia

32. Typical Titration Graph

33. Preparation of 0.1N Perchloric Acid Preparation of 0.1N Perchloric Acid in Glacial Acetic Acid: Mix 8.5 ml 70% of perchloric acid with 500 ml of glacial acetic acid and 21 ml of acetic anhydride, cool, and add glacial acetic acid to make 1000 ml. Allow the prepared solution to stand for 1 day for the excess acetic anhydride to be combined, and determine the water content 0.02% and 0.5% (if > 0.5%, add acetic anhydride).

34. Perchloric Acid Preparation Perchloric acid (70 to 72%) addition in solvent contain water is exothermic and may decompose, handle carefully. Conversion of acetic anhydride to acetic acid requires 15-45 min Allow to cool to room temperature before adding glacial acetic acid Avoid adding an excess of acetic anhydride especially when primary and secondary amines are to be assayed, because it may react and convert acetylation .

35. Standardization of Perchloric Acid Accurately weigh about 700 mg of potassium biphthalate, previously crushed lightly and dried at 120° for 2 h, and dissolve it in 50 mL of glacial acetic acid in a 250-mL flask. Add 2 drops of crystal violet TS, and titrate with the perchloric acid solution until the violet color changes to blue- green. Standardization: perform potentiometrically as specified Sample shall be tested the same type of end point determination (visual or potentiometry) followed for standardization. % RSD of Triplicate NMT 0.2% Note: Use benzoic acid as primary standard where required as specified in USP

36. Preparation of Tetrabutylammonium hydroxide Preparation of 0.1 M tetrabutylammoniumhydroxide in Toluene- Methanol. 1. Dissolve 40g of tetrabutylammoniumiodide in 30ml of absolute methanol and 20g of powdered silver oxide. 2. Shake vigorously for 1 hour. 3. Test supernatant liquid for residual iodine, if unreacted iodine is present add additional 2g of silver oxide & repeat until solution is free from iodine 4. Filter the solution in sintered glass filter and rinse with of 50ml dry toluene repeat 2 or 3 times. 5. Add washing toluene into filtrate ( step-3) and dilute to 1000ml with anhydrous Toluene and mix. 6. Stored the solution in a well closed tight container Precaution: Protect the solution from moisture and carbon dioxide.

37. Preparation of Potassium methoxide Preparation of 0.1M Potassium methoxide in Toluene-Methanol. 1. Take around 50ml Methanol & 50ml Toluene into flask and add carefully 5.6 gm of potassium metal (freshly cut) and stir to dissolve completely. Methanol and Toluene should be anhydrous /dry. 2. Add Methanol into the above solution to get the solution clear and add 50ml Toluene, now solution could be turned hazy. 3. Repeat the addition of Methanol and Toluene until a solution becomes 1000ml. 4. Ensure that final solution should be clear. 5. Store the above solution in an air tight container. Note: To prepare sodium methoxide use 2.3gm of sodium metal instead of potassium.

38. Standardization Base Titrants Standardization of 0.1 M tetrabutylammoniumhydroxide: Accurately weigh 60mg of Benzoic acid in dimethlyformamide (DMF) 30ml. Add 5 drops of Thymol blue (0.2w/v in methanol). Titrate the mixture against 0.1 M tetrabutylammonium hydroxide. Blanket with nitrogen to protect from atmospheric carbon dioxide. Standardization of 0.1 M Potassium methoxide in toluene-methanol Accurately weigh 60mg benzoic acid and transfer to reaction vessel containing 10ml of dimethylformamide(DMF). Added 20 ml DMF and 3-5 drops of Thymol blue indicator. Titrate the mixture against 0.1M potassium methoxide in toluene-methanol Instead manual titration, perform potentiometric titration for more accurate result.

39. Assay Determination of API (USP) Typical test method as per UPS of API Assay: (98.0%– 100.5%) Dissolve about 150 mg of Pseudoephedrine Sulfate, accurately weighed, in 50 mL of glacial acetic acid. Titrate with 0.1 N perchloric acid VS, determining the endpoint potentiometrically. Perform a blank determination, and make any necessary correction. Each mL of 0.1 N perchloric acid is equivalent to 42.85 mg of (C10H15NO)2. H2S04.

40. Electrolytes for Electrode Suitable electrolyte is important for accurate result : Electrolyte shall be filled appropriately. Eg ◦ LiCl saturated in ethanol ◦ KCl saturated in ethanol ◦ TEA-Br on Ethylene glycol Filling of electrolyte: ◦ Remove the aqueous KCl completely ◦ Rinsing with water if any residual KCl ◦ Rinsing with the non-aqueous solvent to remove water ◦ Fill the electrode with the suitable electrolyte.

41. Precautions to Minimize the Error Stirring rate should be adequate. As a general rule, it should be as high as possible. But avoid the formation of a vortex during stirring. If endpoint recognition criteria (ERC) is too small can result in an incorrect equivalence point Use the titration parameters “signal drift” and “min./max. waiting time” to adjust the titration rate. Sensor is suitable for the titration. If sensors is old or dirty, it can result in longer response times. Check that the anti-diffusion tip is functioning correctly. The titrant should not be stirred directly onto the electrode, but rather away from it.

42. Precautions to Minimize the Error Selection of appropriate Electrode and Electrolyte filled shall be appropriate. Burette tip / Diaphragm shall be free from clogging Titration vassal shall be closed while titration Avoid expose to atmospheric air, CO2 may interfere Titration vassal shall not contain water or previous residue Sample shall be completely transferred and dissolved No Air bubbles in the dosing cylinder and/or in the tubing Air bubbles in the measuring vessel and/or on the sensor during photometric titrations

43. Blank Correction Blank correction is usually obtained by residual blank titration. Repeated in every detail except that the substance being assayed is omitted. An appropriate blank correction shall be included in calculation Difference between the volume consumed in the residual blank titration and volume consumed for same titration If blank is high or suspected, repeat the analysis

44. Calculation o Normality: Eq.wt/1000ml or meq/mL o Morality: Mole/1000ml o V1 N1 = V2N2 o N1 = V2N2/V1 Normality = Wt of sample x 1000 / Eq. Weight x Vol Wtof sample (mg) = V x N x Eq. weight Assay = Qtyestimated in sample x 100/ weight of sample Assay = V x N x Eq. wt x 100/ wt of sample x 1000

45. Errors in Titrimetry Error in methods: The endpoint method may not show a change exactly at the equivalence point due to the reactions involved Titration Error = Volume at endpoint – Volume at equivalence point Lower assay: Negative error due endpoint is early /before equivalency point. Higher Assay: Positive error due to later endpoint / after equivalency point

46. Error Estimation Possible Error due to titrant volume or sample weight: 10 ml titre volume = 100 % ◦ If difference is 0.1ml error is 1% or 0.2ml = 2% 5ml titre volume = 100 % difference in 0.1ml = 2% error 25 ml titre volume = 100 % ie difference 0.1ml = 0.4% error Error can be minimized : Ensure optimum titre volume is around 25ml for manual titration Use autotitrator to keep lower dispensing increment. ie burette with 0.001ml or 0.01ml least count is preferred to reduce the error Higher weight of sample

47. Possible Error in Weighing Misreading of the balance • Balance sprit level is not eccentric. • Not cleaning the surface of the balance first • Touching the weighed object with moist hands • Leaving the balance doors open during weighing • Error due to incorrect calibration of balance • Not cooling the sample down to near room temperature, • Not removing a static charge from the sample, • Excess vibration or air currents nearby equipment, • Prolonged time sample left on pan adds/loses moisture.

48. Prevention of Contamination What are the Possibility of Contamination by an Analyst; Contaminate a sample during weighing by placing a contaminated spatula. Placing the sample on or into a contaminated holder during weighing. Dropping some lint/hair/skin or sneeze into the sample while weighing. Opening up a bottle of chemicals near the sample being weighed. When performing trace analysis, it is possible for just a microgram even massive fingerprint!

49. Units and Measurements # Description Unit 1 Molarity Moles of solute/ Liter millimolesof solute/ milliliter 2 Normality Equivalent weight of solute/ Liter (mole / Equivalency in liter) 3 Percent w/v Percent w/w Grams of substance in 100ml Grams of substance in 100gm 4 Parts per million milligram / liter milligram / kilogram microgram / ml (or gram) 5 Parts per billion microgram / liter microgram / kilogram nanogram/ ml (or gram Unit Conversion to factor Parts per million % ppm / 10000 ppm*1000 ppb

50. References USP – General Chapter -Titrimetry <541> 1. European Pharmacopoeia- General Chapter 2.2.20 2. Non-aqueous titration of acids and bases with potentiometric endpoint indication- Peter Bruttel - Metrohm 3. E. G. Wollish, C. W. Pifer, and Morton. Schmall. -Titration in Nonaqueous Solvent to Pharmaceuticals- Analytical Chemistry 4. Vogel's Text Book of Qualitative Inorganic Analysis 6thEdition 5. Analytical Chemistry A Qualitative & Quantitative Approach- Deepak Chowrasia & Dr. Nisha Sharma 6.