calculations for preparation of solutions
- 1. CALCULATIONS FOR PREPARATION OF SOLUTIONS CHEMICALS USED IN THE LABORATORY • All the chemicals used in the laboratory should be of standard grade • Chemicals are of two grades 1. Laboratory reagents (LR) – Purity is about 95-98 percent. – Mostly used for ordinary solutions 2. Analytical reagents (Anal R, A.R or guaranteed reagents) – Purity is about 100 percent. – Mostly used for standard solutions
- 2. Examples of some standard firms manufacturing the chemicals • British Drug House • E.Merk • Sigma • Sarabhai Merck ( S.M.) etc.
- 3. • The solutions of the reagents must be accurately prepared in distilled water and • Preserved as prescribed in the analytical procedure. Preparation of solutions
- 4. 1. Percent solution • If the chemical is solid it is prepared by weight by volume and • If liquid volume by volume • According to the required percent, ‘X’ grams of the solid reagent is dissolved in 100 ml of water or other solvent • similarly for liquids ‘X’ ml of the liquid is diluted to 100 ml water or other solvent Types of solutions required in the laboratory
- 5. 2. Molar solution • It is prepared by dissolving one molecular weight of the chemical in 1 litre of the solvent • Some times fractions of molar solutions such as milli mole (10-3M) or micromole (10-6M) solutions are required • If one ml of molar solution is diluted to 1 litre it will be one milli mole • If 1 ml of milli mole solution is diluted to 1 litre it will be one micro molar solution
- 6. 3. Normal solutions • When one equivalent weight of a substance is dissolved in 1 litre of water, it is called one normal solution. • For the majority of substances the equivalent weight is calculated as follows: Eq. wt. = Molecular wt. of the substance Replaceable number of H or OH ions For example, the Eq. wt. of • NaOH = 40 / 1 : = 40g • KOH = 56 / 1 : = 56 g • Na2CO3 = 106 / 2 = 53 : = 53g • HCl36.45 / 1 : = 36.45 g • H2SO4 = 98 /2 : = 49 g
- 7. Standardization of acid solutions • The exact normal solutions of solid unhygroscopic substances can be prepared by dissolving one equivalent weight of the substance in one litre of solvent • In biochemical tests generally normal solutions of alkali ( NaOH, KOH ) and acids ( HCl, H2 SO4 ) are required • Since alkalis are hygroscopic and acids in liquid form, their exact normal solutions can not be prepared directly • For ordinary purposes their approximate normal solutions are prepared and used but for exact normality they are standardized by titration
- 8. • The volume of concentrated acid to be diluted to 1 litre with water for preparing approximate one normal solution can be calculated in two ways: (i) If the specific gravity and percentage of purity of the acids are known, the volume is calculated by the formula VoI of acid required for 1N = Equivalent wt. of the acid x100 Percent purity x Sp. gr. For Example, 1N H2 SO4 = 49 x 100 = 27.89 ml = 28.0ml 95.6 x 1.84 Acid solutions of approximate normality
- 9. (ii) Divide 1000 by the normality of the concentrated acid and the volume obtained is diluted to 1 litre with water • e.g., the normality of conc. HCl is 11 • So 1000 / 11 = 91 ml which is diluted to 1 litre for 1 N solution.
- 10. Table: approximate normal acid solutions • . Chemicals Sp. gr. Normality of concentrated chemical Vol. of conc. Chemical (ml) to be diluted to 1 litre with water Sulphuric acid 1.84 36 28 Hydrochloric acid 1.18 11 91 Nitric acid 1.42 16 63 Perchloric acid 1.54 9 111 Acetic acid 1.05 17 59 Ammonia 0.88 17 59
- 11. Precautions • Concentrated acids and liquor ammonia should be handled with great care • The bottles should be opened near window where ventilation is available • While diluting concentrated acids, particularly sulphuric acid, they must be added to water gradually with care • Water should never be added to concentrated sulphuric acid
- 12. Preparation of exact 1 N -HCl • For common use 1N –HCL is prepared by diluting 91 ml of concentrated HCL to 1 litre with water. For exact 1N-HCL, more than calculated volume of HCL is diluted to 1 litre and titrated against a base solution as follows: (1) Dilute 100 ml of conc. HCL with water to 1 litre, mix well. (2) prepare exact 1 N solution of sodium carbonate by dissolving 5.30g anhydrous sodium carbonate in 100ml water (Eq wt. 53g)
- 13. (3)Phenolphthalein indicator • Dissolve 250mg indicator in 50ml of 50 percent alcohol. (4)Titration • Take 10ml of acid solution and 10ml water in a small beaker • Add 2-3 drops of the indicator and • Titrate with sodium carbonate solution from a 25ml burette till a faint red colour is obtained • Note the volume (x ml) of base consumed at the end point. Preparation of exact 1 N –HCl Contd…
- 14. (5)Calculate the exact normality of the acid by formula • Normality of base x vol. of base = Normality of acid x volume of acid • So, the normality of HCL=1 x X / 10 • Normality of base is 1 volume of base is x ml. Preparation of exact 1 N –HCl Contd… •After calculating the exact normality of the acid, it is proportionally diluted with water to obtain exact 1 normal solution.
- 15. Exact 1N –NaOH solution • The equivalent wt. of sodium hydroxide is 40g • So 40g is dissolved in 1 litre of water for approximate 1N solution and used. • But for exact normality it is titrated against oxalic acid solution.
- 16. Titration against oxalic acid solution 1. Dissolve 50g sodium hydroxide in 1 litre of water and mix well. 2. Prepare exact 1N solution of oxalic acid by dissolving 6.3g of it in 100ml water. 3. Take 10ml of oxalic acid and 10ml water in a beaker, add 2-3 drops of phenolphthalein indicator. Titrate against the sodium hydroxide from a burette till a faint red colour is obtained. 4. Calculate the exact normality of sodium hydroxide solution as in the case of acid and dilute proportionately with water to obtain exact 1N solution
- 17. • If the equivalent weight of a substance is x gram and if x mg of the substance is dissolved in 1 litre of water it is called one mEq/L solution. • In biochemical tests the results of anions and cations of blood are expressed in mEq/L. (i) To convert the results obtained in mg/100 ml to mEq/L the formula given below is used:- mEq/L = mg /100ml x 10 x valency of ion atomic weight of ion Milli equivalent solution -(mEq/L)
- 18. Divide the values in mg/100 ml by the following factors: Sodium(mg/l00ml) by 2.3 Potassium(m g/l 00 ml) by 3.9 Calcium (mg/l00 ml) by 2.0 Magnesium(m g/l 00 ml) by 1.2 Chloride( mg chloride! 10 0 ml) by 3.55 Chloride(NaCVI00ml) by 5.85 Bicarbonate(volume/l00 ml) by 2.24 Phosphate (mg phosphorus/l 00 ml) by 1.72 Protein( g/100 ml) by 0.41 ii) To convert mEq/L values into mg per 100 ml, multiply by the above factors.
- 19. Isotonic solution • When two solutions are separated by a semi permeable membrane, the solvent will flow from the solution of low osmotic pressure to the solution of higher osmotic pressure until both solutions have equal osmotic pressure. • Such pair of solutions which have same osmotic pressure ( osmolarity) are called isotonic solutions. • When isotonic solutions are mixed together there is no exchange of their contents between the two solutions.
- 20. Standardization of method Prepare the reagents accurately in distilled water and preserve them as prescribed. The standard solutions must he prepared with Anal. pure grade chemicals. Set up 10 tubes with same volume of the standard solution and develop colour as given in the procedure. Measure the absorbance. Calculate mean and standard deviation of the absorbance and keep in the record for the future check up.
- 21. 3. Beer's law – Prepare a set of standard solutions of different concentrations e.g. glucose 50, 100, 150,200 and 250 mg/l00 mI. Develop the colour as given in the procedure and measure their absorbance. Prepare a curve by plotting the absorbance against the concentration. A straight line is found upto that concentration to which Beer's law is obeyed. (4) Analyse a number of food samples Check that the values obtained are in agreement with the normal range reported in literature
- 22. • Once the method is standardized, the main requirement is to maintain a careful and satisfactory performance of the routine analysis. • The standard should be set up with every batch of tests and it should be noted that the absorbance of the standard obtained is practically the same which was obtained during standardization and does not vary significantly from batch to batch or day to day. • The constant value of the absorbance of the standard effects that the reagents, instrumental conditions and performance of the tests are satisfactory and the results reproducible. • If the absorbance of the standard varies significantly, this signals that some thing is wrong and the cause of it should be checked.
- 23. Quality control • The laboratory tests often vary significantly from day to day and differ from laboratory to laboratory even by the same method • In order to maintain an acceptable degree of precision and accuracy of the results, certain measures are adopted in the analysis which is called quality control
- 24. Common practice of quality control in laboratory Use of standard solutions • This is a very common and convenient practice • With each batch of test samples, a known standard solution is assayed and the results of unknown samples are calculated against the absorbance of standard solution • The standard solution may be prepared of a single substance or a mixture of several standard substances used for different tests • The idea behind the use of mixture control is to take in to account the effect of various substances normally present in the samples on the analysis of a particular substance although it is not always feasible • If the standard solution is treated in the test exactly like test samples, it is a better safeguard against error than the standard solution which is used for colour development only in the final stage of the analysis