CHEMICAL QUALITY ASSURANCE

Module 3. Chemical analyses of milk and milk products

Lesson 6
PREPARATION AND STANDARIZATION OF SOLUTIONS

6.1 Introduction

Standard solutions are the solutions with known strength. The calibration of other solutions and reagents depends upon the accurate strength of these solutions. These solutions are prepared by using certain substances having typical characteristics. These substances are known as standard substances. In this chapter, the properties of these substances and how to prepare standard solution has been discussed.

6.2 Standard Substances

There are two types of substances which are generally employed for preparing standard solutions

• Primary standard substances
• Secondary standard substances.

6.2.1 Primary standard substances

Those substances which can easily be obtained in pure and crystalline form e.g. Oxalic acid, sodium carbonate etc, are called primary standard substances.

6.2.1.1 Characteristics of primary standard substances

• It should be easy to obtain, to purify and to preserve.
• It must not be hygroscopic.
• It should not decompose at ordinary temperature.
• It should be readily soluble under the conditions in which it is used.
• Its reaction with other reagents should be quantitative and practically quick.
• It should have high eq. wt. so that the error due to weighing is minimized.
• It should be fairly cheap.

Fig 6.1 Sodium carbonate

6.2.1.2 Primary standard substances used for acid- alkali titration

a) Sodium carbonate

Since, Sodium carbonate can be easily obtained in pure state, its standard solution is prepared by directly dissolving a known wt of it in water and making the solution a known volume. Eq. wt. of sodium carbonate (anhydrous) being 53, its N/10 solution would contain 5.3 gm/litre and N/20 solution would contain 2.65 gm/litre.

These standard solutions are used for finding the strength of solutions of HCl, H₂SO₄ etc whose standard solutions cannot be prepared directly.

b) Oxalic acid (COOH)₂

Oxalic acid is available in pure state and its standard solutions can, therefore, be prepared by the direct method. Eq. wt. of hydrated oxalic acid (C₂H₂O₄.2H₂O), being 63 its N/10 solution would contain 6.3 gm/litre, and N/20 solution would contain 3.15 gm/ litre.

These standard solutions are employed to find the strength of solutions of alkalies (NaOH and KOH) whose standard solutions cannot be prepared by the direct method.

6.2.1.3 Preparation of some primary standard solutions

Standard solutions are prepared by using standard substances. Here a known quantity of standard substances depending upon the requirement is dissolved in a known amount of water and desired volume is made. Since, these substances have a constant weight, high purity, non hygroscopic property, so the solution obtained is of known and definite concentration. The examples of such solutions are as follows:

a) Example 1. Standard N/10 oxalic acid solution (Primary standard)

To prepare N/10 solution of oxalic acid, weigh 6.3 gm of oxalic acid & dissolve in distilled water & finally make up the volume to one liter in a volumetric flask. The standard solution of oxalic acid (Primary standard) is used to find the strength of solutions of alkalies like NaOH, KOH (Secondary standards) whose standard solutions can not be made by direct weighing.

b) Example 2. N/10 standard Na₂CO₃ solution (Primary standard)

It can also be obtained in pure form and its anhydrous form is available. Here also, its standard solution can be prepared by direct weighing & dissolving in water to make up to known volume.

To prepare N/10 Na₂CO₃, weight exactly 5.3 gm of pure anhydrous salt, dissolve in distilled water and make up to 1 litre in volumetric flask. Its standard solution is used to find out the strength of solutions of acids like HCl, H₂SO₄, HNO₃ etc whose standard solutions cannot be prepared directly.

6.2.2 Secondary standard substances

Those substances or reagents which cannot be obtained in a sufficient pure state, e.g. NaOH, KOH, HCl, H₂SO₄ are called secondary standard substances.

6.2.2.1 Preparation of some secondary standard solutions

N/10 NaOH: Prepare concentrated stock solution (say 50%) of NaOH by dissolving equal parts of NaOH pellets (50 gm) & water (50 gm) in a flask. Keep it tightly stoppered for 3-4 days. Use the clean, supernatant liquid for preparing N/10 solution. Approximately 8 ml of this stock solution (50%) is required per litre of distilled water. This will give approximate solution. Now take 10 ml of standard N/10 oxalic acid (primary standard) solution in conical flask and add 2- 3 drops of phenolphthalein indicator. Take unknown solution i.e. approximate N/10 NaOH solution in burette and add to the conical flask containing standard oxalic acid solution by continuous mixing by swirling the flask till the appearance of pink color. NaOH is taken in burette and standard oxalic acid in conical flask as shown below in figure.

Fig 6.2 Sodium hydroxide pellets



Fig. 6.3 (a) Standard solutions

Note down the volume of approximate N/10 NaOH solution used in the titration of 10 ml of standard oxalic acid. Calculate the normality of the unknown sodium hydroxide solution by using following equation:

N₁V₁ = N₂V₂
(Base) = (Acid)

N1 = Normality of NaOH solution. ( ?)
V1 = Volume of NaOH solution used. (ml)
N2 = Normality of standard oxalic acid solution. (0.1 N)
V2 = Volume of standard oxalic acid solution. (10 ml)

If the volume of approximate NaOH used in the titration is less than 10 ml, means the solution is strong and its normality is not N/10, so dilute the basic solution and again standardize with standard oxalic acid solution till normality of approximate solution is same as that of standard solution.

6.3 Approximate Strength of Concentrated Acids Generally Available

Table 6.1 Concentrated acids

6.3.1 N/10 HCl

a) Prepare approximately 0.1 N solution on the basis of the strength given on the label by diluting it 120 times with distilled water. Then standardize it against standard N/10 Na₂CO₃ using methyl orange as an indicator. Alternatively, it can also be prepared as given below in “b”.

b) Prepare approximately 0.1 N solution on the basis of the strength given on the label by diluting it 120 times with distilled water. Then standardize it against standard N/10 NaOH which is already standardized against N/10 oxalic acid using Phenolphthalein indicator.

Fig. 6.3 (b) Standard solutions

6.3.2 N/10 H₂SO₄

Caution:

Concentrated H₂SO₄ is very corrosive in nature, therefore, it should be handled carefully.
Always remember: “ADD ACID TO WATER” under cold conditions. This is done to avoid bumping due to the heat generated.

For the preparation of N/10 H₂SO₄, take 10 ml of concentrated H₂SO₄ (usually about 36 N), dilute 36 times by adding acid in small quantity to distilled water in a cold water bath, to make it 1N and then dilute this 1N solution further 10 times to make it N/10. Then standardize against standard N/10 NaOH or N/10 KOH using phenolphthalein indicator.

6.3.3 N/10 HNO₃

Take 10 ml of concentrated HNO₃ (usually about 16 N), dilute 16 times by adding acid to distilled water to make it 1N and then dilute this 1N solution further 10 times to make it N/10. Then standardize against standard N/10 NaOH or N/10 KOH using phenolphthalein indicator.

6.4 Preparation of Some Other Reagents

6.4.1 Preparation of chromic acid (Cleaning solution)

Fig. 6.4 Chromic acid

Caution:

During preparation of chromic acid, observe the precaution for handling of concentrated H₂SO₄, that is, “ADD ACID TO WATER” under cold conditions.

Dissolve 50 gm of K₂Cr₂O₇ in 50 ml of water in a beaker. Keep the beaker in cold water and add slowly 500 ml of concentration H₂SO₄ and cool. It is a very corrosive solution and care should be taken to avoid its coming in contact with the skin. It is almost a saturated solution of K₂Cr₂O₇ in concentrated H₂SO₄.

Before using chromic acid, it is necessary to clean the glassware with detergent for the economic use of chromic acid.

6.4.2 Preparation of gerber sulphuric acid

During preparation of Gerber sulphuric acid, observe the precaution for handling of concentrated H₂SO₄, that is, “ADD ACID TO WATER” under cold conditions.

Gerber sulphuric acid is used to dissolve casein in milk. If dilute H₂SO₄ is used, casein is precipitated but not dissolved, whereas, if concentrated H₂SO₄ is used, it causes charring of organic matter. Therefore, the concentration of sulphuric acid is so adjusted that it is just strong enough to dissolve the casein without charring the fat. The acid also produces necessary heat to keep the fat in the liquid state.

Gerber H₂SO₄ has strength of about 90-91% corresponding to a Sp. Gr. of 1.807 to 1.812, whereas concentrated H₂SO₄ is generally 97-99% (Av 98%) with Sp. Gr. of 1.835.

Therefore, for practical purposes generally 900 ml of concentrated H₂SO₄ is added to 100 ml water to give 1 litre of Gerber acid.

6.4.2.1 Preparation

Take required vol. of water (say 100 ml) in a flask and keep it in a basin of ice-cold water. Carefully, add the required quantity of concentrated H2SO4 (say 900 ml) in small quantities at a time keeping the container sufficiently cold. Mix gently.

6.4.3 Testing the amyl alcohol (a by product of fuel oil refinery) used for fat determination

Iso-amyl alcohol (also called Iso-butyl carbinol) used in Garber fat test shall be clear, colorless and free from impurities particularly fatty mater. Perform various tests to know its purity.

1. Density: At 27°C, Density shall be between 0.803 to 0.805 gm/ml.
2. Boiling point: Boiling point shall be 128-129°C (can be checked using boiling point apparatus).
3. 95% of the liquid shall get distilled between 130-132°C.
4. Test for absence of furfural and other impurities: 5 ml iso amyl alcohol + 5 ml H₂SO₄ (97%) → Observe the color → shall not show more than a yellow or light brown color.
5. Test for absence of fatty matter: Carry out a blank Gerber fat test using distilled water in place of milk. If any fat separation is observed → indicates impurities of some fatty matter.

6.4.4 Preparation of 0.1 N sodium thiopsulphate solution (Na₂S₂O₃.5H₂O)

Dissolve approximately 24.8 gm of sodium thiosulphate crystals in previously boiled and cooled distilled water and make the volume to 1000 ml. Store the solution in a cool place in a dark colored bottle. After storing the solution for about two weeks, filter if necessary and standardize as follows:

Weigh accurately about 5.0 gm of finely ground potassium dichromate which has been previously dried to a constant weight at 105 ± 2° in to a clean 1.0 litre volumetric flask. Dissolve in water make up to the mark; shake thoroughly and keep the solution in dark place. Pipette 25.0 ml of this solution into a clean glass stoppered 250 ml conical flask. Add 5.0 ml of concentrated hydrochloric acid and 15.0 ml of 10% potassium iodide solution. Allow to stand in dark for 5 minutes and titrate the mixture with the solution of sodium thiosulphate using starch solution as an indicator towards the end. The end point is taken when blue color changes to green. Calculate the normality (N) of the sodium thiosulphate as follows:

25W

N = ----------------

49.03 V

W = weight in g of the potassium dichromate
V = volume in ml of sodium thiosulphate solution required for the titration.

6.4.5 Preparation of 0.1 N standard solution of silver nitrate

Dissolve slightly more than the required quantity (17.2 g instead of 16.989 g) of reagent grade silver nitrate in distilled water and dilute to one litre in a volumetric flask. Weigh accurately 0.5844 g of NaCl (dried at 110°C before weighing) and transfer to a 100 ml volumetric flask and add 50 ml of halogen free water to dissolve the material. Make up the volume with distilled water to the mark and mix the contents. Pipette out 10 ml of the prepared standard sodium chloride solution in 100 ml conical flask and add 2-3 drops of potassium chromate indicator (5% solution in water). Titrate with silver nitrate solution until a perceptible reddish brown color appears. Carry out a blank titration using 10 ml of distilled water instead of sodium chloride solution and deduct the blank reading from the reading for the standard sodium chloride solution.

Calculate the normality of the silver nitrate solution using normality equation:

N₁V₁ = N₂V₂

Where,

N₁ = Normality of standard sodium chloride solution (0.1N)
V₁ = Volume in ml of sodium chloride used for titration (10 ml).
N₂ = Normality of prepared silver nitrate solution.
V₂ = Volume in ml of prepared silver nitrate solution used for titration.

6.4.6 Preparation and standardization of EDTA solutions

1. Preparation of 0.01 M EDTA solution : Dissolve 3.8 g of disodium ethylene diamine dihydrogen tetraacetate (EDTA, M.Wt. 372.25) in distilled water and volume is made to 1 litre. Mix it well, store in polyethylene reagent bottle. It is standardized against 0.01 M CaCO₃ or CaCl₂.
2. Preparation of 0.01 M CaCl₂ solution: Prepare standard Ca solution (1 ml = 1 mg CaCO₃, M.wt. 100) by weighing 1 g CaCO₃ into 500 ml conical flask or beaker and adding dilute HCl through funnel until CaCO₃ is dissolved. Add 20 ml water, boil to expel CO₂ and cool. Add few drops of methyl red indicator and adjust colour intermediate orange (brownish red) with dilute NH₄OH or HCl as required. Transfer quantitatively to 1 L volumetric flask and make up volume to the mark. Shake it well and store it well and store in air-tight reagent bottle.
3. Erichrome Black T indicator: Dissolve 0.5 g of Erichrome black T in 100 ml of triethanolamine. Or 0.4 g in 100 ml methanol.
4. Buffer solution: Dissolve 16.9 g NH₄Cl in 143 ml NH₄OH, and dilute to 250 ml with water. Store in tightly stoppered Pyrex of plastic bottle. Dispense from bulb-operated pipette. Discard after 1 month or when 1-2 ml added to sample fails to produce pH 10.0±0.1 at end point titration.

6.4.7 Standardization of EDTA solution

Rinse and then fill burette with prepared EDTA solution. Pipette 25 ml of standard CaCO₃ solution into 250 ml Erlenmeyer flask, add 1 ml ammonia buffer (to raise the pH as reaction takes place at high pH) and 3-4 drops of Erichrome black T indicator. Titrate the EDTA solution until colour changes from wine red to dark blue with no reddish tinge remaining. Calculate the molarity of EDTA (M₁V₁ = M₂V₂), if excess follows the procedure for the standardization, recheck the molarity and it should be 0.01 M.

6.4.8 Preparation of Fehling solution

Fehling solution used for the estimation of reducing sugars is generally prepared fresh by mixing equal quantities of Fehling’s A and Fehling’s B which are prepared separately as follows:

a) Fehling’s A

Dissolve 34.639 g Cu(SO)₄.5H₂O in distilled water and add 0.5 ml concentrated H₂SO₄, mix and make the volume to 500 ml. Filter if necessary.

b) Fehling’s B

Dissolve 173 g of Rochelle salt (Na K tartarate) and 50 g of NaOH in distilled water. Allow to stand for two days. Filter if necessary.

6.4.8.1 Standardisation of fehling’s solution

1. Pipette 5 ml of Fehling’s solution A and 5 ml of Fehling’s solution B using two separate pipettes in a 250 ml Erlenmeyer flask. Fill up a burette with the standard lactose solution and connect the burette end with an offset tube to keep the burette tube out of steam.
2. Heat the content of the flask to boiling over burner or heater and maintain moderate boiling for 2 min. To prevent bumping add some inert boiling chips. Add 3 to 4 drops of methylene blue indicator (0.2% in water) without removing from the flame. Titrate the content of the flask against standard lactose solution (0.5%) from the burette until the blue colour disappears and the bright brick-red colour of precipitated Cu₂O appears (at the end point the Cu₂O suddenly settles down giving a clear supernatant). Note the volume of lactose solution required for the standardization of Fehling’s solution. After this preliminary titration, further titration or titrations should be carried out, adding practically the whole of the standard lactose solution volume (one ml less than required as observed in first titration) required for the titration before commencing the heating. Let the contents boil for 2 minutes. Now, add 3-4 drops of methylene blue indicator, continue heating and complete the titration within 3 min from the commencement of boiling. Let V₁ ml be the titre for this experiment.

Note: Carefully note the first disappearance of blue colour. Once missed, it is difficult to ascertain the end point. Maintain the boiling at a uniform rate during the titration.

Multiply the titre value by mg/ml lactose of the standard solution to obtain total lactose required to reduce the copper and term the value a "Factor F".

6.4.9 Preparation of pH indicator solutions

6.4.9.1 Phenolphthalein indicator solution

Weigh 1.0 gm phenolphthalein and place the powder in a 100 ml volumetric flask containing about 50ml of 95% ethanol. Stopper and shake vigorously for a few minutes, then add 20ml more ethanol and shake until a clear solution is formed and make the volume to 100 ml.

6.4.9.2 Methyl orange indicator solution

Dissolve 1.0 gm of methyl orange powder in distilled water and dilute to one litre. Filter, if necessary.