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VOLUMETRIC ANALYSIS CORE PRACTICAL (chemistry)

By Maruthu pandi .M,\

Madurai,TN,Indian

1) ESTIMATION OF AMINO ACID (GLYCINE) BY SORENSON’S FORMAL TITRIMETRIC METHOD

Aim To estimate the amount of amino acid (Glycine) present in the whole of the given

unknown solution Principle Amino acid reacts with excess of formaldehyde to give free hydrogen ion and act

as acidic solution. This acidic solution is titrated against standard alkali (Sodium hydroxide) using phenolphthalein as indicator.

Reagents required i. Standard oxalic acid solution (0.1 N). ii. Sodium hydroxide solution .iii. Phenolphthalein as indicator.iv. Formaldehyde .v. Amino acid (Glycine)

PROCEDURETitration – I Standard oxalic acid Vs Sodium hydroxide solution Weighed accurately 1.576 g of crystalline oxalic acid and transfer into a

250 ml of standard flask then the volume is made up to 250 ml using distilled water. Pipette out

exactly 20 ml of this solution into a clean conical flask and two drops of phenolphthalein as indicator is

added. This is titrated against the Sodium hydroxide solution taken in the burette. The end point is the

appearance of pale permanent pink colour. The titrations are repeated for concordant values. From the

titre value the normality of Sodium hydroxide solution is calculated.

PROCEDURETitration – I Standard oxalic acid Vs Sodium hydroxide solution Weighed accurately 1.576 g of crystalline oxalic acid and transfer into a

250 ml of standard flask then the volume is made up to 250 ml using distilled water. Pipette out

exactly 20 ml of this solution into a clean conical flask and two drops of phenolphthalein as indicator is

added. This is titrated against the Sodium hydroxide solution taken in the burette. The end point is the

appearance of pale permanent pink colour. The titrations are repeated for concordant values. From the

titre value the normality of Sodium hydroxide solution is calculated.

PROCEDURETitration – III (Blank value) Blank Vs Standardized Sodium hydroxide solution Pipette out exactly 20 ml of distilled water into a clean

conical flask to this 5 ml of formaldehyde and followed by two drops of phenolphthalein as indicator is

added. The contents are mixed well for 5 minutes. This is titrated against the standardized Sodium

hydroxide solution taken in the burette. The end point is the appearance of pale permanent pink colour.

The titrations are repeated for concordant values.

ESTIMATION OF AMINO ACID (GLYCINE) BY SORENSON’S FORMAL TITRIMETRIC METHOD

i. CALCULATION (Titration – I Standard oxalic acid Vs Sodium hydroxide)ii. Weight of oxalic acid = 1.576 g iii. Volume of Standard oxalic acid solution (V 1) = 20 ml iv. Normality of Standard oxalic acid solution (N1 ) = 0.1 N v. Volume of Sodium hydroxide solution (V2) = -------- ml vi. Normality of Sodium hydroxide solution (N2) = ? vii. We know that, V1N1 = V2N2 viii. N2 = V1N1 ix. V2 x. N2 = 20 x 0.1/--------- xi. N2 = ----------------- xii. Normality of Sodium hydroxide solution (N2) = ---------------------------

CALCULATION Volume of Sodium hydroxide solution (V 1) = -------------- ml Normality of Sodium hydroxide solution (N1) = -------------- N Volume of amino acid (Glycine) solution (V2) = 20 ml Normality of amino acid (Glycine) solution (N2) = ? We know that, V1N1 = V2N2 N2 = V1N1 V2 N2 = ---------------- Normality of amino acid (Glycine) (N2) = ------------------ N Equivalent weight of Glycine - 75 The amount of amino acid (Glycine) present in the whole of the given unknown solution = Normality of amino acid (Glycine) x Equivalent weight of Glycine x 100 1000 = ----------------------------- grams

TEST VALUE – BLANK VALUE Test value – Blank value will give the actual amount of Sodium

hydroxide consumed by the amino acid solution. From this value the strength of amino acid is calculated, and from this strength the amino acid present in the whole of the given unknown solution is calculated

Equivalent weight of Glycine = 75 Result The amount amino acid (Glycine) present in the whole of the

given unknown solution is = ----------------------- grams

3) ESTIMATION OF SUGAR (GLUCOSE) BY BENEDICT’S TITRIMETRIC METHOD

Aim To estimate the amount of Sugar (Gluose) present in the whole of the given unknown solution Principle When glucose is heated with an alkaline solution of copper (Cu2+) ions, the copper (Cu2+) ions

is reduced to cupric (Cu+) ion, which is precipitated as copper oxide CuO2. This is the basic for the estimation of reducing sugar.

Reagent required Benedict’s quantitative Reagent Crystalline copper sulphate – 18 grams, anhydrous sodium carbonate – 200 grams, potassium

thio cyanate – 125 grams, sodium nitrite – 200 grams are added to 250 ml of distilled water and dissolved with the aid of heat. The contents are finally made up to 800 ml using distilled water.

Standard glucose solution Weighed accurately 250 mg of Glucose and transfer into a 250 ml of standard flask then the

volume is made up to 250 ml using distilled water. (concentration – 1 mg/1 ml)

PROCEDURETitration – I Standard Glucose solution Vs Benedict’s quantitative Reagent Pipette out exactly 5 ml of Benedict’s quantitative reagent into a

clean conical flask. To this add 1 gram of sodium carbonate. The contents are mixed well and heated in boiling water bath till the first bubble

appearance. This is titrated against the Standard Glucose solution taken in the burette. The end point is the disappearance of blue colour. The titrations are repeated for concordant values

Precaution i. Excess amount Sodium carbonate will cause frothing ii.

Benedict’s quantitative Reagent is kept at boiling throughout in the titration iii. Porcelain bits are added to prevent bumping

PROCEDURETitration – II Unknown Glucose Vs Benedict’s Reagent The given unknown Glucose solution is made up to 100 ml standard flask

using distilled water. The burette is rinsed with unknown Glucose solution and filled with the

same unknown Glucose solution. Pipette out exactly 5 ml of Benedict’s quantitative Reagent into a

clean conical flask to this 1gram of sodium carbonate added. The contents are mixed well and

heated till the first bubble appearance. This is titrated against the unknown Glucose solution taken in

the burette. The end point is the disappearance of blue colour. The titrations are repeated for

concordant values.

CALCULATIONcalculation X - Volume of standard Glucose Y - Volume of unknown Glucose ie. --------------------- ml of standard Glucose --------------------- ml of unknown Glucose 100 ml of unknown solution contain = X Y = -------------------- mg

Result The amount of Sugar (Glucose) present in the whole of the given unknown Solution ------------------------ grams

3)ESTIMATION OF ASCORBIC ACID USING 2, 6 DICHLORO PHENOL INDOPHENOL

Aim To estimate the amount of ascorbic acid present in the whole of the

given unknown solution

Principle Ascorbic acid is oxidized by the colourD dye 2, 6 – Dichloro phenol

indophenol to dehydro ascorbic acid. At the same time the dye is reduced to colourless

compound, so that, the end point of the reaction can be easily determined.

REAGENT REQUIRED

2, 6 – Dichloro phenol indophenols Dissolve 42 grams of Sodium bi carbonate and 52 grams of Dichloro phenol

indophenols in 50 ml of distilled water and finally dilute to 250 ml using distilled water.

Stock standard ascorbic acid solution 100 mg of ascorbic acid is weighed exactly and carefully transfer in to 100 ml

standard flask and made up to 100 ml using 0.6 % oxalic acid. Working standard ascorbic acid solution 10 ml of Stock standard ascorbic acid solution is pipette out in to a 100 ml

standard flask and made up to 100 ml using 0.6 % oxalic acid.

PROCEDURE

Titration – I Standard Ascorbic acid Vs Dye Pipette out exactly 10 ml of working standard ascorbic

acid solution into a clean conical flask and it is titrated against the dye taken in the burette. The end point is the appearance of pale permanent pink colour. The titrations are repeated for concordant values.

PROCEDURE

Titration – II Unknown Ascorbic acid Vs Dye The given unknown ascorbic acid solution is made up to

100 ml standard flask using 0.6 % oxalic acid. Pipette out exactly 10 ml of this unknown ascorbic acid solution into a clean conical flask and it is titrated against the dye taken in the burette. The end point is the appearance of pale permanent pink colour. The titrations are repeated for concordant values.

CALCULATION Working standard ascorbic acid = 10 mg ascorbic acid/100

ml of 0.6 % oxalic acid [Concentration – 0.1 mg/1 ml] Ie. 1 ml of standard contains 0.1 mg of ascorbic acid 10 ml standard contains [0.1 mg x 10] – 1 mg of ascorbic

acid 10 ml standard ascorbic acid contains -------------------------- ml

of Dye Ie. 1 mg of ascorbic acid consumes ---------------------------- ml

of Dye

But X of ascorbic acid consumes ------------------------ ml of Dye The amount of ascorbic acid present in the 100 ml of unknown solution = = -------------------- mg The amount of ascorbic acid present in the whole of the given unknown solution = ------------------ mg x 10 = ------------------- mg

Result The amount of ascorbic acid present in the whole of the given unknown Solution

------------------------ mg

4) DETERMINATION OF ACID NO OF FAT

Aim To estimate the amount of Acid no of the given Fat Principle During storage of fat become rancid. As a result the peroxide formation of the double

bond by atmospheric oxygen and or hydrolyzed by micro organism with liberation of free fatty acids. The amount of acid present gives the indication of age and quality of the fat.

Acid value is the number of milligrams of KOH required to neutralize the free fatty acids in one gram of a

fat or oil. It is a measure of free fatty acid contents in a fat or oil. Reagents required i. Fat ii. Fat solvent iii. Standard 0.1 N oxalic acid iv. Potassium hydroxide solution v.

Phenolphthalein as indicator. vi. Methanol / Ethanol

PROCEDURETitration – I Standard oxalic acid Vs Potassium hydroxide solution Weigh accurately 1.575 g of oxalic acid and transfer into a 250 ml of

standard flask then the volume is made up to 250 ml using distilled water. Pipette out exactly 20 ml of this solution into a clean conical flask and two drops of phenolphthalein as indicator is added.

This is titrated against the Potassium hydroxide solution taken in the burette. The end point is the appearance of pale permanent pink colour. The titrations are repeated for concordant values. From the titre value the normality of Potassium hydroxide solution is calculated.

PROCEDURETitration – II Test Value Weigh about one gram of edible oil and carefully

transfer in to a clean dry conical flask. Then 50 ml of alcohol is added followed by two drops of phenolphthalein as indicator is added. The contents are mixed well for 20 minutes.

This is titrated against the standardized Potassium hydroxide solution taken in the burette. The end point is the appearance of pale permanent pink colour and persisting up to 20 – 30 seconds.

PROCEDURE

Titration – III Blank Value 50 ml of alcohol is taken in a conical flask and three drops of

phenolphthalein as indicator is added. The contents are mixed well. This is titrated against the standardized Potassium hydroxide solution taken in the burette. The end point is the appearance of pale permanent pink colour and persisting up to 20 – 30 seconds.

TITRATION – I Volume of Standard oxalic acid solution (V 1) = 20 ml Normality of Standard oxalic acid solution (N1 ) = 0.1 N Volume of Potassium hydroxide solution (V2) = -------- ml Normality of Potassium hydroxide solution (N2) = ? We know that, V1N1 = V2N2 N2 = V1N1 V2 N2 = 20 x 0.1/--------- N2 = ----------------- Normality of Potassium hydroxide solution (N2) = --------------------------- N

100 ml of 1 N Potassium hydroxide contain 56 grams of Potassium hydroxide

X = 56 x Strength of Potassium hydroxide x Test value – Blank value 1000

= --------------------- grams Acid no of Fat = ---------------------- grams x 1000 Weight of Oil = ----------------------------- Acid no of

Fat(Oil) Result Acid No of the given Fat = ------------------------------

5)DETERMINATION OF IODINE NO OF FAT Aim To estimate the amount of Iodine no of the given Fat

Principle Iodine no of fat is defined as the no of grams of iodine absorbed by 100 gram of fat or oil. It is a

measure of degree unsaturation of the fatty acids in a fat or oil. Unsaturated fatty acids, either free or combined in lipids react with halogens like bromine and iodine which get decolorized. These halogens add at the carbon – carbon double bond.

Hane’s method is used for the determination of Iodine number. About 1 gram of the fat is taken in a well cleaned dry iodine flask. To this 20 ml of chloroform is added to dissolve the fat. The contents are

shaken well and kept for 30 minutes. Then 20 ml of potassium iodide is added to liberate the iodine and it is titrated against standard sodium thio cyanate solution. From this titration iodine number of fat is calculated.

REAGENTS REQUIRED

i. Hane’s solution ii. Fat iii. Standard 0.1 N potassium dichromate solution iv. Sodium thio cyanate solution v. 10 % potassium iodide vi. 1 % Starch vii. Chloroform

Preparation of Hane’s solution 3.3 grams of iodine is dissolved in 200 ml of acetic acid by constant

shaking and heating. It is cooled to room temperature, to this 50 ml of glacial acetic acid containing 0.75

grams of Bromine is added and mixed well and stored in brown bottle.

PROCEDURE

Titration – I Standard potassium dichromate solution Vs Sodium thio cyanate solution Weighed accurately 1.225 g of potassium dichromate solution and transfer into a

250 ml of standard flask then the volume is made up to 250 ml using distilled water. Pipette out exactly 20 ml of this solution into a clean conical flask to this 5 ml of Conc. Hydrochloric acid is added, followed by 10 ml of 10 % potassium iodide is added. This contents are mixed well and titrated against the Sodium thio cyanate solution taken in the burette, the titration is continued until a pale brown colour is appears.

At the time 1 ml of 1 % Starch solution is added. And the titration is continued till to get the end point appearance of emerald green colour, it is the end point. The titrations are repeated for concordant values. From the titre value the normality of Sodium thio cyanate solution is calculated.

PROCEDURETitration – II Determination of iodine no of fat (Test value) Weigh about one gram of edible oil

and carefully transfer in to a clean dry iodine flask. Then 20 ml of Chloroform is added, the contents are mixed well to dissolve the oil. To this 20 ml of Hane’s solution is added, shaken well and kept in dark for 30 minutes with occasional shaking. Then the flask is taken out to this 20 ml of 10 % potassium iodide is added to liberate iodine. Except the iodine that is absorbed by the oil.

To this mixture 100 ml of distilled water is added, so the liberated iodine is nicely disturbed in the solvent then it is titrated against the Sodium thio cyanate solution taken in the burette, the titration is continued until a pale brown colour is appears. At the time 1 ml of 1 % Starch solution is added. And the titration is continued till to get the end point disappearance of blue colour it is the end

TITRATION – I

Volume of Standard potassium dichromate solution (V 1) = 20 ml Normality of Standard potassium dichromate solution (N1) = 0.1 N Volume of Sodium thio cyanate solution (V2) = -------- ml Normality of Sodium thio cyanate solution (N2) = ? We know that, V1N1 = V2N2 N2 = V1N1 V2 N2 = 20 x 0.1/--------- N2 = ----------------- Normality of Sodium thio cyanate solution (N2) = --------------------------- N

Blank value Blank value is also done without oil Equivalent weight of iodine = 127 Weight of the oil Weight of the weighing bottle + Oil = Weight of the weighing bottle = ( - ) Weight of the Oil transferred = Blank value - Test value = -------------------- ml Determination of iodine no of fat = Equivalent weight of iodine x Blank value - Test value x Normality of Sodium thio cyanate

solution x 100 1000 x Weight of the Oil = ---------------------------------- Iodine no of fat (oil)

Result Iodine No of the given Fat (Oil) = ------------------------------