BY BY

SRAVAN. SSRAVAN. S

QUANTITATIVE QUANTITATIVE DETERMINATION OF DETERMINATION OF

ALDEHYDES AND KETONESALDEHYDES AND KETONES

INDEXINDEX

INTRODUCTION INTRODUCTION

METHODS OF QUNATITATIVE METHODS OF QUNATITATIVE DETERMINATIONDETERMINATION

SPECTROSCOPIC METHODSSPECTROSCOPIC METHODS

CHROMATOGRAPHIC METHODSCHROMATOGRAPHIC METHODS

Aldehydes and ketones can be determined by using Aldehydes and ketones can be determined by using following 3 reagentsfollowing 3 reagents

1.1. Hydroxylamine hydrochloride – PyridineHydroxylamine hydrochloride – Pyridine

2.2. Sodium sulphite – Sulphuric acidSodium sulphite – Sulphuric acid

3.3. 2,4 – Dinitro phenyl hydrazine2,4 – Dinitro phenyl hydrazine

METHOD - 1METHOD - 1

Hydroxylamine hydrochloride+carbonyl Hydroxylamine hydrochloride+carbonyl compoundscompounds

HCL+OXIMEHCL+OXIME PyridinePyridine Pyridine hydrochloridePyridine hydrochloride

ALDEHYDEALDEHYDE

H HH H

R-C=O + NHR-C=O + NH22OH.HCL R-C=NOH + HOH.HCL R-C=NOH + H22O + HCLO + HCL

H HH H

R-C=O + NHR-C=O + NH22OH.HCL + COH.HCL + C55HH55N R-C=NOH + CN R-C=NOH + C55HH55N.HCL N.HCL + H+ H22OO

KETONEKETONE

RR

R-C=O+ NH2OH.HCL + C5H5N R-C=O+ NH2OH.HCL + C5H5N

RR

R-R-C=NOH+C5H5N.HCL+H2OC=NOH+C5H5N.HCL+H2O

C5H5NHCL + NaOH C5H5N + NaCL + H2OC5H5NHCL + NaOH C5H5N + NaCL + H2O

The reaction of the carbonyl compound with The reaction of the carbonyl compound with hydroxyl amine reaches equilibrium which is hydroxyl amine reaches equilibrium which is displaced to the right in presence of pyridine and displaced to the right in presence of pyridine and excess of hydroxyl amine hydrochloride.excess of hydroxyl amine hydrochloride.

The rate of reaction depends upon the structure The rate of reaction depends upon the structure of –R group,lower aldehydes like and ketones give of –R group,lower aldehydes like and ketones give quantitative reaction in about 30min at room quantitative reaction in about 30min at room temp, wher as ketone like acetophenone, temp, wher as ketone like acetophenone, benzophenone benzoin requires 2hrs heating at benzophenone benzoin requires 2hrs heating at 100 degrees centigrade.100 degrees centigrade.

METHOD-IIMETHOD-II

Carbonyl compounds +sodium bisulphateCarbonyl compounds +sodium bisulphate

Crystalline bisulphiteCrystalline bisulphite

It is determined Iodometrically or by titrationIt is determined Iodometrically or by titration

R-CHO+NaHSOR-CHO+NaHSO3 3 RCH(OH)SORCH(OH)SO33NaNa

METHOD -IIIMETHOD -III

2,4-dinitrophenyl hydrazine+carbonyl compound2,4-dinitrophenyl hydrazine+carbonyl compound

2,4-dinitrophenyl hydrazone 2,4-dinitrophenyl hydrazone NHNH2 NHNH2 NONO2 2 NO NO22

R RR RR-C=O + R-C=N-NH- R-C=O + R-C=N-NH-

NO NO22

NO2 NO2

O O

Determination of ketone Determination of ketone using 2,4-dinitrophenyl using 2,4-dinitrophenyl

hydrazinehydrazine Take 0.2g of camphor in a stopper flask, add Take 0.2g of camphor in a stopper flask, add

25ml of aldehyde free alcohol to dissolve the 25ml of aldehyde free alcohol to dissolve the sample, with swirling add 75ml of 2,4-sample, with swirling add 75ml of 2,4-dinitrophenyl hydrazine reagent and reflux on a dinitrophenyl hydrazine reagent and reflux on a water bath for 4hrswater bath for 4hrs

Remove the alcohol by distillation, cool add Remove the alcohol by distillation, cool add 200ml of 2% v/v solution of Sulphuric acid 200ml of 2% v/v solution of Sulphuric acid

2,4-dinitrophenyl hydrazone is collected in 2,4-dinitrophenyl hydrazone is collected in sintered glass cruciblesintered glass crucible

The ppt is washed with 10ml quantities of The ppt is washed with 10ml quantities of water till filtrate is neutral to litmus.water till filtrate is neutral to litmus.

Dry the ppt at 80Dry the ppt at 80°c cool and weigh °c cool and weigh

The percentage of camphor can be The percentage of camphor can be calculated by the factor given belowcalculated by the factor given below

Each gram = 0.458g of camphorEach gram = 0.458g of camphor

% of camphor = % of camphor = W x 0.458 x 100 W x 0.458 x 100

1 x A1 x A

Determination of Aldehyde Determination of Aldehyde by sodium sulphite reagentby sodium sulphite reagent

250ml of Sodium sulphite solution in a suitable 250ml of Sodium sulphite solution in a suitable stoppered flask, add 50ml of 1N sulphuric acidstoppered flask, add 50ml of 1N sulphuric acid

To the above sol add accurately weighed quantity To the above sol add accurately weighed quantity of about 20-30 mini-equivalent of aldehyde and of about 20-30 mini-equivalent of aldehyde and shake vigorously for 14min. shake vigorously for 14min.

Intorduce the glass-calomel electrode system in Intorduce the glass-calomel electrode system in the beaker and connect to pthe beaker and connect to pH H meter. Titrate the meter. Titrate the excess of acid with standard 1N alkali solution.excess of acid with standard 1N alkali solution.

%% of Aldehyde = of Aldehyde = (y-x) x N x M x 100(y-x) x N x M x 100 W x 1000W x 1000

UV-VISIBLE SPECTROSCOPYUV-VISIBLE SPECTROSCOPY

The carbonyl group contains a pair of The carbonyl group contains a pair of ΠΠ electrons electrons and 2 pairs of non-bonding n or p-electrons, in and 2 pairs of non-bonding n or p-electrons, in addition to a pair of addition to a pair of σσ electrons electrons

Saturated ketones and aldehydes exhibit Saturated ketones and aldehydes exhibit 3absorption peaks .They are 3absorption peaks .They are

1.1.ΠΠ – – ΠΠ** 2. n – 2. n – σσ** 3. R - band3. R - band

Carbonyl groups present in aldehydes and Carbonyl groups present in aldehydes and ketones have significant effect upon the n-ketones have significant effect upon the n-ΠΠ* * transitions. transitions.

The R-band gets shifted to shorter The R-band gets shifted to shorter wavelength with a slight change in intensity. wavelength with a slight change in intensity. The shift in absorption is due to a combination The shift in absorption is due to a combination of inductive and resonance effects.of inductive and resonance effects.

Substitution may change the energy levels of Substitution may change the energy levels of both the ground and excited statesboth the ground and excited states

INFRARED SPETROSCOPYINFRARED SPETROSCOPY

A Strong absorption band in the region around A Strong absorption band in the region around 1700 is due to c=o stretching1700 is due to c=o stretching

The carbonyl groups of aldehydes generally The carbonyl groups of aldehydes generally absorb at 10-15cmabsorb at 10-15cm-1-1higher frequencies than the higher frequencies than the corresponding ketones.corresponding ketones.

Thus 3-pentanone shows a c=o stretch at Thus 3-pentanone shows a c=o stretch at 1715cm1715cm-1-1 ,aliphatic aldehydes absorb near 1740- ,aliphatic aldehydes absorb near 1740-1720cm1720cm-1.-1.

Majority of aldehydes show aldehyde C-H Majority of aldehydes show aldehyde C-H stretching absorptions in the region 2900-2695 stretching absorptions in the region 2900-2695 cmcm-1-1. Two moderately intense bands at are . Two moderately intense bands at are frequently observed in the region 2900-frequently observed in the region 2900-2695cm2695cm-1-1and diferentiate aldehyde class or and diferentiate aldehyde class or compounds from other carbonyl compoundscompounds from other carbonyl compounds

The important difference between an aldehyde The important difference between an aldehyde and ketone is there an aldehyde has a H-and ketone is there an aldehyde has a H-bonded to the carbonyl compounds.bonded to the carbonyl compounds.

The important difference b/w an aldehyde and The important difference b/w an aldehyde and ketone is that an aldehyde has a H-bonded to ketone is that an aldehyde has a H-bonded to the carbonyl carbon.the carbonyl carbon.

The aromatic aldehyde , ex: p-tolualdehyde The aromatic aldehyde , ex: p-tolualdehyde shows its aldehydic C-H stretch at 2830 and shows its aldehydic C-H stretch at 2830 and 2736cm2736cm-1-1

The carbonyl stretching frequency of a pure The carbonyl stretching frequency of a pure sample of saturated aliphatic aldehyde around sample of saturated aliphatic aldehyde around 1725cim1725cim-1 -1 that of a ketone at 1710cmthat of a ketone at 1710cm-1-1 . .

IR FREQUENCIES FOR IR FREQUENCIES FOR ALDEHYDES AND KETONESALDEHYDES AND KETONES

ALDEHYDESALDEHYDES

Group/Vibrations Frequencies cmGroup/Vibrations Frequencies cm-1-1

C = O StretchingC = O Stretchinga.a. Saturated acyclic 1740 – 1720Saturated acyclic 1740 – 1720b.b. Alpha,beta-unstaurated,Alpha,beta-unstaurated, acyclic 1710 – 1685acyclic 1710 – 1685c.c. Aryl 1715 – 1695Aryl 1715 – 1695 C – H stretching 2900 – 2695C – H stretching 2900 – 2695 C – H bending 975 - 780C – H bending 975 - 780

KETONES:KETONES:

C =O StretchingC =O Stretching

a.a. Saturated acyclic 1725 – 1705Saturated acyclic 1725 – 1705

b.b. Alpha,beta unsaturated ,Alpha,beta unsaturated ,

acyclic 1685 – 1665acyclic 1685 – 1665

c.c. Aryl 1685 – 1665Aryl 1685 – 1665

C – H stretching 3000 - 2900C – H stretching 3000 - 2900

MASS SPECTORMETRYMASS SPECTORMETRY

Some important features of the mass spectra of Some important features of the mass spectra of aliphatic aldehydes and ketones arealiphatic aldehydes and ketones are

a.a. The Intensity of the molecular ion peak The Intensity of the molecular ion peak decreases as the alkyl chain length increasesdecreases as the alkyl chain length increases

b.b. The major fragmentation processes are The major fragmentation processes are άά and and ββ clevage .clevage .

c.c. In aldehydes and ketones containing In aldehydes and ketones containing γγ--hydrogen atom.McLafferty rearrangement ion is hydrogen atom.McLafferty rearrangement ion is most significant. In an aldehyde, which is not most significant. In an aldehyde, which is not άά--substituted , apeak due to this is fored at m/e substituted , apeak due to this is fored at m/e 44. It may be base peak.44. It may be base peak.

In lower aldehydes, In lower aldehydes, άά- clevage is prominent - clevage is prominent with retention of clevage on oxygen.with retention of clevage on oxygen.

In Aromatic aldehydes and ketones parent ion In Aromatic aldehydes and ketones parent ion peak is intense.peak is intense.

In ketones, the loss of larger group is preferred In ketones, the loss of larger group is preferred by a by a άά- clevage.- clevage.

HPLCHPLC

KETONEKETONE:: Formulation: Thymol,methylsalicylate, Formulation: Thymol,methylsalicylate, Camphor and salicylic acid. Camphor and salicylic acid.

Sample preparation: Sample suspended in 100ml of Sample preparation: Sample suspended in 100ml of solvent, warmed in water bath(60c) and shaken for solvent, warmed in water bath(60c) and shaken for 10min. Centrifuged and supernatant used for analysis.10min. Centrifuged and supernatant used for analysis.

Standard Preparation: Solution of each ingredient is Standard Preparation: Solution of each ingredient is prepared in solvent. [water-THF- methanol]prepared in solvent. [water-THF- methanol]

Internal Standard: Diphenhydramine Hcl - Internal Standard: Diphenhydramine Hcl - 100100µg/ml in solvent.µg/ml in solvent.

HPLC System: Shimadzu LC system.HPLC System: Shimadzu LC system.

Column: Shimpak CLC – DDS Column, Column: Shimpak CLC – DDS Column, 150 x 6mm150 x 6mm

Chromatographic conditions: Compositon of Chromatographic conditions: Compositon of mobile phase and its Pmobile phase and its PHH: 200mM sodium : 200mM sodium perchlorate in 10mM sodium phosphate. Pperchlorate in 10mM sodium phosphate. PHH: 2.6 – : 2.6 – Methanyl acetonitrileMethanyl acetonitrile

Flow rate: 1.5ml/minFlow rate: 1.5ml/min

Volume Injected: 20 Volume Injected: 20 µlµl

Wave length: UV – 210nmWave length: UV – 210nm Retention time: Retention time:

Salicylic acid - 3Salicylic acid - 3

Diphenhydramine Hcl - 3.9Diphenhydramine Hcl - 3.9

Methyl Salicylate - 6.1Methyl Salicylate - 6.1

Camphor - 7.2Camphor - 7.2

Thymol - 9.5Thymol - 9.5

ALDEHYDEALDEHYDE Formulation: Betamethasone valerate, clotrimazole, Formulation: Betamethasone valerate, clotrimazole,

Benzaldehyde.Benzaldehyde.

Sample Preparation: weigh 0.64mg of Sample Preparation: weigh 0.64mg of Betamethasone or 10mg of clotrimazole, add 2ml of Betamethasone or 10mg of clotrimazole, add 2ml of standard solution and 8ml of methanol. Heat in standard solution and 8ml of methanol. Heat in waterbath(60c) for 10min,cool,centrifuge and filter.waterbath(60c) for 10min,cool,centrifuge and filter.

Standard Preparation: Betamethasone dipropionate - Standard Preparation: Betamethasone dipropionate - 320320µg/mlµg/ml

clotrimazole - 5mg/mlclotrimazole - 5mg/ml Both the solutions are prepared in methanol, mixed Both the solutions are prepared in methanol, mixed

standard solutionstandard solution

Column: Suplex PKB – 100 column, 250 x 4.6mmColumn: Suplex PKB – 100 column, 250 x 4.6mm Conditions: Composition of MP and its PConditions: Composition of MP and its PHH

Acetonitrile – Tetrahydrofuran – 15mmAcetonitrile – Tetrahydrofuran – 15mm Acetate buffer – PAcetate buffer – PHH – 4 – 4 Flow rate: 2ml/minFlow rate: 2ml/min Volume Injected: 10Volume Injected: 10µlµl Wave length: U.V – 254nmWave length: U.V – 254nm Retention time: Benzaldehyde – 3.3Retention time: Benzaldehyde – 3.3 clotrimazole - 7.7clotrimazole - 7.7

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