QUALITATIVE TESTS, QUANTITATIVE TESTS, DIFFERENT FORMS OF ALKALOIDS AND THEIR CLASSIFICATION, OF ALKALOIDS.

1. Assignment no-3
Qualitative and quantitative analysis of al
Presented to:
Dr. Inder Pal Singh
Presented by:
D. Gnanabhaskar
16TMM2364
NIPER, Mohali. SAS NAGAR

2. Introduction
Forms of alkaloids&
their classification
Qualitative and quantitative analysis
CONTENTS
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INTRODUCTION
Because alkaloids are so heterogeneous chemically and there are so many of them, they
cannot be identified in plant extracts using a single chromatographic criterion. In
general, because of the wide range of solubility and other properties of the alkaloids,
any general screening procedure for alkaloids in plants may fail to detect particular
compounds.
J. B Harborne, “Phytochemical Methods” 1978(3)203-213

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Forms of alkaloids& their classification
Sr.no Alkaloidal form Example
1 Salts with special acids Meconic acid in Opium& Quinic acid in Cinchona
2 Steroidal Glycosidal
alkaloid.
Solanine in Solanum
&chaconine
3 Terpenoidal Alkaloids. Taxol (diterpinoidal alkaloid)
J. B Harborne, “Phytochemical Methods” 1978(3)203-213

5. Dr. Mazen A. El-Sakka “Phytochemistry of Alkaloids” 2010(3) Al Azhar University Faculty of Pharmacy Department of Pharmacognosy. 5
Classification
According to Hegnauer's classification, which is based on both, the type of nitrogen and the biochemical
origin, three main types of alkaloids are distinguished:
1. True alkaloids: that are derived from amino acids and have nitrogen in a heterocyclic ring.
2. Protoalakloids :that are derived from amino acid and do not have nitrogen in heterocyclic ring.
3. Pseudo alkaloids: that are not derived from amino acids but have nitrogen heterocyclic ring.
Different systems of classification may be adopted based on:
1. The pharmacological action (biological activity.)
2. The chemical structure (type of nitrogen, heterocyclic or non heterocyclic and type of ring structure.)
3. The biochemical origin (biosynthetic pathway of production in the plant.)
4. The taxonomical origin (plant families rich in alkaloids.)

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Volumetric methods:
These are based on reaction of alkaloidal bases with acids (Acid-Base titration).
They include:
Aqueous titration: This is carried by either:
1- Direct titration of the alcoholic solution of the alkaloidal residue with standard acid, or
2- Back titration by dissolving the residue in a known amount of standard acid and back titration of
residual acid against standard alkali.
Non-aqueous titration: This method is suitable for determination of weak bases e.g. Caffeine.
ed.
Quantitative Determination of Alkaloids

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Gravimetric methods:
They can be performed by either:
1- Direct Weighing of the alkaloidal mixtures
2- Precipitation of the total alkaloids and determination of the weight of the precipitate obtain
Spectrophotometric Methods.(Spectrophotometric Method for Estimation of Alkaloids Perceptible with Dragendorff’s
Reagent in Plant Materials)
Polarimetric Method.(The colorimetric determination of various alkaloids Authors F. J. Bandelin)
Fluorimetric Method.(Simplified fluorimetric determination of digitalis alkaloids Datta V. Naik.)
Chromatographic Methods.

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9. 4/17/2017 J. B Harborne, “Phytochemical Methods” 1978(3)203-213 9
Extract dried tissue with 10% acetic acid in ethanol, leaving to stand for at least 4h. Concentrate the extract to one-
quarter of the original volume and precipitate of the alkaloid by drop wise addition of conc.NH40H. Collect by
centrifugation, washing with 1% NH4OH. Dissolve residue in a few drops of ethanol or chloroform.
Chromatograph an aliquot on sodium citrate-buffered paper in n-butanol-aqueous citric acid. Chromatograph another
aliquot on silica gel G plates in methanol-conc.NH40H (200: 3). Detect the presence of alkaloids on the paper and
plate, first of all by any fluorescence in UV
light and then by application of three spray reagents - Dragendorff, iodoplatinate and Marquis. RFs and colours of
twelve of the commonest
alkaloids are shown in below table.
Confirm the presence of a particular alkaloid by measuring the UV spectrum of a sample dissolved in 0.1 M H2SO4,
Typical maxima values range from 250 to 303nm Alkaloids with aromatic rings in their structures may also absorb at
longer wavelengths, e.g. colchicine"-max 243 and 351 nm.
Sample preparation:

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11. 4/17/2017 J. B Harborne, “Phytochemical Methods” 1978(3)203-213
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This reagent was invented by the German Pharmacologist, Johann Georg Dragendorff (1836–
1898). Dragendroff It is a color reagent to detect alkaloids in a test sample. Alkaloids, if present in
the solution of sample, will react with Dragendorff’s reagent and produce an orange or orange
red precipitate.
Composition: Dragendroff’s reagent is a solution of potassium bismuth iodide prepared from
basic bismuth nitrate(Bi(NO3)3), tartaric acid, and potassium iodide (KI).
Dragendorff’s test
VIDEO OF THE TEST DOWNLOAD TO SEE

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Wagner's test
.
When reagent treated with sample Reddish brown
precipitate confirms the presence of alkaloids
Wagner’s Reagent (Potassium Triiodide):
Iodine = 1.3 g
Potassium = 2.0 g
Distilled water to make = 100.00 ml

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SAMPLE
HgCl2
KI It is freshly prepared by dissolving a mixture of mercuric chloride (1.36 g)
and of potassium iodide (5.00 g) in water (100.0 ml).
Most alkaloids are precipitated from neutral or slightly acidic solution by
Mayer's reagent (potassiomercuric iodide solution) to give a cream coloured
precipitate
Mayer's test

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HCl
AMMONIA
VAPORS
Caffeine
AMMONIA
SOLN/VAPORS
Murexide test
VIDEO OF THE TEST DOWNLOAD TO SEE

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Hagers
reagent
2ml
Sample
filtrate
Yellow
colored
precipitate
Hagers test
J. B Harborne, “Phytochemical Methods” 1978(3)203-213

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Solid alkaloid + fuming HNO3 → Evaporate to dryness, dissolve residue in acetone, add methanolic
solution of KOH → Violet colour.
Vitali-Morin’s test-Tropane alkaloids
VIDEO OF THE TEST DOWNLOAD TO SEE

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Libermann’s test
This test is very useful in distinguishing various alkaloids. It is 10%
solution of KMnO4 in H2SO4. Whenever a microdrop of this solution is
applied to given sample of alkaloid, the following different colour will
show the alkaloidspresence.
1. Atropine, Hyoscyamine, Hyoscine, Ephedrineq Orange colour.
2. Papaverine, Emetine Black colour.
3. Cocaine, Physostigmine Red colour.
4. Colchicine Yellow colour.
5. Yohimbine Blue colour.

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1. Dissolve 5-10 mg of the alkaloid in a few drops of hot amyl alcohol or ethanol and allow it cool gradually.
The appearance of jelly-like product gives the characteristic test of the solanum alkaloids.
2. When a few mg of the alkaloids is treated with antimony trichloride solution in dry chloroform, it gives
rise to a distinct red colouration.
The solanum alkaloids, in general, produces an instant red-violet colour with formaldehyde (HCHO) and
sulphuric acid (H2SO4). This particular test is so distinct and sensitive that it is used for the quantitative
estimation of these alkaloids colorimetrically.
Steroidal alkaloids

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1. J. B Harborne, “Phytochemical Methods” 1978(3)203-213
2. William Charles Evans “Trease and Evans' Pharmacognosy,” (16)301-370
3. Dr. Mazen A. El-Sakka “Phytochemistry Alkaloids” 2010(3) Al Azhar University Faculty of Pharmacy Department
of Pharmacognosy.
4. Janet travell, “The potassium mercuric iodide reagent for alkaloids.” From the Laboratory of Pharmacology,
University of Illinois, College of Medicine, andassisted by a grant from the American pharmaceutical association
July, 1934, 689-698.
5. Datta v. Naik “Simplified fluorimetric determination of digitalis alkaloids” J Aalytica Chemica Acrta,, 74 ( 1975)
29-33.
References