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Prof. Dr. Amani S. Awaad
Professor of PharmacognosyPharmacognosy Department,
College of Pharmacy Salman Bin Abdulaziz
University,
Al-Kharj. KSA.
Email: [email protected]
Pharmacognosy -2
PHG 322
Definition and Description of Alkaloid
*Physical & chemical properties
Pharmacological Activity And Uses of alkaloids
*Naming of Alkaloids
Extraction of Alkloids
Classifications of the alkaloids
Alkaloids
No definitive difference between an
alkaloid and naturally occurring
complex amines.
Alkaloids:
the term “alkaloid” (alkali-
like) is commonly used to
designate basic
heterocyclic nitrogenous
compounds of
plant origin that are
physiologically active
Definition
Deviation from Definition:
Basicity: Some alkaloids are not basic
e.g. Colchicine, Piperine,
Quaternary alkaloids.
Nitrogen: The nitrogen in some
alkaloids is not in a heterocyclic ring
e.g. Ephedrine, Colchicine,
Mescaline.
Plant Origin: Some alkaloids are
derived from Bacteria, Fungi,
Insects, Frogs, Animals.
• Alkaloids occur only exceptionally in the bacteria (pyocyaninefrom Pseudomonas aeruginosa)
• They are rarely in the fungi (psilocin from the hallucinogenic mushrooms of Central America, ergolines from Claviceps and other actinomycetes, sporidesmins, roquifortine, and others).
• The Pteridophytes rarely contain alkaloids, and among them the Lycopodiaceae represent the main exception (alkaloids derived from lysine); the same comment applies to the Gymnosperms (alkaloids from Cephalotaxus).
• Thus, alkaloids are compounds essentially found in the Angiosperms, and some authors estimate that 10 to 15% of these synthesize those products
Occurrence, distribution, Localization & Functionof alkaloids
Alkaloids cont…
Occurrence
Occurrence, distribution, Localization & Function of alkaloids
• Certain families have a marked tendency to elaborate alkaloids: this is true for the Monocotyledons, Dicotyldons
• Within these families, some genera produce alkaloids and others do not. Sometimes, they are found in all of the genera (Papaveraceae), although this is far less common.
• Alkaloids concentrations have a wide range of variation: from ppm as in the case of the anticancer alkaloids of Catharanthus roseus to more that 15% in the bark of the trunk of Cinchona ledgeriana.
• rarely do alkaloid-containing plants contain only one alkaloid: sometime they do contain virtually only one constituent (e.g. hyoscyamine from the leaves of belladonnae), but, most often, they yield a complex mixture, which may be dominated by one major constituent.
• It is not uncommon to find several dozen alkaloids in one drug.
Alkaloids cont…
Distribution
Occurrence, distribution, Localization & Function of alkaloids
Alkaloids cont…
Localization
In the plant, alkaloids occur as soluble salts (citrates, malates, tartrates, meconates, isobutyrates, benzoates), or in combination with tannins.
Microchemistry shows that alkaloids are most often localized in the peripheral tissues: external layers of the bark of the stems and roots, or seed tegument.
Most of these compounds have a basic character and Most often synthesis takes place at specific sites (growing root, latcicferous cells, and chloroplasts)
The compounds are subsequently transported to the storage sit, which may be specialized or not. They are normally stored in the cell vacuoles,
Occurrence, distribution, Localization & Function of alkaloids
Alkaloids cont…
Function
Poisonous agents that protect the plants against insects, fungus,
predators
• End products of nitrogen detoxification
• Regulate growth factors
• Reserve substances which can supply nitrogen when needed
As in the case of many other secondary metabolites,
almost nothing is known of the role of alkaloids in plants
Although some authors believe that alkaloids are terminal metabolites, in other words unusable waste, this is highly unlikely: in several cases, they have been shown to act as intermediate metabolites.
The question remains unanswered.
Naming of AlkaloidsAlkaloids cont…
1-The genus of the plant, such as Atropine from Atropa belladona.
2-The plant species, such as Cocaine from Erythroxylon coca.
3-The common name of the drug, such as Ergotamine from ergot.
4-The name of the discoverer, such as Pelletierine that was discovered by Pelletier.
5-The physiological action, such as Emetine that acts as emetic.
6-A prominent physical character, such as Hygrine that is hygroscopic.
7-Other. such as morphine derived from ancient Greek mythology – Morpheus – god of dreams
Trivial names should end by "ine". These names may refer to:
Prefixes and suffixes:
Prefixes:
• "Nor-" designates N-demethylation or N-demethoxylation, e.g.norpseudoephedrine and nornicotine.
• "Apo-" designates dehydration e.g. apomorphine.
• "Iso-, pseudo-, neo-, and epi-" indicate different types ofisomers.
Suffixes:
• "-dine" designates isomerism as quinidine and cinchonidine.
• "-ine" indicates, in case of ergot alkaloids, a lowerpharmacological activity e.g. ergotaminine is less potent thanergotamine.
Physical-Chemical Properties of Alkaloids
A)Physical Properties
II- Color:
The majority of alkaloids are colorless but some are colored e.g.: Colchicine and berberine are yellow. Canadine is
orange. The salts of sanguinarine are copper-
III- Solubility:
Both alkaloidal bases and their salts are soluble in alcohol.
*Generally, the bases are soluble in organic solvents and insoluble in water
Exceptions:Bases soluble in water: caffeine, ephedrine, codeine, colchicine, pilocarpine and quaternary ammonium bases.
Bases insoluble or sparingly soluble in certain organic solvents: morphine in ether, theobromine and theophylline(in benzene).
*Salts are usually soluble in water and, insoluble or sparingly soluble in organic solvents.
Exceptions:
Salts insoluble in water: quinine monosulphate.
Salts soluble in organic solvents: lobeline and apoatropine hydrochlorides are soluble in chloroform.
I- Condition:
Most alkaloids are crystalline solids. Few alkaloids are amorphous solids e.g. emetine. Some are liquids that are
either:(Volatile e.g. nicotine and coniine, or Non-volatile e.g. pilocarpine and hyoscine).
Physical-Chemical Properties of Alkaloids
A)Physical Properties
Optically active isomers may show different physiological activities.
IV- Isomerization:
l-ephedrine is 3.5 times more
active than d-ephedrine.
l-ergotamine is 3-4 times more active than
d-ergotamine.
d- Tubocurarine is more
active than the
corresponding l- form.
Quinine (l-form) is antimalarial
and its d- isomer quinidine is
antiarrythmic.
The racemic (optically inactive) dl-
atropine is physiologically active.
Physical-Chemical Properties of Alkaloids
B) Chemical PropertiesI- Nitrogen:
Primary amines R-NH2 e.g. Norephedrine
Secondary amines R2-NH e.g. Ephedrine
Tertiary amines R3-N e.g. Atropine
Quaternary ammonium salts R4-N e.g d-Tubocurarine
II- Basicity:
R2-NH > R-NH2 > R3-N
Saturated hexacyclic amines is more basic than aromatic amines.
According to basicity Alkaloids are classified into
Weak bases e.g. Caffeine
Strong bases e.g. Atropine
Amphoteric: Phenolic Alkaloids e.g. Morphine
Alkaloids with Carboxylic groups e.g. Narceine
Neutral alkaloids e.g. Colchicine
III- Oxygen
Most alkaloids contain Oxygen and are solid in nature e.g. Atropine.
Some alkaloids are free from Oxygen and are mostly liquids e.g. Nicotine, Coniine
Physical-Chemical Properties of Alkaloids
Bitter tasting,
Most alkaloids are well-defined crystalline substances which unite with acids to form salts.
In plants, they may exist in the free state, as salts or as N-oxides
Most solid bases rotate the plane of polarized light, have high
melting points.
Alkaloids which do not contain O2 are liquid at room
temperature (nicotine), while those that do are solids in
rare cases they are coloured.
Strychnine N-oxide
Alkaloids cont…
* Although the concept of alkaloid is relatively recent, the knowledge of the toxicity
and properties of the plants containing them dates back to ancient times: opium, coca,
aconite, belladonna, colchicum, as well as cinchona, ipecac, and curare have been used
for centuries, if not millennia
History
*It was probably Derosne, who, while extracting a mixture of narcotine and morphine from opium in 1803, was the first to isolate a vegetable alkali.
*In 1806, Serturner recognized the alkaline nature of the somniferous principle of opium, which he named morphine about ten years later.
*Shortly afterwards, between 1817 and 1820, two French pharmacists, Pelletier and Caventou, "discovered" an impressive series of active compounds:
Caffeine, emetine, from ipecac, strychnine from nux vomica, quinine and cinchonine from cinchona bark, followed a little later by coniine.
* The first isolations of alkaloids were in the nineteenth centaury. Prior to
approximately 300 years ago the first one was NICOTINE .
Alkaloids cont…
History
Chemists attempted to elucidate the structure of these molecules veryearly on: in the simplest cases, they were successful (coniine, Schiff,1870).
but in other cases they had to wait until the second half of the twentiethcentury:
the polycyclic edifice of strychnine "resisted" the endeavors of chemicalinvestigators for nearly 130 years!
Today, advanced NMR techniques and X-ray diffraction spectrometryallow the elucidation of the most complex structures.
The synthesis of these compounds also represented a challenge forchemists early on: from the synthesis of coniine at the end of thenineteenth century to that of morphine in 1952,
this challenge played- and it continues to play- a key role in thedevelopment of organic chemistry
• Alkaloids are particularly interesting substances because of their multiple pharmacological activities:
On the CNS, whether they are: depressants (morphine, scopolamine) or stimulants (strychnine, caffeine);
On the autonomic nervous system:
sympathomimetics (ephedrine) or sympatholytic (yohimbine, certain ergot alkaloids), parasympathomimetic (eserine, pilocarpine), anticholinergics (atropine, hyoscyamine), or ganglioplegics (sparteine, nicotine).
Pharmacological Activity And Uses of alkaloids
In addition they have the following uses, - alkaloids include curare, - local anesthetics (cocaine), - and agents to treat fibrillation (quinidine),- antitumor agents (vinblastine, ellipticine),- antimalarials (quinine),- antibacterials (berberine), - and amebicides (emetine).
Although some are only used as galenicals (belladonna, datura, henbane),many others are only used as starting materials for industrial extraction
Tests for Alkaloids
They give a precipitate(from neutral or slightly acidic) with heavy metal iodides. Caffeine, a purine derivative, does not precipitate like most alkaloids
Mayer's reagent (potassiomercuric iodide
solution) Cream coloured precipitate
Wagner’s reagent (iodine in potassium
iodide) red-brown precipitate
Alkaloids cont…
Dragendorff's reagent (solution of potassium
bismuth iodide) orange coloured precipitate
Hagers reagent (picric acid)
yellow precipitate
The specificity of these reagents is
not absolute:
proteins, α-pyrones, some coumarins,
hydroxyflavones, lignans, and other
compounds can give false positive
reactions with Dragendorff's reagent
Extraction
TLC plate
Extraction method normally depends on the raw material, the purpose of extraction & the scale on which is to be performed.
For research purposes: chromatography allows for quick and reliable results.
If larger amounts of alkaloids need to be extracted, one of the following methods can be used.
Extraction is based on the basicity of alkaloids and on the
fact that they normally occur in plants as salts (i.e.: on the
solubility of bases and salts in water and organic solvents).
Herbs often contain other materials which can interfere with
extraction such as large amounts of fat, waxes, terpenes,
pigments and other lipophilic substances (e.g by forming
emulsions) – avoided by defatting the crushed herb (using
petroleum ether and hexane
Alkaloids cont…
General Method
-Powdered, defatted herb is mixed with an alkalineaqueous solution.
-Free bases are then extracted with organic solvents(chloroform, ethyl acetate, alcohol).
-Organic solvent containing alkaloids (bases) is separatedfrom residue & concentrated by distillation underpressure if needed
Step I:
Alkaloids cont…
Step II:
-Solvent is stirred with an acidic aqueous solution: alkaloids go into the solution as salts. Impurities remain in the organic phase.
-Aqueous solution of alkaloid salts is washed with an apolarsolvent (hexane)
-Alkalinized with a base using an organic solvent not miscible with water.
-Alkaloids precipitate and dissolve in the organic phase.
-Extraction of aqueous phase continues till all alkaloids have moved into the organic phase (tested when Mayer’s reaction on the aqueous phase becomes negative).
-This purification step may be carried out in a separation funnel or in centrifugal extractors
Step II:
General MethodAlkaloids cont…
Step III:
Organic solvent containing alkaloid bases is decanted, freed from water traces (drying over anhydrous salt e.g. sodium sulphate) and evaporated under reduced pressure.
A dry residue remains: total basic alkaloids.
their are two Methods possible
1-Plant powder is extracted directly with acidified water
2-Plant powder is extracted with acidified alcoholic or a hydroalcoholic solution. This is then followed by distillation under
vacuum (eliminates that alcohol, leaving behind and acidic aqueous solution of alkaloid salts)
Extraction of liquid alkaloids
Alkaloids cont…
Specific Method
Diagram for extraction of alkaloids
alcohol extraction
alkaloids
Sugars
Soluble peptides
Protein
Plant pigments
Cellulose
fibers
Extract
residue
Water layer
CHCl3 layerAll undesired materials
Alkaloids as salt
1. anhydrous sodium sulphatee
2. concentrate
Plant powder
1.Consetration
2.Desolve in dil. acid
Add Alkaline NH4OH
CHCl3 layer
Water layerQuaternary ammonium salt
Row alkaloids
Alkaloids cont…
Extraction of Alkaloids
• Process remained unchanged >1,000 years
Plant Material
Acid solution
EtOAc: neutral/weakly basic alkaloids
1) Methanol
2) Concentrate
3) Partition EtOAc/2% acid
Petroleum ether extracts non-polar fats and waxes
Residue: polar material
Wash with petroleum ether
Basic aqueous solution of quaternary alkaloids
1) Ammonia
2) Partition with EtOAcEtOAc: basic alkaloids
Test of Alkaloids
I-Qualitative test for Alkaloids:
They are used to:
1- Indicate the absence or presence of Alkaloids
2- Test for complete of extraction
Disadvantages: The specificity of these reagents is not absolute:
proteins, α-pyrones, some coumarins, hydroxyflavones, lignans,
and other compounds can give false positive reactions with
Dragendorff's reagent
Colour Reagents
1- Froehd’s Reagent: Phosphomolybdic acid
2- Marqui’s Reagent: Formaldehyde/ Conc. H2SO4
3- Mandalin’s Reagent: Sulphovanidic acid
4- Erdmann’s Reagent: Conc. HNO3/Conc. H2SO4
5- Mecke's Reagent: Selenious acid / conc. H2SO4
6- Shaer's Reagent: Hydrogen peroxide / conc. H2SO4
7- Rosenthaler's Reagent: Potassium arsenate / conc. H2SO4
8- Conc. HNO3
Precipitation Reagents:
Mayer's reagent (potassiomercuric iodide
solution) Cream coloured precipitate
Hagers reagent (picric acid) yellow
precipitate
Dragendorff's reagent (solution of potassium
bismuth iodide) orange coloured precipitate
Wagner’s reagent (iodine in potassium
iodide) red-brown precipitate
Test of Alkaloids
II-Quantative test for Alkaloids:
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, or2- 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.
Gravimetric methods:These methods are recommended for determination of:
1- Very weak bases which can not be determined by volumetric methods e.g. caffeine andcolchicine.2- Mixtures of alkaloids that are obtained from the same plant but differ greatly in their molecularweight e.g. Cinchona and Rawolfia alkaloids.
Colourimetric Method:
e.g. Morphine + NaNO2/HCl
Ergot + p-dimethylaminobenzaldehyde
Spectrophotometric Methods.
Polarimetric Method.
Fluorimetric Method.
Chromatographic Methods
Isolation of Alkaloids
All methods of alkaloid extraction yield impure compounds, so alkaloids therefore have to be separated.
TLC and HPLC are most commonly used.
Chromatography is used for complex alkaloids and if only
small amounts of alkaloids are needed.
Large-scale extractions may be sent to a factory for purification & separation (for cinchona and cocoa alkaloids).
Separation and final purification may be done using fractional precipitation or fractional crystallization of salts.
Chromatograpy is used for complex alkaloids and if only small amounts of alkaloids are needed
Alkaloids Classification
Alkaloids have a large variety in their botanical & biochemical origin, in chemical structure and pharmaceutical action. Therefore a range of different classification systems exist for grouping alkaloids.
They can therefore be classified according to their
1-General classification
2-Biological uses
3-Botanical Source (According to the plant source of alkaloids.)
Type of Amines.(Primary, Secondary, Tertiary alkaloids.)
4-Biosynthetic or biogenetic pathway (Based on the biogenetic pathway that form the alkaloi)
5-Basic Chemical Skeleton ( 2 divisions)
• i. Atypical/non-hetrocyclic alkaloids (protoalkaloids or biological amines)
• ii. Typical/hetrocyclic alkaloids (divided into 14 groups according to their ring structure)
Alkaloids cont…
Alkaloids Classification
Alkaloids cont…
1-Hagnauer system of classification
1) True (Typical) alkaloids that are derived from
amino acids and have nitrogen in a
heterocyclic ring. e.g Atropine
2) Proto alkaloids that are derived from amino
acids and do not have nitrogen in a heterocyclic
ring. e.g Ephedrine
3) Pseudo alkaloids that are not derived from
amino acids but have nitrogen in a heterocyclic
ring. e.g terpenoidal alkaloids
1-Vasodilator
(eg. Ephedrine,
Ergonovine)
3-Local
anesthetic
(eg. Cocaine)
4-Hallucinating (eg.
Mescaline, Psilocybin,
LSD)
2-Biological origin
(eg. Ephedrine, Ergonovine)
2-Sedatives
Alkaloids cont…
Alkaloids Classification
2-Biosynthetic pathway
Alkaloids Classification
Alkaloids cont…
Biosynthetic pathways to alkaloids
vary greatly depending on the
nitrogen source. Alkaloids are
biosynthesized from various kinds
of amino acids
1-Ornithine derived alkaloids
2-Tryptophan derived alkaloids
3-Lysine derived alkaloids
4-Phenylalanine derived alkaloids
5-Tyrosine derived alkaloids
6-Histidine derived alkaloids
7-Alkaloids derived from substance
other than amino acids.
4-Chemical classification
i. Atypical/non- heterocyclic alkaloids (proto alkaloids or biological amines)
Alkaloids cont…
Alkaloids Classification
Terpenoids & Steroidal (Steroids, Aconite)
Tropolone(Colchicine)
phenylalkylamine(Mescaline, ephedrine, hordenine, capsaicinoids, cathinone )
ii. Typical/hetrocyclic alkaloids (divided into 8groups
according to their ring structure)
Alkaloids cont…
Alkaloids Classification
4-Chemical classification
Opium Alkaloids
Carboline
1-Imidazole: e.g. pilocarpine
2-Indole: e.g.ergometrine
3- Isoquinoline: e.g.Morphine
4-Quinoline : e.g.quinine and quinidine
5-Purine : e.g. caffeine
6-Pyridine & piperidine: e.g. lobeline, nicotine
7-pyrrolyzidine e.g.sthrahidrine, platyphylline
8-Tropane e.g. Atropine.
Amino Acid Precursors
N
N
CH3
nicotine
from ornithine NH2
OCH3
H3CO
H3CO
mescaline
fromtyrosine
N
H3C
CO2CH3
O Ph
Ococaine
from ornithine
N
O
N
O
strychnine
from tryptophan
O
HO
N
CH3
HO
morphine
fromtyrosine N
NH
HO2C CH3
lysergic acid
from tryptophan NH
N
HO
H3CO
H
from tryptophan
NH
N
HO
MeO2C
NMeO
N
OH
CO2CH3
O
O
R
R= -CH3 vinblastineR= -CHO vincristine
from tryptophan
N OH
H3C
Lycopodine
from lysine
NH
OH
Histrionicotoxin
N
COOH
Nicotinic acid
N
PYRIDINE
HOOC
NH2
COOH
Glutamic acid
HOOC
O
COOH
-Ketoglutaric acid
H2NH2N
COOH
Ornithine
N
PYRROLIDINE
N
PYRROLIZINE
N
TROPANE
HOOC
NH2
COOH
-Aminoadipic acid
HOOC
NH2
COOH
Aspartic acid
NH2
HOOC
NH2
COOH
Diaminopimelic acid H2NH2N COOH
Lysine
N
N
INDOLIZIDINE
N
QUINOLIZIDINE
COOH
HO
OH
OH
SHIKIMIC ACID
COOH
OH
O COOH
CHORISMIC ACID
COOH
NH2
ANTHRANILIC ACID
N
QUINOLINE
N
N
QUINAZOLINE
N
O
BENZAXAINE
O
HN
OP
OH
OH
COOH
HOOC
COOH
OH
PREPHENIC ACIDAmino-acids and
origin of the chief
heterocyclic rings
NH2
COOH
R
PHENYLALANINE-TYROSINE
N
ISOQUINOLINE
NH
COOH
NH2
TRYPTOPHAN
N
INDOLE
N
NH
NH2
COOH
HISTIDINE
N
N
IMIDAZOLE
NUCLEOTIDESN
NN
N
PURINE
