A. W.
Sangstor
University College
(1 Determination of Alkaloid Structures
I
of the West lndies I. Isolation, characterization, a n d
Jamaica
physical methods
There are several well-known books on
the chemistry of the alkaloids (I), but these tend to
discuss the detailed chemistry of particular groups of
alkaloids. It is the purpose of these articles to outline
the general methods of approach in alkaloid chemistry.'
The term "alkaloid" (alkali-like) is applied to the
large group of basic nitrogen-containing natural prod-
ucts of vegetable origin. They are usually colorless
(berberine is yellow), well-crystalline solids, but some
from the tobacco, hemlock, and pomegranate root bark
are liquids,. The bases are usually optically active
(mainly levorotatory) though a few such as coniine,
cinchonine, laudanosine, pilocarpine, and pelletierine
are dextrorotatory,andothers like berberine,papaverine,
and pilocereine are optically inactive. I n most cases
the nitrogen atom forms an integral part of a cyclic
structure. Classifications of alkaloids usually exclude
simple amines, purines, hetaines derived from amino
acids of proteins, choliie, and other bases of biological
importance which in contradistinction to the typical
alkaloids are not limited to one or a few species of
plants.
Alkaloids occur most abundantly in the higher orders
of the plant kingdom and have been found in over 90
different families of flowering plants (2).
The majority of these families belong to dicotyl-
edenous orders and the number of monocotyleden~us
families showing the presence of alkaloids is relatively
The present article deals with methods of isohtion and char-
acterization and the applications of physics1 methods to the de-
termination of alkaloid structures. The second paper, to be
published in October, will describe chemical methods of
.structural determination.
454 / Journal of Chemical Education
small. The Gymnosperms and Pteridophytes are poorly
represented and so far as is known alkaloids are virtually
absent from the lower groups of plants with the excep
tion of one or two families of the fungi (e.g., ergot) (8).
Alkaloids generally occur in the plant tissue a t points
of intense cell activity such as the leaves, roots, bark,
and seeds and are generally regarded as plant met,abolic .
byproducts. There is considerable variation in the
alkaloidal content in the plant depending on such factors
as time of reaping and methods of cultivation and selec-
tion.
A knowledge of the botanical origin of an alkaloid
can be useful in giving indications of the structural type
of alkaloid which is likely to be present. Plants of
the same family generally tend to produce alkaloids of
the same structural type. This is a generalization which
has to be applied with caution for there are alkaloids
such as nicotine and berberine which are found in
several different plant families. Also plants such as the
opium poppy give a number of different structural
types of alkaloids though the latter are probably bio-
genetically related.
Alkaloids occur usually as salts of the common plant
acids. I n certain plants, however, a specific acid may
be found associated with certain alkaloids such as
quinic acid in the cinchona group, aconitic acid in the
aconitines, and meconic acid in the opium group. A
few alkaloids such as narceine and narcotine are found
free in nature, probably because of their low basicity.
A few alkaloids as found in the solanum and veratrum
groups occur as glycosides of such sugars as glucose,
rhamnose, and galactose. Other alkaloids as in the
 Table 1. The Occurrence and Use of Some Common Alkaloids
Alkaloid Source Use
Arecoline Betel nuts Masticatory; vermifuge
Atropine Belladonna Mydriatic in ophthalmalogy
Caffeine" Coffee, ten Stimulant
Cocaine Coca. leaf Local anaesthetic
Codeine Opium Pain reliever
Colchicine Crocus Plant genetics
Cusparine Angustura bark Stimulant
Emetine Ipecac roots Emetic; amoebic dysentery
Ephedri?e "Ma Huang" Asthma
Ergonomne Fgot Obstetrics, oxytocic
Febrifugine Ch'ang Shan" ~nti-ma1a;ial
Mescaline "Mescal buttons" Hallucinatory principle
a Sometimes classed as an alkaloid cf. (Ic).
tropane, aconitine, senecio, veratrum, and yohimbine
groups are encountered as esters of various organic
acids of varying complexity. A few others occur as
amides such as piperine and the ergot alkaloids.
The study of the chemistry of the alkaloids has been
particularly att,ractive to chemists partly on account
of their wide occurrence and diversity of physiological
action on the animal organism and partly on account
of the complex structural and synthetic problems in-
volved in the elucidation of their structure. Many of
these investigations have been classics in the realms of
structure elucidation and synthesis. The occurrence
and uses of some of the commoner alkaloids are shown
in Table 1.
Isolation Methods
The presence of alkaloids in plants is first established
by tests on extracts from the plant with various alka-
loidal reagents (Sa). The presence of a precipitate or
turbidity with such common reagents as Mayer's
(potassium mercuric iodide), Wagner's (iodine-potas-
sium iodide), Dragendofi's (potassium bismuth iodide),
Sonnenschein's (phosphomolybdic acid), and Scheibler's
(phosphotungstic arid) indicates the presenre of alka-
loids.
"Mayer'e resgmt is most commonly used as it is generally the
most sensitive. About 1 g of plant material is digested with
5-10 ml of dilute HCI and the filtrate tested with a. few drops
of the reagent. A creamy precipitate or turbidity gmcrally
indicates the- presence of alkaloids."
The extraction of the total alkaloids from the plant,
material is generally accomplished either by extraction
with acids or with organic solvents. Various modifi-
cations have been worked out empirically for the ex-
traction of particular alkaloids. Solvent extraction is
usually the more common procedure. The dried plant.
material is usually subjected to a preliminary extraction
with light petroleum to remove plant oils. Most alka-
loids and their salts are insoluble in petroleum. The
alkaloid bases are then liberated from the salts present
in the plant material by treatment with ba.se (calcium
hydroxide, ammonium hydroxide, or sodium carbonate)
and then extracted with a suitable organic solvent surh
as chloroform, ether, benzene, ethanol, methanol, or
methylene chloride. The alkaloids are then extracted
with acid from the organic solvent or pasty residue that
remains after removal of the solvent. The acid solu-
Alkaloid Source Use
Morphine Opium Pain relief. narcotic
Nicotine Tobacco Stimulant: insecticide
Papaverine Opium Spasmolytio drug
Physo~tigmine Calabar bean Gastrointc~tinel and bladder
stimulant
Pilocarpine Jaborandi Diaphoretic (sweat producing)
Quininr Cinchona bark Malaria fever
Reserpine Rauwolfia Tranquilizer
Scopolamine- 1)atura Narcotic
Sparteine Broom Cardiac treatment
Strychnine Strvchnos Pest exterminator
Tubocurarine Chondrodendron Nerve paralysis
Yohimhine Yohimbe bark Aphrodisiac (sex stimulation)
tion is made basic and again extracted with an organic
solvent usually ether or chloroform to give a crude mix-
ture of bases. Some alkaloid hydrochlorides are soluble
in chloroform and this property can be used for purposes
of separation (5b). A modification of the above process
is the extraction of the plant material with an aqueous-
organic solvent mixture and subsequent separation and
drying of the organic layer. Non-basic alkaloids can
usually be obtained after removal of any acid or alka-
line constituents of the organic layer.
Acid extraction, though less frequently used, has
been applied with success to the isolation of alkaloids
from undried plant material in the field using ion ex-
change techniques. Alternatively the dried plant
material is extracted with a dilute acid such as sulfuric
or tartaric. Tartaric acid has the advantage of tending
to give a purer product. Where there is danger of
decomposition of the plant base, the extraction is done
in the cold. The aqueous solution is hasified and either
extracted with an organic solvent or steam-distilled to
remove volatile components. Quaternary alkaloids
which are usually quite soluble in water are generally
recovered by precipitation with mercuric chloride or
Mayer's reagent and the resulting mercury complex de-
composed with hydrogen sulfide. Some alkaloids which
occur as N-oxides are quite soluble in water. They
may be obtained by reduction with acid zinc dust fol-
lowed by chloroform extraction. These methods are
summarized in Table 2 with appropriate examples.
Separation and Purification Methods
In the early days of alkaloid chemistry the separation
of mixtures of alkaloids often involved tedious frac-
tional rrystallizations or precipitations. Nevertheless,
excellent separations were achieved. However the
complexity of some alkaloid mixtures is such that for
a time only the more abundant components could be
examined. The advent of more elegant techniques of
separation such as counter-current distribution, ad-
sorption, and partition chromatography have made pos-
sible the study of many of the minor components (10).
Ion exchange, though potentially a good method, has
had little application in separation. These techniques
are too well known to require elaboration. A summary
of the main methods of separation with examples of
their application is given in Table 3 and Figures 1and 2.
Some criterion of the effectiveness of a separation is
required. Paper chromatography and to a lesser ex-
Volume 37, Number 9, September 1960 / 455
 Table 2. Examples of Alkaloid Isolation Methods
Acid Extraction Methods Solvent.k>xtraction Methods

Cyclic Percolation of dried plant material with Extraction Extraction Addition of

extraction dilute acid, or hot extraction base and ex-
of undried tractor with
plant and organic sol-
absorption
on ion
exchange
lcolumn ( 4 ) 1 I

Concentration and basification I I Separation

of solvent
Precipitation Acidification
with alcohol of quaternary

%-
& removal of
and ammonia alkaloids a i t b loils with I
mercuric chloride components petroleum

Removal of and extrac-

tion with

'i
distillate solvent organic
solvent
of base by from ~olution
extraction

I Crude Alkaloids

tent paper electrophoresis have been useful in this con-
nection (Table 3, Fig. 3).
The individual alkaloid is characterized by constancy
of its melting point, boiling point, optical rotation,
ultraviolet spectra, infrared spectra, and any other suit-
able physical properties, and also by the preparation
and characterization of derivatives and salts. Often
the purification of an alkaloid through one of its salts
is more convenient; a useful test for homogeneity is
the inter-convertability of the alkaloid and its salts.
Quantitative methods of estimation of alkaloids which
are generally either colorimetric, volumetric, or gravi-
metric depend largely on the types of functional groups
present (Sd).
ELUATEFRACTIONS
Figure 1 . Partition chromatographic seporotion of pomegranate root
bork d k d o i d s [by permission from the J. Phorm. and Phormocol.) 1141.
456 / Journal of Chernicol Education
Structural Investigation b y Physical M e t h o d s
Physical methods are becoming increasingly impor-
tant in the structural elucidation of chemical com-
pounds. A few examples of each method must suffice
to illustrate the approach. It should also be em-
phasized that physical and chemical methods go hand
in hand in structural investigation.
Ultraviolet Spectra
UV spectra have provided the keys to the solution
of many problems in alkaloid chemistry. W spectra
may he useful in pointing to the type of structure
likely to be found. Broadly speaking the spectra of
alkaloids may be divided into three main classes (26).
No UV absorption in the range 200-1000 m#. Examples are
the saturated ring systems such as the pyrrolidine and piperi-
dine alkaloids.
Simple aromatic chromophoric systems such as substituted
benzene or pyridine with medium intensity absorption a t rerelsr
t i d y short wavelengths (230-300 mr),
Alkaloids with complex ehromophores, either aromatic
nuclei in conjugation vith other chromophores or fused aro-
matic systems.
Examples of the use of spectral methods in structure
determination are seen with the alkaloids sempervirine
(27), strychnine ($a), and emetine (29).
The use of UV spectra in the assignment of structures
is illustrated by the following example from the mor-
phine series. The structure of the two dihydro the-
baines was corrected by the use of UV spectra (30).
The structure of the lithium aluminum hydride re-
duction product was shown to be I1 since the extinction
values indicated a conjugated system. The structure
of the sodium and ethanol reduction product was shown
to be I11 since the extinction value showed a marked
lowering indicating the absence of a conjugated double Table 3. Alkaloid Separation Methods
bond system. General
Example of Refer- refer-
Method rtnnlicat,ion ence ence
Fractional
crystallisation
Quinine alkaloids . .
(11)
Fractional Hemlock alkaloids Xe)
distillation
Senamtion bv Verrttrum alkaloids 12
ksction
Adsorption
- by acetylation
Senecio alkaloids 15 19, 20
chromatography
Partition Pomegranate root 14 19
chromatography bark alkaloids (Fig. 1)
Counter-current Veratrum alkaloids 15 dl(a, b )
distribution (Fig. 2)
Ion exchange Yohimbine alkaloids 16 W
Paver Curare alkaloids 17 19. 25
Ehromatography (Fig. 3)
Paper electrophoresis Stryohnos alkaloids 18 B , 2 4 25

I. Thehaine 11. LiA1H4product 111. Na/EtOHproduct

r = 6500 r = 11,000 r = 2000
A = 283 mp X = 284 mp A = 282 mp
The UV fluorescence spectra of some alkaloids have
also been determined (31).
Infrared Spectra
Infrared spectra have been extensively used in the
elucidation of the structures of organic molecules
(9% 99, 94).
Some of the commoner functions found in alkaloids
and their characteristic infrared absorption frequencies
are shown in Table 4. The infrared spectra of alkaloids
have been used for comparative purposes especially in
the fields of drugs and narcotics (95), and also in fol-
lowing the course of synthetic and degradative ex-
periments.
Thus the infrared spectra were used to reformulate
the course of the synthesis of N-methylisomorphinane,
IV. One of the intermediates in the synthesis was the
postulated nitrile, V. This was shown to be incorrect
as the infrared spectrum showed no nitrile bands a t
2240 cm-', but showed bands at 3400 em-', (secondary
amine), 1664 em-' (cyclic amide), 1676 cm-', and 1692
em-' (cyclic conjugated carbonyl). The latter ob-
sewations could be reconciled by a structure VI for
the intermediate (96).
Basic Sfrengtl~sand Polarogmphic Measurements
The determination of pK values has become increas-
ingly important in structure determination and par-
ticularly in the elucidation of structural details. An
example of this application is in the placing of the
double bond in the structure of retronecine, VII, the
alkanolamiue hydrolytic cleavage product of the al-
kaloid monocrotaline (4ia). The a-p or 8-7 position-
ing of the double bond relative to the nitrogen atom
was correctly postulated on the basis of pK measure-
ments and subsequently confirmed by other methods.
Retronecine is a weaker base than the corresponding
saturated amine platynecine, VIII. This is compatible
with a p-y unsaturation whereasau w-8 type of unsatura-
tion would lead to an increase in basic strength due to
the facile transformation:

-N-C=C-
'
I

A
+ H+ - +
-N=C-
'
H
A-
1
H

VIII
pK, 10.22

The application of pK values to the steric arrange-

ment in some terpenoid alkaloids has been discussed
(43).
The presence of a tropolone ring in colchicine was
postulated on the basis of the similarities in the polaro-
grams of colchicine and ythujaplicin (49).
X-Ray Crystallography
No.E
TU~ The application of X-ray crystallographic methods
Figure 2. Twenty-four-tube counter-current distribution of veratrum
to the complex three-dimensional structures often en-
alkaloids (by permission from Anal. Chem.) 175). countered in alkaloid studies has been very rewarding
Volume 37, Number 9, September 1960 / 457
 Nuclear Magnetic Resononce Spectro
The increasing interest in NMR spectra lies in the
fact that many details of structure can be decided
unambiguously by this method though this may not
be possible using ultraviolet or infrared spectra.
Also simple model compounds can be utilized to obtain
a "fingerprint" of the portion of the spectrum under
consideration. This is analogous to the use of model
compounds in UV and I R work but in some cases the
model suitable for NMR comparison may be more
readily available.
An example of the application of the method in the
complete proof of the structure of the alkaloid Luna-
crine, XII, is outlined below.
0

XI1
I I I I I
The structure was elucidated by a detailed analysis
I 6 1 4 1 T- of seven well defined groups of lines in the NMR
Figure 3. Paper chrornotogrorn of curare alkaloids (from Helv. Chim.
A < f d (17).
spectrum which were assigned as follows (49): (a)
the hydrogen atom at position 5 of the quinolone system;
(b) the aromatic hydrogen atoms at 6 and 7; (c) the a-
(41b). Classical examples of the application are the hydrogen atom of the dihydrofurano ring; (d) the
studies of the alkaloids strychnine and colchicine.
Thus the X-ray spectral method made it possible to Table 4. Some Infrared Absorption Characteristics of
distinguish between the structures I X and X for the Alkaloids
alkaloid colchicine and showed that the structure X was
Infrared absorption om-1
the correct one. S = Strong
Intensity: M = Medium Refer-
NHCOCH, Group W = Weak ence
-0H- 36253540 ( S ) non-associated 3b, 37
3400-3200 (S) associated
Phenolic acetate 1767-1764(S) 38
Alcoholic acetate 1742-1735 (S)
C=O 1850-162O(S) 37
IX 0 C=N- 1660-1480 (S) 3s
x OCHs
C=C 1680-1620 ( S M I 32
Recently the structure of the complex alkaloid gelse- near 3030 (W-M-S) Sb, 37
mine, XI, has been independently postulated by both 1640-1600 (S)
near 1500 (W-M-S)
chemical and X-ray methods (44).
32
..
3480-3460 ( 8 ) indale 37
\NH 3480-3440 (S) 2' amines
/
3280-3200
3380-3205
(S) associated
(S) 2nd amine 1 39
\$/
Rotation01 Evidence
Ammonium 244k2350 (W)
/ H' 40
Optical rotational evidence has been used in a few c3
cases to determine stereochemical configurations. Thus =N-
Immonium 2100-1980 (W)
rotational evidence was used to establish stereochemical
correlations between yohimbine and known steroid
4
NHsf2" Aminesalt 1620-1560 (S-M) deformation
structures (45), and the absolute configuration of eme- mode
tine has been deduced from optical rotational differences Hydrochloride salts 3000-2500 (M-W) Series of bands Sb
(46).
Optical rotatory dispersion has had limited applica- methoxyl- and N-methyl-hydrogen atoms; (e) the
tion in the field of alkaloids, largely due to lack of suit- &hydrogen atoms of the dihydrofurano ring; (f) the
able instruments, though some correlations between tertiary hydrogen atom of the isopropyl group; and
the diterpenoid alkaloids and the diterpenes have been (g) the C-methyl-hydrogen atoms.
made (471, and recently the absolute configuration of Other examples of the application of NMR spectra
emetine has been independently determined with the in structure determination are illustrated by studies on
aid of optical rotatory dispersion curves (48). the alkaloids lycodine (50a) and gelsemine (506).
458 / Journal o f Chemical Education
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Volume 37, Number 9, September 1960 / 459