CHEMOTAXONOMY

1.1. CHEMISTRY OF NATURAL PRODUCT: SCIENCE OF ALL TIMES

Natural products, as the term implies, are those chemical compounds derived from living

organisms and the study of natural products is the investigation of their structure,

formation, use and purpose in the organism. Drugs derived from natural products are

usually secondary metabolites and their derivatives. Today those must be pure and highly

characterized compounds. Since prehistoric times, the humans have relied on natural

products as a primary source of medicine. Plants and animals were used to bring back the

health of sick and frail. Plant were found to be beneficial as food, fodder, medicine etc. but

also harmful as being poisonous and toxic (Fuller and Hemrick, 1985). The application of

herbs for external and internal use has always been a major factor in practice of

medicine (Steiner, 1986). The experience and knowledge gained in using the traditional

medicines in different regions over the millennia resulted in the complex science of modern

medication.

The various approaches to drug discovery from nature are:

Ethnobotanical: Ethnic and traditional medicine

Random screening: Bioassay guided routes

Chemotaxonomic: Screening of relatives

1.2. TAXONOMY OF PLANTS

Biological classification or scientific classification, is a method by which

taxonomists group and categorize plants by biological type, such as genus or species.

The classification of plants by grouping data according to morphological similarities is

probably the oldest and most widely-used of all the approaches (Quinlan, 1993).

However many approaches evolved over time towards the taxonomy of plants.

1.2.1. CAUSES OF TAXONOMIC COMPLEXITY

Morphological variants of a species results because of different factors. In addition,

many species exhibit considerable genetic variation, both florally and vegetatively. This

variation may occur in different populations of the same species, or may characterize

different infraspecific categories of a species. Sometimes, the character may be genetically

controlled in one species, but phenotypically plastic in another. Quaternary climatic

changes have had a profound impact on speciation, structuring of genetic diversity and

the shaping of the present-day distributions of plant and animal taxa (Avise, 2000; Hewitt,

1996, 2000, 2004; Vuilleumier, 1971). Oscillations of population sizes, bottle necks,

founder events and other population historical events associated with climatic shifts

have further contributed to differentiation among regional population groups. As a

combined effect of range shifts and population differentiation, divergent lineages have

occasionally formed contact zones, leading to reticulate speciation by means of

hybridization and polyploidization (Grant, 1981; Stebbins, 1984). Polyploid speciation has

long been recognized as an important process in plant evolution (Mntzing, 1936;

Stebbins 1950; Grant, 1981). Recent genomic studies have made it clear that

angiosperms possess genomes with considerable gene redundancy, indicating that most

(if not all) plants have undergone one or more episodes of polyploidization (Soltis et al.,

2003).

1.3. CHEMOTAXONOMY

Chemotaxonomy is also called chemosystematics or biochemical systematics. The

science of chemical taxonomy is used on the classification of plants on the basis of their

chemical constituents which are deeply concerned with the molecular characteristics.

The method of chemical taxonomy is simple in principal and is based on the investigations

of the distribution of chemical compounds or groups of biosynthetically related

compounds in series of related plants. Different plants sometimes contain substances

which although belong to different chemical compounds appear to be biosynthetically

analogous. Such plants may contain similar enzyme systems, and the compounds produced

by such enzymes are indicative of the relationships that exist between the plants. However,

the chemotaxonomic studies include the investigation of the patterns of the compounds

existing in plant. Climatic conditions have a major influence on the distribution of plants

containing certain substances e.g. fats, volatile oils, alkaloids, flavonoids etc. It is well

known that for tropics, and perhaps for all climates, the chemical products are highly

organized. According to Reichert (1919), it is possible to identify many plants by their

starch grains. Stress has been given on the importance of -Cyanins and -Xanthins in plant

taxonomy. -Cyanins are commonly met within the families of order centrospermae. The

other chemicals are also found specifically in particular orders or families of flowering

plants e.g. Isoquinoline (Alkaloids) is found in the families of Ranales; retanone in the

families of leguminales, biflavonoils in casuarinas equisetifolia (casuarinaceae). The

presence of such chemicals in different groups of plants has great taxonomic significance

(Stuessy, 2008). Taxonomic studies for various plant taxa by using different parameters

have been successfully carried out in Pakistan including that of cereals (Ashraf et al.,

2003), Legumes (Ahmad et al., 2007) and Maize (Nawaz and Ashraf, 2007).

1.3.1. APPLICATION OF CHEMOTAXONOMY (Stuessy, 2008)

There are a few angiospermic taxa which are characterized by specific

compounds of general occurrence. For example leaving aside the family of

caryophyllaceae, the rest of the families such as chenopodiaceae, amaranthaceae,

aizoacaceae etc. of the taxon caryophyllales (centrospermea) contain -cyanin a colored

substance but differs from anthocyanins. It appears that, with the exception

caryophyllaceae, these families are closely related and therefore caryophyllaceae may be

isolated. -cyanin also occurs in cactaceae and therefore, the members of caryophyllales

are phylogenetically related. There are certain other chemical connections between

cactaceae and members of caryophyllales e.g. common presence of isoquinole

alkaloids in Salsola of Chenopodiaceae and cactaceae. Another example that may be

cited is in the family crucifereae, where unsaturated acid erucic acid is prominent and

also in Tropaelum erucic acid is present; it indicates the relationship between Geraniales

and Rhocadales. In umbellifereae and Araliaceae petroselinic acid (a structural isomer of

Oleic acid) occurs and these two families are related and belong to the same order. The

other examples are from Magnoliales Ranals taxa, where it is shown that

magnoliaceae, lauraceae, Ranulculaceae, Annoraceae, the alkaloid isoquinoline is present,

this supports that these families are loosely related. On the other hand

Asclepiadaceae and Gentianaceae are allied due to the common occurrence of pyridine. The

lilliaceae and Amarylldeceae are closely associated and this is supported by the

presence of Isoquinoline in both. A number of citations regarding chemotaxonomy

and secondary metabolites further.

1.4. FAMILY SOLANACEAE

The Solanaceae, to which the genus Solanum belongs, is a cosmopolitan family

which is widely distributed throughout tropical and temperate regions of the world, with

centers of diversity occurring in Central and South America and Australia. It is

composed of approximately 84 genera and 3000 species. The name of the family comes from

the Latin word Solanum, meaning "the nightshade plant", but the further etymology of that

word is unclear; it has been suggested it originates from the Latin verb solari meaning "to

soothe". This would presumably refer to alleged soothing pharmacological properties of

some of the psychoactive species found in the family. It is more likely, however, that

the name comes from the perceived resemblance that some of the flowers bear to the

sun and its rays, and in fact a species of S. nigrum Complex (Solanum retroflexum) is

known as the sunberry. The family is also informally known as the nightshade or potato

family (Yasin, 1985).

1.6. SOLANUM NIGRUM: TAXONOMIC COMPLICATIONS

Solanum nigrum is the most variable species of the genus Solanum. The species related

to S. nigrum have been reclassified innumerable times. Characters used by later

taxonomists to separate and describe additional taxa often differed very slightly from those

given for species by earlier workers. These Solanum species display varying amounts of

phenotypic variation, particularly in their vegetative features such as plant habit, leaf

size and form, and stem winging. In addition, senescence is often accompanied by smaller

and fewer flowers and fruits (Ganapathi and Rao, 1986).

S. nigrum was first delimited in four taxa with polynomials by Dillenius. Linnaeus

subsequently modified Dilleniuss work, describing these in six varieties under the

binomial S. nigrum (Edmonds and Chweya, 1997). Since then, the plants morphologically

related to S. nigrum have been reclassified many times. Over 300 post-Linnean specific

and infraspecific names have now been published, and synonymy is extensive within

the section. However, no satisfactory revision of the whole section has yet been devised.

The boundaries between many of the species are still ill-defined, with many of the new

taxa proving to be no more than slight morphological variants of those already

described. The situation is further complicated by the researchers who either treated

different members of the section as varieties of S. nigrum or considered them as

different species on the basis of morphological differences (e.g. Edmonds and

Chweya, 1997; Schilling and Andersen, 1990; Stebbins and Paddock, 1949; Symon,

1970). These Solanum species display varying amounts of phenotypic variation,

particularly in their vegetative features such as plant habit, leaf size and form, and

stem winging. In addition, senescence is often accompanied by smaller and fewer flowers

and fruits than usual. Natural hybridization is probably more widespread in this section

than generally supposed. It is now named as Solanum nigrum Complex because it is

composed of a large number (about 30) of morphologically distinct taxa (Schilling and

Andersen, 1990). Only during the revision of Solanum section Solanum appear in 1979

for Flora Europaea 3, drawn turned out that in Europe two different forms of the species

coexist. The most widespread form was considered subspecies S. nigrum ssp.

nigrum, the second, rarely encountered species as S. nigrum ssp. schultesii classified

(Edmonds and Chweya, 1997).

Deadly nightshade, identified as S. nigrum, causes belladonna poisoning (Hubbs, 1947),

with symptoms including widely dilated pupils (characteristic of the atropine group, but

either unexpressed, or expressed very mildly, in poisonings by plants whose major

poisoning principle is of the solanine group). Deadly nightshade is identified as S. nigrum,

but black nightshade is botanically unidentified and gives a strong cat's-eye test for atropine

(Case, 1955). According to a research at University of Pennsylvania, unripe berries are said

to be more toxic than ripe berries. Berries are more toxic than leaves which, in turn, are

more toxic than stems or roots. Overall plant glycoalkaloid content is often higher in the

autumn than in the spring. These problems clearly state the importance of proper

identification and detailed composition analysis of each taxa of S. nigrum Complex.

The taxa of S. nigrum Complex are difficult to distinguish because

1. They are morphologically similar.

2. These species are all highly phenotypically plastic.

Three taxa belonging to S. nigrum Complex viz.: S. americanum Mill., S. nigrum L.

and S. villosum Mill. had been reported in Pakistan (Schilling and Andersen, 1990).

S. chenopodioides Lam. and S. retroflexum Dunal are two other species that were found

growing wild in and around Botanic Garden, GC University, Lahore. Morphologically S.

nigrum is different from S. villosum in the respect that the former has black matured berries

with peduncles longer than pedicels while latter has orange/orange-red matured berries and

peduncles shorter than or equal to the pedicels. Classification of S. nigrum and S.

villosum as varieties or distinct species started taxonomic controversy between Linnaeus

and Miller (Edmonds and Chweya, 1997). Though S. americanum Mill., S.

chenopodioides Lam. and S. retroflexum Dunal have morphological resemblance with S.

nigrum, yet no chemotaxonomic relationship has so far been established due to lack of a

comprehensive study of their chemical composition.

1.7. BOTANICAL ASPECTS OF THE INVESTIGATED TAXA

The five locally available taxa investigated were:

1. Solanum americanum

2. Solanum chenopodioides

3. Solanum nigrum

4. Solanum retroflexum

5. Solanum villosum

These Solanum species display varying amounts of phenotypic variation, particularly in

their vegetative features such as plant habit, leaf size and form, and stem winging. In

addition, senescence is often accompanied by smaller and fewer flowers and fruits than

usual, while the gene for anthocyanin pigmentation in flowers seems to be dependent on

light intensity and temperature for its expression, in some species. It is therefore often

difficult to define the limits within which such features are genetically fixed (Baylis, 1958;

Henderson, 1974; Edmonds, 1977).

Natural hybridization is probably more widespread in this section than generally

supposed. Though this is probably followed by subsequent genetic breakdown in F1

or F2 generations (Edmonds, 1977), it may also be followed by back-crossing to the

parental species. This would result in morphogenetically complex population variation:

the collection of specimens from such populations would explain some of the

difficulties encountered in the morphological differentiation of these species in the

herbarium (Edmonds, 1979).

1.11. AIMS AND OBJECTIVES

There are various approaches to the taxonomic studies of the plants based on the structural,

cytological and chemical constituents. The ancient classification of the plants was mainly

carried out on the comparative morphological and anatomical concepts of the natural

plant flora.

However with the rapid progress in the isolation, purification, identification, elucidation of

structure and the configuration of natural plant products, the phytochemists and

ethnobotanists believe that it is possible to characterize and classify the plants on the

basis of their chemical constituents. The chemical constituents are formed within the plants

by definite biosynthetic pathways aided by the specific enzymes.